Article

Bioprospecting of Marine Organisms: Exploring Antibacterial

Activities in Aqueous and Organic Extracts
Vinícius Paulino Pinto Menezes 1 , Aldeni Moreira da Silva Filho 1 , Aline Jeferson Costa 1 , Elielton Nascimento 2 ,
Ulisses Santos Pinheiro 2 , Renata Pinheiro Chaves 1 , Alexandre Lopes Andrade 3 , Mayron Alves de Vasconcelos 3,4 ,
Edson Holanda Teixeira 3 , Alexandre Holanda Sampaio 1 , Celso Shiniti Nagano 1 and Rômulo Farias Carneiro 1, *

1 Laboratório de Biotecnologia Marinha—BioMar, Departamento de Engenharia de Pesca, Universidade

Federal do Ceará, Av. Humberto Monte, s/n, Campus do Pici, bloco 871, Fortaleza 60440-970, CE, Brazil;
[Link]@[Link] (V.P.P.M.); aldenibastos4@[Link] (A.M.d.S.F.); alinejcosta@[Link] (A.J.C.);
renatapinheirochaves@[Link] (R.P.C.); alexholandasampaio@[Link] (A.H.S.)
2 Departamento de Zoologia, Universidade Federal de Pernambuco, Av. Prof. Moraes Rego, 1235, Cidade
Universitária, Recife 50670-901, PE, Brazil; biologoefn@[Link] (E.N.); uspinheiro@[Link] (U.S.P.)
3 Laboratório Integrado de Biomoléculas—LIBS, Departamento de Patologia e Medicina Legal, Universidade
Federal do Ceará, Av. Monsenhor Furtado, s/n, Fortaleza 60430-160, CE, Brazil;
[Link]@[Link] (A.L.A.); mayronvasconcelos@[Link] (M.A.d.V.);
edsonlec@[Link] (E.H.T.)
4 Faculdade de Educação de Itapipoca, Universidade Estadual do Ceará, Av. da Universidade, s/n, Madalenas,
Itapipoca 62500-000, CE, Brazil
* Correspondence: romulofc2603@[Link]

Abstract: This study evaluated the antibacterial activity of aqueous and organic extracts
from 78 marine organisms, including seaweeds and sponges, collected along the coast of
Ceará, Brazil. Extracts were obtained by maceration using distilled water and 50% acetoni-
trile and tested against Staphylococcus aureus, Staphylococcus epidermidis (Gram-positive),
and Escherichia coli (Gram-negative) using the disk diffusion method, and inhibition zone
diameters were measured. Antibacterial activity was observed in 30.7% of the extracts, with
organic extracts showing higher efficacy. Several sponge species, particularly those from
Academic Editor: Haihua Liang the genus Aplysina, as well as Amphimedon compressa, Amphimedon viridis, Mycale sp., and
Received: 14 March 2025
Pseudosuberites sp., demonstrated notable inhibitory effects. While seaweed aqueous ex-
Revised: 12 April 2025 tracts showed no activity, some organic extracts—especially from Amansia multifida—were
Accepted: 15 April 2025 effective against Gram-positive strains. In general, Gram-positive bacteria were more
Published: 18 April 2025 susceptible than Gram-negative bacteria. These findings underscore the biotechnological
Citation: Menezes, V.P.P.; Silva Filho, potential of marine organisms from the Brazilian coast as promising sources of antibacterial
A.M.d.; Costa, A.J.; Nascimento, E.; compounds and support future efforts focused on the isolation, characterization, and toxi-
Pinheiro, U.S.; Chaves, R.P.; Andrade,
cological evaluation of active metabolites for pharmaceutical and industrial applications.
A.L.; Vasconcelos, M.A.d.; Teixeira,
E.H.; Sampaio, A.H.; et al.
Keywords: antibacterial activity; bioactive extracts; marine organisms
Bioprospecting of Marine Organisms:
Exploring Antibacterial Activities in
Aqueous and Organic Extracts.
Microorganisms 2025, 13, 940.
[Link] 1. Introduction
microorganisms13040940
Marine organisms are a prolific source of natural products, producing structurally
Copyright: © 2025 by the authors. diverse bioactive compounds shaped by environmental conditions distinct from those in
Licensee MDPI, Basel, Switzerland. terrestrial ecosystems. These compounds, typically derived from secondary metabolism, are
This article is an open access article
not directly involved in growth or maintenance but play key roles in mediating ecological
distributed under the terms and
interactions. Many of these molecules exhibit significant biological activities and have
conditions of the Creative Commons
Attribution (CC BY) license
demonstrated therapeutic potential against various pathogens [1–3].
([Link] Marine-derived compounds such as peptides, alkaloids, polyketides, and terpenes
licenses/by/4.0/). have attracted considerable attention due to their broad spectrum of bioactivity and chemi-

