Transmission-reducing and -enhancing monoclonal antibodies against Plasmodium vivax gamete surface protein Pvs48/45.

Gamete surface protein P48/45 has been shown to be important for male gamete fertility and a strong candidate for the development of a malaria transmission-blocking vaccine (TBV). However, TBV development for Plasmodium vivax homolog Pvs48/45 has been slow because of a number of challenges: availability of conformationally suitable recombinant protein; the lack of an in vivo challenge model; and the inability to produce P. vivax gametocytes in culture to test transmission-blocking activity of antibodies. To support ongoing efforts to develop Pvs48/45 as a potential vaccine candidate, we initiated efforts to develop much needed reagents to move the field forward. We generated monoclonal antibodies (mAbs) directed against Pvs48/45 and characterized putative functional domains in Pvs48/45 using recombinant fragments corresponding to domains D1-D3 and their biological functionality through ex vivo direct membrane feeding assays (DMFAs) using P. vivax parasites from patients in a field setting in Brazil. While some mAbs partially blocked oocyst development in the DMFA, one mAb caused a significant enhancement of the infectivity of gametocytes in the mosquitoes. Individual mAbs exhibiting blocking and enhancing activities recognized non-overlapping epitopes in Pvs48/45. Further characterization of precise epitopes recognized by transmission-reducing and -enhancing antibodies will be crucial to design an effective immunogen with optimum transmission-reducing potential.

Molecular and morphometric study of Brazilian populations of Psychodopygus davisi.

In this study, we analysed the molecular and morphometric differences of several populations of the putative sand fly vector Psychodopygus davisi (Root, 1934) (Diptera, Psychodidae, Phlebotominae) in Brazil. We amplified the 658 base pair fragments of the DNA barcoding region-cytochrome c oxidase subunit 1 (COI) gene-for 57 specimens of P. davisi and three specimens of Psychodopygus claustrei (Abonnenc, Léger & Fauran, 1979). We merged our data with public sequences of the same species available from GenBank. Then, the combined dataset-87 sequences and 20 localities-was analysed using population structure analysis and different species delimitation approaches. Geometric morphometry of wings was performed for 155 specimens of P. davisi populations from the North, Midwest and Southeast Brazilian regions, analysing the differences in centroid sizes and canonical variates. Molecular analysis indicated high intraspecific genetic distance values for P. davisi (maximum p distance = 5.52%). All algorithms identified P. davisi and P. claustrei as distinct molecular taxonomic units, despite the low interspecific distance (p distance to the nearest neighbour = 4.79%). P. davisi sequences were split into four genetic clusters by population structure analysis and at least five genetic lineages using intermediate scenarios of the species delimitation algorithms. The species validation analysis of BPP strongly supported the five-species model in our dataset. We found high genetic diversity in this taxon, which is in agreement with its wide geographic distribution in Brazil. Furthermore, the wing analysis showed that specimens from the Southeast Region of Brazil are different from those in the North and the Midwest. The evolutionary patterns of P. davisi populations in Brazil suggest the presence of candidate species, which need to be validated in future studies using a more comprehensive approach with both genomic data and morphological characters.

You are what you eat: a systematic review exploring the interaction between Brazilian sand flies and their vertebrate food sources.

Sand flies play a crucial role as vectors of bacteria, viruses, and protists, with Leishmania being the most notable among them, transmitted to vertebrate hosts during blood feeding. Understanding the feeding behaviours of sand flies is imperative for gaining insights into their eco-epidemiological roles in the transmission of these infectious agents. This systematic review aimed to answer the question 'What are the blood-feeding sources identified in Brazilian sand flies?' to provide an analysis of their blood-feeding habits. The diverse range of at least 16 vertebrate orders identified as blood sources for 54 sand fly species across different geographic regions was summarised, and the factors potentially associated with the risk of bias in the included studies were analysed. The findings broaden the discussion concerning methods used to identify blood meal sources and shed light on the implications of sand fly feeding behaviours for the transmission dynamics of Leishmania.

