Residual malaria transmission: Magnitude and drivers of persistent Plasmodium infections despite high coverage of control interventions in Burkina Faso, West Africa.

This study collected baseline data on malaria vectors to characterize the drivers and the factors of persistent malaria transmission in two villages in the western part of Burkina Faso. Mosquitoes were collected in each village using the Human landing catch and pyrethrum spray catch and identified using the morphological keys. Molecular analyses were performed for the identification of An. gambiae complex species, the detection of Plasmodium infection and kdr-995F mutation. Anopheles mosquito larvae were also collected in the same villages, reared to adult's stage for the WHO tube and cone tests performing. The physical integrity of the LLINs already used by people in each village was assessed using the proportional hole index (pHI). An. gambiae s.l. was the main malaria vector accounting for 79.82% (5560/6965) of all collected mosquitoes. The biting pattern of An. gambiae s.l. was almost constant during the survey with an early aggressiveness before 8 p.m. and later biting activity after 6 a.m. The EIR varied from 0.13 to 2.55 infected bites per human per night (average: 1.03 infected bites per human per night). An. gambiae s.l. populations were full susceptible to Chlorpyrifos-methyl (0.4%) and Malathion (5%) with high kdr-995F mutation frequencies (>0.8). The physical integrity assessment showed high proportion of good nets in Santidougou compared to those collected in Kimidougou. This study highlighted a persistence of malaria transmission despite the intense use of vector control tools as LLINs and IRS by correlating mosquito biting time and human behavior. It provided a baseline guide for the monitoring of the residual malaria transmission in sub-Saharan Africa and encouraging the development of new alternative strategies to support the current malaria control tools.

Quantifying and characterizing hourly human exposure to malaria vectors bites to address residual malaria transmission during dry and rainy seasons in rural Southwest Burkina Faso.

BACKGROUND: To sustain the efficacy of malaria vector control, the World Health Organization (WHO) recommends the combination of effective tools. Before designing and implementing additional strategies in any setting, it is critical to monitor or predict when and where transmission occurs. However, to date, very few studies have quantified the behavioural interactions between humans and Anopheles vectors in Africa. Here, we characterized residual transmission in a rural area of Burkina Faso where long lasting insecticidal nets (LLIN) are widely used. METHODS: We analysed data on both human and malaria vectors behaviours from 27 villages to measure hourly human exposure to vector bites in dry and rainy seasons using a mathematical model. We estimated the protective efficacy of LLINs and characterised where (indoors vs. outdoors) and when both LLIN users and non-users were exposed to vector bites. RESULTS: The percentage of the population who declared sleeping under a LLIN the previous night was very high regardless of the season, with an average LLIN use ranging from 92.43 to 99.89%. The use of LLIN provided > 80% protection against exposure to vector bites. The proportion of exposure for LLIN users was 29-57% after 05:00 and 0.05-12% before 20:00. More than 80% of exposure occurred indoors for LLIN users and the estimate reached 90% for children under 5 years old in the dry cold season. CONCLUSIONS: LLINs are predicted to provide considerable protection against exposure to malaria vector bites in the rural area of Diébougou. Nevertheless, LLIN users are still exposed to vector bites which occurred mostly indoors in late morning. Therefore, complementary strategies targeting indoor biting vectors in combination with LLIN are expected to be the most efficient to control residual malaria transmission in this area.

Comparative physiological plasticity to desiccation in distinct populations of the malarial mosquito Anopheles coluzzii.

