Diversity of microbial communities of the intestinal mucosa and intestinal contents of fish from Lake Chany (Western Siberia)

The diversity of microbial communities associated with the intestinal mucosa and the intestinal contents of fish with different nutritional profiles have been investigated. Differences in the intestinal microbiota of fish with different dietary preferences have been revealed using group-specific primers for the main large taxa of bacteria. Representatives of the phyla Planctomycetes, Vernicomicrobia, and Cyanobacteria have been detected in the intestinal contents of all investigated nonpredatory (crucian carp, roach, and dace) and predatory (pikeperch, pike, and perch) fish. Representatives of the phylum Firmicutes were detected in the intestinal mucosa and intestinal contents of predatory fish species, but not in those of the nonpredatory fish.     

Histological observation of the urogenital papillae in the bi-directional sex-changing gobiid fish, Trimma okinawae

The gobiid fish Trimma okinawae changes its sex bi-directionally according to its social status. Morphological changes in the urinogenital papillae (UGP) of this fish have been reported during sex change. However, there have been no detailed observations of such changes. Here, we histologically examined the UGP structure of male- and female-phase fish. UGPs of fish in female and male phase contained both oviducts and sperm ducts. Both ducts were coalesced into one duct within the posterior region of the UGP. Female-phase fish had many longitudinal folds in the hypertrophied tunica mucosa of the oviduct, which was found to be responsible for the transport of eggs and the removal of follicular cells from the oocyte. In contrast, male-phase fish had an immature oviduct and a mature sperm duct in the UGP. In the male-phase fish, the co-existence of spermatozoa and fibrillar secretions was observed in the sperm duct during spermiation.     

Effect of pH on the activity of proteinases in intestinal mucosa,chyme, and microbiota of fish from the Cuciurgan reservoir

Proteolytic activities of the intestinal mucosa, chyme, and enteral microbiota have been studied in a wide range of pH values in five fish species from the Cuciurgan reservoir (Moldova). Differences in pH dependence of the intestinal proteinase activity of fish are determined by their feeding type. The maximum activity of proteinases is found in the pumpkinseed Lepomis gibbosus. The minimum activity of proteinases has been demonstrated by the zander Zander lucioperca. The pH optimum of the mucosa and chyme in all fish species (except for the European perch Perca fluviatilis) is 10. The pH optimum of the intestinal microflora varies from 6.0 (in the common carp Cyprinus carpio) to 10 (in the crucian carp Carassius carassius), whereas that in the perch from the Cuciurgan and Rybinsk reservoirs is 7. The majority of fish species, mostly Zander lucioperca and Lepomis gibbosus, are characterized by high proteniase activity of the microbiota, in the pH ranging from 6 to 9. It is assumed that proteinases in the enteral microbiota of fish are able to make up for the relatively low activity of those synthesized by their digestive system in the range of low pH values.     

Impact of polychlorinated biphenyls on the activity of intestinal proteinases and carbohydrases in juvenile roach Rutilus rutilus (L.)

The chronic effects that polychlorinated biphenyls (PCBs) have on the activities of proteinases and carbohydrates in intestinal mucosa and chyme in juvenile roach Rutilus rutilus (L.) have been studied for the first time. Upon consuming food with PCB 50.8 ng/g wet weight for 218 days, the exposition of fingerlings in aquariums with dirt bottoms (contents PCB 425.6 ng/g dry weight) reduces the proteolytic activity of mucosa to a greater degree than amylolytic activity; the activity of sucrase changes in different directions. In 2-year-old fish receiving PCB only with food (50.8 ng/g wet weight bottom), proteolytic and amylolytic activities of the mucosa are reduced 18 and 35% in comparison with the control (52 days); the sucrose activity is reduced 13% (52 days) and 22% (169 days). In most cases the relation of amylolytic activity to proteolytic activity in fish of the experimental groups is less than in the control. The enzyme activities in chyme change in different directions, depending on the terms and conditions of the experiment.     

