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BACKGROUND: Wild boars (Sus scrofa) may cause substantial damage to crops and can spread zoonotic parasites to domestic animals, posing a risk to health and animal production. Metastrongylus spp. can negatively affect the wild boar population, increasing piglet mortality. In addition to that, studies with Metastrongylus genetic characterization are still scarce in Brazil. The present study aims to characterize Metastrongylus spp. from wild boars hunted in the states of São Paulo, Paraná, and Rio Grande do Sul, Brazil, using traditional morphological description and DNA sequences in an integrative taxonomic approach. METHODS: After nematode collection from 58 wild boars, the parasites were morphologically identified and genetically characterized by the amplification of 18S ribosomal DNA (rDNA), 28S rDNA, internal transcribed spacer (ITS) region, and cox-1 mitochondrial DNA (mtDNA). Descriptors of infection were determined and Pearson's Chi-square test was applied to compare the prevalence of infections among the identified parasite species, host age group (juveniles and adults), and sex. The Mann-Whitney U test was performed to compare the mean intensity between the age groups and sex. RESULTS: Metastrongylus salmi, Metastrongylus apri, and Metastrongylus pudendotectus were identified in 77.6% (45/58) of the necropsied wild boars. Metastrongylus salmi was the most prevalent and abundant species (70.7%, 11.1), followed by M. pudendotectus (18.9%, 4.3) and M. apri (17.2%, 2.2). Metastrongylus pudendotectus showed the highest mean intensity and range (25.2, 1-93), followed by M. salmi (15.7, 1-58) and M. apri (12.6, 3-27). We found a significantly higher prevalence of Metastrongylus spp. and M. salmi in adult wild boars, probably associated with a more prolonged time of exposure to intermediate host species. The phylogenetic analysis revealed that ITS2 region and cox-1 mtDNA are the most suitable genetic markers for Metastrongylus species characterization. Genetic variability between M. apri and M. salmi isolates was verified. CONCLUSIONS: We expand the knowledge about the Metastrongylus community in the non-captive wild boar population from Brazil as well as the importance of this exotic species in the maintenance of Metastrongylus spp. in its areas of occurrence. The novel genetic sequences obtained may help further studies to understand the genetic diversity in other nematode populations from Brazil and other countries.
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Metastrongyloidea , Parasitos , Doenças dos Suínos , Suínos , Animais , Brasil/epidemiologia , Filogenia , DNA Ribossômico/genética , DNA Mitocondrial/genética , Sus scrofa , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/parasitologiaRESUMO
Introduction: Bats, along with their ectoparasites, harbor a wide diversity of symbiotic and potential pathogenic bacteria. Despite the enormous diversity of bats (181 species), few studies aimed to investigate the bacterial microbiome of Brazilian chiropterans and associated ectoparasites. This study aimed to characterize the bacterial microbiome of non-hematophagous bats and associated Streblidae flies and Macronyssidae and Spinturnicidae mites in the state of Mato Grosso do Sul, midwestern Brazil. Methods: Oral and rectal swabs were collected from 30 bats (Artibeus lituratus [n = 13], Artibeus planirostris [n = 9], Eptesicus furinalis [n = 5], Carollia perspicillata [n = 2], and Platyrrhinus lineatus [n = 1]). In addition, a total of 58 mites (15 Macronyssidae and 43 Spinturnicidae) and 48 Streblidae bat flies were collected from the captured bats. After DNA extraction and purification, each sample's bacterial composition was analyzed with metagenomic sequencing. Results: The microbiome composition of both oral and rectal bat swab samples showed that Gammaproteobacteria was the most abundant bacterial class. Spiroplasma, Wolbachia and Bartonella represented the most abundant genera in Streblidae flies. While Wolbachia (Alphaproteobacteria) was the most abundant genus found in Spinturnicidae, Arsenophonus (Gammaproteobacteria) was found in high abundance in Macronyssidae mites. In addition to characterizing the microbiome of each sample at the class and genus taxonomic levels, we identified medically significant bacteria able to infect both animals and humans in oral (Streptococcus and Anaplasma) and rectal swabs (Enterobacter, Klebsiella, Escherichia, Enterococcus, Streptococcus), Macronyssidae (Anaplasma, Bartonella, Ehrlichia) and Spinturnicidae (Anaplasma, Bartonella) mites as well as Streblidae flies (Spiroplasma, Bartonella). Discussion and conclusion: Besides expanding the knowledge on the bacterial microbiome of non-hematophagous bats and Streblidae flies from Brazil, the present work showed, for the first time, the bacterial community of bat-associated Macronyssidae and Spinturnicidae mites.
