Assessing the Burden of COVID-19 among Children Aged 6-14 Years in Karnataka, India: A Cross-sectional Survey.

Background: India experienced three coronavirus disease (COVID-19) waves, with the third attributed to the highly contagious Omicron variant. Before the national vaccination rollout for children above 6, understanding severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunoglobulin G (IgG) positivity in the pediatric population was essential. This study aims to assess the burden of Covid-19 infection and to estimate the seroprevalence in children aged 6 to 14 years in the state of Karnataka. Material and Methods: We surveyed 5,358 children aged 6-14 across Karnataka using 232 health facilities, from June 6 to 14, 2022. We determined the sample size using the PPS (Population Proportional to Size) technique and employed cluster sampling. We tested all participants for SARS-CoV-2 IgG with an enzyme-linked immunosorbent assay (ELISA) kit and SARS-CoV-2 RNA with reverse transcription-polymerase chain reaction (RT-PCR). We sequenced samples with a cycle threshold (CT) value below 25 using whole genomic sequencing (WGS). Result: We found an adjusted seroprevalence of IgG at 75.38% statewide, and we found 0.04% of children RT-PCR positive for COVID-19. We determined a case-to-infection ratio of 1:37 and identified the SARS-CoV-2 strains as Omicron, BA.5, and BA.2.10. Conclusion: The study showed a high seroprevalence of IgG among children with low active infection. Omicron, BA. 5, and BA. 2.10 variants were detected through WGS.

Corrigendum to "Second round statewide sentinel-based population survey for estimation of the burden of active infection and anti-SARS-CoV-2 IgG antibodies in the general population of Karnataka, India, during January-February 2021" [IJID Regions Vol 1(2021) pages 107-116].

[This corrects the article DOI: 10.1016/j.ijregi.2021.10.008.].

Clinical, epidemiological, and molecular investigation of Kyasanur forest disease from Karnataka state, India during 2018-2019.

BACKGROUND: In this study, we carried out an investigation of Kyasanur Forest Disease (KFD) suspected human cases reported in Karnataka state, India from December 2018 to June 2019. METHODS: The clinical samples of KFD suspected cases (n = 1955) from 14 districts of Karnataka were tested for KFD using real-time RT-PCR and IgM ELISA. Further, the KFD-negative samples were tested for IgM antibodies against dengue and chikungunya viruses. Monkey samples (n = 276) and tick pools (n = 11582) were also screened using real-time RT-PCR. KFD-positive samples were further analysed using next-generation sequencing along with clinico-epidemiological analysis. RESULTS: Of all, 173 (8.8%) cases tested positive for KFD either by real-time RT-PCR (n = 124), IgM ELISA (n = 53) or both tests (n = 4) from seven districts. Among KFD-negative cases, IgM antibody positivity was observed for dengue (2.6%), chikungunya (5.8%), dengue and chikungunya coinfection (3.7%). KFD cases peaked in January 2019 with fever, conjunctivitis, and myalgia as the predominant symptoms and a mortality of 4.6%. Among confirmed cases, 41% received a single dose and 20% received two doses of the KFD vaccine. Of the seven districts with KFDV positivity, Shivamogga and Hassan districts reported KFD viral RNA positivity in humans, monkeys, and ticks. Sequencing analysis of 2019 cases demonstrated a difference of less than 1.5% amino acid compared to prototype KFDV. CONCLUSION: Although the KFD has been endemic in many districts of Karnataka state, our study confirms the presence of KFDV for the first time in two new districts, i.e. Hassan and Mysore. A comparative analysis of KFDV infection among the KFD-vaccinated and non-vaccinated populations demonstrated an insignificant difference.

Molecular epidemiology of hand, foot, and mouth disease in Karnataka, India in 2022.

