Dr Md Javed Foysal

Cyanobacterial blooms are a concerning issue that threaten ecosystems, ecology and animal health. Bloom frequency has increased tremendously in recent times due to pollution, eutrophication of waterways, climate change, and changes in microbial community dynamics within the aquatic environment. Information about the spatiotemporal variation in microbial communities that drive a cyanobacterial bloom is very limited. Here, we analysed the spatiotemporal diversity and composition of bacterial communities, with a focus on cyanobacteria, during the bloom phase in a natural reservoir in Eastern Australia using high throughput amplicon sequencing. Sampling points and season had no influence on the richness and evenness of microbial communities during the bloom period, however some compositional differences were apparent across the seasons. Cyanobacteria were highly abundant during summer and autumn compared to winter and spring. The dominant cyanobacterial taxa were Planktothrix, Cyanobium and Microcystis and were found to be significantly abundant during summer and autumn. While cyanobacterial abundance soared in summer (25.4 %), dominated by Planktothrix (12.2 %) and Cyanobium (8.0 %), the diversity was highest in autumn (24.9 %) and consisted of Planktothrix (7.8 %), Nodularia (5.3 %), Planktothricoides (4.6 %), Microcystis (3.5 %), and Cyanobium (2.3 %). The strongly correlated non-photosynthetic Gastranaerophilales found in the sediment and water, suggested vertical transmission from the animal gut through faeces. To our knowledge, this is the first report of Planktothrix-driven toxic cyanobacterial bloom in Australia. Our study expands current understanding of the spatiotemporal variation in bacterial communities during a cyanobacterial bloom and sheds light on setting future management strategies for its control.

New aquafeed ingredients produced by a circular economy approach are the opportunity for sustainable and resilient aquaculture. In the light of this approach, the mixture of abalone waste and Sargassum spp (9:1) fermented by Saccharomyces cereviceae and Lactobacillus casei (Yakult®) (FMAS) were used to replace 0% (FMAS0), 25% (FMAS-25), 50% (FMAS-50), 75% (FMAS-75), and 100% (FMAS-100) of fishmeal (FM) protein in marron, Cherax cainii diet. The marron was fed these diets in triplicate for 90 days. Growth, feed utilization and protein efficiency ratio were unchanged in marron-fed all test diets. Improvement in apparent protein digestibility was aligned with an increase in the size and number of B-cells in the hepatopancreas. Most of the immune responses, except for haemocyte clotting time, hyaline cells and neutral red retention time (NRR time) were unchanged by 42- and 90-days feeding trials compared to those of the control group. 90 days post-feeding marron with FMAS25 showed a lower haemocyte clotting time than the post 42 days feeding marron with the same diet. Hyaline cells increased in marron fed FMAS75 for 90 days compared to marron fed the same diet for 42 days. The challenge test involved injecting marron with Vibrio mimicus resulted in a 100% survival rate after 96 h of exposure. During the challenge test, phagocytosis activity in 24 and 48-h post-challenged marron fed FMAS75 decreased which recovered after 96 h post-challenge. Marron fed FMAS50 also recorded a significantly higher proportion of granular cells after 24 h and NRR time at 96 h compared with that of other treatments. Given the above indicators of bio-growth, feed efficiency and immune responses, total replacement of FM protein of marron practical feed with FMAS are considered feasible and optimum to maintain health status and resistance to disease.

Pomegranate peel extract (PPE) has been widely identified as a rich source of therapeutically active phyto-compounds, known for their beneficial effects in aquaculturable species. In this context, a 60-day feeding trial was carried out to evaluate the effect of supplementation of pomegranate peel extract (PPE) on growth performance, haemato-immunological responses and cytokine gene expression in Labeo rohita (rohu) fingerlings challenged with Aeromonas hydrophila. A total of 240 rohu fingerlings (3.25 ± 0.52 g) were randomly distributed into four treatment groups in triplicates with 15 fish per tank. The groups were fed with four different diets containing graded levels of PPE viz., 0 (control), 0.5, 1.0 and 1.5%. The results demonstrated significantly enhanced weight gain% and specific growth rate (SGR) in the group fed with 0.5% of dietary PPE. The cumulative survival post challenge to A. hydrophila was enhanced to 72.90% in 0.5% PPE fed group as compared to 20.01% in the control group. Haemato-immunological parameters in terms of respiratory burst activity (NBT), serum total protein, globulin and lysozyme activity displayed highest levels in L. rohita fed with the 0.5% PPE diet. Further, the dietary group supplemented with PPE at 0.5% level exhibited upregulated mRNA transcript levels of pro-inflammatory cytokines viz. IL-1ß, TNF-a in different lymphoid organs such as gut, kidney and liver of L. rohita fingerlings post-challenge. In contrast, anti-inflammatory cytokine IL-10, was significantly down-regulated in infected rohu fed with 0.5% PPE. Taken together, the best optimum dosage or inclusion level at 0.5% PPE, obtained through the study, represented remarkable favourable variations in growth, haemato-immunological parameters and cytokine gene expression of L. rohita fingerlings. Thus, the dietary supplementation of PPE at 0.5% could effectively enhance different aspects of innate immunity alongwith disease resistance to A. hydrophila infection in L. rohita fingerlings.

Valorising waste from the processing of fishery and aquaculture products into functional additives, and subsequent use in aquafeed as supplements could be a novel approach to promoting sustainability in the aquaculture industry. The present study supplemented 10% of various fish protein hydrolysates (FPHs), obtained from the hydrolysis of kingfish (KH), carp (CH) and tuna (TH) waste, with 90% of poultry by-product meal (PBM) protein to replace fishmeal (FM) completely from the barramundi diet. At the end of the trial, intestinal mucosal barriers damage, quantified by villus area (VA), lamina propria area (LPA), LPA ratio, villus length (VL), villus width (VW), and neutral mucin (NM) in barramundi fed a PBM-based diet was repaired when PBM was supplemented with various FPHs (p < 0.05, 0.01, and 0.001). PBM-TH diet further improved these barrier functions in the intestine of fish (p < 0.05 and 0.001). Similarly, FPHs supplementation suppressed PBM-induced intestinal inflammation by controlling the expression of inflammatory cytokines (tnf-a and il-10; p < 0.05 and 0.001) and a mucin-relevant production gene (i-mucin c; p < 0.001). The 16S rRNA data showed that a PBM-based diet resulted in dysbiosis of intestinal bacteria, supported by a lower abundance of microbial diversity (p < 0.001) aligned with a prevalence of Photobacterium. PBM-FPHs restored intestine homeostasis by enhancing microbial diversity compared to those fed a PBM diet (p < 0.001). PBM-TH improved the diversity (p < 0.001) further by elevating the Firmicutes phylum and the Ruminococcus, Faecalibacterium, and Bacteroides genera. Muscle atrophy, evaluated by fiber density, hyperplasia and hypertrophy and associated genes (igf-1, myf5, and myog), occurred in barramundi fed PBM diet but was repaired after supplementation of FPHs with the PBM (p < 0.05, 0.01, and 0.001). Similarly, creatine kinase, calcium, phosphorous, and haptoglobin were impacted by PBM-based diet (p < 0.05) but were restored in barramundi fed FPHs supplemented diets (p < 0.05 and 0.01). Hence, using circular economy principles, functional FPHs could be recovered from the fish waste applied in aquafeed formulations and could prevent PBM-induced intestinal dysbiosis and muscular atrophy. (Figure presented.)

Enterococcus faecalis is associated with streptococcosis like infection in fish. A whole-genome sequence study was conducted to investigate the virulence factor and antibiotic-resistance genes in three fish pathogenic E. faecalis. Genomic DNA was extracted from three strains of E. faecalis isolated from streptococcosis infected Nile tilapia (strains BF1B1 and BFFF11) and Thai sarpunti (strain BFPS6). The whole genome sequences of these three strains were performed using a MiSeq sequencer (Illumina, Inc.). All three strains conserved 69 virulence factor such as genes associated with protection against oxidative stress, bacterial cell wall synthesis, gelatinase toxin, multiple biofilm-associated genes and capsule producing genes. Moreover, 39 antibiotic-resistance genes against sixteen major groups of antibiotics were identified in the genome sequences of all three strains. The most commonly used antibiotic Tetracycline resistance genes were found only in BFPS6 strain, whereas, Bacteriocin synthesis genes were identified in both BFFF11 and BFPS6 strain. Phylogenetic analysis revealed that strains BF1B1 and BFFF1 form a different cluster than BFPS6. This is one of the first whole-genome sequence study of fish pathogenic E. faecalis, unfold new information on the virulence factor and Antibiotic resistance genes linked to pathogenicity in fish.

Replacement of fishmeal (FM) from aquafeed has gained tremendous attention over the past 20 years due to environmental and ecological ill-impacts of large-scale capture of wild fishes. Insects, black soldier fly (BSF) possess balanced nutritional properties, hence, considered as an ideal candidate for the replacement of FM. A number of studies have been performed on the effects of BSF as a fishmeal alternative on the gut microbiota of aquatic animals using high throughput sequencing (HTS). However, due to the inconsistency in the success of BSF inclusions, a systematic meta-analysis approach was imperative to conclude the usefulness of BSF inclusion in aquafeed. In this context, the present study systematically analysed the HTS data obtained from gut microbiota of BSF, and rainbow trout (RT) fed BSF from various life cycles with different formulations. Results revealed that Actinobacteria and Firmicutes were most abundant in the gut of BSF and fish fed BSF diets while Proteobacteria was the dominant bacteria in the fish gut fed FM. The augmentation of Actinobacteria and Firmicutes in the fish gut was mainly due to higher abundance of Actinomyces, Bacillus, Lactobacillus and Enterococcus at the genus level. More diversified gut microbiota was observed in fish fed larvae and pre-pupae BSF, while defatted BSF was linked to a specific group of bacteria. Feeding BSF was shown to reduce the core microbiota of rainbow trout by increasing Firmicutes. Overall, the findings of this study could be used as future references for fish and shellfish nutrition, specifically in formulating aquafeed with BSF incorporation.