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Bartonella henselae is a zoonotic pathogen responsible for causing Cat Scratch Disease (CSD) and other clinical manifestations in humans. Domestic cats are the main reservoirs of this Bartonella species. Previous studies have suggested that certain genotypes of B. henselae seem to be more associated with human infections. The present study aimed to genotype B. henselae isolates from domestic cats' blood samples in the state of Goiás, midwestern Brazil. The association of quantitative real-time PCR (qPCR) based on the nuoG gene from Bartonella spp. of blood samples, before and after incubation in pre-enrichment liquid medium (BAPGM) and isolation on chocolate agar, showed a positivity frequency of 42% (42/100) for Bartonella spp. Twelve B. henselae isolates obtained on agar chocolate from six cats' blood samples (two isolates from each animal) were characterized by Multi-locus Sequencing Typing (MLST) and revealed to belong to Sequence Types ST1 and ST5. One of the cats (1/6) presented both STs, demonstrating that domestic cats can be coinfected with different variants of B. henselae. The STs detected in this study are distributed worldwide and have already been detected in humans with clinical manifestations of bartonellosis. This is the first report of the zoonotic variants ST1 and ST5 of B. henselae in domestic cats from Brazil.
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Infecções por Bartonella , Bartonella henselae , Bartonella , Doenças do Gato , Coinfecção , Gatos , Animais , Humanos , Bartonella henselae/genética , Tipagem de Sequências Multilocus , Ágar , Brasil/epidemiologia , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/veterinária , Bartonella/genética , Doenças do Gato/epidemiologia , DNA Bacteriano/genéticaRESUMO
The genus Bartonella (Rhizobiales: Bartonellaceae) encompasses facultative intracellular Gram-negative alphaproteobacteria that parasitize mainly erythrocytes and endothelial cells, as well as macrophages, monocytes and dendritic cells. Although they can infect numerous mammal species and arthropod vectors worldwide, reports of Bartonella infections in marsupials are scarce. In fact, such agents have only been detected in marsupials and/or associated ectoparasites in Australia and the United States of America until the present moment. The present study aimed to isolate and characterize molecularly, morphologically and phenotypically Bartonella infecting free-living marsupials sampled in the Brazilian Pantanal, the largest wetland in South America. Two marsupials were captured in December 2018 and six marsupials in February 2019, totaling eight small mammals sampled: five (62.5%) Thylamys macrurus and three (37.5%) Monodelphis domestica. All blood samples were submitted to qPCR for Bartonella spp. based on the nuoG gene, a pre-enrichment liquid culture and a chocolate agar solid culture. Bartonella sp. was isolated from 3 T. macrurus and one M. domestica. One Bartonella isolate obtained from a T. macrurus blood sample (strain 117A) that showed to be closely related to the Bartonella vinsonii complex and Bartonella machadoae was selected for whole genome sequencing using a hybrid approach based on Illumina NovaSeq and Nanopore sequencing platforms. This strain showed a genome of 2.35 Mbp, with an average C + G content of 38.8%, coding for 2013 genes, and a 29 kb plasmid with an average C + G content of 34.5%. In addition, this strain exhibited an average nucleotide identity (ANI) of 85% with Bartonella species belonging to the B. vinsonii group and 91% with B. machadoae. Phylogenomic analysis based on 291 protein coding genes shared by the genomes of 53 Bartonella species positioned this strain closely to B. machadoae. This new isolated species was named Bartonella harrusi sp. nov., which was characterized as having small capnophilic, microaerophilic and aerobic rods with an absence of pili and flagella. In conclusion, the present work describes the biochemical, phenotypic and genomic characteristics of Bartonella harrusi, a new species isolated from the T. macrurus blood samples of the Brazilian Pantanal. Finally, a review of the taxonomic classification of members of the genus Bartonella is proposed, based on the ANI values accessed by whole genome sequencing analyses.