BACKGROUND: In West Africa, populations of the malaria vector mosquito, Anopheles coluzzii, are seasonally exposed to strong desiccating conditions during the dry season. Their dynamics strictly follows the pace of the availability of suitable larval development sites (water collections). Accordingly, mosquitoes can reproduce all year long where permanent breeding is possible, or stop reproduction and virtually disappear at the onset of the dry season when surface water dries up, like observed in temporary habitats of dry savannah areas. This highlights the strong adaptive abilities of this mosquito species, which relies at least in part, upon physiological and molecular mechanisms of specific signatures. METHODS: Here, we analysed a range of physiological and molecular responses expressed by geographically different populations of An. coluzzii inhabiting permanent and temporary breeding sites from the north and the south-west of Burkina Faso. Four mosquito colonies, namely (i) Oursi, built from females breeding in permanent habitats of the north; (ii) Déou, from temporary northern habitats; (iii) Soumousso from south-western temporary breeding sites; and (iv) Bama, from permanent habitats of the same south-western zone, were reared in climatic chambers under contrasted environmental conditions, mimicking temperature, relative humidity and light regimen occurring in northern Burkina Faso. Female mosquitoes were analysed for the seasonal variation in their amounts of proteins, triglycerides and free-circulating metabolites. The expression level of genes coding for the adipokinetic (AKH-I) and the AKH/corazonin-related peptides (ACP) were also assessed and compared among populations and environmental conditions. RESULTS: Our analysis did not reveal an apparent pattern of physiological and molecular variations strictly correlated with either the larval ecotype or the geographical origin of the mosquitoes. However, specific distinct responses were observed among populations, suggesting that dry season survival may rely on more complex ecological parameters at a micro-habitat scale. Interestingly, the physiological and molecular data support the hypothesis that different aestivation abilities exist among populations of An. coluzzii inhabiting contrasted ecological settings. In particular, the striking metabotypes differentiation and the AKH mRNA expression level observed in females from temporary northern populations may suggest the existence of a "strong" aestivation strategy in these specimens. CONCLUSION: Our work provides insights into the physiological and molecular basis of dry and rainy season responses in An. coluzzii, and highlights the important diversity of the mechanisms involved. Such results represent key data for understanding the ecophysiological mechanisms underpinning the strong adaptive potential of this malaria vector species, which undoubtedly contributes to the spreading of mosquito distribution areas in space and time.

Monitoring Dry Season Persistence of Anopheles gambiae s.l. Populations in a Contained Semi-Field System in Southwestern Burkina Faso, West Africa.

To gain insight into the dry season survival strategies of Anopheles gambiae s.l., a new contained semi-field system was developed and used for the first time in Burkina Faso, West Africa. The system consisted of a screened greenhouse within which the local environment was reproduced, including all ecological requirements for mosquito development cycle completion. The system was seeded with the progenies of female Anopheles gambiae, Anopheles coluzzii, and Anopheles arabiensis collected in the vicinity of the greenhouse during the rainy season. After successful establishment in the semi-field system, mosquito populations were monitored over a 1-yr period by regular surveys of larval and adult specimens. We provided evidence for the persistence of adult mosquitoes throughout the dry season, in the absence of any suitable larval development site. During the hot and dry periods, adult insects were observed in artificial shelters (clay pots, building blocks, and dark corners). The mosquito population rapidly built up with the return of the rainy season in the area, when artificial breeding sites were refilled in the enclosure. However, only An. coluzzii and, later, An. arabiensis were detected in the subsequent rainy season, whereas no An. gambiae specimen was found. Our findings suggest that An. coluzzii and An. arabiensis may be able to aestivate throughout the dry season in Southwestern Burkina Faso, whereas An. gambiae might adopt a different dry-season survival strategy, such as long-distance re-colonization from distant locations. These results may have important implications for malaria control through targeted vector control interventions.

Combining two-dimensional gel electrophoresis and metabolomic data in support of dry-season survival in the two main species of the malarial mosquito Anopheles gambiae.

In dry savannahs of West-Africa, the malarial mosquitoes of the Anopheles gambiae sensu stricto complex annually survive the harsh desiccating conditions of the dry season. However, the physiological and biochemical mechanisms underlying how these mosquitoes survive such desiccating conditions are still undefined, and controversial. In this context, we provide the first work examining both proteomic and metabolomic changes in the two molecular forms of A. gambiae s.s (M and S forms) experimentally exposed to the rainy and dry season conditions as they experience in the field. Protein abundances of the mosquitoes were measured using a two-dimensional fluorescence difference gel electrophoresis (2D DIGE) coupled with a matrix-assisted laser desorption/ionisation-time of flight (MALDI-TOF) and tandem mass spectrometry (MS) for protein identification. These assays were conducted by Applied Biomics (http://www.appliedbiomics.com, Applied Biomics, Inc. Hayward, CA, USA), and the mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository with the dataset identifier PXD000294. The metabolomic analysis was conducted using both Acquity UPLC(®) system (for amino acid identification), and a gas-chromatography-mass spectrometry platform (for sugars identification). Metabolomic fingerprintings were assessed in the University of Rennes 1, UMR CNRS 6553 EcoBio (France). A detailed interpretation of the obtained data can be found in Hidalgo et al. (2014) [1] (Journal of Insect Physiology (2014)).

