Antifouling activity of isonitrosoacetanilides against microfouling and macrofouling.

Biofouling is responsible for structural and economic damage to man-made surfaces. Antifouling paints with biocides have been applied to structures to avoid organism adhesion; however, they have high toxicity and are not able to prevent all biofouling processes, necessitating the periodic mechanical removal of organisms and paint reapplication. Thus, there is an urgent demand for novel, effective, and environmentally friendly antifouling alternatives. As isonitrosoacetanilide is the precursor for many compounds with antibacterial activity, we believe that it could have antifouling activity against microfouling and, consequently, against macrofouling. The aim of this work was to investigate the antifouling potential of six isonitrosoacetanilide compounds and their toxicity. The compounds were employed at different concentrations (0.625-1.25-2.5-5-10 µg mL-1) in this study. The biofilm and planktonic bacteria inhibition and biofilm eradication potential were evaluated by crystal violet assay, while Amphibalus amphitrite barnacle settlement was evaluated by cyprid settlement assay. Toxicity evaluation (LC50 and EC50) was performed with A. amphitrite nauplii II and cyprid larvae. At least one of the tested concentrations of 4-Br-INA, 4-CH3-INA, and 2-Br-INA compounds showed nontoxic antifouling activity against microfouling (antibiofilm) and macrofouling (antisettlement). However, only 4-CH3-INA and 2-Br-INA also showed biofilm eradication potential. These compounds with antibiofilm activity and nontoxic effects could be combined with acrylic base paint resin or added directly into commercial paints in place of toxicant biocides to cover artificial structures as friendly antifouling agents.

Biofouling initial succession on offshore artificial substrate under subtropical conditions.

This study presents the initial stages of the macrofouling community on artificial substrate exposed to the offshore subtropical marine environment, and the contribution of depth (3 and 22m), exposure time (1-2-4-7-10-13-weeks), UV-radiation, rainfall, temperature, pH, salinity, water chlorophyll-a, and zooplankton supply to organism establishment. Steel substrates were placed horizontally on the structure of a pipeline monobuoy off the southern shore of Brazil (Tramandaí beach), and the ecological succession was monitored by six random removals per depth during the summer-autumn of 2011. Approximately 88.5% of the quantified settled individuals comprised fouling fauna and 11.5% vagile and sedentary fauna, although the taxa richness was higher for non-sessile invertebrates. Species richness and organism density up to four weeks were significantly higher at 3m-depth. After this period, a higher density of organisms was found at 22m, while during the whole study the species richness and diversity remained higher at 3m-depth. Zooplankton composition did not show a simultaneous temporal relationship with invertebrate recruitment at any depth; however, increasing the exposure time, the similarity between the planktonic and benthic communities also increased. Meroplankton, tychoplankton, and holoplankton were recorded on the substrates. This study showed that the depth of available substrates affects the macrofouling establishment, which is mainly associated with UV-radiation, exposure time, and ecological interspecific interactions.

Salt marshes as the final watershed fate for meso- and microplastic contamination: A case study from Southern Brazil.

Plastics pose a major threat to aquatic ecosystems especially in smaller size fractions. Salt marshes play a crucial role in maintaining the coastal zone and aquatic food web, yet their contamination, including by plastic materials, is still poorly investigated. This work investigated meso- (MEP, 5-25 mm) and microplastic (MIP, 1 µm-5 mm) contamination of a salt marsh, which reached average levels of 279.63 ± 410.12 items kg-1, 366.92 ± 975.18 items kg-1, and 8.89 ± 8.75 items L-1 in surface sediment, sediment cores and water, respectively. Photomicrographs revealed a complex fouling community on plastics surface for both different salt marsh zones and plastic formats. Abundance of plastics in sediment was higher in the dryer, vegetated zones compared to flooded, unvegetated zones. This is consistent with the role of vegetation as a trap for solid litter and final fate of plastic deposition, but also with local hydrodynamics influencing deposition pattern. Plastics were detected up to 66 cm-depth, presenting higher levels at surface sediments. It was also possible to identify the main groups of microorganisms (1638 bacterial cells, 318 microalgae cells, and 20049.93 µm2 of filamentous fungi) composing the Plastisphere communities on all plastic items recorded in the different zones. These results are a pioneer contribution, highlighting that regional salt marshes participate in sequestration and longstanding accumulation of plastic particles in estuarine environments, before exportation to the ocean.

