Unveiling the Enigmatic nature of six neglected Amazonian Leishmania (Viannia) species using the hamster model: Virulence, Histopathology and prospection of LRV1.

American tegumentary leishmaniasis (ATL) is highly endemic in the Amazon basin and occurs in all South American countries, except Chile and Uruguay. Most Brazilian ATL cases are due to Leishmania (Viannia) braziliensis, however other neglected Amazonian species are being increasingly reported. They belong to the subgenus L. (Viannia) and information on suitable models to understand immunopathology are scarce. Here, we explored the use of the golden hamster Mesocricetus auratus and its macrophages as a model for L. (Viannia) species. We also studied the interaction of parasite glycoconjugates (LPGs and GIPLs) in murine macrophages. The following strains were used: L. (V.) braziliensis (MHOM/BR/2001/BA788), L. (V.) guyanensis (MHOM/BR/85/M9945), L. (V.) shawi (MHOM/BR/96/M15789), L. (V.) lindenbergi (MHOM/BR/98/M15733) and L. (V.) naiffi (MDAS/BR/79/M5533). In vivo infections were initiated by injecting parasites into the footpad and were followed up at 20- and 40-days PI. Parasites were mixed with salivary gland extract (SGE) from wild-captured Nyssomyia neivai prior to in vivo infections. Animals were euthanized for histopathological evaluation of the footpads, spleen, and liver. The parasite burden was evaluated in the skin and draining lymph nodes. In vitro infections used resident peritoneal macrophages and THP-1 monocytes infected with all species using a MOI (1:10). For biochemical studies, glycoconjugates (LPGs and GIPLs) were extracted, purified, and biochemically characterized using fluorophore-assisted carbohydrate electrophoresis (FACE). They were functionally evaluated after incubation with macrophages from C57BL/6 mice and knockouts (TLR2-/- and TLR4-/-) for nitric oxide (NO) and cytokine/chemokine production. All species, except L. (V.) guyanensis, failed to generate evident macroscopic lesions 40 days PI. The L. (V.) guyanensis lesions were swollen but did not ulcerate and microscopically were characterized by an intense inflammatory exudate. Despite the fact the other species did not produce visible skin lesions there was no or mild pro-inflammatory infiltration at the inoculation site and parasites survived in the hamster skin/lymph nodes and even visceralized. Although none of the species caused severe disease in the hamster, they differentially infected peritoneal macrophages in vitro. LPGs and GIPLs were able to differentially trigger NO and cytokine production via TLR2/TLR4 and TLR4, respectively. The presence of a sidechain in L. (V.) lainsoni LPG (type II) may be responsible for its higher proinflammatory activity. After Principal Component analyses using all phenotypic features, the clustering of L. (V.) lainsoni was separated from all the other L. (Viannia) species. We conclude that M. auratus was a suitable in vivo model for at least four dermotropic L. (Viannia) species. However, in vitro studies using peritoneal cells are a suitable alternative for understanding interactions of the six L. (Viannia) species used here. LRV1 presence was found in L. (V.) guyanensis and L. (V.) shawi with no apparent correlation with virulence in vitro and in vivo. Finally, parasite glycoconjugates were able to functionally trigger various innate immune responses in murine macrophages via TLRs consistent with their inflammatory profile in vivo.

Sporocysts of Sarcocystis bertrami (syn. Sarcocystis fayeri) shed by dogs: Molecular analysis, morphometry and pattern of excretion.

Sarcocystis bertrami (synonym: Sarcocystis fayeri) is a coccidian parasite that infects horses and donkeys in several countries. Dogs are known as definitive hosts of the parasite, however, the patent period is not well defined, and S. bertrami shed by dogs has never been confirmed by molecular methods. Here we investigated the shedding of S. bertrami by experimentally infected dogs and examined the excreted parasites by morphological and molecular tools. Three dogs of small breeds (one Yorkshire terrier and two miniature Pinschers) were acquired with ages of 30 and 60 days and were exclusively fed commercial dog food. Two dogs consumed equine muscle tissues containing cysts of S. bertrami. The third dog served as negative control and was simultaneously fed commercial dog food. The two animals that received equine tissues shed sporocysts and/or oocysts in their feces after prepatent periods of 13 and 23 days. The patent periods were 47 and 14 days. Sporocysts showed average dimensions of 14.19 µm (± 0.53) x 10.06 µm (± 0.44). The control dog did not shed sporocysts or oocysts of the parasite. Interestingly, patent periods had never been reported, and for one dog, the patent period (47 days) was longer than that reported for other Sarcocystidae parasites. PCRs to the gene 18S and to the internal transcribed spacer 1 (ITS1) of the rDNA were successfully performed with DNA extracted from sporocysts. ITS1 sequences were also obtained from the equine tissue cysts used to infect the dogs. Nucleotide sequences of cloned fragments of 18S from sporocysts, and ITS1 from both stages (tissue cysts and sporocysts) matched with S. bertrami (18S: 97.50-99.88 %; ITS1: 88.76-95.21 %), although high molecular diversity was observed with data from these loci. PCR to cox1 using sporocysts' DNA failed to amplify any product. The possibility of the existence of an additional and undescribed Sarcocystis species in the excreted sporocysts, besides S. bertrami, cannot be excluded from this experiment. To our knowledge, this is the first molecular confirmation of S. bertrami in canine feces. Sporocyst dimensions and prepatent periods observed in this study were similar to those previously described for S. bertrami and S. fayeri. In conclusion, the molecular, morphological and biological data generated here fit in previous descriptions for both S. bertrami and S. fayeri.