Crude saliva of Amblyomma cajennense sensu stricto (Acari: Ixodidae) reduces locomotor activity and increases the hemocyte number in the females of Aedes aegypti (Diptera: Culicidae).

Aedes aegypti are vector insects of arboviruses such as dengue, Zika, and chikungunya. All available vector control methods have limited efficacy, highlighting the urgent need to find alternative ones. Evidence shows that arachnids like ticks are sources of biologically active compounds. Moreover, chemical modulation of the locomotor and immune systems of vector insects can be used to control arbovirus transmission. The present study evaluated the effectiveness of crude saliva of female Amblyomma cajennense sensu stricto (s.s.) ticks in reducing locomotor activity and inducing an immune response in Ae. aegypti females. Additionally, the study evaluated the protein constitution of tick saliva. For this purpose, the crude saliva obtained from several semi-engorged A. cajennense females was used. A volume of 0.2 nL of crude tick saliva was administered to mosquitoes by direct intrathoracic microinjection. The effect of the tick's saliva on the locomotor activity of the mosquito was observed using Flybox, a video-automated monitoring system, and the hemolymph hemocyte levels were quantified by reading slides under a light microscope. The protein concentration of the crude tick saliva was 1.27 µg/µL, and its electrophoretic profile indicates the presence of proteins with a molecular weight ranging between ∼17 and 95 kDa. Microplusins, ixodegrins, cystatin, actins, beta-actin, calponin, albumin, alpha-globulins, and hemoglobin were the main proteins identified by proteomics in the saliva of A. cajennense. The microinjected saliva had low toxicity for Ae. aegypti females and significantly reduced their locomotor activity, especially in the transition between the light and dark phases. The crude tick saliva did not change the period and rhythmicity of the circadian cycle. The tick saliva significantly increased the number of hemocytes two days after injection and reduced it after five days. These results suggest that further evaluation of the biological properties of tick saliva proteins against Ae. aegypti would be of interest.

Evaluation of insemination, blood feeding, and Plasmodium vivax infection effects on locomotor activity patterns of the malaria vector Anopheles darlingi (Diptera: Culicidae).

Circadian behavioral patterns in mosquitoes can be observed through their locomotor activity, which includes fundamental behaviors such as foraging, mating, and oviposition. These habits, which are fundamental to the life cycle of Anopheles mosquitoes, are closely related to pathogen transmission to humans. While rhythmic cycles of locomotor activity have been described in Anopheles species, no studies have been conducted on Anopheles darlingi species, the main malaria vector in the Amazon region. The aim of this study was to investigate how insemination status, blood meal, and Plasmodium vivax infection affect the locomotor activity of An. darlingi. The experiments were performed with 3- to 10-day-old An. darlingi females, which had been fed with 15% honey solution. These mosquitoes were obtained from the Malaria Vector Production and Infection Platform (PIVEM)/FIOCRUZ-RO. The experimental groups were divided into four categories: virgin vs. inseminated, unfed virgin vs. blood-fed virgin, unfed inseminated vs. blood-fed inseminated, and infected blood vs. uninfected blood. Locomotor activity was monitored using the Flybox equipment, capturing images that were subsequently converted into video to measure the insect activity, using PySoLo software. The periodicity and rhythmicity of mosquito locomotor activity were analyzed using MatLab® software. The locomotor activity of An. darlingi females showed a nocturnal and bimodal pattern under LD conditions. When comparing the insemination states and blood meal, there was a reduction in the locomotor activity in inseminated and blood-fed females. However, the P. vivax+ infection did not increase locomotor activity of An. darlingi species.

Diversity of free-living ticks and serological evidence of spotted fever group Rickettsia and ticks associated to dogs, Porto Velho, Western Amazon, Brazil.

