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Environment influences the composition, distribution, and behavior of the vectors and mammalian hosts involved in the transmission of visceral leishmaniasis (VL), affecting the epidemiology of the disease. In Brazil, the urbanization process and canine cases of VL are indicators for local health authorities. This study aimed to investigate the occurrence of the canine visceral leishmaniasis (CVL) in Maranhão State, Brazil. Blood samples collected from 960 dogs from six municipalities and six different ecosystems (Baixada Maranhense, Mangue, Mata dos Cocais, Amazônia, Cerrado, and Restinga) to serological tests (enzyme-linked immunosorbent assay [ELISA], indirect fluorescence antibody test [IFAT], and chromatographic immunoassay methods [Dual Path Platform technology, DPP(®)]) and parasitological diagnosis. From serological tests, 11.14% (107) of the dogs were positive for CVL, with 59.16% (568), 14.5% (148), and 131% (126) positives to ELISA, DPP, and IFAT tests, respectively. Only seven animals (0.73%) were positive in a parasitological test. We also performed parasite isolation and phylogenetic characterization. All isolates of dogs obtained from Maranhão were grouped in a single branch with Leishmania infantum chagasi from Brazil. The ecosystem Amazonia presented the highest positivity rates to CVL in serological and parasitological tests. Brazilian biomes/ecosystems suffer large degradation and may favor, depending on climatic conditions, the installation of new diseases. In the case of VL, dogs are reservoirs of parasites and sentinels for human infection.

Trypanosoma and Leishmania are obligate parasites that cause important diseases in human and domestic animals. Wild mammals are the natural reservoirs of these parasites, which are transmitted by hematophagous arthropods. The present study aimed to detect the natural occurrence of trypanosomatids through serological diagnosis, PCR of whole blood and blood culture (hemoculture), and phylogenetic relationships using small subunit ribosomal DNA (SSU rDNA), cytochrome b, and glycosomal glyceraldehyde 3-phosphate dehydrogenase (gGAPDH) genes. Samples from 131 wild animals, including rodents, marsupials, and bats, were sampled in six areas in the state of Maranhão, in a transition zone of semiarid climates northeast of the equatorial humid Amazon. Serological analysis for Leishmania (Leishmania) infantum chagasi was performed in opossums by indirect fluorescent antibody test (IFAT), and all animals were serologically negative. Nine positive hemocultures (6.77%) were isolated and cryopreserved and from mammals of the Didelphimorphia and Chiroptera orders and positioned in phylogenies on the basis of sequences from different genes with reference strains of Trypanosoma cruzi marinkellei and T. cruzi. From primary samples (blood and tissues) only one bat, Pteronotus parnellii, was positive to SSU rDNA and gGAPDH genes and grouped with the L. infantum chagasi branch. The studies conducted in Maranhão State provide knowledge of parasite diversity. It is important to determine the presence of trypanosomatids in wild mammals with synanthropic habits.

With the aim of studying some tick-borne diseases, a total of 327 dogs (114 from Tapirapé and 213 from Karajá indigenous ethnicity, Mato Grosso, MT, Brazil) were sampled. Serum samples were submitted to the indirect fluorescence antibody test (IFAT) to detect antibodies against Babesia vogeli, Ehrlichia canis, and Rickettsia spp. Possible associations of risk factors and the occurrence of seroreativity to tick-borne agents and tick infestations were analyzed through chi-squared tests. Among 327 dogs, 46 (13.15%) were seropositive for B. vogeli and 47 (14.37%) for E. canis. The B. vogeli seroprevalence was higher for Karajá and for adult dogs (p>0.05). No association was found for E. canis seroprevalence. From 103 serum samples tested with rickettsial antigens, 90 (87%) dogs were seropositive to Rickettsia spp., with highest reactivity to Rickettsia amblyommii. Canine seropositivity to Rickettsia spp. was associated (p<0.05) with ethnicity (higher seroprevalence in Tapirapé dogs), age (higher in adults), and hunting (higher among hunting dogs). From the 327 dogs, 39 (11.9%) were infested by ticks (Amblyomma cajennense sensu stricto, Amblyomma ovale, Amblyomma oblongoguttatum, Amblyomma tigrinum, and Rhipicephalus sanguineus). Infestations by Amblyomma spp. ticks were higher in dogs from Tapirapé community and in hunting dogs (p<0.05). Regarding R. sanguineus, infestations were higher (p<0.05) among young dogs.

