Genetic characterization, structural analysis, and detection of positive selection in small heat shock proteins of Cypriniformes and Clupeiformes.

In the current investigation, a total of 42 full-length, non-redundant small heat shock proteins (sHsp) were detected in Cyprinus carpio, Labeo rohita, Danio rerio, Salmo salar, Oncorhynchus mykiss, and Clupea harengus. The sHsp genes were classified into three groups based on phylogenetic analysis. All the sHsps were shown to have higher aliphatic index values, which is an indication that these proteins are more thermally stable. The hydrophilic nature of sHsps was deduced from the fact that all fish species had negative GRAVY scores. In all of the representative fish species, sHsp genes were assigned to distinct chromosomes in an inconsistent and unequal manner. Segmental duplications are the main events that have contributed to the expansion of the sHsp genes in all species. We were also able to determine the selective pressure that was placed on particular codons and discovered several significant coding sites within the coding region of sHsps. Eventually, diversifying positive selection was found to be connected with evolutionary changes in sHsp proteins, which showed that gene evolution controlled the fish adaption event in response to environmental conditions. Clarification of the links between sHsps and environmental stress in fish will be achieved through rigorous genomic comparison, which will also yield substantial new insights.

Multifunctional integration of tungsten oxide (WO3) coating: A versatile approach for enhanced performance of antibiotics against single mixed bacterial infections.

Nanomaterials containing tungsten (TNMs), characterized by diverse nanostructures had been extensively used in biomedical sector. Despite numerous reports focusing on TNM applications in specific biomedical areas, there is a noticeable absence of comprehensive studies that focused on detailed characterization of nanomaterials along with their biological applications. The present work described the structural, morphological, and antimicrobial properties of tungsten oxide (WO3) nanoparticles coated by antibiotics (nanobiotics), and their application on single and mixed bacterial culture. The nanobiotics included in this study were WO3 coated with ampicillin (W+A), WO3 coated with penicillin (P+W), and WO3 coated with ciprofloxacin (C+W). Techniques such as X-ray diffraction (XRD), scanning electron microscopy (SEM), X-ray spectroscopy (EDX), Fourier transforms infrared spectroscopy (FTIR), Rrman spectroscopy, and UV-visible spectroscopy were used to characterize synthesized nanoparticles. The minimum inhibitory concentration of C+W nanobiotic against S. aureus, E. coli, and mixed culture (S. aureus +E. coli) was lower than that of P+W and A+W. The impact of incubation period showed significant differences for each of nanobiotic against S. aureus, E. coli, and mixed culture. However, there were also non-significant differences among incubation periods for antibacterial activity of nanobiotics. It was pertinent to note that percentage variation in susceptibility of S. aureus with respect to mixed culture remained higher as compared to E. coli, indicating it stronger candidate imposing resistance. This paper thus suggested the strategy of coating of antibiotics with with WO3 nanoparticles as an ideal combination for resistance modulation against single and mixed culture bacteria.

Bioaccumulation Pattern and Health Risk Assessment of Heavy Metals in Cirrhinus mrigala at Panjnad Headworks, Bahawalpur, Pakistan.

Heavy metal accumulation in freshwater ecosystem has become one of the major aquatic environmental concerns for freshwater flora and fauna due to their higher stability and bioaccumulation as well as bio-magnification properties. Furthermore, passing through the food web, these heavy metals affect human populations ultimately. This study assessed the heavy metal accumulation in Cirrhinus mrigala in spring, autumn, and winter at different locations (I, II, and III) of Panjnad headwork. Furthermore, the human health risk assessment for the consumption of C. mrigala from the sampling locations was also carried out. Fish were collected from upper (I), middle (II), and lower (III) stream of Panjnad on a monthly basis. The current study evaluated the accumulation of Aluminum (Al), Arsenic (As), Barium (Ba), and Lead (Pb) in various fish organs (liver, kidney, gills, fins, skin, muscles and bones) and assessed their potential hazard to human health through health risk assessment indicators. The results demonstrated a significant difference (p < 0.05) in heavy metal accumulation in different fish organs, seasons, and locations. The accumulation of Al, As, Ba, and Pb were considerably higher in liver and kidney as compared to the other body organs and followed a trend of liver > kidney > gills > fins > skin > bones > muscle and the overall mean concentrations of metals in different body tissues of C. mrigala were in the order of Al > As > Ba > Pb. The results also concluded that C. mrigala caught from the Panjnad headwork is not safe for human consumption due to higher values of TTHQIng (3.76), THQIng for Ba (3.27) and CRIng for As (6.4742).

