Semi-field evaluation of freestanding transfluthrin passive emanators and the BG sentinel trap as a "push-pull control strategy" against Aedes aegypti mosquitoes

Spatial repellents that drive mosquitoes away from treated areas, and odour-baited traps, that attract and kill mosquitoes, can be combined and work synergistically in a push-pull system. Push-pull systems have been shown to reduce house entry and outdoor biting rates of malaria vectors and so have the potential to control other outdoor biting mosquitoes such as Aedes aegypti that transmit arboviral diseases. In this study, semi-field experiments were conducted to evaluate whether a push-pull system could be used to reduce bites from Aedes mosquitoes.; The push and pull under investigation consisted of two freestanding transfluthrin passive emanators (FTPE) and a BG sentinel trap (BGS) respectively. The FTPE contained hessian strips treated with 5.25 g of transfluthrin active ingredient. The efficacies of FTPE and BGS alone and in combination were evaluated by human landing catch in a large semi-field system in Tanzania. We also investigated the protection of FTPE over six months. The data were analyzed using generalized linear mixed models with binomial distribution.; Two FTPE had a protective efficacy (PE) of 61.2% (95% confidence interval (CI): 52.2-69.9%) against the human landing of Ae. aegypti. The BGS did not significantly reduce mosquito landings; the PE was 2.1% (95% CI: -2.9-7.2%). The push-pull provided a PE of 64.5% (95% CI: 59.1-69.9%). However, there was no significant difference in the PE between the push-pull and the two FTPE against Ae. aegypti (P = 0.30). The FTPE offered significant protection against Ae. aegypti at month three, with a PE of 46.4% (95% CI: 41.1-51.8%), but not at six months with a PE of 2.2% (95% CI: -9.0-14.0%).; The PE of the FTPE and the full push-pull are similar, indicative that bite prevention is primarily due to the activity of the FTPE. While these results are encouraging for the FTPE, further work is needed for a push-pull system to be recommended for Ae. aegypti control. The three-month protection against Ae. aegypti bites suggests that FTPE would be a useful additional control tool during dengue outbreaks, that does not require regular user compliance

Dramatic decreases of malaria transmission intensities in Ifakara, south-eastern Tanzania since early 2000s.

BACKGROUND: Ongoing epidemiological transitions across Africa are particularly evident in fast-growing towns, such as Ifakara in the Kilombero valley, south-eastern Tanzania. This town and its environs (population ~ 70,000) historically experienced moderate to high malaria transmission, mediated mostly by Anopheles gambiae and Anopheles funestus. In early 2000s, malaria transmission [Plasmodium falciparum entomological inoculation rate (PfEIR)] was estimated at ~ 30 infectious bites/person/year (ib/p/yr). This study assessed the PfEIR after 15 years, during which there had been rapid urbanization and expanded use of insecticide-treated nets (ITNs). METHODS: Randomly-selected 110 households were sampled across Ifakara town and four adjacent wards. Mosquitoes were trapped nightly or monthly (June.2015-May.2016) using CDC-light-traps indoors, Suna® traps outdoors and human landing catches (HLC) indoors and outdoors. All Anopheles mosquitoes were morphologically identified and analysed by ELISA for Plasmodium circumsporozoite proteins. Mosquito blood meals were identified using ELISA, and sub-samples of An. gambiae and An. funestus examined by PCR to distinguish morphologically-similar siblings. Insecticide resistance was assessed using WHO-susceptibility assays, and some Anopheles were dissected to examine ovariole tracheoles for parity. RESULTS: After 3572 trap-nights, one Plasmodium-infected Anopheles was found (an An. funestus caught outdoors in Katindiuka-ward by HLC), resulting in overall PfEIR of 0.102 ib/p/yr. Nearly 80% of malaria vectors were from Katindiuka and Mlabani wards. Anopheles gambiae densities were higher outdoors (64%) than indoors (36%), but no such difference was observed for An. funestus. All An. funestus and 75% of An. gambiae dissected were parous. Anopheles gambiae complex consisted entirely of Anopheles arabiensis, while An. funestus included 84.2% An. funestus s.s., 4.5% Anopheles rivulorum, 1.4% Anopheles leesoni and 9.9% with unamplified-DNA. Anopheles gambiae were susceptible to bendiocarb and malathion, but resistant to pyrethroids, DDT and pirimiphos-methyl. Most houses had brick walls and/or iron roofs (> 90%), and 52% had screened windows. CONCLUSION: Malaria transmission in Ifakara has decreased by > 99% since early-2000s, reaching levels nearly undetectable with current entomological methods. These declines are likely associated with ITNs use, urbanization and improved housing. Remaining risk is now mostly in peri-urban wards, but concerted efforts could further decrease local transmission. Parasitological surveys are required to assess actual prevalence, incidence and importation rates