Microorganisms 2025, 13, 940 [Link]

Microorganisms 2025, 13, 940 2 of 12

cal diversity. In particular, antibacterial compounds from marine sources are of growing
interest as potential tools to combat antibiotic resistance [4,5]. Currently, 33 marine bioac-
tive compounds are at different stages of drug development—including preclinical and
clinical phases I–III—and 15 have been approved by regulatory agencies in the United
States, Australia, Japan, and China. Notably, the nucleoside Ara-A, isolated from a marine
sponge, has been approved for antiviral treatment [6,7]. Furthermore, 11 compounds in
the preclinical pipeline—primarily from the shikimate, peptide, polyketide, alkaloid, and
terpene classes—have shown antibacterial activity and are derived from marine sponges,
algae, bacteria, bryozoans, and soft corals [6,7].
The growing inefficacy of conventional antibiotics due to bacterial resistance poses
a major global health threat. Resistance may arise through the acquisition of resistance
genes or the occurrence of spontaneous mutations. The inappropriate or excessive use of
antibiotics accelerates this process, leading to treatment failures and an increased burden
on healthcare systems, along with higher mortality rates [8]. In this context, the discovery
of novel antibacterial molecules is critical to developing alternative therapeutic strategies.
Recent studies have explored the antibacterial properties of extracts from a wide range
of marine organisms, including invertebrates and macroalgae, targeting both common and
multidrug-resistant bacterial strains [9–13]. These investigations underscore the importance
of marine bioprospecting for the identification of secondary metabolites with biotechnolog-
ical potential. Among screening methodologies, the disk diffusion assay remains a widely
employed approach for the preliminary evaluation of antibacterial activity, owing to its
simplicity, low cost, and ability to generate rapid and semi-quantitative results. When using
this method, sterile paper disks impregnated with test extracts are placed on agar plates
inoculated with bacterial cultures. After incubation, the zones of inhibition surrounding the
disks are measured to assess antimicrobial potential against specific bacterial strains [14].
In this study, we investigated the antibacterial activity of aqueous and organic extracts
from marine organisms collected along the coast of Ceará, Northeastern Brazil. As part
of a broad bioprospecting effort, we aimed to identify extracts capable of inhibiting both
Gram-positive and Gram-negative bacteria. The results are expected to support future
efforts directed at achieving the isolation, structural characterization, and development of
novel bioactive compounds with potential applications in biotechnology and medicine.

2. Materials and Methods

2.1. Material Collection
Marine macroalgae and sponges were manually collected from the beaches of Pacheco
and Paracuru and by scuba diving at Parque da Pedra da Risca do Meio, along the coast
of Ceará, Brazil (Tables S1–S3). Fragments of the collected organisms were individually
placed in plastic tubes and kept cool in thermal boxes during transport to the Marine
Biotechnology Laboratory (BioMar-Lab), Department of Fisheries Engineering, Federal
University of Ceará. All collections and the use of biological material were authorized by
the Brazilian environmental regulatory systems SISBIO (Biodiversity Authorization and
Information System, ID: 33913-10, 33913-11) and SISGEN (National System for Genetic
Heritage and Associated Traditional Knowledge Management, ID: AC14AF9, A9D15EA,
A1792FE, AC71058, A625FEE, ACC97AD). Macrolgae were identified at the Department
of Fisheries Engineering, Federal University of Ceará, and sponges were identified at the
Department of Zoology, Federal University of Pernambuco.
Macroalgae were identified at the Department of Fisheries Engineering, Federal Uni-
versity of Ceará, and sponges were identified at the Department of Zoology, Federal
University of Pernambuco.
Microorganisms 2025, 13, 940 3 of 12

