In vitro impact of ethanolic extract of Bryonia laciniosa seed on Gir bull spermatozoa: a comprehensive evaluation through transcriptome profiling.

Aim/objectives: This study examines the in vitro impact of an ethanolic extract derived from Bryonia laciniosa seeds on the Gir bull (Bos indicus) spermatozoa. The objective is to thoroughly assess the effects of the seed extract on the physiological parameters of bull spermatozoa, followed by evaluating its effects on X and Y-bearing spermatozoa and its impact on gene expression through transcriptome profiling. Material method: For this study, one Gir bull was selected, and 12 ejaculates were collected at one-week time intervals. Sperm cells were isolated from each ejaculate and incubated with varying concentrations of the ethanolic extract. The physiological parameters of the spermatozoa were assessed using Computer Assisted Semen Analysis (CASA) and compared with control groups to evaluate the extract's effects on sperm quality and motility. Results and discussion: At a concentration of 18 mg/mL B. laciniosa extract, we noticed a statistically significant 16.4% increase in sperm motility (p = 0.0065). In order to understand the specific effect on X and Y-bearing spermatozoa, motile and non-motile sperm separated by glass wool column method and further evaluated for quantification of X and Y-bearing sperm in all samples by ddPCR. To understand the effect of B. laciniosa extract on spermatozoa at the molecular level, whole transcriptome profiling was carried out using Illumina MiSeq. Transcriptome profiling revealed 81 genes that were expressed differently between the group treated with the extract and the control group. The current investigation revealed an increase in the expression of TLX1, CRYGB, KLF13, and ZAR1 transcripts, which play a role in embryonic development. In addition, several genes have been identified that are involved in sperm motility, such GSK3B, LAPRS, MAPK1, CAMK2B, and AQP7. The findings exhibited the therapeutic effectiveness of B. laciniosa seeds in augmenting fertility through a synergistic blend of activities, including enhanced sperm motility and positive influence on embryogenesis.

Understanding of molecular basis of histological graded horn cancer by transcriptome profiling.

Horn cancer is most devastating and prominent cancer in Indian zebu cattle that affects socio-economic condition of small-scale farmers who depends on their cattle for farm work. Development in the field for genomics through next generation sequencing and bioinformatics advancement have helped to identify genes which have a role in horn cancer development. Histopathological examination of cancerous tissues of horn revealed myxomatous changes, well, moderate and poorly differentiated squamous cell carcinoma. Differential gene expression analysis showed 40, 11, 66 and 29 upregulated genes and 10, 14, 08 and 07 down-regulated genes in myxomatous, well, moderate and poorly differentiated squamous cell carcinoma as compared to normal. Significant differentially expressed genes are related to cell development, cell proliferation, cell-cell communication, cell signaling and angiogenesis which are linked to Akt pathway, mTOR pathway and Wnt pathway. Activity of these genes and related pathways have already been established about their role in development of cancer. Among the candidate genes; keratin family, keratin family related gene, chemokine signaling and cytokines signaling associated genes could be a prominent target for the development of stage specific prognosis marker after further detailed study at large sample population level. CSTA, PTN, SPP1 genes have upregulation in all stages of cancer and they have enrolled as biomarkers for horn cancer.

Characterizing rhizosphere microbiota of peanut (Arachis hypogaea L.) from pre-sowing to post-harvest of crop under field conditions.

The rhizosphere, a narrow zone of soil near plant roots, is a hot spot for microbial activity. Rhizosphere microbiota directly or indirectly benefit plants by supplementing nutrients, producing beneficial chemicals, or suppressing pathogens. Plants attract and modulate bacteria within the rhizosphere by releasing exudates. Plants also tend to select the rhizosphere microbiota based on their needs; a phenomenon termed as "rhizosphere effect". In this study, we characterized the rhizosphere microbiota of peanut plants across the crop development cycle from pre-sowing of seeds to post-harvest of crop under field conditions. The rhizosphere and bulk soil samples from different crop developmental stages were also compared. The composition of bulk soil microbiota resembled microbiota of pre-sowing and post-harvest soil and was markedly different from rhizosphere soil samples. Rhizosphere samples were enriched with multiple organisms mostly from the Proteobacteria, Firmicutes and Bacteroidota phyla. Differences in diversity were observed among the rhizosphere samples but not in bulk soil across different crop development stages. Pseudomonas_M indica was highly enriched during the germination of seeds. Furthermore, Plant Growth Promoting (PGP) bacteria like Bacillus were enriched during the middle stages of crop development but there was a decline in PGP organisms in the matured crop stage. We also observed a significant association of pH and Electrical Conductivity (EC) with the profiles of microbial community. Overall, this study portrayed the changes in rhizosphere microbiota of peanut during different developmental stages of crop and may help to design stage specific bio-strategies such as bio-fertilizer to improve crop yield.

