Design of an agro-industrial by-products-based media for the production of probiotic bacteria for fish nutrition.

Probiotic production in commercial culture media is expensive, so, it is necessary to design culture media based on "low-cost" components like agro-industrial by-products. Therefore, this study aimed to design an agro-industrial by-product-based culture media using whey, sugarcane molasses, and palm kernel cake as components to produce Lactococcus lactis A12, Priestia megaterium M4, and Priestia sp. M10 isolated from Nile tilapia (Oreochromis niloticus) associated gut microbiota. Higher bacterial concentrations were achieved at high whey concentrations and low concentrations of sugarcane molasses and palm kernel cake (PKC) using agitation. The optimal conditions were whey, 3.84% w/v; sugarcane molasses, 7.39% w/v; PKC, 0.77% w/v; and agitation speed, 75 RPM. Bacterial growth under optimal conditions was compared to that in commercial Brain-Heart Infusion (BHI) broth. L. lactis A12 showed similar growth in the optimal media and BHI. The estimated cost of the culture media based on component prices was USD $ 3.01/L, which is 86.93% lower than BHI broth (USD $ 23.04/L). It was possible to design a "low-cost agro-industrial by-product-based culture media to produce L. lactis A12 and the two Priestia species under monoculture conditions.

Assessment of Bacillus species capacity to protect Nile tilapia from A. hydrophila infection and improve growth performance.

The present study evaluated the capacity of three Bacillus species to improve health status and growth performance of Nile Tilapia fed with high levels of soybean meal and challenged with Aeromonas hydrophila. In vitro experiments showed that ß-hemolysin and metalloprotease enzymes were produced by A. hydrophila throughout the exponential growth phase. In vivo experiments showed that 107 colony-forming units (CFUs)/ml of this pathogen killed 50% of control group fishes in 13 days. To evaluate the influence of Bacillus strains on health status and growth performance in Nile Tilapia, 180 fishes (33.44 + 0.05 g) were distributed in 12 tanks of 200 L each, and animals were fed twice per day until satiety. 1) Control group without Bacillus, 2) Bacillus sp1, 3) Bacillus sp2, and 4) Bacillus sp3 groups were formulated containing 106 CFU/g. After 40 days of feeding, the fishes were intraperitoneally injected with 1 ml of A. hydrophila at 2 × 107 CFU/ml, and mortality was recorded. The results showed that cumulative mortality rate was significantly (p< 0.05) lower in the Bacillus sp1 (25%), sp2 (5%), and sp3 (15%) groups, than the control group (50%). Weight gain was also significantly better (p< 0.05) in the Bacillus sp1 (36%), sp2 (67%), and sp3 (55%) groups with respect to the control group (30%). In conclusion, functional diet formulated with high levels of soybean meal and supplemented with Bacillus sp2 could be an alternative to protect Nile tilapia cultures from A. hydrophila infections and improve fish growth performance.

Co-infection of Lactococcus garvieae and Aeromonas hydrophila in cultured Nile Tilapia in Kerala, India.

Co-infection of Lactococcus garvieae and Aeromonas hydrophila, has been confirmed from diseased Nile Tilapia (Oreochromis niloticus), Chithralada strain cultured in a freshwater rearing pond of Alappuzha district of Kerala, India. The aetiological agents behind the disease outbreak were bacteriologically proven and confirmed by 16SrRNA sequencing and phylogenetic analysis. PCR detection of the virulent genes, showed existence of adhesin and hemolysin in L. garvieae and aerolysin in A. hydrophila strain obtained. To fulfil Koch's postulates, challenge experiments were conducted and median lethal dose (LD50) of L. garvieae and A. hydrophila was calculated as 1 × 105.91 CFU per mL and 1 × 105.2 CFU per mL respectively. Histopathologically, eyes, spleen, and kidney were the predominantly infected organs by L. garvieae and A. hydrophila. Out of the 13 antibiotics tested to check antibiotic susceptibility, L. garvieae showed resistance to almost 7 antibiotics tested, with a resistance to Ciprofloxacin while A. hydrophila was found resistant to Streptomycin and Erythromycin. Understanding the complex interaction between Gram-positive and Gram-negative bacteria in the disease process and pathogenesis in fish host will contribute to efficient treatment strategies. As a preliminary investigation into this complex interaction, the present study is aimed at phenotypic and genotypic characterization, pathogenicity evaluation, and antibiotic susceptibility of the co-infecting pathogens in a diseased sample of freshwater-farmed Nile tilapia.

