Evaluation of antibody responses to tsetse fly saliva in domestic animals in the sleeping sickness endemic foci of Bonon and Sinfra, Côte d'Ivoire.

After intensive control efforts, human African trypanosomiasis (HAT) was declared eliminated in Côte d'Ivoire as a public health problem in December 2020 and the current objective is to achieve the interruption of the transmission (zero cases). Reaching this objective could be hindered by the existence of an animal reservoir of Trypanosoma (T.) brucei (b.) gambiense. In the framework of a study led in 2013 to assess the role of domestic animals in the epidemiology of HAT in the two last active foci from Côte d'Ivoire (Bonon and Sinfra), plasmas were sampled from four species of domestic animals for parasitological (microscopic examination by the buffy coat technique (BCT)), serological (immune trypanolysis (TL)) and molecular (specific PCR: TBR for T. brucei s.l., TCF for T. congolense forest type, TVW for T. vivax and PCR for T. b. gambiense) testing. In order to improve the understanding of the involvement/role of these animals in the transmission of T. b. gambiense, we have quantified in this study the IgG response to whole saliva extracts of Glossina palpalis gambiensis in order to perform an association analysis between anti-saliva responses and the positivity of diagnostic tests. Cattle and pigs had significantly higher rates of anti-tsetse saliva responses compared to goats and sheep (p < 0.01). In addition, the anti-tsetse saliva responses were strongly associated with the parasitology (BCT+), serology (TL+) and PCR (TBR+ and TCF+) results (p < 0.001). These associations indicate a high level of contacts between the positive/infected animals and tsetse flies. Our findings suggest that protecting cattle and pigs against tsetse bites could have a significant impact in reducing transmission of both animal and human trypanosome species, and advocates for a "One health" approach to better control African trypanosomosis in Côte d'Ivoire.

Molecular epidemiology of Animal African Trypanosomosis in southwest Burkina Faso.

BACKGROUND: Animal African Trypanosomosis (AAT) is a parasitic disease of livestock that has a major socio-economic impact in the affected areas. It is caused by several species of uniflagellate extracellular protists of the genus Trypanosoma mainly transmitted by tsetse flies: T. congolense, T. vivax and T. brucei brucei. In Burkina Faso, AAT hampers the proper economic development of the southwestern part of the country, which is yet the best watered area particularly conducive to agriculture and animal production. It was therefore important to investigate the extent of the infection in order to better control the disease. The objective of the present study was to assess the prevalence of trypanosome infections and collect data on the presence of tsetse flies. METHODS: Buffy coat, Trypanosoma species-specific PCR, Indirect ELISA Trypanosoma sp and trypanolysis techniques were used on 1898 samples collected. An entomological survey was also carried out. RESULTS: The parasitological prevalence of AAT was 1.1%, and all observed parasites were T. vivax. In contrast, the molecular prevalence was 23%, of which T. vivax was predominant (89%) followed by T. congolense (12.3%) and T. brucei s.l. (7.3%) with a sizable proportion as mixed infections (9.1%). T. brucei gambiense, responsible of sleeping sickness in humans, was not detected. The serological prevalence reached 49.7%. Once again T. vivax predominated (77.2%), but followed by T. brucei (14.7%) and T. congolense (8.1%). Seven samples, from six cattle and one pig, were found positive by trypanolysis. The density per trap of Glossina tachinoides and G. palpalis gambiensis was 1.2 flies. CONCLUSIONS/SIGNIFICANCE: Overall, our study showed a high prevalence of trypanosome infection in the area, pointing out an ongoing inadequacy of control measures.

In vivo analysis of trypanocidal drug resistance in sahelian goats infected by Trypanosoma vivax strains collected in northern Togo.

