Variations of the intestinal gut microbiota of farmed rainbow trout, Oncorhynchus mykiss (Walbaum), depending on the infection status of the fish.

AIMS: The aim of the present study was to investigate the composition of the intestinal microbiota during the acute stage of a bacterial infection to understand how dysbiosis of the gut may influence overall taxonomic hierarchy and diversity, and determine if there exists a bacterial taxon(s) that serve as markers for healthy or diseased rainbow trout (Oncorhynchus mykiss). METHODS AND RESULTS: From July to September 2015, 29 specimens of 3-year-old (an average weight from 240·9 ± 37·7 to 850·7 ± 70·1 g) rainbow trout O. mykiss were studied. Next-generation high-throughput sequencing of the 16S ribosomal RNA genes was applied to stomach and intestinal samples to compare the impact of infection status on the microbiota of rainbow trout O. mykiss (Walbaum) from the northwest part of Eurasia (Karelian region, Russia). The alpha diversity (Chao1, Simpson and Shannon index) of the microbial community of healthy rainbow trout was significantly higher than in unhealthy fish. The greatest contribution to the gut microbial composition of healthy fish was made by OTU's belonging to Bacillus, Serratia, Pseudomonas, Cetobacterium and Lactobacillus. Microbiota of unhealthy fish in most cases was represented by the genera Serratia, Bacillus and Pseudomonas. In microbiota of unhealthy fish there were also registered unique taxa such as bacteria from the family Mycoplasmataceae and Renibacterium. Analysis of similarities test revealed the significant dissimilarity between the microbiota of stomach and intestine (P ≤ 0·05). CONCLUSIONS: A substantial finding was the absence of differences between microbial communities of the stomach and intestine in the unhealthy groups if compared with healthy fish. SIGNIFICANCE AND IMPACT OF THE STUDY: These results demonstrated alterations of the gut microbiota of farmed rainbow trout, O. mykiss during co-infections and can be useful for the development of new strategies for disease control programs.

Diet and other environmental factors shape the bacterial communities of fish gut in an eutrophic lake.

AIMS: The aim of this work was to study the gut microbial diversity from eight species of wild fish with different feeding habits, digestive physiology (gastric vs agastric) and provide comparative structural analysis of the microbial communities within their environment (food items, water, sediments and macrophytes). METHODS AND RESULTS: The microbiota of fish gut and their prey items were studied using next generation high-throughput sequencing of the 16S ribosomal RNA genes. A scatter plot based on PCoA scores demonstrated the microbiota formed three groups: (i) stomach and intestinal mucosa (IM), (ii) stomach and intestinal content (IC), and (iii) prey and environment. Comparisons using ANOSIM showed significant differences among IC of omnivorous, zoobenthivorous, zooplanktivorous-piscivorous fishes (P ≤ 0·1). No significant difference was detected for mucosa from the same groups (P > 0·1). CONCLUSIONS: Neither the interspecies differences in fish diet nor their phylogenetic position had any effect on the microbiome of the IM, but diet did influence the composition of the microbiota of the IC. SIGNIFICANCE AND IMPACT OF THE STUDY: The data demonstrate that fish harboured specific groups of bacteria that do not completely reflect the microbiota of the environment or prey.

Inactivation of proteolytic enzymes by cestodes.

Inhibitor activity of cestodes from intestines of different hosts (sea birds, salt-water fish, and freshwater fish) was investigated. Alcataenia larina, Arctotaenia tetrabothrioides, Tetrabothrius erostris, T. minor, Wardium cirrosa, Bothriocephalus scorpii, Eubothrium rugosum, and Triaenophorus nodulosus were able to inhibit the activity of the commercial trypsin and activity of proteinase homogenates of the intestinal mucosa of the hosts. It was suggested that the inhibitor produced by the cestodes is specific for trypsin and protects them from the digestive enzymes of the host.

[Characteristics of the Effect of Cestodes Parasitizing the Fish Intestine on the Activity of the Host Proteinases].

The activity and spectrum of proteinases in the intestines of host fishes change upon infestation with cestodes. Serine proteinases are found to make a greater contribution to the total proteolytic activity. The reduction of proteolytic activity is associated with adsorption of the enzymes of the host on the surface of cestodes, and the increase in the activity is caused by the injury of the intestinal mucosa by the attachment apparatuses of cestodes. The inhibition of proteainase activity indicates the possible participation of microbiota enzymes in protein hydrolyses.

A comparative study on microbiota from the intestine of Prussian carp (Carassius gibelio) and their aquatic environmental compartments, using different molecular methods.

