AQF 306: Fish Parasites and Diseases

General Information

Reading list: literature materials will be made available by the lecturers through the class
representative at no cost. Students should therefore make the effort not only to read them but also
make their own copies

Attendance: Attendance and active participation in class discussions are very essential.

Continuous Assessment: Continuous Assessment Tests (which may perhaps be in form

quiz, practical, group assignment) in addition to regular class assignments are obligatory.

Absence: In the event that one must unavoidably miss a class, please send a written note to the
lecturer. The lecturer too, will as well inform the class ahead of time if he must predictably be
absent. For the convenience of the students and the lecturer, students are encouraged to see the
lecturer in the office and preferably by appointment.

Course Synopsis
Identification, morphology, taxonomy, life history of fish parasites. The ecological and
pathological effects of parasites and diseases of fish. Epidemiology of parasites populations in
water body. Common bacterial, fungal and viral fish diseases and their control. Other enemies of
fish. International restriction binding the transportation of fish across the country boundaries.
Fish pond and public health. 15h(T), 45h(P);C

15h (T), 45h (P); Compulsory

SCHEME FOR LECTURES

Part I

Lecture 1 Identification, morphology, taxonomy, life history of fish parasites.

Lecture 2 The ecological and pathological effects of parasites and diseases of fish.

Lecture 3 Epidemiology of parasites populations in water body.

Part II

Lecture 4 Common bacterial, fungal and viral fish diseases and their control.

Lecture 5 Other enemies of fish.

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Lecture 6 International restriction binding the transportation of fish across the country
boundaries.

Part III

Lecture 7 Practical Identification of fish fungi

Lecture 8 Practical Identification of fish bacterial

Lecture 9 Practical Identification of fish virus

Lecture 10 Practical Identification of fish parasites

Lecture 11 Practical Other enemies of fish

Part IV

Lecture 12 Assessments/ test/group work/exam

This course will be handled by two lecturers:

Dr. M.I. Abubakar and Dr. .I. Adeshina.

CHAPTER ONE
1.1 INTRODUCTION
Fish Pathology is a branch of Ichthyology that use the principles and
methods of microbiology, mycology, virology, bacteriology, toxicology,
pharmacology, biochemistry, anatomy and physiology, hematology and
histopathology. It is a multidimensional discipline.
Pathology is the study of disease “Patho” (disease) and “logy” (study). It is
devoted to the study of the structural, biochemical and functional changes in
cells, tissues and organs that underlie disease by the use of molecular,
microscopic, immunologic and morphologic techniques. Pathology attempts
to explain the ways and wherefores of the signs and symptoms manifested
by morbid organism while providing a rational basis for treatment and
therapy. Pathology is divided into general and systemic pathology. The
former is concerned with the reactions of cells and tissues to abnormal
stimuli and inherited defects, which are the main causes of diseases. The
latter (Systemic) examines the alterations in specialized organs and tissues
that are responsible for disorders that involve these disease/organs.

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 The history of pathology can be traced back to the period of antiquity when
people began examining bodies. The oldest civilized people (Chinese, India
and Egyptians) were under the impression that disease occurred when
demon or evil spirits were displeased. The medicine men were concerned
with appeasing these evil spirits. The Egyptians began to influence medicine
around 4000BC. The Greek Culture had a profound effect on the scientific
approach to medicine. Greek physicians elucidated the principles of exact
and careful clinical observations.
Pathology began to develop as a subject during the 19 th century through
teachers and physicians that studied related disciplines. They referred to it as
“Pathological anatomy” or “Morbid anatomy”. However, pathology as a
field of medicine was not recognized until the late 19 th and early 20th
century. Pathology has found its root extended to the Ichthyology with
introduction of “Fish Pathology” as a course of study. There is however,
paucity of information in fish pathology, useful information are obtained
from allied areas and published journals which are greatly inadequate.
This book will therefore, fill the vacuum and thus, be useful for lecturers,
fish farmer, researchers and students of Polytechnics and Universities
pursuing courses in OND, HND, B. Sc., M. Sc. and PhD in fish pathology.
1.2 BASIC CONCEPTS AND DEFINITIONS IN PATHOLOGY
Pathology addresses four components of disease: cause (aetiology),
mechanisms of development (pathogenesis), structural alterations of cells
(morphologic changes) and the consequences of changes (clinical
manifestation). It is pertinent to define the various concepts used in
pathology for better understanding.

Pathology: - Scientific study of the

molecular, cellular, tissue,
organ and system response
to injurious agents or
adverse influences.
Disease: - State of critical departure from the normal with clinical
manifestations of signs or symptoms. It is a state in which an
organism exhibits an anatomical, physiological, psychological
or biochemical deviation from the normal.
Aetiology: - Study of the cause of a disease. An aetiologic agent is the
factor (fungi, virus, bacterium, etc) responsible for lesions or a
disease state.

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Predisposing causes of diseases: - Are those factors which make an
organism more susceptible to a disease (damp weather, poor
environment; etc).
Exciting Causes of Disease:- are those factors which are directly
responsible for a disease (bacteria, fungus, viruses, hypoxia,
chemical agents, etc).
Clinical Sign: - is any functional evidence of disease which can be
determined objectively or by the observer (swelling, situated
growth, paleness, pop eye,). It is observable by the pathologist.
Chemical:- is any substance obtained by a chemical process or producing a
chemical effect on a living system.
Enzyme:- is a substance produced by living cells which aids in speeding
up the process of chemical reactions in the body.
Clinical Symptom:- is any functional evidence of a disease that can be
determined subjectively or by the organism only as in
convulsion, opaque eye, swollen belly, etc.
Prognosis:- is an outcome of a disease in an organism. It is the clinician’s
estimate of the severity and possible result of a disease.
Diagnosis:- is the exact determination of the nature or kind of a disease
expressed. Diagnosis is made on the basis of the specific
disease entity involved while a clinical diagnosis is made on the
basis of clinical signs observed in the ailing organism.
Pathogenesis:- refers to the progressive development (sequence of events)
of a disease from the time it is initiated to its final stage in
recovery or death.
Serum:- is the part of the blood that is left after blood has been allowed
to clot and the blood cells have been removed, it is often watery
and yellow in colour.
Plasma:- is also a supernatant (that part of the blood left above sediment)
after centrifuging uncoagulated whole blood.
Chemical Asphyxiant:- is a substance that has the ability to deprive tissue
of oxygen or render the body incapable of
utilizing an adequate oxygen supply.
Carcinogen:- any agent that can initiate or promote the development of
malignant or potentially malignant tumours, malignant
neoplastic proliferation of cells or that possess such material is
called carcinogen.
Carcinogenicity:- is the ability to cause or stimulate cancer.
Acute exposure:- this is a single exposure or multiple exposures occurring
over one or two days.
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Acute or immediate toxicity:- this is the rapid appearance of toxic effects
just after acute exposure.
Chronic Exposure:- is multiple exposures continuing over a longer period
of time (more than three months).
Chronic Toxicity:- is the manifestation of the toxic effects after a prolonged
or chronic exposure to a toxicant.
Clinical Toxicologists:- carry out toxicity testing (as part of clinical trials)
in clinics or laboratory.
Drug:- is any substance that brings about a change in biologic function;
mainly through its chemical actions (fewer drugs have physical
effects).
Environment:- includes all the surroundings of an organism particularly the
air, land, water and all plants and animals living therein and the
interrelationships, which exist among these or any of them.
Invitro Testing:- Study that involves the use of a part or sample of the
organism separated from the entire system.
Invivo Testing:- Study that involves the use of whole organism.
Poisons or Toxicants:- are drugs that produce harmful effects when given at
a certain dose.
Toxin:- these are poisons of biologic origin, that is, from plants or
animals.
Toxicity:- this is the capacity of a chemical agents to cause injury.
Systemic Toxicity:- is the toxic effect that required the absorption and
distribution of the toxicant to a part distant from site of entry
where effects are produced.
Xenobiotics:- These are substances absorbed across the lungs, skin or
ingested intentionally or accidentally.
Toxicology:-Scientific study of substances with harmful effects on the
living systems, from individual cells to complex ecosystems.
Marine Toxicology:- is the study of pollutants and other chemical products
in large waters and their effects on the living organisms therein.
Hyperplasia:- this is an increase in the number of cells in an organ or tissue,
usually resulting in increased mass of the organ or tissue.
Atrophy:- this is a reduced site of an organ or tissue resulting from a
decrease in cell size and number.
Neoplasia:- this means “new growth” and a new growth is called a
neoplasm.
Moribund Fish:- Fish that are alive, but showing signs of the suspected
disease.
Morbidity:- is the condition of having a disease.
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 Disease Fish:- These are fish whose behaviour or physical conditions
indicate the presence of a disease agent.
Congenital Disease:- Congenital disease are diseases or deformities with
which the fish is born e.g twisted backbones, missing gill
covers and fins.
Traumatic Disease:- These are produced by injuries, usually caused by
other fishes during fights, or sometimes when the fishes jump
out on being netted.
Degenerative Diseases:- These are caused by increasing age and fishes
develop heart failure, kidney failure which lead to dropsy,
skeletal deformities, swim bladder and balance troubles.
1.3 MICROBIOLOGY: Is the study of micro organisms. These are organisms
that too small to be seen clearly by the unaided eyes. Micro organisms
include bacteria, fungi, algae, protozoa and viruses. Most micro organisms
are unicellular, in unicellular organisms all the life processes are performed
by a single cell.
1.4 CHARACTERISTICS OF MICRO-ORGANISMS
To understand and appreciate micro-organisms, it is imperative that a good
knowledge of their characteristics will aid pathologist in studying and
identifying the micro-organisms concerned.
1.5 Mycology: Study of Fungi
1.5.1 CHARACTERISTICS OF FUNGI
Fungi are plants with thread like non-photosynthetic threads called hypha
(ae). The non-photosynthetic thread (hyphae) are either septate with cross-
walls (non-coenocytic) or without crosswalls (co-enocytic). A collection of
these hyphae are referred to as mycelium. Fungal hyphae are bounded by a
cell made up of glucans and chitin. The eukaryote contains a nucleus with a
nuclear membrane enclosing the chromosomes. Also present are
mitochondria and endoplasmic reticulum. These characteristics differentiate
fungi from bacteria.
Generally, fungi are known to have the following characteristics:
- The body or vegetative structure is called a thallus.
- They are chlorophyless, eukaryotic unicellular or multicellular plants.
- They reproduce asexually by dispersing different types of spores and
sexual reproduction can be homothalic, heterothallic or by conjugation.
- Cause diseases in plant (parasitic) and animals (dermatophytic).
- They have ecological impact as molds, yeasts, lichens, and parasites.
- They are heterotrophs, acquiring their nutrients by absorption, consisting
of saprophytic, decomposers, parasitic species and mutualistics forms.
- They are important decomposers of food and other useful objects.
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1.6 CHARACTERISTICS OF VIRUSES
Virus is a Latin word which means poison or toxin. They are obligate,
intracellular parasites which utilize the synthesizing machinery of their host
cells for:
1. The expression and replication of their genetic material;
2. The formation of specialized elements that permit the transfer of the
replicated genetic material to other host cells.
SIZES:
Units of Measurement.
1mm = 10-3m
1µm = 10-6m (10-3mm)
1nm = 10-9m (10-3pm)
(1Ao(Angstron unit) = 10-8m i.e. 10nm)
Dimension of bacteria given in terms of µm.
Dimensions of viruses given in terms of nm (nanometers).
Viruses range in size from 18nm diameter (parvoviruses) to 300nm
diameter (poxviruses). In comparison, the Staphylococcus organism is
approximately 1 µm (1000nm) in diameter. The limit of resolution of the
standard light microscope is ⊆ 200nm. Thus, all but the largest viruses
are sub-microscopic and can only be seen by electron microscopy (ϵ .m).
Morphology:-
Viruses have one of 3 types of basic shape:
- Isometric (Cubic symmetry)
- Helical
- Complex – poxviruses

Capsomeres form Capsomeres form Icosahedrons

hollow tube in shape of helix.

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Isometric viruses may be enveloped or non-enveloped. Helical vertebrate viruses
are all enveloped, the helical capsid being folded back on its envelope.
- Classification:
a) Tissue/cells affected
- myxoviruses (‘myxa (or) – mucus)
- adenoviruses (‘aden’ or) – gland)

b) Type of Lesion:
- poxviruses (‘pox (Eng) – plural of ‘Pock’ – pustule)
- Iriodoviruses (‘iridescent’ – appearance of infected insect larvae)
c) Shape/Size/Composition:
- rhabdoviruses (‘rhabdos’ (G) – rod)
- Parvoviruses (‘parvus’ (L) – small)
- Picarnaviruses (‘pico’ (G) – micro-micro + RNA)
d) Signa:
- Reoviruses – respiratory, enteric,
orphan
- Papovaviruses – Pavilloma, Polyoma and vacuolating agent (SV40
virus) (Simian virus) 40.
- Arboviruses – arthropod – borne.
Fish Viruses:
Principal Pathogens:
- Herpeviridae – Channel Catfish virus
(CCV)
- Iridoviridae – Lymphocystis virus (LV)
- Birnaviridae – Infections pancreatic neuroses virus (IPNV)
Rhabdoviridae:- Viral haemorrhagic septicaemia virus (VHSV)
- Infectious haematopoetic necrosis virus (IHNV)
- Spring viraemia of Carpvirus (SVCV)
- Pike fry rhabdovirus (PFR)
- Eel rhabdoviruses (EVA and EVEX)
Herpesviridae:-- Herpesvirus Salmonis
-NeVTA (similar to H.

salmonis)
Oncorhynchus masou virus
(OMV)
Walleye herpes virus
Iridoviridae:-Cod (Atlantic) ulcus syndrome
virus

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Reoviridae:- Golden Shiner virus (GSV)
Birnaviridae:-Menhaden ‘spinning disease’
virus
-Eel virus, European (EVE)
Rhabdoviridae:-- Rio Grande perch (Cichlid)
Rhabdovirus
-Perch rhabdovirus
Unclassified:-Eel Papillomatosis virus.

Viruses can either be Living or Non-Living

A They are living particles because they have the ability to:
- Multiply within appropriate cell, a process resembling reproduction;
- To undergo mutation;

B They are non-living particles because they have no:

- Metabolic system;
- Intrinsic motility;
- Ability to respond to stimuli;

- Can not be cultured outside the host

i.e. they are obligate parasite, they live in the host cells intracellularly.
1.6 GENERAL CHARACTESTICS OF BACTERIA
Single bacteria cells are too tiny to be seen with the naked eyes. However, a
bacteria colony could comprise hundreds of cells and can therefore be
visible on solid media. Such a colony has some peculiar characters which
help as a first step in the identification of the bacterium, such characters
include the following:
i) The shape of the colony which could be circular, irregular or rhizoid.

Circular Irregular Rhizoid

ii) The elevation of the colony could be flat, raised, convex or umbonate.

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 iii) The edge of the colony could be entire, undulate, lobate, dentate or
rhizoid.

a) The rod-shaped one (straight or curved) termed the bacilli.

b) The spherical or ellipsoidal ones termed the cocci, some of which
occur singly and others stick together in pairs, tetrads, packets or
chains.
c) The spiral ones termed the spirilla.
Morphology of bacteria
Morphologically bacteria can resemble
Cocci (singular: Coccus)
Bacilli (rods) (singular: rod, bacillus)
Vibrios (singular: vibrio)
Spirilla (singular: sprillum
Spirochetes (singular: Spirochate)
Cocci:- These are round or oval bacteria measuring about 0.5 – 1.0µm in
diameter. When multiplying, cocci may form pairs, chains or
irregular groups.
Cocci in Pairs are called diplococcic, e.g. meningococci and gonococci.
Cocci in chains are called streptococci e.g. Streptococcis pyogenes.
Cocci in irregular groups are called staphylococci, e.g. Staphylococcens aureus.

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Gram reaction: Staphylococci and Streptococci are Gram positive, whereas
diplococcic can be Gram positive or Gram negative.
Rods (bacilli): These are stick-like bacteria with rounded, tapered (fusiform),
square, or swollen ends. They measure 1 – 10µm in length by 0.3-1.0 µm in width.
The short rods with rounded ends are often called coccobacilli. When multiplying,
bacterial rods do not usually remain attached to one another, but separate.
Occasionally, however, they may:
- form chains, e.g Streptobacillus species.
- form branching chains, e.g lactobacilli
- mass together, e.g Mycobacterium leprae
- remain attached at various angles resembling Chinese letters e.g
Corynebacterium diphtheria. Gram reaction: many are Gram negative.
Vibrios:- These are small slightly curved rods measuring 3-4µm in length by 0.5
µm in width. Most vibrios are motile with a single flagellum at one end.
They show rapid darting motility e.g Vibrio cholerae.
Gram Reaction:- Vibrios are Gram negative.
Spirilla:- These are small, regularly coiled, rigid organisms measuring about 3-4
µm in length. Each coil measure about 1.0 µm. Spirilla are motile with
groups of flagella at both ends. An example of spirillum is Spirillum minus.
Gram Reaction:- Spirilla are Gram negative
Spirochaetes:- These are flexible coiled, motile organisms. They progress by
rapid body movements. Most are not easily stained by the Gram method.
Spirochaetes are divided into three main groups:
Freponemes:- Are thin delicate spirochaetes with regular tight coils, measuring
from 6 – 15 µm by 0.2 µm in width. Examples include Treponema
pallidum and Treponema pertenue.
Borreliae:- Are large spirochaetes with irregular open coils, measuring 10 – 20
µm in length by about 0.5 µm in width. Examples include Barreled dutton;
and Barrelia vincent:
Leptospires:- Are thin spirochaetes with many tightly packed coils that are
difficult to distinguish. They measure 6 – 20 µm in length by 0.1 µm in
width and have hooked ends. The leptospire of medical importance is
Leptospira interrogans.
Rickettsiae:
Although classified as bacteria, rickettsiae resemble viruses in that they replicate
only in living cells and are unable to survive as free-living organisms. They can
just be seen with the light microscope (red particles in Giemsa preparations).
Unlike viruses, rickettsiae contain both RNA and DNA, multiply by binary fission
and have cell walls composed of peptidoglycan. They show sensitivity to
antiseptic and some antibiotics.
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Chlamydiae:
Chlamydiae are small (250 – 500nm) Gram negative bacteria but resemble viruses
in being unable to replicate outside of living cells. They contain both DNA and
RNA and have their own enzyme systems. The energy required for metabolic
activities is supplied by the host cell.
Specifically, Bacteria have the following characteristics:
- They are either Gram negative or Gram positive.
- They are micro-organisms without a true nuclear (Prokaryotes) or plastids
and lacking any sexual reproduction process.
- Bacteria reproduce asexually by binary fission.
- Bacteria are saprophytes, parasites or autotrophs.
- Bacteria can be cocci, rod-like, spiral
- Bacteria can also be single cells or in chains.
- Some bacteria are naked i.e. without cell wall.
- Bacteria are larger than virus ranging from 0.5-1.5µm in diameter.
- They have all the characteristics common to other organisms. They are
cellular, posses comparable metabolism and are microscopic especially with
the oil immersion (X100) objective.
1.7 CHARACTERISTICS OF ALGAE
- They are of various colours ranging from green, blue-green, red,
brown or golden.
- They are filamentous, thread like, photosynthetic plants ranging from
unicellular as in Chlamydomonas sp.
- They are the eukaryotic – with distinct nucleus.
- Some are flagellated while others are not.
- They are aquatic either freshwater or as in marine phytoplankton.
- All algae form the bases of aquatic food webs that support enormous
abundance and diversity of life.
- All algae posses’ chlorophyll, the primary pigment that traps wave
lengths of light.
- These pigments include other forms of chlorophyll (green)
carotenoids (yellow/orange), xantophylls (brown) and phycobilius
(red or blue varieties).
- The mixture of pigments produced by the chloroplasts give
characteristic colour to algae.
CHAPTER TWO
2.0 ISOLATION OF MICRO-ORGANISMS
When a micro-organism grows in the laboratory medium, it is referred to as
a culture. Different organisms growing on the same kind of medium may
appear quite different, thus knowledge of the appearance or the cultural
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 characteristics of a species is useful for the recognition of types of organism
and may also serve as an aid in the identification of species. However,
organisms must be obtained in a pure culture before the cultural
morphological characteristics of a species can be determined. A pure culture
consists of a population of cells all derived from a single parent cell. Micro-
organisms inhabit various parts of the body e.g. fin, gills, intestinal tract and
the skin.
2.1 ISOLATION OF FUNGI
Fungi are isolated from fish substrates by techniques that vary in
complexity. Generally, media are used to culture fungi meant for isolation
and/or identification. Reagents are used for preparing materials for
microscopic examination while stains are used to aid vision.