Molecular Characterization and Phylogenetic Analysis of Casein Gene Family in Camelus ferus.

Camel milk is known for its exceptional medical uses. It has been used since ancient times to treat infant diarrhea, hepatitis, insulin-dependent diabetes (IDDM), lactose intolerance, alcohol-induced liver damage, allergies, and autism. It has the power to treat several diseases, with cancer being the most significant. This study investigated the evolutionary relationship, physiochemical characteristics, and comparative genomic analysis of the casein gene family (CSN1S1, CSN2, CSN1S2, and CSN3) in Camelus ferus. Molecular phylogenetics showing the camelid species clustered casein nucleotide sequences into four groups: CSN1S1, CSN2, CSN1S2, and CSN3. The casein proteins from camels were evaluated and found to be unstable, thermostable, and hydrophilic. CSN1S2, CSN2, and CSN3 were acidic, but CSN1S1 was basic. CSN1S1 showed positive selection for one amino acid (Q), CSN1S2 and CSN2 for three (T, K, Q), and CSN3 showed no positive selection. We also compared high-milk-output species such as cattle (Bos Tarus) and low-milk-yield species such as sheep (Ovies Aries) with camels (Camel ferus) and discovered that YY1 sites are more frequent in sheep than in camels and very low in cattle. We concluded that the ratio of YY1 sites in these species may affect milk production.

Effect of hemoglobin on Nile tilapia (Oreochromis niloticus) kidney (NTK) cell line damage.

Bacteria or viral outbreaks can cause tilapia hemorrhage, ensuring considerable volume of hemoglobin (Hb) into the tissue. However, the hemoglobin toxicity on tissue and high doses also effect on tissue this phenomena is still under consideration. Therefore, current study exploited Nile tilapia kidney (NTK) cells to deeply expose the toxic effect of Hb on NTK cells. Toxicity of Hb on NTK cells was determined in terms of cells growth, expression of iron metabolism and inflammation-related genes, consequently examined antioxidant-related enzymes genes expression, intracellular iron and reactive oxygen species (ROS) contents, and apoptosis-related genes expression. The results showed that Hb and heme significantly inhibited NTK cells growth and up-regulated iron metabolism-related genes expression in different degrees. The Hb and heme activated the expression of pro-inflammatory cytokines (TNF-α, tumor necrosis factor-α; IL-1ß, interleukin 1ß; IL-6, interleukin 6), the anti-inflammatory factor (IL-10, interleukin 10) and the chemotactic factors (IL-4, interleukin 4; IL-8, interleukin 8) through NF-κB pathway, meanwhile activated the expression of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px). Moreover, the Hb significantly increased intracellular iron and ROS contents while the expression of apoptosis-related genes was significantly activated by both Hb and heme. Current investigation suggested that high oxidative activity of Hb could activate iron metabolism- and inflammation-related genes expression, and increase intracellular iron and ROS levels, lead to up-regulated the expression of apoptosis genes in NTK cells.

Genomic and Therapeutic Analyses of Staphylococcus aureus Isolated from Cattle Reproductive Tract.

