First Isolation and Genetic Characterization of Avian Nephritis Virus 4 from Commercial Poultry in India.

Avian nephritis virus (ANV), which belongs to the family Astroviridae, is associated with different clinical manifestations (enteric and kidney disorders) in poultry. Despite being a significant pathogen of the avian industry worldwide, information regarding genetic features of these viruses in India is scarce. In this study, 386 intestinal samples collected from 37 slaughterhouses in two north Indian states (Rajasthan and Haryana) were screened for ANV with reverse transcription PCR (RT-PCR) targeting the conserved ORF1b gene, followed by nucleotide sequencing of the amplified product. RT-PCR and sequencing confirmed the presence of ANV in 32 clinical samples (8.29%), with concurrent infections of infectious bronchitis virus, chicken astrovirus, and fowl adenoviruses observed in some clinical samples (n = 4). Virus isolations were successful from four out of 12 ANV-positive clinical samples passaged via the yolk-sac route in specific-pathogen-free embryonated chicken eggs. Additionally, the near-complete genomes of two viruses were determined through sequencing. Phylogenetic analysis based on full-length capsid protein sequences classified the viruses into ANV genotype 4 (ANV4), and to the best of our knowledge this is the first report of ANV4 from India. This study revealed the presence and circulation of new strains of ANV in Indian poultry. Genetic profiling and isolation of the viruses in this study will not only aid in the development of diagnostic tools and vaccines for ANV but also offer valuable insights into its epidemiology.

Primer aislamiento y caracterización genética del virus de la nefritis aviar 4 en la avicultura comercial de la India El virus de la nefritis aviar (ANV), que pertenece a la familia Astroviridae, se asocia con diferentes manifestaciones clínicas (trastornos entéricos y renales) en la avicultura. A pesar de ser un patógeno importante de la industria avícola en todo el mundo, la información sobre las características genéticas de estos virus en la India es escasa. En este estudio, se analizaron 386 muestras intestinales recolectadas en 37 plantas de procesamiento en dos estados del norte de la India (Rajasthan y Haryana) para detectar al virus de la nefritis aviar mediante un método de RT-PCR dirigido al gene conservado ORF1b, seguido de la secuenciación de nucleótidos del producto amplificado. El método de RT-PCR y la secuenciación confirmaron la presencia del virus de la nefritis aviar en 32 muestras clínicas (8.29%), observándose infecciones concurrentes por el virus de la bronquitis infecciosa, astrovirus del pollo y adenovirus de las aves en algunas muestras clínicas (n = 4). Los aislamientos del virus tuvieron éxito en cuatro de las 12 muestras clínicas positivas para el virus de la nefritis aviar inoculadas a través de la ruta del saco vitelino en huevos de gallina embrionados libres de patógenos específicos. Además, se determinaron los genomas casi completos de dos virus mediante secuenciación. El análisis filogenético basado en secuencias completas de proteínas de la cápside clasificó los virus en el genotipo 4 del virus de la nefritis aviar (ANV4) y hasta donde se sabe, este es el primer informe del virus de la nefritis aviar 4 en la India. Este estudio reveló la presencia y circulación de nuevas cepas del virus de la nefritis aviar en la avicultura comercial de la India. El perfil genético y el aislamiento de los virus en este estudio no solo ayudarán en el desarrollo de herramientas de diagnóstico y vacunas para el virus de la nefritis aviar, sino que también ofrecerán información valiosa sobre su epidemiología.

Rapid Detection of Explicit Volatile Organic Compounds for Early Diagnosis of Lung Cancer Using MoSi2N4 Monolayer.