2.2 WATER AGAR (WA)

This is a simple culture medium. It is not very rich hence it only supports
limited mycelia growth. It is useful when limited mycelia growth is desired
to reduce contamination thereby enhancing isolation from plates. It is useful
for isolating fungi from infected fish tissues.
Agar 15 – 20g
Distilled water 1 litre
The quantity of agar depends on brand, purity and intended used, based on
past experience.
2.3 CZAPEK-DOX AGAR (CZA)
This synthetic medium is often compounded as follows:
NaNo3 3.0g
K2HPO2 1.0g
MgSO4.7H2D 0.5g
Kcl 0.5g
FeSO4.7H2O 0.01g
Sucrose 30.0g
Agar 15.0g
Distilled Water 1 litre
This medium is essentially used for the culture of species of Aspergillus and
penicillum.

2.4 POTATO DEXTROSE AGAR (PDA)

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 Though readymade powder formulation of PDA may be available, most
workers find freshly prepared medium from potato and dextrose more
satisfying.
Peeled, sliced potatoes 200g
Dextrose 10-20g
Agar 12-15g
Distilled water 1 litre
Sliced potato is added to water and allowed to simmer for 30 – 60 minutes
after which it is filtered through layers of cheese - cloth. Agar and other
ingredients are added to the filtrate before auto cleaving. To discourage
bacterial contamination, 100mg of streptomycin sulphate, prepared as stock
solution in sterile distilled water, may be added to 1 litre of autoclaved
medium under sterile hood at 45 oC before pouring into plates. With this
addition, the medium is known as PDAs. This medium is very useful for
growing a large variety of fungi.
2.5 LACTOPHENOL
This mounting solution is variously referred to as Amann’s mounting fluid
or Amann’s lactophenol solution. It is widely used in microscopic mounts
of semi-permanent slide preparations. It is prepared as follows

Phenol 20ml
Lactic acid 20ml
Glycerine 40ml
Distilled water 20ml
For hyaline (transparent or nearly transparent) fungi about 0.1% Catton blue
may be added to lacto phenol.
2.6 POTASSIUM HYDROXIDE
For best result, a freshly prepared solution of 2-10% KOH is used when
mounts are to be made from dried specimens. It should be noted that KOH
must never come in contact with the lens of the microscope. If it happens,
the lens must be cleaned promptly and thoroughly.
In most cases, the tissue is allowed to stay in KOH for a few minutes for re-
hydration to take place, then removed and observed under cover slip with
addition of mounting solution if desired. Alternately, a small piece of the
tissue may be put on a slide and a couple of KOH drops added. This is
raised to simmer over alcohol flame. It is then removed from the flame.
The tissue may then be transferred to a fresh slide and observed under a
cover slip with added mounting solution if desired.
2.7 DESCRIPTION OF SOME SPECIES OF FUNGI
2.7.1 Trichophyton rubrum

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 Clinic:- One of the most common fungi pathogenic for animal. Most
frequent microorganism of mycoses of the body.
Geographical Occurrence:- Worldwide
Reservoir:- Man, animals and rodents. Infectious cutaneous scales, fin and
etc.
Demonstration:- a) KOH preparation;
b) Culture: Inoculation of material with a sterile
mycological hook on to kimming agar.
Macro culture:-
Growth: Moderately quick, after 7 days radiating mycelium with
elevated centre; after 2 weeks typically recognizable
Growth form:- Velvety to fluffy surface, central hat-shaped elevation,
sometimes fine radial grooves, also resembling wheel spokes;
Colour:- Upper side white to light yellow, rarely somewhat reddish,
underside cockade-like yellow- red to dark red, to red violet,
also virtually colourless. Trichophyton rubum colours the agar
brown to dark brown.
Periphery:- Fine radiating extension, slight colour diffusion possible.
Microscopy:
Hyphae:- Ramified and septate, straight or curved;
Microconidia:- Roundish, pear-or Cone-shaped;
Macroconidia:- Longish, smooth-walled, roller-shaped, multiseptate,
blunted ends, frequently only found as isolated examples or not
at all.
2.7.2 Microsporum canis
Clinic: - Closely resembles infection of M. audouni scaling, round foci on
the head and skin.
Geographical Occurrence:- Worldwide, more frequently in North America
and Europe, also in Africa and Asia.
Reservoir:- cats, dogs, horses, monkeys, fish
Demonstration:-
a) Wood light: the foci
fluoresce yellow-green;
b) KOH preparation;
c) Culture: inoculation of
the material with a sterile
mycological hook onto
kimming agar.
Macro Culture:
Growth: Relatively quick-growing, sometimes visible after 48 hours.
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 Growth Form: Fluffy to woolly, sometimes chalky to granular; upper
side white to yellow to light brownish under side yellow-
orange-brown;
Periphery: Radial extensions with a yellow margin,
Microscopy:
Hyphae: Ramified and septate,
usually quite straight;
Micro conidia: Roundish, pear-shaped or
oval;
Macro conidia: Rough-walled, fusiform,
multicellular; if numerous, macroconidia are formed, the
surface of the culture appears sandy.
2.7.3 Candida parapsilosis
Clinic: Candida mycosis.
Geographical Occurrence: Worldwide
Reservoir: Man and Animal including fish.
Demonstration: (a) KOH preparation as for dermatophytes. Round and oval
cells as well as threads are perceptible;
(b) Culture: inoculate smear swab directly on to agar, or for
enhancement, place in beer wart medium. Specimens are
inoculated onto kimmig agar or, for differentiation, on to rice
agar. Growth takes place at room temperature, but can be
accelerated at 37oc.
Macro Culture:
Growth Form: flat, finely wrinkled colonies, irregular fringe;
Colour: cream to yellow-white, semi-glossy.
Microscopy: (Rice agar plate)
Blastospores: Oval to long oval shape 2.5 -4µx2.5 - 9 μ arranged like a
flower rosette;
Pseudomycelium: Present in addition to septate mycelium;
Chlamydospores: none
Arthrospores: none
Biochemical Fermentation Assimilation
Behaviour:
Glucose + +
Galacctose +/- +
Saccharoce - + (-7)
Maltose - +
Lactose - -
Peptone +
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 KNO3 -
2.7.4 Aspergillus species
Clinic: Aspergillus/Aspergilloma is possible as involvement of the
CNS. Aspergillus flavus can form toxins. It is the most
important producer of the highly toxics carcinogenic aflatoxin
B1.
Geographical
Occurrence: Ubiquitous saprophytes.
Reservoir: in man, animal and fish
Demonstration: (a) direct preparation;
(b) Culture: inoculate the material with a sterile mycological
hook onto kimming agar;
(c) Organic material: histological examination with PAS
staining.

Macro Culture:
Growth: recognizable within a few days;
Growth Form: velvety to flaky surface due to marked
sporulation;
Colour: A. flavus: White-
yellow

[Link]: grey-
green.
A. niger: black.
Microscopy:
Hypae: Septate;
Conidiophores:Borne laterally on the hyphae, non-septate; numerous
sterigmata proceed from the apical club-shaped swellings
(head-shaped) fructification organs);
Conidia: Borne in chains on the
sterigmata.
2.7.5 Mucor species
Clinic: Mucomycosis,
It is observed in consumptive diseases.
Geographical occurrence: Ubiquitous
Reservoir: Mainly vegetable
organic material, such as moist bread; fruit or vegetable
juices; stable dung; less frequently in animal and man.

17
 Demonstration: (a) direct preparation;
(b) Culture; inoculation of the material with a sterile
mycological hook onto kimming agar;
(c) Organ material: histological examination with PAS
staining.
Macro Culture:
Growth: quick, covering the agar surface in just a few days;
Growth Form: long-fibred, rough woolly network of hyphae;
Colour: Initially white, later grey with numerous black dots
(sporangia);
Periphery: the entire petric dish is filled after a few days.
Microscopy:
Hyphae: thick, non-septate;
Sporangiophores: departing laterally from
the mycelium, ramified, spherical at the end, sporangia filled
with spores;
Conidia: elliptical, contained in large number in the sporangia.

2.7.6 Isolation of Virus

Procedure:
(i) Prepare suspension of fish tissue in balanced salt solution
(ii) Filter to remove cell debris and bacteria
(iii) Inoculate filterate on growing monolayer of cells
(iv) Allow ½ to 1 hour for attachment of virus to cells
(v) Add suitable nutrient medium and incubate
2.7.7 Growth of Viruses
Viruses do not grow. The synthesizing machinery of the living, host cell is
modified to reproduce virus particles which are released from the cell to
infect further cells. Typical sequences of events are as follows:

18
 Attachment of virus to cell membrane receptors

Absorption of Virus into cell

Uncoating by cell enzymes (removal of protective capsid)

Transfer of genetic code to cell DNA

Synthesis of virus components by cell

Assembly of Virus components Release by: (1) “budding” from

cell
Surface or
(2) bursting from killed cell.
2.7.8 Identification of Viruses
Virus can only be seen under Electronic microscope. It can only be seen
on:-
(i) Observation of effects of virus on infected animals or cell culture and
(ii) Neutralization of these effects with specific antiserum.

2.7.9. Possible observable effects;

(1) In animals: initiation of disease with characteristic clinical signs or
pathology.
(2) In cell cultures: may have
(a) Cytopathic effect (CPE) in cell structure e.g. destruction of cells.
Alteration of cells – foamy appearance (vacuole formation)
- giant cells (fusion of cells)
- inclusion bodies (virus “factories”)
Virus may also be detected within the
host cell by special staining techniques
(e.g fluorescent antibody) or E.M.
examination.
[Link] FLUORESCENT ANTIBODY TEST:
Some dyes have the property of absorbing light of one wavelength and
emitting light of another, and one of these dyes is fluoresce isothiocyanate

19
 which has also the property of combining with certain proteins and which
can, accordingly, be used to combine with antibodies. If a dye-conjugated
antibody is applied to the homologus antigen it combines with the antigen
which itself becomes fluorescent.
This serological staining technique has application in the study and
identification of very minute parasites such as blood protozoa-Piroplasms
and plasmodia – when recognition of various stages in the life of these
parasites can be made under the microscope using ultra-violet light, and a
quantitative measure of antibody present can be made.
Fluorescent antibody technique (FAT) is used for the diagnosis of many
infectious diseases of bacterial, viral and other origins. The basic principle
of the test is based on detection of viral, bacterial or other antigens in tissues,
smears, cell culture etc, by antibodies which have been conjugated with a
fluorescent dye. The attachment of antigen (section or smear or colony of
organisms) with antibody is detected by the observation of fluorescence
isothiocyonate (FITC) etc, to which the antibody is attached. In negative
test, the antibody within the dye is washed out and no fluorescence is seen in
the smear or tissue section.
[Link] Isolation of Bacteria
The direct inoculation of solid media is the method of choice for the primary
isolation of fish bacteria. Because of the prevalence of mixed infections, it
is important that culture plates be inoculated in such a way that isolated
colonies of bacteria are obtained directly from the sample of fish material.
Either a selective or non-selective approach or a combination of both may be
adopted in the choice of media for isolation purposes.
The selective approach uses media designed to isolate or identify a particular
genus or species of organism and is of value where specimens are to be
screened for the presence of specific bacteria only. The non-selective
approach uses media which supports the growth of a wide range of bacteria
and may result in a considerable amount of time and material being spent on
identification procedures. It has the advantage of providing pathologist with
a comprehensive picture of the bacteriological status of the fish.
[Link] A Selective Media
[Link] Isolation of Pseudomonas spp
Commercially available media containing cetrimide (pseudosel Agar – BBL;
pseudomonas C-F-C Agar-Oxoid), which effectively inhibits the growth of
most Gram negative bacteria, may be used for the selective isolation of
pseudomonads. These media also contain MgCl 2 and K2SO4 which enhance
pigment production by these organisms.

20
[Link] Isolation of Aeromonas hydrophila
Rimler-shotts- agar which contains specific amino acids, maltose and
novobiocin, has been produced to facilitate the rapid identification of A.
hydrophila. This organisms forms yellow colonies (maltose fermentation)
without black centres (no H2S production) on the medium after 24h
incubation at 37oC is required to prevent possible growth of A. salmonocida
which will produce similarly pigmented colonies at lowers temperatures.
Yersima ruckeri and Citrobacter freundii will also produce yellow colonies
and an oxidize test is necessary to differentiate the isolates.
[Link] Isolation of Vibrio spp
Thiosulphate Citrate Bile self sucrose Agar (TCBS) (BBL; Difco; Oxoid)
promotes the rapid growth of vibrios bacteria (except V. ordalii) whilst
inhibiting that of other bacteria. The organisms produce green or yellow
colonies depending upon species and although slight growth of other
bacteria may also occur. The colonies are usually easily distinguished from
those formed by Vibrio sp.

21
[Link] B Non-Selective Media
(1) General Purpose Media
Majority of fish bacteria are non-fastidious in their nutrient requirements and
most general purpose nutrient or blood agar media may be used for their
isolation, although the tryptone-soya peptone agar (TSA) formulations
available from many commercial sources (Trypticase soy Agar – BBL;
Tryptic soy Agar- Difes; Tryptone soya Agar- Oxoid) is now the medium of
choice in most laboratories. This will support the growth of all aeromonada,
freshwater Vibrios, pseudomonads, flavobacteria, Enterobacteriacea,
pasteurellae, streptococci, micrococci and nocardiae likely to be encountered
and has the particular advantage of clearly demonstrating the formation of
the diffusible brown pigment characteristically produced by the majority of
strains of A. salmonicida isolated from salmonid fish.
Although TSA and some nutrient agar formulation contain 0.5% (W/V)
NaCl, certain marine vibrios, and Vibrio anguillarum in particular, require a
higher salt concentration for primary isolation. Media should thus be
supplemented with NaCl to a final concentration of 2% (W/V) when fish
from marine or other salt water environments are to be examined. Marine
Agar (Difco), which contains a supplement which duplicates the major
mineral components of seawater, may be used for this purpose. Once
isolated, the high salt requirement may be lost and subculturing can often be
carried out on non-supplemented TSA medium.
[Link] (2) Low Nutrient Media
Myxobacteria are widely distributed group of fish pathogens which are not
recoverable or grow poorly on TSA and similar media. These organisms
require comparatively low levels of both nutrients and agar and freshwater
strains are most frequently isolated on cytophaga agar although general
purpose media diluted to contain about one tenth of nutrients and 1% agar
are suitable alternatives. Few marine strains of myxobacteria tolerate
transfer to freshwater and vice-versa and isolation from marine fish should
be attempted using cytophaga agar containing 50-70% filtered seawater or
marine agar.

22
[Link] (3) Enriched Media
These are only required for the isolation of H. piscium, R. salmoninarum
and Mycobacteria sp., although some of the latter organisms are capable of
growth on simple substrates.
- Haemophilus piscium has requirement for adenosine triphosphate or co-
carboxylase (diphosphothiamine) which may be provided using Mueller
Hinton Agar (Difco; Oxoid) containing 2 μg/ml commercially available
co-corboxylase (sigma).
- R. Salmoninarum is an extremely fastidious organism with an absolute
requirement for L – Cysteine. A number of media containing 10-12%
serum or blood and 0.1% (W/V) L-Cysteine have been developed to
isolate the bacterium directly from tissue but the formulation referred to
as KDM2 or Evelyn’s medium is generally recommended. Beside its
fastidious nutritional requirements, R. Salmoninarum is also a slow
growing organism and although growth from heavily infected material
usually occurs within 4 – 10 days, dilute inoculate may require up to 6
weeks incubation before visible colonies appear.

3.0 CHAPTER THREE

3.1 MICROSCOPIC AND HISTOLOGICAL EXAMINATION
Microscopic examination involves making an impression smear or a wet
mount of material from an affected area, placing it on a clean slide, and using a
microscope, examining it for the presence of microbes. Microscopic examination is
a rapid and inexpensive diagnostic tool. Identification, however; will require
further testing – Figure 1.

23
Source: Noun (2015).

Fig. 1: Light microscope.

The light microscope is made up basically of a stand which is composed of
the arm and the base or foot. It is to this stand that the other parts of the
microscope are attached:
- The base, which carries the rest of the microscope, is very stable and
rests well on a flat surface such that the microscope does not shake while
in use.
- The arm, carrying all the other parts of the microscope is attached to the
base by means of a hinged joint.
- The body tube is attached to the upper end of the am on which it is
moved up and down with the coarse adjustment when focusing.
- The draw tube is the extension tube which can be pulled out and pushed
back into the upper part of the body tube. It carries the eyepiece and
gives room for the alteration of the distance between the eyepiece and the
objective.
- The fixed tube of the monocular microscope is replaced in a binocular
microscope by a binocular tube. The ocular sleeves of the binocular tube
can be moved horizontally by sliding in relation to each other for
adjustment to the distance between the eyes of the operator.
- The revolving nose piece is attached to the bottom of the body tube and it
is on to this that the removable objectives are fixed. Rotating the
nosepiece brings each objective into position with a click.
- The stage – is attached to the lower end of the arm. It has a central
opening through which light rays from the condenser reach the specimen.
It is supplied with clips for holding the slide down. This may be replaced
by the mechanical stage with which the slide can be hold and moved
smoothly in two directions at right angles. In some microscopes, it is the
stage and not the body tube that is moved up and down with the coarse
adjustment when focusing. There are three lens systems in the compound
microscope; the eyepiece, the objectives and the condenser.
- The condenser is a member of the substage assembly which is attached to
the arm below the stage. Other members of the substage are the iris
diaphragm and the filter holder. The sub-stage can be moved up and
down to adjust the level of illumination on the specimen. The condenser
collects light rays from the light source and focuses these onto the
specimen on the slide. The iris diaphragm controls the numerical
aperture of the condenser.

24
 - The eyepiece slots into the upper end of the draw tube in the monocular
microscope or into each of the ocular sleeves of the binocular
microscope. It may vary in power from about X4 to X25.
- The objectives are attached to the noise piece microscope are usually
equipped with three objectives – the low-power object with a focal length
of 16mm and magnification 10X, the high-power objective with a focal
length of 4mm and magnification 40X and the oil immersion objective
with a focal length of 1.8mm and magnification 100X. The light source
can be built into the microscope or it can be separated. Where it is built
on, the light rays pass straight into the condenser and where it is separate,
a mirror, which is attached to the base of the arm, is used to direct the
light rays into the condenser. The condenser throws the light onto the
specimen on the stage. While the light passes through the objective and
the body tube to the eyepiece.

