Status of insecticide resistance in the dengue vector Aedes aegypti in India: A review.

The study aims to present an overview of the extent of insecticide resistance in the dengue vector Aedes aegypti in India. Published data on insecticide resistance in this species were systematically searched through online databases like PubMed, Google, and Google Scholar. From each study, data were extracted and analyzed to understand the spatial and temporal patterns. Particular emphasis was given to the commonly used insecticides for mosquito control. Forty-three studies met the inclusion criteria, of which 13 had adult bioassay data, 13 larval bioassay data and 17 contained both. Data demonstrated high resistance to DDT and resistance is also widespread against carbamates. There is mounting evidence of increased tolerance to pyrethroids and organophosphorus compounds, viz permethrin, deltamethrin, lambda-cyhalothrin, malathion, and temephos. The emergence of resistance to all the insecticide classes further justifies the need for annual resistance monitoring and to maintain a nationwide database that can provide a reference for formulating effective control strategies.

Morphological and molecular characterization of Aedes aegypti variant collected from Tamil Nadu, India.

BACKGROUND & OBJECTIVES: Accurate mosquito species identification is the basis of entomological surveys and effective vector control. Mosquito identification is either done morphologically using diagnostic features mentioned in taxonomic keys or by molecular methods using cytochrome oxidase subunit 1 (coxI) and Internal transcribed spacer 2 (ITS2). METHODS: We performed a larval survey for Aedes mosquitoes from eight different geographical regions in Tamil Nadu, India. The mosquitoes collected during the survey were characterized using both morphological and molecular markers. RESULTS: During an entomological survey from eight different geographical regions in Southern India, a morphological variety named Aedes aegypti var. luciensis was observed. The variant mosquitoes were characterized using both morphological and molecular markers. The variant mosquitoes differed only in the dark scaling of 5th segment of hind-tarsi. Around one third to two third of the 5th segment in variant mosquitoes was dark which has been described as white in identification keys. No other significant difference was observed in adults or immature stages. The variation was heritable and coexisting in the field with the type form mosquitoes. Comparison of the genetic profile of coxI and ITS2 were similar in variant and the type form indicating both of them to be conspecific. INTERPRETATION & CONCLUSION: The morphological variant mosquitoes were found genetically similar to the Ae. aegypti type form. However, considering its high prevalence and coexistence with Ae. aegypti type form in different geographical regions, detailed studies on bionomics, ecology, genetics, behavior as well as its plausible role in disease transmission are warranted.

Changing paradigm in the epidemiology of Japanese encephalitis in a non-endemic region.

BACKGROUND & OBJECTIVES: Japanese encephalitis (JE) is a mosquito-borne zoonotic disease. The JE virus (JEV) does not cause any disease among its natural hosts and transmission continues through mosquitoes belonging to Culex vishnui subgroup. This study was aimed to investigate the prevalence of JEV in mosquitoes and humans in the Thanjavur district, a non-endemic region for JE, in Tamil Nadu, by using standard available assays. METHODS: : A sero-surveillance study was conducted in Thanjavur district among the normal rural school children in the 5-12 yr age group, during the JE season (October) and post-JE season (February) from 2011 to 2013 for the detection of JEV infection. Vector abundance studies were carried out from 2011 to 2014. JE seropositivity and its association between the seasons were analysed statistically. RESULTS: : The occurrence of JE infection among children aged 5-12 yr was very high in the study area. The infection rates for JE in two consecutive seasons for 2011-12 and 2012-13 were 32.2 and 65.2%, respectively. The Cx. tritaeniorhynchus sp. dominated the catch, and was majorly responsible for the transmission. There was a significant difference in the human infection rate compared to the years 1991-92 and 1992-93; and a marked decrease in the cattle to pigs ratio (123 : 1) compared to the studies in 1991-93. INTERPRETATION & CONCLUSION: : The study unearthed the prevailing situation of JE among children, who are at higher risk of developing the disease during the transmission season. The decrease in the cattle to pigs ratio might be the one of the reasons for increase in the JEV infection among the children population compared to 20 years before. This trend requires urgent attention as it could be prevented with effective surveillance systems and vaccines.

Entomological investigations into an epidemic of Japanese encephalitis (JE) in northern districts of West Bengal, India (2011-2012).

