Genotyping of South American clinical isolates of Pythium insidiosum based on single nucleotide polymorphism-based multiplex PCR

We aimed to genotype the South American clinical isolates of Pythium insidiosum using the single nucleotide polymorphisms (SNP) of the ribosomal DNA sequences (rDNA). Previously, an SNP-based multiplex-PCR was able to distinguish three different clades of P. insidiosum isolates. Thus, we used this assay to evaluate South American clinical isolates of P. insidiosum (n=32), standard strains from Costa Rica (n=4), Thailand (n=3), Japan (n=1), and India (n=1), a standard strain of Pythium aphanidermatum, and Brazilian environmental isolates of Pythium torulosum, Pythium rhizo-oryzae and Pythium pachycaule voucher (n=3). It was possible to allocate each American P. insidiosum isolate to clade I, the isolates of India, Japan, and Thailand to clade II, and the Thai isolate to clade III. P. aphanidermatum, P.torulosum, P.rhizo-oryzae and P.pachycaule voucher isolates were not amplified. For the first time, a P. insidiosum isolate from Uruguay, South America, was included in molecular analyzes. By SNP-based multiplex-PCR, it was possible to perform the identification and genotyping of the South American isolates of P. insidiosum, demonstrating similar genetic characteristics of these isolates. Key words: Pythium insidiosum, Pythiosis, molecular detection, genotype, single nucleotide polimorphisms.O objetivo deste estudo foi genotipar isolados clínicos de Pythium insidiosum da América do Sul utilizando polimorfismos de nucleotídeo único (SNP) de sequências de rDNA. Anteriormente, um multiplex-PCR baseado em SNP foi capaz de distinguir P. insidiosum em três diferentes clados. Dessa forma, utilizamos este método para avaliar isolados clínicos de P. insidiosum da América do Sul (n=32), cepas padrão da Costa Rica (n=4), Tailândia (n=3), Japão (n=1) e Índia (n=1), uma cepa padrão de Pythium aphanidermatum e isolados ambientais brasileiros de Pythium torulosum; Pythium rhizo-oryzae e Pythium pachycaule voucher (n=3). Os isolados analisados foram alocados aos clados: I (americanos), II (isolados da Índia, Japão e Tailândia), e III (um isolado tailandês). P. aphanidermatum, P.torulosum, P.rhizo-oryzae e P.pachycaule voucher não foram amplificados. Pela primeira vez, um isolado de P. insidiosum do Uruguai foi incluído em análises moleculares. Através da multiplex-PCR baseada em SNP, foi possível realizar a identificação e genotipagem dos isolados sulamericanos de P. insidiosum, demonstrando características genéticas semelhantes entre esses isolados

Soroprevalência da infecção por pythium insidiosum em equinos no estado do Rio Grande do Sul

