scholarly journals Detección molecular de leptospiras patógenas en roedores sinantrópicos y silvestres capturados en Yucatán, México

Biomédica ◽  
2018 ◽  
Vol 38 ◽  
pp. 51-58 ◽  
Author(s):  
Marco Torres-Castro ◽  
Bayron Cruz-Camargo ◽  
Rodrigo Medina-Pinto ◽  
Bibiana Reyes-Hernández ◽  
Carlos Moguel-Lehmer ◽  
...  
Keyword(s):  
16S Rrna ◽  
Leptospira Interrogans ◽  
Leptospira Spp

Introducción. La leptospirosis es una enfermedad zoonótica endémica en México, ocasionada por la bacteria del género Leptospira, la cual constituye un problema de salud pública y veterinaria. Los roedores son los reservorios más relevantes de Leptospira spp., debido a que la bacteria se establece y se reproduce en su tejido renal y es excretada por la orina.Objetivo. Identificar la presencia de Leptospira spp. en tejido renal de roedores capturados en Yucatán, México.Materiales y métodos. Se capturaron roedores sinantrópicos y silvestres en el municipio rural de Cenotillo, Yucatán, México. Se tomó un riñón de cada roedor y se extrajo el ADN total. La identificación de Leptospira spp. se hizo mediante la detección de dos fragmentos del gen 16S rRNA con una reacción en cadena de la polimerasa (PCR) de punto final. Los productos positivos se secuenciaron y se analizaron con herramientas de alineamiento.Resultados. Se capturaron 92 roedores pertenecientes a siete especies distintas. La PCR arrojó 5,4 % (5/92) de positividad global. El análisis del alineamiento de los aislamientos de los roedores infectados demostró 100 % de cobertura e identidad con la especie Leptospira interrogans. Esta es la primera evidencia molecular de la circulación de Leptospira spp. en Heteromys gaumeri capturados en Yucatán, México.Conclusión. Se evidenció que los roedores de Yucatán, México, son reservorios de Leptospira spp. y participan en el ciclo de infección de la leptospirosis en la región.

scholarly journals Controle da leptospirose em bovinos de leite com vacina autógena em Santo Antônio do Monte, Minas Gerais

2012 ◽  
Vol 32 (7) ◽  
pp. 633-639 ◽  
Author(s):  
Denise Chiareli ◽  
Maria R.V. Cosate ◽  
Elvio C. Moreira ◽  
Rômulo C. Leite ◽  
Francisco C.F. Lobato ◽  
...  
Keyword(s):  
16S Rrna ◽  
Minas Gerais ◽  
Leptospira Interrogans ◽  
Leptospira Spp ◽  
Gene 16S Rrna

Um surto de leptospirose foi observado em bovinos leiteiros em Santo Antônio do Monte, Minas Gerais. O rebanho apresentava reações positivas anti-leptospira sorovar Hardjo no teste de microaglutinação (MAT) e havia sido vacinado anteriormente com vacina experimental contendo a sorovariedade Hardjo. O MAT revelou 48,06% dos bovinos positivos para sorovariedade Hardjo genótipo Hardjobovis, 36,82% para sorovariedade Hardjo genótipo Hardjoprajitno. Os animais apresentavam aborto e mastite com presença de sangue no leite. A presente pesquisa teve como objetivos isolar as sorovariedades existentes a partir da urina de vacas sorologicamente positivas, elaborar uma vacina experimental com as sorovariedades isoladas no rebanho, avaliar a eficiência do programa de vacinação por um período de dois anos por meio da sorologia do rebanho. Foi isolada Leptospira spp. a partir da urina de duas vacas com sinais sugestivos da doença. As amostras isoladas foram identificadas pela sorologia com anticorpos monoclonais e seqüenciamento do gene 16S rRNA como pertencentes à espécie Leptospira interrogans, sorogrupo Sejroe, sorovariedade Hardjo e genótipo Hardjoprajitno. O uso da vacina autógena foi eficaz no controle da leptospirose no rebanho no período de dois anos. Os resultados da sorologia revelaram ausência de animais positivos na última prova realizada no rebanho.