Trypanosoma (T.) vivax is one of the animal trypanosomes species causing calf mortality and economic losses in Togo. Despite its importance as the most widely distributed trypanosome species, T. vivax has received little attention because it is difficult to cultivate most field isolates in rodents. No molecular diagnostic tools for the identification of drug-resistant in T. vivax are currently available. Herein, four field isolates of T. vivax from Togo were cryopreserved and assessed for susceptibility to diminazene aceturate (DA) and isometamidium chloride (ISM) in goats. For field isolate preparation, 1 ml of blood from an infected goat was diluted in 111 µl of phosphate-buffered-saline and stored in liquid nitrogen. The in vivo experiment drug test was performed using twenty Sahelian goats with six-month of age and weighing 14.5 ± 1.6 kg. These experimental goats were purchased from a tsetse free-area Dori, a Sahelian region of Burkina Faso. The cryopreserved T. vivax isolates with unknowns, DA, and ISM sensitivity was inoculated to five goats and one goat was used as control. Each animal was inoculated by intravenously route 1 × 105 trypanosomes from the donor goat. Relapses were earlier in the first phase of treatment (14.85 ± 1.08 days) compared with the second phase (20 ± 3.39 days). The overall mean PCV of the control group decreased from 32% to 17% at day-60 (P-value < 0.001). Three isolates were phenotypically resistant to 0.5 mg per kg body weight (BW) ISM and one for 3.5 mg per kg BW of DA. There were no relapses with the 7 mg per kg BW dose DA. This study shows the resistance of T. vivax to two main trypanocidal drugs in different villages of Mango. The results suggest the extension of surveillance strategies to remote villages in Togo and will guide the veterinarian or herder in choosing a mass treatment strategy. Further studies will be needed to better understand the molecular basis of the observed resistance.

An atlas to support the progressive control of tsetse-transmitted animal trypanosomosis in Burkina Faso.

BACKGROUND: African animal trypanosomosis (AAT), transmitted by tsetse flies, is arguably the main disease constraint to integrated crop-livestock agriculture in sub-Saharan Africa, and African heads of state and governments adopted a resolution to rid the continent of this scourge. In order to sustainably reduce or eliminate the burden of AAT, a progressive and evidence-based approach is needed, which must hinge on harmonized, spatially explicit information on the occurrence of AAT and its vectors. METHODS: A digital repository was assembled, containing tsetse and AAT data collected in Burkina Faso between 1990 and 2019. Data were collected either in the framework of control activities or for research purposes. Data were systematically verified, harmonized, georeferenced and integrated into a database (PostgreSQL). Entomological data on tsetse were mapped at the level of individual monitoring traps. When this was not possible, mapping was done at the level of site or location. Epidemiological data on AAT were mapped at the level of location or village. RESULTS: Entomological data showed the presence of four tsetse species in Burkina Faso. Glossina tachinoides, present from the eastern to the western part of the country, was the most widespread and abundant species (56.35% of the catches). Glossina palpalis gambiensis was the second most abundant species (35.56%), and it was mainly found in the west. Glossina morsitans submorsitans was found at lower densities (6.51%), with a patchy distribution in the southern parts of the country. A single cluster of G. medicorum was detected (less than 0.25%), located in the south-west. Unidentified tsetse flies accounted for 1.33%. For the AAT component, data for 54,948 animal blood samples were assembled from 218 geographic locations. The samples were tested with a variety of diagnostic methods. AAT was found in all surveyed departments, including the tsetse-free areas in the north. Trypanosoma vivax and T. congolense infections were the dominant ones, with a prevalence of 5.19 ± 18.97% and 6.11 ± 21.56%, respectively. Trypanosoma brucei infections were detected at a much lower rate (0.00 ± 0.10%). CONCLUSIONS: The atlas provides a synoptic view of the available information on tsetse and AAT distribution in Burkina Faso. Data are very scanty for most of the tsetse-free areas in the northern part of the country. Despite this limitation, this study generated a robust tool for targeting future surveillance and control activities. The development of the atlas also strengthened the collaboration between the different institutions involved in tsetse and AAT research and control in Burkina Faso, which will be crucial for future updates and the sustainability of the initiative.

Evaluation of the Re-emergence Risk of Human African Trypanosomiasis in the Southwestern Burkina Faso, A Gold-Bearing Mutation Area.

PURPOSE: The boom in Burkina Faso's artisanal gold mining since 2007 has attracted populations from Côte d'Ivoire and Guinea, which are the West African countries most affected by human African trypanosomiasis (HAT) and therefore increases its risk of re-emergence. Our aim was to update the HAT data in Burkina Faso in the risk of the re-emergence context with the advent of artisanal gold mining. METHODS: The study was carried out in the southwestern Burkina Faso where entomological surveys were conducted using biconical traps in March 2017. Follow by an active medical survey in April 2017, which was targeted the gold panners in 7 villages closer to artisanal gold sites, using CATT, mini-anion exchange centrifugation technique, trypanolysis test (TL) and ELISA test to measure human/tsetse contacts. The buffy coat technique and the TL were also applied in pigs to check their reservoir role of human trypanosomes. RESULTS: Our results have shown no case of HAT among 958 individuals tested and all the 50 pigs were also negative, but the level of antibodies against tsetse saliva evidenced by ELISA revealed low human/tsetse contact. Moreover, gold panners practise agriculture and breeding in an infected tsetse area, which are increased the risk. CONCLUSION: Our results illustrate that the risk of re-emergence is low. The passive surveillance system implemented in 2015 in southwestern Burkina Faso is needed to increase the sentinel sites to better cover this area by taking into account the gold mining. Finally, awareness-raising activities are needed among populations about HAT.