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The present study aimed to investigate, by molecular techniques, the occurrence of Anaplasmataceae, Bartonellaceae, Rickettsiaceae, Mycoplasmataceae, Coxiellaceae, and Babesiidae/Theileriidae agents in blood samples of free-living wild boars (Sus scrofa) and associated ticks in south-eastern Brazil. For this purpose, 67 blood samples and 265 ticks (264 Amblyomma sculptum and one Amblyomma ovale) were analysed. In the screening for Anaplasmataceae agents by a PCR assay based on the 16S rRNA gene, 5.97% blood samples and 50.54% ticks were positive. In the PCR assay for Ehrlichia spp. based on the dsb gene, 9.24% of ticks were positive. Despite the low occurrence, a possible new 16S rRNA genotype of Anaplasma sp. was detected in a wild boar's blood sample. According to phylogenetic analyses based on the 16S rRNA, gltA, and sodB genes and ITS (23S-5S rRNA) intergenic region, it was found that A. sculptum and A. ovale ticks collected from wild boars carry Ehrlichia genotypes phylogenetically associated with Ehrlichia ewingii, Ehrlichia ruminantium, and new Ehrlichia genotypes previously detected in horses, peccaries, and ticks collected from jaguars. In the screening for haemoplasmas by a qPCR based on the 16S rRNA gene, 88.06% of blood samples and 8.69% of ticks were positive. Mycoplasma suis, Mycoplasma parvum, and a possible new haemoplasma genotype were detected in wild boars in south-eastern Brazil. In the screening for Bartonella spp. using a nuoG-based qPCR assay, 3.8% of tick samples were positive. Phylogenetic inferences positioned four nuoG and one r gltA Bartonella sequences into the same clade as Bartonella machadoae. No blood or tick samples from wild boars showed to be positive in the qPCR for Coxiella burnetii based on the IS1111 gene. On the other hand, only 1.6% of ticks were positive in the nested PCR assay for piroplasmids based on the 18S rRNA gene. A 18S rRNA sequence detected in a pool of A. sculptum nymphs was phylogenetically close to Cytauxzoon felis sequences previously detected in cats from the United States. Rickettsia sp. closely related to Rickettsia bellii was detected in a pool of A. sculptum nymphs. This is the first report of haemoplasmas, B. machadoae, and Cytauxzoon spp. in A. sculptum. Wild boars and associated ticks do not seem to participate in the epidemiological cycle of C. burnetii in the region studied. This invasive mammal species may act as a potential disperser of ticks infected with Ehrlichia spp., Bartonella spp., haemotropic mycoplasmas, and Cytauxzoon, and may bring important epidemiological implications in the transmission of bartonelosis, ehrlichiosis, haemoplasmosis, and cytauxzoonosis to humans and animals, more specifically to horses, rodents, pigs, and cats.
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Bartonella , Rickettsia , Carrapatos , Anaplasma/genética , Animais , Bartonella/genética , Brasil/epidemiologia , Gatos , DNA Intergênico , Ehrlichia/genética , Genótipo , Humanos , Filogenia , RNA Ribossômico 16S/genética , RNA Ribossômico 18S , RNA Ribossômico 5S , Rickettsia/genética , Sus scrofa , Suínos , Carrapatos/microbiologiaRESUMO
It has been estimated that 75% of emerging infectious diseases comprise zoonoses, whose majority have free-living animals as reservoirs and are mainly transmitted by arthropod vectors. Although rodents represent important Bartonella reservoirs, there are few studies on the genotypic characterization of Bartonella species commonly found in this taxon and from different Brazilian biomes. Therefore, the present study aimed to investigate the occurrence, isolate and molecularly, morphologically and phenotypically characterize a new Bartonella species infecting free-living rodents sampled in the Brazilian Pantanal, the largest wetland in South America. For this purpose, 129 free-living rodents (79 Thrichomys fosteri, 4 Clyomys laticeps, and Oecomys mamorae) were captured. While blood samples were collected from 57 T. fosteri, 4 C. laticeps and 32 O. mamorae; spleen samples were collected from 22 T. fosteri and 14 O. mamorae. Blood and spleen samples were submitted to a qPCR for Bartonella spp. targeting the nuoG gene, using DNA samples extracted directly from blood/spleen, after passage in pre-enrichment liquid culture, and from colonies obtained from solid culture on chocolate agar. Combining all techniques, occurrence of 24.8% for Bartonella sp. was found among the sampled rodents. One Bartonella isolate (strain 56A) obtained from a T. fosteri's blood sample was closely related to the Bartonella vinsonii complex and selected for Whole Genome Sequencing (WGS) hybrid approach using Illumina NovaSeq and Nanopore sequencing platforms. This strain exhibits a circular 2.7 Mbp genome with an average C+G content of 39% and encoding to 2239 genes. In the phylogenomics based on 291 shared protein-coding genes, this strain was positioned in a unique clade, closely related to Bartonella vinsonii subsp. vinsonii, B. vinsonii subsp. berkhoffii and B. visonii subsp. arupensis. An Average Nucleotide Identity of 85% was found between the obtained isolate and Bartonella species belonging to B. vinsonii complex. These findings supported the separation of this strain, now formally named as Bartonella machadoae sp. nov., from the Bartonella vinsonii complex. In addition, Bartonella machadoae sp. nov. was characterized by capnophilic, microaerophilic and aerobic small rods with absence of pili and flagella. Phylogenetic and distance analyses based on five concatenated molecular markers suggest that Bartonella machadoae may parasite rodents from different Brazilian biomes. In conclusion, we described biochemical, phenotypic and genomic characteristics of Bartonella machadoae nov. sp. isolated from blood samples of T. fosteri rodents from the Brazilian Pantanal.
