Influence of roundup herbicide on the activities of peptidases in the intestines of various fish species

It has been revealed that, as a rule, Roundup herbicide inhibits activities of peptidases in the intestine mucosa and chyme in various fish species, both benthivorous (common carp Cyprinus carpio, silver bream Blicca bjoerkna, roach Rutilus rutilus, and crucian carp Carassius carassius) and piscivorous (perch Perca fluviatilis, pike Esox lucius, and sander Sander lucioperca). The tolerance of peptidases to the herbicide is species-specific. The magnitude and vector of the effects of Roundup depend on its concentration and localization of an enzyme. In the mucosa, the enzymes of common carp and pike are most tolerant, while the enzymes of common carp, silver bream, and pike are most tolerant in the chyme.     

Distribution of the enzyme activity in the intestine of beluga <Emphasis Type="Italic">Huso huso</Emphasis> and Russian sturgeon <Emphasis Type="Italic">Acipenser gueldenstaedtii</Emphasis> (Acipenseridae)

Data on distribution of digestive enzymes along the intestinal tract of mature specimens of the beluga Huso huso and Russian sturgeon Acipenser gueldenstaedtii are presented. The absence of a distinct proximo-distal gradient of the enzyme activity can be explained by short length of the intestine of sturgeons. Considerable differences are found in the distribution pattern of enzymes along the intestinal tract of beluga and Russian sturgeon. The effect of the method of calculation of the enzyme activity on the data on the proximo- distal gradient is considered.     

Reaction of microorganisms to the digestive fluid of earthworms

The reaction of soil bacteria and fungi to the digestive fluid of the earthworm Aporrectodea caliginosa was studied. The fluid was obtained by centrifugation of the native enzymes of the digestive tract. The inhibition of growth of certain bacteria, spores, and fungal hyphae under the effect of extracts from the anterior and middle sections of the digestive tract of A. caliginosa was discovered for the first time. In bacteria, microcolony formation was inhibited as early as 20–30 s after the application of the gut extracts, which may indicate the nonenzymatic nature of the effect. The digestive fluid exhibited the same microbicidal activity whether the earthworms were feeding on soil or sterile sand. This indicates that the microbicidal agents are formed within the earthworm’s body, rather than by soil microorganisms. The effect of the digestive fluid from the anterior and middle divisions is selective in relation to different microorganisms. Of 42 strains of soil bacteria, seven were susceptible to the microbicidal action of the fluid (Alcaligenes faecalis 345-1, Microbacterium sp. 423-1, Arthrobacter sp. 430-1, Bacillus megaterium 401-1, B. megaterium 413-1, Kluyvera ascorbata 301-1, Pseudomonas reactans 387-2). The remaining bacteria did not die in the digestive fluid. Of 13 micromycetes, the digestive fluid inhibited spore germination in Aspergillus terreus and Paecilomyces lilacinus and the growth of hyphae in Trichoderma harzianum and Penicillium decumbens. The digestive fluid stimulated spore germination in Alternaria alternata and the growth of hyphae in Penicillium chrysogenum. The reaction of the remaining micromycetes was neutral. The gut fluid from the posterior division of the abdominal tract did not possess microbicidal activity. No relation was found between the reaction of microorganisms to the effects of the digestive fluid and the taxonomic position of the microorganisms. The effects revealed are similar to those shown earlier for millipedes and wood lice in the following parameters: quick action of the digestive fluid on microorganisms, and the selectivity of the action on microorganisms revealed at the strain level. The selective effect of the digestive gut fluid of the earthworms on soil microorganisms is important for animal feeding, maintaining the homeostasis of the gut microbial community, and the formation of microbial communities in soils.     

