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A cross-sectional study was done on 100 consecutive paediatric patients presenting with acute encephalitis syndrome.The clinico-laboratory features of all patients were recorded in a prestructured performa. Cerebrospinal fluid and serum samples were tested for: Japanese encephalitis (JE) virus; Chandipura virus; coxsackie virus; dengue virus; enterovirus 76; and West Nile virus. Twenty-two (22.0%) patients were confirmed JE cases and 17% had parasitic or bacteriological aetiology.The remaining 61cases (61.0%) in which no viral aetiological agent was found were grouped as non-JE cases. Peripheral vascular failure, splenomegaly and hypotonia were distinguishing clinical features found in the non-JE patients. A highmortality of 26.5% was seen in patients with confirmed or presumptive viral encephalitis (22/83). A fatal outcome was independently associated with peripheral vascular failure and pallor at the time of admission. Early recognition of these signsmay help clinicians tomanage these cases. [ABSTRACT FROM AUTHOR]

Abstract Background Fast, specific identification and surveillance of pathogens is the cornerstone of any outbreak response system, especially in the case of emerging infectious diseases and viral epidemics. This process is generally tedious and time-consuming thus making it ineffective in traditional settings. The added complexity in these situations is the non-availability of pure isolates of pathogens as they are present as mixed genomes or hologenomes. Next-generation sequencing approaches offer an attractive solution in this scenario as it provides adequate depth of sequencing at fast and affordable costs, apart from making it possible to decipher complex interactions between genomes at a scale that was not possible before. The widespread application of next-generation sequencing in this field has been limited by the non-availability of an efficient computational pipeline to systematically analyze data to delineate pathogen genomes from mixed population of genomes or hologenomes. Findings We applied next-generation sequencing on a sample containing mixed population of genomes from an epidemic with appropriate processing and enrichment. The data was analyzed using an extensive computational pipeline involving mapping to reference genome sets and de-novo assembly. In depth analysis of the data generated revealed the presence of sequences corresponding to Japanese encephalitis virus. The genome of the virus was also independently de-novo assembled. The presence of the virus was in addition, verified using standard molecular biology techniques. Conclusions Our approach can accurately identify causative pathogens from cell culture hologenome samples containing mixed population of genomes and in principle can be applied to patient hologenome samples without any background information. This methodology could be widely applied to identify and isolate pathogen genomes and understand their genomic variability during outbreaks.

Background & objectives: An outbreak of acute encephalitis syndrome was reported from Vidarbha region of Maharashtra state, India, during July 2012. Anti-IgM antibodies against Chandipura virus (CHPV) were detected in clinical samples. Sandfly collections were done to determine their role in CHPV transmission. Methods: Twenty nine pools of Sergentomyia spp. comprising 625 specimens were processed for virus isolation in Vero E6 cell line. Diagnostic RT-PCR targeting N-gene was carried out with the sample that showed cytopathic effects (CPE). The PCR product was sequenced, analysed and the sequences were deposited in Genbank database. Results: CPE in Vero E6 cell line infected with three pools was detected at 48 h post infection. However, virus could be isolated only from one pool. RT-PCR studies demonstrated 527 nucleotide product that confirmed the agent as CHPV. Sequence analysis of the new isolate showed difference in 10-12 nucleotides in comparison to earlier isolates. Interpretation & conclusions: This is perhaps the first isolation of CHPV from Sergentomyia spp. in India and virus isolation during transmission season suggests their probable role in CHPV transmission. Further studies need to be done to confirm the precise role of Sargentomyia spp. in CHPV transmission. [ABSTRACT FROM AUTHOR]

Background: India aims to eliminate rubella and congenital rubella syndrome (CRS) by 2023. We conducted serosurveys among pregnant women to monitor the trend of rubella immunity and estimate the CRS burden in India following a nationwide measles and rubella vaccination campaign., Methods: We surveyed pregnant women at 13 sentinel sites across India from Aug to Oct 2022 to estimate seroprevalence of rubella IgG antibodies. Using age-specific seroprevalence data from serosurveys conducted during 2017/2019 (prior to and during the vaccination campaign) and 2022 surveys (after the vaccination campaign), we developed force of infection (FOI) models and estimated incidence and burden of CRS., Results: In 2022, rubella seroprevalence was 85.2% (95% CI: 84.0, 86.2). Among 10 sites which participated in both rounds of serosurveys, the seroprevalence was not different between the two periods (pooled prevalence during 2017/2019: 83.5%, 95% CI: 82.1, 84.8; prevalence during 2022: 85.1%, 95% CI: 83.8, 86.3). The estimated annual incidence of CRS during 2017/2019 in India was 218.3 (95% CI: 209.7, 226.5) per 100, 000 livebirths, resulting in 47,120 (95% CI: 45,260, 48,875) cases of CRS every year. After measles-rubella (MR) vaccination campaign, the estimated incidence of CRS declined to 5.3 (95% CI: 0, 21.2) per 100,000 livebirths, resulting in 1141 (95% CI: 0, 4,569) cases of CRS during the post MR-vaccination campaign period., Conclusion: The incidence of CRS in India has substantially decreased following the nationwide MR vaccination campaign. About 15% of women in childbearing age in India lack immunity to rubella and hence susceptible to rubella infection. Since there are no routine rubella vaccination opportunities for this age group under the national immunization program, it is imperative to maintain high rates of rubella vaccination among children to prevent rubella virus exposure among women of childbearing age susceptible for rubella., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The interpretation for Zika virus serology results is challenging due to high antibody cross reactivity with other flaviviruses. This limits availability of reliable and accurate methods for serosurveillance studies to understand the disease burden. Therefore, we conducted study to harmonize anti-Zika IgG antibody detection assays with 1 st WHO International Standard (16/352) and working standard (16/320) for anti-Zika virus antibody.Additionally, evaluated NuGen TM ZIKA-IgG and NovaLisa®ZIKA virus IgG-Capture ELISA using a panel of 278 seraFurther, 106 samples positive for other-flavi viruses were taken for assessing cross-reactivity of the assay, all serums were further tested by Zika-PRNT. The results of this study indicates satisfactory performance of both the assays. Serological and neutralization assays were calibrated according to the international standards. This will help in understanding antibody dynamics in serosurveillance and vaccine studies. However the performance of the kits with possibilities of cross-reactivity will have to be verified by coupling ZIKV and DENV specific ELISA., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Nucleic acid amplification tests (NAATs) have revolutionized reliable detection of dengue virus (DENV) during acute phase of infection. The study evaluated performance of CDC DENV-1-4 real-time assay, trioplex RT-PCR and heminested conventional RT-PCR assay in the diagnosis of DENV. The three NAATs were performed on 107 consecutive samples collected from patients suspected of DENV infection during acute phase of illness. Their performance was compared against composite reference standard, consisting of DENV NS1 antigen ELISA and DENV IgM ELISA. 88/107 study samples were positive by DENV ELISA, either NS1Ag (80), IgM (3) or both (5). The overall sensitivity of CDC DENV-1-4 RT-PCR assay, trioplex RT-PCR assay and conventional multiplex RT-PCR was 68.18%, 54.55% and 38.64%, respectively in diagnosing dengue during acute phase, with an area under the curve of 0.841, 0.773 and 0.693 respectively when compared against composite reference standard. The sensitivity was 82.93%, 73.17% and 51.22%, respectively within three days of illness and 60%, 42.86% and 28.57%, respectively between 4 and 5th day of illness. All the three molecular assays had 100% specificity. Maximum concordance values of 86.9% were recorded among CDC DENV-1-4 rRT-PCR assay and trioplex assay with kappa value of 0.74, suggestive of substantial agreement. CDC DENV-1-4 rRT-PCR assay can be used as a reliable and accurate test for diagnosis of DENV during acute phase of illness., Competing Interests: Conflict of interestAuthors declare that they have no conflict of interest., (© The Author(s), under exclusive licence to Indian Virological Society 2023. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

