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Dengue, Zika and chikungunya are Aedes-borne viral diseases that have become great global health concerns in the past years. Several countries in Africa have reported outbreaks of these diseases and despite Ghana sharing borders with some of these countries, such outbreaks are yet to be detected. Viral RNA and antibodies against dengue serotype-2 have recently been reported among individuals in some localities in the regional capital of Ghana. This is an indication of a possible silent transmission ongoing in the population. This study, therefore, investigated the entomological transmission risk of dengue, Zika and chikungunya viruses in a forest and domestic population in the Greater Accra Region, Ghana. All stages of the Aedes mosquito (egg, larvae, pupae and adults) were collected around homes and in the forest area for estimation of risk indices. All eggs were hatched and reared to larvae or adults for morphological identification together with larvae and adults collected from the field. The forest population had higher species richness with 7 Aedes species. The predominant species of Aedes mosquitoes identified from both sites was Aedes aegypti (98%). Aedes albopictus, an important arbovirus vector, was identified only in the peri-domestic population at a prevalence of 1.5%, significantly higher than previously reported. All risk indices were above the WHO threshold except the House Index for the domestic site which was moderate (19.8). The forest population recorded higher Positive Ovitrap (34.2% vs 26.6%) and Container (67.9% vs 36.8%) Indices than the peri-domestic population. Although none of the mosquito pools showed the presence of dengue, chikungunya or Zika viruses, all entomological risk indicators showed that both sites had a high potential arboviral disease transmission risk should any of these viruses be introduced. Continuous surveillance is recommended in these and other sites in the Metropolis to properly map transmission risk areas to inform outbreak preparedness strategies. [ABSTRACT FROM AUTHOR]

Aedes-borne viral diseases mainly Yellow Fever (YF), Dengue (DEN), Zika (ZIK) and Chikungunya (CHK) have contributed to many deaths' in the world especially in Africa. There have been major outbreaks of these diseases in West Africa. Although, YF outbreaks have occurred in Ghana over the years, no outbreak of DEN, ZIK and CHK has been recorded. However, the risk of outbreak is high due to its proximity to West African countries where outbreaks have been recently been recorded. This study surveyed the mosquito fauna to assess the risk of transmission of Yellow fever (YFV), Dengue (DENV), Chikungunya (CHKV) and Zika (ZIKV) viruses in Larabanga and Mole Game Reserve areas in Northern Ghana. The immature and adult stages of Aedes mosquitoes were collected from Larabanga and Mole Game Reserve area. There was a significant (P>0.001) number of mosquitoes collected during the rainy season than the dry season. A total of 1,930 Aedes mosquitoes were collected during the rainy season and morphologically identified. Of these, 1,915 (99.22%) were Aedes aegypti and 15 (0.22%) were Aedes vittatus. During the dry season, 27 Ae. aegypti mosquitoes were collected. A total of 415 Ae. aegypti mosquitoes were molecularly identified to subspecies level of which Ae. (Ae) aegypti aegypti was the predominant subspecies. Both Ae. aegypti aegypti and Ae aegypti formosus exist in sympatry in the area. All Aedes pools (75) were negative for DENV, ZIKV and CHKV when examined by RT- PCR. Three Larval indices namely House Index, HI (percentage of houses positive for Aedes larvae or pupae), Container Index, CI (the percentage of containers positive for Aedes larvae or pupae) and Breteau Index, BI (number of positive containers per 100 houses inspected) were assessed as a measure for risk of transmission in the study area. The HI, CI and BI for both sites were as follows; Mole Game Reserve (HI, 42.1%, CI, 23.5% and BI, 100 for rainy season and 0 for all indices for dry season) and Larabanga (39%, 15.5% and 61 for rainy season and 2.3%, 1.3% and 2.3 for dry season). The spatial distribution of Aedes breeding sites in both areas indicated that Aedes larvae were breeding in areas with close proximity to humans. Lorry tires were the main source of Aedes larvae in all the study areas. Information about the species composition and the potential role of Aedes mosquitoes in future outbreaks of the diseases that they transmit is needed to design efficient surveillance and vector control tools. [ABSTRACT FROM AUTHOR]

