Insecticide susceptibility status of human biting mosquitoes in Muheza, Tanzania

Background: There has been a rapid emergence in insecticide resistance among mosquito population to commonly used public health insecticides. This situation presents a challenge to chemicals that are currently used to control mosquitoes in sub-Saharan African. Furthermore, there is limited information on insecticide susceptibility status of human-biting mosquitoes in some areas of Tanzania. This study aimed to determine insecticide susceptibility status of human biting mosquitoes in a rural area of north-eastern Tanzania.Methods: The study was conducted in two villages in Muheza district, Tanzania. Insecticide susceptibility bioassays were performed according to the World Health Organization standard operating procedures on two to five-day old human biting mosquitoes. The mosquitoes of each species were exposed to four classes of insecticides commonly used for malaria vector control. Mosquito mortality rates (%) were determined after 24 hours post insecticide exposure.Results: Mosquito species tested were Anopheles gambiae s.l., An. funestus, Aedes aegypti, and Culex quinquefasciatus species. Real-time PCR have showed that the main sibling species of An. gambiae complex and An. funestus group were An. gambiae s. s. (58.2%) and An. funestus s. s. (91.1%), respectively. All mosquitoes, except Ae. aegypti formosus were susceptible to pirimiphos-methyl (0.25%). An. gambiae s. l. was found to be resistant to permethrin (0.75%) but showed possibility of resistance to DDT (4%) and bendiocarb (0.1%). Our findings have shown that, An. funestus was fully susceptible to all insecticide tested.Conclusion: The present study has revealed different levels of insecticide susceptibility status to four classes of commonly used insecticides in the most common mosquito vectors of human diseases in north-eastern Tanzania. The findings of the present study call for integrated vector control interventions.

Surveillance of artemether-lumefantrine associated Plasmodium falciparum multidrug resistance protein-1 gene polymorphisms in Tanzania.

BACKGROUND: Resistance to anti-malarials is a major public health problem worldwide. After deployment of artemisinin-based combination therapy (ACT) there have been reports of reduced sensitivity to ACT by malaria parasites in South-East Asia. In Tanzania, artemether-lumefantrine (ALu) is the recommended first-line drug in treatment of uncomplicated malaria. This study surveyed the distribution of the Plasmodium falciparum multidrug resistance protein-1 single nucleotide polymorphisms (SNPs) associated with increased parasite tolerance to ALu, in Tanzania. METHODS: A total of 687 Plasmodium falciparum positive dried blood spots on filter paper and rapid diagnostic test strips collected by finger pricks from patients attending health facilities in six regions of Tanzania mainland between June 2010 and August 2011 were used. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to detect Pfmdr1 SNPs N86Y, Y184F and D1246Y. RESULTS: There were variations in the distribution of Pfmdr1 polymorphisms among regions. Tanga region had exceptionally high prevalence of mutant alleles, while Mbeya had the highest prevalence of wild type alleles. The haplotype YFY was exclusively most prevalent in Tanga (29.6%) whereas the NYD haplotype was the most prevalent in all other regions. Excluding Tanga and Mbeya, four, most common Pfmdr1 haplotypes did not vary between the remaining four regions (χ² = 2.3, p = 0.512). The NFD haplotype was the second most prevalent haplotype in all regions, ranging from 17% - 26%. CONCLUSION: This is the first country-wide survey on Pfmdr1 mutations associated with ACT resistance. Distribution of individual Pfmdr1 mutations at codons 86, 184 and 1246 varies throughout Tanzanian regions. There is a general homogeneity in distribution of common Pfmdr1 haplotypes reflecting strict implementation of ALu policy in Tanzania with overall prevalence of NFD haplotype ranging from 17 to 26% among other haplotypes. With continuation of ALu as first-line drug this haplotype is expected to keep rising, thus there is need for continued pharmacovigilance studies to monitor any delayed parasite clearance by the drug

Molecular monitoring of Plasmodium falciparum super-resistance to sulfadoxine-pyrimethamine in Tanzania.