Staphylococcus aureus is emerging as a ubiquitous multidrug-resistant pathogen circulating among animals, humans, and their environment. The current study focused on molecular epidemiology and evidence-based treatment against S. aureus from bovine endometritis. For this study, n = 304 cattle were screened for endometritis using ultrasonography while presenting case history, and clinical signs were also considered. S. aureus was isolated from endometritis-positive uterine samples which were further put to molecular identification, phylogenetic analysis, susceptibility to antibiotics, and testing of novel drug combinations in both in vitro and field trials. The findings of the study revealed 78.20% of bovine endometritis samples positive for S. aureus, while nuc gene-based genotyping of S. aureus thermal nuclease (SA-1, SA-2, and SA-3) showed close relatedness with S. aureus thermal nuclease of Bos taurus. Drug combinations showed 5.00 to 188.88% rise in zones of inhibitions (ZOI) for drugs used in combination compared to the drugs used alone. Gentamicin in combination with amoxicillin and enrofloxacin with metronidazol showed synergistic interactions in an in vitro trial. Co-amoxiclav with gentamicin, gentamicin with enrofloxacin, and metronidazole with enrofloxacin showed 100%, 80%, and 60% efficacy in treating clinical cases in field trials, respectively. As a result, the study came to the conclusion the higher prevalence of endometritis-based S. aureus, genetic host shifts, narrow options for single drugs, and need for novel drug combinations to treat clinical cases.

Divergent Analyses of Genetic Relatedness and Evidence-Based Assessment of Therapeutics of Staphylococcus aureus from Semi-intensive Dairy Systems.

Use of antibiotics without following standard guidelines is routine practice in developing countries which is giving rise to genetic divergence and increased drug resistance. The current study analyzed genetic divergence and drug resistance by S. aureus and therapeutic efficacy of novel antibiotic combinations. The study revealed that 42.30% (minimum 20%-maximum 70%) of milk samples are positive for S. aureus. Study also revealed seven SNPs in the S. aureus nuc gene (c.53A>G, c.61A>G, c.73T>C, c.93C>A, c.217C>T, c.280T>C, and c.331T>A). Local isolates Staph-2 and Staph-3 were closely related to Bos taurus nuc gene (bovine S. aureus), while Staph-1 was closely related to Homo sapiens (human S. aureus) indicating shifting of host. Change of two amino acids and staphylococcal nuclease conserved domain was observed in all local isolates of S. aureus. The isoelectric points predicted by protParam of Staph-1, Staph-2, and Staph-3 proteins were 9.30, 9.20, and 9.20, respectively. The antibiotic susceptibility profile of S. aureus presented highest resistance against penicillin (46.67%) and glycopeptide (43.33%). When a single antibiotic regimen was adopted in a field trial, the highest efficacy was reported in the case of oxytetracycline (80%) while lowest was presented by azithromycin. Among antibiotics' combined regimen, the highest efficacy (80%) was presented by gentamicin with oxytetracycline: cefotaxime with vancomycin; and ciprofloxacin with vancomycin. The current study concluded rising percentages of S. aureus from dairy milk, proofs of genetic host shifts, and altered responses of in on field therapeutics.

Effect of milrinone on the meiosis resumption and cytoplasm maturation of buffalo oocytes.

Buffalo has many excellent economic traits and it is one of the greatest potential livestock. Compared with cattle, buffalo has poorer reproductivity, it is of great significance to improve the development potential of oocytes. Buffalo oocyte in vitro maturation (IVM) has been widely used in production, but the poor development ability of bovine oocytes IVM limits the development of buffalo reproductivity. Milrinone as a phosphodiesterase inhibitor could affect the maturation of oocytes in goat and mice, but there have been few reported studies in water buffalo. To optimize buffalo oocyte in vitro maturation systems, the effects of phosphodiesterase inhibitor (milrinone) on pre-maturation culture of buffalo oocytes were investigated in this study. Buffalo cumulus-oocyte complexes (COCs) were cultured in medium with different concentrations (0, 12, 25, 50 and 100 mol/l) of milrinone for different times (0, 4, 8, 12, 16, 22 and 24 h). The results showed that the buffalo COCs nuclear maturation process could be inhibited by milrinone (25-100 mol/l) in a dose-dependent manner. The inhibitory effect of milrinone on in vitro maturation of buffalo oocytes did not decrease with the extension of time. This indicated that milrinone can be used as a nuclear maturation inhibitor during the maturation process in buffalo oocytes. In addition, milrinone can inhibit the effect of follicle stimulating hormone (FSH)-induced IVM of buffalo oocytes, but with time FSH partially eliminated the inhibition. Therefore, inhibition of milrinone on the nuclear maturation of buffalo oocytes was reversible, and buffalo oocytes can mature normally after the inhibition is lessened.