Background: The phenotypical profile of cardiovascular malformations in patients with congenital rubella syndrome (CRS) is varied. We aimed to describe the profile of cardiac defects among CRS patients detected in the sentinel CRS surveillance in India during 2016-22., Methods: Sentinel sites enrolled infants with suspected CRS based on presence of cardiac defects, hearing impairment, eye signs, or maternal history of febrile rash illness. Suspected CRS cases underwent detailed systemic examination, including echocardiography and serological investigation for rubella. Cardiac defects were categorized as 'Simple' or 'Complex' as per the National Heart, Lung, and Blood Institute classification. We compared the distribution of cardiac defects among laboratory confirmed CRS cases and seronegative discarded cases., Findings: Of the 4578 suspected CRS cases enrolled by 14 sites, 558 (12.2%) were laboratory confirmed. 419 (75.1%) laboratory confirmed cases had structural heart defects (simple defects: n = 273, 65.2%, complex defects: n = 144, 34.4%), with ventricular septal defect (42.7%), atrial septal defect (39.4%), patent ductus arteriosus (36.5%), and tetralogy of Fallot as the commonest defects (4.5%). Laboratory confirmed CRS cases had higher odds of left to right shunt lesions (OR = 1.58, 95% CI: 1.15-2.17). This was mainly on account of a significant association of PDA with CRS (OR = 1.77, 95% CI: 1.42-2.21). Mortality was higher among CRS patients with complex heart defects (HR = 2.04, 95% CI: 1.26-3.30)., Interpretation: Three-fourths of the laboratory confirmed CRS cases had structural heart defects. CRS patients with complex cardiac defects had higher mortality. Detecting CRS infection early and providing timely intervention for cardiovascular defects is critical for the management of CRS patients., Funding: Ministry of Health and Family Welfare, Govt of India, through Gavi, the Vaccine Alliance., Competing Interests: None declared., (© 2023 The Author(s).)

The coronavirus (COVID-19) pandemic, along with its various strains, has emerged as a global health crisis that has severely affected humankind and posed a great challenge to the public health system of affected countries. The replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mainly depends on RNA-dependent RNA polymerase (RdRp), a key enzyme that is involved in RNA synthesis. In this regard, we designed, synthesized, and characterized hybrid thiouracil and coumarin conjugates (HTCAs) by ether linkage, which were found to have anti-SARS-CoV-2 properties. Our in vitro real-time quantitative reverse transcription PCR (RT-qPCR) results confirmed that compounds such as 5d , 5e , 5f , and 5i inhibited the replication of SARS-CoV-2 with EC 50 values of 14.3 ± 0.14, 6.59 ± 0.28, 86.3 ± 1.45, and 124 ± 2.38 μM, respectively. Also, compound 5d displayed significant antiviral activity against human coronavirus 229E (HCoV-229E). In addition, some of the HTCAs reduced the replication of SARS-CoV-2 variants such as D614G and B.617.2. In parallel, HTCAs in uninfected Vero CCL-81 cells indicated that no cytotoxicity was noticed. Furthermore, we compared the in silico interaction of lead compounds 5d and 5e toward the cocrystal structure of Suramin and RdRp polymerase with Remdesvir triphosphate, which showed that compounds 5d , 5e , and Remdesvir triphosphate (RTP) share a common catalytical site of RdRp but not Suramin. Additionally, the in silico ADMET properties predicted for the lead HTCAs and RTP showed that the maximum therapeutic doses recommended for compounds 5d and 5e were comparable to those of RTP. Concurrently, the pharmacokinetics of 5d was characterized in male Wistar Albino rats by administering a single oral gavage at a dose of 10 mg/kg, which gave a Cmax value of 0.22 μg/mL and a terminal elimination half-life period of 73.30 h. In conclusion, we established a new chemical entity that acts as a SARS-CoV-2 viral inhibitor with minimal or no toxicity to host cells in the rodent model, encouraging us to proceed with preclinical studies., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Background: In India, facility-based surveillance for congenital rubella syndrome (CRS) was initiated in 2016 to estimate the burden and monitor the progress made in rubella control. We analyzed the surveillance data for 2016-2021 from 14 sentinel sites to describe the epidemiology of CRS., Method: We analyzed the surveillance data to describe the distribution of suspected and laboratory confirmed CRS patients by time, place and person characteristics. We compared clinical signs of laboratory confirmed CRS and discarded case-patients to find independent predictors of CRS using logistic regression analysis and developed a risk prediction model., Results: During 2016-21, surveillance sites enrolled 3940 suspected CRS case-patients (Age 3.5 months, SD: 3.5). About one-fifth (n = 813, 20.6%) were enrolled during newborn examination. Of the suspected CRS patients, 493 (12.5%) had laboratory evidence of rubella infection. The proportion of laboratory confirmed CRS cases declined from 26% in 2017 to 8.7% in 2021. Laboratory confirmed patients had higher odds of having hearing impairment (Odds ratio [OR] = 9.5, 95% confidence interval [CI]: 5.6-16.2), cataract (OR = 7.8, 95% CI: 5.4-11.2), pigmentary retinopathy (OR = 6.7, 95 CI: 3.3-13.6), structural heart defect with hearing impairment (OR = 3.8, 95% CI: 1.2-12.2) and glaucoma (OR = 3.1, 95% CI: 1.2-8.1). Nomogram, along with a web version, was developed., Conclusions: Rubella continues to be a significant public health issue in India. The declining trend of test positivity among suspected CRS case-patients needs to be monitored through continued surveillance in these sentinel sites., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Background: Acute encephalitis syndrome (AES) is a major public health concern in India, and the Japanese Encephalitis (JE) virus is the most common cause of viral encephalitis in Asia affecting children under the age of 15 years. In India, despite the introduction of the JE vaccine (SA-14-14-2) in the immunization programme, JE continues to account for 15-20% of AES cases to date. This study evaluates the immunogenicity of live attenuated SA-14-14-2 JE vaccine in terms of persistence of the humoral response after two doses., Methods: A cross-sectional study was conducted among 266 children belonging to one of the JE endemic regions of Uttar Pradesh, India. Blood samples were taken from children (2-10 years) and grouped according to the duration (in years) after two doses of the vaccine (5 groups with a class interval of 2 years). Informed written consent was obtained from the parents/guardians. All the samples collected were tested for the presence of anti-JEV-specific IgG antibodies by enzyme-linked immunosorbent assay (ELISA) and further confirmed by micro neutralization test (MNT) and immunofluorescence assays., Results: Of the 266 samples tested by ELISA for anti-JEV-specific IgG antibodies, 260 (97.74%) were negative and 6 (2.26%) were equivocal. The geometric mean immune status ratio across the five groups, 0-2 years (n = 59), 2-4 years (n = 73), 4-6 years (n = 65), 6-8 years (n = 48) and 8-10 years (n = 21) post-two doses of SA-14-14-2 JE vaccine was 1.143, 1.059, 1.138, 1.075 and 1.130, respectively, and the geometric mean titre obtained from MNT across the five groups was 10.77, 8.400, 8.453, 9.517 and 9.674, respectively., Conclusion: The study showed a decreasing trend of anti-JEV specific IgG antibody titres across the five groups based on the duration following two doses of SA-14-14-2 vaccine. The results emphasize the significance of booster doses of vaccine for children living in endemic areas., (© International Society of Travel Medicine 2022. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The apprehension of needles related to injection site pain, risk of transmitting bloodborne pathogens, and effective mass immunization have led to the development of a needle-free injection system (NFIS). Here, we evaluated the efficacy of the NFIS and needle injection system (NIS) for the delivery and immunogenicity of DNA vaccine candidate ZyCoV-D in rhesus macaques against SARS-CoV-2 infection. Briefly, 20 rhesus macaques were divided into 5 groups (4 animals each), that is, I (1 mg dose by NIS), II (2 mg dose by NIS), III (1 mg dose by NFIS), IV (2 mg dose by NFIS) and V (phosphate-buffer saline [PBS]). The macaques were immunized with the vaccine candidates/PBS intradermally on Days 0, 28, and 56. Subsequently, the animals were challenged with live SARS-CoV-2 after 15 weeks of the first immunization. Blood, nasal swab, throat swab, and bronchoalveolar lavage fluid specimens were collected on 0, 1, 3, 5, and 7 days post infection from each animal to determine immune response and viral clearance. Among all the five groups, 2 mg dose by NFIS elicited significant titers of IgG and neutralizing antibody after immunization with enhancement in their titers postvirus challenge. Besides this, it also induced increased lymphocyte proliferation and cytokine response. The minimal viral load post-SARS-CoV-2 challenge and significant immune response in the immunized animals demonstrated the efficiency of NFIS in delivering 2 mg ZyCoV-D vaccine candidate., (© 2023 Wiley Periodicals LLC.)