Litter contamination at a salt marsh: An ecological niche for biofouling in South Brazil.

The presence of solid litter and its consequences for coastal ecosystems is now being investigated around the world. Different types of material can be discarded in areas such as salt marshes, and various fouling organisms can associate with such items forming the Plastisphere. This study investigated the distribution of solid litter along zones (dry, middle, flooded) of a salt marsh environment in the Patos Lagoon Estuary (South Brazil) and the association of biofouling organisms with these items. Solid litter quantities were significantly higher in the dry zone when compared to the middle and flooded zones, showing an accumulation area where the water rarely reaches. Most items were made of plastic, as shown for many other coastal areas, and originated from food packaging, fishery and shipping activities and personal use. Although not statistically significant, there was a tendency of increased biofouling towards the flooded zone. Thirteen groups were found in association with solid litter items, mainly algae, amphipods, and gastropods. The preference for salt marsh zones, types of material and items' colour was highly variable among groups of organisms, which can be related to their varied physiological requirements. In summary, significant plastic contamination of salt marshes of the Patos Lagoon was associated with a heterogeneous distribution of fouling communities.

Acartia tonsa Dana 1849 as a Model Organism: Considerations on Acclimation in Ecotoxicological Assays.

The copepod Acartia tonsa was standardized as model organism in acute toxicity bioassays due to its key position in coastal food chains and high sensitivity. Once bioassays are performed according to a protocol their results may become tools for the protection of aquatic ecosystems. However, there are divergences in bioassays methods using A. tonsa. This study aimed to investigate: (i) the need for acclimation of A. tonsa collected from the environment for use in acute toxicological bioassays; and (ii) differences in sensitivity between copepods collected from the environment and laboratory-grown copepods. Laboratory-grown copepods are more sensitive to SDS than A. tonsa from the environment. The acclimation time of 30 h helped organisms to recover from stress of collection/handling and changing environment/conditions. Therefore, laboratory-grown copepods showed to be more sensitive than organisms from environment; and for ecotoxicological bioassays acclimating A. tonsa collected from the environment for 30 h can be adopted.

Surface coatings select their micro and macrofouling communities differently on steel.

Previous studies have shown the effect of surface coatings on biofouling; however, they did not take into account the interaction of the micro and macrofouling communities, the effect of substrate orientation and the zooplankton-zoobenthic coupling together. Therefore, the aim of this study was to evaluate the effect of Zn- and Cu2O-based coatings on micro and macrofouling on steel surfaces, while also observing the role of substrate orientation and zooplankton supply. An experiment was carried out in the Patos Lagoon Estuary in southern Brazil for three months between spring and summer, where ASTM-36 steel plates represented different coatings (Zn- and/or Cu2O-based) and orientations (vertical and horizontal). To assess the zooplankton supply, sampling was carried out weekly using a 200 µm plankton net. Zn-based coating positively affected microfouling density compared to uncoated surfaces. The same pattern was observed with macrofouling, associated with vagile fauna preference, which represented 70% of the settled macrofoulers. Cu2O-based antifouling painted surfaces showed the highest microfouling density inhibition, while Zn + Cu2O-based coating did not affect the bacteria adhesion but showed lower density compared to Zn-based coating alone. The coatings combination showed the highest invertebrate inhibition. In this way, the macrofouling community was more sensitive than microfouling was to the antifouling coatings tested. The substrate orientation only affected macrofouling, horizontal surfaces being more attractive than vertical. Meroplankton, tychoplankton and holoplankton were recorded on the surfaces, although their representation in plankton was not proportional to the recruits recorded on the substrates. This was probably due to fast dispersion, the interactions of other factors and/or ecological succession stage. Surface coating, substrate orientation, and zooplankton supply interacted with the biofouling process on steel in different ways depending on the organism evaluated. Therefore, copper oxide- and zinc-based coatings were not suitable as coatings to avoid the total biofouling establishment.

