Pyrethroid Resistance Intensity in Anopheles gambiae s.l. from Different Agricultural Production Zones in Benin, West Africa.

Agricultural production activities usually occur in Benin with the use of a huge amount of insecticides including pyrethroids for pest control. It is therefore important to regularly monitor pyrethroid resistance intensity in Anopheles gambiae s.l., the main malaria vector. This study was conducted in cereal, cotton, rice growing, and urban market gardening areas throughout the country in 2018 and 2019. Females An. gambiae s.l. field-collected as larvae were exposed to deltamethrin 1 × (0.05%), 2 × (0.1%), 5 × (0.25%), and 10 × (0.5%) and permethrin 1 × (0.75%), 2 × (1.5%), 5 × (3.75%), and 10 × (7.5%). Synergist assays were also performed using World Health Organization articles combining piperonyl butoxide (PBO) (4%) + deltamethrin 1 × and, PBO (4%) + Permethrin 1 × . Molecular species and L1014F kdr mutation were identified using PCR. Expression of metabolic enzymes was also assessed through biochemical tests. After exposure to permethrin and deltamethrin 10 × , An. gambiae s.l. displayed mortality rates <98%. Synergist assays induced significantly higher mortality rates than pyrethroids alone (p < 0.05). An. gambiae s.l. complex was composed of An. gambiae s.s., Anopheles coluzzii, and Anopheles arabiensis, with mean frequency of the L1014F kdr mutation >75%. Overexpression of nonspecific α and ß esterases was observed in the cereal, cotton, and urban market gardening areas, while an overexpression of mixed function oxidases was observed in the cotton and rice growing areas. Overall, An. gambiae s.l. showed high resistance intensity to both deltamethrin and permethrin. The synergist and biochemical tests performed suggest that PBO long-lasting insecticidal nets may provide a greater control of pyrethroid-resistant mosquitoes.

Coverage, Usage, Physical Integrity, and Bio-efficacy of Olyset Nets in the Plateau Region, South Benin Following the 2011 Nationwide Distribution.

The present study investigated in 8 villages of the Plateau region the coverage, usage, physical integrity, and bio-efficacy of the Olyset nets distributed nationwide by the Benin's National Malaria Control Programme in July 2011. The questionnaire administered as well as the observations made in the households allowed estimating the coverage and usage rates of the 2011 Olyset nets. While their physical integrity was assessed through standard WHO methodology, their bio-efficacy was evaluated through gas chromatography, and WHO cone testing performed with the Kisumu susceptible strain. Mosquito collections through human landing catches (HLCs) were also performed in torn nets to assess if a loss of protection of sleepers occurred as the nets fabric integrity got more damaged. Nine months postdistribution, the coverage and usage rates of the 2011 Olyset nets were 67.4% (95% CI: 65.8-68.9) and 73.3% (95% CI: 70.7-75.8) respectively. About 28% of the 2011 Olyset nets were torn. A drastic drop of the insecticide quantity on the fibers of the nets [from 7.08 µg (95% CI: 5.74-8.42) to 0.2 µg (95% CI: 0.01-0.38)] as well as mortality rates <80% were observed with most nets evaluated. Moreover, the biting rates of An. gambiae s.l. (Diptera: Culicidae) inside torn nets increased in line with their fabric integrity loss. These data support the conclusion that future deployment of nets in the field must be strengthened by community sensitization on their correct use in order to postpone as much as possible appearance of holes and loss of insecticidal activity and encourage repairing of torn nets.