Long-lasting insecticidal nets retain bio-efficacy after 5 years of storage: implications for malaria control programmes.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are the most sustainable and effective malaria control tool currently available. Global targets are for 80% of the population living in malaria endemic areas to have access to (own) and use a LLIN. However, current access to LLINs in endemic areas is 56% due to system inefficiencies and budget limitations. Thus, cost-effective approaches to maximize access to effective LLINs in endemic areas are required. This study evaluated whether LLINs that had been stored for 5 years under manufacturer's recommended conditions may be optimally effective against Anopheles mosquitoes, to inform malaria control programmes and governments on the periods over which LLINs may be stored between distributions, in an effort to maximize use of available LLINs. METHODS: Standard World Health Organization (WHO) bioassays (cone and tunnel test) were used to evaluate the bio-efficacy and wash resistance of Olyset® and DawaPlus® 2.0 (rebranded Tsara® Soft) LLINs after 5 years of storage at 25 °C to 33.4 °C and 40% to 100% relative humidity. In addition, a small scale Ifakara Ambient Chamber test (I-ACT) was conducted to compare the bio-efficacy of one long stored LLINs to one new LLIN of the same brand, washed or unwashed. LLINs were evaluated using laboratory reared fully susceptible Anopheles gambiae sensu stricto (s.s.) (Ifakara strain) and pyrethroid resistant Anopheles arabiensis (Kingani strain). RESULTS: After 5 years of storage, both unwashed and washed, Olyset® and DawaPlus® 2.0 (Tsara® Soft) LLINs passed WHO bio-efficacy criteria on knockdown (KD60) ≥ 95%, 24-h mortality ≥ 80% and ≥ 90% blood-feeding inhibition in WHO assays against susceptible An. gambiae s.s. DawaPlus® 2.0 LLINs also passed combined WHO bioassay criteria against resistant An. arabiensis. Confirmatory I-ACT tests using whole nets demonstrated that long-stored LLINs showed higher efficacy than new LLINs on both feeding inhibition and mortality endpoints against resistant strains. CONCLUSIONS: Even after long-term storage of around 5 years, both Olyset® and DawaPlus® 2.0 LLINs remain efficacious against susceptible Anopheles mosquitoes at optimal storage range of 25 °C to 33.4 °C for temperature and 40% to 100% relative humidity measured by standard WHO methods. DawaPlus® 2.0 (Tsara® Soft) remained efficacious against resistant strain

Single blinded semi-field evaluation of MAÏA® topical repellent ointment compared to unformulated 20% DEET against Anopheles gambiae, Anopheles arabiensis and Aedes aegypti in Tanzania.