Targeting Leishmania Promastigotes and Amastigotes Forms through Amino Acids and Peptides: A Promising Therapeutic Strategy.

Millions of people worldwide are affected by leishmaniasis, caused by the Leishmania parasite. Effective treatment is challenging due to the biological complexity of the parasite, drug toxicity, and increasing resistance to conventional drugs. To combat this disease, the development of specific strategies to target and selectively eliminate the parasite is crucial. This Review highlights the importance of amino acids in the developmental stages of Leishmania as a factor determining whether the infection progresses or is suppressed. It also explores the use of peptides as alternatives in parasite control and the development of novel targeted treatments. While these strategies show promise for more effective and targeted treatment, further studies to address the remaining challenges are imperative.

Synthetic Peptides Selected by Immunoinformatics as Potential Tools for the Specific Diagnosis of Canine Visceral Leishmaniasis.

Diagnosing canine visceral leishmaniasis (CVL) in Brazil faces challenges due to the limitations regarding the sensitivity and specificity of the current diagnostic protocol. Therefore, it is urgent to map new antigens or enhance the existing ones for future diagnostic techniques. Immunoinformatic tools are promising in the identification of new potential epitopes or antigen candidates. In this study, we evaluated peptides selected by epitope prediction for CVL serodiagnosis in ELISA assays. Ten B-cell epitopes were immunogenic in silico, but two peptides (peptides No. 45 and No. 48) showed the best performance in vitro. The selected peptides, both individually and in combination, were highly diagnostically accurate, with sensitivities ranging from 86.4% to 100% and with a specificity of approximately 90%. We observed that the combination of peptides showed better performance when compared to peptide alone, by detecting all asymptomatic dogs, showing lower cross-reactivity in sera from dogs with other canine infections, and did not detect vaccinated animals. Moreover, our data indicate the potential use of immunoinformatic tools associated with ELISA assays for the selection and evaluation of potential new targets, such as peptides, applied to the diagnosis of CVL.

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.

Pathogen-associated molecular patterns (PAMPs) derived from Leishmania and bacteria increase gene expression of antimicrobial peptides and gut surface proteins in sand flies.

The interaction between pathogens and vectors' physiology can impact parasite transmission. Studying this interaction at the molecular level can help in developing control strategies. We study leishmaniases, diseases caused by Leishmania parasites transmitted by sand fly vectors, posing a significant global public health concern. Lipophosphoglycan (LPG), the major surface glycoconjugate of Leishmania, has been described to have several roles throughout the parasite's life cycle, both in the insect and vertebrate hosts. In addition, the sand fly midgut possesses a rich microbiota expressing lipopolysaccharides (LPS). However, the effect of LPG and LPS on the gene expression of sand fly midgut proteins or immunity effectors has not yet been documented. We experimentally fed Lutzomyia longipalpis and Phlebotomus papatasi sand flies with blood containing purified LPG from Leishmania infantum, Leishmania major, or LPS from Escherichia coli. The effect on the expression of genes encoding gut proteins galectin and mucin, digestive enzymes trypsin and chymotrypsin, and antimicrobial peptides (AMPs) attacin and defensins was assessed by quantitative PCR (qPCR). The gene expression of a mucin-like protein in L. longipalpis was increased by L. infantum LPG and E. coli LPS. The gene expression of a galectin was increased in L. longipalpis by L. major LPG, and in P. papatasi by E. coli LPS. Nevertheless, the gene expression of trypsins and chymotrypsins did not significantly change. On the other hand, both L. infantum and L. major LPG significantly enhanced expression of the AMP attacin in both sand fly species and defensin in L. longipalpis. In addition, E. coli LPS increased the expression of attacin and defensin in L. longipalpis. Our study showed that Leishmania LPG and E. coli LPS differentially modulate the expression of sand fly genes involved in gut maintenance and defence. This suggests that the glycoconjugates from microbiota or Leishmania may increase the vector's immune response and the gene expression of a gut coating protein in a permissive vector.

