Distribution and Diversity of Sand Fly Species (Diptera: Psychodidae, Phlebotominae) in Two Geoclimatic Zones of Chad

Leishmaniasis transmitted by sandflies is an important vector-borne disease. In Chad, information on sandflies is outdated, and so this study was designed to update the sandfly fauna. Sandflies were collected in five health districts representing two geoclimatic zones using sticky traps and pyrethrum sprays in indoor and peridomestic habitats between September 2019 and December 2020. All collected sandfly specimens were identified based on species-level morphological characters. A total of 2,015 specimens belonging to 13 species of sandflies (one Phlebotomus and twelve Sergentomyia) were collected and identified. The vector of human cutaneous leishmaniasis, Phlebotomus duboscqi, represents 0.65% of all collected sandflies and is the only representative of the Phlebotomus genus to be collected predominantly inside human dwellings. Phlebotomus orientalis, the vector of visceral leishmaniasis previously collected in Chad in 1976 was not found in this study. Sergentomyia clydei, Sergentomyia schwetzi, Sergentomyia antennata and Sergentomyia africana were the most abundant species collected with 44.71%; 22.73%; 16.03% and 11.17%, respectively. Sergentomyia schwetzi and Sergentomyia dubia, the two species involved in the transmission of canine leishmaniasis, were found in five and four study sites respectively. According to our results, the sandfly fauna of the two geoclimatic zones of Chad is comprised of 13 species. Our data showed that, unlike P. orientalis which was not found, P. duboscqi is present in four of the five sites surveyed. Therefore, these areas are at risk and remain potential foci of cutaneous leishmaniasis. However, the need for further studies such as vector species detection, their seasonal fluctuations and their vector competence.

Repeated Sand Fly Bites of Infected BALB/c Mice Enhance the Development of Leishmania Lesions

Sand fly saliva has considerable immunomodulatory effects on Leishmania infections in mammalian hosts. Studies on several Leishmania – sand fly - host combinations have demonstrated that co-inoculation with Leishmania parasites enhances pathogenicity, while pre-exposure of hosts to sand fly bites provides significant protection against infection. However, the third scenario, the effect of sand fly saliva on parasite development in hosts infected before exposure to sand flies, remains an understudied aspect of Leishmania–host–vector interaction. Here we studied the effect of exposure of L. major-infected BALB/c mice to repeated sand fly bites. Mice infected intradermally with sand fly-derived Leishmania were repeatedly bitten by Phlebotomus duboscqi females every two weeks. The lesion development was recorded weekly for ten weeks post-infection and parasite load and distribution in various organs were tested post mortem using qPCR. Repeated sand fly bites significantly enhanced the development of cutaneous lesions; they developed faster and reached larger size than in unexposed mice. Multiple sand fly bites also increased parasites load in inoculated ears. On the other hand, the distribution of parasites in mice body and their infectiousness to vectors did not differ significantly between groups. Our study provides the first evidence that multiple and repeated exposures of infected BALB/c mice to sand fly bites significantly enhance the progress of local skin infection caused by Leishmania major and increase tissue parasite load, but do not affect the visceralization of parasites. This finding appeals to adequate protection of infected humans from sand fly bites, not only to prevent transmission but also to prevent enlarged lesions.

Distribution and Diversity of Sand Fly Species (Diptera: Psychodidae, Phlebotominae) in Two Geoclimatic Zones of Chad

Introduction: Leishmaniasis transmitted by sandflies is an important vector-borne disease. In Chad, information on sandflies is outdated, and so this study was designed to update the sandfly fauna. Methods: Sandflies were collected in five health districts representing two geoclimatic zones using sticky traps and pyrethrum sprays in indoor and peridomestic habitats between September 2019 and December 2020. All collected sandfly specimens were identified based on species-level morphological characters. Results: A total of 2,015 specimens belonging to 13 species of sandflies (one Phlebotomus and twelve Sergentomyia) were collected and identified. The vector of human cutaneous leishmaniasis, Phlebotomus duboscqi, represents 0.65% of all collected sandflies and is the only representative of the Phlebotomus genus to be collected predominantly inside human dwellings. Phlebotomus orientalis, the vector of visceral leishmaniasis previously collected in Chad in 1976 was not found in this study. Sergentomyia clydei, Sergentomyia schwetzi, Sergentomyia antennata and Sergentomyia africana were the most abundant species collected with 44.71%; 22.73%; 16.03% and 11.17%, respectively. Sergentomyia schwetzi and Sergentomyia dubia, the two species involved in the transmission of canine leishmaniasis, were found in five and four study sites respectively. Conclusion: According to our results, the sandfly fauna of the two geoclimatic zones of Chad is comprised of 13 species. Our data showed that, unlike P. orientalis which was not found, P. duboscqi is present in four of the five sites surveyed. Therefore, these areas are at risk and remain potential foci of cutaneous leishmaniasis. However, the need for further studies such as vector species detection, their seasonal fluctuations and their vector competence.

