Leishmania (Sauroleishmania) tarentolae versus pathogenic species: comparative evaluation of protease activity, glycoconjugates, resistance to complement and metabolome composition.

BACKGROUND: Leishmania tarentolae is a non-pathogenic species found in lizards representing an important model for Leishmania biology. However, several aspects of this Sauroleishmania remain unknown to explain its low level of virulence. OBJECTIVES: We reported several aspects of L. tarentolae biology including glycoconjugates, proteolytic activities and metabolome composition in comparison to pathogenic species (Leishmania amazonensis, Leishmania braziliensis, Leishmania infantum and Leishmania major). METHODS: Parasites were cultured for extraction and purification of lipophosphoglycan (LPG), immunofluorescence probing with anti-gp63 and resistance against complement. Parasite extracts were also tested for proteases activity and metabolome composition. FINDINGS: Leishmania tarentolae does not express LPG on its surface. It expresses gp63 at lower levels compared to pathogenic species and, is highly sensitive to complement-mediated lysis. This species also lacks intracellular/extracellular activities of proteolytic enzymes. It has metabolic differences with pathogenic species, exhibiting a lower abundance of metabolites including ABC transporters, biosynthesis of unsaturated fatty acids and steroids, TCA cycle, glycine/serine/threonine metabolism, glyoxylate/dicarboxylate metabolism and pentose-phosphate pathways. MAIN CONCLUSIONS: The non-pathogenic phenotype of L. tarentolae is associated with alterations in several biochemical and molecular features. This reinforces the need of comparative studies between pathogenic and non-pathogenic species to elucidate the molecular mechanisms of virulence during host-parasite interactions.

Diversity, Leishmania detection, and blood meal sources of sand flies from Iguatama, Minas Gerais, Brazil.

This study investigated the sand fly fauna of the municipality Iguatama, in the Midwest Region of Minas Gerais state, Brazil, including Leishmania infection rates and blood meal sources. Sand flies were collected during four periods over the course of a single year, encompassing both dry and rainy seasons, using CDC light traps placed in peridomiciles where dogs were seropositive for visceral leishmaniasis (VL). A total of 762 sand fly specimens, representing 12 species across seven genera, were collected. Lutzomyia longipalpis was the most abundant species, comprising 57.6% of the collected specimens, followed by Nyssomyia neivai (19.6%) and Nyssomyia whitmani (10.5%). Species richness and diversity varied among collection periods, with the highest diversity observed in January 2019. Molecular analysis detected Leishmania DNA in 12.5% of the sand fly specimens, with Le. infantum being the predominant species. Blood meal analysis revealed feeding on multiple vertebrate species, including humans, rats, dogs, and chickens. The presence of Leishmania DNA in sand flies, and the identification of human blood meals, highlight the potential role of these species in VL transmission. These findings underscore the importance of continued surveillance and control measures to prevent the spread of VL and reduce transmission risk in the region.

Leishmania (Sauroleishmania) tarentolae versus pathogenic species: comparative evaluation of protease activity, glycoconjugates, resistance to complement and metabolome composition

BACKGROUND Leishmania tarentolae is a non-pathogenic species found in lizards representing an important model for Leishmania biology. However, several aspects of this Sauroleishmania remain unknown to explain its low level of virulence. OBJECTIVES We reported several aspects of L. tarentolae biology including glycoconjugates, proteolytic activities and metabolome composition in comparison to pathogenic species (Leishmania amazonensis, Leishmania braziliensis, Leishmania infantum and Leishmania major). METHODS Parasites were cultured for extraction and purification of lipophosphoglycan (LPG), immunofluorescence probing with anti-gp63 and resistance against complement. Parasite extracts were also tested for proteases activity and metabolome composition. FINDINGS Leishmania tarentolae does not express LPG on its surface. It expresses gp63 at lower levels compared to pathogenic species and, is highly sensitive to complement-mediated lysis. This species also lacks intracellular/extracellular activities of proteolytic enzymes. It has metabolic differences with pathogenic species, exhibiting a lower abundance of metabolites including ABC transporters, biosynthesis of unsaturated fatty acids and steroids, TCA cycle, glycine/serine/threonine metabolism, glyoxylate/dicarboxylate metabolism and pentose-phosphate pathways. MAIN CONCLUSIONS The non-pathogenic phenotype of L. tarentolae is associated with alterations in several biochemical and molecular features. This reinforces the need of comparative studies between pathogenic and non-pathogenic species to elucidate the molecular mechanisms of virulence during host-parasite interactions.

