Swarna Bhasma Induces Antigen-Presenting Abilities of Macrophages and Helps Antigen Experienced CD4+ T Cells to Acquire Th1 Phenotypes Against Leishmania donovani Antigens.

In leishmaniasis, the protective immunity is largely mediated by proinflammatory cytokine producing abilities of T cells and an efficient parasite killing by phagocytic cells. Notwithstanding a substantial progress that has been made during last decades, the mechanisms or factors involved in establishing protective immunity against Leishmania are not identified. In ancient Indian literature, metallic "bhasma," particularly that of "swarna" or gold (fine gold particles), is indicated as one of the most prominent metal-based therapeutic medicine, which is known to impart protective and curative properties in various health issues. In this work, we elucidated the potential of swarna bhasma (SB) on the effector properties of phagocytes and antigen-activated CD4+ T cells in augmenting the immunogenicity of L. donovani antigens. The characterization of SB revealing its shape, size, composition, and measurement of cytotoxicity established the physiochemical potential for its utilization as an immunomodulator. The activation of macrophages with SB enhanced their capacity to produce nitric oxide and proinflammatory cytokines, which eventually resulted in reduced uptake of parasites and their proliferation in infected cells. Further, in Leishmania-infected animals, SB administration reduced the generation of IL-10, an anti-inflammatory cytokine, and enhanced pro-inflammatory cytokine generation by antigen activated CD4+ T cells with increased frequency of double (IFNγ+/TNFα+) and triple (IFNγ+TNFα+IL-2+) positive cells and abrogated disease pathogeneses at the early days of infection. Our results also suggested that cow-ghee (A2) emulsified preparation of SB, either alone or with yashtimadhu, a known natural immune modulator which enhances the SB's potential in enhancing the immunogenicity of parasitic antigens. These findings suggested a definite potential of SB in enhancing the effector functions of phagocytes and CD4+ T cells against L. donovani antigens. Therefore, more studies are needed to elucidate the mechanistic details of SB and its potential in enhancing vaccine-induced immunity.

Leishmania antigens activated CD4+ T cells expressing CD200R receptors are the prime IL-10 producing phenotype and an important determinant of visceral leishmaniasis pathogenesis.

The excessive production of IL-10, an anti-inflammatory cytokine, by Leishmania antigen-activated T cells is supposed to be a key player in the onset and progression of visceral leishmaniasis (VL). The IL-10-producing sources in VL remain unidentified and uncharacterized. In this study, we reveal that antigen-activated CD4+ T cells, i.e., CD44+CD4+ T cells expressing CD200R receptors, are the prime IL-10-producing phenotypes in Leishmania donovani infection-induced pathogenesis. These phenotypes are separate from CD25+Foxp3+CD4+ T regulatory cells, which are classical IL-10-producing phenotypes. In order to ascertain the role of CD200R and CD25 receptors in IL-10 overexpression-associated VL pathogenesis, we abrogated CD200R and CD25 receptor-mediated signaling in the infected mice. The splenic load of parasites and the size of the liver and spleen were significantly reduced in CD200-blocked mice as compared to CD25-blocked mice. Further, the CD200 blocking polarized CD4+ T cells to pro-inflammatory cytokines-producing phenotypes, as we observed a higher frequency of IFN-γ, TNF-α, and IL-12 positive cells as compared to controls including the CD25 blocking. Our findings suggest that in L. donovani infection-induced pathogenesis the expression of CD200R on antigen-activated T cells helps them to acquire IL-10-producing abilities as part of its one of the survival strategies. However, more studies would be warranted to better understand CD200R receptors role in VL pathogenesis and to develop the next generation of therapeutic and prophylactic control measures.

Adjuvantation of whole-killed Leishmania vaccine with anti-CD200 and anti-CD300a antibodies potentiates its efficacy and provides protection against wild-type parasites.