Zeolite is known to uptake toxic metals and filter nitrogenous waste from aquaculture effluents. The present study aimed to investigate the impacts of zeolite in three different applications namely, dietary zeolite (DZ), suspended zeolite (SZ) in the water column, and a combination of both (DZSZ) relative to unexposed freshwater crayfish, marron (control). At the end of the 56-days trial, the impact was assessed in terms of characterization of microbial communities in the culture environment and the intestine of marron. Alongside the microbial communities, the innate immune response of marron was also evaluated. The 16S rRNA data showed that marrons exposed to the suspended zeolite had a significant increase of bacterial diversity in the gut, including the restoration of marron core operational taxonomic units (OTUs), relative to other forms of exposures (DZ, DZSZ) and the control. Suspended zeolite alone also increased the number of unshared OTUs and genera, and improved predicted metabolic functions for the biosynthesis and digestion of proteins, amino acids, fatty acids, and hormones. In the tank sediment, the shift of microbial communities was connected more strongly with the time of experiment than the type of zeolite exposure. In the second case, only control marron had a different microbial ordination in terms of rare taxa present in the community. Nevertheless, the modulation in the gut environment was found more prominent in DZ, relative to modulation in the tank sediments. The taxa-environment correlation identified Rhodoferax as the most potential bacteria in removing nitrogenous waste from the rearing environment. Further analysis showed that SZ resulted in the upregulation of genes associated with the innate immune response of marron. Overall results suggest that SZ can be used to enrich microbial communities in the gut and tank sediments and better immune performance of marron.

Trace element supplementation to the freshwater environment can influence the plankton density and species diversity, contributing to the nutrition of aquaculture species, especially during the juvenile stage. An experiment was conducted under laboratory conditions to evaluate the effects of supplementing different mixtures of manganese, silica and phosphorus on the plankton density and species diversity and their impact on cultured juvenile marron (Cherax cainii, Austin and Ryan, 2002). Manganese, silica and phosphorus in concentrations of 0.0024, 0.41, 0.05 mg*L-1; 0.0041, 0.82, 0.12 mg*L-1; and 0.0058, 1.26, 0.25 mg*L-1 respectively termed as low, medium and high were supplemented to tank water containing a phytoplankton density of 3.77 ± 0.16 × 106 cells*L-1 and 292.9 ± 17.6 individuals*L-1 of zooplankton, and plankton growth was observed every 24 h for 6 days. Afterwards, a 3-month trial was conducted studying the effects of these trace element concentrations and resulting plankton densities on marron growth, survival, moulting, gut microbiota and health indices. Silica supplementation at high concentration increased the diatom abundance, silica and phosphorus supplementation at higher concentration that resulted in a significant increase in plankton density and species diversity, leading to improved marron health indices than the control and the tanks receiving a low concentration. Marron-specific growth rate, weight gain and dissolved copper concentration in haemolymph were significantly higher in tanks with higher supplementation and higher plankton density. Marron survival, moult interval and total haemocyte count were not affected by the supplementation. Marron gut microbiota at higher trace element concentration supplementation showed a significant increase in abundance of phosphate solubilizing bacteria.

Immune mechanisms are major players in ensuring the normal functioning of testicular functions. However, apart from their role in active defence against pathogens, prior studies have also suggested a possibility for reproduction-related (non-immune) functions of certain immune elements. This study employs a comparative transcriptomics approach followed by network analysis for tracking the variation in the immuno-reproductive milieu of Clarias magur testis in spawning versus pre-spawning phase. The results show a significant modulation of both reproduction and immune-relevant genes in spawning versus pre-spawning phase. The functional enrichment of the upregulated reproduction-relevant gene network also shows immune-related biological processes which indicates a probability of involvement of these candidates in spermatogenesis-related events for switching from pre-spawning to spawning phase. The upregulated immune network is highly dense with 40 hubs, 10 cluster sub-networks and 142 functionally enriched pathways in comparison to its downregulated counterpart with only 5 hubs, 1 cluster and 1 enriched pathway. These findings indicate that the synchronisation in modulation of both reproductive and immune-related factors is critical for progression of testicular events guiding the switch from pre-spawning to spawning phase. The reproductive phase-dependent variation in plasma sex steroid levels and the selected genes for quantitative PCR also corroborated this hypothesis. The study also serves as a preliminary screening step for probable immune candidates that may be involved in reproductive functions of testis in addition to defence.

The present study aimed to characterize and compare the skin and gut microbial communities of rohu at various post-harvest stages of consumption using quantitative real-time polymerase chain reaction and 16S rRNA-based amplicon sequencing. Real-time PCR amplification detected higher copy numbers for coliform bacteria¿Escherichia coli, Salmonella enterica and Shigella spp. in the marketed fish¿compared to fresh and frozen samples. The 16S rRNA data revealed higher alpha diversity measurements in the skin of fish from different retail markets of Dhaka city. Beta ordination revealed distinct clustering of bacterial OTUs for the skin and gut samples from three different groups. At the phylum level, Proteobacteria was most abundant in all groups except the Fusobacteria in the control fish gut. Although Aeromonas was found ubiquitous in all types of samples, diverse bacterial genera were identified in the marketed fish samples. Nonetheless, low species richness was observed for the frozen fish. Most of the differentially abundant bacteria in the skin samples of marketed fish are opportunistic human pathogens enriched at different stages of postharvest handling and processing. Therefore, considering the microbial contamination in the aquatic environment in Bangladesh, post-harvest handling should be performed with proper methods and care to minimize bacterial transmission into fish.

Yogurt is an ethnic fermented food that dominates as an integral part of human diet for its health benefits. The nutritional quality of yogurt is diminished due to the presence of adulterants, metal and microbial contaminants. Upon consumption, these materials cause metabolic disorders and food-borne diseases. This study was designed to explore the physicochemical parameters, minerals, heavy metals, and bacteriome diversity in Bangladeshi sour yogurt prepared from cow and buffalo milk. Bacterial culture followed by 16S rRNA sequencing was performed to assess the bacterial consortia in yogurt. The higher amount of total solid (TS), solid-not-fat (SNF), pH, calcium and iron, and lower value of moisture, potassium and magnesium were observed at significant level (P < 0.05). Lactobacillus and other pathogenic bacteria like Shigella, Aeromonas, Enterobacter and Escherechia were isolated where Lactobacillus predominated. From amplicon sequencing, cow yogurt samples were observed to be predominated by Lactobacillus and Streptococcus while Enterobacter, Lactococcus, Aeromonas and Acinetobacter were the top abundant bacterial genera in buffalo yogurt. Crucial factors affecting the yogurt quality viz. nutritional profile, and bacterial community structure were explored for the first time in this study that will inevitably aid in the safe production of yogurt and public health measures globally.

To reduce the reliance on fishmeal (FM), other protein sources have been evaluated on cultured animals. In a 60-days feeding trial, marrons (Cherax cainii) were fed a FM diet and five test diets containing 100% of plant-based protein sources such as soybean, lupin and valorised animal-based proteins such as poultry-by-product, black soldier fly and tuna hydrolysate. At the end of the trial, DNA samples from marron gut and rearing water were investigated through DNA-based 16S rRNA gene sequencing. Plant-based diets increased abundance for Aeromonas, Flavobacterium and Vogesella, whereas animal and insect proteins influenced diverse bacterial groups in the gut linked to various metabolic activities. Insect meal in the water favoured the growth of Firmicutes and lactic acid bacteria, beneficial for the marron health. Aeromonas richness in the gut and reared water signified the ubiquitous nature of the genus in the environment. The higher bacterial diversity in the gut and water with PBP and BSF was further supported by qPCR quantification of the bacterial single-copy gene, rpoB. The overall results suggested that PBP and BSF can exhibit positive and influential effects on the gut and water microbial communities, hence can be used as sustainable ingredients for the crayfish aquaculture.

Coronavirus disease-2019 (COVID-19) is an infectious disease caused by SARS-CoV-2 virus. The microbes inhabiting the oral cavity and gut might play crucial roles in maintaining a favorable gut environment, and their relationship with SARS-CoV-2 infection susceptibility and severity is yet to be fully explored. This study investigates the diversity and species richness of gut and oral microbiota of patients with COVID-19, and their possible implications toward the severity of the patient's illness and clinical outcomes. Seventy-four (n = 74) clinical samples (gut and oral) were collected from 22 hospitalized patients with COVID-19 with various clinical conditions and 15 apparently healthy people (served as controls). This amplicon-based metagenomic sequencing study yielded 1,866,306 paired-end reads that were mapped to 21 phyla and 231 classified genera of bacteria. Alpha and beta diversity analyses revealed a distinct dysbiosis of the gut and oral microbial communities in patients with COVID-19, compared to healthy controls. We report that SARS-CoV-2 infection significantly reduced richness and evenness in the gut and oral microbiomes despite showing higher unique operational taxonomic units in the gut. The gut samples of the patients with COVID-19 included 46 opportunistic bacterial genera. Escherichia, Shigella, and Bacteroides were detected as the signature genera in the gut of patients with COVID-19 with diarrhea, whereas a relatively higher abundance of Streptococcus was found in patients with COVID-19 having breathing difficulties and sore throat (BDST). The patients with COVID-19 had a significantly lower abundance of Prevotella in the oral cavity, compared to healthy controls and patients with COVID-19 without diabetes, respectively. The altered metabolic pathways, including a reduction in biosynthesis capabilities of the gut and oral microbial consortia after SARS-CoV-2 infection, were also observed. The present study may, therefore, shed light on interactions of SARS-CoV-2 with resilient oral and gut microbes which might contribute toward developing microbiome-based diagnostics and therapeutics for this deadly pandemic disease.

The gut microbiome of fish contains core taxa whose relative abundances are modulated in response to diet, environmental factors, and exposure to toxicogenic chemicals, influencing the health of the host fish. Recent advances in genomics and metabolomics have suggested the potential of microbiome analysis as a biomarker for exposure to toxicogenic compounds. In this 35-day laboratory study, 16S RNA sequencing and multivariate analysis were used to explore changes in the gut microbiome of juvenile Lates calcarifer exposed to dietary sub-lethal doses of three metals: vanadium (20 mg/kg), nickel (480 mg/kg), and iron (470 mg/kg), and to two oils: bunker C heavy fuel oil (HFO) (1% w/w) and Montara, a typical Australian medium crude oil (ACO) (1% w/w). Diversity of the gut microbiome was significantly reduced compared to negative controls in fish exposed to metals, but not petroleum hydrocarbons. The core taxa in the microbiome of negative control fish comprised phyla Proteobacteria (62%), Firmicutes (7%), Planctomycetes (3%), Actinobacteria (2%), Bacteroidetes (1%), and others (25%). Differences in the relative abundances of bacterial phyla of metal-exposed fish were pronounced, with the microbiome of Ni-, V-, and Fe-exposed fish dominated by Proteobacteria (81%), Firmicutes (68%), and Bacteroidetes (48%), respectively. The genus Photobacterium was enriched proportionally to the concentration of polycyclic aromatic hydrocarbons (PAHs) in oil-exposed fish. The probiotic lactic acid bacterium Lactobacillus was significantly reduced in the microbiota of fish exposed to metals. Transcription of cytokines IL-1, IL-10, and TNF-a was significantly upregulated in fish exposed to metals but unchanged in oil-exposed fish compared to negative controls. However, IL-7 was significantly downregulated in fish exposed to V, Ni, Fe, and HFOs. Fish gut microbiome exhibits distinctive changes in response to specific toxicants and shows potential for use as biomarkers of exposure to V, Ni, Fe, and to PAHs present in crude oil.

