Fish Gut Samples Research Articles

Metagenomic approach revealed the mobility and co-occurrence of antibiotic resistomes between non-intensive aquaculture environment and human

BackgroundAquaculture is an important food source worldwide. The extensive use of antibiotics in intensive large-scale farms has resulted in resistance development. Non-intensive aquaculture is another aquatic feeding model that is conducive to ecological protection and closely related to the natural environment. However, the transmission of resistomes in non-intensive aquaculture has not been well characterized. Moreover, the influence of aquaculture resistomes on human health needs to be further understood. Here, metagenomic approach was employed to identify the mobility of aquaculture resistomes and estimate the potential risks to human health.ResultsThe results demonstrated that antibiotic resistance genes (ARGs) were widely present in non-intensive aquaculture systems and the multidrug type was most abundant accounting for 34%. ARGs of non-intensive aquaculture environments were mainly shaped by microbial communities accounting for 51%. Seventy-seven genera and 36 mobile genetic elements (MGEs) were significantly associated with 23 ARG types (p < 0.05) according to network analysis. Six ARGs were defined as core ARGs (top 3% most abundant with occurrence frequency > 80%) which occupied 40% of ARG abundance in fish gut samples. Seventy-one ARG-carrying contigs were identified and 75% of them carried MGEs simultaneously. The qacEdelta1 and sul1 formed a stable combination and were detected simultaneously in aquaculture environments and humans. Additionally, 475 high-quality metagenomic-assembled genomes (MAGs) were recovered and 81 MAGs carried ARGs. The multidrug and bacitracin resistance genes were the most abundant ARG types carried by MAGs. Strikingly, Fusobacterium_A (opportunistic human pathogen) carrying ARGs and MGEs were identified in both the aquaculture system and human guts, which indicated the potential risks of ARG transfer.ConclusionsThe mobility and pathogenicity of aquaculture resistomes were explored by a metagenomic approach. Given the observed co-occurrence of resistomes between the aquaculture environment and human, more stringent regulation of resistomes in non-intensive aquaculture systems may be required.4-S8N7kaEyTj6WBvQ6hX9pVideo

Microplastic in fishes: the first report from a Himalayan River - Alaknanda.

The present study aimed to understand microplastic (MP) ingestion by five fish species with different feeding habits namely, Schizothorax richardsonii and Crossocheilus latius (herbivore),Cyprinus carpio (omnivore), Tor chelenoid (herbi-omnivore), and Botia horii (carnivore). The fishes were sampled from River Alaknanda (one of the headwaters of River Ganga) patch at Srinagar, Garhwal, Uttarakhand. The fish gut samples were digested with 30% hydrogen peroxide and vacuum filtered through glass microfiber filter. Each filter paper was observed microscopically to count MPs, and then, selected MPs were chemically characterized using Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy (ATR-FTIR). The results revealed that all the species were contaminated with MPs and the rate of MP ingestion varied with feeding habits. The herbi-omnivore fish, Tor chelenoid, was observed to have the highest MP ingestion. Among the analyzed particles, fibers (66%) were the dominant type of MPs. The MPs were chemically characterized as High-Density Polyethylene (HDPE), Polypropylene (PP), and Polyester. This study contributes as a reference for the forthcoming researches, as it is the pioneer work on the ingestion of MP by fishes of a Himalayan River.

The Gill-Associated Bacterial Community Is More Affected by Exogenous Chlorella pyrenoidosa Addition than the Bacterial Communities of Water and Fish Gut in GIFT Tilapia (Oreochromis niloticus) Aquaculture System.