3.2 Preparation of Temporary Slides

General Method of Making a Temporary Preparation
A temporary preparation of cells is usually observed as a wet mount. This is
prepared by placing a cell suspension in liquid on the slide or if the material
to be studied is dried, by placing it directly on the slide and adding water,
glycerin or stain to it. The material is then covered by a cover slip. You
have to be careful not trap air bubbles while lowering the cover slip over the
material as it will cause interference in observing the material.
The procedure is shown below in figure 2.

25
 Source: Noun (2015).

Fig. 2: The technique for lowering the

cover slip over the slide

3.3 OPERATING THE MICROSCOPE

The proper way to operate the microscope is to start focus with the lowest
power objective lens (OB) while you look through the side and move the
lens down as close to the slide without touching it. Next, you look through
the eyepiece lens and focus upward only until the image is sharp. If you
cannot get it in focus, repeat the process over. When you have a sharp
image at low power lens. You turn the revolving noise piece (RN) to click
the next higher power lens and do a minor adjustments with the fine focus
knob. Continue with subsequent objective lenses and fine focusing each
time.
(B)HISTOLOGICAL EXAMINATION
This is the examination of tissue that has been preserved in a fixative,
embedded in wax, sliced extremely thinly, placed on a clean slide, stained,
and examined using microscopy. Special stains applied to the tissue sections
can provide more specific information. Histological examination can prove
to be valuable tool in the diagnostic methods used in the field of fish health.
(B) Basic Tissue: Micro-techniques
Microscopic study of fish parts demands cutting of the part into thin sections
which may be paraffin-embedded sections or fresh sections.
(1) PARAFFIN SECTIONS
(a) Fixation
26
In anatomical studies, tissues are
examined either fresh or fixed. One of the most commonly used
fixatives for light microscopic work is referred to as FAA – Formal –
Acetic – Alcohol with the following constituents:
Formalin - 5cc
Glacial acetic acid - 5cc
70% Ethanol - 90cc
The fixation stabilizes the tissue components for subsequent chemical
and physical treatments.
(b) Dehydration and Infiltration
The fixed tissues are then dehydrated in series of ethanol solutions of
increasing concentration, and the water removed from the tissues, is
replaced with xylene and paraffin wax successively:
1. Rinse off excess fixative in a little 70% ethano;l
2. Dehydrate in ethanol series namely 70%, 90% and 100%
(absolute) ethanol for 10 – 20min, each;
3. The ethanol is then subsequently replaced with xylene by passing
the dehydrated tissues through a series of xylene/ethanol solutions,
namely 30% 50%, 70%, 90% and pure xylene for about 5min,
each;
4. Immerse the tissues in saturated solution of paraffin wax in xylene
and leave on a warm plate over night;
5. Replace the xylene/paraffin wax solution with pure molten wax
and maintain at about 56oC for a few hours. This is to get the
tissues infiltrated with paraffin wax.
(c) Embedment
The infiltrated tissues are then embedded in solid paraffin wax as
follows:
1. Lubricate the interior of a small petridish or an embryo cup, with
glycerine;
2. Pour some molten paraffin wax and transfer the block of tissue into
it in the desired orientation;
3. Hold the Petri-dish or embryo cup flat on the surface of a tray of
cold water. Blow gently until a thin film of solidifying wax is
observed;
4. Let the dish or cup go and sink into the cold water to effect rapid
cooling and solidification of the wax.
(d) Microtoming
This is cutting of the tissues into thin sections using a microtome, an
instrument equipped with knife and specimen holder facilities.
27
1. Cut a block or pyramid of solid paraffin wax around the specimen,
and affix to the microtome ensuring that the top surface is parallel
to the knife edge.
2. Make thin series of paraffin sections.
3. Smear the microscope slide with little Haupt’s adhesive.
(a) Gelatin - 1g
(b) Distilled Water - 100ml
(c) Glycerine - 15ml
(d) Phenol - 2g
4. Transfer sections in some drops of water on the microscope slides,
placed on a warm plate. Allow them to dry properly
(E) Staining;
This is an application of stains or dyes to the section to improve
contrast. Before commencing the staining, ensure that the paraffin
wax is removed as follows:
1. Remove paraffin wax by immersing slides in pure xylene (2-3min).
2. Remove xylene in absolute ethanol (1-2min)
3. Hydrate the sections in series of ethanol solutions of decreasing
concentrations, viz: 90%, 70%, 50%, 30%, 10% and water if the
stain is aqueous. Staining can be of single stain or double stains.
Staining with Safranin-Fast green (Double Stain Method, after Sass,
1958).
Stock Solutions:
1. 1% aqueous Safranin
2. 0.5% fast green in 95% ethanol
3. Hydrate to water 90%, 70%, 50%, 30%, 10%, water (5mins each).
Procedure:
1. Stain in 1% aqueous safranin 1-12hr (time depends on specimen).
2. Rinse in water until no safranin bleeds from specimen.
3. Dehydrate rapidly through 30%, 50%, 70%, to 95% ethanol (1min
each).
4. Counter stain 0.5% fast green for 5-30 seconds (dissolved in 95%
Absolute ethanol).
5. Dehydrate rapidly through 2-3 changes of absolute ethanol.
6. Transfer to carbol-xylene (1 volume of malted c.p phenol and 3
volumes of xylene) for 5 min.
7. Give 3 changes in xylene, 5 min. per change
8. Mount in synthetic mountant (Canada balsam or D.P.X). It is
advantageous to use DPX because it dries up easily. To hasten the

28
 process of drying up, slides may be kept in oven at lower
temperature for some times.
(iii) FREE-HAND SECTIONS
These are sections cut by means of sharp razor blades, held with hand.
In this method and indeed in paraffin method described earlier, there
may be a need to soften tissue to facilitate sectioning. For free-hand
sectioning, the following procedure is needed:
1. Immerse the block of tissue in water for a few minutes.
2. Hold the tissue upright, between the thumb and the index finger.
Moisten the transfer surface of the specimens as well as the edge of
the blade in water.
3. Make brisk, random and transverse cuts into the tissue. Transfer
sections into a vial of water and select thin, translucent sections.
4. Stain the sections in 1% aqueous safanin solution.
5. Mount in glycerine or glycerine jelly.
(a) Gelatine - 1g
(b) Distilled Water- 60ml
(c) Glycerine - 70ml
(d) Phenol Crystals - 0.1g
(iii) CHEMICAL MACERATION OF TISSUE
Chemical maceration of tissue is a technique for softening and
separation of cells in a tissue into individual component cells.
Procedures:
1. Cut a tiny block of tissue end, immerse in a few mills of Jane’s
solution (5% potassium chlorate in 50% nitric acid) in a test-tube
or boiling tube.
2. Heat gently for 4 – 5 minutes until the fumes have ceased.
3. Allow to cool and decant excess acid.
4. Carefully stain in 1% safranin for 5 – 10 min.
5. Transfer a little of the macerated tissue to a microscope slide.
6. Carefully stain in 1% safranin for 5-10min.
7. Rinse off excess stain with water.
8. Using a mounted needle gently, and carefully tease out the clump
of tissue into individual cells.
9. Apply a drop of glycerine and a clean cover slip.

29
 CHAPTER FOUR
4.0 BIOCHEMICAL TESTS FOR BACTERIAL
The ability of bacterial to change specific substrates and synthesize diverse
products is essential when characterizing bacterial species. Bacterial
cultures may appear similar in their morphological and cultural
characteristics, but may exhibit dramatic differences in their metabolic
reactions.
Qualitative tests, generally consisting of a nutrient medium plus a substrate
in which the organism is cultured, are designed to easily detect important
bio-chemical features. An “indicator” is often added to show the
accumulation of a metabolic product. Often changes in the physical nature
of the medium itself are adequate evidence of a substrate (e.g., liquefaction
of gelatin).
More definitive measurements of metabolic products may be desired,
establishing better criteria for a taxonomic group. In the routine test to
determine the ability of two cultures to ferment glucose, for example, both
are found to produce acid; the result for each is the same. Upon further
testing to identify the acidic constituents, very different results could be
obtained. One culture may produce only a single acid, the other a different
acid or even several different acids.
The biochemical activities of a microbiological culture have applications
beyond that of characterizing a bacterial species the role of micro-organisms
in the aquatic environment is attributable to the ability of microorganisms to
degrade various substrates and to synthesize new product.
4.1 Catalase Test (for non-mycobacterium)
This test is used to differentiate those bacteria that produce the enzyme
catalase, such as Staphylococcus aurens, Listeria moncytogenes,
Campylobactor hepaticus, Vibrio metschnikovii and Yersinia from non-
catalase producing bacteria such as Streptococci.
Principle
Catalase acts as a catalyst in the breakdown of hydrogen peroxide to oxygen
and water. An organism is tested for catalase production by bringing it into
cantact with hydrogen peroxide. Bubbles of oxygen are released if the
organism is a catalase producer. The culture should not be more than 24
hours old.
Required
Hydrogen peroxide, 3% H2O2 (10 volume solution).
Method
1. Pour 2 – 3 ml of the hydrogen peroxide solution into a test tube.

30
 2. Using a sterile wooden stick or a glass rod (not a nichrome wire loop),
remove several colonies of the test organism and immerse in the
hydrogen peroxide solution.
3. Look for immediate bubbling.
Results:
Active bubbling - Positive catalse test
No bubbles - Negative catalase test
4.2 Citrate utilization test
This test is used in the identification of Enterobacteria e.g. Edwardsiella
tarda. The test is based on the ability of an organism to use citrate as its
only source of carbon.
Ways of performing a citrate test
- Using a Rosco citrate identification tablet. This is the most economical
method when only a few tests are performed. The tablets have a long
shell-life and good stability in tropical climates.
- Simmon’s citrate agar can also be used but the dehydrated medium is
only available in 500g pack size from manufacturers.
Citrate Utilization Using a Rosco citrate tablets
Citrate identification tablets (Code 565-21) are available from Rosco
Diagnostica.
1. Prepare a dense bacterial suspension of the test organism in 0.25ml sterile
physiological saline in small tube.
2. Add a citrate tablet and stopper the tube.
3. Incubate at the appropriate temperature.
4. Observe after 48 hours and then further incubate for another 4-7 days for
any slow growing organisms.
Results:
Red Colour - Positive citrate
test
Yellow-orange colour- Negative citrate test
Ingredients for Simmon’s citrate medium are:
Sodium chloride - 5g
Megnesium sulphate - 0.2g
Monoammonium phosphate- 1g
Dipotassium phosphate - 1g
Sodium citrate - 2g
Agar - 20g
Distilled water - 1,000ml
PH - 7.0

31
 Combine all ingredients, mix thoroughly, adjust PH, and add 40ml of 1:500
bromothymol blue indicator solution. Dispense in 5-ml aliquots. Sterilize at
121oC for 15 minutes. Slant the tubes so as to obtain a 1 – inch butt and 1.5
– inch slant and allow to cool. Store at 4-6oC for a maximum of 6 months.
Note: Add 15g sodium chloride to yield 2% simmon’s citrate for the
inoculation of organisms from marine or brackish environment.
4.3 Coagulate Test
This test is used to identify S. aureus which produced the enzyme coagulate.
Principle:
Coagulate causes plasma to clot by converting fibrinogen to fibrin. Two
types of coagulate are produced by most strains of S. aureus:
- Free coagulate which converts fibrinogen to fibrin by activating a
coagulate – reacting factor-present in plasma. Free coagulate is detected
by clotting in the tube test.
- Bound coagulate (clumping factor) which converts fibrinogen directly to
fibrin without requiring a coagulate reacting factor. It can be detected by
the clumping of bacterial cells in the rapid slide test.
A tube test must always be performed when the result of a slide test is not
clear, or when the slide test is negative and Staphylococcus has been isolated
from a serious infection. A tube test is required to detect some MRSA
(Methicillin resistance S. aureus) strains. Before performing a coagulate
test, examine a Gram stained smear to confirm that the organism is a Gram
positive coccus.
Required
EDTA anti coagulated human plasma or rabbit plasma. The plasma should
be allowed to warm to room temperature before being used.
Slide test method (detects bound Coagulate)
(1) Mark a glass slide with a wax pencil to make two halves.
(2) Put a drop of normal saline on each half.
(3) Make emulsion of culture of bacteria taken from solid culture media, in
the saline drops so that it appears whey-like.
(4) Add a drop of rabbit plasma to the suspension in one of the mixture, and
mix it with a wire loop.
Results:
Clumping within
10 seconds - S. aureus
No clumping within
10 seconds - No bound
Coagulate
Note: Virulent Strains of Yersima pestis
32
 Controls:
Positive Coagulate Control: Staphylococcus aureus
Negative Coagulate Control: Escherichia coli or Staphylococcus epidemids.
Tube test method (detects free
coagulate).
1. Take three small test tubes and label
T = Test organism (18 – 24 hours
broth culture)
Pos = Positive Control (18 – 24 h S. aureus broth culture)
Neg = Negetive control (sterile broth)
2. Pipette 0.2ml of plasma into each tube
3. Add 0.8ml of the test broth culture to tube T.
Add 0.8ml of the S. aureus culture to the tube labeled ‘pos’.
Add 0.8ml of sterile broth to the tube labeled ‘Neg’.
4. After mixing gently, incubate the three tubes at 35 – 37 oC. Examine for
clotting after 1 hour.
If no clotting has occurred, examine after 3 hours.
If the test is still negative, leave the tube at room temperature overnight
and examine again.
Results:
- Clotting of the tube contents or fibrin clot tube - S. aureus
- No clotting or fibrin clot - Negative test
Commercially produced agglutination tests to identify S. aureus several
latex agglutination test kits have been developed to identify S. aureus based
on the detection of clumping factor, and or protein A.
4.4 DNase Test
This test is used to identify S. aureus which produces deoxyribonuclease
(DNase) enzymes.
The DNase test is particularly useful when plasma is not available to
perform a coagulate test or when the results of a coagulate test are difficult
to interpret.
Principle
Deoxyribonuclease hydrolyzes deoxyribonucleic acid (DNA). The test
organism is cultured on a medium which consists DNA. After overnight
incubation, the colonies are tested for DNase production by flooding the
plate with a weak hydrochloric acid solution. The acid precipitates
unhydrolyzed DNA. DNase – Producing colonies are therefore surrounded
by clear areas due to DNA hydrolysis.
Required
- DNase agar plate No. 33
33
 Up to size organism may be tested on the same plate
- Hydrochloric acid Reagent No. 43
1mol/1 (1N)
Method
(1) Divide a DNase plate into the required number of strips by marking the
underside of the plate.
(2) Using a sterile loop or swab, spot-inoculate the test and control
organisms. Make sure each test area is labeled clearly.
(3) Incubate the plate at 35 – 37oC overnight.
(4) Cover the surface of the plate with 1 mol/1 hydrochloric acid solution.
Tip off the excess acid.
(5) Look for clearing around the colonies within 5 minutes of adding the
acid.
Results
Clearing around the colonies - DNase
Positive strain.
No clearing around the colonies-DNase
Negative strain.
Control
Positive DNase control: Staphylococcus
aureus.
Negative DNase control: Staphylococcus
epidemidis.

4.5 Indole test

Testing for indole production is important in the identification of
Enterobacteria. Most strains of E. coli, [Link], M. morganii and
providential species breakdown the amino acid tryptophan with the release
of indole.
Principle
The test organism is cultured in a medium which contain tryptophan. Indole
production is detected by kovac’s or Ehrlich’s reagent which contain 4 (p)-
dimethylamino-benzaldehyde. This reacts with the indole to produce a red
coloured compound. Kovac’s reagent is recommended in preference to
Ehrlich’s reagent for the detection of indole from enterobacteria.
Ways of performing an indole test
- As a single test using tryptone water and kovac’s reagent.
- As a combined beta-glucuronidase-indole test using a Rosco
PGUA/indole identification tablet. This is useful in identify Salmonellae
and Shigellae.
34
4.5.1 Detecting indole using tryptone water
1. Inoculate the test organism in a bijou bottle containing 3ml of sterile
tryptone water.
2. Incubate at 35 – 37oC for up to 48h.
3. Test for indole by adding 0.5ml of kovac’s reagent. Shake gently.
Examine for a red colour in the surface layer within 10minutes.
Results
Red surface layer - Positive indole
test.
No red surface layer - Negative indole test.
4.5.2 Detecting indole using Rosco PGUA/indole tablet.
PGUA/indole tablets (code 590-21) are available from Rosco Diagnostica.
The tablets have a long shelf life (3-4yrs).
1. Prepare dense suspension of the test organism in 0.25ml physiological
saline in a small tube.
2. Add a PGUA/indole tablet and close the tube incubate at 35 – 37 oC for 3
– 4 hours overnight.
3. First read the beta-glucuronidase (PGUA) reaction.
Results:
Yellow colour - Positive PGUA
test.
Colourless - Negative PGUA
test.
4. Add 3 drops of kovac’s reagent (Rosco code 970-31) or other kovac’s
reagent) and shake.
5. Wait 3 minutes before reading the indole reaction. Examine the colour of
the surface layer.
Results:
Red Surface layer – Positive indole
test.
Yellow Surface layer- Negative indole
test.
Note: About 94% of E. coli strains are PGUA Positive and 99% are
indole positive.

35
4.5.3 Detecting indole using Rosco LD/indole tablet.
LD/indole tablets (code 584-21) are available from Rosco Diagnostica. The
tablets have a long shelf life (3-4 years).
1. Prepare a dense suspension of the test organism in 0.25ml physiological
saline in a small tube.
2. Add 3 drops of paraffin oil” on LD/indole talblet oil” and close the tube.
*The oil over layer provides the anaerobic conditions required for the
LDC reaction.
3. Incubate at 35 – 37oC for 3-4 hours overnight.
4. First read the lysine decarboxylase (LDC) reaction.
Results:
Blue/violet colour - Positive LDC
test.
Yellow, green or grey colour –
Negative LDC test.
*If examining after overnight incubation, a positive test is indicated by a
strong blue or violet colour.
5. Add 3 drops of kovac’s reagent (Rosco code: 920 – 31 or other kovac’s
reagent) and shake.
6. Wait 3 minutes before reading the indole reaction. Examine the colour of
the surface layer.
Results:
Red surface layer - Positive indole
test.
Yellow surface layer- Negative indole test.
4.6 Oxidese (Cytochrome oxidase test)
The oxidase test is used to assist on the identification of Pseudomonas,
Neisseria, Vibrio, Brucella, and Pasteurella species, all of which produce
the enzyme cytochrome oxidase.
Principle:
A piece of filter paper is soaked with a few drop of oxidase reagent. A
colony of the test organism is then smeared on the filter paper.
Alternatively, an oxidase reagent strip can be used. When the organism is
oxidase producing, the phenylenediamine in the reagent will be oxidized to a
deep purple colour.
Occasionally the test is performed by flooding the culture plate with oxidase
reagent but the technique is not recommended for routine use because the
reagent rapidly kills bacteria. It can however, be useful when attempting to
isolate N. gonorrhea colonies from mixed cultures in the absence of a

36
 selective medium. The oxidase positive colonies must be removed and
subcultured within 30 seconds of flooding the plate.
Important: Acidity inhibits oxidase enzyme activity; therefore the oxidase
test must not be performed on colonies that produce fermentation on
carbohydrate containing media such as TCBS or MacConkey agar. Sub
inoculation on nutrient agar is required before the oxidase test can be
performed. Colonies tested from a medium that contains nitrate may give
unreliable oxidase results.
Required
Oxidase reagent freshly (Reagent No. 64) prepared or use an oxidase
reagent strip.
Note: Fresh oxidase reagent is easily oxidized. When oxidized, it appears
blue and must not be used.
4.6.1 Method (fresh reagent).
1. Place a piece of filter paper in a clean Petridish and add 2 or 3 drops
of freshly prepared oxidase reagent.
2. using a piece of stick or glass rod, remove a colony of the test
organism and smear it on the filter paper.
3. Look for the development of a blue-purple colour within a few
seconds.
Results:
Blue-Purple clour - Positive oxidase
test.

