Correction: Identification and In-Silico study of non-synonymous functional SNPs in the human SCN9A gene.

[This corrects the article DOI: 10.1371/journal.pone.0297367.].

Identification and In-Silico study of non-synonymous functional SNPs in the human SCN9A gene.

Single nucleotide polymorphisms are the most common form of DNA alterations at the level of a single nucleotide in the genomic sequence. Genome-wide association studies (GWAS) were carried to identify potential risk genes or genomic regions by screening for SNPs associated with disease. Recent studies have shown that SCN9A comprises the NaV1.7 subunit, Na+ channels have a gene encoding of 1988 amino acids arranged into 4 domains, all with 6 transmembrane regions, and are mainly found in dorsal root ganglion (DRG) neurons and sympathetic ganglion neurons. Multiple forms of acute hypersensitivity conditions, such as primary erythermalgia, congenital analgesia, and paroxysmal pain syndrome have been linked to polymorphisms in the SCN9A gene. Under this study, we utilized a variety of computational tools to explore out nsSNPs that are potentially damaging to heath by modifying the structure or activity of the SCN9A protein. Over 14 potentially damaging and disease-causing nsSNPs (E1889D, L1802P, F1782V, D1778N, C1370Y, V1311M, Y1248H, F1237L, M936V, I929T, V877E, D743Y, C710W, D623H) were identified by a variety of algorithms, including SNPnexus, SNAP-2, PANTHER, PhD-SNP, SNP & GO, I-Mutant, and ConSurf. Homology modeling, structure validation, and protein-ligand interactions also were performed to confirm 5 notable substitutions (L1802P, F1782V, D1778N, V1311M, and M936V). Such nsSNPs may become the center of further studies into a variety of disorders brought by SCN9A dysfunction. Using in-silico strategies for assessing SCN9A genetic variations will aid in organizing large-scale investigations and developing targeted therapeutics for disorders linked to these variations.

Screening of Toxoplasmosis in Owned and Stray Dogs of District Faisalabad, Pakistan through Latex Agglutination and Indirect ELISA.

INTRODUCTION: Toxoplasma gondii (an intracellular protozoan) causes toxoplasmosis in warm-blooded animals, including humans and dogs. The present study was carried out to investigate the seroprevalence of canine toxoplasmosis in the owned and stray populations of dogs in Faisalabad District, Punjab, Pakistan. MATERIALS AND METHODS: Commercially available Latex Agglutination Test (LAT) kits were used for the screening of samples (139 stray and 150 owned), followed by confirmation through ELISA. For the statistical analyses, chi-square was used to correlate the prevalence of toxoplasmosis with various factors. RESULTS: The overall prevalence of toxoplasmosis, determined by the LAT, was 22.5% and, by ELISA, was 21.8%. A nonsignificant association of toxoplasmosis was determined among owned and stray dogs. Among owned dog breeds, Bulldogs showed 28.30% prevalence, and among stray dogs, the highest prevalence was determined in Bhakarwal dogs (39.29%). Young and female dogs showed a slightly higher prevalence of toxoplasmosis than adults and males, respectively. CONCLUSIONS: The present study determined by LAT and ELISA in owned dogs showed the same results, while a little variation was found in the stray dogs. It is concluded that both owned and stray dogs are infected with toxoplasmosis in Faisalabad District, and based on this, it is recommended that province-wide epidemiological studies be carried out to examine the prevalence of Toxoplasma and develop policies in order to control toxoplasmosis.

Perspectives from remote sensing to investigate the COVID-19 pandemic: A future-oriented approach.

