Dengue Fever in Mizoram: Aedes Aegypti & Aedes Albopictus Vector Insights | BMC Infectious Diseases
Study Area
Table of Contents
Mizoram is a small state in northeastern India. It has a total area of 21,087 square kilometers and shares 722 km of international borders with Myanmar and Bangladesh. The state is bordered by Assam, Tripura, and Manipur. Mizoram has a humid subtropical climate, receiving an annual rainfall of 254 centimeters with 46% relative humidity.
Data Sources
Data on dengue cases in Mizoram from 2012 to 2023 was collected from the Integrated Disease Surveillance Programme (IDSP), Government of Mizoram. Patient information included age, sex, address, and diagnosis date. Rainfall data was sourced from published literature. Disease incidence trends were analyzed, and a map showing dengue cases at the district level was created. Incidence rates were expressed as cases per 100,000 persons per year, with a focus on seasonality and age and sex distribution.
Mosquito Sample Collection
From July to December 2023, adult mosquitoes were collected in the Aizawl Municipal Area. This semi-urban area features a hot, muggy climate with periods of rain. Collection sites were close to residential neighborhoods and business complexes. Thirty-eight sites in ten localities with high dengue prevalence were selected. Mosquitoes were collected using flashlights, gravid traps, and mouth aspirators. The study protocol was approved by the Institutional Animal Ethics Committee.
Morphological identification of mosquitoes followed standard taxonomic keys. Female mosquitoes were grouped into pools of about 15, based on species and collection site. Specimens were preserved in RNAlater for later DENV detection.
DNA Extraction
Genomic DNA was isolated from mosquito legs using the QIAamp DNA Mini Kit. A minimum of six DNA extracts were prepared from each mosquito species for molecular analysis.
PCR Amplification of COX1 Gene
A segment of the COX1 gene was amplified using specific primers in a ProFlex PCR System. The PCR reaction mix was prepared and subjected to a series of temperature cycles. The PCR products were stored for further analysis.
The QIAamp Viral RNA Mini Kit was used to isolate viral RNA from mosquito pools. The heads and thoraxes of Aedes mosquitoes were grounded in a lysis buffer, followed by several washing and centrifugation steps. RNA was stored for future studies, and cDNA synthesis was performed for DENV detection and serotyping.
Real-time PCR Detection of DENV
The detection of DENV was conducted using SYBR Green-based real-time PCR in a BioRad real-time PCR system. Specific primer pairs were utilized, and cycling conditions were set for amplification. A control was included in each run. The Minimum Infection Rate (MIR) was calculated to assess DENV presence among the mosquito populations.
DENV Serotyping by PCR Amplification
The C-prM gene of DENV was amplified using specific primers for sequence analysis. Cycling conditions were set, and serotypes were identified through visualization of PCR products using agarose gel electrophoresis.
DNA Sequencing and Phylogenetic Analysis
PCR products of the COX1 gene and the C-prM gene were sequenced using the Sanger method. Sequences were aligned for analysis, and sequence identity was determined using NCBI’s GenBank nucleotide database. Phylogenetic trees were constructed using the Maximum Likelihood method.
Statistical Analysis
The seasonality of dengue disease prevalence was examined during the study period using statistical software for descriptive analysis.