BACKGROUND & OBJECTIVES: Japanese encephalitis (JE) is one of the most important arboviral diseases of human beings with outbreaks in many parts of Southeast Asia including India. We present the entomological findings of an outbreak occurred in northern part of West Bengal during 2011-2012 with special emphasis on the role of JE vectors in different seasons. METHODS: Adult mosquito collections were made with the help of mouth aspirators, aided by flash lights during day time resting inside human and animal habitations as indoor, and resting outside field grasses, bushes, underneath of culverts and bridges as outdoor, and in and around the pig enclosures and cattle sheds during dusk period in JE affected villages from Cooch Behar, Dakshin Dinajpur, Darjeeling and Jalpaiguri districts in North West Bengal. In all study villages, a long handled with enamel bowl dipper was used to obtain immature stages of mosquitoes from various breeding habitats. RESULTS: A total of 19 different types of mosquito breeding habitats were examined for vectors of JE. From these habitats, 23.7 per cent were positive for breeding during the study period. Overall, nine different species were recorded through emergence, but none was positive for JE virus when subjected for detection of virus. Adult mosquitoes of more than 50 per cent of the potential JE vector species obtained through dusk and the rest through indoor and outdoor collections in all seasons. Altogether, 27 different species were recorded. Most of these were JE vectors. INTERPRETATION & CONCLUSIONS: Our results showed that in addition to Cx. vishnui subgroup, detection of JE virus antigen in Cx. quinquefasciatus indicated the possible maintenance of JE virus in nature through poor vector mosquitoes throughout the year. Since, all potential vector species reported elsewhere in India were also found in this region and fluctuated in density in different seasons, a proper integrated vector control programme needs to be implemented to control JE transmission.

Identification and prioritisation of potential vaccine candidates using subtractive proteomics and designing of a multi-epitope vaccine against Wuchereria bancrofti.

This study employed subtractive proteomics and immunoinformatics to analyze the Wuchereria bancrofti proteome and identify potential therapeutic targets, with a focus on designing a vaccine against the parasite species. A comprehensive bioinformatics analysis of the parasite's proteome identified 51 probable therapeutic targets, among which "Kunitz/bovine pancreatic trypsin inhibitor domain-containing protein" was identified as the most promising vaccine candidate. The candidate protein was used to design a multi-epitope vaccine, incorporating B-cell and T-cell epitopes identified through various tools. The vaccine construct underwent extensive analysis of its antigenic, physical, and chemical features, including the determination of secondary and tertiary structures. Docking and molecular dynamics simulations were performed with HLA alleles, Toll-like receptor 4 (TLR4), and TLR3 to assess its potential to elicit the human immune response. Immune simulation analysis confirmed the predicted vaccine's strong binding affinity with immunoglobulins, indicating its potential efficacy in generating an immune response. However, experimental validation and testing of this multi-epitope vaccine construct would be needed to assess its potential against W. bancrofti and even for a broader range of lymphatic filarial infections given the similarities between W. bancrofti and Brugia.

Detection of Wolbachia endobacteria in Culex quinquefasciatus by Gimenez staining and confirmation by PCR.

BACKGROUND & OBJECTIVES: Wolbachia are common intracellular bacteria that are found in arthropods and nematodes. These endosymbionts are transmitted vertically through host eggs and alter host biology in diverse ways, including the induction of reproductive manipulations, such as feminization, parthenogenesis, male killing and sperm-egg incompatibility. Since they can also move horizontally across species boundaries, Wolbachia is gaining importance in recent days as it could be used as a biological control agent to control vector mosquitoes or for paratransgenic approaches. However, the study of Wolbachia requires sophisticated techniques such as PCR and cell culture facilities which cannot be affordable for many laboratories where the diseases transmitted by arthropod vectors are common. Hence, it would be beneficial to develop a simple method to detect the presence of Wolbachia in arthropods. METHOD: In this study, we described a method of staining Wolbachia endobacteria, present in the reproductive tissues of mosquitoes. The reliability of this method was compared with Gram staining and PCR based detection. RESULTS: The microscopic observation of the Gimenez stained smear prepared from the teased ovary of wild caught and Wolbachia (+) Cx. quinquefasciatus revealed the presence of pink coloured pleomorphic cells of Wolbachia ranging from cocci, comma shaped cells to bacillus and chain forms. The ovaries of Wolbachia (-) cured mosquito did not show any cell. Although Gram's staining is a reliable differential staining for the other bacteria, the bacterial cells in the smears from the ovaries of wild caught mosquitoes did not take the stain properly and the cells were not clearly visible. The PCR amplified product from the pooled remains of wild caught and Wolbachia (+) Cx. quinquefasciatus showed clear banding, whereas, no banding was observed for the negative control (distilled water) and Wolbachia (-) Cx. quinquefasciatus. INTERPRETATION & CONCLUSION: The Gimenez staining technique applied, could be used to detect the members of the endobacteria Wolbachia easily, even in a simple laboratory without any special facilities or even in the field condition and for handling large number of samples in a shorter duration.