Introduction: Supplementary immunization activities (SIAs) aim to interrupt measles transmission by reaching susceptible children, including children who have not received the recommended two routine doses of MCV before the SIA. However, both strategies may miss the same children if vaccine doses are highly correlated. How well SIAs reach children missed by routine immunization is a key metric in assessing the added value of SIAs., Methods: Children aged 9 months to younger than 5 years were enrolled in cross-sectional household serosurveys conducted in five districts in India following the 2017-2019 measles-rubella (MR) SIA. History of measles containing vaccine (MCV) through routine services or SIA was obtained from documents and verbal recall. Receipt of a first or second MCV dose during the SIA was categorized as "added value" of the SIA in reaching un- and under-vaccinated children., Results: A total of 1,675 children were enrolled in these post-SIA surveys. The percentage of children receiving a 1st or 2nd dose through the SIA ranged from 12.8% in Thiruvananthapuram District to 48.6% in Dibrugarh District. Although the number of zero-dose children prior to the SIA was small in most sites, the proportion reached by the SIA ranged from 45.8% in Thiruvananthapuram District to 94.9% in Dibrugarh District. Fewer than 7% of children remained measles zero-dose after the MR SIA (range: 1.1-6.4%) compared to up to 28% before the SIA (range: 7.3-28.1%)., Discussion: We demonstrated the MR SIA provided considerable added value in terms of measles vaccination coverage, although there was variability across districts due to differences in routine and SIA coverage, and which children were reached by the SIA. Metrics evaluating the added value of an SIA can help to inform the design of vaccination strategies to better reach zero-dose or undervaccinated children., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Introduction: Dengue, chikungunya and Japanese encephalitis are the most common arthropod-borne viral diseases in India. Due to overlapping clinical symptoms, accurate, high-quality and timely laboratory-based differential diagnosis is essential for control and containment of outbreaks. This is most commonly done by detection of IgM antibodies in serum using enzyme-linked immunosorbent assays. The Resource Centre for Virus Research and Diagnostic Laboratories (VRDLs) in Pune, India organized an external quality assurance (EQA) study to check the accuracy of serological diagnostics in the VRDL network., Methods: Three panels, one each for anti-dengue virus, anti-chikungunya virus and anti-Japanese encephalitis virus IgM antibodies, comprising six human serum samples (two positive and four negative) were distributed to test the sensitivity, specificity and reproducibility of serological testing in 124 VRDLs across India in 2018-19 and 2019-20., Results: Among the 124 VRDLs, the average concordance for both 2018-19 and 2019-20 was 98%. In 2018-19, 78.33%, 13.33% and 6.66% of VRDLs reported 100% concordance, 91-99% concordance and 81-90% concordance with the reference results, respectively, and 1.66% of VRDLs had concordance <80%. In 2019-20, 79.68%, 14.06% and 4.68% of VRDLs reported 100% concordance, 91-99% concordance and 81-90% concordance with the reference results, respectively, and 1.56% of VRDLs had concordance <80%., Conclusion: The EQA programme was beneficial for assessing and understanding the performance of the VRDLs. The study data indicate good proficiency in serological diagnosis of dengue, chikungunya and Japanese encephalitis in the VRDL network laboratories. Further expansion of the EQA programme to cover other viruses of public health importance will increase confidence among the VRDL network, and generate evidence of high-quality testing., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors.)

The immunity acquired after natural infection or vaccinations against SARS-CoV-2 tend to wane with time. Here, we compared the protective efficacy of COVAXIN® following two- and three-dose immunizations against the Delta variant and also studied the efficacy of COVAXIN® against Omicron variants in a Syrian hamster model. Despite the comparable neutralizing antibody response against the homologous vaccine strain in both the two-dose and three-dose immunized groups, considerable reduction in the lung disease severity was observed in the 3 dose immunized group after Delta variant challenge. In the challenge study using the Omicron variants, i.e., BA.1.1 and BA.2, lesser virus shedding, lung viral load and lung disease severity were observed in the immunized groups. The present study shows that administration of COVAXIN® booster dose will enhance the vaccine effectiveness against the Delta variant infection and give protection against the BA.1.1 and BA.2 variants., (© 2022 The Author(s).)