Trypanosomes of anurans and fish are grouped into the Aquatic Clade which includes species isolated from fish, amphibians, turtles and platypus, usually transmitted by leeches and phlebotomine sand flies. Trypanosomes from Brazilian frogs are grouped within the Aquatic Clade with other anuran trypanosome species, where there seems to be coevolutionary patterns with vertebrate hosts and association to Brazilian biomes (Atlantic Forest, Pantanal and Amazonia Rainforest). We characterised the anuran trypanosomes from two different areas of the Cerrado biome and examined their phylogenetic relationships based on the SSU rRNA gene. A total of 112 anurans of six species was analysed and trypanosome prevalence evaluated through haemoculture was found to be 7% (8 positive frogs). However, only three isolates (2.7%) from two anuran species were recovered and cryopreserved. Analysis including SSU rDNA sequences from previous studies segregated the anuran trypanosomes into six groups, the previously reported An01 to An04, and An05 and An06 reported herein. Clade An05 comprises the isolates from Leptodactylus latrans (Steffen) and Pristimantis sp. captured in the Cerrado biome and Trypanosoma chattoni Mathis & Leger, 1911. The inclusion of new isolates in the phylogenetic analyses provided evidence for a new group (An06) of parasites from phlebotomine hosts. Our results indicate that the diversity of trypanosome species is underestimated since studies conducted in Brazil and other regions of the world are still few.

During 2009-2012, wild animals and their ticks were sampled in two areas within the Amazon biome of Brazil, in the states of Mato Grosso and Pará. Animal tissues, blood, and ticks were molecularly tested for Rickettsia and Coxiella DNA. A total of 182 wild animals were sampled, comprising 28 mammalian, five avian, and three reptilian species. Animal tissues or blood were all negative for Rickettsia or Coxiella DNA. A total of 454 ticks (22 larvae, 226 nymphs, 127 males, 79 females) were collected from 52 (28.6%) animals, and identified into 15 species: Amblyomma cajennense, A. naponense, A. humerale, A. nodosum, A. goeldii, A. oblongoguttatum, A. longirostre, A. calcaratum, A. coelebs, A. pacae, A. geayii, A. rotundatum, A. auricularium, A. ovale, and Haemaphysalis juxtakochi. While no Coxiella DNA was identified in ticks, six Rickettsia species were detected in the ticks. "Candidatus Rickettsia amblyommii" was the most common agent, detected in four tick species, A. cajennense, A. auricularium, A. longirostre, and A. humerale. The second most common agent, R. bellii, was detected in A. humerale and A. naponense. Rickettsia rhipicephali was detected in H. juxtakochi, and R. felis in A. humerale. Two possible new Rickettsia species were detected in A. naponense ticks, namely, a novel spotted fever group agent close-related to R. africae in Pará, and a novel Canadensis group agent in Mato Grosso. Results of the present study expand our knowledge on the tick fauna, and on the yet infantile knowledge of tick-borne rickettsiae in the Amazon biome.

In this study a novel Rickettsia from the spotted fever group, isolated from Ornithodoros erraticus soft ticks collected from pigpens in the south of Portugal, is described. After initial screening revealed Rickettsia-positive ticks, isolation attempts were then performed. Successful isolates were achieved by shell-vial technique using Vero E6 cells at 28°C. Molecular characterization of the isolate was performed based on analysis of five rickettsial genes gltA, ompA, ompB, sca1 and htr with their subsequent concatenation along with other rickettsial species resulting in a clustering of the new isolate with Rickettsia felis and Rickettsia hoogstraalii. The degree of nucleotide sequence similarity with other rickettsiae fulfills the criteria for classification of our isolate as a novel species. The name Rickettsia lusitaniae sp. nov. (=CEVDI PoTiRo) is proposed for this new species found in O. erraticus., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