Effects of a multi-strain Bacillus probiotic on the intestinal microbiome, haemato-immunology, and growth performance of Nile tilapia.

The study evaluated dietary supplementation with a feed additive composed of multi-strain Bacillus for Nile tilapia Oreochromis niloticus. In vitro and in vivo assays employing culture-based microbiological methods and metagenomics were performed. Additionally, the study assessed the haemato-immunology, intestinal microbiome, and growth performance of the animals. For this, 30 juvenile Nile tilapia were used in the in vitro assay and 180 (60 + 120) in the in vivo assays. In the in vitro assay, we found evidence of adhesion of the probiotic bacteria to the intestinal mucus of fish, corroborated in the 15-day in vivo assay, in which the count of B. licheniformis was significantly higher in fish fed with probiotic when compared to fish of the control group. Furthermore, in the 50-day in vivo trial, a metagenomic analysis provided evidence for the modulation of the intestine microbiome of Nile tilapia by dietary supplementation of the probiotic. In addition, there was an increase in species richness, higher abundance of potentially probiotic autochthonous species and a lower abundance of Aeromonas sp. when the animals were fed the supplemented diet. Finally, no significant differences were observed in growth performance and haemato-immunological analyses, suggesting no harm to fish health when the product was supplemented for 15 and 50 days. The in vitro results indicate that the multi-strain probiotics were able to adhere to the intestinal mucus of Nile tilapia. Additionally, a modulation of the intestinal microbiome was evidenced in the in vivo assay.

Pathogenic bacteria prevalence in cultured Nile tilapia in Southwest Mexico: A real-time PCR analysis.

The present study investigates molecular-based PCR techniques to estimate the prevalence of fish pathogens in southwest Mexico where recurrent mortality in the tilapia cultures has been observed. Sample of internal organs and lesions of Nile tilapia were taken and analysed in 2018, 2019, 2020 and 2022 to detect bacterial pathogens using PCR. No samples were taken in 2021 due to the COVID-19 pandemic. The real-time PCR conditions were optimized to allow a qualitative reliable detection of the bacteria from fixed fish tissue. A total of 599 pond- and cage-cultured tilapia from the southwestern Mexican Pacific (Guerrero, Oaxaca and Chiapas states) were analysed. In this tropical region, during 2018 and 2019 water temperatures of the tilapia cultures were generally with the optimal range to grow Nile tilapia, although extreme values were recorded on some farms. Most of the tilapia sampled were apparently healthy. No Francisella sp. was detected in any sample, and Staphylococcus sp. was the most prevalent (from 0% to 64%) bacteria from the three states over time. Low prevalence of Aeromonas sp. was found, from 0% to 4.3%, although the fish pathogen Aeromonas dhakensis was not detected. Sterptococcus iniae was only detected in Chiapas in 2019 at a low prevalence (1.4%), while the major tilapia pathogen S. agalactiae was detected at a high prevalence (from 0% to 59%) in the three Mexican states. This is the first detection of these pathogenic bacteria in rural farms using real-time PCR and constitutes a great risk for tilapia aquaculture in Mexico, as well as a potential dispersion of these pathogens to other aquaculture areas.

Study of the Antioxidant, Antimicrobial, and Wound Healing Properties of Raw Hydrolyzed Extract from Nile Tilapia Skin (Oreochromis niloticus).