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Infecções por Bartonella , Bartonella , Animais , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/veterinária , Brasil/epidemiologia , DNA Bacteriano/análise , DNA Bacteriano/genética , Filogenia , Roedores , Áreas AlagadasRESUMO
Although Bartonella spp. is described in cats worldwide, little is known about the occurrence and genetic diversity of Bartonella spp. in cats from South America. To date, it has only been detected in cats from Brazil, Chile and Argentina. This study aimed to undertake a molecular survey and explore the genetic diversity of Bartonella spp. in domestic cats from Paraguay. A TaqMan real-time quantitative PCR (qPCR) targeting the nuoG gene (83â¯bp) for Bartonella spp. was used to screen 125 blood samples from cats in Asuncion, Paraguay. nuoG qPCR-positive samples were further submitted to conventional PCR assays based on the ITS (453- 717â¯bp), gltA (767â¯bp), ftsZ (515â¯bp), rpoB (333â¯bp), ribC (585-588â¯bp), and pap-31 (564â¯bp) loci. Positive samples were sequenced for species identification, phylogenetic, and haplotype analyses. Bartonella D.N.A. was present in 20.8% (26/125) cat blood samples, with low levels of Bartonella nuoG D.N.A. cPCR products targeting gltA, ftsZ, ITS, and rpoB loci from sixteen cats were successfully sequenced. However, all nouG qPCR-positive samples were negative for the ribC and pap-31 genes. Bartonella henselae [62.5% (10/16)] and Bartonella clarridgeiae [37.5% (6/16)] were identified among the sequenced samples. Upon phylogenetic analysis, B. henselae and B. clarridgeiae from Paraguay clustered with sequences detected in domestic and wild cats, dogs, and cat fleas worldwide. Two to four haplotypes of B. henselae and B. clarridgeiae in cats from Paraguay were observed, with some being exclusive and others shared with worldwide distributed haplotypes. Here, we report B. henselae and B. clarridgeiae for the first time in cats from Paraguay. Its circulation in cats suggests the need to consider Bartonellae when testing clinical samples from suspected infectious diseases in humans from Paraguay.
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Infecções por Bartonella/veterinária , Bartonella henselae/genética , Bartonella/genética , Doenças do Gato/epidemiologia , Variação Genética , Animais , Infecções por Bartonella/epidemiologia , Gatos , Paraguai/epidemiologia , FilogeniaRESUMO
Anaplasma marginale is an obligate intracellular Gram-negative bacterium that is parasitic to erythrocytes and is the main agent of bovine anaplasmosis. This disease causes severe anemia and reduces weight gain and milk production, thus giving rise to major economic losses relating to livestock worldwide. The genetic diversity of this bacterium has been characterized based on sequences of major surface proteins (MSPs), especially MSP1α. This has enabled identification of several geographical strains, according to different amino acid sequences. The aim of this study was to investigate the genetic diversity of A. marginale in naturally infected Angus beef cattle during a disease outbreak in southeastern Brazil. Four blood samples were collected over a four-month period from each of 20 animals on a cattle farm in Itú, São Paulo, Brazil. Serum samples were subjected to indirect ELISA to detect anti-A. marginale IgG antibodies. The 80 whole-blood samples obtained were subjected to DNA extraction, quantitative real-time PCR (qPCR) for the msp1ß gene, semi-nested PCR (snPCR) for the msp1α gene, cloning of the target fragment and sequencing using the Sanger method. The sequences obtained were analyzed for genetic diversity using the RepeatAnalyzer software. Both iELISA tests, using recombinant MSP5 and the Anaplasma antibody test kit (VMRD), revealed high seroprevalence: 91.25% and 97.5%, respectively. In qPCR, 100% of the samples were positive, with between 103 and 107 DNA copies/µL. In the snPCR based on the msp1α gene, 57.5% (46/80) of the samples were positive. Microsatellite analysis on the 36 sequences obtained showed the presence of genotypes H (58.3%), F (25%), E (19.4%), C (2.7%) and G (2.7%). The RepeatAnalyzer software identified 36 strains in the study region, among which some had not previously been described in the literature (13 27 13 27 13 F; 16 FF; τ 27; 63 29 104 29; LJ1 13 LJ1 13; 16 F 17; 16 F 91). High genetic diversity of A. marginale bacteria was found on this farm in Itú, São Paulo.
