An effective method for extraction and polymerase chain reaction (PCR) amplification of DNA from formalin preserved tissue samples of snow leopard

Formalin-preserved biological samples obtained from endangered species are valuable in assessing genetic diversity. To make use of snow leopard samples preserved in formalin over a period of two to seven years, we optimized the method of extracting DNA from these samples. We used (a) phenol chloroform : isoamyl alcohol, (b) the Qiagen DNeasy Blood and Tissue Kit (Qiagen, Germany), (c) the Qiagen DNeasy Blood and Tissue Kit after treating the samples with NaOH for three days and (d) the Qiagen DNeasy Blood and Tissue Kit after treating the samples with phosphate buffered saline (PBS) for three days. The usefulness of the extracted DNA was assessed on the basis of mitochondrial (150 to 550 bp) and nuclear (95 to 229 bp) markers. There was no PCR amplification with the first two methods. The PCR amplification with the NaOH and PBS treatment had a success rate of 30 to 100% for both mitochondrial and nuclear markers. The PBS method is the best method for extraction of DNA from formalin-preserved samples of longer period (two to seven years) because of higher success rate in amplifying mitochondrial gene of ca. 550 bp (60%) than the NaOH method (28%). The overall amplification of microsatellite markers in such samples was also higher in samples treated with PBS (43 to 100%) than NaOH (0 to 100%). The PCR products obtained were confirmed through DNA sequencing to be of snow leopard origin. The optimized protocol will enable genetic studies to be conducted on tissue samples of other species that have been preserved in formalin. The protocol will be particularly useful for species that are elusive and from which it is difficult to collect fresh tissue samples.Keywords: Formalin, polymerase chain reaction (PCR), mtDNA, microsatellites, snow leopardAfrican Journal of Biotechnology Vol. 12(22), pp. 3399-340

Establishment of the mechanism of purification and levigation of green chemistry-assisted biocomposites of red ochre (Gairika): synthesis, characterization, and antibacterial, prebiotic, antioxidant, and antacid activities of the traditional Ayurvedic medicine Laghu Sutashekhara Rasa

Gairika (red ochre) has a long history of influencing human civilization. Gairika is a rich source of nutrients used for reproductive and brain health. Gairika is mentioned as an antacid drug in Indian Ayurvedic medicine under Laghu Sutashekhara Rasa (LSR). However, a detailed study on LSR has not been reported to date. In the present study, LSR was prepared, and a pharmaceutical SOP (standardization procedure) was reported to obtain batch-to-batch reproducibility. LSR was characterized using FTIR, XRD, SEM-EDX, and TGA analyses. LSR was tested in vitro for its antacid activity. Advanced instrumentation revealed that LSR formation produced symmetrical particles (5–8 µm) with kaolin, kaolinite, quartz, goethite, and hematite, along with the phytoconstituents of Goghrita (clarified cow’s butter), Shunthi, and Nagawalli, as confirmed by GC-MS/MS analysis. The FTIR study revealed the formation of a chelating complex of goethite and hematite along with their phytoconstituents. XRD analysis confirmed the presence of kaolin, kaolinite, quartz, goethite, and hematite. Using in vitro antacid experiments, LSR and Shunthi demonstrated significant antacid activity as compared to antacid drugs and standards in the market, such as CaCO3. The DPPH assay revealed IC50 values of 12.16 ± 1.23 mg/mL, which is 0.0029 of Trolox-equivalent antioxidant activity. The inhibition (18 ± 4 mm) against pathogens (S. aureus, E. coli, P. aeruginosa, and B. subtilis) and the prominent growth of gut microbiota-supported strains (S. boulardii, L. paracasei, and L. plantarum) observed on LSR formulation were indicative of LSR application as a prebiotic. Here, the mechanism of purification and levigation mentioned in the classical literature of LSR was established. Overall, purification of Gairika with cow ghee and levigation with Nagawalli may enhance the solubility, bioavailability, and shelf-life of LSR through hydration and co-crystallization mechanisms. This is the first comprehensive report on the pharmaceutical validation of LSR and its characterization. The results of the present study could contribute to the development and reliable reproduction of LSR and the utility of environmental red ochre as a medicine in combination with Shunthi (Zingiber officinale Roxb.), as prescribed under Indian Ayurvedic medicine

Genomic characterization and epidemiology of an emerging SARS-CoV-2 variant in Delhi, India

Delhi, the national capital of India, experienced multiple severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreaks in 2020 and reached population seropositivity of >50% by 2021. During April 2021, the city became overwhelmed by COVID-19 cases and fatalities, as a new variant, B.1.617.2 (Delta), replaced B.1.1.7 (Alpha). A Bayesian model explains the growth advantage of Delta through a combination of increased transmissibility and reduced sensitivity to immune responses generated against earlier variants (median estimates: 1.5-fold greater transmissibility and 20% reduction in sensitivity). Seropositivity of an employee and family cohort increased from 42% to 87.5% between March and July 2021, with 27% reinfections, as judged by increased antibody concentration after a previous decline. The likely high transmissibility and partial evasion of immunity by the Delta variant contributed to an overwhelming surge in Delhi

Tigers of Sundarbans in India: Is the Population a Separate Conservation Unit?