From 2005 to 2012, ticks were collected from different hosts at different localities of the state of Rondônia. The following 16 ixodid tick species were identified: Ixodes fuscipes, Amblyomma auricularium, Amblyomma coelebs, Amblyomma dubitatum, Amblyomma geayi, Amblyomma humerale, Amblyomma latepunctatum, Amblyomma longirostre, Amblyomma naponense, Amblyomma nodosum, Amblyomma oblongoguttatum, Amblyomma ovale, Amblyomma romitii, Amblyomma rotundatum, Amblyomma scalpturatum, and Amblyomma varium. From these, A. auricularium, A. dubitatum, and A. geayi are reported for the first time in the state of Rondônia. We provide the following tick-host associations that have not been reported anywhere: A. longirostre on Pteroglossus bitorquatus, A. rotundatum on Hydrodynastes gigas, and A. latepunctatum and A. scalpturatum on Hydrochoerus hydrochaeris. An adult male specimen of A. rotundatum is reported on Boa constrictor, comprising only the fourth male specimen to be recorded for this obligate parthenogenetic tick species. We also report the presence of the argasid species Ornithodoros kohlsi for the first time in Brazil, based on larval specimens collected on bats Molossops (Neoplatymops) mattogrossensis in Monte Negro, Rondônia. The present study increases the Brazilian tick fauna to 65 species, from which 34 species (52 %) are now registered to Rondônia. Such high diversity of ticks in a relatively small state, associated with increasing environmental alteration due to deforestation and human occupation, makes Rondônia a potential source of tick-borne diseases.

In the order Chiroptera, more than 30 trypanosome species belonging to the subgenera Herpetosoma, Schizotrypanum, Megatrypanum, and Trypanozoon have been described. The species Trypanosoma cruzi , Trypanosoma cruzi marinkellei, and Trypanosoma dionisii are the most common in bats and belong to the Schizotrypanum subgenus. Bats from 2 different biomes, Pantanal and Amazonia/Cerrado in the state of Mato Grosso, Brazil, were evaluated according to the presence of trypanosome parasites by means of hemoculture and PCR in primary samples (blood samples). A total of 211 bats from 20 different species were caught and the trypanosome prevalence, evaluated through hemoculture, was 9.0% (19), 15.5% (13), and 4.8% (6) in the municipalities of Confresa (Amazonia/Cerrado biome) and Poconé (Pantanal biome). Among the 123 primary samples obtained from the bats, only 3 (2.4%) were positive. Phylogenetic analysis using trypanosomatid barcoding (V7V8 region of SSU rDNA) identified all the isolates and primary samples as T. c. marinkellei. The sequences of the isolates were segregated according to the bat host genus or species and suggest that co-evolutionary patterns exist between hosts and parasites. Further studies in different Brazilian regions and biomes need to be conducted in order to gain real understanding of the diversity of trypanosomes in bats.

The Borrelia burgdorferi sensu lato (s.l.) group comprises genetically related spirochetes, mostly associated with tick species belonging to the Ixodes ricinus complex in the Northern Hemisphere. The present study evaluated borrelial infection in the tick Ixodes pararicinus, which is the only representative species of the I. ricinus complex in Uruguay. A total of 137 I. pararicinus ticks were collected from deer, cattle, or vegetation in 2 Uruguayan Departments. A part of these ticks was tested directly by PCR targeting the borrelial gene flagellin (fla), whereas another part of the ticks was inoculated into Barbour-Stoenner-Kelly (BSK)-H medium in an attempt to isolate Borrelia. Overall, Borrelia infection was detected in 9 males and 1 nymphal tick pool. These ticks were found to be infected by unique fla haplotypes, which were shown through phylogenetic analysis to represent possibly 2 new B. burgdorferi s.l. genospecies, 1 associated with B. bissettii, the other phylogenetically closest to B. americana. These results were reinforced by PCR and DNA sequencing analyses of portions of 2 additional borrelial genes, rrfA-rrlB intergenic spacer region (IGS) and 16S rDNA (rrs). Weekly examinations of BSK cultures by dark-field microscopy failed to demonstrate live Borrelia through a 100-day incubation period. However, Borrelia DNA was detected by fla-PCR in culture media from 2 vials up to 90 days after inoculation. To the best of our knowledge, this is the first report of B. burgdorferi s.l. infecting ticks in South America.