One of the major reasons behind the limited success of vaccine candidates against all forms of leishmaniasis is the inability of parasitic antigens to induce robust cell-mediated immunity and immunological memory. Here we find, for the first time, that the adjuvantation of whole-killed Leishmania vaccine (Leishvacc) with anti-CD200 and anti-CD300a antibodies enhances CD4+ T cells mediated immunity in vaccinated mice and provides protection against wild-type parasites. The antibody adjuvantation, either alone or with a TLR4 agonist monophosphoryl A (MPL-A), induced the production of pro-inflammatory cytokines viz., IFN-γ, TNF-α, and IL-2 by antigen experienced CD4+ T cells, and also enhanced their rate of conversion into their memory phenotypes against Leishvacc antigens. The antibody adjuvanted vaccine also promoted the generation of IgG2a-mediated protective humoral immunity in vaccinated mice. Further, the mice vaccinated with antibodies adjuvanted vaccine showed strong resilience against metacyclic forms of L. donovani parasites as we observed reduced clinical features such as splenomegaly, hepatomegaly, granulomatous tissues in the liver, and parasitic load in their spleen. The findings of this study demonstrate that the anti-CD200 and anti-CD300a antibodies have potential to increase the protective efficacy of the whole-killed Leishmania vaccine, and opens up a new gateway to diversify the roles of immune checkpoints in vaccine development against leishmaniasis.

"Trained Immunity" from Mycobacterium spp. exposure (BCG vaccination and environmental) may have an impact on the incidence of early childhood leukemia.

Preventive variables for childhood leukemia incidence (LI) remain unknown. Past assertions that childhood vaccinations, especially BCG, may be potentially protective have remained disputed for over five decades because of the lack of a unifying framework to explain variable outcomes in different studies. An examination of the early childhood LI for 2020 in European Region countries with supposedly similar underlying confounders but differential childhood vaccination coverage displays negative covariation with prevailing Mycobacterium spp. exposure in BCG-vaccinated children. The childhood LI in 0-4-year-old populations with >90% childhood BCG vaccination coverage is found to be strongly but negatively correlated with prevailing tuberculin immunoreactivity [r(24): -0.7868, p-value: < 0.0001]. No such correlation existed for the LI in 0-4-year-old populations without BCG vaccinations, though weak associations are hinted at by the available data for MCV2, PCV3, and DTP3 vaccinations. We hypothesize that early childhood BCG vaccination "priming" and subsequent "trained immunity" augmentation by "natural" boosting from Mycobacterium spp. exposure play a preventive and protective role in childhood LI. The non-consideration of prevailing "trained immunity" could have been a cause behind the conflicting outcomes in past studies. Exploratory studies, preferably performed in high-burden countries and controlling for the trained-immunity correlate and other potential confounders, would be warranted in order to establish a role for BCG vaccination and early-life immune training (or lack thereof) in childhood LI and help put the current controversy to rest.

BCG vaccination policy, natural boosting and pediatric brain and CNS tumor incidences.

Bacille Calmette-Guérin (BCG) vaccination supposedly imparts and augments "trained immunity" that cross-protects against multiple unrelated pathogens and enhances general immune surveillance. Gradual reductions in tuberculosis burden over the last 3-5 decades have resulted in the withdrawal of BCG vaccination mandates from developed industrialized countries while reducing to a single neonatal shot in the rest. Concurrently, a steady increase in early childhood Brain and CNS (BCNS) tumors has occurred. Though immunological causes of pediatric BCNS cancer are suspected, the identification of a causal protective variable with intervention potential has remained elusive. An examination of the countries with contrasting vaccination policies indicates significantly lower BCNS cancer incidence in 0-4-year-olds (per hundredthousand) of countries following neonatal BCG inoculations (n=146) vs. non-BCG countries (n=33) [Mean: 1.26 vs. 2.64; Median: 0.985 vs. 2.8; IQR: 0.31-2.0 vs. 2.4-3.2; P=<0.0001 (two-tailed)]. Remarkably, natural Mycobacterium spp. reexposure likelihood is negatively correlated with BCNS cancer incidence in 0-4-year-olds of all affected countries [r(154): -0.6085, P=<0.0001]. Seemingly, neonatal BCG vaccination and natural "boosting" are associated with a 15-20-fold lower BCNS cancer incidence. In this opinion article, we attempt to synthesize existing evidence implying the immunological basis of early childhood BCNS cancer incidence and briefly indicate possible causes that could have precluded objective analysis of the existing data in the past. We draw the attention of the stakeholders to consider the comprehensive evaluation of immune training as a potential protective variable through well-designed controlled clinical trials or registry-based studies as feasible for its potential applications in reducing childhood BCNS cancer incidence.

Toxin-antitoxin systems in bacterial pathogenesis.