The Sundarbans tiger inhabits a unique mangrove habitat and are morphologically distinct from the recognized tiger subspecies in terms of skull morphometrics and body size. Thus, there is an urgent need to assess their ecological and genetic distinctiveness and determine if Sundarbans tigers should be defined and managed as separate conservation unit. We utilized nine microsatellites and 3 kb from four mitochondrial DNA (mtDNA) genes to estimate genetic variability, population structure, demographic parameters and visualize historic and contemporary connectivity among tiger populations from Sundarbans and mainland India. We also evaluated the traits that determine exchangeability or adaptive differences among tiger populations. Data from both markers suggest that Sundarbans tiger is not a separate tiger subspecies and should be regarded as Bengal tiger (P. t. tigris) subspecies. Maximum likelihood phylogenetic analyses of the mtDNA data revealed reciprocal monophyly. Genetic differentiation was found stronger for mtDNA than nuclear DNA. Microsatellite markers indicated low genetic variation in Sundarbans tigers (He= 0.58) as compared to other mainland populations, such as northern and Peninsular (Hebetween 0.67- 0.70). Molecular data supports migration between mainland and Sundarbans populations until very recent times. We attribute this reduction in gene flow to accelerated fragmentation and habitat alteration in the landscape over the past few centuries. Demographic analyses suggest that Sundarbans tigers have diverged recently from peninsular tiger population within last 2000 years. Sundarbans tigers are the most divergent group of Bengal tigers, and ecologically non-exchangeable with other tiger populations, and thus should be managed as a separate "evolutionarily significant unit" (ESU) following the adaptive evolutionary conservation (AEC) concept.Wildlife Institute of India, Dehra Dun (India)

SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion

Abstract: The B.1.617.2 (Delta) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in the state of Maharashtra in late 2020 and spread throughout India, outcompeting pre-existing lineages including B.1.617.1 (Kappa) and B.1.1.7 (Alpha)1. In vitro, B.1.617.2 is sixfold less sensitive to serum neutralizing antibodies from recovered individuals, and eightfold less sensitive to vaccine-elicited antibodies, compared with wild-type Wuhan-1 bearing D614G. Serum neutralizing titres against B.1.617.2 were lower in ChAdOx1 vaccinees than in BNT162b2 vaccinees. B.1.617.2 spike pseudotyped viruses exhibited compromised sensitivity to monoclonal antibodies to the receptor-binding domain and the amino-terminal domain. B.1.617.2 demonstrated higher replication efficiency than B.1.1.7 in both airway organoid and human airway epithelial systems, associated with B.1.617.2 spike being in a predominantly cleaved state compared with B.1.1.7 spike. The B.1.617.2 spike protein was able to mediate highly efficient syncytium formation that was less sensitive to inhibition by neutralizing antibody, compared with that of wild-type spike. We also observed that B.1.617.2 had higher replication and spike-mediated entry than B.1.617.1, potentially explaining the B.1.617.2 dominance. In an analysis of more than 130 SARS-CoV-2-infected health care workers across three centres in India during a period of mixed lineage circulation, we observed reduced ChAdOx1 vaccine effectiveness against B.1.617.2 relative to non-B.1.617.2, with the caveat of possible residual confounding. Compromised vaccine efficacy against the highly fit and immune-evasive B.1.617.2 Delta variant warrants continued infection control measures in the post-vaccination era

A cross-sectional, clinical study to evaluate mobility of teeth during pregnancy using periotest

Background and Objective: Over a century, an increased prevalence of gingival diseases associated with increasing plasma sex steroid hormone levels has been reported. These situations present unique challenges to the oral health-care professional. It is believed that hormonal fluctuations such as those associated with pregnancy, menstruation, and use of hormonal contraceptives lead to an increase in tooth mobility. However, this effect of female sex hormones on periodontal ligament and tooth-supporting alveolar bone has rarely been investigated. Hence, this study was conducted to understand the effect of hormonal changes during pregnancy on tooth mobility. Materials and Methods: Mobility of index teeth 16, 13, 21, 23, 24, 36, 33, 41, 43, and 44 was measured with periotest for fifty pregnant females at first trimester (12th week), second trimester (24th week), and third trimester (35th week). Simplified oral hygiene index, plaque index, Gingival index (GI), and probing depth were also evaluated during the three trimesters of pregnancy for each subject participating in the study. Statistical Analysis: The results of the study were subjected to statistical analysis. Data analysis was done by applying Z-test for comparing difference between two sample means. Results: A small but statistically significant influence on the periotest value was seen during the three trimesters of pregnancy. GI scores significantly increased throughout pregnancy despite no significant change in plaque levels. Conclusion: Pregnancy had a significant influence on tooth mobility. Highest value of tooth mobility was seen in the last month of pregnancy. The maximum severity of gingivitis was also seen during the third trimester of pregnancy