BACKGROUND: Hand, foot, and mouth disease (HFMD) is an enteroviral disease that occurs as outbreaks and sporadic cases in India. In this study, we investigated and characterized the aetiology of HFMD cases that occurred in Karnataka, South India from April to October 2022. METHODS: Throat swabs, vesicular swabs, urine, and blood samples from suspected cases were analysed by reverse transcription polymerase chain reaction (RT-PCR) for the detection of enteroviruses. Molecular typing of the enterovirus-positive samples was carried out by amplifying the partial virion protein 1(VP1) gene sequence, followed by sequencing and phylogenetic analysis. RESULTS: Out of the 187 samples received from 82 cases, 93 (50%) tested positive (55/82 cases, 67%) for enteroviruses, with the majority of the HFMD cases reported in paediatric population of less than 5 years (36/55, 65.4%), while 3 cases (3/55, 5.4%) were adults. Out of the 55 enterovirus-positive cases, 31 showed partial VP1 region amplification and 19 of these cases were typed as coxsackievirus A16 (CV-A16) (13/19, 68.4%) and CV-A6 (6/19, 31.6%). The CV-A16 strains identified belonged to subclade B1c while two CV-A6 strains belonged to subclade E2. On molecular testing for other viruses causing fever-rash symptoms, 4/27 (15%) enterovirus-negative cases were detected as herpes simplex virus (1 case) and varicella zoster virus (3 cases) positive. CONCLUSION: The main causative agent of HFMD in Karnataka in 2022 was CV-A16, followed by CV-A6. Apart from the common paediatric HFMD cases, adult cases were also reported during this period. Further studies involving laboratory and clinical investigations are essential for monitoring and managing HFMD in the community.

Trend of coronavirus disease 2019 pandemic in Bengaluru, Karnataka, India.

The study reports the trend of coronavirus disease 2019 (COVID-19) infection observed from 2020 to 2022 in the city of Bengaluru, Karnataka, India, across each week in relation to the corresponding severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sewage positivity data and other non-COVID-19 respiratory viral infections such as influenza and respiratory syncytial virus (RSV). The data on COVID-19 cases were procured from public domains, whereas the molecular testing of sewage samples and clinical samples for influenza and RSV was performed at our unit. The pattern of sewage positivity matched the waves of COVID-19, and few sewage samples in 2021 and 2022 tested positive before the onset of clinical cases. Influenza and RSV cases were reported during the periods of low COVID-19 cases. Thus, sewage surveillance is effective in monitoring the circulating SARS-CoV-2 virus, while laboratory surveillance of SARS-CoV-2, influenza, and RSV cases is pertinent to identify the common viral etiology of respiratory infections in the community.

Profile of cardiac lesions among laboratory confirmed congenital rubella syndrome (CRS) infants: a nationwide sentinel surveillance, India, 2016-22.

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.

Molecular characterization & recombination analysis of complete enterovirus-88 isolated from acute flaccid paralysis cases in India.

Background & objectives: Focus on non-polio enteroviruses (NPEVs) causing acute flaccid paralysis (AFP) due to myelitis has increased with the containment of the poliovirus. Enterovirus-B88 (EV-B88) has been associated with the AFP cases in Bangladesh, Ghana, South Africa, Thailand and India. In India, EV-B88 infection was linked to AFP a decade ago; however, to date, no complete genome has been made available. In this study, the complete genome sequence of EV-B88 was identified and reported from two different States (Bihar and Uttar Pradesh) in India using the next-generation sequencing technique. Methods: Virus isolation was performed on the three AFP suspected cases as per the WHO-recommended protocol. Samples showing cytopathic effects in the human Rhabdocarcinoma were labelled as NPEVs. Next-generation sequencing was performed on these NPEVs to identify the aetiological agent. The contiguous sequences (contigs) generated were identified, and reference-based mapping was performed. Results: EV-B88 sequences retrieved in our study were found to be 83 per cent similar to the EV-B88 isolate from Bangladesh in 2001 (strain: BAN01-10398; Accession number: AY843306.1). Recombination analyses of these samples demonstrate recombination events with sequences from echovirus-18 and echovirus-30. Interpretation & conclusions: Recombination events in the EV-B serotypes are known, and this work reconfirms the same for EV-B88 isolates also. This study is a step in increasing the awareness about EV-B88 in India and emphasizes future studies to be conducted in the identification of other types of EV present in India.

A case with SARS-CoV-2 reinfection from India.

A healthcare worker presented with fever, cough, headache and tested positive by SARS-CoV-2 real time reverse transcriptase polymerase chain reaction (qRT-PCR). He got admitted to hospital and recovered after 14 days. After 2 months, as a screening protocol considering the high risk setup he got tested and again found to be positive for SARS-CoV-2 by qRT-PCR. Our patient had detectable levels of Anti-SARS-CoV-2 IgG antibodies during the reinfection but found negative for Neutralizing antibodies (NAb). Our findings suggest that the person after the initial infection might not develop the desired protective immunity to prevent the reinfection as demonstrated by absence of NAb.

Identification of Phasi Charoen-Like Phasivirus in Field Collected Aedes aegypti from Karnataka State, India.

