Veena Mittal

Chikungunya fever: a re-emerging viral infection.

Chikungunya (CHIK) fever is a re-emerging viral disease characterized by abrupt onset of fever with severe arthralgia followed by constitutional symptoms and rash lasting for 1-7 days. The disease is almost self-limiting and rarely fatal. Chikungunya virus (CHIKV) is a RNA virus belonging to family Togaviridae, genus Alphavirus. Molecular characterization has demonstrated two distinct lineages of strains which cause epidemics in Africa and Asia. These geographical genotypes exhibit differences in the transmission cycles. In contrast to Africa where sylvatic cycle is maintained between monkeys and wild mosquitoes, in Asia the cycle continues between humans and the Aedes aegypti mosquito. CHIKV is known to cause epidemics after a period of quiescence. The first recorded epidemic occurred in Tanzania in 1952-1953. In Asia, CHIK activity was documented since its isolation in Bangkok, Thailand in 1958. Virus transmission continued till 1964. After hiatus, the virus activity re-appeared in the mid-1970s and declined by 1976. In India, well-documented outbreaks occurred in 1963 and 1964 in Kolkata and southern India, respectively. Thereafter, a small outbreak of CHIK was reported from Sholapur district, Maharashtra in 1973. CHIKV emerged in the islands of South West Indian Ocean viz. French island of La Reunion, Mayotee, Mauritius and Seychelles which are reporting the outbreak since February, 2005. After quiescence of about three decades, CHIKV re-emerged in India in the states of Andhra Pradesh, Karnataka, Maharashtra, Madhya Pradesh and Tamil Nadu since December, 2005. Cases have also been reported from Rajasthan, Gujarat and Kerala. The outbreak is still continuing. National Institute of Communicable Diseases has conducted epidemiological, entomological and laboratory investigations for confirmation of the outbreak. These have been discussed in detail along with the major challenges that the country faced during the current outbreak.

Phylogenetic studies reveal existence of multiple lineages of a single genotype of DENV-1 (genotype III) in India during 1956–2007

BackgroundDengue virus type 1 (DENV-1) have been mostly circulating silently with dominant serotypes DENV-2 and DENV-3 in India. However recent times have marked an increase in DENV-1 circulation in yearly outbreaks. Many studies have not been carried out on this virus type, leaving a lacunae pertaining to the circulating genotypes, since its earliest report in India. In the present study, we sequenced CprM gene junction of 13 DENV-1 isolated from Delhi and Gwalior (North India) between 2001–2007 and one 1956 Vellore isolate as reference. For comparison, we retrieved 11 other Indian and 70 global reference sequences from NCBI database, making sure that Indian and global isolates from all decades are available for comparative analysis.ResultsThe region was found to be AT rich with no insertion or deletion. Majority of the nucleotide substitutions were silent, except 3 non-conservative amino acid changes (I → T, A → T and L → S at amino acid positions 59,114 and 155 respectively) in the Indian DENV-1 sequences, sequenced in this study. Except two 1997–98 Delhi isolates, which group in genotype I; all other Indian isolates group in genotype III. All Indian genotype III DENV-1 exhibited diversity among them, giving rise to at least 4 distinct lineages (India 1–4) showing proximity to isolates from diverse geographic locations.ConclusionThe extensive phylogenetic analysis revealed consistent existence of multiple lineages of DENV-1 genotype III during the last 5 decades in India.

Emergence of an independent lineage of dengue virus type 1 (DENV-1) and its co-circulation with predominant DENV-3 during the 2006 dengue fever outbreak in Delhi

OBJECTIVES The sudden emergence of dengue virus type 1 (DENV-1) and its co-circulation with predominant DENV-3 was the hallmark of the 2006 dengue fever outbreak in Delhi. Viruses that circulated between 1996 and 2005 in the City have been well characterized, but the genomic diversity in 2006 strains is not known. The present study was undertaken to reveal the emerging molecular genotype(s) and evolutionary trend of the viruses responsible for the dengue fever outbreak in Delhi during 2006. STUDY DESIGN The CprM gene junction of the DENV isolates from the 2006 Delhi dengue fever outbreak were subjected to nucleotide sequencing. Comparative phylogenetic analysis was done using DENV-1 and DENV-3 sequences retrieved from the global database. RESULTS Multiple sequence alignment revealed only substitutions, with no insertions or deletions. A dendrogram indicated emergence of a distinct lineage of DENV-1 (having similarity with the Comoros/Singapore 1993 and Delhi 1982 strains, but quite different from the Delhi 2005 lineage) and microevolution of the pre-circulating DENV-3. These findings point towards the circulation of two independent lineages of DENV-1 in Delhi during 2005 and 2006. CONCLUSIONS It is feared that the introduction of an independent lineage of the outbreak-associated strain of DENV-1 and its co-circulation with the deeply-rooted strain of DENV-3 in Delhi may result in yet another, possibly more severe outbreak in the near future.

Continued persistence of a single genotype of dengue virus type-3 (DENV-3) in Delhi, India since its re-emergence over the last decade.

