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Sample records for leptospira interrogans mediante

  1. Global Proteome Analysis of Leptospira interrogans

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Comparative global proteome analyses were performed on Leptospira interrogans serovar Copenhageni grown under conventional in vitro conditions and those mimicking in vivo conditions (iron limitation and serum presence). Proteomic analyses were conducted using iTRAQ and LC-ESI-tandem mass spectrometr...

  2. Global Proteome Analysis of Leptospira interrogans

    PubMed Central

    2009-01-01

    Comparative global proteome analyses were performed on Leptospira interrogans serovar Copenhageni grown under conventional in vitro conditions and those mimicking in vivo conditions (iron limitation and serum presence). Proteomic analyses were conducted using iTRAQ and LC-ESI-tandem mass spectrometry complemented with two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. A total of 563 proteins were identified in this study. Altered expression of 65 proteins, including upregulation of the L. interrogans virulence factor Loa22 and 5 novel proteins with homology to virulence factors found in other pathogens, was observed between the comparative conditions. Immunoblot analyses confirmed upregulation of 5 of the known or putative virulence factors in L. interrogans exposed to the in vivo-like environmental conditions. Further, ELISA analyses using serum from patients with leptospirosis and immunofluorescence studies performed on liver sections derived from L. interrogans-infected hamsters verified expression of all but one of the identified proteins during infection. These studies, which represent the first documented comparative global proteome analysis of Leptospira, demonstrated proteome alterations under conditions that mimic in vivo infection and allowed for the identification of novel putative L. interrogans virulence factors. PMID:19663501

  3. Mouse model for sublethal Leptospira interrogans infection.

    PubMed

    Richer, Luciana; Potula, Hari-Hara; Melo, Rita; Vieira, Ana; Gomes-Solecki, Maria

    2015-12-01

    Although Leptospira can infect a wide range of mammalian species, most studies have been conducted in golden Syrian hamsters, a species particularly sensitive to acute disease. Chronic disease has been well characterized in the rat, one of the natural reservoir hosts. Studies in another asymptomatic reservoir host, the mouse, have occasionally been done and have limited infection to mice younger than 6 weeks of age. We analyzed the outcome of sublethal infection of C3H/HeJ mice older than age 10 weeks with Leptospira interrogans serovar Copenhageni. Infection led to bloodstream dissemination of Leptospira, which was followed by urinary shedding, body weight loss, hypothermia, and colonization of the kidney by live spirochetes 2 weeks after infection. In addition, Leptospira dissemination triggered inflammation in the kidney but not in the liver or lung, as determined by increased levels of mRNA transcripts for the keratinocyte-derived chemokine, RANTES, macrophage inflammatory protein 2, tumor necrosis factor alpha, interleukin-1β, inducible nitric oxide synthase, interleukin-6, and gamma interferon in kidney tissue. The acquired humoral response to Leptospira infection led to the production of IgG mainly of the IgG1 subtype. Flow cytometric analysis of splenocytes from infected mice revealed that cellular expansion was primarily due to an increase in the levels of CD4(+) and double-negative T cells (not CD8(+) cells) and that CD4(+) T cells acquired a CD44(high) CD62L(low) effector phenotype not accompanied by increases in memory T cells. A mouse model for sublethal Leptospira infection allows understanding of the bacterial and host factors that lead to immune evasion, which can result in acute or chronic disease or resistance to infection (protection).

  4. Mouse model for sublethal Leptospira interrogans infection.

    PubMed

    Richer, Luciana; Potula, Hari-Hara; Melo, Rita; Vieira, Ana; Gomes-Solecki, Maria

    2015-12-01

    Although Leptospira can infect a wide range of mammalian species, most studies have been conducted in golden Syrian hamsters, a species particularly sensitive to acute disease. Chronic disease has been well characterized in the rat, one of the natural reservoir hosts. Studies in another asymptomatic reservoir host, the mouse, have occasionally been done and have limited infection to mice younger than 6 weeks of age. We analyzed the outcome of sublethal infection of C3H/HeJ mice older than age 10 weeks with Leptospira interrogans serovar Copenhageni. Infection led to bloodstream dissemination of Leptospira, which was followed by urinary shedding, body weight loss, hypothermia, and colonization of the kidney by live spirochetes 2 weeks after infection. In addition, Leptospira dissemination triggered inflammation in the kidney but not in the liver or lung, as determined by increased levels of mRNA transcripts for the keratinocyte-derived chemokine, RANTES, macrophage inflammatory protein 2, tumor necrosis factor alpha, interleukin-1β, inducible nitric oxide synthase, interleukin-6, and gamma interferon in kidney tissue. The acquired humoral response to Leptospira infection led to the production of IgG mainly of the IgG1 subtype. Flow cytometric analysis of splenocytes from infected mice revealed that cellular expansion was primarily due to an increase in the levels of CD4(+) and double-negative T cells (not CD8(+) cells) and that CD4(+) T cells acquired a CD44(high) CD62L(low) effector phenotype not accompanied by increases in memory T cells. A mouse model for sublethal Leptospira infection allows understanding of the bacterial and host factors that lead to immune evasion, which can result in acute or chronic disease or resistance to infection (protection). PMID:26416909

  5. Pathogenic Leptospira interrogans exoproteins are primarily involved in heterotrophic processes.

    PubMed

    Eshghi, Azad; Pappalardo, Elisa; Hester, Svenja; Thomas, Benjamin; Pretre, Gabriela; Picardeau, Mathieu

    2015-08-01

    Leptospirosis is a life-threatening and emerging zoonotic disease with a worldwide annual occurrence of more than 1 million cases. Leptospirosis is caused by spirochetes belonging to the genus Leptospira. The mechanisms of disease manifestation in the host remain elusive, and the roles of leptospiral exoproteins in these processes have yet to be determined. Our aim in this study was to assess the composition and quantity of exoproteins of pathogenic Leptospira interrogans and to construe how these proteins contribute to disease pathogenesis. Label-free quantitative mass spectrometry of proteins obtained from Leptospira spirochetes cultured in vitro under conditions mimicking infection identified 325 exoproteins. The majority of these proteins are conserved in the nonpathogenic species Leptospira biflexa, and proteins involved in metabolism and energy-generating functions were overrepresented and displayed the highest relative abundance in culture supernatants. Conversely, proteins of unknown function, which represent the majority of pathogen-specific proteins (presumably involved in virulence mechanisms), were underrepresented. Characterization of various L. interrogans exoprotein mutants in the animal infection model revealed host mortality rates similar to those of hosts infected with wild-type L. interrogans. Collectively, these results indicate that pathogenic Leptospira exoproteins primarily function in heterotrophic processes (the processes by which organisms utilize organic substances as nutrient sources) to maintain the saprophytic lifestyle rather than the virulence of the bacteria. The underrepresentation of proteins homologous to known virulence factors, such as toxins and effectors in the exoproteome, also suggests that disease manifesting from Leptospira infection is likely caused by a combination of the primary and potentially moonlight functioning of exoproteins.

  6. Pathogenic Leptospira interrogans Exoproteins Are Primarily Involved in Heterotrophic Processes

    PubMed Central

    Eshghi, Azad; Pappalardo, Elisa; Hester, Svenja; Thomas, Benjamin; Pretre, Gabriela

    2015-01-01

    Leptospirosis is a life-threatening and emerging zoonotic disease with a worldwide annual occurrence of more than 1 million cases. Leptospirosis is caused by spirochetes belonging to the genus Leptospira. The mechanisms of disease manifestation in the host remain elusive, and the roles of leptospiral exoproteins in these processes have yet to be determined. Our aim in this study was to assess the composition and quantity of exoproteins of pathogenic Leptospira interrogans and to construe how these proteins contribute to disease pathogenesis. Label-free quantitative mass spectrometry of proteins obtained from Leptospira spirochetes cultured in vitro under conditions mimicking infection identified 325 exoproteins. The majority of these proteins are conserved in the nonpathogenic species Leptospira biflexa, and proteins involved in metabolism and energy-generating functions were overrepresented and displayed the highest relative abundance in culture supernatants. Conversely, proteins of unknown function, which represent the majority of pathogen-specific proteins (presumably involved in virulence mechanisms), were underrepresented. Characterization of various L. interrogans exoprotein mutants in the animal infection model revealed host mortality rates similar to those of hosts infected with wild-type L. interrogans. Collectively, these results indicate that pathogenic Leptospira exoproteins primarily function in heterotrophic processes (the processes by which organisms utilize organic substances as nutrient sources) to maintain the saprophytic lifestyle rather than the virulence of the bacteria. The underrepresentation of proteins homologous to known virulence factors, such as toxins and effectors in the exoproteome, also suggests that disease manifesting from Leptospira infection is likely caused by a combination of the primary and potentially moonlight functioning of exoproteins. PMID:25987703

  7. NEW WILDLIFE HOSTS OF Leptospira interrogans IN CAMPECHE, MEXICO

    PubMed Central

    ESPINOSA-MARTÍNEZ, Deborah V.; SÁNCHEZ-MONTES, Daniel Sokani; LEÓN-PANIAGUA, Livia; RÍOS-MUÑOZ, César A.; BERZUNZA-CRUZ, Miriam; BECKER, Ingeborg

    2015-01-01

    Leptospira interrogans has been identified to cause leptospirosis, a widespread zoonotic disease that has been identified in domestic and wild animals. This work analyzed kidneys from two species of wild rodents from the state of Campeche, Mexico. Analyses were made by PCR using specific primers for detection of Leptospira interrogans DNA. The rodent species that tested positive were Heteromys gaumeri and Ototylomys phyllotis, both of which are new hosts for the bacteria in Southeastern Mexico. These records provide new insights into the disease’s transmission that should be studied carefully in order to identify other potential host species, including humans, which are at risk of becoming infected if they are in contact with infected wildlife. PMID:25923901

  8. Complete genome sequence of Leptospira interrogans serovar Bratislava, strain PigK151

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Leptospira contains pathogens serologically classified into over 250 serovars, intermediate pathogens and saprophytes with genetic classification into 21 different species. Worldwide, leptospirosis is one of the most widespread zoonoses. L. interrogans serovar Bratislava has been isolated ...

  9. [Modified EMJH medium for cultivation of Leptospira interrogans serogroup ballum].

    PubMed

    González, A; Borrero, R; Ruiz, J; Batista, N; Fernández, Y; Valdés, Y; González, M

    2006-01-01

    Strains within the Ballum serogroup of spirochete Leptospira show fastidious growth with more exigent nutritional requirements than those of other Leptospira pathogenic strains. The influence of 37 nutritional compounds on the growth of Leptospira interrogans serogroup Ballum was investigated employing the synthetic EMJH medium as the base for the study. Microbial growth was estimated spectrophotometrically and direct counts were performed with a Petroff-Hausser counting chamber. Virulence stability was evaluated by calculating the mean lethal dose in hamsters. Antigenicity stability was evaluated by Western blotting using a specific antiserum. Cell yields commonly obtained in EMJH were triplicated without virulence or antigenicity depletions after culturing in a modified EMJH medium with an increased concentration of Tween 80, and the incorporation of sodium acetate and beef extract. Neither the increased concentration of at least 6 components of EMJH nor the incorporation of a variety of new nutrients stimulated cell yields or the growth rate of the microorganism. The results allow us to make use of an enriched culture medium that promotes high cell yields of this fastidious serogroup most prevalent in humans in Cuba. PMID:17037250

  10. Draft Genome Sequence of Leptospira interrogans Serovar Bataviae Strain LepIMR 22 Isolated from a Rodent in Johor, Malaysia

    PubMed Central

    Amran, Fairuz; Mohamad, Saharuddin; Mat Ripen, Adiratna; Ahmad, Norazah; Goris, Marga G. A.; Muhammad, Ayu Haslin; Noor Halim, Nurul Atiqah

    2016-01-01

    Leptospira interrogans serovar Bataviae was recently identified as one of the persistent Leptospira serovars in Malaysia. Here, we report the draft genome sequence of the L. interrogans serovar Bataviae strain LepIMR 22 isolated from kidney of a rodent in Johor, Malaysia. PMID:27609924

  11. Draft Genome Sequence of Leptospira interrogans Serovar Bataviae Strain LepIMR 22 Isolated from a Rodent in Johor, Malaysia.

    PubMed

    Amran, Fairuz; Mohd Khalid, Mohd Khairul Nizam; Mohamad, Saharuddin; Mat Ripen, Adiratna; Ahmad, Norazah; Goris, Marga G A; Muhammad, Ayu Haslin; Noor Halim, Nurul Atiqah

    2016-01-01

    Leptospira interrogans serovar Bataviae was recently identified as one of the persistent Leptospira serovars in Malaysia. Here, we report the draft genome sequence of the L. interrogans serovar Bataviae strain LepIMR 22 isolated from kidney of a rodent in Johor, Malaysia. PMID:27609924

  12. Isolation and molecular characterization of Leptospira interrogans and Leptospira borgpetersenii isolates from the urban rat populations of Kuala Lumpur, Malaysia.

    PubMed

    Benacer, Douadi; Mohd Zain, Siti Nursheena; Amran, Fairuz; Galloway, Renee L; Thong, Kwai Lin

    2013-04-01

    Rats are considered the principal maintenance hosts of Leptospira. The objectives of this study were isolation and identification of Leptospira serovars circulating among urban rat populations in Kuala Lumpur. Three hundred urban rats (73% Rattus rattus and 27% R. norvegicus) from three different sites were trapped. Twenty cultures were positive for Leptospira using dark-field microscopy. R. rattus was the dominant carrier (70%). Polymerase chain reaction (PCR) confirmed that all isolates were pathogenic Leptospira species. Two Leptospira serogroups, Javanica and Bataviae, were identified using microscopic agglutination test (MAT). Pulsed-field gel electrophoresis (PFGE) identified two serovars in the urban rat populations: L. borgpetersenii serovar Javanica (85%) and L. interrogans serovar Bataviae (15%). We conclude that these two serovars are the major serovars circulating among the urban rat populations in Kuala Lumpur. Despite the low infection rate reported, the high pathogenicity of these serovars raises concern of public health risks caused by rodent transmission of leptospirosis.

  13. Crystallization and preliminary X-ray diffraction studies of ferredoxin reductase from Leptospira interrogans

    SciTech Connect

    Nascimento, Alessandro S.; Ferrarezi, Thiago; Catalano-Dupuy, Daniela L.; Ceccarelli, Eduardo A.; Polikarpov, Igor

    2006-07-01

    Crystals adequate for X-ray diffraction analysis have been prepared from L. interrogans ferredoxin-NADP{sup +} reductase. Ferredoxin-NADP{sup +} reductase (FNR) is an FAD-containing enzyme that catalyzes electron transfer between NADP(H) and ferredoxin. Here, results are reported of the recombinant expression, purification and crystallization of FNR from Leptospira interrogans, a parasitic bacterium of animals and humans. The L. interrogans FNR crystals belong to a primitive monoclinic space group and diffract to 2.4 Å resolution at a synchrotron source.

  14. Antibodies against Leptospira interrogans in California sea lion pups from Gulf of California.

    PubMed

    Godínez, C R; Zelaya de Romillo, B; Aurioles-Gamboa, D; Verdugo-Rodríguez, A; Rodríguez-Reyes, E A; De la Peña-Moctezuma, A

    1999-01-01

    One hundred and twenty-five serum samples from California sea lion (Zalophus californianus californianus) pups, and one from an adult female from eight reproductive rookeries located in seven islands in the Gulf of California (Mexico), were collected during the 1994-96 reproductive seasons. These were tested for antibodies to 19 serovars of Leptospira interrogans using a Microscopic Agglutination Test (MAT). Forty-one samples (32%) had antibody levels from 1:20 to 1:320 to one or more serovars. The most frequently detected serotypes were Leptospira interrogans hardjo (n = 13), cynopteri (8), ballum (6), and szwajizak (5). Serovars with the highest prevalence were Leptospira interrogans hardjo and serjoe (1:320), ballum (1:160), and cynopteri, girppotyphosa, and tarassovi (1:80). Based on these results, exposure of sea lions to L. interrogans serovar hardjo seems to be relatively common among colonies located in the islands of the Gulf of California in contrast with those located on the Pacific coast, where the most frequently detected serovar is L. interrogans serovar pomona.

  15. First Genome Sequence of Leptospira interrogans Serovar Pomona, Isolated from a Bovine Abortion.

    PubMed

    Varni, Vanina; Koval, Ariel; Nagel, Ariel; Ruybal, Paula; Caimi, Karina; Amadio, Ariel F

    2016-05-19

    Leptospirosis is a widespread zoonosis and a re-emergent disease of global distribution with major relevance in veterinary production. Here, we report the whole-genome sequence of Leptospira interrogans serovar Pomona strain AKRFB, isolated from a bovine abortion during a leptospirosis outbreak in Argentina.

  16. First Genome Sequence of Leptospira interrogans Serovar Pomona, Isolated from a Bovine Abortion

    PubMed Central

    Varni, Vanina; Koval, Ariel; Nagel, Ariel; Ruybal, Paula

    2016-01-01

    Leptospirosis is a widespread zoonosis and a re-emergent disease of global distribution with major relevance in veterinary production. Here, we report the whole-genome sequence of Leptospira interrogans serovar Pomona strain AKRFB, isolated from a bovine abortion during a leptospirosis outbreak in Argentina. PMID:27198013

  17. First Genome Sequence of Leptospira interrogans Serovar Pomona, Isolated from a Bovine Abortion.

    PubMed

    Varni, Vanina; Koval, Ariel; Nagel, Ariel; Ruybal, Paula; Caimi, Karina; Amadio, Ariel F

    2016-01-01

    Leptospirosis is a widespread zoonosis and a re-emergent disease of global distribution with major relevance in veterinary production. Here, we report the whole-genome sequence of Leptospira interrogans serovar Pomona strain AKRFB, isolated from a bovine abortion during a leptospirosis outbreak in Argentina. PMID:27198013

  18. Transcriptional response of Leptospira interrogans to iron limitation and characterization of a PerR homolog

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospira interrogans is the causative agent of leptospirosis, a zoonosis of global significance. Iron is essential for growth of most bacterial species. Since availability of iron is low in the host, pathogens have evolved complex iron acquisition mechanisms to survive and establish infection. In ...

  19. Complete Genome Sequence of Leptospira interrogans Serovar Bratislava, Strain PigK151

    PubMed Central

    Alt, David P.; Bayles, Darrell O.; Cameron, Caroline; Adler, Ben; Bulach, Dieter M.; Seemann, Torsten; Lehane, Michael J.; Haines, Lee R.; Darby, Alistair C.; Hall, Neil; Radford, Alan D.; Zuerner, Richard L.

    2015-01-01

    Leptospira interrogans serovar Bratislava infection occurs in multiple domestic and wildlife species and is associated with poor reproductive performance in swine and horses. We present the complete genome assembly of strain PigK151 comprising two chromosomes, CI (4.457 Mbp) and CII (358 kbp). PMID:26112787

  20. Characterization of a virulent Leptospira interrogans strain isolated from an abandoned swimming pool.

    PubMed

    Forster, Karine M; Hartwig, Daiane D; Seixas, Fabiana K; McBride, Alan J A; Monte, Leonardo G; Recuero, Ana Lúcia C; Brod, Claudiomar S; Hartleben, Cláudia P; Amaral, Marta; Dellagostin, Odir A

    2013-01-01

    Pathogenic Leptospira spp. are the etiological agents of leptospirosis, an important disease of both humans and animals. In urban settings, L. interrogans serovars are the predominant cause of disease in humans. The purpose of this study was to characterize a novel Leptospira isolate recovered from an abandoned swimming pool. Molecular characterization through sequencing of the rpoB gene revealed 100% identity with L. interrogans and variable-number tandem-repeat (VNTR) analysis resulted in a banding pattern identical to L. interrogans serogroup Icterohaemorrhagiae, serovar Copenhageni or Icterohaemorrhagiae. The virulence of the strain was determined in a hamster model of lethal leptospirosis. The lethal dose 50% (LD50) was calculated to be two leptospires in female hamsters and a histopathological examination of infected animals found typical lesions associated with severe leptospirosis, including renal epithelium degeneration, hepatic karyomegaly, liver-plate disarray and lymphocyte infiltration. This highly virulent strain is now available for use in further studies, especially evaluation of vaccine candidates.

  1. Leptospira interrogans Catalase Is Required for Resistance to H2O2 and for Virulence

    PubMed Central

    Eshghi, Azad; Lourdault, Kristel; Murray, Gerald L.; Bartpho, Thanatchaporn; Sermswan, Rasana W.; Picardeau, Mathieu; Adler, Ben; Snarr, Brendan; Zuerner, Richard L.

    2012-01-01

    Pathogenic Leptospira spp. are likely to encounter higher concentrations of reactive oxygen species induced by the host innate immune response. In this study, we characterized Leptospira interrogans catalase (KatE), the only annotated catalase found within pathogenic Leptospira species, by assessing its role in resistance to H2O2-induced oxidative stress and during infection in hamsters. Pathogenic L. interrogans bacteria had a 50-fold-higher survival rate under H2O2-induced oxidative stress than did saprophytic L. biflexa bacteria, and this was predominantly catalase dependent. We also characterized KatE, the only annotated catalase found within pathogenic Leptospira species. Catalase assays performed with recombinant KatE confirmed specific catalase activity, while protein fractionation experiments localized KatE to the bacterial periplasmic space. The insertional inactivation of katE in pathogenic Leptospira bacteria drastically diminished leptospiral viability in the presence of extracellular H2O2 and reduced virulence in an acute-infection model. Combined, these results suggest that L. interrogans KatE confers in vivo resistance to reactive oxygen species induced by the host innate immune response. PMID:22927050

  2. Phenotypic and Molecular Characterization of Leptospira interrogans Isolated from Canis familiaris in Southern Brazil.

    PubMed

    Jorge, Sérgio; Monte, Leonardo G; De Oliveira, Natasha R; Collares, Thais F; Roloff, Bárbara C; Gomes, Charles K; Hartwig, Daiane D; Dellagostin, Odir A; Hartleben, Cláudia P

    2015-10-01

    Leptospirosis is a zoonotic disease caused by pathogenic spirochetes from the genus Leptospira, which includes 20 species and more than 300 serovars. Canines are important hosts of pathogenic leptospires and can transmit the pathogen to humans via infected urine. Here, we report the phenotypic and molecular characterization of Leptospira interrogans isolated from Canis familiaris in Southern Brazil. The isolated strain was characterized by variable-number tandem-repeats analysis as L. interrogans, serogroup Icterohaemorrhagiae. In addition, the isolate was recognized by antibodies from human and canine serum samples previously tested by microscopic agglutination test. Ultimately, the expression of membrane-associated antigens (LipL32 and leptospiral immunoglobulin-like proteins) from pathogenic leptospires using monoclonal antibodies was detected by indirect immunofluorescence assay. In conclusion, identification of new strains of Leptospira can help in the diagnosis and control of leptospirosis. PMID:26100241

  3. High-Resolution Typing of Leptospira interrogans Strains by Multispacer Sequence Typing

    PubMed Central

    Zilber, Anne-Laure; Picardeau, Mathieu; Ayral, Florence; Artois, Marc; Demont, Pierre; Kodjo, Angeli

    2014-01-01

    Leptospirosis is a worldwide zoonosis which is responsible for the typical form of Weil's disease. The epidemiological surveillance of the Leptospira species agent is important for host prevalence control. Although the genotyping methods have progressed, the identification of some serovars remains ambiguous. We investigated the multispacer sequence typing (MST) method for genotyping strains belonging to the species Leptospira interrogans, which is the main agent of leptospirosis worldwide. A total of 33 DNA samples isolated from the reference strains of L. interrogans serogroups Icterohaemorrhagiae, Australis, Canicola, and Grippotyphosa, which are the most prevalent serogroups in France, were analyzed by both the variable-number tandem-repeat (VNTR) and MST methods. An MST database has been constructed from the DNA of these reference strains to define the MST profiles. The MST profiles corroborated with the VNTR results. Moreover, the MST analysis allowed the identification at the serovar level or potentially to the isolate level for strains belonging to L. interrogans serovar Icterohaemorrhagiae, which then results in a higher resolution than VNTR (Hunter-Gaston index of 0.94 versus 0.68). Regarding L. interrogans serogroups Australis, Canicola, and Grippotyphosa, the MST and VNTR methods similarly identified the genotype. The MST method enabled the acquisition of simple and robust results that were based on the nucleotide sequences. The MST identified clinical isolates in correlation with the reference serovar profiles, thus permitting an epidemiological surveillance of circulating L. interrogans strains, especially for the Icterohaemorrhagiae serogroup, which includes the most prevalent strains of public health interest. PMID:24478489

  4. Heterogenic colonization patterns by Leptospira interrogans in Rattus norvegicus from urban slums

    PubMed Central

    Santos, Ana Amélia Nunes; Figueira, Cláudio Pereira; dos Reis, Mitermayer Galvão; Costa, Federico; Ristow, Paula

    2015-01-01

    Abstract We evaluated the renal colonization by Leptospira interrogans in Rattus norvegicus (rats), as it is the major natural reservoir of urban leptospirosis. We caught 72 R. norvegicus, out of which 32 were found to be positive for L. interrogans by immunofluorescence assay. From these rats, we selected 17 and divided them into six groups based on the mass-age/sex. We performed the immunohistochemistry test against L. interrogans in the kidney sections of the rats and systematically counted the colonized tubules (CTs) in 20 fields. The proportion of positive fields varied from 5% to 95%. The number of CTs in 20 fields varied from 0.5 to 85.5. These differences were not related to age or sex of the animals. The characterization of leptospiral colonization patterns in the natural reservoirs is important to better understand the host-pathogen interactions in leptospirosis. PMID:26691476

  5. Heterogenic colonization patterns by Leptospira interrogans in Rattus norvegicus from urban slums.

    PubMed

    Santos, Ana Amélia Nunes; Figueira, Cláudio Pereira; dos Reis, Mitermayer Galvão; Costa, Federico; Ristow, Paula

    2015-01-01

    We evaluated the renal colonization by Leptospira interrogans in Rattus norvegicus (rats), as it is the major natural reservoir of urban leptospirosis. We caught 72 R. norvegicus, out of which 32 were found to be positive for L. interrogans by immunofluorescence assay. From these rats, we selected 17 and divided them into six groups based on the mass-age/sex. We performed the immunohistochemistry test against L. interrogans in the kidney sections of the rats and systematically counted the colonized tubules (CTs) in 20 fields. The proportion of positive fields varied from 5% to 95%. The number of CTs in 20 fields varied from 0.5 to 85.5. These differences were not related to age or sex of the animals. The characterization of leptospiral colonization patterns in the natural reservoirs is important to better understand the host-pathogen interactions in leptospirosis. PMID:26691476

  6. Leptospira interrogans induces uterine inflammatory responses and abnormal expression of extracellular matrix proteins in dogs.

    PubMed

    Wang, Wei; Gao, Xuejiao; Guo, Mengyao; Zhang, Wenlong; Song, Xiaojing; Wang, Tiancheng; Zhang, Zecai; Jiang, Haichao; Cao, Yongguo; Zhang, Naisheng

    2014-10-01

    Leptospira interrogans (L. interrogans), a worldwide zoonosis, infect humans and animals. In dogs, four syndromes caused by leptospirosis have been identified: icteric, hemorrhagic, uremic (Stuttgart disease) and reproductive (abortion and premature or weak pups), and also it caused inflammation. Extracellular matrix (ECM) is a complex mixture of matrix molecules that is crucial to the reproduction. Both inflammatory response and ECM are closed relative to reproductive. The aim of this study was to clarify how L. interrogans affected the uterus of dogs, by focusing on the inflammatory responses, and ECM expression in dogs uterine tissue infected by L. interrogans. In the present study, 27 dogs were divided into 3 groups, intrauterine infusion with L. interrogans, to make uterine infection, sterile EMJH, and normal saline as a control, respectively. The uteruses were removed by surgical operation in 10, 20, and 30 days, respectively. The methods of histopathological analysis, ELISA, Western blot and qPCR were used. The results showed that L. interrogans induced significantly inflammatory responses, which were characterized by inflammatory cellular infiltration and high expression levels of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in uterine tissue of these dogs. Furthermore, L. interrogans strongly down-regulated the expression of ECM (collagens (CL) IV, fibronectins (FN) and laminins (LN)) in mRNA and protein levels. These data indicated that strongly inflammatory responses, and abnormal regulation of ECM might contribute to the proliferation of dogs infected by L. interrogans. PMID:25153777

  7. Leptospira interrogans induces uterine inflammatory responses and abnormal expression of extracellular matrix proteins in dogs.

    PubMed

    Wang, Wei; Gao, Xuejiao; Guo, Mengyao; Zhang, Wenlong; Song, Xiaojing; Wang, Tiancheng; Zhang, Zecai; Jiang, Haichao; Cao, Yongguo; Zhang, Naisheng

    2014-10-01

    Leptospira interrogans (L. interrogans), a worldwide zoonosis, infect humans and animals. In dogs, four syndromes caused by leptospirosis have been identified: icteric, hemorrhagic, uremic (Stuttgart disease) and reproductive (abortion and premature or weak pups), and also it caused inflammation. Extracellular matrix (ECM) is a complex mixture of matrix molecules that is crucial to the reproduction. Both inflammatory response and ECM are closed relative to reproductive. The aim of this study was to clarify how L. interrogans affected the uterus of dogs, by focusing on the inflammatory responses, and ECM expression in dogs uterine tissue infected by L. interrogans. In the present study, 27 dogs were divided into 3 groups, intrauterine infusion with L. interrogans, to make uterine infection, sterile EMJH, and normal saline as a control, respectively. The uteruses were removed by surgical operation in 10, 20, and 30 days, respectively. The methods of histopathological analysis, ELISA, Western blot and qPCR were used. The results showed that L. interrogans induced significantly inflammatory responses, which were characterized by inflammatory cellular infiltration and high expression levels of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in uterine tissue of these dogs. Furthermore, L. interrogans strongly down-regulated the expression of ECM (collagens (CL) IV, fibronectins (FN) and laminins (LN)) in mRNA and protein levels. These data indicated that strongly inflammatory responses, and abnormal regulation of ECM might contribute to the proliferation of dogs infected by L. interrogans.

  8. Whole Genome Sequencing Allows Better Understanding of the Evolutionary History of Leptospira interrogans Serovar Hardjo

    PubMed Central

    Llanes, Alejandro; Restrepo, Carlos Mario; Rajeev, Sreekumari

    2016-01-01

    The genome of a laboratory-adapted strain of Leptospira interrogans serovar Hardjo was sequenced and analyzed. Comparison of the sequenced genome with that recently published for a field isolate of the same serovar revealed relatively high sequence conservation at the nucleotide level, despite the different biological background of both samples. Conversely, comparison of both serovar Hardjo genomes with those of L. borgpetersenii serovar Hardjo showed extensive differences between the corresponding chromosomes, except for the region occupied by their rfb loci. Additionally, comparison of the serovar Hardjo genomes with those of different L. interrogans serovars allowed us to detect several genomic features that may confer an adaptive advantage to L. interrogans serovar Hardjo, including a possible integrated plasmid and an additional copy of a cluster encoding a membrane transport system known to be involved in drug resistance. A phylogenomic strategy was used to better understand the evolutionary position of the Hardjo serovar among L. interrogans serovars and other Leptospira species. The proposed phylogeny supports the hypothesis that the presence of similar rfb loci in two different species may be the result of a lateral gene transfer event. PMID:27442015

  9. Whole Genome Sequencing Allows Better Understanding of the Evolutionary History of Leptospira interrogans Serovar Hardjo.

    PubMed

    Llanes, Alejandro; Restrepo, Carlos Mario; Rajeev, Sreekumari

    2016-01-01

    The genome of a laboratory-adapted strain of Leptospira interrogans serovar Hardjo was sequenced and analyzed. Comparison of the sequenced genome with that recently published for a field isolate of the same serovar revealed relatively high sequence conservation at the nucleotide level, despite the different biological background of both samples. Conversely, comparison of both serovar Hardjo genomes with those of L. borgpetersenii serovar Hardjo showed extensive differences between the corresponding chromosomes, except for the region occupied by their rfb loci. Additionally, comparison of the serovar Hardjo genomes with those of different L. interrogans serovars allowed us to detect several genomic features that may confer an adaptive advantage to L. interrogans serovar Hardjo, including a possible integrated plasmid and an additional copy of a cluster encoding a membrane transport system known to be involved in drug resistance. A phylogenomic strategy was used to better understand the evolutionary position of the Hardjo serovar among L. interrogans serovars and other Leptospira species. The proposed phylogeny supports the hypothesis that the presence of similar rfb loci in two different species may be the result of a lateral gene transfer event.