As scientific technology and space science progress, remote sensing has emerged as an innovative solution to ease the challenges of the COVID-19 pandemic. To examine the research characteristics and growth trends in using remote sensing for monitoring and managing the COVID-19 research, a bibliometric analysis was conducted on the scientific documents appearing in the Scopus database. A total of 1,509 documents on this study topic were indexed between 2020 and 2022, covering 165 countries, 577 journals, 5239 institutions, and 8,616 authors. The studies related to remote sensing and COVID-19 have a significant increase of 30% with 464 articles. The United States (429 articles, 28.42% of the global output), China (295 articles, 19.54% of the global output), and the United Kingdom (174 articles, 11.53%) appeared as the top three most contributions to the literature related to remote sensing and COVID-19 research. Sustainability, Science of the Total Environment, and International Journal of Environmental Research and Public Health were the three most productive journals in this research field. The utmost predominant themes were COVID-19, remote sensing, spatial analysis, coronavirus, lockdown, and air pollution. The expansion of these topics appears to be associated with cross-sectional research on remote sensing, evidence-based tools, satellite mapping, and geographic information systems (GIS). Global pandemic risks will be monitored and managed much more effectively in the coming years with the use of remote sensing technology.

Effect of forage to concentrate ratio on growth performance and feeding behavior of Thalli lambs.

The objective of the study was to evaluate the effect of forage to concentrate ratio (F:C) on growth performance and feeding behavior of Thalli lambs. For this purpose, twenty-one male Thalli lambs with initial live body weight (26.30 ± 3.02 kg) were selected and divided into three experimental groups in a randomized complete block design. The experimental duration was 12 weeks. The experimental diets were F:C (100:0), F:C (80:20), and F:C (60:40). Alfalfa hay was used as forage source. Lambs fed F:C (60:40) and F:C (80:20) diets showed higher dry matter intake (DMI), total dry matter intake (TDMI), organic matter intake (OMI), weight gain, growth rate, better feed conversion ratio, and feed efficiency as compared to the lambs fed F:C (100:0) (P < 0.05). The results also showed that crude protein (CP), ether extract (EE), and total digestible nutrient (TDN) intake were maximum in F:C (60:40)-fed lambs (P < 0.05). However, higher crude fiber (CF), neutral detergent fiber (NDF), and acid detergent fiber (ADF) intakes were seen in the F:C (100:0)-fed lambs (P < 0.05). The results of nutrient digestibilities revealed the improved OM, DM, CP, and ADF digestibilities in lambs fed with F:C (60:40) and F:C (80:20) (P < 0.05). The digestibility of NDF and CF were not affected by the all-dietary treatments (P > 0.05). Behavior results explored that eating time, and chewing time was higher in F:C (100:0)-fed lambs (P < 0.05), while standing time, lying time, and abnormal behavior were increased in lambs fed with F:C (60:40) and F:C (80:20) (P < 0.05). Based on the results, it is concluded that the addition of concentrate at 20% and 40% improves growth performance and increases DMI, TDMI, OMI, and nutrient digestibility. However, standing time, lying time, and abnormal behavior increase by feeding of the 20% and 40% concentrate. Therefore, it is suggested that the concentrate should be supplied in restricted amounts to improve lambs' welfare and normal behavior without influencing their performance.

Structural and Functional Impact of Damaging Nonsynonymous Single Nucleotide Polymorphisms (nsSNPs) on Human VPS35 Protein Using Computational Approaches.

Parkinson's disease is the second most common progressive neurodegenerative movement disorder. Mutations in retromer complex subunit and VPS35 represent the second most common cause of late-onset familial Parkinson's disease. The mutation in VPS35 can disrupt the normal protein functions resulting in Parkinson's disease. The aim of this study was the identification of deleterious missense Single Nucleotide Polymorphisms (nsSNPs) and their structural and functional impact on the VPS35 protein. In this study, several insilico tools were used to identify deleterious and disease-associated nsSNPs. 3D structure of VPS35 protein was constructed through MODELLER 9.2, normalized using FOLDX, and evaluated through RAMPAGE and ERRAT whereas, FOLDX was used for mutagenesis. 25 ligands were obtained from literature and docked using PyRx 0.8 software. Based on the binding affinity, five ligands i.e., PG4, MSE, GOL, EDO, and CAF were further analyzed. Molecular Dynamic simulation analysis was performed using GROMACS 5.1.4, where temperature, pressure, density, RMSD, RMSF, Rg, and SASA graphs were analyzed. The results showed that the mutations Y67H, R524W, and D620N had a structural and functional impact on the VPS35 protein. The current findings will help in appropriate drug design against the disease caused by these mutations in a large population using in-vitro study.