Effects of Dietary <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> on Growth Performance,Digestive Enzymes and Gut Morphology of <Emphasis Type="Italic">Litopenaeus vannamei</Emphasis>

A 15-day feeding trial was conducted to investigate the effect of dietary Lactobacillus plantarum on growth performance, digestive enzyme activities and gut morphology of juvenile Pacific white shrimp, Litopenaeus vannamei (initial body weight = 7.96 ± 0.59 g). Four microbound diets were formulated to contain fermentation supernatant (FS), live bacteria (LB), dead bacteria (DB), and cell-free extract (CE) of L. plantarum. Results indicated that final weight was significantly higher in FS, DB, and CE group in comparison to the control group (P < 0.05). The maximum weight gain rate (WGR) and specific growth rate (SGR) of the CE diet group were significantly higher than that of other groups (P < 0.05). The FCR of CE diet group was lower than that of the control, LB, DB, and FS diets groups (P < 0.05). The highest digestive enzyme activities (amylase, lipase, and pepsin activity) in the hepatopancreas and gut of shrimp were observed in the CE diet group. Histological study revealed that dietary CE diet could significantly increase the enterocytes height of shrimp. The administration of cell-free extract of L. plantarum could effectively improve the growth performance of L. vannamei via the improvement of digestive enzyme activities and the enterocytes height of shrimp. The results of this study will be essential to promote application of probiotics in shrimp aquaculture.     

<Emphasis Type="Italic">Diphyllobothrium dendriticum</Emphasis> (Cestoda: Diphyllobothriidae) in the intestinal tract of the herring gull <Emphasis Type="Italic">Larus argentatus</Emphasis>: Localization and trophic parameters

We studied the activities both of digestive enzymes in the small intestine of the herring gull (Larus argentatus) and a tapeworm Diphyllobothrium dendriticum (Cestoda: Diphyllobothriidae) residing in the intestine. It was found that D. dendriticum infects the medial section of the small intestine of the herring gull. Such localization of D. dendriticum is caused by the maximal activity of proteases and glycosidases and by the high rate of membrane and cavitary digestion in this section. The activity of protease and glycosidase in gulls infected with D. dendriticum is decreased. The activity of proteases in the fractions desorbed from the tegument surface of D. dendriticum is significantly higher than that of glycosidases.     

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.     

Effects of Probiotics on Survival,Growth and Digestive Enzymes Activities in Freshwater Prawn <Emphasis Type="Italic">Macrobrachium rosenbergii</Emphasis> (De Man 1879)

A study was conducted to examine the effects of three probiotics, Lactobacillus sporogenes, Bacillus subtilis and Saccharomyces cerevisiae on the survival, growth and digestive enzymes activities of the freshwater prawn Macrobrachium rosenbergii post larvae (PL). The probiotics, L. sporogenes (4 %), B. subtilis (3 %) and S. cerevisiae (4 %) were taken and mixed with basal diet. Diet without probiotics served as control. These probiotics diets were fed to M. rosenbergii PL for a period of 60 days. After the feeding trail, the growth parameters such as survival, weight gain, specific growth rate and protein efficiency rate were found to be significantly (P < 0.05) higher in 4 % S. cerevisiae incorporated diet fed PL when compared with control. In the case of feed conversion rate just the reverse was seen (P < 0.05) at this concentration. This indicates its superior quality among different concentrations of probiotics tested. Activities of digestive enzymes, such as protease, amylase and lipase were significantly (P < 0.05) higher at this concentration (4 % S. cerevisiae). Some of essential and non-essential amino acids also significantly elevated in probiotics supplemented diet fed prawns. This study indicated that probiotics, S. cerevisiae incorporated diets were beneficial for M. rosenbergii in terms of increasing growth, enzyme and amino acid production.     