Vector population control using insecticides is a key element of current strategies to prevent malaria transmission in Africa. The introduction of effective insecticides, such as the organophosphate pirimiphos-methyl, is essential to overcome the recurrent emergence of resistance driven by the highly diverse Anopheles genomes. Here, we use a population genomic approach to investigate the basis of pirimiphos-methyl resistance in the major malaria vectors Anopheles gambiae and A. coluzzii. A combination of copy number variation and a single non-synonymous substitution in the acetylcholinesterase gene, Ace1, provides the key resistance diagnostic in an A. coluzzii population from Côte d'Ivoire that we used for sequence-based association mapping, with replication in other West African populations. The Ace1 substitution and duplications occur on a unique resistance haplotype that evolved in A. gambiae and introgressed into A. coluzzii, and is now common in West Africa primarily due to selection imposed by other organophosphate or carbamate insecticides. Our findings highlight the predictive value of this complex resistance haplotype for phenotypic resistance and clarify its evolutionary history, providing tools to for molecular surveillance of the current and future effectiveness of pirimiphos-methyl based interventions. Author summary: Control of mosquito populations via insecticidal tools or interventions is a mainstay of campaigns to reduce malaria transmission. However, especially in sub-Saharan Africa, continued insecticidal selection pressure on the most important species of Anopheles malaria mosquitoes has favoured the evolutionary selection of increasingly effective resistance mechanisms. We investigate the genetic basis of resistance to the organophosphate pirimiphos-methyl, the dominant insecticide now used for indoor residual spraying campaigns in Africa. Genome-wide association analysis of a population from Cote d'Ivoire showed that resistant specimens share a unique combination of mutations in one gene, the acetylcholinesterase enzyme, which constitute the prime cause of pirimiphos-methyl resistance. Further testing of these mutations in diagnostic assays involving two major malaria vectors, A. coluzzii and A. gambiae, validate their use as informative predictors of pirimiphos-methyl resistance. Using data from a large collection of whole genome sequenced specimens from a broader range of locations (Burkina-Faso, Côte d'Ivoire, Ghana, and Guinea), our evolutionary analyses demonstrate that these mutations emerged in A. gambiae and transferred into A. coluzzii by inter-specific hybridisation. Our results show how resistance mechanisms in key malaria vectors have developed and spread, and provide validated tools for molecular surveillance to inform public health campaigns. [ABSTRACT FROM AUTHOR]

Dengue fever is known to be one of the most common arthropod-borne viral infectious diseases of public health importance. The disease is now endemic in more than 100 countries in Africa, the Americas, the Eastern Mediterranean, Southeast Asia and the Western Pacific with an estimated two fifths of the world's population being at risk. The notable endemic viral hemorrhagic fevers (VHFs) found in West Africa, including yellow fever, Lassa fever, Rift Valley fever, dengue fever and until recently Ebola have been responsible for most outbreaks with fatal consequences. These VHFs usually produce unclear acute febrile illness, especially in the acute phase of infection. In this study we detected the presence of 2 different serotypes (DENV-2 and DENV-3) of Dengue virus in 4 sera of 150 patients clinically suspected of Ebola virus disease during the Ebola Virus Disease (EVD) outbreak in West Africa with the use of serological and molecular test assays. Sequence data was successfully generated for DENV-3 and phylogenetic analysis of the envelope gene showed that the DENV-3 sequences had close homology with DENV-3 sequences from Senegal and India. This study documents molecular evidence of an indigenous Dengue fever viral infection in Ghana and therefore necessitates the need to have an efficient surveillance system to rapidly detect and control the dissemination of the different serotypes in the population which has the potential to cause outbreaks of dengue hemorrhagic fevers. [ABSTRACT FROM AUTHOR]