Microalgae has been widely used in aquaculture to improve both the water environment and fish growth; however, the current understanding of the effects of microalgae addition on the key players involved in regulating the water environment and fish health, such as microorganisms, remains limited. Here, a 50-day mesocosm experiment was set up to simulate the culture of Genetic Improvement of Farmed Tilapia (GIFT, Oreochromis niloticus) with an average weight of 14.18 ± 0.93 g and an average length of 82.77 ± 2.80 mm. Different amounts of Chlorella pyrenoidosa were added into these artificial systems to investigate dynamics of bacterial communities in aquaculture water, fish gill, and gut using amplicon-based high-throughput sequencing technology. Our results showed that Chlorella pyrenoidosa addition increased diversity and network complexity of gill-associated bacterial communities rather than those of the water and gut. Furthermore, more biomarkers in the gill-associated bacterial communities were detected in response to Chlorella pyrenoidosa addition than the water and fish gut samples. These findings highlighted the high sensitivity of gill-associated bacterial communities in response to the Chlorella pyrenoidosa addition, implying Chlorella pyrenoidosa addition could play important roles in regulating the fish mucosal immunity by altering the gill-associated microbiota.

Using the health index of the environmental microbiota to predict the health status of aquatic wildlife

In aquatic animals, the gut microbiota has a significant function in host health and is impacted by the environmental microbiota. To conveniently monitor, assess and predict aquatic animals’ health, a general and effective microbiota health index (microHI) would be helpful, and the correlation between the aquatic environmental microHI and wildlife gut microHI could be expected. Here, we proposed a general microHI based on the microbial phenotypes of mobile element content, potential pathogenicity and oxidative stress tolerance, and then tested the reliability of the fish gut microHI and its correlation with the environmental microHI in the middle Yangtze River. The results showed that there were positive correlations among the abundance ratios of microbes with the microbial phenotypes of mobile element content, potential pathogenicity and oxidative stress tolerance, suggesting that defining the microHI using the average abundance ratios of microbes without these microbial phenotypes is suitable. Fish gut samples with a lower microHI were more likely to have been obtained from specimens with a disordered microbiota (always means unhealthy host), indicating that the gut microHI has the potential to be used to evaluate the health status of aquatic animals. In the middle Yangtze River, the aquatic environmental microHI had hundred-kilometers-scale high spatial heterogeneity but month-scale low temporal heterogeneity from June to September 2020, and in the section with a low aquatic environmental microHI, the fish gut samples were more likely to have relatively low gut microHIs, indicating that the environmental microHI has the potential to be used to predict the health status of aquatic animals at the community level. The current work initially showed the possibility of using environmental microHI to predict aquatic animals’ health although it was not very exact and precise. We encourage more researches to assess the quantitative relationship between the aquatic environmental microHI and aquatic animal gut microHI.

Isolation, estimation of bacterial load, and characterization of gut microbes of three Garra species from Bhavani river, Western ghats,Tamil Nadu, India

Microbiomes play an important role in fish physiology, particularly in digestion, by metabolizing largely indigestible feed components for the host or synthesis of essential micronutrients. The gut flora has a dynamic effect on the host’s gut and systemic immunity. This study is the preliminary exploration of gut microflora of three species Garra mcclellandi, Garra stenorhynchus, and Garra gotyla stenorhynchus . The total bacterial load ranges from 66×108 CFU/g to 81×108 CFU/g in the gut sample. It was found that the maximum load in G. stenorhynchus with 81×108 CFU/g followed by Gg. stenorhynchus with 72×108 CFU/ml minimum in G. mcclellandi with 66×108 CFU/ml. The isolates were subjected to various biochemical characterizations such as the Indole test, Voges Proskauer test, Methyl Red test, Carbohydrate Utilization Test, Citrate Utilization Test, Starch Hydrolysis, and Catalase Production activity. The three fish gut samples are screened for antagonistic properties for certain human pathogens which show zero antagonistic activities. The present preliminary study shows variation in the gut microflora and these microbes may be used for the development of probiotics and drug production.