Surface-attached and suspended bacterial community structure as affected by C/N ratios: relationship between bacteria and fish production

Bacteria play crucial roles in the combined system of substrate addition and C/N control, which has been demonstrated to improve aquaculture production. However, the complexity of surface-attached bacteria on substrates and suspended bacteria in the water column hamper further application of this system. This study firstly applied this combined system into the culture of grass carp, and then explored the relationship between microbial complexes from surface-attached and suspended bacteria in this system and the production of grass carp. In addition, this study investigated bacterial community structures as affected by four C/N ratios using Illumina sequencing technology. The results demonstrated that the weight gain rate and specific growth rate of grass carp in the CN20 group (C/N ratio 20:1) were the highest (P < 0.05), and dietary supplementation of the microbial complex had positive effects on the growth of grass carp (P < 0.05). Sequencing data revealed that, (1) the proportions of Verrucomicrobiae and Rhodobacter (surface-attached), sediminibacterium (suspended), and emticicia (surface-attached and suspended) were much higher in the CN20 group compared with those in the other groups (P < 0.05); (2) Rhodobacter, Flavobacterium, Acinetobacter, Pseudomonas, Planctomyces, and Cloacibacterium might be important for the microbial colonization on substrates; (3) as the C/N ratio increased, proportions of Hydrogenophaga (surface-attached and suspended), Zoogloea, and Flectobacillus (suspended) increased, but proportions of Bacillus, Clavibacter, and Cellvibro (surface-attached and suspended) decreased. In summary, a combined system of substrate addition and C/N control increased the production of grass carp, and Verrucomicrobiae and Rhodobacter in the surface-attached bacterial community were potential probiotic bacteria that contributed to the enhanced growth of grass carp.     

Alteration of thermoregulation behavior in juvenile fish in relation to satiation level

Thermoregulation behavior of satiated and hungry juvenile sterlet Acipenser ruthenus, the Siberian sturgeon Acipenser baeri, and the goldfish Carassius auratus was studied. Significant alterations of thermoregulation behavior appear in juvenile fish one day after food deprivation. Hungry fish preferred, on average, lower temperatures, their temperature range was broader, and the locomotor activity was higher than in satiated fish.     

Probiotic Potential of Autochthonous Bacteria Isolated from the Gastrointestinal Tract of Four Freshwater Teleosts

In this study, a total of 121 bacterial strains were isolated from the gastrointestinal tract of four teleostean species, namely striped snakehead (Channa striatus), striped dwarf catfish (Mystus vittatus), orangefin labeo (Labeo calbasu) and mrigal carp (Cirrhinus mrigala), among which 8 isolates showed promising antibacterial activity against four potential fish pathogens, Aeromonas hydrophila, Aeromonas salmonicida, Aeromonas sobria and Pseudomonas fluorescens and were non-hemolytic. The isolates were further screened in response to fish bile tolerance and extracellular digestive enzyme activity. Two bacterial strains MVF1 and MVH7 showed highest tolerance and extracellular enzymes activities, and selected for further studies. Antagonistic activity of these two isolates was further confirmed by in vitro growth inhibition assay against four selected fish pathogens in liquid medium. Finally, these two bacterial strains MVF1 and MVH7 were selected as potential probiotic candidates and thus identification by partial 16S rRNA gene sequence analysis. The bacterial isolates MVF1 and MVH7 were identified as two strains of Bacillus sp.     

Properties of Probiotics <Emphasis Type="Italic">Kocuria</Emphasis> SM1 and <Emphasis Type="Italic">Rhodococcus</Emphasis> SM2 Isolated from Fish Guts

This study characterized probiotics Kocuria SM1 and Rhodococcus SM2, which were recovered from the intestinal microbiota of rainbow trout (Oncorhynchus mykiss, Walbaum). The cultures were Gram-positive, non-motile, catalase-positive and oxidase-negative cocci or rods. Cell multiplication of SM1 and SM2 was observed at 4–37 °C (45 °C for SM1), in 0–20% (w/v) NaCl and at pH 2–11. The viability was not affected when exposed to pepsin at pH 2.0 and 3.0, and pancreatin at pH 8.0. Neither isolates were chrome azurol S-positive for siderophore production. Of the 19 common enzymes analysed using the API-ZYM system, only 8 were evident in the culture of SM1 compared to 11 enzymes for SM2. The secondary metabolites of both probiotics were inhibitory to Acinetobacter baumannii, Vibrio anguillarum and V. ordalii; SM2 inhibited Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus. SM2 was resistant to penicillin and sulphatriad, out of six antimicrobial agents; SM1 was resistant to sulphatriad. These results suggest that Kocuria SM1 and Rhodococcus SM2 are able to grow over a wide range of temperature, salinity and pH, including in conditions that mimic the gastrointestinal environment of fish and produce extracellular enzymes that may have a role in the host digestive processes. Importantly, Rhodococcus SM2 displays a high degree of bacteriocinogenic potential against multi-drug-resistant human pathogens that have never been documented among the gut microbiota of fish.     