Oreochromis niloticus (Nile tilapia) skin is a by-product of Brazilian fish farming, rich in collagen. The present study aims to evaluate the wound healing, antioxidant, and antimicrobial potential of the raw hydrolyzed extract of Nile tilapia skin, as well as the identification of the main compounds. The in vitro activity was performed using antioxidant, antimicrobial and scratch wound healing assays. An in vivo experiment was performed to evaluate the wound healing potential. On days 1, 7, 14 and 21, the lesions were photographed to assess wound retraction and on the 7th , 14th and 21st  days the skins were removed for histological evaluation and the blood of the animals was collected for glutamic oxaloacetic transaminase and glutamic pyruvic transaminase determination. The chemical study was carried out through liquid chromatography-tandem mass spectrometry and de novo sequencing of peptides. The in vitro assays showed a reduction of the gap area in 24 h, dose-dependent antimicrobial activity for both bacteria, and antioxidant activity. The chemical analysis highlighted the presence of active biopeptides. The histological evaluation showed that the raw hydrolyzed extract of Nile tilapia skin has a healing potential, and does not present toxicological effects; therefore, is promising for the treatment of wounds.

Enterogyrus spp. (Monogenea: Ancyrocephalinae) and Aeromonas jandaei co-infection associated with high mortality following transport stress in cultured Nile tilapia.

Monogenean infection of the internal organs is extremely rare when compared to external infections. This study describes mass mortality of Nile tilapia (Oreochromis niloticus L.) originating from co-infection with Enterogyrus spp. and Aeromonas jandaei following transport stress. The first fish deaths occurred on day 1 post-transport, while cumulative mortality reached approximately 90% by day 10 post-stocking. An atypical amount of pale (whitish) faeces floating on the surface of the water as well as typical clinical signs of motile Aeromonas septicemia, were reported. Adult monogeneans and countless eggs of monogeneans were found in the stomachs and the intestines of both moribund and dead fish, respectively. Two strains of A. jandaei were isolated from the kidneys. Scanning electron microscope microphotographs of the stomach revealed the presence of numerous monogeneans penetrating deep into the gastric tissue, and diffuse lesions filled with bacilliform bacteria. Histopathological examination showed multifocal eosinophilic infiltrate, gastric gland and epithelial necrosis with sloughed necrotic debris in the lumen. This is the first report of co-infection by Enterogyrus spp. and A. jandaei in Nile tilapia and the first report of Enterogyrus coronatus, Enterogyrus foratus, and Enterogyrus malbergi parasitizing tilapia in Brazil. These findings indicate that synergic co-infection by Monogenean stomach parasites (E. coronatus, E. foratus, and E. malbergi) and A. jandaei may induce high mortalities in tilapia following transport stress.

ß-glucan mimics tissue damage signaling and generates a trade-off between head kidney and spleen to activate acquired immunity in vaccinated tilapia (Oreochromis niloticus).

The association of vaccines with immunostimulants such as ß-glucan, promote the production of cytokines, competent immune cells and antibodies. However, differences between ß-glucan types and trials make it difficult to understand ß-glucan's mechanism of action. In this study, three trials were carried out with control and fish fed ß-glucan, the first trial occurred at 15 days; the second trial occurred at 30 days when we associated ß-glucan and vaccine; and the third trial occurred at 15 days post-challenge with Streptococcus agalactiae in tilapia (O. niloticus) in order to investigate immune-related gene expression in the head kidney and spleen using real-time qPCR. We found increases in HSP70, IL-6, IL-1ß, TNF-α, IL-10, Lys and C3 predominantly in the head kidney, except for IgM expression, which prevailed in the spleen, under vaccinated + ß-glucan action. This demonstrates the trade-off presented by the head kidney and spleen after immunostimulation in order to produce acquired immunity, as well as an increase in HSP70 expression in vaccinated + ß-glucan fish. The results suggest that ß-glucan stimulates the immune response through damage-associated molecular patterns (DAMPs) recognition. Therefore, these dynamics of the immune response promote a more robust defense against disease.

Effects of sodium butyrate and Lippia origanoides essential oil blend on growth, intestinal microbiota, histology, and haemato-immunological response of Nile tilapia.