Rondônia is the only state in the North Region of Brazil to have registered confirmed cases of Brazilian Spotted Fever (BSF). The present study investigated the epidemiological cycle of Rickettsia spp. by surveying free-living ixodofauna and tick parasitism of dogs in the municipality of Porto Velho, Rondônia State. Ticks and dogs were tested for the presence of Rickettsia spp. DNA and dog serum was tested for reactivity to anti-Rickettsia spp. antibodies. Tick collection and dog blood sampling were performed in peri-urban and rural environments at 11 locations. Eight free-living Amblyomma species and one Haemaphysalis species were collected: A. scalpturatum, A. naponense, A. oblongoguttatum, A. coelebs, A. latepunctatum, A. pacae, A. ovale, Amblyomma sp., and H. juxtakochi. Three tick species were found parasitizing dogs: Rhipicephalus sanguineus sensu lato, A. oblongoguttatum and A. ovale. Molecular analysis did not identify the presence of the gltA gene fragment in any tick specimen. Results from an indirect immunofluorescent assay (IFA) showed that 20.8% of peri-urban and 15.4% of rural dog sera exhibited reactivity to Rickettsia rhipicephali, Rickettsia amblyommatis, Rickettsia bellii and Rickettsia parkeri antigens. Antibody prevalence in dogs was 16.4%. This study is the first to describe the prevalence of Rickettsia spp. infection in dogs from Porto Velho municipality. Our findings enhance current knowledge of Rickettsia spp. circulation in the Western Amazon.

Description of an automatic copulation induction system used to establish a free-mating laboratory colony of Nyssorhynchus deaneorum from Brazil.

BACKGROUND Nyssorhynchus deaneorum is a potential malaria vector because it has been shown to be competent to transmit Plasmodium vivax and Plasmodium falciparum, and because it exhibits antropophilic and endophilic behaviors in some regions of the Amazon. This profile makes Ny. deaneorum a useful mosquito for experiments that model Plasmodium-vector interactions in the Amazon. OBJECTIVE Herein we describe how a free-mating colony of Ny. deaneorum has been established using an automated light stimulation system. METHODS Mosquitoes were captured in São Francisco do Guaporé, Rondônia. The F1 generation was reared until adult emergence at which point copulation was induced using an automatic copulation induction system (ACIS). FINDINGS After four generations, natural mating and oviposition began to occur without light stimulation. The number of pupae and adult mosquitoes increased from the F5 to F10 generations. The new Ny. deaneorum colony exhibited susceptibility to P. vivax. MAIN CONCLUSIONS Automated light stimulation is an effective method for establishing an Ny. deaneorum colony under laboratory conditions as it produces enough adults to create a stenogamic colony. The establishment of a stable, P. vivax-susceptible colony of Ny. deaneorum makes it possible to model parasite-vector interactions and to test novel drug therapies that target parasite development in mosquitoes.

Diversity, natural infection and blood meal sources of phlebotomine sandflies (Diptera, Psychodidae) in the western Brazilian Amazon.

BACKGROUND: The state of Rondônia (RO) is a hot spot for human cases of cutaneous leishmaniasis. Many sandfly species in RO are putative vectors of leishmaniasis. OBJECTIVES: This study examines the diversity patterns and the presence of Leishmania DNA and blood meal sources of sandflies in RO. METHODS: A sandfly survey was performed between 2016 and 2018 in 10 municipalities categorised into three different environment types: (i) Conservation Unit (CUN) - comprised of preserved ombrophilous forests; (ii) Forest Edge (FE) - small forest fragments; and (iii) Peridomicile (PE) - areas around dwellings. FINDINGS: A total of 73 species were identified from 9,535 sandflies. The most abundant species were Psychodopygus davisi (1,741 individuals), Nyssomyia antunesi (1,397), Trichophoromyia auraensis (1,295) and Trichophoromyia ubiquitalis (1,043). Diversity was the highest in CUN, followed by the FE and PE environments. One pool of Ps. davisi tested positive for Leishmania braziliensis, reinforcing the possibility that Ps. davisi acts as a vector. The cytochrome b (cytb) sequences were used to identify three blood meal sources: Bos taurus, Homo sapiens and Tamandua tetradactyla. MAIN CONCLUSIONS: Our results demonstrated that sandflies can switch between blood meal sources in differing environments. This study enhances the knowledge of the vector life cycle in RO and provides information relevant to leishmaniasis surveillance.