Genetic structure and connectivity among Aedes aegypti populations within Madurai city in Southern India.

We investigated the genetic variability and differentiation among 12 Ae. aegypti populations collected within the Madurai city in Tamil Nadu state of Southern India. Genotyping of 12 microsatellite markers in 353 individual samples showed moderate levels of genetic diversity among 12 populations. UPGMA tree, hierarchical clustering, Bayesian clustering and Discriminant Analysis on Principal Components roughly divided these populations into two genetic clusters: main city populations and the populations located at the border of the corporation limit. Significant positive correlation between genetic and geographic distance was observed among 12 populations, however, the correlation was non-significant within each genetic cluster. Population assignment and divMigrate graph depicted less migration between two groups. Overall, the findings of this study provided an overview of Ae. aegypti population structure within an urban setting in India that have implications in effective implementation of vector control in the city area.

Nation-wide vector surveillance on Zika and Dengue did not indicate transmission of the American lineage-pandemic Zika virus in India.

OBJECTIVES: Following the Public Health Emergency of International Concern declared on Zika by the World Health Organization during 2016, the Indian Council of Medical Research carried out nationwide vector surveillance for Zika and Dengue viruses (ZIKV and DENV) in India as a preparedness measure in 2016-19. METHODS: High-risk zones distributed to 49 Districts in 14 states/union territories were included in the study. Seven ICMR institutions participated, following a standard operating protocol. Aedes specimens sampled weekly were processed by multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) for ZIKV/DENV and random samples crosschecked with real-time RT-PCR for ZIKV. RESULTS: Altogether, 79 492 Aedes specimens in 6492 pools were processed; 3 (0.05%) and 63 (0.97%) pools, respectively, were found positive for ZIKV and DENV. ZIKV infections were recorded in Aedes aegypti sampled during the 2018 sporadic Zika outbreak in Jaipur, Rajasthan. However, these belonged to the Asian lineage of the virus, already circulating in the country. Both Ae. aegypti and Aedes albopictus distributed to 8 states/union territories were found to be infected with DENV. Both sexes of Ae. albopictus were infected, indicating transovarial transmission. CONCLUSION: This investigation evinced no active transmission of the American lineage-pandemic Zika virus in India during the pandemic period.

Prevalence of Ascogregarina spp. in the container breeding Aedes albopictus from Chikungunya fever affected areas of Kerala State, India.

The prevalence of protozoan, Ascogregarina sp. had been determined in the container breeding mosquitoes, Aedes albopictus. Since, the cyst of Ascogregarina might play role in the maintenance of the Chik virus during silent period, the presence of Ascogregarina has gained importance in recent days. The prevalence was found to be 71.62.

Longitudinal studies of Japanese encephalitis virus infection in vector mosquitoes in Kurnool district, Andhra Pradesh, South India.

A 4-yr (2002-2006) entomological study was carried out in Kurnool district, Andhra Pradesh state, south India, to identify the mosquito vectors of Japanese encephalitis virus (family Flaviviridae, genus Flavivirus, JEV). In total, 37,139 female mosquitoes belonging five genera and 18 species resting on vegetation were collected in villages and periurban areas at dusk. Mosquito species composition and pattern of JEV infection in mosquitoes varied in periurban and rural areas. In periurban area, Culex gelidus Theobald was abundant, making up 49.7% of total catch followed by Culex tritaeniorhynchus Giles (44.5%). In rural area, Cx. tritaeniorhynchus was predominant, making up 78.9% of total catch followed by Culex quinquefasciatus Say (10.8%), Anopheles subpictus Grassi (7.1%), and Cx. gelidus (1.1%). In light trap collections, Cx. gelidus and Cx. tritaeniorhynchus predominated in periurban and rural areas, respectively. Of 50,145 mosquitoes screened JEV isolations were made only from Cx. gelidus and Cx. tritaeniorhynchus. Based on high abundance and frequent JEV isolation, Cx. tritaeniorhynchus was found to be the principal vector in both areas, whereas Cx. gelidus plays a secondary vector role in periurban areas only.

Natural vertical transmission of dengue viruses by Aedes aegypti in Chennai, Tamil Nadu, India.

