Kavita Arunagiri

Dengue epidemiology in Thanjavur and Trichy district, Tamilnadu - Jan 2011-Dec 2011

BACKGROUND Dengue infection is emerging as a serious public health problem in Tamil Nadu. An enhanced surveillance system can generate information on the epidemiology of the disease, which is essential for planning and development of relevant control/preventive measures against Dengue. MATERIALS AND METHODS A prospective descriptive study was undertaken between January 2011 to December 2011, by testing suspected Dengue patients attending Thanjavur Medical College and Trichy Hospital (TMCH, a major Government referral hospital in Thanjavur District, Tamil Nadu, India) to define the magnitude of Dengue burden, the natural history of this disease in terms of clinical presentation and outcome of the infections in hospitalized Dengue patients. The sera collected from suspected patients were analyzed for Dengue specific IgM and IgG antibodies by IgM antibody capture enzyme linked immunosorbent assay (ELISA) using NIV kit and IgGPanBio Duo Rapid Immunochromatographic Card Test (Brisbane, Australia). The clinical case definition by World Health Organization was adopted to categorize the Dengue cases. RESULTS The total number of samples screened during the period was 200, out of which 79 (39.5%) were positive for IgM and IgG antibodies and 65 (32%) for IgM antibodies only. By clinical evaluation, Dengue fever was diagnosed in 43 patients, 18 had hemorrhagic manifestations and four patients had progressed to DSS. Though (DSS+DHF) was present in 22 patients, all of them recovered well. CONCLUSION In developing countries like India, building of laboratory with advanced capacity for diagnosis and combat-mode ready preparedness for the management of Dengue cases in emergency situation may reduce Dengue-related mortality.

Development and evaluation of flavi-immunoglobulinM capture enzyme-linked immunosorbent assay

In this study, we report the evaluation of In-house flavi virus immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA), which can be used as a screening test to determine the infecting flavivirus serotype over the current serological methods. A panel of 88 sera (inclusive of well characterized dengue, Japanese Encephalitis (JE) and West Nile virus (WNV) positive and negative samples tested and confirmed by commercial kit) was used for evaluation of the kit. The sensitivity and specificity of the In-house capture assay versus the commercial kit for the sero-diagnosis of dengue was 100% and 87% respectively, for JE IgM, it was found to be 90% and 100% respectively, and for West Nile it was 87.5% and 90.9%. Based on the study, we concluded that this flavivirus-serotyping ELISA provides rapid results and may be used as an accurate alternate to other serological tests for the specific diagnosis of flavivirus infections.

Genetic analysis of HA gene of pandemic H1N1 2009 influenza viruses circulating in India.

The H1N1 2009 influenza pandemic took the health care workers by surprise in spite of warning about influenza pandemic. Influenza A virus has the ability to overcome immunity from previous infections through the acquisition of genetic changes by shift or drift. Thus, understanding the evolution of the viruses in human is important for the surveillance and the selection of vaccine strains. A total of 23 pandemic A/H1N1 2009 viral HA gene sequences were downloaded from NCBI submitted during March and May 2010 by NIV and were analysed. Along with that the vaccine strain A/California/07/2009 was also downloaded from NCBI. All the sequences were used to analyse the evolution of the haemagglutinin (HA) by phylogenetic analysis. The HA gene could be divided into four groups with shift from 1 to lV revealing that the HA genes of the influenza A viruses evolved in a sequential way, in comparison to vaccine strain A/California/07/2009. Amino acid sequence analysis of the HA genes of the A/H1N1 2009 isolates, revealed mutations at positions 100, 220 and additional mutations in different positions 114, 171, 179, 190, 208, 219, 222, 239, 240, 247, 251, 260 and 285 .The mutations identified showed the adaptation of the new virus to the host that could lead to genetic changes inherent to the virus resulting in a reassortant which could be catastrophic, hence continuous monitoring of strains is mandatory.