Intricate balance of dually-localized catalase modulates infectivity of Leptomonas seymouri (Kinetoplastea: Trypanosomatidae).

Nearly all aerobic organisms are equipped with catalases, powerful enzymes scavenging hydrogen peroxide and facilitating defense against harmful reactive oxygen species. In trypanosomatids, this enzyme was not present in the common ancestor, yet it had been independently acquired by different lineages of monoxenous trypanosomatids from different bacteria at least three times. This observation posited an obvious question: why was catalase so "sought after" if many trypanosomatid groups do just fine without it? In this work, we analyzed subcellular localization and function of catalase in Leptomonas seymouri. We demonstrated that this enzyme is present in the cytoplasm and a subset of glycosomes, and that its cytoplasmic retention is H2O2-dependent. The ablation of catalase in this parasite is not detrimental in vivo, while its overexpression resulted in a substantially higher parasite load in the experimental infection of Dysdercus peruvianus. We propose that the capacity of studied flagellates to modulate the catalase activity in the midgut of its insect host facilitates their development and protects them from oxidative damage at elevated temperatures.

Karyotypic changes and diversification time in Epinephelidae groupers (Perciformes). Implications on reproductive isolation.

Groupers (Epinephelidae and Serranidae) have attracted special attention to fish farming, and their species offer good opportunities for successful hybridizations. Cytogenetic data allow a better understanding of the role of karyotypic diversification in the acquisition of post-zygotic reproductive isolation (RI). Thus, chromosomal analyses were performed on E. striatus (Caribbean Sea), E. coioides and E. tauvina (Indo-Pacific Region), using standard procedures and mapping of six repetitive DNA classes by the in situ hybridization. The three species have 2n=48 chromosomes. The karyotypes of E. coioides and E. striatus are composed only of acrocentric chromosomes (FN=48), while E. tauvina has 8 submetacentric chromosomes (FN=56). Heterochromatin has a preferential centromeric distribution, and the microsatellite repeats are dispersed throughout the chromosomes of all species. The 18S and 5S rDNA sites are unique but show a colocalization arrangement in E. tauvina and E. striatus. The chromosomal organization suggests that the three species still maintain a significant amount of syntenic regions. The range of the karyotype divergence and the RI levels showed low, but goes turn proportionally greater in relation to the divergence time between the parental species. The slow acquisition of postzygotic RI is consistent with the high karyotype homogeneity presented by Epinephelidae family.

Corrigendum: Evolutionary tracks of chromosomal diversification in surgeonfishes (Acanthuridae: Acanthurus) along the world's biogeographic domains.

[This corrects the article DOI: 10.3389/fgene.2021.760244.].

The effect of micro-osteoperforations on the rate of maxillary incisors' retraction in orthodontic space closure: a randomized controlled clinical trial.

BACKGROUND: This single-centered randomized controlled clinical trial aimed to evaluate the effectiveness of micro-osteoperforations (MOPs) in accelerating the orthodontic retraction of maxillary incisors. METHODS: Forty-two patients aged 16-40 were recruited and randomly assigned into two groups, one which underwent MOPs (MOPG) in the buccal and palatal region of all maxillary incisors immediately before the start of retraction and one which did not (CG). Eligibility criteria included the orthodontic need for maxillary first premolars extraction and space closure in two phases. The primary outcome of the study consisted of measuring the rate of space closure and, consequently, the rate of incisors' retraction using digital model superimposition 14 days later and monthly thereafter for the next 4 months. The secondary outcomes included measuring anchorage loss, central incisors' inclination, and root length shortening, analyzed using cone beam computed tomography scans acquired before retraction and 4 months after retraction. Randomization was performed using QuickCalcs software. While clinical blinding was not possible, the image's examinator was blinded. RESULTS: Twenty-one patients were randomly assigned to each group. However, due to various reasons, a total of 37 patients (17 male and 20 female) were analyzed (mean age: 24.3 ± 8.1 years in the MOPG; 22.2 ± 4.2 years in the CG) during the trial. No statistically significant difference was found between the MOPG and the CG regarding the incisors' retraction measured at different time points at the incisal border (14 days, 0.4 mm vs. 0.5 mm; 1 month, 0.79 mm vs. 0.77 mm; 2 months, 1.47 mm vs. 1.41 mm; 3 months, 2.09 mm vs. 1.88 mm; 4 months, 2.62 mm vs. 2.29 mm) and at the cervical level (14 days, 0.28 mm vs. 0.30 mm; 1 month, 0.41 mm vs. 0.32 mm; 2 months, 0.89 mm vs. 0.61 mm; 3 months, 1.36 mm vs. 1.10 mm; 4 months, 1.73 mm vs. 1.39 mm). Similarly, no statistically significant differences were detected in the space closure, anchorage loss, central incisors' inclination, and radicular length between groups. No adverse effect was observed during the trial. CONCLUSIONS: MOPs did not accelerate the retraction of the maxillary incisors, nor were they associated with greater incisor inclination or root resorption. Trial registration ClinicalTrials.gov NCT03089996. Registered 24 March 2017- https://clinicaltrials.gov/ct2/show/NCT03089996 .