In this study, we investigate the adsorption  of MoSi2N4) and MoSi2N4-VN towards five potential lung cancer volatile organic compounds (VOCs). Density functional theory calculations reveal that MoSi2N4 weakly adsorb the mentioned VOCs, whereas introduction of nitrogen vacancies significantly enhances the adsorption energies ([[EQUATION]]), both in gas phase and aqueous medium. The MoSi2N4-VN monolayers exhibit a reduced bandgap and facilitate charge transfer upon VOCs adsorption, resulting in enhanced [[EQUATION]] values of -0.83, -0.76, -0.49, -0.61, and -0.50 eV for 2,3,4-trimethyl hexane, 4-methyl octane, o-toluidine, Aniline, and Ethylbenzene, respectively. Bader charge analysis and spin-polarized density of states (SPDOS) elucidate the charge redistribution and hybridization between MoSi2N4-VN and the adsorbed VOCs. The work function of MoSi2N4-VN is significantly reduced upon VOCs adsorption due to induced dipole moments, enabling smooth charge transfer and selective VOCs sensing. Notably, MoSi2N4-VN monolayers exhibit sensor responses ranging from 16.2% to 26.6% towards the VOCs, with discernible selectivity. Importantly, the recovery times of the VOCs desorption is minimal, reinforcing the suitability of MoSi2N4-VN as a rapid, and reusable biosensor platform for efficient detection of lung cancer biomarkers. Thermodynamic analysis based on Langmuir adsorption model shows improved adsorption and detection capabilities MoSi2N4-VN under diverse operating conditions of temperatures and pressures.

Antiviral and Cytoprotective Effect of Zinc (Yasad Bhasma) Based Nanoformulations Against Bovine Coronavirus.

Ayurvedic medicine utilizes metal-based preparations, known as bhasmas, to treat various health conditions. Yasad bhasma (YB), a zinc-based ayurvedic preparation, shows promise as a potential candidate for developing zinc-based nanomedicines with anti-inflammatory and antioxidant properties. In this study, we synthesized a formulation combining YB and hydroxychloroquine (HC) as a zinc ionophore (YBHC) and investigated its biocompatibility and antiviral effects against buffalo calf coronavirus (BCoV) in Vero cells. Our results demonstrated that the formulation exhibited good conformity and enhanced cell proliferation compared to untreated cells. Additionally, no cytopathic effects were observed in BCoV-infected Vero cells treated with YBHC and YB, while infected control cells exhibited cytopathic effects. YB showed cytoprotection by promoting epithelial tissue turnover. We further explored whether YB/YBHC exerted a lysosomotropic effect to produce antiviral effects on coronavirus-adapted Vero cells, but no cell internalization was observed. In addition to the synergistic antiviral effect of YB and HC, YB may play a vital role in rejuvenating affected tissues.

Milk somatic cell DNA isolation and characterization of κ-casein gene in Halari donkey milk.

Halari donkey breed is one of the indigenous breeds of India and its population is rapidly decreasing. The Jenny milk is more similar to human milk, exhibits a wide range of probiotic diversity and hypo-allergenicity, especially among infants suffering from cow and buffalo milk protein allergy. Some studies indicated low levels of κ-casein fraction of casein protein in donkey milk. The k-casein gene was not amplified from the DNA derived from the milk somatic cells of Halari donkeys. The Halari donkey milk has low somatic cells count. We report the first isolation of DNA from milk somatic cells of Halari donkeys with subsequent characterization of k-casein gene. Through our work, we showed that the milk somatic cells can be used as a non-invasive source for DNA isolation towards molecular studies. It also proved the presence of k-casein gene in Halari donkey milk by its amplification from isolated DNA.

Heterologous Seminal Plasma Reduces the Intracellular Calcium and Sperm Viability of Cryopreserved Stallion Spermatozoa.

Despite the vital role of seminal plasma (SP) in maintaining sperm function and aiding gamete interaction in many species, SP is usually removed before cryopreservation of stallion sperm to improve cryosurvival of sperm. The present study assessed if the vital sperm functional parameters of genetically superior stallions producing poor quality semen can be enhanced by the supplementation of heterologous SP from the stallion producing high quality semen. Spermatozoa from poor quality semen producing stallions were divided into three aliquots: two aliquots were supplemented with SP obtained from good quality semen producing stallions at the rate of 20% and 30%, respectively, whereas the third aliquot remained as control (0% SP) and incubated at 37°C for 30 minutes. Sperm membrane integrity, mitochondrial membrane potential (MMP), mitochondrial superoxide (mtROS) generation, and intracellular calcium status were assessed at different time intervals during incubation by flow cytometry. It was observed that the dead sperm population increased (p < 0.01) during incubation in both the 20% and 30% SP-supplemented groups. However, no significant changes were observed in MMP in both the control and treatment groups at different time intervals. Interestingly, it was found that sperm mtROS production increased (p < 0.01) during incubation in the SP-supplemented groups compared with the control group. The proportion of live spermatozoa with high intracellular calcium was reduced (p < 0.01) during incubation in the SP-incubated groups. Collectively, heterologous SP addition could not repair the damages caused by the cryopreservation and further resulted in deterioration of semen quality as observed in our study by reducing viability, increasing reactive oxygen species (ROS) production possibly due to high proportion of dead cells, or some factors (yet to be identified) that are inducive of oxidative stress in stallion spermatozoa.