scholarly journals Detection of Leptospira spp. in kidney tissues isolated from rats in the Napu and Bada Highlands of Poso District, Central Sulawesi Province

2020 ◽  
Vol 14 (1) ◽  
pp. 17-26
Author(s):  
Gunawan Gunawan ◽  
Tri Wibawa ◽  
Mahardika Agus Wijayanti ◽  
Hayani Anastasia
Keyword(s):  
16S Rrna ◽  
Urban Areas ◽  
Target Gene ◽  
Dna Analysis ◽  
Pathogenic Leptospira ◽  
Rural And Urban Areas ◽  
Rural And Urban ◽  
Leptospira Spp ◽  
Pcr Method ◽  
Central Sulawesi

Leptospirosis is still a global health problem because it affects human health in rural and urban areas, both in industrialized and developing countries. The aim of the study was to detect Leptospira spp. bacteria in kidney tissues isolated from rats in the Napu and Bada Highlands of Poso District, Central Sulawesi Province. Kidneys sample from 63 rats were collected from Napu and Bada Highlands of Poso District, Central Sulawesi Province in MayJune 2018. Polymerase Chain Reaction (PCR) was used to detect Leptospira. The molecular characterizations were conducted based on the 16SrRNA and LipL32 genes. Data were analyzed descriptively to describe the presence of pathogenic Leptospira DNA. Analysis phylogenetic was performed using MEGA 6.2 software. A total of 63 rats was successfullycaught during the study consisting of males and female for 36 (57.1%) and 27 (42.9%), respectively. The species of rats were R. exulans, R. tanezumi, R. argentiventer, R. norvegicus,  M. Musculus, Paruromys dominator, Maxomys sp., and Rattus sp. The pathogenic of Leptospira DNA was detected in rats with R. argentiventer and Paruromys dominatorspecies using the 16S rRNA and LipL32 gene. Sample sequences using LipL32 target gene is a close similarity with L. interrogans serovar Hardjo, serovar Autumnalis, Lai, Icterohaemorrhagiae, Balico, Grippotyphosa, Mini, Canicola, Hebdomadis; L. noguchii serovar Pomona and L. kirschneri whereas the sample sequence using 16S rRNA targetgene showed similarity with L. interrogans serovar Canicola, Copenhagen, Autumnalis, Pyrogenes, Javanica, Icterohaemorrhagiae, Manilae, Bratislava, Linhae, Hebdomadis, and L. kirschneri serovar Grippotyphosa. The PCR method with the target gene 16SrRNA and LipL32 are able to detect Leptospira spp. in rats R. argentiventer and P. dominator species Keywords: Leptospira, 16S rRNA, LipL32, PCR, Kidney’s Rat   Leptospirosis masih merupakan masalah kesehatan global karena mempengaruhikesehatan manusia di daerah pedesaan dan perkotaan, baik di negara industri maupun mnegara berkembang. Tujuan penelitian adalah untuk mendeteksi bakteri Leptospira spp di jaringan ginjal dari tikus di Dataran Tingi Napu dan Bada Kabupaten Poso, Provinsi Sulawesi Tengah. Ginjal tikus sebanyak 63 sampel dikoleksi dari Dataran Tinggi Napu dan Bada Kabupaten Poso, Provinsi Sulawesi Tengah pada bulan Mei – Juni 2018. PCR digunakan untuk mendeteksi Leptospira. Karakterisasi molekuler dilakukan berdasarkan gen 16SrRNA dan LipL32. Data dianalisis secara deskriptif untuk menggambarkan keberadaaN Leptospira yang patogenik. Analisis filogenetik dilakukan dengan menggunakan perangkat lunak Mega 6.2. Sebanyak 63 tikus berhasil ditangkap selama penelitian yang terdiri dari jantan dan betina, masing masing 36 ekor (75,1%) dan 27 ekor (42,9%). Spesies tikus adalah R. exulans, R. tanezumi, R. argentiventer, R. norvegicus, M. Musculus, Paruromys dominator, Maxomys sp, dan Rattus sp. DNA Leptospira patogenik terdeteksi pada tikus dengan spesies R. argentiventer dan Paruromys dominator menggunakan gen 16SrRNA dan LipL32 Sekuen sampel dengan target gen LipL32 menunjukkan kesamaan dengan L. interrogans serovar Hardjo, serovar Autumnalis, Lai, Icterohaemorrhagiae, Balico, Grippotyphosa, Mini, Canicola, Hebdomadis; L. noguchii serovar Pomona dan L. kirschneri. Sedangkan sekuen sampel dengan target gen 16S rRNA menunjukkan kesamaan dengan L. interrogans serovar Canicola,Copenhagen, Autumnalis, Pyrogenes, Javanica, Icterohaemorrhagiae, Manilae, Bratislava, Linhae, Hebdomadis, dan L. kirschneri serovar Grippotyphosa. Metode PCR dengan target gen 16SrRNA dan LipL32 mampu mendeteksi Leptospira spp. pada tikus dengan spesies R. argentiventer dan P. dominator. Kata kunci: Leptospira,  16S rRNA, LipL32,  PCR,  Ginjal Tikus