This study aimed to verify the effects of dietary supplementation with sodium butyrate and Lippia origanoides, combined and isolated, on the health and zootechnical performance of Nile tilapia juveniles Oreochromis niloticus. A total of 120 fish (5.38 ± 0.65 g) were randomly distributed in 12 experimental units and fed different experimental diets for 30 days, namely: commercial diet without supplementation (Unsupplemented); commercial diet supplemented with 0.5% sodium butyrate (Butyrate); commercial diet supplemented with 0.125% L. origanoides (Lippia) and commercial diet supplemented with a mixture of 0.5% sodium butyrate and 0.125% L. origanoides (Butyrate + Lippia). After preparing the experimental diets there was an increase in the pH of diet Butyrate when compared to the other diets. After 30 days the fish supplemented with Butyrate + Lippia showed reduction significate in the mean corpuscular haemoglobin, concentration of total heterotrophic bacteria in the intestine, and lymphocyte infiltrates in the liver. Besides that, the supplementation with Butyrate + Lippia promoted an increased number of intestinal villi compared to the fish Unsupplemented ones. Additionally, fish fed a diet containing only Lippia presented an increase in the villus perimeter in the posterior region of the gut and in the red blood cell number. Animals supplemented only with sodium butyrate demonstrated increased lactic acid bacterium in the gut and macrosteatosis in the liver, besides decreased melanomacrophages in the spleen. The use of sodium butyrate associated with essential oil had positive effects on the intestinal microbiota, intestinal structure, liver, and spleen integrity, suggesting a greater efficiency of the compounds when used together in the nutrition of Nile tilapia juveniles.

Parallel and non-parallel changes of the gut microbiota during trophic diversification in repeated young adaptive radiations of sympatric cichlid fish.

BACKGROUND: Recent increases in understanding the ecological and evolutionary roles of microbial communities have underscored the importance of their hosts' biology. Yet, little is known about gut microbiota dynamics during the early stages of ecological diversification and speciation. We sequenced the V4 region of the 16s rRNA gene to study the gut microbiota of Nicaraguan Midas cichlid fish (Amphilophus cf. citrinellus). Specifically, we tested the hypothesis that parallel divergence in trophic ecology in extremely young adaptive radiations from two crater lakes is associated with parallel changes of their gut microbiota. RESULTS: Bacterial communities of fish guts and lake water were highly distinct, indicating that the gut microbiota is shaped by host-specific factors. Among individuals of the same crater lake, differentiation in trophic ecology was weakly associated with gut microbiota differentiation, suggesting that diet, to some extent, affects the gut microbiota. However, differences in trophic ecology were much more pronounced across than within species whereas similar patterns were not observed for taxonomic and functional differences of the gut microbiota. Across the two crater lakes, we could not detect conclusive evidence for parallel changes of the gut microbiota associated with trophic ecology. CONCLUSIONS: A lack of clearly differentiated niches during the early stages of ecological diversification might result in non-parallel changes of gut microbial communities, as observed in our study system as well as in other recently diverged fish species. Video Abstract.

Fish Skin and Gut Microbiomes Show Contrasting Signatures of Host Species and Habitat.

Teleost fish represent an invaluable repertoire of host species to study the factors shaping animal-associated microbiomes. Several studies have shown that the phylogenetic structure of the fish gut microbiome is driven by species-specific (e.g., host ancestry, genotype, or diet) and habitat-specific (e.g., hydrochemical parameters and bacterioplankton composition) factors. However, our understanding of other host-associated microbial niches, such as the skin mucus microbiome, remains limited. The goal of our study was to explore simultaneously the phylogenetic structure of the fish skin mucus and gut microbiome and compare the effect of species- and habitat-specific drivers on the structure of microbial communities in both tissues. We sampled 114 wild fish from 6 populations of 3 ecologically and phylogenetically contrasting Amazonian teleost species. Water samples were collected at each site, and 10 physicochemical parameters were characterized. The skin mucus, gut, and water microbial communities were characterized using a metabarcoding approach targeting the V3-V4 regions of the 16S rRNA. Our results showed a significant distinction between the phylogenetic profile and diversity of the microbiome from each microbial niche. Skin mucus and bacterioplankton communities were significantly closer in composition than gut and free-living communities. Species-specific factors mostly modulated gut bacterial communities, while the skin mucus microbiome was predominantly associated with environmental physicochemistry and bacterioplankton community structure. These results suggest that the variable skin mucus community is a relevant target for the development of microbial biomarkers of environmental status, while the more conserved gut microbiome is better suited to study long-term host-microbe interactions over evolutionary time scales.IMPORTANCE Whether host-associated microbiomes are mostly shaped by species-specific or environmental factors is still unresolved. In particular, it is unknown to what extent microbial communities from two different host tissues from the same host respond to these factors. Our study is one of the first to focus on the microbiome of teleost fish to shed a light on this topic as we investigate how the phylogenetic structure of microbial communities from two distinct fish tissues are shaped by species- and habitat-specific factors. Our study showed that in contrast to the teleost gut microbiome, skin mucus communities are highly environment dependent. This result has various implications: (i) the skin mucus microbiome should be used, rather than the gut, to investigate bacterial biomarkers of ecosystem perturbance in the wild, and (ii) the gut microbiome is better suited for studies of the drivers of phylosymbiosis, or the coevolution of fish and their symbionts.