Natural and non-toxic products from Fabaceae Brazilian plants as a replacement for traditional antifouling biocides: an inhibition potential against initial biofouling.

In this study, we screened for the antifouling activity of 15 species plant extracts from Brazilian the Brazilian Caatinga Fabaceae against the initial colonization of natural marine bacterial biofilm. We also investigated the potential toxicity of extracts against planktonic and benthic non-target organisms. Aqueous extracts of plants collected in the Caatinga biome (PE, Brazil) were prepared and tested at different concentration levels (0, 0.5, 1, 2, 4, and 8 mg mL-1). Natural marine bacterial consortium was inoculated in multi-well plates and incubated with the different treatments for 48 h. The biofilm and planktonic bacterial density and biomass inhibition were evaluated along with biofilm biomass eradication. The extracts that showed the highest bacterial biofilm inhibition were evaluated for toxicity against microalgae and crustaceans. The biofilm and planktonic bacterial inhibition potential were evaluated through flow cytometry and spectrophotometry. The selected treatments were evaluated for their toxicity using the microalgae Chaetoceros calcitrans, the copepod Nitokra sp., and the brine shrimp Artemia salina as bioindicators. Our work demonstrates the biotechnological potential of Fabaceae plant compounds as a safe antifouling alternative. Anadenanthera colubrina var. cebil fruits and Apuleia leiocarpa leaf extracts showed antibiofilm activity (≥ 80%), while Myroxylon peruiferum and Dioclea grandiflora leaf extracts showed antibiotic activity. These extracts were safe to planktonic and benthic non-target organisms. The results of this study point to potential substitutes to highly toxic antifouling paints and shed light on the prospect of a yet to be explored biome for more sustainable alternatives in biofouling research.

Euphausiacea diversity in a trans-oceanic transect through the South Atlantic Ocean: the first Atlantic record of Thysanopoda astylata Brinton, 1975.

Information about euphausiids in central South Atlantic Ocean is scarce; hence, we investigated species composition and distribution of euphausiids along a longitudinal transect in this region, with an emphasis on Thysanopoda. Zooplankton samples were collected from 44 stations during the first Transatlantic Commission (Brazil-Africa). Euphausiids comprised 21,390 individuals across larval stages (nauplius, calyptopis, and furcilia) and adults. Furcilia and adults were classified to species level when possible, with a total of 19 identified species. Overall, Euphausia species frequency of occurrence and abundance were highest in samples collected near the African coast, while Thysanopada species dominated near the Brazilian coast. Of the euphausiids caught, 158 were identified as Thysanopoda, including 2 specimens of T. astylata, 6 T. aequalis, 3 T. pectinata, 2 T. monacantha, 2 T. tricuspida, and 1 T. egregia; 118 damaged specimens could only be identified as Thysanopoda spp., and 24 as T. aequalis / T. astylata complex because of the lack of diagnostic structures. Thysanopada egregia was present in samples collected down to 96 m, which increases the vertical range for this species. This report constitutes the first record of Thysanopoda astylata Brinton, 1975 in Atlantic waters.

Could some procedures commonly used in bioassays with the copepod Acartia tonsa Dana 1849 distort results?