Toxin-Antitoxin (TA) systems are abundant in prokaryotes and play an important role in various biological processes such as plasmid maintenance, phage inhibition, stress response, biofilm formation, and dormant persister cell generation. TA loci are abundant in pathogenic intracellular micro-organisms and help in their adaptation to the harsh host environment such as nutrient deprivation, oxidation, immune response, and antimicrobials. Several studies have reported the involvement of TA loci in establishing successful infection, intracellular survival, better colonization, adaptation to host stresses, and chronic infection. Overall, the TA loci play a crucial role in bacterial virulence and pathogenesis. Nonetheless, there are some controversies about the role of TA system in stress response, biofilm and persister formation. In this review, we describe the role of the TA systems in bacterial virulence. We discuss the important features of each type of TA system and the recent discoveries identifying key contributions of TA loci in bacterial pathogenesis.

Leishmania donovani induces CD300a expression to dampen effector properties of CD11c+ dendritic and antigen activated CD8+ T cells.

CD8+ T cells are an important regiment of adaptive immunity that play a decisive role in elimination of many species of Leishmania parasite from the host. In visceral leishmaniasis, caused by L. donovani, the loss of CD8+ T cells function has been found associated with augmented pathogenesis. The factors determining CD8+ T cells activation and function against Leishmania antigens are largely unknown. In this study, we investigated the role of an immune inhibitory receptor, CD300a, on the effector properties of dendritic cells and CD8+ T cells. We observed that the Leishmania regulates the effectors function of CD8+ T cells by increasing CD300a expression on CD11c+ dendritic cells. The abrogation of CD300a signaling in parasites infected animals induced CD8+ T cell abilities to produce IFN-γ, TNF-α and also helped them to acquire desired multifunctionality. The CD300a receptor blocking also enhanced the number of CD8+ T cells memory phenotypes at the early days of infection, suggesting its potential beneficial role in vaccine induced immunity. We also observed significantly enhanced levels of pro-inflammatory cytokines in the spleen of CD300a blocked infected animals with concomitant reduced spleen parasite load. Additionally, the abrogation of CD300a signals in the infected animals helped in establishing Th1 type protective humoral immunity with significantly elevated levels of IgG2a antibodies. Since CD8+ T cells are an important determinant of vaccine induced immunity against leishmaniasis, the findings corroborate the potential of CD300a in vaccine induced immunity and thus require further attention.

Higher BCG-induced trained immunity prevalence predicts protection from COVID-19: Implications for ongoing BCG trials.

Endeavors to identify potentially protective variables for COVID-19 impact on certain populations have remained a priority. Multiple attempts have been made to attribute the reduced COVID-19 impact on populations to their Bacillus-Calmette-Guérin (BCG) vaccination coverage ignoring the fact that the effect of childhood BCG vaccination wanes within 5 years while most of the COVID-19 cases and deaths have occurred in aged with comorbidities. Since the supposed protection being investigated could come from heterologous 'trained immunity' (TI) conferred by exposure to Mycobacterium spp. (i.e., environmental and BCG), it is argued that the estimates of the prevalence of TI in populations currently available as latent tuberculosis infection (LTBI) prevalence would be a better variable to evaluate such assertions. Indeed, when we analyze the European populations (24), and erstwhile East and West Germany populations completely disregarding their BCG vaccination coverage, the populations with higher TI prevalence consistently display reduced COVID-19 impact as compared to their lower TI prevalence neighbors. The TI estimates of the populations not the BCG coverage per se, negatively correlated with pandemic phase-matched COVID-19 incidences (r(24): -0.79 to -0.57; p-value < .004), mortality (r(24): -0.63 to -0.45; p-value < .03), and interim case fatality rates (i-CFR) data. To decisively arrive at dependable conclusions about the potential protective benefit gained from BCG vaccination in COVID-19, the ongoing or planned randomized controlled trials should consciously consider including measures of TI as: (a) all individuals immunized do not respond equally, (b) small study groups from higher background TI could fail to indicate any protective effect.

Potential for Further Mismanagement of Fever During COVID-19 Pandemic: Possible Causes and Impacts.

Fever remains an integral part of acute infectious diseases management, especially for those without effective therapeutics, but the widespread myths about "fevers" and the presence of confusing guidelines from different agencies, which have heightened during the coronavirus disease 2019 (COVID-19) pandemic and are open to alternate interpretation, could deny whole populations the benefits of fever. Guidelines suggesting antipyresis for 37.8-39°C fever are concerning as 39°C boosts the protective heat-shock and immune response (humoral, cell-mediated, and nutritional) whereas ≥40°C initiates/enhances the antiviral responses and restricts high-temperature adapted pathogens, e.g., severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), strains of influenza, and measles. Urgent attention is accordingly needed to address the situation because of the potential public health consequences of the existence of conflicting guidelines in the public domain. We have in this article attempted to restate the benefits of fever in disease resolution, dispel myths, and underline the need for alignment of national treatment guidelines with that of the WHO, to promote appropriate practices and reduce the morbidity and mortality from infectious diseases, such as COVID-19.