Background: A wide range of insect-specific viruses (ISVs) have been reported worldwide. There are no studies from India that have reported ISVs. The current study describes the identification of Phasi Charoen-like virus (PCLV) from Aedes aegypti mosquito-pools from six districts of Karnataka state, India. Materials and Methods: During the Chikungunya virus (CHIKV) outbreak in the Bangalore Urban district in 2019, using conventional PCR, it was found that both human and mosquito samples were positive for CHIKV. For retrieve the complete genome sequence, mosquito samples were subjected to next generation sequencing (NGS) analysis and PCLV was also found. During 2019, as part of a vector-borne disease surveillance, we received 50 mosquito pool samples from 6 districts of the state, all of them were subjected to NGS to identify PCLV. Results: The A. aegypti mosquito-pools samples were subjected to the NGS platform that led to identification of an ISV, PCLV. PCLV was identified in 26 A. aegypti mosquito-pools collected from 6 districts. We also found mixed infection of PCLV with the Dengue virus (DENV; genotypes 1 and 3) and CHIKV from five pools. The nucleotide identity for the L gene of Indian PCLV sequences ranged between 97.1% and 98.3% in comparison with the Thailand sequences. Conclusions: To the best of our knowledge, this is the first report of PCLV dual infection with DENV and CHIKV in India. The present study confirms the presence of PCLV in A. aegypti mosquitoes from Karnataka state. The study adds India in the global geographical distribution of PCLV.

Clinical Characterization and Genomic Analysis of Samples from COVID-19 Breakthrough Infections during the Second Wave among the Various States of India.

From March to June 2021, India experienced a deadly second wave of COVID-19, with an increased number of post-vaccination breakthrough infections reported across the country. To understand the possible reason for these breakthroughs, we collected 677 clinical samples (throat swab/nasal swabs) of individuals from 17 states/Union Territories of the country who had received two doses (n = 592) and one dose (n = 85) of vaccines and tested positive for COVID-19. These cases were telephonically interviewed and clinical data were analyzed. A total of 511 SARS-CoV-2 genomes were recovered with genome coverage of higher than 98% from both groups. Analysis of both groups determined that 86.69% (n = 443) of them belonged to the Delta variant, along with Alpha, Kappa, Delta AY.1, and Delta AY.2. The Delta variant clustered into four distinct sub-lineages. Sub-lineage I had mutations in ORF1ab A1306S, P2046L, P2287S, V2930L, T3255I, T3446A, G5063S, P5401L, and A6319V, and in N G215C; Sub-lineage II had mutations in ORF1ab P309L, A3209V, V3718A, G5063S, P5401L, and ORF7a L116F; Sub-lineage III had mutations in ORF1ab A3209V, V3718A, T3750I, G5063S, and P5401L and in spike A222V; Sub-lineage IV had mutations in ORF1ab P309L, D2980N, and F3138S and spike K77T. This study indicates that majority of the breakthrough COVID-19 clinical cases were infected with the Delta variant, and only 9.8% cases required hospitalization, while fatality was observed in only 0.4% cases. This clearly suggests that the vaccination does provide reduction in hospital admission and mortality.

An Epidemiological Analysis of SARS-CoV-2 Genomic Sequences from Different Regions of India.

The number of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) cases is increasing in India. This study looks upon the geographic distribution of the virus clades and variants circulating in different parts of India between January and August 2020. The NPS/OPS from representative positive cases from different states and union territories in India were collected every month through the VRDLs in the country and analyzed using next-generation sequencing. Epidemiological analysis of the 689 SARS-CoV-2 clinical samples revealed GH and GR to be the predominant clades circulating in different states in India. The northern part of India largely reported the 'GH' clade, whereas the southern part reported the 'GR', with a few exceptions. These sequences also revealed the presence of single independent mutations-E484Q and N440K-from Maharashtra (first observed in March 2020) and Southern Indian States (first observed in May 2020), respectively. Furthermore, this study indicates that the SARS-CoV-2 variant (VOC, VUI, variant of high consequence and double mutant) was not observed during the early phase of virus transmission (January-August). This increased number of variations observed within a short timeframe across the globe suggests virus evolution, which can be a step towards enhanced host adaptation.

The burden of active infection and anti-SARS-CoV-2 IgG antibodies in the general population: Results from a statewide sentinel-based population survey in Karnataka, India.