Novel protein candidates for serodiagnosis of African animal trypanosomosis: Evaluation of the diagnostic potential of lysophospholipase and glycerol kinase from Trypanosoma brucei.

African trypanosomosis, a parasitic disease caused by protozoan parasites transmitted by tsetse flies, affects both humans and animals in sub-Saharan Africa. While the human form (HAT) is now limited to foci, the animal form (AAT) is widespread and affects the majority of sub-Saharan African countries, and constitutes a real obstacle to the development of animal breeding. The control of AAT is hampered by a lack of standardized and easy-to used diagnosis tools. This study aimed to evaluate the diagnostic potential of TbLysoPLA and TbGK proteins from Trypanosoma brucei brucei for AAT serodiagnosis in indirect ELISA using experimental and field sera, individually, in combination, and associated with the BiP C-terminal domain (C25) from T. congolense. These novel proteins were characterized in silico, and their sequence analysis showed strong identities with their orthologs in other trypanosomes (more than 60% for TbLysoPLA and more than 82% for TbGK). TbLysoPLA displays a low homology with cattle (<35%) and Piroplasma (<15%). However, TbGK shares more than 58% with cattle and between 45-55% with Piroplasma. We could identify seven predicted epitopes on TbLysoPLA sequence and 14 potential epitopes on TbGK. Both proteins were recombinantly expressed in Escherichia coli. Their diagnostic potential was evaluated by ELISA with sera from cattle experimentally infected with T. congolense and with T.b. brucei, sera from cattle naturally infected with T. congolense, T. vivax and T.b. brucei. Both proteins used separately had poor diagnostic performance. However, used together with the BiP protein, they showed 60% of sensitivity and between 87-96% of specificity, comparable to reference ELISA tests. In conclusion, we showed that the performance of the protein combinations is much better than the proteins tested individually for the diagnosis of AAT.

Variant antigen diversity in Trypanosoma vivax is not driven by recombination.

African trypanosomes (Trypanosoma) are vector-borne haemoparasites that survive in the vertebrate bloodstream through antigenic variation of their Variant Surface Glycoprotein (VSG). Recombination, or rather segmented gene conversion, is fundamental in Trypanosoma brucei for both VSG gene switching and for generating antigenic diversity during infections. Trypanosoma vivax is a related, livestock pathogen whose VSG lack structures that facilitate gene conversion in T. brucei and mechanisms underlying its antigenic diversity are poorly understood. Here we show that species-wide VSG repertoire is broadly conserved across diverse T. vivax clinical strains and has limited antigenic repertoire. We use variant antigen profiling, coalescent approaches and experimental infections to show that recombination plays little role in diversifying T. vivax VSG sequences. These results have immediate consequences for both the current mechanistic model of antigenic variation in African trypanosomes and species differences in virulence and transmission, requiring reconsideration of the wider epidemiology of animal African trypanosomiasis.

Drug quality analysis of isometamidium chloride hydrochloride and diminazene diaceturate used for the treatment of African animal trypanosomosis in West Africa.

BACKGROUND: Diminazene diaceturate (DA) and isometamidium chloride hydrochloride (ISM) are with homidium bromide, the main molecules used to treat African Animal Trypanosomosis (AAT). These drugs can be purchased from official suppliers but also from unofficial sources like local food markets or street vendors. The sub-standard quality of some of these trypanocides is jeopardizing the efficacy of treatment of sick livestock, leading thus to economic losses for the low-resource farmers and is contributing to the emergence and spread of drug resistance. The objective of this study was to assess the quality of trypanocidal drugs sold in French speaking countries of West Africa. In total, 308 drug samples including 282 of DA and 26 of ISM were purchased from official and unofficial sources in Benin, Burkina Faso, Côte d'Ivoire, Mali, Niger and Togo. All samples were analysed at LACOMEV (Dakar, Senegal), a reference laboratory of the World Organisation for Animal Health, by galenic inspection and high performance liquid chromatography. RESULTS: The results showed that 51.90% of the samples were non-compliant compared to the standards and were containing lower quantity of the active ingredient compared to the indications on the packaging. The non-compliances ranged from 63.27% in Togo to 32.65% in Burkina Faso (61.82% in Benin, 53.84% in Mali, 50% in Côte d'Ivoire, 47.36% in Niger). The rates of non-compliance were not statistically different (P = 0.572) from official or unofficial suppliers and ranged from 30 to 75% and from 0 to 65% respectively. However, the non-compliance was significantly higher for ISM compared to DA (P = 0.028). CONCLUSIONS: The high non-compliance revealed in this study compromises the efficacy of therapeutic strategies against AAT, and is likely to exacerbate chemoresistance in West Africa. Corrective actions against sub-standard trypanocides urgently need to be taken by policy makers and control authorities.