Clinical and Laboratory Response of Domiciled Dogs with Visceral Leishmaniasis Treated with Miltefosine and Allopurinol.

Canine visceral leishmaniasis (CVL) remains a significant disease worldwide. In Brazil, its treatment is performed using miltefosine, which has demonstrated promising outcomes in dogs. This study represents the first attempt to treat and monitor dogs with CVL in natural conditions over the course of one year. The dogs were divided into two groups: G1 received miltefosine and allopurinol for 28 days, while G2 received miltefosine for 28 days, followed by allopurinol for one year. The follow-up involved clinical, hematological, and biochemical evaluations, as well as the detection of Leishmania DNA in skin and bone marrow samples. By the end of the follow-up, dogs in G2 exhibited improved staging compared to their initial conditions, whereas those in G1 showed worsened staging. Leishmania DNA in skin and bone marrow decreased between 6 and 12 months after treatment. Our observations indicate that the treatment using miltefosine reduces the detection of the parasite in the skin and bone marrow for up to one year following its administration. The continuous use of allopurinol contributes to control of the disease in dogs. These findings provide valuable insights into the response of dogs treated in natural conditions, offering essential information for veterinarians and public health authorities.

First report of putative Leishmania RNA virus 2 (LRV2) in Leishmania infantum strains from canine and human visceral leishmaniasis cases in the southeast of Brazil.

BACKGROUND: Leishmania RNA virus 1 (LRV1) is commonly found in South American Leishmania parasites belonging to the subgenus Viannia, whereas Leishmania RNA virus 2 (LRV2) was previously thought to be restricted to the Old-World pathogens of the subgenus Leishmania. OBJECTIVES: In this study, we investigated the presence of LRV2 in strains of Leishmania (L.) infantum, the causative agent of visceral leishmaniasis (VL), originating from different hosts, clinical forms, and geographical regions. METHODS: A total of seventy-one isolates were screened for LRV2 using semi-nested reverse transcription-polymerase chain reaction (RT-PCR) targeting the RNA-dependent RNA polymerase (RdRp) gene. FINDINGS: We detected LRV2 in two L. infantum isolates (CUR268 and HP-EMO) from canine and human cases, respectively. MAIN CONCLUSIONS: To the best of our knowledge, this is the first detection of LRV2 in the New World.

Leishmania species infection of bats: A systematic review.

A wide variety of mammals, including domestic and wild species, have been considered potential hosts and reservoirs for Leishmania. Bats have longevity, dispersal capacity, and adaptability to synotropic environments, characteristics that may favor their role in maintaining the life cycle of parasites. Therefore, the objective of this study was to carry out a worldwide systematic review of the occurrence of Leishmania species in bats, as well as to identify associations between eating habits and the type of sample collected with the occurrence of the infection. Data were obtained from a bibliographic search for studies that used molecular methods to identify parasites, employing the keywords "bats" AND "Leishmania" and their synonyms. We found 68 original studies, of which 20 were included in this review. Most studies were conducted in Brazil (60 %) and only 10 % were conducted in Old World countries. In all, 48 bat species were recorded that hosted seven Leishmania species, resulting in 62 different host-parasite interactions, and the Leishmania infantum interaction with bat species presented higher frequency. There was no significant difference between Leishmania species richness, infection percentage, and type of sample analyzed, but in general, it is observed that the use of different biological samples seems to expand the possibility of parasite detection. The patterns observed here indicate that bats can become infected with a wide variety of Leishmania species and likely play an important role in maintaining the parasite's life cycle. Thus, we suggest that studies aimed at understanding the transmission cycle of leishmaniasis include the investigation of bats as potential hosts or reservoirs of Leishmania.