Pseudo scapula alata in adolescent: Unwonted diagnosis

Winging of scapula is comprehensively known for the serratus anterior palsy and its dynamic presentation. However, on the contrary, pseudo winging may occur due to nonneuromuscular cause like osteochondroma. It is a rare case of pseudo winging of scapula due to osteochondroma in a pubescent girl. This article is for knowledge to share the experience of static scapula alata due to osteochondroma. It was managed by excision and confirmed by histopathological study. Patient relieved completely from her complaints

Biofunctionalized Gold Nanoparticle-Conducting Polymer Nanocomposite Based Bioelectrode for CRP Detection

An electrochemical impedance immunosensing method for the detection and quantification of C-reactive protein (alpha CRP) in phosphate buffered saline (PBS) is demonstrated. The protein antibody, Ab-alpha CRP, has been covalently immobilized on a platform comprising of electrochemically deposited 3-mercaptopropionic acid-capped gold nanoparticles Au(MPA)-polypyrrole (PPy) nanocomposite film of controlled thickness onto an indium tin oxide-coated glass plate. The free carboxyl groups present on the nanocomposite film have been used to site-specifically immobilize the Ab-alpha CRP biomolecules through a stable acyl amino ester intermediate generated by N-(3-dimethylaminopropyl)-N'-ethyl carbodiimide hydrochloride and N-hydroxysuccinimide. The nanocomposite film was characterized by atomic force microscopy, high-resolution transmission electron microscopy, Fourier transform infrared spectroscopy, and electrochemical techniques. The bioelectrode was electrochemically analyzed using modified Randles circuit in terms of constant phase element (CPE), electron transfer resistance (R (et)), and Warburg impedance (Z (w)). The value of n, a CPE exponent used as a gauge of heterogeneity, for the Au-PPy nanocomposite film was found to be 0.56 which is indicative of a rather rough morphology and porous structure. A linear relationship between the increased a dagger R (et) values and the logarithmic value of protein antigen, Ag-alpha CRP, concentrations was found in the range of 10 ng to 10 mu g mL(-1) with a R (et) sensitivity of 46.27 a"broken vertical bar cm(2)/decade of [Ag-alpha CRP] in PBS (pH 7.4

Prevalence of oral premalignant lesions and conditions in patients with tobacco and tobacco-related habits reporting to a dental institution in Aurangabad

Aims: The purpose of this study is to determine, through a questionnaire, the prevalence of oral premalignant lesions and conditions in the population of Aurangabad city, having adverse oral habits of using tobacco and tobacco-related products in various forms, to analyze the adverse health effects of these habits with respect to the duration and frequency of consumption, and to analyze the patient motivation toward the cessation of this habit. Materials and Methods: The study was conducted in the form of a questionnaire, comprising 13 questions, to gather the sociodemographic data, information on habits of using various forms of tobacco and areca-nut (Q.1-5), symptoms associated with various habits and the treatment taken for the same (Q.6-9), and the attempt toward tobacco cessation (Q.10-13). The subjects who were diagnosed with premalignant lesions or other conditions, based on the clinical examination, underwent this questionnaire survey. Seven hundred and thirty-five subjects were included in this study during a period of 12 months. The obtained data was subjected to statistical analysis using the Epi Info™ 3.5.3. The mean percentage proportion was used for data analysis. Results: The results showed that areca nut was the most popular product among young adults. The survey data suggested that only few of the patients had tried to stop these adverse habits at some point in their lives. The most common reason for this was, advice given by the dentist after the patients were made aware of these lesions. Conclusion: The authors conclude that although the number of cases due to adverse oral habits is rising, if awareness is created by a dentist among such patients, it can bring a ray of hope in changing these malevolent trends

Electrochemical impedance spectroscopy characterization of mercaptopropionic acid capped ZnS nanocrystal based bioelectrode for the detection of the cardiac biomarker-myoglobi

3-Mercaptopropionic acid (MPA) capped ZnS nanocrystals (ZnS(MPA)) are covalently attached to a self assembled monolayer (SAM) of 3-aminopropyltriethoxysilane (APTES) on an indium-tin-oxide (ITO) coated glass plate. The protein antibody, anti-myoglobin (Ab-Mb), is covalently linked to free carboxyl groups present on ZnS(MPA) nanocrystals via carbodiimide coupling reaction to form a bioelectrode (Ab-Mb(BSA)/ZnS(MPA)/APTES/ITO-glass). This bioelectrode has been characterized using atomic force microscopy (AFM), contact angle measurements, cyclic voltammetry and electrochemical impedance spectroscopy (EIS). The optimal equivalent circuit model that matches the impedimetric responses of the bioelectrode describes three distinct regions: the electrolyte solution resistance (R-s), the double layer capacitance (C-dl) and the specific charge transfer resistance (R-et). The EIS measurements revealed that the Ret increases considerably with no significant change in C-dl after immunoreaction with protein specific antigen myoglobin. Ag-Mb, so that the prepared bioelectrode can be used for the detection of Ag-Mb. The bioelectrode exhibits an electrochemical impedance response to Ag-Mb, in a linear range from 10 ng to 1 mu g mL(-1) phosphate buffer solution (pH 7.4) with a R-et sensitivity of 117.36 Omega cm(2) per decade