Human metapneumovirus infection in haematopoietic stem cell transplant recipients: a case series.

Human metapneumovirus (hMPV) is an enveloped virus that causes serious respiratory tract infection among immunocompromised populations especially haematopoietic stem cell transplant (HSCT) recipients. Here, we describe 3 cases of hMPV infection which led to mortality among post HSCT adults. 66 post HSCT adults enrolled between January 2017 and March 2019 at Dr. B. R. Ambedkar Institute Rotary Cancer Hospital, AIIMS, New Delhi, were followed up for a period varying from 16 days to 18 months for any episode of respiratory illness until March 2019. Real time reverse transcriptase polymerase chain reaction (rRT-PCR) was used to detect the virus from appropriate specimens when symptoms of acute respiratory infection appeared. Samples from 88 out of a total of 172 episodes of suspected acute respiratory infection could be tested by rRT-PCR. Of these, 9 episodes were positive for hMPV. Three patients with hMPV associated lower respiratory tract infection (LRTI) expired within 30 days of HSCT. The possible risk factors associated with mortality included LRTI, infection during early post-transplant period (first week following HSCT), absolute lymphocyte count less than 200/µl, absolute neutrophil count less than 500/µl, use of steroid within 30 days prior to infection and need for mechanical ventilation.

In-silico Designing and Testing of Primers for Sanger Genome Sequencing of Dengue Virus Types of Asian Origin.

Rarity in reporting whole genome sequence of Dengue virus from dengue endemic countries leaves lacunae in understanding regional pattern of virus mutation and ultimately leading to non-understanding of transmission pattern and clinical outcomes emerging at regional levels. Due to inter-serotype genomic similarity and intra-serotype genomic diversity, appropriate designing of primer pairs appears as an exhaustive exercise. Present paper reports new Dengue virus type-specific primer which may help in characterizing virus specific to Asian origin. Genomes of dengue virus serotypes of Asian region were searched and using advanced bioinformatics tools, serotype specific primers were designed and tested for their targeted amplification efficiency. 19 primers sets for DENV-1, 18 primer sets for DENV-2, 17 for DENV-3 and 18 for DENV-4 were designed. In-silico and experimental testing of the designed primers were performed on virus isolated from both clinical isolates and passaged cultures. While all 17 and 18 primer sets of DENV-3 and DENV-2 respectively yielded good quality sequencing results; in case of DENV-4, 16 out of 18 primer sets and in DENV-1, 16 out of 19 primer sets yielded good results. Average sequencing read length was 382 bases and around 82% nucleotide bases were Phred quality QV20 bases (representing an accuracy of circa one miscall every 100 bases) or higher. Results also highlighted importance of use of primer development algorithm and identified genomic regions which are conservative, yet specific for developing primers to achieve efficiency and specificity during experiments.

Age of initial cohort of dengue patients could explain the origin of disease outbreak in a setting: a case control study in Rajasthan, India.

Dengue fever (DF) and dengue hemorrhagic fever (DHF) is a public health problem with 390 million cases reported in world annually. In Rajasthan, DF with DHF is being reported for about two decades. For undertaking interventions into disease transmission, locating origin of transmission is very important. Present paper reports retrospective analysis of the hospital reported cases of dengue during the year 2013-2014 undertaken in Barmer, Rajasthan. To address task of investigating outbreak, detailed analysis of the data on serological test results (Mac-ELISA assay of NS1, IgG and IgM) performed by local hospital, Balotra was made. The domestic breeding containers were examined for the presence of larvae and adult forms of Aedes aegypti by visiting individual households as well as common places of human aggregation like schools and hospitals. The analysis showed that first dengue cases started from the lot of school going children and then followed by adults and finally during peak period of infection only children around 1-2 years got infected. The subsequent entomological investigations during the outbreak showed school as principal source of mosquito breeding. Present investigations highlight that schools (March to April) play the role of primary sites of disease transmission and should be preferred for undertaking vector control operations to prevent dengue transmission from getting aggravated.

