Understanding the resurgence of chikungunya virus during 2020-2021 in Pune, India, based on genomic analyses: A seven year study.

A surge in chikungunya was observed during 2020-21 in Pune district of Maharashtra, India. Whole genome sequencing and phylogenetic analysis of 21 samples/sequences revealed them as Indian ocean lineage of East Central South African genotype. Two distinct sequence clusters were found to circulate during 2020-21; one with E1:K211E and E2:V264A mutations while the other had E1:I317V mutation along with E1:K211E and E2: V264A mutations. The former, the predominant cluster (n = 18), clustered with chikungunya virus (CHIKV) strains of pre 2014 period while the latter (n = 3) clustered with 2016-2018 period Indian strains. Though E1: A226V was not detected in any of the 21 sequences, several unique mutations were detected in the strains which might have played key roles in the enhanced virus transmission during the period. The study highlights parallel evolution, introduction from the neighboring regions and cocirculation of two sequence clusters of CHIKV in Pune. The complete genome data can be useful to determine how the circulating strains differ from candidate vaccines and might help to predict the protective efficacy of chikungunya vaccine candidates.

In Vitro Antiviral Activity of Potential Medicinal Plant Extracts Against Dengue and Chikungunya Viruses.

Dengue and chikungunya are two important mosquito-borne infections which are known to occur extensively in tropical and subtropical areas. Presently, there is no treatment for these viral diseases. In vitro antiviral screening of 25 extracts prepared from the plants of Vitex negundo, Plumeria alba, Ancistrocladus heyneanus, Bacopa monnieri, Anacardium occidentale, Cucurbita maxima, Simarouba glauca, and Embelia ribes using different solvents and four purified compounds (anacardic acid, chloroquinone, glaucarubinone, and methyl gallate) were carried out for their anti-dengue virus (DENV) and anti-chikungunya virus (CHIKV) activities. Maximum nontoxic concentrations of the chloroform, methanol, ethyl acetate, petroleum ether, dichloromethane, and hydroalcoholic extracts of eight plants were used. The antiviral activity was assessed by focus-forming unit assay, quantitative real-time RT-PCR, and immunofluorescence assays. Extracts from Plumeria alba, Ancistrocladus heyneanus, Bacopa monnieri, and Cucurbita maxima showed both anti-DENV and CHIKV activity while extract from Vitex negundo showed only anti-DENV activity. Among the purified compounds, anacardic acid, chloroquinone and methyl gallate showed anti-dengue activity while only methyl gallate had anti-chikungunya activity. The present study had identified the plant extracts with anti-dengue and anti-chikungunya activities, and these extracts can be further characterized for finding effective phytopharmaceutical drugs against dengue and chikungunya.

SARS-CoV-2 and Influenza Virus Co-Infection Cases Identified through ILI/SARI Sentinel Surveillance: A Pan-India Report.

SARS-CoV-2/influenza virus co-infection studies have focused on hospitalized patients who usually had grave sequelae. Here, we report SARS-CoV-2/influenza virus co-infection cases from both community and hospital settings reported through integrated ILI/SARI (Influenza Like Illness/Severe Acute Respiratory Infection) sentinel surveillance established by the Indian Council of Medical Research. We describe the disease progression and outcomes in these cases. Out of 13,467 samples tested from 4 July 2021-31 January 2022, only 5 (0.04%) were of SARS-CoV-2/influenza virus co-infection from 3 different sites in distinct geographic regions. Of these, three patients with extremes of age required hospital admission, but none required ICU admission or mechanical ventilation. No mortality was reported. The other two co-infection cases from community settings were managed at home. This is the first report on SARS-CoV-2/Influenza virus co-infection from community as well as hospital settings in India and shows that influenza viruses are circulating in the community even during COVID-19. The results emphasize the need for continuous surveillance for multiple respiratory pathogens for effective public health management of ILI/SARI cases in line with the WHO (World Health Organization) recommendations.