Exposure of Nile Tilapia (Oreochromis niloticus) Fingerlings to a Saxitoxin-Producing Strain of Raphidiopsis (Cylindrospermopsis) raciborskii (Cyanobacterium) Reduces Growth Performance and Increases Mortality Rate.

Blooms of toxin-producing cyanobacteria have been more frequent and lasting because of the eutrophication of freshwater ecosystems, including those used for aquaculture. The aim of the present study was to investigate the effects of chronic exposure to a saxitoxin-producing strain of Raphidiopsis (Cylindrospermopsis) raciborskii on the performance of Nile tilapia (Oreochromis niloticus) fingerlings over a 60-d period. The fingerlings were cultivated under the following conditions: 1) water without cyanobacterium (WATER), 2) R. raciborskii in ASM-1 culture medium (CYANO), and 3) ASM-1 culture medium without cyanobacterium (ASM). Exposure to the CYANO treatment led to a significant increase in the mortality rate (p < 0.05) and a significant reduction in growth (p < 0.05) compared to fingerlings submitted to the ASM and WATER treatments, in which similar survival and growth were found (p > 0.05). Saxitoxin toxicity was dependent on the weight of the fingerling (p < 0.05), with maximum mortality caused by the ingestion of 13.66 µg saxitoxin equivalent L-1 g-1 . The present results clearly show the harm caused by saxitoxins to the production of Nile tilapia fingerlings in the early growth phase. These findings underscore the importance of maintaining adequate water quality in aquaculture activities to minimize the risk of saxitoxin-producing cyanobacterial blooms and avoid economic losses among producers. Environ Toxicol Chem 2020;39:1409-1420. © 2020 SETAC.

Quorum quenching potential of Enterococcus faecium QQ12 isolated from gastrointestinal tract of Oreochromis niloticus and its application as a probiotic for the control of Aeromonas hydrophila infection in goldfish Carassius auratus (Linnaeus 1758).

Quorum quenching (QQ), the obstruction of quorum sensing, is the most attractive way to break down the N-acyl-homoserine lactones (AHL) molecules. This work was focused at isolating AHL degrading bacteria from gastrointestinal tract of Oreochromis niloticus, with abilities appropriate for use as probiotic in aquaculture. The presence of an autoinducer inactivation (aiiA) homolog gene and AHL inactivation assay showed that Enterococcus faecium QQ12, which was one among the 20 isolates, could rapidly degrade synthetic C6-HSL in vitro and hampered violacein production by Chromobacterium violaceum. It had excellent biodegrading ability of natural N-AHL produced by Aeromonas hydrophila, suggesting that it can be used as a potential quencher bacterium for disrupting the virulence of A. hydrophila. It was susceptible to all the five antibiotics tried out. The isolate grew well at pH 3.0-7.0, was resistant to high level of bile salts (0-0.9%) and 0.5% of phenol. QQ12 also exhibited high degree of auto-aggregation and co-aggregation, confirming that it possessed good probiotic attributes. Goldfish fed diet incorporated with 108 and 1010 CFU g-1 of the QQ12 for 30 days showed 76.66-86.66% survival when challenged with A. hydrophila. The study indicates that Enterococcus faecium QQ12 could be used as a non-antibiotic feed additive in aquaculture to control bacterial diseases.