Genome-Wide Single-Nucleotide Polymorphism-Based Genomic Diversity and Runs of Homozygosity for Selection Signatures in Equine Breeds.

The horse, one of the most domesticated animals, has been used for several purposes, like transportation, hunting, in sport, or for agriculture-related works. Kathiawari, Marwari, Manipuri, Zanskari, Bhutia, Spiti, and Thoroughbred are the main breeds of horses, particularly due to their agroclimatic adaptation and role in any kind of strong physical activity, and these characteristics are majorly governed by genetic factors. The genetic diversity and phylogenetic relationship of these Indian equine breeds using microsatellite markers have been reported, but further studies exploring the SNP diversity and runs of homozygosity revealing the selection signature of breeds are still warranted. In our study, the identification of genes that play a vital role in muscle development is performed through SNP detection via the whole-genome sequencing approach. A total of 96 samples, categorized under seven breeds, and 620,721 SNPs were considered to ascertain the ROH patterns amongst all the seven breeds. Over 5444 ROH islands were mined, and the maximum number of ROHs was found to be present in Zanskari, while Thoroughbred was confined to the lowest number of ROHs. Gene enrichment of these ROH islands produced 6757 functional genes, with AGPAT1, CLEC4, and CFAP20 as important gene families. However, QTL annotation revealed that the maximum QTLs were associated with Wither's height trait ontology that falls under the growth trait in all seven breeds. An Equine SNP marker database (EqSNPDb) was developed to catalogue ROHs for all these equine breeds for the flexible and easy chromosome-wise retrieval of ROH along with the genotype details of all the SNPs. Such a study can reveal breed divergence in different climatic and ecological conditions.

Synergistic Effect of Zinc-Chitosan Nanoparticles and Hydroxychloroquine to Inhibit Buffalo Coronavirus.

Zinc ions can hinder the synthesis of proteins required for accomplishing several stages of the viral life cycle. The intracellular zinc concentration can be increased by using zinc ionophores which transport zinc ions into the cells and hinder viral replication. (Hydroxy)chloroquine is an example of a zinc ionophore, but both zinc and (hydroxy)chloroquine can be toxic to the host organism. The nanocarriers may serve as camouflage to evade the adverse effects of drugs, chemicals, and nanoparticles on the host. We formulated ZnO nanoparticles with flower-like morphology (ZnONFs). It was further decorated with chitosan along with hydroxychloroquine (as a zinc ionophore) (CHCZnO NPs). We have chosen the cationic polymer chitosan since it is biocompatible, biodegradable and binds easily with the cells, and enhances the transport of drugs across cell membranes. The formulation was investigated for size, shape, surface charge, and interaction of chemicals used. We evaluated the formulations for cytotoxicity, and biocompatibility in embryonated chicks and their efficacy against bovine coronavirus (BCoV) isolated from a buffalo calf, and pneumo-enteric coronaviruses isolated from a buffalo calf with promising results in comparison to ZnONFs/hydroxychloroquine alone. Furthermore, we elucidate the mechanism underlying the lysosomotropic effect of various formulations on Vero cells infected with the buffalo coronavirus.

Identification and characterization of single nucleotide polymorphisms in DMRT3 gene in Indian horse (Equus caballus) and donkey (Equus asinus) populations.