Effects of potato peels inclusion with exogenous enzymes in broiler diet on growth performance, nutrients digestibility and carcass characteristics.

The purpose of the research was to investigate the effects of including potato peels (PP) in broilers diet with exogenous enzymes supplementation on feed intake (FI), body weight gain (BWG), nutrient digestibility and carcass parameters. For this purpose, five iso-caloric and iso-nitrogenous rations were formulated with different levels of PP (0, 5, 10, 15 and 20%). Experimental diets were supplemented with a blend of exogenous enzymes (Xylanase, Mannanase, Protease, Cellulase). A total of four hundred male broiler chicks (day old) of 38 ± 3 gram were randomly distributed into 5 experimental groups in such a way that each group had 8 replicates and each replicate had 10 birds. Results revealed that the inclusion of PP @ 5% with exogenous enzymes had similar FI, BWG and nutrients digestibility with control. However, the inclusion of P @ of 10, 15, and 20% with exogenous enzymes in the broiler diet adversely affected the BWG, feed conversion ratio and nutrient digestibility (p < 0.05). Results of carcass parameters were not affected (p > 0.05) by dietary treatments. Based on findings, it is concluded that 5% addition of PP with exogenous enzymes could be practiced in broiler diet without any adverse effect on the performance of commercial broilers.

Long Intergenic Non-Coding RNAs in the Mammary Parenchyma and Fat Pad of Pre-Weaning Heifer Calves: Identification and Functional Analysis.

Enhanced plane of nutrition at pre-weaning stage can promote the development of mammary gland especially heifer calves. Although several genes are involved in this process, long intergenic non-coding RNAs (lincRNAs) are regarded as key regulators in the regulated network and are still largely unknown. We identified and characterized 534 putative lincRNAs based on the published RNA-seq data, including heifer calves in two groups: fed enhanced milk replacer (EH, 1.13 kg/day, including 28% crude protein, 25% fat) group and fed restricted milk replacer (R, 0.45 kg/day, including 20% crude protein, 20% fat) group. Sub-samples from the mammary parenchyma (PAR) and mammary fat pad (MFP) were harvested from heifer calves. According to the information of these lincRNAs' quantitative trait loci (QTLs), the neighboring and co-expression genes were used to predict their function. By comparing EH vs R, 79 lincRNAs (61 upregulated, 18 downregulated) and 86 lincRNAs (54 upregulated, 32 downregulated) were differentially expressed in MFP and PAR, respectively. In MFP, some differentially expressed lincRNAs (DELs) are involved in lipid metabolism pathways, while, in PAR, among of DELs are involved in cell proliferation pathways. Taken together, this study explored the potential regulatory mechanism of lincRNAs in the mammary gland development of calves under different planes of nutrition.

Somatostatin and its receptors: functional regulation in the development of mice Sertoli cells.

Recently, Sertoli cells have been ascertained as the target for the regulatory peptide somatostatin (SST). Therefore, the present study investigated the expression of somatostatin receptors, their age-related alterations and homologous regulation by in vitro treatment with SRIF14 on mice Sertoli cells; furthermore, it dealt with SRIF14 action on growth progression, apoptotic activity and related gene expressions in these cells. We found that mice Sertoli cells expressed all SST1-5 receptors with differential intensities. Age-related real-time PCR of all somatostatin receptors identified abundance of SSTR2 and SSTR5 mRNA level during Sertoli cell developmental period. Furthermore, higher level of these two receptors was independent of SRIF14, as treatment with SRIF14 failed to induce both receptor expressions when compared with control. Somatostatin treatment elicited a dose-dependent decrease in forskolin stimulated cAMP production. Low (100pM and 10nM) and high dosage (1µM) groups of SRIF14 significantly promoted apoptosis, while all treatment groups led to dose dependent cessation (P<0.05) of G1 phase of cell cycle as further validated by increase in casp3, decrease in bcl2, elevation of P21 (all by western blot) and decrease in Igf1 expressions, similarly, SST treatment caused a dose dependent suppression in the mRNA level of kitl gene, which is important in the regulation of spermatogenesis. These findings suggest that somatostatin and its receptors (SSTR2 and SSTR5) are important markers in the regulation and development of Sertoli cell; furthermore, it portrays physiological inhibitory role in Sertoli cell development by inducing apoptosis and cell cycle arrest.