An epidemiological survey was carried out by performing an Enzyme Linked Immuno Sorbent Assay (ELISA) test to determine the seroprevalence of Pythium insidiosum infection in equine in Rio Grande do Sul State (RS), Brazil. The serological study covered seven geographical regions of RS, classified according to the Instituto Brasileiro de Geografia e Estatística (IBGE). The samples were obtained from official veterinary service (Serviço Veterinário Oficial, SVO) linked to the Secretaria da Agricultura, Pecuária e Agronegócio of RS (SEAPA-RS) to proceed the investigation of equine infectious anemia in 2014. Samples were collected during the months of September and October of 2013, covering the seven geographical regions of RS, and totalized 1,002 serum samples. The seroprevalence for P. insidiosum in RS was 11.1% (CI95% 9.23 to 13.22). The relative risk (RR) of the presence of antibodies anti-P. insidiosum was in the regions Southeast 11.17 (CI95%, 4.65 to 26.8), Porto Alegre 4.62 (CI95%, 1.70 to 12.55), Southwest 11.17 (CI95%, 4.65 to 26.8) and Northwestern 3.72 (CI95%, 1.52 to 9.09). The highest prevalence (69.1%) was observed in females with RR of 1.59 (CI95%, 1.11 to 2.27). When the presence of dams was evaluated, the seropositivity was evident in 74.4%, presenting an association of 2.13 (CI95%, 1.16 to 3.91) compared to farms without dams. In properties with veterinary assistance, the frequency of 72.7% and RR of 3.04 (CI95%,, 1,85 to 4,98) of seropositivity were observed. Due to the importance of pythiosis in horse herds, this study highlights the presence of anti-P. insidiosum antibodies in horses in RS, Brazil.Um levantamento soroepidemiológico foi realizado através do teste de ELISA indireto para determinar a soroprevalência da infecção por Pythium insidiosum em equinos no estado do Rio Grande do Sul (RS), classificadas de acordo com o Instituto Brasileiro de Geografia e Estatística (IBGE). As amostras utilizadas eram provenientes do cadastro das propriedades do Serviço Veterinário Oficial (SVO), da Secretaria da Agricultura, Pecuária e Agronegócio do RS (SEAPA-RS), coletadas para o inquérito da anemia infecciosa equina de 2014. As coletas foram realizadas durante os meses de setembro e outubro de 2013, abrangendo as sete mesorregiões geográficas do RS, e totalizaram 1.002 amostras de soro. Do total das amostras testadas, 11.1% (CI95% 9.23 to 13.22) foram soropositivas para P. insidiosum. Constatou-se o risco relativo (RR) da presença de anticorpos anti-P. insidiosum nas regiões Sudeste 11,17(IC95%, 4,65- 26,8), Porto Alegre 4,62 (IC95%, 1,70-12,55), Sudoeste 11,17 (IC95%, 4,65-26,8) e Noroeste 3,72 (IC95%, 1,52-9,09). Observou-se a maior soroprevalência (69,1%) em fêmeas com RR de 1,59 (IC95%, 1,11- 2,27). Quanto à presença de açudes, evidenciou-se soropositividade em 74,4% das propriedades, apresentando associação de 2,13 (IC95%,1,16-3,91) em comparação com propriedades sem açude. Em propriedades com assistência veterinária, foi verificada a frequência de 72,7% e RR de 3,04 (IC95%,1,85-4,98). Diante da relevância da pitiose em rebanhos equinos, destaca-se a presença de anticorpos anti-P. insidiosum em equinos no estado do RS

Perfil epidemiológico e terapêutico de pacientes com criptococose atendidos em hospital escola de Santa Maria/RS

Cryptococcosis is a fungal disease caused by yeasts of the complex Cryptococcus neoformans and Cryptococcus gattii. The infection usually occurs by inhaling the basidiospores of the fungus present in the environment. Severe cases occur due to infection of the central nervous system. The disease has a worldwide distribution, mainly associated to the immunocompromised individuals with human immunodeficiency virus (HIV). This study aimed to verify the epidemiological profile and therapeutic protocols of patients diagnosed with cryptococcosis at the Hospital Universitário de Santa Maria (Santa Maria, RS), from March 2010 to March 2017. A total of 46 clinical records were evaluated, the profile of the patients was predominantly Caucasian, aged between 31 and 50 years, with HIV, presenting a CD4 + cell count <100 cells / μL and with a prevalent clinical form neurocryptococcosis. The combination of amphotericin B and fluconazole was the therapeutic protocol adopted in the majority of patients studied.Criptococose é uma doença fúngica causada por leveduras do complexo Cryptococcus neoformans e Cryptococcus gattii. A infecção geralmente ocorre através da inalação dos basidiósporos do fungo presente no ambiente. Casos graves ocorrem devido à infecção do sistema nervoso central. A enfermidade possui distribuição mundial, associada principalmente aos indivíduos imunocomprometidos, portadores do vírus da imunodeficiência humana (HIV). Este estudo objetivou verificar o perfil epidemiológico e os protocolos terapêuticos dos pacientes diagnosticados com criptococose no Hospital Universitário de Santa Maria (Santa Maria, RS), no período de março de 2010 a março de 2017. Avaliaram-se 46 prontuários clínicos, sendo que o perfil dos pacientes observados foi predominantemente do sexo masculino, etnia branca, faixa etária de 31 a 50 anos, portadores de HIV, apresentando taxa de células CD4+<100 células/µL e com forma clínica prevalente, a neurocriptococose. A associação de anfotericina B e fluconazol foi o protocolo terapêutico adotado na maioria dos pacientes estudados

