Comparison of Real-time RT-PCR cycle threshold (Ct) values with clinical features and severity of COVID-19 disease among hospitalized patients in the first and second waves of COVID-19 pandemic in Chennai, India.

Introduction: Real-time reverse transcriptase-polymerase chain reaction (rRT-PCR) of nasopharyngeal/ oropharyngeal swab has been the gold standard test for detection of SARS-CoV-2 infection The relationship between cycle threshold (Ct) values of rRT-PCR and severity of disease remain disputable and not clearly defined in COVID-19. Methodology: This is a single-centered retrospective observational study conducted at Government Corona Hospital (GCH), Guindy, Chennai. In the present study, we compared the Ct value of rRT-PCR from nasopharyngeal swab specimens with a diverse range of symptoms and disease severity among 240 individuals who were hospitalized with COVID-19, viz., mild cases (MC; n = 160), moderately severe cases (MSC; n = 46) and severe cases (SC; n = 34) in the first and second waves of COVID-19 pandemic. Results: The study included 240 hospitalized COVID-19 patients with a median age of 52 years (range 21 to 90 years). MC, MSC, and SC all had median Ct values of 25.0 (interquartile range - IQR 20.0 to 30.5), 29.5 (IQR 23.0 to 34.0), and 29.0 (IQR 24 to 37.5) for the ORF1ab gene. The Ct value differed significantly between mild vs moderate, and mild vs severe cases. The Ct value of SC group with co-morbidity of type 2 diabetes have a significant difference compared to non-diabetes group (p value <0.05). There was a significant difference in the median Ct value of ORF1ab gene among the MSC group and MC but not in the SC group in the first and second waves of the pandemic (p<0.05). Conclusion: We conclude that SARS-CoV-2 Ct values of rRT-PCR alone does not have a role in aiding severity stratification among patients with COVID-19 since the viral dynamics and Ct value may vary due to the emerging variants that occur in different waves of the pandemic.

CHSI costing study-Challenges and solutions for cost data collection in private hospitals in India.

INTRODUCTION: Ayushman Bharat Pradhan Mantri Jan Aarogya Yojana (AB PM-JAY) has enabled the Government of India to become a strategic purchaser of health care services from private providers. To generate base cost evidence for evidence-based policymaking the Costing of Health Services in India (CHSI) study was commissioned in 2018 for the price setting of health benefit packages. This paper reports the findings of a process evaluation of the cost data collection in the private hospitals. METHODS: The process evaluation of health system costing in private hospitals was an exploratory survey with mixed methods (quantitative and qualitative). We used three approaches-an online survey using a semi-structured questionnaire, in-depth interviews, and a review of monitoring data. The process of data collection was assessed in terms of time taken for different aspects, resources used, level and nature of difficulty encountered, challenges and solutions. RESULTS: The mean time taken for data collection in a private hospital was 9.31 (± 1.0) person months including time for obtaining permissions, actual data collection and entry, and addressing queries for data completeness and quality. The longest time was taken to collect data on human resources (30%), while it took the least time for collecting information on building and space (5%). On a scale of 1 (lowest) to 10 (highest) difficulty levels, the data on human resources was the most difficult to collect. This included data on salaries (8), time allocation (5.5) and leaves (5). DISCUSSION: Cost data from private hospitals is crucial for mixed health systems. Developing formal mechanisms of cost accounting data and data sharing as pre-requisites for empanelment under a national insurance scheme can significantly ease the process of cost data collection.

Cost of Surgical Care at Public Sector District Hospitals in India: Implications for Universal Health Coverage and Publicly Financed Health Insurance Schemes.

BACKGROUND: In low- and middle-income countries (LMICs), provisioning for surgical care is a public health priority. Ayushman Bharat Pradhan Mantri-Jan Aarogya Yojana (AB PM-JAY) is India's largest national insurance scheme providing free surgical and medical care. In this paper, we present the costs of surgical health benefit packages (HBPs) for secondary care in public district hospitals. METHODS: The costs were estimated using mixed (top-down and bottom-up) micro-costing methods. In phase II of the Costing of Health Services in India (CHSI) study, data were collected from a sample of 27 district hospitals from nine states of India. The district hospitals were selected using stratified random sampling based on the district's composite development score. We estimated unit costs for individual services-outpatient (OP) visit, per bed-day in inpatient (IP) and intensive care unit (ICU) stays, and surgical procedures. Together, this was used to estimate the cost of 250 AB PM-JAY HBPs. RESULTS: At the current level of utilization, the mean cost per OP consultation varied from US$4.10 to US$2.60 among different surgical specialities. The mean unit cost per IP bed-day ranged from US$13.40 to US$35.60. For the ICU, the mean unit cost per bed-day was US$74. Further, the unit cost of HBPs varied from US$564 for bone tumour excision to US$49 for lid tear repair. CONCLUSIONS: Data on the cost of delivering surgical care at the level of district hospitals is of critical value for evidence-based policymaking, price-setting for surgical care and planning to strengthen the availability of high quality and cost-effective surgical care in district hospitals.