Prevalence of Toxoplasma gondii and Neospora caninum antibodies in sera of 325 dogs in 11 villages inhabited by the Tapirapé and Karajá ethnic groups in the south of the Brazilian Amazon was determined by the use of an indirect fluorescence antibody test. Antibodies (cutoff 1:16) to T. gondii were found in 169 (52%) and to N. caninum (cut-off 1:50) in 32 (9.8%) of 325 dogs. Seropositivity for both parasitic infections was widely prevalent in dogs from these villages and was higher in older dogs, indicating post-natal transmission.

In the present study, the presence of tick-associated bacteria and protozoa in Ornithodoros rostratus ticks (adults, nymphs, and eggs) from the Pantanal region of Brazil were determined by molecular detection. In these ticks, DNA from protozoa in the genera Babesia and Hepatozoon, and bacteria from the genera Rickettsia, Borrelia, Anaplasma, and Ehrlichia were not detected. Conversely, all tested ticks (100%) yielded PCR products for 3 Coxiella genes (16S rRNA, pyrG, cap). PCR and phylogenetic analysis of 3 amplified genes (16S rRNA, pyrG, cap) demonstrated that the agent infecting O. rostratus ticks was a member of the genus Coxiella. This organism grouped with Coxiella symbionts of other soft tick species (Argasidae), having different isolates of C. burnetii as a sister group, and these 2 groups formed a clade that grouped with another clade containing Coxiella symbionts of hard tick species (Ixodidae). Analysis of tick mitochondrial 16S rRNA gene database composed mostly of tick species previously shown to harbor Coxiella symbionts suggests a phylogenetic congruence of ticks and their Coxiella symbionts. Furthermore, these results suggest a very long period of coevolution between ticks and Coxiella symbionts and indicates that the original infection may have occurred in an ancestor common to the 2 main tick families, Argasidae (soft ticks) and Ixodidae (hard ticks). However, this evolutionary relationship must be confirmed by more extensive testing of additional tick species and expanded populations., (Copyright © 2012 Elsevier GmbH. All rights reserved.)

A new species of argasid tick (Acari: Argasidae) is described from immature and adult specimens collected from several localities in Brazil. A complete morphological account is provided for all postembryonic life stages, i.e., larva, nymph, female, and male. Ornithodoros cavernicolous n. sp. is the 113(th) in the genus. Morphologically, the new species shares common features, e.g., presence of well-developed cheeks and legs with micromammillate cuticle, with other bat-associated argasid ticks included in the subgenus Alectorobius. In particular, the new species is morphologically related to Ornithodoros azteci Matheson, with which it forms a species group. Phylogenetic analysis based on the 16S rRNA gene sequences supports the placement of the new species within a large clade that includes other New World bat-associated argasids. However, the new species seems to represent an independent lineage within the genus Ornithodoros.

Based on chaetotaxy of the dorsal shield, the taxonomic status of many species of Ornithonyssus has been considered invalid, resulting in the synonymy of all Brazilian Ornithonyssus from small terrestrial wild mammals into one of the following four species: Ornithonyssus bacoti (Hirst, 1913), Ornithonyssus matogrosso (Fonseca, 1954), Ornithonyssus pereirai (Fonseca, 1935) or Ornithonyssus wernecki (Fonseca, 1935). Despite the revision of this genus in 1980, including all known species worldwide, the knowledge of Ornithonyssus in Brazil has not progressed for more than 40 years. Considering the potential importance of these haematophagous mites in transmitting rickettsial disease agents to animals and humans, we have revised Ornithonyssus species collected from small mammals in Brazil by means of morphological and molecular studies. Types and other material deposited in the Acari Collection of the Instituto Butantan (IBSP) were examined in addition to recently collected specimens. Morphological and genetic analysis of the 16S rDNA mitochondrial gene revealed that small terrestrial mammals in Brazil are parasitized by six species of Ornithonyssus mites: Ornithonyssus brasiliensis (Fonseca, 1939), O. matogrosso, O. monteiroi (Fonseca, 1941), O. pereirai, O. vitzthumi (Fonseca, 1941), and O. wernecki. An illustrated key to females of the valid Brazilian species of Ornithonyssus is included, based on optical and scanning electron microscopy.