Evidence-Based Tracking of MDR E. coli from Bovine Endometritis and Its Elimination by Effective Novel Therapeutics.

Antibiotic-resistant bacteria have become the predominant etiology of endometritis and thus require effective treatment approaches. We used ultrasonography coupled with clinical signs and presented complaints of reproductive issues to investigate the epidemiology, phylogenetic analysis, antimicrobial resistance, and development of novel therapeutics against Escherichia coli isolated from endometritis in bovine (n = 304 from 10 commercial dairy farms). The prevalence of bovine endometritis in this study was 43.75%, while among these, 72.18% samples were positive for E. coli. Nucleotide analysis performed through BLAST and MEGAX showed 98% similarity to the nucleotide sequence of the reference E. coli strain (accession number CP067311.1). The disk diffusion assay revealed pathogen resistance to most antibiotics. Pattern of MIC order of resistance was as follows: enrofloxacin < gentamicin < co-amoxiclav < streptomycin < amoxicillin < metronidazole < oxytetracycline. Field trials revealed the highest recovery rate (in terms of clearance of endometritis and establishment of pregnancy) in case of gentamicin + enrofloxacin (100%) and gentamicin alone (100%), followed by co-amoxiclav + gentamicin (84.61%), oxytetracycline alone (78.57%), and metronidazole + enrofloxacin (33.33%). Hence, the current study reported a higher prevalence of multidrug-resistant E. coli showing considerable similarity with reference strain, and finally, the effective response of novel antibiotics to treat cases.

The Feasibility of Using Yellow Mealworms (Tenebrio molitor): Towards a Sustainable Aquafeed Industry.

The success of the aquafeed industry mainly depends on the availability of raw ingredients with high nutritional value, such as fishmeal (FM). However, the increased demand for FM elevates its prices and leads to high feed costs. Thus, there is an urgent need to find suitable alternatives for FM in fish diets to achieve sustainability in aquaculture. Currently, attention is being paid to the possibility of using insect meals as FM substitutes in aquafeed because of their relatively high nutritional quality. TM is one of those insects that can be regarded as a unique candidate because of its relatively high nutritional value. TM are rich sources of essential amino acids (methionine), lipids, and fatty acids, which vary based on the developmental stage of the worms. Although TM have an abundant amount of chitin as a fiber source and other anti-nutritional factors, numerous studies have investigated the efficacy of partial or complete substitution of FM by T. molitor in fish diets. In this context, we reviewed the current research findings on the achievable inclusion levels of T. molitor versus FM substitution in the diets of several finfish and shellfish species. We discussed the potential use of T. molitor as an FM substitute in fish diets and evaluated its effects on growth, biometric indices, and body composition. Besides, the hematological parameters, immunological responses, antioxidative efficacy, intestinal health status, and sensory criteria of fish fed T. molitor-based diets were also assessed.

Evolutionary Trajectory for the Emergence of Novel Coronavirus SARS-CoV-2.

Over the last two decades, the world experienced three outbreaks of coronaviruses with elevated morbidity rates. Currently, the global community is facing emerging virus SARS-CoV-2 belonging to Betacoronavirus, which appears to be more transmissible but less deadly than SARS-CoV. The current study aimed to track the evolutionary ancestors and different evolutionary strategies that were genetically adapted by SARS-CoV-2. Our whole-genome analysis revealed that SARS-CoV-2 was the descendant of Bat SARS/SARS-like CoVs and bats served as a natural reservoir. SARS-CoV-2 used mutations and recombination as crucial strategies in different genomic regions including the envelop, membrane, nucleocapsid, and spike glycoproteins to become a novel infectious agent. We confirmed that mutations in different genomic regions of SARS-CoV-2 have specific influence on virus reproductive adaptability, allowing for genotype adjustment and adaptations in rapidly changing environments. Moreover, for the first time we identified nine putative recombination patterns in SARS-CoV-2, which encompass spike glycoprotein, RdRp, helicase and ORF3a. Six recombination regions were spotted in the S gene and are undoubtedly important for evolutionary survival, meanwhile this permitted the virus to modify superficial antigenicity to find a way from immune reconnaissance in animals and adapt to a human host. With these combined natural selected strategies, SARS-CoV-2 emerged as a novel virus in human society.

