Molecular analysis of intestinal microbiota of rainbow trout (Oncorhynchus mykiss)

The aim of this study was to evaluate different molecular tools based on the 16S rRNA gene, internal transcribed spacer, and the rpo B gene to examine the bacterial populations present in juvenile rainbow trout intestines. DNA was extracted from both pooled intestinal samples and bacterial strains. Genes were PCR-amplified and analysed using both temporal temperature gradient gel electrophoresis (TTGE) and restriction fragment length polymorphism methods. Because of the high cultivability of the samples, representative bacterial strains were retrieved and we compared the profiles obtained from isolated bacteria with the profile of total bacteria from intestinal contents. Direct analysis based on rpo B-TTGE revealed a simple bacterial composition with two to four bands per sample, while the 16S rRNA gene-TTGE showed multiple bands and comigration for a few species. Sequencing of the 16S rRNA gene- and rpo B-TTGE bands revealed that the intestinal microbiota was dominated by Lactococcus lactis, Citrobacter gillenii, Kluyvera intermedia, Obesumbacterium proteus , and Shewanella marinus . In contrast to 16S rRNA gene-TTGE, rpo B-TTGE profiles derived from bacterial strains produced one band per species. Because the single-copy state of rpo B leads to a single band in TTGE, the rpo B gene is a promising molecular marker for investigating the bacterial community of the rainbow trout intestinal microbiota.     

Microbial diversity of intestinal contents and mucus in rainbow trout (Oncorhynchus mykiss)

AIMS: The aim of this study was to understand the microbial community of intestinal contents and mucosal layer in the intestine of rainbow trout by means of culture-dependent conventional and independent molecular techniques. METHODS AND RESULTS: Forty-one culturable microbial phylotypes, and 39 sequences from 16S rRNA and two from 18S rRNA genes, were retrieved. Aeromonadaceae, Enterobacteriaceae and Pseudomonadaceae representatives were the dominant cultured bacteria. Genomic DNA isolated from intestinal contents and mucus was used to generate 104 random clones, which were grouped into 32 phylotypes at 99% minimum similarity, most of which were affiliated with Proteobacteria (>70% of the total). However, unlike library C (intestinal contents), the phyla Bacteroidetes and Fusobacteria were not found in intestinal mucus (library M), indicating that the microbiota in the gut mucus was different from that of the intestinal contents. Twelve sequences were retrieved from denaturing gradient gel electrophoresis analysis, and dominant bands were mostly related to Clostridium. CONCLUSIONS: Many novel sequences that have not been previously recognized as part of the intestinal flora of rainbow trout were retrieved. SIGNIFICANCE AND IMPACT OF THE STUDY: The fish gut harbours a larger bacterial diversity than previously recognized, and the diversity of gut mucus is different from that of intestinal contents.     

Changes in microbiota of rainbow trout caused by sediments contamination

The abundance, composition and hydrocarbon-degrading bacteria, as possible biomarkers of contamination with oil hydrocarbons, of autochthonous and alochtonous microbiota of the digestive tract of rainbow trout have been estimated. The samples of the bottom sediments for microbiological tests have been collected and a response of natural bacterial communities in the digestive tract of rainbow trout and nutritional changes has been investigated. Experimental fish have been fed with a mixture of three substances with the aim to assess the influence of hydrocarbon-degrading bacteria contained in the sediments on the microbiota of rainbow trout’s digestive tracts. The abundance values of rainbow trout intestinal heterotrophic bacteria were found to change depending on alochtonous microbiota of different bottom sediments given to the experimental fish with food in vitro. According to the results of our research, it is likely that the changes in the abundance values of the microbiota of the digestive tract of fish and in the proportions of functional groups of the bacteria allow us to determine changes in the functional activity of bacteria depending on food composition. Any relative increase or decrease of abundance or activity of alochtonous microbiota allows the prediction of toxic effects of the contaminants on animals and the environment.     