Detection of extended spectrum beta lactam producing Escherichia coli isolated from Cyprinus carpio in Mosul city

Extended-spectrum b-lactamase (ESBL)-producing Escherichia coli constitute an emerging health problem globally, fish act as a potential reservoir for ESBLs E. coli and serve as a vehicle of transmission of ESBL resist genes to others strains of bacteria. A total of 75 samples of Cyprinus carpio were collected from the local fish market of Mosul city during the period between) October 2021 to February 2022(. each sample was placed separately in a sterile plastic bag and transported directly under cooling conditions to the microbiology lab, College of Veterinary Medicine. ESBLs producing Escherichia coli were isolated and characterized using MacConkey agar medium supplemented with 2 mcg/ml cefotaxime. Twenty-six isolates 34% of fish gut samples were obtained. A polymerase chain reaction was carried out to confirm the results of the isolation using special primers for E. coli (ECO223-f, ECO223-r). Resistance genes assay were performed using the primers Cefotaxamase (CTX-M) and sulphydryl variable (SHV). All isolates showed that possessed the CTX-M gene 100%, while none of the isolates possessed the SHV gene. This study showed that fish play a major role in the transmission of broad-spectrum beta-lactam-resistant Escherichia coli to humans as a result of handling or marketing it, or by consuming contaminated or infected fish.

Do Changes in Prey Community in the Environment Affect the Feeding Selectivity of Silver Carp (Hypophthalmichthys molitrix) in the Pearl River, China?

Silver carp (Hypophthalmichthys molitrix) is a world-wide species and its ecosystem level influence is controversial. Therefore, understanding the feeding behaviour of silver carp is important for assessing ecosystem functioning in many freshwater habitats. In this study, we used 18S rRNA sequencing to identify the spatiotemporal taxonomic composition of water and fish gut samples. Significant seasonal and spatial variations were observed in the taxonomic compositions of water and fish gut samples (p &lt; 0.05). Five selected environmental factors (temperature, total phosphorus, chlorophyll a, ammonia, and distance from the first dam downstream) significantly affected the taxonomic composition of water (p &lt; 0.05). Additionally, the temperature was the most important factor affecting seasonal (winter and summer) variations in the prey community composition of water, whereas the distance from the dam was the key factor contributing to spatial differences in the community. Significant differences were observed in the taxonomic composition between the water and fish gut samples (p &lt; 0.05). Feeding selectivity analysis revealed that the selection ratios of the top 20 prey taxa underwent slight spatial and temporal changes, but silver carp always preferred to feed on Peridiniales, unidentified Streptophyta, unidentified Trebouxiophyceae, and unidentified Chlorophyceae. These results indicate that changes in the prey community in the environment affect the taxonomic composition of the fish gut but do not affect feeding preferences.

Northern pike (Esox lucius) young‐of‐year gut microbiota and associations with wetland nursery water microbial communities in the St. Lawrence River

AbstractThe core‐microbiome hypothesis postulates that key fish gut microbiota develop independently from the environment they exist in. Fish nursery habitats serve as critical environments early in ontogeny that support fish growth and development, but their relationship to the development of gut microbiota is poorly understood. We compared gut microbiota of first growing season northern pike (Esox lucius) and source water from two spatially distinct nursery wetlands to examine their potential association and as a field test of the core‐microbiome hypothesis. Microbial communities within the pike hindguts and associated water column samples were characterized through metabarcoding and sequencing of the 16S rRNA gene. As expected, community level non‐metric multidimensional scaling analysis revealed significant separation in microbial community composition between fish and water samples. Unifrac distances indicated no significant dissimilarity among fish gut and source water combinations compared to fish gut samples pairwise tested with non‐source water samples, suggesting similar patterns of in vivo selection of the gut microbiome. This was consistent among habitats for three of four dominant hindgut bacterial genera (Aeromonas spp., Clostridium sensu stricto, and Plesiomonas spp.). Cetobacterium spp. was an exception, where they dominated fish guts in one wetland system. Because these four‐core gut microbiota were only minor components of the source water environment, it is possible that environmental exposure is a selective process in the developing fish gut. Further experimental and field research with examination of the influence of diet is needed to isolate mechanisms involved in gut microbiome development in freshwater fishes.