No blue-purple colour- Negative

oxidase test.
Note: Ignore any blue-purple colour that develops after 10 seconds.
4.6.2 Method using an oxidase reagent strips.
1. Moisten the strip with a drop of sterile water.
2. Using a piece of stick or glass rod, remove a colony of the test
organism and rub it on the strip.
3. Look for a red-purple colour within 20 seconds.
Red-purple colour - Positive
oxidase test.
Controls:
Positive oxidase control: Pseudomonas
aeruginosa
Negative oxidase control: Escherichia
coli
4.7 Urease Test
37
 Testing for urease enzyme activity is important in differentiating
enterobacteria. Proteus strains are strong urease productions. Y.
enterocolitica also shows urease activity. Salmonellae and shigellae do not
produce urease.
Principle
The test organism is cultured in a medium which contains urea and the
indicator phenol red. When the strain is urease producing, the enzyme will
break down the urea (by hydrolysis) to give ammonia and carbondioxide.
With the release of ammonia, the medium become alkaline as shown by a
change in colour of the indicator to pink-red.
Ways of performing a urease test
i) Using a Rosco urease identification tablet.
ii) Using modified christensen’s urea broth.
4.7.1 Urease test using a Rosco urease tablet.
Urease identification tablets are available from Rosco Diagnostica (code 575
– 21). The tablets have a long shelf – life (3-4 years).
1. Prepare a dense ‘milky’ suspension of the test organism in 0.25ml
physiological saline in a small tube.
2. Add a urease tablet, close the tube and incubate at 35 – 37 oC for up to 4
hours or overnight.
Proteins and M. morganii organism give a positive reaction within 4
hours.
Results:
Red/purple colour - Positive urease test.
Yellow/orange - Negative urease test.
4.7.2 Urease test: Using Christansen’s (Modified) Urea broth.
1. Inoculate heavily the test organism in a bijou bottle containing 3ml
sterile christensen’s modified urea broth.
2. Incubate at 35 – 37oC for 3-12 hours in a water bath for a quicker
result.
3. Look for a pink colour in the medium.
CHAPTER FIVE
5.0 Sample Collection to Detect Bacterial Infection
If bacteria are suspected as the cause of a disease outbreak, a number of
similarly affected fish should be examined. Although the commonly
recommended number of fish to examine is 5 – 10 fish, there are many
factors that may determine the size of the sample, these include the disease
history of the fish stock, the signs of infection, the number of fish available
for sampling, the value of the fish, and the workload the diagnostic
laboratory is able to handle.
38
Fish that are alive, but with signs of the suspected disease (moribund fish),
are the most useful specimens to sample. If the disease is being caused by
bacteria, these fish tend to harbor large numbers of this disease – causing
bacteria. Samples should be taken from all damaged or affected tissues.
Fresh mortalities may also be examined. It is important to remember that
decay and the invasion of bacteria associated with rotting begin immediately
after the death of the fish. The rate at which this occurs depends on the
environmental conditions, especially the temperature of the water. Fresh
mortalities should be removed immediately from the water and kept
refrigerated or stored on ice prior to examination. Detecting pathogen in fish
stocks that appear healthy requires planning. A random sample
representative of the whole population can be obtained during transport,
treatment or vaccination, stocking and grading, or at slaughter. Subclinical
infections become more apparent when fish are stressed.
Fish stocks that may be more susceptible to disease at specific times may
require multiple samples to increase the chances of the infectious agent
being detected (i.e., prior to spawning for a vertically transmitted disease
such as bacterial kidney disease; following transfer to seawater for
anadromous fish for a disease such as furunculosis, or when water quality
deteriorates causing external infections such as columnaria disease). If an
infectious agent is present, disease is more likely to occur under these
circumstances, and therefore it becomes easier to detect. Stress – testing
experiments may also be used as necessary to increase the chances of
detection (e.g., furunculosis).
Knowledge of the basic biology of both the fish and the infectious agent
suspected is very important when identifying an optimal sampling time for
prevalence estimation. Environmental and species differences can markedly
affect the growth rate of salmonids. In addition, some pathogens are more
readily detected in fish of a certain size, where others are more readily
detected in fish of a certain age.
Two components are necessary to implement an effective detection program.
The number of fish being screened must be representative of the entire
population, and information on the fish being screened must be collected
(i.e., fish species, age, or growth).
More fish are required for examination to detect a rare infection than are
needed to detect a more common infection.
Choosing a sample size for defection purposes depends on the expected
prevalence of the infection within the population, whether the infection is
likely to be rare or common, and the size of the population sampled. The

39
assumption should be made that the infection is rarer than expected and that
the population to be sampled is larger than anticipated.
Assuming that the detection method being used is 100% efficient all the time
and that an infected fish will always be detected, then unless the entire
population is examined, the ability to detect an infection also depends on an
element of chance because of the sampling process. One usually accepts a
5% risk (called alpha) that the sample is not representative of the population
(e.g., 5 samples in 100, alpha = 0.05, would still not detect an infection when
it is, in fact, present in the population).
To detect an infection in a single sample, it is important that a minimum
sample size is determined before sampling starts. It is not worth collecting a
sample that is so small that it will miss an infection more than 5% of the
time. Furthermore, infections may spread very fast, especially in the warmer
season or in tropical regions, and this makes it impossible to compare
samples collected on different occasions. As a rule, pooling of samples
should be avoided. Therefore, a sample must be collected that is big enough
for the purpose defined before hand to escape the temptation of grouping
several little samples that relate to different populations.
Samples sizes for detection purposes are determined from three quantities;
the apparent prevalence expected in the population; the accepted level of risk
(alpha) attached to sampling: and the sensitivity of the diagnostic method
used for detection (see Table 1).
Table 1:
Sample size required (95% confidence level) to detect at least one carrier
fish in populations with an assumed minimum carrier prevalence of 2, 5 and
10%.
Assumed Carrier Prevalence
Population
Size 2% 5% 10%
50 46 29 20
100 76 43 23
250 110 49 25
500 127 54 26
1,000 136 55 27
2,500 142 56 27
5,000 145 57 27
10,000 146 57 27
100,000 147 57 27
Over 100,000 150 60 30

40
 Note: For intermediate population size, use sample size for next larger
population listed.
Source: Nicolas, F (1974).

5.1 PACKAGING LIVE FISH FOR TRANSPORT

1. Select live fish exhibiting the clinical signs most common to the
population of diseases fish.
2. Fill a strong plastic bag (preferably double bagged) about one-third
full with freshly aerated water to accommodate the live fish.
3. Place the fish in the bag. Smaller fish can survive up to 48 hours in a
plastic bag with cool water, adequate oxygen, and the appropriate
biomass per liter of water. The rule of thumb is 10 – 12 fry or five
fingerlings per liter of water. To maintain cold-water fishes,
particularly in warm weather and for long transit tunes, place crushed
ice plastic bags, seal them, and place then directly in the water with
the fish. Ensure that the crushed ice is not made from chlorinated
water.
4. Change the water and bag with pure oxygen if available, or use
compressed air.
5. Tightly seal the bag to ensure that there are no leaks. Once sealed, the
double bag should provide a good air cushion above the water-line.
6. Label the bag clearly by attaching a water proof label to the bag. Do
not write directly on the bag.
7. Line a strong water tight container made of plastics, thick Styrofoam,
or insulated metal with a plastic bag.
8. Layer the bottom and sides of the container with crushed ice or glycol
ice packs.
9. Carefully position the plastic bag containing the fish in the prepared
container and place the lid on top of the container.
10. Seal the container with strong packing tape, ensuring there are no
leaks.
11. Place the disease history sheet containing all information pertinent to
the sample in an addressed envelope and attach the envelope to the
outside of the container.
12. Label the container “Rush! perishable live fish”.
13. Check airline flight times and courier times to ensure the submission
will arrive at the diagnostic laboratory within 12 – 24 hours.

41
 14. Call or fax ahead to ensure the receiving laboratory knows when your
shipment will arrive.
15. Include the waybill number and mode of transportation. It is also
helpful to provide the company name of the airline or courier service.
5.2 PACKAGING DEAD FISH FOR TRANSPORT
The proper sampling of a population of sick fish is an important part of any
disease investigation. The selection and packaging of specimens play an
important part in the evaluation of a disease problem. Improper packaging
can result in costly delays in identifying the cause of the problem within the
population. Avoid repeat sampling by ensuring the specimens arrive at the
laboratory in a firm, fresh condition.
1. If shipment of live fish is not possible, or if the fish are larger than
25.4cm (10 inches), send recently euthanized fish.
2. If the fish weigh more than 500g (>1 1b), they should be packaged in
separate plastic bags. If the fish weigh less than 500g (<11b), then a
maximum of three fish can be packaged per bag.
3. Seal the bags tightly.
4. Label the bag clearly by attaching a water-proof label to the outside of
the bag. Do not write directly on the bag.
5. Line a strong, water tight container made of plastic, thick Styrofoam, or
insulated metal with a plastic bag.
6. Layer the bottom and sides of the container with crushed ice or glycol ice
packs. Crushed ice is superior to cubed ice or glycol ice pack, because
the cooling is more evenly distributed ensuring that each fish is properly
cooled.
7. Carefully position the plastic bags containing the fish in the prepared
container.
8. Place a layer of crushed ice or glycol packs on top of the samples, and
put on the lid of the container.
9. Seal the container with strong packing tape, ensuring there are no leaks.
[Link] the disease history sheet containing all information pertinent to the
sample in an addressed envelope and attach the envelope to the outside of
the container.
[Link] the container “Rush! Perishable specimens”.
[Link] airline flight times and courier times to ensure the submission will
arrive at the diagnostic laboratory within 12 – 24 hours.
[Link] or fax ahead to ensure the receiving laboratory knows when your
shipment will arrive.
[Link] the waybill numbers and mode of transportation. It is also
helpful to provide the company name of the airline or courier service.
42
 CHAPTER SIX
6.0 FISH DISEASE DIAGNOSIS FORM
Date: ……………….. Case No:………..
Name:…………………………………….
Address: …………….. GSM No:……….
…………………………………………...
HISTORY
FreshWater :…Marine:…System Size:…
gal (l)ac(ha).
Species affected…………………………
Species in System………………………
No of fish in system:……No of fish
affected:…….. % of fish affected:……….
Average fish size:……… in (cm)/oz (g)
Age(s) of affected fish:…………………..
When mortality started:……….. When
mortality ended:………. .
When morbidity started:……… When
morbidity ended:…………
How long has the system been set up? ….
… Temperature:…………………………
Types of life support present:………….
Any new introduction? Y:…..N:……. If
yes, When and What?...............................
…………………………………………...
…………………………………………...
Water source:……….Pipe: metal:………
Plastic:……….. water appearance
(cloudy, colored?): ……………………..
History of routine maintenance, including
Water changes and water quality
checks:…………………………………...
…………………………………………...
Behavioural changes? Y….. N…………
Describe:…………………………………
…………………………………………...
Respiratory rates: (normal …… fast…..
Slower……..)
Appearance of fish:……………………..
43
Appetite (normal … less …… more…):..
Other clinical signs:……………………..
…………………………………………...
CLINICAL WORK-UP
Water quality:
Do:……..mg/L Temperature:……PH….
Ammonia………mg/L TAN:…….. mg/L
UIA:……..mg/L, Nitrite:……mg/L, Salinity:………ppt.
Water sample preserved for further analysis:…………………………………
Physical examination
Behavior:………..Respiration (depth and rate):……….
……………………………
Skin:……………………………………..
Gills:……………………………………..
Biopsies and Cultures
Skin Biopsy:…………………………….
Gill Biopsy:………………………………
Blood smears taken? Y..N….Results:….
Bacterial Cultures taken? Y…N…………
Viral cultures taken? Y…….N………….
Fungal cultures taken? Y…..N………….
Organs cultured; kidney…… other……..
Results of cultures:………………………
Necropsy
Peritoneal cavity/visceral fat:……………
Gonads:………………………………….
Liver/gall bladder:………………………..
Stomach/intestines:………………………
Spleen:…………………………………...
Swim bladder:……………………………
Kidney:…………………………………..
Heart:…………………………………….
Brain:…………………………………….
Other:…………………………………….
Tissues preserved for histology or other further analysis:
…………………………
…………………………………………...
Problems Identified Treatments
1. ………………… 1. ……………..
2. ………………… 2. ……………..
44
 3. ………………… 3. ……………..
Results of treatment:…………………….
…………………………………………...
6.1 Aseptic Examination of Fish
Aseptic techniques are important and must be mastered since accurate and
dependable diagnosis of disease is directly related to the quality of the
necropsy and collection of specimens.
6.1.1 Equipment Required
- Dissecting board;
- 70% ethyl alcohol;
- Dissecting instruments;
- Syringes or Vacutainer tubes with
no additive;
- Needle or Vacutainer needles;
- Artificial culture media;
- Gram stain kit;
- Lens cleaner;
- Transport swabs;
- Paper towels;
- Latex gloves;
- Small beaker;
- Inoculating needle and loop;
- Light microscope with oil immersion;
- Blotting paper;
- Glass slides;
- Crushed ice or glycol packs;
- Flame source;
- Anesthetic;
- Disinfecting soap;
- Water proof marker;
- Hematocrit tube;
- Sterile swabs;
- Immersion oil;
- Lens paper;
- Shipping container.
6.2 SPECIMEN PREPARATION
If the specimen is alive, it must be euthanized prior to sampling. Euthanasia
means a gentle death and should be regarded as an act of human killing with
the minimum of pain, fear, and distress. This involves arranging facilities in
advance as well as ensuring a minimum of disturbance to the animals,
45
 considering safety of personnel, and using a method in which you have
confidence, i.e., one that is going to work quickly and reliably. Acceptable
methods of euthanasia include physical and chemical methods.
6.2.1 Physical Method (e.g., concussion): This involves a blow to the back of the
head and, if carried out by experienced personnel, is a humane method of
euthanasia.
6.2.2 Chemical Method e.g., anesthetic): Agents can be administered by
dissolving the chemical in the tank water. Drugs may also be administered
by intramuscular or intraperitorieal infection. For euthanasia, anesthetic
drugs are generally used at double or treble the recommended anesthetic
dose. A concentration of 150mg/l of tricaine methane sulfonate (TMS)
powder or MS222) will humanely euthanize most species of fish.
Benzocaine is a cheaper and equally effective alternative when used at a rate
of 0.2g/8 litres of water. Benzocaine is poorly soluble in aqueous solutions.
It should be dissolved in a small quantity of acetone before being added to
water. The use of anesthetic agents is contra-indicated if a parasitological
examination is also required as the anesthetic will also kill any live parasites
on the specimen and render them much more difficult to observe.
Prepare the examination area by cleaning and disinfecting the area with a
disinfecting agent. Be sure that there is plenty of work space. Arrange
materials and equipment to suit the surrounding and allow for easy access.
Wash your hands thoroughly with a disinfectant soap. Put on a pair of
lactase gloves. Following examination of the external body surface and
gills, the body cavity is opened aseptically to expose the internal organs.
The ventral body surface should be swabbed with disinfectant and an
incision made with a sterile scalpel through the body wall in the mid-ventral
line opposite the base of the pectoral fins. Using blunt-ended scissors or a
scalpel blade with the sharpened edge directed away from the fish, the
incision is extended antennary to the symphysis of the mandible and
posteriorly until just short of the vent taking care not to puncture the
intestine. Any adhesion between the body wall and underlying viscera are
carefully separated and the two sides of the body wall pinned back to the
dissecting surface. Alternatively, after opening the body cavity one side of
the body wall may be removed by making a roughly semi-circular incision
along the boundary of the cavity from the vent to the base of the pectoral fin
(fig. 3).

46
Fig 3: Diagram of internal anatomy of fish.

6.3 BLOOD COLLECTION

Three procedures are commonly used to obtain blood in larger fish: caudal
venous puncture, heart puncture, and dorsal aorta puncture.
6.3.1 Caudal Venous Puncture
1. Insert a needle under the skin of the ventral midline of the caudal
peduncle of an anesthetized or freshly euthanized fish.
2. Ease the needle towards the vertebral column until you reach the base
of the column.
3. Withdraw the needle a fraction of a millimeter and obtain the blood
sample.
6.3.2 Heart Puncture
1. Hold the needle perpendicular to the skin and insert the needle slightly
below the tip of the V-shaped notch formed by the gill cover and the
isthmus of an anesthetized or freshly euthanized fish.
2. Collect the blood as the needle enters the bulbous arteriosus.
6.3.3 Dorsal Aorta Puncture
(1) Insert the needle, along the dorsal midline of the mouth, just past the
juncture of the second gill arch of an anesthetized or freshly
euthanized fish.
(2) Collect a blood sample

CHAPTER SEVEN
7.0 FISH DISEASES
Unlike the situation in higher animals, most of the organisms associated with
disease in fish are naturally occurring and widely distributed pathogens
which utilize the organic and mineral matter in the aquatic environment for
their growth and multiplication.
The occurrence and magnitude of pathogenic infections in fishes are closely
related to the sanitary conditions prevalent in the water as well as the general
health of the fishes themselves. Hence, incidence of diseases can be avoided
or minimized by good pond management. Actually, most of the diseases

47
 depends mainly on the two factors, one for which the aquatic environment is
important and the other stems from cold-bloodedness of the fish.
The transition from commensal to pathogens is invariably triggered by the
imposition of one or more “stress” factors which increase the susceptibility
of the fish to pathogenic infections. Poor water quality and temperature
changes are probably the most commonly encountered factors which
predispose to clinical disease but overcrowding, trauma, transportation,
nutritional deficiencies, parasitism and primary viral infections may all
adversely affect resistance. It is important that this should be borne in mind
when assessing the significance of laboratory isolates and prescribing
treatments, since chemical or antibiotics therapy or prophylactic vaccination
measures are seldom entirely satisfactory unless proper attention is given to
correcting the underlying causes.
7.1 Host – Parasites - Environment Inter-relationship. The relationship
between a pathogen and the host causes serious loss only when fish (i.e., the
host) and pathogen are present in an environment which favours the disease.
Fish disease do not occur as a single cause but are the end result of
interaction of the etiological (disease causing) agent, a susceptible fish and a
predisposing environmental condition. These three factors must occur
concurrently in order for an infectious disease outbreak to occur.
Experience fish farmers take precautions to prevent the concurrent
occurrence of all three factors and this is regarded as “preventive medicine”
in fisheries and aquaculture (Fig. 4).