2.2. Aqueous Extraction

Different tissues from the marine organisms were macerated with or without liquid
nitrogen, depending on tissue consistency, and then homogenized in distilled water at a
ratio of 1:3 (w/v) for macroalgae and 1:2 for sponges. Homogenization was performed
using a refrigerated shaker (TH 6430B, Thoth Equipments, Piracicaba, Brazil) at 170 rpm
for 4 h at 25 ◦ C. The resulting extracts were centrifuged at 9000× g for 15 min, and the
supernatants were transferred to new tubes and stored at −20 ◦ C until further use.

2.3. Organic Extraction

Tissues were processed as described for aqueous extraction. The macerated material
was homogenized in 50% acetonitrile at the same ratios (1:3 for macroalgae and 1:2 for
sponges) and agitated at 170 rpm for 4 h at 25 ◦ C. The aliquots (2 mL) of the resulting
extracts were transferred to microtubes and concentrated by acetonitrile evaporation using
a vacuum concentrator (Labconco, Kansas City, MO, USA) at 35 ◦ C for 2 h. The concentrated
organic extracts were stored at −20 ◦ C until further analysis.

2.4. Bacteria and Culture Conditions

To assess antibacterial activity, Escherichia coli ATCC 11303, Staphylococcus aureus ATCC
25923, and Staphylococcus epidermidis ATCC 12228 were obtained from the microbial collec-
tion of the Integrated Biomolecules Laboratory (LIBS), Department of Fisheries Engineering,
Federal University of Ceará. Bacterial colonies were cultured on Petri dishes containing
Tryptic Soy Agar (TSA; Sigma-Aldrich, St. Louis, MO, USA) and then inoculated into tubes
with 15 mL of Tryptic Soy Broth (TSB; Sigma-Aldrich, MO, USA). Tubes were incubated
at 37 ◦ C for 18 h using an incubator (KM-CC17RU1A, Panasonic, Newark, NJ, USA) to
promote bacterial growth.

2.4.1. Antibiogram Assay

The bacterial concentration was adjusted to 2 × 108 colony-forming units (CFUs)·mL− 1
using a spectrophotometer at 620 nm based on calibration curves previously established
for each strain. Antibacterial activity was assessed using a modified disk diffusion method
based on Nugroho et al.’s protocol [12], with modifications. All samples and controls were
exposed to ultraviolet light for 15 min to ensure sterility. Sterile 6 mm paper disks (Laborclin,
Pinhais, Brazil) were placed on TSA plates previously seeded with bacterial suspensions,
and 10 µL of each extract was applied to the disks. Negative controls included 10 µL of
distilled water and 50% acetonitrile, while ampicillin (50 µg) served as a positive control.
Plates were incubated at 37 ◦ C for 20 h, after which inhibition zones were measured. The
presence of inhibition zones indicated antibacterial activity, while their absence suggested
bacterial resistance and a lack of activity in the tested extract.

2.4.2. Statistical Analysis

All experiments were conducted in triplicate in three independent runs. Data were
analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test
using GraphPad Prism software (version 9.0; San Diego, CA, USA). Statistical significance
was considered at p < 0.05.

3. Results
A total of 78 marine organisms were collected, of which the majority (65.4%) were
marine algae. Among these, red algae were the most prevalent, representing 56.9% of all
algae and 37.2% of the total specimens. Marine sponges comprised the remaining 34.6% of
the samples. Disk diffusion assays were performed on both aqueous and organic extracts,
Microorganisms 2025, 13, 940 4 of 12