The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.

Kinetoplastid parasites-trypanosomes and leishmanias-infect millions of humans and cause economically devastating diseases of livestock, and the few existing drugs have serious deficiencies. Benzoxaborole-based compounds are very promising potential novel anti-trypanosomal therapies, with candidates already in human and animal clinical trials. We investigated the mechanism of action of several benzoxaboroles, including AN7973, an early candidate for veterinary trypanosomosis. In all kinetoplastids, transcription is polycistronic. Individual mRNA 5'-ends are created by trans splicing of a short leader sequence, with coupled polyadenylation of the preceding mRNA. Treatment of Trypanosoma brucei with AN7973 inhibited trans splicing within 1h, as judged by loss of the Y-structure splicing intermediate, reduced levels of mRNA, and accumulation of peri-nuclear granules. Methylation of the spliced leader precursor RNA was not affected, but more prolonged AN7973 treatment caused an increase in S-adenosyl methionine and methylated lysine. Together, the results indicate that mRNA processing is a primary target of AN7973. Polyadenylation is required for kinetoplastid trans splicing, and the EC50 for AN7973 in T. brucei was increased three-fold by over-expression of the T. brucei cleavage and polyadenylation factor CPSF3, identifying CPSF3 as a potential molecular target. Molecular modeling results suggested that inhibition of CPSF3 by AN7973 is feasible. Our results thus chemically validate mRNA processing as a viable drug target in trypanosomes. Several other benzoxaboroles showed metabolomic and splicing effects that were similar to those of AN7973, identifying splicing inhibition as a common mode of action and suggesting that it might be linked to subsequent changes in methylated metabolites. Granule formation, splicing inhibition and resistance after CPSF3 expression did not, however, always correlate and prolonged selection of trypanosomes in AN7973 resulted in only 1.5-fold resistance. It is therefore possible that the modes of action of oxaboroles that target trypanosome mRNA processing might extend beyond CPSF3 inhibition.

New insights from Gorongosa National Park and Niassa National Reserve of Mozambique increasing the genetic diversity of Trypanosoma vivax and Trypanosoma vivax-like in tsetse flies, wild ungulates and livestock from East Africa.

BACKGROUND: Trypanosoma (Duttonella) vivax is a major pathogen of livestock in Africa and South America (SA), and genetic studies limited to small sampling suggest greater diversity in East Africa (EA) compared to both West Africa (WA) and SA. METHODS: Multidimensional scaling and phylogenetic analyses of 112 sequences of the glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) gene and 263 sequences of the internal transcribed spacer of rDNA (ITS rDNA) were performed to compare trypanosomes from tsetse flies from Gorongosa National Park and Niassa National Reserve of Mozambique (MZ), wild ungulates and livestock from EA, and livestock isolates from WA and SA. RESULTS: Multidimensional scaling (MDS) supported Tvv (T. vivax) and TvL (T. vivax-like) evolutionary lineages: 1) Tvv comprises two main groups, TvvA/B (all SA and WA isolates plus some isolates from EA) and TvvC/D (exclusively from EA). The network revealed five ITS-genotypes within Tvv: Tvv1 (WA/EA isolates), Tvv2 (SA) and Tvv3-5 (EA). EA genotypes of Tvv ranged from highly related to largely different from WA/SA genotypes. 2) TvL comprises two gGAPDH-groups formed exclusively by EA sequences, TvLA (Tanzania/Kenya) and TvLB-D (MZ). This lineage contains more than 11 ITS-genotypes, seven forming the lineage TvL-Gorongosa that diverged from T. vivax Y486 enough to be identified as another species of the subgenus Duttonella. While gGAPDH sequences were fundamental for classification at the subgenus, major evolutionary lineages and species levels, ITS rDNA sequences permitted identification of known and novel genotypes. CONCLUSIONS: Our results corroborate a remarkable diversity of Duttonella trypanosomes in EA, especially in wildlife conservation areas, compared to the moderate diversity in WA. Surveys in wilderness areas in WA may reveal greater diversity. Biogeographical and phylogenetic data point to EA as the place of origin, diversification and spread of Duttonella trypanosomes across Africa, providing relevant insights towards the understanding of T. vivax evolutionary history.