Streptococcosis is one of the major threats to Nile tilapia (Oreochromis niloticus) in most regions of the world. Recently, Enterococcus faecalis has been widely reported to be involved in streptococcosis in O. niloticus in Asia and Africa. This study aimed to isolate beneficial marine bacteria to evaluate their effects on growth, hematological parameters, nonspecific immunity, the gut bacteriome, and streptococcosis prevention efficacy in O. niloticus. A total of 36 marine soil bacteria were isolated, and in vitro screening was conducted to determine their antibacterial activities against fish pathogens. Two antagonistic bacteria were identified based on 16S rRNA gene sequencing, Bacillus haynesii CD223 and Advenella mimigardefordensis SM421. These bacteria were incorporated into fish feed and fed to O. niloticus for 90 days. The application of these strains via incorporation into fish feed significantly promoted growth, improved hematological parameters and immunoglobulin M (IgM) levels, modulated the gut bacteriome by reducing the load of pathogenic Enterococcus spp., and developed disease prevention efficacy in O. niloticus. Furthermore, in vivo assays revealed that the inclusion of extracellular products (ECPs) (at 250 mg mL21) of CD223 and SM421 with feed significantly enhanced the rate of survival (100%) of O. niloticus from streptococcosis compared to the controls (only 30%). The ECPs of these bacteria also prevented 90 to 100% of fish from developing streptococcosis. These strains could be promising for safe use in O. niloticus farming to prevent and control the emergence of streptococcosis caused by E. faecalis.

This study investigated the diversity of bacterial community in healthy and streptococcosis infected Nile tilapia (Oreochromis niloticus) to identify the primary causative agents associated with the disease using metagenomics, phylogenetic analysis and in vivo challenge test. A total of 24 fishes, both healthy and diseased were collected during summer from three different districts and six different ponds of Bangladesh. Alpha-beta diversity analysis of the next generation sequence data showed distinctly different (P <.05) microbial communities in control and diseased fishes. In diseased fish, we found significant (P <.05) increase abundance of Enterococcus in the skin lesion, and Leifsonia, Photobacterium, Aeromonas, Pseudomonas in the gut. Interestingly, three different Enterococcus species, E. faecalis, E. hirae and E. faecium were identified the causative agents of streptococcosis through 16S rRNA based phylogenetic analysis. In-vivo challenge test also revealed the high pathogenicity and mortality of these species to experimental tilapia fingerlings. Further study revealed a significant correlation between the pathogenicity and sequence divergence in characterized Enterococcus spp. isolates. Taken together, our study for the first time demonstrated the dominance of multiple Enterococcus species as causative agents of streptococcosis in Nile tilapia in Bangladesh. Our findings should be valuable for the diagnosis and treatment of streptococcosis infection in tilapia.

This study was conducted to investigate the combined effects of probiotic yeast, Saccharomyces cerevisiae, and lactic acid bacteria, Lactobacillus casei on the growth performance, biochemical response, cytokine gene expression, gut histomorphology and microbial composition of juvenile barramundi, Lates calcarifer. Barramundi juveniles with an initial mean weight of 12.17 ± 0.15 g were randomly allocated into 8 tanks (300L) at a density of 20 fish per tank and fed either a basal diet (control), or the same basal diet supplemented with 1% of S. cerevisiae and 1% of L. casei, collectively termed ¿probiotic¿, for 56 days in quadruplicate. Results indicate that dietary probiotic supplementation produced significant (P < 0.05) improvement in final body weight, specific growth rate and feed conversion ratio compared to the control. A significant increase in serum protein was evident in fish fed the probiotic supplemented diet, while none of the other measured blood and serum parameters were influenced by the probiotic supplementation. Probiotic supplementation also resulted in an up-regulation of immune responsive inflammatory cytokine genes (IL-10 and TNF-a) and exhibited a higher number of gut mucosal goblet cells and increased microvillous length. The sequence data of the distal gut showed significant positive influence of the probiotic diet on alpha-beta microbial diversities and decreased abundance of pathogenic Staphylococcus and Corynebacterium. These results confirm that the co-supplementation of S. cerevisiae and L. casei improved the growth, immune response and gut health by increasing the diversity of microbiota in juvenile barramundi.

Promoting a circular economy via the transformation of food waste into alternative and high-value protein sources for aquaculture diets is a novel approach to developing alternative raw materials to fishmeal (FM). This approach can reduce the ecological impact on the aquatic environment and simultaneously can provide an option for sustainable food waste management. In this context, we report a 56-day trial of feeding barramundi, Lates calcarifer on four iso-nitrogenous and iso-lipidic diets where the control (0PBM-0HI) was a FM-based diet and the other test diets replaced FM protein with mixtures of a poultry by-product meal (PBM) and a full-fat Hermetia illucens (HI) larvae meal reared on fish waste: the test diets were 85% PBM + 15% HI (85PBM-15HI), 80% PBM + 20% HI (80PBM-20HI) and 75% PBM + 25% HI (75PBM-25HI). Fish fed PBM-HI-based diets showed an equal growth rate and amino acid profile when compared to the control group. Among all serum metabolites, alanine aminotransferase and glutamate dehydrogenase decreased in fish fed PBM-HI-based diets, whilst total protein levels improved in the same diets. Serum lysozyme and bactericidal activity were unchanged which supported the observation of similar infection rates against V. harveyi. Except for the kidney and intestine, catalase activity in the serum and liver increased in fish-fed PBM-HI-based diets. In assessing the gastrointestinal mucosal morphology, the goblet cells producing neutral mucins were higher in PBM-HI-fed fish than the control. PBM-HI diets also enhanced bacterial richness and diversity and increased abundance for Lactobacillus, Clostridium, and Ruminococcus. In summary, combining full-fat HI with PBM allowed complete replacement of FM with no negative effects on growth whilst improving gut health. Such diets would be beneficial for the aquaculture industry, both ecologically and economically, as well as providing value-adding to animal waste as alternative protein sources for aquafeed production.

Feeding freshwater crayfish species with different diets not only affects the water quality but also induces the abundance of various microbial communities in their digestive tracts. In this context, very limited research has been undertaken to understand the impacts of various protein incorporated aqua-diets on the characteristics of water and its microbial communities. In this study, we have critically analysed the water quality parameters including pH, dissolved oxygen, nitrate, nitrite, ammonia and phosphorus, as well as bacterial communities under marron (Cherax cainii) aquaculture, fed fishmeal (FM) and poultry by-product meal (PBM)-based diets for 60¿days. The results unveiled that over the time, feeding has significant impacts on organic waste accumulation, especially ammonia, nitrate, nitrite and phosphate, while no effects were observed on pH and dissolved oxygen. Analysis of 16S rRNA sequence data of water sample indicated significant (P < 0.05) shift of microbial abundance in post-fed FM and PBM water with the evidence of microbial transmission from the gut of marron. Post-fed marron resulted in a significant correlation of Hafnia, Enterobacter, Candidatus Bacilloplasma and Aquitella with the quality and microbial population of water. The results of this study generated valuable knowledge database of microbes-water relationship for better health management practices and production of marron aquaculture fed with FM and PBM diets in under restricted feeding regime with the feeding ratios provided.

Vibriosis caused by luminous Vibrio species is one of the biggest challenges to shrimp industry in Bangladesh. This study aimed to characterize whole microbial communities from Vibrio-infected black tiger shrimp (Penaeus monodon) using 16S rRNA-based amplicon sequencing. A total of 36 disease-free and infected shrimp were collected from six different hatcheries in Bagerhat, Bangladesh. A final pool of 12 samples (n¿=¿6) was created by homogenization of the hepatopancreas samples from three shrimps collected from each hatchery for the same group. The amplicon sequencing data revealed significant (p¿<.05) decrease of alpha diversity measurements and subsequent effects (p¿<.05) on the hepatopancreas microbiota in the infected group, compared to control shrimp. Proteobateria and Aeromonas were the most dominant bacteria at phylum and genus level in both groups and identified as core microbiota in the community. Two bacterial groups at phyla level and eight at genus level were found associated with the alteration of hepatopancreas microbial communities and associated gene functions in vibriosis-infected shrimp, revealed by differential abundance and KEGG pathway analysis. The overwhelming abundance of Citroibacter, Shewanella and Candidatus lineages in vibriosis-infected shrimp needs further investigations.

Probiotic supplements are being used to improve the growth and immune performance of aquaculture species over the last couple of decades. In recent times, black soldier fly (BSF) is considered as one of the promising sources of alternative protein to fishmeal protein in aqua-diets. Since the freshwater crayfish, marron (Cherax cainii), a Western Australian's native and iconic freshwater crayfish species, grows fairly slow under commercial farming environment, this study was aimed to investigate the supplemental effect of BSF and BSF with probiotic bacteria Lactobacillus plantarum (BSFLP) on overall health and immune performance of marron after 56 days of feeding under laboratory conditions. The post-trial data revealed insignificant influences of any diets on growth performance, however, both BSF and BSFLP based diets significantly improved some haemolymph parameters and gut health of marron. High throughput sequence data revealed that both BSF and BSFLP diets significantly improved the diversity of microbial communities including some beneficial bacteria for crustaceans in the hindgut of marron. Further analysis showed that both BSF and BSFLP diets upregulated the expression of some genes in the gut tissue and haemocytes associated with the innate immune response of marron at 48 h post injection. The up-regulation of some immune genes in BSFLP diet group was found significantly linked to OTU abundance for Lactobacillus. The findings of this study could be helpful for improving overall health status of marron.

