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Background: Immunity to mosquito salivary proteins could provide protection against multiple mosquito-borne diseases and significantly impact public health. We evaluated the safety and immunogenicity of AGS-v PLUS, a mosquito salivary peptide vaccine, in healthy adults 18-50 years old., Methods: We conducted a randomized, double-blind, placebo-controlled Phase 1 study of AGS-v PLUS administered subcutaneously on Days 1 and 22 at the Center for Vaccine Development and Global Health, Baltimore, MD, USA. Participants were block randomized 1:1:1:1:1 to two doses saline placebo, two doses AGS-v PLUS, AGS-v PLUS/ISA-51 and saline placebo, two doses AGS-v PLUS/ISA-51, or two doses AGS-v PLUS/Alhydrogel. Primary endpoints were safety (all participants receiving ≥1 injection) and antibody and cytokine responses (all participants with day 43 samples), analysed by intention to treat., Findings: Between 26 August 2019 and 25 February 2020, 51 participants were enrolled and randomized, 11 into the single dose AGS-v PLUS/ISA-51 group and ten in other groups. Due to COVID-19, 15 participants did not return for day 43 samplings. Participants experienced no treatment-emergent or serious adverse events. All solicited symptoms in 2/10 placebo recipients and 22/41 AGS-v PLUS recipients after dose one and 1/10 placebo recipients and 22/41 AGS-v PLUS recipients after dose two were mild/moderate except for one severe fever the day after vaccination (placebo group). Only injection site pain was more common in vaccine groups (15/51 after dose 1 and 11/51 after dose 2) versus placebo. Compared to placebo, all vaccine groups had significantly greater fold change in anti-AGS-v PLUS IgG and IFN-ɣ from baseline., Interpretation: AGS-v PLUS had favourable safety profile and induced robust immune responses. Next steps will determine if findings translate into clinical efficacy against mosquito-borne diseases., Funding: UK Department of Health and Social Care., Competing Interests: Declaration of interests The views expressed in this publication are those of the author(s) and not necessarily those of Innovate UK or the UK's Department of Health and Social Care. Imutex Limited is a joint venture with PepTcell Limited (trading as SEEK) in which Open Orphan has a 49% shareholding, and PepTcell has 51%. Dr. Olga Pleguezuelos is an employee of ConserV Bioscience, a subsidiary of PepTcell Limited (trading as SEEK) and a shareholder in PepTcell Limited (trading as SEEK). Gregory Stoloff is an employee and shareholder in PepTcell. Imutex Ltd owns the intellectual property rights around AGS-v and AGS-v PLUS. All remaining authors declare no conflicts of interest. OP and GS are also named inventors in the following issued patents: AU 2008297213, AU 2013200258, BE 2783694, CH 2783694, CN 20101815526, DE 602008054283.1, DK 2783694, EP 2783694, ES 2783694, FI 2783694, FR 2783694, GB 2783694, IE 2783694, IN 296636, IT 2783694, JP 5981883, JP 6227018, MC 2783694, MT 2783694, MX 310667, NO 2783694, NZ 602090, OA 14958, PL 2783694, PT 2783694, RU 2466737, SE 2783694, SG 187401, TR 2783694, TW 548646, US 8,986,703, ZA 2010/00914., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Anopheles gambiae sensu stricto, one of the principal vectors of malaria, has been divided into two subspecific groups, known as the M and S molecular forms. Recent studies suggest that the M form found in Cameroon is genetically distinct from the M form found in Mali and elsewhere in West Africa, suggesting further subdivision within that form., Methods: Chromosomal, microsatellite and geographic/ecological evidence are synthesized to identify sources of genetic polymorphism among chromosomal and molecular forms of the malaria vector Anopheles gambiae s.s., Results: Cytogenetically the Forest M form is characterized as carrying the standard chromosome arrangement for six major chromosomal inversions, namely 2La, 2Rj, 2Rb, 2Rc, 2Rd, and 2Ru. Bayesian clustering analysis based on molecular form and chromosome inversion polymorphisms as well as microsatellites describe the Forest M form as a distinct population relative to the West African M form (Mopti-M form) and the S form. The Forest-M form was the most highly diverged of the An. gambiae s.s. groups based on microsatellite markers. The prevalence of the Forest M form was highly correlated with precipitation, suggesting that this form prefers much wetter environments than the Mopti-M form., Conclusion: Chromosome inversions, microsatellite allele frequencies and habitat preference all indicate that the Forest M form of An. gambiae is genetically distinct from the other recognized forms within the taxon Anopheles gambiae sensu stricto. Since this study covers limited regions of Cameroon, the possibility of gene flow between the Forest-M form and Mopti-M form cannot be rejected. However, association studies of important phenotypes, such as insecticide resistance and refractoriness against malaria parasites, should take into consideration this complex population structure.