with 10 aqueous extracts (12.8%) and 23 organic extracts (29.5%) exhibiting antibacterial
activity against at least one tested bacterial strain (Tables S4 and S5).
Among the aqueous extracts, 37% of sponge-derived samples showed antibacterial
activity, while none of the algal aqueous extracts were active. In contrast, organic extracts
displayed higher antibacterial potential, with 48.1% of sponge and 19.6% of algal extracts
showing inhibitory effects. Activity against the Gram-negative bacterium Escherichia coli
was observed in 10.2% of the tested species. In comparison, Gram-positive strains were
more frequently inhibited, with 24.4% of extracts effective against Staphylococcus aureus and
23.1% against Staphylococcus epidermidis.
Among the aqueous extracts, the most pronounced inhibition zones were observed
in the extract from Pseudosuberites sp., with diameters of 15.00 ± 1.00 mm (E. coli),
19.33 ± 2.31 mm (S. aureus), and 14.67 ± 2.31 mm (S. epidermidis) (Figure 1). Six sponge
extracts—Amphimedon compressa, Aplysina fistularis, Aplysina fulva, Aplysina lactuca, Mycale
sp., and Pseudosuberites sp.—were active against all three bacterial strains, representing 60%
of the active aqueous samples. In contrast, Agelas sventres displayed the lowest activity,
inhibiting only S. aureus (9.00 ± 1.00 mm). Tedania ignis and Topsentia ophiraphidites showed
no activity against E. coli. Examples of inhibition halos from aqueous extracts are shown in
Figure 2.
The genus Aplysina showed the highest number of positive results in both extraction
types, with inhibition zones ranging from 8.33 ± 0.58 to 23.00 ± 2.00 mm and consistent
activity against all of the tested bacterial strains (Figures 1 and 3). Other sponges, such as
Mycale sp. and Topsentia ophiraphidites, also exhibited notable activity, particularly in organic
extracts. Four sponge species—Aiolochroia crassa, Aplysina cauliformes, Erylus formosus, and
Ircinia felix—that were inactive in aqueous extractions showed activity in their organic
counterparts. Conversely, only Tedania ignis lost activity upon organic extraction, while the
other nine species maintained or increased their antibacterial performance.
None of the aqueous algal extracts exhibited antibacterial activity. However, 19.6% of
organic algal extracts were effective, primarily against Staphylococcus strains, with inhibition
zones ranging from 8.00 ± 0.00 to 12.00 ± 1.00 mm (Figure 3). A notable exception was
Amansia multifida, which produced a prominent inhibition zone of 24.00 ± 4.00 mm against
S. aureus. Only two algal species—Dictyota mertensii and Gracilariopsis sp.—exhibited
activity against both Staphylococcus strains, with inhibition zones ranging from 8.00 ± 0.00
to 9.33 ± 0.58 mm.
Organic extracts from Amansia multifida (24.00 ± 4.00 mm), Dictyota dichotoma
(8.00 ± 0.00 mm), Sargassum vulgare (11.00 ± 2.00 mm), and Valonia aegagropila
(12.00 ± 1.00 mm) were active exclusively against S. aureus. In contrast, extracts from
Corallina panizzoi (9.00 ± 0.00 mm), Cryptonemia crenulata (10.00 ± 0.00 mm), Gracilaria ramo-
sissima (8.67 ± 0.58 mm), and Laurencia sp. (9.00 ± 0.00 mm) were active only against
S. epidermidis. Among the macroalgae, 42.8% of brown algae exhibited antibacterial activity,
followed by 20.7% of red algae and only 6% of green algae.
 Microorganisms2025,
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Antibacterialactivity
[Link]
Figure activityofofaqueous
aqueousextracts
extractsfrom
frommarine
marineorganisms.
[Link]
Meaninhibition
inhibitionzone
zone
diameters (mm) include the disk diameter (6 mm). Disks were loaded with 10 µL of aqueous extract,
diameters (mm) include the disk diameter (6 mm). Disks were loaded with 10 µL of aqueous extract,
distilled water (negative control), or ampicillin (50 µg; positive control). Panels show results against
distilled water (negative control), or ampicillin (50 µg; positive control). Panels show results against
(A) Escherichia coli, (B) Staphylococcus aureus, and (C) Staphylococcus epidermidis. Different letters within
(A) Escherichia coli, (B) Staphylococcus aureus, and (C) Staphylococcus epidermidis. Different letters
each panel indicate statistically significant differences (p < 0.05).
within each panel indicate statistically significant differences (p < 0.05).
 Microorganisms2025,
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13,940
940 6 of 1312
6 of