The absence of eosinophils is associated with early metastatic lesions in Leishmania amazonensis-infected mice.

BACKGROUND: Eosinophils are granulocytes that rapidly increase frequency in the bloodstream during helminthic infections and allergic responses. They are found in tissue infected by Leishmania during early disease, but their role during infection is not entirely understood. OBJECTIVES: We aim to compare the disease due to Leishmania amazonensis in BALB/c and Δdbl-GATA1 mice, which lack eosinophils. METHODS: BALB/c and Δdbl-GATA1 mice infected with L. amazonensis were observed for several weeks. The parasite load and dissemination pattern were assessed. FINDINGS: The Δdbl-GATA1 mice developed an anticipated dissemination of L. amazonensis and a worsening disease. No differences were found in the lesion development or the parasite load in the footpad among Δdbl-GATA1 mice and BALB/c eight weeks after infection. However, nine weeks after infection, massive growth of metastatic lesions appeared in several parts of the skin in Δdbl-GATA1 mice, weeks earlier than BALB/c. We observed increased parasites in the bloodstream, probably an essential dissemination route. Thirteen weeks after infection, metastatic lesions were found in all Δdbl-GATA1 mice. MAIN CONCLUSION: These results suggest a protective role of eosinophils in delaying the disease caused by L. amazonensis, although several limitations of this mice strain must be considered.

The absence of eosinophils is associated with early metastatic lesions in Leishmania amazonensis-infected mice

BACKGROUND Eosinophils are granulocytes that rapidly increase frequency in the bloodstream during helminthic infections and allergic responses. They are found in tissue infected by Leishmania during early disease, but their role during infection is not entirely understood. OBJECTIVES We aim to compare the disease due to Leishmania amazonensis in BALB/c and Δdbl-GATA1 mice, which lack eosinophils. METHODS BALB/c and Δdbl-GATA1 mice infected with L. amazonensis were observed for several weeks. The parasite load and dissemination pattern were assessed. FINDINGS The Δdbl-GATA1 mice developed an anticipated dissemination of L. amazonensis and a worsening disease. No differences were found in the lesion development or the parasite load in the footpad among Δdbl-GATA1 mice and BALB/c eight weeks after infection. However, nine weeks after infection, massive growth of metastatic lesions appeared in several parts of the skin in Δdbl-GATA1 mice, weeks earlier than BALB/c. We observed increased parasites in the bloodstream, probably an essential dissemination route. Thirteen weeks after infection, metastatic lesions were found in all Δdbl-GATA1 mice. MAIN CONCLUSION These results suggest a protective role of eosinophils in delaying the disease caused by L. amazonensis, although several limitations of this mice strain must be considered.

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.

Trypanosoma sp. infection in Boa constrictor snakes: morphological, hematological, clinical biochemistry, molecular, and phylogenetic characteristics.

There are few reports of Trypanosoma in snakes, as well as little information about its pathogenicity in these animals. Thus, the present study aimed to characterize Trypanosoma found in Boa constrictor snakes, to verify the influence of the parasitism on hematological and clinical biochemistry parameters, and to perform a phylogenetic study of the isolates. Blood samples from sixty-one boas were analyzed for the presence of trypanosomatids and by hematological and clinical biochemistry assays. The flagellates that were found in this analysis were used for cell culture, morphometry, and molecular analysis. Later, molecular typing phylogenetic studies were performed. Nine positive animals (14.75%) were identified by microscopy analysis. The hematological results showed that parasitized animals presented significantly lower levels of packed cell volume, hemoglobin, mean corpuscular volume, and mean corpuscular hemoglobin. In the leukogram, eosinophils and heterophils counts were higher in parasitized animals. Considering the molecular analyses, the isolates presented a higher identity of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the 18S small subunit ribosomal RNA (SSU rRNA) gene fragments with Trypanosoma serpentis. The phylogenetic tree, using the GAPDH, clustered all isolates with T. serpentis and Trypanosoma cascavelli. This is the first description of T. serpentis parasitizing boas and of the clinical changes caused by trypanosomatid infection in snakes.