Background: The leather industry generates huge volume of waste each year. Keratin is the principal constituents of this waste that is resistant to degradation. Some bacteria have the ability to degrade keratin through synthesis of a protease called keratinase that can be used as sources of animal feed and industrial production of biodiesel, biofertilizer, and bioplastic. Majority of the studies focused on keratin degradation using gram-positive bacteria. Not much of studies are currently available on production of keratinase from gram-negative bacteria and selection of best parameters for the maximum production of enzyme. The aim of this study was to isolate and characterize both groups of bacteria from soil for keratinase and optimize the production parameters. Results: A total of 50 isolates were used for initial screening of enzyme production in skim milk, casein, and feather meal agar. Out of 50, five isolates showed significantly higher enzyme production in preliminary screening assays. Morphological and biochemical characterization revealed 60% of the isolates as gram-negative bacteria including two highest enzyme-producing isolates. The isolates were identified as Pseudomonas aeruginosa through sequencing of 16S rRNA gene. Maximum production of enzyme from P. aeruginosa YK17 was achieved with 2% chicken feather, beef extract, and ammonium nitrate as organic and inorganic nitrogen sources and glucose as a carbon source. Further analysis revealed that 3% inoculum, 40 °C growth temperature and 72-h incubation, resulted in maximum production of keratinase. Conclusion: The overall results showed significant higher production of enzyme by the P. aeruginosa YK17 that can be used for the degradation of recalcitrant keratin waste and chemical dehairing in leather industries, thereby preventing environmental pollution.

MicroRNAs (miRNAs) are small non-coding RNAs that control the expression of genes by targeting specific mRNAs. Data related to miRNAs are limited for fish species, with only 16 out of the 30¿000 fish species enlisted in miRBase. In the present review, we have summarized the recent findings on the implication of miRNAs in the teleost reproduction with an emphasis on commercially important candidates. The information related to various miRNAs and their roles during different developmental stages of gonads in nine important species has been compiled. We have focused on the trend of sexual dimorphism in the gonadal miRNA repertoire of teleost species. Species-specific variability is observed in the expression pattern of gonadal miRNAs in both male and female fish. It is noteworthy that our summarization of teleost miRNAs in reproduction has highlighted the gaps in functional information on the identified miRNAs (both conserved and novel). It is only after functional validation of the miRNA targets, one can use these findings to enhance reproductive health and production of the commercial teleost species to boost overall aquaculture sector.

Escherichia coli associated infections are major threats in poultry industry owing to severe economic losses each year. This study was conducted to identify E. coli isolates, to evaluate their antibiotic sensitivity and to find out their virulence patterns from infected broilers of Sylhet city in Bangladesh. Using polymerase chain reaction, a total 20 isolates were identified as E. coli from 11 chickens, exhibiting symptoms like colibacillosis and/or diarrhea. All isolates were positive for type-1 fimbrial adhesion (fimH), followed by putative avian hemolysin (hlyF) in 17 isolates; while none of the isolates was amplified with intimin (eaeA). Among 10 tested antibiotics, 100% of the isolates (n = 20) showed resistance to ampicillin, amoxicillin and tetracycline; but they were 100% sensitive to gentamicin. Organ specific correlations of antibiotic sensitivity were obtained among the isolates through principal component analysis (PCA) and Agglomerative Hierarchical Clustering (AHC). The 16S rRNA data of two multi-drug resistant isolates revealed closed clustering with clinical E. coli strains which could be indication of their zoonotic potential. In conclusion, the results depict higher prevalence of fimH and hlyF genes and drug resistance patterns of E. coli isolates from broilers in Sylhet city of Bangladesh.

Streptococcosis is one of the most important diseases of a wide range of freshwater and marine fishes. Objectives of the present study were to isolate and identify the pathogens of streptococcosis like infection in silver barb (Barbonymus gonionotus) using phenotypic, molecular, and artificial infection challenge test; investigate the antibiogram profile of the pathogenic isolates; and sequence the whole-genome of a virulent strain. Bacteria were isolated from different organs of diseased fish on Streptococcus selective media. Phenotypic identification of randomly selected isolates was performed through morphological, physiological, and biochemical tests. Molecular identification was done by 16S rRNA gene sequence analyses. Artificial infection challenge test was performed in B. gonionotus through intraperitoneal injection. To investigate the antibiogram profile, 15 commercial antibiotic disks and 24 medicinal plant extracts were screened by disk diffusion-assay. The whole-genome of a selected strain BFPS6 was sequenced by using an Illumina MiSeq platform. In this study, a total of 20 out of 30 randomly selected isolates were phenotypically identified as Enterococcus sp. The 16S rRNA gene sequence homology of 10 randomly selected isolates revealed that these isolates exhibited 99¿100% sequence homology with Enterococcus faecalis. The lowest and highest median lethal doses (LD50) of the pathogenic isolates in B. gonionotus were obtained 5.54 × 106 CFU/ml and 8.03 × 108 CFU/ml for the isolates BFPS6 and BFPS13, respectively. The isolates showed resistance to multiple antibiotics but susceptibility to three medicinal plant extracts (Spondias mombin, Allium sativum, and Syzygium aromaticum). The genome size and average G+C content of the strain BFPS6 were 2866,855 bp and 37.5%, respectively. The genome contains 2743 CDS, 55 tRNA, 5 rRNA, 9 repeat regions, and one CRISPR sequence. This is the first report on identification and whole-genome sequence of E. faecalis isolated from B. gonionotus suffering from streptococcosis like infection.

The effects of feeding different levels of poultry by-product meal (PBM) replacing fishmeal (FM) protein, supplemented with tuna hydrolysate (TH) and Hermetia illucens (HI) larvae, on the growth, fillet quality, histological traits, immune status, oxidative biomarker levels and gut microbiota of juvenile barramundi, Lates calcarifer were investigated for six weeks. Barramundi were fed four isonitrogenous and isolipidic diets in which a FM based diet was used as the Control diet (Diet1) and compared with other non-FM diets containing 80%, 85% and 90% PBM along with the concurrent supplementation of 5% and/or 10% TH and HI larvae meal. These treatment diets were designated as 80PBM10TH+10HI (Diet2), 85PBM5TH+10HI (Diet3) and 90PBM5TH+5HI (Diet4). The growth and condition factor of fish fed 80PBM10TH+10HI and 85PBM5TH+10HI were significantly higher than the Control. Total saturated, monounsaturated and polyunsaturated fatty acid retention in the fish muscle increased in fish fed PBM-based diets, supplemented with TH and HI larvae meal, with no adverse effect on post-harvest characteristics such as texture and colour of fish fillets. Improvement in serum total bilirubin and total protein content was found in all fish fed TH and HI larvae supplemented PBM. Similarly, immune response showed a significant increase in fish fed non-FM test diets than the Control. In the distal intestine, supplementation of any quantities of TH and HI larvae to PBM led to an increase in the microvilli density and neutral mucins while the number of goblet cells in the skin were unchanged. Liver, kidney, and spleen histology demonstrated a normal structure with no obvious changes in response to all test diets. Bacterial diversity increased in fish fed Diets 2 and 3 with a high abundance of Proteobacteria in Diets 1 and 4 and Firmicutes in Diets 2 and 3. The fish on test diets showed a lower abundance of genus Vibrio. Fish fed TH and HI larvae supplemented PBM diets showed lower infection rate to V. harveyi than the Control. Collectively, concurrent supplementation of TH and HI larvae could improve the quality of PBM diets with positive effects on growth, fillet quality, intestinal health, immunity, and disease resistance.

Yogurt is one of the most frequently consumed dairy products for nutritional benefits. Although yogurt is enriched with probiotics, it is susceptible to spoilage because of the presence of pathogenic microbes. Spoiled yogurt if consumed can cause food-borne diseases. This study aimed to assess the nutritional composition and microbiome diversity in yogurt manufactured in Bangladesh. Microbial diversity was analyzed through high-throughput sequencing of bacterial 16S rRNA gene and fungal internal transcribed spacer (ITS) region. From nutritional analysis,¿significantly (P < 0.05) higher pH, fat, moisture, total solid and solid-non-fat contents (%) were observed in sweet yogurt. Following the classification of Illumina sequences, 84.86% and 72.14% of reads were assigned to bacterial and fungal genera, respectively, with significantly higher taxonomic richness in sour yogurt prepared from buffalo. A significant difference in bacterial (Ppermanova = 0.001) and fungal (Ppermanova = 0.013) diversity between sweet and sour yogurt was recorded. A total of 76 bacterial and 70 fungal genera were detected across these samples which were mostly represented by Firmicutes (92.89%) and Ascomycota (98%) phyla, respectively. This is the first study that accentuates nutritional profiles and microbiome diversity of Bangladeshi yogurt¿which are crucial in determining both active and passive health effects of yogurt consumption in individuals.

Present study aimed to investigate the physiological and molecular response of 28-days starved marron (Cherax cainii), an economically important freshwater crayfish species. Thirty two marron were randomly distributed into two distinct groups in quadruplicated tanks with a density of 4 marron per tank. The feeding- group of marron was fed the marron basal diet on every day while the starved- group was deprived of feed for four weeks. The results showed a significant (P <.05) reduction in protein, energy and fat contents in the tail muscle of marron due to starvation. The microbiome data showed significant modulation of bacterial abundance at both genus and species level in post-starved marron where core microbiota was replaced by Vibrio. The predicted KEGG metabolic pathway based on 16S rRNA data showed a notable shifting of biosynthesis capability towards nutrients stress response activity in starved marron. Significant impact of starvation on the relative expression level of immune genes was observed in starved marron where pro and anti-inflammatory cytokine genes were significantly down-regulated in fourth weeks. This study provides an insights into the immune response and health of marron exposed to starvation under farming conditions.

Streptococcosis has been emerged as major threat for mass mortality of cultured tilapia in Bangladesh. Present study aimed to investigate the effects of probiotic bacteria Lactobacillus plantarum on biochemical composition, gut microbiota, innate immune response and disease resistance of Oreochromis niloticus against Enterococcus faecalis, the causative agent of streptococcosis in tilapia farming. A total of 60 fish were randomly distributed into six different aquaria with two distinct treatment groups, viz. control and probiotic. After 7 days acclimation, control group was fed with commercial tilapia feed while probiotic group was served with 1.02 × 109 CFU/mL per kg of L. plantarum supplemented diet for 56 days. At the end of trial, the results showed no significant changes in growth and biochemical composition of fish meat, however, the probiotic diet significantly modulated the gut microbiota, improved innate immune response and disease resistance of experimental tilapia. Significant increase in the abundance of Lactobacillus and Lactococcus and upregulation of cytokine genes were detected in post-feeding probiotic fish gut, wherein differentially abundant bacteria were found positively correlated with the immune gene expression. Furthermore, dietary supplementation of probiotic bacteria revealed significant reduction of mortality following injection with 2 × 105 CFU/mL of E. faecalis. The results of this study, suggest that dietary supplementation of L. plantarum could be used as potent probiotic to control streptococcosis in commercial tilapia farming.