Figure 2. Representative
Figure Representative results
results of
of disk
disk diffusion
diffusionassays
assaysfor foraqueous
aqueousextracts
extractsofofmarine
marineorganisms
organisms
against E. coli, S. aureus, and
against and S.
[Link].
[Link]
Panels(A–D):
(A–D):[Link];
coli;(E–H):
(E–H):[Link];
aureus; (I–L):
(I–L):S.S.
epidermidis.
epidermidis.
Red
Red arrows indicate inhibition
inhibition zones
zonesproduced
producedby byampicillin
ampicillin(50 (50µg); whitearrows
µg);white arrowsindicate
indicate inhibi-
inhibi-
tion
tion zones
zones produced by the extracts.
extracts. (A): Aplysinafistularis;
(A):Aplysina fistularis;(B): Aplysinafulva,
(B):Aplysina fulva,Pseudosuberites
Pseudosuberites sp.;
sp.;
(C): Amphimedon compressa; (D): ampicillin (50 µg); (E): Tedania ignis; (F): Ircinia felix;
(C): Amphimedon compressa; (D): ampicillin (50 µg); (E): Tedania ignis; (F): Ircinia felix; (G): Aplysina (G): Aplysina
fulva,
fulva, Mycale sp.,Topsentia
Mycalesp., Topsentiaophiraphidites;
ophiraphidites;(H):
(H):ampicillin
ampicillin (50(50
µg); Aplysina
(I):(I):
µg); fistularis;
Aplysina Tedania
(J): (J):
fistularis; ignis;
Tedania
(K): Aplysina lactuca; (L): Aplysina fulva, Mycale sp., Topsentia ophiraphidites, Pseudosuberites
ignis; (K): Aplysina lactuca; (L): Aplysina fulva, Mycale sp., Topsentia ophiraphidites, Pseudosuberites sp. sp.