The effect of antimicrobial compounds on the gastrointestinal microflora of rainbow trout, Salmo gairdneri Richardson

Populations of aerobic and anaerobic heterotrophic bacteria occurring in the gastrointestinal tract of healthy rainbow trout were estimated using a dilution plate technique. Data revealed a progressive decline in numbers of aerobic bacteria along the digestive tract from oesophagus to lower intestine. However, the highest numbers were recovered from the intestinal contents and faeces. Anaerobes were generally restricted to the upper intestine and intestinal contents. The aerobic component of the bacterial microflora from the digestive tract was equated with Acinetobacter calcoaceticus, Aeromonas hydrophila, Bacillus circulans, Bac. megaterium , coryneforms, Grampositive irregularly shaped rods, Flavobacterium sp., Kurthia sp., Microhacterium sp., Providencia stuartii, Pseudomonas spp., Ps.fluorescens and Ps. pseudoalcaligenes . Evidence from scanning electron microscopy pointed to a general lack of colonization of the gut wall: instead, microorganisms were abundant in the intestinal contents. Antimicrobial compounds, i.e. oxolinic acid, oxytetracycline and sulphafurazole (which are commonly used to combat infections by Gramnegative bacterial fish pathogens), caused an increase in bacterial numbers throughout the digestive tract, with maximal numbers in the lower intestine. The bacteria, comprising an essentially different range of taxa, were generally resistant to the antibiotics in use. Conversely, erythromycin and penicillin G, which are used to treat some diseases caused by Gram-positive bacteria, caused a rapid reduction in bacterial numbers within the gastrointestinal tract.     

Bacterial community composition of the gut microbiota of <Emphasis Type="Italic">Cylindroiulus fulviceps</Emphasis> (diplopoda) as revealed by molecular fingerprinting and cloning

Bacterial clone libraries of the gut microbiota of nurtured and starved Cylindroiulus fulviceps specimens displayed the predominance of the phyla Bacteroidetes (55 and 37 %, respectively) and Proteobacteria (40 and 35 %, respectively) and a high similarity to bacteria previously detected in the intestinal tract of termites and beetles, which are known to harbor symbiotic bacteria essential for digestive activity. Bacterial isolates were dominated by Proteobacteria (74 %), followed by members of the phyla Actinobacteria, Firmicutes and Bacteroidetes. PCR-DGGE fingerprints of the gut samples showed that intestinal bacteria were affected by starvation, although the change was not significant.     

Comparison between cultured small-intestinal and fecal microbiotas in beagle dogs

The microbiota of the small intestine is poorly known because of difficulties in sampling. In this study, we examined whether the organisms cultured from the jejunum and feces resemble each other. Small-intestinal fluid samples were collected from 22 beagle dogs with a permanent jejunal fistula in parallel with fecal samples. In addition, corresponding samples from seven of the dogs were collected during a 4-week period (days 4, 10, 14, and 28) to examine the stability of the microbiota. In the jejunal samples, aerobic/facultative and anaerobic bacteria were equally represented, whereas anaerobes dominated in the fecal samples. Despite lower numbers of bacteria in the jejunum (range, 10(2) to 10(6) CFU/g) than in feces (range, 10(8) to 10(11) CFU/g), some microbial groups were more prevalent in the small intestine: staphylococci, 64% versus 36%; nonfermentative gram-negative rods, 27% versus 9%; and yeasts, 27% versus 5%, respectively. In contrast, part of the fecal dominant microbiota (bile-resistant Bacteroides spp., Clostridium hiranonis-like organisms, and lactobacilli) was practically absent in the jejunum. Many species were seldom isolated simultaneously from both sample types, regardless of their overall prevalence. In conclusion, the small intestine contains a few bacterial species at a time with vastly fluctuating counts, opposite to the results obtained for the colon, where the major bacterial groups remain relatively constant over time. Qualitative and quantitative differences between the corresponding jejunal and fecal samples indicate the inability of fecal samples to represent the microbiotas present in the upper gut.     

