Mating of unfed, engorged, and partially to fully gravid Aedes aegypti (Diptera: Culicidae) female mosquitoes in producing viable eggs.

BACKGROUND: Understanding the relationship between blood-feeding and mating is important in effectively managing the most well-adapted vector insect, Aedes aegypti (Linnaeus). Although extensive studies have investigated the behavioural aspects of Aedes such as blood-feeding, mating, and their relationship, several knowledge gaps still exist. Therefore, the present study was undertaken to determine the possibility of successful mating by unfed, engorged, and partially to fully gravid (up to 5 days after blood-feeding with fully developed eggs) female Ae. aegypti mosquitoes and production of viable eggs. METHODS: Mating of sexually mature adult Aedes aegypti was allowed in three different ways. In control 1, the females were allowed to mate before taking blood meal, and in control 2, the females were not at all allowed to mate. In the experiment, the females were separated into six categories, viz. D-0 to D-5. In D-0, the females were allowed to mate immediately after the bloodmeal and, in D-1, the females were allowed to mate on the first day of blood feeding, likewise, the females of D-2, D-3, D-4 and D-5 were allowed to mate on 2nd, 3rd, 4th and 5th day of blood feeding. Ovitrap was uniformly kept on the 4th day of blood feeding for the cages D-0 to D-3 for 1 h and then removed and for the cages D-4, and D-5, the ovitrap was kept on 4th and 5th day of blood feeding for 1h immediately after mating. The total number of eggs and the total number of hatching were counted. In the subsequent days, the entire experiment was replicated two times with different cohorts of mosquitoes, and the mean value of three experiments was used to draw Excel bars with 5% error bars and also for the statistical analysis. RESULTS: It was found that mating just before oviposition was sufficient to produce 1581 eggs (70% compared with control) and fertilize 1369 eggs (85% compared with total eggs laid), which is far higher than the 676 non-hatching (unfertilized) eggs (30%) laid by unmated females. Although mating is not essential for producing eggs, our study shows that even brief exposure to the semen and seminal fluids greatly enhances the oviposition and hatching efficiency, even if the mating occurs just before oviposition. However, those females mating before blood-feeding and those mating after blood-feeding produced 2266 and 2128 eggs, with hatching rates of 96.78% and 95.54%, respectively. Hence, the retention time of seminal fluid in the female seems to influence the number of eggs laid and the number of eggs hatched. CONCLUSIONS: In general, mating is possible in Ae. aegypti even minutes before oviposition and is sufficient to produce a greater number of viable eggs.

West Nile virus in India: An update on its genetic lineages.

West Nile virus (WNV) is a rapidly spreading mosquito-transmitted zoonotic flavivirus. Mosquitoes belonging to the genus Culex are incriminated as the principal vectors of the virus, which causes West Nile fever (WNF) in humans. Manifestations of WNF include a mild, self-limiting, flu-like illness, which in severe cases (rare) may progress to encephalitis, resulting in life-threatening consequences. WNV is geographically distributed worldwide, covering Africa, the Americas, Europe, and Asia (except Antarctica). The virus exists in a bird-mosquito transmission cycle in nature, with humans and horses as incidental/accidental hosts. The virus can infect a large variety of hosts worldwide, i.e., about 300 birds and around 70 different mosquito species belonging to several genera. For a long time, it was believed that WNV was not highly virulent and caused only mild infection globally. However, the recent frequent and increasing incidence of clinically severe WNV infections, such as encephalitis in humans and horses with significant mortality, has been reported in the Americas, Europe, and several East Asian countries. The emergence of lineage 2 strains endemic to Africa, with epidemic potential in humans and horses in Europe, is considered a serious global health concern. Although WNV is known to circulate in India since 1952, its re-emergence with severe neuro-invasive pathogenic potential in humans in Assam, Kerala, West Bengal and Tamil Nadu states signals urgent efforts to understand the dynamics of circulating strains with regard to its vector, hosts, and environment. This could be done by prioritizing "One Health" approach for developing effective preventive and control strategies. In view of the global interest, we present an overview of the circulating genetic lineages of WNV in India in comparison with the global scenario. In addition, we stress on holistic approaches of "One Health" strategy as the current need of the hour for designing effective preventive and control strategies in the country.

Impact of COVID-19 lockdown during 2020 on the occurrence of vector-borne diseases in India.