Sperm capacitation combined with removal of the sperm acrosome and plasma membrane enhances paternal nucleus remodelling and early development of bovine androgenetic embryos.

The androgenetic embryo is a useful model for functional analysis of the paternal genome during embryogenesis. However, few studies have focused on the factors involved in the suppressed developmental competence of such embryos or why sperm cloning-derived androgenetic embryos fail to develop beyond the morula stage in large domestic animals. To overcome this developmental failure, we tried to improve sperm decondensation, as well as to enhance embryonic development by sperm capacitation and removal of the acrosome and plasma membrane before injection of the spermatozoa. Before injection of the spermatozoa, we quantified the effects of sperm capacitation combined with sperm pretreatment on the acrosome and plasma membrane status. We also evaluated sperm decondensation potential, sperm viability and chromatin integrity. Immunostaining data showed that the sperm acrosome and plasma membrane could be more efficiently removed after capacitation. Dithiothreitol-induced sperm decondensation potential was improved with capacitation and removal of the acrosome and plasma membrane. Although most spermatozoa lost viability after pretreatment, their chromatin remained integrated. The patterns of paternal chromatin remodelling within uncleaved androgenetic embryos and the nucleus morphology of cleaved embryos indicated that capacitation combined with membrane disruption could make injected spermatozoa decondense synchronously not only with each other, but also with the developmental pace of the ooplasm. We successfully produced androgenetic blastocysts, and efficiency increased with sperm pretreatment. In conclusion, sperm decondensation and the early development of androgenetic embryos were enhanced with sperm capacitation and removal of the acrosome and plasma membrane prior to sperm injection.

Vasoactive intestinal peptide suppresses ovarian granulosa cell apoptosis in vitro by up-regulating Bcl-2 gene expression.

Vasoactive intestinal peptide (VIP) is an endogenous peptide showing a rich profile of biological activities. Within ovaries, VIP directly regulates the ovarian functions, including granulosa cells (GCs) development. In the present study, the effects of VIP on proliferation and apoptosis in goose granulosa cells were demonstrated and its underlying mechanism investigated. A strategy of RNAi-mediated "gene silencing" of Bcl-2 (RV-Bcl-2), over-expression of Bcl-2 (JLV-Bcl-2) synthesis, and exogenous VIP was used to treat goose GCs. The results showed the amounts of Bcl-2 protein were negatively correlated with apoptosis of goose GCs in all experimental groups. Compared with other control groups, apoptosis was decreased in goose GCs following treatment of 100 nM VIP, and the amount of Bcl-2 protein was increased (P < 0.05) increased. However, VIP failed to exert an effect on cell proliferation (P > 0.05). In conclusion, the exogenous VIP plays an important role in inhibiting apoptosis of goose GCs via inducing Bcl-2 gene expression.

Factors affecting yield and composition of camel milk kept under desert conditions of central Punjab, Pakistan.

This study was planned to study the herd composition, farming system, and reproductive traits and to evaluate the effect of season, stage of lactation and parity on milk production, and composition of camels kept under pastoral environment of central Punjab, Pakistan. Based on purposive sampling method, 50 herds belonging to small, medium, and large-sized herds were selected. From these herds, 1,137 she-camels were entered in this study and their composite milk samples were collected and analyzed through standard procedures to determine the milk yield and percentages of milk contents. The results showed that the male camels constituted a lesser percentage (p < 0.05: 43.08; 380/882) in the total herd composition compared to that of she-camels (56.92; 502/882). The mean daily milk yield was 8.17 ± 0.09 L and mean percentage of fat was 3.79 ± 0.13%, protein was 3.66 ± 0.07%, lactose was 5.15 ± 0.09%, ash was 0.81 ± 0.02%, acidity was 0.20 ± 0.01%, solids not fat (SNF) was 9.63 ± 0.15%, total solids was 13.42 ± 0.21, and moisture was 86.58 ± 0.43. Mean daily milk yield was significantly higher (p < 0.01) during rainy season followed by winter season, warm dry summer, and hot summer season. Milk fat and protein contents were the highest in hot dry summer, while lactose contents were higher during rainy season. The stage of lactation and parity confirmed to impinge significantly (p < 0.01) on protein, lactose, and SNF. The present study will be helpful to design measures for the eradication of reproductive constraints and for the improvement of milk yield and composition in order to make camel rearing an economical proposition.