Background: Yellow fever is endemic in some countries in Africa, and Aedes aegpyti is one of the most important vectors implicated in the outbreak. The mapping of the nation-wide distribution and the detection of insecticide resistance of vector mosquitoes will provide the beneficial information for forecasting of dengue and yellow fever outbreaks and effective control measures. Methodology/Principal Findings: High resistance to DDT was observed in all mosquito colonies collected in Ghana. The resistance and the possible existence of resistance or tolerance to permethrin were suspected in some colonies. High frequencies of point mutations at the voltage-gated sodium channel (F1534C) and one heterozygote of the other mutation (V1016I) were detected, and this is the first detection on the African continent. The frequency of F1534C allele and the ratio of F1534C homozygotes in Ae. aegypti aegypti (Aaa) were significantly higher than those in Ae. aegypti formosus (Aaf). We could detect the two types of introns between exon 20 and 21, and the F1534C mutations were strongly linked with one type of intron, which was commonly found in South East Asian and South and Central American countries, suggesting the possibility that this mutation was introduced from other continents or convergently selected after the introgression of Aaa genes from the above area. Conclusions/Significance: The worldwide eradication programs in 1940s and 1950s might have caused high selection pressure on the mosquito populations and expanded the distribution of insecticide-resistant Ae. aegypti populations. Selection of the F1534C point mutation could be hypothesized to have taken place during this period. The selection of the resistant population of Ae. aegypti with the point mutation of F1534C, and the worldwide transportation of vector mosquitoes in accordance with human activity such as trading of used tires, might result in the widespread distribution of F1534C point mutation in tropical countries. [ABSTRACT FROM AUTHOR]

The development of resistance to insecticides has become a classic exemplar of evolution occurring within human time scales. In this study we demonstrate how resistance to DDT in the major African malaria vector Anopheles gambiae is a result of both target-site resistance mechanisms that have introgressed between incipient species (the M- and S-molecular forms) and allelic variants in a DDT-detoxifying enzyme. Sequencing of the detoxification enzyme, Gste2, from DDT resistant and susceptible strains of An. gambiae, revealed a non-synonymous polymorphism (I114T), proximal to the DDT binding domain, which segregated with strain phenotype. Recombinant protein expression and DDT metabolism analysis revealed that the proteins from the susceptible strain lost activity at higher DDT concentrations, characteristic of substrate inhibition. The effect of I114T on GSTE2 protein structure was explored through X-ray crystallography. The amino acid exchange in the DDT-resistant strain introduced a hydroxyl group nearby the hydrophobic DDT-binding region. The exchange does not result in structural alterations but is predicted to facilitate local dynamics and enzyme activity. Expression of both wild-type and 114T alleles the allele in Drosophila conferred an increase in DDT tolerance. The 114T mutation was significantly associated with DDT resistance in wild caught M-form populations and acts in concert with target-site mutations in the voltage gated sodium channel (Vgsc-1575Y and Vgsc-1014F) to confer extreme levels of DDT resistance in wild caught An. gambiae. [ABSTRACT FROM AUTHOR]

The development of resistance to insecticides has become a classic exemplar of evolution occurring within human time scales. In this study we demonstrate how resistance to DDT in the major African malaria vector Anopheles gambiae is a result of both target-site resistance mechanisms that have introgressed between incipient species (the M- and S-molecular forms) and allelic variants in a DDT-detoxifying enzyme. Sequencing of the detoxification enzyme, Gste2, from DDT resistant and susceptible strains of An. gambiae, revealed a non-synonymous polymorphism (I114T), proximal to the DDT binding domain, which segregated with strain phenotype. Recombinant protein expression and DDT metabolism analysis revealed that the proteins from the susceptible strain lost activity at higher DDT concentrations, characteristic of substrate inhibition. The effect of I114T on GSTE2 protein structure was explored through X-ray crystallography. The amino acid exchange in the DDT-resistant strain introduced a hydroxyl group nearby the hydrophobic DDT-binding region. The exchange does not result in structural alterations but is predicted to facilitate local dynamics and enzyme activity. Expression of both wild-type and 114T alleles the allele in Drosophila conferred an increase in DDT tolerance. The 114T mutation was significantly associated with DDT resistance in wild caught M-form populations and acts in concert with target-site mutations in the voltage gated sodium channel (Vgsc-1575Y and Vgsc-1014F) to confer extreme levels of DDT resistance in wild caught An. gambiae.