Extracellular Vesicles Released by Leishmania (Leishmania) amazonensis Promastigotes with Distinct Virulence Profile Differently Modulate the Macrophage Functions.

Leishmania spp. is the aetiologic agent of leishmaniasis, a disease endemic in several developing countries. The parasite expresses and secretes several virulence factors that subvert the macrophage function and immune response. Extracellular vesicles (EVs) can carry molecules of the parasites that show immunomodulatory effects on macrophage activation and disease progression. In the present work, we detected a significantly higher expression of lpg3 and gp63 genes in Leishmania amazonensis promastigotes recovered after successive experimental infections (IVD-P) compared to those cultured for a long period (LT-P). In addition, we observed a significantly higher percentage of infection and internalized parasites in groups of macrophages infected with IVD-P. Macrophages previously treated with EVs from LT-P showed higher percentages of infection and production of inflammatory cytokines after the parasite challenge compared to the untreated ones. However, macrophages infected with parasites and treated with EVs did not reduce the parasite load. In addition, no synergistic effects were observed in the infected macrophages treated with EVs and reference drugs. In conclusion, parasites cultured for a long period in vitro and recovered from animals' infections, differently affected the macrophage response. Furthermore, EVs produced by these parasites affected the macrophage response in the early infection of these cells.

Acute, fatal Sarcocystis falcatula infection in rose-ringed parakeets (Psittacula krameri).

Sarcocystosis is an important avian disease that affects several intermediate host species. Birds not endemic from Americas, like Old World psittacine species, appear to be more susceptible to lethal infection than New World psittacine species. The aim of this study was to investigate the sudden death of rose-ringed parakeets (Psittacula krameri) in an exotic private parrot's aviary. Macroscopically, the most prevalent findings were severe lung congestion, slight superficial myocardial hemorrhagic lesions, enlarged liver and congestion of meningeal vessels. The initial diagnosis of sarcocystosis was made in all birds by microscopic observations of intravascular pulmonary schizonts, as well hepatitis, myocarditis, and nephritis. Immunohistochemistry for detection of Sarcocystis sp. antigen revealed an intense immunoreactivity in the lungs. Molecular identification of Sarcocystis falcatula were obtained by nested PCR and sequencing of amplified fragments of internal transcribed spacer 1 (ITS1) and three surface antigen-coding genes (SAG2, SAG3 and SAG4). SAG-based phylogenies showed a close relatedness of the isolate described here and S. falcatula previously detected in naturally infected native birds, which suggests that the isolates that affected ringnecks are a common isolate that circulates in Brazil.

Sarcocystis sp. shed by the common boa snake (Boa constrictor) in Brazil.

The genus Sarcocystis contains around 200 species and 25 of these infect snakes. Two Sarcocystis spp. shed by snakes have called special attention of the scientific community. S. nesbitti, which is shed by scrub pythons (Simalia amethistina), causes myopathy in humans that consume water or food contaminated with the parasite. Sporocysts of S. singaporensis, excreted by reticulated pythons (Malayopython reticulatus), is letal for rats and was successfully tested in the biological control of these rodents. A high biodiversity of snakes is found in Brazil, however, scarce information is available about Sarcocystis spp. in Brazilian snakes. Herein, we investigated Sarcocystis sp. in feces of the common boa (Boa constrictor) from Salvador, as it is widely distributed in Brazil and it is also bred in other countries. Feces of 65 boas were examined, and Sarcocystis sp. was found in 1/65 (1.53%) snakes. All snakes were alive, and for this reason, intestinal scrapping, which is the most sensitive method to detect the parasite, was not performed. Morphometric evaluation of sporocysts showed significant differences in their sizes. PCR and multilocus sequencing of four genetic markers (cox1, 18S, ITS1, and 28S) revealed that sporocysts corresponded to a new Sarcocystis species. Sequences of cox1 and 18S had identities of 100% and higher than 98%, respectively, with sequences obtained from the rodent Lagostomus maximus in Argentina. ITS1 and 28S sequences did not match with any known Sarcocystis sp. No ITS1 and 28S sequences were available for the Sarcocystis sp. found in the Argentinian L.maximus. Bioassay using the boa sporocysts was conducted in three mouse lineages and in Rattus norvegicus, but no parasitic stages were detected in these rodents. We concluded that the common boa is probably the definitive host of a new species of Sarcocystis sp. that has L. maximus or related rodents as intermediate hosts.

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.