Exocytotic release of vasoactive intestinal polypeptide and serotonin from mucosal nerve fibres and endocrine cells of the intestine of the goldfish (Carassius auratus) and the tilapia (Oreochromis mossambicus): an ultrastructural study

In earlier studies we determined the effect, presence and ultrastructure of vasoactive intestinal polypeptide (VIP) and 5- hydroxytryptamine (5-HT)-containing nerve fibres in the tilapia and goldfish intestinal mucosa. 5-HT-labelled varicosities were found close to the epithelial cells; however, synaptic membrane specializations have never been observed. VIP-like immunoreactive nerve fibres appear to be located less frequently close to the goldfish epithelium, as in the tilapia intestine, in which the distance between the VIP- or 5-HT-labelled varicosities and the epithelial cells was also rather large (more than 2 μm). To establish a possible role of VIP and 5-HT as neurotransmitters involved in the regulation of fish intestinal epithelium, both electron microscopical and immunoelectron microscopical methods were used to visualize the release of 5-HT and VIP from nerve fibres. We found exocytoses from VIP-ergic and serotonergic varicosities in the muscle layers of both fish. Directly underneath the intestinal epithelium of the goldfish, it was demonstrated that 5-HT could be released from scarce varicosities. The release of 5-HT in the tilapia intestinal mucosa could only be observed from endocrine cells     

Exocytotic release of vasoactive intestinal polypeptide and serotonin from mucosal nerve fibres and endocrine cells of the intestine of the goldfish (Carassius auratus) and the tilapia (Oreochromis mossambicus): an ultrastructural study

In earlier studies we determined the effect, presence and ultrastructure of vasoactive intestinal polypeptide (VIP) and 5- hydroxytryptamine (5-HT)-containing nerve fibres in the tilapia and goldfish intestinal mucosa. 5-HT-labelled varicosities were found close to the epithelial cells; however, synaptic membrane specializations have never been observed. VIP-like immunoreactive nerve fibres appear to be located less frequently close to the goldfish epithelium, as in the tilapia intestine, in which the distance between the VIP- or 5-HT-labelled varicosities and the epithelial cells was also rather large (more than 2 μm). To establish a possible role of VIP and 5-HT as neurotransmitters involved in the regulation of fish intestinal epithelium, both electron microscopical and immunoelectron microscopical methods were used to visualize the release of 5-HT and VIP from nerve fibres. We found exocytoses from VIP-ergic and serotonergic varicosities in the muscle layers of both fish. Directly underneath the intestinal epithelium of the goldfish, it was demonstrated that 5-HT could be released from scarce varicosities. The release of 5-HT in the tilapia intestinal mucosa could only be observed from endocrine cells     

Effects of temperature,pH, and heavy metals (copper,zinc) upon proteinase activities in digestive tract mucosa of typical and facultative piscivorous fish

Species-specific differences and effects of natural (temperature, pH) and anthropogenic (zinc, copper) factors upon activities of casein-and hemoglobinlytic proteinases have been studied in the gut mucosa in pike (Esox lucius), zander (Stizostedion lucioperca), burbot (Lota lota) and perch (Perca fluviatilis). It is revealed that the intestine mucosa proteinase activities differ considerably from those ones in the fish stomach mucosa and that studied factors affect these activities to different extents. The stomach mucosa proteinases are most tolerant to the effects of studied factors, which is especially true for pike. The least tolerant are the intestine caseinlytic proteinases (except for the enzymes in perch). The effects of temperature, pH, and heavy metals depend on fish species, factors’ combinations (mono-factor, bi-factor, or multi-factor action) and on factor variability.     

Evidence for somatostatin,gastrin and pancreatic polypeptide-like substances in the mucosa cells of the gut in fishes with and without stomach

Gastrin, pancreatic polypeptide and somatostatin immunoreactive cells in the gut of two fish with stomachs (perch and catfish) and a stomachless fish (carp) were studied by immunocytochemistry. In the gastric mucosa of perch and catfish, cells showing gastrin and somatostatin-like immunoreactivity are found, scattered among the surface mucous cells and mucous neck cells. No pancreatic polypeptide (P.P.) immunoreactive cells are detected in the gastric mucosa. Cells showing gastrin and P.P.-like immunoreactivity are observed in the intestinal mucosa of perch, catfish and carp. In this location no somatostatin immunoreactive cells are found.     