Immune response in Mansonella ozzardi infection modulated by IL-6/IL-10 axis in Amazon region of Brazil.

Mansonellosis is an endemic disease in the South and Central America. In Brazil, one of the etiological agents is Mansonella ozzardi. This filarial infection is yet poorly understood, with a controversial morbity, presenting since a oligosymptoms, malaria-like signs or without complaint in humans. The knowledge of the human immune response to microfilariae infection is limited mainly by different evolutionary cycles of the parasite in the host. In addition, the prevalence of this filarial parasite infection is high in several regions of Amazonas State. A cross-sectional study was conducted in an endemic area for microfilariae of M. ozzardi (MF) infection in the Amazonas State, Brazil. Proinflammatory and regulatory cytokines (IL-2, IL-4, IL-6, IL-10, TNF, IFN-gamma, and IL-17A) were measured in cryopreserved serum using the Cytometric Bead Array techniques (CBA) in 54 patients diagnosed with M. ozzardi infection and 55 individuals without the infection were included in the study (Controls). The IL-4, IL-6 and IL-10 level increased in infected patients with MF infection, while IL-17A increased in control only. When we compared controls to patients with high or low parasite load, the increased level of IL-6 and IL-10 were maintained. IL-6 contributes to the proinflammatory activity and IL-10 modulates Th1, Th2 and Th17 immune response. Furthermore, IL-4 was detected as a marker in the MF infection and MF patients with low parasite load, indicating the action of the Th2 cell response. The complex network of cytokines acting during M. ozzardi infection depends on a fine balance to determine a host protective effect or filarial persistence. Therefore, these results suggest that the immune response in MF infection is modulated by IL-6/IL-10 axis.

Sensitivity of diagnostic methods for Mansonella ozzardi microfilariae detection in the Brazilian Amazon Region.

BACKGROUND: The human filarial worm Mansonella ozzardi is highly endemic in the large tributaries of the Amazon River. This infection is still highly neglected and can be falsely negative when microfilariae levels are low. OBJECTIVES: This study investigated the frequency of individuals with M. ozzardi in riverine communities in Coari municipality, Brazilian Amazon. METHODS: Different diagnostic methods including polymerase chain reaction (PCR), blood polycarbonate membrane filtration (PCMF), Knott's method (Knott), digital thick blood smears (DTBS) and venous thick blood smears (VTBS) were used to compare sensitivity and specificity among the methods. Data were analysed using PCMF and Bayesian latent class models (BLCM) as the gold standard. We used BLCM to calculate the prevalence of mansonelliasis based on the results of five diagnostic methods. FINDINGS: The prevalence of mansonelliasis was 35.4% by PCMF and 30.1% by BLCM. PCR and Knott methods both possessed high sensitivity. Sensitivity relative to PCMF was 98.5% [95% confidence interval (CI): 92.0 - 99.7] for PCR and 83.5% (95% CI: 72.9 - 90.5) for Knott. Sensitivity derived by BLCM was 100% (95% CI 93.7 - 100) for PCMF, 100% (95% CI: 93.7 - 100) for PCR and 98.3% (95% CI: 90.6 - 99.9) for Knott. The odds ratio of being diagnosed as microfilaremic increased with age but did not differ between genders. Microfilariae loads were higher in subjects aged 30 - 45 and 45 - 60 years. MAIN CONCLUSIONS: PCMF and PCR were the best methods to assess the prevalence of mansonelliasis in our samples. As such, using these methods could lead to higher prevalence of mansonelliasis in this region than the most commonly used method (i.e., thick blood smears).

Trichophoromyia auraensis is a putative vector

The sandfly Trichophoromyia auraensis has recently evolved as a proven vector of Leishmania (Viannia) endemic to state of Acre in the north of Brazil. This note is intended to propose a correction in the report of the first occurrence of natural infection of Leishmania (Viannia) in this species. We and the other scientific groups reinforced that Tr. auraensis is a possible vector involved in the transmission of American cutaneous leishmaniasis in Acre, Brazil.