Degradation capacities of bacteria and yeasts isolated from the gut of <Emphasis Type="Italic">Dendroctonus rhizophagus</Emphasis> (Curculionidae: Scolytinae)

Bark beetles (Curculionidae: Scolytinae) feed on the xylem and phloem of their host, which are composed of structural carbohydrates and organic compounds that are not easily degraded by the insects. Some of these compounds might be hydrolyzed by digestive enzymes produced by microbes present in the gut of these insects. In this study, we evaluated the enzymatic capacity of bacteria (Acinetobacter lwoffii, Arthrobacter sp., Pseudomonas putida, Pseudomonas azotoformans, and Rahnella sp.) and yeasts (Candida piceae, Candida oregonensis, Cyberlindnera americana, Zygoascus sp., and Rhodotorula mucilaginosa) isolated from the Dendroctonus rhizophagus gut to hydrolyze cellulose, xylan, pectin, starch, lipids, and esters. All isolates, with the exception of C. piceae, showed lipolytic activity. Furthermore, P. putida, P. azotoformans, C. americana, C. piceae, and R. mucilaginosa presented amylolytic activity. Esterase activity was shown by A. lwoffii, P. azotoformans, and Rahnella sp. Cellulolytic and xylanolytic activities were present only in Arthrobacter sp. and P. azotoformans. The pectinolytic activity was not recorded in any isolate. This is the first study to provide evidence on the capacity of microbes associated with the D. rhizophagus gut to hydrolyze specific substrates, which might cover part of the nutritional requirements for the development, fitness, and survival of these insects.     

Proteinases of kilka <Emphasis Type="Italic">Clupeonella cultriventris</Emphasis> of the Rybinsk Reservoir

The effect of temperature on activity of proteinases of the stomach mucosa, intestinal mucosa, chime, and the entire body of kilka Cluperonella cultriventris is described. Significant differences are revealed in temperature characteristics (t ^o-function, Q ₁₀, and E _act) of the investigated enzymes. The temperature has a larger influence on the activity of intestinal proteinases than the activity of proteinases of the stomach, chime, and the whole kilka organism. The lowest value of E _act within the temperature range of life of this species is characteristic of chime proteinases. Hemoglobinlytic proteinases of various tissues of kilka that substantiate, at pH 3.0 and 5.0 (the action zone of cathespsins), the processes of induced autolysis contribute to efficient alimentation of other fish species. The assumption is made that a high activity level of proteinases of the digestive tract, comparable with that of other fish species of the upper Volga, enables kilka that have assimilated to the Rybinsk Reservoir to feed efficiently within the temperature range characteristic of the water-bodies of this zone.     

Alterations in Digestive Enzymes of Three Freshwater Teleostean Fishes by Almix Herbicide: A Comparative Study

Three freshwater teleostean fishes viz., Anabas testudineus (Bloch), Heteropneustes fossilis (Bloch) and Oreochromis niloticus (Linnaeus) were exposed to almix (66.67 mg/l) herbicide for 30 days to investigate the activity of digestive enzymes (amylase, lipase and protease) in stomach, intestine and liver. Amylase activity showed significantly high (p < 0.05) in all the fishes compared to control value and highest activity was observed in liver of A. testudineus (721.99 %) and minimum in intestine of H. fossilis (195.37 %). Lipase activity was also significantly increased (p < 0.05) in all the tissues; but highest in intestine of O. niloticus (235.51 %) and minimum in intestine of A. testudineus (130.51 %). Protease activity also showed similar trends of enhancement; it was maximum in stomach of O. niloticus (362.69 %), whereas in liver of H. fossilis it was rather less (173.72 %). Increased activity of digestive enzymes resulting from tissue damage ultimately affected the fish health due to impairment of digestive physiology confirming the herbicidal contamination on fish species. The sensitivity to the almix herbicide was pronounced in the order of O. niloticus > A. testudineus > H. fossilis.     

