Comparative muscle transcriptome of Mali and Hampshire breeds of pigs: a preliminary study.

Muscle development is an important priority of pig breeding programs. There is a considerable variation in muscularity between the breeds, but the regulation mechanisms of genes underlying myogenesis are still unclear. Transcriptome data from two breeds of pigs with divergent muscularity (Mali and Hampshire) were integrated with histology, immunofluorescence and meat yield to identify differences in myogenesis during the early growth phase. The muscle transcriptomics analysis revealed 17,721 common, 1413 and 1115 unique transcripts to Hampshire and Mali, respectively. This study identified 908 differentially expressed genes (p < 0.05; log2FC > ±1) in the muscle samples, of which 550 were upregulated and 358 were downregulated in Hampshire pigs, indicating differences in physiological process related to muscle function and development. Expression of genes related to myoblast fusion (MYMK), skeletal muscle satellite cell proliferation (ANGPT1, CDON) and growth factors (HGF, IGF1, IGF2) were higher in Hampshire than Mali, even though transcript levels of several other myogenesis-related genes (MYF6, MYOG, MSTN) were similar. The number of fibers per fascicle and the expression of myogenic marker proteins (MYOD1, MYOG and PAX7) were more in Hampshire as compared to Mali breed of pig, supporting results of transcriptome studies. The results suggest that differences in muscularity between breeds could be related to the regulation of myoblast fusion and myogenic activities. The present study will help to identify genes that could be explored for their utility in the selection of animals with different muscularities.

Androgen receptor gene deficiency results in the reduction of steroidogenic potential in porcine luteal cells.

Luteal steroidogenesis is critical to implantation and pregnancy maintenance in mammals. The role of androgen receptors (AR) in the progesterone (P4) producing luteal cells of porcine corpus luteum (CL) remains unexplored. The aim of the present study was to establish AR gene knock out (KO) porcine luteal cell culture system model by CRISPR/Cas9 genome editing technology and to study the downstream effects of AR gene deficiency on steroidogenic potential and viability of luteal cells. For this purpose, genomic cleavage detection assay, microscopy, RT-qPCR, ELISA, annexin, MTT, and viability assay complemented by bioinformatics analysis were employed. There was significant downregulation (p < 0.05) in the relative mRNA expression of steroidogenic marker genes STAR, CYP11A1, HSD3B1 in AR KO luteal cells as compared to the control group, which was further validated by the significant (p < 0.05) decrease in the P4 production. Significant decrease (p < 0.05) in relative viability on third passage were also observed. The relative mRNA expression of hypoxia related gene HIF1A was significantly (p < 0.05) downregulated in AR KO luteal cells. Protein-protein interaction analysis mapped AR to signaling pathways associated with luteal cell functionality. These findings suggests that AR gene functionality is critical to luteal cell steroidogenesis in porcine.

Factors Responsible for Prehospital Delay in Patients with Acute Coronary Syndrome in Bangladesh.

Background: Acute coronary syndrome (ACS) remains a cause of high morbidity and mortality among adults, despite advances in treatment. Treatment modality and outcomes of ACS mainly depend on the time yielded since the onset of symptoms. Prehospital delay is the time between the onset of myocardial ischemia/infarction symptoms and arrival at the hospital, where either pharmacological or interventional revascularization is available. This delay remains unacceptably long in many countries worldwide, including Bangladesh. The current study investigates several sociodemographic characteristics as well as clinical, social, and treatment-seeking behaviors, with an aim to uncover the factors responsible for the decision time to get medical help and home-to-hospital delay. Materials and Methods: A prospective cross-sectional study was conducted between July 2019 and June 2020 in 21 district hospitals and 6 medical college hospitals where cardiac care facilities were available. The population selected for this study was patients with ACS who visited the studied hospitals during the study period. Following confirmation of ACS, a semi-structured data sheet was used to collect the patient data and was subsequently analyzed. Results: This study evaluated 678 ACS patients from 30 districts. The majority of the patients were male (81.9%), married (98.2%), rural residents (79.2), middle-aged (40-60 years of age) (55.8%), low-income holders (89.4%), and overweight (56.9%). It was found that 37.5% of the patients received their first medical care after 12 h of first symptom presentation. The study found that the patients' age, residence, education, and employment status were significant factors associated with prehospital delay. The patients with previous myocardial infarction (MI) and chest pain arrived significantly earlier at the hospital following ACS onset. Location of symptom onset, first medical contact with a private physician, distance from symptom onset location to location of first medical contact, the decision about hospitalization, ignorance of symptoms, and mode of transportation were significantly associated with prehospital delay. Conclusions: Several factors of prehospital delay of the ACS patients in Bangladesh have been described in this study. The findings of this study may help the national health management system identify the factors related to treatment delay in ACS and thus reduce ACS-related morbidity and mortality.