An experiment was conducted to study the effect of microbial levan supplemented diet on innate and adaptive immune responses as well as stress biomarkers of pathogen challenged Labeo rohita fingerlings. Fish were randomly divided into two groups in triplicates and fed with a pre-standardized dose of 1.25% (w/w%) levan and without levan (control group) for 60 days. Post completion of the feeding trial, fish from both treatment groups were challenged with a pathogenic strain of A. hydrophila and samples were collected on different time points till 96 h. The results of the study showed that supplementation of microbial levan significantly upregulated the mRNA levels of TLR22, ß-2 M and IFN-¿, and downregulated TGF-ß in the intestine, gill, kidney and liver in a time-dependent manner. Significant decrease in the expression of TGF-ß in the hepatic cells was noticed at later time points of 24, 48 and 96 h post-challenge with the highest of 2.66 fold at 48 h. Upregulation of the ß-2 M gene at 48 h and 96 h was noticed in the intestine, gill, liver and kidney. Maximum expression of IFN-¿ was observed at highest time points of 96 h in both intestine and gill, whereas 3.4-fold in the liver at 6 h and 2.6-fold in the kidney at 24 h was noticed. TGF-ß expression analysis displayed the significant downregulation with a maximum decrease of 2.6 fold in the gill at 12 h, while a 2.3-fold and 2.5-fold decrease were noticed at 48 h in the kidney and liver, respectively. Total immunoglobulin level and myeloperoxidase content of A. hydrophila infected rohu increased upto 24 h in the levan-fed group compared to the control. Remarkable decrease of stress biomarkers such as serum cortisol, blood glucose, and HSP70 in the liver, muscle and gill, was observed due to dietary feeding of levan at 24 h. In totality, the present study revealed the significant upregulation of immune responsive genes (TLR22, ß-2 M and IFN-¿), downregulation of regulatory gene (TGF-ß) and decrease of HSP70, cortisol and blood glucose with dietary microbial levan supplementaion in infected Labeo rohita fingerlings. These findings on the modulation of immune responsive gene and stress parameters provide an understanding about the molecular basis of the function of prospective prebiotic, microbial levan.

Seven isonitrogenous (47.0% crude protein), isolipidic (14.0% crude lipid) and isocaloric (22.0 MJ Kg-1 gross energy) diets were formulated and tested in a feeding trial to evaluate the efficacy of raw and enzyme-treated alga, Spirulina platensis (SP) to replace fishmeal (FM) on growth, digestibility, biochemical response and health status of juvenile barramundi, Lates calcarifer. The control diet had fishmeal as the main protein source and was progressively substituted with raw (RSP) or enzyme-treated SP (ESP) protein at 10%, 20%, and 40% of FM. Diets were fed in triplicated groups of barramundi with an initial body weight of 9.20 ± 0.3 g/fish, thrice a day for 8 weeks. The results showed that the weight gain (WG) and feed conversion ratio (FCR) were not negatively affected by the replacement of dietary ESP, while dietary 40% RSP level of FM significantly reduced (P <.05) the WG and FCR than the control. The apparent digestibility coefficients (ADCs) of protein and lipid were similar up to 20% replacement but decreased at 40% replacement by ESP and RSP. A significant (P >.05) reduction of intestinal fold height, microvillus height, and thickness of muscular wall were recorded in fish fed RSP20, RSP40, and ESP40 diets than the control. The histopathological alterations in liver and muscle appeared in fish fed both ESP and RSP diets at 40% replacement of FM. Except for the fish fed 10% of RSP diet, all other fish, the exposure to transportation stress increased the plasma glucose levels although the plasma cortisol levels were not changed in various feeding levels. Overall, this study demonstrates that up to 20% RSP or 40% ESP unaffected the growth performance of juvenile barramundi while replacement of 40% FM with SP either raw or enzyme-treated impacted the health status in terms of the liver, muscle and intestinal morphology of fish.

Water quality has significant impacts on the health and immune responses of aquaculture species. This study aimed to analyse and compare the effects of two biological filters namely, gravel and, Bio-Ball with a recently developed filter called Water-cleanser on regulation of water quality parameters, health and immune response of marron reared in plastic tanks for 60 days. Results showed that addition of Bio-Ball significantly (P < 0.05) reduced the concentration of ammonia, nitrate and phosphate while Water-cleanser showed the ability to reduce ammonia and nitrate from water in aquaculture tanks. Although the biological filters had no significant effect on marron growth but inclusion of Bio-Ball and Water-cleanser positively influenced the biochemical composition of tail muscle and some haemolymph parameters of marron. The next generation sequence data demonstrated higher bacterial diversity in the hindgut of marron with Water-cleanser, followed by Bio-Ball and gravel, respectively. In addition, the predicted metabolic pathways revealed a significantly higher bacterial activity and gene function correlated to metabolism and biosynthesis of protein, energy and secondary metabolites in Bio-Ball and Water-cleanser. Bio-Ball and Water-cleanser were also associated with up-regulation of innate immune responsive genes of marron gut. Overall, Bio-Ball and Water-cleanser proved to have higher water remediation and immune response modulation capabilities, and therefore could be used as preferred filters for growth of beneficial bacteria in crayfish culture.

Bacterial communities strongly influence the digestion, health and immune status of fish. This study investigates the microbial distribution of the anterior, middle and distal gut sections of three economically important carp species in Bangladesh, rohu, catla and mrigal (commonly known as Indian major carps), using 16S rRNA-based Illumina sequencing technology. The alpha-diversity measurement with one-way ANOVA indicated high species richness, Shannon and Simpson indices in the middle and distal gut, while the anterior gut of IMCs had the lowest diversity. At the phylum level, there was high abundance of Proteobacteria in the GITs of rohu and mrigal, whereas Fusobacteria was dominant in the anterior and middle guts of catla. At the genus level, diverse microbial communities were identified across the three GIT sections, with six indicator genera found in rohu, catla and mrigal, as revealed by linear discriminant analysis (LDA) at a 0·05 level of significance. Of the 218 genera identified, only 33 were common across the anterior, middle and distal guts of all three species. Bacterial diversity was significantly higher (P¿<¿0·05) in mrigal, followed by catla and rohu, respectively. Alongside the common bacteria Aeromonas, Enterobacter and Serratia, the overwhelming abundance of Cetobacterium, Shewanella and Plesiomonas warrants further investigation. Significance and Impact of the Study: This study investigates the microbial communities of the gastrointestinal tracts (GITs) of three Indian major carp (IMC) species¿rohu, catla and mrigal, obtained from a polyculture pond under the same feeding regime. Diverse microbial communities were found, with significantly different relative abundances and diversities of phyla and genera. The results provide valuable information on GIT microbial communities that may be useful for nutrition and health management in IMCs.

This study evaluated the effects of dietary supplementation of probiotic bacteria, Bacillus mycoides on the gut microbiota and health status of marron (Cherax cainii) under laboratory conditions. A total of 40 marron were randomly distributed into eight different tanks and two different groups in quadruplicates. The control group were fed only basal marron pellet while the second group were fed 108 CFU/g of probiotic bacteria in the diet. After 60 days of feeding trial, the health and immune indices of marron including protein and energy in the tail muscle, and total haemocyte counts in the haemolymph were positively influenced (p < 0.05) by the probiotic diet. The 16S rRNA sequence data revealed distinctly different microbial communities in the hindgut of marron with two different feeds after trial where Vibrio and Holdemania were the significantly abundant bacteria in the control and probiotic fed groups, respectively. In addition, the qRT-PCR analysis revealed upregulation of cytokine genes associated with the immunity and health status of crayfish indicated the beneficial effects of probiotic bacteria on the immune status of marron. Our results suggest that the diet supplemented with probiotic bacteria B. mycoides positively influenced the health status, immune indices and microbial composition of the marron gastrointestinal tract.

The purpose of the study was to investigate the impacts of dietary narrow-leafed lupin Lupinus angustifolius kernel meal (LKM) on growth performance, feed utilization, digestibility, haematology, as well as liver and intestinal health of juvenile cobia Rachycentron canadum. Five isonitrogenous (crude protein around 460.0 g/kg) diets were formulated to replace 0, 200, 400, 600 and 800 g/kg of fishmeal protein with the protein from LKM designated as LKM0, LKM20, LKM40, LKM60 and LKM80, respectively. Triplicate groups of cobia were fed the respective diets for a period of 56 days. At the end of the feeding trial, growth indices expressed as final body weight (FBW) and specific growth rate (SGR) decreased following the linear and quadratic models with increasing levels of LKM. Feed intake, protein retention and energy retention followed the same trend, whereas, feed conversion ratio (FCR) and hepatosomatic index (HSI) followed a reverse trend with FBW and SGR. A significant decrease in the apparent digestibility coefficient of protein, lipid and dry matter was observed in cobia when more than 400 g/kg protein from fishmeal was substituted by LKM protein. Haematological indices such as red blood cell and haemoglobin declined significantly in fish fed with LKM60 and LKM80, while a significant increase was observed in aspartate transaminase and alanine aminotransferase. Histopathological alterations including higher lipid deposition, necrosis and vacuolization were found in the liver of fish fed higher (>600 g/kg) fishmeal protein replacement levels. Similarly, intestinal structure was affected by higher replacement level of fishmeal protein (>600 g/kg) as fold height and goblet cells decreased significantly with increasing levels of LKM. The results indicate that 200 g/kg fishmeal protein can be replaced by protein from LKM and substitution beyond that level adversely affected the growth performance of cobia.