Routine fluorescence in situ hybridization in soil

The use of fluorescence in situ hybridization (FISH) to identify and enumerate soil bacteria has long been hampered by the autofluorescence of soil particles masking the bacterial signals and because the need of counting hundreds of bacteria in order to achieve statistically reliable data is time consuming. Recently, it was demonstrated that Nycodenz facilitates FISH in soil by concentrating bacteria on membrane filters and avoiding autofluorescent soil particles. We present a routine protocol for FISH in soil including the use of Nycodenz. The protocol allows fast and easy enumeration of hundreds of bacteria. We propose the use of silicon grease coated slides to treat in parallel seven samples per hybridization. Further, we developed a semi-automated approach for the enumeration of bacteria by implementing macros concatenating all steps of the image analyzes in the Image J software. Using Nycodenz, software-assisted bacterial counts statistically matched eye-counts of the same images and it was possible to count 880 DAPI stained bacteria per ten images. Fifty-five percent of these bacteria were co-labelled with the FISH probe specific for the Domain Bacteria, in accordance with recent FISH studies of bacterial populations in bulk soil. With a soil slurry protocol used for comparison, soil particles impaired automatic counts of the bacteria and FISH analysis, and only 88 DAPI stained bacteria per ten images could be counted by eye. With the Nycodenz protocol, 5 mM Na(2)EDTA used as an extractant increased the number of bacteria observed by 49%. In contrast, Tween 20 (1% or 5%) had no significant effect and increased the variability between the samples. Overall, the proposed procedure allows to process a high number of samples and to achieve a time efficient FISH characterization of soil bacterial communities.     

Microbial diversity in Maras salterns, a hypersaline environment in the Peruvian Andes

Maras salterns are located 3,380 m above sea level in the Peruvian Andes. These salterns consist of more than 3,000 little ponds which are not interconnected and act as crystallizers where salt precipitates. These ponds are fed by hypersaline spring water rich in sodium and chloride. The microbiota inhabiting these salterns was examined by fluorescence in situ hybridization (FISH), 16S rRNA gene clone library analysis, and cultivation techniques. The total counts per milliliter in the ponds were around 2 x 10(6) to 3 x 10(6) cells/ml, while the spring water contained less than 100 cells/ml and did not yield any detectable FISH signal. The microbiota inhabiting the ponds was dominated (80 to 86% of the total counts) by Archaea, while Bacteria accounted for 10 to 13% of the 4',6'-diamidino-2-phenylindole (DAPI) counts. A total of 239 16S rRNA gene clones were analyzed (132 Archaea clones and 107 Bacteria clones). According to the clone libraries, the archaeal assemblage was dominated by microorganisms related to the cosmopolitan square archaeon "Haloquadra walsbyi," although a substantial number of the sequences in the libraries (31% of the 16S rRNA gene archaeal clones) were related to Halobacterium sp., which is not normally found in clone libraries from solar salterns. All the bacterial clones were closely related to each other and to the gamma-proteobacterium "Pseudomonas halophila" DSM 3050. FISH analysis with a probe specific for this bacterial assemblage revealed that it accounted for 69 to 76% of the total bacterial counts detected with a Bacteria-specific probe. When pond water was used to inoculate solid media containing 25% total salts, both extremely halophilic Archaea and Bacteria were isolated. Archaeal isolates were not related to the isolates in clone libraries, although several bacterial isolates were very closely related to the "P. halophila" cluster found in the libraries. As observed for other hypersaline environments, extremely halophilic bacteria that had ecological relevance seemed to be easier to culture than their archaeal counterparts.     