Equines' ability in racing and riding as well as gaitedness have influenced the human civilization. Aim of this study was to identify and characterize the novel polymorphisms or SNPs in DMRT3 gene in Indian horse and donkey breeds. In this study, the DMRT3 gene was sequenced and characterized in 72 Indian horses' and 33 Indian donkeys' samples. One SNP (A > C) at 878 was found in studied horses while identical SNPs (A > C) at two different nucleotide positions i.e., 878 and 942 in DMRT3 gene (chromosome 23) were observed in studied Indian donkey breeds. Horses and donkeys both have a non-synonymous mutation (A > C) at nucleotide 878 (codon 61) that converts a Stop codon (TAG > TCG) to coding codon Serine, whereas donkeys have a synonymous mutation at nucleotide 942 (codon 82) that converts Serine (TCA > TCC) into Serine. A phylogenetic tree indicated that the DMRT3 gene was equally distributed among the equine breeds. Most of the donkey breeds have been shown high levels of genetic diversity while horse breeds and Halari donkey showed the least genetic diversity. Mutation in DMRT3 has a major impact on gaitedness in horses and is presented at a high frequency in gaited breeds and in horses breed for harness racing.

Evidence of lumpy skin disease virus infection in camels.

Countries in the Indian subcontinent are currently facing a deadly epidemic of lumpy skin disease (LSD).  LSD is primarily a disease of cattle. Buffaloes may sometimes develop mild illness, however, other domestic animals are considered resistant to LSD. We confirmed the LSDV infection in camels as evidenced by skin nodules on the body surface of the affected camels, isolation of LSD virus (LSDV) and amplification of LSDV-specific gene segments from the skin nodules (PCR), nucleotide sequencing of the viral genome and, demonstration of anti-LSDV antibodies in serum. Phylogenetic analysis based on nucleotide sequencing of ORF011, ORF012 and ORF036 revealed that the virus (LSDV/Camel/India/2022/Bikaner) is related to the historical NI-2490/Kenya/KSGP-like field strains which are predominantly circulating in the Indian subcontinent. This is the first report wherein LSDV has been to infect camels.

Evaluation of the safety, immunogenicity and efficacy of a new live-attenuated lumpy skin disease vaccine in India.

Lumpy skin disease (LSD) was reported for the first time in India in 2019 and since then, it has become endemic. Since a homologous (LSD-virus based) vaccine was not available in the country, goatpox virus (GPV)-based heterologous vaccine was authorized for mass immunization to induce protection against LSD in cattle. This study describes the evaluation of safety, immunogenicity and efficacy of a new live-attenuated LSD vaccine developed by using an Indian field strain, isolated in 2019 from cattle. The virus was attenuated by continuous passage (P = 50) in Vero cells. The vaccine (50th LSDV passage in Vero cells, named as Lumpi-ProVacInd) did not induce any local or systemic reaction upon its experimental inoculation in calves (n = 10). At day 30 post-vaccination (pv), the vaccinated animals were shown to develop antibody- and cell-mediated immune responses and exhibited complete protection upon virulent LSDV challenge. A minimum Neethling response (0.018% animals; 5 out of 26,940 animals) of the vaccine was observed in the field trials conducted in 26,940 animals. There was no significant reduction in the milk yield in lactating animals (n = 10108), besides there was no abortion or any other reproductive disorder in the pregnant animals (n = 2889). Sero-conversion was observed in 85.18% animals in the field by day 30 pv.

Milk from Halari Donkey Breed: Nutritional Analysis, Vitamins, Minerals, and Amino Acids Profiling.

This current research set out to characterize Halari donkey milk by investigating its nutritional constituents, including its proximate analysis, water activity, titratable acidity, energy, and microbiological analysis. A comprehensive profiling of vitamins, minerals, and amino acids was also carried out. It was found that the composition of Halari donkey milk was consistent with previously published donkey milk literature and was comparable to that of human milk. Halari donkey milk has low 0.86 ± 0.04% fat content, 2.03 ± 0.03% protein content, 0.51 ± 0.05% ash content, and high 5.75 ± 0.15% lactose content making it sweet and palatable. The energy content of Halari donkey milk was 40.39 ± 0.31 kcal/100 g, and the water activity ranged from 0.973 to 0.975. Titratable acidity was 0.03 ± 0.01%. Halari donkey milk can be considered acceptable and microbiologically safe, having low total plate count and yeast and mould counts. Mineral testing revealed that Halari donkey milk included significant amounts of magnesium, sodium, calcium, potassium, phosphorus, and zinc. The concentration of different vitamins and amino acids such as isoleucine and valine also contribute to the nutritional value of Halari donkey milk.