Increased Risk of American Tegumentary Leishmaniasis in an Urban and Rural Area of Caratinga, Brazil between 2016 and 2021.

We used spatial analysis tools to examine the epidemiological situation and spatial distribution of American tegumentary leishmaniasis in the municipality of Caratinga between 2016 and 2021. In addition, potential sandfly vectors were captured. All information used in this study was retrieved from public health archives and confirmed in the state health services databases. All cases were analyzed using Geographic Information Systems software. In addition, sandfly collections and molecular detection of Leishmania were carried out in areas with the highest number of cases. During the analyzed period, American tegumentary leishmaniasis (ATL) cases increased and remained high in the last years. The hotspots included urban areas of Caratinga city and the districts of Patrocínio of Caratinga and Sapucaia. The species Nyssomyia whitmani, Nyssomyia intermedia, and Migonemyia migonei were the most abundant species and the ITS1-polymerase chain reaction technique detected Leishmania DNA in these species. On the basis of our analyses, the urbanization of ATL in Caratinga has taken place in recent years. Because of the increase in the number of human cases and the presence of vectors, it is recommended that health authorities focus on control measures in hotspots.

First report of putative Leishmania RNA virus 2 (LRV2) in Leishmania infantum strains from canine and human visceral leishmaniasis cases in the southeast of Brazil

BACKGROUND Leishmania RNA virus 1 (LRV1) is commonly found in South American Leishmania parasites belonging to the subgenus Viannia, whereas Leishmania RNA virus 2 (LRV2) was previously thought to be restricted to the Old-World pathogens of the subgenus Leishmania. OBJECTIVES In this study, we investigated the presence of LRV2 in strains of Leishmania (L.) infantum, the causative agent of visceral leishmaniasis (VL), originating from different hosts, clinical forms, and geographical regions. METHODS A total of seventy-one isolates were screened for LRV2 using semi-nested reverse transcription-polymerase chain reaction (RT-PCR) targeting the RNA-dependent RNA polymerase (RdRp) gene. FINDINGS We detected LRV2 in two L. infantum isolates (CUR268 and HP-EMO) from canine and human cases, respectively. MAIN CONCLUSIONS To the best of our knowledge, this is the first detection of LRV2 in the New World.

Characterization of Differentially Abundant Proteins Among Leishmania (Viannia) braziliensis Strains Isolated From Atypical or Typical Lesions.

Leishmania (Viannia) braziliensis is the main etiological agent of cutaneous and mucocutaneous leishmaniasis in Latin America. Non-ulcerated atypical tegumentary leishmaniasis cases caused by L. braziliensis have been reported in several regions of the American continent, including the Xacriabá indigenous reserve in São João das Missões/Minas Gerais, Brazil. Parasites isolated from these atypical clinical lesions are resistant to antimony-based therapeutics. In the present study, proteins displaying differential abundance in two strains of L. braziliensis isolated from patients with atypical lesions compared with four strains isolated from patients with typical lesions were identified using a quantitative proteomics approach based on tandem mass tag labeling (TMT) and mass spectrometry. A total of 532 (P<0.05) differentially abundant proteins were identified (298 upregulated and 234 downregulated) in strains from atypical lesions compared to strains from typical lesions. Prominent positively regulated proteins in atypical strains included those that may confer greater survival inside macrophages, proteins related to antimony resistance, and proteins associated with higher peroxidase activity. Additionally, we identified proteins showing potential as new drug and vaccine targets. Our findings contribute to the characterization of these intriguing L. braziliensis strains and provide a novel perspective on Atypical Cutaneous Leishmaniasis (ACL) cases that have been associated with therapeutic failures.