Background: In July 2021, an outbreak of chikungunya virus (CHIKV) was reported in a rural region of Maharashtra state, India., Methods: Serum samples of symptomatic cases (n=33) were screened for dengue virus (DENV), CHIKV and Zika virus (ZIKV) by molecular and serological assays., Results: The first case of ZIKV infection from Maharashtra was detected and confirmed by molecular and serological assays. Complete genome sequencing revealed that the ZIKV sequence belongs to the Asian genotype and had a closer homology with pre-epidemic strains present before 2007., Conclusions: ZIKV surveillance needs to be strengthened in the regions experiencing dengue and chikungunya outbreaks., (© The Author(s) 2022. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

This is a comprehensive report on immunogenicity of COVAXIN ® booster dose against ancestral and Variants of Concern (VOCs) up to 12 months. It is well known that neutralizing antibodies induced by COVID-19 vaccines wane within 6 months of vaccination leading to questions on the effectiveness of two-dose vaccination against breakthrough infections. Therefore, we assessed the persistence of immunogenicity up to 6 months after a two or three-dose with BBV152 and the safety of a booster dose in an ongoing phase 2, double-blind, randomized controlled trial (ClinicalTrials.gov: NCT04471519). We report persistence of humoral and cell mediated immunity up to 12 months of vaccination, despite decline in the magnitude of antibody titers. Administration of a third dose of BBV152 increased neutralization titers against both homologous (D614G) and heterologous strains (Alpha, Beta, Delta, Delta Plus and Omicron) with a slight increase in B cell memory responses. Thus, seronversion rate remain high in boosted recipients compared to non-booster, even after 6 months, post third dose against variants. No serious adverse events observed, except pain at the injection site, itching and redness. Hence, these results indicate that a booster dose of BBV152 is safe and necessary to ensure persistent immunity to minimize breakthrough infections of COVID-19, due to newly emerging variants.Trial registration: Registered with the Clinical Trials Registry (India) No. CTRI/2021/04/032942, dated 19/04/2021 and on Clinicaltrials.gov: NCT04471519., (© 2022. The Author(s).)

Background: SARS-CoV-2 Omicron variant is rampantly spreading across the globe. We assessed the pathogenicity and immune response generated by BA.1.1 sub-lineage of SARS-CoV-2 [Omicron (R346K) variant] in 5 to 6-week old Syrian hamsters and compared the observations with that of Delta variant infection., Methods: Virus shedding, organ viral load, lung disease and immune response generated in hamsters were sequentially assessed., Findings: The disease characteristics of the Omicron (R346K) variant were found to be similar to that of the Delta variant infection in hamsters like viral replication in the respiratory tract and interstitial pneumonia. The Omicron (R346K) infected hamsters demonstrated lesser body weight reduction and viral RNA load in the throat swab and nasal wash samples in comparison to the Delta variant infection. The viral load in the lungs and nasal turbinate samples and the lung disease severity of the Omicron (R346K) infected hamsters were found comparable with that of the Delta variant infected hamsters. Neutralizing antibody response against Omicron (R346K) variant was detected from day 5 and the cross-neutralization titre of the sera against other variants showed severe reduction ie., 7 fold reduction against Alpha and no titers against B.1, Beta and Delta., Interpretation: This preliminary data shows that Omicron (R346K) variant infection can produce moderate to severe lung disease similar to that of the Delta variant and the neutralizing antibodies produced in response to Omicron (R346K) variant infection shows poor neutralizing ability against other co-circulating SARS-CoV-2 variants like Delta which necessitates caution as it may lead to increased cases of reinfection., Funding: This study was supported by Indian Council of Medical Research as an intramural grant (COVID-19) to ICMR-National Institute of Virology, Pune., Competing Interests: Declaration of interests The authors declare no competing financial interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) Delta variant has evolved to become the dominant SARS-CoV-2 lineage with multiple sub-lineages and there are also reports of re-infections caused by this variant. We studied the disease characteristics induced by the Delta AY.1 variant and compared it with the Delta and B.1 variants in Syrian hamsters. We also assessed the potential of re-infection by these variants in Coronavirus disease 2019 recovered hamsters 3 months after initial infection. The variants produced disease characterized by high viral load in the respiratory tract and interstitial pneumonia. The Delta AY.1 variant produced mild disease in the hamster model and did not show any evidence of neutralization resistance due to the presence of the K417N mutation, as speculated. Re-infection with a high virus dose of the Delta and B.1 variants 3 months after B.1 variant infection resulted in reduced virus shedding, disease severity and increased neutralizing antibody levels in the re-infected hamsters. The reduction in viral load and lung disease after re-infection with the Delta AY.1 variant was not marked. Upper respiratory tract viral RNA loads remained similar after re-infection in all the groups. The present findings show that prior infection could not produce sterilizing immunity but that it can broaden the neutralizing response and reduce disease severity in case of reinfection.