Phlebotomine sandfly (Diptera: Psychodidae) diversity and their Leishmania DNA in a hot spot of American Cutaneous Leishmaniasis human cases along the Brazilian border with Peru and Bolivia.

In this study, we identified the phlebotomine sandfly vectors involved in the transmission of American Cutaneous Leishmaniasis (ACL) in Assis Brasil, Acre, Brazil, which is located on the Brazil-Peru-Bolivia frontier. The genotyping of Leishmania in phlebotomines was performed using polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism. A total of 6,850 sandflies comprising 67 species were captured by using CDC light traps in rural areas of the municipality. Three sandfly species were found in the state of Acre for the first time: Lutzomyia georgii, Lu. complexa and Lu. evangelistai. The predominant species was Lu. auraensis/Lu. ruifreitasi and Lu. davisi (total 59.27%). 32 of 368 pools were positive for the presence of Leishmania DNA (16 pools corresponding to Lu. davisi, and 16 corresponding to Lu. auraensis/Lu. ruifreitasi), with a minimal infection prevalence of 1.85% in Lu. davisi and 2.05% in Lu. auraensis/Lu. ruifreitasi. The Leishmania species found showed maximum identity with L. (Viannia) guyanensis and L. (V.) braziliensis in both phlebotomine species. Based on these results and similar scenarios previously described along the Brazil/Peru/Bolivia tri-border, the studied area must take into consideration the possibility of Lu. davisi and Lu. auraensis/Lu. ruifreitasi as probable vectors of ACL in this municipality.

Mansonella ozzardi in the municipality of Tefé, Amazonas, Brazil, 60 years after the first report: an epidemiologic study.

The prevalence of mansonelliasis was studied in the municipality of Tefé, state of Amazonas, Brazil. The prevalence (thick blood smear method) was 13.6% (147/1,078), higher in the Solimões River region (16.3%) than in the Tefé River region (6.3%). In the sampled communities in the Solimões River region, a higher density of cases was observed, as indicated by a kernel analysis (odds ratio 0.34; 95% confidence interval: 0.20-0.57). Males had a higher prevalence (χ2 = 31.292, p < 0.001) than women. Mansonella ozzardi prevalence was higher in retirees and farmers (28.9% and 27%, respectively). Prevalence also significantly increased with age (χ2 = -128.17, p < 0.001), with the highest numbers occurring in persons older than 67 years.

Evaluation of combination vaccines targeting transmission of Plasmodium falciparum and P. vivax.

Transmission-blocking vaccines interrupting malaria transmission within mosquitoes represent an ideal public health tool to eliminate malaria at the population level. Plasmodium falciparum and P. vivax account for more than 90% of the global malaria burden, co-endemic in many regions of the world. P25 and P48/45 are two leading candidates for both species and have shown promising transmission-blocking activity in preclinical and clinical studies. However, neither of these target antigens as individual vaccines has induced complete transmission inhibition in mosquitoes. In this study, we assessed immunogenicity of combination vaccines based on P25 and P48/45 using a DNA vaccine platform to broaden vaccine specificity against P. falciparum and P. vivax. Individual DNA vaccines encoding Pvs25, Pfs25, Pvs48/45 and Pfs48/45, as well as various combinations including (Pvs25 + Pvs48/45), (Pfs25 + Pfs48/45), (Pvs25 + Pfs25), and (Pvs48/45 + Pfs48/45), were evaluated in mice using in vivo electroporation. Potent antibody responses were induced in mice immunized with individual and combination DNA vaccines, and specific antibody responses were not compromised when combinations of DNA vaccines were evaluated against individual DNA vaccines. The anti-Pvs25 IgG from individual and combination groups revealed concentration-dependent transmission-reducing activity (TRA) in direct membrane feeding assays (DMFA) using blood from P. vivax-infected donors in Brazil and independently in ex vivo MFA using Pvs25-transgenic P. berghei. Similarly, anti-Pfs25 and anti-Pfs48/45 IgGs from mice immunized with Pfs25 and Pfs48/45 DNA vaccines individually and in various combinations revealed antibody dose-dependent TRA in standard membrane feeding assays (SMFA) using culture-derived P. falciparum gametocytes. However, antibodies induced by immunization with Pvs48/45 DNA vaccines were ineffective in DMFA and require further vaccine construct optimization, considering the possibility of induction of both transmission-blocking and transmission-enhancing antibodies revealed by competition ELISA. These studies provide a rationale for combining multiple antigens to simultaneously target transmission of malaria caused by P. falciparum and P. vivax.