Rare occurrence of natural transovarial transmission of dengue virus and elimination of infected foci as a possible intervention method.

Transovarial transmission of dengue virus has been studied in 33 districts of Rajasthan, India. Small proportion (1.09%) of breeding containers positive for the virus and their elimination has been demonstrated as a possible intervention method of disease control. Dengue virus was isolated from individual mosquitoes employing Indirect Fluorescence Antibody Test and Reverse Transcriptase Polymerase Chain Reaction. Out of 1,30,525 containers examined only 1432(1.09%) showed transovarially transmitted virus activity. Elimination of larvae from all the 1432 virus positive containers resulted in substantial control over prospective transmission of dengue. The study highlights rarity of transovarial transmission under natural conditions and sensitizes whether elimination of vertically infected foci could be used as a new intervention method.

Multiple dengue virus types harbored by individual mosquitoes.

The existing knowledge on pathogenesis and aetiology of DHF establishes that Dengue Hemorrhagic Fever (DHF) and Dengue Shock Syndrome (DSS) are caused by two subsequent infections of two different serotypes of dengue affecting a common human population with a time gap. Present studies have been undertaken on 212 laboratory reared infected individual mosquitoes from larvae collected from 31 dengue endemic towns of Rajasthan, India. Type specific DEN viruses were detected from individual mosquitoes employing RT-PCR. In 78.7% of 212 infected individual mosquitoes studied, vertically transmitted multiple DENV types were observed. We report for the first time that single mosquitoes contain multiple dengue virus types.

Distribution of dengue virus types in Aedes aegypti in dengue endemic districts of Rajasthan, India.

BACKGROUND & OBJECTIVE: Dengue haemorrhagic fever (DHF) is the major cause of sustained morbidity/ mortality among human cases of dengue in dengue endemic areas of Rajasthan. Screening of mosquitoes collected from disease endemic settings and typing the virus could provide significant epidemiological information for prospective risk of DHF. We therefore carried out a study on different dengue virus types as occurring in field collected Aedes aegypti mosquitoes from four dengue endemic districts of Rajasthan, India. METHODS: Adult Ae. aegypti were collected from the human dwellings of urban, peri-urban and rural settings of four dengue endemic districts of Rajasthan, India. The field collected adults were fed on 4 per cent glucose solution and kept in the laboratory for 3-4 days. The adult field collected Ae. aegypti, were subjected to indirect fluorescence antibody test (IFAT) following standard procedure. Commercially acquired monoclonal antibodies against DEN types 1, 2, 3 and 4 were used. The remnants of IFA test subjected mosquitoes were made into viral suspension which was inoculated into the cell culture medium and mouse brain to confirm the presence of virus as shown by IFA test. RESULTS: Of the 498 adult Ae. aegypti tested, 78 (15.6%) were positive by IFA test. Among urban areas, desert area (Jodhpur) showed highest (21.6%) mosquito infectivity followed by 7.1 per cent in forest and river area (Kota) and least (3.2%) in semi-arid area (Jaipur). Among rural settings also, desert area showed maximum (25.0%) natural infection in mosquitoes followed by rural setting-1 of semi-arid area (24.1%). Among urban setting of desert area, all the four dengue types viz., DEN-1, 2, 3 and 4 were detected. In semi-arid area, urban settings showed presence of DEN-3, whereas among rural settings, rural-1 showed all the four DEN types, rural-2 showed DEN-1 and DEN-3, rural-4 showed DEN-3 and DEN-4, and rural-3 showed no mosquito infections. In forest and river area, among urban settings only, three DEN types, 1, 2 and 4 were observed. INTERPRETATION & CONCLUSION: In desert and semi-arid areas of Rajasthan, where people possess tendency of over- and sustained storage of domestic water, present observations on occurrence of all four dengue virus types may have important bearing on the epidemiology of DHF in the area.

Entomological studies for surveillance and prevention of dengue in arid and semi-arid districts of Rajasthan, India.