  10. First Observation of Leptospira interrogans in the Lungs of Rattus norvegicus

    PubMed Central

    Belli, Patrick; Artois, Marc; Djelouadji, Zoheira

    2016-01-01

    We report the first two cases of pulmonary presence of leptospires in apparently healthy rats captured in a city park in Lyon (France). Only renal carriage of Leptospira has been described in the literature. Blood serology was performed in parallel with molecular and histological analyses of the kidney and lung samples. We isolated leptospires from the kidneys of two out of three seropositive wild rats. These results were confirmed by specific detection of pathogenic Leptospira by real-time PCR. Moreover, Leptospira DNA was detected in lung tissues. Immunohistochemistry and Warthin-Starry staining revealed that leptospires were present on the surface of the ciliated epithelium of the bronchi. Using PCR of the rrs (16S) gene and Multispacer Sequence Typing, DNA extracts of the kidney and lung were identified as belonging to Leptospira interrogans serovar Icterohaemorrhagiae “CHU Réunion.” This first observation of the presence Leptospira in the lung with simultaneous renal carriage will require further study in future on several target organs to gain a better understanding of the Leptospira infection in wild rat. PMID:27800495

  11. Asymptomatic and chronic carriage of Leptospira interrogans serovar Pomona in California sea lions (Zalophus californianus).

    PubMed

    Prager, K C; Greig, Denise J; Alt, David P; Galloway, Renee L; Hornsby, Richard L; Palmer, Lauren J; Soper, Jennifer; Wu, Qingzhong; Zuerner, Richard L; Gulland, Frances M D; Lloyd-Smith, James O

    2013-05-31

    Since 1970, periodic outbreaks of leptospirosis, caused by pathogenic spirochetes in the genus Leptospira, have caused morbidity and mortality of California sea lions (Zalophus californianus) along the Pacific coast of North America. Yearly seasonal epizootics of varying magnitude occur between the months of July and December, with major epizootics occurring every 3-5 years. Genetic and serological data suggest that Leptospira interrogans serovar Pomona is the infecting serovar and is enzootic in the California sea lion population, although the mechanism of persistence is unknown. We report asymptomatic carriage of Leptospira in 39% (33/85) of wild, free-ranging sea lions sampled during the epizootic season, and asymptomatic seroconversion with chronic asymptomatic carriage in a rehabilitated sea lion. This is the first report of asymptomatic carriage in wild, free-ranging California sea lions and the first example of seroconversion and asymptomatic chronic carriage in a sea lion. Detection of asymptomatic chronic carriage of Leptospira in California sea lions, a species known to suffer significant disease and mortality from the same Leptospira strain, goes against widely-held notions regarding leptospirosis in accidental versus maintenance host species. Further, chronic carriage could provide a mechanism for persistent circulation of Leptospira in the California sea lion population, particularly if these animals shed infectious leptospires for months to years. PMID:23419822

  12. Asymptomatic and chronic carriage of Leptospira interrogans serovar Pomona in California sea lions (Zalophus californianus).

    PubMed

    Prager, K C; Greig, Denise J; Alt, David P; Galloway, Renee L; Hornsby, Richard L; Palmer, Lauren J; Soper, Jennifer; Wu, Qingzhong; Zuerner, Richard L; Gulland, Frances M D; Lloyd-Smith, James O

    2013-05-31

    Since 1970, periodic outbreaks of leptospirosis, caused by pathogenic spirochetes in the genus Leptospira, have caused morbidity and mortality of California sea lions (Zalophus californianus) along the Pacific coast of North America. Yearly seasonal epizootics of varying magnitude occur between the months of July and December, with major epizootics occurring every 3-5 years. Genetic and serological data suggest that Leptospira interrogans serovar Pomona is the infecting serovar and is enzootic in the California sea lion population, although the mechanism of persistence is unknown. We report asymptomatic carriage of Leptospira in 39% (33/85) of wild, free-ranging sea lions sampled during the epizootic season, and asymptomatic seroconversion with chronic asymptomatic carriage in a rehabilitated sea lion. This is the first report of asymptomatic carriage in wild, free-ranging California sea lions and the first example of seroconversion and asymptomatic chronic carriage in a sea lion. Detection of asymptomatic chronic carriage of Leptospira in California sea lions, a species known to suffer significant disease and mortality from the same Leptospira strain, goes against widely-held notions regarding leptospirosis in accidental versus maintenance host species. Further, chronic carriage could provide a mechanism for persistent circulation of Leptospira in the California sea lion population, particularly if these animals shed infectious leptospires for months to years.

  13. Characterization of a virulent Leptospira interrogans strain isolated from an abandoned swimming pool

    PubMed Central

    Forster, Karine M.; Hartwig, Daiane D.; Seixas, Fabiana K.; McBride, Alan J.A.; Monte, Leonardo G.; Recuero, Ana Lúcia C.; Brod, Claudiomar S.; Hartleben, Cláudia P.; Amaral, Marta; Dellagostin, Odir A.

    2013-01-01

    Pathogenic Leptospira spp. are the etiological agents of leptospirosis, an important disease of both humans and animals. In urban settings, L. interrogans serovars are the predominant cause of disease in humans. The purpose of this study was to characterize a novel Leptospira isolate recovered from an abandoned swimming pool. Molecular characterization through sequencing of the rpoB gene revealed 100% identity with L. interrogans and variable-number tandem-repeat (VNTR) analysis resulted in a banding pattern identical to L. interrogans serogroup Icterohaemorrhagiae, serovar Copenhageni or Icterohaemorrhagiae. The virulence of the strain was determined in a hamster model of lethal leptospirosis. The lethal dose 50% (LD50) was calculated to be two leptospires in female hamsters and a histopathological examination of infected animals found typical lesions associated with severe leptospirosis, including renal epithelium degeneration, hepatic karyomegaly, liver-plate disarray and lymphocyte infiltration. This highly virulent strain is now available for use in further studies, especially evaluation of vaccine candidates. PMID:24159300

  14. Immunological characteristics of the glycolipid antigen of Leptospira interrogans serovar lai.

    PubMed Central

    Masuzawa, T; Nakamura, R; Shimizu, T; Iwamoto, Y; Morita, T; Yanagihara, Y

    1989-01-01

    The protective antigen (PAg), a glycolipid substance, was extracted from Leptospira interrogans serovar lai strain 017 with a chloroform-methanol-water (1:2:0.8 [vol/vol/vol]) solution and partially purified by silica gel column chromatography. The PAg was not detected by Coomassie brilliant blue staining in sodium dodecyl sulfate-polyacrylamide gel electrophoresis but was observed as a smearlike band, which corresponded to a 24- to 30-kilodalton standard protein, by silver staining. The outer envelope (OE) fraction showed the same band, suggesting that the PAg was one of the chemical components of the OE. The immunogenicity and protective activity of the PAg were compared with those of the OE. The PAg as well as the OE and whole cells was able to induce agglutinating antibody against L. interrogans. Furthermore, the immune sera exhibited opsonic activity against L. interrogans, as observed by measurement of chemical luminescence derived from reactive oxygen. The PAg exhibited protective activity in hamsters challenged with lethal doses of L. interrogans. Therefore, the antigen may be useful as a component vaccine against leptospiral infection. Images PMID:2744857

  15. Leptospira interrogans serovar Copenhageni Harbors Two lexA Genes Involved in SOS Response

    PubMed Central

    Fonseca, Luciane S.; da Silva, Josefa B.; Milanez, Juliana S.; Monteiro-Vitorello, Claudia B.; Momo, Leonardo; de Morais, Zenaide M.; Vasconcellos, Silvio A.; Marques, Marilis V.; Ho, Paulo L.; da Costa, Renata M. A.

    2013-01-01

    Bacteria activate a regulatory network in response to the challenges imposed by DNA damage to genetic material, known as the SOS response. This system is regulated by the RecA recombinase and by the transcriptional repressor lexA. Leptospira interrogans is a pathogen capable of surviving in the environment for weeks, being exposed to a great variety of stress agents and yet retaining its ability to infect the host. This study aims to investigate the behavior of L. interrogans serovar Copenhageni after the stress induced by DNA damage. We show that L. interrogans serovar Copenhageni genome contains two genes encoding putative LexA proteins (lexA1 and lexA2) one of them being potentially acquired by lateral gene transfer. Both genes are induced after DNA damage, but the steady state levels of both LexA proteins drop, probably due to auto-proteolytic activity triggered in this condition. In addition, seven other genes were up-regulated following UV-C irradiation, recA, recN, dinP, and four genes encoding hypothetical proteins. This set of genes is potentially regulated by LexA1, as it showed binding to their promoter regions. All these regions contain degenerated sequences in relation to the previously described SOS box, TTTGN 5CAAA. On the other hand, LexA2 was able to bind to the palindrome TTGTAN 10TACAA, found in its own promoter region, but not in the others. Therefore, the L. interrogans serovar Copenhageni SOS regulon may be even more complex, as a result of LexA1 and LexA2 binding to divergent motifs. New possibilities for DNA damage response in Leptospira are expected, with potential influence in other biological responses such as virulence. PMID:24098496

  16. Leptospira interrogans serovar copenhageni harbors two lexA genes involved in SOS response.

    PubMed

    Fonseca, Luciane S; da Silva, Josefa B; Milanez, Juliana S; Monteiro-Vitorello, Claudia B; Momo, Leonardo; de Morais, Zenaide M; Vasconcellos, Silvio A; Marques, Marilis V; Ho, Paulo L; da Costa, Renata M A

    2013-01-01

    Bacteria activate a regulatory network in response to the challenges imposed by DNA damage to genetic material, known as the SOS response. This system is regulated by the RecA recombinase and by the transcriptional repressor lexA. Leptospira interrogans is a pathogen capable of surviving in the environment for weeks, being exposed to a great variety of stress agents and yet retaining its ability to infect the host. This study aims to investigate the behavior of L. interrogans serovar Copenhageni after the stress induced by DNA damage. We show that L. interrogans serovar Copenhageni genome contains two genes encoding putative LexA proteins (lexA1 and lexA2) one of them being potentially acquired by lateral gene transfer. Both genes are induced after DNA damage, but the steady state levels of both LexA proteins drop, probably due to auto-proteolytic activity triggered in this condition. In addition, seven other genes were up-regulated following UV-C irradiation, recA, recN, dinP, and four genes encoding hypothetical proteins. This set of genes is potentially regulated by LexA1, as it showed binding to their promoter regions. All these regions contain degenerated sequences in relation to the previously described SOS box, TTTGN 5CAAA. On the other hand, LexA2 was able to bind to the palindrome TTGTAN10TACAA, found in its own promoter region, but not in the others. Therefore, the L. interrogans serovar Copenhageni SOS regulon may be even more complex, as a result of LexA1 and LexA2 binding to divergent motifs. New possibilities for DNA damage response in Leptospira are expected, with potential influence in other biological responses such as virulence.

  17. Gene inactivation of a chemotaxis operon in the pathogen Leptospira interrogans.

    PubMed

    Lambert, Ambroise; Wong Ng, Jérôme; Picardeau, Mathieu

    2015-01-01

    Chemotaxis may have an important role in the infection process of pathogenic Leptospira spp.; however, little is known about the regulation of flagellar-based motility in these atypical bacteria. We generated a library of random transposon mutants of the pathogen L. interrogans, which included a mutant with insertion in the first gene of an operon containing the chemotaxis genes cheA, cheW, cheD, cheB, cheY and mcp. The disrupted gene encodes a putative histidine kinase (HK). The HK mutant was motile and virulent, but swarm plate and capillary assays suggested that chemotaxis was reduced in this mutant. Further analysis of bacterial trajectories by videomicroscopy showed that the ability of this mutant to reverse was significantly impaired in comparison to wild-type strain. Our data therefore show that this operon is required for full chemotaxis of Leptospira spp.

  18. Isolation and characterization of Leptospira interrogans from pigs slaughtered in São Paulo State, Brazil

    PubMed Central

    Miraglia, Fabiana; Moreno, Andréa Mike; Gomes, Cleise Ribeiro; Paixão, Renata; Liuson, Esequiel; Morais, Zenaide Maria; Maiorka, Paulo; Seixas, Fabiana Kömmling; Dellagostin, Odir Antonio; Vasconcellos, Silvio Arruda

    2008-01-01

    With the aim of isolating Leptospira spp., blood serum, kidney, liver and genital tract of 137 female swine (40 sows and 97 gilts) and also urine samples from 22 sows were collected in a slaughterhouse in the State of São Paulo, from April 2003 to August 2004. Four isolates were obtained from animals that presented microagglutination test (MAT) titers ≥ 100 for the serovar Pomona and one was obtained from an animal negative by MAT in which Leptospira was isolated from the liver and reproductive tract. The presence of leptospiral DNA was investigated by PCR, and positive results were found in kidneys of 11 females, liver of two, genital tract of two and urine of one of them. Nephrosis, interstitial multifocal nephritis, moderate to severe changing, hyalines cylinders and hemorrhagic focuses, hepatic and uterine horns congestion were histological lesions observed in higher frequency in animals positive for leptospira. The silver impregnation (Warthin Starry) confirmed the presence of spirochetes in renal tubules of four females with positive leptospira cultures from kidneys. The serogroup of the five isolates was identified as Pomona by cross agglutination with reference polyclonal antibodies. Molecular characterization of the isolates was carried out by variable-number tandem-repeats analysis. All the isolates revealed a pattern distinct from the L. interrogans Pomona type strain, but identical to a previously identified pattern from strains isolated in Argentina belonging to serovar Pomona. PMID:24031254

  19. Generation of mammalian host-adapted Leptospira interrogans by cultivation in peritoneal dialysis membrane chamber implantation in rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospira interrogans can infect a myriad of mammalian hosts, including humans (Bharti, Nally et al. 2003, Ko, Goarant et al. 2009). Following acquisition by a suitable host, leptospires disseminate via the bloodstream to multiple tissues, including the kidneys, where they adhere to and colonize th...

  20. Molecular characterization, serotyping, and antibiotic susceptibility profile of Leptospira interrogans serovar Copenhageni isolates from Brazil.

    PubMed

    Miraglia, Fabiana; Matsuo, Minekazo; Morais, Zenaide Maria; Dellagostin, Odir Antonio; Seixas, Fabiana Kömmling; Freitas, Julio César; Hartskeerl, Rudy; Moreno, Luisa Zanolli; Costa, Bárbara Letícia; Souza, Gisele Oliveira; Vasconcellos, Silvio Arruda; Moreno, Andrea Micke

    2013-11-01

    Leptospira interrogans serogroup Icterohaemorrhagiae is the major serogroup infecting humans worldwide, and rodents and dogs are the most significant transmission sources in urban environments. Knowledge of the prevalent serovars and their maintenance hosts is essential to understand the epidemiology of leptospirosis. In this study, 20 Leptospira isolates were evaluated by pulsed-field gel electrophoresis (PFGE), variable number tandem-repeat analysis (VNTR), serotyping, and determination of antimicrobial resistance profile. Isolates, originated from bovine, canine, human, and rodent sources, were characterized by microscopic agglutination test with polyclonal and monoclonal antibodies and were identified as L. interrogans serogroup Icterohaemorrhagiae serovar Copenhageni. MICs of antimicrobials often used in veterinary medicine were determined by broth microdilution test. Most of tested antibiotics were effective against isolates, including penicillin, ampicillin, and ceftiofur. Higher MIC variability was observed for fluoroquinolones and neomycin; all isolates were resistant to trimethoprim/sulfamethoxazole and sulphadimethoxine. Isolates were genotyped by PFGE and VNTR; both techniques were unable to discriminate between serovars Copenhageni and Icterohaemorrhagiae, as expected. PFGE clustered all isolates in 1 pulsotype, indicating that these serovars can be transmitted between species and that bovine, rodent, and dogs can maintain them in the environment endangering the human population. PMID:24054736

  1. Molecular Characterization of Thermoinduced Immunogenic Proteins Q1p42 and Hsp15 of Leptospira interrogans

    PubMed Central

    Nally, Jarlath E.; Artiushin, Sergey; Timoney, John F.

    2001-01-01

    Leptospira interrogans is a mammalian pathogen which must adapt to a range of new environmental conditions including temperature change when it infects new hosts. In vitro studies of organisms cultured at 30°C and shifted to 37°C for 5 to 7 days have confirmed that synthesis of several proteins involved in equine infection is regulated in response to temperature change (J. E. Nally, J. F. Timoney, and B. Stevenson, Infect. Immun. 69:400–404, 2001). In order to specifically identify antigenic proteins upregulated at 37°C, groups of three ponies were immunized with organisms shifted to 37°C for 5 to 7 days or with organisms maintained at 30°C. A lambda ZAP II genomic DNA library was screened with the pool of antisera to organisms shifted to 37°C. Clones reactive with this pool but unreactive with the pool of pony antisera to organisms cultured at 30°C were selected for further analysis. Sequence analysis of the first two clones identified open reading frames for proteins designated Qlp42 and Hsp15. Qlp42 is predicted to be an outer membrane lipoprotein. Its synthesis was upregulated when cultures were shifted from 30 to 37°C and downregulated when cultures were shifted from 37 to 30°C. Although the predicted molecular mass of Qlp42 is 39.8 kDa for the mature protein, Qlp42-specific equine antiserum was reactive with two bands of 30 and 29.5 kDa. Hsp15 is a stress response protein and a member of the Hsp20/α-crystallin family. PCR detected homologues of qlp42 and hsp15 in pathogenic serovars of L. interrogans but not in the nonpathogenic Leptospira biflexa. Enzyme-linked immunosorbent assays of antibody in convalescent sera from mares naturally infected with L. interrogans suggest that Qlp42 is expressed during leptospiral infection. PMID:11705941

  2. Leptospira interrogans lpxD Homologue Is Required for Thermal Acclimatization and Virulence.

    PubMed

    Eshghi, Azad; Henderson, Jeremy; Trent, M Stephen; Picardeau, Mathieu

    2015-11-01

    Leptospirosis is an emerging disease with an annual occurrence of over 1 million human cases worldwide. Pathogenic Leptospira bacteria are maintained in zoonotic cycles involving a diverse array of mammals, with the capacity to survive outside the host in aquatic environments. Survival in the diverse environments encountered by Leptospira likely requires various adaptive mechanisms. Little is known about Leptospira outer membrane modification systems, which may contribute to the capacity of these bacteria to successfully inhabit and colonize diverse environments and animal hosts. Leptospira bacteria carry two genes annotated as UDP-3-O-[3-hydroxymyristoyl] glucosamine N-acyltransferase genes (la0512 and la4326 [lpxD1 and lpxD2]) that in other bacteria are involved in the early steps of biosynthesis of lipid A, the membrane lipid anchor of lipopolysaccharide. Inactivation of only one of these genes, la0512/lpxD1, imparted sensitivity to the host physiological temperature (37°C) and rendered the bacteria avirulent in an animal infection model. Polymyxin B sensitivity assays revealed compromised outer membrane integrity in the lpxD1 mutant at host physiological temperature, but structural analysis of lipid A in the mutant revealed only minor changes in the lipid A moiety compared to that found in the wild-type strain. In accordance with this, an in trans complementation restored the phenotypes to a level comparable to that of the wild-type strain. These results suggest that the gene annotated as lpxD1 in Leptospira interrogans plays an important role in temperature adaptation and virulence in the animal infection model.

  3. Leptospira interrogans lpxD Homologue Is Required for Thermal Acclimatization and Virulence

    PubMed Central

    Eshghi, Azad; Henderson, Jeremy; Trent, M. Stephen

    2015-01-01

    Leptospirosis is an emerging disease with an annual occurrence of over 1 million human cases worldwide. Pathogenic Leptospira bacteria are maintained in zoonotic cycles involving a diverse array of mammals, with the capacity to survive outside the host in aquatic environments. Survival in the diverse environments encountered by Leptospira likely requires various adaptive mechanisms. Little is known about Leptospira outer membrane modification systems, which may contribute to the capacity of these bacteria to successfully inhabit and colonize diverse environments and animal hosts. Leptospira bacteria carry two genes annotated as UDP-3-O-[3-hydroxymyristoyl] glucosamine N-acyltransferase genes (la0512 and la4326 [lpxD1 and lpxD2]) that in other bacteria are involved in the early steps of biosynthesis of lipid A, the membrane lipid anchor of lipopolysaccharide. Inactivation of only one of these genes, la0512/lpxD1, imparted sensitivity to the host physiological temperature (37°C) and rendered the bacteria avirulent in an animal infection model. Polymyxin B sensitivity assays revealed compromised outer membrane integrity in the lpxD1 mutant at host physiological temperature, but structural analysis of lipid A in the mutant revealed only minor changes in the lipid A moiety compared to that found in the wild-type strain. In accordance with this, an in trans complementation restored the phenotypes to a level comparable to that of the wild-type strain. These results suggest that the gene annotated as lpxD1 in Leptospira interrogans plays an important role in temperature adaptation and virulence in the animal infection model. PMID:26283339

  4. Heat stability of protective antigen of Leptospira interrogans serovar lai.

    PubMed Central

    Masuzawa, T; Nakamura, R; Shimizu, T; Yanagihara, Y

    1990-01-01

    Protective antigen (PAg; glycolipid antigen; molecular size, 23 to 30 kilodaltons), the serogroup-specific antigen partially purified from leptospiral cells, is one of the most important protective antigens. The heat stability of PAg was compared with that of whole-cell (WC) antigen by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblotting, protective activity, opsonin-inducing activity, agglutinating antibody-inducing activity, and an inhibition test in an enzyme-linked immunosorbent assay. A band of 23 to 30 kilodaltons of PAg, which was seen in untreated PAg and WC, shifted to a position with a molecular size of ca. 20 kilodaltons after heat treatment of PAg at 80 degrees C for 30 min and WC at 100 degrees C for 30 min. In the enzyme-linked immunosorbent assay inhibition test with monoclonal antibody LW2 and a sonicated antigen of WC, the inhibition rate of PAg and WC to sonicated WC was reduced by heat treatment at 80 degrees C for 30 min and at 100 degrees C for 30 min, respectively. Agglutinating antibody-inducing activities and opsonin-inducing activities of PAg and WC in mice were reduced by heat treatment under the same conditions; these activities were assayed by a microscopic agglutination test and by chemical luminescence response in serum from immunized mice, respectively. Protective activity of heated PAg and heated WC in cyclophosphamide-pretreated mice agreed with the results of immunogenicity in mice. These results indicate that the Leptospira PAg is one of the important protective antigens and is altered by heat treatment at 80 degrees C. Furthermore, the immunogenicity and antigenicity of the PAg present in WC are more stable than that of the extracted PAg, and the coexistence of other cellular components with PAg might protect and stabilize PAg from the heat treatment. Images PMID:2332463

  5. Isolation of Leptospira interrogans from suburban rats in Tandil, Buenos Aires, Argentina.

    PubMed

    Scialfa, E; Bolpe, J; Bardón, J C; Ridao, G; Gentile, J; Gallicchio, O

    2010-01-01

    The main objective of this study was to investigate the role of wild rodents as Leptospira spp. reservoirs in a suburban area of Tandil city, Buenos Aires province (Argentina), where a person had died due to pulmonary leptospirosis. The specific objectives were: to estimate the rodent density near the patient's home, to determine the serological prevalence and isolation of leptospirosis from wild rodents, and to identify the isolated strains. The area examined was a suburban neighbourhood in Tandil near the Langueyú stream, where the patient's house is located. Rattus norvegicus were trapped on the stream banks during two nights and a high capture rate (70%), was obtained. All rats (42) were examined serologically by the microscopic agglutination test (MAT), and 22 of them (52.3%) reacted with Leptospira serovars castellonis, canicola, grippotyphosa, icterohaemorrhagiae and hebdomadis at a titer of 1:50. The kidneys from 25 animals were cultured, and 24 isolates of L. interrogans (96%) were obtained. The isolated strains were identified as Icterohaemorrhagiae serogroup by MAT performed with rabbit hyperimmune reference sera. These findings showed a high density of suburban rodents highly infected with pathogenic leptospira, sharing environment in close contact with humans with evidence of leptospiral disease.

  6. Vaccination with leptospiral outer membrane lipoprotein LipL32 reduces kidney invasion of Leptospira interrogans serovar canicola in hamsters.

    PubMed

    Humphryes, P C; Weeks, M E; AbuOun, M; Thomson, G; Núñez, A; Coldham, N G

    2014-04-01

    The Leptospira interrogans vaccines currently available are serovar specific and require regular booster immunizations to maintain protection of the host. In addition, a hamster challenge batch potency test is necessary to evaluate these vaccines prior to market release, requiring the use of a large number of animals, which is ethically and financially undesirable. Our previous work showed that the N terminus of the outer membrane protein LipL32 was altered in Leptospira interrogans serovar Canicola vaccines that fail the hamster challenge test, suggesting that it may be involved in the protective immune response. The aim of this study was to determine if vaccination with LipL32 protein alone could provide a protective response against challenge with L. interrogans serovar Canicola to hamsters. Recombinant LipL32, purified from an Escherichia coli expression system, was assessed for protective immunity in five groups of hamsters (n = 5) following a challenge with the virulent L. interrogans serovar Canicola strain Kito as a challenge strain. However, no significant survival against the L. interrogans serovar Canicola challenge was observed compared to that of unvaccinated negative controls. Subsequent histological analysis revealed reduced amounts of L. interrogans in the kidneys from the hamsters vaccinated with recombinant LipL32 protein prior to challenge; however, no significant survival against the L. interrogans serovar Canicola challenge was observed compared to that of unvaccinated negative controls. This finding corresponded to a noticeably reduced severity of renal lesions. This study provides evidence that LipL32 is involved in the protective response against L. interrogans serovar Canicola in hamsters and is the first reported link to LipL32-induced protection against kidney invasion.

  7. ATCC 43642 replaces ATCC 23581 as the type strain of Leptospira interrogans (Stimson 1907) Wenyon 1926. Opinion 91. Judicial Commission of the International Committee on Systematics of Prokaryotes.

    PubMed

    Tindall, B J

    2014-10-01

    The Judicial Commission affirms that, according to information presented to it, the type strain of Leptospira interrogans (Stimson 1907) Wenyon 1926 designated on the Approved Lists of Bacterial Names (ATCC 23581) has been shown not to represent an authentic culture of strain RGA (a member of the serovar Icterohaemorrhagiae) and ATCC 43642, derived from an authentic strain of strain RGA, a member of the serovar Icterohaemorrhagiae, is designated the type strain of Leptospira interrogans (Stimson 1907) Wenyon 1926.

  8. Ecology of Leptospira interrogans in Norway Rats (Rattus norvegicus) in an Inner-City Neighborhood of Vancouver, Canada

    PubMed Central

    Himsworth, Chelsea G.; Bidulka, Julie; Parsons, Kirbee L.; Feng, Alice Y. T.; Tang, Patrick; Jardine, Claire M.; Kerr, Thomas; Mak, Sunny; Robinson, John; Patrick, David M.

    2013-01-01

    Background Leptospira interrogans is a bacterial zoonosis with a worldwide distribution for which rats (Rattus spp.) are the primary reservoir in urban settings. In order to assess, monitor, and mitigate the risk to humans, it is important to understand the ecology of this pathogen in rats. The objective of this study was to characterize the ecology of L. interrogans in Norway rats (Rattus norvegicus) in an impoverished inner-city neighborhood of Vancouver, Canada. Methodology/Principal Findings Trapping was performed in 43 city blocks, and one location within the adjacent port, over a 12 month period. Kidney samples were tested for the presence of L. interrogans using PCR and sequencing. A multivariable model was built to predict L. interrogans infection status in individual rats using season and morphometric data (e.g., weight, sex, maturity, condition, etc.) as independent variables. Spatial analysis was undertaken to identify clusters of high and low L. interrogans prevalence. The prevalence of L. interrogans varied remarkably among blocks (0–66.7%), and spatial clusters of both high and low L. interrogans prevalence were identified. In the final cluster-controlled model, characteristics associated with L. interrogans-infection in rats included weight (OR = 1.14, 95% CI = 1.07–1.20), increased internal fat (OR = 2.12, 95% CI = 1.06–4.25), and number of bite wounds (OR = 1.20, 95% CI = 0.96–1.49). Conclusions/Significance Because L. interrogans prevalence varied with weight, body fat, and bite wounds, this study suggests that social structure and interactions among rats may influence transmission. The prevalence and distribution of L. interrogans in rats was also highly variable even over a short geographic distance. These factors should be considered in future risk management efforts. PMID:23818996

  9. Comparative seroprevalence of Leptospira interrogans in Colombian mammals along a climatic gradient.

    PubMed

    Astudillo, Viviana González; Hernández, Dave Wehdeking; Stadlin, Juliana Peña; Bernal, Leonardo Arias; Rodríguez, Dora Adriana Lombo; Hernández, Miryam Astudillo

    2012-12-01

    Leptospirosis is a widespread zoonotic disease with well-established impacts on human health in tropical and subtropical regions. Although Leptospira spp. are known to readily infect many wildlife species, the understanding of interspecies and climatic variability in patterns of infection in Neotropical mammals is limited. To improve the understanding of this interplay, 85 mammals representing 17 species were sampled from four Colombian zoos along a climatic gradient. Prevalence of the 21 primary serovars against Leptospira interrogans was determined using the microagglutination test. Individuals were considered positive for a given serovar if antibodies were observable at a 1:100 dilution or greater. Overall prevalence was 9.52%, with positive titers to serovar hurstbridge in Carnivora (Canidae); serovar sarmin in Primata (Atelidae); and serovars australis, mini, autumnalis, pomona, icterohaemorrhagiae, and seramanga in Primata (Cebidae). Prevalence was positively correlated with humidity and temperature, with significantly higher prevalence at the site characterized by high humidity, severe flooding because of rainfall, and warm weather throughout the year. All positive animals were classified as clinically asymptomatic, meaning that antibodies from a current or past infection were detected but no overt symptoms were apparent. The diversity of serovars observed and the taxon-specific nature of these associations suggest that the epidemiology of Leptospira transmission is likely to be complex and multidimensional. The strong association observed between prevalence and climate suggests that the important role of climate as an indicator of Leptospira infection risk in humans may also be applicable to wildlife. Future studies in both wild and captive populations of Neotropical wildlife will further elucidate this disease interplay. PMID:23272343

  10. Leptospira Interrogans Induces Fibrosis in the Mouse Kidney through Inos-Dependent, TLR- and NLR-Independent Signaling Pathways

    PubMed Central

    Fanton d'Andon, Martine; Quellard, Nathalie; Fernandez, Béatrice; Ratet, Gwenn; Lacroix-Lamandé, Sonia; Vandewalle, Alain; Boneca, Ivo G.; Goujon, Jean-Michel; Werts, Catherine

    2014-01-01

    Background Leptospira (L.) interrogans are bacteria responsible for a worldwide reemerging zoonosis. Rodents carry L. interrogans asymptomatically in their kidneys and excrete bacteria in the urine, contaminating the environment. Humans get infected through skin contact and develop a mild or severe leptospirosis that may lead to renal failure and fibrosis. L. interrogans provoke an interstitial nephritis, but the induction of fibrosis caused by L. interrogans has not been studied in murine models. Innate immune receptors from the TLR and NLR families have recently been shown to play a role in the development and progression of tissue fibrosis in the lung, liver and kidneys under different pathophysiological situations. We recently showed that TLR2, TLR4, and NLRP3 receptors were crucial in the defense against leptospirosis. Moreover, infection of a human cell line with L. interrogans was shown to induce TLR2-dependent production of fibronectin, a component of the extracellular matrix. Therefore, we thought to assess the presence of renal fibrosis in L. interrogans infected mice and to analyze the contribution of some innate immune pathways in this process. Methodology/principal findings Here, we characterized by immunohistochemical studies and quantitative real-time PCR, a model of Leptospira-infected C57BL/6J mice, with chronic carriage of L. interrogans inducing mild renal fibrosis. Using various strains of transgenic mice, we determined that the renal infiltrates of T cells and, unexpectedly, TLR and NLR receptors, are not required to generate Leptospira-induced renal fibrosis. We also show that the iNOS enzyme, known to play a role in Leptospira-induced interstitial nephritis, also plays a role in the induction of renal fibrosis. Conclusion/significance To our knowledge, this work provides the first experimental murine model of sustained renal fibrosis induced by a chronic bacterial infection that may be peculiar, since it does not rely on TLR or NLR receptors

  11. Leptospira interrogans serovar hardjo Infection in Cattle in the South Okanagan District of British Columbia

    PubMed Central

    Kingscote, Barbara F.