Nutritional Immunity, Zinc Sufficiency, and COVID-19 Mortality in Socially Similar European Populations.

The impact of zinc (Zn) sufficiency/supplementation on COVID-19-associated mortality and incidence (SARS-CoV-2 infections) remains unknown. During an infection, the levels of free Zn are reduced as part of "nutritional immunity" to limit the growth and replication of pathogen and the ensuing inflammatory damage. Considering its key role in immune competency and frequently recorded deficiency in large sections of different populations, Zn has been prescribed for both prophylactic and therapeutic purposes in COVID-19 without any corroborating evidence for its protective role. Multiple trials are underway evaluating the effect of Zn supplementation on COVID-19 outcome in patients getting standard of care treatment. However, the trial designs presumably lack the power to identify negative effects of Zn supplementation, especially in the vulnerable groups of elderly and patients with comorbidities (contributing 9 out of 10 deaths; up to >8,000-fold higher mortality). In this study, we have analyzed COVID-19 mortality and incidence (case) data from 23 socially similar European populations with comparable confounders (population: 522.47 million; experiencing up to >150-fold difference in death rates) and at the matching stage of the pandemic (March 12 to June 26, 2020; first wave of COVID-19 incidence and mortality). Our results suggest a positive correlation between populations' Zn-sufficiency status and COVID-19 mortality [r (23): 0.7893-0.6849, p-value < 0.0003] as well as incidence [r (23):0.8084-0.5658; p-value < 0.005]. The observed association is contrary to what would be expected if Zn sufficiency was protective in COVID-19. Thus, controlled trials or retrospective analyses of the adverse event patients' data should be undertaken to correctly guide the practice of Zn supplementation in COVID-19.

Clinicopathological Analysis and Demographic Features of Ocular Malignancies.

OBJECTIVE: This study aimed to evaluate the epidemiological and clinicopathological spectrum of ocular malignancies among patients presenting to a teaching hospital in Northern India. METHODS: A total of 246 histopathologically diagnosed patients with ocular malignancies were included in the study. Tumor type and size, primary origin and location of tumor, clinical staging, radiological findings, histopathological type, and treatment outcomes were assessed. RESULTS: Overall, males over 55 years of age were most commonly affected and the majority of cases were primary ocular or adnexal malignancies (n = 226; 91.87%). The eyelids and periocular structures (n = 92; 37.40%) were the most commonly involved site, followed by the orbit (n = 72; 29.27%), ocular surface (n = 46; 18.70%) and intraocular region (n = 36; 14.63%). The majority of the patients (n = 68; 27.64%) were managed by primary surgical excision and reconstruction. However, 46 patients (18.70%) with advanced lesions underwent neoadjuvant chemotherapy followed by surgical excision and more extensive orbital lesions were treated by exenteration followed by adjuvant chemotherapy (n=48; 19.51%), while patients with metastatic tumor were given palliative chemotherapy/external beam radiation therapy (n= 46; 18.70%). Overall, 45.12% of patients were cured completely, 15.45% showed a partial response to the treatment, 13.04% had progressive disease and 16.67% demonstrated disease recurrence. CONCLUSION: A clinicopathological analysis of ocular malignancies at a teaching hospital in Northern India indicated the preponderance of primary ocular malignancies, with eyelid sebaceous gland carcinomas being the most common pathological diagnosis. Most of our patients had advanced and extensive disease among them majority belonged to the rural background and poor socio-economic status.