BACKGROUND: N,N-Diethyl-3-methylbenzamide (DEET) topical mosquito repellents are effective personal protection tools. However, DEET-based repellents tend to have low consumer acceptability because they are cosmetically unappealing. More attractive formulations are needed to encourage regular user compliance. This study evaluated the protective efficacy and protection duration of a new topical repellent ointment containing 15% DEET, MAÏA® compared to 20% DEET in ethanol using malaria and dengue mosquito vectors in Bagamoyo Tanzania. METHODS: Fully balanced 3 × 3 Latin square design studies were conducted in large semi-field chambers using laboratory strains of Anopheles gambiae sensu stricto, Anopheles arabiensis and Aedes aegypti. Human volunteers applied either MAÏA® ointment, 20% DEET or ethanol to their lower limbs 6 h before the start of tests. Approximately 100 mosquitoes per strain per replicate were released inside each chamber, with 25 mosquitoes released at regular intervals during the collection period to maintain adequate biting pressure throughout the test. Volunteers recaptured mosquitoes landing on their lower limbs for 6 h over a period of 6 to 12-h post-application of repellents. Data analysis was conducted using mixed-effects logistic regression. RESULTS: The protective efficacy of MAÏA® and 20% DEET was not statistically different for each of the mosquito strains: 95.9% vs. 97.4% against An. gambiae (OR = 1.53 [95% CI 0.93-2.51] p = 0.091); 96.8% vs 97.2% against An. arabiensis (OR = 1.08 [95% CI 0.66-1.77] p = 0.757); 93.1% vs 94.6% against Ae. aegypti (OR = 0.76 [95% CI 0.20-2.80] p = 0.675). Average complete protection time (CPT) in minutes of MAÏA® and that of DEET was similar for each of the mosquito strains: 571.6 min (95% CI 558.3-584.8) vs 575.0 min (95% CI 562.1-587.9) against An. gambiae; 585.6 min (95% CI 571.4-599.8) vs 580.9 min (95% CI 571.1-590.7) against An. arabiensis; 444.1 min (95% CI 401.8-486.5) vs 436.9 min (95% CI 405.2-468.5) against Ae. aegypti. CONCLUSIONS: MAÏA® repellent ointment provides complete protection for 9 h against both An. gambiae and An. arabiensis, and 7 h against Ae. aegypti similar to 20% DEET (in ethanol). MAÏA® repellent ointment can be recommended as a tool for prevention against outdoor biting mosquitoes in tropical locations where the majority of the people spend an ample time outdoor before going to bed

Observing the distribution of mosquito bites on humans to inform personal protection measures against malaria and dengue vectors

Background Understanding mosquito biting behaviours is important for designing and evaluating protection methods against nuisance biting and mosquito-borne diseases (e.g. dengue, malaria and zika). We investigated the preferred biting sites by Aedes aegypti and Anopheles arabiensis on adult volunteers in standing or sleeping positions; and estimated the theoretical protection limits affordable from protective clothing or repellent-treated footwear. Methods Adult volunteers dressed in shorts and t-shirts were exposed to infection-free laboratory-reared mosquitoes inside screened chambers from 6am to noon (for day-biting Ae. aegypti) or 6pm to midnight (night-biting An. arabiensis). Attempted bites on different body parts were recorded. Comparative observations were made on same volunteers while wearing sandals treated with transfluthrin, a vapour-phase pyrethroid that kills and repels mosquitoes. Results An. arabiensis bites were mainly on the lower limbs of standing volunteers (95.9% of bites below the knees) but evenly-distributed over all exposed body surfaces when the volunteers were on sleeping positions (only 28.8% bites below knees). Ae. aegypti bites were slightly concentrated on lower limbs of standing volunteers (47.7% below knees), but evenly-distributed on sleeping volunteers (23.3% below knees). Wearing protective clothing that leave only hands and head uncovered (e.g. socks + trousers + long-sleeved shirts) could theoretically prevent 78–83% of bites during sleeping, and at least 90% of bites during non-sleeping hours. If the feet are also exposed, protection declines to as low as 36.3% against Anopheles. The experiments showed that transfluthrin-treated sandals reduced An. arabiensis by 54–86% and Ae. aegypti by 32–39%, but did not change overall distributions of bites. Conclusion Biting by An. arabiensis and Ae. aegypti occur mainly on the lower limbs, though this proclivity is less pronounced in the Aedes species. However, when hosts are on sleeping positions, biting by both species is more evenly-distributed over the exposed body surfaces. High personal protection might be achieved by simply wearing long-sleeved clothing, though protection against Anopheles particularly requires covering of feet and lower legs. The transfluthrin-treated footwear can reduce biting risk, especially by An. arabiensis. These findings could inform the design and use of personal protection tools (both insecticidal and non-insecticidal) against mosquitoes and mosquito-borne diseases

Protecting migratory farmers in rural Tanzania using eave ribbons treated with the spatial mosquito repellent, transfluthrin.