Background: Considering the potential threat from emerging Severe Acute Respiratory Syndrome-Corona Virus-2 (SARS-CoV-2) variants and the rising COVID-19 cases, SARS-CoV-2 genomic surveillance is ongoing in India. We report herewith the isolation of the P.2 variant (B.1.1.28.2) from international travelers and further its pathogenicity evaluation and comparison with D614G variant (B.1) in hamster model., Methods: Virus isolation was performed in Vero CCL81 cells and genomic characterization by next generation sequencing. The pathogenicity and host immune response of the isolate was assessed in Syrian hamster model and compared with B.1 variant., Results: B.1.1.28.2 variant was isolated from nasal/throat swabs of international travelers returned to India from United Kingdom and Brazil. The B.1.1.28.2 variant induced body weight loss, viral replication in the respiratory tract and caused severe lung pathology in infected Syrian hamster model in comparison, with B.1 variant infected hamsters. The sera from B.1.1.28.2 infected hamsters efficiently neutralized the D614G variant virus whereas 6-fold reduction in the neutralization was seen in case of D614G variant infected hamsters' sera with the B.1.1.28.2 variant., Conclusions: B.1.1.28.2 lineage variant could be successfully isolated and characterization could be performed. Pathogenicity of the isolate was demonstrated in Syrian hamster model and the findings of neutralization reduction is of great concern and point towards the need for screening the vaccines for efficacy., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: We report the clinical efficacy against COVID-19 infection of BBV152, a whole virion inactivated SARS-CoV-2 vaccine formulated with a toll-like receptor 7/8 agonist molecule adsorbed to alum (Algel-IMDG) in Indian adults., Methods: We did a randomised, double-blind, placebo-controlled, multicentre, phase 3 clinical trial in 25 Indian hospitals or medical clinics to evaluate the efficacy, safety, and immunological lot consistency of BBV152. Adults (age ≥18 years) who were healthy or had stable chronic medical conditions (not an immunocompromising condition or requiring treatment with immunosuppressive therapy) were randomised 1:1 with a computer-generated randomisation scheme (stratified for the presence or absence of chronic conditions) to receive two intramuscular doses of vaccine or placebo administered 4 weeks apart. Participants, investigators, study coordinators, study-related personnel, the sponsor, and nurses who administered the vaccines were masked to treatment group allocation; an unmasked contract research organisation and a masked expert adjudication panel assessed outcomes. The primary outcome was the efficacy of the BBV152 vaccine in preventing a first occurrence of laboratory-confirmed (RT-PCR-positive) symptomatic COVID-19 (any severity), occurring at least 14 days after the second dose in the per-protocol population. We also assessed safety and reactogenicity throughout the duration of the study in all participants who had received at least one dose of vaccine or placebo. This report contains interim results (data cutoff May 17, 2021) regarding immunogenicity and safety outcomes (captured on days 0 to 56) and efficacy results with a median of 99 days for the study population. The trial was registered on the Indian Clinical Trials Registry India, CTRI/2020/11/028976, and ClinicalTrials.gov, NCT04641481 (active, not recruiting)., Findings: Between Nov 16, 2020, and Jan 7, 2021, we recruited 25 798 participants who were randomly assigned to receive BBV152 or placebo; 24 419 received two doses of BBV152 (n=12 221) or placebo (n=12 198). Efficacy analysis was dependent on having 130 cases of symptomatic COVID-19, which occurred when 16 973 initially seronegative participants had at least 14 days follow-up after the second dose. 24 (0·3%) cases occurred among 8471 vaccine recipients and 106 (1·2%) among 8502 placebo recipients, giving an overall estimated vaccine efficacy of 77·8% (95% CI 65·2-86·4). In the safety population (n=25 753), 5959 adverse events occurred in 3194 participants. BBV152 was well tolerated; the same proportion of participants reported adverse events in the vaccine group (1597 [12·4%] of 12 879) and placebo group (1597 [12·4%] of 12 874), with no clinically significant differences in the distributions of solicited, unsolicited, or serious adverse events between the groups, and no cases of anaphylaxis or vaccine-related deaths., Interpretation: BBV152 was highly efficacious against laboratory-confirmed symptomatic COVID-19 disease in adults. Vaccination was well tolerated with no safety concerns raised in this interim analysis., Funding: Bharat Biotech International and Indian Council of Medical Research., Competing Interests: Declaration of interests This work was funded by Bharat Biotech International and co-funded by the Indian Council of Medical Research. RE, KMV, SPr, SRe, VKA and VS are employees of Bharat Biotech International, with no stock options or incentives. KE is the chairman and managing director of Bharat Biotech International and owns equity in the company. WB is an independent statistical development consultant. VP, PY, GS, PA, NG, BB, SK, and SPa are employees of the Indian Council of Medical Research. All other authors declare no competing interests., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The pandemic of COVID-19 has caused enormous fatalities worldwide. Serological assays are important for detection of asymptomatic or mild cases of COVID-19, and sero-prevalence and vaccine efficacy studies. Here, we evaluated and compared the performance of seven commercially available enzyme-linked immunosorbent assay (ELISA)s for detection of anti-severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) immunoglobulin G (IgG). The ELISAs were evaluated with a characterized panel of 100 serum samples from qRT-PCR confirmed COVID-19 patients, collected 14 days post onset disease, 100 SARS-CoV-2 negative samples and compared the results with that of neutralization assay. Results were analysed by creating the receiver operating characteristic curve of all the assays in reference to the neutralization assay. All kits, were found to be suitable for detection of IgG against SARS-CoV-2 with high accuracy. The DiaPro COVID-19 IgG ELISA showed the highest sensitivity (98%) among the kits. The assays demonstrated high sensitivity and specificity in detecting the IgG antibodies against SARS-CoV-2. However, the presence of IgG antibodies does not always correspond to neutralizing antibodies. Due to their good accuracy indices, these assays can also aid in tracing mild infections, in cohort studies and in pre-vaccine evaluations., (© 2021 Wiley Periodicals LLC.)

Objectives: Monitoring the antibody responses to SARS-CoV-2 infection and its correlation to clinical spectrum of disease is critical in understanding the disease progression and protection against re-infection. We assessed the nucleocapsid (N) and receptor-binding-domain of spike (SRBD) protein specific IgG and neutralizing antibody (NAb) responses in COVID-19 patients up to 8 months and its correlation with diverse disease spectrum., Methods: During the first wave of the SARS-CoV-2 pandemic, from 284 COVID-19 patients, 608 samples were collected up to 8 months post infection. The patients were categorized as asymptomatic, symptomatic and severe. The N and SRBD IgG and NAb titers were evaluated and correlated with clinical data., Results: A steep increase in antigen specific antibody titers was observed till 40 days post onset of the disease (POD), followed by a partial decline till 240 days. Severe disease was associated with a stronger SRBD IgG response and higher NAb titers. The persistence of antibody response was observed in 76% against N, 80% against SRBD and 80% for NAbs of cases up to 8 months POD., Conclusion: RBD and N protein specific IgG persisted till 240 days POD which correlated with NAb response, irrespective of individual`s symptomatic status indicating overall robust protection against re-infection., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: Large data on the clinical characteristics and outcome of COVID-19 in the Indian population are scarce. We analysed the factors associated with mortality in a cohort of moderately and severely ill patients with COVID-19 enrolled in a randomised trial on convalescent plasma., Design: Secondary analysis of data from a Phase II, Open Label, Randomized Controlled Trial to Assess the Safety and Efficacy of Convalescent Plasma to Limit COVID-19 Associated Complications in Moderate Disease., Setting: 39 public and private hospitals across India during the study period from 22 April to 14 July 2020., Participants: Of the 464 patients recruited, two were lost to follow-up, nine withdrew consent and two patients did not receive the intervention after randomisation. The cohort of 451 participants with known outcome at 28 days was analysed., Primary Outcome Measure: Factors associated with all-cause mortality at 28 days after enrolment., Results: The mean (SD) age was 51±12.4 years; 76.7% were males. Admission Sequential Organ Failure Assessment score was 2.4±1.1. Non-invasive ventilation, invasive ventilation and vasopressor therapy were required in 98.9%, 8.4% and 4.0%, respectively. The 28-day mortality was 14.4%. Median time from symptom onset to hospital admission was similar in survivors (4 days; IQR 3-7) and non-survivors (4 days; IQR 3-6). Patients with two or more comorbidities had 2.25 (95% CI 1.18 to 4.29, p=0.014) times risk of death. When compared with survivors, admission interleukin-6 levels were higher (p<0.001) in non-survivors and increased further on day 3. On multivariable Fine and Gray model, severity of illness (subdistribution HR 1.22, 95% CI 1.11 to 1.35, p<0.001), PaO 2 /FiO 2 ratio <100 (3.47, 1.64-7.37, p=0.001), neutrophil lymphocyte ratio >10 (9.97, 3.65-27.13, p<0.001), D-dimer >1.0 mg/L (2.50, 1.14-5.48, p=0.022), ferritin ≥500 ng/mL (2.67, 1.44-4.96, p=0.002) and lactate dehydrogenase ≥450 IU/L (2.96, 1.60-5.45, p=0.001) were significantly associated with death., Conclusion: In this cohort of moderately and severely ill patients with COVID-19, severity of illness, underlying comorbidities and elevated levels of inflammatory markers were significantly associated with death., Trial Registration Number: CTRI/2020/04/024775., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

B.1.617 is becoming a dominant Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) lineage worldwide with many sublineages, of which B.1.617.2 is designated as a variant of concern. The pathogenicity of B.1.617.2 (Delta) and B.1.617.3 lineage of SARS-CoV-2 was evaluated and compared with that of B.1, an early virus isolate with D614G mutation in a Syrian hamster model. Viral load, antibody response, and lung disease were studied. There was no significant difference in the virus shedding pattern among these variants. High levels of SARS-CoV-2 sub genomic RNA were detected in the respiratory tract of hamsters infected with the Delta variant for 14 days, which warrants further transmission studies. The Delta variant induced lung disease of moderate severity in about 40% of infected animals, which supports the attributed disease severity of the variant. Cross neutralizing antibodies were detected in animals infected with B.1, Delta, and B.1.617.3 variant, but neutralizing capacity was significantly lower with B.1.351 (Beta variant).