Identification of a Tsal152-75 salivary synthetic peptide to monitor cattle exposure to tsetse flies.

BACKGROUND: The saliva of tsetse flies contains a cocktail of bioactive molecules inducing specific antibody responses in hosts exposed to bites. We have previously shown that an indirect-ELISA test using whole salivary extracts from Glossina morsitans submorsitans was able to discriminate between (i) cattle from tsetse infested and tsetse free areas and (ii) animals experimentally exposed to low or high numbers of tsetse flies. In the present study, our aim was to identify specific salivary synthetic peptides that could be used to develop simple immunoassays to measure cattle exposure to tsetse flies. METHODS: In a first step, 2D-electrophoresis immunoblotting, using sera from animals exposed to a variety of bloodsucking arthropods, was performed to identify specific salivary proteins recognised in cattle exposed to tsetse bites. Linear epitope prediction software and Blast analysis were then used to design synthetic peptides within the identified salivary proteins. Finally, candidate peptides were tested by indirect-ELISA on serum samples from tsetse infested and tsetse free areas, and from exposure experiments. RESULTS: The combined immunoblotting and bioinformatics analyses led to the identification of five peptides carrying putative linear epitopes within two salivary proteins: the tsetse salivary gland protein 1 (Tsal1) and the Salivary Secreted Adenosine (SSA). Of these, two were synthesised and tested further based on the absence of sequence homology with other arthropods or pathogen species. IgG responses to the Tsal152-75 synthetic peptide were shown to be specific of tsetse exposure in both naturally and experimentally exposed hosts. Nevertheless, anti-Tsal152-75 IgG responses were absent in animals exposed to high tsetse biting rates. CONCLUSIONS: These results suggest that Tsal152-75 specific antibodies represent a biomarker of low cattle exposure to tsetse fly. These results are discussed in the light of the other available tsetse saliva based-immunoassays and in the perspective of developing a simple serological tool for tsetse eradication campaigns to assess the tsetse free status or to detect tsetse reemergence in previously cleared areas.

Administration of ivermectin to peridomestic cattle: a promising approach to target the residual transmission of human malaria.

BACKGROUND: The success of current control tools in combatting malaria vectors is well established. However, sustained residual transmission of Plasmodium parasites persists. Mass drug administration (MDA) to humans of the endectocide ivermectin for vector control is receiving increasing attention. However, vectors feeding upon animals escape this promising approach. Zoophagy of mosquitoes sustains both the vector population and endemic population of vector-borne pathogens. Therefore, only a strategy that will combine ivermectin MDAs targeted at humans and their peridomestic animals could be successful at controlling residual malaria transmission. METHODS: Burkinabé cattle have been treated with injectable therapeutic dose of ivermectin (0.2 mg/kg of body weight) to render blood meals toxic to field representative populations of Anopheles coluzzii carrying the kdr mutation. Direct skin-feeding assays were performed from 2 to 28 days after injection (DAI) and mosquitoes were followed for their survival, ability to become gravid and fecundity. Membrane feeding assays were further performed to test if an ivermectin blood meal taken at 28 DAI impacts gametocyte establishment and development in females fed with infectious blood. RESULTS: The mosquitocidal effect of ivermectin is complete for 2 weeks after injection, whether 12 days cumulative mortalities were of 75 and 45 % the third and fourth weeks, respectively. The third week, a second ivermectin blood meal at sub-lethal concentrations further increased mortality to 100 %. Sub-lethal concentrations of ivermectin also significantly decreased egg production by surviving females, increasing further the detrimental effect of the drug on vector densities. Although females fitness was impaired by sub-lethal ivermectin blood meals, these did not diminish nor increase their susceptibility to infection. CONCLUSION: This study demonstrates the potential of integrated MDA of ivermectin to both human and peridomestic cattle to target vector reservoirs of residual malaria transmission. Such integration lies in 'One-Health' efforts being implemented around the globe, and would be especially relevant in rural communities in Africa where humans are also at risk of common zoonotic diseases.

Microsatellite analysis supports clonal propagation and reduced divergence of Trypanosoma vivax from asymptomatic to fatally infected livestock in South America compared to West Africa.