Diversity of Culturable Bacteria Associated with Ascocarps of a Chinese White Truffle,Tuber panzhihuanense (Ascomycota)*

Culturable bacterial communities inhabiting ascocarps of Tuber panzhihuanense were investigated. Isolates obtained on tryptone soy agar (TSA) were screened with high performance capillary electrophoresis (HPCE) according to differences in size of 16S rDNA V3. Target isolates were identified by analysis of the whole length of 16S rDNA gene. The results revealed that the ascocarps of T. panzhihuanense harbored a great number of culturable bacteria which belonging to 20 species and 11 genera in 5 phyla. Most isolates (4968%) were affiliated to the γ Proteobacteria, dominated by Pseudomonas lurida. The second major subclass was α Proteobacteria (3742%), with Phyllobacterium and a nitrogen fixing bacterium Bradyrhizobium japonicum also occurring as dominant taxa. The remaining bacterial isolates contained members of Actinobacteria (322%) and Firmicutes (774%) of which Bacillus was the commonest bacterium. A novel Tuber associated culturable bacterium species, Terriglobus roseus, was isolated and detected for the first time in Tuber ascocarps.     

中国白块菌——攀枝花块菌子囊果内可培养细菌的多样性研究

对新近发现的块菌属一新种——攀枝花白块菌（Tuber panzhihuanense）子囊果中可培养细菌的多样性进行了研究。采用胰蛋白大豆培养基（TSA）对菌株进行分离。用毛细管电泳（HPCE）对所有获得的菌株的16S rDNA V3高变区进行筛选获得不同条带大小的菌株,对筛选出的菌株的16S rDNA进行测序,并进行细菌多样性分析和研究。结果显示,攀枝花块菌子囊果内可培养细菌在数量及种类上都表现出很高的多样性,所有细菌分属于5个门的11个属和20个种。在所分离到的变形菌门的细菌中,数量最多的菌株（4968%）属于γ Proteobacteria,其中假单胞菌属的Pseudomonas lurida为优势类群;其次为α Proteobacteria,占3742%,其中以固氮菌 Bradyrhizobium japonicum和Phyllobacterium spp.为优势类群。其余的菌株属于放线菌门（Actinobacteria） （322%）和厚壁菌门（Firmicutes） （774%）,厚壁菌门中以芽孢杆菌属（Bacillus）为代表菌群。酸杆菌门中的Terriglobus roseus（194%）首次从块菌中分离获得。     

Aquacultured Rainbow Trout (Oncorhynchus mykiss) Possess a Large Core Intestinal Microbiota That Is Resistant to Variation in Diet and Rearing Density

As global aquaculture fish production continues to expand, an improved understanding of how environmental factors interact in fish health and production is needed. Significant advances have been made toward economical alternatives to costly fishmeal-based diets, such as grain-based formulations, and toward defining the effect of rearing density on fish health and production. Little research, however, has examined the effects of fishmeal- and grain-based diets in combination with alterations in rearing density. Moreover, it is unknown whether interactions between rearing density and diet impact the composition of the fish intestinal microbiota, which might in turn impact fish health and production. We fed aquacultured adult rainbow trout (Oncorhynchus mykiss) fishmeal- or grain-based diets, reared them under high- or low-density conditions for 10 months in a single aquaculture facility, and evaluated individual fish growth, production, fin indices, and intestinal microbiota composition using 16S rRNA gene sequencing. We found that the intestinal microbiotas were dominated by a shared core microbiota consisting of 52 bacterial lineages observed across all individuals, diets, and rearing densities. Variations in diet and rearing density resulted in only minor changes in intestinal microbiota composition despite significant effects of these variables on fish growth, performance, fillet quality, and welfare. Significant interactions between diet and rearing density were observed only in evaluations of fin indices and the relative abundance of the bacterial genus Staphylococcus. These results demonstrate that aquacultured rainbow trout can achieve remarkable consistency in intestinal microbiota composition and suggest the possibility of developing novel aquaculture strategies without overtly altering intestinal microbiota composition.     