Blood collection using dried blood spots (DBS) provides an easier alternative to venipuncture for sample collection, transport, and storage but requires additional processing that can cause variability in results. Whole-blood samples spotted on four DBS devices and respective paired serum samples were tested for antimeasles and antirubella IgG antibody concentrations by enzyme immunoassay. Elution protocols for DBS devices were optimized for comparability relative to serum samples using 12 adult volunteers. Stability of DBS collected on HemaSpot HF was assessed under various temperature conditions (+4, 22 to 25, and 45°C) at six time points (0, 7, 15, 30, 60, and 90 days) in a controlled laboratory setting using six adult volunteers. Devices were shipped and stored for 30 days at four settings with variable temperature and humidity conditions to assess the impact on antibody concentrations. Three DBS devices demonstrated comparable antibody concentrations with paired sera following optimization. Antibodies recovered from DBS were stable for at least 90 days at 4°C and for 30 days at ambient temperature (22 to 25°C) using the HemaSpot HF device. A drastic decline in antibody concentrations was observed at 45°C, resulting in quantitative and qualitative discrepancies by day 7. HemaSpot HF devices shipped to field sites and stored at ambient temperature and humidity resulted in quantitative, but not qualitative, variability. Measurement of antimeasles and antirubella IgG antibodies with DBS devices is an accurate alternative to testing serum, provided elution protocols are optimized. Stability of HemaSpot HF devices at ambient temperature enables broader use in surveys when serum processing and cold storage are not feasible. IMPORTANCE Dried blood spot (DBS) collection offers various advantages over conventional methods of blood collection, especially when collecting and transporting samples for a serosurvey. Yet use of DBS requires additional processing steps in the laboratory that can add to variability in results. We optimized a protocol to elute IgG antibodies against measles and rubella viruses in four DBS devices, demonstrating high concordance with paired venous sera for most devices. Extensive stability studies with various temperature and storage conditions in the laboratory and in the field were conducted using HemaSpot HF DBS devices prior to its use in one of the largest community-based measles and rubella serological surveys in the world.

Background: BBV152 is a whole-virion inactivated SARS-CoV-2 vaccine (3 μg or 6 μg) formulated with a toll-like receptor 7/8 agonist molecule (IMDG) adsorbed to alum (Algel). We previously reported findings from a double-blind, multicentre, randomised, controlled phase 1 trial on the safety and immunogenicity of three different formulations of BBV152 (3 μg with Algel-IMDG, 6 μg with Algel-IMDG, or 6 μg with Algel) and one Algel-only control (no antigen), with the first dose administered on day 0 and the second dose on day 14. The 3 μg and 6 μg with Algel-IMDG formulations were selected for this phase 2 study. Herein, we report interim findings of the phase 2 trial on the immunogenicity and safety of BBV152, with the first dose administered on day 0 and the second dose on day 28., Methods: We did a double-blind, randomised, multicentre, phase 2 clinical trial to evaluate the immunogenicity and safety of BBV152 in healthy adults and adolescents (aged 12-65 years) at nine hospitals in India. Participants with positive SARS-CoV-2 nucleic acid and serology tests were excluded. Participants were randomly assigned (1:1) to receive either 3 μg with Algel-IMDG or 6 μg with Algel-IMDG. Block randomisation was done by use of an interactive web response system. Participants, investigators, study coordinators, study-related personnel, and the sponsor were masked to treatment group allocation. Two intramuscular doses of vaccine were administered on day 0 and day 28. The primary outcome was SARS-CoV-2 wild-type neutralising antibody titres and seroconversion rates (defined as a post-vaccination titre that was at least four-fold higher than the baseline titre) at 4 weeks after the second dose (day 56), measured by use of the plaque-reduction neutralisation test (PRNT 50 ) and the microneutralisation test (MNT 50 ). The primary outcome was assessed in all participants who had received both doses of the vaccine. Cell-mediated responses were a secondary outcome and were assessed by T-helper-1 (Th1)/Th2 profiling at 2 weeks after the second dose (day 42). Safety was assessed in all participants who received at least one dose of the vaccine. In addition, we report immunogenicity results from a follow-up blood draw collected from phase 1 trial participants at 3 months after they received the second dose (day 104). This trial is registered at ClinicalTrials.gov, NCT04471519., Findings: Between Sept 5 and 12, 2020, 921 participants were screened, of whom 380 were enrolled and randomly assigned to the 3 μg with Algel-IMDG group (n=190) or 6 μg with Algel-IMDG group (n=190). Geometric mean titres (GMTs; PRNT 50 ) at day 56 were significantly higher in the 6 μg with Algel-IMDG group (197·0 [95% CI 155·6-249·4]) than the 3 μg with Algel-IMDG group (100·9 [74·1-137·4]; p=0·0041). Seroconversion based on PRNT 50 at day 56 was reported in 171 (92·9% [95% CI 88·2-96·2] of 184 participants in the 3 μg with Algel-IMDG group and 174 (98·3% [95·1-99·6]) of 177 participants in the 6 μg with Algel-IMDG group. GMTs (MNT 50 ) at day 56 were 92·5 (95% CI 77·7-110·2) in the 3 μg with Algel-IMDG group and 160·1 (135·8-188·8) in the 6 μg with Algel-IMDG group. Seroconversion based on MNT 50 at day 56 was reported in 162 (88·0% [95% CI 82·4-92·3]) of 184 participants in the 3 μg with Algel-IMDG group and 171 (96·6% [92·8-98·8]) of 177 participants in the 6 μg with Algel-IMDG group. The 3 μg with Algel-IMDG and 6 μg with Algel-IMDG formulations elicited T-cell responses that were biased to a Th1 phenotype at day 42. No significant difference in the proportion of participants who had a solicited local or systemic adverse reaction in the 3 μg with Algel-IMDG group (38 [20·0%; 95% CI 14·7-26·5] of 190) and the 6 μg with Algel-IMDG group (40 [21·1%; 15·5-27·5] of 190) was observed on days 0-7 and days 28-35; no serious adverse events were reported in the study. From the phase 1 trial, 3-month post-second-dose GMTs (MNT 50 ) were 39·9 (95% CI 32·0-49·9) in the 3μg with Algel-IMDG group, 69·5 (53·7-89·9) in the 6 μg with Algel-IMDG group, 53·3 (40·1-71·0) in the 6 μg with Algel group, and 20·7 (14·5-29·5) in the Algel alone group., Interpretation: In the phase 1 trial, BBV152 induced high neutralising antibody responses that remained elevated in all participants at 3 months after the second vaccination. In the phase 2 trial, BBV152 showed better reactogenicity and safety outcomes, and enhanced humoral and cell-mediated immune responses compared with the phase 1 trial. The 6 μg with Algel-IMDG formulation has been selected for the phase 3 efficacy trial., Funding: Bharat Biotech International., Translation: For the Hindi translation of the abstract see Supplementary Materials section., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: To mitigate the effects of COVID-19, a vaccine is urgently needed. BBV152 is a whole-virion inactivated SARS-CoV-2 vaccine formulated with a toll-like receptor 7/8 agonist molecule adsorbed to alum (Algel-IMDG) or alum (Algel)., Methods: We did a double-blind, multicentre, randomised, controlled phase 1 trial to assess the safety and immunogenicity of BBV152 at 11 hospitals across India. Healthy adults aged 18-55 years who were deemed healthy by the investigator were eligible. Individuals with positive SARS-CoV-2 nucleic acid and/or serology tests were excluded. Participants were randomly assigned to receive either one of three vaccine formulations (3 μg with Algel-IMDG, 6 μg with Algel-IMDG, or 6 μg with Algel) or an Algel only control vaccine group. Block randomisation was done with a web response platform. Participants and investigators were masked to treatment group allocation. Two intramuscular doses of vaccines were administered on day 0 (the day of randomisation) and day 14. Primary outcomes were solicited local and systemic reactogenicity events at 2 h and 7 days after vaccination and throughout the full study duration, including serious adverse events. Secondary outcome was seroconversion (at least four-fold increase from baseline) based on wild-type virus neutralisation. Cell-mediated responses were evaluated by intracellular staining and ELISpot. The trial is registered at ClinicalTrials.gov (NCT04471519)., Findings: Between July 13 and 30, 2020, 827 participants were screened, of whom 375 were enrolled. Among the enrolled participants, 100 each were randomly assigned to the three vaccine groups, and 75 were randomly assigned to the control group (Algel only). After both doses, solicited local and systemic adverse reactions were reported by 17 (17%; 95% CI 10·5-26·1) participants in the 3 μg with Algel-IMDG group, 21 (21%; 13·8-30·5) in the 6 μg with Algel-IMDG group, 14 (14%; 8·1-22·7) in the 6 μg with Algel group, and ten (10%; 6·9-23·6) in the Algel-only group. The most common solicited adverse events were injection site pain (17 [5%] of 375 participants), headache (13 [3%]), fatigue (11 [3%]), fever (nine [2%]), and nausea or vomiting (seven [2%]). All solicited adverse events were mild (43 [69%] of 62) or moderate (19 [31%]) and were more frequent after the first dose. One serious adverse event of viral pneumonitis was reported in the 6 μg with Algel group, unrelated to the vaccine. Seroconversion rates (%) were 87·9, 91·9, and 82·8 in the 3 μg with Algel-IMDG, 6 μg with Algel-IMDG, and 6 μg with Algel groups, respectively. CD4 + and CD8 + T-cell responses were detected in a subset of 16 participants from both Algel-IMDG groups., Interpretation: BBV152 led to tolerable safety outcomes and enhanced immune responses. Both Algel-IMDG formulations were selected for phase 2 immunogenicity trials. Further efficacy trials are warranted., Funding: Bharat Biotech International., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The COVID-19 pandemic is a global health crisis that poses a great challenge to the public health system of affected countries. Safe and effective vaccines are needed to overcome this crisis. Here, we develop and assess the protective efficacy and immunogenicity of an inactivated SARS-CoV-2 vaccine in rhesus macaques. Twenty macaques were divided into four groups of five animals each. One group was administered a placebo, while three groups were immunized with three different vaccine candidates of BBV152 at 0 and 14 days. All the macaques were challenged with SARS-CoV-2 fourteen days after the second dose. The protective response was observed with increasing SARS-CoV-2 specific IgG and neutralizing antibody titers from 3 rd -week post-immunization. Viral clearance was observed from bronchoalveolar lavage fluid, nasal swab, throat swab and lung tissues at 7 days post-infection in the vaccinated groups. No evidence of pneumonia was observed by histopathological examination in vaccinated groups, unlike the placebo group which exhibited interstitial pneumonia and localization of viral antigen in the alveolar epithelium and macrophages by immunohistochemistry. This vaccine candidate BBV152 has completed Phase I/II (NCT04471519) clinical trials in India and is presently in phase III, data of this study substantiates the immunogenicity and protective efficacy of the vaccine candidates.