BACKGROUND/PURPOSE The re-emergence of an epidemic strain of dengue virus type-3 (DENV-3) in Delhi in 2003 and its persistence in subsequent years marked a changing trend in dengue virus circulation in this part of India. Its evolving phylogeny over the past decade has not been studied in detail as yet. METHODS Reverse transcription polymerase chain reaction and sequencing of the CprM gene junction of DENV-3 from different outbreaks since 2003 was carried out. Thirty CprM DENV-3 sequences from this study were compared with 46 other previously reported CprM DENV-3 sequences from India and other countries. Multiple sequence alignment and phylogenetic trees were constructed to determine the extent of genetic heterogeneity and trace the phylogeny of DENV-3. RESULTS Thirty CprM DENV-3 sequences (Accession numbers AY706096-99, DQ645945-52, EU181201-14, and EU846234-36) were submitted to GenBank. The CprM junction was found to be AT rich (approximately 53%). Nucleotide sequence alignment revealed only nucleotide substitutions. Phylogenetic analysis indicated sustained evolution of a distinct Indian lineage of DENV-3 genotype III in Delhi. CONCLUSION Active circulation of DENV-3 genotype III over the last decade in Delhi was evident and worrying. This genotype has been implicated in several outbreaks in South-East Asia and other parts of the world.

Current research and clinical trials for a vaccine against Chikungunya virus

Correspondence: Mala Chhabra National Centre for Disease Control, 22-Sham Nath Marg, New Delhi 110054, India Tel +91 11 2390 9236 Fax +91 11 2392 2677 Email malachhabra@yahoo.co.in Abstract: Chikungunya infection is a self-limiting Aedes mosquito-borne arboviral disease with variable clinical manifestations, ranging from asymptomatic illness to a very severe and crippling arthralgia. Until recently, Chikungunya was a little known disease that re-emerged in 2005–2006, leading to major outbreaks on the Indian Ocean Islands and in South East Asia, and eventually extending its range to temperate regions. It drew global attention due to its explosive onset, extensive geographic distribution, and high morbidity. Since re-emergence, an estimated one million symptomatic cases with 0.1% fatality per year have been reported globally. A lack of herd immunity, vector control, and globalization and trade are clearly a problem in the spread of this disease. The Chikungunya virus (CHIKV) has also acquired biologically important mutations during its evolution, increasing its geographic reach. This disease has resulted in a loss of productivity in affected communities. The absence of a vaccine or an effective antiviral therapy makes dealing with this disease challenging for those involved in public health. There is an emergent need for an effective vaccine against CHIKV infection. The candidates that have been tested include attenuated living, nonliving and genetically engineered vaccines. Several of these vaccine candidates are in preclinical and clinical trials. This review outlines the current knowledge about chikungunya infection and vaccine development.

Use of multilocus microsatellite typing (MLMT) for the genetic analysis of Indian isolates of Leishmania donovani

In India, the transmission of the Leishmania donovani that causes human visceral leishmaniasis (VL) or kala-azar is thought to be entirely anthroponotic, with cases of postkala-azar dermal leishmaniasis (PKDL) believed to be the sole source of the parasites carried to humans by sandflies (Thakur and Kumar, 1992). The incidence of VL and the number of current cases of the disease world-wide are estimated at 0.5 million cases/year and 2.5 million, respectively, with 90% of all cases occurring in just five countries: Bangladesh, Brazil, Nepal, India and Sudan (Bora, 1999). In India, the disease is endemic in the eastern parts of the country, primarily in the state of Bihar, where .90% of all Indian cases occur. The parasites in the genus Leishmania have been classified into different species primarily on the basis of clinical, biological, geographical and epidemiological criteria. Currently, the most comprehensive method for identification of the species and strains is by multilocus iso-enzyme electrophoresis (MLEE). This technique, which allows various zymodemes to be distinguished, is, however, too slow, laborious and expensive for high-throughput analyses. Moreover, it is not easy to compare data-sets from the different laboratories in which MLEE-based analyses are performed. Genotyping is a potential alternative to MLEE that offers greater discriminatory power. Standard PCR, PCR–RFLP and sequencing allow interand intra-specific variations in polymorphic and multicopy DNA sequences (such as ‘microsatellites’) to be explored. Microsatellites are tandem repeat stretches of short nucleotide sequences (of just one to six base pairs) that show high variability, mainly as the result of variation in the length of the allelic repeats (Ellegren, 2000). Highly polymorphic and co-dominant microsatellite markers have proven to be very useful for studies on population dynamics, and, over the last decade, such markers have been developed for the parasites in the L. donovani complex (Bulle et al., 2002; Jamjoom et al., 2002; Ochsenreither et al., 2006; Kuhls et al., 2007). The results of a recent study based on multilocus microsatellite typing (MLMT) revealed considerable genetic homogeneity among L. donovani isolates from the Indian subcontinent (Alam et al., 2009). For the present study, MLMT based on six microsatellite loci was used to explore the genetic variation among 93 L. donovani isolates from Bihar state (82 from VL cases and 11 from PKDL cases, collected between 2004 and 2008). Although the primer sets used appear to be specific to L donovani/ L. infantum (Rossi et al., 1994; Jamjoom et al., 2002), the test isolates were compared against reference strains of L. major (two) and L. tropica (three) as well as reference strains of L donovani/L. infantum [L. donovani collected in India (four), Kenya (one) and Ethiopia (two), and L. infantum collected in France (two), Portugal (one) and Brazil (one)].