BACKGROUND: Mechanical transmission of the major livestock pathogen Trypanosoma vivax by other biting flies than tsetse allows its spread from Africa to the New World. Genetic studies are restricted to a small number of isolates and based on molecular markers that evolve too slowly to resolve the relationships between American and West African populations and, thus, unable us to uncover the recent history of T. vivax in the New World. METHODS: T. vivax genetic diversity, population structure and the source of outbreaks was investigated through the microsatellite multiloci (7 loci) genotype (MLGs) analysis in South America (47isolates from Brazil, Venezuela and French Guiana) and West Africa (12 isolates from The Gambia, Burkina Faso, Ghana, Benin and Nigeria). Relationships among MLGs were explored using phylogenetic, principal component and STRUCTURE analyses. RESULTS: Although closely phylogenetically related, for the first time, genetic differences were detected between T. vivax isolates from South America (11 genotypes/47 isolates) and West Africa (12 genotypes/12 isolates) with no MLGs in common. Diversity was far greater across West Africa than in South America, where genotypes from Brazil (MLG1-6), Venezuela (MLG7-10) and French Guiana (MLG11) shared similar but not identical allele composition. No MLG was exclusive to asymptomatic (endemic areas) or sick (outbreaks in non-endemic areas) animals, but only MLGs1, 2 and 3 were responsible for severe haematological and neurological disorders. CONCLUSIONS: Our results revealed closely related genotypes of T. vivax in Brazil and Venezuela, regardless of endemicity and clinical conditions of the infected livestock. The MLGs analysis from T. vivax across SA and WA support clonal propagation, and is consistent with the hypothesis that the SA populations examined here derived from common ancestors recently introduced from West Africa. The molecular markers defined here are valuable to assess the genetic diversity, to track the source and dispersion of outbreaks, and to explore the epidemiological and pathological significance of T. vivax genotypes.

In silico identification of a candidate synthetic peptide (Tsgf118-43) to monitor human exposure to tsetse flies in West Africa.

BACKGROUND: The analysis of humoral responses directed against the saliva of blood-sucking arthropods was shown to provide epidemiological biomarkers of human exposure to vector-borne diseases. However, the use of whole saliva as antigen presents several limitations such as problems of mass production, reproducibility and specificity. The aim of this study was to design a specific biomarker of exposure to tsetse flies based on the in silico analysis of three Glossina salivary proteins (Ada, Ag5 and Tsgf1) previously shown to be specifically recognized by plasma from exposed individuals. METHODOLOGY/PRINCIPAL FINDINGS: Synthetic peptides were designed by combining several linear epitope prediction methods and Blast analysis. The most specific peptides were then tested by indirect ELISA on a bank of 160 plasma samples from tsetse infested areas and tsetse free areas. Anti-Tsgf118-43 specific IgG levels were low in all three control populations (from rural Africa, urban Africa and Europe) and were significantly higher (p<0.0001) in the two populations exposed to tsetse flies (Guinean HAT foci, and South West Burkina Faso). A positive correlation was also found between Anti-Tsgf118-43 IgG levels and the risk of being infected by Trypanosoma brucei gambiense in the sleeping sickness foci of Guinea. CONCLUSION/SIGNIFICANCE: The Tsgf118-43 peptide is a suitable and promising candidate to develop a standardize immunoassay allowing large scale monitoring of human exposure to tsetse flies in West Africa. This could provide a new surveillance indicator for tsetse control interventions by HAT control programs.

Dynamics of tsetse natural infection rates in the Mouhoun river, Burkina Faso, in relation with environmental factors.

In Burkina Faso, the cyclical vectors of African animal trypanosomoses (AAT) are riverine tsetse species, namely Glossina palpalis gambiensis Vanderplank (G.p.g.) and Glossina tachinoides Westwood (G.t.) (Diptera: Glossinidae). Experimental work demonstrated that environmental stress can increase the sensitivity of tsetse to trypanosome infection. Seasonal variations of the tsetse infection rates were monitored monthly over 17 months (May 2006-September 2007) in two sites (Douroula and Kadomba). In total, 1423 flies were dissected and the infection of the proboscis, middle intestine and salivary glands was noted. All the positive organs were analyzed using monospecific polymerase chain reaction (PCR) primers. To investigate the role of different environmental factors, fly infection rates were analyzed using generalized linear mixed binomial models using the species, sex, and monthly averages of the maximum, minimum and mean daily temperatures, rainfalls, Land Surface Temperature day (LSTd) and night (LSTn) as fixed effects and the trap position as a random effect. The overall infection rate was 10% from which the predominant species was T. congolense (7.6% of the flies), followed by T. vivax (2.2% of the flies). The best model (lowest AICc) for the global infection rates was the one with the maximum daily temperature only as fixed effect (p < 0.001). For T. congolense, the best model was the one with the tsetse species, sex, maximum daily temperature and rainfalls as fixed effect, where the maximum daily temperature was the main effect (p < 0.001). The number of T. vivax infections was too low to allow the models to converge. The maturation rate of T. congolense was very high (94%), and G. t. harbored a higher maturation rate (p = 0.03). The results are discussed in view of former laboratory studies showing that temperature stress can increase the susceptibility of tsetse to trypanosomes, as well as the possibility to improve AAT risk mapping using satellite images.