BACKGROUND & OBJECTIVES: Rajasthan is one of the dengue endemic states of India. Very few studies have been published on entomological aspects of dengue in this state. Owing to water scarcity, inhabitants in desert areas overstore domestic water which leads to the persistence of dengue vectors within the domestic premises. Area specific knowledge on breeding, key containers and seasonal rhythms of vector population is essential for preparing an effective prevention plan against dengue. Present paper reports results of entomological investigations on dengue vectors in arid and semi-arid districts of Rajasthan. METHODS: Longitudinal studies were undertaken during 2004-06 in one arid and two semi-arid dengue endemic districts of Rajasthan. Adult and larval Aedes were collected from the randomly selected houses in representative towns and villages with associated details of container types and water storage practices of inhabitants. RESULTS: In urban areas during all the seasons adult house index (AHI) of Aedes aegypti was maximum in desert zone (25) and least in semi-arid area with saline river III (1). The difference of AHI during three seasons was statistically significant (chi2 = 16.1, p < 0.01 for urban; and chi2 = 50.71, p < 0.001 for rural). Breeding of Ae. aegypti among urban settings was maximum in desert zone. During all the seasons cement tanks were the key breeding habitats for Ae. aegypti in desert as well as semi-arid areas. INTERPRETATION & CONCLUSION: Water storage habits during summer season emerged to be the risk factor of vector abundance in urban areas of arid and semi-arid settings. A carefully designed study of key containers targeting cement tanks as the primary habitats of mosquito control may lead to commendable results for dengue prevention.

Distribution and seasonality of vertically transmitted dengue viruses in Aedes mosquitoes in arid and semi-arid areas of Rajasthan, India.

BACKGROUND & OBJECTIVES: Transovarial transmission of dengue virus is a crucial etiological phenomenon responsible for persistence of virus during inter-epidemic period of the disease. Distribution and seasonality of this phenomenon in disease endemic areas may contribute to explain emergence of dengue and its subsequent prevention. The study on seasonal and area distribution of transovarial transmission of dengue virus in Aedes aegypti, Ae. albopictus and Ae. vittatus has been made in desert and non-desert districts of Rajasthan, India from 2006 to 2007. The observations revealed role of different Aedes species in transmission and retention of dengue virus. METHODS: The larvae of Ae. aegypti, Ae. albopictus and Ae. vittatus were collected during each of the study seasons from rural and urban areas of three districts-Jodhpur, Jaipur and Kota. The larvae were collected from domestic and peri-domestic containers and from tree holes of peri-urban foci such as gardens and parks and were reared into adults in the laboratory at room temperature. The laboratory reared adults were subjected to Indirect fluorescence antibody test (IFAT). The laboratory-reared adult mosquitoes showing positive IFA were treated as the sample showing vertically transmitted dengue virus. RESULTS: Pooled data for all the four seasons revealed maximum (15.7%) mosquito infectivity in Ae. albopictus followed by Ae. aegypti (12.6%) in Jodhpur district. In Jaipur district, Ae. vittatus showed highest infection (20%) of vertically transmitted virus followed by Ae. albopictus (18.7%) and least in Ae. aegypti (13.3%). In Kota district, pooled data for all the four seasons showed maximum vertical infection of mosquitoes in Ae. albopictus (14.2%). INTERPRETATION & CONCLUSION: Transovarial transmission of dengue virus by available vector species in a dengue endemic setting could be the key etiological phenomenon responsible for re-emergence of the disease from inter-epidemic to epidemic phase of disease onset. The observations in the present study suggest that during winter season which is not the active transmission season of dengue in Rajasthan, Ae. albopictus has shown maximum percentage of vertically transmitted virus. Our observation substantiates with the earlier studies that how Ae. albopictus is horbouring virus during inter-epidemic period of dengue. Another important lead emerging through present study is the high mosquito infectivity of Ae. aegypti during summer and rainy seasons especially from desert districts, Jodhpur and semi district. This observation suggests that in Rajasthan, owing to tendency of overstorage of domestic water by the inhabitants, mosquito and vertically transmitted virus get pronounced during summer season which could precedes the active transmission season of dengue during following rainy season.