BACKGROUND: Many subsistence farmers in rural southeastern Tanzania regularly relocate to distant farms in river valleys to tend to crops for several weeks or months each year. While there, they live in makeshift semi-open structures, usually far from organized health systems and where insecticide-treated nets (ITNs) do not provide adequate protection. This study evaluated the potential of a recently developed technology, eave ribbons treated with the spatial repellent transfluthrin, for protecting migratory rice farmers in rural southeastern Tanzania against indoor-biting and outdoor-biting mosquitoes. METHODS: In the first test, eave ribbons (0.1 m × 24 m each) treated with 1.5% transfluthrin solution were compared to untreated ribbons in 24 randomly selected huts in three migratory communities over 48 nights. Host-seeking mosquitoes indoors and outdoors were monitored nightly (18.00-07.00 h) using CDC light traps and CO2-baited BG malaria traps, respectively. The second test compared efficacies of eave ribbons treated with 1.5% or 2.5% transfluthrin in 12 huts over 21 nights. Finally, 286 farmers were interviewed to assess perceptions about eave ribbons, and their willingness to pay for them. RESULTS: In the two experiments, when treated eave ribbons were applied, the reduction in indoor densities ranged from 56 to 77% for Anopheles arabiensis, 36 to 60% for Anopheles funestus, 72 to 84% for Culex, and 80 to 98% for Mansonia compared to untreated ribbons. Reduction in outdoor densities was 38 to 77% against An. arabiensis, 36 to 64% against An. funestus, 63 to 88% against Culex, and 47 to 98% against Mansonia. There was no difference in protection between the two transfluthrin doses. In the survey, 58% of participants perceived the ribbons to be effective in reducing mosquito bites. Ninety per cent were willing to pay for the ribbons, the majority of whom were willing to pay but less than US$2.17 (5000 TZS), one-third of the current prototype cost. CONCLUSIONS: Transfluthrin-treated eave ribbons can protect migratory rice farmers, living in semi-open makeshift houses in remote farms, against indoor-biting and outdoor-biting mosquitoes. The technology is acceptable to users and could potentially complement ITNs. Further studies should investigate durability and epidemiological impact of eave ribbons, and the opportunities for improving affordability to users

Associations Between Methylation of Paternally Expressed Gene 3 (PEG3), Cervical Intraepithelial Neoplasia and Invasive Cervical Cancer.

Cytology-based screening for invasive cervical cancer (ICC) lacks sensitivity and specificity to discriminate between cervical intraepithelial neoplasia (CIN) likely to persist or progress from cases likely to resolve. Genome-wide approaches have been used to identify DNA methylation marks associated with CIN persistence or progression. However, associations between DNA methylation marks and CIN or ICC remain weak and inconsistent. Between 2008-2009, we conducted a hospital-based, case-control study among 213 Tanzania women with CIN 1/2/3 or ICC. We collected questionnaire data, biopsies, peripheral blood, cervical scrapes, Human papillomavirus (HPV) and HIV-1 infection status. We assessed PEG3 methylation status by bisulfite pyrosequencing. Multinomial logistic regression was used to estimate odds ratios (OR) and confidence intervals (CI 95%) for associations between PEG3 methylation status and CIN or ICC. After adjusting for age, gravidity, hormonal contraceptive use and HPV infection, a 5% increase in PEG3 DNA methylation was associated with increased risk for ICC (OR = 1.6; 95% CI 1.2-2.1). HPV infection was associated with a higher risk of CIN1-3 (OR = 15.7; 95% CI 5.7-48.6) and ICC (OR = 29.5, 95% CI 6.3-38.4). Infection with high risk HPV was correlated with mean PEG3 differentially methylated regions (DMRs) methylation (r = 0.34 p<0.0001), while the correlation with low risk HPV infection was weaker (r = 0.16 p = 0.047). Although small sample size limits inference, these data support that PEG3 methylation status has potential as a molecular target for inclusion in CIN screening to improve prediction of progression. Impact statement: We present the first evidence that aberrant methylation of the PEG3 DMR is an important co-factor in the development of Invasive cervical carcinoma (ICC), especially among women infected with high risk HPV. Our results show that a five percent increase in DNA methylation of PEG3 is associated with a 1.6-fold increase ICC risk. Suggesting PEG3 methylation status may be useful as a molecular marker for CIN screening to improve prediction of cases likely to progress