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Anaplasma marginale , Anaplasmose , Doenças dos Bovinos , Superinfecção , Anaplasma marginale/genética , Anaplasmose/microbiologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Brasil/epidemiologia , Bovinos , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/microbiologia , Surtos de Doenças/veterinária , Variação Genética , Filogenia , Superinfecção/epidemiologiaRESUMO
Bartonella henselae is the causative agent for the infectious disease Cat Scratch Disease (CSD), which can be fatal. Domestic and wild felines are known to be its main mammal reservoirs. The present study aimed to investigate the occurrence and genetic diversity of Bartonella spp. in cats sampled in São Paulo (SP) and Minas Gerais (MG) States, Southeastern Brazil. Based on a quantitative real-time PCR (qPCR) assay, a Bartonella sp. nuoG gene fragment was detected in 39.9% (122/306) of the blood samples (46/151 cats of SP; 76/155 cats of MG). The blood samples were submitted to a pre-enrichment culture technique that allowed the detection of 12 additional positive samples, which showed to be negative in the qPCR using DNA blood samples as templates. Furthermore, five B. henselae isolates were obtained from qPCR-negative samples for both blood and pre-enrichment culture. Seven out of 24 Ctenocephalides felis fleas were positive for Bartonella spp. in the qPCR assay; 4/7 positive fleas were collected from Bartonella-negative cats. Twenty-three rpoB B. henselae cloned sequences were obtained from nine cats' blood samples, showing the occurrence of 13 different genotypes. Median-joining network and SplitsTree distance analysis showed that the obtained sequences represented distinct B. henselae genotypes when compared to those previously deposited in GenBank. Intra-host diversity was found, since different rpoB genotypes of B. henselae were detected in individual single cats. Bartonella henselae isolates showed two allelic profiles (ST37 in cats from MG state and ST9 in SP state) by MLST (Multilocus Sequence Typing) based on sequencing of eight molecular markers. The present study is the first molecular report of Bartonella sp. in cats from Minas Gerais State. In summary, this body of work showed the occurrence of different B. henselae rpoB genotypes at an intra-reservoir host level. Based on qPCR from blood samples and pre-enrichment liquid culture and isolation, occurrence of 33.1% (50/151) and 56.8% (88/155) for Bartonella sp. was found in cats from SP and MG states, respectively. Two different allelic profiles of B. henselae were found in cats from the states of São Paulo (ST9) and Minas Gerais (ST37), suggesting a clonal evolution of Bartonellae in a certain geographical region.
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Infecções por Bartonella , Bartonella henselae , Doenças do Gato , Animais , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/veterinária , Bartonella henselae/classificação , Bartonella henselae/genética , Brasil/epidemiologia , Doenças do Gato/epidemiologia , Doenças do Gato/microbiologia , Doença da Arranhadura de Gato , Gatos , DNA Bacteriano/genética , Variação Genética , Tipagem de Sequências MultilocusRESUMO
Mycoplasma suis and Mycoplasma parvum bind strongly to erythrocytes and may cause clinical hemoplasmosis in swine, affecting several age groups. Mycoplasma spp. infected animals may be asymptomatic carriers and/or show nonspecific clinical signs. In Brazil, information on genetic diversity associated with porcine hemoplasmas (PH) has not been described yet. Therefore, this study has aimed to detect, quantify and characterize the genetic diversity of PH in finishing pigs from technified farms in the state of Goiás, central-western Brazil. Ethylenediaminetetraacetic acid-blood samples from 450 swine belonging to 30 different farms from Goiás state were collected at the slaughterhouse. Quantitative real-time PCR (qPCR) assays were performed for the molecular detection and quantification of PH 16S rRNA gene fragments. Cloning and sequencing of 16S and 23S rRNA amplicons were performed to evaluate the genetic diversity. Moreover, a questionnaire was applied to each farm manager to obtain epidemiological information about the herd. The results on qPCR showed herd occurrence of 68.89% for PH. Quantification values (starting quantity [SQ]) ranged from 8.43 × 10-1 to 4.69 × 106 copies/µl, and 52.71% of the samples presented SQ values equal or lower than 1 × 103 copies/µl. Risk factors were not evaluated once all farms had at least one positive animal. However, Spearman's coefficient test revealed that the occurrence of PH was inversely associated with the number of farrows per week, weaned piglets per week, and weight at slaughter. Phylogenetic analysis based on maximum likelihood and Bayesian methods showed that the 16S rRNA and 23S rRNA gene sequences obtained from five samples formed a single cluster closely related to M. parvum. Genotype analysis using DNASP software confirmed seven and four different 16S and 23S rRNA genotypes among the cloned amplicons, indicating that there are several genotypes of M. parvum circulating in individual pigs and among pig farms in central-western Brazil.