scholarly journals Avaliação sorológica da leptospirose e brucelose em pacientes moradores da área rural do município de Guaraci, Paraná, Brasil

2001 ◽  
Vol 34 (3) ◽  
pp. 299-300
Author(s):  
João Luis Garcia ◽  
Italmar Teodorico Navarro
Keyword(s):  
Brucella Abortus ◽  
Leptospira Interrogans ◽  
Leptospira Spp

No presente estudo foram avaliadas as soropositividades para Leptospira interrogans e Brucella abortus, através das provas de soroaglutinação microscópica e aglutinação em tubo, respectivamente, em pacientes da área rural que procuraram o Posto de Saúde do município de Guaraci, Paraná, Brasil. Das 115 amostras de soros testadas, três (2,6%) apresentaram-se sororeagentes para Leptospira spp. Com relação ao estudo sorológico da brucelose não foram verificados títulos positivos.

scholarly journals Diagnóstico molecular de Leptospira spp em matrizes suínas descartadas

2007 ◽  
Vol 44 (1) ◽  
pp. 18 ◽  
Author(s):  
Sérgio José de Oliveira ◽  
Fabrício Bortolanza ◽  
Daniel Thompsen Passos ◽  
José Antonio Simões Pires-Neto ◽  
Luiz Cesar Bello Fallavena ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Proteus Mirabilis ◽  
Rio Grande ◽  
Leptospira Interrogans ◽  
Rio Grande Do Sul ◽  
Leptospira Spp ◽  
And Staphylococcus Aureus

O Diagnóstico de leptospirose foi efetuado através de método molecular, histopatológico e sorológico em 30 matrizes suínas, descartadas, no Rio Grande do Sul, Brasil. Os objetivos foram comparar a eficiência dos 3 métodos, verificar a sensibilidade de um método de PCR que utiliza um primer único baseado na seqüência de um elemento repetitivo do genoma de Leptospira interrogans, bem como verificar a possível detecção de leptospiras em vários tecidos, incluindo o trato genital. Os animais foram selecionados com base no teste de aglutinação microscópica para incluir tanto animais negativos como positivos e com baixos e altos títulos sorológicos. As maiores freqüências (90 % dos aniamis positivos) e títulos (100 to 800) foram observados para L. interrogans serovar bratislava. Leptospiras foram detectadas por histopatologia em apenas 9 matrizes, todas com altos títulos (pelo menos 100). Um produto de PCR de 438 bp foi observado em todos os animais (fragmentos de 25 rins, 24 úteros e 9 ovidutos). Produtos de PCR similares foram obtidos em DNA de culturas de leptospiras patogênicas, enquanto a não patogênica, L. patoc apresentou um padrão distinto. Nenhum produto de amplificação de DNA de Leptospira spp foi detectado em DNA de culturas de Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella sp, Streptococcus sp and Staphylococcus aureus, ou de sangue de dois leitões. O método molecular foi, assim, específico e o mais eficiente para detectar baixos níveis de patógeno, sendo capaz de diferenciar leptospiras patogênicas e não patogênicas.