Prevalence and Determinants of Antibiotic Self-Administration Among Adult Antibiotic Users: A Cross-Sectional Study.

Introduction: Each year, antibiotics save hundreds of thousands of lives; nonetheless, antibiotic self-administration is a major concern all over the world. This study aimed to investigate the prevalence of antibiotic self-administration among two-month adult antibiotic users as well as the factors contributing to this prevalence. Method and Participants: This cross-sectional study was conducted among 295 Bangladeshi adults between May 22nd and June 15th, 2021, during the COVID-19 pandemic. Descriptive statistics included frequency distribution, while inferential statistics included the Pearson chi-square test. For data analysis, the statistical software STATA-16 was used. Results: In this study, the prevalence of antibiotic self-administration was 17.97%. Antibiotic self-administration was found to be significantly more prevalent among those who were unable to take antibiotic on time, incomplete doses, did not know over prescriptions may cause antibiotic resistance, and could not correctly recognize amoxicillin and azithromycin are antibiotics. Conclusion: Due to the increased rate of antibiotic self-administration among adults in Bangladesh, the responsible authority should give more attention towards the factors responsible for antibiotic self-administration and revise their current policy to ensure the safe and effective use of antibiotics.

Endemic HBV among hospital in-patients in Bangladesh, including evidence of occult infection.

Bangladesh is one of the top-ten most heavily burdened countries for viral hepatitis, with hepatitis B (HBV) infections responsible for the majority of cases. Recombinant and occult HBV infections (OBI) have been reported previously in the region. We investigated an adult fever cohort (n=201) recruited in Dhaka, to determine the prevalence of HBV and OBI. A target-enrichment deep sequencing pipeline was applied to samples with HBV DNA >3.0 log10 IU ml-1. HBV infection was present in 16/201 (8 %), among whom 3/16 (19 %) were defined as OBI (HBsAg-negative but detectable HBV DNA). Whole genome deep sequences (WGS) were obtained for four cases, identifying genotypes A, C and D. One OBI case had sufficient DNA for sequencing, revealing multiple polymorphisms in the surface gene that may contribute to the occult phenotype. We identified mutations associated with nucleos(t)ide analogue resistance in 3/4 samples sequenced, although the clinical significance in this cohort is unknown. The high prevalence of HBV in this setting illustrates the importance of opportunistic clinical screening and DNA testing of transfusion products to minimise OBI transmission. WGS can inform understanding of diverse disease phenotypes, supporting progress towards international targets for HBV elimination.

Post-calving umbilical cord tissue offcut: A potential source for the isolation of bovine mesenchymal stem cells.

BACKGROUND AND AIM: Veterinary health care is an emergent area in animal sciences and innovative therapeutic approaches happen to be imperative in the present days. In view of the importance of cattle health and production, it is necessary to take up contemporary approach of stem cell therapy in this sector also. This study aimed to standardize an explant culture method of bovine umbilical tissue offcut to isolate mesenchymal stem cells (MSCs) because considerable efforts are required for ensuring easy accessibility and availability of MSCs in bulk quantity, as well as in establishing and characterizing the cell lines. MATERIALS AND METHODS: The umbilical cord (UC) tissue matrix offcut was collected after calving. A simplified in vitro cell isolation technique was followed to collect the emerged out cells from the explants of UC. Further, we expanded these isolated cells in vitro, observed its growth kinetics, and characterized to confirm as per the criterion of bovine MSCs. RESULTS: A considerable exponential growth rate of the UC-derived cells was noticed. In addition to their confirmation as MSCs, the cells also exhibited plastic adherent property and maintained the spindle-shaped morphology throughout the in vitro culture. The cultured cells were found positive MSC-specific surface markers CD105, CD90, and CD73 and were negative for hematopoietic cell marker CD45. Cytochemical studies revealed the ability of the cells to differentiate into osteogenic, chondrogenic, and adipogenic lineages. CONCLUSION: This simplified method of isolation and culture of bovine multipotent MSCs from the UC offcut collected after calving could be extrapolated for the greater availability of the cells for prospective therapeutic applications.