    1985-01-01

    An outbreak of leptospirosis due to Leptospira interrogans serovar hardjo in the South Okanagan District of British Columbia was investigated. The infection was associated primarily with bulls, but serovar hardjo was isolated from both bulls and cows at slaughter. Kidney and cerebrospinal fluid were found to contain leptospires, independently of the presence and level of serum agglutinins. Treatment of a bull twice in six months with dihydrostreptomycin failed to diminish an agglutinin titer (1/200) which persisted for two years without reexposure of the bull. A serological survey of cull cows sold through a central auction mart revealed the presence of hardjo agglutinins in 15.4% of 1300 sera representing 163 herds in 20 locations. Thirty percent of these herds contained reactor cattle. The number of premises from which reactor cattle came in a given locality varied from 4% to 67.7%. Measures to control leptospirosis in the study are suggested. PMID:17422584

  12. Leptospira interrogans serovars Bratislava and Muenchen animal infections: Implications for epidemiology and control.

    PubMed

    Arent, Z; Frizzell, C; Gilmore, C; Allen, A; Ellis, W A

    2016-07-15

    Strains of Leptospira interrogans belonging to two very closely related serovars - Bratislava and Muenchen - have been associated with disease in domestic animals, in particular pigs, but also in horses and dogs. Similar strains have also been recovered from various wildlife species. Their epidemiology is poorly understood. Two hundred and forty seven such isolates, from UK domestic animal and wildlife species, were examined by restriction endonuclease analysis in an attempt to elucidate their epidemiology. A representative sub-sample of 65 of these isolates was further examined by multiple-locus variable-number tandem repeat analysis and 22 by secY sequencing. Ten restriction pattern types were identified. The majority of isolates fell into one of three restriction endonuclease analysis pattern types designated B2a, B2b and M2a. B2a was ubiquitous and was isolated from 10 species and represented the majority of the horse and all dog isolates. B2b was very different, being isolated only from pigs, indicating that this type was maintained by pigs. The pattern M2a was reported for the majority of isolates from pigs but also was common in small rodents isolates. Five restriction pattern types were found only in wildlife suggesting that they are unlikely to pose a disease threat to domestic animals. Multiple-locus variable-number tandem repeat analysis identified six clusters. The REA types B2a and B2b were all found in one MLVA cluster while the majority of the M2a strains examined occurred in another cluster. The secY sequencing detected only one sequence type, clustered with other serovars of Leptospira interrogans. PMID:27283852

  13. Lsa63, a newly identified surface protein of Leptospira interrogans binds laminin and collagen IV.

    PubMed

    Vieira, Monica L; de Morais, Zenaide M; Gonçales, Amane P; Romero, Eliete C; Vasconcellos, Silvio A; Nascimento, Ana L T O

    2010-01-01

    Leptospira interrogans is the etiological agent of leptospirosis, a zoonotic disease that affects populations worldwide. We have identified in proteomic studies a protein that is encoded by the gene LIC10314 and expressed in virulent strain of L. interrogans serovar Pomona. This protein was predicted to be surface exposed by PSORT program and contains a p83/100 domain identified by BLAST analysis that is conserved in protein antigens of several strains of Borrelia and Treponema spp. The proteins containing this domain have been claimed antigen candidates for serodiagnosis of Lyme borreliosis. Thus, we have cloned the LIC10314 and expressed the protein in Escherichia coli BL21-SI strain by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. This protein is conserved among several species of pathogenic Leptospira and absent in the saprophytic strain L. biflexa. We confirm by liquid-phase immunofluorescence assays with living organisms that this protein is most likely a new surface leptospiral protein. The ability of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC10314, named Lsa63 (Leptospiral surface adhesin of 63kDa), binds strongly to laminin and collagen IV in a dose-dependent and saturable fashion. In addition, Lsa63 is probably expressed during infection since it was recognized by antibodies of serum samples of confirmed-leptospirosis patients in convalescent phase of the disease. Altogether, the data suggests that this novel identified surface protein may be involved in leptospiral pathogenesis.

  14. Whole-Genome Sequence of Leptospira interrogans Serovar Hardjo Subtype Hardjoprajitno Strain Norma, Isolated from Cattle in a Leptospirosis Outbreak in Brazil

    PubMed Central

    Soares, S. C.; Mendes, T. A.; Raittz, R. T.; Moreira, E. C.; Leite, R.; Fernandes, G. R.; Haddad, J. P. A.; Ortega, J. Miguel

    2015-01-01

    Leptospirosis is caused by pathogenic bacteria of the genus Leptospira spp. This neglected re-emergent disease has global distribution and relevance in veterinary production. Here, we report the whole-genome sequence and annotation of Leptospira interrogans serovar Hardjo subtype Hardjoprajitno strain Norma, isolated from cattle in a livestock leptospirosis outbreak in Brazil. PMID:26543126

  15. Geographical dissemination of Leptospira interrogans serovar Pomona during seasonal migration of California sea lions.

    PubMed

    Zuerner, Richard L; Cameron, Caroline E; Raverty, Stephen; Robinson, John; Colegrove, Kathleen M; Norman, Stephanie A; Lambourn, Dyanna; Jeffries, Steven; Alt, David P; Gulland, Frances

    2009-05-28

    Leptospirosis is one of the most widespread bacterial zoonoses in the world and affects most mammalian species. Although leptospirosis is well documented and characterized in terrestrial species, less information is available regarding the distribution and impact of leptospirosis in marine mammals. Additionally, the role of animal migrations on the geographical spread of leptospirosis has not been reported. Periodic epizootic outbreaks of acute leptospirosis among California sea lions (Zalophus californianus) have been reported since 1971. In this study, we collected samples from California sea lions stranded along the Pacific coast of North America during the most recent epidemic in 2004, and maintained leptospirosis surveillance of the California sea lion population along the California coast through 2007. Several isolates of Leptospira interrogans serovar Pomona were obtained from kidney and urine samples collected during this study, a finding consistent with serological evidence that California sea lions are persistently exposed to this leptospiral serovar. Combined, these data support a model whereby California sea lions are maintenance hosts for L. interrogans serovar Pomona, yet periodically undergo outbreaks of acute infection. During the 2004 outbreak, the incidence of new leptospirosis cases among California sea lions coincided with the seasonal movement of male sea lions from rookeries along the coast of central and southern California north as far as British Columbia. These data show that seasonal animal movement contributes to the distribution of leptospirosis across a large geographical region. PMID:19186009

  16. Characterization of a bifunctional enzyme with (p)ppGpp-hydrolase/synthase activity in Leptospira interrogans.

    PubMed

    He, Ping; Deng, Cong; Liu, Boyu; Zeng, LingBing; Zhao, Wei; Zhang, Yan; Jiang, XuCheng; Guo, XiaoKui; Qin, JinHong

    2013-11-01

    Alarmone Guanosine 5'-diphosphate (or 5'-triphosphate) 3'-diphosphate [(p)ppGpp] is the key component that globally regulates stringent control in bacteria. There are two homologous enzymes, RelA and SpoT in Escherichia coli, which are responsible for fluctuations in (p)ppGpp concentration inside the cell, whereas there exists only a single RelA/SpoT enzyme in Gram-positive bacteria. We have identified a bifunctional enzyme with (p)ppGpp-hydrolase/synthase activity in Leptospira interrogans. We show that the relLin gene (LA_3085) encodes a protein that fully complements the relA/spoT double mutants in E. coli. The protein functions as a (p)ppGpp degradase as well as a (p)ppGpp synthase when the cells encounter amino acid stress and deprivation of carbon sources. N-terminus HD and RSD domains of relLin (relLinN ) were observed to restore growth of double mutants of E. coli. Finally, We demonstrate that purified RelLin and RelLinN show high (p)ppGpp synthesis activity in vitro. Taken together, our results suggest that L. interrogans contain a single Rel-like bifunctional protein, RelLin , which plays an important role in maintaining the basal level of (p)ppGpp in the cell potentially contributing to the regulation of bacterial stress response.

  17. Geographical dissemination of Leptospira interrogans serovar Pomona during seasonal migration of California sea lions.

    PubMed

    Zuerner, Richard L; Cameron, Caroline E; Raverty, Stephen; Robinson, John; Colegrove, Kathleen M; Norman, Stephanie A; Lambourn, Dyanna; Jeffries, Steven; Alt, David P; Gulland, Frances

    2009-05-28

    Leptospirosis is one of the most widespread bacterial zoonoses in the world and affects most mammalian species. Although leptospirosis is well documented and characterized in terrestrial species, less information is available regarding the distribution and impact of leptospirosis in marine mammals. Additionally, the role of animal migrations on the geographical spread of leptospirosis has not been reported. Periodic epizootic outbreaks of acute leptospirosis among California sea lions (Zalophus californianus) have been reported since 1971. In this study, we collected samples from California sea lions stranded along the Pacific coast of North America during the most recent epidemic in 2004, and maintained leptospirosis surveillance of the California sea lion population along the California coast through 2007. Several isolates of Leptospira interrogans serovar Pomona were obtained from kidney and urine samples collected during this study, a finding consistent with serological evidence that California sea lions are persistently exposed to this leptospiral serovar. Combined, these data support a model whereby California sea lions are maintenance hosts for L. interrogans serovar Pomona, yet periodically undergo outbreaks of acute infection. During the 2004 outbreak, the incidence of new leptospirosis cases among California sea lions coincided with the seasonal movement of male sea lions from rookeries along the coast of central and southern California north as far as British Columbia. These data show that seasonal animal movement contributes to the distribution of leptospirosis across a large geographical region.

  18. Immunological and molecular characterization of Leptospira interrogans isolated from a bovine foetus.

    PubMed

    Monte, Leonardo Garcia; Ridieri, Karine Forster; Jorge, Sérgio; Oliveira, Natasha Rodrigues; Hartwig, Daiane Drawanz; Amaral, Marta Gonçalves; Hartleben, Cláudia Pinho; Dellagostin, Odir Antonio

    2015-06-01

    Cattle are commonly infected with pathogenic leptospires, and similarly to rodents, they excrete the bacteria in their urine and can transmit the pathogen from animal to animal or animal to human. Thus, surveillance and monitoring systems for detection of new Leptospira serovars are important for the control of leptospirosis. Here, we report the isolation of a spirochete from a stillborn bovine foetus and its characterization by immunological and molecular techniques. A variable number tandem repeat profile using seven discriminatory primers identified the spirochete as belonging to species Leptospira interrogans serogroup Australis serovar Muenchen. A phenotypic analysis using monoclonal antibodies (mAbs) against leptospiral membrane-associated proteins confirmed the expression of important virulence and pathogenicity factors (LipL32 and LigBrep). Out of 120 reference sera tested, 22 positive (36.66%) and 9 negative (15%) also reacted with the new isolate. Furthermore, the serovar Muenchen isolate was virulent in hamster model. The animal inoculated developed acute lethal infection characterized by hepatic, pulmonary and renal lesions. Local isolates exhibited unique characteristics that differed from those of reference strains; therefore, isolation of leptospires is useful in the surveillance of local pathogenic serovars. In conclusion, the data obtained from this study can contribute to the epidemiological understanding and control of leptospirosis in southern Brazil.

  19. First isolation of Leptospira interrogans from Lycalopex griseus (South American gray fox) in Argentina shows new MLVA genotype.

    PubMed

    Scialfa, Exequiel; Brihuega, Bibiana; Venzano, Agustín; Morris, Winston Eduardo; Bolpe, Jorge; Schettino, Mateo

    2013-01-01

    To identify carriers of Leptospira spp. in Argentina, wild animals were trapped in Buenos Aires Province during three nights, capturing 12 Didelphis albiventris (white-eared opossum), six Chaetophractus villosus (big hairy armadillo), five Lycalopex griseus (South American gray fox), and two Conepatus chinga (Molina's hog-nosed skunk). All were tested by microscopic agglutination test, and five (two gray foxes, two armadillos, and one skunk) were positive for Leptospira interrogans serovars Canicola and Icterohaemorrhagiae, L. borgpetersenii serovar Castellonis, and L. kirschneri serovar Grippotyphosa, at titers of 1:50 and 1:100. Kidney tissue from all animals was cultured, and one isolate of L. interrogans from a gray fox was obtained. Hamsters inoculated with the isolate died after 6 days with no macroscopic lesions at necropsy. However, histologic examination revealed glomerulonephritis, interstitial nephritis, and pneumonia. The Leptospira strain from the South American gray fox was analyzed serologically and its pathogenicity was established. Genotyping through multiple-locus variable-number tandem repeat analysis showed that the strain was a new genotype related to the L. interrogans serogroup Icterohaemorrhagiae. PMID:23307384

  20. First isolation of Leptospira interrogans from Lycalopex griseus (South American gray fox) in Argentina shows new MLVA genotype.

    PubMed

    Scialfa, Exequiel; Brihuega, Bibiana; Venzano, Agustín; Morris, Winston Eduardo; Bolpe, Jorge; Schettino, Mateo

    2013-01-01

    To identify carriers of Leptospira spp. in Argentina, wild animals were trapped in Buenos Aires Province during three nights, capturing 12 Didelphis albiventris (white-eared opossum), six Chaetophractus villosus (big hairy armadillo), five Lycalopex griseus (South American gray fox), and two Conepatus chinga (Molina's hog-nosed skunk). All were tested by microscopic agglutination test, and five (two gray foxes, two armadillos, and one skunk) were positive for Leptospira interrogans serovars Canicola and Icterohaemorrhagiae, L. borgpetersenii serovar Castellonis, and L. kirschneri serovar Grippotyphosa, at titers of 1:50 and 1:100. Kidney tissue from all animals was cultured, and one isolate of L. interrogans from a gray fox was obtained. Hamsters inoculated with the isolate died after 6 days with no macroscopic lesions at necropsy. However, histologic examination revealed glomerulonephritis, interstitial nephritis, and pneumonia. The Leptospira strain from the South American gray fox was analyzed serologically and its pathogenicity was established. Genotyping through multiple-locus variable-number tandem repeat analysis showed that the strain was a new genotype related to the L. interrogans serogroup Icterohaemorrhagiae.

  1. Multiple-locus variable-number tandem repeat analysis and clinical characterization of Leptospira interrogans canine isolates.

    PubMed

    Koizumi, Nobuo; Muto, Maki Mizutani; Izumiya, Hidemasa; Suzuki, Motoi; Ohnishi, Makoto

    2015-03-01

    Canine leptospirosis occurs worldwide; however, information on the relationship between Leptospira serotypes/genotypes and virulence in dogs remains limited. We investigated the molecular characteristics of Leptospira interrogans canine isolates belonging to three serogroups using multiple-locus variable-number tandem repeat analysis (MLVA) and the effects of each serotype/genotype on the clinical characteristics of leptospirosis in dogs. MLVA using 11 loci of the three major L. interrogans serogroups in Japan, Australis (32 strains from 21 dogs), Autumnalis (12; 7) and Hebdomadis (66; 39), revealed more divergent genetic heterogeneity within each serogroup than multilocus sequence typing (MLST), and they formed two, three and five clusters (CLs), respectively. Lethal infections were caused by all Leptospira serogroup isolates (70.3 % with Hebdomadis, 83.3 % with Australis and 100 % with Autumnalis) or Leptospira isolates belonging to all the CLs (57.1-100 %) without any significant differences. A significant difference in hyperaemia and haemorrhage of mucus membrane was observed between serogroups Australis and Autumnalis (P = 0.03). Leptospira isolates of Australis CL2 caused no hyperaemia and haemorrhage from mucus membrane, whereas those of Australis CL1, Autumnalis CL3 and Hebdomadis CL1 and CL3 did (P<0.05). Significant differences in creatinine (Cre) levels were observed between serogroups Australis and Hebdomadis (P = 0.02). In addition, significant differences in blood urea nitrogen levels were observed between serogroups Australis and Hebdomadis (P = 0.004) and Australis and Autumnalis (P = 0.02). Based on MLVA types, a significant difference in Cre levels was observed between Hebdomadis CL1 and CL4 (P = 0.0018). Our results indicated that MLVA had a higher discriminatory power and was more concordant with serotyping than MLST. Although all Leptospira serotypes and genotypes caused lethal infections in dogs, the L. interrogans

  2. Protective Immunity and Reduced Renal Colonization Induced by Vaccines Containing Recombinant Leptospira interrogans Outer Membrane Proteins and Flagellin Adjuvant.

    PubMed

    Monaris, D; Sbrogio-Almeida, M E; Dib, C C; Canhamero, T A; Souza, G O; Vasconcellos, S A; Ferreira, L C S; Abreu, P A E

    2015-08-01

    Leptospirosis is a global zoonotic disease caused by different Leptospira species, such as Leptospira interrogans, that colonize the renal tubules of wild and domestic animals. Thus far, attempts to develop effective leptospirosis vaccines, both for humans and animals, have failed to induce immune responses capable of conferring protection and simultaneously preventing renal colonization. In this study, we evaluated the protective immunity induced by subunit vaccines containing seven different recombinant Leptospira interrogans outer membrane proteins, including the carboxy-terminal portion of the immunoglobulinlike protein A (LigA(C)) and six novel antigens, combined with aluminum hydroxide (alum) or Salmonella flagellin (FliC) as adjuvants. Hamsters vaccinated with the different formulations elicited high antigen-specific antibody titers. Immunization with LigA(C), either with alum or flagellin, conferred protective immunity but did not prevent renal colonization. Similarly, animals immunized with LigA(C) or LigA(C) coadministered with six leptospiral proteins with alum adjuvant conferred protection but did not reduce renal colonization. In contrast, immunizing animals with the pool of seven antigens in combination with flagellin conferred protection and significantly reduced renal colonization by the pathogen. The present study emphasizes the relevance of antigen composition and added adjuvant in the efficacy of antileptospirosis subunit vaccines and shows the complex relationship between immune responses and renal colonization by the pathogen. PMID:26108285

  3. Protective Immunity and Reduced Renal Colonization Induced by Vaccines Containing Recombinant Leptospira interrogans Outer Membrane Proteins and Flagellin Adjuvant

    PubMed Central

    Monaris, D.; Sbrogio-Almeida, M. E.; Dib, C. C.; Canhamero, T. A.; Souza, G. O.; Vasconcellos, S. A.; Ferreira, L. C. S.

    2015-01-01

    Leptospirosis is a global zoonotic disease caused by different Leptospira species, such as Leptospira interrogans, that colonize the renal tubules of wild and domestic animals. Thus far, attempts to develop effective leptospirosis vaccines, both for humans and animals, have failed to induce immune responses capable of conferring protection and simultaneously preventing renal colonization. In this study, we evaluated the protective immunity induced by subunit vaccines containing seven different recombinant Leptospira interrogans outer membrane proteins, including the carboxy-terminal portion of the immunoglobulinlike protein A (LigAC) and six novel antigens, combined with aluminum hydroxide (alum) or Salmonella flagellin (FliC) as adjuvants. Hamsters vaccinated with the different formulations elicited high antigen-specific antibody titers. Immunization with LigAC, either with alum or flagellin, conferred protective immunity but did not prevent renal colonization. Similarly, animals immunized with LigAC or LigAC coadministered with six leptospiral proteins with alum adjuvant conferred protection but did not reduce renal colonization. In contrast, immunizing animals with the pool of seven antigens in combination with flagellin conferred protection and significantly reduced renal colonization by the pathogen. The present study emphasizes the relevance of antigen composition and added adjuvant in the efficacy of antileptospirosis subunit vaccines and shows the complex relationship between immune responses and renal colonization by the pathogen. PMID:26108285

  4. Production and characterization of monoclonal antibodies specific for Leptospira borgpetersenii serovar hardjo type hardjobovis and Leptospira interrogans serovar hardjo type hardjoprajitno.

    PubMed Central

    Surujballi, O; Howlett, C; Henning, D

    1999-01-01

    Murine monoclonal antibodies were produced by immunizing BALB/c mice with a killed whole-cell antigen prepared from Leptospira borgpetersenii serovar hardjo type hardjobovis. Six of these antibodies recognized epitopes on the homologous antigen and on whole-cell antigen prepared from Leptospira interrogans serovar hardjo type hardjoprajitno. These antibodies did not cross-react with whole-cell antigens prepared from L. borgpetersenii serovar sejroe, 10 other pathogenic Leptospira serovars, or the saprophytic Leptospira biflexa serovar patoc. Three other monoclonal antibodies reacted with antigens prepared from the 2 hardjo serovars and serovar sejroe but not with antigens from the 10 other pathogenic serovars, or serovar patoc. The epitopes recognized by all of the hardjo-specific antibodies and 2 of the 3 hardjo/sejroe-specific antibodies were susceptible to sodium meta-periodate oxidation. All of the antibodies were characterized by Western blots with the hardjobovis whole-cell antigen. Each of the 9 monoclonal antibodies was inhibited from binding to the hardjobovis antigen by bovine sera which were obtained from cattle experimentally infected with hardjobovis and from field cattle, with anti-serovar hardjo microscopic agglutination test antibody titres ranging from 100 to 12800. Some of these antibodies may be suitable for incorporation into competitive enzyme immunoassays for the specific detection of antibodies to either of the hardjo serovars. Images Figure 1. PMID:9918336

  5. Human leptospirosis cases and the prevalence of rats harbouring Leptospira interrogans in urban areas of Tokyo, Japan.

    PubMed

    Koizumi, Nobuo; Muto, Maki; Tanikawa, Tsutomu; Mizutani, Hiroshi; Sohmura, Yoshiko; Hayashi, Eiji; Akao, Nobuaki; Hoshino, Mayu; Kawabata, Hiroki; Watanabe, Haruo

    2009-09-01

    Thirteen patients with leptospirosis were identified, as confirmed by laboratory analysis during the last 5 years in our laboratory, who came from urban areas of Tokyo, Japan. All of the patients came into contact with rats before the onset of illness. Seventeen per cent of Norway rats captured in the inner cities of Tokyo carried leptospires in their kidneys. Most of these rat isolates were Leptospira interrogans serovar Copenhageni/Icterohaemorrhagiae. Antibodies against these serovars and their DNA were detected in the patients. This suggests that rats are important reservoirs of leptospirosis, and that rat-borne leptospires occur in urban areas of Tokyo.

  6. Variable Nucleotide Tandem-Repeat Analysis Revealing a Unique Group of Leptospira interrogans Serovar Pomona Isolates Associated with California Sea Lions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospira interrogans serovar Pomona is commonly isolated from a variety of wildlife and domesticated livestock. It is difficult to assess whether disease outbreaks with serovar Pomona in given animal populations are due to endemic infections or accidental exposure. Unlike many leptospiral serovars...

  7. Management practices as risk factors for the presence of bulk milk antibodies to Salmonella, Neospora caninum and Leptospira interrogans serovar hardjo in Irish dairy herds.

    PubMed

    O' Doherty, E; Berry, D P; O' Grady, L; Sayers, R

    2014-06-01

    A survey of management practices in 309 Irish dairy herds was used to identify risk factors for the presence of antibodies to Salmonella, Neospora caninum and Leptospira interrogans serovar hardjo in extensively managed unvaccinated dairy herds. A previous study documented a herd-level seroprevalence in bulk milk of 49%, 19% and 86% for Salmonella, Neospora caninum and leptospira interrogans serovar hardjo, respectively in the unvaccinated proportion of these 309 herds in 2009. Association analyses in the present study were carried out using multiple logistic regression models. Herds where cattle were purchased or introduced had a greater likelihood of being positive to leptospira interrogans serovar hardjo (P<0.01) and Salmonella (P<0.01). Larger herds had a greater likelihood of recording a positive bulk milk antibody result to leptospira interrogans serovar hardjo (P<0.05). Herds that practiced year round calving were more likely to be positive to Neospora caninum (P<0.05) compared to herds with a spring-calving season, with no difference in risk between herds that practiced split calving compared to herds that practiced spring calving. No association was found between presence of dogs on farms and prevalence of Neospora caninum possibly due to limited access of dogs to infected materials including afterbirths. The information from this study will assist in the design of suitable control programmes for the diseases under investigation in pasture-based livestock systems. PMID:24661904

  8. Leptospira interrogans at the human-wildlife interface in northern Botswana: a newly identified public health threat.

    PubMed

    Jobbins, S E; Sanderson, C E; Alexander, K A

    2014-03-01

    Leptospirosis is the most widespread zoonosis in the world. In northern Botswana, humans live in close proximity to a diversity of wildlife and peridomestic rodents and may be exposed to a variety of zoonotic pathogens. Little is known regarding the occurrence and epidemiology of L. interrogans in Africa despite the recognized global importance of this zoonotic disease and the threat it poses to public health. In Botswana, banded mongooses (Mungos mungo) live in close proximity to humans across protected and unprotected landscapes and may be a useful sentinel species for assessing the occurrence of zoonotic organisms, such as L. interrogans. We utilized PCR to screen banded mongoose kidneys for leptospiral DNA and identified 41.5% prevalence of renal carriage of L. interrogans (exact binomial 95% CI 27.7-56.7%, n = 41). Renal carriage was also detected in one Selous' mongoose (Paracynictis selousi). This is the first published confirmation of carriage of L. interrogans in either species. This is also the first report of L. interrogans occurrence in northern Botswana and the only report of this organism in a wildlife host in the country. Pathogenic Leptospira are usually transmitted indirectly to humans through soil or water contaminated with infected urine. Other avenues, such as direct contact between humans and wildlife, as well as consumption of mongooses and other wildlife as bushmeat, may pose additional exposure risk and must be considered in public health management of this newly identified zoonotic disease threat. There is a critical need to characterize host species involvement and pathogen transmission dynamics, including human-wildlife interactions that may increase human exposure potential and infection risk. We recommend that public health strategy be modified to include sensitization of medical practitioners to the presence of L. interrogans in the region, the potential for human infection, and implementation of clinical screening. This study

  9. Leptospira interrogans at the human-wildlife interface in northern Botswana: a newly identified public health threat.

    PubMed

    Jobbins, S E; Sanderson, C E; Alexander, K A

    2014-03-01

    Leptospirosis is the most widespread zoonosis in the world. In northern Botswana, humans live in close proximity to a diversity of wildlife and peridomestic rodents and may be exposed to a variety of zoonotic pathogens. Little is known regarding the occurrence and epidemiology of L. interrogans in Africa despite the recognized global importance of this zoonotic disease and the threat it poses to public health. In Botswana, banded mongooses (Mungos mungo) live in close proximity to humans across protected and unprotected landscapes and may be a useful sentinel species for assessing the occurrence of zoonotic organisms, such as L. interrogans. We utilized PCR to screen banded mongoose kidneys for leptospiral DNA and identified 41.5% prevalence of renal carriage of L. interrogans (exact binomial 95% CI 27.7-56.7%, n = 41). Renal carriage was also detected in one Selous' mongoose (Paracynictis selousi). This is the first published confirmation of carriage of L. interrogans in either species. This is also the first report of L. interrogans occurrence in northern Botswana and the only report of this organism in a wildlife host in the country. Pathogenic Leptospira are usually transmitted indirectly to humans through soil or water contaminated with infected urine. Other avenues, such as direct contact between humans and wildlife, as well as consumption of mongooses and other wildlife as bushmeat, may pose additional exposure risk and must be considered in public health management of this newly identified zoonotic disease threat. There is a critical need to characterize host species involvement and pathogen transmission dynamics, including human-wildlife interactions that may increase human exposure potential and infection risk. We recommend that public health strategy be modified to include sensitization of medical practitioners to the presence of L. interrogans in the region, the potential for human infection, and implementation of clinical screening. This study

  10. Profiling of Leptospira interrogans, L. santarosai, L. meyeri and L. borgpetersenii by SE-AFLP, PFGE and susceptibility testing--a continuous attempt at species and serovar differentiation.

    PubMed

    Moreno, Luisa Z; Miraglia, Fabiana; Lilenbaum, Walter; Neto, José S F; Freitas, Julio C; Morais, Zenaide M; Hartskeerl, Rudy A; da Costa, Barbara L P; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-01-01

    Leptospirosis is a widespread systemic zoonosis, considered as reemerging in certain developing countries. Although the cross agglutinin absorption test is still considered the standard method for Leptospira identification, it presents several disadvantages. The aim of this study was to characterize Leptospira spp. isolated from various hosts by genotyping and broth microdilution susceptibility testing in an attempt to differentiate Leptospira species, serogroups and serovars. Forty-seven isolates were studied. They were previously serotyped, and species confirmation was performed by 16S rRNA sequencing. Single-enzyme amplified fragment length polymorphism (SE-AFLP) and pulsed-field gel electrophoresis (PFGE) analysis enabled the distinction of L. interrogans from L. santarosai, L. meyeri and L. borgpetersenii in two main clusters. Among L. interrogans, it was possible to differentiate into two new clusters the serogroup Icterohaemorrhagiae from the serogroups Canicola and Pomona. L. santarosai isolates presented higher genetic variation than the other species in both techniques. Interestingly, the minimum inhibitory concentration (MIC) cluster analysis also provided Leptospira serogroup differentiation. Further studies are necessary regarding serovar Bananal isolates, as they presented the highest MIC values for most of the antimicrobials tested. All studied techniques successfully distinguished Leptospira species and serogroups. Despite being library-dependent methods, these approaches are less labor intensive and more economically viable, particularly SE-AFLP, and can be implemented in most reference laboratories worldwide to enable faster Leptospira typing. PMID:26956446

  11. Profiling of Leptospira interrogans, L. santarosai, L. meyeri and L. borgpetersenii by SE-AFLP, PFGE and susceptibility testing—a continuous attempt at species and serovar differentiation

    PubMed Central

    Moreno, Luisa Z; Miraglia, Fabiana; Lilenbaum, Walter; Neto, José SF; Freitas, Julio C; Morais, Zenaide M; Hartskeerl, Rudy A; da Costa, Barbara LP; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-01-01

    Leptospirosis is a widespread systemic zoonosis, considered as reemerging in certain developing countries. Although the cross agglutinin absorption test is still considered the standard method for Leptospira identification, it presents several disadvantages. The aim of this study was to characterize Leptospira spp. isolated from various hosts by genotyping and broth microdilution susceptibility testing in an attempt to differentiate Leptospira species, serogroups and serovars. Forty-seven isolates were studied. They were previously serotyped, and species confirmation was performed by 16S rRNA sequencing. Single-enzyme amplified fragment length polymorphism (SE-AFLP) and pulsed-field gel electrophoresis (PFGE) analysis enabled the distinction of L. interrogans from L. santarosai, L. meyeri and L. borgpetersenii in two main clusters. Among L. interrogans, it was possible to differentiate into two new clusters the serogroup Icterohaemorrhagiae from the serogroups Canicola and Pomona. L. santarosai isolates presented higher genetic variation than the other species in both techniques. Interestingly, the minimum inhibitory concentration (MIC) cluster analysis also provided Leptospira serogroup differentiation. Further studies are necessary regarding serovar Bananal isolates, as they presented the highest MIC values for most of the antimicrobials tested. All studied techniques successfully distinguished Leptospira species and serogroups. Despite being library-dependent methods, these approaches are less labor intensive and more economically viable, particularly SE-AFLP, and can be implemented in most reference laboratories worldwide to enable faster Leptospira typing. PMID:26956446

  12. Expression and characterization of recombinant leptospiral outer membrane protein LipL32 from Leptospira interrogans serovar autumnalis.

    PubMed

    Boonsathorn, Naphatsawan; Konghom, Ganokrot; Mongkolsiri, Kaveewan; Jirapongwattana, Chanin; Balachandra, Kruavon; Naigowit, Pimjai; Sawanpanyalert, Pathom

    2009-01-01

    Leptospira interrogans serovar autumnalis, a causative agent of leptospirosis in Thailand, was isolated from a patient for DNA extraction and amplification of LipL32 gene by polymerase chain reaction (PCR). The 782 bp PCR product was obtained, which was inserted into pAE plasmid with polyhistidine (His6 tag) to construct pAE-LipL32. This recombinant plasmid was transfected into E. coli BL21 (DE3). His6-LipL32 was purified by Ni-NTA affinity chromatography. The recombinant protein was used as antigen for testing with sera from leptospirosis and syphilis patients by dot-ELISA technique. It reacted positively with leptospirosis patient sera and negatively with syphilis and healthy sera.