灌喂氧化鱼油使草鱼肠道黏膜胆固醇胆汁酸合成基因通路表达上调

以草鱼(Ctenopharyngodon idella)为试验对象, 灌喂氧化鱼油7d后, 采集肠道黏膜组织并提取总RNA, 采用RNA-seq方法, 进行氧化鱼油组和正常鱼油组草鱼肠道黏膜基因差异表达水平、基因注释和IPA基因通路分析, 并测定了血清中胆固醇、甘油三酯、高密度脂蛋白和低密度脂蛋白含量. 研究结果显示, 草鱼肠道黏膜在受到氧化鱼油损伤后, 胆固醇和胆汁酸生物合成通路代谢酶、调节胆固醇和胆汁酸合成或转运的代谢酶或蛋白基因差异表达, 部分基因差异表达达到显著性上调水平. 实验结果表明, 草鱼肠道黏膜具备完整的乙酰辅酶A胆固醇胆汁酸的合成代谢基因通路. 肠道黏膜在受到氧化鱼油损伤后, 以乙酰辅酶A为原料的胆固醇生物合成代谢通路基因表达增强, 胆固醇由细胞外转运到细胞内的逆转运途径基因通路表达下调, 胆固醇由细胞内向细胞外转运基因通路表达上调; 以胆固醇为原料的胆汁酸经典合成代谢途径基因通路表达上调, 而胆汁酸的补充合成途径基因表达下调. 在灌喂氧化鱼油后, 血清胆固醇、低密度脂蛋白、甘油三酯含量分别增加了28.84%、29.56%和12.13%, 而高密度脂蛋白含量下降了8.15%.       

氧化鱼油对草鱼肠道黏膜抗氧化应激通路基因表达水平的影响

为了探讨氧化鱼油对草鱼肠道黏膜损伤后, 参与抗氧化应激的基因通路及其通路基因表达活性的变化,以草鱼为试验对象, 灌喂氧化鱼油7d后, 采集肠道黏膜组织并提取总RNA, 采用RNA-seq方法, 进行了氧化鱼油组和正常鱼油组草鱼肠道黏膜基因注释、IPA基因通路分析和基因表达活性差异分析。结果显示, 组织切片观察发现氧化鱼油导致草鱼肠道黏膜出现严重的损伤; 肠道黏膜中具有较为完整的Keap1-Nrf2-ARE基因调控通路。肠道黏膜在受到氧化鱼油的氧化损伤作用后, 激活了细胞的抗氧化损伤保护机制, 使NRF2介导的氧化应激反应通路基因差异表达显著性地上调, 并导致了下游的GSH/GSTs通路基因差异表达显著性上调, 促进了GSH的生物合成和GSTs的抗氧化作用; 导致Keap1-Nrf2-ARE信号通路下游的热休克蛋白和泛素-蛋白酶体通路基因差异表达显著性上调, 清除受损伤蛋白质, 保护细胞结构完整性。研究表明, 上述三类抗氧化应激通路构成了对肠道黏膜损伤细胞、损伤蛋白质的降解系统和清除系统, 显示其对肠道黏膜组织和黏膜细胞的保护、修复发挥了重要的作用。     

Effects of fasting on plasma lipids and cholesterol esterification in plasma, liver and intestinal mucosa in the char (Salmo alpinus L.)

1. CoA-dependent cholesterol esterification, measured as esterification of 3H-cholesterol, was demonstrated in homogenates of liver and intestinal mucosa of the char (Salmo alpinus L.). 2. Plasma concentration of triacylglycerols, unesterified and total cholesterol were significantly reduced to 43, 58 and 72% of the control values, respectively, after 6 weeks fasting. 3. The rate of cholesterol esterification in plasma and liver homogenate was significantly lower in the fasted fish compared to the controls, but the esterification activity in the homogenate of intestinal mucosa increased twofold in the fish fasted for 6 weeks.     

Diurnal Dynamics of Activity of Peptidases at Early Ontogenesis of Bleak <Emphasis Type="Italic">Alburnus alburnus</Emphasis> (L.) from Coastal Shallows of the Rybinsk Reservoir

The diurnal dynamics of the activity of peptidases in fish larvae by the example of bleak Alburnus alburnus (L.) as the dominant species in the open and protected shallows of the Rybinsk Reservoir has been studied for the first time. Two peaks of enzyme activity in the intestinal mucosa of fish larvae are revealed: morning and night. The data on species composition and daily dynamics of number of fish larvae in the shallows of different types are presented.     