Molecular survey of Leishmania spp. in skin samples of capybaras (Hydrochoerus hydrochaeris) from different areas of Brazil / Investigação molecular de Leishmania spp. em amostras de pele de capivaras (Hydrochoerus hydrochaeris) de diferentes regiões do Brasil

Leishmaniases comprise a spectrum of diseases caused by protozoan parasites of the genus Leishmania, with some species of rodents being incriminated as reservoirs. The capybara is the largest extant rodent species in the world and is widely distributed in South America. The occurrence of infection by Leishmania spp. was investigated in capybaras captured in Brazil during 2015­2019 from established populations in five highly anthropic areas of the state of São Paulo and two natural areas of the states of Mato Grosso and Mato Grosso do Sul. A total of 186 individuals were captured and subjected to abdominal skin biopsy. All skin samples were Leishmania kDNA-negative, suggesting that capybaras have no role in the transmission cycles of Leishmania species in the studied areas despite the well-known role of other rodents in the life cycle of Leishmania spp.(AU)

As leishmanioses compreendem um espectro de doenças causadas por protozoários do gênero Leishmania e algumas espécies de roedores são incriminadas como reservatórios de Leishmania spp. As capivaras compreendem a maior espécie de roedores existentes e são amplamente distribuídas na América do Sul. Para investigar a ocorrência de infecção por Leishmania spp. em capivaras, durante os anos de 2015-2019 capivaras foram capturadas em cinco áreas antrópicas do estado de São Paulo e em duas áreas naturais dos estados do Mato Grosso e do Mato Grosso do Sul, todos esses ambientes com populações de capivaras estabelecidas. Um total de 186 indivíduos foram capturados e submetidos à biópsia de pele abdominal. Todas as amostras de pele foram negativas para o alvo kDNA, assim, os dados sugerem que nas áreas estudadas as capivaras não têm papel no ciclo de transmissão de espécies de Leishmania spp., apesar do papel bem conhecido de outros roedores no ciclo de vida de Leishmania spp.(AU)

Leishmania diversity in bats from an endemic area for visceral and cutaneous leishmaniasis in Southeastern Brazil.

This study aimed to determine the occurrence of Leishmania infection in bats in urban and wild areas in an endemic municipality for visceral and cutaneous leishmaniasis in Minas Gerais, Brazil. Between April 2014 to April 2015, 247 bats were captured and classified into 26 species belonging to Phyllostomidae (90.7%), Vespertilionidae (8.1%) and Molossidae (1.2%) families. Blood samples from 247 bats were collected and submitted to nested-PCR, targeting the variable V7-V8 region of the SSU rRNA gene, followed by sequencing of the PCR product. The overall infection rate of Leishmania spp. in bats was 4.4%. Of the eleven bats infected, ten were frugivorous bats: Artibeus planirostris (8/11), Artibeus lituratus (1/11) and Artibeus cinereus (1/11) and one a nectarivorous bat (Glossophaga soricina). None of the individuals exhibited macroscopic alterations in the skin, spleen or liver. Phylogenetic analysis separated Leishmania species in clades corresponding to the subgenera Viannia, Leishmania, and Mundinia, and supported that the isolates characterized in the present study clustered closely with Leishmania (Viannia) sp., Leishmania (Leishmania) infantum and Leishmania (Leishmania) amazonensis. Here we report for the first time the bat Artibeus cinereus as a host of Leishmania (Leishmania) amazonensis. In the study we found that the mean abundance of bats did not differ in wild habitats and urban areas and that bat-parasite interactions were similarly distributed in the two environments. On the other hand, further studies should be conducted in more recent times to verify whether there have been changes in these parameters.

Cross-sectional study on Brucella spp., Leptospira spp. and Salmonella spp. in bats from Montes Claros, Minas Gerais, Brazil.

The understanding on the role of bats in the ecology of zoonotic diseases, especially its relevance as a carrier of pathogens, is important for the determination of preventive measures considering the One Health context. The present study aimed to investigate the presence of Brucella spp., Leptospira spp. and Salmonella spp. in blood (n = 163), liver (n = 35) and spleen (n = 62) samples from bats captured in Montes Claros, Minas Gerais, Brazil. Only Salmonella spp. was found in a blood sample of an insectivorous female bat of the species Lasiurus blossevilli, evidencing the capacity of this animal species to host this pathogen. In conclusion, our results in bats from Montes Claros indicate that they do not act as hosts for Brucella spp. and Leptospira spp., although being potential carriers of Salmonella spp. in a low prevalence.