AIMS: The aim of this study was to evaluate, via various molecular methods, the possible correlations between microbial community structure of Prussian carp and the environmental compartments of their habitat. METHODS AND RESULTS: Microbial communities in the intestine and environmental compartments were studied using PCR-screening, cloning and next-generation high-throughput sequencing of the 16S ribosomal RNA genes. The 16S rDNA metagenomic sequencing showed higher bacterial diversity in comparison with clone libraries, while group-specific PCR showed positive detection of nine bacteria phyla. Proteobacteria, Bacteroidetes, Firmicutes, Cyanobacteria and Actinobacteria were most abundant both in the intestine and habitat environments. The comparative analyses reveal that the bacterial community in the Prussian carp intestine is most similar to that identified from the chironomid. CONCLUSIONS: This study demonstrated some differences between molecular methods and showed advantages and limitations associated with them. These differences have the potential to reduce bias in results obtained from analysis of the community structure. The advantages of each molecular technique can be used for a better understanding of microbial diversity. The microbiota of Prussian carp intestine is most similar to those from the chironomids. SIGNIFICANCE AND IMPACT OF THE STUDY: We investigated the diversity of the intestinal microbiota in an economically important aquaculture species, the Prussian carp (Carassius gibelio). The results provide significant information to discuss possible functions of these bacteria for further understanding of Prussian carp health.

Feeding habits and ontogenic changes in digestive enzyme patterns in five freshwater teleosts.

Feeding habits and the activity of digestive enzymes (total alkaline proteases, α-amylase and lipase) from dace Leuciscus leuciscus, roach Rutilus rutilus, Prussian carp Carassius auratus gibelio, perch Perca fluviatilis and pikeperch Sander lucioperca fry were studied in the Malye Chany Lake-Kargat Estuary (western Siberia, Russia). The diet of fry from all studied species was mainly composed of chironomid larvae and zooplanktonic organisms (i.e. cladocera and copepoda), whereas carnivorous species such as P. fluviatilis and S. lucioperca also preyed on fry from other fishes while detritus and microalgae were also important in the diet of ommivorous species. When comparing diet similarity (Sørensen-Dice index, Q(S)) among fry at different stages of development, both omnivorous and carnivorous species showed a high level of similarity (0.67 < Q(S) < 0.89 and 0.73 < Q(S)< 0.89, respectively). Diet similarity values were in agreement with the overall digestive activity profile analysed by cluster analysis. Diet similarity suggested potential trophic competition when zooplanktonic and benthic prey began to decline towards autumn. The analysis of pancreatic digestive enzymes revealed a correlation among their activities and fry feeding habits with α-amylase:total proteases (A:P) values higher than 1 in omnivorous species and lower (A:P ≤ 1) in carnivorous species.

[Effect of Caryophyllaeus laticeps (Cestoda, Caryophyllidea) upon activity of digestive enzymes in bream].

The activities of the main digestive hydrolases were comparatively studied in bream infected and noninfected with cestodes Caryophyllaeus laticeps (Pallas, 1781). It was shown that enzyme activities are distributed in the fish intestine in an irregular manner; the gradient of protease and lipase activities along the gut is presented. Following the infection of bream by cestodes, the activities of the studied enzymes decreased and the percentages of activities of various proteinase subclasses changed. No relation between the distribution of worms along the intestine and the levels of activities of digestive hydrolases was revealed.

[The contribution of carbohydrases produced by symbiotic microflora to the digestive processes in cestodes and their host fishes].

The bacteria capable of producing the enzymes hydrolyzing carbohydrates of various degrees of complexity (from starch to sucrose) were found to be associated with the intestinal mucosa of fishes and tegument of cestodes. Presence of the bacteria displaying the sucrase activity is especially important for macroorganisms, as bacteriogenous glucose can be used by all members of the arising community. The greatest contribution to the hydrolysis of carbohydrates (both in host and parasite) is obviously made by those microorganisms which are more closely connected with the digestive-transport surfaces and are hardly removable from the intestines by peristalsis. The levels of total amylolytic activity of bacteriogenous enzymes and activity of their a-amylase under the experimental conditions are comparable to those of the enzymes involved in membrane digestion of the host and parasite, which can be evidence of the significant contribution of enzymes produced by symbiotic microflora to the digestive processes in macroorganisms.

[Symbiotic microflora in fishes of different ecological groups].

Published data on the taxonomic composition of the intestinal bacterial flora in fishes living under different conditions (marine and freshwater fishes) and differing in the type of feeding are reviewed. The results confirm the conclusion that the composition of intestinal microflora differs depending on fish ecology. A limited number of bacterial taxa found in the intestines of some fish species may be evidence not only for a low diversity of these bacteria but also for insufficient knowledge about them. Considerable differences in the composition of intestinal microflora in marine and freshwater fishes are described.

[The role of alpha-amylase of symbiotic microflora in digestion by lower cestodes and their fish hosts].

The symbiotic microflora associated with the digestive-transport surfaces of the pike intestine and the parasitic cestodes Triaenophorus nodulosus proved capable of the initial stages of carbohydrate hydrolysis mediated by alpha-amylase. The products of hydrolysis by alpha-amylase can be used by both the host and the parasite, which decreases energy expenditures of the macroorganisms. The levels of the bacterial alpha-amylase activity are comparable to those of the analogous enzyme absorbed on the mucosa of the intestine and on the cestode tegument, which indicates a considerable contribution of enzymes of the symbiotic microflora to digestion by the host and the parasite. Apparently, this contribution depends on the fish diet.