Development and Characterization of Polymeric Microsponge as a New Vehicle to Deliver Urea Topically.

BACKGROUND: Topical delivery of therapeutic agents is considered beneficial due to various advantages like ease of administration, avoidance of the first-pass effect, and improved patient compliance. Therefore, scientists around the globe are exploring this route for the delivery of drugs nowadays. OBJECTIVE: The present patent investigation aimed to prepare, optimize, and characterize the urealoaded microsponges for efficient topical delivery in vitro. METHODS: Urea-loaded ethylcellulose microsponges were prepared using quasi emulsion solvent diffusion technique and optimized using Box-Behnken design (BBD). Furthermore, they were characterized in-vitro using various techniques like scanning electron microscopy (SEM), differential scanning calorimetry (DSC), Fourier-transform infrared spectroscopy (FT-IR), and X-ray diffraction analysis (XRD). In-vitro drug release and release kinetics analysis was also performed. RESULTS: Urea-loaded microsponges were spherical and porous. Optimized urea loaded microsponges showed a minimum size (39.78 ± 1.98 µm), high entrapment (74.56 ± 2.8%), acceptable polydispersity index (PDI) (0.224 ± 0.081) and zeta potential (-21.9 ± 2.9 mV). These microsponges were capable of sustaining the release of urea for 24 h (91.21 ± 5.20%), and the mechanism of release was the combination of diffusion and erosion. CONCLUSION: The developed microsponge system could be beneficial for topical delivery of urea as it could reduce the dosing frequency of urea and increase patient compliance through its sustained release.

ROCK1/MLC2 inhibition induces decay of viral mRNA in BPXV infected cells.

Rho-associated coiled-coil containing protein kinase 1 (ROCK1) intracellular cell signaling pathway regulates cell morphology, polarity, and cytoskeletal remodeling. We observed the activation of ROCK1/myosin light chain (MLC2) signaling pathway in buffalopox virus (BPXV) infected Vero cells. ROCK1 depletion by siRNA and specific small molecule chemical inhibitors (Thiazovivin and Y27632) resulted in a reduced BPXV replication, as evidenced by reductions in viral mRNA/protein synthesis, genome copy numbers and progeny virus particles. Further, we demonstrated that ROCK1 inhibition promotes deadenylation of viral mRNA (mRNA decay), mediated via inhibiting interaction with PABP [(poly(A)-binding protein] and enhancing the expression of CCR4-NOT (a multi-protein complex that plays an important role in deadenylation of mRNA). In addition, ROCK1/MLC2 mediated cell contraction, and perinuclear accumulation of p-MLC2 was shown to positively correlate with viral mRNA/protein synthesis. Finally, it was demonstrated that the long-term sequential passage (P = 50) of BPXV in the presence of Thiazovivin does not select for any drug-resistant virus variants. In conclusion, ROCK1/MLC2 cell signaling pathway facilitates BPXV replication by preventing viral mRNA decay and that the inhibitors targeting this pathway may have novel therapeutic effects against buffalopox.

Resistance Evolution against Host-directed Antiviral Agents: Buffalopox Virus Switches to Use p38-ϒ under Long-term Selective Pressure of an Inhibitor Targeting p38-α.

Host-dependency factors have increasingly been targeted to minimize antiviral drug resistance. In this study, we have demonstrated that inhibition of p38 mitogen-activated protein kinase (a cellular protein) suppresses buffalopox virus (BPXV) protein synthesis by targeting p38-MNK1-eIF4E signaling pathway. In order to provide insights into the evolution of drug resistance, we selected resistant mutants by long-term sequential passages (P; n = 60) in the presence of p38 inhibitor (SB239063). The P60-SB239063 virus exhibited significant resistance to SB239063 as compared to the P60-Control virus. To provide mechanistic insights on the acquisition of resistance by BPXV-P60-SB239063, we generated p38-α and p38-ϒ (isoforms of p38) knockout Vero cells by CRISPR/Cas9-mediated genome editing. It was demonstrated that unlike the wild type (WT) virus which is dependent on p38-α isoform, the resistant virus (BPXV-P60-SB239063) switches over to use p38-ϒ so as to efficiently replicate in the target cells. This is a rare evidence wherein a virus was shown to bypass the dependency on a critical cellular factor under selective pressure of a drug.