Carios mimon is an argasid tick common on Chiroptera, originally described from larvae collected on bats Mimon crenulatum from Bolivia and Eptesicus brasiliensis from Uruguay. Later it was also registered from Argentina and recently included among the Brazilian tick fauna. In Brazil, this species is very aggressive to man, resulting in intense inflammatory response and pain. It is known only by the larval description and its morphology resembles that from other species currently included into the genus Carios, formerly classified into the subgenus Alectorobius, genus Ornithodoros. Here we describe adults and redescribe the larva of C. mimon, based on light and scanning electron microscopy. Remarks about its morphological similarity with other species of this genus are also discussed. Molecular analysis inferred from a portion of the 16S rRNA mitochondrial gene placed C. mimon in a cluster supported by maximal bootstrap value (100%) with other argasid species (mostly bat parasites in the New World), which have been classified into either the genus Ornithodoros or Carios, depending on the Argasidae classification adopted by different authors.

The genetic diversity and phylogeographical patterns of Trypanosoma species that infect Brazilian bats were evaluated by examining 1043 bats from 63 species of seven families captured in Amazonia, the Pantanal, Cerrado and the Atlantic Forest biomes of Brazil. The prevalence of trypanosome-infected bats, as estimated by haemoculture, was 12.9%, resulting in 77 cultures of isolates, most morphologically identified as Trypanosoma cf. cruzi, classified by barcoding using partial sequences from ssrRNA gene into the subgenus Schizotrypanum and identified as T. cruzi (15), T. cruzi marinkellei (37) or T. cf. dionisii (25). Phylogenetic analyses using nuclear ssrRNA, glycosomal glyceraldehyde 3-phosphate dehydrogenase (gGAPDH) and mitochondrial cytochrome b (Cyt b) gene sequences generated three clades, which clustered together forming the subgenus Schizotrypanum. In addition to vector association, bat trypanosomes were related by the evolutionary history, ecology and phylogeography of the bats. Trypanosoma cf. dionisii trypanosomes (32.4%) infected 12 species from four bat families captured in all biomes, from North to South Brazil, and clustered with T. dionisii from Europe despite being separated by some genetic distance. Trypanosoma cruzi marinkellei (49.3%) was restricted to phyllostomid bats from Amazonia to the Pantanal (North to Central). Trypanosoma cruzi (18.2%) was found mainly in vespertilionid and phyllostomid bats from the Pantanal/Cerrado and the Atlantic Forest (Central to Southeast), with a few isolates from Amazonia., (2009 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.)

In this study, we provide phylogenetic and biogeographic evidence that the Trypanosoma cruzi lineages T. cruzi I (TCI) and T. cruzi IIa (TCIIa) circulate amongst non-human primates in Brazilian Amazonia, and are transmitted by Rhodnius species in overlapping arboreal transmission cycles, sporadically infecting humans. TCI presented higher prevalence rates, and no lineages other than TCI and TCIIa were found in this study in wild monkeys and Rhodnius from the Amazonian region. We characterised TCI and TCIIa from wild primates (16 TCI and five TCIIa), Rhodnius spp. (13 TCI and nine TCIIa), and humans with Chagas disease associated with oral transmission (14 TCI and five TCIIa) in Brazilian Amazonia. To our knowledge, TCIIa had not been associated with wild monkeys until now. Polymorphisms of ssrDNA, cytochrome b gene sequences and randomly amplified polymorphic DNA (RAPD) patterns clearly separated TCIIa from TCIIb-e and TCI lineages, and disclosed small intra-lineage polymorphisms amongst isolates from Amazonia. These data are important in understanding the complexity of the transmission cycles, genetic structure, and evolutionary history of T. cruzi populations circulating in Amazonia, and they contribute to both the unravelling of human infection routes and the pathological peculiarities of Chagas disease in this region.