Genomic correlates of variability in immune response to an oral cholera vaccine.

Cholera is endemic to many countries. Recent major outbreaks of cholera have prompted World Health Organization to recommend oral cholera vaccination as a public-health strategy. Variation in percentage of seroconversion upon cholera vaccination has been recorded across populations. Vaccine-induced responses are influenced by host genetic differences. We have investigated association between single-nucleotide polymorphic (SNP) loci in and around 296 immunologically relevant genes and total anti-lipopolysaccharide (LPS) antibody response to a killed whole-cell vaccine, comprising LPS from multiple strains of Vibrio cholerae. Titers derived from standard vibriocidal assays were also analyzed to gain further insights on validated SNP associations. Vaccination was administered to 1000 individuals drawn from India. Data on two independent random subsets, each comprising ∼500 vaccinees, were used for discovery of genomic associations and validation, respectively. Significant associations of four SNPs and haplotypes in three genes (MARCO, TNFAIP3 and CXCL12) with AR were discovered and validated, of which two in TNFAIP3 and CXCL12 were also significantly associated with immunity (fourfold increase in vibriocidal titers). CXCL12 is a neutrophil and lymphocyte chemoattractant that is upregulated in response to V. cholerae infection. LPS in the vaccine possibly provides signals that mimic those of the live bacterium. TNFAIP3 promotes intestinal epithelial barrier integrity and provides tight junction protein regulation; possible requirements for adequate response to the vaccine. LPS is a potent activator of innate immune responses and a ligand of MARCO. Variants in this gene have been found to be associated with LPS response, but not with high vibriocidal titer level.

Mycobacterium avium subspecies paratuberculosis diagnosis and geno-typing: genomic insights.

Effective control of paratuberculosis and investigations of potential link to Crohn's disease have been hampered by the lack of effective assays for easy and accurate diagnosis of Mycobacterium avium subspecies paratuberculosis (Map). Map is extremely fastidious and depends on iron chelator (Mycobactin). Map strains from humans and sheep are very difficult to isolate and may require years to emerge. Therefore, small numbers of Map isolates have been maintained in available collections. This situation has limited the study of biodiversity of Map. Though, much is known about environmental and host factors that contribute to paratuberculosis disease, but little is known about bacterial genetic mechanism of infection. Diagnostic and strain typing markers still demand improvements. Complete genome sequence of Map K10 strain is available in public domain for comparative genomics with other mycobacteria and clinical isolates of Map. It is anticipated that the genome sequence will help in carrying molecular diagnosis and strain typing with respect to Map forward at rapid pace. This paper reviews the current diagnostic and strain typing markers, which may be useful in typing of clinical isolates in near future.

Genetic determinants of immune-response to a polysaccharide vaccine for typhoid.

UNLABELLED: Differences in immunological response among vaccine recipients are determined both by their genetic differences and environmental factors. Knowledge of genetic determinants of immunological response to a vaccine can be used to design a vaccine that circumvents immunogenetic restrictions. The currently available vaccine for typhoid is a pure polysaccharide vaccine, immune response to which is T-cell independent. Little is known about whether genetic variation among vaccinees associates with variation in their antibody response to a polysaccharide vaccine. We conducted a study on 1,000 individuals resident in an area at high-risk for typhoid; vaccinated them with the typhoid vaccine, measured their antibody response to the vaccine, assayed >2,000 curated SNPs chosen from 283 genes that are known to participate in immune-response; and analyzed these data using a strategy to (a) minimize the statistical problems associated with testing of multiple hypotheses, and (b) internally cross-validate inferences, using a half-sample design, with little loss of statistical power. The first stage analysis, using the first half-sample, identified 54 SNPs in 43 genes to be significantly associated with immune response. In the second-stage, these inferences were cross-validated using the second half-sample. First-stage results of only 8 SNPs (out of 54) in 7 genes (out of 43) were cross-validated. We tested additional SNPs in these 7 genes, and found 8 more SNPs to be significantly associated. Haplotypes constructed with these SNPs in these 7 genes also showed significant association. These 7 genes are DEFB1, TLR1, IL1RL1, CTLA4, MAPK8, CD86 and IL17D. The overall picture that has emerged from this study is that (a) immune response to polysaccharide antigens is qualitatively different from that to protein antigens, and (b) polymorphisms in genes involved in polysaccharide recognition, signal transduction, inhibition of T-cell proliferation, pro-inflammatory signaling and eventual production of antimicrobial peptides are associated with antibody response to the polysaccharide vaccine for typhoid. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11568-010-9134-1) contains supplementary material, which is available to authorized users.