Diversity, vertical stratification and detection of Leishmania DNA in sand flies (Diptera: Psychodidae) collected in the Mapinguari National Park, Brazilian Amazon.

This study aimed to assess the diversity patterns of sand fly fauna across different strata and detect Leishmania DNA in these insects in the Mapinguari National Park in Rondônia and Amazonas states, Northern Brazil. Sand flies were collected with "HP" light traps in the canopy (15 m) and at ground level (1 m) on two trails, during August and November 2021 and March and August 2022. Polymerase chain reaction and DNA sequencing were used to identify Leishmania species. A total of 8,040 individuals (2,303♂ - 28.64 %, 5,737♀ - 71.36 %) were collected and 53 species and 13 genera were identified. The most abundant species were Psychodopygus chagasi (21.00 %), Trichophoromyia ubiquitalis (13.61 %) Psychodopygus davisi (11.69 %), Nyssomyia fraihai (10.58 %), Nyssomyia antunesi (6.68 %) and Nyssomyia richardwardi (5.55 %). Species diversity was greater in the canopy (Shannon index H' = 10.8 common species) when compared to ground level (H' = 10.5 common species). We observed a minimum infection rate of 0.45 % (22/4,868 females), in which Leishmania braziliensis DNA was found in Ps. chagasi and Le. lainsoni DNA in Ps. chagasi, Ny. richardwardi, Ps. ayrozai and Th. ubiquitalis. The minimum infection rate of Leishmania in the canopy was 0.47 % (19/4,031) and in the ground was 0.52 % (3/567). In the present study, we observed Le. lainsoni DNA in females of Ny. richardwardi for the first time. The data presented in this study contribute to understanding sand fly diversity and its distribution between the states of Rondônia and Amazonas. They may be useful for implementing targeted control measures to reduce the spread of leishmaniasis and implement entomological surveillance strategies.

Entomological inferences highlight the risk of Leishmania transmission in the urban area of Porto Velho, Rondônia, Brazil.

Entomological investigations were conducted for the first time in urban forest remnants of Porto Velho, state of Rondônia, Brazil, to explore the transmission dynamics of Leishmania. Sand fly collections were carried out at ten sites, encompassing both canopy and ground strata, from October to December 2021. A total of 1,671 sand flies were collected, representing 42 species within 12 genera. Nyssomyia Antunesi (n = 384) and Psychodopygus davisi (n = 111) were the most abundant species. Molecular analyses targeting the V7V8 region (18S gene) unveiled the presence of sequences 100% identical to Leishmania infantum in females of Bichromomyia flaviscutellata (1), Nyssomyia Antunesi complex (6), Nyssomyia umbratilis (1), Nyssomyia sp. (1), Psychodopygus ayrozai (1), Ps. davisi (3), Psychodopygus paraensis (1), and Sciopemyia sordellii (1). Sequences 100% similar to Trypanosoma minasense were found in two samples of the Nyssomyia Antunesi complex, and two samples of Sc. sordellii presented 100% identity to a Trypanosoma sp. strain, previously identified in this same sand fly in Rondônia. Sequencing of Cytb fragment suggested Homo sapiens, Dasypus novemcinctus and Tamandua tetradactyla as the blood source for distinct sand flies. The identification of sequences similar to L. infantum in sand flies collected in urban forest fragments is noteworthy, correlating with the recent local and regional occurrence of autochthonous cases of human visceral leishmaniasis. However, further studies are imperative to ascertain the presence of hosts/reservoirs and evaluate the risk of L. infantum transmission to humans.