Common garlic (Allium sativum L.) has potent Anti-Bacillus anthracis activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Gastrointestinal anthrax, a disease caused by Bacillus anthracis, remains an important but relatively neglected endemic disease of animals and humans in remote areas of the Indian subcontinent and some parts of Africa. Its initial symptoms include diarrhea and stomachache. In the current study, several common plants indicated for diarrhea, dysentery, stomachache or as stomachic as per traditional knowledge in the Indian subcontinent, i.e., Aegle marmelos (L.) Correa (Bael), Allium cepa L. (Onion), Allium sativum L. (Garlic), Azadirachta indica A. Juss. (Neem), Berberis asiatica Roxb. ex DC. (Daruharidra), Coriandrum sativum L. (Coriander), Curcuma longa L. (Turmeric), Cynodon dactylon (L.) Pers. (Bermuda grass), Mangifera indica L. (Mango), Morus indica L. (Black mulberry), Ocimum tenuiflorum L. (Ocimum sanctum L., Holy Basil), Ocimum gratissimum L. (Ram Tulsi), Psidium guajava L. (Guava), Zingiber officinale Roscoe (Ginger), were evaluated for their anti-Bacillus anthracis property. The usage of Azadirachta indica A. Juss. and Curcuma longa L. by Santals (India), and Allium sp. by biblical people to alleviate anthrax-like symptoms is well documented, but the usage of other plants is traditionally only indicated for different gastrointestinal disturbances/conditions. AIM OF THE STUDY: Evaluate the above listed commonly available edible plants from the Indian subcontinent that are used in the traditional medicine to treat gastrointestinal diseases including those also indicated for anthrax-like symptoms for the presence of potent anti-B. anthracis activity in a form amenable to use by the general population in the endemic areas. MATERIALS AND METHODS: Aqueous extracts made from fourteen plants indicated above were screened for their anti-B. anthracis activity using agar-well diffusion assay (AWDA) and broth microdilution methods. The Aqueous Garlic Extract (AGE) that displayed most potent anti-B. anthracis activity was assessed for its thermostability, stability under pH extremes encountered in the gastrointestinal tract, and potential antagonistic interaction with bile salts as well as the FDA-approved antibiotics used for anthrax control. The bioactive fractions from the AGE were isolated by TLC coupled bioautography followed by their characterization using GC-MS. RESULTS: Garlic (Allium sativum L.) extract was identified as the most promising candidate with bactericidal activity against B. anthracis. It consistently inhibited the growth of B. anthracis in AWDA and decreased the viable colony-forming unit counts in liquid-broth cultures by 6-logs within 6-12 h. The AGE displayed acceptable thermostability (>80% anti-B. anthracis activity retained on incubation at 50 °C for 12 h) and stability in gastric pH range (2-8). It did not antagonize the activity of FDA-approved antibiotics used for anthrax control. GC-MS analysis of the TLC separated bioactive fractions of AGE indicated the presence of previously unreported constituents such as phthalic acid derivatives, acid esters, phenyl group-containing compounds, steroids etc. CONCLUSION: The Aqueous Garlic Extract (AGE) displayed potent anti-B. anthracis activity. It was better than that displayed by Azadirachta indica A. Juss. (Neem) and Mangifera indica L., while Curcuma longa L. (Turmeric) did not show any activity under the assay conditions used. Further work should be undertaken to explore the possible application of AGE in preventing anthrax incidences in endemic areas.

CD300a Receptor Blocking Enhances Early Clearance of Leishmania donovani From Its Mammalian Host Through Modulation of Effector Functions of Phagocytic and Antigen Experienced T Cells.

The parasites of the genus Leishmania survive and proliferate in the host phagocytic cells by taking control over their microbicidal functions. The parasite also promotes differentiation of antigen-specific anti-inflammatory cytokines producing effector T cells, which eventually results in disease pathogenesis. The mechanisms that parasites employ to dominate host adaptive immunity are largely unknown. For the first time, we report that L. donovani, which causes visceral leishmaniasis in the Indian subcontinent, upregulates the expression of an immune inhibitory receptor i.e., CD300a on antigen presenting and phagocytic cells to dampen their effector functions. The blocking of CD300a signals in leishmania antigens activated macrophages and dendritic cells enhanced the production of nitric oxide, pro-inflammatory cytokines along with MHCI/II genes expression, and reduced parasitic uptake. Further, the abrogation of CD300a signals in Leishmania infected mice benefited antigen-experienced, i.e., CD4+CD44+ and CD8+CD44+ T cells to acquire more pro-inflammatory cytokines producing phenotypes and helped in the early clearance of parasites from their visceral organs. The CD300a receptor blocking also enhanced the conversion of CD4+ T effectors cells to their memory phenotypes i.e., CCR7high CD62Lhigh up to 1.6 and 1.9 fold after 14 and 21 days post-infection, respectively. These findings implicate that CD300a is an important determinant of host phagocytic cells functions and T cells differentiation against Leishmania antigens.