Comparison between Cultured Small-Intestinal and Fecal Microbiotas in Beagle Dogs

The microbiota of the small intestine is poorly known because of difficulties in sampling. In this study, we examined whether the organisms cultured from the jejunum and feces resemble each other. Small-intestinal fluid samples were collected from 22 beagle dogs with a permanent jejunal fistula in parallel with fecal samples. In addition, corresponding samples from seven of the dogs were collected during a 4-week period (days 4, 10, 14, and 28) to examine the stability of the microbiota. In the jejunal samples, aerobic/facultative and anaerobic bacteria were equally represented, whereas anaerobes dominated in the fecal samples. Despite lower numbers of bacteria in the jejunum (range, 10² to 10⁶ CFU/g) than in feces (range, 10⁸ to 10¹¹ CFU/g), some microbial groups were more prevalent in the small intestine: staphylococci, 64% versus 36%; nonfermentative gram-negative rods, 27% versus 9%; and yeasts, 27% versus 5%, respectively. In contrast, part of the fecal dominant microbiota (bile-resistant Bacteroides spp., Clostridium hiranonis-like organisms, and lactobacilli) was practically absent in the jejunum. Many species were seldom isolated simultaneously from both sample types, regardless of their overall prevalence. In conclusion, the small intestine contains a few bacterial species at a time with vastly fluctuating counts, opposite to the results obtained for the colon, where the major bacterial groups remain relatively constant over time. Qualitative and quantitative differences between the corresponding jejunal and fecal samples indicate the inability of fecal samples to represent the microbiotas present in the upper gut.     

The probiotic potential against vibriosis of the indigenous microflora of rainbow trout

The antibacterial properties of the indigenous microflora of rainbow trout ( Oncorhynchus mykiss Walbaum) and the potential use of inhibitory bacteria as fish probiotics were investigated. A total of 1018 bacteria and yeasts were isolated on tryptone soy agar (TSA) from skin, gills and intestine. Forty-five of these inhibited growth of the fish pathogenic bacterium Vibrio anguillarum in a well diffusion assay. The antagonism was most prominent among Pseudomonas spp., as 28 (66%) of the antagonistic bacteria belonged to this genus, despite constituting only 15% of the total tested flora. As pseudomonads are typically siderophore producers, chrome azurol S (CAS) agar was used as a semi-selective medium for isolation of antagonistic bacteria. On this medium, 75% of the iron-chelating strains were inhibitory to V. anguillarum . Eight strains out of a subset of 11 antagonists caused a 3–6 log unit reduction in the density of V. anguillarum [measured by polymerase chain reaction (PCR) detection in a most probable number (MPN) regimen] in a broth co-culture assay. Survival of rainbow trout infected with vibriosis was improved 13–43% by six out of nine antagonistic strains tested in vivo. All disease-protecting strains were pseudomonads, isolated from CAS plates, whereas two Carnobacterium spp. that were antagonistic in in vitro well diffusion assays did not alter the accumulated mortality of rainbow trout. The addition of live bacterial cultures to fish-rearing water may thus improve survival of the fish; however, in vitro antagonism could not completely predict an in vivo effect. Further studies on the underlying mechanism of activity are required to design appropriate selection criteria for fish probiotic bacteria.     

Community analysis of the bacterial assemblages in the winter cover and pelagic layers of a high mountain lake by in situ hybridization

The bacterial community structure in the winter cover and pelagic zone of a high mountain lake was analyzed by in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotide probes. Cells fixed on membrane filters were hybridized with a probe specific for the domain Bacteria as well as with probes for the alpha, beta, and gamma subclasses of the class Proteobacteria and the Cytophaga-Flavobacterium group. The fraction of bacteria detectable after hybridization with the bacterial probe EUB ranged from 40 to 81% of 4(prm1),6-diamidino-2-phenylindole (DAPI) counts. The bacterial assemblage varied considerably between and within different habitats (snow, slush, and lake water) but was in most cases dominated by members of the beta subclass (6.5 to 116% of bacteria detectable with probe EUB). The sum of bacteria hybridizing with group-specific probes was usually lower than the fraction detectable with probe EUB. Image analysis was used to characterize morphology and the size-specific biomass distribution of bacterial assemblages, which clearly separated the three habitats. Although the measured secondary production parameters and the fraction of 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride-reducing bacteria varied by more than an order of magnitude in the different slush and pelagic layers, detectability with the fluorescent probe EUB was constantly high. Physiological strategies of bacteria under nutrient limitation and at low temperatures are discussed in the context of the ribosome content of single cells. This study confirms the suitability of fluorescently labeled rRNA-targeted probes for the characterization of bacterial population structures even in oligotrophic habitats.     