Complete Genome Sequence of Mumps Virus Isolated from Karnataka State, India.

We report the first whole-genome sequence of mumps virus isolated from a two-year-old girl with bilateral parotitis from a Chikkahallivana village in the Davangere district of Karnataka State, India. The genome of the Davangere mumps isolate was 15,384 bp in length and identical to previously published mumps virus (MuV) genomes from India. BLAST results show 99.1% identity with previously sequenced genotype C viruses isolated from the states of Maharashtra, Tamil Nadu, and Uttar Pradesh.

Measles virus genotypes circulating in India, 2011-2015.

The Government of India is accepted to participate in the measles elimination and rubella control goal 2020, hence genetic characterization of measles viruses (MeV) becomes essential. At National Reference Laboratory (National Institute of Virology, Pune), the throat swabs/urine specimens (n = 380) or PCR products (n = 219) obtained from the suspected measles cases were referred for the molecular testing and subsequently, MeV nucleoprotein (N) gene sequencing/genotyping. In addition, 2,449 suspected measles cases, mainly from the Maharashtra state were referred for the laboratory diagnosis. A detailed study was performed on N gene sequences obtained during last two decades. Indian MeV sequences obtained during 2011-2015 were compared with 1996-2010 sequences and genetic divergence was studied. Circulation of measles genotypes B3 (n = 3), D4 (n = 49), and D8 (n = 351) strains were observed in 19 States and three Union Territories of India. In addition, 64 measles viruses were isolated from 253 throat swab or urine specimens obtained from the suspected measles cases. During 2011-2015, 67.9% (1,663/2,449) suspected measles cases were laboratory confirmed. Molecular studies showed circulation of measles genotype B3 in India along with prominently circulating genotypes D4 and D8 except D7 strains. The genetic diversion within Indian B3, D4, and D8 genotypes was 0.3%, 1.1%, and 2.1%, respectively. The genetic divergence of Indian B3, D4, and D8 measles strains with the WHO reference sequences was 2.5%, 2.6%, and 1.8%, respectively. It is crucial data for national immunization program. More measles/rubella genotyping studies are necessary to track transmission and to support measles elimination and rubella control. J. Med. Virol. 89:753-758, 2017. © 2016 Wiley Periodicals, Inc.

Measles & rubella outbreaks in Maharashtra State, India.

BACKGROUND & OBJECTIVES: Under the outbreak-based measles surveillance in Maharashtra State the National Institute of Virology at Pune receives 3-5 serum samples from each outbreak and samples from the local hospitals in Pune for laboratory diagnosis. This report describes one year data on the measles and rubella serology, virus isolation and genotyping. METHODS: Maharashtra State Health Agencies investigated 98 suspected outbreaks between January-December 2013 in the 20 districts. Altogether, 491 serum samples were received from 20 districts and 126 suspected cases from local hospitals. Samples were tested for the measles and rubella IgM antibodies by commercial enzyme immunoassay (EIA). To understand the diagnostic utility, a subset of serum samples (n=53) was tested by measles focus reduction neutralization test (FRNT). Further, 37 throat swabs and 32 urine specimens were tested by measles reverse transcription (RT)-PCR and positive products were sequenced. Virus isolation was performed in Vero hSLAM cells. RESULTS: Of the 98 suspected measles outbreaks, 61 were confirmed as measles, 12 as rubella and 21 confirmed as the mixed outbreaks. Four outbreaks remained unconfirmed. Of the 126 cases from the local hospitals, 91 were confirmed for measles and three for rubella. Overall, 93.6 per cent (383/409) confirmed measles cases were in the age group of 0-15 yr. Measles virus was detected in 18 of 38 specimens obtained from the suspected cases. Sequencing of PCR products revealed circulation of D4 (n=9) and D8 (n=9) strains. Four measles viruses (three D4 & one D8) were isolated. INTERPRETATION & CONCLUSIONS: Altogether, 94 measles and rubella outbreaks were confirmed in 2013 in the State of Maharasthra indicating the necessity to increase measles vaccine coverage in the State.