Distribution of Antibiotic-Resistant Enterobacteriaceae Pathogens in Potable Spring Water of Eastern Indian Himalayas: Emphasis on Virulence Gene and Antibiotic Resistance Genes in Escherichia coli.

Every year millions of people die due to fatal waterborne diseases around the world especially in developing countries like India. Sikkim, a northeastern state of India, greatly depends on natural water sources. About 80% of the population of Sikkim depends on natural spring water for domestic as well as agricultural use. Recent waterborne disease outbreaks in the state raises a concerning question on water quality. In this study, we analyzed water quality especially for the detection of Enterobacteriaceae members from four districts of the state. Isolation with selective culture media techniques and taxonomic characterization of Enterobacteriaceae bacteria with 16S rRNA gene showed the prevalence of Escherichia coli (37.50%), Escherichia fergusonii (29.41%), Klebsiella oxytoca (36.93%), Citrobacter freundii (37.92%), Citrobacter amalonaticus (43.82%), Enterobacter sp. (43.82%), Morganella morganii (43.82%), Hafnia alvei (32.42%), Hafnia paralvei (38.74%), and Shigella flexneri (30.47%) in the spring water of Sikkim. Antibiotic susceptibility test (AST) showed resistance of the isolates to common antibiotics like ampicillin, amoxicillin as well as to third generation antibiotics like ceftazidime and carbapenem. None of the isolates showed resistance to chloramphenicol. E. coli isolated from spring water of Sikkim showed presence of different virulence genes such as stx1 (81.81%), elt (86.66%), and eae (66.66%) along with resistance gene for ampicillin (CITM) (80%), quinolones (qnrB) (44.44%), tetracycline (tetO) (66.66%), and streptomycin (aadA1) (66.66%). The data indicates a high incidence rate of multiple antibiotic resistant enteric bacteria in the spring water of Sikkim. Additionally, the presence of enteric bacteria in the water samples indicates widespread fecal contamination of the spring water.

"Trained immunity" from Mycobacterium spp. exposure or BCG vaccination and COVID-19 outcomes.

Protective variables for Coronavirus Disease 2019 (COVID-19) are unknown. "Trained immunity" of the populace as a result of Bacille Calmette-Guérin (BCG) vaccination policy implementation and coverage had been suggested to be one of the factors responsible for the differential impact of COVID-19 on different countries. Several trials are underway to evaluate the potential protective role of BCG vaccination in COVID-19. However, the lack of clarity on the use of appropriate controls concerning the measures of "trained immunity" or the heterologous cell-mediated immunity conferred by BCG vaccination has been a cause of concern leading to more confusion as exemplified by a recently concluded trial in Israel that failed to find any protective correlation with regard to BCG vaccination. Whereas, when we analyze the COVID-19 epidemiological data of European countries without any regard for BCG vaccination policy but with similar age distribution, comparable confounding variables, and the stage of the pandemic, the prevalence of tuberculin immunoreactivity-a measure of cell-mediated immunity persistence as a result of Mycobacterium spp. (including BCG vaccine) exposure of the populations-is found consistently negatively correlated with COVID-19 infections and mortality. We seek to draw attention toward the inclusion of controls for underlying "trained immunity" and heterologous cell-mediated immunity prevalence that may be preexisting or resulting from the intervention (e.g., BCG vaccine) in such trials to arrive at more dependable conclusions concerning potential benefit from them.

Revisiting the role of vitamin D levels in the prevention of COVID-19 infection and mortality in European countries post infections peak.

Various studies are underway to identify protective variables for the COVID-19 pandemic. We hypothesized that if indeed the vitamin D levels would be protective in the European population, as recently proposed, the correlation would become more robust when the countries had passed the infection peak as on May 12 2020, compared to April 8 2020, when the majority had not. Comparative analysis of data from the mentioned stages indicated a significant increase in negative correlation of vitamin D levels with COVID-19 cases per million population in later stage (r(20): -0.5504; R2 = 0.3029; p value: 0.0119 vs r(20): -0.4435; R2 = 0.1967; p value: 0.0501), whereas the correlation with deaths per million population became insignificant (r(20): -0.3935; R2 = 0.1549; p value: 0.0860 vs r(20): -0.4378; R2 = 0.1917; p value: 0.0535). Considering divergence of vitamin D levels from the mean in subgroups, e.g. children, women, aged, dedicated exploratory studies with carefully chosen matched target groups is advisable.