Fig. 4: Relationship between a pathogen, host and Environment

Source: Ezeri (2014).
Serious losses only occur when factors (1+2) are present in an environment
(3) which favours the disease (i.e., 1+2+3). When there is no favourable
environment (i.e., 1+2) no disease outbreak will occur. When there is no
48
 pathogen i.e. (1+3) and when there is no fish (i.e. 1) in the ponds (i.e. 2+3)
no disease outbreak will occur.

ADVERSE ENVIRONMENT BETTER ENVIRONMENT

FAVOURS PATHOGEN FAVOURS FISH PREVENT
PROLIFERATION AND QUICK PROLIFERATION
CAUSE STRESS TO FISH PATHOGEN

THOGEN ENVIRONMENT INTERACTIONS

HOST PATHOGEN
RESULTING
ENVIRONMENT
DISEASEINTERACTION
OUTBREAK RESULTING IN “NO DISEASE” CONDITION

H – SUSCEPTIBLE HOST
AE – ADVERSE ENVIRONMENT
P – VIRULENT PATHOGEN

7.3 TYPES AND CAUSES OF FISH DISEASES

For the fact that disease organisms are present in water, ever ready to
become problem causing pathogens especially when fishes are weakened by
stress. Variations in normal water parameters would make the aquatic
environment conducive for fish to contact disease. Holding stress within the
tolerance limit of a particular fish species that is being cultured is part of
good management.
Fish diseases can be broken down into two broad categories, infectious and
non infectious.
7.3.1 Infectious: - (diseases caused by a living organism).

49
 - Fungi – Fungi are a specific group of plants that lack chlorophyll and
are mainly secondary invaders of fish. Usually they can grow only on
dead organic matter. Therefore, fungal infections indicate there is
something else wrong with the fish.
- Viruses – Submicroscopic particles that live within the cells of living
organisms are called viruses. Sophisticated laboratory techniques are
required for diagnosis. Their location makes them almost impossible
to treat with chemicals or drugs.
- Bacteria – There are many different kinds that can cause serious
losses of catfish. Most are internal, although a few occur on the skin
and gills. Specialized laboratory techniques are necessary for their
isolation and identification. Most are about 3/25,000 inches long
(3microns), though some may be 10-12microns long.
- Parasite – An organism that lives in or on another at the expense of
its host is a parasite. There are many different kinds of parasite, both
internal and external ranging from the very small (8/25,000 inch = 8
microns) to some that are 5 – 6 inches long. Most problems are
caused by protozoans (single-cell animals) that live on the gills.
7.3.2 Non Infectious: - (diseases caused by other than living organisms).
- Nutritional – Causes by too much or too little food or nutrients
- Environmental – Oxygen depletions, gas bubble disease, toxic algae,
brown blood diseases ammonia etc.
The environmental changes so rapidly or to such a degree that the fish
are not able to adjust to the changes.
- Physiological – A change in blood PH of 0.2PH level due to over
exertion, feeding in relation to time of harvest, or a malfunction of the
organ system.
- Chemical Toxicants – Pesticides, insecticide etc.
7.4 Symptoms and Clinical Signs of Disease
Diseased fish usually exhibit noticeable signs which are either physical or
behavioral in nature. Usually the first indication that fish may be sick is a
reduction in feeding activity. It is, therefore, very important that the person
feeding the fish be an experienced fish culturist to detect any change in
feeding behaviour. Any unusual behaviour or abnormal physical appearance
should be a “red flag” that something is wrong and should be checked
immediately. Failure to do so could result in the loss of some or all of the
fish in the pond.
7.4.1 The following behavioural signs indicate that certain diseases may be
present in fish:
(1) Swim sluggishly in an uncoordinated zigzag manner.
50
 (2) Scratching, flashing or rubbing against objects in the pond.
(3) Gasp at the surface of the water for air (anoxia).
(4) Crowding or gathering in vegetation, shallow water or at water inflow,
hiding under objects to avoid light as a result of low oxygen, toxins
and many others.
(5) Loss of appetite, so fish refuse to eat (anorexia).
(6) Failure to flee when exposed to fright stimuli. Low oxygen
consumption, metabolic factors and many diseases could be
responsible for this behaviour.
(7) Accumulation of liquid in body cavity.
(8) Topping or piping at water surface; floating head-up, moribund
(dying) could result from low oxygen; toxins, external parasites and
bacterial infection could cause such behaviour.
7.4.2 Some Physical signs that suggest the presence of disease in pond.
(1) Leisons or sore in the body;
(2) Loss of scales and body
discolouration;
(3) Gills turns pale colour;
(4) Ragged or torn tins;
(5) Loss of weight;
(6) Pale gills;
(7) Bleached skin colour due to
vitamin E deficiency and low
oxygen intake;
(8) Other diseases could result in
clamped fins, heavy breathing,
inactivity, scratching against tanks, pond walls, scales printing
outward like a pine cone, blooded belly, loss of colour, cotton patches
on the body, staying in vertical position and mutilated barbells;
(9) Popeye (exophthalmia).

CHAPTER EIGHT
8.0 Fungal Diseases
Fungus infections are very common in freshwater fishes. The fungal
infection is characterized by the appearance of turfts or “cottonwood” on the
skin (dermatomycosis). Highly infectious, the mould parasite spreads with
astonishing rapidity. The mould comprises the growing hyphae of the
fungus. The hyphae wall contains cellulose and tubular space filled with
cytoplasm. However, the problem in the diagnosis is the difficulties in

51
 proper identification of the pathogen, ubiquitous nature, the secondary
infection behaviour and its saprophytic growth pattern.
8.1 (1) Saprolegniasis
The most common species of fungus affecting fish is Saprolegnia
parasitica. Fungus is a secondary infection and bacterial attack is the
primary cause.

Clinical and Pathological Signs

(1) Deformation of bones.
(2) Blindness,
(3) Congestion at the base of pectoral and anal fin.
(4) Inflammation of the liver or intestine.
(5) Fish becomes weak and lethargic and gradually dies after ulceration or
exfoliatioin of the skin.
(6) Gills may also be affected in late stages.
Curing Methods
Treatment includes the following steps:
(1) Dip the affected fish in 3% sodium chloride solution or in 1:2000
solution of copper sulphate or in 1:1000 Kmno 4 solution for about 5
to 10 minutes
(2) Avoid injuries to fish skin
(3) Disinfecting of ponds with fungicidal and fungi static agents
(Malachite green, potassium dichromate).
(ii) Branchiomyces sanguinis
Another disease caused by fungus is the
Gillrot. This is caused by Branchiomyces
sangunis. The parasite usually choke the
vein of gill filaments.
Clinical and Pathological Signs
- Reddish and fading of gill filaments.
- Gasping and suffocation which may lead to the death of the fish.
- Dropping out leaving exposed cartilaginous supporting skeleton.
Curing Methods
- Dip the affected fishes in 3 to 5% sodium chloride (common salt)
solution or in 5ppm Kmno4 solution and keep the fishes in the solution
for about 5 minutes.
- Treat the pond with quicklime or copper sulphate to kill the fungus
spores.
- Add quicklime every few weeks until there is no sign of the disease.
(iii) Aspergillosis
52
 This is caused by the species of Aspergillus niger, Aspergillus flavus,
Aspegillus fumigatus and Aspergillus candidas.
Clinical and Pathological Signs
- Inflammation of visceral organs.
- Systemic necrosis of tissues.
- Darkness of skin due to toxicosis.
- Granuloma formation.
- Emaciation.
Curing Method
- Ponds disinfection with fungicides.
- Avoid water contamination with mold feed.
CHAPTER NINE
9.0 VIRAL DISEASES
Viral diseases are usually of epidemic proportion because of their mode of
transmission. It could be vertical (one generation to next), horizontal
(transmission through contemporaries), Direct contact (fish to fish of all
ages), mechanical (water or non vector) and vector (other organism).
Some of the known fish viral diseae include the following:
- Viral Haemorrhagic septicemia (VHS)
- Infectious Pancreatic Necrosis (IPN)
- Infectious Haematopocitic Necrosis (IHN)
- Channel Catfish virus Disease (CCVD)
- Lymphocystis (LDV)
- Spring Viraemia of Carp (SVC)

9.1 Viral haemorrhagic septicaemia of trout (VHS)

Homopoietic Tissue
Spleen
Liver
Gills

Fig 5: Pathogenesis of VHS

53
Viral haemorrhagic sepheaemia is a disease of Salmonids. Nine species are
presently known to be susceptible to VHS infection leading to clinical
diseases: rainbow trout (Salmo gairdner), brown trout (S. trutta), lake trout
(Salvelinus thymallus), white fish (Coregonus sp). sea bass (Dicentrarchus
labrax) and Turbot (Scophthalmus maximus) and seabream (Chrysophris
aurata). This has been known under different synonyms, such as
Forellensenche, Egtved-virus-Disease or infectious degeneration of kidney
and liver. It is also known as INUL-Krankheit, infectious anaemia,
enterohepato-renal syndrome, and Danish virus. It is now known as viral
haemorrhagic septicemia of trout (VHS).
Clinical and Pathological Signs
- Mortalities occur very rapidly and most fish in a pond are affected.
- The ill fish have irregular reflexes and become dark.
- They stop eating.
- Prominent exophthalmos is very typical.
- The gills are pale and show signs of anaemia.
- At one time, one organ is attacked at another time; another is affected,
though most often the kidneys and the liver are affected.
- There is apathy: - The fish move about very little and rest on the surface
close to the banks.
- When they swim, their movement is sharp and disordered and their
position in the water is abnormal.
- They gyrate and take on a dark colour.
- The belly swells and a foul smelling yellowish liquid is found in the body
cavity.
- The anus protrudes and there are slight haemorrhages in the muscles and
on the swim bladder.
- The liver is grey brown and friable while the posterior parts of the kidney
swell.
- A neutral or alkaline liquid is found in the stomach.
- Intestine turns red and is inflamed.
- The fin fray and there are sores on the skin.
Curing Methods
When farms are affected, it is possible to reduce the virulence of the VHS
disease and the death rate in the following ways:
(1) Bringing in fish from uncontaminated farms;
(2) Isolation of the diseased fish;
(3) Removal of dead or seriously affected ones;
(4) Disinfect the ponds if the disease is very bad;
(5) Avoid high stocking densities and over feeding;
54
 (6) Food distributed should not be too fatty but rich in vitamins containing
appropriate protein and fillers;
(7) If the disease is chronic, feeding must be temporarily stopped and then
followed by the progressive distribution of light and varied food.
9.2 Infectious Pancreatic Necrosis of Trout (IPN)
This has been found in both rainbow trout and Atlantic salmon in both fresh
and sea water. The virus brings sudden and widespread death to young trout
at the start of artificial feeding.
Clinical and Pathological Signs
- Affected fish revolve on their longitudinal axis;
- They swim around rapidly and in a disordered fashion;
- Immobilized near the banks or on the bottom of the pond;
- Become dark in colour;
- Their eyes protrude;
- Viscous whitish liquid in the stomach and intestine;
- Liver and spleen become pale;
- Gall bladder shows severe necrosis;
Curing Methods
- Feeding of povidoneiodine is recommended.

9.3 Lipoid Degeneration of the Liver

This is closely related to viral haemorrhagic septicaemia. It is mainly a
disease of hatchery salmonids and is due exclusively to poor feeding.
It presents identical symptoms with those of VHS. The major difference
between lipoid degeneration of the liver and viral haemorrhagi septicaemia
is that in VHS, haemorrhage concerns the whole body, all the muscles; swim
bladder and the gills. In lipoid degeneration, the liver turns yellow-brown. If
trout suffering from VHS have an affected liver, it is spotted and turns wine
red or grey in colour. The two diseases can exist at the same time.
9.4 Infectious Haematopoeitic Necrosis (IHN)
Infectious hematopoetic necrosis virus, a rhabdovirus, is a major cause of
mortality in salmonids in fresh water. Viral strains vary in pathogenicity for
different salmonids. Coho salmon and brook, brown, and Cutthroat are
considered refractory, although the virus has been isolated from
asymptomatic cohosalmon and from brook and Cutthroat trout.
Clinical and Pathological Signs
IHN causes profound changes in cellular and chemical blood constituents,
primarily because of renal damage. The most clinically useful change is the
presence of remnants of necrotic cells (necrobiotic bodies), probably
erythrocytes, in kidney smears.
55
 Fish are anemic and leukopenic, and evidence of osmotic imbalance.
Curing Methods
- Disinfection and quarantine is the only proven means of controlling IHN
epidemics.
- Extreme caution should be taken to avoid spreading virus to
uncontaminated areas, both within and outside the hatchery.
- Avoidance is the most useful prophylactic measure.
9.5 Channel Catfish Virus Diseases (CCVD)
Channel catfish virus is the most important viral disease affecting channel
catfish. It is restricted to the Channel catfish – producing areas of the United
States except for its accidental introduction into Honduras. It is a highly
species – specific herpesvirus and only naturally affects Channel catfish,
although it can experimentally infects some other ictaurids (blue catfish,
Channel X blue catfish hybrid) and possibly some Clariid catfishes.
Clinical and Pathological Signs
(1) Clinical signs include hanging head up in the water.
(2) Disorientation (corkscrew spiral swimming).
(3) Abdominal distension.
(4) Exophthalmos.
(5) Hemorrhages on the body, gills and at the bases of the fins.
Curing Methods
- Disinfection and quarantine is the most effective means of controlling
CCVD epidemic.
- All fish swimming on outbreak should be destroyed.
- Stress reduction is considered to be important in managing the disease,
including preventing overcrowding, adequate oxygen and nutrition, and
not handling fish when temperature exceeds 20oC.
- Controlling concurrent secondary infections is mandatory.
9.6 Lymphocystis
Lymphocystis is a chronic, self limiting disease affecting many cultured and
wild marine and freshwater fish. It has been reported from over 125 species
in 34 families. Lymphocystis is a disease of higher teleosts and does not
affect salmonids, catfish or cyprinids. An iridovirus, it is the most common
viral infection of aquarium fish. While it causes only low mortality,
lymphocystis is disfiguring and can render affected fish unsalable.
Clinical and Pathological Signs
The lymphocystis virus infects the dermal fibroblasts, producing
tremendously hypertrophied cells that are often just visible to the naked eye.

56
Early or mild stages of the disease appear as a salt – like dusting of the body,
which may later coalesce into larger neoplastic-like masses of hypertrophied
cells.
Curing Methods
- Fish should be watched closely for secondary infections and medicated
accordingly.
- Affected fish should be quarantined, preferably for at least 1 month after
recovery.
- Stress reduction and avoidance of skin trauma are essential to control.
9.7 Spring Viraemia of Carp SVC:- is caused by the Rhabdovirus
carpio which was isolated by Fijan and co-workers in 1971. Affected
species are Cyprinus carpio and pike try.
The incubation time is between 5 and 60 days. Outbreaks of the disease are
usually observed in the spring time. This observation is also the cause for
the term “spring viraemia”. Carp of all ages may be affected.
Clinical and Pathological Signs
The clinical picture is characterized by ceasing to feed, disturbed reflexes,
darkening, exophthalmos and ascites.
Petchial or larger bleeding may be visible on the skin and gills.
At necropsy, fibrinous or clear ascites and haemorrhages in muscles can be
found.
Catharralic enteritis is very common.
Haemorrhages in the serosa of the swim bladder have been observed.
- There can be necrosis in different organs such as haemopoietic tissue,
spleen and liver.
Prevention and Control
The most important factor is to improve the condition of fish before and
after wintering.
- Chloramphenicol can be used at the beginning of winter time.

57
 CHAPTER TEN
10.0 BACTERIAL DISEASE
Bacterial diseases are characterized into three types:
(1) Internal hemorrhaging or bleeding (hemorrhagic septicemia);
(2) Skin ulcers or sores (focal dermomyonecrosis leading to ulceration; and
(3) Chronic loss with no signs of disease (Chronic proliferative response).
10.1 Systemic Bacterial Pathogens are those bacteria that can cause septicemic
disease once they have made their home in the internal organs and tissues.
They have several routes of entry into the fish, such as through the gut or
intestinal wall; through the layers of the skin (integument); through skin
lesions or sores (abrasions); through natural surface openings such as the
nose, lateral line, and sensory pores; and by being brought in attached to
parasites. Once these bacteria break through to the internal structure of the
fish, they are distributed to the organs through the blood stream via the
blood vessels and generally get trapped in the organs that have fixed
macrophages such as the spleen, kidney, heart, méninges of the brain and the
central venous sinusoids of the gills.
Phagocytosis or trapping of bacteria does not always mean elimination of
these bacteria by the fish’s immune system. Therefore, the kidney, spleen,
brain, and gills are often sites where the pathogen multiplies due to relatively
rich nutrient media present and causes regional and systemic disease.
10.2 External Bacterial Pathogens; are those bacteria that cause skin or
integument damage, such as fin rot. They are considered environmental
pathogens rather than systemic pathogens. The sequences of events that
generally lead to an external infection include the following:
1. There are increased numbers of bacteria in the water where the fish
are being held.
2. There are increased numbers of bacteria within the mucus cuticle that
covers the integument.
3. The mucous defense system becomes overwhelmed and unable to rid
itself of the bacteria.
4. The bacteria colonize the epithelial surface below the mucous cuticle
and become stabilized in a bio film.
5. The bio film grows and becomes a mechanism for concentrating
bacterial toxins.
6. Skin and gill erosion and damage occurs.
10.3 Columnaries Infection (Myxobacterial
Disease, Peduncle Disease, saddle back, Fin Rot, Cotton wool
Disease, Block patch Necrosis)

58
 Flexibacter columnaris is an important fish pathogen. It can rapidly infect a
population and cause large mortalities. Flexibacter columnaris is usually
pathogenic at higher than 15oC (59oF). Both mortality and acuteness of
disease increase with temperature.
Clinical and Pathological Signs
- Columnaris is primarily an epithelial disease.
- It causes erosive/necrotic skin and gill lesions that may become systemic.
- It often presents as whitish plaques that may have a red periphery on the
head, back (saddle back lesion) and / or fins (fin rot), especially the
caudal fin.
- Fragments of the fin rays may remain after the epithelium has slaughed,
leaving a ragged appearance.
- Lesions rapidly progress to ulcers, which may be yellow or orange due to
masses of pigmented bacteria.
- Ulceration spread by radial expansion and may penetrate into deeper
tissues, producing a bacteremia.
Curing Methods
- Prolonged immersion in potassium permanganate or copper sulfate.
10.4 Furunculosis (Ulcer Diseases)
Furunculosis is caused by the Gram-negative non-motile bacterium called
Aeromonas salmonicida. It is one of the most serious infectious diseases of
salmomid fish. The disease is named after the raised liquefactive muscle
lesion (furuncles) which sometimes occur in chronically infected fish though
these lesions are rarely seen in acute infections which are characterized by a
rapidly fatal septicaemia. The host range of A. salmonicida also includes
many non-salmonid freshwater and manne fish species.
Clinical and Pathological Signs
- Clinical signs of Aeromonas salmonicida infection range from superficial
or deep skin lesions without systemic involvement (Ulcer disease) to a
typical gram-negative bacteria septicemia.
- The classical form of Aeromonas salmonicids infection primarily affects
salmonids. Clinical signs of furunculosis depend on the time – course of
infection, with gross signs more apparent with increasing chronicity.
- The acute form is the most common, especially in growing fish. It
presents as a typical bacterial hemorrhagic septicemia, with bacteria
disseminated in many tissues; fish often die in 2 to 3 days.
- The subacute/chronic form is less common than the acute form. Mostly
seen in adults, it presents as a more chronic form of bacterial
hemorrhagic septicemia, which may include exophthalmos, bloody

59
 discharge from nerves and vent, multifocal hemorrhages in the viscera
and muscle.
- The gills may be pale from anemia or may have hemorrhages.
- The gastrointestinal tract may have necrotic enteritis and catarrhal
exudates.
- Furuncle develops from localization of hematogenous bacteria in the
muscle or skin, not from an external skin infection.
10.4.1Ulcer Disease
Ulcer disease is the most common form of A. salmonicida infection in
nonsalmonids; salmonids can also be affected. Unlike furunculosis, ulcer
disease is typically localized to the skin and only becomes systemic late in
the disease.
Clinical and Pathological Signs
- Skin lesions range from whitish discoloration to shallow hemorrhagic
ulcers to deep lesions that expose underlying muscle or bone.
- Because of their chronicity, lesions are often secondarily infected with
water molds, protozoa, and other bacteria.
- Fish may have hemorrhage on the body and the base of the fins.
- In eels, infections begin as depigmented foci that spread to form large
patches of necrotic skin up to 16 cm2 in area.
- The infection commonly affects the head, producing cranial swelling and
corneal edema. Aeromonas hydrophila can cause similar lesions.