Physical integrity and survivorship of long-lasting insecticidal nets distributed to households of the same socio-cultural community in Benin, West Africa.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are designed to survive and sustain their physical barrier for 3 years in household conditions. However, studies have shown that most of these nets are usually torn or no longer present in the households in less than 3 years. This study was initiated in Benin to compare the survivorship and physical integrity of seven types of LLINs in a same socio-geographic area. METHODS: In August 2017, 1890 households were selected in 9 villages in the municipality of Zagnanado in central Benin. Each one of the selected households received one of the seven LLIN products: Aspirational®, DawaPlus® 2.0, OlysetNet®, PermaNet® 2.0, PermaNet® 3.0, Royal Sentry® and Yorkool®. Overall, 270 LLINs of each type were freely distributed in Zagnanado, at a rate of 30 LLINs per type per village. These bed nets have been monitored and evaluated every 6 months to identify the most resilient and preferred LLINs in the community. Net survivorship was assessed using the rate of net loss and physical condition. RESULTS: The survivorship of all types of LLIN was estimated at 92% (95% CI 90.33-92.96) after 6 months and 70% (95% CI 67.25-71.81) after a year of use. At 12 months, all bed nets monitored were below the NetCalc model threshold of 92.8% for an LLIN with a lifespan of 3 years. Only 1.73% of all types of LLIN had a visible loss of integrity after 6 months with a median proportionate hole index (PHI) estimated at zero. The percentage significantly increased after 12 months with 10.41% of damaged nets (all types of LLINs). The median PHI for each brand of net was 23, 196, 141, 23, 23, 121 and 72, respectively for Aspirational®, DawaPlus® 2.0, OlysetNet®, PermaNet® 2.0, PermaNet® 3.0, Royal Sentry® and Yorkool®. A significant difference was noted between the PHI at 6 and 12 months (p < 0.0001). After 12 months, the DawaPlus®2.0, OlysetNet® and Royal Sentry® suffered significantly more damage compared to the others (p < 0.001). CONCLUSION: The results of this study showed that after a year of use, the survivorship of the 7 LLIN products in households was lower than expected. However, all the LLIN products successfully met WHO standards for physical integrity after 12 months of use. The monitoring continues. The next steps will help to identify the most sustainable LLINs.

Lessons learned, challenges and outlooks for decision-making after a decade of experience monitoring the impact of indoor residual spraying in Benin, West Africa.

BACKGROUND: Since 2008, Indoor Residual Spraying (IRS) has been performed in Benin in 19 districts, including 4 in southern Benin, 9 in Atacora, and 8 in Atacora, Alibori and Donga in northern Benin. However, Benin still struggles with questions about IRS cost-benefit and epidemiological impact. Lessons learned and challenges from 10 years of IRS in Benin to be shared with the stakeholders involved in vector control implementation for decision-making. METHODS: Entomological parameters have been assessed entomological parameters in IRS communes since 2008. In all IRS intervention communes, decreases in human biting rate (HBR) of Anopheles gambiae, blood feeding inhibition and entomological inoculation rate (EIR) as compared to control district have been measured. RESULTS: EIR was reduced by 80-90%, which is encouraging, but should be observed with caution because: (i) the reduction may be insufficient to decrease epidemiological indicators given that the residual EIR in IRS districts is still higher than it is in some regions of stable malaria; (ii) the reduction in EIR is based on comparisons with control communes, but it is difficult to select control areas with the same environmental characteristics as intervention areas; (iii) despite the reduction, half of all mosquitoes that entered IRS-treated houses succeeded in taking human blood meals. Further, there are behaviours among Benin's population that limit IRS efficacy, including recent data showing that > 90% of people are not protected by IRS between 7 and 10 p.m. This is due to the fact that they remain outdoors and that most people are not protected from mosquito bites after 10 p.m. because they either sleep outdoors without IRS protection or indoors without an ITN. Moreover, people have large amounts of clothing hanging on walls where mosquitoes can rest instead of IRS-treated walls. Finally, other components are important to consider in implementing IRS among which: (i) Vector resistance management strategies are sometimes poorly understood; this is actually different from the need to replace one insecticide with another after the emergence of resistance; (ii) African countries should prepare to finance IRS themselves. CONCLUSION: To curtail residual malaria transmission, additional interventions able to target vectors escaping IRS should be prioritized.

Resistance status of Anopheles gambiae s.l. to insecticides following the 2011 mass distribution campaign of long-lasting insecticidal nets (LLINs) in the Plateau Department, south-eastern Benin.