The availability of a safe and effective vaccine would be the eventual measure to deal with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) threat. Here, we have assessed the immunogenicity and protective efficacy of inactivated SARS-CoV-2 vaccine candidates BBV152A, BBV152B, and BBV152C in Syrian hamsters. Three dose vaccination regimes with vaccine candidates induced significant titers of SARS-CoV-2-specific IgG and neutralizing antibodies. BBV152A and BBV152B vaccine candidates remarkably generated a quick and robust immune response. Post-SARS-CoV-2 infection, vaccinated hamsters did not show any histopathological changes in the lungs. The protection of the hamster was evident by the rapid clearance of the virus from lower respiratory tract, reduced virus load in upper respiratory tract, absence of lung pathology, and robust humoral immune response. These findings confirm the immunogenic potential of the vaccine candidates and further protection of hamsters challenged with SARS-CoV-2. Of the three candidates, BBV152A showed the better response., Competing Interests: The authors declare no competing interests., (© 2021 The Author(s).)

Background: First Zika virus (ZIKV) positive case from North India was detected on routine surveillance of Dengue-Like Illness in an 85-year old female. Objective of the study was to conduct an investigation for epidemiological, clinical and genomic analysis of first ZIKV outbreak in Rajasthan, North India and enhance routine ZIKV surveillance., Method: Outbreak investigation was performed in 3 Km radius of the index case among patient contacts, febrile cases, and pregnant women. Routine surveillance was enhanced to include samples from various districts of Rajasthan. Presence of ZIKV in serum and urine samples was detected by real time PCR test and CDC trioplex kit. Few ZIKV positive samples were sequenced using the next-generation sequencing method for genomic analysis., Result: On outbreak investigation 153/2043 (7.48%) cases were found positive: 1/153 (0.65%) among contacts, 90/153 (58.8%) in fever cases, 62/153(40.5%) in pregnant females. In routine surveillance, 6/4722 (0.12%) serum samples were ZIKV positive.Majority of patients had mild signs and symptoms, no case of microcephaly and Guillain- Barre Syndrome was seen, 25 (40.3%) pregnant females delivered healthy babies, four (6.4%) reported abortion and three (4.8%) had intrauterine death, one (1.6%) child had colorectal malformation and died after few days of birth. ZIKV was found to belong to Asian lineage, mutation related to enhanced neuro-virulence and transmission in animal models was not found., Conclusion: ZIKV was endogenous to India belonging to Asian Lineage. Disease profile of the ZIKV was asymptomatic to mild. No major anomaly was observed in infants born to ZIKV positive mothers; however, long term follow up of these children is required. There is need to scale up surveillance in the virology lab network of India for early detection and control., Summary Line: Zika virus infection was endogenous due to Asian Lineage with mild disease, no case of microcephaly or Guillain- Barre Syndrome was seen but children need to be followed for anomalies and surveillance of ZIKV needs to be enhanced in the country., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Zika virus is a mosquito-borne flavivirus that has emerged recently and affected in many countries. Since its discovery in Uganda in 1947, two major outbreaks were reported from Yap Islands in 2007 and French Polynesia in 2013. In 2015, the first case of ZIKV infection was confirmed from Brazil followed by a report of cases from American and Caribbean countries. In February 2016, the World Health Organization declared ZIKV infection a Public Health Emergency of International Concern. India reported the first Zika case in 2017. Subsequently, 157 laboratory-confirmed cases of ZIKV including 63 pregnant women were reported from Rajasthan, India in 2018. Since 2014, many countries took initiatives to boost their public health system to combat ZIKV. However, there is still scope for the improvement. This review describes ZIKV outbreaks, diagnostic challenges, surveillance and control measures in India and the future perspective to deal with the ZIKV outbreak in India., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background & Objectives: The genome of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), belonging to the family Coronaviridae, encodes for structural, non-structural, and accessory proteins, which are required for replication of the virus. These proteins are encoded by different genes present on the SARS-CoV-2 genome. The expression pattern of these genes in the host cells needs to be assessed. This study was undertaken to understand the transcription pattern of the SARS-CoV-2 genes in the Vero CCL-81 cells during the course of infection., Methods: Vero CCL-81 cells were infected with the SARS-CoV-2 virus inoculum having a 0.1 multiplicity of infection. The supernatants and cell pellets were harvested after centrifugation at different time points, post-infection. The 50% tissue culture infective dose (TCID 50 )and cycle threshold (C t ) values of the E and the RdRp-2 genes were calculated. Next-generation sequencing of the harvested sample was carried out to observe the expression pattern of the virus by mapping to the SARS-CoV-2 Wuhan HU-1 reference sequence. The expressions were in terms of the reads per kilobase million (RPKM) values., Results: In the inital six hours post-infection, the copy numbers of E and RdRp-2 genes were approximately constant, which raised 10 log-fold and continued to increase till the 12 h post-infection (hpi). The TCID 50 was observed in the supernatant after 7 hpi, indicating the release of the viral progeny. ORF8 and ORF7a, along with the nucleocapsid transcript, were found to express at higher levels., Interpretation & Conclusions: This study was a step towards understanding the growth kinetics of the SARS-CoV-2 replication cycle. The findings indicated that ORF8 and ORF7b gene transcripts were expressed in higher amounts indicating their essential role in viral replication. Future studies need to be conducted to explore their role in the SARS-CoV-2 replication., Competing Interests: None