Novel insights into the metabolic and biochemical underpinnings assisting dry-season survival in female malaria mosquitoes of the Anopheles gambiae complex.

The mechanisms by which Anopheles gambiae mosquitoes survive the desiccating conditions of the dry season in Africa and are able to readily transmit malaria soon after the rains start remain largely unknown. The desiccation tolerance and resistance of female An. gambiae M and S reared in contrasting environmental conditions reflecting the onset of dry season ("ods") and the rainy season ("rs") was determined by monitoring their survival and body water loss in response to low relative humidity. Furthermore, we investigated the degree to which the physiology of 1-h and 24-h-old females is altered at "ods" by examining and comparing their quantitative metabotypes and proteotypes with conspecifics exposed to "rs" conditions. Results showed that distinct biochemical rearrangements occurred soon after emergence in female mosquitoes that enhance survival and limit water loss under dry conditions. In particular, three amino acids (phenylalanine, tyrosine, and valine) playing a pivotal role in cuticle permeability decreased significantly from the 1-h to 24-h-old females, regardless of the experimental conditions. However, these amino acids were present in higher amounts in 1-h-old female An. gambiae M reared under "ods" whereas no such seasonal difference was reported in S ones. Together with the 1.28- to 2.84-fold increased expression of cuticular proteins 70 and 117, our data suggests that cuticle composition, rigidity and permeability were adjusted at "ods". Increased expression of enzymes involved in glycogenolytic and proteolytic processes were found in both forms at "ods". Moreover, 1-h-old S forms were characterised by elevated amounts of glycogen phosphorylase, isocitrate dehydrogenase, and citrate synthase, suggesting an increase of energetic demand in these females at "ods".

Metabolomic and ecdysteroid variations in Anopheles gambiae s.l. mosquitoes exposed to the stressful conditions of the dry season in Burkina Faso, West Africa.

This study explored the metabolic adjustments prompted by a switch between the rainy and dry season conditions in the African malaria mosquitoes Anopheles gambiae (M and S molecular forms) and Anopheles arabiensis. Mosquitoes were reared in contrasted experimental conditions reflecting environmental variation in Burkina Faso. Thirty-five metabolites (including sugars, polyols, and amino acids) were monitored in newly emerged males and females, and their ecdysteroid titers were determined. Metabolomic signatures were remarkably similar across species, when specimens of same age and sex were reared under identical experimental conditions. In males and females, amino acids (including glycine, leucine, phenylanine, serine, threonine, and valine) were accumulated in 1-h-old mosquitoes, then decreased 24 h after emergence, probably reflecting adult maturation and the amino acid-consuming process of cuticle sclerotisation. In turn, elevated amounts of alanine and proline in 24-h-old mosquitoes may assist the development of flight ability. Lower concentration of tricarboxylic acid cycle intermediates and isoleucine characterized older females reared under dry season conditions, suggesting metabolic and reproduction depression. In all cases, ecdysteroid concentration was much higher in males than in females, with significant seasonal variation in males. This might reflect a unique role of these hormones in shaping reproductive strategies and population demography in the An. gambiae s.l. species complex, further contributing to local adaptation in a highly fluctuating environment.

Swarming behaviour in natural populations of Anopheles gambiae and An. coluzzii: review of 4 years survey in rural areas of sympatry, Burkina Faso (West Africa).

The swarming behaviour of natural populations of Anopheles gambiae and An. coluzzii (formerly known as An. gambiae S and M forms, respectively) were investigated through longitudinal surveys conducted between July 2006 and October 2009 in two rural areas of south-western Burkina Faso where these forms are sympatric. In both sites, the majority of swarms were recorded above visual markers localised among houses. In Soumousso, a wooded area of savannah, 108 pairs caught in copula from 205 swarms were sampled; in VK7, a rice growing area, 491 couples from 250 swarms were sampled. If segregated swarms were the norm in both sites, many visual markers were shared by the two forms of An. gambiae. Furthermore, mixed swarms were collected annually in frequencies varying from one site to another, though no mixed inseminations were recorded, corroborating the low hybrid rate previously reported in the field. The occurrence of inter-specific mate-recognition mechanisms, which allow individuals to avoid hybridisation, is discussed.