Many organizations have suggested the use of the Calanoid copepod Acartia tonsa in protocols for acute toxicity tests. Nevertheless, these protocols present some problems, such as using 60-180µm meshes to separate specific stages of A. tonsa or carrying out the tests using small volumes that reflect high densities of A. tonsa that do not occur in nature, which could lead to distorted results. In addition, ecotoxicological studies may use statistical approaches that are inadequate for the type of data being analysed. For these reasons, some methodological approaches for bioassays using A. tonsa need to be clarified and revised. In this study, we present information about (i) the retention of copepodite stages of A. tonsa on 180, 330 and 500µm net meshes; (ii) tested storage volumes of 1 organism per 5, 10 or 20mL in each test container (TC); and (iii) considerations about the statistics employed. The results demonstrated that a net mesh of 180µm is capable of retaining all copepodite stages (CI to CVI), contrasting with the recommendation of using a 180µm mesh to separate out adults only. Coarser meshes (330 and 500µm) can also retain different proportions of all copepodite stages, but cannot separate out one developmental stage only. Twenty-five millilitres of medium in an open TC, commonly employed in bioassays simulating densities of 1 organism 5mL-1, completely evaporated, and the results showed that the TCs need to be covered (e.g., PVC film) and filled with a minimum of 100mL of culture medium (simulating densities of 1 organism 20mL-1) to avoid evaporation and increases in salinity. The current use of ANOVA in ecotoxicological studies with proportions of surviving organisms should also be reconsidered since the data are discrete and have a binomial distribution; general linear models (GLMs) are considered more adequate. The information presented here suggests some adjustments that hopefully will enable the improvement of the procedures and methods employed in studies of acute toxicity using the copepod A. tonsa.

What determines sclerobiont colonization on marine mollusk shells?

Empty mollusk shells may act as colonization surfaces for sclerobionts depending on the physical, chemical, and biological attributes of the shells. However, the main factors that can affect the establishment of an organism on hard substrates and the colonization patterns on modern and time-averaged shells remain unclear. Using experimental and field approaches, we compared sclerobiont (i.e., bacteria and invertebrate) colonization patterns on the exposed shells (internal and external sides) of three bivalve species (Anadara brasiliana, Mactra isabelleana, and Amarilladesma mactroides) with different external shell textures. In addition, we evaluated the influence of the host characteristics (mode of life, body size, color alteration, external and internal ornamentation and mineralogy) of sclerobionts on dead mollusk shells (bivalve and gastropod) collected from the Southern Brazilian coast. Finally, we compared field observations with experiments to evaluate how the biological signs of the present-day invertebrate settlements are preserved in molluscan death assemblages (incipient fossil record) in a subtropical shallow coastal setting. The results enhance our understanding of sclerobiont colonization over modern and paleoecology perspectives. The data suggest that sclerobiont settlement is enhanced by (i) high(er) biofilm bacteria density, which is more attracted to surfaces with high ornamentation; (ii) heterogeneous internal and external shell surface; (iii) shallow infaunal or attached epifaunal life modes; (iv) colorful or post-mortem oxidized shell surfaces; (v) shell size (<50 mm2 or >1,351 mm2); and (vi) calcitic mineralogy. Although the biofilm bacteria density, shell size, and texture are considered the most important factors, the effects of other covarying attributes should also be considered. We observed a similar pattern of sclerobiont colonization frequency over modern and paleoecology perspectives, with an increase of invertebrates occurring on textured bivalve shells. This study demonstrates how bacterial biofilms may influence sclerobiont colonization on biological hosts (mollusks), and shows how ecological relationships in marine organisms may be relevant for interpreting the fossil record of sclerobionts.

Evaluation of antibiotics as a methodological procedure to inhibit free-living and biofilm bacteria in marine zooplankton culture.

There is a problem with keeping culture medium completely or partially free from bacteria. The use of prokaryotic metabolic inhibitors, such as antibiotics, is suggested as an alternative solution, although such substances should not harm non-target organisms. Thus, the aim of this study was to assess the effectiveness of antibiotic treatments in inhibiting free-living and biofilm bacteria and their half-life in artificial marine environment using the copepod Acartia tonsa as bioindicador of non-harmful antibiotic combinations. Regarding to results, the application of 0.025 g L-1 penicillin G potassium + 0.08 g L-1 streptomycin sulphate + 0.04 g L-1 neomycin sulphate showed great potential for use in marine cultures and scientific experiments without lethal effects to non-target organisms. The effect of this combination starts within the first six hours of exposure and reduces up to 93 % the bacterial density, but the half-life is short, requiring replacement. No adverse changes in water quality were observed within 168 hours of exposure. As a conclusion, we can infer that this treatment was an effective procedure for zooplankton cultures and scientific experiments with the aim of measuring the role of free-living and biofilm in the marine community.