Comparison of environmental microbiomes in an antibiotic resistance-polluted urban river highlights periphyton and fish gut communities as reservoirs of concern

Natural waterways near urban areas are heavily impacted by anthropogenic activities, including their microbial communities. A contaminant of growing public health concern in rivers is antibiotic resistant genes (ARGs), which can spread between neighboring bacteria and increase the potential for transmission of AR bacteria to animals and humans. To identify the matrices of most concern for AR, we compared ARG burdens and microbial community structures between sample types from the Scioto River Watershed, Ohio, the United States, from 2017 to 2018. Five environmental matrices (water, sediment, periphyton, detritus, and fish gut) were collected from 26 river sites. Due to our focus on clinically relevant ARGs, three carbapenem resistance genes (blaKPC, blaNDM, and blaOXA-48) were quantified via DropletDigital™ PCR. At a subset of nine urbanized sites, we conducted16S rRNA gene sequencing and functional gene predictions. Carbapenem resistance genes were quantified from all matrices, with blaKPC being the most detected (88 % of samples), followed by blaNDM (64 %) and blaOXA-48 (23 %). Fish gut samples showed higher concentrations of blaKPC and blaNDM than any other matrix, indicating potential ARG bioaccumulation, and risk of broader dissemination through aquatic and nearshore food webs. Periphyton had higher concentrations of blaNDM than water, sediment, or detritus. Microbial community analysis identified differences by sample type in community diversity and structure. Sediment samples had the most diverse microbial communities, and detritus, the least. Spearman correlations did not reveal significant relationships between the concentrations of the monitored ARGs and microbial community diversity. However, several differentially abundant taxa and microbial functions were identified by sample type that is definitive of these matrices' roles in the river ecosystem and habitat type. In summary, the fish gut and periphyton are a concern as AR reservoirs due to their relatively high concentration of carbapenem resistance genes, diverse microbial communities, and natural functions that promote AR.

High-throughput Method for Novel Medium Development for Culture of Anaerobic Gut Bacteria.

Gut microbiota play important roles in the health of their host and detailed investigation of these organisms requires in vitro culture. Culturing strictly anaerobic bacteria can be a challenge as the gut environment they inhabit is nutritionally complex. Use of complex media containing nutritionally rich but undefined gut fluid reduces the accuracy of physiological and metabolomic studies. Here we present a high-throughput protocol for comparing growth rates of fastidiously anaerobic bacteria on different media. These protocols can be used to develop a solid medium made up of commercially sourced ingredients, providing replicable growth conditions for previously uncultured anaerobic bacteria. As many fastidious bacteria grow poorly in a liquid broth, these protocols measure bacterial growth rate on solid media. These protocols speed up and simplify the growth rate measurement process by using a multiwell format and equations in place of physical McFarland standards to calculate approximate cell density. Bacterial strains belonging to the families Erysipelotrichaceae and Lachnospiraceae (phylum Firmicutes) isolated from the hindgut of Kyphosus sydneyanus were used to demonstrate the efficacy of these protocols. Bacterial growth rates were compared between a nutritionally rich medium with gut fluid versus a novel replicable medium with mannitol. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Preparation of solid YCFA growth medium Basic Protocol 2: Collection of fish gut samples and plating to single isolates Basic Protocol 3: Genetic identification of single isolates with colony PCR and 16S rRNA gene sequencing Basic Protocol 4: Measurement of bacterial growth rates on solid media.

Comparison of the Intestinal Bacteria Between Black Seabass Centropristis striata Reared in Recirculating Aquaculture System and Net Pen.