Characterizing rumen microbiota and CAZyme profile of Indian dromedary camel (Camelus dromedarius) in response to different roughages.

In dromedary camels, which are pseudo-ruminants, rumen or C1 section of stomach is the main compartment involved in fiber degradation, as in true ruminants. However, as camels are adapted to the harsh and scarce grazing conditions of desert, their ruminal microbiota makes an interesting target of study. The present study was undertaken to generate the rumen microbial profile of Indian camel using 16S rRNA amplicon and shotgun metagenomics. The camels were fed three diets differing in the source of roughage. The comparative metagenomic analysis revealed greater proportions of significant differences between two fractions of rumen content followed by diet associated differences. Significant differences were also observed in the rumen microbiota collected at different time-points of the feeding trial. However, fraction related differences were more highlighted as compared to diet dependent changes in microbial profile from shotgun metagenomics data. Further, 16 genera were identified as part of the core rumen microbiome of Indian camels. Moreover, glycoside hydrolases were observed to be the most abundant among all Carbohydrate-Active enzymes and were dominated by GH2, GH3, GH13 and GH43. In all, this study describes the camel rumen microbiota under different dietary conditions with focus on taxonomic, functional, and Carbohydrate-Active enzymes profiles.

Transcriptomic analysis to affirm the regulatory role of long non-coding RNA in horn cancer of Indian zebu cattle breed Kankrej (Bos indicus).

Long non-coding RNA (lncRNA) was previously considered as a non-functional transcript, which now established as part of regulatory elements of biological events such as chromosome structure, remodeling, and regulation of gene expression. The study presented here showed the role of lncRNA through differential expression analysis on cancer-related coding genes in horn squamous cell carcinoma of Indian zebu cattle. A total of 10,360 candidate lncRNAs were identified and further analyzed for its coding potential ability using three tools (CPC, CPAT, and PLEK) that provide 8862 common lncRNAs. Pfam analysis of these common lncRNAs gave 8612 potential candidates for lncRNA differential expression analysis. Differential expression analysis showed a total of 59 significantly differentially expressed genes and 19 lncRNAs. Pearson's correlation analysis was used to identify co-expressed mRNA-lncRNAs to established relation of the regulatory role of lncRNAs in horn cancer. We established a positive relation of seven upregulated (XLOC_000016, XLOC_002198, XLOC_002851, XLOC_ 007383, XLOC_010701, XLOC_010272, and XLOC_011517) and one downregulated (XLOC_011302) lncRNAs with eleven genes that are related to keratin family protein, keratin-associated protein family, cornifelin, corneodesmosin, serpin family protein, and metallothionein that have well-established role in squamous cell carcinoma through cellular communication, cell growth, cell invasion, and cell migration. These biological events were found to be related to the MAPK pathway of cell cycle regulation indicating the role of lncRNAs in manipulating cell cycle regulation during horn squamous cell carcinomas that will be useful in identifying molecular portraits related to the development of horn cancer.

Effects of substrate binding site residue substitutions of xynA from Bacillus amyloliquefaciens on substrate specificity.

BACKGROUND: The aromatic residues of xylanase enzyme, W187, Y124, W144, Y128 and W63 of substrate binding pocket from Bacillus amyloliquefaciens were investigated for their role in substrate binding by homology modelling and sequence analysis. These residues are highly conserved and play an important role in substrate binding through steric hindrance. The substitution of these residues with alanine allows the enzyme to accommodate nonspecific substrates. RESULTS: Wild type and mutated genes were cloned and overexpressed in BL21. Optimum pH and temperature of rBAxn exhibited pH 9.0 and 50 °C respectively and it was stable up to 215 h. Along with the physical properties of rBAxn, kinetic parameters (Km 19.34 ± 0.72 mg/ml; kcat 6449.12 ± 155.37 min- 1 and kcat/Km 333.83 ± 6.78 ml min- 1 mg- 1) were also compared with engineered enzymes. Out of five mutations, W63A, Y128A and W144A lost almost 90% activity and Y124A and W187A retained almost 40-45% xylanase activity. CONCLUSIONS: The site-specific single mutation, led to alteration in substrate specificity from xylan to CMC while in case of double mutant the substrate specificity was altered from xylan to CMC, FP and avicel, indicating the role of aromatic residues on substrate binding, catalytic process and overall catalytic efficiency.