Among the sponge-derived organic extracts, the strongest inhibition zones were
Among the sponge-derived organic extracts, the strongest inhibition zones were ob-
observed in Aplysina fulva (23.00 ± 2.00 mm) and Mycale sp. (22.67 ± 4.16 mm) against
served in Aplysina fulva (23.00 ± 2.00 mm) and Mycale sp. (22.67 ± 4.16 mm) against S. au-
S. aureus. Activity against E. coli was observed in extracts from Amphimedon compressa
reus. Activity against E. coli was observed in extracts from Amphimedon compressa and Am-
and Amphimedon viridis, species of the genus Aplysina, Mycale sp., and Pseudosuberites sp.,
phimedon viridis, species of the genus Aplysina, Mycale sp., and Pseudosuberites sp., with
with inhibition zones ranging from 9.00 ± 1.00 to 17.67 ± 0.58 mm. Five sponge extracts
inhibition zones ranging from 9.00 ± 1.00 to 17.67 ± 0.58 mm. Five sponge extracts (Am-
(Amphimedon compressa,
phimedon compressa, Amphimedon
Amphimedon viridis,
viridis, Aplysina
Aplysina fulva,
fulva, Aplysina
Aplysina lactuca,
lactuca, andand Mycale
Mycale sp.)
sp.)
inhibited all three bacterial strains, accounting for 21.7% of the active organic extracts
inhibited all three bacterial strains, accounting for 21.7% of the active organic extracts and and
38.5%
38.5% ofof the
the sponge
sponge samples
samplestested.
tested. Representative
Representativeresults
resultsfrom
fromthese
theseassays
assaysareareshown
shownin
Figure 4.
in Figure 4.
In
In total,
total, seven
seven sponge
sponge species
species exhibited
exhibitedantibacterial
antibacterialactivity
activityagainst
againstall
allthree
threebacterial
bacte-
strains tested, representing 9% of the organisms collected. Of these, four species
rial strains tested, representing 9% of the organisms collected. Of these, four species werewere
active
in bothinaqueous
active and organic
both aqueous extractions,
and organic corresponding
extractions, to 5% to
corresponding of 5%
theof
total
thesamples.
total samples.
Microorganisms2025,
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Figure Antibacterialactivity
Figure3.3. Antibacterial activityofoforganic
organicextracts
extractsfrom
from marine
marine organisms.
organisms. Mean
Meaninhibition
inhibitionzone
zone
diameters (mm) include the disk diameter (6 mm). Disks were loaded with 10 µL of
diameters (mm) include the disk diameter (6 mm). Disks were loaded with 10 µL of organic extract, organic extract,
distilledwater
distilled water(negative
(negativecontrol),
control),or orampicillin
ampicillin(50
(50µg; positivecontrol).
µg;positive control).Panels
Panelsshow
showresults
resultsagainst
against
(A) [Link],
(A)E. coli,(B)
(B)[Link],
aureus,and (C)[Link].
and(C) [Link]fferent
Differentletters
letterswithin
withineach
eachpanel
panelindicate
indicatestatistically
statistically
significant differences (p < 0.05).
significant differences (p < 0.05).
Microorganisms2025,
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Representativeresults
[Link]
Figure resultsofofdisk
diskdiffusion
diffusionassays
assaysfor
fororganic
organicextracts
extractsof ofmarine
marineorganisms
organisms
againstE.
against [Link],
coli,[Link],
aureus,and
[Link].
[Link] (A–E):[Link];
Panels(A–E): coli;(F–J):
(F–J):[Link];
aureus;(K,L):
(K,L):[Link].
epidermidis.
Redarrows
Red arrowsindicate
indicateinhibition
inhibitionzones
zonesfrom
fromampicillin
ampicillin(50(50µg); whitearrows
µg);white arrowsindicate
indicateinhibition
inhibitionzones
zones
from the extracts. (A): Aplysina fulva; (B): Mycale sp., Aplysina fistularis; (C): Aplysina cauliformes,
from the extracts. (A): Aplysina fulva; (B): Mycale sp., Aplysina fistularis; (C): Aplysina cauliformes, Pseu-
Pseudosuberites sp.; (D): ampicillin (50 µg); (E): Aplysina lactuca; (F): ampicillin (50 µg); (G): Amansia
dosuberites sp.; (D): ampicillin (50 µg); (E): Aplysina lactuca; (F): ampicillin (50 µg); (G): Amansia mul-
multifida, Dictyota mertensii, D. dichotoma, Sargassum vulgare, Gracilaria ramosissima, Valonia aegagropila,
tifida, Dictyota mertensii, D. dichotoma, Sargassum vulgare, Gracilaria ramosissima, Valonia aegagropila,
Aplysina fulva; (H): Topsentia ophiraphidites, Ircinia felix, Aplysina lactuca; (I): Aiolochroia crassa, Mycale
Aplysina fulva; (H): Topsentia ophiraphidites, Ircinia felix, Aplysina lactuca; (I): Aiolochroia crassa, Mycale
sp., Aplysina fistularis, A. cauliformes; (J): Aplysina fulva; (K): Mycale sp., Topsentia ophiraphidites, Aplysina
sp., Aplysina
lactuca; fistularis,fulva,
(L): Aplysina A. cauliformes; (J): Aplysina fulva; (K): Mycale sp., Topsentia ophiraphidites, Ap-
Erylus formosus.
lysina lactuca; (L): Aplysina fulva, Erylus formosus.
4. Discussion
4. Discussion
Bioprospecting marine organisms for antibacterial activity remains a critical strategy in
Bioprospecting
the search for novel marine
bioactive organisms
compounds for antibacterial activity remains
with biotechnological a critical potential.
and therapeutic strategy
Extraction
in the searchmethods
for novelplay a fundamental
bioactive compounds rolewith
in the recovery of such
biotechnological and compounds,
therapeuticranging
poten-
from
tial. simple aqueous
Extraction methodsextractions to organicrole
play a fundamental solvents with varying
in the recovery polarities
of such compounds,[9,11,13,15].
rang-
In the
ing frompresent
simple study, alongside
aqueous distilledtowater,
extractions organicwe solvents
employedwith 50% varying
acetonitrile due to
polarities
its compatibility
[9,11,13,15]. In the with both
present polaralongside
study, and non-polar
distilledsubstances, as well as 50%
water, we employed its documented
acetonitrile
effectiveness
due in reversed-phase
to its compatibility with both HPLCpolarforand
biomolecule
non-polarrecovery
substances,[16–18]. Thisaschoice
as well was
its docu-
intendedeffectiveness
mented to support future fractionation and
in reversed-phase HPLC compound identification
for biomolecule efforts.
recovery [16–18]. This
choiceOur wasfindings
intended revealed
to supportthatfuture
30.7% fractionation
of the tested marine organisms
and compound exhibited antibacte-
identification efforts.
rial activity, particularly
Our findings revealed inthat
organic
30.7% extracts. Whilemarine
of the tested aqueous extracts from
organisms seaweeds
exhibited were
antibacte-
inactive,
rial sponge
activity, extracts in
particularly demonstrated activity
organic extracts. across
While both solvent
aqueous extractstypes.
fromThese results
seaweeds cor-
were
roborate sponge
inactive, previous reportsdemonstrated
extracts [13,15,19,20] and underscore
activity across the
both relevance of solvent
solvent types. polarity
These in
results
recovering lipophilic
corroborate secondary
previous reports metabolites,
[13,15,19,20] andmany of which
underscore theare linked toofantimicrobial
relevance solvent polar- ef-
fects.
ity For instance,
in recovering although
lipophilic Haliclona metabolites,
secondary species showed many activity in studies
of which usingtomethanol
are linked antimicro- or
dichloromethane
bial [21–25],although
effects. For instance, our acetonitrile-based
Haliclona species extraction
showeddid not yield
activity activeusing
in studies compounds
meth-
fromor
anol these sponges, highlighting
dichloromethane [21–25], theourinfluence of solvent extraction
acetonitrile-based choice. did not yield active
Geographical
compounds from thesevariation
sponges,mayhighlighting
also influence themetabolite
influence profiles.
of solvent For example, Aplysina
choice.
fulvaGeographical
and Amphimedon viridis
variation were
may alsopreviously reported asprofiles.
influence metabolite inactiveForwhen extracted
example, with
Aplysina
acetone
fulva and[26] but showedviridis
Amphimedon activity in our
were acetonitrile
previously extracts.
reported as Similarly, the aqueous
inactive when extract
extracted of
with
acetone [26] but showed activity in our acetonitrile extracts. Similarly, the aqueous extract
Microorganisms 2025, 13, 940 9 of 12