Background & Objectives: Since the beginning of the year 2020, the pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impacted humankind adversely in almost all spheres of life. The virus belongs to the genus Betacoronavirus of the family Coronaviridae. SARS-CoV-2 causes the disease known as coronavirus disease 2019 (COVID-19) with mild-to-severe respiratory illness. The currently available diagnostic tools for the diagnosis of COVID-19 are mainly based on molecular assays. Real-time reverse transcription-polymerase chain reaction is the only diagnostic method currently recommended by the World Health Organization for COVID-19. With the rapid spread of SARS-CoV-2, it is necessary to utilize other tests, which would determine the burden of the disease as well as the spread of the outbreak. Considering the need for the development of such a screening test, an attempt was made to develop and evaluate an IgG-based ELISA for COVID-19., Methods: A total of 513 blood samples (131 positive, 382 negative for SARS-CoV-2) were collected and tested by microneutralization test (MNT). Antigen stock of SARS-CoV-2 was prepared by propagating the virus in Vero CCL-81 cells. An IgG capture ELISA was developed for serological detection of anti-SARS-CoV-2 IgG in serum samples. The end point cut-off values were determined by using receiver operating characteristic (ROC) curve. Inter-assay variability was determined., Results: The developed ELISA was found to be 92.37 per cent sensitive, 97.9 per cent specific, robust and reproducible. The positive and negative predictive values were 94.44 and 98.14 per cent, respectively., Interpretation & Conclusions: This indigenously developed IgG ELISA was found to be sensitive and specific for the detection of anti-SARS-CoV-2 IgG in human serum samples. This assay may be used for determining seroprevalence of SARS-CoV-2 in a population exposed to the virus., Competing Interests: None

Background: Government of India is committed to eliminate measles and control rubella/congenital rubella syndrome (CRS) by 2020. In 2016, CRS surveillance was established in five sentinel sites. We analyzed surveillance data to describe the epidemiology of CRS in India., Methodology/principal Findings: We used case definitions adapted from the WHO-recommended standards for CRS surveillance. Suspected patients underwent complete clinical examination including cardiovascular system, ophthalmic examination and assessment for hearing impairment. Sera were tested for presence of IgM and IgG antibodies against rubella. Of the 645 suspected CRS patients enrolled during two years, 137 (21.2%) were classified as laboratory confirmed CRS and 8 (1.2%) as congenital rubella infection. The median age of laboratory confirmed CRS infants was 3 months. Common clinical features among laboratory confirmed CRS patients included structural heart defects in 108 (78.8%), one or more eye signs (cataract, glaucoma, pigmentary retinopathy) in 82 (59.9%) and hearing impairment in 51. (38.6%) Thirty-three (24.1%) laboratory confirmed CRS patients died over a period of 2 years. Surveillance met the quality indicators in terms of adequacy of investigation, adequacy of sample collection for serological diagnosis as well as virological confirmation., Conclusions/significance: About one fifth suspected CRS patients were laboratory confirmed, indicating significance of rubella as a persistent public health problem in India. Continued surveillance will generate data to monitor the progress made by the rubella control program in the country., Competing Interests: The authors have declared that no competing interests exist.

Background & Objectives: An outbreak of respiratory illness of unknown aetiology was reported from Hubei province of Wuhan, People's Republic of China, in December 2019. The outbreak was attributed to a novel coronavirus (CoV), named as severe acute respiratory syndrome (SARS)-CoV-2 and the disease as COVID-19. Within one month, cases were reported from 25 countries. In view of the novel viral strain with reported high morbidity, establishing early countrywide diagnosis to detect imported cases became critical. Here we describe the role of a countrywide network of VRDLs in early diagnosis of COVID-19., Methods: The Indian Council of Medical Research (ICMR)-National Institute of Virology (NIV), Pune, established screening as well as confirmatory assays for SARS-CoV-2. A total of 13 VRDLs were provided with the E gene screening real-time reverse transcription-polymerase chain reaction (rRT-PCR) assay. VRDLs were selected on the basis of their presence near an international airport/seaport and their past performance. The case definition for testing included all individuals with travel history to Wuhan and symptomatic individuals with travel history to other parts of China. This was later expanded to include symptomatic individuals returning from Singapore, Japan, Hong Kong, Thailand and South Korea., Results: Within a week of standardization of the test at NIV, all VRDLs could initiate testing for SARS-CoV-2. Till February 29, 2020, a total of 2,913 samples were tested. This included both 654 individuals quarantined in the two camps and others fitting within the case definition. The quarantined individuals were tested twice - at days 0 and 14. All tested negative on both occasions. Only three individuals belonging to different districts in Kerala were found to be positive., Interpretation & Conclusions: Sudden emergence of SARS-CoV-2 and its potential to cause a pandemic posed an unsurmountable challenge to the public health system of India. However, concerted efforts of various arms of the Government of India resulted in a well-coordinated action at each level. India has successfully demonstrated its ability to establish quick diagnosis of SARS-CoV-2 at NIV, Pune, and the testing VRDLs., Competing Interests: None