Determination of diversity and function of the bacteria in fish gut is essential to understanding the interaction between intestinal bacteria and their host organism. This study compared intestinal bacterial community of black seabass (Centropristis striata) hatched by the same breeding farm but reared in different aquaculture systems, an indoor recirculating aquaculture system (RAS) and an inshore net pen (INP). The fish were fed with formulated feed manufactured by same feed company. Bacteria in fish gut, formulated feed and seawater were identified by 16S rRNA high throughout sequencing (HTS). Total 1484 OTUs, which belonged to 34 phyla and 79 genera, were identified from fish gut, formulated feed and seawater. In fish gut, 24 phyla and 43 genera were identified. Proteobacteria, Fusobacteria, and Firmicutes dominated at the phylum level in fish gut in INP, while Proteobacteria and Firmicutes dominated in fish gut in RAS. Photobacterium, Vibrio, and Cetobacterium dominated at the genus level in fish gut in both INP and RAS. One OTU of Photobacterium occurred in all the fish gut samples, suggesting this bacterium might be the main component of the core microbiota. No significant difference was found in bacterial diversity in fish gut between INP and RAS, suggesting genetic background should be a primary factor determining intestinal bacterial community of black seabass. Bacterial diversity in seawater was high relative to that in fish gut and formulated feed, regardless in INP or RAS. The common OTU between fish gut and seawater was more than that between fish gut and formulated feed in INP, while the common OTU between fish gut and seawater was slightly less than that between fish gut and formulated feed in RAS. These results reveal that the bacteria in formulated feed and seawater could influence the bacteria in fish gut, and their priority in shaping intestinal bacterial community depended on the bacterial composition in feed and seawater. This study reveals that intestinal bacterial community of black seabass was influenced by both genetic background and environmental factors.

Physiological parameters and gut microbiome associated with different dietary lipid levels in hybrid yellow catfish (Tachysurus fulvidraco♀× Pseudobagrus vachellii♂)

Yellow catfish are intensively farmed in China and are often fed a high-fat diet (HFD) with the aim of using less protein. However, an excess of dietary lipids is likely to affect the gut microflora, which strongly affects immunity and nutrient digestion. To determine the effects of different lipid levels on the growth, physiological parameters and gut microbiome of hybrid yellow catfish, we conducted an 8-week feeding experiment with a low-fat diet (2% lipids, LFD), a normal-fat diet (9% lipids, NFD), and a HFD (15% lipids) (120 fish per group). The HFD group showed higher serum alanine aminotransferase and aspartate transaminase activities, which suggests that excess dietary lipids cause liver damage. A total of 1138 operational taxonomic units, 11 phyla, and 117 genera were identified from fish gut samples. Neither the HFD nor the LFD strongly affected the microbial composition in gut samples. Compared with fish in the NFD, those in the HFD and LFD showed significantly decreased intestinal microbial diversity. The composition of macronutrients in the different diets affected the composition of intestinal microflora, mainly the phyla Fusobacteria, Proteobacteria, and Firmicutes. The HFD and the LFD favored the growth of Fusobacteria, while the HFD and LFD resulted in decreased abundance of Firmicutes and Proteobacteria, respectively. These findings shed light on the complex relationships among diet, intestinal microorganisms, and host metabolism. When using an HFD for farmed fish, its effects on the gut microbiome should be considered to avoid illness and poor growth.

Microbiomes of fish, sediment and seagrass suggest connectivity of coral reef microbial populations.

The benthic environments of coral reefs are heavily shaped by physiochemical factors, but also the ecological interactions of the animals and plants in the reef ecosystem. Microbial populations may be shared within the ecosystem of sediments, seagrasses and reef fish. In this study, we hypothesize that coral reef and seagrass environments share members of the microbial community that are rare in some habitats and enriched in others, and that animals may integrate this connectivity. We investigated the potential connectivity between the microbiomes of sediments, seagrass blades and roots (Syringodium isoetifolium), and a seagrass-specialist parrotfish (C. spinidens) guts in reef areas of Fiji. We contrasted these with sediment samples from the Florida Keys, gut samples from surgeonfish (A. nigricauda, Acanthurinae sp. unknown, C. striatus), and ocean water microbiomes from the Atlantic, Pacific and Indian Oceans to test the robustness of our characterizations of microbiome environments. In general, water, sediment and fish gut samples were all distinct microbiomes. Sediment microbiomes were mostly similar between Fiji and Florida, but also showed some regional similarities. In Fiji, we show connectivity of a shared microbiome between seagrass, fish and sediments. Additionally, we identified an environmental reservoir of a surgeonfish symbiont, Epulopiscium. The connection of these ecosystem components suggests that the total microbiome of these environments may vary as their animal inhabitants shift in a changing ocean.