  13. Seasonal prevalence of antibodies to Leptospira interrogans in Antillean manatees from a landlocked lake in Tabasco, Mexico.

    PubMed

    Aragón-Martínez, Arianna; Olivera-Gómez, León D; Jiménez-Domínguez, Darwin

    2014-07-01

    Factors that alter the dynamics of ecologic systems can influence transmission of infectious diseases and may lead to decreases in natural populations. Leptospirosis is a cosmopolitan disease of zoonotic importance that affects most mammals. At the southern Gulf of Mexico, Antillean manatees (Trichechus manatus manatus) inhabit highly variable environments, with extended floods during the rainy season and drought conditions during the dry season that affect food availability and the thermal environment for manatees. We tested for changes in prevalence and titers of antibodies to 12 serovars of Leptospira interrogans in manatees between dry and rainy seasons. We determined titers for L. interrogans through microscopic agglutination tests (MAT) from 10 manatees, six during the dry season (DS), and six during the rainy season (RS) in Laguna de las Ilusiones, a landlocked lake hosting a population of about 20 manatees. All individuals were antibody positive (titers ≥ 100) to at least one serovar. The serovars bataviae, bratislava, canicola, and icterohaemorrhagiae had overall prevalences ≥ 50%; bataviae, bratislava, and canicola had prevalences ≥ 50% during both seasons. Serovars icterohaemorrhagiae and pyrogenes had prevalences ≥ 50% during DS and pomona, tarassovi, wolfii, and autumnalis during RS. Significant differences in prevalence between seasons were found for pomona, tarassovi, and autumnalis. Titers of tarassovi, wolfii, autumnalis, and bataviae were significantly higher during RS. There was a high prevalence of L. interrogans during the RS independent of high availability of plant foods, coinciding with the epizootiology of the bacteria that are endemic to tropical regions. Another factor possibly influencing prevalence is high anthropogenic pressure at the lake, causing an increase in potential sources of infection. Because of possible cross-reaction in MAT, further research is needed on the molecular discrimination of serovars in animals in the

  14. Seasonal prevalence of antibodies to Leptospira interrogans in Antillean manatees from a landlocked lake in Tabasco, Mexico.

    PubMed

    Aragón-Martínez, Arianna; Olivera-Gómez, León D; Jiménez-Domínguez, Darwin

    2014-07-01

    Factors that alter the dynamics of ecologic systems can influence transmission of infectious diseases and may lead to decreases in natural populations. Leptospirosis is a cosmopolitan disease of zoonotic importance that affects most mammals. At the southern Gulf of Mexico, Antillean manatees (Trichechus manatus manatus) inhabit highly variable environments, with extended floods during the rainy season and drought conditions during the dry season that affect food availability and the thermal environment for manatees. We tested for changes in prevalence and titers of antibodies to 12 serovars of Leptospira interrogans in manatees between dry and rainy seasons. We determined titers for L. interrogans through microscopic agglutination tests (MAT) from 10 manatees, six during the dry season (DS), and six during the rainy season (RS) in Laguna de las Ilusiones, a landlocked lake hosting a population of about 20 manatees. All individuals were antibody positive (titers ≥ 100) to at least one serovar. The serovars bataviae, bratislava, canicola, and icterohaemorrhagiae had overall prevalences ≥ 50%; bataviae, bratislava, and canicola had prevalences ≥ 50% during both seasons. Serovars icterohaemorrhagiae and pyrogenes had prevalences ≥ 50% during DS and pomona, tarassovi, wolfii, and autumnalis during RS. Significant differences in prevalence between seasons were found for pomona, tarassovi, and autumnalis. Titers of tarassovi, wolfii, autumnalis, and bataviae were significantly higher during RS. There was a high prevalence of L. interrogans during the RS independent of high availability of plant foods, coinciding with the epizootiology of the bacteria that are endemic to tropical regions. Another factor possibly influencing prevalence is high anthropogenic pressure at the lake, causing an increase in potential sources of infection. Because of possible cross-reaction in MAT, further research is needed on the molecular discrimination of serovars in animals in the

  15. Real-Time PCR Reveals Rapid Dissemination of Leptospira interrogans after Intraperitoneal and Conjunctival Inoculation of Hamsters.

    PubMed

    Wunder, Elsio A; Figueira, Claudio P; Santos, Gisele R; Lourdault, Kristel; Matthias, Michael A; Vinetz, Joseph M; Ramos, Eduardo; Haake, David A; Picardeau, Mathieu; Dos Reis, Mitermayer G; Ko, Albert I

    2016-07-01

    The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant infections in humans and other accidental hosts. Hematogenous dissemination is important for infection by the pathogen but remains poorly understood because few animal model studies have used sensitive tools to quantify the bacteria. We evaluated the kinetics of leptospiral infection in Golden Syrian hamsters by a sensitive quantitative real-time PCR (TaqMan) with lipl32 as the target gene. The dissemination and bacterial burden were measured after intraperitoneal infection with a high dose (10(8)) or low dose (2.5 × 10(2)) of leptospires. We also examined the conjunctival challenge route to mimic the natural history of infection. Quantification of leptospires in perfused animals revealed that pathogens were detected in all organs of intraperitoneally infected hamsters, including the eye and brain, within 1 h after inoculation of 10(8) virulent L. interrogans bacteria. Peaks of 10(5) to 10(8) leptospires per gram or per milliliter were achieved in blood and all tissues between day 4 and day 8 after intraperitoneal inoculation of high- and low-dose challenges, respectively, coinciding with macroscopic and histological changes. The conjunctival route resulted in a delay in the time to peak organ burden in comparison to intraperitoneal infection, indicating that although infection could be established, penetration efficiency was low across this epithelial barrier. Surprisingly, infection with a large inoculum of high-passage-number attenuated L. interrogans strains resulted in dissemination to all organs in the first 4 days postinfection, albeit with a lower burden, followed by clearance from the blood and organs 7 days postinfection and survival of all animals. These results demonstrate that leptospiral dissemination and tissue invasion occur. In contrast, development of a critical level of tissue burden and pathology are dependent on the virulence of the infecting

  16. Response of Leptospira interrogans to Physiologic Osmolarity: Relevance in Signaling the Environment-to-Host Transition

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transmission of pathogenic Leptospira between mammalian hosts usually involves dissemination via soil or water contaminated by the urine of reservoir hosts. The ability of Leptospira to adapt to the variety of conditions found inside and outside of the host is reflected in the relatively large geno...

  17. Post-translational modification of LipL32 during Leptospira interrogans infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospirosis, a re-emerging disease of global importance caused by pathogenic Leptospira spp., is considered the world’s most widespread zoonotic disease. Rats serve as asymptomatic carriers of pathogenic Leptospira and are critical for disease spread. In such reservoir hosts, leptospires colonize ...

  18. Analysis of LexA binding sites and transcriptomics in response to genotoxic stress in Leptospira interrogans

    PubMed Central

    Schons-Fonseca, Luciane; da Silva, Josefa B.; Milanez, Juliana S.; Domingos, Renan H.; Smith, Janet L.; Nakaya, Helder I.; Grossman, Alan D.; Ho, Paulo L.; da Costa, Renata MA

    2016-01-01

    We determined the effects of DNA damage caused by ultraviolet radiation on gene expression in Leptospira interrogans using DNA microarrays. These data were integrated with DNA binding in vivo of LexA1, a regulator of the DNA damage response, assessed by chromatin immunoprecipitation and massively parallel DNA sequencing (ChIP-seq). In response to DNA damage, Leptospira induced expression of genes involved in DNA metabolism, in mobile genetic elements and defective prophages. The DNA repair genes involved in removal of photo-damage (e.g. nucleotide excision repair uvrABC, recombinases recBCD and resolvases ruvABC) were not induced. Genes involved in various metabolic pathways were down regulated, including genes involved in cell growth, RNA metabolism and the tricarboxylic acid cycle. From ChIP-seq data, we observed 24 LexA1 binding sites located throughout chromosome 1 and one binding site in chromosome 2. Expression of many, but not all, genes near those sites was increased following DNA damage. Binding sites were found as far as 550 bp upstream from the start codon, or 1 kb into the coding sequence. Our findings indicate that there is a shift in gene expression following DNA damage that represses genes involved in cell growth and virulence, and induces genes involved in mutagenesis and recombination. PMID:26762976

  19. Analysis of LexA binding sites and transcriptomics in response to genotoxic stress in Leptospira interrogans.

    PubMed

    Schons-Fonseca, Luciane; da Silva, Josefa B; Milanez, Juliana S; Domingos, Renan H; Smith, Janet L; Nakaya, Helder I; Grossman, Alan D; Ho, Paulo L; da Costa, Renata M A

    2016-02-18

    We determined the effects of DNA damage caused by ultraviolet radiation on gene expression in Leptospira interrogans using DNA microarrays. These data were integrated with DNA binding in vivo of LexA1, a regulator of the DNA damage response, assessed by chromatin immunoprecipitation and massively parallel DNA sequencing (ChIP-seq). In response to DNA damage, Leptospira induced expression of genes involved in DNA metabolism, in mobile genetic elements and defective prophages. The DNA repair genes involved in removal of photo-damage (e.g. nucleotide excision repair uvrABC, recombinases recBCD and resolvases ruvABC) were not induced. Genes involved in various metabolic pathways were down regulated, including genes involved in cell growth, RNA metabolism and the tricarboxylic acid cycle. From ChIP-seq data, we observed 24 LexA1 binding sites located throughout chromosome 1 and one binding site in chromosome 2. Expression of many, but not all, genes near those sites was increased following DNA damage. Binding sites were found as far as 550 bp upstream from the start codon, or 1 kb into the coding sequence. Our findings indicate that there is a shift in gene expression following DNA damage that represses genes involved in cell growth and virulence, and induces genes involved in mutagenesis and recombination. PMID:26762976

  20. Leptospira interrogans survey by PCR in wild rodents coming from different urban areas of Palermo, Italy.

    PubMed

    Vitale, Maria; Di Bella, Carobelo; Agnello, Stefano; Curro, Victoria; Vicari, Domenico; Vitale, Fabrizio

    2007-01-01

    DNA extracted from the kidneys of rodents captured in different urban areas of Palermo, Italy, had been analysed for the presence of pathogenic L. interrogans sensu latu DNA. PCR analysis had shown that in rodents captured close to green areas and small river up to 40 % animals give positive PCR results. Not many cases of human leptospirosis are reported in Sicilian island in which hot season is usually dry. But considering climate change toward subtropical aspect in Sicily, with hot humid summer and sudden thunderstorm, screening for L. interrogans sensu latu prevalence can be useful for leptospirosis risk analysis on human population. PMID:23427420

  1. Leptospira interrogans survey by PCR in wild rodents coming from different urban areas of Palermo, Italy.

    PubMed

    Vitale, Maria; Di Bella, Carobelo; Agnello, Stefano; Curro, Victoria; Vicari, Domenico; Vitale, Fabrizio

    2007-01-01

    DNA extracted from the kidneys of rodents captured in different urban areas of Palermo, Italy, had been analysed for the presence of pathogenic L. interrogans sensu latu DNA. PCR analysis had shown that in rodents captured close to green areas and small river up to 40 % animals give positive PCR results. Not many cases of human leptospirosis are reported in Sicilian island in which hot season is usually dry. But considering climate change toward subtropical aspect in Sicily, with hot humid summer and sudden thunderstorm, screening for L. interrogans sensu latu prevalence can be useful for leptospirosis risk analysis on human population.

  2. Development and initial evaluation of an indirect enzyme-linked immunosorbent assay for the detection of Leptospira interrogans serovar hardjo antibodies in bovine sera.

    PubMed Central

    Surujballi, O P; Marenger, R M; Eaglesome, M D; Sugden, E A

    1997-01-01

    Outer sheath antigen from Leptospira interrogans serovar hardjo type hardjoprajitno and acetic acid extracted antigens from serovar hardjo types hardjoprajitno and hardjobovis were evaluated in an immunoassay for ability to detect hyperimmune rabbit serum to serovar hardjo. The degree of cross-reactivity with hyperimmune rabbit sera to L. interrogans serovars pomona, copenhageni, grippotyphosa, canicola and sejroe, and Leptospira biflexa serovar patoc was also measured for each antigen. All of the antigens reacted with the antiserum to L. interrogans serovar hardjo. The outer sheath antigen however, also showed wide cross-reactivity with the antisera to all of the serovars of L. interrogans tested and with the antiserum to L. biflexa serovar patoc. The acetic acid extracted antigen from either type hardjoprajitno, or type hardjobovis, showed a high degree of specificity for serovar hardjo antiserum. The hardjobovis acetic acid extracted antigen was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting, and was incorporated into an indirect ELISA for detection of anti-serovar hardjo antibodies in bovine serum. This ELISA showed a relative specificity of 100% with 156 bovine sera which were negative at a dilution of 1:100 in the microscopic agglutination test (MAT) for L. interrogans serovars hardjo, pomona, sejroe, icterohaemorrhagiae, copenhageni, canicola, and grippotyphosa. The relative sensitivity of this assay with 192 bovine sera which had serovar hardjo MAT titres of > or = 100 was 95.3% (95% confidence limit = 2.99%). The degree of cross-reactivity with 289 bovine sera which had serovar pomona MAT titres of > or = 100 (with no detectable serovar hardjo MAT titres) was approximately 1.0%. This assay was: easily standardized, scored objectively, repeatable, semi-automated and used a non-hazardous antigen that can be routinely prepared in gram amounts. Images Figure 2. PMID:9342449

  3. The multifunctional LigB adhesin binds homeostatic proteins with potential roles in cutaneous infection by pathogenic Leptospira interrogans.

    PubMed

    Choy, Henry A; Kelley, Melissa M; Croda, Julio; Matsunaga, James; Babbitt, Jane T; Ko, Albert I; Picardeau, Mathieu; Haake, David A

    2011-01-01

    Leptospirosis is a potentially fatal zoonotic disease in humans and animals caused by pathogenic spirochetes, such as Leptospira interrogans. The mode of transmission is commonly limited to the exposure of mucous membrane or damaged skin to water contaminated by leptospires shed in the urine of carriers, such as rats. Infection occurs during seasonal flooding of impoverished tropical urban habitats with large rat populations, but also during recreational activity in open water, suggesting it is very efficient. LigA and LigB are surface localized proteins in pathogenic Leptospira strains with properties that could facilitate the infection of damaged skin. Their expression is rapidly induced by the increase in osmolarity encountered by leptospires upon transition from water to host. In addition, the immunoglobulin-like repeats of the Lig proteins bind proteins that mediate attachment to host tissue, such as fibronectin, fibrinogen, collagens, laminin, and elastin, some of which are important in cutaneous wound healing and repair. Hemostasis is critical in a fresh injury, where fibrinogen from damaged vasculature mediates coagulation. We show that fibrinogen binding by recombinant LigB inhibits fibrin formation, which could aid leptospiral entry into the circulation, dissemination, and further infection by impairing healing. LigB also binds fibroblast fibronectin and type III collagen, two proteins prevalent in wound repair, thus potentially enhancing leptospiral adhesion to skin openings. LigA or LigB expression by transformation of a nonpathogenic saprophyte, L. biflexa, enhances bacterial adhesion to fibrinogen. Our results suggest that by binding homeostatic proteins found in cutaneous wounds, LigB could facilitate leptospirosis transmission. Both fibronectin and fibrinogen binding have been mapped to an overlapping domain in LigB comprising repeats 9-11, with repeat 11 possibly enhancing binding by a conformational effect. Leptospirosis patient antibodies react

  4. Characterization of Leptospira interrogans Serovars by Polymorphism Variable Number Tandem Repeat Analysis

    PubMed Central

    Rezasoltani, Sama; Dabiri, Hossein; Khaki, Pejvak; Rostami Nejad, Mohammad; Karimnasab, Nasim; Modirrousta, Shiva

    2015-01-01

    Background: Leptospirosis is recognized as a re-emerging infectious disease; therefore, understanding the epidemiology of the disease is vital for designing intervention programs and diminishing its transmission. Recently, Multilocus variable number tandem repeat analysis (MLVA) is used for segregating and identifying Leptospira serovars. The method has potential application in investigating the molecular epidemiology of Leptospira. Objectives: The propose of this study was genomic identification of pathogenic Leptospires in Iran by MLVA. Materials and Methods: Leptospira serovars were obtained from National Reference Laboratory of Leptospira at Razi Vaccine and Serum Research Institute, Karaj, Iran. Serovars were cultured into the liquid EMJH medium and incubated at 28˚C for 7 days. DNA of serovars was extracted using the phenol-chloroform method. PCR was performed with 5 selected variable number tandem repeat analysis (VNTR) loci. The amplified products were analyzed by agarose gel electrophoresis. The size of the amplified products was estimated by 100 bp ladder and sequencing analysis. Results: The saprophytic serovar showed no amplified fragments. PCR products in all pathogenic serovars were observed. The 12 reference serovars used for the development of technique displayed distinct patterns. Conclusions: Results showed that MLVA technique with its range of polymorphism is a good marker for identification of pathogenic serovars. Some VNTR loci are more powerful than the other ones with regard to differentiation. Serovars from the same geographical area have more genetic similarity than same serovars from different places. MLVA is a suitable technique for epidemiological survey. PMID:26568805

  5. Asymptomatic and chronic carriage of Leptospira interrogans serovar Pomona in California sea lions (Zalophus californianus)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since 1970, periodic outbreaks of leptospirosis, caused by pathogenic spirochetes in the genus Leptospira, have caused morbidity and mortality of California sea lions (Zalophus californianus) along the Pacific coast of North America. Yearly seasonal epizootics of varying magnitude occur between the ...

  6. Comparative genomic analysis of eight Leptospira strains from Japan and the Philippines revealing the existence of four putative novel genomic islands/islets in L. interrogans serovar Lai strain 56601.

    PubMed

    Youn, Jung-Ho; Hayashida, Kyoko; Koizumi, Nobuo; Ohnishi, Makoto; Sugimoto, Chihiro

    2014-12-01

    Leptospirosis is one of the most widespread zoonotic diseases worldwide and can be considered an emerging health problem to both human and animal. Despite the importance of the disease, complete genome sequences are currently available for only three Leptospira interrogans strains: 56601, Fiocruz L1-130, and IPAV. Therefore, intra- and inter-species comparative genomic analyses of Leptospira are limited. Here, to advance current knowledge of the genomic differences within Leptospira species, next-generation sequencing technology was used to examine the genomes of eight L. interrogans strains belonging to six different serogroups isolated from humans and dogs in Japan and the Philippines. The genomic sequences were mapped to that of the reference strain, L. interrogans serovar Lai strain 56601. The results revealed the presence of four novel genomic islands/islets (GIs) in strain 56601. This study provides a deeper insight into the molecular basis and evolutionary perspective of the virulence of leptospires. PMID:25449997

  7. Distribution of Leptospira interrogans by Multispacer Sequence Typing in Urban Norway Rats (Rattus norvegicus): A Survey in France in 2011-2013

    PubMed Central

    Bicout, Dominique J.; Kodjo, Angeli; Artois, Marc; Djelouadji, Zoheira

    2015-01-01

    Background Urban leptospirosis has increasingly been reported in both developing and developed countries. The control of the disease is limited because our understanding of basic aspects of the epidemiology, including the transmission routes of leptospires among rat populations, remains incomplete. Through the ability to distinguish among Leptospira strains in rats, multispacer sequence typing (MST) could provide a modern understanding of Leptospira epidemiology; however, to our knowledge, the distribution of Leptospira strains among urban rat colonies has not been investigated using MST. Aims and Methodology The objective of this study was to identify the Leptospira strains present in rats (Rattus norvegicus) in Lyon (France) using MST and to characterize their spatial distribution. Kidneys and urine were collected from rats trapped live in seven locations in the city and in one suburban location. Each location was considered to represent a rat colony. Bacterial cultures and quantitative polymerase chain reaction (qPCR) assays were performed, and the L. interrogans DNA identified was then genotyped using MST. The distributions of Leptospira strains were spatially described. Key Results Among 84 wild rats, MST profiles were obtained in 35 of 37 rats that had a positive result for L. interrogans by bacterial culture and/or qPCR analyses. All of the MST profiles were related to reference strains previously isolated from human patients that belong to the serogroup Icterohaemorrhagiae and the serovars [strain(s)] Copenhageni [Wijinberg or M20] (n = 26), Icterohaemorrhagiae [CHU Réunion] (n = 7), Icterohaemorrhagiae [R1] (n = 1) and Copenhageni [Shibaura 9] (n = 1). Each colony was infected with leptospires having the same MST profile. Major Conclusions This study demonstrated that MST could be used for the purpose of field studies, either on culture isolates or on DNA extracted from kidneys and urine, to distinguish among L. interrogans isolates in rats. MST could

  8. Methylation and in vivo expression of the surface-exposed Leptospira interrogans outer-membrane protein OmpL32

    PubMed Central

    Eshghi, Azad; Pinne, Marija; Haake, David A.; Zuerner, Richard L.; Frank, Ami

    2012-01-01

    Recent studies have revealed that bacterial protein methylation is a widespread post-translational modification that is required for virulence in selected pathogenic bacteria. In particular, altered methylation of outer-membrane proteins has been shown to modulate the effectiveness of the host immune response. In this study, 2D gel electrophoresis combined with MALDI-TOF MS identified a Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 protein, corresponding to ORF LIC11848, which undergoes extensive and differential methylation of glutamic acid residues. Immunofluorescence microscopy implicated LIC11848 as a surface-exposed outer-membrane protein, prompting the designation OmpL32. Indirect immunofluorescence microscopy of golden Syrian hamster liver and kidney sections revealed expression of OmpL32 during colonization of these organs. Identification of methylated surface-exposed outer-membrane proteins, such as OmpL32, provides a foundation for delineating the role of this post-translational modification in leptospiral virulence. PMID:22174381

  9. Occurrence of antibodies anti -Toxoplasma gondii, Neospora caninum and Leptospira interrogans in a captive deer herd in Southern Brazil.

    PubMed

    Zimpel, Cristina Kraemer; Grazziotin, Ana Laura; de Barros Filho, Ivan Roque; Guimaraes, Ana Marcia de Sa; dos Santos, Leonilda Correia; de Moraes, Wanderlei; Cubas, Zalmir Silvino; de Oliveira, Marcos Jose; Pituco, Edviges Maristela; Lara, Maria do Carmo Custódio de Souza Hunold; Villalobos, Eliana Monteforte Cassaro; Silva, Lília Marcia Paulin; Cunha, Elenice Maria Sequetin; Castro, Vanessa; Biondo, Alexander Welker

    2015-01-01

    A large number of Brazilian zoos keep many endangered species of deer, however, very few disease surveillance studies have been conducted among captive cervids. Blood samples from 32 Brazilian deer (Blastocerus dichotomus, Mazama nana and Mazama americana) kept in captivity at Bela Vista Biological Sanctuary (Foz do Iguaçu, Brazil) were investigated for 10 ruminant pathogens, with the aims of monitoring deer health status and evaluating any potential zoonotic risk. Deer serum samples were tested for Brucella abortus, Leptospira (23 serovars), Toxoplasma gondii, Neospora caninum, bovine viral diarrhea virus, infectious bovine rhinotracheitis virus, foot-and-mouth disease virus, western equine encephalitis virus, eastern equine encephalitis virus and Venezuelan equine encephalitis virus. Antibodies against T. gondii (15.6%), N. caninum (6.2%) and L. interrogans serogroup Serjoe (3.1%) were detected. The serological results for all other infectious agents were negative. The deer were considered to be clinically healthy and asymptomatic regarding any disease. Compared with studies on free-ranging deer, the prevalences of the same agents tested among the captive deer kept at the Sanctuary were lower, thus indicating good sanitary conditions and high-quality management practices at the zoo. PMID:26689185

  10. Experimental infections of brush-tailed possums, common wombats and water rats with Leptospira interrogans serovars balcanica and hardjo.

    PubMed

    Durfee, P T; Presidente, P J

    1979-06-01

    Of 12 brush-tailed possums (Trichosurus vulpecula) inoculated with Leptospira interrogans serovar balcanica 11 developed migroagglutination (MA) antibody to jardjo antigen by 14 days postincubation (PI). Leptospiruria was observed in 2 possums 117 to 145 days PI. Of 6 possums inoculated with serovar hardjo 4 developed low short-lived titres by day 18 PI. Two of 3 wombats (Vombatus ursinus) inoculated with balcanica had high MA titres (greater than or equal to 1:128) by day 16 PI and leptospiruria occurred by day 16. One wombat inoculated with hardjo developed a low MA titre. Low transitory MA titres to hardjo were found in 1 of 3 water rats (Hydromys chrysogaster) after inoculation with balcanica and 1 of 2 given hardjo. Histopathological examination of kidneys revealed mild to moderately severe focal interstitial nephritis in 4 of 8 possums, in 2 wombats and in 2 water rats following experimental infection with balcanica. Similar lesions were observed in 2 of 4 possums, 1 wombat and 2 water rats following experimental infection with hardjo. PMID:533478

  11. Occurrence of antibodies anti -Toxoplasma gondii, Neospora caninum and Leptospira interrogans in a captive deer herd in Southern Brazil.

    PubMed

    Zimpel, Cristina Kraemer; Grazziotin, Ana Laura; de Barros Filho, Ivan Roque; Guimaraes, Ana Marcia de Sa; dos Santos, Leonilda Correia; de Moraes, Wanderlei; Cubas, Zalmir Silvino; de Oliveira, Marcos Jose; Pituco, Edviges Maristela; Lara, Maria do Carmo Custódio de Souza Hunold; Villalobos, Eliana Monteforte Cassaro; Silva, Lília Marcia Paulin; Cunha, Elenice Maria Sequetin; Castro, Vanessa; Biondo, Alexander Welker

    2015-01-01

    A large number of Brazilian zoos keep many endangered species of deer, however, very few disease surveillance studies have been conducted among captive cervids. Blood samples from 32 Brazilian deer (Blastocerus dichotomus, Mazama nana and Mazama americana) kept in captivity at Bela Vista Biological Sanctuary (Foz do Iguaçu, Brazil) were investigated for 10 ruminant pathogens, with the aims of monitoring deer health status and evaluating any potential zoonotic risk. Deer serum samples were tested for Brucella abortus, Leptospira (23 serovars), Toxoplasma gondii, Neospora caninum, bovine viral diarrhea virus, infectious bovine rhinotracheitis virus, foot-and-mouth disease virus, western equine encephalitis virus, eastern equine encephalitis virus and Venezuelan equine encephalitis virus. Antibodies against T. gondii (15.6%), N. caninum (6.2%) and L. interrogans serogroup Serjoe (3.1%) were detected. The serological results for all other infectious agents were negative. The deer were considered to be clinically healthy and asymptomatic regarding any disease. Compared with studies on free-ranging deer, the prevalences of the same agents tested among the captive deer kept at the Sanctuary were lower, thus indicating good sanitary conditions and high-quality management practices at the zoo.

  12. Features of Two New Proteins with OmpA-Like Domains Identified in the Genome Sequences of Leptospira interrogans

    PubMed Central

    Teixeira, Aline F.; de Morais, Zenaide M.; Kirchgatter, Karin; Romero, Eliete C.; Vasconcellos, Silvio A.; Nascimento, Ana Lucia T. O.

    2015-01-01

    Leptospirosis is an acute febrile disease caused by pathogenic spirochetes of the genus Leptospira. It is considered an important re-emerging infectious disease that affects humans worldwide. The knowledge about the mechanisms by which pathogenic leptospires invade and colonize the host remains limited since very few virulence factors contributing to the pathogenesis of the disease have been identified. Here, we report the identification and characterization of two new leptospiral proteins with OmpA-like domains. The recombinant proteins, which exhibit extracellular matrix-binding properties, are called Lsa46 - LIC13479 and Lsa77 - LIC10050 (Leptospiral surface adhesins of 46 and 77 kDa, respectively). Attachment of Lsa46 and Lsa77 to laminin was specific, dose dependent and saturable, with KD values of 24.3 ± 17.0 and 53.0 ± 17.5 nM, respectively. Lsa46 and Lsa77 also bind plasma fibronectin, and both adhesins are plasminogen (PLG)-interacting proteins, capable of generating plasmin (PLA) and as such, increase the proteolytic ability of leptospires. The proteins corresponding to Lsa46 and Lsa77 are present in virulent L. interrogans L1-130 and in saprophyte L. biflexa Patoc 1 strains, as detected by immunofluorescence. The adhesins are recognized by human leptospirosis serum samples at the onset and convalescent phases of the disease, suggesting that they are expressed during infection. Taken together, our data could offer valuable information to the understanding of leptospiral pathogenesis. PMID:25849456

  13. Experimental infections of brush-tailed possums, common wombats and water rats with Leptospira interrogans serovars balcanica and hardjo.

    PubMed

    Durfee, P T; Presidente, P J

    1979-06-01

    Of 12 brush-tailed possums (Trichosurus vulpecula) inoculated with Leptospira interrogans serovar balcanica 11 developed migroagglutination (MA) antibody to jardjo antigen by 14 days postincubation (PI). Leptospiruria was observed in 2 possums 117 to 145 days PI. Of 6 possums inoculated with serovar hardjo 4 developed low short-lived titres by day 18 PI. Two of 3 wombats (Vombatus ursinus) inoculated with balcanica had high MA titres (greater than or equal to 1:128) by day 16 PI and leptospiruria occurred by day 16. One wombat inoculated with hardjo developed a low MA titre. Low transitory MA titres to hardjo were found in 1 of 3 water rats (Hydromys chrysogaster) after inoculation with balcanica and 1 of 2 given hardjo. Histopathological examination of kidneys revealed mild to moderately severe focal interstitial nephritis in 4 of 8 possums, in 2 wombats and in 2 water rats following experimental infection with balcanica. Similar lesions were observed in 2 of 4 possums, 1 wombat and 2 water rats following experimental infection with hardjo.

  14. Risk factors associated with prevalence of antibodies to Leptospira interrogans in a metapopulation of black-tailed prairie dogs in Mexico.

    PubMed

    Montiel-Arteaga, Ana; Atilano, Daniel; Ayanegui, Alejandra; Ceballos, Gerardo; Suzán, Gerardo

    2015-01-01

    Interest in the study of infectious diseases of wildlife has grown in recent decades and now focuses on understanding host-parasite dynamics and factors involved in disease occurrence. The black-tailed prairie dog (Cynomys ludovicianus) is a useful species for this type of investigation because it lives in heterogeneous landscapes where human activities take place, and its populations are structured as a metapopulation. Our goal was to determine if colony area, density, and proximity to human settlements are associated with prevalence of antibodies to Leptospira interrogans in black-tailed prairie dogs of northwestern Chihuahua State, Mexico. We captured 266 prairie dogs in 11 colonies in 2009 and analyzed 248 serum samples with the microscopic agglutination test (MAT) for antibody to any of the 12 pathogenic serovars of L. interrogans. Serologically positive test results for only serovars Bratislava, Canicola, Celledoni, and Tarassovi were considered for statistical analysis. Almost 80% of sera were positive for at least one pathogenic serovar (MAT titer ≥1∶80). The highest recorded antibody prevalences were to serovars Bratislava and Canicola. Correlation analysis showed a negative relationship between L. interrogans antibody prevalence and colony area (r = -0.125, P<0.005), suggesting that animals living in larger colonies were at a lower risk than those in smaller colonies. The correlation between the serovar Canicola and distance was negative (r = -0.171, P<0.007), and this relationship may be explained by the presence of domestic dogs associated with human dwellings. This is the first study of Leptospira spp. antibody prevalence in prairie dogs, and it provides valuable insights into the dynamics of leptospirosis in threatened wildlife species. Further studies are needed to evaluate the impact of Leptospira serovars in metapopulations of prairie dogs and other domestic and wild mammals in grassland communities. PMID:25380365

  15. Prozone effects in microscopic agglutination tests for leptospirosis in the sera of mice infected with the pathogenic Leptospira interrogans serovar Canicola

    PubMed Central

    Shimabukuro, Fabio Hiroto; da Costa, Veruska Maia; da Silva, Rodrigo Costa; Langoni, Hélio; da Silva, Aristeu Vieira; de Carvalho, Lídia Raquel; Domingues, Paulo Francisco

    2013-01-01

    Mice experimentally infected with a pathogenic strain of Leptospira interrogans serovar Canicola produced false negative results (prozone effect) in a microscopic agglutination test (MAT). This prozone effect occurred in several serum samples collected at different post-infection times, but it was more prominent in samples collected from seven-42 days post-infection and for 1:50 and 1:100 sample dilutions. This phenomenon was correlated with increased antibody titres in the early post-infection phase. While prozone effects are often observed in serological agglutination assays for the diagnosis of animal brucellosis and human syphilis, they are not widely reported in leptospirosis MATs. PMID:23903987

  16. Interaction of Leptospira interrogans with Human Proteolytic Systems Enhances Dissemination through Endothelial Cells and Protease Levels

    PubMed Central

    Vieira, Monica L.; Alvarez-Flores, Miryam P.; Kirchgatter, Karin; Romero, Eliete C.; Alves, Ivy J.; de Morais, Zenaide M.; Vasconcellos, Silvio A.; Chudzinski-Tavassi, Ana M.