Effects of dietary supplementation with a microalga (Schizochytrium sp.) on the hemato-immunological, and intestinal histological parameters and gut microbiota of Nile tilapia in net cages.

Nutritional improvements in intensive aquaculture production systems is necessary for the reduction of stress, maximum utilization of nutritional components, and expression of the genetic potential of fish. The objective of this study was to evaluate the hemato-immunological, and histological parameters and gut microbiota of Nile tilapia fed with the microalga Schizochytrium sp. Males of Nile tilapia were distributed among eight net cages (6 m3), and fed for 105 days with two diets: control (CON), without Schizochytrium sp., and supplemented (SUP), with 1.2% Schizochytrium sp. in the diet. The final weight, mortality, hematocrit, total erythrocyte count (RBC), hemoglobin, hematimetric indices, white blood cell count (WBC), total protein, and serum lysozyme were measured. Alterations in intestinal morphology were evaluated. The gut microbiota was evaluated with next-generation sequencing. No significant differences (p>0.05) were found in the final weight and mortality between diets. Regarding the hematological parameters, a difference (p<0.05) was detected only in RBC, with there being lower values in the SUP, although this group also showed a tendency toward having an increased mean corpuscular hemoglobin level. There were no differences (p>0.05) in total protein and serum lysozyme concentrations or in WBCs between diets, except for lymphocytes, which presented lower values (p<0.05) in the SUP, suggesting immunomodulation by the polyunsaturated fatty acids present in the microalga. There was no difference (p>0.05) in the intestinal morphology between diets. Metagenomic data indicated greater richness (represented by the Chao index) and a higher abundance of the bacterial phylum Firmicutes in the gut microbiota of the tilapia fed with the SUP diet, demonstrating that the digestion and use of the components of the microalga could influence the microbial community. The results indicated that the microalga had modulatory effects on blood cells and the intestinal microbiota, without affecting the structure and integrity of the intestinal villi.

First report of Paracoccidioides brasiliensis infection in fish.

The thermodimorphic fungus Paracoccidioides brasiliensis is the etiological agent of paracoccidioidomycosis (PCM), a deep mycosis endemic in Latin American countries that affects mainly male rural workers. Infection by P. brasiliensis has also been reported in several species of terrestrial animals; however, the capacity of the fungus to infect aquatic organisms is poorly known. The aim of this study was to detect P. brasiliensis in a fish species, Nile tilapia (Oreochromis niloticus), the most farmed and widely distributed fish in endemic areas for human PCM in Brazil. As a first step, the humoral immune response against the fungus was evaluated in an experimental group of three fish immunized with inactivated P. brasiliensis yeast cells. For the seroepidemiological study, serum samples of Nile tilapia raised in cages (n = 109) and in ponds (n = 105), collected from a fish slaughterhouse, were analyzed for P. brasiliensis antibodies by ELISA using gp43 as antigen. All the inoculated fish produced antibodies against the fungus. The seropositivity observed in fish raised in cages and ponds was 17.4 and 5.7%, respectively. Due to the higher seropositivity observed in caged fish, 100 tissue samples (encephalon, liver, and kidney), from another group of tilapia raised in cages, were analyzed by polymerase chain reaction (PCR; Pb-ITSR and Pb-ITSE). Three tissue samples (liver n = 1, kidney n = 1, and enchepahlon n = 1) from three different fish resulted positive to PCR. This is the first report to show serological and molecular evidence of P. brasiliensis infection in a fish species.