scholarly journals Leptospiral Culture without 5’-Fluorouracil Revealed Improved Leptospira Isolation from Febrile Patients in North-Eastern Malaysia

2020 ◽  
Vol 17 (4) ◽  
pp. 1307
Author(s):  
Amira Wahida Mohamad Safiee ◽  
Mohammad Ridhuan Mohd Ali ◽  
Mohd Hashairi Fauzi ◽  
Alwi Muhd Besari ◽  
Chan Yean Yean ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Whole Blood ◽  
Rapid Test ◽  
Febrile Illness ◽  
Culture Method ◽  
Clinical Samples ◽  
Rrna Gene ◽  
Leptospira Interrogans ◽  
Acute Febrile Illness ◽  
Leptospira Spp

Objectives: Isolation of Leptospira by culture represents a definitive growth and confirmation of the disease, yet it is hampered with its nature of slow growth. With slight modification of culture method, the study aims to isolate and characterize Leptospira spp. from patients with acute febrile illness. Methods: A total of 109 blood samples were collected from patients with acute febrile illness that presented at the Emergency Department of Hospital Universiti Sains Malaysia, Malaysia. Clinical samples were subjected to Leptospira IgM Rapid test, microscopic agglutination test (MAT), isolation by culture method, and direct real-time PCR test. For leptospiral isolation, the samples (whole blood and deposit from spun plasma) were cultured into modified Ellinghausen McCullough Johnson Harris (EMJH) media with and without 5’-fluorouracil (5-FU). In every culture positive sample, partial 16S rRNA gene sequencing was performed for molecular identification of the isolates. Phylogenetic analysis was carried out to determine the genetic relatedness among the isolates. An inhibition of 5-FU study was performed on Leptospira interrogans serovar Canicola with different concentrations to compare the growth detection of the tested Leptospira with or without 5-FU within 7 days of incubation. Results: Leptospirosis was diagnosed in 14.7% of patients with acute febrile illness. Two Leptospira spp. (n = 2/109, 1.85%) were successfully isolated from whole blood and deposit from spun plasma samples. B004 and B208 samples were positive at day 11 and day 7, respectively, in EMJH media without addition of 5-FU. Sample B004 was identified as Leptospira interrogans and B208 as Leptospira weilli. Phylogenetic analysis confirmed that both of them were within pathogenic group and they were not related. The 5-FU inhibition study revealed that additional of 5-FU at final concentration of 200 µg/mL to EMJH media demonstrated an inhibitory effect on the growth of the tested strain Conclusion: Isolation of Leptospira spp. using EMJH media without addition of 5’-fluorouracil resulted in a better outcome. Two pathogenic Leptospira isolates were successfully cultivated from patients with acute febrile illness that were genetically not related.

scholarly journals TYPING OF LEPTOSPIRA SPP. STRAINS BASED ON 16S rRNA

2016 ◽  
pp. 35-39
Author(s):  
Yu. V. Ostankova ◽  
A. V. Semenov ◽  
N. A. Stoyanova ◽  
N. K. Tokarevich ◽  
N. E. Lyubimova ◽  
...  
Keyword(s):  
16S Rrna ◽  
Molecular Genetic ◽  
International Database ◽  
Genetic Characteristic ◽  
Complex Type ◽  
Pathogenic Leptospira ◽  
Strain Collection ◽  
16S Rna ◽  
Leptospira Spp ◽  
16S Sequence

Aim. Comparative typing of Leptospira spp. strain collection based on analysis of 16S RNA fragment. Materials and methods. 2 pairs of primers were used for PCR, that jointly flank 1423 b.p. sized fragment. Sequences of Leptospira spp. strain 16S rRNA, presented in the international database, were used for phylogenetic analysis. Results. A high similarity, including interspecies, of the 16S fragment in Leptospira spp. strains was shown independently of the source, serovar and serogroup. Heterogeneity ofthe primary matrix, spontaneous mutations of hotspots and erroneous nucleotide couplings, characteristic for 16S sequence of pathogenic Leptospira spp. strains, are discussed. Molecular-genetic characteristic of certain reference Leptospira spp. strains by 16S sequence is obtained. Conclusion. Results of the studies give evidence on expedience of introduction into clinical practice of identification of Leptospira spp. by 16S sequence directly from the clinical material, that would allow to significantly reduce identification time, dismiss complex type-specific sera and other labor-intensive methods.