    2013-01-01

    We have recently reported the ability of Leptospira to capture plasminogen (PLG) and generate plasmin (PLA) bound on the microbial surface in the presence of exogenous activators. In this work, we examined the effects of leptospiral PLG binding for active penetration through the endothelial cell barrier and activation. The results indicate that leptospires with PLG association or PLA activation have enhanced migration activity through human umbilical vein endothelial cell (HUVEC) monolayers compared with untreated bacteria. Leptospira cells coated with PLG were capable of stimulating the expression of PLG activators by HUVECs. Moreover, leptospires endowed with PLG or PLA promoted transcriptional upregulation matrix metalloprotease 9 (MMP-9). Serum samples from patients with confirmed leptospirosis showed higher levels of PLG activators and total MMP-9 than serum samples from normal (healthy) subjects. The highest level of PLG activators and total MMP-9 was detected with microscopic agglutination test (MAT)-negative serum samples, suggesting that this proteolytic activity stimulation occurs at the early stage of the disease. Furthermore, a gelatin zymography profile obtained for MMPs with serum samples from patients with leptospirosis appears to be specific to leptospiral infection because serum samples from patients with unrelated infectious diseases produced no similar degradation bands. Altogether, the data suggest that the Leptospira-associated PLG or PLA might represent a mechanism that contributes to bacterial penetration of endothelial cells through an activation cascade of events that enhances the proteolytic capability of the organism. To our knowledge, this is the first proteolytic activity associated with leptospiral pathogenesis described to date. PMID:23478319

  17. Prevalence of Leptospira interrogans antibodies in free-ranging Tayassu pecari of the Southern Pantanal, Brazil, an ecosystem where wildlife and cattle interact.

    PubMed

    de Freitas, Tatiana P Tavares; Keuroghlian, Alexine; Eaton, Donald P; de Freitas, Emanuel Barbosa; Figueiredo, Aline; Nakazato, Luciano; de Oliveira, Jacqueline M; Miranda, Flávia; Paes, Rita Cassia S; Monteiro, Leticia A R Carneiro; Lima, José Vergílio B; da C Neto, Aparecida A; Dutra, Valéria; de Freitas, Julio Cesar

    2010-12-01

    We surveyed a wild population of white-lipped peccaries (Tayassu pecari) in the Brazilian Pantanal for evidence of Leptospira interrogans. Serum samples from 71 free-ranging T. pecari were obtained between 2003 and 2005 in the southern Pantanal of Mato Grosso do Sul state. We used microscopic microagglutination to test for antibodies against 14 L. interrogans serovars (antibody titers ≥ 1:100 were considered seropositive). Seventy percent of captured animals tested positive for leptospirosis antibodies. Antibodies against icterohaemorrhagiae and autumnalis serovars were the most prevalent. We used log-linear analyses to test for associations among seropositivity, age class, and sex of captured animals. Seropositivity was strongly associated with animal age class, but independent of sex. Forty-six percent of animals less than 2 years old, 63% of adults during peak reproductive years, and 100% of the oldest age class were seropositive. A nonparametric multivariate procedure (MRPP) showed that the composition of serovar antibody types changed with age, and ANOVA models demonstrated that antibody titers increased with age, suggesting long-term exposure to a greater number and variety (i.e., serovar types) of L. interrogans infections. This study presents the first quantitative survey of antibodies against L. interrogans serovars in a T. pecari population of the Pantanal. The high prevalence of leptospirosis antibodies in free-ranging white-lipped peccaries and the potential impacts on reproduction and population dynamics emphasize the need for further studies investigating the roles of Pantanal wildlife and livestock in the transmission and maintenance of L. interrogans in the environment. PMID:20596776

  18. Refolding of the recombinant protein OmpA70 from Leptospira interrogans from inclusion bodies using high hydrostatic pressure and partial characterization of its immunological properties.

    PubMed

    Fraga, Tatiana R; Chura-Chambi, Rosa M; Gonçales, Amane P; Morais, Zenaide M; Vasconcellos, Sílvio A; Morganti, Ligia; Martins, Elizabeth A L

    2010-07-20

    Leptospira is the etiological agent of leptospirosis, a life-threatening disease that affects human populations worldwide. Available vaccines have demonstrated limited effectiveness, and therapeutic interventions are complicated by the difficulty of establishing an early diagnosis. The genome of Leptospira strains was sequenced, and bioinformatic analyses revealed potential vaccine and serodiagnosis candidates. The present work studied OmpA70, a putative outer membrane protein from Leptospira interrogans serovar Copenhageni that combines structural features of Loa22, the first genetically defined virulence factor in Leptospira, and Lp49, a protein that reacts with sera from early and convalescent patients. Recombinant OmpA was produced in Escherichia coli in an insoluble form. Considering the importance of the structural integrity of a protein to confer immune protection, high hydrostatic pressure (HHP) was used to refold OmpA70 aggregated as inclusion bodies. HHP was applied in association with redox-shuffling reagents (oxidized and reduced glutathione) and guanidine hydrochloride or l-arginine. About 40% of the protein was refolded by applying 200MPa for 16h in concentrations of l-arginine above 0.4M. Circular dichroism revealed the presence of secondary structure. OmpA70 has immunogenic and antigenic properties as high antibody titers were seen after immunization with this protein, and sera from infected hamsters reacted with soluble OmpA70.

  19. Identification of Seroreactive Proteins of Leptospira interrogans Serovar Copenhageni Using a High-Density Protein Microarray Approach

    PubMed Central

    Lessa-Aquino, Carolina; Borges Rodrigues, Camila; Pablo, Jozelyn; Sasaki, Rie; Jasinskas, Algis; Liang, Li; Wunder, Elsio A.; Ribeiro, Guilherme S.; Vigil, Adam; Galler, Ricardo; Molina, Douglas; Liang, Xiaowu; Reis, Mitermayer G.; Ko, Albert I.; Medeiros, Marco Alberto; Felgner, Philip L.

    2013-01-01

    Background Leptospirosis is a widespread zoonotic disease worldwide. The lack of an adequate laboratory test is a major barrier for diagnosis, especially during the early stages of illness, when antibiotic therapy is most effective. Therefore, there is a critical need for an efficient diagnostic test for this life threatening disease. Methodology In order to identify new targets that could be used as diagnostic makers for leptopirosis, we constructed a protein microarray chip comprising 61% of Leptospira interrogans proteome and investigated the IgG response from 274 individuals, including 80 acute-phase, 80 convalescent-phase patients and 114 healthy control subjects from regions with endemic, high endemic, and no endemic transmission of leptospirosis. A nitrocellulose line blot assay was performed to validate the accuracy of the protein microarray results. Principal findings We found 16 antigens that can discriminate between acute cases and healthy individuals from a region with high endemic transmission of leptospirosis, and 18 antigens that distinguish convalescent cases. Some of the antigens identified in this study, such as LipL32, the non-identical domains of the Lig proteins, GroEL, and Loa22 are already known to be recognized by sera from human patients, thus serving as proof-of-concept for the serodiagnostic antigen discovery approach. Several novel antigens were identified, including the hypothetical protein LIC10215 which showed good sensitivity and specificity rates for both acute- and convalescent-phase patients. Conclusions Our study is the first large-scale evaluation of immunodominant antigens associated with naturally acquired leptospiral infection, and novel as well as known serodiagnostic leptospiral antigens that are recognized by antibodies in the sera of leptospirosis cases were identified. The novel antigens identified here may have potential use in both the development of new tests and the improvement of currently available assays for

  20. Comparative studies of strains Ictero No. I and RGA as the type strain of Leptospira interrogans: agglutinin absorption test, protein and antigen profiles, and enzyme activities.

    PubMed

    Hata, K; Ono, E; Yanagawa, R

    1988-01-01

    Strain Ictero No. I, the first isolate of Leptospira, isolated by Inada and Ido in 1914, was found to be sufficiently qualified to be the type strain of Leptospira interrogans rather than strain RGA. In an agglutinin absorption test, anti-Ictero No. I serum was not absorbed completely with strain RGA, and 25% of the homologous titer remained unabsorbed, while anti-RGA serum was completely absorbed with strain Ictero No. I. Thus, strain Ictero No. I was not serologically identical with strain RGA, and the two strains were considered to be different serovars. A protein band with a molecular weight of approximately 33,000 daltons was detected in strain Ictero No. I but not in strain RGA by SDS-PAGE. By Western blotting, this protein band was detectable with anti-Ictero No. I serum but not with anti-RGA serum. The presence of the 33K protein in strain Ictero No. I, but not in strain RGA, was confirmed by radioimmunoprecipitation using [125I]-labeled antigens, indicating that the protein antigen was surface-exposed. Only 8 of the 89 enzymes activities were different between strains Ictero No. I and RGA (line Sapporo). From the above results, we propose that strain Ictero No. I should be designated as the type strain of L. interrogans instead of strain RGA.

  1. SEROPREVALENCE OF NINE LEPTOSPIRA INTERROGANS SEROVARS IN WILD CARNIVORES, UNGULATES, AND PRIMATES FROM A ZOO POPULATION IN A METROPOLITAN REGION OF CHILE.

    PubMed

    Moreno-Beas, Eduardo; Abalos, Pedro; Hidalgo-Hermoso, Ezequiel

    2015-12-01

    Serum samples from 130 individuals representing 42 species of carnivores, ungulates, and primates from a population of captive mammals in Metropolitan Region in Chile were tested for antibodies against nine serovars of Leptospira interrogans using the microscopic agglutination test. Ten percent of the animals were seropositive to one or more serovars. Seroprevalence was significantly higher in ungulates (20.4%) compared to carnivores (3.8%) and primates (3.4%). There were no significant differences in seroprevalence among sex and age ranges. The most frequent serovar detected was Autumnalis, present in 53.4% of antibody-positive animals. Most positive animals had titers of ≤1 : 200, except for a maned wolf ( Chrysocyon brachyurus ) with titers of 1 : 400 against serovar Hardjo. To the authors' knowledge, this is the first report of Leptospira exposure detected in native endangered pudu ( Pudu puda ) and the first confirmation of exposure to L. interrogans in captive wild mammals in Chile. Leptospirosis should be considered as a differential diagnosis in future disease presentation for hepatitis or abortions in captive mammals in Chile.

  2. Safety and efficacy of a new octavalent combined Erysipelas, Parvo and Leptospira vaccine in gilts against Leptospira interrogans serovar Pomona associated disease and foetal death.

    PubMed

    Jacobs, A A C; Harks, F; Hoeijmakers, M; Collell, M; Segers, R P A M

    2015-07-31

    The safety and protective efficacy of a new octavalent combination vaccine containing inactivated Erysipelothrix rhusiopathiae, Parvovirus, and Leptospira interrogans (sensu lato) serogroups Canicola, Icterohaemorrhagiae, Australis (Bratislava), Grippotyphosa, Pomona and Tarassovi - Porcilis(®) Ery+Parvo+Lepto - was evaluated in laboratory studies and under field conditions. The safety (2× overdose and repeated dose) was tested in 26 gilts. In this study, neither vaccine related temperature increase nor other systemic reactions were observed after intramuscular vaccination. No local reactions were observed except for one animal that had a small local reaction (2cm diameter) that lasted for 5 days after the third vaccination. Efficacy was tested in 40 gilts. A group of 20 gilts was vaccinated at 20 and 24 weeks of age with Porcilis(®) Ery+Parvo+Lepto and a group of 20 age- and source-matched animals served as the control group. The gilts were inseminated at 41 weeks or 66 weeks of age and were challenged with serovar Pomona 10 weeks after insemination, corresponding to 6 months (n=2×10) and 12 months (n=2×10) after the last vaccination. After both the 6- and 12-month challenges the control animals developed clinical signs (fever, lethargy and anorexia) and leptospiraemia as determined by positive blood culture. In addition, both the 6- and 12-month challenges resulted in death of 21% and 27% of the total number of foetuses in the control groups, respectively. Clinical signs and leptospiraemia were statistically significantly lower in vaccinated gilts after both the 6- and 12-month challenges. In addition, foetal death was statistically significantly lower (3% and 2%, respectively) in vaccinated gilts after both the 6- and 12 month challenges. The vaccine was tested further under field conditions on a Portuguese farm with a history of an increasing abortion rate associated with a Leptospira serovar Pomona infection (confirmed by PCR and serology). This study was

  3. Safety and efficacy of a new octavalent combined Erysipelas, Parvo and Leptospira vaccine in gilts against Leptospira interrogans serovar Pomona associated disease and foetal death.

    PubMed

    Jacobs, A A C; Harks, F; Hoeijmakers, M; Collell, M; Segers, R P A M

    2015-07-31

    The safety and protective efficacy of a new octavalent combination vaccine containing inactivated Erysipelothrix rhusiopathiae, Parvovirus, and Leptospira interrogans (sensu lato) serogroups Canicola, Icterohaemorrhagiae, Australis (Bratislava), Grippotyphosa, Pomona and Tarassovi - Porcilis(®) Ery+Parvo+Lepto - was evaluated in laboratory studies and under field conditions. The safety (2× overdose and repeated dose) was tested in 26 gilts. In this study, neither vaccine related temperature increase nor other systemic reactions were observed after intramuscular vaccination. No local reactions were observed except for one animal that had a small local reaction (2cm diameter) that lasted for 5 days after the third vaccination. Efficacy was tested in 40 gilts. A group of 20 gilts was vaccinated at 20 and 24 weeks of age with Porcilis(®) Ery+Parvo+Lepto and a group of 20 age- and source-matched animals served as the control group. The gilts were inseminated at 41 weeks or 66 weeks of age and were challenged with serovar Pomona 10 weeks after insemination, corresponding to 6 months (n=2×10) and 12 months (n=2×10) after the last vaccination. After both the 6- and 12-month challenges the control animals developed clinical signs (fever, lethargy and anorexia) and leptospiraemia as determined by positive blood culture. In addition, both the 6- and 12-month challenges resulted in death of 21% and 27% of the total number of foetuses in the control groups, respectively. Clinical signs and leptospiraemia were statistically significantly lower in vaccinated gilts after both the 6- and 12-month challenges. In addition, foetal death was statistically significantly lower (3% and 2%, respectively) in vaccinated gilts after both the 6- and 12 month challenges. The vaccine was tested further under field conditions on a Portuguese farm with a history of an increasing abortion rate associated with a Leptospira serovar Pomona infection (confirmed by PCR and serology). This study was

  4. Live imaging of bioluminescent leptospira interrogans in mice reveals renal colonization as a stealth escape from the blood defenses and antibiotics.

    PubMed

    Ratet, Gwenn; Veyrier, Frédéric J; Fanton d'Andon, Martine; Kammerscheit, Xavier; Nicola, Marie-Anne; Picardeau, Mathieu; Boneca, Ivo G; Werts, Catherine

    2014-12-01

    Leptospira (L.) interrogans are bacteria responsible for a worldwide reemerging zoonosis. Some animals asymptomatically carry L. interrogans in their kidneys and excrete bacteria in their urine, which contaminates the environment. Humans are infected through skin contact with leptospires and develop mild to severe leptospirosis. Previous attempts to construct fluorescent or bioluminescent leptospires, which would permit in vivo visualization and investigation of host defense mechanisms during infection, have been unsuccessful. Using a firefly luciferase cassette and random transposition tools, we constructed bioluminescent chromosomal transformants in saprophytic and pathogenic leptospires. The kinetics of leptospiral dissemination in mice, after intraperitoneal inoculation with a pathogenic transformant, was tracked by bioluminescence using live imaging. For infective doses of 106 to 107 bacteria, we observed dissemination and exponential growth of leptospires in the blood, followed by apparent clearance of bacteria. However, with 2×108 bacteria, the septicemia led to the death of mice within 3 days post-infection. In surviving mice, one week after infection, pathogenic leptospires reemerged only in the kidneys, where they multiplied and reached a steady state, leading to a sustained chronic renal infection. These experiments reveal that a fraction of the leptospiral population escapes the potent blood defense, and colonizes a defined number of niches in the kidneys, proportional to the infective dose. Antibiotic treatments failed to eradicate leptospires that colonized the kidneys, although they were effective against L. interrogans if administered before or early after infection. To conclude, mice infected with bioluminescent L. interrogans proved to be a novel model to study both acute and chronic leptospirosis, and revealed that, in the kidneys, leptospires are protected from antibiotics. These bioluminescent leptospires represent a powerful new tool to

  5. Live Imaging of Bioluminescent Leptospira interrogans in Mice Reveals Renal Colonization as a Stealth Escape from the Blood Defenses and Antibiotics

    PubMed Central

    Ratet, Gwenn; Veyrier, Frédéric J.; Fanton d'Andon, Martine; Kammerscheit, Xavier; Nicola, Marie-Anne; Picardeau, Mathieu; Boneca, Ivo G.; Werts, Catherine

    2014-01-01

    Leptospira (L.) interrogans are bacteria responsible for a worldwide reemerging zoonosis. Some animals asymptomatically carry L. interrogans in their kidneys and excrete bacteria in their urine, which contaminates the environment. Humans are infected through skin contact with leptospires and develop mild to severe leptospirosis. Previous attempts to construct fluorescent or bioluminescent leptospires, which would permit in vivo visualization and investigation of host defense mechanisms during infection, have been unsuccessful. Using a firefly luciferase cassette and random transposition tools, we constructed bioluminescent chromosomal transformants in saprophytic and pathogenic leptospires. The kinetics of leptospiral dissemination in mice, after intraperitoneal inoculation with a pathogenic transformant, was tracked by bioluminescence using live imaging. For infective doses of 106 to 107 bacteria, we observed dissemination and exponential growth of leptospires in the blood, followed by apparent clearance of bacteria. However, with 2×108 bacteria, the septicemia led to the death of mice within 3 days post-infection. In surviving mice, one week after infection, pathogenic leptospires reemerged only in the kidneys, where they multiplied and reached a steady state, leading to a sustained chronic renal infection. These experiments reveal that a fraction of the leptospiral population escapes the potent blood defense, and colonizes a defined number of niches in the kidneys, proportional to the infective dose. Antibiotic treatments failed to eradicate leptospires that colonized the kidneys, although they were effective against L. interrogans if administered before or early after infection. To conclude, mice infected with bioluminescent L. interrogans proved to be a novel model to study both acute and chronic leptospirosis, and revealed that, in the kidneys, leptospires are protected from antibiotics. These bioluminescent leptospires represent a powerful new tool to

  6. "Features of two proteins of Leptospira interrogans with potential role in host-pathogen interactions"

    PubMed Central

    2012-01-01

    Background Leptospirosis is considered a re-emerging infectious disease caused by pathogenic spirochaetes of the genus Leptospira. Pathogenic leptospires have the ability to survive and disseminate to multiple organs after penetrating the host. Leptospires were shown to express surface proteins that interact with the extracellular matrix (ECM) and to plasminogen (PLG). This study examined the interaction of two putative leptospiral proteins with laminin, collagen Type I, collagen Type IV, cellular fibronectin, plasma fibronectin, PLG, factor H and C4bp. Results We show that two leptospiral proteins encoded by LIC11834 and LIC12253 genes interact with laminin in a dose - dependent and saturable mode, with dissociation equilibrium constants (KD) of 367.5 and 415.4 nM, respectively. These proteins were named Lsa33 and Lsa25 (Leptospiral surface adhesin) for LIC11834 and LIC12253, respectively. Metaperiodate - treated laminin reduced Lsa25 - laminin interaction, suggesting that sugar moieties of this ligand participate in this interaction. The Lsa33 is also PLG - binding receptor, with a KD of 23.53 nM, capable of generating plasmin in the presence of an activator. Although in a weak manner, both proteins interact with C4bp, a regulator of complement classical route. In silico analysis together with proteinase K and immunoflorescence data suggest that these proteins might be surface exposed. Moreover, the recombinant proteins partially inhibited leptospiral adherence to immobilized laminin and PLG. Conclusions We believe that these multifunctional proteins have the potential to participate in the interaction of leptospires to hosts by mediating adhesion and by helping the bacteria to escape the immune system and to overcome tissue barriers. To our knowledge, Lsa33 is the first leptospiral protein described to date with the capability of binding laminin, PLG and C4bp in vitro. PMID:22463075

  7. Infections by Leptospira interrogans, Seoul virus, and Bartonella spp. among Norway rats (Rattus norvegicus) from the urban slum environment in Brazil.

    PubMed

    Costa, Federico; Porter, Fleur Helena; Rodrigues, Gorete; Farias, Helena; de Faria, Marcus Tucunduva; Wunder, Elsio A; Osikowicz, Lynn M; Kosoy, Michael Y; Reis, Mitermayer Galvão; Ko, Albert I; Childs, James E

    2014-01-01

    Norway rats (Rattus norvegicus) are reservoir hosts for zoonotic pathogens that cause significant morbidity and mortality in humans. Studies evaluating the prevalence of zoonotic pathogens in tropical Norway rat populations are rare, and data on co-infection with multiple pathogens are nonexistent. Herein, we describe the prevalence of leptospiral carriage, Seoul virus (SEOV), and Bartonella spp. infection independently, in addition to the rates of co-infection among urban, slum-dwelling Norway rats in Salvador, Brazil, trapped during the rainy season from June to August of 2010. These data were complemented with previously unpublished Leptospira and SEOV prevalence information collected in 1998. Immunofluorescence staining of kidney impressions was used to identify Leptospira interrogans in 2010, whereas isolation was used in 1998, and western blotting was used to detect SEOV antibodies in 2010, whereas enzyme-linked immunosorbent assay (ELISA) was used in 1998: in 2010, Bartonella spp. were isolated from a subsample of rats. The most common pathogen in both years was Leptospira spp. (83%, n=142 in 1998, 63%, n=84 in 2010). SEOV was detected in 18% of individuals in both 1998 and 2010 (n=78 in 1998; n=73 in 2010), and two species of Bartonella were isolated from 5 of 26 rats (19%) tested in 2010. The prevalence of all agents increased significantly with rat mass/age. Acquisition of Leptospira spp. occurred at a younger mass/age than SEOV and Bartonella spp. infection, suggesting differences in the transmission dynamics of these pathogens. These data indicate that Norway rats in Salvador serve as reservoir hosts for all three of these zoonotic pathogens and that the high prevalence of leptospiral carriage in Salvador rats poses a high degree of risk to human health.

  8. Control of Gene Expression in Leptospira spp. by Transcription Activator-Like Effectors Demonstrates a Potential Role for LigA and LigB in Leptospira interrogans Virulence.

    PubMed

    Pappas, Christopher J; Picardeau, Mathieu

    2015-11-01

    Leptospirosis is a zoonotic disease that affects ∼1 million people annually, with a mortality rate of >10%. Currently, there is an absence of effective genetic manipulation tools for targeted mutagenesis in pathogenic leptospires. Transcription activator-like effectors (TALEs) are a recently described group of repressors that modify transcriptional activity in prokaryotic and eukaryotic cells by directly binding to a targeted sequence within the host genome. To determine the applicability of TALEs within Leptospira spp., two TALE constructs were designed. First, a constitutively expressed TALE gene specific for the lacO-like region upstream of bgaL was trans inserted in the saprophyte Leptospira biflexa (the TALEβgal strain). Reverse transcriptase PCR (RT-PCR) analysis and enzymatic assays demonstrated that BgaL was not expressed in the TALEβgal strain. Second, to study the role of LigA and LigB in pathogenesis, a constitutively expressed TALE gene with specificity for the homologous promoter regions of ligA and ligB was cis inserted into the pathogen Leptospira interrogans (TALElig). LigA and LigB expression was studied by using three independent clones: TALElig1, TALElig2, and TALElig3. Immunoblot analysis of osmotically induced TALElig clones demonstrated 2- to 9-fold reductions in the expression levels of LigA and LigB, with the highest reductions being noted for TALElig1 and TALElig2, which were avirulent in vivo and nonrecoverable from animal tissues. This study reconfirms galactosidase activity in the saprophyte and suggests a role for LigA and LigB in pathogenesis. Collectively, this study demonstrates that TALEs are effective at reducing the expression of targeted genes within saprophytic and pathogenic strains of Leptospira spp., providing an additional genetic manipulation tool for this genus. PMID:26341206

  9. Effect of exposure to Neospora caninum, Salmonella, and Leptospira interrogans serovar Hardjo on the economic performance of Irish dairy herds.

    PubMed

    O' Doherty, E; Sayers, R; O' Grady, L; Shalloo, L

    2015-04-01

    The objective of the current study was to quantify the effects of exposure to Salmonella, Neospora caninum, and Leptospira interrogans serovar Hardjo (L. hardjo) on dairy farm profitability and to simulate the effect of vaccination for Salmonella and L. hardjo on dairy farm profitability. The production effects associated with exposure to each of these pathogens in study herds were defined under 3 categories: (1) milk production effects, (2) reproduction effects (including culling), and (3) mortality effects. The production effects associated with exposure to Salmonella, N. caninum, and L. hardjo were incorporated into the Moorepark Dairy Systems Model. In the analysis, herds negative for exposure to Salmonella, N. caninum, and L. hardjo were assumed baseline herds, with all results presented relative to this base. In simulations examining the effect of vaccination for Salmonella and L. hardjo on farm profitability, vaccinated herds (vaccination costs included) were considered as baseline herds and results were presented relative to this base. Total annual profits in unvaccinated herds were reduced by €77.31, €94.71, and €112.11 per cow at milk prices of €0.24, €0.29, and €0.34/L, respectively, as a result of exposure to Salmonella. In the current study, herds positive for exposure to Salmonella recorded a 316-kg reduction in milk yield, whereas no association was detected between exposure to N. caninum or L. hardjo and milk production. Exposure to both N. caninum and L. hardjo was associated with compromised reproductive performance. Herds positive for exposure to N. caninum and Salmonella had greater rates of adult cow mortality and calf mortality, respectively. Vaccination for both Salmonella and L. hardjo was associated with improved performance in study herds. Exposure to N. caninum resulted in a reduction in annual farm profits of €11.55, €12, and €12.44 per cow at each milk price, whereas exposure to L. hardjo resulted in a reduction in

  10. Analysis of a Spontaneous Non-Motile and Avirulent Mutant Shows That FliM Is Required for Full Endoflagella Assembly in Leptospira interrogans.

    PubMed

    Fontana, Célia; Lambert, Ambroise; Benaroudj, Nadia; Gasparini, David; Gorgette, Olivier; Cachet, Nathalie; Bomchil, Natalia; Picardeau, Mathieu

    2016-01-01

    Pathogenic Leptospira strains are responsible for leptospirosis, a worldwide emerging zoonotic disease. These spirochetes are unique amongst bacteria because of their corkscrew-like cell morphology and their periplasmic flagella. Motility is reported as an important virulence determinant, probably favoring entry and dissemination of pathogenic Leptospira in the host. However, proteins constituting the periplasmic flagella and their role in cell shape, motility and virulence remain poorly described. In this study, we characterized a spontaneous L. interrogans mutant strain lacking motility, correlated with the loss of the characteristic hook-shaped ends, and virulence in the animal model. Whole genome sequencing allowed the identification of one nucleotide deletion in the fliM gene resulting in a premature stop codon, thereby preventing the production of flagellar motor switch protein FliM. Genetic complementation restored cell morphology, motility and virulence comparable to those of wild type cells. Analyses of purified periplasmic flagella revealed a defect in flagella assembly, resulting in shortened flagella compared to the wild type strain. This also correlated with a lower amount of major filament proteins FlaA and FlaB. Altogether, these findings demonstrate that FliM is required for full and correct assembly of the flagella which is essential for motility and virulence. PMID:27044038

  11. Analysis of a Spontaneous Non-Motile and Avirulent Mutant Shows That FliM Is Required for Full Endoflagella Assembly in Leptospira interrogans

    PubMed Central

    Fontana, Célia; Lambert, Ambroise; Benaroudj, Nadia; Gasparini, David; Gorgette, Olivier; Cachet, Nathalie; Bomchil, Natalia; Picardeau, Mathieu

    2016-01-01

    Pathogenic Leptospira strains are responsible for leptospirosis, a worldwide emerging zoonotic disease. These spirochetes are unique amongst bacteria because of their corkscrew-like cell morphology and their periplasmic flagella. Motility is reported as an important virulence determinant, probably favoring entry and dissemination of pathogenic Leptospira in the host. However, proteins constituting the periplasmic flagella and their role in cell shape, motility and virulence remain poorly described. In this study, we characterized a spontaneous L. interrogans mutant strain lacking motility, correlated with the loss of the characteristic hook-shaped ends, and virulence in the animal model. Whole genome sequencing allowed the identification of one nucleotide deletion in the fliM gene resulting in a premature stop codon, thereby preventing the production of flagellar motor switch protein FliM. Genetic complementation restored cell morphology, motility and virulence comparable to those of wild type cells. Analyses of purified periplasmic flagella revealed a defect in flagella assembly, resulting in shortened flagella compared to the wild type strain. This also correlated with a lower amount of major filament proteins FlaA and FlaB. Altogether, these findings demonstrate that FliM is required for full and correct assembly of the flagella which is essential for motility and virulence. PMID:27044038

  12. In Vivo-Expressed Proteins of Virulent Leptospira interrogans Serovar Autumnalis N2 Elicit Strong IgM Responses of Value in Conclusive Diagnosis

    PubMed Central

    Raja, Veerapandian; Shanmughapriya, Santhanam; Kanagavel, Murugesan; Artiushin, Sergey C.; Velineni, Sridhar; Timoney, John F.

    2015-01-01

    Leptospirosis is a serious zoonosis that is underdiagnosed because of limited access to laboratory facilities in Southeast Asia, Central and South America, and Oceania. Timely diagnosis of locally distributed serovars of high virulence is crucial for successful care and outbreak management. Using pooled patient sera, an expression gene library of a virulent Leptospira interrogans serovar Autumnalis strain N2 isolated in South India was screened. The identified genes were characterized, and the purified recombinant proteins were used as antigens in IgM enzyme-linked immunosorbent assay (ELISA) either singly or in combination. Sera (n = 118) from cases of acute leptospirosis along with sera (n = 58) from healthy subjects were tested for reactivity with the identified proteins in an ELISA designed to detect specific IgM responses. We have identified nine immunoreactive proteins, ArgC, RecA, GlpF, FliD, TrmD, RplS, RnhB, Lp28.6, and Lrr44.9, which were found to be highly conserved among pathogenic leptospires. Apparently, the proteins ArgC, RecA, GlpF, FliD, TrmD, and Lrr44.9 are expressed during natural infection of the host and undetectable in in vitro cultures. Among all the recombinant proteins used as antigens in IgM ELISA, ArgC had the highest sensitivity and specificity, 89.8% and 95.5%, respectively, for the conclusive diagnosis of leptospirosis. The use of ArgC and RecA in combination for IgM ELISA increased the sensitivity and specificity to 95.7% and 94.9%, respectively. ArgC and RecA thus elicited specific IgM responses and were therefore effective in laboratory confirmation of Leptospira infection. PMID:26607308

  13. Comparison of Bacterial Burden and Cytokine Gene Expression in Golden Hamsters in Early Phase of Infection with Two Different Strains of Leptospira interrogans.