Elevada frequência de Aeromonas spp. e genes de virulência em cultivos de tilápia-do-nilo (Oreochromis niloticus) em tanques-rede, na região semiárida de Pernambuco, Brasil / High frequency of Aeromonas spp. and virulence genes in Nile Tilapia farms (Oreochromis niloticus) in semi-arid regions of Pernambuco, Brazil

Objetivou-se avaliar a ocorrência de Aeromonas spp. em peixes e amostras de água na região semiárida de Pernambuco e avaliar a frequência de aerolissina (aerA), enterotoxina citotóxica (act), enterotoxina citotônica (alt) e serina protease (ahp) nesses isolados. Foram analisados 70 peixes vivos e oito mortos com sinais clínicos de aeromoniose e 16 amostras de água. Aeromonas spp. foram identificadas por análises microbiológicas (provas bioquímicas) e molecular, usando-se primers específicos para a região 16S rRNA, e a distribuição dos quatro fatores de virulência (aerA, alt, act e ahp) foi investigada por ensaio de PCR. Cento e cinquenta e cinco (84,7%) isolados foram confirmados como Aeromonas spp. na análise molecular. Os genes de virulência mais frequentes foram act (53,55%) e aerA (51,61%). De acordo com o tipo de amostra, observou-se maior frequência do gene aerA (87,5% P=0,0474) em isolados de peixes mortos e a menor frequência do gene act (47,73% P=0,0002) em peixes vivos. Este estudo demonstrou a presença de aeromoniose no cultivo de tilápias em tanques-rede, nos municípios de Jatobá e Petrolândia, na região semiárida de Pernambuco. A detecção de aerA, act e alt pode ser utilizada na tipagem de virulência de Aeromonas spp.(AU)

The purpose of this study was to evaluate the occurrence of Aeromonas spp. from fishes and tilapia net-cage farm water in semi-arid regions of Pernambuco and to evaluate the frequency of the aerolysin (aerA), cytotoxic enterotoxin (act), cytotonic enterotoxin (alt) and serine protease (ahp) genes in Aeromonas isolates. 70 live and eight dead fish with aeromoniosis clinical signs and 16 water samples were analyzed. Aeromonas spp. isolated were identified by microbiological (biochemical evidence) and molecular analysis using specific primers for 16SrRNA region, while the distribution of four virulence factors, including aerA, alt, act and ahp, was investigated by PCR assay. One hundred fifty-five (84.7%) isolates were confirmed as Aeromonas spp. by molecular analysis. The most frequent virulence genes in isolates were act (53.55%) and aerA (51,61%). According to the kind of sample, the higher frequency of aerA gene (87.5% P= 0.0474) was observed in isolates from dead fish and the lowest frequency of act gene (47.73% P= 0.0002) from live fish. This study found the presence of aeromoniosis on tilapia farming in net-cages on Jatobá and Petrolândia counties in the semiarid Pernambuco region. The detection of aerA, act and alt can be used for virulence typing of Aeromonas spp. isolates.(AU)

Elevada frequência de Aeromonas spp. e genes de virulência em cultivos de tilápia-do-nilo (Oreochromis niloticus) em tanques-rede, na região semiárida de Pernambuco, Brasil / High frequency of Aeromonas spp. and virulence genes in Nile Tilapia farms (Oreochromis niloticus) in semi-arid regions of Pernambuco, Brazil

Objetivou-se avaliar a ocorrência de Aeromonas spp. em peixes e amostras de água na região semiárida de Pernambuco e avaliar a frequência de aerolissina (aerA), enterotoxina citotóxica (act), enterotoxina citotônica (alt) e serina protease (ahp) nesses isolados. Foram analisados 70 peixes vivos e oito mortos com sinais clínicos de aeromoniose e 16 amostras de água. Aeromonas spp. foram identificadas por análises microbiológicas (provas bioquímicas) e molecular, usando-se primers específicos para a região 16S rRNA, e a distribuição dos quatro fatores de virulência (aerA, alt, act e ahp) foi investigada por ensaio de PCR. Cento e cinquenta e cinco (84,7%) isolados foram confirmados como Aeromonas spp. na análise molecular. Os genes de virulência mais frequentes foram act (53,55%) e aerA (51,61%). De acordo com o tipo de amostra, observou-se maior frequência do gene aerA (87,5% P=0,0474) em isolados de peixes mortos e a menor frequência do gene act (47,73% P=0,0002) em peixes vivos. Este estudo demonstrou a presença de aeromoniose no cultivo de tilápias em tanques-rede, nos municípios de Jatobá e Petrolândia, na região semiárida de Pernambuco. A detecção de aerA, act e alt pode ser utilizada na tipagem de virulência de Aeromonas spp.(AU)