Curing Methods
- During outbreaks, all moribund fish, especially those with ulcers, should
be promptly removed and disposed of properly.
- Oral oxytetracycline, furazolidone, oxolinic acid and potentiated
sulfonamides have been used successfully.
- Disinfection and quarantine, followed by stocking specific – pathogen
free fish and eggs, can eliminate the infection from facilities, so long as
stocks are not re-exposed to water that has infected feral fish.
10.5 Enteric Red mouth (ERM)
ERM is caused by the Gram-negative motile bacterium (Yersinia ruckeri). It
is principally a disease of rainbow trout but all salmonid species are now
considered potential hosts.
ERM is a subacute to acute systemic infection and takes its name from the
characteristic reddening of the mouth and opercula which is caused by
subcutaneous hemorrhaging. Other signs include inflammation and erosion
of the jaws and palate, haemorrhaging at the base of fins and exopthalmia.
Internally, haemorrhages may occur in muscle and
60
 intestine which may also contain a yellow fluid.

Curing Methods
Yersinia ruckeri has been controlled by a broad spectrum of antiobiotics
including sulphamerazine, oxytetracycline and oxolinic acid.
10.6 Vibriosis
Vibriosis is a bacterial disease of salt-water and migratory fish.
The most commonly encountered fish pathogen (vibrio species) is V.
angullarum and it is this species which is responsible for the majority of
loses worldwide, and was probably the causative agent of the disease. The
next most important member of the Vibrio species is V. ordalii. Other vibrio
species reported to be pathogenic to fish are as follows:
- V. alginolyticus
- V. carchariae
- V. cholarae
- V. damsel
- [Link]
- V. salmonicida
Clinical and Pathological Signs
Symptoms of vibriosis are similar to those caused by Aeromonas
salmonicida.
The external pathology produced by V. angullarum includes haemorrhaging
at the base of the fins, around the vent and gills and inside the month-
petechiae, necrotic lesions and diffuse haemorrhages can appear on the body
surfaces.
- Internally, the intestine is often inflamed with petechiae present on the
viscera and musculature.
- The intestine may be distended and filled with clear viscous fluid.
Curing Methods
- Antibiotics and antimicrobials are commonly used as food additives.
- Commonly used antibiotic is tetracycline.
10.7 Bacterial Kidney Disease
Bacterial kidney disease (BKD) is caused by a Gram-positive bacteria called
Reni bacterium salmonarium, it is short rod (0.8-1.0x0.3-0.5 μm). It is non-
motile, asporogenous, non-acid fast and not encapsulated.
Renibacterium salmonarum is an important pathogen of salmonids,
especially rainbow, brown and brook trout and Coho and Chinock salmon.
Clinical and Pathological Signs

61
 Fish with severe BKD may have no external signs. Affected fish may
present with dark colouration, exophthalmos, pale gills, abdominal
distension, or hemorrhages at the vent or base of the fins.
- The major target organ is the kidney, which has white, nodular masses
- Nodules may also occur in other viscera, especially spleen.
- There may be fluid in the abdomen.
Curing Methods
- Macrolide antibiotic e.g. erythromycin is the most effective agents in
treating diseased fish.
- Only erythromycin, thiocyanate, and phosphate were effective
propylactically or therapeutically against BKD.
10.8 Enteric Septicemia
The bacterium called Edwardsiella ictaluri is the aetiological agent of
Enteric septicemia of Channel catfish (ESC) which has become the most
serious infectious disease problem of cultured Channel catfish (Ictalurus
punctatus).
Clinical and Pathological Signs
- Clinically affected fish may occasionally hang head up in the water and
exhibit cockscrew spiral swimming, usually followed by death.
- Fish may have abdominal distension, exophthalmos or pale gills.
- Hemorrhage and necrosis of the liver, and splenic and renal hypertrophy.
- There may be petechial hemorrhages in the muscles.
Curing Methods
- Oxytetracycline and ormetroprim-sulfadimenthoxine are very effective.
10.9 Dropsy
The bacterium responsible for dropsy is called Aeromonas punctata.
Clinical and Pathological Signs
- General Edema.
- Accumulation of watery fluid, yellow or pink in colour in the body
cavity.
- Withering away of fins.
- Intestines are inflamed.
- Liver and kidneys are affected.
Curing Methods
- Dip the fishes in 5ppm solution of KMno4 for about 2 minutes.
- No food should be given during treatment.
10.10 Eye Disease
The bacterium causing eye disease is the Aeromonas liquefaciens. It affects
medium and large sized carps.
Clinical and Pathological Signs
62
 - The infected sites are – eyes, optic nerves and brain of the fish.
- During the initial stages, cornea of the eye becomes vascularized and
later becomes opaque.
- The eyeball gets putrefied, leading to death.
Curing Method
- During the initial stage of infection, chloromycetin (8 – 10mg/L) both for
one hour given for 2 – 3 days is good for treatment.
- The entire pond should be disinfected by 1ppm solution of kmno4.

CHAPTER ELEVEN
11.0 Protozoan Disease
A number of protozoans cause numerous external and internal diseases
particularly in the young fish. Fry and fingerlings are their large-scale
victims.
11.1 Ichthyopthiriasis
The causative organism for ichthyopthiriasis is Ichthyophthirius multifillis.
It is a fairly widespread disease around the world. It infects different parts
of the body externally.
Clinical and Pathological Signs
- Appearance of numerous small white dot or white spots over the entire
skin, gills, or fins.
- The spots or cysts range between 0.1 – 1.0mm in diameter.
- The spot is the small chamber in which the parasites are accumulated.
Curing Methods
- Affected ponds should be disinfected with salt.
- Dip the infected fish in 1.5000 formalin solutions for 7 to 10 days.
11.2 Trichodina indica
It affects majorly carps.
Costia necatrix causes coast.
Bodomonas rebae cause a lot of problems
in Catla and Mrigal.
Myxobolus Sp causes ulceration with
knots in the skin of carp.

Clinical and Pathological Signs

- Presence of bluish coating on the skin of fish.
- Large amount of mucus is also produced.
Curing Methods
63
 - Bath in 3% Nacl for about 10 minutes.
11.3 Worm Diseases
The flatworm (trematodes). Tapeworms (cestodes). Round worms
(nematodes) and Thorny – headed worms (aecenthocephalans) are all
parasites of fish. They affect the internal organs of the fish body,
particularly the intestines.
11.3.1Trematodes
A. Order:Monogenea (Heterocotylead)
(1) Ancyrocephalus curvianuhoratus Enyenihi
(2) Cichlidogyrus tilapia
(3) Dactylogyrus sp Diplectarium elongatum
(4) Diplectarum enyenihi
(5) Diplectanum jamestowni
(6) Gyrodactylus cichlidarum
(7) Haliotrema sp
(8) Macrogyrodactylus clarii
(9) Mazocracoides ethmalotis Enyenihi
(10)Mazocracoides kwariensis Enyenihi
(11)Microcotyle calabarensis Enyenihi
(12)Microcotyle Okpossi Enyenihi
(13)Protancylodiscoides chryichtyes
(14)Quadriacenthus clariades

B. Order: Digena (Malacocotylea)

(1) Acanthostomium ansconditum
(2) Acanthostrum Spinicaps
(3) Acetodextra Sp
(4) Allocreadium Sp
(5) Brevicaecum miloticus centrovarium Sp
(6) Clinostomum tilapiae
(7) Euchlinestomum galilaeus
(8) Lasiotocus chaetodipteri Thomas
(9) Lasiotocus cynoglossi major Thomes
(10) Monodhelmis torpeddenus Dollfus
(11) Neascus Sp
(12) Pedunculotrema capecoastensis – Fischthal and Thomas
(13) Plagiorchis mutabile
(14) Sandonia Sudanensis
(15) Siphodera ghanensis – Fischthal and Kuntz
(16) Strephano cathemum africanum
64
 (17) Diplostomum Sp
Out of the above listed monogenetic
Trematodes, Gyrodactylus cichlidarum
and Dactylogyrus diplectanum infect
fishes in a big way. Gyrodactylus infects
skin and gills, whereas Dactylogyrus
affects only gills.
Clinical and Pathological Signs
- Fading of colour
- Loss of scales
- Excessive mucus on caudal peduncle and fins.
Curing Methods
- Dip the infected fishes in 1:2000 solution of acetic acid or Nacl for about
5 minutes.
Digenetic trematodes
Diplostomum sp causes Diplostomiasis or black spot disease.
Clinical and Pathological Signs
- The major sign is the presence of small black spots or cysts (1.3mm in
diameter) all over the body of the fish.
Curing Methods
- Isolate the infected fishes.
- Dip the effected fishes in 3:100,000 picric acid solution for a period of
one hour.
11.3.2 Cestodes
These are platyhelminthes with a thick cuticle, suckers or hooks with which
they attach to their hosts as they are endoparasites of aquatic and terrestrial
vertebrates.
CLASS CESTODARIA
Family Camallanidae
1. Procamallanus sp
Family Caryophylacida
2. Wenyonia acuminata
3. Wenyonia synodontis
Family Cucullanidae
4. Cucullanus sp
5. Dichlyne sp
Family Kathlaniidae
6. Spironoura sp
Family Proteoceptialidae
7. Proteophtialus sp
65
 Family Ptychobothriidae
8. Diphyllobathnium sp
9. Monobathrium sp
[Link] sp
[Link] sp
Family Rhabdochonidae
[Link] sp
Tapeworms affect the alimentary canal and gall bladder. It is relatively
more common in wild fish than in cultivated fish. The commonest parasite
is Ligla intestinalis with size up to 15 to 40cm.
Clinical and Pathological Signs
- Infected fish shows a swelling of the belly
- The gonads are adversely affected with loss of fertility.
- It usually leads to death.
Curing Methods
Prevention is the major way of avoiding Ligla intestinalis.
11.3.3 NEMATODES
NEMATODA
These are the true round worms with cylindrical elongate bodies in which
the mouth, located anteriorly is surrounded by three (3) lips – one dorsal and
two ventro-lateral. There are as many free living as there are parasitic
nematodes.
Fish parasitologists and benthic hydrobiologists have provided a list of
nematodes:
Free-living nematodes
1. Dorylalmus sp
Parasitic Nematodes
2. Anisakis sp
3. Capillaria sp
4. Contracaecum sp
5. Cucullanus sp
6. Dichelyne sp
7. Hysterothylacium sp
8. Metabroneum sp
9. Philometra polynemi
[Link] globiceps
[Link] laevichonchus
[Link] denudata
[Link] sp
[Link] sp
66
 [Link] sp
[Link] PHYLUM ACANTHOCEPHALA
This is a small group of parasitic acoelomate invertebrates with an elongated
and cylindrical body in which the anterior is recognized by the presence of
rows of hooks surrounding the eversible pharynx. Their common name is
“SPINY HEADED WORMS”. Investigators of fish parasites have provided
a list of six (6) Acanthocephalans.
Order – Acanthocephala
1. Acanthella sp
2. Acanthogyrus sp
3. Neochinorhynchus sp
4. Polymorphus sp
5. Quadrigyrus sp
6. Rhadinorhynchus pristia
They have all been found to infect fishes. They affect the alimentary canal
of fishes but very little is known about their pathological effects. In addition,
Heliconema, proleptus, camallanus are all dangerous to the life of fish.
11.4 Annelida
The annelids are metamerically segmented coelomate inverterbrates bound
by a soft and thin cuticle through which segmentally arranged CHAETAE
emerge. There are three main groups of free living annelid in our water
bodies the leeches, Oligochaetes and polychaetes.
Hirudinaria (Leeches)
It infects major carps and feed on the blood of the host fish.
Clinical and Pathological Signs
- It causes irritation and abnormal movements of the hosts.
Curing Methods
- Dip the infected fish in 1:1000 solution of acetic acid and disinfect with
1:10000 kmn04 solution.
- 5 ppm Gammexane was observed to cure carps infected with leeches.
11.5 Crustacaen Diseases
Parasitic crustaeeans with indistinct body segmentation with lateral
expensions of the head and a pair of compound eyes.
The fish lice (Argulus, lernea and Ergasilus) belonging to crustaeea are
important fish ectoperasite.
11.5.1 1. Argulus africanus
2. Argulus schoutedeni
3. Chonopeltis brevis
4. Chonopeltis minutes
11.5.2 Family Ergasillidae
67
 1. Caligus haemuloni
2. Ergasilus africana
3. Ergasilus latus
4. Ergasilus lizae
5. Ergasilus kandti
6. Ergasilus monodi
7. Ergasilus nodosus
8. Lamproglema clarias
9. Lamproglema hamprechi
10. Lamproglema monodi
11. Lamproglema werneri
12. Lamproglema wilsoni
13. Lepeophtheirus monocanthus
14. Lerneae hardinji
15. Neoacila orbignyi
16. paraergasilus lagoonalis
They weaken fish by feeding on its blood.
Argulus sp and Lerneae sp attach themselves to the fish with their body
buried in the scale pockets and with paired egg sacs protruding free.
Argulus sp attaches itself to the body of the fish by means of suckers and
hooks, but it can also swim freely in water.
NB: Ergasilus is generally found attached to the gills of fishes.
Lerneae also known as anchor worm is an external parasite which attaches
itself to the host fish anywhere on the body by means of anchor – like
appendages present on the head of the parasite. The anterior portion of the
parasite is buried deep into the skin of the fish, and only the posterior region
may be visible externally.
Clinical and Pathological Signs
- Loosing of scales.
- Appearance of red spots at the sites of infection.
- Stunted growth.
Curing Methods
- Mechanical removal of the parasites.
- Dip the infected fishes in a weak kmno4 solution for 2 – 3 minutes.
11.6 Mollusc Larvae
Glochidium larvae cause heavy infection in fishes and may result in the
death of a fish.
Clinical and Pathological Signs
- Gill arches are damaged.
- Filaments are also damaged.
68
 - Appearance of tumours
Curing Methods
- Mechanical removal is the best choice.
CHAPTER TWELVE
12.0 Non – Infectious Disease
12.1 Asphyxiations:
Asphyxia refers to conditions in which hypoxia is combined with hyper
capnia (an increased CO2 tension in the arterial blood). Hypoxia refers to
any condition in which there is an inadequate supply of oxygen to the
tissues. Gas disease in fish is caused by under super saturation of dissolved
oxygen.
Clinical and Pathological Signs
- Bubbles appear beneath the skin, around the eyes, on the fins, in blood
capillaries and in the alimentary canal.
- Fish swims at an angle of about 45 and head downwards.

Curing Methods
- Transfer the fishes or change their environment.
12.2 Miscellaneous Disease
12.2.1 Algae Toxicity/Bloom
This sometimes occurs during blooms of certain blue – green algae. Low
oxygen and high PH, hydrogen sulphide, ammonia and nitrite are water
quality parameters that can cause environmental diseases in fish. Some algae
may cause mortality among fishes by chocking the gills or by releasing toxic
substances in the water.
The spread of the algal bloom all over the surface of water usually causes
obstruction to the sunlight.
- Percentage of dissolved oxygen in water is also lowered. Thus, it creates
mortality of the fish.
Clinical and Pathological Signs
- Gills are choked.
Curing Methods
- Algae are removed by beating the surface of water with bamboo sticks.
- Transfer all the fish to another pond.
12.3 Nutritional Diseases
Nutritional diseases have been observed from time to time and the most
commonly observed have been Lipoid liver degeneration (LLD) and

69
 thiamine deficiency. The latter is caused by feeding of fish on feed
containing thiaminase or feed deficient in the vitamin.
LLD has usually resulted when fish were fed rancid herring or capelin, but
also when old pelleted dry feed were used, the conditions has been
characterized by a bilateral increased cloudiness and opaqueness of the lens
resulting in blindness to a greater or lesser extent. Due to the blindness,
secondary traumatic eye lesions also occurred resulting in increased
mortality. The disease has been associated with dry feed with low levels of
zinc.
12.4 Problems Related to Poor Protein Quality
Most important consideration is given to Essential Amino Acids. Fish
effectively have absolute requirement for ten Amino – acids: Argimine,
histidine; isoleucine, leucine, lysine, methionine, phenyl-alamine,
Threonine, Tryptophan and valine.
12.4.1 E. A. A. Deficiencies
Generally – E. A. A. deficiency produces loss of appetite and a concomitant
reduction in growth.
Specially:
Tryptophan deficiency results in scoliosis/Cataract. Methionine
deficiency results in Cataracts. Lysine deficiency results in Dorsal fin
erosion.
The problems of amino acid deficiency usually occur as a result of:
(i) Excessive heat processing of ingredients during food manufacturing;
lysine particularly is susceptible.
(ii) The use of second class protein sources, which may be deficient in one or
more of the 10 Essential Amino acids. For example, meat and bone meal are
deficient in lysine and Tryptophan.
(iii) The use of protein sources with imbalanced amino acid profiles.
e.g.; Blood meal appears to be good protein source, having
approximately 85% crude protein. Unfortunately, blood meal has
very high leucine content but low isoleucine; leucine has an
antagonistic effect on isoleucine absorption. So, isoleucine levels in
the diet must be increased to compensate.

70
12.4.2 Problems Related to Lipid Quality
(1) Essential Fatty Acid Requirements.
Essential Fatty Acids are required by fish in order to maintain the structure
and fluidity of the phospholipids membranes.
Coldwater fish species tend to be more demanding in their requirement for
the E.F.A’s than warm water species. The reason for this is that membrane
fluidity is much more difficult to maintain at low temperature than at high
temperature. Carnivorous fish have high polyunsaturated fatty acid content
in their natural diet compare to herbivorous fish species, so they may have a
poorer ability to desaturate and interconvert fatty acids.
Carnivorous fish tend to be more demanding in their fatty acid requirements.
In Rainbow trout; Essential fatty acids deficiency disease is prevented by
addition of 1% linolenic acid to purified diets.
Turbot and Red sea Bream species are incapable of desaturating fatty acids
to acids with full Essential Fatty Acids activity, so they must be provided
with preformed series of polyunsaturated fatty acids in their diets.
E. F. A Deficiency Symptoms
- Reduced growth.
- Caudal fin erosion.
- Shock Syndrome.
- Overstimulation of muscles.
- Fatty livers.
These second aspect of the problems related to lipid quality is that of:
12.4.3(ii) Rancidity or Lipid Oxidation
Rancidity problems arise in relation to feed ingredients which contain lipids
with a significant proportion of C =C double bonds, those which are highly
unsaturated. These highly unsaturated lipids are particularly susceptible to
oxidation on exposure to air during storages and unless suitably protected by
invitro antioxidant such as Ethoxyquin, B.H.T which may eventually
produce problems when rations in which they have been incorporated are fed
to fish.
Fish fed diets containing rancid oils may demonstrate symptoms which
include;
- Dark colouration.
- Anaemia.
- General lethargy.
- Ceroid fluid in liver and intestine.
- Gill clubbing.
- Sunken back syndrome.