Characterization of Partial Sequence of Myostatin Gene Exon 2 along with SNP detection in Indian Horse Breeds (Equus caballus).

India has well documented horse and pony breeds; however, the population is well diversified in different geographical regions. The Myostatin gene is one of the most profoundly studied genetic components for the detection of SNP's for the performance analysis in horses. In the present study, the MSTN exon 2 partial cds were amplified, sequenced and characterized in about 60 samples of eight different breeds of Indian horses. The results indicated the transition of Thymine to Cytosine (T>C) as single nucleotide polymorphisms in the partial sequence of exon 2 of the MSTN gene at two different codon positions (T12C, T13C) on chromosome 18. The haplotypes and phylogeny of the MSTN gene in the selected horse population were also analyzed. The overall and singleton haplotype are two different entities, indicating the variation among breeds is unique while the gene is equally distributed throughout the population. The phylogeny suggests that all the breeds are somehow equally distributed in their specific geographical tracts. It is the first study of MSTN gene variations in Indian horse breeds, which provides insight into predicting athletic performance as well as phylogeny. This study provides useful genetic information on Indian horses that can be used to model the racing performances of the breeds.

A Systematic Review of the Bioactive Components, Nutritional Qualities and Potential Therapeutic Applications of Donkey Milk.

In addition to providing individualized, specific, and ample nutritional compounds, donkey milk (DM) offers immunological modulation during health and disease. Recently, DM has attracted major interest in preparing infant formulas due to its similarity to human milk in terms of high protein and lactose content and low-fat concentration. The antimicrobial, anti-inflammation, antioxidant, and hypo-allergenicity properties of DM in human infants are well-documented. The purpose of this review is to summarize the knowledge of studies done in characterizing the composition of DM, including bioactive macronutrient levels influenced by the lactation status. The manufacture of DM-based food products and promising therapeutic applications in humans will also be discussed. The beneficial health effects of DM have been extensively studied as a valuable alternative source to breast milk. DM has proven to be a suitable nutrient to relieve milk-related allergies in human infants as opposed to cow's milk. Factors that influence the levels of macronutrients in DM include lactation status, processing, and manufacturing techniques. A wide variety of dairy products have been prepared using DM, such as cheese, ice cream, milk powder, novel functional fermented beverages, and milk powder for infant formulas. The bioactive macromolecules of DM exhibit antibacterial, antiviral, and antifungal effects as well as hypo-allergenicity, anti-inflammation, and antioxidant properties.

Exploring the Distinct Distribution of Archaeal Communities in Sites Contaminated with Explosives.

Most of the research on bioremediation and estimation of microbial diversity in waste contaminated sites is focused on the domain Bacteria, whereas details on the relevance of Archaea are still lacking. The present study examined the archaeal diversity and predicted metabolic pathways in two discrete sites (SITE1 and SITE2) contaminated with explosives (RDX and HMX) by amplicon-targeted sequencing of 16S rRNA genes. In total, 14 soil samples were processed, and 35,758 OTUs were observed, among which 981 OTUs were classified as Archaea, representing ~2.7% of the total microbial diversity in our samples. The majority of OTUs belonged to phyla Euryarchaeota (~49%), Crenarchaeota (~24%), and Thaumarchaeota (~23%), while the remaining (~4%) OTUs were affiliated to Candidatus Parvarchaeota, Candidatus Aenigmarchaeota, and Candidatus Diapherotrites. The comparative studies between explosives contaminated and agricultural soil samples (with no history of explosives contamination) displayed significant differences between the compositions of the archaeal communities. Further, the metabolic pathways pertaining to xenobiotic degradation were presumably more abundant in the contaminated sites. Our data provide a first comprehensive report of archaeal communities in explosives contaminated sites and their putative degradation role in such ecosystems which have been as yet unexplored.

High throughput deep proteomic analysis of seminal plasma from stallions with contrasting semen quality.