Occurrence of natural Anopheles arabiensis swarms in an urban area of Bobo-Dioulasso City, Burkina Faso, West Africa.

The swarming behaviour of natural populations of Anopheles arabiensis was investigated by conducting transect surveys on 10 consecutive days, around dusk, from March to April and from September to October 2012 in Dioulassoba, a district of Bobo-Dioulasso city in Burkina Faso (West Africa). Swarms were observed outside, around identified larval breeding sites on the banks of the Houet River, as well as in the open-air courtyards found at the centre of many homes in the region. Swarms were found to occur in open sunlit spaces, mostly located above physical or visual cues somehow visually distinct from the surrounding area. Overall 67 and 78 swarms were observed, respectively, during the dry season (March-April) and the rainy season (September-October) of 2012, between 1.5m and 4.5m above the ground at their centre. 964 mosquitoes were collected and analysed from dry season swarms, of which most were male, and all were An. arabiensis, as were the few resting mosquitoes collected indoors. Larvae collected from breeding sites found on the banks of the Houet River mostly consisted of An. arabiensis and only a minority of Anopheles coluzzii (formerly identified as An. gambiae M form). Of 1694 mosquitoes analysed from 78 swarms in the rainy season collections, a few An. gambiae (formerly known as An. gambiae S form) males were identified, and the remainders were An. arabiensis. The majority of larvae collected during the wet season from the same breeding sites were identified as An. arabiensis followed by An. coluzzii and An. gambiae. The same pattern of species composition was observed in resting mosquitoes, though the proportion of An. arabiensis was less overwhelming. These data support the conclusion that An. arabiensis is the most prevalent species in this area, though the difference in species composition when using different population sampling techniques is noteworthy. Further studies are required for more detailed investigations of male dispersal, feeding behaviour and mating patterns in this urban setting.

Assortative mating in mixed swarms of the mosquito Anopheles gambiae s.s. M and S molecular forms, in Burkina Faso, West Africa.

The molecular form composition of Anopheles gambiae Giles s.s. (Diptera: Culicidae) mating swarms and the associated mating pairs (copulae) were investigated during two rainy seasons (July to October, 2005 and July to November, 2006) in the villages of Soumousso and Vallée du Kou (VK7). Although the habitats of these villages differ markedly, sympatric populations of M and S molecular forms of An. gambiae s.s. occur in both places periodically. The main aim was to assess the degree to which these molecular forms mate assortatively. In Soumousso, a wooded savannah habitat, the majority of swarm samples consisted of only S-form males (21/28), although a few M-form males were found in mixed M- and S-form swarms. In VK7, a rice growing area, the majority of swarm samples consisted of only M-form males (38/62), until October and November 2006, when there were nearly as many mixed-form as single-form swarms. Overall, ∼60% of M- and S-form swarms were temporally or spatially segregated; the two forms were effectively prevented from encountering each other. Of the remaining 40% of swarms, however, only about half were single-form and the rest were mixed-form. Of the 33 copulae collected from mixed-form swarms, only four were mixed-form pairs, significantly fewer than expected by random pairing between forms (χ(2) = 10.34, d.f. = 2, P < 0.01). Finally, all specimens of inseminated females were of the same form as the sperm contained within their spermatheca (n = 91), even for the four mixed-form copulae. These findings indicate that assortative mating occurs within mixed-form swarms, mediated most probably by close-range mate recognition cues.

Distribution of insensitive acetylcholinesterase (ace-1R) in Anopheles gambiae s.l. populations from Burkina Faso (West Africa).