The present study investigated the supplemental effects of tuna hydrolysate (TH) in poultry by-product meal (PBM) and dietary fishmeal (FM) diets on antioxidant enzymatic activities, gut microbial communities and expression of cytokine genes in the distal intestine of juvenile barramundi, Lates calcarifer. Fish were fed with fermented (FPBM + TH) as well as non-fermented PBM (PBM + TH) and FM (FMBD + TH) diets with 10% TH supplementation for 10 weeks. A basal diet prepared without TH supplementation served as control. The results showed that the activity of glutathione peroxidase was significantly higher in FPBM + TH than the control, while the malondialdehyde and catalase activities were unchanged. FPBM + TH diet significantly (P < 0.05) upregulated the pro-inflammatory cytokines including IL-1ß and TNF-a while considerable downregulation (P < 0.05) was observed in the mRNA expression levels of anti-inflammatory cytokine, IL-10 in the distal intestine of fish. The 16SrRNA analysis using V3¿V4 region evidenced the ability of FPBM + TH to modulate the distal intestinal gut microbiome, augmenting the richness of Firmicutes and Fusobacteriaat at phylum level and Bacillus, Lactococcus and Cetobacterium at genus level. All these results have shown that fermented PBM with TH supplementation could improve the antioxidant capacity and inflammatory responses of juvenile barramundi while influencing the microbial communities at both phylum and genera levels.

In an effort to reduce the use of fishmeal (FM), the effect of using protein from poultry by product meal (PBM) along with the supplementation of three different fish protein hydrolysate (FPH) including yellowtail kingfish, carp and tuna hydrolysate (designated as KH, CH and TH, respectively) were evaluated in juvenile barramundi for growth performance, fillet quality, mucosal immunity, serum biochemistry, immune response and infection against Vibrio harveyi. Fish were fed a FM based control diet + three isonitrogenous and isolipidic diets containing 90% of PBM protein supplemented with different types of hydrolysates: 90% PBM +10% KH (90PBM + KH), 90% PBM + 10% CH (90PBM + CH) and 90% PBM + 10% TH (90PBM + TH). Growth performance and indices were unaffected by the hydrolysate supplemented diets when compared to the control. FPH supplemented PBM diets resulted in improved muscle quality by improving poly unsaturated fatty acids (PUFA), ¿n-3, ¿n-6 and ¿n-9, and health related lipid indexes were not affected. The internal architecture of spleen and kidney were not altered by test diets whilst FPH supplemented PBM modulated acidic mucins in intestine and skin of fish. Improved infection rate in response to two weeks post infection with V. harveyi in the FPH supplemented diets was further associated with an increased serum immune response and a concomitant regulation of proinflammatory and inflammatory cytokines in the head kidney. Serum biochemistry including alanine transaminase (ALT), glutamate dehydrogenase (GLDH) and total bilirubin (TB) showed a decreasing trend both in pre-challenge and post-challenge barramundi fed FPH supplemented diets whereas cholesterol level decreased significantly in post-challenge groups fed 90PBM + KH and 90PBM + TH than pre-challenge barramundi. This study signifies that supplementation of 10% with different three FPH, hydrolysed by an alcalase® enzyme in PBM-based diets for barramundi could be good strategies to overcome the negative consequences triggered by animal by-product ingredients.

The present study compared the nutrient compositions of peanut seed (Arachis hypogaea) subjected to three processing techniques; mechanical pressing, fermentation and germination. The study also evaluated the effect of inclusion level (0%, 15 %, 30 % and 60 %) on nutrient digestibility of these meals by juvenile barramundi (Lates calcarifer) and the subsequent discharge of nutrients to the rearing water. A diet containing no peanut meal served as a reference diet. Barramundi of average weight of 31.2 ± 1.2 g were fed on 10 test diets for 4 weeks. Germination processing significantly (p < 0.05) lowered the carbohydrate and hydrolysed fat contents of peanut meal and its protein level. Fermentation and germination significantly affected the protein quality of peanut meal producing a greater proportion of short peptides (<35 kDa) than achieved by mechanical pressing. Fermented peanut meal at any inclusion level resulted in significantly increased ADC of protein, while 60 % inclusion of mechanically pressed and germinated PM reduced the diet ADC of protein. Ionic phosphate (PO4-), chemical oxygen demand (COD) and total suspended solids (TSS) of rearing water of holding barramundi fed peanut meal-based diets were not different to the reference diet measured two, four and six days after water circulation of tanks was stopped. However, after six days, NH4+ decreased in tanks of barramundi fed fermented peanut meal diet while it increased in tanks fed mechanically pressed peanut meal and of those 60 % germinated peanut meal. Overall, it can be concluded that fermentation processing of peanut meal improves its protein digestibility, increases the production of short chain peptides and reduces the concentration of NH4+ entering holding water compared to diet using germinated or mechanically pressed peanut meal.

A feeding trial was carried out to evaluate the effects of substitution of fishmeal (FM) by dietary poultry by-product meal, fermented by Lactobacillus casei and Saccharomyces cerevisiae on growth, intestinal health, microbial composition, immune related cytokines and disease resistance of freshwater crayfish, marron (Cherax cainii) against Vibrio mimicus. Two isonitrogenous and isocaloric diets were formulated by replacing FM protein with fermented poultry by-product meal (FPBM) protein at 0% (Control) and 75% (FPBM), and fed marron for 70 days. The results indicated no significant difference (P > 0.05) in final body weights between two groups of marron, whilst intestinal microvilli number per fold was increased in marron fed FPBM than the control. The 16S rRNA sequences revealed an increased number of Lactobacillus and Streptococcus, and decreased number of Aeromonas at genus level in the distal intestine of marron fed FPBM. Marron fed FPBM showed up-regulated expression of IL-8, IL-10, and IL-17F genes in the distal intestine. Significantly (P < 0.05) increased lysozyme and phagocytic activity, and higher survival was found in marron fed FPBM following a bacterial challenge with Vibrio mimicus. Therefore, it is concluded that FPBM is beneficial to marron in terms of microbial community, immune-related cytokines and disease resistance against V. mimicus.

This study reports the draft genome sequence of a promising fish probiotic, Bacillus subtilis strain WS1A, that possesses antimicrobial activity against Aeromonas veronii and suppressed motile Aeromonas septicemia in Labeo rohita. The de novo assembly resulted in an estimated chromosome size of 4,148,460 bp, with 4,288 open reading frames.

Background and Objective: The mucoviscosity associated gene A (magA) in the hypermucoviscous variants of K. pneumoniae is reported to be associated with invasive infections and considered a virulence factor. We sought to analyze the magA genes in K. pneumoniae isolates in the clinical specimen collected from Bangladesh. Methods: We established a multicenter cohort of patients with Klebsiella infection hospitalized at 05 different hospitals between September 2016 and April 2017. We collected 313 K. pneumoniae isolates from patients who consented to participate in the study. The isolates were evaluated for harboring the magA genes using a single-tube multiplexed polymerase chain reaction. The magA genes were analyzed by PCR-RFLP technique using two enzymes, namely PciI and SmaI. Antibiogram assay using 12 commercially available antibiotic discs was performed on all the isolates. Results: The presence of K. pneumoniae specific gene (ureD) was confirmed in all the isolates. The percentage of isolates harboring the magA gene was 7.34%(23 isolates), the majority of which was collected from the patients admitted in intensive care units (16 isolates, 69.6%), and infectious diseases wards (5 isolates, 21.7%). PCR-RFLP analysis revealed that for 7 out of 23 isolates, where Sma1 could not cleave the magA gene. All the isolates showed resistance to ampicillin, carbenicillin cefradine, chloramphenicol, erythromycin, kanamycin, and sulphamethoxazole, though the extent was varying. However, imipenem showed 100% sensitivity to all the tested isolates. Conclusion: This study demonstrates the presence of the magA gene in multidrug-resistant clinical isolates of K. pneumoniae collected from Bangladesh.

This study aimed to investigate the combined effects of two most potent probiotic bacteria Lactobacillus acidophilus and Lactobacillus plantarum on overall health and immune status of freshwater crayfish, marron under laboratory conditions. A total of 36 marron were distributed into six different tanks and two different feeding groups, control and probiotic-fed group. After acclimation, control group was fed with basal diet while probiotic group was fed 109 CFU/mL per kg of bacterial supplemented feed for 60 days. The results showed no significant differences in weight gain, however, probiotic feed significantly enhanced some hemolymph parameters and biochemical composition of tail muscle. Histology data revealed better hepatopancreas health and higher microvilli counts in the marron gut fed probiotic diet. The probiotic bacteria triggered significant shift of microbial communities at different taxa level, mostly those reported as beneficial for crayfish. The probiotic diet also enriched the metabolic functions and genes associated with innate immune response of crayfish. Further correlation analysis revealed significant association of some taxa with increased activity for hemolymph and immune genes. Therefore, dietary Lactobacillus supplementation can modulate the overall health and immunity as well as gut microbial composition and interaction network between gut microbiota and immune system in crayfish.

The search for suitable fish meal replacements in aqua-diets is a salient agenda in the constant effort of making aquaculture practices more sustainable. In this study, we tested four customised diets composed by systematic inclusion of pre-selected fish meal substitutes, lupin kernel meal, BSF meal, TH and PBM on growth, metabolism, cytokine profile, gut morphology and microbiota of juvenile Lates calcarifer. Five isoproteic and isoenergetic diets were prepared viz. FM100 as a control (without fish meal substitute), while FM75, FM50, FM25 and FM0 indicates replacement of fish meal (FM) at 25%, 50%, 75%, and 100%, respectively by a mixture of four different pre-selected non-fish meal (NFM) ingredients. Fish fed FM100, FM75, FM50, FM25 exhibited consistent growth and haematological response, while the fish fed no fishmeal (FM0) showed significant decline in final body weight (FBW) and specific growth rate (SGR). The poor growth performance was correlated with a decrease in villous width, microvilli height and goblet cells density. A significant shift in abundance profile of Psychrobacter in the gut microbial profile of fish fed FM50 was noticed compared to fish fed FM100. The results of qRT-PCR showed up-regulated expression of innate immune responsive genes in the FM50 group. The adverse impacts on growth performance and gut health of fish fed FM0 suggest that the complete substitution of fishmeal is not advisable and the inclusion range of these alternatives should be decided for a species only after examining their effect on maximal physiological performance.