Effects of hydrolyzed fish protein and autolyzed yeast as substitutes of fishmeal in the gilthead sea bream (Sparus aurata) diet, on fish intestinal microbiome

BackgroundThis study evaluated the effects of partial substitution of dietary fishmeal (FM) with either fish protein hydrolysate (FPH) or autolysed dried yeast (HiCell®, Biorigin, Brazil) on intestinal microbiota of gilthead sea bream (Sparus aurata). A total number of 720 fish of 122.18 ± 6.22 g were fed for 92 days with three different diets in triplicate (3 tanks/diet). A diet based on FM/vegetable meal was used as control. The other two diets were formulated by replacing FM with 5% of either FPH or HiCell®. To analyze the gut microbiota associated to autochthonous and allochthonous microbial communities, the Illumina MiSeq platform for sequencing of 16S rRNA gene and QIIME pipeline were used.ResultsA total number of 102 OTUs (operational taxonomic units) at 97% identity were identified in fish gut samples collected at the end of feeding trial. Fourteen OTUs constituted the core gut microbiota, i.e. those OTUs found in at least nine out of fifteen samples per group and shared regardless of the diet. Eight OTUs were assigned to Firmicutes represented by Lactobacillus, Staphylococcus, and Bacillus genera, and six to Proteobacteria phylum. Dietary dried yeast autolysate modulated the intestinal microbiota by promoting the growth of some beneficial bacteria. At order level, fish fed yeast showed an enrichment in Bacillales and Clostridiales as compared to the control group, whereas fish fed FPH showed a significantly lower amount of bacteria belonging to Alteromonadales and Enterobacteriales than the other two feeding groups. Although we did not observe any effect of 5% FM replacement with alternative nitrogen sources at phylum level, at lower taxonomical levels, the composition of gut microbiota, in terms of relative abundance of specific taxa, was significantly influenced by the dietary treatment.ConclusionsThe metabarcoding analysis revealed a clearly intestinal microbiota modulation in response to dietary autolyzed yeast. The abundance of some beneficial bacteria, i.e. indigestible carbohydrate degrading- and SCFA producing bacteria, was positively affected. Brewer’s yeast autolysate could be a valid alternative protein source to FM as well as a valid functional ingredient for aquafeed production.

MATRIX-ASSISTED LASER DESORPTION IONIZATION-TIME OF FLIGHT MASS SPECTROMETRY BASED IDENTIFICATION OF THE FISH GUT MICROBIOTA

The aim of the study was to investigate the composition of intestinal microflora of freshwater fish in Latvia. A total amount of 28 fish were collected from fishermen (n=20) and retail outlets (n=8), including roach Rutilus rutilus, n=15, crucian carp Carassius carassius, n=5, perch Perca fluvialitis, n=5, bream Blicca bjoerkna, n=3. Microbiological testing consisted of the detection of total viable count (TPC), Enterobacteriaceae and coliforms with subsequent identification with MALDI-TOF Biotyper. TPC, coliforms and Enterobacteriaceae counts ranged from 2.7±0.4 to 5.4±0.3, 2.4±1.5 to 3.7±0.7 and 2.00±1.2 to 3.7±2.5 log cfu/g in gut of wild crucian carp and retailed roach, wild perch and retailed roach, and crucian carp and bream, accordingly. The TPC, coliforms and Enterobacteriaceae counts were significantly higher in retailed fish than in wild fish gut samples (P≤0.05). Gut microbiota were represented by Proteobacteria (93.0%), Firmicutes (3.9%) and Ascomycota (3.1%). The most abundant families were Enterobacteriacea (50.8%) and Pseudomonadaceae (36.7%). Rahnella aquatilis, Serratia fonticola and Pantoe aagglomerans were the most abundant among Enterobacteriaceae while Pseudomonas extremorientalis and P. fragi among the Pseudomonadaceae. Results of the present study show that the gut of freshwater fish were mostly represented by Enterobacteriaceae and Pseudomonadaceae and the presence of fish bacterial pathogens must be considered.