BACKGROUND: Sulfadoxine-pyrimethamine (SP) is recommended for prophylactic treatment of malaria in pregnancy while artemisinin combination therapy is the recommended first-line anti-malarial treatment. Selection of SP resistance is ongoing since SP is readily available in health facilities and in private drug shops in sub-Saharan Africa. This study reports on the prevalence and distribution of Pfdhps mutations A540E and A581G in Tanzania. When found together, these mutations confer high-level SP resistance (sometimes referred to as 'super-resistance'), which is associated with loss in protective efficacy of SP-IPTp. METHODS: DNA samples were extracted from malaria-positive blood samples on filter paper, used malaria rapid diagnostic test strips and whole blood collected from eight sites in seven administrative regions of Tanzania. PCR-RFLP and SSOP-ELISA techniques were used to genotype the A540E and A581G Pfdhps. Data were analysed using SPSS version 18 while Chi square and/or Fischer Exact tests were used to compare prevalence between regions. RESULTS: A high inter-regional variation of Pfdhps-540E was observed (χ(2) = 76.8, p < 0.001). High inter-regional variation of 581G was observed (FE = 85.3, p < 0.001). Both Tanga and Kagera were found to have the highest levels of SP resistance. A high prevalence of Pfdhps-581G was observed in Tanga (56.6 %) in northeastern Tanzania and in Kagera (20.4 %) in northwestern Tanzania and the 540-581 EG haplotype was found at 54.5 and 19.4 %, respectively. Pfdhps-581G was not detected in Pwani and Lindi regions located south of Tanga region. CONCLUSIONS: Selection of SP super-resistant Pfdhps A581G is highest in northern Tanzania. Variation in distribution of SP resistance is observed across the country: northeastern Tanga region and northwestern Kagera region have highest prevalence of SP super-resistance markers, while in Pwani and Lindi in the southeast the prevalence of super-resistance was zero. More studies should be conducted to understand the factors underlying the remarkable heterogeneity in SP resistance in the country

Prevalence of dengue and chikungunya virus infections in north-eastern Tanzania:a cross sectional study among participants presenting with malaria-like symptoms

BACKGROUND: In spite of increasing reports of dengue and chikungunya activity in Tanzania, limited research has been done to document the general epidemiology of dengue and chikungunya in the country. This study aimed at determining the sero-prevalence and prevalence of acute infections of dengue and chikungunya virus among participants presenting with malaria-like symptoms (fever, headache, rash, vomit, and joint pain) in three communities with distinct ecologies of north-eastern Tanzania. METHODS: Cross sectional studies were conducted among 1100 participants (aged 2–70 years) presenting with malaria-like symptoms at health facilities at Bondo dispensary (Bondo, Tanga), Hai hospital (Hai, Kilimanjaro) and TPC hospital (Lower Moshi). Participants who were malaria negative using rapid diagnostic tests (mRDT) were screened for sero-positivity towards dengue and chikungunya Immunoglobulin G and M (IgG and IgM) using ELISA-based kits. Participants with specific symptoms defined as probable dengue and/or chikungunya by WHO (fever and various combinations of symptoms such as headache, rash, nausea/vomit, and joint pain) were further screened for acute dengue and chikungunya infections by PCR. RESULTS: Out of a total of 1100 participants recruited, 91.2 % (n = 1003) were malaria negative by mRDT. Out of these, few of the participants (<5 %) were dengue IgM or IgG positive. A total of 381 participants had fever out of which 8.7 % (33/381) met the defined criteria for probable dengue, though none (0 %) was confirmed to be acute cases. Chikungunya IgM positives among febrile participants were 12.9 % (49/381) while IgG positives were at 3.7 % (14/381). A total of 74.2 % (283/381) participants met the defined criteria for probable chikungunya and 4.2 % (11/263) were confirmed by PCR to be acute chikungunya cases. Further analyses revealed that headache and joint pain were significantly associated with chikungunya IgM seropositivity. CONCLUSION: In north-eastern Tanzania, mainly chikungunya virus appears to be actively circulating in the population. Continuous surveillance is needed to determine the contribution of viral infections of fever cases. A possible establishment of arboviral vector preventive control measures and better diagnosis of pathogens to avoid over-treatment of other diseases should be considered. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12879-016-1511-5) contains supplementary material, which is available to authorized users

Genetic Sequence Variation in the Plasmodium falciparum Histidine-Rich Protein 2 Gene from Field Isolates in Tanzania: Impact on Malaria Rapid Diagnosis