The Toll-like receptors TLR2 and TLR4 do not affect the intestinal microbiota composition in mice

The interaction between intestinal epithelial cells and microbes is partly mediated by Toll-like receptors (TLRs). Sensing of Gram-positive and Gram-negative bacteria by TLR2 and TLR4, respectively, can result in immune system activation and in an exclusion of bacteria from the intestine. To test the impact of these TLRs on bacterial composition, germ-free TLR2/TLR4 double-knock out mice and the corresponding C57BL/10ScSn wild-type mice where associated with fecal bacteria from one single donor mouse. In addition, C3H/HeOuJ and BALB/c mice were used in this study. Fecal bacteria were monitored over 13 weeks with denaturing-gradient gel electrophoresis (DGGE). Colonic bacteria were enumerated by fluorescent in situ hybridization (FISH) and short-chain fatty acids (SCFA) were measured in caecal samples. No effect of the TLRs on intestinal microbiota composition and SCFA concentrations was observed. However, the microbiota composition as reflected by DGGE band patterns differed between C3H and BALB/c mice on the one hand and C57BL/10 mice on the other hand. Corresponding differences between the mouse strains were also observed in cecal propionic, valeric and i-valeric acid concentrations. No differences between the animals were observed in the numbers of bacteria detected by FISH. We conclude that genetic traits but not TLR2 and TLR4 have an impact on the intestinal microbiota composition.     

Yeast colonizing the intestine of rainbow trout (Salmo gairdneri) and turbot (Scophtalmus maximus)

Yeast were isolated from the intestine of farmed rainbow trout (Salmo gairdneri), turbot (Scophtalmus maximus), and free-living flat-fish (Pleuronectes platessa and P. flesus). The average number of viable yeasts recovered from farmed rainbow trout was 3.0 × 10³ and 0.5 × 10² cells per gram homogenized intestine for white and red-pigmented yeasts, respectively. The dominant species were Debaryomyces hansenii, Saccharomyces cerevisiae, Rhodotorula rubra, and R. glutinis. In 5 of 10 free-lving marine fish, > 100 viable yeast cells per gram intestinal mucus were recovered. Red-pigmented yeasts dominated and composed >90% of the isolates. Colonization experiments were performed by inoculating rainbow trout and turbot with fish-specific, isolated yeast strains and by examining the microbial intestinal colonization at intervals. Inoculation of experimental fish with pure cultures of R. glutinis and D. hansenii HF1 yielded colonization at a level several orders of magnitude higher than before the inoculation. Up to 3.8 × 10⁴, 3.1 × 10⁶, and 2.3 × 10⁹ viable yeast cells per gram intestine or feces were recovered in three separate colonization experiments. The high level of colonizing yeasts persisted for several weeks. The concentrations of yeasts in the tank water never exceeded 10³ viable cells per milliliter. No traces of fish sickness as a result of high yeast colonization were recorded during any of the colonization experiments. For periods of the experiments, the concentration of aerobic bacteria in the fish intestine was lower than the intestinal yeast concentration. Scanning electron microscopy studies demonstrated a close association of the yeasts with the intestinal mucosa. The mucosal colonization was further demonstrated by separating intestinal content, mucus, and tissue. All compartments were colonized by >10³ viable yeast cells per gram. No bacteria were detected on the micrographs, indicating that their affinity for the intestinal mucosa was less than that of the yeasts. Correspondence to: Thomas Andtid     

Composition, diversity, and origin of the bacterial community in grass carp intestine