Case Report: Plasmodium vivax Sporozoite Melanization in the Midgut and Salivary Gland of the Malaria Vector Anopheles darlingi.

Anopheles darlingi is the primary malaria vector in the Amazon region and is highly susceptible to both Plasmodium vivax and Plasmodium falciparum parasites. Although anopheline mosquitoes may develop melanotic encapsulation in response to Plasmodium parasites, there is no record of An. darlingi exhibiting a melanization response to P. vivax, the main malaria parasite in the Americas. Here, we report the occurrence of P. vivax sporozoite melanization in An. darlingi mosquitoes.

Detection and Genotyping of Trypanosoma cruzi Samples in Species of Genus Rhodnius from Different Environments in the Brazilian Amazon.

Background: In the Amazon region, several species of triatomines occur in the natural environments. Among them, species of the genus Rhodnius are a risk to human populations due to their high rates of infection with Trypanosoma cruzi. The aim of this study was to identify the T. cruzi genotypes in Rhodnius specimens and their relationship with sylvatic hosts from different environments in the Brazilian Amazon. Methods: A total of 492 triatomines were collected from the municipalities of Monte Negro, Rondônia state, and Humaitá, Amazonas state, 382 of them being nymphs and 110 adults. Genotyping of T. cruzi in six discrete typing units (DTUs) was performed using conventional multilocus PCR. The triatomines that were positive for T. cruzi and engorged with blood were also targeted for amplification of the cytochrome B (cytB) gene to identify bloodmeal sources. Results: Of the 162 positive samples, the identified DTUs were TcI (87.65%) and TcIV (12.35%). It was observed that 102 specimens were engorged with a variety of bloodmeals. Triatomines infected with TcI were associated with DNA of all identified vertebrates, except Plecturocebus brunneus. TcIV was detected in triatomines that fed on Coendou prehensilis, Didelphis marsupialis, Mabuya nigropunctata, P. brunneus, Pithecia irrorata, Sapajus apella, and Tamandua tetradactyla. Conclusion: Results highlight the need to understand the patterns of T. cruzi genotypes in Rhodnius spp. and their association with sylvatic hosts to better elucidate their role in the transmission of Chagas disease in the Amazon region.

Description of a new phlebotomine species (Diptera: Psychodidae, Phlebotominae) and new records of sand flies from the State of Acre, northern Brazil.

Groundbreaking studies of phlebotomine sand fly populations in Assis Brasil, State of Acre, Brazil, resulted in the collection of 13 new records of phlebotomine sand flies and one previously undescribed species. Lutzomyia naiffi sp. nov. is described here. The new species is similar to Lutzomyia columbiana (Ristorcelli & Van Ty) in measurements and other morphological characters.

Inaccurate recording of Mansonella perstans in free-ranging primates outside its endemic area in Brazil?

In the study undertaken by Souza et al. [Primates 64(1):153-159, 2022; https://doi.org/10.1007/s10329-022-01038-5 ], published in the most recent volume of this journal, the blood samples of two Alouatta guariba clamitans (Primates, Atelidae) from two municipalities in the state of Rio Grande do Sul, southern Brazil were reported to be positive for Mansonella perstans. This is the first reported finding of M. perstans in A. guariba clamitans, as well as the first time that M. perstans has been recorded in Brazil outside the Amazon region. We would like to express our concern about this finding, specifically with respect to the geographical distribution of M. perstans in Brazil, as, up until this study, this filaria had only been found in the upper Rio Negro region in São Gabriel da Cachoeira, Amazonas, Brazil. Moreover, species identification was performed using partial sequences of three gene fragments, namely internal transcribed spacer 2, 12S, and 18S, yet neither the phylogenetic trees nor the BLAST alignments of these sequences provided supporting evidence that they belong to M. perstans.