OBJECTIVE: To investigate through countrywide sampling at 20 localities across the three different agro-climatic zones of Burkina Faso, the distribution of the acetylcholinesterase insensitive mutation ace-1(R), which confers resistance to organophosphates (OP) and carbamates (CM) insecticides in An. gambiae s.l. METHODS: Adult mosquitoes were collected by indoor aerosol spraying from August to October 2006. Specimens were identified to species by polymerase chain reaction (PCR) assay and characterized for the ace-1(R) mutation using a PCR-restriction fragment length polymorphism diagnostic. RESULTS: Collected mosquitoes were a mixture of An. gambiae s.s. and An. arabiensis across the Sudan (98.3%vs. 1.7%), Sudan-sahelian (78.6%vs. 21.4%) and the Sahel (91.5%vs. 8.5%) ecotypes. The An. gambiae S-form predominated in the Sudan sites from the West (69%vs. 31% for the M form) but was not found in the Sahel (100% M form). The ace-1(R) mutation was dispersed throughout the Sudan and Sudan-sahelian localities at moderate frequency (<50%) but was absent in the Sahel. It was far more prevalent in S form than M form mosquitoes (0.32 for the S form vs. 0.036 for the M form). No An. arabiensis was detected carrying the mutation. The geographic distribution of ace-1(R) in the Sudan and Sudan-sahelian correlated with the cotton growing areas dispersed throughout the two climatic zones. CONCLUSIONS: These results have special significance as OP and CM insecticides have been proposed as alternatives or additions to pyrethroids which are currently used exclusively in many vector control programmes.

Distribution of pyrethroid and DDT resistance and the L1014F kdr mutation in Anopheles gambiae s.l. from Burkina Faso (West Africa).

This study reports on the distribution of pyrethroid and DDT resistance and the L1014F knockdown resistance (kdr) mutation in Anopheles gambiae s.l. populations from 21 localities in three different climatic zones of Burkina Faso from August to October 2006. The susceptibility of these populations was assessed by bioassay using DDT (4%), permethrin (1%) and deltamethrin (0.05%). Anophelesgambiae were resistant to both permethrin and DDT in the Sudanian regions but were susceptible in the central and sahelian areas and susceptible to deltamethrin at all sites except Orodara, although mortality values in some populations were close to the resistance threshold. The kdr frequency varied from 0.4 to 0.97 in populations from the Sudanian region and was lower in populations from the Sudano-sahelian and sahelian areas (0.047 to 0.54). Compared to the last survey of kdr in An. gambiae populations conducted in 2000, the kdr frequency did not differ in the S form but had increased in the M form (0.6), with an extended distribution into the Sudano-sahelian region. The frequency of kdr was also found to have increased in An. arabiensis populations (0.28), where it was formerly reported in only a single specimen. These results have practical significance for malaria vector control programs.

Year to year and seasonal variations in vector bionomics and malaria transmission in a humid savannah village in west Burkina Faso.

A longitudinal entomological study was carried out from 1999 to 2001 in Lena, a humid savannah village in the western region of Burkina Faso in order to establish malaria vector bionomics and the dynamics of malaria transmission. In the first year, malaria transmission was mainly due to An. gambiae s.s., but during the two later years was due to An. funestus, which were observed in high frequency towards the end of the rainy season. PCR identification of samples of An. gambiae s.1. showed 93% to be An. gambiae s.s. and 7% An. arabiensis. An. funestus constituting more than 60% of the vectors were identified in PCR as An. funestus s.s. The persistence of intense vectorial activity in this village was probably due to the road building in a swampy area creating a semi-permanent swamp that provided large sites for larval mosquitoes. These swampy sites seemed to be more favorable for An. funestus than for An. gambiae s.s. Thus, land development must be monitored and subjected to planning to minimize vector proliferation. Such a system of planning could lead to the restriction or even elimination of the swamp that is the source of larvae developing in the heart of the village.

Distribution of the members of Anopheles gambiae and pyrethroid knock-down resistance gene (kdr) in Guinea-Bissau, West Africa.

