Detecting the impacts of humidity, rainfall, temperature, and season on chikungunya, dengue and Zika viruses in Aedes albopictus mosquitoes from selected sites in Cebu city, Philippines.

BACKGROUND: Aedes albopictus is the secondary vector for dengue virus (DENV) in the Philippines, and also harbors chikungunya (CHIKV) and Zika (ZIKV) viruses. This study aimed to determine the minimum infection rates (MIRs) of CHIKV, DENV serotypes, and ZIKV in Ae. albopictus collected from selected two-site categories by altitude (highland [H] and lowland [L] sites) in Cebu city, Philippines during the wet (WS) and dry seasons (DS) of 2021-2022, and to explore the relationships between these arboviral MIRs and the local weather. METHODS: The viral RNA extracts in pooled and reared adult Ae. albopictus collected during the DS and WS from two-site categories were subjected to RT-PCR to amplify and detect gene loci specific for CHIKV, DENV-1 to DENV-4, and ZIKV and analyzed with the weather data. RESULTS: The range of CHIKV MIRs was higher in the WS (13.61-107.38 infected individuals per 1,000 mosquitoes) than in the DS (13.22-44.12), but was similar between the two-site categories. Rainfall (RF) influenced the CHIKV MIR. The MIR ranges of both DENV-2 (WS: H = 0, L = 0; DS: H = 0-5.92; L = 0-2.6) and DENV-4 (WS: H = 0, L = 0-2.90; DS: H = 2.96-6.13, L = 0-15.63) differed by season but not between the two-site categories. Relative humidity (RH), RF, and temperature did not influence DENVs' MIRs. The MIR range of ZIKV was similar in both seasons (WS: 11.36-40.27; DS: 0-46.15) and two-site categories (H = 0-90.91, L = 0-55.56). RH and temperature influenced ZIKV MIR. CONCLUSIONS: RF influenced CHIKV MIR in Ae. albopictus, whereas RH and temperature influenced that of ZIKV. Season influenced the MIRs of CHIKV and DENVs but not in ZIKV. Ae. albopictus were co-infected with CHIKV, DENVs, and ZIKV in both highland and lowland sites in Cebu city. Recommendations include all-year-round implementation of the Philippine Department of Health's  4S enhanced strategy and installation of water pipelines in rural highlands for vector and disease control. Our findings are relevant to protect public health in the tropics in this climate change.

Novel and Broadly Applicable Microsatellite Markers in Identified Chromosomes of the Philippine Dengue Mosquitoes, Aedes aegypti (Diptera: Culicidae).

Dengue is the leading arboviral infection in the Philippines. Its endemicity in the country is due to the presence of its primary mosquito vector, Aedes aegypti (L.). This species has limited microsatellite markers. This study characterized microsatellite markers screened in silico from intergenic regions of the updated reference genome of Ae. aegypti from Liverpool, U.K. Criteria for good markers are: polymorphic, inherited in a Mendelian codominant manner, no null alleles, selectively neutral, randomly associated, and broadly applicable across different regions. Genotypes were scored using ABI Peak Scanner and were screened for the presence of null alleles. Hardy-Weinberg equilibrium, linkage disequilibrium, and robustness of the markers were determined by GENEPOP using Ae. aegypti samples from selected highland and lowland sites (n = 30 each) in the Philippines and outgroups (Thailand and Vietnam). Mendelian codominant inheritance was examined using F1 offspring of Ae. aegypti family (n = 30 each) derived from samples collected from Cebu city highlands and Maramag, Bukidnon. From the 63 randomly selected markers, nine were polymorphic. Two markers (Aaeg1-3D of chromosome 1 and Aaeg3-4C of chromosome 3) satisfied all criteria, hence, are good broadly useful microsatellite markers. Two other markers (Aaeg2-2E of chromosome 2 and Aaeg3-2A of chromosome 3) met all criteria but deviated from Mendelian codominant inheritance. These new markers of the Philippine Ae. aegypti with their chromosomal locations relative to the other published markers are presented, and will ultimately be useful in a variety of population genetic studies of Ae. aegypti to protect the public health.