Risk factors associated with prevalence and major bacterial causes of mastitis in dromedary camels (Camelus dromedarius) under different production systems.

A cross-sectional study was conducted in desert environment of Jhang (Pakistan) from November 2008 to October 2009 on she-camels kept under pastoralist conditions to determine the prevalence of mastitis, impact of risk factors, and isolate the dominant mastitis-causing bacteria on total of 150 lactating she-camels by using clinical examination and surf field mastitis test. From the 150 she-camels examined, 69 (46%) were positive for mastitis at animal level, 12 (8%) clinical, and 57 (38%) subclinical. Age, parity number, stage of lactation, breed, production system, hygiene of milking process, and presence of lesion on udder/teat were found significantly associated (p<0.05) with the prevalence of mastitis in she-camels. There was the lowest prevalence (33.33%; 15 of 45) of mastitis in she-camels of 5-7 years of age, while the highest (80%; 12 of 15) in the animals aged between 14 to 16 years. Stage of lactation significantly affected (p<0.05) and was found to be associated with the prevalence of mastitis being the highest (54.55%; 18 of 33) during the initial stage of lactation (0 to 1 month) followed by last 2 months (10-12 months) as 54.17% and mid-stages (1-3 and 3-10 months) of lactation as 28.57% (6 of 21) and 37.50% (9 of 24), respectively. According to breed of camels, it was noted that the prevalence of mastitis affected significantly (p<0.05) being the highest in crossbred (Desi × Mareecha) as 51.39% (37 of 72) followed in order by Mareecha and Desi as 43.14% and 37.04%, respectively. Staphylococcus (42.19%) and Streptococcus (15.63%) genera were the dominant isolates identified. Good hygiene in milking process, milking clinically infected she-camels at last, culling chronic mastitis carriers, treating clinically infected she-camels, and dry period therapy could reduce the prevalence of contagious mastitis in the study area.

Action mechanism of inhibin α-subunit on the development of Sertoli cells and first wave of spermatogenesis in mice.

Inhibin is an important marker of Sertoli cell (SC) activity in animals with impaired spermatogenesis. However, the precise relationship between inhibin and SC activity is unknown. To investigate this relationship, we partially silenced both the transcription and translation of the gene for the α-subunit of inhibin, Inha, using recombinant pshRNA vectors developed with RNAi-Ready pSIREN-RetroQ-ZsGreen Vector (Clontech Laboratories, Mountain View, Calif). We found that Inha silencing suppresses the cell-cycle regulators Cyclin D1 and Cyclin E and up-regulates the cell-cycle inhibitor P21 (as detected by Western blot analysis), thereby increasing the number of SCs in the G1 phase of the cell cycle and decreasing the amount in the S-phase of the cell cycle (as detected by flow cytometry). Inha silencing also suppressed Pdgfa, Igf1, and Kitl mRNA levels and up-regulated Tgfbrs, Inhba, Inhbb, Cyp11a1, Dhh, and Tjp1 mRNA levels (as indicated by real-time polymerase chain reaction [PCR] analysis). These findings indicate that Inha has the potential to influence the availability of the ligand inhibin and its antagonist activin in the SC in an autocrine manner and inhibit the progression of SC from G1 to S. It may also participate in the development of the blood-testis barrier, Leydig cells, and spermatogenesis through its effect on Dhh, Tjp1, Kitl, and Pdgfa. Real-time PCR and Western blot analyses of Inha, Inhba, and Inhbb mRNA and Inha levels over time show that Inha plays an important role in the formation of round spermatid during the first wave of spermatogenesis in mice.