Fatty acid biosynthesis and transcriptional regulation of Stearoyl-CoA Desaturase 1 (SCD1) in buffalo milk.

BACKGROUND: Buffalo milk is considered as a highly nutritious food owing to its higher contents of fatty acids (FA) and rich nutrient profile. Higher fat contents of buffalo milk make it suitable for processing to develop various healthy and nutritious products. Moreover, buffalo milk contains more unsaturated FAs (UFA) such as oleic and linolenic acid, which are important from the human health point of view owing to their desirable physiological effects. However, inadequate information is available about the chemical composition and mechanism of FA synthesis in buffalo milk. In this study, we hypothesized that expression of SCD1 gene could alter the biosynthesis of FA in epithelial cells of mammary gland and subsequently affect the FA contents in buffalo milk. We investigated the transcriptional and biological role of Stearoyl-CoA Desaturase 1 (SCD1) in the buffalo mammary epithelial cells (BMECs) during FA and triacylglycerol (TAG) synthesis. RESULTS: Results revealed that unsaturated fatty acid contents were much higher in concentration in buffalo milk as compared to Holstein cow. Significant increase in the expression level of FAS, ACACA, SREBP1, PPARG, GPAT, and AGPAT genes was observed in response to altered expression of SCD1 in buffalo milk. Moreover, change in SCD1 gene in BMECs also mediated the expression of genes related to FA biosynthesis subsequently leading to alter the FA composition. Overexpression of SCD1 significantly increased the expression of genes associated with FA and TAG synthesis leading to enhance FA and unsaturated FA contents in BMECs. However, down-regulation of SCD1 exhibited opposite consequences. CONCLUSION: Our study provides mechanistic insights on transcriptional regulation of SCD1 to alter FA and TAG synthesis through directly or indirectly mediating biosynthesis and metabolic pathways in BMECs. We provide preliminary findings regarding engineering of FA contents in buffalo milk through SCD1 signaling.

Comparison of Long Non-Coding RNA Expression Profiles of Cattle and Buffalo Differing in Muscle Characteristics.

Buffalo meat consist good qualitative characteristics as it contains "thined tender" which is favorable for cardavascular system. However, the regulatory mechanisms of long non-coding RNA (lncRNA), differences in meat quality are not well known. The chemical-physical parameters revealed the muscle quality of buffalo that can be equivalent of cattle, but there are significant differences in shearing force and muscle fiber structure. Then, we examined lncRNA expression profiles of buffalo and cattle skeletal muscle that provide first insights into their potential roles in buffalo myogenesis. Here, we profiled the expression of lncRNA in cattle and buffalo skeletal muscle tissues, and 16,236 lncRNA candidates were detected with 865 up-regulated lncRNAs and 1,296 down-regulated lncRNAs when comparing buffalo to cattle muscle tissue. We constructed coexpression and ceRNA networks, and found lncRNA MSTRG.48330.7, MSTRG.30030.4, and MSTRG.203788.46 could be as competitive endogenous RNA (ceRNA) containing potential binding sites for miR-1/206 and miR-133a. Tissue expression analysis showed that MSTRG.48330.7, MSTRG.30030.4, and MSTRG.203788.46 were highly and specifically expressed in muscle tissue. Present study may be used as a reference tool for starting point investigations into the roles played by several of those lncRNAs during buffalo myogenesis.

Understanding divergent domestication traits from the whole-genome sequencing of swamp- and river-buffalo populations.