    PubMed

    Fujita, Rie; Koizumi, Nobuo; Sugiyama, Hiromu; Tomizawa, Rina; Sato, Ryoichi; Ohnishi, Makoto

    2015-01-01

    Leptospirosis, a zoonotic infection with worldwide prevalence, is caused by pathogenic spirochaetes of Leptospira spp., and exhibits an extremely broad clinical spectrum in human patients. Although previous studies indicated that specific serovars or genotypes of Leptospira spp. were associated with severe leptospirosis or its outbreak, the mechanism underlying the difference in virulence of the various Leptospira serotypes or genotypes remains unclear. The present study addresses this question by measuring and comparing bacterial burden and cytokine gene expression in hamsters infected with strains of two L. interrogans serovars Manilae (highly virulent) and Hebdomadis (less virulent). The histopathology of kidney, liver, and lung tissues was also investigated in infected hamsters. A significantly higher bacterial burden was observed in liver tissues of hamsters infected with serovar Manilae than those infected with serovar Hebdomadis (p < 0.01). The average copy number of the leptospiral genome was 1,302 and 20,559 in blood and liver, respectively, of hamsters infected with serovar Manilae and 1,340 and 4,896, respectively, in hamsters infected with serovar Hebdomadis. The expression levels of mip1alpha in blood; tgfbeta, il1beta, mip1alpha, il10, tnfalpha and cox2 in liver; and tgfbeta, il6, tnfalpha and cox2 in lung tissue were significantly higher in hamsters infected with serovar Manilae than those infected with serovar Hebdomadis (p < 0.05). In addition, infection with serovar Manilae resulted in a significantly larger number of hamsters with tnfalpha upregulation (p = 0.04). Severe distortion of tubular cell arrangement and disruption of renal tubules in kidney tissues and hemorrhage in lung tissues were observed in Manilae-infected hamsters. These results demonstrate that serovar Manilae multiplied more efficiently in liver tissues and induced significantly higher expression of genes encoding pro- and anti-inflammatory cytokines than serovar Hebdomadis

  14. Fine Mapping of the Interaction between C4b-Binding Protein and Outer Membrane Proteins LigA and LigB of Pathogenic Leptospira interrogans.

    PubMed

    Breda, Leandro C D; Hsieh, Ching-Lin; Castiblanco Valencia, Mónica M; da Silva, Ludmila B; Barbosa, Angela S; Blom, Anna M; Chang, Yung-Fu; Yung-Fu, Chang; Isaac, Lourdes

    2015-01-01

    The complement system consists of more than 40 proteins that participate in the inflammatory response and in pathogen killing. Complement inhibitors are necessary to avoid the excessive consumption and activation of this system on host cells. Leptospirosis is a worldwide zoonosis caused by spirochetes from the genus Leptospira. Pathogenic leptospires are able to escape from complement activation by binding to host complement inhibitors Factor H [FH] and C4b-binding protein (C4BP) while non-pathogenic leptospires are rapidly killed in the presence of fresh serum. In this study, we demonstrate that complement control protein domains (CCP) 7 and 8 of C4BP α-chain interact with the outer membrane proteins LcpA, LigA and LigB from the pathogenic leptospire L. interrogans. The interaction between C4BP and LcpA, LigA and LigB is sensitive to ionic strength and inhibited by heparin. We fine mapped the LigA and LigB domains involved in its binding to C4BP and heparin and found that both interactions are mediated through the bacterial immunoglobulin-like (Big) domains 7 and 8 (LigA7-8 and LigB7-8) of both LigA and LigB and also through LigB9-10. Therefore, C4BP and heparin may share the same binding sites on Lig proteins.

  15. Leptospira Protein Expression During Infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We are characterizing protein expression in vivo during experimental leptospirosis using immunofluorescence microscopy. Coding regions for several proteins were identified through analysis of Leptospira interrogans serovar Copenhageni and L. borgpetersenii serovar Hardjo genomes. In addition, codi...

  16. The recombinant LIC10508 is a plasma fibronectin, plasminogen, fibrinogen and C4BP-binding protein of Leptospira interrogans.

    PubMed

    Siqueira, Gabriela H; Teixeira, Aline F; Fernandes, Luis G; de Souza, Gisele O; Kirchgatter, Karin; Romero, Eliete C; Vasconcellos, Silvio A; Vieira, Monica L; Nascimento, Ana Lucia T O

    2016-03-01

    Leptospirosis is a zoonosis caused by pathogenic Leptospira spp. In this study, we report that the recombinant proteins LIC10507, LIC10508 and LIC10509 are recognized by confirmed leptospirosis serum samples at both phases of the disease. The recombinant rLIC10508 and rLIC10507 are plasminogen (PLG)-binding proteins, capable of generating plasmin in the presence of a PLG activator. The proteins bind to PLG in a dose-dependent and saturable manner, fulfilling host-ligand interaction. Furthermore, rLIC10508 interacts with fibrinogen (Fg), plasma fibronectin and C4b binding protein (C4BP). The binding of rLIC10508 to Fg decreases the fibrin clotting in a thrombin-catalyzed reaction. The incubation with 4 μM of protein promoted 40% inhibition upon clotting formation. C4BP bound to rLIC10508 retained its cofactor activity for factor I promoting the cleavage of C4b protein, which may reduce the membrane attack complex formation. Although these proteins have high amino acid sequence similarity, rLIC10508 is the most talented of the three, a behavior that might be explained by its unique putative 3D structure, whereas structures of rLIC10507 and rLIC10509 are very similar. Plasmin generation (rLIC10507 and rLIC10508), together with decreasing fibrin clot formation (rLIC10508) and impairment of the complement system (rLIC10508) may help the bacteria to overcome host defense, facilitating the infection process.

  17. Detection of human leptospirosis as a cause of acute fever by capture ELISA using a Leptospira interrogans serovar Copenhageni (M20) derived antigen

    PubMed Central

    2013-01-01

    Background Leptospirosis is a potentially lethal zoonosis mainly affecting low-resource tropical countries, including Peru and its neighbouring countries. Timely diagnosis of leptospirosis is critical but may be challenging in the regions where it is most prevalent. The serodiagnostic gold standard microagglutination test (MAT) may be technically prohibitive. Our objective in this study was to assess the sensitivity, specificity, and predictive value of an IgM antibody capture enzyme-linked immunoassay (MAC-ELISA) derived from the M20 strain of Leptospira interrogans serovar Copenhageni (M20) by comparison to MAT, which was used as the gold standard method of diagnosis. Methods Acute and convalescent sera from participants participating in a passive febrile surveillance study in multiple regions of Peru were tested by both IgM MAC-ELISA and MAT. The sensitivity, specificity, positive and negative predictive value (PPV, NPV) of the MAC-ELISA assay for acute, convalescent and paired sera by comparison to MAT were calculated. Results The sensitivity, specificity, PPV and NPV of the MAC-ELISA assay for acute sera were 92.3%, 56.0%, 35.3% and 96.6% respectively. For convalescent sera, the sensitivity, specificity, PPV and NPV of the MAC-ELISA assay were 93.3%, 51.5%, 63.6% and 89.5% respectively. For paired sera, the sensitivity, specificity, PPV and NPV of the MAC-ELISA assay were 93.6%, 37.5%, 59.2%, 85.7% respectively. Conclusions The M20 MAC-ELISA assay performed with a high sensitivity and low specificity in the acute phase of illness. Sensitivity was similar as compared with MAT in the convalescent phase and specificity remained low. Paired sera were the most sensitive but least specific by comparison to MAT serodiagnosis. NPV for acute, convalescent and paired sera was high. The limited specificity and high sensitivity of the MAC-ELISA IgM suggests that it would be most valuable to exclude leptospirosis in low-resource regions that lack immediate access to

  18. Leptospirosis in Kuala Lumpur and the comparative evaluation of two rapid commercial diagnostic kits against the MAT test for the detection of antibodies to leptospira interrogans.

    PubMed

    Sekhar, W Y; Soo, E H; Gopalakrishnan, V; Devi, S

    2000-08-01

    The aim of the study was to look into the epidemiology of serodiagnosed cases of leptospirosis at the University Hospital and compare two commercial ELISA Assays to the Microscopic Agglutination Test (MAT). Demographic data for all serodiagnosed cases for the years 1991-1997 were collected. From this data, 104 sera (n = 104) were selected as samples for comparative evaluation of the commercial ELISAs (INDX Dip-S-Ticks and PanBio ELISA) to the MAT test. Thirty two (n = 32) negative control sera were selected from serodiagnosed cases of other differential diagnosis of leptospira infection. The MAT test is a standard test that detects agglutination antibodies to leptospira biflexa, while the INDX Dip-S-Ticks is an ELISA dot test assaying for total anti-leptospira antibodies. The PanBio ELISA is a colorometric assay in test well strips to detect anti-leptospira IgM. The sensitivity, specificity, and efficiency of tests were calculated at a MAT cut-off value of 1:320. Demographic data showed that leptospirosis peaks during March-May and Aug-Nov coinciding with the inter-monsoon period with more men being infected than women and more adults than children. The sensitivity, specificity, and efficiency of test for the INDX Dip-S-Ticks were 83.3%, 93.8% and 87.5% while the values for the PanBio ELISA were 54.2%, 96.9% and 71.3%. The suboptimal PanBio result could be related to the blocking effect of high IgG titres or could be related to the diagnostic MAT cut-off values used in this study. The data hence reflects a pattern of transmission that is related to "wet" occupational risk factors. The commercial assays evaluated, are easier to perform but interpretation of results should be based on level of endemicity. The INDX Dip-S-Ticks allows this flexibility and is a practical alternative to the MAT test.

  19. Leptospirosis in Kuala Lumpur and the comparative evaluation of two rapid commercial diagnostic kits against the MAT test for the detection of antibodies to leptospira interrogans.

    PubMed

    Sekhar, W Y; Soo, E H; Gopalakrishnan, V; Devi, S

    2000-08-01

    The aim of the study was to look into the epidemiology of serodiagnosed cases of leptospirosis at the University Hospital and compare two commercial ELISA Assays to the Microscopic Agglutination Test (MAT). Demographic data for all serodiagnosed cases for the years 1991-1997 were collected. From this data, 104 sera (n = 104) were selected as samples for comparative evaluation of the commercial ELISAs (INDX Dip-S-Ticks and PanBio ELISA) to the MAT test. Thirty two (n = 32) negative control sera were selected from serodiagnosed cases of other differential diagnosis of leptospira infection. The MAT test is a standard test that detects agglutination antibodies to leptospira biflexa, while the INDX Dip-S-Ticks is an ELISA dot test assaying for total anti-leptospira antibodies. The PanBio ELISA is a colorometric assay in test well strips to detect anti-leptospira IgM. The sensitivity, specificity, and efficiency of tests were calculated at a MAT cut-off value of 1:320. Demographic data showed that leptospirosis peaks during March-May and Aug-Nov coinciding with the inter-monsoon period with more men being infected than women and more adults than children. The sensitivity, specificity, and efficiency of test for the INDX Dip-S-Ticks were 83.3%, 93.8% and 87.5% while the values for the PanBio ELISA were 54.2%, 96.9% and 71.3%. The suboptimal PanBio result could be related to the blocking effect of high IgG titres or could be related to the diagnostic MAT cut-off values used in this study. The data hence reflects a pattern of transmission that is related to "wet" occupational risk factors. The commercial assays evaluated, are easier to perform but interpretation of results should be based on level of endemicity. The INDX Dip-S-Ticks allows this flexibility and is a practical alternative to the MAT test. PMID:11256343

  20. Detection of reactive canines to Leptospira in Campeche City, Mexico.

    PubMed

    Blum Domínguez, Selene Del C; Chi Dzib, María Y; Maldonado Velázquez, María G; Nuñez Oreza, Luis A; Gómez Solano, Mónica I; Caballero Poot, Rebeca I; Tamay Segovia, Paulino

    2013-01-01

    Leptospira reactivity in stray and household dogs in Campeche as well as associated risk factors to the seropositivity in household dogs have been herein determined. The survey included 323 dogs, 142 of which were stray dogs and 181 household dogs. Nine Leptospira interrogans serovars were tested by the microagglutination test. Reactivity was 21.3 % (69/323), 17.2 % corresponded to household dogs and 26.7 % to stray dogs. Leptospira Canicola (29 %), Leptospira Hardjo (22.58 %), and Leptospira Icterohaemorrhagiae (16.12 %) were the most common serovars reacting against the serum of household animals, while Leptospira Canicola (15.78 %), Leptospira icterohaemorrhagiae (13.15 %), and Leptospira Pomona (7.89 %) were those reacting in stray dogs. Results showed that all dogs have been in contact with different Leptospira serovars and outdoor exposure is the main infection risk factor. PMID:23560786

  1. Genome-wide transposon mutagenesis in pathogenic Leptospira species.

    PubMed

    Murray, Gerald L; Morel, Viviane; Cerqueira, Gustavo M; Croda, Julio; Srikram, Amporn; Henry, Rebekah; Ko, Albert I; Dellagostin, Odir A; Bulach, Dieter M; Sermswan, Rasana W; Adler, Ben; Picardeau, Mathieu

    2009-02-01

    Leptospira interrogans is the most common cause of leptospirosis in humans and animals. Genetic analysis of L. interrogans has been severely hindered by a lack of tools for genetic manipulation. Recently we developed the mariner-based transposon Himar1 to generate the first defined mutants in L. interrogans. In this study, a total of 929 independent transposon mutants were obtained and the location of insertion determined. Of these mutants, 721 were located in the protein coding regions of 551 different genes. While sequence analysis of transposon insertion sites indicated that transposition occurred in an essentially random fashion in the genome, 25 unique transposon mutants were found to exhibit insertions into genes encoding 16S or 23S rRNAs, suggesting these genes are insertional hot spots in the L. interrogans genome. In contrast, loci containing notionally essential genes involved in lipopolysaccharide and heme biosynthesis showed few transposon insertions. The effect of gene disruption on the virulence of a selected set of defined mutants was investigated using the hamster model of leptospirosis. Two attenuated mutants with disruptions in hypothetical genes were identified, thus validating the use of transposon mutagenesis for the identification of novel virulence factors in L. interrogans. This library provides a valuable resource for the study of gene function in L. interrogans. Combined with the genome sequences of L. interrogans, this provides an opportunity to investigate genes that contribute to pathogenesis and will provide a better understanding of the biology of L. interrogans. PMID:19047402

  2. First Isolates of Leptospira spp., from Rodents Captured in Angola

    PubMed Central

    Fortes-Gabriel, Elsa; Carreira, Teresa; Vieira, Maria Luísa

    2016-01-01

    Rodents play an important role in the transmission of pathogenic Leptospira spp. However, in Angola, neither the natural reservoirs of these spirochetes nor leptospirosis diagnosis has been considered. Regarding this gap, we captured rodents in Luanda and Huambo provinces to identify circulating Leptospira spp. Rodent kidney tissue was cultured and DNA amplified and sequenced. Culture isolates were evaluated for pathogenic status and typing with rabbit antisera; polymerase chain reaction (PCR) and sequencing were also performed. A total of 37 rodents were captured: Rattus rattus (15, 40.5%), Rattus norvegicus (9, 24.3%), and Mus musculus (13, 35.2%). Leptospiral DNA was amplified in eight (21.6%) kidney samples. From the cultures, we obtained four (10.8%) Leptospira isolates belonging to the Icterohaemorrhagiae and Ballum serogroups of Leptospira interrogans and Leptospira borgpetersenii genospecies, respectively. This study provides information about circulating leptospires spread by rats and mice in Angola. PMID:26928840

  3. First Isolates of Leptospira spp., from Rodents Captured in Angola.

    PubMed

    Fortes-Gabriel, Elsa; Carreira, Teresa; Vieira, Maria Luísa

    2016-05-01

    Rodents play an important role in the transmission of pathogenic Leptospira spp. However, in Angola, neither the natural reservoirs of these spirochetes nor leptospirosis diagnosis has been considered. Regarding this gap, we captured rodents in Luanda and Huambo provinces to identify circulating Leptospira spp. Rodent kidney tissue was cultured and DNA amplified and sequenced. Culture isolates were evaluated for pathogenic status and typing with rabbit antisera; polymerase chain reaction (PCR) and sequencing were also performed. A total of 37 rodents were captured: Rattus rattus (15, 40.5%), Rattus norvegicus (9, 24.3%), and Mus musculus (13, 35.2%). Leptospiral DNA was amplified in eight (21.6%) kidney samples. From the cultures, we obtained four (10.8%) Leptospira isolates belonging to the Icterohaemorrhagiae and Ballum serogroups of Leptospira interrogans and Leptospira borgpetersenii genospecies, respectively. This study provides information about circulating leptospires spread by rats and mice in Angola.

  4. [Detection of leptospira by culture of vitreous humor and detection of antibodies against leptospira in vitreous humor and serum of 225 horses with equine recurrent uveitis].

    PubMed

    Dorrego-Keiter, Elisa; Tóth, József; Dikker, Lieke; Sielhorst, Jutta; Schusser, Gerald Fritz

    2016-01-01

    In the ongoing discussion regarding the aetiopathogenesis of equine recurrent uveitis (ERU) it was the aim of the present study to elucidate the relationship of leptospira infection and ERU. In a population of 225 horses leptospira were examined in vitreous humor by culture and leptospira antibody were detected in vitreous humor and serum samples. Preoperative serum samples were collected from 221/225 ERU patients of different age, gender and breed. Undiluted vitreous humor was aseptically taken from 198/225 patients that underwent pars plana vitrectomy at the beginning of surgery and from 27/225 patients' eyeball after enucleation: Serum and vitreous humor were tested for specific leptospiral antibodies by microscopic agglutination test (MAT). Furthermore, vitreous humor was examined by culture. 20 patients which were euthanized due to a live-threatening disease other than ERU served as a control group. A total of 127/221 (57.5%) horses had serum antibodies (≥ 1:100). Most frequently antibodies against L. interrogans serovar Grippotyphosa were detected (79/127), followed by L. interrogans serovar lcterohaemorrhagiae (34/127) and L. interrogans serovar Bratislava (29/127). Only 79/225 horses (35.1%) had leptospiral antibodies in vitreous humor, in which L. interrogans serovar Grippotyphosa (67/79) was identified most frequently followed by L. interrogans serovar Pomona (18/79) and L. interrogans serovar lcterohaemorrhagiae (8/79) which was identified as single or multiple reaction. Isolation of leptospira from vitreous humor was positive in 34/212 horses (16%). 10/20 control horses had a positive antibody titer against leptospira in serum and 2/20 horses in vitreous humor, whereas there was no leptospira detected in culture. The result of 84% negative cultures from vitreous humor of 212 ERU patients is decisive for the diagnosis and therapy of ERU. PMID:27344913

  5. [Detection of leptospira by culture of vitreous humor and detection of antibodies against leptospira in vitreous humor and serum of 225 horses with equine recurrent uveitis].

    PubMed

    Dorrego-Keiter, Elisa; Tóth, József; Dikker, Lieke; Sielhorst, Jutta; Schusser, Gerald Fritz

    2016-01-01

    In the ongoing discussion regarding the aetiopathogenesis of equine recurrent uveitis (ERU) it was the aim of the present study to elucidate the relationship of leptospira infection and ERU. In a population of 225 horses leptospira were examined in vitreous humor by culture and leptospira antibody were detected in vitreous humor and serum samples. Preoperative serum samples were collected from 221/225 ERU patients of different age, gender and breed. Undiluted vitreous humor was aseptically taken from 198/225 patients that underwent pars plana vitrectomy at the beginning of surgery and from 27/225 patients' eyeball after enucleation: Serum and vitreous humor were tested for specific leptospiral antibodies by microscopic agglutination test (MAT). Furthermore, vitreous humor was examined by culture. 20 patients which were euthanized due to a live-threatening disease other than ERU served as a control group. A total of 127/221 (57.5%) horses had serum antibodies (≥ 1:100). Most frequently antibodies against L. interrogans serovar Grippotyphosa were detected (79/127), followed by L. interrogans serovar lcterohaemorrhagiae (34/127) and L. interrogans serovar Bratislava (29/127). Only 79/225 horses (35.1%) had leptospiral antibodies in vitreous humor, in which L. interrogans serovar Grippotyphosa (67/79) was identified most frequently followed by L. interrogans serovar Pomona (18/79) and L. interrogans serovar lcterohaemorrhagiae (8/79) which was identified as single or multiple reaction. Isolation of leptospira from vitreous humor was positive in 34/212 horses (16%). 10/20 control horses had a positive antibody titer against leptospira in serum and 2/20 horses in vitreous humor, whereas there was no leptospira detected in culture. The result of 84% negative cultures from vitreous humor of 212 ERU patients is decisive for the diagnosis and therapy of ERU.

  6. Conservation of the S10-spc-alpha Locus within Otherwise Highly Pastic Genomes Provides Phylogenetic Insight into the Genus Leptospira

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A comparative analysis of the Leptospira interrogans S10-spc-alpha operon was performed by PCR using primer sets covering the whole operon. Correctly sized fragments were obtained by PCR from all of L. interrogans strains for each primer set indicating that the S10-spc-alpha locus is well conserved ...

  7. Sensitivity of pathogenic and free-living Leptospira spp. to UV radiation and mitomycin C

    SciTech Connect

    Stamm, L.V.; Charon, N.W.

    1988-03-01

    The habitats for the two major Leptospira spp. differ. The main habitat of L. biflexa is soil and water, whereas L. interrogans primarily resides in the renal tubules of animals. We investigated whether these two species, along with L. illini (species incertae sedis), differ with respect to their sensitivity to UV radiation. The doses of UV resulting in 37, 10 and 1% survival were determined for representive serovars from each species. L. interrogans serovar pomona was 3.0 to 4.8 times more sensitive to UV than the other Leptospira species under the 37, 10, and 1% survival parameters. In comparison to other bacteria, L. interrogans serovar pomona is among the most sensitive to UV. In a qualitative UV sensitivity assay., L. interrogans serovars were found to be in general more sensitive than L. biflexa serovars. All three species were found to have a photoreactivation DNA repair mechanism. Since organisms that are resistant to UV are often resistant to the DNA cross-linking agent mitomycin C, we tested the relative sensitivity of several Leptospira serovars to this compound. With few exceptions, L. biflexa and L. illini serovars were considerably more resistant to mitomycin C than the L. interrogans serovars. The mitomycin C sensitivity assay could be a useful addition to current characterization tests used to differentiate the Leptospira species.

  8. Draft Genome Sequence of the First Pathogenic Leptospira Isolates from Ecuador.

    PubMed

    Barragan, Veronica; Sahl, Jason W; Wiggins, Kristin; Chiriboga, Jorge; Salinas, Ana; Cantos, Nancy E; Loor, Mariana N; Intriago, Bertha I; Morales, Melba; Trueba, Gabriel; Pearson, Talima

    2016-01-01

    Pathogenic Leptospira spp. cause leptospirosis upon contact with mucosa through wounds or ingestion, leading to headaches, fever, jaundice, kidney or liver failure, or death in about 1.3 million people each year. Here, we present the draft genomes of one L. santarosai isolate and two L. interrogans isolates from Ecuador. PMID:27151788

  9. Draft Genome Sequence of the First Pathogenic Leptospira Isolates from Ecuador

    PubMed Central

    Barragan, Veronica; Sahl, Jason W.; Wiggins, Kristin; Chiriboga, Jorge; Salinas, Ana; Cantos, Nancy E.; Loor, Mariana N.; Intriago, Bertha I.; Morales, Melba; Trueba, Gabriel

    2016-01-01

    Pathogenic Leptospira spp. cause leptospirosis upon contact with mucosa through wounds or ingestion, leading to headaches, fever, jaundice, kidney or liver failure, or death in about 1.3 million people each year. Here, we present the draft genomes of one L. santarosai isolate and two L. interrogans isolates from Ecuador. PMID:27151788

  10. Distribution of Plasmids in Distinct Leptospira Pathogenic Species.

    PubMed

    Wang, Yanzhuo; Zhuang, Xuran; Zhong, Yi; Zhang, Cuicai; Zhang, Yan; Zeng, Lingbing; Zhu, Yongzhang; He, Ping; Dong, Ke; Pal, Utpal; Guo, Xiaokui; Qin, Jinhong

    2015-11-01

    Leptospirosis, caused by pathogenic Leptospira, is a worldwide zoonotic infection. The genus Leptospira includes at least 21 species clustered into three groups--pathogens, non-pathogens, and intermediates--based on 16S rRNA phylogeny. Research on Leptospira is difficult due to slow growth and poor transformability of the pathogens. Recent identification of extrachromosomal elements besides the two chromosomes in L. interrogans has provided new insight into genome complexity of the genus Leptospira. The large size, low copy number, and high similarity of the sequence of these extrachromosomal elements with the chromosomes present challenges in isolating and detecting them without careful genome assembly. In this study, two extrachromosomal elements were identified in L. borgpetersenii serovar Ballum strain 56604 through whole genome assembly combined with S1 nuclease digestion following pulsed-field gel electrophoresis (S1-PFGE) analysis. Further, extrachromosomal elements in additional 15 Chinese epidemic strains of Leptospira, comprising L. borgpetersenii, L. weilii, and L. interrogans, were successfully separated and identified, independent of genome sequence data. Southern blot hybridization with extrachromosomal element-specific probes, designated as lcp1, lcp2 and lcp3-rep, further confirmed their occurrences as extrachromosomal elements. In total, 24 plasmids were detected in 13 out of 15 tested strains, among which 11 can hybridize with the lcp1-rep probe and 11 with the lcp2-rep probe, whereas two can hybridize with the lcp3-rep probe. None of them are likely to be species-specific. Blastp search of the lcp1, lcp2, and lcp3-rep genes with a nonredundant protein database of Leptospira species genomes showed that their homologous sequences are widely distributed among clades of pathogens but not non-pathogens or intermediates. These results suggest that the plasmids are widely distributed in Leptospira species, and further elucidation of their biological

  11. Distribution of Plasmids in Distinct Leptospira Pathogenic Species.

    PubMed

    Wang, Yanzhuo; Zhuang, Xuran; Zhong, Yi; Zhang, Cuicai; Zhang, Yan; Zeng, Lingbing; Zhu, Yongzhang; He, Ping; Dong, Ke; Pal, Utpal; Guo, Xiaokui; Qin, Jinhong

    2015-11-01

    Leptospirosis, caused by pathogenic Leptospira, is a worldwide zoonotic infection. The genus Leptospira includes at least 21 species clustered into three groups--pathogens, non-pathogens, and intermediates--based on 16S rRNA phylogeny. Research on Leptospira is difficult due to slow growth and poor transformability of the pathogens. Recent identification of extrachromosomal elements besides the two chromosomes in L. interrogans has provided new insight into genome complexity of the genus Leptospira. The large size, low copy number, and high similarity of the sequence of these extrachromosomal elements with the chromosomes present challenges in isolating and detecting them without careful genome assembly. In this study, two extrachromosomal elements were identified in L. borgpetersenii serovar Ballum strain 56604 through whole genome assembly combined with S1 nuclease digestion following pulsed-field gel electrophoresis (S1-PFGE) analysis. Further, extrachromosomal elements in additional 15 Chinese epidemic strains of Leptospira, comprising L. borgpetersenii, L. weilii, and L. interrogans, were successfully separated and identified, independent of genome sequence data. Southern blot hybridization with extrachromosomal element-specific probes, designated as lcp1, lcp2 and lcp3-rep, further confirmed their occurrences as extrachromosomal elements. In total, 24 plasmids were detected in 13 out of 15 tested strains, among which 11 can hybridize with the lcp1-rep probe and 11 with the lcp2-rep probe, whereas two can hybridize with the lcp3-rep probe. None of them are likely to be species-specific. Blastp search of the lcp1, lcp2, and lcp3-rep genes with a nonredundant protein database of Leptospira species genomes showed that their homologous sequences are widely distributed among clades of pathogens but not non-pathogens or intermediates. These results suggest that the plasmids are widely distributed in Leptospira species, and further elucidation of their biological

  12. Comparative analyses of transport proteins encoded within the genomes of Leptospira species.

    PubMed

    Buyuktimkin, Bora; Saier, Milton H

    2016-09-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, Leptospira interrogans serovar Lai str. 56601 and Leptospira borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, Leptospira biflexa serovar Patoc str. 'Patoc 1 (Ames)'. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they all have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity arose in Leptospira correlating to progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. PMID:27296707

  13. High virulence in hamsters of four dominant Leptospira serovars isolated from rats in the Philippines.

    PubMed

    Villanueva, Sharon Y A M; Saito, Mitsumasa; Tsutsumi, Yutaka; Segawa, Takaya; Baterna, Rubelia A; Chakraborty, Antara; Asoh, Tatsuma; Miyahara, Satoshi; Yanagihara, Yasutake; Cavinta, Lolita L; Gloriani, Nina G; Yoshida, Shin-ichi

    2014-02-01

    Leptospirosis is caused by pathogenic species of Leptospira. The aim of this study was to determine and characterize the pathogenicity of four dominant Leptospira isolates prevailing among rats in the Philippines. The isolates were Leptospira interrogans serovar Manilae strain K64, L. interrogans serovar Losbanos strain K37, L. interrogans serovar Ratnapura strain K5 and Leptospira borgpetersenii serovar Javanica strain K6. Pathogenicities were studied using hamsters, which reproduce severe human leptospirosis. The minimum lethal doses were 10(0) ( = 1) leptospires for K64, K37 and K5, and 10(1) leptospires for K6. Weight loss amongst the Leptospira-infected hamsters was observed from 1 day before death (K64-, K37- and K5-infected hamsters) to as much as 1 week before death for K6-infected hamsters. Similar and varied gross and microscopic lesions were observed amongst infected hamsters, even for strains belonging to the same species (i.e. L. interrogans). The most significant and common histopathological findings were congestion of the glomerulus, disarrangement of hepatic cords and erythrophagocytosis. Other findings were foamy splenic macrophages for K6, severe petechial pulmonary haemorrhage for K64, and hematuria and severe pulmonary congestion for K37. Immunostaining and culture revealed the presence of leptospires in different organs of the infected hamsters. Based on these results, Leptospira isolates from rats in the Philippines were shown to be highly virulent, causing pulmonary haemorrhage, severe hepato-renal damage and death in hamsters even at lower doses. The present findings on experimental leptospirosis support clinical data showing that patients with severe manifestations of leptospirosis, such as pulmonary haemorrhage, are increasing in the Philippines. These findings may serve as a basis to strengthen the early diagnosis and treatment of human leptospirosis.

  14. Molecular and serological characterization of the first Leptospira santarosai strain isolated from a dog.

    PubMed

    Miotto, Bruno Alonso; Moreno, Luisa Zanolli; Guilloux, Aline Gil Alves; Sousa, Gisele Oliveira de; Loureiro, Ana Paula; Moreno, Andrea Micke; Lilenbaum, Walter; Vasconcellos, Silvio Arruda; Heinemann, Marcos Bryan; Hagiwara, Mitika Kuribayashi

    2016-10-01

    Leptospirosis is a zoonotic disease of global importance caused by pathogenic Leptospira species. Dogs can become asymptomatically infected, acting like reservoir hosts for pathogenic Leptospira, notably Leptospira interrogans serovar Canicola. Identification of such individuals and characterization of leptospires involved in chronic infections may unravel the role of dogs in the epidemiology of particular leptospiral strains. The aim of the present work was to describe the first Leptospira santarosai strain isolated from a dog. The dog was kept in a public shelter in São Paulo city, Brazil, and presented asymptomatic urinary shedding detected by PCR. Prospective evaluation was performed to fully characterize its chronic carrier state. The dog did not present anti-Leptospira titles or clinical/laboratorial abnormalities during the evaluations; nevertheless long-term urinary shedding was confirmed by PCR and leptospires were recovered from two occasions. The isolated strain was molecularly characterized by partial 16S rRNA and secY gene sequencing and MLST analysis. Serogroup identification was performed using polyclonal antibodies. The strain was identified as Leptospira santarosai, serogroup Sejroe. This is the first evidence in the literature of the isolation of L. santarosai in dogs. Our findings show that dogs can persistently harbor leptospires other than L. interrogans.