Certain forms of Anopheles gambiae s.s. actively maintain malaria transmission in the driest areas and months of the year because of considerable drought tolerance. We monitored desiccation resistance of F1 offspring of both the M and S forms of field-collected An. gambiae s.s. Our results indicate that the geographic cline in the distribution of the two forms, as observed in Mali, corresponds to a physiological difference in response to arid environments. In addition, female mosquitoes survived significantly longer than males, enhancing the vector competence for the malaria parasite. Our study supports a genetic link to the drought tolerance phenotype, a phenotype with important consequences to malaria transmission in many places in Africa.

Background: Chromosomal inversions have been considered to be potentially important barriers to gene flow in many groups of animals through their effect on recombination suppression in heterokaryotypic individuals. Inversions can also enhance local adaptation in different groups of organisms and may often represent species-specific differences among closely related taxa. We conducted a study to characterize the 2La inversion karyotypes of An. gambiae sensu stricto mosquitoes sampled from the Kilombero Valley (Tanzania) using a newly designed PCR assay., Results: We frequently encountered a (687 bp) fragment which was only present in the Kilombero Valley populations. Laboratory crossing between An. gambiae s.s. from Njage (Tanzania) and Kisumu (Western Kenya) populations resulted in F1 offspring carrying the observed fragment. Karyotype analysis did not indicate differences in 2La region chromosome morphology between individuals carrying the PCR fragments, the 207 bp fragment, or the 687 bp fragement., Conclusion: The observed insertion/deletion polymorphism within the region amplified by the 2La PCR diagnostic test may confound the interpretation of this assay and should be well considered in order to maintain an acceptable level of reliability in studies using this assay to describe the distribution and frequency of the 2La inversion among natural populations of An. gambiae s.s.

The sand fly Lutzomyia longipalpis (Lutz & Neiva) (Diptera: Psychodidae), is the vector of the Neotropical parasite Leishmania chagasi. Designing methods to control the spread of this pathogen involves maintaining large laboratory colonies of these flies. However, transmission studies have been hampered particularly in regard to significant fluctuations in colony productivity. In the process of optimizing our sand fly rearing protocol, we increased moisture levels in the environment of the adult blood-fed female and found that egg development increased, whereas sand fly longevity decreased. Interestingly, continuous moisture from the fourth day after a bloodmeal significantly increased the longevity of the fly, but it did not significantly decrease the number of eggs developed. These results will help guide investigators aiming to manipulate the longevity and productivity of sand fly colonies.