71
 Protection from the effects of rancid lipids can be conferred by the
incorporation of vitamin E in production rations.
Vitamin E (alpha Tocopherol) requirement;
Generally 40 – 50mg Kg-1 diet
Suggested 125 – 175mg kg-1 diet
Salt 400mg kg-1 diet.
12.4.4Vitamins and Vitamin Deficiencies
Vitamin deficiency problems can often be related to one or more stages in
the production, storage and delivery of feed rations.
[Link] Two Main Groups:
Fat soluble and water soluble.
(i) Fat soluble vitamins deficiency.
SYMPTOMS
Vit. A: Reduced growth, poor food
conversion, (possible weight loss), ‘Pop eye’, skin discoloration,
anaemia, haemorrhagic kidneys/skin/fins, warped opercula,
oedema/ascites.
Vit. D: Sterol. Absorbtion/deposition of calcium phosphate, poor bone
and poor food conversion.
Vit. E: Fat Rancidity – invivo antioxidant. Reduced growth, poor food
conversion, ‘Pop eye’, skin discolouration, anaemia, ceroid fluid
(liver/spleen/kidney), oedema/ascites, muscular dystrophy, clubbed
gills, lordosis.
Vit. K: Prothrombin synthesis in liver, blood clotting, anaemia and
prolong clotting tune.
(ii) Water Soluble Vitamins
deficiency SYMPTOMS:
Vit. C: (Ascorbic acid) – Collagen formation, wound healing etc. loss of
appetite, reduced growth, poor food conversion, skin depigmentation,
anaemia, scoliosis/lordosis, haemorrhages in
skin/fins/liver/kidney/intestine/muscle, mortality.
Vit. B1: (Thiamine) – Phosphate = coenzyme – carbohydrate metabolism.
Reduced growth, poor food conversion, skin discolouration, anaemia,
loss of balance, convulsions, corneal opacity, anorexia, fatty livers
and high mortality.
Vit. B2: (Riboflavin) Loss of appetite, reduced growth, poor food
conversion, cataracts, cloudy lens, dark colouration, haemorrhage in
eyes/nose/opercula, loss of coordination, anaemia , mortality.
Vit. B6: (Pyridoxine)-

72
 Loss of appetite, reduced growth, poor food conversion, ‘Pop eye’,
anaemia, convulsions, hyperumtability, spasm, cataracts, rapid onset
of rigor mortis.
Vit. B12: (Cobalamine)
Loss of appetite, anaemia
Folic acid: Loss of appetite, reduced growth, poor food conversion, anaemia,
pale gills, dark skin, letharzy, “Pop eye”, mortality.
Niacin: Loss of appetite, reduced growth, poor food conversion, swollen
gills, intestinal lesions, anaemia, skin haemorrhages, loss of
coordination, muscle spasm, oedema, mortality.
Pantothenic acid: Loss of appetite, reduced growth, poor food conversion,
clubbed gills, sluggistues, anaemia, flaring of opercula, mortality.
Biotin: Loss of appetite, reduced growth, poor food conversion, anaemia,
enlargement of liver, blue – slime disease (skin), dark colouration,
spastic convulsions, cataracts, mortality.
Choline: Reduced growth, poor food conversion, haemorrhagic
skin/liver/kidney/intestine, fatty livers.
MINERALS AND MINERAL
DEFICIENCIES
Sodium, Potassium, chlorine: Osmorgulation and acid – base balance are
readily absorbed through gut, gills and skin.
Calcium: Skeletal component. Activation of enzyme systems associated
with nerve transmission and contraction of muscle. They are readily
absorbed through gut, gills and skin.
Magnesium: Skeletal component. Enzyme activator, associated with
enzymes involved in energy transfer. They are readily absorbed
through gills, skin and gut.
Phosphorus: skeletal component. Important constituent of phospholipids,
nucleic acids phosphoproteins, A.T.P., etc and several key enzymes,
mostly derived from diet.
Iron: Essential component of haemoglobin, myoglobin, the cytochromes and
various enzymes systems: They are readily absorbed from water.
Absorption depressed by high dietary intake of phosphate. Calcium
phytates, copper, zinc, or a low intake of nickel. Ferrous iron more
readily absorbed than ferric iron . Vit, C enhances absorption of non-
hem iron.
Zinc: Essential component of several metal co-enzyme, and has role as co-
factor for many enzymes. Zinc absorption and availability depressed
by phytates and high dietary intakes of phosphorus, calcium and
copper.
73
 Copper: Essential component of several key enzymes, and essential for
normal haematopoiesis. Absorbed from water. Copper absorption
depressed by phytates, and high dietary intakes of zinc, iron,
molybdenum, cadmium sulphate and calcium carbonate.
Molybdenum: Enzyme activator for enzymes which mediate phosphate
transfer. They are readily absorbed from water. Molybdenum
absorption and availability depressed by phytates and high dietary
calcium intake.
Selenium: Component of Glutathione peroxidase, and functions similarly to
vitamin E. They are readily absorbed through gills and gut.
Cobalt: Essential component of vitamin B12, prevent anaemia.
Fodine: Constituent of thyroid hormones. They are readily absorbed through
gut and gills.

CHAPTER THIRTEEN
13.0 LAWS BINDING TRANSPORTATION OF FISH ACROSS
INTERNATIONAL BORDERS
(1) No foreign vessel is allowed to come and fish within the EEZ
(Exclusive Economy Zone) of another country or take fish to their
country.
(2) Before life fish are allowed into a country. It must be put in
quarantine by the receiving nation to ensure it is declared disease free;
(3) smoked/preserved or ice fish must be declared fit for consumption by
the receiving nation.
(4) In canned fish, apart from been declared fit for consumption, the
expiring date must be indicated on the package and the tin/can.
(5) Those species prohibited in international trade must not be transported
across the country.
(6) Exotic species which when introduced into a country has ecological
advantage over the indigenous species are not usually welcome,
except it will be to the advantage of the receiving nation.
(7) Smuggling fish into a country without legal backing is against the
international law.
(8) Sub-adult/fry fishes are not allowed to be transported across a country
except by legal backing.
13.1 Fish as Source of Human Disease:
Fish may act as source of human disease in two ways:
(1) Direct contamination of fish eaten, fish itself not being affected.
(2) Transmission of disease agents to man, which is called zoonosis.
13.2 Contamination
74
 This may be due to contamination of fish flesh by;
(a) Bio accumulation of chemical pollutants such as heavy metals, pesticides
or herbicides.
(b) Accumulation of biotoxin from bacteria, blue – green algae or
dinoflagellates. These can enter into human food chain.
Accumulation of heavy metals (Methylmercury) in fish has caused
concern in U.S and Canada in 19 th centuries. In 1953, many fishermen
and domestic animals living around Minamata Bay in Japan contracted a
disease characterised by neurological disorder because they had eaten
fish/shellfish taken from the Bay near a plastic factory where mercury
and its salts were used as catalyst and the effluent containing methyl
mercury was been discharged into the Bay.
A biotoxin from dinoflagellate toxin accumulated in muscles (Mytilus sp)
was reported to be responsible for ‘Paralytic shellfish poisoning’ (PSP) in
Britain in 1968.
Biotoxins and naturally occurring poisons in marine dinoflagellates such
Gonyaulax catenella and Crymnodium brevis accumulate in shellfish
(crabs, lobsters, mussels, clams) when eaten.
Symptoms of PSP (Paralytic Shellfish Poisoning).
- Paresthesia (tingling, numbness/burn from the mouth) lips and tongue
and later spread to the face, scalp, neck, fingers and toes).
- Cardiovascular collapse (heart failure).
- Respiratory failure.
Curing Method
- Avoid sea foods from waters laden with toxin dinoflagellate.
13.3 Ichthyosarcotoxic fishes
These are fishes that contain toxin in their flesh, viscera, skin or slimming
mucus covering and when eaten produce biotoxin in man. Scombroid
poison is a good example.
Scombrotoxin causes Scombroid fish poisoning. It is caused by fishes of
sub-order scombroidae (Tuna, mackerel and skip jack).
The poisoning is usually caused by improper preservation of scombroid
fishes which may result in Proteus morgani bacterium acting on instidine in
the muscle of scombroid fish and converting it to saurine. The toxin is
associated with bacterial enzyme action on the fish flesh during spoilage.
Symptoms of Scombroid Poisoning
- Headache.
- Dizziness.
- Epigastric pain.
- Diarrhoea.
75
 - Throat burns.
- Cardiac Palpitation.
- Abdominal pains.
- Respiratory distress.
- Death.
Prevention and Control
- Freezing.
- Canning.
- Immediate Consumption.
- Avoid too long preservation.
13.4 Ciquatera Poisoning
This is contracted through eaten of such fishes like barracudas, garupers,
seabasses, jacks, sharks, eels, snappers, ladyfish, surgeon fish and etc.
Majority of fish involved in ciquatera poisoning are reef or shoreline species
which has become toxic by feeding on herbivorous fishes which inturn feed
upon toxic algae or other toxopheric substances in the coral reefs or related
areas.
Flesh of these fishes is less toxic than their visceral with liver being the most
poisoning part.
The toxin (C35H65NO8) causes respiratory paralysis in man.

76
 Symptoms of Ciquatera Poisoning
- Nauses.
- Paresthesis (tingling sensation) around the mouth, tongue, throat, face;
fingers and toes.
- Abdominal pain.
- Vomiting.
- Diarrhoea and Chill.
Prevention
- Avoidance is the best option.
13.5 Tetradon Poisoning (Tetradon toxin)
This can be contracted through eaten of puffer fish. This is usually found in
Asia. 50 out of 100 species of puffer fish are poisonous.
13.6 Muray eel poisoning
This is found in parts of the world where Muray eel are eaten. The poison
usually paralyses the diaphragm of the eater.
Some other fish species whose skin or flesh may be toxic to men include the
lampreys and hagfish whose slummy skin and flesh may be toxic, chimeras
or ratfish which contain neurotoxin.
Angler sp has poisonous blood.
Other aspect of fish borne infections is biological contaminants:-
Bacterial Infections:- This may be through either Gram-positive or Gram-
negative bacteria.
Bacteria and virus are agents of food poisoning which can be transmitted by
fish.
(b) Viral infection – through food poisoning
(c) Mycotoxins – These are poisonous substances produced by fungi and
are referred to as aflatoxin. Aflatoxins are very hepatocarcinogenic.
Zoonoses (Infective agent)
These are endogenous infections transmitted between animals and man.
These are bacteria and parasites infections transmitted by fish.
(a) Mycobacterium fortitum – It causes skin infections. It can also infect
internal organs. It grows very well at 37OC or between 18 – 42OC.
(b) Mycobacterium Marinum. It causes swimming pool granuloma, chronic
dermal ulcers. It is a disease of aquarium fish. It grows between 33 –
37OC.

13.7 Helminth Zoonosis

Transmission of worm parasites between animal and man, such as cestodes
and nematodes.

77
 Cestodes transmitted to man include Diphyllobothrum sp found in white
worm in the fish muscles and viscera. They are contracted through eaten of
raw or improperly cooked fish.
Preventions
- Freezing at 5OC for over 1 hour
- Dipping in brine solution (3% Nacl).
Digenetic trematodes can also be transmitted to man through eaten of raw or
improperly cooked fish. Thise include members of Heterophye,
stellantcharis, Haplorchis and Anisakidae.
13.8 Occupational Hazards
This may take the forms of:
(1) Bites and stings of dogfish, stangrey, catfish and other dangerous aquatic
animals.
(2) Incidental diseases e.g Leptospires may be transmitted to man through
skin contact with water contaminated by urine of infected rodents and
other animals. Ponds usually serve as breeding ground for disease
vectors like snails, mosquitoes and leeches.

CHAPTER FOURTEEN
Haematological Assessment of Fish Blood
Haematology:-
This is the scientific study of the blood. Whole blood is actually a mixture
of red blood cells, while blood cells and platelets which are suspended in
plasma. Whole blood therefore separates into three distinct layers when it is
spun at high speed in a centrifuge.
The greater density of red blood cells causes them to pack tightly in the
bottom of the tube. The white blood cells and platelets appear as a buffy
coat in the middle of the tube, and blood plasma appears as a clear layer
from the centrifuge (Fig. 6).

78
Fig. 6 centrifuge

Whole blood can be separated into three distinct layers by spinning it at high
speed in a centrifuge. The layers correspond to the red blood cell, white cell
(buffy coat) and plasma. The main function of the haematology laboratory
are to detect blood disorders such as anaemia or leukaemia, to assist in the
accurate characterization and diagnosis of these disorders so that appropriate
treatment can be given and to monitor the progress of treatment.
Haematological tests
This involves the following:
(1) Measurement of haemoglobin and basic investigation of anaemia.
(2) Measurement of pack cell volume (Haematocrit).
(3) Measurement of red cell indices.
(4) Calculation of red cell indices.
(5) White blood cell (WBC) count.
(6) Platelet count.
(7) Differential WBC count.
1. Haemoglobin
This is the main intracellular protein of the red blood cell. Its primary
function is to transport oxygen to the cells and remove carbondioxide.
The haemoglobin molecules consist of two main parts – heme and globin.
Heme is composed of the red pigment, porphyrin and iron which are
capable of combining loosely with oxygen.
Globin is a protein that consists of nearly 600 amino acids organized into
four polypeptide chains. Each chain of globin is associated with a heme
group. Haemoglobin determinations are of greatest use in the evaluation
of anemia, as the oxygen – carrying capacity of the blood is directly
related to the haemoglobin level, rather than to the number of
erythrocytes. In order to interprete result accurately, the haemoglobin
level must be determined in combination with the haematocrit level.

79
 Normally, haemoglobin and haematocrit levels parallel each other and
are commonly used together to express the degree of anemia. The
combined values are also useful in evaluating situations involving blood
loss and related treatment.
The haematocrit is normally three times the haemoglobin level (i,e,
haematocrit = 3 x haemoglobin). If erythrocyles are abnormal in shape
or size, or if haemoglobin manufacture is defective, the relationship
between haemoglobin and haematocrit is disproportionate.

2. Haematocrit (packed cell volume)

Blood consist of a fluid portion (plasma) and a solid portion that includes
red blood cells, white blood cells and platelets. More than 99 per cent of
the total blood cell mass is composed of red blood cells. The haematocrit
measures the proportion of red blood cells in a volume of whole blood
and is expressed as a percentage. With the newer auto mated methods of
cell counting. The haematocrit is calculated indirectly as the product of
the red cell count and mean cell volume (MCV). Normally, the
haematocrit parallels the erythrocyte count. Thus, factors influencing the
erythrocyte count will also affect the results of the haematocrit.
3. Erythrocyte Count
The erythrocyte count is the determination of the number of red blood
cells (RBC) per mm3. In international units this is expressed as the
number of RBCs per liter of blood.

4. Red Cell Indices

Red cell indices are calculated
mean values that reflect the size, weight and haemoglobin content of
individual erythrocytes. They consist of the following:
- Mean corpuscular volume (MCV).
- Mean Corpuscuar Haemoglobin (MCH) and the Mean Corpuscular
Haemoglobin Concentration (MCHC).
- MCV (Mean Corpuscular Volume) indicate the volume of the
haemoglobin in each red blood cells,
- MCH (Mean Corpuscular Haemoglobin): is the weight of the
haemoglobin in each red blood cell.
- MCHC (Mean Corpuscular Haemoglobin Concentration): is the
proportion of haemoglobin contained in each red blood cell.
- Mean Corpuscular Haemoglobin:
This is the average amount of haemoglobin in each cell,
measured in pictograms (Pg = 10-12 gram).
80
 haemoglobin (gm/100 ml) x 10
MCH= = MCH Pg+
Erythrocyte count
Mean Corpuscular Haemoglobin Concentration: This is the amount of
haemoglobin in 100ml of red blood cells.
haemoglobin (gm/100 ml) x 100
MCHC=
haemotocrit
=MCHCg/1.
Mean corpuscular volume: This is the
Average volume of cells, measured in
femtolitres (f1 = 10-8 litres).
haematocrit x 10
MCV = =MCVf1+
erythrocyte count

5. White Blood Cells

Six different types of white blood cells (leucocytes) are found in the blood. While
blood cells are polymorpho-nuclear neutrophils, polymorphonuclear eosinophils,
polymorphonuclear basophils, monocytes, lymphocytes and occasional plasma
cells. There are also large numbers of platelets that are fragments of other types of
white blood cell called magakaryocytes. Neutrophils, eosinophils and basophils
are collectively termed granulocytes because of their granular appearance. The
granulocytes, monocytes and a few lymphocytes are formed in bone marrow while
lymphocytes and plasma cells are formed in the lymph tissue (Fig. 7).

Basophils
Eosinophil

Monocytes
Eosinophil Lymphocylo
Neutrophils

Fig. 7: Relative concentration of white blood cells

Neutrophils and lymphocytes normally comprise around 90% of the white
blood cells. The remaining fraction is made up of a combination of
monocytes, eosinophils and basophils. Neutrophils and macrophages that
are derived from monocytes attack and destroy bacteria and viruses by
81
phagocytosis. Monocytes that enter cells are transformed into macrophages
when they grow in size and develop large numbers of lysosomes that contain
hydrolytic enzymes.
Purposes of haematological tests
A. Indications/purpose for a Complete Blood Count.
(1) Suspected haematological disorder, neoplasm or immunologic
abnormality.
(2) Suspected infection.
(3) Monitoring effect of physical or emotional stress.
(4) Monitoring desired responses to drug therapy and undesired reaction
to drug that may cause blood dyscrasias.
B. Indications/purpose for haemoglobin determination
(1) Routine screening as part of a complete blood count.
(2) Along with a haematocrit, to evaluate known or suspected anemia.
(3) Along with haematocrit, to monitor blood loss and response to blood
replacement.
C. Indications/purpose for Haematocrit determination.
(1) Routine screening as part of a complete blood count
(2) Along with a haemoglobin, to monitor blood loss and response to
blood placement.
(3) Along with a haemoglobin, to evaluate known or suspected anemia
and related treatments.
(4) A decreased haematocrit may indicate hemodilution.
(5) An increased haematocrit may indicate dehydration.
D. Indications/purpose for an Erythrocyte count.
(1) Routine screening as part of a complete blood count.
(2) Suspected hematologic disorder involving red blood cell destruction
(e.g haemolytic anomia).
(3) Monitoring effect of acute or chronic blood loss.
(4) Monitoring response to drug therapy that may alters erythrocyte
count.
E. Indications/purpose for red cell indices
(1) Routine screening as part of a complete blood count.
(2) Identification and classification of anomia.
F. Indications/purposes for Differential white blood cells.
(1) Routine screening as part of a complete blood count.
(2) Abnormal total white cell count to determine the source of elevation
(3) Confirmation of the presence of various disorders associated with
increase and decrease in the several types of white blood cells.
(4) Monitoring physiologic responses to anti neoplastic therapy.
82
 Table 1: Summary of Red Cell Indices in common anomia.
Reference MCV MCH MCHC
Range 80- 27- 32-
98+1 32pg 36g/1
Anemia
Normocytic N N N
normochromi
c

Microcytic ↓ ↓ ↓
hypochromic
e.g from
deficiency

Macrocytic
normochromi ↑ ↑ N
c e.g lack of
vit. B12 and
folate

Note: N = Normal
↓ = Reduced value
↑ = Increased value

Table 2: classification of anemia and Examples

S/n Anemia Examples of MCV MCH MCHC
causes (Pm3) (Pg) (%)
1 Normorcytic Sepsis,
normochromic haemorrhage,
haemolysis, 82-92 25-30 32-36
drug-induce,
aplastic anemia,
radiation

2 Microcytic Renal disease, <80 20-25 27

normochromic infection liver
disease

3 Microcytic Iron deficiency,

83
 hypochromic lead poisoning, 50-80 12-25 25-30
thalassemia

4 Macrocytic Vitamin B12

normochromic and folic acid
deficiency,
some drugs, 95-150 30-50 32-36
pernicious
anemia

Anemia
Anemia is a deficiency of haemoglobin in the blood that can be due to a
reduced number of red blood cells or a low concentration of haemoglobin in
the red blood cells. A number of anemia have been identified:
(1) Hemorrhagic anemia: This occur after
blood loss or during a parasitic infection.