Effect of Cestodal Infestation on the Distribution Pattern of Digestive Enzyme Activities along the Small Intestine of the Kittiwake (<Emphasis Type="Italic">Rissa tridactyla</Emphasis>)

Activities of digestive enzymes in the intestine of adult and nestling kittiwakes (Rissa tridactyla) were studied. Glycosidase and protease activities in the three parts of the small intestine (proximal, medial and distal) as well as their total activity were determined, and the glycosidase/protease activity ratio (G/P coefficient) was calculated. Enzyme activities were found to be distributed unevenly along the intestine. In a proximal to distal direction, glycosidase and protease activities decrease in nestlings and increase in adult kittiwakes. In nestlings, protease activity values are several times higher than in adults. G/P coefficient values vary within the range of 0.33–0.92 in adult animals and 0.22–0.41 in nestlings. Infestation with tapeworms Alcataenia larina (Cestoda, Dilepididae) and Tetrabothrius erostris (Cestoda, Tetrabothriidae) leads to reduced digestive enzyme activities in the small intestine of kittiwake nestlings. In adult animals, infestation with T. erostris leads to a reduced and with A. larina to an increased protease activity.     

Comparison of L-lactate dehydrogenases of different origins produced in the cells of yeast <Emphasis Type="Italic">Schizosaccharomyces pombe</Emphasis>

The expression of L-lactate dehydrogenase genes ldh1 (Bos taurus), ldhA (Homo sapiens), ldhA (Rhizopus oryzae), ldh1 (Lactobacillus plantarum), and ldh1 (Lactobacillus pentosus) in the cells of yeast Schizosaccharomyces pombe VKPM U-3106 has been investigated. The catalytic characteristics of the enzymes encoded by these genes have been compared, and the intensity of lactic acid synthesis by the recombinant strains obtained has been evaluated. The enzymatic activity of L-lactate dehydrogenases from L. plantarum and L. pentosus was the highest (approximately 2 to 2.5 times higher than that of the mammalian enzymes), and these enzymes therefore appear to have the highest potential for the development of lactic-acid producing strains of yeast S. pombe.     

Dietary Supplement of Medicinal Herbal Leaf Powder on Growth Performance,Digestive Enzymes Activities,Energy Utilization and Vitamin Levels of the Freshwater Prawn <Emphasis Type="Italic">Macrobrachium rosenbergii</Emphasis>

Experiments were conducted to evaluate the influence of dietary supplement (@ 5%) dried leaf powder of the medicinal herbs, Ocimum sanctum, Solanum trilobatum and Phyllanthus amarus on survival, growth, activities of digestive enzymes, energy utilization, and vitamin contents of the freshwater prawn, Macrobrachium rosenbergii. Results indicated significant (P < 0.05) improvements in survival, growth, activities of digestive enzymes, energy utilization, and vitamin levels in herb supplemented feeds fed prawns group compared to prawns fed non-herbs feed. Best growth was noted in prawns fed with the feed supplemented by leaf powder of P. amarus, followed by S. trilobatum and O. sanctum. It is concluded that leaf powder of these herbs can be used as dietary supplement for better growth of freshwater prawn.     