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Variação Genética , Infecções por Mycoplasma/veterinária , Mycoplasma/genética , Sus scrofa/microbiologia , Doenças dos Suínos/microbiologia , Animais , Teorema de Bayes , Sangue/microbiologia , Brasil/epidemiologia , Coinfecção/microbiologia , Coinfecção/veterinária , Fazendas , Genes Bacterianos , Genes de RNAr , Genótipo , Mycoplasma/classificação , Infecções por Mycoplasma/epidemiologia , Infecções por Mycoplasma/microbiologia , Filogenia , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Reação em Cadeia da Polimerase em Tempo Real , Suínos , Doenças dos Suínos/epidemiologiaRESUMO
Recently, an increasing number of Bartonella species have been emerged to cause human diseases. Among animal reservoirs for Bartonella spp., bats stand out due to their high mobility, wide distribution, social behaviour and long-life span. Although studies on the role of vampire bats in the epidemiology of rabies have been extensively investigated in Latin America, information on the circulation and genetic diversity of Bartonella species in these bat species is scarce. In the present work, 208 vampire bats, namely Desmodus rotundus (the common vampire bat; n = 167), Diphylla ecaudata (the hairy-legged vampire bat; n = 32) and Diaemus youngii (the white-winged vampire bat; n = 9) from 15 different states in Brazil were sampled. DNA was extracted from liver tissue samples and submitted to real-time PCR (qPCR) and conventional PCR (cPCR) assays for Bartonella spp. targeting five genetic loci, followed by phylogenetic and genotype network analyses. Fifty-one out of 208 liver samples (24.51%) were positive for Bartonella DNA in the ITS real-time PCR assay [40 (78.43%) of them were from D. rotundus from 11 states, and 11 (21.57%) samples from D. ecaudata from three states. Eleven genotypes were found for each gltA and rpoB genes. Several ITS sequences detected in the present study clustered within the lineage that includes B. bacilliformis and B. ancachensis. The Bayesian phylogenetic inference based on the gltA gene positioned the obtained sequences in six different clades, closely related to Bartonella genotypes previously detected in D. rotundus and associated ectoparasites sampled in Latin America. On the other hand, the Bartonella rpoB genotypes clustered together with the ruminant species, B. schoenbuchensis and B. chomelii. The present study describes for the first time the molecular detection of Bartonella spp. in D. ecaudata bats. It also indicates that Bartonella spp. of vampire bats are genetically diverse and geographically widespread in Brazil.
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Bartonella/genética , Quirópteros/microbiologia , DNA Bacteriano/genética , Variação Genética , Animais , Brasil , Reservatórios de Doenças/microbiologia , Genótipo , Filogenia , Reação em Cadeia da PolimeraseRESUMO
There are few studies on the genetic diversity of Anaplasma marginale in Brazilian cattle herds, especially about beef cattle. The objective of this study was to evaluate the genetic diversity of A. marginale, based on the msp1α gene in Bos taurus indicus sampled from the Brazilian Pantanal. Aliquots of blood with and without EDTA were taken from 400 cattle (200 cows and 200 calves) across five extensive farms. The samples were submitted to the indirect immunoenzymatic assay (iELISA), quantitative real-time PCR (qPCR) for the msp1ß gene and to the semi-nested (sn) PCR for the msp1α gene. Positive samples were sequenced by the Sanger method and subjected to diversity analysis using the RepeatAnalyser software. The percentage of positive animals by iELISA, qPCR and (sn) PCR was 72.2% (289/400), 56.7% (227/400) and 23% (52/227), respectively. Cows (154/200) showed to be significantly more seropositive than calves (135/200). In qPCR, the number of calves and average quantification value (138/200; 1.3 × 106) A. marginale msp1α copies per µL proved to be higher when compared to that found for the cows (89/200; 3.9 × 104). The microsatellite analysis of the 26 sequences obtained from the msp1α gene revealed the presence of E (77%), C (15.4%) and B (7.7%) genotypes. Fourteen A. marginale strains were identified in the studied region, with eight that have never before been described in the literature (τ-10-13-13-18; τ-27-18; EV8-EV8-17; α-ß-ß-ß-100; EV7-11-10-15; τ-11-11-27-18; τ-11-10-15; τ-27-13-18). Beef cattle are highly exposed to A. marginale in the Brazilian Pantanal. Moreover, a high genetic diversity of A. marginale, with eight new strains, was found in the studied region. While cows may act as chronic carriers, perpetuating the pathogen within the herd, male beef calves sold to other regions may disperse these strains.