Yearly variations of the genetic structure of Aedes aegypti (Linnaeus) (Diptera: Culicidae) in the Philippines (2017-2019).

Dengue is the fastest emerging arboviral disease in the world, imposing a substantial health and economic burden in the tropics and subtropics. The mosquito, Aedes aegypti, is the primary vector of dengue in the Philippines. We examined the genetic structure of Ae. aegypti populations collected from the Philippine major islands (Luzon, Visayas and Mindanao), each with highland (Baguio city, Cebu city mountains and Maramag, Bukidnon, respectively) and lowland sites (Quezon city; Liloan, Cebu and Cagayan de Oro [CDO] city, respectively) during the wet (2017-2018 and 2018-2019) and dry seasons (2018 and 2019). Mosquitoes (n = 1800) were reared from field-collected eggs and immatures, and were analyzed using 12 microsatellite loci. Generalized linear model analyses revealed yearly variations between highlands and lowlands in the major islands as supported by Bayesian clustering analyses on: 1) stronger selection (inbreeding coefficient, FIS = 0.52) in 2017-2018 than in 2018-2019 (FIS = 0.32) as influenced by rainfall, 2) the number of non-neutral loci indicating selection, and 3) differences of effective population size although at p = 0.05. Across sites except Baguio and CDO cities: 1) FIS varied seasonally as influenced by relative humidity (RH), and 2) the number of non-neutral loci varied as influenced by RH and rainfall indicating selection. Human-mediated activities and not isolation by distance influenced genetic differentiations of mosquito populations within (FST = 0.04) the major islands and across sites (global FST = 0.16). Gene flow (Nm) and potential first generation migrants among populations were observed between lowlands and highlands within and across major islands. Our results suggest that dengue control strategies in the epidemic wet season are to be changed into whole year-round approach, and water pipelines are to be installed in rural mountains to prevent the potential breeding sites of mosquitoes.

Temperature, season, and latitude influence development-related phenotypes of Philippine Aedes aegypti (Linnaeus): Implications for dengue control amidst global warming.

BACKGROUND: Dengue is endemic in the Philippines. Aedes aegypti is the primary vector. This study aimed to determine the hatching behavior and viability of Ae. aegypti first-generation (F1) eggs when exposed to temperature and photoperiod regimes under laboratory conditions. METHODS: Parental eggs were collected from selected highland and lowland sites in the Philippine big islands (Luzon, Visayas, and Mindanao) during the wet (2017-2018) and dry (2018) seasons. F1 egg cohorts were exposed separately in environmental chambers at 18, 25, and 38 °C with respective photoperiods for 6 weeks. Phenotypes (percent pharate larvae [PPL], hatch rates [HRs], and reproductive outputs [ROs]) were determined. RESULTS: Results of multivariate analyses of variance (MANOVA) between seasons showed significant main effects of temperature, season, and big island on all phenotypes across all sites. Significant interaction effects between seasons on all phenotypes across sites were shown between or among (1) season and big island, (2) season and temperature, (3) big island and temperature, (4) season, big island, and temperature, (5) big island, altitude, and temperature, and (6) season, big island, altitude, and temperature. Factors associated with the big islands might include their ecology, available breeding sites, and day lengths due to latitudinal differences, although they were not measured in the field. MANOVA results within each season on all phenotypes across sites showed (1) significant main effects of big island and temperature, and (2) significant interaction effects between big island and temperature within the wet season and (3) between temperature and photoperiod within the dry season. PPL were highest at 18 °C and were formed even at 38 °C in both seasons. Pharate larvae might play an adaptive role in global warming, expanded distribution to highlands, and preponderance to transmit human diseases. HRs in both seasons were highest at 25 °C and lowest at 38 °C. ROs were highest at 25 °C in the wet season and at 18 °C in the dry season. CONCLUSIONS: Temperature and latitude of Philippine big islands influenced the development-related phenotypes of Ae. aegypti in both seasons. The two seasons influenced the phenotypes and their interaction effects with big island and/or temperature and/or altitude. Recommendations include year-round enhanced 4S control strategies for mosquito vectors and water pipeline installation in rural highlands.