A comparison of storage methods for gut microbiome studies in teleosts: Insights from rainbow trout (Oncorhynchus mykiss)

Immediate freezing is perhaps the most preferred method used for preserving gut microbial samples, but research on sample preservation has been principally based around samples from mammalian species, and little is known about the advantages or disadvantages relating to different storage methods for fish guts. Fish gut samples may pose additional challenges due to the different chemical and enzymatic profile, as well as the higher water content, which might affect the yield and purity of DNA recovered. To explore this, we took gut content and mucosal scrape samples from 10 rainbow trout (Oncorhynchus mykiss), and tested whether different preservation methods have any effect on the ability to construct high quality genomic libraries for shotgun and 16S rRNA gene sequencing. Four different storage methods were compared for the gut content samples (immediate freezing on dry ice, 96% ethanol, RNAlater and DNA/RNA shield), while two different methods were compared for mucosal scrape samples (96% ethanol and RNAlater). The samples were thereafter stored at −80 °C. Our findings concluded that 96% ethanol outperforms the other storage methods when considering DNA quantity, quality, cost and labor. Ethanol works consistently well for both gut content and mucosal scrape samples, and enables construction of DNA sequencing libraries of sufficient quantity and with a fragment length distribution suitable for shotgun sequencing. Two main conclusions from our study are i) sample storage optimisation is an important part of establishing a microbiome research program in a new species or sample type system, and ii) 96% ethanol is the preferred method for storing rainbow trout gut content and mucosal scrape samples.

Prevalence and characteristics of -lactamase and plasmid-mediated quinolone resistance genes in Escherichia coli isolated from farmed fish in China

To determine the molecular epidemiology of extended-spectrum β-lactamases (ESBLs) and plasmid-mediated quinolone resistance (PMQR) in Escherichia coli isolated from farmed fish in China. E. coli was isolated from fish gut samples from fish farmed throughout Guangdong province and tested for the presence of the β-lactamase genes and PMQR-encoding genes using PCR and DNA sequence analysis. Co-transfer of plasmids encoding for ESBLs as well as PMQR determinants was explored by conjugation into E. coli. A total of 218 non-duplicate E. coli were recovered from fish gut samples. β-Lactamase genes were identified in 19 (17%) of 112 strains with reduced susceptibility to ampicillin, and PMQR genes were identified in 59 (73.8%) of 80 strains with reduced susceptibility to ciprofloxacin. Only three ESBL genes were identified in three isolates: bla(CTX-M-14), bla(CTX-M-79) and bla(SHV-27). PMQR gene screening identified qnr genes (n = 59) as the most common, including qnrB (n = 33), qnrS (n = 21) and qnrD (n = 5), with aac(6')-Ib-cr (n = 6) being rarely found. The co-carriage of two or three PMQR genes in one strain was found in 7 (11.9%) isolates. The ESBL gene bla(CTX-M-79) was found to be co-carried with qnrS. Co-transfer of qnrS was observed with bla(CTX-M-79). Our study is the first to demonstrate the existence of high levels of mobile genes conferring reduced susceptibility to fluoroquinolones as well as the presence of ESBL genes in fish produced in China, and identifies a significant reservoir of antibiotic resistance genes relevant to human medicine.