The purpose of this study was to evaluate the occurrence of Aeromonas spp. from fishes and tilapia net-cage farm water in semi-arid regions of Pernambuco and to evaluate the frequency of the aerolysin (aerA), cytotoxic enterotoxin (act), cytotonic enterotoxin (alt) and serine protease (ahp) genes in Aeromonas isolates. 70 live and eight dead fish with aeromoniosis clinical signs and 16 water samples were analyzed. Aeromonas spp. isolated were identified by microbiological (biochemical evidence) and molecular analysis using specific primers for 16SrRNA region, while the distribution of four virulence factors, including aerA, alt, act and ahp, was investigated by PCR assay. One hundred fifty-five (84.7%) isolates were confirmed as Aeromonas spp. by molecular analysis. The most frequent virulence genes in isolates were act (53.55%) and aerA (51,61%). According to the kind of sample, the higher frequency of aerA gene (87.5% P= 0.0474) was observed in isolates from dead fish and the lowest frequency of act gene (47.73% P= 0.0002) from live fish. This study found the presence of aeromoniosis on tilapia farming in net-cages on Jatobá and Petrolândia counties in the semiarid Pernambuco region. The detection of aerA, act and alt can be used for virulence typing of Aeromonas spp. isolates.(AU)

Establishment and characterization of a competitive exclusion bacterial culture derived from Nile tilapia (Oreochromis niloticus) gut microbiomes showing antibacterial activity against pathogenic Streptococcus agalactiae.

This study reports the characterization of the microbial community composition, and the establishment and dynamics of a continuous-flow competitive exclusion culture (CFCEC) derived from gut microbiomes of Nile tilapia (Oreochromis niloticus) specimens reared on aquaculture farms in Colombia. 16S rRNA gene amplicon Illumina sequencing was used to identify taxonomical changes in the CFCEC microbial community over time. The CFCEC was developed from adult tilapia from two farms in Colombia, and CFCEC samples were collected over two months. The pH varied from 6.25 to 6.35 throughout culturing, while anaerobic and aerobic cell counts stabilized at day 9, at 109 CFU mL-1 and were maintained to day 68. A variation in the CFCEC bacterial composition was observed over time. Cetobacterium was the most abundant in the first two days and coincided with a higher CFCEC supernatant antimicrobial effect against the fish pathogen Streptococcus agalactiae. Antimicrobial activity against S. agalactiae disappeared by day 3. Changes in bacterial composition continued to day 33 with Lactococcus spp. becoming the most abundant member of the community. In conclusion, the study of the CFCEC from intestinal tract of Nile tilapia (Oreochromis niloticus) by 16S rRNA gene sequencing allowed identification of predominant bacterial genera in the continuous-flow competitive exclusion culture exhibiting antibacterial activity against the fish pathogen Streptococcus agalactiae.

Efeito da utilização de própolis vermelha em salame de peixe no crescimento de bactérias láticas fermentadoras / Effect of the use of red propolis in salami of fish in the growth of fermenting lactic bacteria

O presente estudo teve como objetivo verificar a atividade antimicrobiana do extrato de própolis vermelha em salame de tilápia. Para isso, foram elaborados três tratamentos de salame: T1 = 0,1% de extrato de própolis, T2 = 0,2% de extrato de própolis e T3 = 0,2% de sal de cura (controle). Os tratamentos foram avaliados quanto a quantificação de bactérias mesófilas e bactérias láticas, e o pH durante 15 dias de maturação do salame. O extrato de própolis não interferiu no crescimento das bactérias láticas, mas também não inibiu a presença de bactérias mesófilas. Nos primeiros cinco dias de fermentação o pH diminuiu (pH 5,9 ), aumentando posteriormente ao final da maturação dos salames (pH 6,8). Conclui-se que é preciso testar concentrações mais altas da própolis vermelha em embutidos cárneos fermentados a fim de se obter uma ação mais eficiente contra bactérias mesófilas.