Histamine in endocrine cells in the stomach

Summary Antibodies to histamine were used to examine the localization of the amine in cells of the stomach and upper small intestine of a great variety of species, including cartilaginous and bony fish, amphibia, reptiles (lizard), birds (chicken) and a large number of mammals. In all species gastric histamine was localized in endocrine cells (invariably found in the epithelium) and mast cells (usually with an extra-epithelial localization). The endocrine cells were identified as such by immunostaining with antibodies to chromogranin A and the mast cells were identified by toluidine blue staining. Histamine-immunoreactive endocrine cells were found almost exclusively in the acid-producing part of the stomach; only rarely were such cells observed in the pyloric gland area. They were fairly numerous in the gastric mucosa of the two subclasses of fish as well as in the amphibia and reptile species studied. Here, the majority of the histamine-immunoreactive endocrine cells seemed to have contact with the gastric lumen (open type cells) and were located in the surface epithelium (certain fish only) or together with mucous neck cells at the bottom of the pits. In the chicken, histamine-immunoreactive endocrine cells were numerous and located peripherally in the deep compound glands. They were without contact with the lumen (closed type) and had long basal extensions (paracrine appearance), running close to the base of the oxyntico-peptic cells. In mammals, the number of histamine-immunoreactive endocrine cells in the stomach varied greatly. They were particularly numerous in the rat and notably few in the dog, monkey and man. In all mammals, the histamine-immunoreactive endocrine cells were of the closed type and located basally in the oxyntic glands. They often had a paracrine appearance with long basal processes. Histamine-storing mast cells, finally, were few in both subclasses of fish as well as in the amphibian species and in the lizard. They were fairly numerous in chicken proventriculus (beneath the surface epithelium), few in the oxyntic mucosa of mouse, rat and hamster, moderate in number in hedge-hog, guinea-pig, rabbit, pig and monkey, and numerous in cat, dog and man. In the oxyntic mucosa of the latter three species mast cells sometimes seemed to have an intraepithelial localization which made their distinction from endocrine cells difficult. In newborn cats (1–3 days old) in human foetuses (17–24 weeks gestational age) mast cells were relatively few in the gastric mucosa and the histamine-containing endocrine cells were easier to demonstrate as a consequence. Patients with achlorhydria (and pernicious anemia) or suffering from hypergastrinemia due to gastrinoma had a greatly increased number of histamine-storing endocrine cells in the oxyntic mucosa compared with normal individuals.     

Influence of various concentrations of phenol and its derivatives on the activity of fish intestinal peptidases

The effects of phenol and its derivatives (4-chlorophenol, 4-nirophenol and 2,4-dinitrophenol) on the activity of peptidases in five species of fish differing in feeding modes have been studied. The power of the effects depends on the fish species and localization of the enzyme (mucosa or chyme). As a rule, in the in vitro conditions the phenols in concentrations of 0.06?0.5 μM/L considerably decreases activities of the intestinal peptidases in bream; silver bream; and, especially, in pike. In some cases, phenol and its derivatives induce an inconsiderable increase in peptidase activities. In zander and perch, in fact, no changes in the enzymatic activity under impact of phenols are observed. It is suggested that the effect of phenol and its derivatives depends to a high extent on the species specificity of peptidases: in the fish of fam. Percidae, the enzymes are relatively tolerant; in fam. Cyprinidae and Esocidae, they are sensitive to the studied toxicants.     

Effect of thyroid hormones on activity dynamics of enzymes of intestinal mucosa of juvenile roach <Emphasis Type="Italic">Rutilus rutilus</Emphasis>

The effect of thyroid hormones on activity dynamics of enzymes (proteinases and glycosidases) of intestinal mucosa of juvenile roach Rutilius rutilus was investigated. Application of substances increasing and decreasing the level of thyroid hormones in blood plasma significantly influences the growth rate and the activity of proteinases and glycosidases functioning in the intestinal mucosa. In most cases, the activity level of trypsin-like proteinases and the activity of glycosidases in the fish exposed to triiodothyronine were significantly higher than in the control. The activity level of chymotrypsin-like proteinases in fish form the group with exposure of exogenous triiodothyronine only in the end of the experiment surpassed the values of this parameter in the control fish. In the fish developing at deficiency of thyroid hormones, the growth rate and proteinases activity were significantly lower in comparison with the control.     

Influence of temperature acclimatization on the temperature-dependence and ouabain-sensitivity of goldfish intestinal adenosine triphosphatase

1. Homogenates of goldfish intestinal mucosa were separated into various fractions by differential centrifugation. Both adenosine-triphosphatase and beta-glycerophosphatase activities were found to be concentrated mainly in a membrane fraction which sedimented after 1200000g-min. 2. This membrane adenosine-triphosphatase system was activated by Na(+)+K(+) and inhibited by ouabain. 3. The ouabain-sensitive adenosine-triphosphatase activity was high and the ouabain-insensitive activity low in membrane fractions prepared from fish acclimatized previously to 8 degrees . The opposite was true for fish acclimatized to 30 degrees . 4. The Arrhenius plots of ouabain-sensitive and ouabain-insensitive adenosine-triphosphatase activities, measured from 5 degrees to 30 degrees , showed discontinuities at incubation temperatures that varied with the previous acclimatization temperature of the fish. 5. It is considered that modification of the membrane adenosine-triphosphatase system in goldfish intestinal mucosa may serve to regulate Na(+) transport at different environmental temperatures.     