Complete genome sequence of mumps viruses isolated from patients with parotitis, pancreatitis and encephalitis in India.

Limited information is available regarding epidemiology of mumps in India. Mumps vaccine is not included in the Universal Immunization Program of India. The complete genome sequences of Indian mumps virus (MuV) isolates are not available, hence this study was performed. Five isolates from bilateral parotitis and pancreatitis patients from Maharashtra, a MuV isolate from unilateral parotitis patient from Tamil Nadu, and a MuV isolate from encephalitis patient from Uttar Pradesh were genotyped by the standard protocol of the World Health Organization and subsequently complete genomes were sequenced. Indian MuV genomes were compared with published MuV genomes, including reference genotypes and eight vaccine strains for the genetic differences. The SH gene analysis revealed that five MuV isolates belonged to genotype C and two belonged to genotype G strains. The percent nucleotide divergence (PND) was 1.1% amongst five MuV genotype C strains and 2.2% amongst two MuV genotype G strains. A comparison with widely used mumps Jeryl Lynn vaccine strain revealed that Indian mumps isolates had 54, 54, 53, 49, 49, 38, and 49 amino acid substitutions in Chennai-2012, Kushinagar-2013, Pune-2008, Osmanabad-2012a, Osmanabad-2012b, Pune-1986 and Pune-2012, respectively. This study reports the complete genome sequences of Indian MuV strains obtained in years 1986, 2008, 2012 and 2013 that may be useful for further studies in India and globally.

Circulation of two mumps virus genotypes in an unimmunized population in India.

Two separate outbreaks of fever with parotitis were reported from the Apsinga and Pimpla villages in the Osmanabad district of the Maharashtra State, India during February and March 2012. Meningo-encephalitis was noted in two patients resulting in the death of an 11-year male. Samples of blood and throat swabs were collected from patients with fever and parotitis. Serum samples from suspected (n=62) and convalescent (n=19) patients were tested for mumps virus specific IgM and/or IgG antibodies. Mumps virus specific IgM antibodies were detected in 44 of 62 serum samples (71%). Of the 19 convalescent phase sera 16 had both, anti-mumps virus IgM and IgG antibodies. Twenty-eight throat swabs collected from patients with parotitis were tested by RT-PCR for the SH gene. Twenty-three specimens were found to be positive and nucleotide sequencing of the amplified PCR products revealed circulation of two distinct genotypes that were village specific. Mumps virus genotype C (n=18) was detected in Apsinga village and genotype G (n=5) in Pimpla village. Two mumps virus isolates were also obtained using Vero cells. This is the first report from India confirming simultaneous circulation of mumps virus genotype C in one village and the G genotype in another village only 37 km away.

Molecular epidemiology of measles in India, 2005-2010.

Measles is a childhood disease that causes great morbidity and mortality in India and worldwide. Because measles surveillance in India is in its infancy, there is a paucity of countrywide data on circulating Measles virus genotypes. This study was conducted in 21 of 28 States and 2 of 7 Union Territories of India by MeaslesNetIndia, a national network of 27 centers and sentinel practitioners. MeaslesNetIndia investigated 52 measles outbreaks in geographically representative areas from 2005 through June 2010. All outbreaks were serologically confirmed by detection of antimeasles virus immunoglobulin M (IgM) antibodies in serum or oral fluid samples. Molecular studies, using World Health Organization (WHO)-recommended protocols obtained 203 N-gene, 40 H-gene, and 4 M-gene sequences during this period. Measles genotypes D4, D7, and D8 were found to be circulating in various parts of India during the study period. Further phylogenetic analysis revealed 4 lineages of Indian D8 genotypes: D8a, D8b, D8c, and D8d. This study generated a large, countrywide sequence database that can form the baseline for future molecular studies on measles virus transmission pathways in India. This study has created support and capabilities for countrywide measles molecular surveillance that must be carried forward.