Changes in expression of monocarboxylate transporters, heat shock proteins and meat quality of Large White Yorkshire and Ghungroo pigs during hot summer period.

OBJECTIVE: Present study explores the effect of hot summer period on the glycolytic rate of early post-mortem meat quality of Ghungroo and Large White Yorkshire (LWY) pig and comparative adaptability to high temperature between above breeds by shifting the expression of stress related genes like mono-carboxylate transporters (MCTs) and heat shock proteins (HSPs). METHODS: Healthy pigs of two different breeds, viz., LYW and Ghungroo (20 from each) were maintained during hot summer period (May to June) with a mean temperature of about 38°C. The pigs were slaughtered and meat samples from the longissimus dorsi (LD) muscles were analyzed for pH, glycogen and lactate content and mRNA expression. Following 24 h of chilling, LD muscle was also taken from the carcasses to evaluate protein solubility and different meat quality measurements. RESULTS: LWY exhibited significantly (p<0.01) higher plasma cortisol and lactate dehydrogenase concentration than Ghungroo indicating their higher sensitivity to high temperature. LD muscle from LWY pigs revealed lower initial and ultimate pH values and higher drip loss compared to Ghungroo, indicating a faster rate of pH fall. LD muscle of Ghungroo had significantly lower lactate content at 45 min postmortem indicating normal postmortem glycolysis and much slower glycolytic rate at early postmortem. LD muscle of LWY showed rapid postmortem glycolysis, higher drip loss and higher degrees of protein denaturation. Ghungroo exhibited slightly better water holding capacity, lower cooking loss and higher protein solubility. All HSPs (HSP27, HSP70, and HSP90) and MCTs (MCT1, MCT2, and MCT4) in the LD muscle of pigs inclined to increase more in Ghungroo than LWY when exposed to high temperature. CONCLUSION: Effect of high temperature on the variation of HSPs and MCTs may play a crucial role in thermal tolerance and adaptation to different climatic conditions, pH regulation, muscle acidification, drip loss, protein denaturation and also in postmortem meat quality development.

A comparative study on the expression profile of MCTs and HSPs in Ghungroo and Large White Yorkshire breeds of pigs during different seasons.

Thermal stress has a significant adverse effect on commercial swine production but it is not easy to measure. Animals may adapt to stress conditions by an alteration in the expression of stress-related genes such as heat shock proteins (HSPs) and monocarboxylate transporters (MCTs). The present study presents a comparative analysis of seasonally varied effects on the expression profiles of HSPs (27, 70, and 90) and MCTs (1, 2, and 4) transcripts in thigh muscle and colon tissue of Ghungroo and Large White Yorkshire (LWY) breeds of pig. By real-time polymerase chain reaction, the mRNA expression of HSP27 and HSP90 genes was found to be higher in both thigh muscle and colon tissue in Ghungroo compared to Large White Yorkshire pigs during the summer. However, the relative expression of HSP70 was significantly higher (P < 0.01) in Ghungroo compared to Large White Yorkshire pigs during both seasons in both thigh muscle and colon tissue. The expression of HSP90 was higher in Ghungroo when compared to LWY though the variation was non-significant (P > 0.05) in the colon during different seasons. However, in Ghungroo, the mRNA expression of MCT1 was found to be significantly (P < 0.05) higher in thigh muscle and colon regions during the summer compared to LWY, whereas MCT2 was expressed more in the colon in LWY compared to Ghungroo during the summer. The relative expression of mRNA of MCT4 was found to be significantly (P < 0.05) higher in thigh region in both summer and winter in Ghungroo compared with LWY. Thus, the study demonstrated that both HSPs and MCTs gene expression during thermal stress suggests the possible involvement of these genes in reducing the deleterious effect of thermal stress, thus maintaining cellular integrity and homeostasis in pigs. These genes could be used as suitable markers for the assessment of stress in pigs.