OBJECTIVE: To estimate the burden of active infection and anti-SARS-CoV-2 IgG antibodies in Karnataka, India, and to assess variation across geographical regions and risk groups. METHODS: A cross-sectional survey of 16,416 people covering three risk groups was conducted between 3-16 September 2020 using the state of Karnataka's infrastructure of 290 healthcare facilities across all 30 districts. Participants were further classified into risk subgroups and sampled using stratified sampling. All participants were subjected to simultaneous detection of SARS-CoV-2 IgG using a commercial ELISA kit, SARS-CoV-2 antigen using a rapid antigen detection test (RAT) and reverse transcription-polymerase chain reaction (RT-PCR) for RNA detection. Maximum-likelihood estimation was used for joint estimation of the adjusted IgG, active and total prevalence (either IgG or active or both), while multinomial regression identified predictors. RESULTS: The overall adjusted total prevalence of COVID-19 in Karnataka was 27.7% (95% CI 26.1-29.3), IgG 16.8% (15.5-18.1) and active infection fraction 12.6% (11.5-13.8). The case-to-infection ratio was 1:40 and the infection fatality rate was 0.05%. Influenza-like symptoms or contact with a COVID-19-positive patient were good predictors of active infection. RAT kits had higher sensitivity (68%) in symptomatic people compared with 47% in asymptomatic people. CONCLUSION: This sentinel-based population survey was the first comprehensive survey in India to provide accurate estimates of the COVID-19 burden. The findings provide a reasonable approximation of the population immunity threshold levels. Using existing surveillance platforms coupled with a syndromic approach and sampling framework enabled this model to be replicable.

Genetic and antigenic characterization of wild type rubella viruses isolated from India.

Rubella, is a contagious disease caused by Rubella virus (RuV) that manifests as fever with skin-rashes in children and adults along with complications in pregnant women. WHO-SEAR has set a target for Rubella elimination by 2023. This is the first report of antigenic characterization and genome sequencing of nine RuVs sampled during 1992, 2007-9, and 2015-17 from four Indian states. Comparative analysis of Indian RuVs (2B) with that of global isolates and vaccine strain RA 27/3 (1a) revealed that the observed mutations in structural proteins have no major impact on the 3D structure, function and antigenicity. Indian RuVs formed three major clusters (Pune-1992, Kannur-2009 and Chitradurg-2007) in genome-based phylogeny of global isolates. Neutralizing antibody titers in a panel of serum samples from measles negative cases were significantly higher to the vaccine strain compared to a wild-type 2B isolate (Kannur) with concordance of 91.9%, thereby substantiating the use of current vaccines.

Second round statewide sentinel-based population survey for estimation of the burden of active infection and anti-SARS-CoV-2 IgG antibodies in the general population of Karnataka, India, during January-February 2021.

Objective: Demonstrate the feasibility of using the existing sentinel surveillance infrastructure to conduct the second round of the serial cross-sectional sentinel-based population survey. Assess active infection, seroprevalence, and their evolution in the general population across Karnataka. Identify local variations for locally appropriate actions. Additionally, assess the clinical sensitivity of the testing kit used on account of variability of antibody levels in the population. Methods: The cross-sectional study of 41,228 participants across 290 healthcare facilities in all 30 districts of Karnataka was done among three groups of participants (low, moderate, and high-risk). The geographical spread was sufficient to capture local variations. Consenting participants were subjected to real-time reverse transcription-polymerase chain reaction (RT-PCR) testing, and antibody (IgG) testing. Clinical sensitivity was assessed by conducting a longitudinal study among participants identified as COVID-19 positive in the first survey round. Results: Overall weighted adjusted seroprevalence of IgG was 15.6% (95% CI: 14.9-16.3), crude IgG prevalence was 15.0% and crude active infection was 0.5%. Statewide infection fatality rate (IFR) was estimated as 0.11%, and COVID-19 burden estimated between 26.1 to 37.7% (at 90% confidence). Further, Cases-to-infections ratio (CIR) varied 3-35 across units and IFR varied 0.04-0.50% across units. Clinical sensitivity of the IgG ELISA test kit was estimated as ≥38.9%. Conclusion: We demonstrated the feasibility and simplicity of sentinel-based population survey in measuring variations in subnational and local data, useful for locally appropriate actions in different locations. The sentinel-based population survey thus helped identify districts that needed better testing, reporting, and clinical management. The state was far from attaining natural immunity during the survey and hence must step up vaccination coverage and enforce public health measures to prevent the spread of COVD-19.

Laboratory preparedness for SARS-CoV-2 testing in India: Harnessing a network of Virus Research & Diagnostic Laboratories.

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.