The principal vector of malaria in sub-Saharan Africa, Anopheles gambiae is subdivided into two molecular forms M and S. Additionally, several chromosomal forms, characterized by the presence of various inversion polymorphisms, have been described. The molecular forms M and S each contain several chromosomal forms, including the Savanna, Mopti and Forest forms. The M and S molecular forms are now considered to be the reproductive units within A. gambiae and it has recently been argued that a low recombination rate in the centromeric region of the X chromosome has facilitated isolation between these forms. The status of the chromosomal forms remains unclear however. Therefore, we studied genetic differentiation between Savanna S, Forest S, Forest M and Mopti M populations using microsatellites. Genetic differentiation between Savanna S and Forest S populations is very low (F(ST) = 0.0053 +/- 0.0049), even across large distances. In comparison, the Mopti M and Forest M populations show a relatively high degree of genetic differentiation (F(ST) = 0.0406 +/- 0.0054) indicating that the M molecular form may not be a single entity, but could be subdivided into at least two distinct chromosomal forms. Previously it was proposed that inversions have played a role in the origin of species within the A. gambiae complex. We argue that a possible subdivision within the M molecular form could be understood through this process, with the acquisition of inversions leading to the expansion of the M molecular form into new habitat, dividing it into two distinct chromosomal forms.

The precipitin test was applied to identify the blood meal sources of Lutzomyia intermedia, collected in two distinct habitats: inside houses and in the peridomicile, in Mesquita, Rio de Janeiro State, Brazil, a transmission area of Leishmania (V.) braziliensis. The following antisera were tested: human, avian, equine, rodent, and opossum. From a total of 370 females analyzed, 128 specimens from the domicile and 59 from the peridomicile reacted with specific antisera. The anthropophily of L. intermedia was confirmed in both habitats; likewise, the feeding of this sand fly species on domestic animals, observed in previous entomological surveys, was confirmed by the strong reactivity with avian, canine, and equine antisera. However, feeding on rodents, mammals frequently found inside and around houses, represents further evidence related to the vector competence of L. intermedia, since synanthropic and sylvatic rodents have been considered a putative reservoir of L. (V.) braziliensis.

The genetic relationships among male Lutzomyia (Nyssomyia) intermedia (Lutz & Neiva) (Diptera: Psychodidae) from three populations from the same endemic area of American cutaneous leishmaniasis (ACL) in the state of Rio de Janeiro, Brazil, were compared. The sandflies were collected in three ecologically different habitats: domestic, extra-domestic and sylvatic over a total range of 800 m. Three molecular markers were employed to assess population variation. Based on MLEE markers, it could not be concluded that the three populations do not belong to the same gene pool (F(st) = 0.005). No within-population departure from Hardy-Weinberg equilibrium was detected (P < 0.05) and they presented the same level of gene variation. The number of migrants (Nm) indicated that at least 50 individuals per generation migrated between the three habitats. RAPD-PCR markers revealed that, except for the primer five, all were polymorphic. Phenetic analysis of the genotypes showed the presence of two principal clusters corresponding to: (1) domestic plus extra-domestic and (2) sylvatic. Unique genotypes were observed in each population. The sylvatic population was the most polymorphic, showing the largest number of genotypes and low level of similarity between them. Three mtDNA gene markers were studied by SSCP analysis. The most frequent haplotype for each marker ranged in frequency from 60 to 87% and individuals with unique haplotypes varied from 1 to 5%. Interestingly, the SSCP analysis showed a low level of polymorphism within populations. The disagreement between the different molecular markers observed and the hypothesis that L. intermedia could be participating in the transmission cycle of Leishmania (Viannia) braziliensis in environments ranging from the interior of human dwellings to the forest, are discussed.

Canine visceral leishmaniosis (CVL) may be an important factor preceding human outbreaks of the disease. We report that the prevalence of canine visceral leishmaniosis infection has been increasing in recent years in Anastácio town, located in the central western region of Brazil. Serological investigations showed that 75.3% of dogs presented antibody titres ranging from 1/40 to 1/160 in the indirect immunofluorescence antibody test (IFAT). Bone marrow and lymph node aspirates provided positive cultures and furnished parasites for enzymological and serological typing in 42.5% and 41.1% of the cases, respectively. All the strains were typed as Leishmania (L.) chagasi. This is primarily a canine disease that spills over into the human population as a zoonosis. The study showed the epidemiological features of the infection in a region in which the problem of visceral leishmaniosis has been underestimated.