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Anaplasma marginale/genética , Variação Genética , Genótipo , Filogenia , Anaplasmose/microbiologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Brasil , Bovinos/microbiologia , DNA Bacteriano/sangue , DNA Bacteriano/genética , Feminino , Masculino , Repetições de Microssatélites , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Mycoplasma suis, the etiological agent of swine hemoplasmosis, is an epicellular bacterium that adheres to the surface of pig erythrocytes leading to deformations of the target cells. Little is known about the occurrence of M. suis in wild swine populations around the world, its economic impact on swine herds, and the risk of human infection. The aim of this study was to investigate, by quantitative real-time PCR (qPCR) based on the 16S rRNA gene, the occurrence of M. suis in a captive population of white-lipped peccaries (100 Tayassu pecari) and in free-living wild boars (14 Sus scrofa) in Brazil. None of the white-lipped peccaries were positive for M. suis, whereas seven (50%) wild boars were positive in qPCR assays. The quantification of M. suis-16S rRNA copies/µL ranged from 1.42 × 10° to 3.906 × 101 in positive animals, indicating a low bacteremia and a chronic carrier status in free-living wild boars. In conclusion, M. suis might be a non-frequent pathogen in wild suids maintained in captivity. Despite the low bacteremia, the prevalence of M. suis in wild boar population in Brazil seems to be high.
Assuntos
Infecções por Mycoplasma/epidemiologia , Infecções por Mycoplasma/veterinária , Mycoplasma/genética , Mycoplasma/isolamento & purificação , Animais , Bacteriemia/epidemiologia , Bacteriemia/microbiologia , Bacteriemia/veterinária , Brasil/epidemiologia , DNA Bacteriano/genética , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Tipagem Molecular , RNA Ribossômico 16S/genética , Reação em Cadeia da Polimerase em Tempo Real , Sus scrofa , SuínosRESUMO
The family Streblidae comprises a monophyletic group of Hippoboscoidea, hematophagous dipterans that parasitize bats. Bartonella spp. and Rickettsia spp. have been reported in bats sampled in Europe, Africa, Asia, North, Central and South America. However, there are few reports on the Bartonella and Rickettsia bacteria infecting Hippoboscoidea flies and mites. While Spinturnicidae mites are ectoparasites found only in bats, those belonging to the family Macronyssidae comprise mites that also parasitize other mammal species. This study investigates the occurrence and assesses the phylogenetic positioning of Bartonella spp. and Rickettsia spp. found in Streblidae flies and Spinturnicidae and Macronyssidae mites collected from bats captured in Brazil. From May 2011 to April 2012 and September 2013 to December 2014, 400 Streblidae flies, 100 Macronyssidaes, and 100 Spinturnicidae mites were collected from bats captured in two sites in northeastern Nova Iguaçu, Rio de Janeiro, southeastern Brazil. Forty (19.8%) out of 202 Streblidae flies were positive for Bartonella spp. in qPCR assays based on the nuoG gene. Among the flies positive for the bacterium, six (18%) were Paratrichobius longicrus, seven (29%) Strebla guajiro, two (40%) Aspidoptera phyllostomatis, five (11%) Aspidoptera falcata, one (10%) Trichobius anducei, one (25%) Megistopoda aranea, and 18 (32%) Trichobius joblingi, and collected from bats of the following species: Artibeus lituratus, Carollia perspicillata, Artibeus planirostris, Sturnira lilium, and Artibeus obscurus. Six sequences were obtained for Bartonella (nuoG [n = 2], gltA [n = 2], rpoB [n = 1], ribC = 1]). The phylogenetic analysis based on gltA (750pb) gene showed that the Bartonella sequences clustered with Bartonella genotypes detected in bats and ectoparasites previously sampled in Latin America, including Brazil. Only one sample (0.49%) of the species Trichobius joblingi collected from a specimen of Carollia perspicillata was positive for Rickettsia sp. in cPCR based on the gltA gene (401bp). This sequence was clustered with a 'Candidatus Rickettsia andaenae" genotype detected in an Amblyomma parvum tick collected from a rodent in the southern region of Brazilian Pantanal. The sampled Macronyssidae and Spinturnicidae mites were negative for Bartonella spp. and Rickettsia spp. This study demonstrated the first occurrence of Bartonella spp. and Rickettsia spp. DNA in Streblidae flies collected from bats in Brazil.