An entomological survey conducted in 2002 in Guinea Bissau aimed i) to study the distribution of the members of Anopheles gambiae Giles complex (Diptera: Culicidae) throughout four ecological areas extended from mangrove to savannah ii) to evaluate the insecticide susceptibility status of these malaria vectors exposed to permethrin 0.75% and DDT4%, and finally iii) to investigate the occurrence and the spread of the Leu-Phe knock down resistance (kdr) gene associated with pyrethroid and DDT resistance within these vector populations. Adult female mosquitoes issued from indoor morning collections were tested using WHO procedures, test kits and impregnated papers to assess their insecticide susceptibility status. Tested specimens were identified by PCR assays and characterized for the kdr gene. Malaria vectors were mainly dominated elsewhere by An. gambiae s.s. (both S and M molecular forms) living in sympatry with low proportion of An. melas in the littoral. An. gambiae s.s. tested populations were fully susceptible both to permethrin 0.75% and to DDT 4% irrespective to their location and ecotypes. The Leu-Phe kdr mutation was detected at low frequency only in two sites respectively urban (Bissau) and Guinea-savannah (Gabu) areas. It occurred only in the S molecular form in Gabu (at the frequency of 0.14) and both in the S and M molecular forms in Bissau at the frequency of 0.06 and 0.02 respectively. These results suggested that the populations of An. gambiae s.s., the most frequent malaria vector in Guinea Bissau, still remain cross-susceptible to pyrethroids and DDT This susceptibility status and the frequency of resistance mechanism such as the kdr mutation must be monitored in the future particularly in the urban and savannah areas with continuous and intensive use of insecticides.

Anopheles funestus (Diptera: Culicidae) in a humid savannah area of western Burkina Faso: bionomics, insecticide resistance status, and role in malaria transmission.

An entomological survey was carried out in three humid savannah sites of western Burkina Faso (Bama, Lena, and Soumousso) to (1) update the taxonomy of the Anopheles funestus Giles group, (2) examine the role of each species in malaria transmission, (3) characterize the insecticide resistance status of this malaria vector, and (4) determine the distribution of An. funestus chromosomal forms in these areas. Polymerase chain reaction identification of the members showed the occurrence of An. leesoni Evans in Lena and An. rivulorum-like in Soumousso in addition to An. funestus s.s. Malaria transmission was ensured mainly by An. funestus s.s. both in Soumousso and Lena and by An. gambiae s.s. Giles in Bama, the rice-growing area. The insecticide resistance status performed only on An. funestus indicated that this mosquito was susceptible to pyrethroids irrespective of the study area, but it was resistant to dieldrin. Furthermore, the occurrence of the two chromosomal forms of An. funestus, namely, Kiribina and Folonzo, seemed to follow ecological setups where Kiribina predominated in the irrigated area and Folonzo was more frequent in classic savannah. This study revealed that the problematic of An. funestus taxonomy was closer to that of An. gambiae requiring more structured studies to understand its genetic ecology.

The indoor use of plastic sheeting pre-impregnated with insecticide for control of malaria vectors.

OBJECTIVE: To evaluate the efficacy of permethrin-treated plastic sheeting (ITPS) applied as a lining to the ceiling or walls of rooms against pyrethroid-susceptible and pyrethroid-resistant Anopheles gambiae. METHOD: Overnight tests were carried out in veranda-trap experimental huts in Vallée du Kou, where two molecular forms of A. gambiae, S and M, occur. The S form is mostly pyrethroid resistant due to the kdr mechanism, and the M form is mostly kdr susceptible. A variety of ITPS covered surfaces were tested, ranging from ceiling only to all walls plus ceiling covered. RESULTS: ITPS had a major effect on the mortality of mosquitoes, the proportion killed being dependent upon the surface area covered. Homozygotes for kdr resistance showed lower rates of mortality than did heterozygotes or homozygotes for susceptibility. Deterred entry of mosquitoes and inhibition of blood feeding were also correlated with surface area covered. The mode of action and efficacy of ITPS seems to bear closer resemblance to that induced by indoor residual spraying (IRS) than to that induced by insecticide-treated nets. CONCLUSIONS: ITPS might be conceived as being equivalent to long-lasting or permanent IRS but without some of the operational constraints normally associated with spraying. High coverage of ITPS could potentially have a mass population effect on mosquitoes and give rise to long-term community protection against malaria. A phase III trial is justified to assess the acceptability of ITPS and its efficacy against malaria.

First biologic and genetic characterization of Toxoplasma gondii isolates from chickens from Africa (Democratic Republic of Congo, Mali, Burkina Faso, and Kenya).