Malaria rapid diagnosis test (RDT) is crucial for managing the disease, and the effectiveness of detection depends on parameters such as sensitivity and specificity of the RDT. Several factors can affect the performance of RDT. In this study, we focused on the pfhrp2 sequence variation and its impact on RDTs targeted by antigens encoded by Plasmodium falciparum histidine-rich protein 2 (pfhrp2). Field samples collected during cross-sectional surveys in Tanzania were sequenced to investigate the pfhrp2 sequence diversity and evaluate the impact on HRP2-based RDT performance. We observed significant mean differences in amino acid repeats between current and previous studies. Several new amino acid repeats were found to occur at different frequencies, including types AAY, AHHAHHAAN, and AHHAA. Based on the abundance of types 2 and 7 amino acid repeats, the binary predictive model was able to predict RDT insensitivity by about 69% in the study area. About 85% of the major epitopes targeted by monoclonal antibodies (MAbs) in RDT were identified. Our study suggested that the extensive sequence variation in pfhrp2 can contribute to reduced RDT sensitivity. The correlation between the different combinations of amino acid repeats and the performance of RDT in different malaria transmission settings should be investigated further

Mapping clusters of chikungunya and dengue transmission in northern Tanzania using disease exposure and vector data

Background: Dengue and chikungunya are mosquito-borne viral diseases that are of public health importance throughout the tropical and subtropical regions of the world. Seasonal variations in transmission of these viruses have been suggested owing to the ecology of their mosquito vectors. However, little is known about the epidemiology of the diseases Tanzania. To address this gap, seasonal community-based cross-sectional surveys were undertaken to identify potential clusters of transmission in Hai district in northern Tanzania.Methods: Epidemiological and entomological data from two cross-sectional surveys were used to examine the spatial pattern of dengue and chikungunya transmission. Six villages namely, Boma Ng’ombe, Magadini, Rundugai, Nshara and Kware were involved in the study. Serological measures of dengue and chikungunya virus infections were derived using enzyme-linked immunosorbent assays, and all participants were geo-referenced to the household level using a global positioning system. Potential clusters of individual exposed to dengue and chikungunya virus , as well as clusters of Aedes mosquitoes in the wet and dry seasons were detected using SaTScan. All significant clusters (with p≤0.05) were mapped using ArcGIS.Results: A large, widely dispersed cluster of chikungunya exposed individuals was detected spanning Rundugai and parts of Magadini villages (RR = 2.58,  p= 0.01), while no significant clustering was observed in the dry season. Spatial clusters of Aedes aegypti were detected in Rundugai in both the wet and dry seasons (RR = 2.56, p&lt; 0.001 and RR = 2.24, p=0.05, respectively). In the dry season a small cluster was also detected in Kware (RR = 2.25, p=0.05). No significant clusters of dengue were detected in both seasons.Conclusion: Clusters of chikungunya-exposed individuals and Aedes mosquitoes indicate on-going transmission of chikungunya virus in Hai district of northern Tanzania

Deletions of the Plasmodium falciparum histidine-rich protein 2/3 genes are common in field isolates from north-eastern Tanzania.

Plasmodium falciparum parasites lacking histidine-rich protein 2 and 3 (pfhrp2/3) genes have been reported in several parts of the world. These deletions are known to compromise the effectiveness of HRP2-based malaria rapid diagnostic tests (HRP2-RDT). The National Malaria Control Programme (NMCP) in Tanzania adopted HRP2-RDTs as a routine tool for malaria diagnosis in 2009 replacing microscopy in many Health facilities. We investigated pfhrp2/3 deletions in 122 samples from two areas with diverse malaria transmission intensities in Northeastern Tanzania. Pfhrp2 deletion was confirmed in 1.6% of samples while pfhrp3 deletion was confirmed in 50% of samples. We did not find parasites with both pfhrp2 and pfhrp3 deletions among our samples. Results from this study highlight the need for systematic surveillance of pfhrp2/3 deletions in Tanzania to understand their prevalence and determine their impact on the performance of mRDT

A non-inferiority and GLP-compliant study of broflanilide IRS (VECTRON™ T500), a novel meta-diamide insecticide against Anopheles arabiensis