Apolipoprotein A-I, an antimicrobial protein in Oncorhynchus mykiss: evaluation of its expression in primary defence barriers and plasma levels in sick and healthy fish

Antimicrobial proteins and peptides play an important role in the primary defence barriers in vertebrates and invertebrates. In a previous study it was shown that high-density lipoprotein (HDL) and its major apolipoproteins, ApoA-I and ApoA-II display antimicrobial activity in the carp (Cyprinus carpio L.). The aim of this study was to evaluate if ApoA-I conserves this defensive function in a salmonid fish like the rainbow trout, in spite of the low level of primary sequence conservation between fish ApoA-I. Here it is shown that trout ApoA-I displays an antimicrobial activity in the micromolar range against Gram positive and Gram negative bacteria, including some fish pathogens. In addition, its expression was also demonstrated by immunohistochemistry and RT-PCR in epidermis, gills and intestinal mucosa, which constitute the main primary defence barriers in fish. Finally, no significant difference in the hepatic expression and plasma levels of this abundant apolipoprotein was found in groups of healthy and diseased fish, in clear contrast with mammals where ApoA-I have been considered a negative acute phase reactant. These findings suggest that ApoA-I could constitute an important innate immunity effector in trout and perhaps other teleost fish.     

Immunocytochemical localization of vasoactive intestinal polypeptide in the digestive tracts of a holostean and a teleostean fish

1. The localization of vasoactive intestinal polypeptide (VIP) in the gastrointestinal tracts of a holostean fish, the bowfin (Amia calva) and a teleostean fish, the bluegill (Lepomis macrochirus) was determined using immunocytochemistry.2. In the bowfin, VIP immunoreactivity was observed in both gut nerves and gastrointestinal endocrine cells. In the bluegill, only gut nerves exhibited VIP-like immunoreactivity.3. The presence of VIP endocrine cells in the gastric mucosa of bowfin appears to be unique among vertebrates. VIP-containing endocrine cells of the open type were seen in cardiac, oxyntic, and antral gastric mucosa. There appeared to be morphological differences in VIP endocrine cell shapes in anterior versus posterior stomach regions. No VIP endocrine cells were observed in bowfin intestine.4. We conclude that VIP may have an endocrine/paracrine regulatory role in the bowfin stomach and may be strictly a neurotransmitter/neuromodulator in the bowfin gut. There are many species differences in the distribution of VIP-like peptides between neurons and endocrine cells in the guts of lower vertebrates, complicating analysis of the neural versus endocrine evolutionary origin of gut VIP.     

灌喂氧化鱼油后黄颡鱼胃肠道黏膜黑色素细胞分化和黑色素合成途径基因的差异表达

以黄颡鱼(Pelteobagrus fulvidraco)为实验对象, 灌喂氧化鱼油、鱼油7d后, 提取胃肠道黏膜总RNA, 采用RNA-seq测序并做转录组分析, 分析了黑色素生物合成途径关键酶(酪氨酸酶)及其相关蛋白基因、黑素体运动的3个蛋白基因、α黑素细胞刺激激素途径和WNT/β-catenin、EDN3和EDNRB、KIT及其配体KITL3个信号通路的主要蛋白基因的差异表达活性。结果显示, 黄颡鱼胃肠道黏膜中存在黑色素细胞分化和发育过程、黑色素合成及其调控途径的代谢网络, 通过绘制代谢网络得到了关键性酶或蛋白质的基因信息。在灌喂氧化鱼油后, 控制黑色素合成途径主要基因的表达活性显著下调, 可能导致黑色素合成量的不足; α-MSH激素途径主要基因差异表达上调, 具备促进黑色素细胞分化和发育的调控基础; 而调控黑色素细胞分化和发育的3个信号通路主要基因也有差异表达。因此, 黄颡鱼受灌喂氧化鱼油的影响, 黑色素细胞分化和发育过程受到较大影响, 会影响到鱼体成熟的黑色素细胞的数量, 同时, 黑色素的生物合成量不足将导致引起黄颡鱼体色的变化。     

Histological effects of cyanide, stress and starvation on the intestinal mucosa of Pomacentrus coelestis, a marine aquarium fish species

The histological effects of cyanide, stress and starvation on the gastrointestinal tract of Pomacentrus coelestis , a common marine aquarium fish species, were investigated. Neither anaesthetic cyanide nor stress were found to have any detectable effects on the mucosal lining of the intestine. However, starvation resulted in a significant reduction in the intestine length, the surface area of the intestinal mucosa and the mucosal thickness, all occurring within 13 days.