A follow-up study (2007-2018) on American Tegumentary Leishmaniasis in the municipality of Caratinga, Minas Gerais State, Brazil: Spatial analyses and sand fly collection.

BACKGROUND: The municipality of Caratinga is an important endemic area for American Tegumentary Leishmaniasis (ATL) and no epidemiological studies were performed during the past two decades. Here, we analyzed the epidemiological situation and the geographical distribution of ATL cases in the municipality of Caratinga from 2007 to 2018 using geographic information systems (GIS). Also, we evaluated the impact of several demographic parameters in ATL distribution and the sand flies incriminated in its transmission. METHODS: All demographic information (gender, age, educational level, clinical form, diagnostic criteria and case evolution) used in this study was retrieved from the public health archives and confirmed in the State Health Services databases. All cases were analyzed using GIS software based on ATL distribution. Also, non-systematic sand fly collections and molecular detection of Leishmania were performed in the hotspots. RESULTS AND CONCLUSIONS: During the period, ATL cases continued and increased especially in the past years (2016-2018). Hotspots included urban Caratinga areas and the districts of Patrocínio de Caratinga and Sapucaia. The species Nyssomyia whitmani, Nyssomyia intermedia, Migonemyia migonei and Evandromyia cortelezzii complex were captured. However, ITS1-PCR did not detect Leishmania DNA in those insects. Based on our analyses, urbanization of ATL in Caratinga has occurred in the past years. Due to the increase in the number of cases and vectors presence, it is recommended that health authorities focus on control measures in the most affected areas (Patrocínio of Caratinga and Sapucaia districts and urban Caratinga).

Correlations between tissue parasite load and common clinical signs in dogs naturally infected by Leishmania infantum.

qPCR is being used for the quantification of parasite load in different tissues of dogs infected by Leishmania infantum with or without clinical manifestations. It may be employed in the diagnosis, monitoring of the infection during treatment, and clinical studies for validation of vaccines. Aimed at enhancing the molecular diagnosis and the subsequent monitoring of the infection, this study evaluated the parasite load in several tissues from dogs infected by Leishmania infantum, showing different clinical status. Thus, the qPCR was performed on skin, conjunctival swab, popliteal lymph node, and bone marrow puncture samples taken from 65 dogs naturally infected by L. infantum. Dogs were divided into three groups per clinical score: group 1 (n = 12), included animals with zero points and no clinical manifestations of the disease; group 2 (n = 35), included animals with a score ranging from 1 to 5 points and moderate clinical manifestations; and group 3 (n = 18), included dogs with a score ranging from 6 to 11 points and intensive clinical manifestations. Another analysis was performed classifying the animals into two groups, considering the presence of, or lack of clinical signs of the disease. Analyses of these results showed that the skin was the tissue with a higher parasite load, followed by popliteal lymph node and bone marrow punctures, and conjunctival swab samples having the lowest loads. Furthermore, the skin was also the tissue with the highest parasite load when evaluating the groups individually. Animals in group 3, with intensive clinical manifestations, showed a higher parasite load in different tissues when compared to animals from groups 1 and 2. Finally, animals with clinical manifestations of the disease showed a higher parasite load when compared to dogs with no manifestations. The importance of the dog as a reservoir of L. infantum in nature is reinforced by the demonstration of skin having the highest amount of parasites/µL in this study's analysis, as well as the fact that skin is the main point of access to the parasite vector. Also, a strong and positive correlation between the intensity of clinical manifestations and the increase of parasite load in the skin was observed. In conclusion, skin was the tissue that was demonstrated to be the best option for the molecular diagnosis of L. infantum infection in dogs with varying clinical statuses used in this study.

Chemokines in Leishmaniasis: Map of cell movements highlights the landscape of infection and pathogenesis.