(2) Aplastic anemia: This is due to a

dysfunction in the bone marrow where red blood cells are
made.
(3) Megaloblastic anemia: This is caused
by an inadequate supply of the vitamin B12 and folic acid.

(4) Microcytic anemia: This is due to lack

of Iron, protein, or vitamin B 6 necessary for the synthesis
of haemoglobin.
(5) Haemolytic anemia: This is stems
From abnormatities that make the red blood cells fragile
and have a shortened life span.
(6) Sickle cell anemia: This is due to an
abnormal form of haemoglobin called haemoglobin S
that precipitates into elongated crystals when it is
exposed to low concentration of oxygen. This
precipitation changes the shape of the red blood cell from
that of a biconcave disc to that of a sickle thereby
impending the flow of blood to the tissues.
HEMATOLOGICAL VALUES
Constituent Normal Ranges
Haemoglobin 13.5-18.0g/d1
Haematocrit 40-50%
Erythrocyte (red
blood cell count) 4.2-6.2million/mm3
84
 Mean corpuscular
volume (MCV) 80-90µm3
Mean corpuscular
haemoglobin (MCH) 27-32pg
Mean corpuscular
Haemoglobin
concentration (MCHC) 32-36%
Leukocyte (White blood
cell Count) 5,000-10,000/mm3
Leukocyte differential
Lymphocyte 25-33% of Leukocyte
Count
Monocyte and
Macrophage 3-7% of Leukocyte
Differential
Eosinophil 1-4% of Leukocyte
Differential
Basophil 0-0.75% of Leukocyte
Differential
Platelet 140,000-340,000/ mm3

15. CHAPTER FIFTEEN

15.0 Prevention of Diseases

A fish farmer should aim at preventing (prophylactic) rather than treating
diseases (therapeutic). Good farm management is of primary importance in
avoiding diseases and parasite attacks.
The working procedures are:
1. Pond conditioning: Pathogens and diseases vectors develop in cycles. In
most cases their spores, eggs and cysts survive in pond bottoms. Periodic
drying and living of ponds will kill eggs or spores and their immediate
hosts (e.g. snail and aquatic weed).
2. Treatment of Fish before stocking: Fish should be given bath treatment
before stocking. Transfer of fish from small to bigger ponds reduces the
chance of reproduction in parasites.
3. Stocking Density: Avoid overcrowding of fish at any time particularly
during hot weather. Observe recommended stocking rate for each fish
species and for species combination.
4. Brood stock management: Preventive treatment should be given to brood
stock before they are used for spawning. They should be separated from
young fish hatching as soon as possible.

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5. Provision of good quality and pathogen free water: Ensure that pond and
hatchery water is in sufficient quantity and readily available at all times.
Water should not be passed from one pond to another.
- Control of wild fish and predators: Wild fish live in canals and other
natural water bodies. They are therefore potential hosts and vectors for
disease pathogens. It is therefore recommended that they should be
barred from gaining access into ones farm by:
- Placing wire mesh screens on all water inlets
- Use of chemical on them before stocking the pond.

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15.1 THERAPEUTIC TREATMENTS FOR CONTROLLING PARASITES
AND DISEASES IN CULTURED FISHES
The treatment of fish diseases with various medication and
chemotherapeutic agents is only to some extent, and for the purpose of
holding the disease organisms in check, retard their growth or even kill the
pathogen, but cannot eliminate 100% of disease causing organisms present
in the pond water. If the treatment or cure of fish disease is to be successful,
it is the protective mechanisms of the fish that must overcome the disease
producing organism.
Parasites, diseases and their therapeutic treatment are listed below:
(i) Aeromonas salmonicida: medication: Betadina, chloramphenicols,
furacine,
furnace, furazolidone.
(ii) Flexibacter colummnaris and other external bacteria Medication:
Acriflavine, copper sulfate, diquat, furacin, furnace, Hyamine 3500,
potassium permanganate (Kmn04), oxytetracycline Roccol.
(iii) Edwardscella spp
Medication: oxytetracycline.
(iv) Aeromonas hydrophylla and other bacteria medication:
chloramphenicol, combiotic, furnace, Furazolidone.
(v) Argulus sp, lernaae and Egersilus medication: Baytex, malathion,
masoten.
(vi) Epistylis
Medication: Table salt.
(vii) Fungus:
Medication: copper sulfate, potassium permanagenate
(viii) Trichodina, Bodomonas, and Myxobolus sp
Medication: copper sulfate, formalin, methylene blue, metronicazole,
potassium permanganate
(ix) Leaches:
Medication: Baylex, masoten, Table salt.

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 OTHER ENEMIES OF FISH
List of some invertebrate and predators of fish
Group Parasite/Predators Type of Receptacle

ARTHROPOD (Parasitic) Isopoda (Praniza) Fingerlings production; tank fish

culture
Brachiura (Argulu sp)

Copepodes (Ergasilus sp, Caligus sp

and Lernea sp)

Cladocera (Branchiopoda)

Rotifers

HIRUDINEA (Parasitic) Leeches Pond fish culture

MULLUSKS (Parasitic) Snails, bivalve larvae Pond fish culture

INSECTA (Predator) Odonata nymph (dragon fly) Fingerling production, pond and
tank culture

Fingerling production, pond and

Nepa (water scorpion) tank culture

Belsotoma (water beetle) ˵

Notonecta (back swimmer) ˵

Corixa (water boat man) ˵

Cybister (water skater) ˵

AVIAN (Predator) Piscivorous birds Extensive pond and reservoir fish

culture

AMPHIBIAN (Predator) Frogs and Toads Pond fish culture

REPTILE (Predator) Crocodile, monitor lizard terrapin Pond and reservoir fish culture
and tortoise.

MAMMAL ( Predator) Otters and Swamp Mongoose, Pond and reservoir fish culture

SUBCLASS MALACOSTRACA

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 ORDER ISOPODA

The Isopoda (Greek isos, equal; podos, foot) are dorsoventrally flattened. All the thoracic limbs
of isopods are similar and so are all the abdominal appendages. Examples of isopods include
Ligia gracilipes which lives on the Nigerian seashore. Some isopods are parasitic e.g. Bopyrus
found permanently fixed to the gills under the carapace of decapods crustaceans, prawns.

Order: Odonata (Dragonfly and Damselflies).

Among Odonatans, dragonfly Nymphs (Anisoptera) are predatory and feed upon Carp spawn.

Among aquatic bugs (Hemipterans), a good number are inimical to fish seed e.g., Notoneota,
Anisops, Plea sp (Back swimmers) attack and eat carp spawn as soon as it is released into the
ponds. They do not only attack carp spawn but small fish as well.

Water –scorpions (Ranatra and Laccotrephes or Nepa) usually attack young fishes but if they get
into the spawning ponds in large numbers can cause substantial harm, killing many larvae.

Predatory diving beetles (Coleopterans) belonging to genera like Cybister, Hydaticus,

Sandrocottus, Eretes, Laccophilus, Hyphoporus etc., feed on all kinds of aquatic metazoans, the
larger species commonly attacking small fish, tadpoles and even dragonfly nymph. The larvae of
Cybister sp (water skater) are highly destructive to carp fry (24-40mm) and even fingerlings (10-
13cm). Similarly, the larvae of whirling beetle (Gyrinus sp) and water scavenger beetle
(Sternolophus sp) kill fish spawn and small fishes respectively.

CLASS CRUSTACEA
(PHYLUM ARTHROPODA)
Unlike the arachnids, the crustacean head, thorax and abdomen are well defined and the first pair
of the 6-20 joined appendages are ANTENNAE which perform SENSORY rather than feeding
functions. All crustaceans have two pairs of antennae as well as a pair of jointed appendages per
body segment. The crustacean body is enveloped in calcified cuticle of epidermal origin. In
addition to this, there may or may not be a hardened “coat”, shield or fold known as the
CARAPACE. When present, the carapace may “envelope” the whole crustacean enclosing its
head, trunk and limbs; or it may ’envelope’ the rest of the body without the head and one or two
appendages; it could simply arch over the head and thoracic segments leaving the abdominal
segments free; it could be fused to all the thoracic segments as is seen in the crabs or to some of
the thoracic segments leaving the other thoracic segments and the abdomen free. In shape, the
carapace may be horseshoe shaped, bivalve, roughly trapezoid, flat or slightly curved on top.

Nigerian crustacean fauna exist in 6 subclasses-

1. Branchiopoda. 2. Branchiura. 3. Cirripedia. 4. Copepoda.

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 5. Malacostraca. 6. Ostracoda.

SUBCLASS BRANCHIURA

(CLASS CRUSTACEA: PHYLUM ARTHROPODA)

1. DEFINITION
Parasitic crustaceans with indistinct body segmentation with lateral expansions of the head and a
pair of compound eyes. The other cephalic appendages include a pair each of small antennules
and antennae and a factorial mouth. The branchiurans are ectoparasites of fish.

2. TAXONOMY HISTORY
Information from the reviewed works on the taxonomic history of crustacean parasites of Africa
showed that branchiuran parasites of fish appeared to be very scanty although there is
information on parasitic copepods.

The fish lice (Argulus, lernae and Ergasilus) belonging to crustacean are important fish
ectoparasites.

3. CHECK-LIST
1. Argulus africanus
2. Argulus schoutedeni
3. Chonopeltis brevis
4. Chonopeltis minutes

4. DISTRIBUTION AND ECONOMIC IMPORTANCE

Species of Argulus are known by damage to fish tissues where they attach directly or
provide entrances for bacterial, fungal and viral infections of the fishes they
parasitise on.
They weaken fish by feeding on its blood. Argulus sp attach themselves to the fish
with their body buried in the scale pockets and with paired egg sacs protruding free.

SUBCLASS COPEPODA

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 (CLASS CRUSTACEA: PHYLUM ARTHROPODA)

These are crustaceans whose bodies are divided into an anterior METASOMA and a posterior
UROSOMA at a noticeable ARTICULATION 1/3-2/3 way along the body. The subclass
copepoda consists of crustaceans in seven orders: Calanoida, Caligoida, Cyclopoida,
Harpacticoida, Lernaeopodoida, Monstrilloida and Notodelphyoida. The common members are
in the orders Calanoida, Cyclopoida and Harpacticoida.

SUBCLASS BRANCHIOPODA

Branchiopods are crustaceans with/without a CARAPACE and with at least 4 (four) and at most
27 (twenty seven) pairs of jointed appendages. Living branchiopods belong to four orders-
Anostraca, the Notostraca and Cladocera. In Nigerian aquatic ecosystems they are represented
only by the Conchostraca and Cladocera, whose bodies end in a characteristic post abdomen.

Order Cladocera

Cladocerans are branchiopods with a laterally compressed carapace which encloses only the
trunk but not the head. The second antennae are enlarged and used for swimming. In Nigeria, the
cladoceran fauna consists of 100 species in seven families: - Bosminidae, Chydoridae,
Daphnidae, Macrothricidae, Monidae, Podonidae and Sididae.

Representative cladoceran species

1. Bosmina longirostris (Bosminidae)

2. Graptoleberis testudinaria (Chydoridae)
3. Illyocryptus spinifer (Macrothricidae)
4. Penilia avirostris (Sididae)

HARMFUL CRABS

Crabs make holes in the dikes and consequently there is leakage of water. Sometimes water
flowing through the holes can cause a complete collapse of dike. These holes also provide a
passage for the entry of predatory animals. The ‘cages’ made of nets are also damaged resulting
in escape of fishes from the cages.

PHYLUM ANNELIDA

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The annelids are metamerically segmented coelomate invertebrates bound by a soft and thin
cuticle through which segmentally arranged CHAETAE emerge. There are three main groups of
free living annelids in our water bodies: the leeches, oligochaetes and polychaetes.

Class Hirudinea (leeches)

Leeches are easily recognized by the anterior and posterior suckers with which they cling (“like
leeches”) to the bodies of persons who wade through freshwater bodies in swamps. They are
further recognized by the anterior region and the many-segmented body between the two
terminal suckers. Leeches are common in shallow backwaters, pools and ponds where they could
be found at the bottom or attached to vegetation or fish.

It infects major carps and feed on the blood of the host fish. It causes irritation and abnormal
movement of the hosts.

CLASS AVES (BIRDS)

Birds are found in the littoral, river banks, open water columns, on rocks or vegetational
outcrops or on aquatic vegetation in inland and coastal water bodies. Because they feed on
aquatic invertebrates and vertebrates including fish, they constitute an important component
of the ecosystem and their activities could result in a reduction in biodiversity. However,
increase in aquatic primary productivity as a result of fertilization by an avian feaces
probably compensates for the loss in fish mess consumed by the birds.

ENDANGERED SPECIES
Certain aquatic or shore birds cause a considerable damage to fishery waters and a
Pisciculturist must be aware about them. Examples of predatory birds are:

Family Common Name Scientific Name

Pelicanidae White or Rosy pelican Pelicanus onocrotalus

Phalacrocoracidae Large Cormorants Phatacrocorax carbo

Small Cormorants P. niger

Ardeidae Eastern Grey Heron Ardea cinerea rectirostris

Ciconidae Painted stock Mycteria lucocephalia

Accipitridae Fishing Eagle Ichthyophaga sp

Alcedinidae Kingfisher Ceryle sp

Alcedo sp.

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 The Pelicans, Cormorants, Fishing Eagle, Herons and kingfishers are the worst enemies. The
shallow waters provide ideal conditions for birds to prey on cultured species. Large flocks of
‘Cormorants’ can drive fish into shallow areas by flapping their wings and prey on them in
large number. According to an estimate, ten breeding pairs of Cormorants can catch about
4.5 tons of fish in a year, a ‘Pelican’ can consume between 1-3 tons in a year and a’ Heron’
may consume 100kg fish per year (causing 30-40% losses to fry and juvenile fish in a pond
farm).

CLASS AMPHIBIA

ENDANGERED SPECIES

Frogs and Toads have been reported to cause damage to the larvae and juveniles of fish. Besides,
aquatic snakes, crocodiles, alligators and large lizards also prey upon fish.

CLASS REPTILIA

ENDANGERED SPECIES

Aquatic snakes, crocodiles, alligators and large lizards prey upon fish.

CLASS MAMMALIA

These are vertebrates whose bodies are covered with HAIRS, unlike birds their eggs are
fertilized internally and the young develop within the body of the mammal. Most mammals are
terrestrial while a few are aquatic and amphibious. The mustelids (Family Mustelidae: Order
Carnivora), dolphins, whales (Order Cetacea ), sea lions, walruses, seals (Order Pinnipedia),
dugongs and manatees (Order Sirenia) are amphibious although only hippos and manatees have
been recorded in the aquatic habitats of Nigeria.

ENDANGERED SPECIES

The three species- the hippo and pigmy hippo (Cheropsis liberensis) and the manatee
(Trichechus senegalensis ) are endangered species in Nigeria.

‘Otters’ (Lutra sp and Aeonyx sp) are found to be the most destructive as they attack relatively
large fish at night.

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LITERATURE MATERIALS

A.M. Idi-ogede (2016): Fish Pathology in Perspective. Published by Boga Press.

38 Adelaja Street. Behind Uncle Joe Bus Stop. Makola-Ibadan, Oyo State, Nigeria.
Pp 178.

Atlas, R. M. (1995): Microorganisms in our

world. Mosby Book Inc. pp5 -13.

Dimmick, N.J. and Primrose, S. B (1987):

Introduction to modern virology. Blackwell scientific publications. Oxford
London. Edinburgh Boston Melbourne.

Ellis, A.E (1988): Fish vaccination: United

State. Edition Published by Academic press Inc.

Essentials of Pathology by Emmanuel Rubin.

Third edition

Fawole, M. O. and Oso, B.A (1998):

Laboratory, Manual of microbiology: Spectrums, Books, Limited. Ibadan.
Pp46.

G.N.O. Ezeri (2014): Fish Diseases: A major

hinderance to realization of “Fish for All”. 44th Inagural lecture. Federal
University of Agriculture, Abeokuta. Pp98.

-
Iwuji, S.C (2010): Basics and Application of General pharmacology. Owerri,
Nigeria: Milestone publisher

June, H and Juanita, W (2006): Manual of laboratory test. Published by A.I.T.B.S

India.

94
Kennath, A and Kathleen, V.A (1997): An easier and better way to learn
physiology: Princeton Review publishing L.L.C. Random. House. Inc. New
– York.
Kimberley, A. W(2004): Fin fish and shell fish Bacteriology manual: Techniques
and procedures. Iowa State, press, Blakwell publishing company.

Lon, E (1965): History of pathology. New York. Dover king.

Medigon, M.T., Martinko, J.M., Dunlap, P.V and clark, D.P (2009): Book Biology
of Microorganism: pearson Education Inc. 12th Edition Pp 2- 6.

Minica, C (2000): District laboratory practice in Tropical countries. Cambridge

University press. United Kingdom.

Nicolas, F.C (1983): Isolation and Identification of Fish bacterial pathogen.

Noga, E.J (2000): Fish Disease. Diagnosis and Treatment. Blackwell Publishing.
Iowa University, press edition, 2000- 367 pp.

Pelczar, M. J., Chan, E.C.S and Krieg, R.N (2001): microbiology: McGraw0hill,
5th edition pp 1 – 17.

Runnels, M and Monluse, D: Principles of Veterinary pathology (7th ed.) pp 6-21.

Salako, E.A (1994): Principles of General Mycology: Nigerian Educational

Research and Development Council. Vantage publishers limited, Aba
Samond, Oil Airport, Ibadan.

Shotts, E.B (1987): Bacterial diseases of fish associated with human health. Vet
clin. North America. Small animal practical 17(i): 241-7.

Snieszko, S. F (1974): The effects of environmental stress in outbreak of infectious

disease of fishes. J. fish boil. 6: 197 – 208.
Tortora, G.J., B.R. Funke and C.L (1982):
Microbiology- An introduction. Benjamin Cummings Publishing Company.
California. 726pp.

William, C. K (2001): “Paracelsus Dose Response” Handbook of pesticide

Toxicology. Academic press oct.

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Lecturer: Dr. Abubakar M. Idi-Ogede.

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