Ircinia felix exhibited Gram-positive activity in our study. These differences may reflect the
environmental or ecological factors that modulate secondary metabolite production [16,27].
Among all of the tested taxa, sponges from the genus Aplysina demonstrated the
most consistent and potent antibacterial activity, in agreement with previous studies [28].
Other sponge genera, including Agelas, Amphimedon, and Mycale, also showed significant
activity, particularly in organic extracts. This aligns with prior reports highlighting their
alkaloid-rich profiles and broad-spectrum bioactivity [29–33]. In contrast, Pseudosuberites
sp. displayed higher activity in aqueous extracts, reinforcing the importance of solvent
compatibility and species-specific metabolite solubility [34].
Notably, extracts from Topsentia ophiraphidites and Tedania ignis were inactive against
E. coli, consistent with some previous findings but contrasting others [21,34,35]. These
differences further underscore the influence of extraction methods and solvents on antibac-
terial efficacy.
Regarding macroalgae, organic extracts were more effective, particularly against Gram-
positive bacteria. Amansia multifida organic extract showed the most pronounced activity,
corroborating earlier findings using hexane-based extractions [36]. Extracts from Dictyota
spp. and Sargassum spp. also showed inhibitory effects, which have been attributed to sterol
content and other lipophilic compounds [37,38]. In contrast, several green and red algae,
including Ulva fasciata, Caulerpa prolifera, and Gracilaria domingensis, lacked antibacterial
activity in our study, supporting prior reports [39].
Overall, our results reinforce the potential of marine organisms—particularly
sponges—as valuable sources of antibacterial compounds. Several extracts demonstrated
significant activity, especially those obtained with organic solvents, highlighting the in-
fluence of solvent polarity on the extraction of bioactive metabolites [32–35,40,41]. The
genus Aplysina stood out as a particularly promising candidate, along with the species of
Amphimedon, Agelas, and Mycale, all of which yielded broad-spectrum antibacterial activity.
A general trend observed across our assays was the greater susceptibility of Gram-
positive bacteria to the tested extracts. This can likely be attributed to structural differences
in the bacterial cell envelope: the simpler peptidoglycan-rich wall of Gram-positive bacteria
may facilitate the entry of bioactive compounds, whereas the outer membrane of Gram-
negative bacteria acts as a permeability barrier that limits compound access [42–44].
These findings are especially relevant in light of the escalating public health
threat posed by antimicrobial resistance, which has been linked to millions of deaths
globally each year [45]. The identification of new antibacterial sources from marine
biodiversity—particularly from underexplored tropical regions—contributes meaningfully
to the global effort to discover alternative therapeutic agents.
While this study was designed as an initial screening effort, it provides essential
insights into the antibacterial potential of marine organisms and lays the groundwork for
future investigations. The use of disk diffusion assays allowed for the rapid assessment of
a large number of extracts, enabling the identification of promising candidates for a more
detailed analysis. Building on these results, future studies will aim to isolate and chemically
characterize the active compounds, assess their stability and potential cytotoxicity, and
explore their mechanisms of action. Such efforts will be instrumental in advancing these
findings toward practical applications in biotechnology and antimicrobial therapy.