Individuals co-exposed to sand fly saliva and filarial parasites exhibit altered monocyte function

Background In Mali, cutaneous leishmaniasis (CL) and filariasis are co-endemic. Previous studies in animal models of infection have shown that sand fly saliva enhance infectivity of Leishmania parasites in naïve hosts while saliva-specific adaptive immune responses may protect against cutaneous and visceral leishmaniasis. In contrast, the human immune response to Phlebotomus duboscqi (Pd) saliva, the principal sand fly vector in Mali, was found to be dichotomously polarized with some individuals having a Th1-dominated response and others having a Th2-biased response. We hypothesized that co-infection with filarial parasites may be an underlying factor that modulates the immune response to Pd saliva in endemic regions. Methodology/Principal findings To understand which cell types may be responsible for polarizing human responses to sand fly saliva, we investigated the effect of salivary glands (SG) of Pd on human monocytes. To this end, elutriated monocytes were cultured in vitro, alone, or with SG, microfilariae antigen (MF ag) of Brugia malayi, or LPS, a positive control. The mRNA expression of genes involved in inflammatory or regulatory responses was then measured as were cytokines and chemokines associated with these responses. Monocytes of individuals who were not exposed to sand fly bites (mainly North American controls) significantly upregulated the production of IL-6 and CCL4; cytokines that enhance leishmania parasite establishment, in response to SG from Pd or other vector species. This selective inflammatory response was lost in individuals that were exposed to sand fly bites which was not changed by co-infection with filarial parasites. Furthermore, infection with filarial parasites resulted in upregulation of CCL22, a type-2 associated chemokine, both at the mRNA levels and by its observed effect on the frequency of recruited monocytes. Conclusions/Significance Together, our data suggest that SG or recombinant salivary proteins from Pd alter human monocyte function by upregulating selective inflammatory cytokines.

Protein methyltransferase 7 deficiency in Leishmania major increases neutrophil associated pathology in murine model

Leishmania major is the main causative agent of cutaneous leishmaniasis in the Old World. In Leishmania parasites, the lack of transcriptional control is mostly compensated by post-transcriptional mechanisms. Methylation of arginine is a conserved post-translational modification executed by Protein Arginine Methyltransferase (PRMTs). The genome from L. major encodes five PRMT homologs, including the cytosolic protein associated with several RNA-binding proteins, LmjPRMT7. It has been previously reported that LmjPRMT7 could impact parasite infectivity. In addition, a more recent work has clearly shown the importance of LmjPRMT7 in RNA-binding capacity and protein stability of methylation targets, demonstrating the role of this enzyme as an important epigenetic regulator of mRNA metabolism. In this study, we unveil the impact of PRMT7-mediated methylation on parasite development and virulence. Our data reveals that higher levels of LmjPRMT7 can impair parasite pathogenicity, and that deletion of this enzyme rescues the pathogenic phenotype of an attenuated strain of L. major. Interestingly, lesion formation caused by LmjPRMT7 knockout parasites is associated with an exacerbated inflammatory reaction in the tissue correlated with an excessive neutrophil recruitment. Moreover, the absence of LmjPRMT7 also impairs parasite development within the sand fly vector Phlebotomus duboscqi. Finally, a transcriptome analysis shed light onto possible genes affected by depletion of this enzyme. Taken together, this study highlights how post-transcriptional regulation can affect different aspects of the parasite biology.

Morphological Characterization of the Antennal Sensilla of the Afrotropical Sand Fly, Phlebotomus duboscqi (Diptera: Psychodidae)

We investigated by scanning electron microscopy the morphology, distribution, and abundance of antennal sensilla of females Phlebotomus duboscqi sand fly, an important vector of zoonotic cutaneous leishmaniasis at Afrotropical region. Thirteen well-differentiated sensilla were identified, among six types of cuticular sensilla. The probable function of these sensillary types is discussed in relation to their external structure and distribution. Five sensillary types were classified as olfactory sensilla, as they have specific morphological characters of sensilla with this function. Number and distribution of sensilla significantly differed between antennal segments. The results of the present work, besides corroborating in the expansion of the morphological and ultrastructural knowledge of P. duboscqi, can foment future electrophysiological studies for the development of volatile semiochemicals, to be used as attractants in traps for monitoring and selective vector control of this sand fly.