scholarly journals Biofilm Production of Leptospira spp. Strains

2018 ◽  
Vol 46 (1) ◽  
pp. 5
Author(s):  
Dayane Olímpia Gomes ◽  
Laura Gonçalves da Silva Chagas ◽  
Gabriela Bim Ramos ◽  
Andreia Zago Ciuffa ◽  
Laís Miguel Rezende ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Propidium Iodide ◽  
Biofilm Formation ◽  
Matrix Composition ◽  
Leptospira Interrogans ◽  
Biofilm Production ◽  
Leptospira Spp ◽  
The Matrix ◽  
Polypropylene Material ◽  
Pooled Sample

Background: Leptospirosis is a zoonosis that affects many species of mammals and occurs endemically in Brazil. The biofilm matrix provides structure and protection to the biofilm cells working as a physical barrier to antibiotic agents, which are attached or consumed by the matrix components. However, this attribute varies according to the matrix, antimicrobial agent and biofilm age. Leptospira may change morphologically according to environmental conditions, including cell aggregation and biofilm formation. Leptospira can colonize the ducts of kidney from hosts for a long time, forming a biofilm, which is believed to be an important factor for their maintenance in animals and in the environment. Thus, the objective of this research was to determine the biofilm formation capacity of four strains of Leptospira interrogans.Materials, Methods & Results: The strains were typified by WHO/FAO/OIE and National Collaborating Center for Reference and Research on Leptospirosis (Kit Biomedical Research, Amsterdam, Netherlands). Leptospira interrogans strains, two isolated from cattle and two isolated from dogs were biofilms tested for adhesion on polystyrene plates, extracellular matrix composition and confocal microscopy. In the plating adhesion test, the suspension was inoculated into 96-well sterile polystyrene microplates with flat bottom at a ratio of 1:200 in EMJH medium, followed by 24 h incubation at 28°C, with medium renewal after 12 h. After this period the wells were washed three times with sterile PBS and following incubation; the plates were dried in the oven at 60°C for 30 min and added 200 μL of 1% violet crystal for five min. Subsequently, the plates were washed with distilled water, after complete removal, 200 μL of acetic acid 33% was added and the readings were performed at 570 nm in the ELISA reader. The proteins and polysaccharides were quantified in a scraped pooled sample diluted in 0.85% sterile saline solution to achieve an optimal amount for testing used reagents of the BCA kit. The polysaccharide content was determined by adding into a tube, an aliquot of 0.5 mL from the pooled sample, 0.5 mL of phenol and then immediately 2.5 mL of sulfuric acid. The solution was homogenized and left to react for 15 min at room temperature. The reading was performed at 490 nm in ELISA reader. The strains were compared regarding polysaccharides and protein matrices using analysis of variance (ANOVA) and Tukey test. At confocal microscopy the strains were incubated with the tested polypropylene material for 24 h. The materials were washed with sterile phosphate buffer and stained with propidium iodide. The reading was performed using a Laser Scanning Confocal Microscope (Zeiss 710) with laser excitation (488 nm) and 580-680 nm emission filters for propidium iodide (red marking). All strains displayed strong adherence on microplate and the amount of polysaccharides in biofilm was not statistically different among the studied strains, but the amount of protein was significantly different in strain 4 (P > 0.5). The confocal microscopy showed the adherence of the Leptospira spp. strains to polypropylene material after washing.Discussion: Biofilm production plays an important role in the maintenance of a chronic infection by Leptospira interrogans with renal colonization. The exopolysaccharide (EPS) has various functions, such as checking insolubility in water; giving the three-dimensional conformation of the biofilm; protecting cells from physical (mechanical action, irradiation and temperature variations), chemical.

scholarly journals PRIMEIRO ISOLAMENTO DE Leptospira kirschneri SOROVAR CANICOLA E Leptospira interrogans SOROVAR PYROGENES EM AMOSTRAS DE URINA DE BOVINOS ABATIDOS