Co-infection of methicillin-resistant Staphylococcus epidermidis, methicillin-resistant Staphylococcus aureus and extended spectrum ß-lactamase producing Escherichia coli in bovine mastitis--three cases reported from India.

Emergence of antimicrobial resistance among bovine mastitis pathogens is the major cause of frequent therapeutic failure and a cause of concern for veterinary practitioners. This study describes intra-mammary infection of methicillin-resistant Staphylococcus epidermidis (MRSE), methicillin-resistant Staphylococcus aureus (MRSA) and extended spectrum ß-lactamase (ESBL) producing Escherichia coli in two Holstein Friesian crossbred cows with subclinical mastitis and one non-descript cow with clinical mastitis in two different districts of West Bengal, India. In total, three MRSE, one MRSA and three ESBL producing E. coli were isolated from these cases. Both the crossbreds were detected with MRSE (HFSE1 and HFSE2) and ESBL producing E. coli (HFEC1 and HFEC2), whereas, simultaneous infection of three pathogens viz. MRSA (NDSA1), MRSE (NDSE1) and ESBL producing E. coli (NDEC1) was found in the non-descript cow. The methicillin-resistant isolates possessed mecA gene and exhibited resistance to various antibiotics such as amikacin, tetracycline and glycopeptides. The ESBL producers were positive for blaCTX-M and blaTEM genes; in addition, HFEC1 and HFEC2 were positive for blaSHV and possessed the genes for class I integron (int1), sulphonamide resistance (sul1), quinolone resistance (qnrS) and other virulence factors (papC, iucD and ESTA1). All the ESBL producers exhibited resistance to a variety of antibiotics tested including third- and fourth-generation cephalosporins and were also intermediately resistant to carbapenems. This is the first ever report on simultaneous occurrence of MRSE, MRSA and ESBL producing E. coli in bovine mastitis indicating a major concern for dairy industry and public health as well.

Generation of parthenogenetic goat blastocysts: effects of different activation methods and culture media.

The present study was carried out to investigate the effects of different activation methods and culture media on the in vitro development of parthenogenetic goat blastocysts. Calcium (Ca2+) ionophore, ethanol or a combination of the two, used as activating reagents, and embryo development medium (EDM), modified Charles Rosenkrans (mCR2a) medium and research vitro cleave (RVCL) medium were used to evaluate the developmental competence of goat blastocysts. Quantitative expression of apoptosis, stress and developmental competence-related genes were analysed in different stages of embryos. In RVCL medium, the cleavage rate of Ca2+ ionophore-treated oocytes (79.61 ± 0.86) was significantly (P < 0.05) higher than in ethanol (74.90 ± 1.51) or in the combination of both Ca2+ ionophore and ethanol. In mCR2a or EDM, hatched blastocyst production rate of Ca2+ ionophore-treated oocytes (8.33 ± 1.44) was significantly higher than in ethanol (6.46 ± 0.11) or in the combined treatment (6.70 ± 0.24). In ethanol, the cleavage, blastocyst and hatched blastocyst production rates in RVCL medium (74.90 ± 1.51, 18.30 ± 1.52 and 8.24 ± 0.15, respectively) were significantly higher than in EDM (67.81 ± 3.21, 14.59 ± 0.27 and 5.59 ± 0.42) or mCR2a medium (65.09 ± 1.57, 15.36 ± 0.52 and 6.46 ± 0.11). The expression of BAX, Oct-4 and GlUT1 transcripts increased gradually from 2-cell stage to blastocyst-stage embryos, whereas the transcript levels of Bcl-2 and MnSOD were significantly lower in blastocysts. In addition, different activation methods and culture media had little effect on the pattern of variation and relative abundance of the above genes in different stages of parthenogenetic activated goat embryos. In conclusion, Ca2+ ionophore as the activating agent, and RVCL as the culture medium are better than other tested options for development of parthenogenetic activated goat blastocysts.

Expression and localization of locally produced growth factors regulating lymphangiogenesis during different stages of the estrous cycle in corpus luteum of buffalo (Bubalus bubalis).