Efficacy of azithromycin and miltefosine in experimental systemic pythiosis in immunosuppressed mice

We evaluated the efficacy of azithromycin (50 mg/kg, every 12 h [q12h] orally) and miltefosine (25 mg/kg, q24h orally) treatments in an experimental model of vascular/disseminated pythiosis in immunosuppressed mice. Azithromycin was the only treatment able to reduce mortality. The histopathological findings showed acute vascular inflammation, pathogen dissemination, necrotizing myositis, neuritis, and arteritis. The results suggest that azithromycin, but not miltefosine, may have clinical relevance in the treatment of vascular/disseminated pythiosis

Epidemiological, clinical and diagnostic aspects of sheep conidiobolomycosis in Brazil

ABSTRACT: Conidiobolomycosis is an emerging disease caused by fungi of the cosmopolitan genus Conidiobolus . Particular strains of Conidiobolus coronatus, Conidiobolus incongruus and Conidiobolus lamprauges , mainly from tropical or sub-tropical origin, cause the mycosis in humans and animals, domestic or wild. Lesions are usually granulomatous and necrotic in character, presenting two clinical forms: rhinofacial and nasopharyngeal. This review includes the main features of the disease in sheep, with an emphasis on the epidemiology, clinical aspects, and diagnosis of infections caused by Conidiobolus spp. in Brazil. In this country, the disease is endemic in the Northeast and Midwest, affecting predominantly woolless sheep breeds and occasioning death in the majority of the studied cases. The species responsible for infections of sheep are C. coronatus and C. lamprauges and the predominant clinical presentation is nasopharyngeal. These fungal infections are very important, since they compromise the health status of the sheep flock and cause serious economic losses to the sheep industry. Thus, research is needed to investigate faster tools for diagnosis and effective methods for the control and treatment of conidiobolomycosis

Isolation and molecular characterization of symbiotic fungus from Acromyrmex ambiguus and Acromyrmex heyeri ants of Rio Grande do Sul State, Brazil

Leaf-cutting ants of the genera Atta and Acromyrmex determine serious agricultural problems and live on symbiosis with Leucoagaricus gongylophorus. The aim of this study is to identify morphological and molecularly, as well as to verify the genotypic variability of the symbiotic fungus cultivated by A. heyeri and A. ambiguus from three different regions of the state of Rio Grande do Sul, Brazil. Fungus gardens were collected and fragments of mycelia were grown in selective medium. Total DNA was extracted and amplification of the ITS region was performed by PCR using universal primers. After DNA sequencing, the chromatograms were assembled and phylogenetic analyzes were performed by the Neighbor-Joining method. A total of six isolates of L. gongylophorus were obtained and their identities were confirmed by molecular analyses. Phylogenetic analysis of the ITS region showed a tree with two distinct groups regarding the fungus isolates from A. heiyeri and A. ambiguous. In this study, it was verified that A. heyeri and A. ambiguous, cultivate the same fungus. Additionally, the molecular marker used in this study showed variations in L. gongylophorus, evidencing two distinct branches in the phylogenetic tree, according to the ant species that cultivate L. gongylophorus. However, other studies involving the inclusion of a great number of isolates of L. gongylophorus, as well as the use of other molecular markers to validate the possible variations in the phylogenetic relationship of this symbiotic fungus are required

Echinococcus granulosus sensu lato e Taenia hydatigena em suínos no sul do Brasil