BACKGROUND: In 2011, Benin's National Malaria Control Programme (NMCP) organized a nationwide mass distribution campaign of LLINs throughout the country. Following this intervention, it was important to assess whether the level of susceptibility of malaria vectors to insecticides had remained the same as compared to the pre-intervention period. The current study investigated this. METHODS: Larval collections were conducted in Ifangni, Sakété, Pobè and Kétou districts located in Plateau department, Southeastern Benin before (2009) and after (2012-2013) LLIN distribution. Anopheles gambiae sensu lato (s.l.) larvae from the 4 study districts were reared to adulthood and WHO susceptibility tests were conducted. The insecticides tested were deltamethrin (0.05%), permethrin (0.75%), bendiocarb (0.1%) and DDT (4%). Molecular species identification as well as, the characterization of the kdr L1014F mutation were also performed in the An. gambiae s.l. complex using PCR method. RESULTS: Overall, a significant decrease in mortality rates of An. gambiae s.l. to deltamethrin (0.05%), permethrin (0.75%) and DDT (4%) was observed post-LLIN distribution, respectively: (100% vs 80.9%, p < 0.0001), (77.5% vs 70%, p = 0.01) and, (47.8% vs 4.4%, p < 0.0001). By contrast, susceptibility of vectors to bendiocarb (0.1%) remained the same (100% mortality in the WHO susceptibility tube tests) pre- and post-intervention. An increase in the kdr L1014F frequency was observed post-LLIN distribution [F(kdr) = 0.91)] compared to the pre-intervention period [F(kdr) = 0.56], p < 0.0001. Anopheles coluzzii and An. gambiae were the two molecular species identified in the study area. CONCLUSION: The decrease susceptibility to pyrethroids and DDT as well as, the increase in the frequency of the kdr L1014F mutation after the intervention stressed at the time, the need for the development and implementation of effective insecticide resistance management strategies. At present, an update of the vectors resistance status in the area is also necessary for decision-making.

Baseline entomologic data on malaria transmission in prelude to an indoor residual spraying intervention in the regions of Alibori and Donga, Northern Benin, West Africa.

BACKGROUND: Despite the success of indoor residual insecticide spraying (IRS) in Africa, particularly in Benin, some gaps of information need to be filled to optimize the effectiveness of this intervention in the perspective of the country's effort to eliminate malaria. In anticipation to the 2018 IRS campaign in two targeted regions of northern Benin, this study aimed, to collect baseline information on vector composition, spatio-temporal variation and peak malaria transmission in the Alibori and Donga, two targeted regions of northern Benin. Information collected will help to better plan the implementation and later on the impact assessment of this IRS campaign. METHODS: The study was carried out in four districts of the two IRS targeted regions of northern Benin. Human landing catches and pyrethrum spray catches protocols were used to assess the biting rate (HBR) and, biting/resting behaviour of malaria vector populations. After morphological identification of collected Anopheles, the heads and thoraxes of Anopheles gambiae sensu lato (s.l.) were analysed by the ELISA CSP tests to estimate the sporozoite index (SI). The entomological inoculation rate was calculated as the product of mosquito biting rate (HBR) and the SI. RESULTS: The biting rates of An. gambiae s.l., the major vector in this study sites, varied significantly from region to region. It was higher: in rural than in urban areas, in rainy season than in dry season, indoors than outdoors. Overall, SI was comparable between sites. The highest EIRs were observed in the Donga region (16.84 infectious bites/man/month in Djougou district and 17.64 infectious bites/man/month in Copargo district) and the lowest in the Alibori region (10.74 infectious bites/man/month at Kandi district and 11.04 infectious bites/man/month at Gogounou district). CONCLUSION: This study showed the heterogeneous and various nature of malaria epidemiology in Northern Benin. Indeed, the epidemiological profile of malaria transmission in the Alibori and Donga regions is made of a single season of transmission interrupted by a dry season. This period of transmission is relatively longer in Donga region than in Alibori. This information can be used to guide the extension of IRS in the Alibori and in the Donga, by primarily targeting areas with short periods of transmission, and easy to cover.