Recent experiments using expression, immunolocalization, and cell culture approaches have provided leading insights into regulation of luteal angiogenesis by different growth factor systems and its role in the function of corpus luteum (CL) in buffalo. On the contrary, lymphangiogenesis and its regulation in the CL are still poorly understood. The aim of this study was to evaluate the expression and localization of lymphangiogenic factors (vascular endothelial growth factor [VEGF]-C and VEGFD), their receptor (VEGFR3), and lymphatic endothelial marker (LYVE1) in bubaline CL during different stages of the estrous cycle and to investigate functional role of VEGFC and VEGFD in luteal lymphangeogenesis. The mRNA and protein expression of VEGFC, VEGFD, and VEGFR3 was significantly greater in mid and late luteal phases, which correlated well with the expression of LYVE1. The lymphangiogenic factors were localized in luteal cells, exclusively in the cytoplasm. Immunoreactivity of VEGFC was greater during midluteal phase and that of VEGFD was greater during the mid and late luteal phases. Luteal cells were cultured in vitro and treated for different time duration (24, 48, and 72 hours) with VEGFC and VEGFD each at 50, 100, and 150 ng/mL concentration and VEGFC with VEGFD at 100 ng/mL concentration. The temporal increase in LYVE1 mRNA expression was significant (P < 0.05) in VEGFC and VEGFC with VEGFD treatment and no significant change was seen in VEGFD treatment. Thus, it seems likely that VEGFD itself has little role in lymphangiogenesis but along with VEGFC it might have a synergistic effect on VEGFR3 receptors for inducing lymphangiogenesis. In summary, the present study provided evidence that VEGFC and VEGFD, and their receptor VEGFR3, are expressed in bubaline CL and are localized exclusively in the cell cytoplasm, suggesting that these factors have a functional role in lymphangiogenesis of CL in buffalo.

Survivability of parthenogenetic embryos following in vivo transfer in naturally synchronized Capra hircus.

The present study was conducted to see the in vivo developmental potency of caprine parthenogenetic embryos generated in a modified way. The good quality caprine oocytes were matured in presence of cytochalasin B (CCB) and then activated by 7% ethanol followed by 2 mM 6-dimethyl amino purine (6-DMAP) and embryo development was recorded. Early stage parthenogenetic embryos (two to four cells) were surgically transferred in recipients (10). The pregnancy diagnosis was done by nonreturn to oestrus, ultrasonography (USG), and progesterone estimation. The levels of progesterone were above normal values (1 ng/ml) of pregnancy and fall below the level of pregnancy just before retuned to oestrus. Progesterone profile revealed that out of ten recipients (G1-G10), four goats (G1, G2, G3, and G5) returned to oestrus after 43 ± 7.29 (Mean ± SE) d of embryo transfer and six goats (G4, G6, G7, G8, G9, and G10) did not return to cycle even after 70 d of embryo transfer. In three recipients (G4, G5, and G6), the USG on day 40 revealed that there was fluid filled uterine body with solid fetus-like structure. These might be dead fetus and had started resorption. The progesterone profile also corroborated the assumption of pregnancy in these animals. Authors believe that this may be the first report on in vivo diploid parthenogenetic embryo development in caprine species.

Expression profile of HSP genes during different seasons in goats (Capra hircus).

The present study has demonstrated the expression of HSP60, HSP70, HSP90, and UBQ in peripheral blood mononuclear cells (PBMCs) during different seasons in three different age groups (Groups I, II, and III with age of 0-2, 2-5, and >5 years, respectively) of goats of tropical and temperate regions. Real-time polymerase chain reaction was applied to investigate mRNA expression of examined factors. Specificity of the desired products was documented using analysis of the melting temperature and high-resolution gel electrophoresis to verify that the transcripts are of the exact molecular size predicted. The mRNA expression of HSP60, HSP90, and UBQ was significantly higher (P < 0.05) in all age groups during peak summer season as compared with peak winter season in both tropical and temperate region goats. HSP70 mRNA expression was significantly higher (P < 0.05) during summer season as compared with winter season in tropical region goats. However, in the temperate region, in goats from all the three age groups studied, a non-significant difference of HSP70 expression between summer and winter seasons was noticed. In conclusion, results demonstrate that (1) HSP genes are expressed in caprine PBMCs and (2) higher expression of HSPs during thermal stress suggest possible involvement of them to ameliorate deleterious effect of thermal stress so as to maintain cellular integrity and homeostasis in goats.