The development of resistance to insecticides has become a classic exemplar of evolution occurring within human time scales. In this study we demonstrate how resistance to DDT in the major African malaria vector Anopheles gambiae is a result of both target-site resistance mechanisms that have introgressed between incipient species (the M- and S-molecular forms) and allelic variants in a DDT-detoxifying enzyme. Sequencing of the detoxification enzyme, Gste2, from DDT resistant and susceptible strains of An. gambiae, revealed a non-synonymous polymorphism (I114T), proximal to the DDT binding domain, which segregated with strain phenotype. Recombinant protein expression and DDT metabolism analysis revealed that the proteins from the susceptible strain lost activity at higher DDT concentrations, characteristic of substrate inhibition. The effect of I114T on GSTE2 protein structure was explored through X-ray crystallography. The amino acid exchange in the DDT-resistant strain introduced a hydroxyl group nearby the hydrophobic DDT-binding region. The exchange does not result in structural alterations but is predicted to facilitate local dynamics and enzyme activity. Expression of both wild-type and 114T alleles the allele in Drosophila conferred an increase in DDT tolerance. The 114T mutation was significantly associated with DDT resistance in wild caught M-form populations and acts in concert with target-site mutations in the voltage gated sodium channel (Vgsc-1575Y and Vgsc-1014F) to confer extreme levels of DDT resistance in wild caught An. gambiae. [ABSTRACT FROM AUTHOR]

High-malaria burden countries in sub-Saharan Africa are shifting from malaria control towards elimination. Hence, there is need to gain a contemporary understanding of how indoor residual spraying (IRS) with non-pyrethroid insecticides when combined with long-lasting insecticidal nets (LLINs) impregnated with pyrethroid insecticides, contribute to the efforts of National Malaria Control Programmes to interrupt transmission and reduce the reservoir of Plasmodium falciparum infections across all ages. Using an interrupted time-series study design, four age-stratified malariometric surveys, each of ~2,000 participants, were undertaken pre- and post-IRS in Bongo District, Ghana. Following the application of three-rounds of IRS, P. falciparum transmission intensity declined, as measured by a >90% reduction in the monthly entomological inoculation rate. This decline was accompanied by reductions in parasitological parameters, with participants of all ages being significantly less likely to harbor P. falciparum infections at the end of the wet season post-IRS (aOR = 0.22 [95% CI: 0.19-0.26], p-value < 0.001). In addition, multiplicity of infection (MOI var ) was measured using a parasite fingerprinting tool, designed to capture within-host genome diversity. At the end of the wet season post-IRS, the prevalence of multi-genome infections declined from 75.6% to 54.1%. This study demonstrates that in areas characterized by high seasonal malaria transmission, IRS in combination with LLINs can significantly reduce the reservoir of P. falciparum infection. Nonetheless despite this success, 41.6% of the population, especially older children and adolescents, still harboured multi-genome infections. Given the persistence of this diverse reservoir across all ages, these data highlight the importance of sustaining vector control in combination with targeted chemotherapy to move high-transmission settings towards pre-elimination. This study also points to the benefits of molecular surveillance to ensure that incremental achievements are not lost and that the goals advocated for in the WHO's High Burden to High Impact strategy are realized.

The Anopheles gambiae sensu lato species complex consists of a number of cryptic species with different habitats and behaviours. These morphologically indistinct species are identified by chromosome banding. Several molecular diagnostic techniques for distinguishing between An. coluzzii and An. gambiae are still under improvement. Although, the current SINE method for identification between An. coluzzii and An. gambiae works reliably, this study describes a refinement of the SINE method to increase sensitivity for identification of An. coluzzii, An. gambiae and An. arabiensis based on amplicon dissociation curve characteristics. Field-collected samples, laboratory-reared colonies and crossed specimens of the two species were used for the design of the protocol. An. gambiae, An. coluzzii, and hybrids of the two species were sampled from Ghana and An. arabiensis from Kenya. Samples were first characterised using conventional SINE PCR method, and further assayed using SYBR green, an intercalating fluorescent dye. The three species and hybrids were clearly differentiated using the melting temperature of the dissociation curves, with derivative peaks at 72°C for An. arabiensis, 75°C for An. gambiae and 86°C for An. coluzzii. The hybrids (An. gambiae / An. coluzzii) showed both peaks. This work is the first to describe a SYBR green real time PCR method for the characterization of An. arabiensis, An. gambiae and An. coluzzii and was purposely designed for basic melt-curve analysis (rather than high-resolution melt-curve) to allow it to be used on a wide range of real-time PCR machines., Competing Interests: Joseph Chabi, Louis Kouadio N’Dri, Alex Datsomor and Dora Okyere were employed by Noguchi Memorial Institute for Medical Research (NMIMR) within Vestergaard-NMIMR project. Helen Pates Jamet and Melinda Hadi were employed by Vestergaard. This commercial affiliation of some authors does not alter our adherence to PLOS ONE policies on sharing data and materials.