Aerolysin (aer) is one of the most important and abundant virulence factors in the infection of fish by Aeromonas veronii. A comprehensive study on the molecular characterization and pathogenicity of the aer gene from 34 A.¿veronii isolates from diseased carp and catfish was carried out and its interactome was analysed to observe the functional correlations between aer and other proteins within the A.¿veronii network. The PCR-based amplification of aer from the 34 isolates of A.¿veronii showed more aer-positive isolates from catfish with a high pathogenic potential in the in vivo challenge test than the carp fish. The analysis of aer gene sequence from challenged fish revealed significant sequence divergence according to the types and geographical distribution of the fish. The networking analysis of aer from the model A.¿veronii B565 revealed histidine kinase (cheA) as the most functional interacting partner. The study of the interaction between aer from the experimental A.¿veronii and cheA demonstrated that the A chain of cheA plays a more important role than the corresponding B chain during contact, and a linker sequence of 15 residues controlled the entire interaction process. Therefore, cheA could be an excellent drug target for controlling A.¿veronii infection of fish.

Length¿weight relationships (LWRs) were determined for three Carangid fish, [Alepes vari (Cuvier, 1833), Uraspis uraspis (Günther, 1860) and Carangiodes oblongus (Cuvier, 1833)] inhabiting in the Bay of Bengal coast, Bangladesh. A total of 147 individuals of three species were sampled from four different sampling points of the Bay of Bengal coast, Bangladesh and examined from April 2017 to October 2017. Fishermen operated beach seine nets (mesh size BSN, 30¿mm) and drift gillnet (10.5¿cm) twice in each sampling campaign to catch the studied species. The calculated value for parameter ¿b¿ of LWRs were 3.003, 2.945 and 2.980 for A. vari, U.uraspis and C. oblongus, respectively.

This study aimed to characterize the bacterial communities in rearing water treated with commercial plastic biological ball filters named as Bio-ball in marron culture for 60¿days. Inclusion of Bio-ball in the aquaculture tanks showed improvement in water quality parameters and enrichment of bacterial communities in terms of operational taxonomic units. The water treated with Bio-ball showed significantly less nitrate, nitrite, ammonia, phosphorus and high dissolved oxygen concentration than untreated control group. At phylum level, Proteobacteria was dominant in both control and treated water, whereas Firmicutes was found to be significantly (P¿<¿0·05) enriched in Bio-ball treated water. Among the classified genus, Aquabacterium and Polunucleobacter were most dominant in control and Bio-ball treated water respectively. Linear Discriminant Analysis Effect Size exhibited 31 indicator bacterial genus, 10 in control and 21 in treated condition, suggesting the enrichment of microbial lineages with addition of Bio-ball. The bacteria Haliscomenobacter, Hypnocyclicus, Pajaroellobacter and Vibrio were found to be significantly (P¿<¿0·001) correlated with higher pH, nitrate, nitrite, phosphorus and ammonia in control tanks, whereas Corynebacterium was linked to higher temperature in treated water. Overall results suggest that Bio-ball filter media significantly improved the water quality and microbial populations in aquaculture tanks. Significance and Impact of the Study: The results of this study revealed the positive impacts of Bio-ball in enrichment of microbial flora associated with the degradation process of nitrogenous and organic compounds. Bio-ball also showed the capability to prevent the colonization of harmful bacteria, and favoured the growth of beneficial microbes in aquatic system. This study therefore could pave the ways of increasing the aquaculture production by improving the water quality.

A two phased feeding trial was conducted to evaluate the effects of alternative protein sources on the immunophysiological responses of marron. During the phase I, marron were fed with five alternative protein supplemented diets for 90 days, while in phase II, the same marron were exposed to elevated temperature (30 °C) and their immunophysiological responses were investigated post exposure. Five isoproteic (crude protein 30%) and isoenergetic diets were prepared by containing fishmeal, poultry by-product meal, feather meal, lupin meal, and meat and bone meal as the main protein source. A hundred and fifty juvenile marron (Cherax cainii) of the average weight 9.09 ± 0.21 g were randomly distributed into 15 tanks (three replicates per feeding treatments). In the Phase I, general immune response parameters, such as, total haemocyte count (THC), proportion of hyaline cells, neutral red retention time (NRRT), phagocytic rate (PR), heamolymph bacteraemia, and condition indices of marron were investigated. The highest (P < 0.05) THC among dietary protein sources was obtained in marron fed with PbM at the end of experiment. Marron fed with FeM protein sources resulted in the highest survival rate followed by PbM fed group. Longer microvilli length (3.83 ± 0.18 µm) was demonstrated in marron fed with PbM diet. Diets containing FM and PbM protein sources revealed significantly (P < 0.05) lower number of microvilli/group than diets containing FeM and LM. The results demonstrated that different dietary protein sources in the marron diets did not detect significant (P > 0.05) change of the condition indices throughout the experiment period, however highest Hiw and Hid was recorded in marron fed with PBM at day 45. The PR of marron fed dietary protein from PbM did not change significantly after temperature exposure. Increased NRRT, PR and haemolymph bacteraemia was observed with dietary feeding of FM at the end of the trial. However, results revealed that PbM could be an alternative protein source for culture of marron as reflected in terms of increased THC, longer microvillus length and improved susceptibility to high temperature exposure. Overall, result could serve as useful baseline data in developing cost effective potential diets for marron aquaculture.

The shrimp white spot syndrome virus (WSSV) causes significant damage to aquaculture production worldwide but a vaccine, eliciting the immunogenicity of shrimps against WSSV has yet to be developed. Thus, a programmed immunoinformatics study was conducted to find out the potential immunogens based on genome-wide screening of WSSV envelope proteins. The measurements of the phylogenetic and evolutionary distances found the common geographical routes of three countries, where the proteins from other six countries were clustered together. Among all the four major envelope proteins i.e., VP19, VP24, VP26, and VP28; AAO69663.1 from VP26 showed the highest antigenicity and thus selected for further studies. The properties of the secondary and tertiary structure including the modelled 3D protein revealed that the protein had all the properties required for a protective immunogen. The peptide regions ranging from 99 to 115 and 98 to 106, representing the sequences ¿VTAPRTDPAGTGAENSN¿ and ¿TVTAPRTDP¿ were found to be most effective regions for B-cell linear and cytotoxic T lymphocyte (CTL), respectively. The CTL epitope also showed a strong and stable interaction with the MHC class I and class II molecules, reported to be found in fish. Therefore, the present epitope could be used as a potential vaccine candidate against WSSV.

This study was conducted to characterize the causative agent of streptococcosis in tilapia (Oreochromis niloticus) and control of Streptococcus infection by means of garlic (Allium sativum) supplementation. The morphological, biochemical and polymerase chain reaction amplification confirmed 11 isolates belong to Streptococcus iniae from the infected fish eyes and tissue samples. Random screening of 12 well-known medicinal plant parts against S.¿iniae revealed the garlic extract as the most effective herbal recovery against Streptococcus infection. In vivo challenge test with dietary supplementation of garlic powder significantly improved survival rates of fish against S.¿iniae infections, and modulate the microbial community and cytokine gene expression profiling in the intestine of the experimental tilapia. Among the two garlic supplemented treatments, 1.0¿g garlic supplemented diet significantly increased (p¿<¿0.05) the survival rates of tilapia and the gut bacterial operational transitional units abundance for Proteobacteria and Tenericutes, the phyla associated with healthy intestinal flora. The bacterial diversity index also found high with garlic supplemented diets. Significant upregulations of IL-10 and IL-17F gene expression in the intestinal tissue were observed with 1.0¿g garlic supplemented diet where IL-8 and IL-1ß expression levels were relatively static. The dietary supplementation of garlic, therefore, could be effective in the prevention of S.¿iniae infection in fish.

Aims: This study conducted bacterial community, virulence and antibiogram profiling inside the hindgut and skin of freshly caught hilsa fish and those sold at markets. Methods and Results: The results of 16S rRNA-based high-throughput sequencing showed a higher number of bacterial genera in marketed fish samples than in fresh fish samples. The total operational taxonomic units, genus counts and diversity index were significantly higher (P¿>¿0·05) in marketed fish, which also had abundant pathogenic bacterial groups. Skin samples had a lower profusion of pathogenic bacteria than gut samples. A total of 52 bacterial isolates from nine species were identified in this study, of which 25 were from a Chittagong market and 22 were from a Dhaka market, whereas only five were from fresh hilsa. The polymerase chain reaction amplification of 12 species-specific virulence genes in the 52 isolates, namely, aer, hly, chxA, toxB, rtxC, sfa, uge, norB, trx, toxA, ipaH, sigA and coa, indicated a high number of positive samples containing Vibrio cholerae, Aeromonas spp., Klebsiella pneumoniae, Escherichia coli and Staphylococcus aureus. Antibiogram profiling of these bacteria against 10 commercial antibiotics showed high-resistance patterns of the isolates against sulfamethoxazole, kanamycin, neomycin, ampicillin and tetracycline. Conclusion: The results reveal the spread of multidrug-resistant bacteria in hilsa fish marketed for human consumption in Bangladesh. Significance and Impact of the Study: This study highlights the risk of spreading environmentally and clinically pathogenic bacteria in fish sold for human consumption in Bangladesh. Such bacteria come from aquatic pollution and poor handling, storage and transportation practices that may predispose fish to major outbreaks of infectious and waterborne diseases.

Aim: Klebsiella pneumoniae is considered to be one of the most frequent bacterial species associated with urinary tract infections (UTIs) and recurrent UTIs (RUTIs) worldwide. The present study aimed to comprehensively characterize K. pneumoniae isolates from women suffering from UTI and RUTIs. Methodology and results: A total of 15 clinical isolates, collected from different hospitals in Bangladesh, were tested for biochemical features, and amplified by PCR. Antibiogram assay was performed by disk-diffusion assay. Phylogenetic and functional features were analyzed using bioinformatics platform. XLSTAT was used for principal component analysis (PCA). PCR amplification using Klebsiella hemolysin gene (khe) confirmed the presence of K. pneumoniae in agarose gel with expected product size of 486 kb. Antibiogram assay revealed all K. pneumoniae isolates to be completely resistant to six out of ten relevant drugs namely ampicillin, cephradine, chloramphenicol, erythromycin, kanamycin and sulfamethoxazole used for treating UTIs in Bangladesh. Sequencing of 16S rRNA gene of clinically significant K. pneumoniae isolates showed a high level of sequence divergence among the isolates from UTI and RUTIs as well as functional features such as SNP variants and restriction sites. Conclusion, significance and impact of study: We surmise that the results could be used as a pipeline for further research in the identification of K. pneumoniae associated with UTI and RUTIs, and treatment of infection.