  15. Molecular and serological characterization of the first Leptospira santarosai strain isolated from a dog.

    PubMed

    Miotto, Bruno Alonso; Moreno, Luisa Zanolli; Guilloux, Aline Gil Alves; Sousa, Gisele Oliveira de; Loureiro, Ana Paula; Moreno, Andrea Micke; Lilenbaum, Walter; Vasconcellos, Silvio Arruda; Heinemann, Marcos Bryan; Hagiwara, Mitika Kuribayashi

    2016-10-01

    Leptospirosis is a zoonotic disease of global importance caused by pathogenic Leptospira species. Dogs can become asymptomatically infected, acting like reservoir hosts for pathogenic Leptospira, notably Leptospira interrogans serovar Canicola. Identification of such individuals and characterization of leptospires involved in chronic infections may unravel the role of dogs in the epidemiology of particular leptospiral strains. The aim of the present work was to describe the first Leptospira santarosai strain isolated from a dog. The dog was kept in a public shelter in São Paulo city, Brazil, and presented asymptomatic urinary shedding detected by PCR. Prospective evaluation was performed to fully characterize its chronic carrier state. The dog did not present anti-Leptospira titles or clinical/laboratorial abnormalities during the evaluations; nevertheless long-term urinary shedding was confirmed by PCR and leptospires were recovered from two occasions. The isolated strain was molecularly characterized by partial 16S rRNA and secY gene sequencing and MLST analysis. Serogroup identification was performed using polyclonal antibodies. The strain was identified as Leptospira santarosai, serogroup Sejroe. This is the first evidence in the literature of the isolation of L. santarosai in dogs. Our findings show that dogs can persistently harbor leptospires other than L. interrogans. PMID:27282095

  16. Hedgehogs and Mustelid Species: Major Carriers of Pathogenic Leptospira, a Survey in 28 Animal Species in France (20122015)

    PubMed Central

    Raton, Vincent; Zilber, Anne-Laure; Gasqui, Patrick; Faure, Eva; Baurier, Florence; Vourc’h, Gwenaël; Kodjo, Angeli; Combes, Benoît

    2016-01-01

    Human leptospirosis is a zoonotic and potentially fatal disease that has increasingly been reported in both developing and developed countries, including France. However, our understanding of the basic aspects of the epidemiology of this disease, including the source of Leptospira serogroup Australis infections in humans and domestic animals, remains incomplete. We investigated the genetic diversity of Leptospira in 28 species of wildlife other than rats using variable number tandem repeat (VNTR) and multispacer sequence typing (MST). The DNA of pathogenic Leptospira was detected in the kidney tissues of 201 individuals out of 3,738 tested individuals. A wide diversity, including 50 VNTR profiles and 8 MST profiles, was observed. Hedgehogs and mustelid species had the highest risk of being infected (logistic regression, OR = 66.8, CI95% = 30.9–144 and OR = 16.7, CI95% = 8.7–31.8, respectively). Almost all genetic profiles obtained from the hedgehogs were related to Leptospira interrogans Australis, suggesting the latter as a host-adapted bacterium, whereas mustelid species were infected by various genotypes, suggesting their interaction with Leptospira was different. By providing an inventory of the circulating strains of Leptospira and by pointing to hedgehogs as a potential reservoir of L. interrogans Australis, our study advances current knowledge on Leptospira animal carriers, and this information could serve to enhance epidemiological investigations in the future. PMID:27680672

  17. Transcriptional Response of Leptospira interrogans to Different Iron Sources

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospirosis is a globally important zoonotic disease. Humans can become infected via exposure to infected animals or contaminated water or soil. Iron is an essential element for many cellular processes and its sequestration in the host environment constitutes an immune defence mechanism. Pathoge...

  18. Transcriptional Response of Leptospira interrogans to Different Iron Sources

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Australian Bacterial Pathogenesis Program, Department of Microbiology, Monash University, Victoria 3800, Australia (1), Australian Research Council Centre of Excellence in Structural and Functional Microbial Genomics, Department of Microbiology, Monash University, Victoria 3800, Australia (2), Davi...

  19. Household Transmission of Leptospira Infection in Urban Slum Communities

    PubMed Central

    Maciel, Elves A. P.; de Carvalho, Ana Luiza F.; Nascimento, Simone F.; de Matos, Rosan B.; Gouveia, Edilane L.; Reis, Mitermayer G.; Ko, Albert I.

    2008-01-01

    Background Leptospirosis, a spirochaetal zoonotic disease, is the cause of epidemics associated with high mortality in urban slum communities. Infection with pathogenic Leptospira occurs during environmental exposures and is traditionally associated with occupational risk activities. However, slum inhabitants reside in close proximity to environmental sources of contamination, suggesting that transmission during urban epidemics occurs in the household environment. Methods and Findings A survey was performed to determine whether Leptospira infection clustered within households located in slum communities in the city of Salvador, Brazil. Hospital-based surveillance identified 89 confirmed cases of leptospirosis during an outbreak. Serum samples were obtained from members of 22 households with index cases of leptospirosis and 52 control households located in the same slum communities. The presence of anti-Leptospira agglutinating antibodies was used as a marker for previous infection. In households with index cases, 22 (30%) of 74 members had anti-Leptospira antibodies, whereas 16 (8%) of 195 members from control households had anti-Leptospira antibodies. Highest titres were directed against L. interrogans serovars of the Icterohaemorrhagiae serogroup in 95% and 100% of the subjects with agglutinating antibodies from case and control households, respectively. Residence in a household with an index case of leptospirosis was associated with increased risk (OR 5.29, 95% CI 2.13–13.12) of having had a Leptospira infection. Increased infection risk was found for all age groups who resided in a household with an index case, including children <15 years of age (P = 0.008). Conclusions This study identified significant household clustering of Leptospira infection in slum communities where recurrent epidemics of leptospirosis occur. The findings support the hypothesis that the household environment is an important transmission determinant in the urban slum setting

  20. The Characteristics of Ubiquitous and Unique Leptospira Strains from the Collection of Russian Centre for Leptospirosis

    PubMed Central

    Voronina, Olga L.; Kunda, Marina S.; Aksenova, Ekaterina I.; Ryzhova, Natalia N.; Semenov, Andrey N.; Petrov, Evgeny M.; Didenko, Lubov V.; Lunin, Vladimir G.; Ananyina, Yuliya V.; Gintsburg, Alexandr L.

    2014-01-01

    Background and Aim. Leptospira, the causal agent of leptospirosis, has been isolated from the environment, patients, and wide spectrum of animals in Russia. However, the genetic diversity of Leptospira in natural and anthropurgic foci was not clearly defined. Methods. The recent MLST scheme was used for the analysis of seven pathogenic species. 454 pyrosequencing technology was the base of the whole genome sequencing (WGS). Results. The most wide spread and prevalent Leptospira species in Russia were L. interrogans, L. kirschneri, and L. borgpetersenii. Five STs, common for Russian strains: 37, 17, 199, 110, and 146, were identified as having a longtime and ubiquitous distribution in various geographic areas. Unexpected properties were revealed for the environmental Leptospira strain Bairam-Ali. WGS of this strain genome suggested that it combined the features of the pathogenic and nonpathogenic strains and may be a reservoir of the natural resistance genes. Results of the comparative analysis of rrs and rpoB genes and MLST loci for different Leptospira species strains and phenotypic and serological properties of the strain Bairam-Ali suggested that it represented separate Leptospira species. Conclusions. Thus, the natural and anthropurgic foci supported ubiquitous Leptospira species and the pool of genes important for bacterial adaptivity to various conditions. PMID:25276806

  1. Diversification of an emerging pathogen in a biodiversity hotspot: Leptospira in endemic small mammals of Madagascar.

    PubMed

    Dietrich, Muriel; Wilkinson, David A; Soarimalala, Voahangy; Goodman, Steven M; Dellagi, Koussay; Tortosa, Pablo

    2014-06-01

    Biodiversity hotspots and associated endemism are ideal systems for the study of parasite diversity within host communities. Here, we investigated the ecological and evolutionary forces acting on the diversification of an emerging bacterial pathogen, Leptospira spp., in communities of endemic Malagasy small mammals. We determined the infection rate with pathogenic Leptospira in 20 species of sympatric rodents (subfamily Nesomyinae) and tenrecids (family Tenrecidae) at two eastern humid forest localities. A multilocus genotyping analysis allowed the characterization of bacterial diversity within small mammals and gave insights into their genetic relationships with Leptospira infecting endemic Malagasy bats (family Miniopteridae and Vespertilionidae). We report for the first time the presence of pathogenic Leptospira in Malagasy endemic small mammals, with an overall prevalence of 13%. In addition, these hosts harbour species of Leptospira (L. kirschneri, L. borgpetersenii and L. borgpetersenii group B) which are different from those reported in introduced rats (L. interrogans) on Madagascar. The diversification of Leptospira on Madagascar can be traced millions of years into evolutionary history, resulting in the divergence of endemic lineages and strong host specificity. These observations are discussed in relation to the relative roles of endemic vs. introduced mammal species in the evolution and epidemiology of Leptospira on Madagascar, specifically how biodiversity and biogeographical processes can shape community ecology of an emerging pathogen and lead to its diversification within native animal communities.

  2. Improvement of trivalent leptospira vaccine by removal of anaphylactic agents.

    PubMed

    Moazenijula, Gholamreza; Jabbari, A R; Geravand, M Moradi; Banihashemi, R; Hajizadeh, A

    2011-12-01

    The anaphylactic reactions in cattle following leptospira vaccination mostly booster dose in different parts of Iran have been reported. The serum proteins as allergic substances are components of liquid phase of the vaccine. Therefore, the vaccine was modified by washing the whole cultures by centrifugations. The modified vaccine was safe in laboratory animals and cattle as well as under field conditions. Microagglutination test revealed a similar pattern of antibody response to the three Leptospira interrogans serovars (Canicola, Grippotyphosa, and Sejro hardjo) in all vaccinated cattle groups while was higher than the response of control animals. The results of the present investigation revealed that we can minimize postvaccination shock in vaccinated cattle populations with removing the shock proteins. PMID:21698521

  3. Functional thioredoxin reductase from pathogenic and free-living Leptospira spp.

    PubMed

    Sasoni, Natalia; Iglesias, Alberto A; Guerrero, Sergio A; Arias, Diego G

    2016-08-01

    Low molecular mass thiols and antioxidant enzymes have essential functions to detoxify reactive oxygen and nitrogen species maintaining cellular redox balance. The metabolic pathways for redox homeostasis in pathogenic (Leptospira interrogans) and free-living (Leptospira biflexa) leptospires species were not functionally characterized. We performed biochemical studies on recombinantly produced proteins to in depth analyze kinetic and structural properties of thioredoxin reductase (LinTrxR) and thioredoxin (LinTrx) from L. interrogans, and two TrxRs (LbiTrxR1 and LbiTrxR2) from L. biflexa. All the TrxRs were characterized as homodimeric flavoproteins, with LinTrxR and LbiTrxR1 catalyzing the NADPH dependent reduction of LinTrx and DTNB. The thioredoxin system from L. interrogans was able to use glutathione disulfide, lipoamide disulfide, cystine and bis-γ-glutamyl cysteine and homologous peroxiredoxin as substrates. Classic TrxR activity of LinTrxR2 had not been evidenced in vitro, but recombinant Escherichia coli cells overexpressing LbiTrxR2 showed high tolerance to oxidative stress. The enzymatic systems herein characterized could play a key role for the maintenance of redox homeostasis and the function of defense mechanisms against reactive oxidant species in Leptospira spp. Our results contribute to the general knowledge about redox biochemistry in these bacteria, positioning TrxR as a critical molecular target for the development of new anti-leptospiral drugs. PMID:27178006

  4. Functional thioredoxin reductase from pathogenic and free-living Leptospira spp.

    PubMed

    Sasoni, Natalia; Iglesias, Alberto A; Guerrero, Sergio A; Arias, Diego G

    2016-08-01

    Low molecular mass thiols and antioxidant enzymes have essential functions to detoxify reactive oxygen and nitrogen species maintaining cellular redox balance. The metabolic pathways for redox homeostasis in pathogenic (Leptospira interrogans) and free-living (Leptospira biflexa) leptospires species were not functionally characterized. We performed biochemical studies on recombinantly produced proteins to in depth analyze kinetic and structural properties of thioredoxin reductase (LinTrxR) and thioredoxin (LinTrx) from L. interrogans, and two TrxRs (LbiTrxR1 and LbiTrxR2) from L. biflexa. All the TrxRs were characterized as homodimeric flavoproteins, with LinTrxR and LbiTrxR1 catalyzing the NADPH dependent reduction of LinTrx and DTNB. The thioredoxin system from L. interrogans was able to use glutathione disulfide, lipoamide disulfide, cystine and bis-γ-glutamyl cysteine and homologous peroxiredoxin as substrates. Classic TrxR activity of LinTrxR2 had not been evidenced in vitro, but recombinant Escherichia coli cells overexpressing LbiTrxR2 showed high tolerance to oxidative stress. The enzymatic systems herein characterized could play a key role for the maintenance of redox homeostasis and the function of defense mechanisms against reactive oxidant species in Leptospira spp. Our results contribute to the general knowledge about redox biochemistry in these bacteria, positioning TrxR as a critical molecular target for the development of new anti-leptospiral drugs.

  5. Genotypes of Leptospira spp. strains isolated from dogs in Buenos Aires, Argentina.

    PubMed

    Grune Loffler, Sylvia; Passaro, Diego; Samartino, Luis; Soncini, Analía; Romero, Graciela; Brihuega, Bibiana

    2014-01-01

    Leptospirosis is an infectious disease of wide global distribution, which is endemic in Argentina. The objective of this study was to obtain the genetic profiles of Leptospira spp. strains isolated from clinical cases of dogs in the province of Buenos Aires by the multiple-locus variable-number tandem repeat analysis (MLVA). Eight isolated canine strains were genotyped by MLVA, obtaining the identical profile of Leptospira interrogans serovar Canicola Hond Utrecht IV in the strains named Dogy and Mayo. The strains named Bel, Sarmiento, La Plata 4581 and La Plata 5478 were identical to the profile of the genotype of L. interrogans serovar Portlandvere MY 1039.The strain named Avellaneda was identical to the genotype profile of L. interrogans serovar Icterohaemorrhagiae RGA and the strain named SB had the same profile as the L. interrogans serovar Pomona Baires genotype and was similar to the profile of serovar Pomona Pomona genotype. It would be useful to include a larger number of isolates from different dog populations in various provinces of Argentina and to characterize the genetic profiles of the strains circulating in the country. The information obtained will be useful for the control of leptospirosis in the dog population.

  6. Genotypes of Leptospira spp. strains isolated from dogs in Buenos Aires, Argentina.

    PubMed

    Grune Loffler, Sylvia; Passaro, Diego; Samartino, Luis; Soncini, Analía; Romero, Graciela; Brihuega, Bibiana

    2014-01-01

    Leptospirosis is an infectious disease of wide global distribution, which is endemic in Argentina. The objective of this study was to obtain the genetic profiles of Leptospira spp. strains isolated from clinical cases of dogs in the province of Buenos Aires by the multiple-locus variable-number tandem repeat analysis (MLVA). Eight isolated canine strains were genotyped by MLVA, obtaining the identical profile of Leptospira interrogans serovar Canicola Hond Utrecht IV in the strains named Dogy and Mayo. The strains named Bel, Sarmiento, La Plata 4581 and La Plata 5478 were identical to the profile of the genotype of L. interrogans serovar Portlandvere MY 1039.The strain named Avellaneda was identical to the genotype profile of L. interrogans serovar Icterohaemorrhagiae RGA and the strain named SB had the same profile as the L. interrogans serovar Pomona Baires genotype and was similar to the profile of serovar Pomona Pomona genotype. It would be useful to include a larger number of isolates from different dog populations in various provinces of Argentina and to characterize the genetic profiles of the strains circulating in the country. The information obtained will be useful for the control of leptospirosis in the dog population. PMID:25444128

  7. Genotypes of pathogenic Leptospira spp isolated from rodents in Argentina

    PubMed Central

    Loffler, Sylvia Grune; Pavan, Maria Elisa; Vanasco, Bibiana; Samartino, Luis; Suarez, Olga; Auteri, Carmelo; Romero, Graciela; Brihuega, Bibiana

    2014-01-01

    Leptospirosis is the most widespread zoonosis in the world and significant efforts have been made to determine and classify pathogenic Leptospira strains. This zoonosis is maintained in nature through chronic renal infections of carrier animals, with rodents and other small mammals serving as the most important reservoirs. Additionally, domestic animals, such as livestock and dogs, are significant sources of human infection. In this study, a multiple-locus variable-number tandem repeat analysis (MLVA) was applied to genotype 22 pathogenic Leptospira strains isolated from urban and periurban rodent populations from different regions of Argentina. Three MLVA profiles were identified in strains belonging to the species Leptospira interrogans (serovars Icterohaemorrhagiae and Canicola); one profile was observed in serovar Icterohaemorrhagiae and two MLVA profiles were observed in isolates of serovars Canicola and Portlandvere. All strains belonging to Leptospira borgpetersenii serovar Castellonis exhibited the same MLVA profile. Four different genotypes were isolated from urban populations of rodents, including both mice and rats and two different genotypes were isolated from periurban populations. PMID:24676656

  8. Genotypes of pathogenic Leptospira spp isolated from rodents in Argentina.

    PubMed

    Loffler, Sylvia Grune; Pavan, Maria Elisa; Vanasco, Bibiana; Samartino, Luis; Suarez, Olga; Auteri, Carmelo; Romero, Graciela; Brihuega, Bibiana

    2014-04-01

    Leptospirosis is the most widespread zoonosis in the world and significant efforts have been made to determine and classify pathogenic Leptospira strains. This zoonosis is maintained in nature through chronic renal infections of carrier animals, with rodents and other small mammals serving as the most important reservoirs. Additionally, domestic animals, such as livestock and dogs, are significant sources of human infection. In this study, a multiple-locus variable-number tandem repeat analysis (MLVA) was applied to genotype 22 pathogenic Leptospira strains isolated from urban and periurban rodent populations from different regions of Argentina. Three MLVA profiles were identified in strains belonging to the species Leptospira interrogans (serovars Icterohaemorrhagiae and Canicola); one profile was observed in serovar Icterohaemorrhagiae and two MLVA profiles were observed in isolates of serovars Canicola and Portlandvere. All strains belonging to Leptospira borgpetersenii serovar Castellonis exhibited the same MLVA profile. Four different genotypes were isolated from urban populations of rodents, including both mice and rats and two different genotypes were isolated from periurban populations. PMID:24676656

  9. Though not reservoirs, dogs might transmit Leptospira in New Caledonia.

    PubMed

    Gay, Noellie; Soupé-Gilbert, Marie-Estelle; Goarant, Cyrille

    2014-04-01

    Leptospira has been a major public health concern in New Caledonia for decades. However, few multidisciplinary studies addressing the zoonotic pattern of this disease were conducted so far. Here, pig, deer and dog samples were collected. Analyses were performed using molecular detection and genotyping. Serological analyses were also performed for dogs. Our results suggest that deer are a reservoir of L. borgpetersenii Hardjobovis and pigs a reservoir of L. interrogans Pomona. Interestingly, 4.4% of dogs were renal carriers of Leptospira. In dog populations, MAT results confirmed the circulation of the same Leptospira serogroups involved in human cases. Even if not reservoirs, dogs might be of significance in human contamination by making an epidemiological link between wild or feral reservoirs and humans. Dogs could bring pathogens back home, shedding Leptospira via their urine and in turn increasing the risk of human contamination. We propose to consider dog as a vector, particularly in rural areas where seroprevalence is significantly higher than urban areas. Our results highlight the importance of animal health in improving leptospirosis prevention in a One Health approach. PMID:24747539

  10. Genotyping of Leptospira directly in urine samples of cattle demonstrates a diversity of species and strains in Brazil.

    PubMed

    Hamond, C; Pestana, C P; Medeiros, M A; Lilenbaum, W

    2016-01-01

    The aim of this study was to identify Leptospira in urine samples of cattle by direct sequencing of the secY gene. The validity of this approach was assessed using ten Leptospira strains obtained from cattle in Brazil and 77 DNA samples previously extracted from cattle urine, that were positive by PCR for the genus-specific lipL32 gene of Leptospira. Direct sequencing identified 24 (31·1%) interpretable secY sequences and these were identical to those obtained from direct DNA sequencing of the urine samples from which they were recovered. Phylogenetic analyses identified four species: L. interrogans, L. borgpetersenii, L. noguchii, and L. santarosai with the most prevalent genotypes being associated with L. borgpetersenii. While direct sequencing cannot, as yet, replace culturing of leptospires, it is a valid additional tool for epidemiological studies. An unexpected finding from this study was the genetic diversity of Leptospira infecting Brazilian cattle.

  11. Detection of virulence factors and molecular typing of pathogenic Leptospira from capybara (Hydrochaeris hydrochaeris).

    PubMed

    Jorge, Sérgio; Monte, Leonardo G; Coimbra, Marco Antonio; Albano, Ana Paula; Hartwig, Daiane D; Lucas, Caroline; Seixas, Fabiana K; Dellagostin, Odir A; Hartleben, Cláudia P

    2012-10-01

    Leptospirosis is a globally prevalent zoonosis caused by pathogenic Leptospira spp.; several serologic variants have reservoirs in synanthropic rodents. The capybara is the largest living rodent in the world, and it has a wide geographical distribution in Central and South America. This rodent is a significant source of Leptospira since the agent is shed via urine into the environment and is a potential public health threat. In this study, we isolated and identified by molecular techniques a pathogenic Leptospira from capybara in southern Brazil. The isolated strain was characterized by partial rpoB gene sequencing and variable-number tandem-repeats analysis as L. interrogans, serogroup Icterohaemorrhagiae. In addition, to confirm the expression of virulence factors, the bacterial immunoglobulin-like proteins A and B expression was detected by indirect immunofluorescence using leptospiral specific monoclonal antibodies. This report identifies capybaras as an important source of infection and provides insight into the epidemiology of leptospirosis.

  12. Rapid Isolation and Susceptibility Testing of Leptospira spp. Using a New Solid Medium, LVW Agar

    PubMed Central

    Wuthiekanun, Vanaporn; Amornchai, Premjit; Paris, Daniel H.; Langla, Sayan; Thaipadunpanit, Janjira; Chierakul, Wirongrong; Smythe, Lee D.; White, Nicholas J.; Day, Nicholas P. J.; Peacock, Sharon J.

    2013-01-01

    Pathogenic Leptospira spp., the causative agents of leptospirosis, are slow-growing Gram-negative spirochetes. Isolation of Leptospira from clinical samples and testing of antimicrobial susceptibility are difficult and time-consuming. Here, we describe the development of a new solid medium that facilitates more-rapid growth of Leptospira spp. and the use of this medium to evaluate the Etest's performance in determining antimicrobial MICs to drugs in common use for leptospirosis. The medium was developed by evaluating the effects of numerous factors on the growth rate of Leptospira interrogans strain NR-20157. These included the type of base agar, the concentration of rabbit serum (RS), and the concentration and duration of CO2 incubation during the initial period of culture. The highest growth rate of NR-20157 was achieved using a Noble agar base supplemented with 10% RS (named LVW agar), with an initial incubation at 30°C in 5% CO2 for 2 days prior to continuous culture in air at 30°C. These conditions were used to develop the Etest for three species, L. interrogans (NR-20161), L. kirschnerii (NR-20327), and L. borgpetersenii (NR-20151). The MICs were read on day 7 for all samples. The Etest was then performed on 109 isolates of pathogenic Leptospira spp. The MIC90 values for penicillin G, doxycycline, cefotaxime, ceftriaxone, and chloramphenicol were 0.64 units/ml and 0.19, 0.047, 0.5, and 2 μg/ml, respectively. The use of LVW agar, which enables rapid growth, isolation of single colonies, and simple antimicrobial susceptibility testing for Leptospira spp., provides an opportunity for new areas of fundamental and applied research. PMID:23114772

  13. Similarities in Leptospira Serogroup and Species Distribution in Animals and Humans in the Indian Ocean Island of Mayotte

    PubMed Central

    Desvars, Amélie; Naze, Florence; Vourc'h, Gwenaël; Cardinale, Eric; Picardeau, Mathieu; Michault, Alain; Bourhy, Pascale

    2012-01-01

    Our objective was to identify local animal reservoirs of leptospirosis to explain the unusual features of Leptospira strains recently described among patients on the island of Mayotte. By means of a microscopic agglutination test using local clinical isolates, we found that 11.2% of black rats were seropositive to Leptospira, whereas 10.2% of flying foxes, 2% of lemurs, 93.1% of domestic dogs, and 87.5% of stray dogs were seropositive. As observed in humans, Mini was the main serogroup circulating in animals, whereas serogroup Icterohaemorrhagiae was absent. Using quantitative polymerase chain reaction, we also showed that 29.8% of rats carried leptospires in their kidneys. The sequencing of 16S rRNA gene sequences of Leptospira found in black rat kidneys identified four genomospecies (Leptospira borgpetersenii, Leptospira interrogans, Leptospira kirschneri, and L. borgpetersenii group B), which established black rats as the major source of leptospirosis transmission to humans. The origins of such a genetic diversity in Leptospira strains are discussed. PMID:22764304

  14. Interaction of Bovine Peripheral Blood Polymorphonuclear Cells and Leptospira Species; Innate Responses in the Natural Bovine Reservoir Host

    PubMed Central

    Wilson-Welder, Jennifer H.; Frank, Ami T.; Hornsby, Richard L.; Olsen, Steven C.; Alt, David P.

    2016-01-01

    Cattle are the reservoir hosts of Leptospira borgpetersenii serovar Hardjo, and can also be reservoir hosts of other Leptospira species such as L. kirschneri, and Leptospira interrogans. As a reservoir host, cattle shed Leptospira, infecting other animals, including humans. Previous studies with human and murine neutrophils have shown activation of neutrophil extracellular trap or NET formation, and upregulation of inflammatory mediators by neutrophils in the presence of Leptospira. Humans, companion animals and most widely studied models of Leptospirosis are of acute infection, hallmarked by systemic inflammatory response, neutrophilia, and septicemia. In contrast, cattle exhibit chronic infection with few outward clinical signs aside from reproductive failure. Taking into consideration that there is host species variation in innate immunity, especially in pathogen recognition and response, the interaction of bovine peripheral blood polymorphonuclear cells (PMNs) and several Leptospira strains was evaluated. Studies including bovine-adapted strains, human pathogen strains, a saprophyte and inactivated organisms. Incubation of PMNs with Leptospira did induce slight activation of neutrophil NETs, greater than unstimulated cells but less than the quantity from E. coli P4 stimulated PMNs. Very low but significant from non-stimulated, levels of reactive oxygen peroxides were produced in the presence of all Leptospira strains and E. coli P4. Similarly, significant levels of reactive nitrogen intermediaries (NO2) was produced from PMNs when incubated with the Leptospira strains and greater quantities in the presence of E. coli P4. PMNs incubated with Leptospira induced RNA transcripts of IL-1β, MIP-1α, and TNF-α, with greater amounts induced by live organisms when compared to heat-inactivated leptospires. Transcript for inflammatory cytokine IL-8 was also induced, at similar levels regardless of Leptospira strain or viability. However, incubation of Leptospira strains

  15. Interaction of Bovine Peripheral Blood Polymorphonuclear Cells and Leptospira Species; Innate Responses in the Natural Bovine Reservoir Host.

    PubMed

    Wilson-Welder, Jennifer H; Frank, Ami T; Hornsby, Richard L; Olsen, Steven C; Alt, David P

    2016-01-01

    Cattle are the reservoir hosts of Leptospira borgpetersenii serovar Hardjo, and can also be reservoir hosts of other Leptospira species such as L. kirschneri, and Leptospira interrogans. As a reservoir host, cattle shed Leptospira, infecting other animals, including humans. Previous studies with human and murine neutrophils have shown activation of neutrophil extracellular trap or NET formation, and upregulation of inflammatory mediators by neutrophils in the presence of Leptospira. Humans, companion animals and most widely studied models of Leptospirosis are of acute infection, hallmarked by systemic inflammatory response, neutrophilia, and septicemia. In contrast, cattle exhibit chronic infection with few outward clinical signs aside from reproductive failure. Taking into consideration that there is host species variation in innate immunity, especially in pathogen recognition and response, the interaction of bovine peripheral blood polymorphonuclear cells (PMNs) and several Leptospira strains was evaluated. Studies including bovine-adapted strains, human pathogen strains, a saprophyte and inactivated organisms. Incubation of PMNs with Leptospira did induce slight activation of neutrophil NETs, greater than unstimulated cells but less than the quantity from E. coli P4 stimulated PMNs. Very low but significant from non-stimulated, levels of reactive oxygen peroxides were produced in the presence of all Leptospira strains and E. coli P4. Similarly, significant levels of reactive nitrogen intermediaries (NO2) was produced from PMNs when incubated with the Leptospira strains and greater quantities in the presence of E. coli P4. PMNs incubated with Leptospira induced RNA transcripts of IL-1β, MIP-1α, and TNF-α, with greater amounts induced by live organisms when compared to heat-inactivated leptospires. Transcript for inflammatory cytokine IL-8 was also induced, at similar levels regardless of Leptospira strain or viability. However, incubation of Leptospira strains

  16. Evaluation of Cell Binding Activities of Leptospira ECM Adhesins

    PubMed Central

    Robbins, Gregory T.; Hahn, Beth L.; Evangelista, Karen V.; Padmore, Lavinia; Aranda, Patrick S.; Coburn, Jenifer

    2015-01-01

    Pathogenic spirochetes of the genus Leptospira are the causative agents of leptospirosis, a zoonotic infection that occurs globally. The bacteria colonize the renal proximal tubules of many animals and are shed in the urine. Contact with the urine, or with water contaminated with the urine of infected animals can cause infection of new host animals, including humans. Mechanisms of colonization of the proximal tubule and other tissues are not known, but specific interactions between bacterial adhesins and host substrates are likely to be critical in this process. Several extracellular matrix (ECM) adhesins have been previously identified, but more recently, it has been shown that Leptospira bind more efficiently to cells than ECM. In this work, recombinant forms of five putative Leptospira ECM adhesins, namely LipL32, Loa22, OmpL1, p31/LipL45, and LenA were evaluated for binding to cells as well as an expanded variety of ECM components. Reproducible and significant adhesin activity was demonstrated only for OmpL1, which bound to both mammalian cell lines tested and to glycosaminoglycans (GAGs). While determination of biologically significant bacterial adhesion activity will require generation of site-directed mutant strains, our results suggest that OmpL1 is a strong candidate for future evaluation regarding the roles of the adhesin activity of the protein during L. interrogans infection. PMID:25875373

  17. Leptospira spp. in Rodents and Shrews in Germany

    PubMed Central

    Mayer-Scholl, Anne; Hammerl, Jens Andre; Schmidt, Sabrina; Ulrich, Rainer G.; Pfeffer, Martin; Woll, Dietlinde; Scholz, Holger C.; Thomas, Astrid; Nöckler, Karsten

    2014-01-01

    Leptospirosis is an acute, febrile disease occurring in humans and animals worldwide. Leptospira spp. are usually transmitted through direct or indirect contact with the urine of infected reservoir animals. Among wildlife species, rodents act as the most important reservoir for both human and animal infection. To gain a better understanding of the occurrence and distribution of pathogenic leptospires in rodent and shrew populations in Germany, kidney specimens of 2973 animals from 11 of the 16 federal states were examined by PCR. Rodent species captured included five murine species (family Muridae), six vole species (family Cricetidae) and six shrew species (family Soricidae). The most abundantly trapped animals were representatives of the rodent species Apodemus flavicollis, Clethrionomys glareolus and Microtus agrestis. Leptospiral DNA was amplified in 10% of all animals originating from eight of the 11 federal states. The highest carrier rate was found in Microtus spp. (13%), followed by Apodemus spp. (11%) and Clethrionomys spp. (6%). The most common Leptospira genomospecies determined by duplex PCR was L. kirschneri, followed by L. interrogans and L. borgpetersenii; all identified by single locus sequence typing (SLST). Representatives of the shrew species were also carriers of Leptospira spp. In 20% of Crocidura spp. and 6% of the Sorex spp. leptospiral DNA was detected. Here, only the pathogenic genomospecies L. kirschneri was identified. PMID:25062275

  18. Development of Leptospira in vitro potency assays: EU/industry experience and perspectives.

    PubMed

    Klaasen, H L B M; van der Veen, M; Molkenboer, M J C H; Bruderer, U

    2013-09-01

    Nobivac® Lepto (MSD Animal Health) is a non-adjuvanted canine leptospirosis vaccine containing inactivated whole cells of Leptospira interrogans serogroup Canicola serovar Portlandvere and L. interrogans serogroup Icterohaemorrhagiae serovar Copenhageni. The current standard in vivo potency test is a hamster challenge test associated with major drawbacks such as animal suffering and poor reproducibility. Here, the quantification of antigenic mass by ELISA as a new in vitro potency test is described, supporting the 3Rs concept (replacement, reduction, and refinement of animal tests) and in accordance with European Pharmacopoeia Monograph 0447 (Canine Leptospirosis Vaccine [Inactivated]). The two corresponding sandwich ELISAs are based on monoclonal antibodies specific for immunodominant leptospiral lipopolysaccharide epitopes. Protection in passive immunization experiments demonstrate that these monoclonal antibodies recognize key protective antigens in currently licensed human and veterinary whole cell Leptospira vaccines. The high precision and robustness renders the two ELISAs much more reliable correlates of potency in dogs than the hamster potency test. The recent approval of these assays for a new canine leptospirosis vaccine is an important contribution to the 3Rs in quality control testing of Leptospira vaccines. PMID:23867758

  19. Comparative Genomic Analyses of Transport Proteins Encoded Within the Genomes of Leptospira Species

    PubMed Central

    Buyuktimkin, Bora; Saier, Milton H.