Impact of land-use on malaria transmission in the Plateau region, southeastern Benin.

BACKGROUND: The goal of the study is to investigate if local agricultural practices have an impact on malaria transmission in four villages located in the same geographical area within a radius of 15 kilometers. Among the villages, one (Itassoumba) is characterized by the presence of a large market garden and fishpond basins, the three others (Itakpako, Djohounkollé and Ko-koumolou) are characterized by traditional food-producing agriculture. METHODS: Malaria transmission was evaluated using human-landing catches, both indoors and outdoors, two nights per month for 12 months. Field collected females An. gambiae s.l. were exposed for 1 hour to 0.75% permethrin and 0.05% deltamethrin using WHO insecticide susceptibility test kits and procedures. The presence of the kdr mutation was analyzed by PCR. RESULTS: Anopheles gambiae s.s form M (93.65%), was identified as the main malaria vector. Its susceptibility level to pyrethroids was the same (p > 0.05) in all villages. kdr mutation frequencies are 81.08 in Itakpako, 85 in Itassoumba, 79.73 in Djohounkollé and 86.84 in Ko-Koumolou (p = 0.63). The entomological inoculation rate ranged from 9.62 to 21.65 infected bites of An. gambiae per human per year in Djohounkollé, Itakpako and Ko-Koumolou against 1159.62 in Itassoumba (p < 0.0001). CONCLUSION: The level of resistance of An. gambiae to pyrethroids was the same in the four villages. The heterogeneous character of malaria epidemiology was confirmed. The creation of fishponds basins and the development of market-gardening activities increased drastically the malaria transmission in Itassoumba.

Impact of operational effectiveness of long-lasting insecticidal nets (LLINs) on malaria transmission in pyrethroid-resistant areas.

BACKGROUND: A dynamic study on the transmission of malaria was conducted in two areas (R⁺ area: Low resistance area; R⁺⁺⁺ area: High resistance area) in the department of Plateau in South Eastern Benin, where the population is protected by Long Lasting Insecticidal Nets (LLINs). The aim of this study was to determine if the resistance of malaria vectors to insecticides has an impact on their behavior and on the effectiveness of LLINs in the reduction of malaria transmission. METHODS: Populations of Anopheles gambiae s.l. were sampled monthly by human landing catch in the two areas to evaluate human biting rates (HBR). Collected mosquitoes were identified morphologically and female Anopheles mosquitoes were tested for the presence of Plasmodium falciparum antigen as assessed using ELISA. The entomological inoculation rate (EIR) was also calculated (EIR = HBR x sporozoitic index [S]). We estimated the parity rate by dissecting the females of An. gambiae. Finally, window catch and spray catch were conducted in order to assess the blood feeding rate and the exophily rate of vectors. RESULTS: After 6 months of tracking the mosquito's behavior in contact with the LLINs (Olyset) in R⁺⁺⁺ and R⁺ areas, the entomological indicators of the transmission of malaria (parity rate and sporozoitic index) were similar in the two areas. Also, An. gambiae populations showed the same susceptibility to P. falciparum in both R⁺ and R⁺⁺⁺ areas. The EIR and the exophily rate are higher in R⁺ area than in R⁺⁺⁺ area. But the blood-feeding rate is lower in R⁺ area comparing to R⁺⁺⁺. CONCLUSION: The highest entomological inoculation rate observed in R⁺ area is mostly due to the strong aggressive density of An. gambiae recorded in one of the study localities. On the other hand, the highest exophily rate and the low blood-feeding rate recorded in R⁺ area compared to R⁺⁺⁺ area are not due to the resistance status of An. gambiae, but due to the differences in distribution and availability of breeding sites for Anopheles mosquitoes between areas. However, this phenomenon is not related to the resistance status, but is related to the environment instead.