The prevalence of Toxoplasma gondii in free-ranging chickens (Gallus domesticus) is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. In the present study, prevalence of T. gondii in chickens from Democratic Republic of Congo, Mali, Burkina Faso, and Kenya is reported. The prevalence of T. gondii antibodies in sera of 50 free-range chickens from Congo was 50% based on the modified agglutination test (MAT); antibody titers were 1:5 in 7, 1:10 in 7, 1:20 in 6, 1:40 in 1, and 1:160 or more in 4 chickens. Hearts, pectoral muscles, and brains of 11 chickens with titers of 1:20 or more were bioassayed individually in mice; T. gondii was isolated from 9, from the hearts of 9, brains of 3, and muscles of 3 chickens. Tissues of each of the 14 chickens with titers of 1:5 or 1:10 were pooled and bioassayed in mice; T. gondii was isolated from 1 chicken with a titer of 1:10. Tissues from the remaining 25 seronegative chickens were pooled and fed to 1 T. gondii-free cat. Feces of the cat were examined for oocysts, but none was seen. The results indicate that T. gondii localizes in the hearts more often than in other tissues of naturally infected chickens. Genotyping of these 10 isolates using the SAG2 locus indicated that 8 were isolates were type III, 1 was type II, and 1 was type I. Two isolates (1 type I and 1 type III) were virulent for mice. Toxoplasma gondii was isolated by mouse bioassay from a pool of brains and hearts of 5 of 48 chickens from Mali and 1 of 40 chickens from Burkina Faso; all 6 isolates were avirulent for mice. Genetically, 4 isolates were type III and 2 were type II. Sera were not available from chickens from Mali and Burkina Faso. Toxoplasma gondii antibodies (MAT 100 or more) were found in 4 of 30 chickens from Kenya, and T. gondii was isolated from the brain of 1 of 4 seropositive chickens; this strain was avirulent for mice and was type II. This is the first report on isolation and genotyping of T. gondii from any source from these 4 countries in Africa.

The spread of the Leu-Phe kdr mutation through Anopheles gambiae complex in Burkina Faso: genetic introgression and de novo phenomena.

During extensive sampling in Burkina Faso and other African countries, the Leu-Phe mutation producing the kdr pyrethroid resistance phenotype was reported in both Anopheles gambiae ss and A. arabiensis. This mutation was widely distributed at high frequency in the molecular S form of A. gambiae while it has been observed at a very low frequency in both the molecular M form and A. arabiensis in Burkina Faso. While the mutation in the M form is inherited through an introgression from the S form, its occurrence is a new and independent mutation event in A. arabiensis. Three nucleotides in the upstream intron of the kdr mutation differentiated A. arabiensis from A. gambiae ss and these specific nucleotides were associated with kdr mutation in A. arabiensis. Ecological divergences which facilitated the spread of the kdr mutation within the complex of A. gambiae ss in West Africa, are discussed.

First report of a kdr mutation in Anopheles arabiensis from Burkina Faso, West Africa.

The leu-phe kdr mutation was detected in a specimen of Anopheles arabiensis during an extensive survey of pyrethroid resistance in An. gambiae s.l. in Burkina Faso. The detection of this mutation in An. arabiensis, which had so far been observed only in An. gambiae s.s., is important at both epidemiologic and fundamental levels. It can be useful to understand the history of this gene throughout the range of An. gambiae complex.

Natural swarming behaviour of the molecular M form of Anopheles gambiae.

In Anopheles gambiae, as in most species of mosquitoes, mating is initiated in flight. The males aggregate in aerial swarms and conspecific females individually fly to these swarms where they mate with males. In this study, we investigated the swarming behaviour of A. gambiae and conducted 2 surveys in the rice field area of the Vallée du Kou in Burkina Faso in 1999 and 2002. A high number of anopheline mosquitoes were observed in this area and both molecular M and S forms of A. gambiae were found in sympatry. Swarms formed a few minutes after sunset in different places and no obvious markers were associated with their occurrence. However, swarms occurred close to cow herds generally in open flat areas, 2-3 m above the ground. Overall, 2829 anopheline mosquitoes were collected from 21 swarms composed primarily of males. A few specimens of Culex quinquefasciatus were collected from 3 swarms. Although both molecular M and S forms were found in sympatry in the village, swarms were composed almost exclusively of the molecular M form. This suggests that there are alternative swarming habits for both molecular M and S forms of A. gambiae in nature.