Detection of malaria parasites in dried human blood spots using mid-infrared spectroscopy and logistic regression analysis

Background: Epidemiological surveys of malaria currently rely on microscopy, polymerase chain reaction assays (PCR) or rapid diagnostic test kits for Plasmodium infections (RDTs). This study investigated whether mid-infrared (MIR) spectroscopy coupled with supervised machine learning could constitute an alternative method for rapid malaria screening, directly from dried human blood spots. Methods: Filter papers containing dried blood spots (DBS) were obtained from a cross-sectional malaria survey in 12 wards in southeastern Tanzania in 2018/19. The DBS were scanned using attenuated total reflection-Fourier Transform Infrared (ATR-FTIR) spectrometer to obtain high-resolution MIR spectra in the range 4000 cm−1 to 500 cm−1. The spectra were cleaned to compensate for atmospheric water vapour and CO2 interference bands and used to train different classification algorithms to distinguish between malaria-positive and malaria-negative DBS papers based on PCR test results as reference. The analysis considered 296 individuals, including 123 PCR-confirmed malaria positives and 173 negatives. Model training was done using 80% of the dataset, after which the best-fitting model was optimized by bootstrapping of 80/20 train/test-stratified splits. The trained models were evaluated by predicting Plasmodium falciparum positivity in the 20% validation set of DBS. Results: Logistic regression was the best-performing model. Considering PCR as reference, the models attained overall accuracies of 92% for predicting P. falciparum infections (specificity = 91.7%; sensitivity = 92.8%) and 85% for predicting mixed infections of P. falciparum and Plasmodium ovale (specificity = 85%, sensitivity = 85%) in the field-collected specimen. Conclusion: These results demonstrate that mid-infrared spectroscopy coupled with supervised machine learning (MIR-ML) could be used to screen for malaria parasites in human DBS. The approach could have potential for rapid and high-throughput screening of Plasmodium in both non-clinical settings (e.g., field surveys) and clinical settings (diagnosis to aid case management). However, before the approach can be used, we need additional field validation in other study sites with different parasite populations, and in-depth evaluation of the biological basis of the MIR signals. Improving the classification algorithms, and model training on larger datasets could also improve specificity and sensitivity. The MIR-ML spectroscopy system is physically robust, low-cost, and requires minimum maintenance

Fine-scale distribution of malaria mosquitoes biting or resting outside human dwellings in three low-altitude Tanzanian villages.