[Final stages of carbohydrate metabolism in a cestode Eubothrium rugosum (Cestoda: Pseudophyllidea)]. / Kharakteristika zakliuchitel'nykh étapov uglevodnogo obmena u tsestody Eubothrium rugosum (Cestoda: Pseudophyllidea).

Characteristics of final stages of carbohydrate metabolism in a cestode Eubothrium rugosum from the burbot intestine have been studied. It is found out that in the course of in vitro incubation the worms produce lactic acid to the environment with a concomitant acidification of the medium. The rate of pH changes and lactate production eventually fell during the incubation. The medium pH being the result of helminth's metabolism varies within the normal physiological limits of the host intestine pH and pH optima for its basic digestive enzymes.

[Physiological specificity of the interaction between Traenophorus nodulosus (Cestoda) and its host--fishes]. / Fiziologicheskaia spetsifika vzaimootnoshenii mezhdu Triaenophorus nodulosus (Cestoda) i ikh khoziaevami--rybami.

The influence of the T. nodulosus plerocercoid invasion on the liver of the intermediate fish hosts (perch) was studied. The water, protein, glycogen and glucose contents was examined in adult worms inhabiting the intestine of definitive hosts (pike), the rate of glucose accumulation along the strobile of parasite and an ability of adult worms to hydrolyze the main nutrients was also investigated. As a result of the plerocercoid infection, the glucose and glycogen contents in the liver of perch juveniles decreased, while the wet weight of liver increased. It was found, that the water and protein contents from the anterior to posterior section of strobile in T. nodulosus adults was not significantly different, while the glucose contents and the rate of its accumulation decreased. The data confirming the possibility to realize the final stages of protein and carbohydrate hydrolisis with the participation of membrane digestion on the surface of helminths and proving the ability of cestodes. The problems of relations of T. nodulosus with intermediate and definitive hosts at the level of the whole organisms are discussed.

[The parasite-host relations in the system of Ligula intestinalis (L.) (Cestoda, Pseudophyllidae)--the bream]. / Nekotorye aspekty parazito-khoziainnykh otnoshenii v sisteme Ligula intestinalis (L.) (Cestoda, Pseudophyllidae)--leshch.

Proteins and reducing sugars were assayed. The total proteolytic and amylolytic activities were determined in flushes of body cavity of the breams infected and noninfected by plerocercoids Ligula intestinalis. Infection with L. intestinalis markedly affected the composition of the medium and the activity of enzymes functioning in the body cavity of the bream: protein content increased, total proteolytic activity remained practically unchanged, while the content of reducing sugars and total amylolytic activity decreased. The strength of fixation of amylolytic enzymes on the surface of L. intestinalis tegument was tested. The dynamics of desorption of these enzymes from the tegument of plerocercoids and adult cestodes parasitizing in the fish intestine. The contents of protein and glycogen do not depend on the cestode length and strobile region.

Digestion and digestive-transport surfaces in cestodes and their fish hosts.

The structural and functional organization of digestive-transport surfaces in some lower cestodes and their fish hosts was studied. It has been shown that the ultrastructure of cestode microtriches and fish enterocyte microvilli being the basis of membrane-linked digestion is quite similar. These organelles increase the digestive-transport surfaces both in helminths and fishes. However, the hydrolytic enzyme activity in helminths is usually 2-4 times lower than that of the fishes. Desorption (adsorption) characteristics of various hydrolases in helminths and fishes are also different. In helminths the easily desorbed fraction of each enzyme is always more abundant than in fishes. In contrast, the intensity of transport processes in helminths is higher when compared with fishes. The adaptation of digestive-transport surfaces and enzyme systems to feeding conditions is discussed.

[Iso-osmotic regulation in cestodes from freshwater fishes]. / Izoosmoticheskaia reguliatsiia u tsestod iz presnovodnykh ryb.

The content of sodium, potassium, calcium, and magnesium ions was determined in cestodes Ligula intestinalis, Eubothrium rugosum, and Triaenophorus nodulosus. In freshwater fishes the total concentration of Na, K, Ca, Mg ions in cestode tissue water is practically similar to that of fishes muscles and in serum. It was suggested that examined cestodes have isoosmotic regulation of the water-salt balance.

[The hydrolysis and transport of sugars in the cestode Caryophyllaeus laticeps and in the intestines of its host, the bream]. / Gidroliz i transport sakharov u tsestody Caryophyllaeus laticeps i v kishechnike ee khoziaina--leshcha.

A comparative study of hydrolysis and transport of carbohydrates by the cestode C. laticeps and in the intestine of its host, bream, has revealed a considerable influence of the host's enzymes on the characteristics of the parasite's enzyme system. The cestodes have been found to possess effective transport systems of glucose absorbing it in an active way while in the intestine of bream during the glucose absorption prevail the mechanisms of simple and easy diffusion.