Pathogen interactions with the host immune response components are critical for establishing protective immunity and pathological responses against Leishmania parasites. A predominant proinflammatory profile associated with enhanced phagocytosis trigger a cell-mediated immune response that is relevant to infection control. On the other hand, an anti-inflammatory phenotype, correlated with a predominant modulated/regulatory response, favors intracellular proliferation of Leishmania parasites and disease progression. In this context, chemokines play an important role in determining cellular composition at inflammatory sites. Leishmania infection induces the expression of various chemokines and chemokine receptors in the mammalian host, which can subvert the host immune responses. Indeed, the balance and dynamic changes in cytokines and chemokines may control or predict the disease outcome. In this review, we address our current knowledge regarding the chemokines and chemokines receptors' role in the immunopathogenesis of Tegumentary and Visceral Leishmaniasis.

Serological study of feline leishmaniasis and molecular detection of Leishmania infantum and Leishmania braziliensis in cats (Felis catus).

Blood samples and swabs from ocular conjunctiva and mouth were obtained from 64 cats. Of 64 serum samples, 19 were positive for Leishmania antibodies by ELISA (29.80%). Eight cats were positive by PCR (12.5%) in swab samples from mouth and/or ocular mucosa. Poor kappa agreement between serological and molecular results (k = 0.16) was obtained. From five positive PCR samples one was L. braziliensis and four were L. infantum. Phylogenetic analysis performed with the five isolates of Leishmania, showed that samples of L. infantum isolated from the cats were phylogenetically close to those isolated from domestic dogs in Brazil, while the L. braziliensis is very similar to the one described in humans in Venezuela. The study demonstrated that, despite high seropositivity for Leishmania in cats living in the study region, poor agreement between serological and molecular results indicate that positive serology is not indicative of Leishmania infection in cats. Parasite DNA can be detected in ocular conjunctiva and oral swabs from cats, indicating that such samples could be used for diagnosis. Results of phylogenetic analyzes show that L. infantum circulating in Brazil is capable of infecting different hosts, demonstrating the parasite's ability to overcome the interspecies barrier.

Diversity of phlebotomine sand flies and molecular detection of trypanosomatids in Brumadinho, Minas Gerais, Brazil.

This study aimed to describe the sand fly fauna and detect trypanosomatids in these insects from Casa Branca, state of Minas Gerais, Brazil, an endemic area of both visceral (VL) and tegumentary leishmaniasis (TL). Sand flies were collected bimonthly from May 2013 to July 2014, using automatic light traps exposed for three consecutive nights in peridomiciliary areas of nine houses with previous reports of VL and TL. ITS1-PCR and DNA sequencing were performed for trypanosomatids identification. A total of 16,771 sand flies were collected belonging to 23 species. The most abundant species was Nyssomyia whitmani (Antunes & Coutinho, 1939) (70.9%), followed by Lutzomyia longipalpis (Lutz & Neiva, 1912) (15.2%) and Migonemyia migonei (França, 1920) (9.1%). Leishmania amazonensis DNA was detected in Ny. whitmani (four pools) and Le. braziliensis DNA was detected in Psychodopygus lloydi (one pool). In seven pools of Ny. whitmani and in one pool of Lu. longipalpis positive for Leishmania DNA, the parasite species was not determined due to the low quality of the sequences. Moreover, DNA of Herpetomonas spp. was detected in Ny. whitmani (two pools) and Cortelezzii complex (one pool). DNA of Crithidia spp. was detected in Ny. whitmani and Ps. lloydi (both one pool). Our results suggest that Ny. whitmani may be involved in the transmission of Le. amazonensis in the study area. The molecular detection of Le. amazonensis suggests the presence of this species in a sylvatic cycle between vertebrate and invertebrate hosts in the region of Casa Branca. Our data also reveal the occurrence of other non-Leishmania trypanosomatids in sand flies in Casa Branca District.

Visceral leishmaniasis: a practical strategy for quantitative molecular diagnosis in naturally infected dogs.