The present study aimed to evaluate the dietary supplementary effects of black soldier fly (Hermetia illucens) (BSF) meal on the bacterial communities in the distal gut, immune response and growth of freshwater crayfish, marron (Cherax cainii) fed poultry-by-product meal (PBM) as an alternative protein source to fish meal (FM). A total of 64 marron were randomly distributed into 16 different tanks with a density of four marron per tank. After acclimation, a 60-days feeding trial was conducted on marron fed isonitrogenouts and isocalorific diets containing protein source from FM, PBM, and a combination of FM + BSF and PBM + BSF. At the end of the trial, weight gain and growth of marron were found independent of any dietary treatment, however, the two diets supplemented with BSF significantly (P 0.05) enhanced haemolymph osmolality, lysozyme activity, total haemocyte counts, and protein and energy contents in the tail muscle. In addition, the analysis of microbiota and its predicted metabolic pathways via 16s rRNA revealed a significantly (P 0.05) higher bacterial activity and gene function correlated to biosynthesis of protein, energy and secondary metabolites in PBM + BSF than other dietary groups. Diets FM + BSF and PBM + BSF were seen to be associated with an up-regulation of cytokine genes in the intestinal tissue of marron. Overall, PBM + BSF diet proved to be a superior diet in terms of improved health status, gut microbiota and up-regulated expression of cytokine genes for marron culture.

This study aimed to investigate the effects of Clostridium butyricum as a dietary probiotic supplement in fishmeal based diet on growth, gut microbiota and immune performance of marron (Cherax cainii). Marron were randomly distributed into two different treatment groups, control and probiotic fed group. After 42 days of feeding trial, the results revealed a significant (P <0:05) increase in growth due to increase in number of moults in marron fed probiotics. The probiotic diet also significantly enhanced the total haemocyte counts (THC), lysozyme activity in the haemolymph and protein content of the tail muscle in marron. Compared to control, the 16S rRNA sequences data demonstrated an enrichment of bacterial diversity in the probiotic fed marron where significant increase of Clostridium abundance was observed. The abundance for crayfish pathogen Vibrio and Aeromonas were found to be significantly reduced post feeding with probiotic diet. Predicted metabolic pathway revealed an increased activity for the metabolism and absorption of carbohydrate, degradation of amino acid, fatty acid and toxic compounds, and biosynthesis of secondary metabolites. C. butyricum supplementation also significantly modulated the expression level of immune-responsive genes of marron post challenged with Vibrio mimicus. The overall results suggest that C. butyricum could be used as dietary probiotic supplement in marron aquaculture.

Populations of gizzard shad, Anodontostoma chacunda from four adjoining rivers (Karnafuli, Feni, Bhulua, and Moheskhali) of the Bay of Bengal, Bangladesh, were studied to investigate phenotypic structure on the basis of body shape morphometrics using a truss network model. A total of 28 distance variables were extracted by interconnecting 12 morphometric landmarks. The transformed morphometric measurements were subjected to discriminate function analysis (DFA), principal component analysis (PCA), cluster analysis (CA) and univariate analysis of variance (ANOVA). Based on the analysis of variance, all morphometric measurements differed significantly (P < 0.001) between all four populations. A 95% confidence ellipse drawn by DF1 and DF2 clearly separated two groups of A. chacunda from the Moheskhali and Karnafuli populations compared to all others. Their separation was confirmed by cross-validation of the morphometrics analysis with DFA, where the highest correct re-classification rate was found in the Moheskhali (98.4%) and Karnafuli (98.1%) populations. CA analysis also confirmed all river populations into three distinct clusters: Moheskhali population in one cluster, Karnafuli population in another cluster and Feni and Bhulua populations together formed a separate cluster. The observed morphometric variability between the different locations was expressed based on the measurements that particularly related to the head and body shape of the species. The coefficient of variation (CV) result revealed a low intra-population variation from all the rivers. The findings confirmed the presence of different stocks of fish in the four adjoining rivers of the Bay of Bengal.

Studies were conducted to characterize Klebsiella pneumoniae isolates from urinary tract infection (UTI) patients in Sylhet city of Bangladesh. At the same time, all isolates were screened for some common virulence genes and four significant isolates were searched for plasmid number and sizes by mini alkaline-lysis method. Among five tested isolates from female UTI patients, gyrase subunit B2 (gyrb2) amplified in all isolates, lipase and nuclease detected in three isolates and serine protease amplifies in two isolates and gave the expected band of 1130 bp, 517 bp, 1055 bp and 211 bp respectively. Two of four isolates showed 9.82 kb plasmid band on agarose gel. Isolates bearing 9.82 kb plasmid were found to be resistant to multiple commercial antibiotics. At the same time all isolates were screened for in-vitro plate assay for proteolytic, lypolytic and hemolytic activity. Isolates with positive plasmid and more than one virulent gene with gyrB2 showed positive result in in-vitro culture plate with clear zone of proteolysis, hemolysis or lipolysis. This study will be helpful for further study in finding correlation or pattern of virulence properties for K. pneumoniae associated UTI in Bangladesh.

Klebsiella pneumoniae is the second leading causative agent of UTI. In this study, a rapid combined polymerase chain reaction and restriction fragment length polymorphism analysis was developed to identify K. pneumoniae in women, infected with urinary tract infection in the Sylhet city of Bangladesh. Analysis of 11 isolates from women at the age range of 20¿55 from three different hospitals were done firstly by amplification with K. pneumoniae specific ITS primers. All of the 11 collected isolates were amplified in PCR and showed the expected 136 bp products. Then, restriction fragment length polymorphism analysis of 11 isolates were conducted after PCR amplification by 16s rRNA universal primers, followed by subsequent digestion and incubation with two restriction enzymes, Pst1 and Alu1. Seven out of 11 isolates were digested by Pst1 restriction enzymes, six isolates digested by Alu1, and while others were negative for both enzymes. Data results reveal that, women at age between 25 and 50 were digested by both enzymes. A woman aged over than 50 was negative while bellow 20 was digested by only Pst1. The results could pave the tactic for further research in the detection of K. pneumoniae from UTI infected women.

Fungal and bacterial pathogens infect a diverse range of hosts including various plant and animal species. Fungal and bacterial diseases, especially of plants and aquatic animals, such as fish, lead to significant damage to crops and aquaculture, respectively, worldwide. The present study was conducted to isolate and characterize potent Bacillus strains with significant antagonistic activity against the major plant and fish pathogenic fungi and bacteria. We randomly collected 22 isolates of Bacillus from the soil, rhizosphere, and sediment from different parts of Bangladesh. Initial characterization, based on in vitro antagonistic activity on the culture plate, resulted in the selection of four gram-positive Bacillus sp. isolates. Among these, the isolate BC01, obtained from soil demonstrated the highest broad-spectrum anti-bacterial and anti-fungal activities. We confirmed the genus of BC01 to be Bacillus by morphological and biochemical tests as well as using molecular data analysis tools, including the study of 16s rDNA, phylogenetic relationship, and evolutionary divergence scores. The isolate significantly inhibited the mycelial growth of the plant pathogen, Penicillium digitatum and fish pathogen, Aphanomyces invadans in vitro. The anti-bacterial effect of the isolate was also evaluated against Pseudomonas spp. and Xanthomonas spp., the two deadliest plant pathogens, and Aeromonas veronii, Pseudomonas fluorescens, and Streptococcus iniae, three major fish pathogens that are primarily responsible for global aquaculture loss. The results of the present study could pave the way for developing potent drugs to combat microbial infection of plants and fish.

Introduction: Alcaligenes faecalis is a common environmental bacteria that often infects human as an opportunistic pathogen. It rarely causes Urinary Tract Infection (UTI) in human; however, infection brings severe outcomes. Also, the treatment of A. faecalis associated infections with common antibiotics can be difficult due to a high level of antibiotic resistance. Aim: Molecular characterisation of A. faecalis isolate from women suffering from Recurrent Urinary Tract Infections (RUTIs). Materials and Methods: The study has been conducted in the USDA-laboratory of the Department of Genetic Engineering and Biotechnology, Shahjalal University of Science and Technology, Sylhet, Bangladesh. The present study conducted from July 2017 to December 2017, characterised an A. faecalis strain from the women suffering from RUTIs by applying 16s rRNA gene sequencing and phylogenetic analysis. Antibiogram profiling was done by means of disk diffusion assay. Analysis of in-silico SNP variants was done using Geneious software. Results: The isolates were resistant to seven out of ten commercial antibiotics used to treat UTI in Bangladesh. Phylogenetic and evolutionary distance data analysis revealed a close proximity of the study AF1 strain with other A. faecalis strains identified from the environment, especially from a previously characterised water sample. In-silico variants search found nine potentials of Single Nucleotide Polymorphism (SNPs) in the studied strain compared to other environmental A. faecalis bacteria characterised from India, South-Korea, Japan, Mexico and Brazil. Conclusion: The present study revealed the transmission of environmental opportunistic pathogens to human and cause chronic infections in Bangladesh.

Development of molecular techniques for detection of virulence gene is an imperative section in determining the pathogenicity and virulence properties of any isolates because these genes act multi-functionally and multi-factorially. Pathogenesis of Salmonellosis depends upon a large number of multi-functional factors controlled by an array of genes that synergize into the actual virulence of Salmonella. In this study the lipA gene encoding an extracellular lipase and protease specific elastase gene primer allows precise detection of lipase and elastase gene of Salmonella typhimurium by Polymerase Chain Reaction (PCR). This is the first report of lipase gene in Salmonella typhimurium. All isolates of Salmonella typhimurium were collected from patients suffering from typhoid and urinary tract infection (UTI). A total of 14 isolates viz., ST1, ST2, ST3, ST4, ST5, ST6, ST7, ST8, ST9, ST10, ST11, ST12, ST13 and ST14 were used in present study in which lipase gene was amplified in 3 isolates viz., ST1, ST2, ST3 and ST10 with expected PCR product of 1055bp whereas elastase gene was amplified in 6 isolates viz., ST1, ST2, ST4, ST11, ST13 and ST14 and gave the expected 467bp PCR product after visualization under gel documentation system. At the same time 14 samples were examined in-vitro for their putative virulence characteristics viz. proteolytic, lipolytic and hemolytic activity. High lipolytic activity was observed for isolates containing lipase whereas high proteolytic activity was observed for isolates containing elastase. Isolates containing both lipase and elastase showed elevated in-vitro virulence properties with significant P value of <0.05.