Decrease in survival and fecundity of Glossina palpalis gambiensis vanderplank 1949 (Diptera: Glossinidae) fed on cattle treated with single doses of ivermectin.

BACKGROUND: Human and Animal Trypanosomes are major problems for the socio-economic growth of developing countries like Burkina Faso. Ivermectin is currently used to treat humans in mass drug administration programs in Africa, and is also commonly used for veterinary purposes. In this study, we tested the effect of ivermectin injected into cattle on the survival and fecundity of Glossina palpalis gambiensis, the main vector of human and animal trypanosomes in West Africa. METHODS: Three cows (local zebu*baoulé crossbreds) were used, and received either no ivermectin (for the control), or ivermectin at therapeutic dose (0.2 mg/kg) and 10 times the therapeutic dose (2 mg/kg) respectively. G. palpalis gambiensis were fed on the cattle for their first bloodmeal, and then either on cattle or on membrane for subsequent meals. RESULTS: Our results showed that survival of Glossina palpalis gambiensis was significantly decreased when they were fed on cattle treated with ivermectin. This decrease in survival ranged from 21% to 83.7% for the therapeutic dose (0.2 mg/kg), up to 8 days after treatment. The effects of a dose of 2 mg/kg were higher with a 78.3% to 93.9% decrease in survival, until 14 days after injection. The therapeutic dose of ivermectin also decreased fecundity, and delayed the first larviposition, but there was no significant effect on hatching rate. CONCLUSION: Ivermectin injected into cattle may constitute an additional potential tool for the control of Glossina palpalis gambiensis and possibly other vector species. Further studies will be needed to assess its effect on trypanosome transmission, and to define more precisely the adequate dose to be used for control purposes.

Treating cattle to protect people? Impact of footbath insecticide treatment on tsetse density in Chad.

BACKGROUND: In Chad, several species of tsetse flies (Genus: Glossina) transmit African animal trypanosomoses (AAT), which represents a major obstacle to cattle rearing, and sleeping sickness, which impacts public health. After the failure of past interventions to eradicate tsetse, the government of Chad is now looking for other approaches that integrate cost-effective intervention techniques, which can be applied by the stake holders to control tsetse-transmitted trypanosomoses in a sustainable manner. The present study thus attempted to assess the efficacy of restricted application of insecticides to cattle leg extremities using footbaths for controlling Glossina m. submorsitans, G. tachinoides and G. f. fuscipes in southern Chad. METHODOLOGY/PRINCIPAL FINDINGS: Two sites were included, one close to the historical human African trypanosomiasis (HAT) focus of Moundou and the other to the active foci of Bodo and Moissala. At both sites, a treated and an untreated herd were compared. In the treatment sites, cattle were treated on a regular basis using a formulation of deltamethrin 0.005% (67 to 98 cattle were treated in one of the sites and 88 to 102 in the other one). For each herd, tsetse densities were monthly monitored using 7 biconical traps set along the river and beside the cattle pen from February to December 2009. The impact of footbath treatment on tsetse populations was strong (p < 10(-3)) with a reduction of 80% in total tsetse catches by the end of the 6-month footbath treatment. CONCLUSIONS/SIGNIFICANCE: The impact of footbath treatment as a vector control tool within an integrated strategy to manage AAT and HAT is discussed in the framework of the "One Health" concept. Like other techniques based on the treatment of cattle, this technology should be used under controlled conditions, in order to avoid the development of insecticide and acaricide resistance in tsetse and tick populations, respectively.

First insights into the cattle serological response to tsetse salivary antigens: a promising direct biomarker of exposure to tsetse bites.