This study aimed to identify the parasitical etiologic agents of visceral cysts in pigs from the central/northern region of Rio Grande do Sul State, Brazil. Fifty-eight cysts were found in livers during veterinary inspection of swine slaughtered from January 2008 to 2012. Collected samples were submitted to macroscopic and molecular analyzes. Polymerase chain reaction (PCR), DNA sequencing and BLAST alignment of sequences was used to molecular characterization of the samples. By PCR 10.3% (6/58) of tested samples were positive for Echinococcus granulosus sensu lato and 56.9% (33/58) for Cysticercus tenuicollis. In this study, it was verified the occurrence of larval forms of E. granulosus sensu lato and Taenia hydatigena in pig herds from the central/northern region of Rio Grande do Sul State. The presence of both parasites is relevant due to the economic losses for the meat industry. Additionally, E. granulosus sensu lato has zoonotic importance and may be infecting pig herds in southern Brazil.Neste estudo, buscou-se identificar os agentes etiológicos de origem parasitária em cistos nas vísceras de suínos oriundos sa região centro/norte do sEstado do Rio Grande do Sul, Brasil. Cinquenta e oito cistos foram encontrados durante a inspeção veterinária em fígados de suínos abatidos entre janeiro de 2008 a 2012. As amostras foram submetidas às análises macroscópicas e moleculares. Reação em cadeia da polimerase (PCR), sequenciamento de DNA e alinhamento das sequências no BLAST (bases de dados do GenBank) foram utilizados na caracterização molecular das amostras. Das amostras analisadas, 10,3% (6/58) foram positivas para Echinococcus granulosus sensu lato e 56,9% (33/58) para Cysticercus tenuicollis. Evidenciou-se a ocorrência de larvas de E. granulosus sensu lato e Taenia hydatigena em rebanhos suínos na região centro/norte do RS. A presença de ambos os agentes é relevante devido às perdas econômicas para a indústria de carnes. Além disso, destaca-se que E. granulosus sensu lato possui importância zoonótica e pode contaminar os rebanhos suínos no sul do Brasil

Echinococcus granulosus sensu lato e Taenia hydatigena em suínos no sul do Brasil

This study aimed to identify the parasitical etiologic agents of visceral cysts in pigs from the central/northern region of Rio Grande do Sul State, Brazil. Fifty-eight cysts were found in livers during veterinary inspection of swine slaughtered from January 2008 to 2012. Collected samples were submitted to macroscopic and molecular analyzes. Polymerase chain reaction (PCR), DNA sequencing and BLAST alignment of sequences was used to molecular characterization of the samples. By PCR 10.3% (6/58) of tested samples were positive for Echinococcus granulosus sensu lato and 56.9% (33/58) for Cysticercus tenuicollis. In this study, it was verified the occurrence of larval forms of E. granulosus sensu lato and Taenia hydatigena in pig herds from the central/northern region of Rio Grande do Sul State. The presence of both parasites is relevant due to the economic losses for the meat industry. Additionally, E. granulosus sensu lato has zoonotic importance and may be infecting pig herds in southern Brazil.Neste estudo, buscou-se identificar os agentes etiológicos de origem parasitária em cistos nas vísceras de suínos oriundos sa região centro/norte do sEstado do Rio Grande do Sul, Brasil. Cinquenta e oito cistos foram encontrados durante a inspeção veterinária em fígados de suínos abatidos entre janeiro de 2008 a 2012. As amostras foram submetidas às análises macroscópicas e moleculares. Reação em cadeia da polimerase (PCR), sequenciamento de DNA e alinhamento das sequências no BLAST (bases de dados do GenBank) foram utilizados na caracterização molecular das amostras. Das amostras analisadas, 10,3% (6/58) foram positivas para Echinococcus granulosus sensu lato e 56,9% (33/58) para Cysticercus tenuicollis. Evidenciou-se a ocorrência de larvas de E. granulosus sensu lato e Taenia hydatigena em rebanhos suínos na região centro/norte do RS. A presença de ambos os agentes é relevante devido às perdas econômicas para a indústria de carnes. Além disso, destaca-se que E. granulosus sensu lato possui importância zoonótica e pode contaminar os rebanhos suínos no sul do Brasil