BACKGROUND & OBJECTIVE: Dengue viruses are spread and maintained in an Aedes aegypti-human- Ae. aegypti cycle in urban areas of the tropics. Dengue viruses are also maintained in nature by vertical transmission by Ae. aegypti. A study was undertaken in Chennai, a known endemic city in south India, to comprehend the natural vertical transmission dynamics in Ae. aegypti and to assess its epidemiological importance. METHODS: Ae. aegypti males collected in resting and landing collections were tested for dengue virus infection by antigen-capture enzyme-linked immunosorbent assay (ELISA) and further examined by insect bioassay, Toxorhynchites splendens inoculation-indirect immunofluorescence technique (Toxo-IFA) using serotype-specific monoclonal antibodies (Mabs), if found positive by ELISA. RESULTS: Of the 509 pools of Ae. aegypti males (n=5408) screened, 15 pools, collected in April, June- July, November-December in 2003 and March, May in 2004, were found positive for dengue virus infection and the minimum infection rate (MIR) among adult males was high in June 2003 (28.0/ 1000). Three positive pools could be serotyped as dengue-2 (2 pools) and dengue-3 (1 pool). INTERPRETATION & CONCLUSION: Dengue virus isolations from wild caught males of Ae. aegypti indicate the occurrence of transovarial transmission. Vertical transmission was mainly observed in summer months when dengue infections in humans were low suggesting that dengue viruses adopt a novel strategy of surviving adverse climatic conditions.

Emergence of dengue in the tribal pockets of Nilgiris district, Tamil Nadu.

As original tribal ways of living have morphed from a forest dweller existence, dengue is no longer an urban infection but is now also found in rural hilly areas. The spread of dengue is enhanced by the frequent movement of people to endemic areas where there is a vector mosquito presence. The impact of the virus is known to be great in the immunologically naive population. Our study reports on the threat of the dengue virus in these hilly areas.

Serological and entomological investigations of an outbreak of dengue fever in certain rural areas of Kanyakumari district, Tamil Nadu.

BACKGROUND AND OBJECTIVES: During the first week of July 2003, suspected cases of dengue fever were reported from three villages in Kanyakumari district in Tamil Nadu. Since the fever outbreak occurred for the first time in these villages, serological, virological and entomological investigations were carried out to confirm the aetiology of outbreak. METHODS: A total of 76 plasma samples were collected from suspected cases of dengue fever and screened for the presence of IgM antibodies by Pan Bio ELISA kit. Toxo-IFA system was used for the isolation of dengue virus from the plasma samples. Vector survey employing ovitraps and adult landing collection were carried out in the study villages. Pooled samples of Aedes mosquito were screened for dengue virus antigen by an in-house antigen capture ELISA test employing dengue virus specific monoclonal antibodies. RESULTS: Of the 76 samples tested, 15 (20%) were found positive for dengue virus specific IgM antibodies. Dengue virus serotype-3 was detected from a plasma sample by Toxo-IFA test using virus specific monoclonal antibodies. Entomological survey revealed the abundance of Aedes albopictus (Skuse) mosquitoes in the study area. One pool consisting of 12 Ae. albopictus males were found positive for dengue virus infection. INTERPRETATION AND CONCLUSION: Based on the IgM antibody capture ELISA results, it was evident that the current infection was caused by dengue virus in the affected areas. All the age groups were affected during this outbreak. Detection of dengue virus serotype-3 in plasma samples further confirmed the aetiology of this outbreak. The high prevalence of the mosquito vector Ae. albopictus (Skuse) was observed. Detection of dengue virus antigen in the male mosquitoes confirms that the virus is maintained in wild populations of Ae. albopictus in these areas.

Identification of sequence motifs involved in Dengue virus-host interactions.

Dengue fever is a rapidly spreading mosquito-borne virus infection, which remains a serious global public health problem. As there is no specific treatment or commercial vaccine available for effective control of the disease, the attempts on developing novel control strategies are underway. Viruses utilize the surface receptor proteins of host to enter into the cells. Though various proteins were said to be receptors of Dengue virus (DENV) using Virus Overlay Protein Binding Assay, the precise interaction between DENV and host is not explored. Understanding the structural features of domain III envelope glycoprotein would help in developing efficient antiviral inhibitors. Therefore, an attempt was made to identify the sequence motifs present in domain III envelope glycoprotein of Dengue virus. Computational analysis revealed that the NGR motif is present in the domain III envelope glycoprotein of DENV-1 and DENV-3. Similarly, DENV-1, DENV-2 and DENV-4 were found to contain Yxxphi motif which is a tyrosine-based sorting signal responsible for the interaction with a mu subunit of adaptor protein complex. High-throughput virtual screening resulted in five compounds as lead molecules based on glide score, which ranges from -4.664 to -6.52 kcal/Mol. This computational prediction provides an additional tool for understanding the virus-host interactions and helps to identify potential targets in the host. Further, experimental evidence is warranted to confirm the virus-host interactions and also inhibitory activity of reported lead compounds.