Management of insecticide resistance in vector control requires development and evaluation of active ingredients (AIs) with new modes of action. VECTRON™ T500 is a wettable powder formulation used for Indoor Residual Spraying (IRS) containing 50% of broflanilide as an AI. This study evaluated the efficacy of VECTRON™ T500 sprayed on blocks of different substrates (concrete, mud and plywood) against pyrethroid susceptible and resistant Anopheles gambiae sensu stricto (s.s.) strains, and wild An. arabiensis. It also assessed the efficacy of VECTRON™ T500 in experimental huts plastered with mud and concrete against wild free-flying An. arabiensis; and non-inferiority to a World Health Organization listed indoor residual spraying product Actellic® 300CS in terms of mortality in Moshi, Tanzania.Monthly cone bioassays on blocks and in experimental huts (against pyrethroid susceptible and resistant An. gambiae s.s.) were conducted over a 12-month period after spraying of VECTRON™ T500 and Actellic® CS300. Collections of wild free-flying An. arabiensis were also done in the sprayed huts. The main outcome for cone bioassays was mortality while for the wild hut trial collections, it was mortality and blood feeding inhibition. Grouped logistic regressions with random effects were used to analyse all dichotomous outcome variables from wild collections.The results showed residual efficacy of VECTRON™ T500 of at least 80% mortality was longest on concrete, followed by plywood and then mud substrates for all mosquito strains. Furthermore, VECTRON™ T500 significantly increased the likelihood of mortality (OR:> 1.37, P<0.001) in wild collections of An. arabiensis compared to Actellic® 300CS. Blood feeding was not significantly different in the wild collection of An. arabiensis between VECTRON™ T500 and Actellic® 300CS arms.These results show that VECTRON™ T500 is efficacious against pyrethroid-resistant An. gambiae s.s. and non-inferior to Actellic® 300CS. Therefore, it should be an important addition to the current arsenal of insecticides used for insecticide resistance management and vector control

Expression of pyrethroid metabolizing P450 enzymes characterizes highly resistant Anopheles vector species targeted by successful deployment of PBO-treated bednets in Tanzania

Long lasting insecticidal nets (LLINs) are a proven tool to reduce malaria transmission, but in Africa efficacy is being reduced by pyrethroid resistance in the major vectors. A previous study that was conducted in Muleba district, Tanzania indicated possible involvement of cytochrome P450 monooxygenases in a pyrethroid resistance in An. gambiae population where pre-exposure to piperonyl butoxide (PBO) followed by permethrin exposure in CDC bottle bioassays led to partial restoration of susceptibility. PBO is a synergist that can block pyrethroid-metabolizing enzymes in a mosquito. Insecticide resistance profiles and underlying mechanisms were investigated in Anopheles gambiae and An. funestus from Muleba during a cluster randomized trial. Diagnostic dose bioassays using permethrin, together with intensity assays, suggest pyrethroid resistance that is both strong and very common, but not extreme. Transcriptomic analysis found multiple P450 genes over expressed including CYP6M2, CYP6Z3, CYP6P3, CYP6P4, CYP6AA1 and CYP9K1 in An. gambiae and CYP6N1, CYP6M7, CYP6M1 and CYP6Z1 in An. funestus. Indeed, very similar suites of P450 enzymes commonly associated with resistant populations elsewhere in Africa were detected as over expressed suggesting a convergence of mechanisms across Sub-Saharan African malaria vectors. The findings give insight into factors that may correlate with pyrethroid PBO LLIN success, broadly supporting model predictions, but revision to guidelines previously issued by the World Health Organization is warranted

Longitudinal evaluation of aflatoxin exposure in two cohorts in south-western Uganda

Aflatoxins (AF) are a group of mycotoxins. AF exposure causes acute and chronic adverse health effects such as aflatoxicosis and hepatocellular carcinoma in human populations, especially in the developing world. In this study, AF exposure was evaluated using archived serum samples from human immunodeficiency virus (HIV)-seronegative participants from two cohort studies in south-western Uganda. AFB1-lysine (AFB-Lys) adduct levels were determined via HPLC fluorescence in a total of 713 serum samples from the General Population Cohort (GPC), covering eight time periods between 1989 and 2010. Overall, 90% (642/713) of the samples were positive for AFB-Lys and the median level was 1.58 pg mg(-1) albumin (range = 0.40-168 pg mg(-1) albumin). AFB-Lys adduct levels were also measured in a total of 374 serum samples from the Rakai Community Cohort Study (RCCS), across four time periods between 1999 and 2003. The averaged detection rate was 92.5% (346/374) and the median level was 1.18 pg mg(-1) albumin (range = 0.40-122.5 pg mg(-1) albumin). In the GPC study there were no statistically significant differences between demographic parameters, such as age, sex and level of education, and levels of serum AFB-Lys adduct. In the RCCS study, longitudinal analysis using generalised estimating equations revealed significant differences between the adduct levels and residential areas (p = 0.05) and occupations (p = 0.02). This study indicates that AF exposure in people in two populations in south-western Uganda is persistent and has not significantly changed over time. Data from one study, but not the other, indicated that agriculture workers and rural area residents had more AF exposure than those non-agricultural workers and non-rural area residents. These results suggest the need for further study of AF-induced human adverse health effects, especially the predominant diseases in the region