Anthrax prevention through vaccine and post-exposure therapy.

INTRODUCTION: Vaccines and therapeutic antibodies are the most crucial components of anthrax prophylaxis (pre- and post-exposure) and treatment. The improvement in the availability and safety profile of vaccines and the therapeutic antibodies has helped immensely in reducing the worldwide burden of anthrax. AREAS COVERED: Current recommendations for anthrax prophylaxis and control, vaccines and therapeutic antibodies, the recent endeavors, particularly, made after 2010 toward making them safer and more efficacious along with our opinion on its future course. Primarily, PubMed and Europe PMC were searched to cover the recent developments in the above-indicated areas. EXPERT OPINION: Some key existing lacunae in our understanding of the working of biologicals-based anthrax-control measures, i.e., vaccines and therapeutic antibodies, should be addressed to improve their overall stability, safety profile, and efficacy. The identification of novel inhibitors targeting different key-molecules and vital-steps contributing to the overall anthrax pathophysiology could make a difference in anthrax control.

Physicochemical Parameters and Alarming Coliform Count of the Potable Water of Eastern Himalayan State Sikkim: An Indication of Severe Fecal Contamination and Immediate Health Risk.

Continuous decline in potable water sources has raised serious concerns over human health. Developing countries are the most affected in this regard due to a lack of proper hygiene maintenance. Sikkim, an Eastern Himalayan state with mountains as the predominant topological features, harbors several perennial natural springs. Spring water is the primary source of potable water for the population in four districts of the state viz. East, West, North and South. Recent outbreaks of water-borne diseases and the relative lack of scientific studies on its potential correlation with the water quality of the area have educed this study. Physicochemical parameters of springs, community reservoirs, and household water were analyzed by ICP-MS and multi probe meter. Using the membrane filtration method, the microbial quality of the water samples during different seasons was assessed, primarily evaluating the presence of fecal indicators viz. Escherichia coli, total coliform and Enterococcus. The seasonal risk category of the water sources was also determined. Most of the physicochemical parameters of the spring water were within the permissible limits of WHO standards. However, water from four districts was recorded with traces of toxic heavy metals like mercury (0.001-0.007 mg/l), lead (0.001-0.007 mg/l), and selenium (0.526-0.644 mg/l), which are above the permissible limits of WHO. All the spring water samples were categorized as Mg-HC O 3 - type and can be predicted as shallow fresh ground water based on the piper analysis. Microbial confirmatory testing indicated severe fecal contamination of water sources with high counts of total coliform (TC), Escherichia coli (EC) and Enterococcus (EN). The highest level of TC was recorded from West Sikkim (37.26 cfu/100 ml) and the lowest in North Sikkim (22.13 cfu/100 ml). The highest level of contamination of E. coli and Enterococcus was found in East Sikkim (EC = 8.7 cfu/100 ml; EN = 2.08 cfu/100 ml) followed by South Sikkim (EC = 8.4 cfu/100 ml; EN = 2.05 cfu/100 ml). There was a significant positive correlation between the contamination levels of the spring water and the community reservoir tank. As far as the seasonal variation is concerned, the rainy season showed the most contamination with coliform correlating with a high incidence of different water-borne diseases (East = 86%; West = 100%; South = 100%; North = 80%).

"Programmed Cell Death: A Process of Death for Survival" - How Far Terminology Pertinent for Cell Death in Unicellular Organisms.

Programmed cell death (PCD) is genetically regulated phenomenon of selective elimination of target cells that are either under pathological conditions or unwanted for organism's normal growth and development due to other reasons. The process although being genetically controlled is physiological in nature that renders some hallmarks like blebs in the cell membrane, lobe formation in nuclear membrane, DNA nicks resulting to DNA ladder of 200 bp, and downstream activation of caspases. Moreover, as the process refers to the death of "targeted cell", the term is exclusively suitable for multicellular organisms. Number of reports advocate similar type of cell death process in unicellular organisms. As cell death in unicellular organisms is also reflected by the signature of PCD obtained in metazoans, such cell death has been grouped under the broad category of PCD. It is pertinent to mention that by definition a unicellular organism is made of a single cell wherein it carries out all of its life processes. Using the term "Programmed Cell Death" with a preset "survival strategy of the organism" for unicellular organisms looks misnomer. Therefore, this correspondence argues and requests recommendation committee on cell death to revisit for the nomenclature of the cell death process in the unicellular organisms.