BACKGROUND: While malaria transmission in Africa still happens primarily inside houses, there is a substantial proportion of Anopheles mosquitoes that bite or rest outdoors. This situation may compromise the performance of indoor insecticidal interventions such as insecticide-treated nets (ITNs). This study investigated the distribution of malaria mosquitoes biting or resting outside dwellings in three low-altitude villages in south-eastern Tanzania. The likelihood of malaria infections outdoors was also assessed. METHODS: Nightly trapping was done outdoors for 12 months to collect resting mosquitoes (using resting bucket traps) and host-seeking mosquitoes (using odour-baited Suna® traps). The mosquitoes were sorted by species and physiological states. Pooled samples of Anopheles were tested to estimate proportions infected with Plasmodium falciparum parasites, estimate proportions carrying human blood as opposed to other vertebrate blood and identify sibling species in the Anopheles gambiae complex and An. funestus group. Environmental and anthropogenic factors were observed and recorded within 100 meters from each trapping positions. Generalised additive models were used to investigate relationships between these variables and vector densities, produce predictive maps of expected abundance and compare outcomes within and between villages. RESULTS: A high degree of fine-scale heterogeneity in Anopheles densities was observed between and within villages. Water bodies covered with vegetation were associated with 22% higher densities of An. arabiensis and 51% lower densities of An. funestus. Increasing densities of houses and people outdoors were both associated with reduced densities of An. arabiensis and An. funestus. Vector densities were highest around the end of the rainy season and beginning of the dry seasons. More than half (14) 58.3% of blood-fed An. arabiensis had bovine blood, (6) 25% had human blood. None of the Anopheles mosquitoes caught outdoors was found infected with malaria parasites. CONCLUSION: Outdoor densities of both host-seeking and resting Anopheles mosquitoes had significant heterogeneities between and within villages, and were influenced by multiple environmental and anthropogenic factors. Despite the high Anopheles densities outside dwellings, the substantial proportion of non-human blood-meals and absence of malaria-infected mosquitoes after 12 months of nightly trapping suggests very low-levels of outdoor malaria transmission in these villages

"In starvation, a bone can also be meat": a mixed methods evaluation of factors associated with discarding of long-lasting insecticidal nets in Bagamoyo, Tanzania.

BACKGROUND: Between 2000 and 2019, more than 1.8 billion long-lasting insecticidal nets (LLINs) were distributed in Africa. While the insecticidal durability of LLINs is around 3 years, nets are commonly discarded 2 years post distribution. This study investigated the factors associated with the decision of users to discard LLINs. METHODS: A mixed-method sequential explanatory approach using a structured questionnaire followed by focus group discussions (FGDs) to collect information on experiences, views, reasons, how and when LLINs are discarded. Out of 6,526 households that responded to the questionnaire of LLINs durability trial, 160 households were randomly selected from the households in four villages in Bagamoyo Tanzania for FGDs but only 155 households participated in the FGDs. Five of the household representatives couldn't participate due to unexpected circumstances. A total of sixteen FGDs each comprising of 8-10 adults were conducted; older women (40-60 years), older men (40-60 years), younger women (18-39 years), younger men (18-39 years). During the FGDs, participants visually inspected seven samples of LLINs that were "too-torn" based on Proportionate Hole Index recommended by the World Health Organization (WHO) guidelines on LLIN testing, the nets were brought to the discussion and participants had to determine if such LLINs were to be kept or discarded. The study assessed responses from the same participants that attended FGD and also responded to the structured questionnaire, 117 participants fulfilled the criteria, thus data from only 117 participants are analysed in this study. RESULTS: In FGDs, integrity of LLIN influenced the decision to discard or keep a net. Those of older age, women, and householders with lower income were more likely to classify a WHO "too-torn" net as "good". The common methods used to discard LLINs were burning and burying. The findings were seen in the quantitative analysis. For every additional hole, the odds of discarding a WHO "too-torn" LLIN increased [OR = 1.05 (95%CI (1.04-1.07)), p < 0.001]. Younger age group [OR = 4.97 (95%CI (3.25-7.32)), p < 0.001], male-headed households [OR = 6.85 (95%CI (4.44 -10.59)), p < 0.001], and wealthy households [OR = 3.88 (95%CI (2.33-6.46)), p < 0.001] were more likely to discard LLINs. CONCLUSION: Integrity of LLIN was the main determinant for discarding or keeping LLINs and the decision to discard the net is associated with socioeconomic status of the household, and the age and gender of respondents. WHO "too torn" nets are encouraged to be used instead of none until replacement, and disposal of nets should be based on recommendation