Prevalence and Molecular Characterization of Circulating Respiratory Syncytial Virus (RSV) in Chennai, South India during 2011-2014

The objective of the study was to determine the prevalence and genotyping of RSV A and B in Chennai, Tamilnadu, south India. Human Respiratory Syncytial Virus (RSV) is one of the most important respiratory viruses causing acute respiratory tract infections amongst children. Based on genotyping of the attachment glycoprotein (G) gene, it is divided into two groups, RSV-A and RSV-B. We tested 850 samples from patients with influenza like illness (ILI) and severe respiratory illness (SARI) during the period 2011-2014 for RSV using a conventional RT-PCR and found 124 (14.5%) samples to be positive for RSV. Further sub typing by nested PCR in which 84(10%) were RSV A, 40 (4.5%) were RSV B, suggesting that RSV-A was the predominant group circulating in south India during the study period. Among patients with RSV infection, is 47.5% were in less than 1 year age group, 29.8 % were between 1 to 5 year age group, 14.5% were between 6 to 14 years age group and 8 % were above 14 years age. Phylogenetic analysis all RSV-A sequences belonged to ON1 within NA1 genotype and is ON1 with in NA1 genotype and RSV B sequences belonged to the genotype BA9 and BA12.

A novel indirect ELISA for diagnosis of dengue fever.

Background & objectives: Dengue fever (DF) is associated with significant morbidity and mortality in the tropical and sub-tropical regions of the world. Since there are no effective antiviral drugs for treatment, clinicians often rely on the accurate diagnosis of dengue fever to begin supportive therapy at early stages of the illness. The objective of this study was to develop an in-house dengue virus serotype 2 (DENV-2) non-structural protein- 5 (NS5) based indirect ELISA. Methods: DENV-2 was raised in Vero cells and the viral proteins were separated and subsequently the NS5 protein was eluted. Serum samples from primary and secondary dengue fever patients; and acute and convalescent samples from Japanese encephalitis (JE) and West Nile virus (WNV) cases were used to validate the ELISA. Results: The assay was found to be 100 per cent specific in detecting DENV-2 specific antibodies from patients serum. However, in terms of sensitivity, the assay could detect IgM antibodies only from 90 per cent of the primary dengue samples. The IgM/IgG ratio of the primary and secondary samples was 7.24 and 0.64, respectively. Interpretation & conclusions: The results indicate that the DENV-2 NS5 ELISA is dengue group specific and can be used to differentiate dengue infection from other circulating Flavivirus infections. This NS5 ELISA can also be used to distinguish between primary and secondary dengue fever on the basis of IgM/IgG ratios. Further studies with larger sample sizes and different DENV serotypes are required to validate the ELISA.

Isolation & molecular characterization of human parainfluenza virus in Chennai, India

Background & objectives: Human parainfluenza virus (HPIV) accounts for a significant proportion of lower respiratory tract infections in children as well as adults. This study was done to detect the presence of different subtypes of HPIV from patients having influenza like illness (ILI). Methods: Throat and nasal swabs from 232 patients with ILI who were negative for influenza viruses were tested by multiplex reverse transcription polymerase chain reaction(mRT-PCR) for the detection of human parainfluenza virus. All samples were inoculated in rhesus monkey kidney (LLC-MK2) cell line. Results: Of the 232 samples, 26(11.2%) were positive by mRT-PCR and nine (34.6%) showed cytopathic effect with syncytium formation for HPIV and all were HPIV-3 serotype, other serotypes like 1,2,4 were negative. The HPIV-3 strains (HN gene) were sequenced and analysed. Two novel mutations were identified at amino acid residues 295 and 297. Interpretation & conclusions: The mRT-PCR assay offers a rapid, sensitive and accurate diagnostic method for detection of HPIV which enables early detection and control. In our study there was a predominance of HPIV among 1-5 yr age group and the school going age group was less affected. Further studies need to be done to characterize HPIV isolated from different parts of the country.

Pandemic H1N1 2009 among Pediatric Age Group in Tamilnadu, June 2009 - Aug 2010

Influenza A viruses causes recurrent outbreaks on local or global scale with potentially severe consequences on human health and the global economy. The new strain of Influenza A virus - H1N1 2009 had caused pandemic disease among human, probably owing to little or no preexisting immunity to the new strain. This is a retrospective analysis of the impact of H1N12009 on the pediatric population of Tamil Nadu during the pandemic period of June 2009- August 2010. Throat and nasal swabs were taken from the suspected cases admitted in pediatric wards and intensive care units (ICUs) and were subjected to Real time RT PCR. A total of 6245 suspected pediatric cases were screened, of which 787 (12.60%) were found to be positive for H1N12009. A majority of cases belonged to the 6–12 age group (35.58%). Male children were more affected than female children. Despite a fall in the number of positives in 2010, there is a concern about the probability of a new reasssortment of H1N1 2009 with other viruses of either human or animal hosts during the next season that could result in a potentially pathogenic strain.