Decreased microbial diversity and <Emphasis Type="Italic">Lactobacillus</Emphasis> group in the intestine of geriatric giant pandas (<Emphasis Type="Italic">Ailuropoda melanoleuca</Emphasis>)

It has been established beyond doubt that giant panda genome lacks lignin-degrading related enzyme, gastrointestinal microbes may play a vital role in digestion of highly fibrous bamboo diet. However, there is not much information available about the intestinal bacteria composition in captive giant pandas with different ages. In this study, we compared the intestinal bacterial community of 12 captive giant pandas from three different age groups (subadults, adults, and geriatrics) through PCR-denaturing gradient gel electrophoresis (DGGE) and real-time PCR analysis. Results indicated that microbial diversity in the intestine of adults was significantly higher than that of the geriatrics (p < 0.05), but not significant compared to the subadults (p > 0.05). The predominant bands in DGGE patterns shared by the twelve pandas were related to Firmicutes and Proteobacteria. Additionally, in comparison to healthy individuals, antibiotic-treated animals showed partial microbial dysbiosis. Real-time PCR analyses confirmed a significantly higher abundance of the Lactobacillus in the fecal microbiota of adults (p < 0.05), while other bacterial groups and species detected did not significantly differ among the three age groups (p > 0.05). This study revealed that captive giant pandas with different ages showed different intestinal bacteria composition.     

Nitrification in intertidal sponge <Emphasis Type="Italic">Cinachyrella cavernosa</Emphasis>

The nitrification process in different sections of the sponges remains unresolved, despite several studies on the nitrogen cycling pathways in the tissues of temperate and Arctic sponges. In this study, the abundance, diversity and activity of the associated nitrifying organisms in intracellular, intercellular, extracellular and cortex of a tropical intertidal sponge, Cinachyrella cavernosa, were investigated using most probable number, next-generation sequencing and incubation method, respectively. The nitrification rate and the abundance of nitrifying bacteria showed significant difference among different sections. The nitrification rate in C. cavernosa was 2–12× higher than the reported values in other sponge species from temperate and Arctic regions. Nitrification rate in sponge cortex was 2× higher than in intercellular and extracellular sections. Ammonium and nitrite oxidisers ranged from 10³ to 10⁴ CFU g^?1 in the sponge with a high number of ammonium and nitrite oxidisers in the cortex. Nitrifiers belonging to Nitrosomonas, Nitrospira, Nitrospina, Nitrobacter and Nitrosopumilus were present in different sections of the sponge, with nitrifying archaea dominating the intracellular section and nitrifying bacteria dominating other sections. This study reports for the first time the nitrification inside the sponge cells. The study also suggests that the intertidal sponge, C. cavernosa, harbours metabolically active nitrifiers in different sections of the sponge body with different rates of nitrification. Thus, nitrifiers play an important role in ammonia detoxification within the sponge and also contribute to the nitrogen budget of the coastal ecosystem.     

Investigation of the midgut structure and ultrastructure in <Emphasis Type="Italic">Cimex lectularius</Emphasis> and <Emphasis Type="Italic">Cimex pipistrelli</Emphasis> (Hemiptera: Cimicidae)

Cimicidae are temporary ectoparasites, which means that they cannot obtain food continuously. Both Cimex species examined here, Cimex lectularius (Linnaeus 1758) and Cimex pipistrelli (Jenyns 1839), can feed on a non-natal host, C. lectularius from humans on bats, C. pipistrelli on humans, but never naturally. The midgut of C. lectularius and C. pipistrelli is composed of three distinct regions—the anterior midgut (AMG), which has a sack-like shape, the long tube-shaped middle midgut (MMG), and the posterior midgut (PMG). The different ultrastructures of the AMG, MMG, and PMG in both of the species examined suggest that these regions must fulfill different functions in the digestive system. Ultrastructural analysis showed that the AMG fulfills the role of storing food and synthesizing and secreting enzymes, while the MMG is the main organ for the synthesis of enzymes, secretion, and the storage of the reserve material. Additionally, both regions, the AMG and MMG, are involved in water absorption in the digestive system of both Cimex species. The PMG is the part of the midgut in which spherites accumulate. The results of our studies confirm the suggestion of former authors that the structure of the digestive tract of insects is not attributed solely to diet but to the basic adaptation of an ancestor.