Immunology of mycobacterial infections: with special reference to Mycobacterium avium subspecies paratuberculosis.

The interplay between mycobacteria and host determines the outcome of infection. After uptake of mycobacteria by macrophages, several possible scenarios may emerge; mycobacteria may be destroyed immediately or there is establishment of persistent infection. This review is focused around mycobacteria-host interactions with reference to Mycobacterium avium subspecies paratuberculosis (MAP) infection and highlights protective mechanisms involved in order to design vaccines and other control strategies.

Genetic variation and haplotype structures of innate immunity genes in eastern India.

This study reports results of an extensive and comprehensive study of genetic diversity in 12 genes of the innate immune system in a population of eastern India. Genomic variation was assayed in 171 individuals by resequencing approximately 75kb of DNA comprising these genes in each individual. Almost half of the 548 DNA variants discovered was novel. DNA sequence comparisons with human and chimpanzee reference sequences revealed evolutionary features indicative of natural selection operating among individuals, who are residents of an area with a high load of microbial and other pathogens. Significant differences in allele and haplotype frequencies of the study population were observed with the HapMap populations. Gene and haplotype diversities were observed to be high. The genetic positioning of the study population among the HapMap populations based on data of the innate immunity genes substantially differed from what has been observed for Indian populations based on data of other genes. The reported range of variation in SNP density in the human genome is one SNP per 1.19kb (chromosome 22) to one SNP per 2.18kb (chromosome 19). The SNP density in innate immunity genes observed in this study (>3SNPskb(-1)) exceeds the highest density observed for any autosomal chromosome in the human genome. The extensive genomic variation and the distinct haplotype structure of innate immunity genes observed among individuals have possibly resulted from the impact of natural selection.

Non-chemical method of DNA recovery and characterization of Mycobacterium avium subspecies paratuberculosis using IS 900 PCR.

In the present study, two methods of DNA isolation-routine, traditional and standard DNA isolation protocol for Mycobacteria (Method 1) and a new non-chemicals and non-enzymes (physical) method (Method 2) of DNA recovery have been compared and evaluated in IS900 PCR for the specific detection of pathogen. Using the new Method 2, DNA has been recovered from few (1 - 3 colonies), extremely minute and stunted colonies. DNA, thus, isolated from these colonies (colonies PCR) and cultured for the first time from the cases of Crohn's disease in human beings, dairy cattle, raw milk and pasteurized commercial milk samples has been characterized in the present study. It is the first report from India.

HEPATOPROTECTIVE EFFECT OF A POLYHERBAL FORMULATION (AYUSH-LIV.04) AGAINST ETHANOL AND CCl(4) INDUCED LIVER DAMAGE IN RATS.

A polyherbal formulation was evaluated for its hepatoprotective activity against ethanol and CCl(4) induced liver damage in rats. The rats were divided into five groups of six animals each and serve as control, toxic, post-treated, herbal control, Liv.52 treated groups respectively. The results showed that the activities of liver marker enzymes in serum namely AST, ALT, ALP, ACP and serum bilirubin level (total) were increased in toxic group animals. But the activities of these enzymes were significantly lowered in post-treated group of rats. Thus, the results suggest antihepatotoxicity of "Ayush-Liv.04".

Utilization of acrylonitrile by bacteria isolated from petrochemical waste waters.

A bacterium, utilising acrylonitrile as a sole source of carbon and nitrogen, was isolated from Indian Petrochemical Corporation Limited (IPCL) waste waters and identified as Arthrobacter sp. This strain could also utilize acetonitrile, acetamide and acrylamide individually as a source of carbon and nitrogen. The metabolic studies with the whole cells indicated the sequential conversion of the nitrile to the respective amide and then to the respective acid and ammonia. The rate of nitrile hydrolysis was slower than the corresponding amide hydrolysis. Acrylic acid, the end product of acrylonitrile breakdown, did not support the growth when provided as a carbon source.

Alcohol exposure and undernutrition: effects on lipid metabolism and alcohol partitioning in rat brain regions in vitro.

The effects of maternal alcohol consumption and undernutrition on lipid metabolism and alcohol partitioning in brain cortical and stem slices of pups were studied under in vitro conditions. Alcohol administration along with a normal diet during gestation and lactation resulted in an increase in the synthesis of cholesterol in cortical and stem slices associated with decreased entry of alcohol. Phospholipid metabolism was not affected in the cortex, whereas the incorporation of [32P] was found to be altered in the brain stem, indicating regional differences with respect to alcohol effects. Undernutrition induced by feeding the mothers a low protein diet, on the other hand, decreased the incorporation of labelled precursors into lipids in cortex and stem. The changes in lipid metabolism observed in the high protein alcohol pups were not evident in the brain regions of undernourished pups exposed to alcohol and the partitioning of alcohol was not altered.