A study of the sand fly fauna was carried out in the endemic area ofAmerican cutaneous leishmaniasis (ACL) in the Municipality of Mesquita, State of Rio de Janeiro, in an area where dwellings (with humans and dogs infected by Leishmania (Viannia) braziliensis) have been recently visited by sloths. Weekly captures were made during two consecutive years in two different sites (1 m and 600 m above sea level) and in three different habitats in each site (domestic, extra-domestic and sylvatic) using CDC light traps and Shannon traps. The following species were identified: Brumptomyia (Brumptomyia) guimaraesi, B. (B.) nitzulescui, Lutzomyia (Lutzomyia) longipalpis, L. migonei, L. serrana, L. (Psathyromyia) lutziana, L. barrettoi, L. (Nyssomyia) intermedia, L. (N.) whitmani, L. (Mycropygomyia) schireiberi, L. quinquefer, L. sallesi, L. (Psychodopygus) ayrozai, L. pascalei, L. (Pintomyia) bianchigalatiae and L. lanei. The collection of B. guimaraesi and B. nitzulescui species indicates the proximity of the study area to the sylvatic habitat. Two important suspected vectors of ACL were found: L. intermedia and L. migonei. L. longipalpis, a proven vector of American visceral leishmaniasis (AVL), was also found in this area. L. intermedia, the predominant sand fly species, was collected by both methods of capture and in the three different sites. It was observed biting man inside houses, but it was less anthropophilic in the sylvatic habitat. Our observations suggest that altitude does not influence the dispersion of L. intermedia population in the Municipality of Mesquita. The data indicate that L. intermedia remains as a putative vector of Leishmania (V.) braziliensis in this locality. An hypothesis of a sylvatic transmission cycle in the area and its correlation with the domestic cycle are discussed in the present paper.

We analyzed 64 quantitative and qualitative morphological characters in different populations of Lutzomyia (Nyssomyia) umbratilis Ward & Fraiha from Venezuela (Gran Sabana, Bolívar/BO) and Brazil (Serra do Navio, Amapá/AP Manaus, Amazonas/AM, Monte Dourado, Pará/ PA, and Peixoto de Azevedo, Mato Grosso/MT). Based on an analysis of qualitative characters, no differences were observed that could distinguish between the populations. Parametric tests were used to verify differences between sample means and variance homogeneity. The analysis-of-variance (ANOVA) test showed the existence of differences between character means in the five populations. In the graphic representation of the Student-Newman-Keuls test (Student-Newman-Keuls), we observed that 77% of the analyzed characters displayed disjunctions between the Venezuelan and Brazilian populations. Considering only the Brazilian population averages, we concluded that they are statistically different. When the Student-Newman-Keuls test was applied, no pattern was found that could separate them. However, in part, the phenetic analysis showed some evidences of heterogeneity in MT population.

Peixoto de Azevedo is located in the north of State of Mato Grosso, where environmental alterations led to an outbreak of American cutaneous leishmaniasis in the 80s. The parasite from patients was characterized as Leishmania (V.) braziliensis. The aim of this study is to contribute to the sand fly ecology of Central-West Brazil. Captures were carried out monthly using CDC light traps. Twenty-six species of sand fly were characterized; among which Lutzomyia (Lutzomyia) spathotrichia, L. runoides and L. (Psychodopygus) llanosmartinsi were recorded in the State of Mato Grosso for the first time. L. (Nyssomyia) whitmani, L. (N.) antunesi, L. (L.) spathotrichia, L. (P.) c. carrerai, L. (P.) complexa, L. (P.) lainsoni and L. (N.) umbratilis constituted 92.4% of the local fauna, among which L. (N.) whitmani and L. (N.) antunesi, accounting for about 53% of the fauna at the stations of capture. On the vertical distribution of sand flies on the Beira-Rio Farm, L. (N.) whitmani and L. (N.) antunesi prevailed at ground level and in the canopy, respectively, whereas on the BR-080, L. (P.) llanosmartinsi was prevalent on the ground and L. (P.) c. carrerai, in the canopy. It is suggested that L. (N.) umbratilis is the local vector.