Seminal plasma proteins and pathways associated with sperm motility have not been elucidated in stallions. Therefore, in the current study, using the high throughput LC/MS-MS approach, we profiled stallion seminal plasma proteins and identified the proteins and pathways associated with sperm motility. Seminal plasma from six stallions producing semen with contrasting sperm motility (n = 3 each high-and low-motile group) was utilized for proteomic analysis. We identified a total of 1687 proteins in stallion seminal plasma, of which 1627 and 1496 proteins were expressed in high- (HM) and low- motile (LM) sperm of stallions, respectively. A total number of 1436 proteins were co-expressed in both the groups; 191 (11%) and 60 (3.5%) proteins were exclusively detected in HM and LM groups, respectively. A total of 220 proteins were upregulated (>1-fold change) and 386 proteins were downregulated in SP from LM group stallions as compared to HM group stallions, while 830 proteins were neutrally expressed in both the groups. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed dysregulation of the important proteins related to mitochondrial function, acrosome, and sperm cytoskeleton in the seminal plasma of stallions producing ejaculates with low sperm motility. High abundance of peroxiredoxins and low abundance of seminal Chaperonin Containing TCP1 Complex (CCT) complex and Annexins indicate dysregulated oxidative metabolism, which might be the underlying etiology for poor sperm motility in LM group stallions. In conclusion, the current study identified the seminal plasma proteomic alterations associated with poor sperm motility in stallions; the results indicate that poor sperm motility in stallions could be associated with altered expression of seminal plasma proteins involved in oxidative metabolism.

S-adenosylmethionine-dependent methyltransferase inhibitor DZNep blocks transcription and translation of SARS-CoV-2 genome with a low tendency to select for drug-resistant viral variants.

We report the in vitro antiviral activity of DZNep (3-Deazaneplanocin A; an inhibitor of S-adenosylmethionine-dependent methyltransferase) against SARS-CoV-2, besides demonstrating its protective efficacy against lethal infection of infectious bronchitis virus (IBV, a member of the Coronaviridae family). DZNep treatment resulted in reduced synthesis of SARS-CoV-2 RNA and proteins without affecting other steps of viral life cycle. We demonstrated that deposition of N6-methyl adenosine (m6A) in SARS-CoV-2 RNA in the infected cells recruits heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), an RNA binding protein which serves as a m6A reader. DZNep inhibited the recruitment of hnRNPA1 at m6A-modified SARS-CoV-2 RNA which eventually suppressed the synthesis of the viral genome. In addition, m6A-marked RNA and hnRNPA1 interaction was also shown to regulate early translation to replication switch of SARS-CoV-2 genome. Furthermore, abrogation of methylation by DZNep also resulted in defective synthesis of the 5' cap of viral RNA, thereby resulting in its failure to interact with eIF4E (a cap-binding protein), eventually leading to a decreased synthesis of viral proteins. Most importantly, DZNep-resistant mutants could not be observed upon long-term sequential passage of SARS-CoV-2 in cell culture. In summary, we report the novel role of methylation in the life cycle of SARS-CoV-2 and propose that targeting the methylome using DZNep could be of significant therapeutic value against SARS-CoV-2 infection.

Metal/metal oxide nanoparticles: Toxicity concerns associated with their physical state and remediation for biomedical applications.

Metal/metal oxide nanoparticles show promise for various applications, including diagnosis, treatment, theranostics, sensors, cosmetics, etc. Their altered chemical, optical, magnetic, and structural properties have differential toxicity profiles. Depending upon their physical state, these NPs can also change their properties due to alteration in pH, interaction with proteins, lipids, blood cells, and genetic material. Metallic nanomaterials (comprised of a single metal element) tend to be relatively stable and do not readily undergo dissolution. Contrarily, metal oxide and metal alloy-based nanomaterials tend to exhibit a lower degree of stability and are more susceptible to dissolution and ion release when introduced to a biological milieu, leading to reactive oxygen species production and oxidative stress to cells. Since NPs have considerable mobility in various biological tissues, the investigation related to their adverse effects is a critical issue and required to be appropriately addressed before their biomedical applications. Short and long-term toxicity assessment of metal/metal oxide nanoparticles or their nano-formulations is of paramount importance to ensure the global biome's safety; otherwise, to face a fiasco. This article provides a comprehensive introspection regarding the effects of metal/metal oxides' physical state, their surface properties, the possible mechanism of actions along with the potential future strategy for remediation of their toxic effects.