The diagnosis of canine visceral leishmaniasis (CVL) has been a problem for public health services due to the variety of clinical signs similar to other diseases and low sensitivity and specificity of available tests. In this sense, our main objective was to develop a simple, rapid, and accurate quantitative real-time PCR (qPCR) diagnosis for CVL. Thus, low-invasive samples from bone marrow (BM), popliteal lymph nodes (PLN), and conjunctival swabs (CS) were selected from negative and VL-positive dogs, using as gold standard, immunological and parasitological tests performed with different tissues. Oligonucleotides for Leishmania infantum kDNA were designed and the limit of quantification and amplification efficiency of the qPCR were determined using tissue-specific standards produced with DNA from those different tissues, mixed with DNA from a known amount of L. infantum promastigotes. Endogenous control was used to validate a comparative Ct method, and tissue parasite concentrations were estimated by comparison with tissue-specific reference standard samples. The overall analysis of the qPCR data suggests the following ranking for tissue choice: PLN > BM > CS. Finally, we have concluded that this molecular approach simplifies and accelerates the quantitative diagnostic process because it is easy to perform, requiring no DNA dosing or standard curve application, and it shows good diagnostic parameters, especially when using popliteal lymph node samples.

Comparative PCR-based diagnosis for the detection of Leishmania infantum in naturally infected dogs.

The applicability of molecular biology/PCR for canine visceral leishmaniasis diagnosis presents challenges, mainly due to the diversity of targets described. The objectives of this study were to compare the sensitivities and reliability of five targets (kDNA/120, kDNA/145, ITS1, hsp70/234 and hsp70/1300) in four different tissue samples (bone marrow, popliteal lymph node, skin and conjunctival swab). Sixty-five dogs (32 males and 33 females) naturally infected with Leishmania infantum and ten dogs without infection were examined. Dogs were characterized by serological and parasitological methods. The parasitological test was considered the gold standard for analysis. All tests presented high specificity 100% (95% CI 0.72-1), and variable sensitivity. The targets kDNA/145, ITS1, kDNA/120, hsp70/234 and hsp70/1300 detected 100% (65/65), 93.4% (61/65), 92.3% (60/65), 84.61% (55/65) and 72.3% (77/65) of positive animals respectively. The performance of PCR methods was analyzed in two different scenarios. The highest sensitivity value identified in all scenarios studied was kDNA/145. Our results suggest that popliteal lymph node and conjunctival swab samples, besides being less invasive collections, represent a good substratum for PCR-based diagnosis, and the target kDNA/145 is the best choice for detecting L. infantum DNA in naturally infected dogs.

Leishmania presence in bats in areas endemic for leishmaniasis in central-west Brazil.

Leishmaniasis involves the participation of several species of both wild and domestic mammal hosts and sandfly vectors, which demonstrates the eco-epidemiological complexity observed in this disease. Bats are among the most abundant types of mammals and the scarcity of research on Leishmania infection in these animals gives evidence of the importance of new studies that aim to clarify this relationship. This study aimed to detect the Leishmania spp. in bats. 146 bats, representing 16 different species belonging to the Molossidae, Vespertilionidae, and Phyllostomidae families, were received and processed for collection of tissues. Skin samples were collected from 100% of the bats, and liver samples were collected from 87% (n = 127). After evaluating the quality of the DNA extracted by means of PCR directed to the IRBP gene, the samples considered suitable for the Leishmania detection test were submitted for PCR directed to Leishmania kDNA, and to confirm positivity, were tested to the SSUrRNA gene-directed Nested-PCR. The Leishmania presence in the species Molossus pretiosus, Nyctinomops macrotis, and Lasiurus cinereus are the first reports this encounter in these species of bats in Brazil. Furthermore, new species of bats as possible hosts for L. infantum are reported, such as Molossus pretiosus, Myotis nigricans, Nyctinomops laticaudatus, Nyctinomops macrotis, and, for L. braziliensis, Lasiurus cinereus and Cynomops planirostris. These findings in bats in an area endemic for leishmaniasis indicate that these animals may be involved in sustaining the disease cycle in this location.