Assuntos
Bartonella/isolamento & purificação , Quirópteros/parasitologia , Ectoparasitoses/parasitologia , Parasitos/microbiologia , Rickettsia/isolamento & purificação , Animais , Bartonella/genética , Brasil , DNA Bacteriano/isolamento & purificação , Dípteros/microbiologia , Ectoparasitoses/veterinária , Ácaros/microbiologia , Filogenia , Reação em Cadeia da Polimerase em Tempo Real , Rickettsia/genética , Análise de Sequência de DNARESUMO
The present work aimed to investigate the genetic diversity of Bartonella in mammals and ectoparasites in Pantanal wetland, Brazil. For this purpose, 31 Nasua nasua, 78 Cerdocyon thous, 7 Leopardus pardalis, 110 wild rodents, 30 marsupials, and 42 dogs were sampled. DNA samples were submitted to a quantitative real-time PCR assay (qPCR). Positive samples in qPCR were submitted to conventional PCR assays targeting other five protein-coding genes. Thirty-five wild rodents and three Polygenis (P.) bohlsi bohlsi flea pools showed positive results in qPCR for Bartonella spp. Thirty-seven out of 38 positive samples in qPCR were also positive in cPCR assays based on ftsZ gene, nine in nuoG-cPCR, and six in gltA-cPCR. Concatenated phylogenetic analyses showed that two main genotypes circulate in rodents and ectoparasites in the studied region. While one of them was closely related to Bartonella spp. previously detected in Cricetidae rodents from North America and Brazil, the other one was related to Bartonella alsatica, Bartonella pachyuromydis, Bartonella birtlesii, Bartonella acomydis, Bartonella silvatica, and Bartonella callosciuri. These results showed that at least two Bartonella genotypes circulate among wild rodents. Additionally, the present study suggests that Polygenis (P.) bohlsi bohlsi fleas could act as possible Bartonella vectors among rodents in Pantanal wetland, Brazil.
Assuntos
Doenças dos Animais/microbiologia , Infecções por Bartonella/veterinária , Bartonella/classificação , Bartonella/genética , Variação Genética , Mamíferos/microbiologia , Áreas Alagadas , Animais , Animais Selvagens/microbiologia , Proteínas de Bactérias/genética , Bartonella/patogenicidade , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/microbiologia , Brasil/epidemiologia , Proteínas do Citoesqueleto/genética , DNA Bacteriano/genética , Vetores de Doenças , Genes Bacterianos/genética , Genótipo , América do Norte/epidemiologia , Filogenia , Roedores/microbiologia , Sifonápteros/microbiologiaRESUMO
The present study determined the prevalence, hematological findings and genetic diversity of Bartonella spp. in domestic cats from Valdivia, Southern Chile. A complete blood count and nuoG gene real-time quantitative PCR (qPCR) for Bartonella spp. were performed in 370 blood samples from cats in Valdivia, Southern Chile. nuoG qPCR-positive samples were submitted to conventional PCR for the gltA gene and sequencing for species differentiation and phylogenetic analysis. Alignment of gltA gene was used to calculate the nucleotide diversity, polymorphic level, number of variable sites and average number of nucleotide differences. Bartonella DNA prevalence in cats was 18·1% (67/370). Twenty-nine samples were sequenced with 62·0% (18/29) identified as Bartonella henselae, 34·4% (10/29) as Bartonella clarridgeiae, and 3·4% (1/29) as Bartonella koehlerae. Bartonella-positive cats had low DNA bacterial loads and their hematological parameters varied minimally. Each Bartonella species from Chile clustered together and with other Bartonella spp. described in cats worldwide. Bartonella henselae and B. clarridgeiae showed a low number of variable sites, haplotypes and nucleotide diversity. Bartonella clarridgeiae and B. koehlerae are reported for the first time in cats from Chile and South America, respectively.