Gut microbiota has become an integral component of the host, and received increasing attention. However, for many domestic animals, information on the microbiota is insufficient and more effort should be exerted to manage the gastrointestinal bacterial community. Understanding the factors that influence the composition of microbial community in the host alimentary canal is essential to manage or improve the microbial community composition. In the present study, 16S rRNA gene sequence-based comparisons of the bacterial communities in the grass carp (Ctenopharyngodon idellus) intestinal contents and fish culture-associated environments are performed. The results show that the fish intestinal microbiota harbors many cellulose-decomposing bacteria, including sequences related to Anoxybacillus, Leuconostoc, Clostridium, Actinomyces, and Citrobacter. The most abundant bacterial operational taxonomic units (OTUs) in the grass carp intestinal content are those related to feed digestion. In addition, the potential pathogens and probiotics are important members of the intestinal microbiota. Further analyses show that grass carp intestine holds a core microbiota composed of Proteobacteria, Firmicutes, and Actinobacteria. The comparison analyses reveal that the bacterial community in the intestinal contents is most similar to those from the culture water and sediment. However, feed also plays significant influence on the composition of gut microbiota.     

Gibel carp Carassius auratus gut microbiota after oral administration of trimethoprim/ sulfamethoxazole

Trimethoprim/sulfamethoxazole is widely used in the treatment of infectious diseases caused by bacterial pathogens in aquaculture. However, the practice of antibiotic administration can promote the emergence of resistant strains of bacteria and result in a wane in efficacy over time. The objective of this study was to assess the effect of oral treatment with trimethoprim/sulfamethoxazole on the gastrointestinal (GI) microbiota of healthy gibel carp and those affected with bacterial enteritis. By using denaturing gradient gel electrophoresis (DGGE), the changes in the predominant bacterial communities were directly depicted for the first time. The main findings were (1) Actinobacteria, Firmicutes and Proteobacteria were the predominant phyla in the healthy gibel carp intestine; (2) administration of antibiotics had a more profound impact on the intestinal microflora of healthy fish than of the diseased ones; and (3) Enterobacteriaceae might be one of the major drug-resistant bacteria in the gibel carp intestine. This study provides an insight into the effect of antibiotic treatment on the establishment and colonization of fish GI microbiota and speculates on some possible drug-resistant bacteria.     

The cutthroat trout Y chromosome is conserved with that of rainbow trout

Five genetic markers previously shown to be located on the sex chromosomes of rainbow trout (Oncorhynchus mykiss) were tested for linkage with the sex locus of Yellowstone cutthroat trout (Oncorhynchus clarki bouvieri) in a genetic cross created from a rainbow x cutthroat male hybrid. We show that the sex locus of both rainbow and cutthroat trout is on the same homologous linkage group. Fluorescence in situ hybridization (FISH) using a probe for the microsatellite marker Omm1665, which maps close to the sex locus of Yellowstone cutthroat trout, was used to identify the Y chromosome of cutthroat trout in the hybrid. The Y chromosome of cutthroat trout is sub-telocentric and lacks a DAPI band found on the short arm of the Y chromosome of some rainbow trout males.     

Association of Flavobacterium psychrophilum strains with intestinal explants of rainbow trout Oncorhynchus mykiss

Flavobacterium psychrophilum is an important pathogen in rainbow trout Oncorhynchus mykiss. The portal of entry for F. psychrophilum is not well known. In this study, the role of the intestine as a colonization site for F. psychrophilum was determined. For this purpose, the ability of a high (Dubois) and a low (99/10A) virulence strain of F. psychrophilum to adhere to intestinal explants of rainbow trout was evaluated. After incubation, samples of the gut were examined bacteriologically, histologically and by electron microscopy. The number of gut-associated F. psychrophilum bacteria was significantly higher for the Dubois than for the 99/10A strain. Histological samples clearly showed numerous bacteria of the high virulence strain associated with the intestinal tissue as opposed to only a few bacteria of the low virulence strain. Additionally, extensive exfoliation of intestinal epithelium was noted after incubation with the high virulence strain, but less with the low virulence strain. These findings were confirmed using scanning electron microscopy and suggest that the intestinal epithelium might represent an important site for colonization of the F. psychrophilum strain and may act as a portal of entry for high virulence F. psychrophilum.