Domesticated buffaloes have been integral to rice-paddy agro-ecosystems for millennia, yet relatively little is known about the buffalo genomics. Here, we sequenced and assembled reference genomes for both swamp and river buffaloes and we re-sequenced 230 individuals (132 swamp buffaloes and 98 river buffaloes) sampled from across Asia and Europe. Beyond the many actionable insights that our study revealed about the domestication, basic physiology and breeding of buffalo, we made the striking discovery that the divergent domestication traits between swamp and river buffaloes can be explained with recent selections of genes on social behavior, digestion metabolism, strengths and milk production.

Transcriptomics of Cherax quadricarinatus hepatopancreas during infection with Decapod iridescent virus 1 (DIV1).

Cherax quadricarinatus is a large-sized, highly fecund, and fast-growing species of freshwater crayfish, and has become one of the world's most intensely studied crustaceans. Decapod iridescent virus 1 (DIV1), a newly described species in the family Iridoviridae, is known to infect various crustaceans, including C. quadricarinatus, and may pose a new threat in the shrimp-farming industry. The present study performed de novo transcriptome sequencing of C. quadricarinatus hepatopancreas during DIV1 infection. A total of 114,784 transcripts and 56,418 genes were obtained; 1070 genes were upregulated and 775 genes were downregulated when compared with the uninfected samples (controls). Three pattern recognition receptor genes (fibrinogen-related protein, C-type lectin, and beta-1,3-glucan-binding protein) were upregulated during DIV1 infection. Among the top-30 upregulated unigenes, 9 unigenes were identified as vitellogenin (Vg) genes, and the top-3 upregulated unigenes were identified as involved in Vg lipid transport, lipid localization, and lipid transporter activity, which were all significantly over-representative GO terms in the GO enrichment analysis of total and upregulated differentially expressed genes (DEGs). Many genes associated with Jak-STAT signaling pathway, Endocytosis, Phagosome, MAPK signaling pathway, Apoptosis and Lysosome were positively modified after DIV1 infection. The predicted protein-protein interaction (PPI) analysis showed NF1 and TUBA, CRM1 and TUBB were involved in protein interactions. This research showed that DIV1 infection has a significant impact on the transcriptome profile of C. quadricarinatus hepatopancreas, and the results enhance our understanding of virus-host interactions. Furthermore, the high number of transcripts generated in the present study will provide information for identifying novel genes in the absence of a full C. quadricarinatus genome sequence.

A Novel Pale-Yellow Coat Color of Rabbits Generated via MC1R Mutation With CRISPR/Cas9 System.

Coat color is of great importance in animal breed characteristics; it is not only a significant productive trait but also an indispensable economic trait, especially in the rabbit industry. In the present study, the relationship between melanocortin 1 receptor (MC1R) genotypes and coat color phenotypes was observed in five rabbit breeds with popular coat colors that are present in China. These breeds comprised the Lianshan black rabbit (BR), Fujian yellow rabbit (YR), New Zealand white rabbit (WR), Gray Giant rabbit (GR), and Checkered Giant rabbit (CR), which were firstly determined, and the results showed that GR had an E allele; WR, CR, and BR had a 6-bp in-frame deletion (c.281_286del6, ED allele); and YR had a 30-bp deletion (c.304_333del30 E allele). To explore the feasibility of obtaining a novel rabbit coat color through the mutation of MC1R with the CRISPR/Cas9 system, two single-guide RNAs (sgRNAs) were designed for the MC1R gene, and the editing efficiency was confirmed by injection of rabbits' zygotes. Unlike the donor rabbits whose coat color was originally black, two novel pale-yellow-coated rabbits were generated in the founders. A total of six novel MC1R gene deletions were identified in the two founder rabbits, in which the longest deletion was more than 700 bp. The histological hematoxylin-and-eosin (H&E) staining results indicated that eumelanin amounts were absent in hair follicles of MC1R-knockout (KO) rabbits, when compared with that of donor BR. In addition, the messenger RNA (mRNA) levels of some key downstream genes in the MC1R pathway were all downregulated in MC1R-KO rabbits compared with BR and YR. These results further indicate that loss-of-function MC1R contributed to blocking the synthesis of eumelanin and created a novel pale-yellow coat color in the MC1R-KO rabbits, and gene editing technology may be a useful tool to generate novel phenotypes in rabbit breeding.