In the context of the Pan African Tsetse and Trypanosomiasis Eradication Campaign, the value of tsetse saliva antibodies as a biomarker of cattle exposure to tsetse flies was evaluated, as this could provide an alternative and complementary tool to conventional entomological methods. Serum immune reactivity to Glossina (G.) palpalis (p.) gambiensis, G. tachinoides and G. morsitans (m.) submorsitans whole saliva extracts (WSE) were monitored in cattle from both tsetse free and tsetse infested areas, and in cows experimentally exposed to tsetse flies and other hematophagous arthropods. In the tsetse infested area, cattle IgG responses to Glossina WSE were significantly higher during the dry season (p<0.0001) when herds are most exposed to tsetse flies and in infected animals (p=0.01) as expected in the case of a biomarker of exposure. Experimental studies further confirmed this as a quick rise of specific IgGs was observed in animals exposed to tsetse flies (within weeks), followed by a rapid clearance after exposure was stopped. In contrast to the two other tsetse species, G. m. submorsitans WSE enabled to detect exposure to all tsetse species and were associated with low level of cross-reactivity to other blood sucking arthropods. Finally, IgG responses to G. m. submorsitans salivary antigens enabled to distinguish different groups of cows according to exposure levels, thus indicating that tsetse saliva antibodies are not only indicators of tsetse exposure but also are correlated to the intensity of tsetse contacts (p=0.0031). Implementation of this new sero-epidemiological marker of cattle exposure to tsetse flies in the framework of tsetse elimination campaigns is discussed.

Identification of Glossina palpalis gambiensis specific salivary antigens: towards the development of a serologic biomarker of human exposure to tsetse flies in West Africa.

The saliva of blood sucking arthropods contains a number of pharmacologically active compounds that induce an antibody response in exposed human individuals. The objectives of the present study were (i) to assess the human IgG response directed against salivary antigens of Glossina palpalis gambiensis, the main vector of Trypanosoma brucei gambiense in West Africa, as a biomarker of human-tsetse contacts; and (ii) to identify specific salivary antigens. Immune reactivity of human plasma collected within active human African trypanosomiasis (HAT) foci (coastal Guinea), historical foci where tsetse flies are still present (South-West Burkina Faso) and a tsetse free area (Bobo-Dioulasso, Burkina Faso), was measured by ELISA against whole saliva extracts. In the active HAT foci and areas where tsetse flies were present in high densities, specific IgG responses were significantly higher (p < 0.0001) to those in Bobo-Dioulasso or in Loropeni, where tsetse flies were either absent or only present at low densities. Furthermore, 2D-electrophoresis combined with mass spectrometry enabled to reveal that several antigens were specifically recognized by plasma from exposed individuals. Among them, four salivary proteins were successfully identified (Ada, 5'Nuc, Ag5 and Tsgf1). These results represent a first attempt to identify Glossina salivary proteins or synthetic peptides to develop a standardized and specific biomarker of tsetse exposure in West Africa.

Climate, cattle rearing systems and African Animal Trypanosomosis risk in Burkina Faso.

BACKGROUND: In sub-Saharan countries infested by tsetse flies, African Animal Trypanosomosis (AAT) is considered as the main pathological constraint to cattle breeding. Africa has known a strong climatic change and its population was multiplied by four during the last half-century. The aim of this study was to characterize the impact of production practices and climate on tsetse occurrence and abundance, and the associated prevalence of AAT in Burkina Faso. METHODOLOGY/PRINCIPAL FINDINGS: Four sites were selected along a South-north transect of increasing aridity. The study combines parasitological and entomological surveys. For the parasitological aspect, blood samples were collected from 1,041 cattle selected through a stratified sampling procedure including location and livestock management system (long transhumance, short transhumance, sedentary). Parasitological and serological prevalence specific to livestock management systems show a gradual increase from the Sahelian to the Sudano-Guinean area (P<0.05). Livestock management system had also a significant impact on parasitological prevalence (P<0.05). Tsetse diversity, apparent densities and their infection rates overall decreased with aridity, from four species, an apparent density of 53.1 flies/trap/day and an infection rate of 13.7% to an absence at the northern edge of the transect, where the density and diversity of other biting flies were on the contrary highest (p<0.001). CONCLUSIONS/SIGNIFICANCE: The climatic pressure clearly had a negative impact on tsetse abundance and AAT risk. However, the persistency of tsetse habitats along the Mouhoun river loop maintains a high risk of cyclical transmission of T. vivax. Moreover, an "epidemic mechanical livestock trypanosomosis" cycle is likely to occur in the northern site, where trypanosomes are brought in by cattle transhuming from the tsetse infested area and are locally transmitted by mechanical vectors. In Burkina Faso, the impact of tsetse thus extends to a buffer area around their distribution belt, corresponding to the herd transhumance radius.