5. Conclusions
This study highlights the biotechnological potential of marine organisms from the
northeastern coast of Brazil as sources of antibacterial compounds. Approximately 30.7%
of the tested extracts exhibited inhibitory activity, with organic extracts demonstrating
greater efficiency. Sponges from the genera Aplysina and Amphimedon, as well as the species
Microorganisms 2025, 13, 940 10 of 12

Mycale sp. and Pseudosuberites sp., were the most promising, while Amansia multifida
was the most notable among marine algae, showing a significant inhibition halo against
Staphylococcus aureus.

Supplementary Materials: The following supporting information can be downloaded at: https://
[Link]/article/10.3390/microorganisms13040940/s1, Table S1—Marine organisms collected
from Parque da Pedra da Risca do Meio, Ceará. Table S2—Marine organisms collected from Pacheco
Beach, Ceará. Table S3—Marine organisms collected from Paracuru Beach, Ceará. Table S4—Results
of the disk diffusion antibiogram assay for aqueous extracts of marine organisms—average inhibition
zone diameter (mm). Table S5—Results of the disk diffusion antibiogram assay for organic extracts of
marine organisms—average inhibition zone diameter (mm).

Author Contributions: Conceptualization, R.P.C., C.S.N. and R.F.C.; Methodology and Investiga-
tion, V.P.P.M., A.M.d.S.F. and A.J.C.; Project Administration, A.L.A. and R.P.C.; Validation, E.N.,
U.S.P. and A.H.S.; Resources and Funding Acquisition, C.S.N., R.F.C., A.H.S., M.A.d.V. and E.H.T.;
Writing—Original Draft Preparation, V.P.P.M. and R.P.C.; Writing—Review and Editing, R.F.C.; Super-
vision, R.P.C. and R.F.C. All authors have read and agreed to the published version of the manuscript.

Funding: This research received no external funding.

Institutional Review Board Statement: All collections and use of biological material were authorized
and certified by the competent environmental institutions SISBIO (Biodiversity Authorization and
Information System, ID: 33913-10, 33913-11, approve date 1 July 2023) and SISGEN (National System
for Genetic Heritage and Associated Traditional Knowledge Management, ID: AC14AF9, A9D15EA,
A1792FE, AC71058, A625FEE, ACC97AD, approve date: 4 May 2021).

Informed Consent Statement: Not applicable.

Data Availability Statement: No new data were created or analyzed in this study. Data sharing is
not applicable to this article.

Acknowledgments: This work was supported by the Brazilian agencies CNPq (Conselho Nacional de
Desenvolvimento Científico e Tecnológico), CAPES (Fundação Coordenação de Aperfeiçoamento de
Pessoal de Nível Superior), FUNCAP (Fundação Cearense de Apoio ao Desenvolvimento Científico e
Tecnológico), and FINEP (Financiadora de Estudos e Projetos). AHS, CSN, EHT, and RFC are Senior
Investigators of CNPq.

Conflicts of Interest: The authors declare no conflicts of interest.

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