Detection of Salmonellae from Fish in a Natural River System

Sediment, water, and fish gut samples taken at three sites near the headwaters of the San Marcos River, Texas, were analyzed for salmonellae Salmonella spp. by culture and molecular techniques. While enrichment cultures from sediment and water samples from the two uppermost sites were negative for salmonellae in polymerase chain reaction analyses, both sediment and water samples were positive at the downstream site. At all sites, salmonellae were present in the guts of different fishes (e.g., largemouth bass Micropterus salmoides, channel catfish Ictalurus punctatus, common carp Cyprinus carpio, and suckermouth catfish Hypostomus plecostomus). The highest percentage of detection (33% of analyzed fish) occurred at the downstream site, whereas detection percentages at the upper two sites were 18% and 17%. Detection of salmonellae was usually limited to one segment of the gut (i.e., upper or lower part). Serovars were highly variable among individuals and differed between the upper and lower gut in the only individual (a common carp) that had salmonellae in both gut segments. In situ hybridization demonstrated that salmonellae were normally associated with particulate material in the gut and occurred in highly variable numbers ranging from an occasional organism to a majority of the gut microbe population. These results demonstrate the presence of different serovars of potentially human pathogenic salmonellae among four ecologically distinct fishes within natural environments. They also suggest that salmonellae are not components of the indigenous microbial community in fish intestines but rather are ingested with particulate material.

Screening and evaluation of probiotics as a biocontrol agent against pathogenic Vibrios in marine aquaculture

The present work aims at finding potential probionts from marine sources as a biocontrol agent against pathogenic Vibrio species in shrimp larval culture. A total of 109 bacterial strains were isolated from seawater, sediment and marine fish-gut samples, and were screened for their antagonistic activity against Vibrio species. Three strains (Q, Q1 and M) isolated from the marine sediment were found antagonistic against Vibrio strains. Based on 16S ribosomal DNA gene sequence analysis, the strain Q was identified as Paenibacillus spp. (EF012164); Q1 as Bacillus cereus (DQ915582); and the M as Paenibacillus polymyxa (DQ915580). Further, the two bacterial species, Paenibacillus spp. and B. cereus were challenged separately at two different concentrations of 10(4) and 10(5) CFU ml(-1) for probiotic activity in the postlarvae of Penaeus monodon against pathogenic Vibrio harveyi and Vibrio spp. The present study identified the probiotic activity of Paenibacillus spp., B. cereus and Pa. polymyxa against the pathogenic Vibrios in the postlarvae of P. monodon. In vivo study reveals that the marine bacterial species can be used as probionts against pathogenic Vibrios in shrimp larval culture practices.

USING FISH TO SAMPLE DIATOM COMPOSITION IN STREAMS: ARE INTESTINAL FLORAS REPRESENTATIVE OF NATURAL SUBSTRATES?

Sellman, S. M., Johansen, J. R. &amp; Coburn, M. M.Department of Biology, John Carroll University, University Heights, OH 44118 USAThe diatom composition of natural substrates at a single site were compared with that of the gut contents of three species of fish: stoneroller minnows, bluntnose minnows, and common shiners. Seven samples of each type were collected on a single day from a single pool in a headwater stream. Cluster analysis showed that diatom species composition clustered by fish species, with common shiners being a totally exclusive cluster, while natural substrates were scattered within the stoneroller and bluntnose clusters. Both richness and Shannon diversity were higher in fish gut samples than in the samples collected by humans, with stonerollers having the greatest diversity. When water quality indices calculated from the diatom floras recovered in these samples were compared, it was surprising to see that these indices consistently were better in fish gut samples than in samples collected by researchers. Common shiners particularly differed, a result possibly due to the fact that they apparently fed off of Cladophora and the highly oxygenated rock wall where the stream cascaded into the pool. In no case was the difference between stonerollers and natural substrates significantly different. Furthermore, samples from natural substrates had an internal similarity not significantly different from their similarity to samples in stoneroller and bluntnose guts. Our data thus consistently indicate that stoneroller minnows are excellent samplers that efficiently collect diverse, representative diatom samples.