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Suzane Manzini ◽  
Wesley José dos Santos ◽  
Lívia Maísa Guiraldi ◽  
Isabella Neves Aires ◽  
Maria Izabel Merino de Medeiros ◽  
...  
Keyword(s):  
Sao Paulo ◽  
Leptospira Interrogans ◽  
São Paulo ◽  
Leptospira Spp

A leptospirose é uma zoonose de grande importância em saúde pública. Cento e quatro (104) amostras de urina de bovinos abatidos na região centro-oeste do Estado de São Paulo, Brasil, foram avaliadas para detectar Leptospira spp. As amostras foram mantidas em PBS e em culturas de EMJH e Fletcher. Observamos crescimento de microrganismos em 13 amostras do meio Fletcher, embora não tenha sido observado no meio EMJH. Por PCR, 25 animais (amostras) foram positivos com os primers LEP1 / LEP2, sendo que 16 amostras foram mantidas em PBS, seis em meio Fletcher e três em meio EMJH. As 25 amostras foram submetidas a PCR com o gene LipL32 para verificação da patogenicidade. Uma amostra amplificada e por sequenciamento resultou em 99,77% de similaridade com Leptospira kirschneri sorovar Canicola e Leptospira interrogans sorovar Pyrogenes. Este é o primeiro relato de isolamento de Leptospira kirschneri sorovar Canicola e Leptospira interrogans sorovar Pyrogenes em amostra de urina bovina na Região Centro-Oeste do estado de São Paulo, Brasil. Concluímos a importância da leptospirose no rebanho avaliado e a atenção para o risco ocupacional, devido à possibilidade de urina contaminada nas pastagens e disseminação da infecção para outros animais e pessoas contactantes.

scholarly journals OCORRÊNCIA DE ANTICORPOS ANTI-TOXOPLASMA GONDII E ANTI-LEPTOSPIRA SPP. EM SUÍNOS ABATIDOS EM TRÊS ABATEDOUROS DOS ESTADOS DE MINAS GERAIS E SÃO PAULO

2007 ◽  
Vol 74 (3) ◽  
pp. 267-270
Author(s):  
G.B. Pezerico ◽  
S.B. Pezerico ◽  
R.C. Silva ◽  
J.L. Hoffmann ◽  
L.B. Camargo ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Minas Gerais ◽  
Sao Paulo ◽  
Leptospira Interrogans ◽  
São Paulo ◽  
Leptospira Spp

RESUMO Diversos inquéritos soroepidemiológicos a respeito da toxoplasmose e leptospirose em suínos foram realizados em diferentes estados brasileiros, demonstrando a importância destas enfermidades para a suinocultura, bem como o potencial da espécie suína como fonte de infecção destas enfermidades para o homem. Este trabalho objetivou estabelecer as ocorrências de anticorpos anti-Toxoplasma gondii e anti-Leptospira spp. em suínos abatidos em três abatedouros, situados nos Estados de Minas Gerais e São Paulo. Foram colhidas 262 amostras de sangue de suínos terminados, provenientes de 16 municípios. Os soros foram submetidos ao método de Aglutinação Direta Modificada (MAD) para pesquisa de anticorpos anti-T. gondii, e à Soroaglutinação Microscópica (SAM), para anticorpos anti-Leptospira spp. Nenhuma amostra foi positiva para T. gondii e duas amostras reagiram para Leptospira interrogans, sorovar Pyrogenes (título 100). Os resultados devem ser em função do emprego do sistema de confinamento total, aliado ao manejo higiênico-sanitário na atividade suinícola e pelo pequeno período de permanência dos animais terminados nas granjas.

scholarly journals Detection of Leptospira spp. in the semen and urine of bulls serologically reactive to Leptospira interrogans serovar hardjo

2005 ◽  
Vol 36 (1) ◽  
Author(s):  
Fernanda Senter Magajevski ◽  
Raul Jose Silva Girio ◽  
Luis Antonio Mathias ◽  
Simone Myashiro ◽  
Margareth Élide Genovez ◽  
...  
Keyword(s):  
Leptospira Interrogans ◽  
Leptospira Spp
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