BACKGROUND & OBJECTIVES: India imposed one of the world's largest nationwide lockdown in the aftermath of COVID-19 pandemic in March 2020 which was partly extended up to December. Some of the impacts of the COVID-19 lockdown on the economy, research, travel, education, and sports were readily apparent; the same was less obvious in the occurrence of vector-borne diseases (VBDs). The objective of this study was to statistically determine the impact of the COVID-19 lockdown on the occurrence of VBDs in India. METHODS: The reported incidence of VBDs such as malaria, dengue, Chikungunya, Japanese encephalitis, and kala-azar in India during the years 2015-2019 was analyzed by fitting Poisson and negative binomial (NB) models for each VBDs separately. The number of cases reported was compared with the number of cases predicted for each year from 2015 to 2020 for all the VBDs under study to infer whether or not the lockdown had any impact on their prevalence in India. RESULTS: The percentage of the actual case was lower by 46, 75, 49, 72, and 38 respectively for malaria, dengue, Chikungunya, Japanese encephalitis, and kala-azar when comparing the lockdown period and the year before lockdown (2020 vs. 2019). The number of cases predicted for the year 2020 based on the trend of the five preceding years (2015-2019) also showed a great variation between actual and predicted cases. The differences in cases were considered as the cases missed in 2020 were largely due to the lockdown. INTERPRETATION & CONCLUSION: The analysis showed that the lockdown had a considerable impact on the occurrence of VBDs.

Molecular Evidence of Rickettsia conorii subsp. raoultii and Rickettsia felis in Haemaphysalis intermedia Ticks in Sirumalai, Eastern Ghats, Tamil Nadu, South India.

Rickettsia is an important pathogenic entity among tick-borne diseases (TBD), which are considered serious emerging public health problems globally. In India, though the widespread distribution of ticks and TBD has been documented, its real burden remains underreported. In a preliminary attempt, rickettsial surveillance was carried out in ticks collected from Sirumalai, Eastern Ghats in Tamil Nadu, India by using pathogen genome-based phylogenetic inferences generated through multi-locus sequence typing (MLST), targeting the genes 16s rRNA, OmpA, OmpB, and gltA by nested PCR. The laboratory evidence confirms the circulation of Rickettsia in Haemaphysalis intermedia species collected from this area. Analysis of the four gene sequences detected demonstrates their closest identity to the spotted fever group (SFG) available in the GenBank database. Further, multiple sequence alignment with other sequences derived from the GenBank database showed close relatedness to Rickettsia conorii subsp. raoultii (16s rDNA-99.32%, OmpA-93.38%, OmpB-97.39%, and gltA-98.57%) and Rickettsia felis (16s rDNA 99.54%, OmpA-100%, OmpB-100% and gltA-99.41%). With this genomic evidence, the circulation of rickettsial pathogens in the pools of H. intermedia ticks infesting livestock in the Sirumalai foothill area has been demonstrated and to complement the microscopic identification of the tick species, DNA barcodes were generated for H. intermedia using the mitochondrial cytochrome c oxidase subunit I gene (COI). Nevertheless, R. raoultii and R. felis were found to be the aetiological agents of tick-borne lymphadenopathy and flea-borne spotted fever in human cases, respectively, further study on the determination of their diversity, distribution, clinical relevance, and potential risk to the local community in these areas is highly warranted.

Molecular Detection of Multiple Genotypes of Orientia tsutsugamushi Causing Scrub Typhus in Febrile Patients from Theni District, South India.

Scrub typhus (St) is a re-emerging mite-transmitted public health problem in Southeast Asia with escalating case incidences in the endemic areas. Though, more than 40 genotypes of the causative agent Orientia tsutsugamushi (Ot) have been documented, the information on the circulating genotypes in India is scanty. A hospital-based retrospective screening was undertaken to map the circulating molecular subtypes of the etiological agent in serologically confirmed scrub typhus (St) human cases, by targeting the GroEL gene of O. tsutsugamushi using the nested polymerase chain reaction method. Nine out of 34 samples (26%) yielded positive results and DNA sequencing analysis of six positive samples out of nine revealed that the sequences were related to three major genotypes, such as Karp (HSB1, FAR1), Kato (Wuj/2014, UT76), and Kawasaki (Kuroki, Boryong, Gilliam, and Hwasung). Additionally, the St-positive samples exhibited 100% and 99.45%; 97.53% and 97.81%; 96.99% nucleotide identity with the closely related Karp, Kato, and Kawasaki-related sequences, respectively. Overall, 94% of the nucleotides were conserved, and the variable site was 20/365 (5.5%). The prevalence of multiple genotypes among human cases further stresses the need to conduct in-depth studies to map the genotypes and their clinical relevance, and the contributing risk factors for the emergence of St cases in this area.

Japanese encephalitis virus in India: An update on virus genotypes.

Japanese encephalitis (JE) is a leading cause of viral encephalitis in Southeast Asia. It is a serious public health issue in India, and cases have been emerging in newer areas of the country. Although vaccination efforts have already been initiated in the country since 2006 and later through the Universal Immunization Programme in 2011, still a significant reduction in the number of cases has to be achieved since an escalating trend of JE incidence has been reported in certain States such as Assam, Uttar Pradesh and West Bengal. Moreover, fresh cases of JE have been reported from certain pockets in Odisha as well. Despite the mass JE vaccination programme implemented in prioritized endemic zones in the country in 2011, a shift in the age group of JE virus (JEV) infection was noticed affecting the adult population in West Bengal. The recent detection of the circulation of genotype I (GI) in Gorakhpur, Uttar Pradesh and the co-circulation of GI and genotype III (GIII) in West Bengal are probably a warning signal for the public health personnel to strengthen the surveillance system in all endemic hotspots in the country. The abrupt emergence of JEV genotype V (GV) in China and Korea in 2009, after its first detection in Malaya in 1952, endemic countries have been cautioned to strengthen their surveillance, because GV has been suspected of getting dispersed efficiently in other parts of Asia. Moreover, the reduced protection efficiency of the JEV GIII-based vaccine against the JEV genotype V further warrants careful evaluation of the ongoing vaccination strategies in the endemic countries, anticipating the possible incursion of GV and its impact on future control strategies. In view of the above facts, the present communication reviews the current knowledge on the molecular epidemiology of JEV in India vis-a-vis the global scenario and discusses the future priorities in JEV research in India for effectively designing control strategies.

Nucleic acid detection strategy using gold nanoprobe of two diverse origin.

In recent years, nanoparticles especially with gold and silver nanoparticles based point of care diagnostic methods is being developed for the lethal diseases like dengue. This study focused to work on the dengue virus detection in a simplest method using gold nanoparticles probe (AuNPs) with thiol tagged single strand DNA (ss-DNA). A sensitive, fluorescence-based detection strategy was designed to examine and quantified the hybridisation process and also elucidated the behaviour of AuNPs before and after interaction of biomolecule. The detection process was focused on aggregation of gold nanoprobe in the presence of complementary strand (target region). Hence the percentage of aggregation was measured and as a result, the limit of detection was found to be 10-6 dilutions. Current detection method was highly sensitive, easy to perform and the reaction timing is rapid between 5 and 10 min, and it can be observed through naked eye.

Kyasanur Forest Disease in India: innovative options for intervention.

Kyasanur Forest Disease (KFD) is a tick-borne hemorrhagic fever of human, caused by Kyasanur forest disease virus (KFDV) in India. The tick, Haemaphysalis spinigera, has been incriminated as the vector of KFDV. In human, KFD clinically presents with high fever, frontal headache, and severe myalgia, followed by bleeding from the nasal cavity, throat, gingivae, and in some cases, gastrointestinal tract. The mortality rate in KFDV infected cases is estimated to be 3-10%. Monkeys infected with the virus also develop the disease and die. Though the incidence of KFD was found to be confined only to the sylvatic area of Shimoga district in Karnataka state in India during 1967, recent reports indicate its expanding potential to the neighboring states such as Kerala, Tamil Nadu, and Goa. The administration of an indigenous, inactivated tissue culture vaccine was found to drastically decrease the percentage of incidence; however, the recurrence of KFD in vaccinated subjects warrants innovative strategies for effective control of the infection. The present communication proposes and discusses innovative intervention strategies for the effective prevention and control of KFD in India.

DNA-triangular silver nanoparticles nanoprobe for the detection of dengue virus distinguishing serotype.

There is always a substantial effort to develop a point of care detection for a severe and acute disease like dengue. In this work, we have described the detection of serotype-specific dengue virus using multicolor triangular silver nanoparticles (TAg) which could be a potential diagnosis method to distinguish between serotype. Functionalized TAg was prepared with polyA10 DNA through pH-mediated process. Further, it was characterized with Transmission Electron Microscopy (TEM), Ultra Visible-visible spectroscopy (UV-vis) and gel electrophoresis and its stability towards NaCl concentration. The colorimetric detection was carried out based on the interaction of TAg-DNA probe with specific complimentary strand which was designed to form a network assembly between DNA probes and sample RNA. The hybridization process was enhanced in presence of NaCl (0.4 M) which makes the hybridization process more stable over probe compared to un-hybridization. Therefore a simple and effective colorimetric method was developed to detect the different dengue serotype RNA with high sensitivity and specificity.