    2015-01-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, L. interrogans serovar Lai str. 56601 and L. borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, L. biflexa serovar Patoc str. ‘Patoc 1 (Ames)’. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity was accompanied by progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. PMID:26247102

  20. Human Leptospirosis Caused by a New, Antigenically Unique Leptospira Associated with a Rattus Species Reservoir in the Peruvian Amazon

    PubMed Central

    Cespedes, Manuel; Diaz, M. Monica; Galloway, Renee L.; Saito, Mayuko; Steigerwalt, Arnold G.; Patra, Kailash P.; Ore, Carlos Vidal; Gotuzzo, Eduardo; Gilman, Robert H.; Levett, Paul N.; Vinetz, Joseph M.

    2008-01-01

    As part of a prospective study of leptospirosis and biodiversity of Leptospira in the Peruvian Amazon, a new Leptospira species was isolated from humans with acute febrile illness. Field trapping identified this leptospire in peridomestic rats (Rattus norvegicus, six isolates; R. rattus, two isolates) obtained in urban, peri-urban, and rural areas of the Iquitos region. Novelty of this species was proven by serological typing, 16S ribosomal RNA gene sequencing, pulsed-field gel electrophoresis, and DNA-DNA hybridization analysis. We have named this species “Leptospira licerasiae” serovar Varillal, and have determined that it is phylogenetically related to, but genetically distinct from, other intermediate Leptospira such as L. fainei and L. inadai. The type strain is serovar Varillal strain VAR 010T, which has been deposited into internationally accessible culture collections. By microscopic agglutination test, “Leptospira licerasiae” serovar Varillal was antigenically distinct from all known serogroups of Leptospira except for low level cross-reaction with rabbit anti–L. fainei serovar Hurstbridge at a titer of 1∶100. LipL32, although not detectable by PCR, was detectable in “Leptospira licerasiae” serovar Varillal by both Southern blot hybridization and Western immunoblot, although on immunoblot, the predicted protein was significantly smaller (27 kDa) than that of L. interrogans and L. kirschneri (32 kDa). Isolation was rare from humans (2/45 Leptospira isolates from 881 febrile patients sampled), but high titers of MAT antibodies against “Leptospira licerasiae” serovar Varillal were common (30%) among patients fulfilling serological criteria for acute leptospirosis in the Iquitos region, and uncommon (7%) elsewhere in Peru. This new leptospiral species reflects Amazonian biodiversity and has evolved to become an important cause of leptospirosis in the Peruvian Amazon. PMID:18382606

  1. Use of PCR to identify Leptospira in kidneys of big brown bats (Eptesicus fuscus) in Kansas and Nebraska, USA.

    PubMed

    Harkin, Kenneth R; Hays, Michael; Davis, Rolan; Moore, Michael

    2014-07-01

    Bats have been implicated as potential carriers of Leptospira as a result of surveys, mostly in Australia and South America. We measured the prevalence of pathogenic leptospires in kidneys of bats from Kansas and Nebraska. From 7 August 2012 to 21 August 2012, we extracted DNA from kidneys of 98 big brown bats (Eptesicus fuscus) submitted and found negative for rabies. The DNA was processed in a two-step, seminested PCR assay with a dual-labeled Taqman probe specific for pathogenic leptospires. As a negative control, we used a saprophytic leptospire (Leptospira biflexa Patoc) and, as a pathogenic control, Leptospira interrogans Canicola. All bat kidneys were negative for pathogenic leptospires, suggesting that it is unlikely that the big brown bat, one of the most prevalent bat species in North America, is a reservoir for transmission of leptospires to dogs or humans. PMID:24807182

  2. Prevalence and Genotype Allocation of Pathogenic Leptospira Species in Small Mammals from Various Habitat Types in Germany.

    PubMed

    Obiegala, Anna; Woll, Dietlinde; Karnath, Carolin; Silaghi, Cornelia; Schex, Susanne; Eßbauer, Sandra; Pfeffer, Martin

    2016-03-01

    Small mammals serve as most important reservoirs for Leptospira spp., the causative agents of Leptospirosis, which is one of the most neglected and widespread zoonotic diseases worldwide. The knowledge about Leptospira spp. occurring in small mammals from Germany is scarce. Thus, this study's objectives were to investigate the occurrence of Leptospira spp. and the inherent sequence types in small mammals from three different study sites: a forest in southern Germany (site B1); a National Park in south-eastern Germany (site B2) and a renaturalised area, in eastern Germany (site S) where small mammals were captured. DNA was extracted from kidneys of small mammals and tested for Leptospira spp. by real-time PCR. Positive samples were further analysed by duplex and conventional PCRs. For 14 positive samples, multi locus sequence typing (MLST) was performed. Altogether, 1213 small mammals were captured: 216 at site B1, 456 at site B2 and 541 at site S belonging to following species: Sorex (S.) araneus, S. coronatus, Apodemus (A.) flavicollis, Myodes glareolus, Microtus (Mi.) arvalis, Crocidura russula, Arvicola terrestris, A. agrarius, Mustela nivalis, Talpa europaea, and Mi. agrestis. DNA of Leptospira spp. was detected in 6% of all small mammals. At site B1, 25 small mammals (11.6%), at site B2, 15 small mammals (3.3%) and at site S, 33 small mammals (6.1%) were positive for Leptospira spp. Overall, 54 of the positive samples were further determined as L. kirschneri, nine as L. interrogans and four as L. borgpetersenii while five real-time PCR-positive samples could not be further determined by conventional PCR. MLST results revealed focal occurrence of L. interrogans and L. kirschneri sequence type (ST) 117 while L. kirschneri ST 110 was present in small mammals at all three sites. Further, this study provides evidence for a particular host association of L. borgpetersenii to mice of the genus Apodemus. PMID:27015596

  3. Prevalence and Genotype Allocation of Pathogenic Leptospira Species in Small Mammals from Various Habitat Types in Germany

    PubMed Central

    Karnath, Carolin; Silaghi, Cornelia; Schex, Susanne; Eßbauer, Sandra; Pfeffer, Martin

    2016-01-01

    Small mammals serve as most important reservoirs for Leptospira spp., the causative agents of Leptospirosis, which is one of the most neglected and widespread zoonotic diseases worldwide. The knowledge about Leptospira spp. occurring in small mammals from Germany is scarce. Thus, this study’s objectives were to investigate the occurrence of Leptospira spp. and the inherent sequence types in small mammals from three different study sites: a forest in southern Germany (site B1); a National Park in south-eastern Germany (site B2) and a renaturalised area, in eastern Germany (site S) where small mammals were captured. DNA was extracted from kidneys of small mammals and tested for Leptospira spp. by real-time PCR. Positive samples were further analysed by duplex and conventional PCRs. For 14 positive samples, multi locus sequence typing (MLST) was performed. Altogether, 1213 small mammals were captured: 216 at site B1, 456 at site B2 and 541 at site S belonging to following species: Sorex (S.) araneus, S. coronatus, Apodemus (A.) flavicollis, Myodes glareolus, Microtus (Mi.) arvalis, Crocidura russula, Arvicola terrestris, A. agrarius, Mustela nivalis, Talpa europaea, and Mi. agrestis. DNA of Leptospira spp. was detected in 6% of all small mammals. At site B1, 25 small mammals (11.6%), at site B2, 15 small mammals (3.3%) and at site S, 33 small mammals (6.1%) were positive for Leptospira spp. Overall, 54 of the positive samples were further determined as L. kirschneri, nine as L. interrogans and four as L. borgpetersenii while five real-time PCR-positive samples could not be further determined by conventional PCR. MLST results revealed focal occurrence of L. interrogans and L. kirschneri sequence type (ST) 117 while L. kirschneri ST 110 was present in small mammals at all three sites. Further, this study provides evidence for a particular host association of L. borgpetersenii to mice of the genus Apodemus. PMID:27015596

  4. Prevalence and Genotype Allocation of Pathogenic Leptospira Species in Small Mammals from Various Habitat Types in Germany.

    PubMed

    Obiegala, Anna; Woll, Dietlinde; Karnath, Carolin; Silaghi, Cornelia; Schex, Susanne; Eßbauer, Sandra; Pfeffer, Martin

    2016-03-01

    Small mammals serve as most important reservoirs for Leptospira spp., the causative agents of Leptospirosis, which is one of the most neglected and widespread zoonotic diseases worldwide. The knowledge about Leptospira spp. occurring in small mammals from Germany is scarce. Thus, this study's objectives were to investigate the occurrence of Leptospira spp. and the inherent sequence types in small mammals from three different study sites: a forest in southern Germany (site B1); a National Park in south-eastern Germany (site B2) and a renaturalised area, in eastern Germany (site S) where small mammals were captured. DNA was extracted from kidneys of small mammals and tested for Leptospira spp. by real-time PCR. Positive samples were further analysed by duplex and conventional PCRs. For 14 positive samples, multi locus sequence typing (MLST) was performed. Altogether, 1213 small mammals were captured: 216 at site B1, 456 at site B2 and 541 at site S belonging to following species: Sorex (S.) araneus, S. coronatus, Apodemus (A.) flavicollis, Myodes glareolus, Microtus (Mi.) arvalis, Crocidura russula, Arvicola terrestris, A. agrarius, Mustela nivalis, Talpa europaea, and Mi. agrestis. DNA of Leptospira spp. was detected in 6% of all small mammals. At site B1, 25 small mammals (11.6%), at site B2, 15 small mammals (3.3%) and at site S, 33 small mammals (6.1%) were positive for Leptospira spp. Overall, 54 of the positive samples were further determined as L. kirschneri, nine as L. interrogans and four as L. borgpetersenii while five real-time PCR-positive samples could not be further determined by conventional PCR. MLST results revealed focal occurrence of L. interrogans and L. kirschneri sequence type (ST) 117 while L. kirschneri ST 110 was present in small mammals at all three sites. Further, this study provides evidence for a particular host association of L. borgpetersenii to mice of the genus Apodemus.

  5. Prevalence of antibodies against Leptospira sp. in snakes, lizards and turtles in Slovenia

    PubMed Central

    2013-01-01

    Background Leptospiral infections in poikilothermic (cold blooded) animals have received very little attention and the literature concerning natural infections of these animals is limited. The aim of this study was to determine the prevalence of leptospiral antibodies in reptiles, imported into Slovenia and intended to be pets in close contact with humans. A total of 297 reptiles (22 snakes, 210 lizards and 65 turtles) were tested for specific antibodies against serovars of Leptospira interrogans sensu stricto using the microscopic agglutination test (MAT). Live cultures of different serovars were used as antigens. MAT was performed according to standard procedures and the degree of reaction was interpreted by estimating the percentage of agglutinated leptospires. Samples showing titres of ≥ 50 against one or more serovars were considered as positive. Results Antibodies against seven pathogenic serovars of L. interrogans sensu stricto were detected in 46 of 297 reptiles. Among 22 snakes, specific antibodies against pathogenic serovars of three Leptospira species (L. interrogans, L. kirschneri and L. borgpetersenii) at titre levels from 1:50 to 1:400 were detected in 6 snakes. In 31 of 210 lizards, specific antibodies were found in titres from 1:50 to 1:1000 and, finally, among 65 turtles (terrapins and tortoises), 9 had specific antibodies at titre levels between 1:50 and 1:1600. Animals imported from non-EU countries showed significantly higher prevalence (25.0%; 95 confidence interval: 16.7–33.3%) than animals from EU member states (10.4%; confidence interval: 6.1–14.7%). Conclusions Reptiles may be considered as potential reservoirs of L. interrogans sensu stricto. Origin of the animals is a risk factor for presence of leptospiral antibodies, especially in lizards. Special attention should be focused on animals from non-EU member states. PMID:24020619

  6. Serologic and Molecular Studies of Leptospira and Leptospirosis among Rats in the Philippines

    PubMed Central

    Villanueva, Sharon Y. A. M.; Ezoe, Hirokazu; Baterna, Rubelia A.; Yanagihara, Yasutake; Muto, Maki; Koizumi, Nobuo; Fukui, Takashi; Okamoto, Yoshihiro; Masuzawa, Toshiyuki; Cavinta, Lolita L.; Gloriani, Nina G.; Yoshida, Shin-ichi

    2010-01-01

    Rats are known to be the most important reservoirs and transmission sources of leptospirosis. However, the status of leptospirosis in the Philippines regarding reservoirs and transmission remains unknown. A survey was conducted in Metro Manila and Laguna that analyzed samples obtained from 106 rats. Using the microscopic agglutination test, we found that 92% of rat serum samples were positive for anti-Leptospira antibodies; the most common infecting serovars were Manilae, Hebdomadis, and Losbanos. On the basis of pulsed-field gel electrophoresis and gyrase B gene sequence analyses, four groups of rat kidney isolates were found: L. interrogans serovar Manilae, serovar Losbanos, and serogroup Grippotyphosa, and L. borgpetersenii serogroup Javanica. Most isolates were lethal after experimental infection of golden Syrian hamsters. Results showed that these four Leptospira serovars and serogroups are circulating among rats, and that these animals may be one of the possible transmission sources of leptospirosis in the Philippines. PMID:20439972

  7. Distribution and Diversity of Pathogenic Leptospira Species in Peri-domestic Surface Waters from South Central Chile

    PubMed Central

    Mason, Meghan R.; Encina, Carolina; Sreevatsan, Srinand; Muñoz-Zanzi, Claudia

    2016-01-01

    Background Leptospirosis is a neglected zoonosis affecting animals and humans caused by infection with Leptospira. The bacteria can survive outside of hosts for long periods of time in soil and water. While identification of Leptospira species from human cases and animal reservoirs are increasingly reported, little is known about the diversity of pathogenic Leptospira species in the environment and how surveillance of the environment might be used for monitoring and controlling disease. Methods and Findings Water samples (n = 104) were collected from the peri-domestic environment of 422 households from farms, rural villages, and urban slums participating in a broader study on the eco-epidemiology of leptospirosis in the Los Rios Region, Chile, between October 2010 and April 2012. The secY region of samples, previously detected as pathogenic Leptospira by PCR, was amplified and sequenced. Sequences were aligned using ClustalW in MEGA, and a minimum spanning tree was created in PHYLOViZ using the goeBURST algorithm to assess sequence similarity. Sequences from four clinical isolates, 17 rodents, and 20 reference strains were also included in the analysis. Overall, water samples contained L. interrogans, L. kirschneri, and L. weilii, with descending frequency. All species were found in each community type. The distribution of the species differed by the season in which the water samples were obtained. There was no evidence that community-level prevalence of Leptospira in dogs, rodents, or livestock influenced pathogen diversity in the water samples. Conclusions This study reports the presence of pathogenic Leptospira in the peri-domestic environment of households in three community types and the differences in Leptospira diversity at the community level. Systematic environmental surveillance of Leptospira can be used for detecting changes in pathogen diversity and to identify and monitor contaminated areas where an increased risk of human infection exists. PMID

  8. Whole Genome Analysis of Leptospira licerasiae Provides Insight into Leptospiral Evolution and Pathogenicity

    PubMed Central

    Selengut, Jeremy D.; Harkins, Derek M.; Patra, Kailash P.; Moreno, Angelo; Lehmann, Jason S.; Purushe, Janaki; Sanka, Ravi; Torres, Michael; Webster, Nicholas J.; Vinetz, Joseph M.; Matthias, Michael A.

    2012-01-01

    The whole genome analysis of two strains of the first intermediately pathogenic leptospiral species to be sequenced (Leptospira licerasiae strains VAR010 and MMD0835) provides insight into their pathogenic potential and deepens our understanding of leptospiral evolution. Comparative analysis of eight leptospiral genomes shows the existence of a core leptospiral genome comprising 1547 genes and 452 conserved genes restricted to infectious species (including L. licerasiae) that are likely to be pathogenicity-related. Comparisons of the functional content of the genomes suggests that L. licerasiae retains several proteins related to nitrogen, amino acid and carbohydrate metabolism which might help to explain why these Leptospira grow well in artificial media compared with pathogenic species. L. licerasiae strains VAR010T and MMD0835 possess two prophage elements. While one element is circular and shares homology with LE1 of L. biflexa, the second is cryptic and homologous to a previously identified but unnamed region in L. interrogans serovars Copenhageni and Lai. We also report a unique O-antigen locus in L. licerasiae comprised of a 6-gene cluster that is unexpectedly short compared with L. interrogans in which analogous regions may include >90 such genes. Sequence homology searches suggest that these genes were acquired by lateral gene transfer (LGT). Furthermore, seven putative genomic islands ranging in size from 5 to 36 kb are present also suggestive of antecedent LGT. How Leptospira become naturally competent remains to be determined, but considering the phylogenetic origins of the genes comprising the O-antigen cluster and other putative laterally transferred genes, L. licerasiae must be able to exchange genetic material with non-invasive environmental bacteria. The data presented here demonstrate that L. licerasiae is genetically more closely related to pathogenic than to saprophytic Leptospira and provide insight into the genomic bases for its infectiousness

  9. Targeted mutagenesis in pathogenic Leptospira species: disruption of the LigB gene does not affect virulence in animal models of leptospirosis.

    PubMed

    Croda, Julio; Figueira, Claudio Pereira; Wunder, Elsio A; Santos, Cleiton S; Reis, Mitermayer G; Ko, Albert I; Picardeau, Mathieu

    2008-12-01

    The pathogenic mechanisms of Leptospira interrogans, the causal agent of leptospirosis, remain largely unknown. This is mainly due to the lack of tools for genetically manipulating pathogenic Leptospira species. Thus, homologous recombination between introduced DNA and the corresponding chromosomal locus has never been demonstrated for this pathogen. Leptospiral immunoglobulin-like repeat (Lig) proteins were previously identified as putative Leptospira virulence factors. In this study, a ligB mutant was constructed by allelic exchange in L. interrogans; in this mutant a spectinomycin resistance (Spc(r)) gene replaced a portion of the ligB coding sequence. Gene disruption was confirmed by PCR, immunoblot analysis, and immunofluorescence studies. The ligB mutant did not show decrease virulence compared to the wild-type strain in the hamster model of leptospirosis. In addition, inoculation of rats with the ligB mutant induced persistent colonization of the kidneys. Finally, LigB was not required to mediate bacterial adherence to cultured cells. Taken together, our data provide the first evidence of site-directed homologous recombination in pathogenic Leptospira species. Furthermore, our data suggest that LigB does not play a major role in dissemination of the pathogen in the host and in the development of acute disease manifestations or persistent renal colonization. PMID:18809657

  10. Whole Genome Shotgun Sequencing Shows Selection on Leptospira Regulatory Proteins During in vitro Culture Attenuation.

    PubMed

    Lehmann, Jason S; Corey, Victoria C; Ricaldi, Jessica N; Vinetz, Joseph M; Winzeler, Elizabeth A; Matthias, Michael A

    2016-02-01

    Leptospirosis is the most common zoonotic disease worldwide with an estimated 500,000 severe cases reported annually, and case fatality rates of 12-25%, due primarily to acute kidney and lung injuries. Despite its prevalence, the molecular mechanisms underlying leptospirosis pathogenesis remain poorly understood. To identify virulence-related genes in Leptospira interrogans, we delineated cumulative genome changes that occurred during serial in vitro passage of a highly virulent strain of L. interrogans serovar Lai into a nearly avirulent isogenic derivative. Comparison of protein coding and computationally predicted noncoding RNA (ncRNA) genes between these two polyclonal strains identified 15 nonsynonymous single nucleotide variant (nsSNV) alleles that increased in frequency and 19 that decreased, whereas no changes in allelic frequency were observed among the ncRNA genes. Some of the nsSNV alleles were in six genes shown previously to be transcriptionally upregulated during exposure to in vivo-like conditions. Five of these nsSNVs were in evolutionarily conserved positions in genes related to signal transduction and metabolism. Frequency changes of minor nsSNV alleles identified in this study likely contributed to the loss of virulence during serial in vitro culture. The identification of new virulence-associated genes should spur additional experimental inquiry into their potential role in Leptospira pathogenesis.

  11. Whole Genome Shotgun Sequencing Shows Selection on Leptospira Regulatory Proteins during in vitro Culture Attenuation

    PubMed Central

    Lehmann, Jason S.; Corey, Victoria C.; Ricaldi, Jessica N.; Vinetz, Joseph M.; Winzeler, Elizabeth A.; Matthias, Michael A.

    2016-01-01

    Leptospirosis is the most common zoonotic disease worldwide with an estimated 500,000 severe cases reported annually, and case fatality rates of 12–25%, due primarily to acute kidney and lung injuries. Despite its prevalence, the molecular mechanisms underlying leptospirosis pathogenesis remain poorly understood. To identify virulence-related genes in Leptospira interrogans, we delineated cumulative genome changes that occurred during serial in vitro passage of a highly virulent strain of L. interrogans serovar Lai into a nearly avirulent isogenic derivative. Comparison of protein coding and computationally predicted noncoding RNA (ncRNA) genes between these two polyclonal strains identified 15 nonsynonymous single nucleotide variant (nsSNV) alleles that increased in frequency and 19 that decreased, whereas no changes in allelic frequency were observed among the ncRNA genes. Some of the nsSNV alleles were in six genes shown previously to be transcriptionally upregulated during exposure to in vivo-like conditions. Five of these nsSNVs were in evolutionarily conserved positions in genes related to signal transduction and metabolism. Frequency changes of minor nsSNV alleles identified in this study likely contributed to the loss of virulence during serial in vitro culture. The identification of new virulence-associated genes should spur additional experimental inquiry into their potential role in Leptospira pathogenesis. PMID:26711524

  12. Detection of pathogenic Leptospira bacteria in pinniped populations via PCR and identification of a source of transmission for zoonotic leptospirosis in the marine environment.

    PubMed

    Cameron, Caroline E; Zuerner, Richard L; Raverty, Stephen; Colegrove, Kathleen M; Norman, Stephanie A; Lambourn, Dyanna M; Jeffries, Steven J; Gulland, Frances M

    2008-05-01

    Leptospirosis, caused by the spirochete Leptospira, is a geographically widespread disease that affects a broad range of mammals, including marine mammals. Among pinniped populations, periodic epizootics of leptospirosis are responsible for significant die-offs. Along the west coast of North America, the most recent leptospirosis epizootic occurred in 2004, during which samples were collected from cases ranging from California to British Columbia. The primary objective of this study was to use this well-defined sample set to determine the feasibility of using PCR techniques to diagnose Leptospira infection among pinniped populations in comparison with diagnostic methodologies commonly used for marine mammals. Successful amplification was achieved from a variety of samples, including freshly collected urine, urine stored at -80 degrees C for less than 6 months, and kidney (freshly collected, frozen, and decomposed), as well as feces- and urine-contaminated sand collected in the vicinity of a live-stranded animal. Pathological examination of tissue collected from Leptospira-infected animals revealed the presence of leptospiral antigen in the kidneys. The use of species-specific primer pairs revealed a pattern of host specificity for Leptospira interrogans in sea lions and Leptospira kirschneri in elephant seals. These studies indicate PCR is a sensitive and specific diagnostic tool for the detection of Leptospira infection in pinnipeds and reveal a potential source for epizootic, enzootic, and zoonotic spread of leptospirosis in a marine environment. PMID:18367568

  13. Detection of Pathogenic Leptospira Bacteria in Pinniped Populations via PCR and Identification of a Source of Transmission for Zoonotic Leptospirosis in the Marine Environment▿

    PubMed Central

    Cameron, Caroline E.; Zuerner, Richard L.; Raverty, Stephen; Colegrove, Kathleen M.; Norman, Stephanie A.; Lambourn, Dyanna M.; Jeffries, Steven J.; Gulland, Frances M.

    2008-01-01

    Leptospirosis, caused by the spirochete Leptospira, is a geographically widespread disease that affects a broad range of mammals, including marine mammals. Among pinniped populations, periodic epizootics of leptospirosis are responsible for significant die-offs. Along the west coast of North America, the most recent leptospirosis epizootic occurred in 2004, during which samples were collected from cases ranging from California to British Columbia. The primary objective of this study was to use this well-defined sample set to determine the feasibility of using PCR techniques to diagnose Leptospira infection among pinniped populations in comparison with diagnostic methodologies commonly used for marine mammals. Successful amplification was achieved from a variety of samples, including freshly collected urine, urine stored at −80°C for less than 6 months, and kidney (freshly collected, frozen, and decomposed), as well as feces- and urine-contaminated sand collected in the vicinity of a live-stranded animal. Pathological examination of tissue collected from Leptospira-infected animals revealed the presence of leptospiral antigen in the kidneys. The use of species-specific primer pairs revealed a pattern of host specificity for Leptospira interrogans in sea lions and Leptospira kirschneri in elephant seals. These studies indicate PCR is a sensitive and specific diagnostic tool for the detection of Leptospira infection in pinnipeds and reveal a potential source for epizootic, enzootic, and zoonotic spread of leptospirosis in a marine environment. PMID:18367568

  14. A replicative plasmid vector allows efficient complementation of pathogenic Leptospira strains.

    PubMed

    Pappas, Christopher J; Benaroudj, Nadia; Picardeau, Mathieu

    2015-05-01

    Leptospirosis, an emerging zoonotic disease, remains poorly understood because of a lack of genetic manipulation tools available for pathogenic leptospires. Current genetic manipulation techniques include insertion of DNA by random transposon mutagenesis and homologous recombination via suicide vectors. This study describes the construction of a shuttle vector, pMaORI, that replicates within saprophytic, intermediate, and pathogenic leptospires. The shuttle vector was constructed by the insertion of a 2.9-kb DNA segment including the parA, parB, and rep genes into pMAT, a plasmid that cannot replicate in Leptospira spp. and contains a backbone consisting of an aadA cassette, ori R6K, and oriT RK2/RP4. The inserted DNA segment was isolated from a 52-kb region within Leptospira mayottensis strain 200901116 that is not found in the closely related strain L. mayottensis 200901122. Because of the size of this region and the presence of bacteriophage-like proteins, it is possible that this region is a result of a phage-related genomic island. The stability of the pMaORI plasmid within pathogenic strains was tested by passaging cultures 10 times without selection and confirming the presence of pMaORI. Concordantly, we report the use of trans complementation in the pathogen Leptospira interrogans. Transformation of a pMaORI vector carrying a functional copy of the perR gene in a null mutant background restores the expression of PerR and susceptibility to hydrogen peroxide comparable to that of wild-type cells. In conclusion, we demonstrate the replication of a stable plasmid vector in a large panel of Leptospira strains, including pathogens. The shuttle vector described will expand our ability to perform genetic manipulation of Leptospira spp.

  15. Direct detection and differentiation of pathogenic Leptospira species using a multi-gene targeted real time PCR approach.

    PubMed

    Ferreira, Ana Sofia; Costa, Pedro; Rocha, Teresa; Amaro, Ana; Vieira, Maria Luísa; Ahmed, Ahmed; Thompson, Gertrude; Hartskeerl, Rudy A; Inácio, João

    2014-01-01

    Leptospirosis is a growing public and veterinary health concern caused by pathogenic species of Leptospira. Rapid and reliable laboratory tests for the direct detection of leptospiral infections in animals are in high demand not only to improve diagnosis but also for understanding the epidemiology of the disease. In this work we describe a novel and simple TaqMan-based multi-gene targeted real-time PCR approach able to detect and differentiate Leptospira interrogans, L. kirschneri, L. borgpeteresenii and L. noguchii, which constitute the veterinary most relevant pathogenic species of Leptospira. The method uses sets of species-specific probes, and respective flanking primers, designed from ompL1 and secY gene sequences. To monitor the presence of inhibitors, a duplex amplification assay targeting both the mammal β-actin and the leptospiral lipL32 genes was implemented. The analytical sensitivity of all primer and probe sets was estimated to be <10 genome equivalents (GE) in the reaction mixture. Application of the amplification reactions on genomic DNA from a variety of pathogenic and non-pathogenic Leptospira strains and other non-related bacteria revealed a 100% analytical specificity. Additionally, pathogenic leptospires were successfully detected in five out of 29 tissue samples from animals (Mus spp., Rattus spp., Dolichotis patagonum and Sus domesticus). Two samples were infected with L. borgpetersenii, two with L. interrogans and one with L. kirschneri. The possibility to detect and identify these pathogenic agents to the species level in domestic and wildlife animals reinforces the diagnostic information and will enhance our understanding of the epidemiology of leptopirosis. PMID:25398140

  16. Direct Detection and Differentiation of Pathogenic Leptospira Species Using a Multi-Gene Targeted Real Time PCR Approach

    PubMed Central

    Ferreira, Ana Sofia; Costa, Pedro; Rocha, Teresa; Amaro, Ana; Vieira, Maria Luísa; Ahmed, Ahmed; Thompson, Gertrude; Hartskeerl, Rudy A.; Inácio, João

    2014-01-01

    Leptospirosis is a growing public and veterinary health concern caused by pathogenic species of Leptospira. Rapid and reliable laboratory tests for the direct detection of leptospiral infections in animals are in high demand not only to improve diagnosis but also for understanding the epidemiology of the disease. In this work we describe a novel and simple TaqMan-based multi-gene targeted real-time PCR approach able to detect and differentiate Leptospira interrogans, L. kirschneri, L. borgpeteresenii and L. noguchii, which constitute the veterinary most relevant pathogenic species of Leptospira. The method uses sets of species-specific probes, and respective flanking primers, designed from ompL1 and secY gene sequences. To monitor the presence of inhibitors, a duplex amplification assay targeting both the mammal β-actin and the leptospiral lipL32 genes was implemented. The analytical sensitivity of all primer and probe sets was estimated to be <10 genome equivalents (GE) in the reaction mixture. Application of the amplification reactions on genomic DNA from a variety of pathogenic and non-pathogenic Leptospira strains and other non-related bacteria revealed a 100% analytical specificity. Additionally, pathogenic leptospires were successfully detected in five out of 29 tissue samples from animals (Mus spp., Rattus spp., Dolichotis patagonum and Sus domesticus). Two samples were infected with L. borgpetersenii, two with L. interrogans and one with L. kirschneri. The possibility to detect and identify these pathogenic agents to the species level in domestic and wildlife animals reinforces the diagnostic information and will enhance our understanding of the epidemiology of leptopirosis. PMID:25398140

  17. Epizootiological survey of small mammals as Leptospira spp. reservoirs in Eastern Croatia.

    PubMed

    Stritof Majetic, Zrinka; Galloway, Renee; Ruzic Sabljic, Eva; Milas, Zoran; Mojcec Perko, Vesna; Habus, Josipa; Margaletic, Josip; Pernar, Renata; Turk, Nenad

    2014-03-01

    In this survey we investigated a population of small mammals in Eastern Croatia in order to determine Leptospira carriage rates and identify circulating serovars. Out of 67 trapped animals, 20 (29.9%) isolates were obtained. Identification of isolates using microscopic agglutination test, pulsed field gel electrophoresis and multi locus sequence typing revealed that 10 (50.0%) isolates belong to serogroup Pomona, serovar Mozdok, 6 (30.0%) isolates to serogroup Australis, serovar Jalna, 2 (10.0%) isolates to serogroup Sejroe, serovar Saxkoebing, and 1 (5.0%) isolate to serogroup Grippotyphosa, serovar Grippotyphosa. One isolate from serogroup Bataviae was unable to be identified to the serovar level. Amplification of a 331-bp region of the locus LA0322 using real-time polymerase chain reaction determined that 12 (60.0%) isolates belong to L. kirschneri, 6 (30.0%) isolates to L. interrogans, and 2 (10.0%) isolates to L. borgpetersenii. Leptospira carriage rate was high (29.9%), which corresponds to a high incidence of human and domestic animal leptospirosis in Eastern Croatia. Furthermore, 90.0% of the isolates belong to serogroups Pomona, Australis and Sejroe which are also the most prevalent serogroups in humans in this area. These findings suggest that small mammals might be an important source of Leptospira spp. infection in Eastern Croatia.

  18. Comparison of Immunoprotection of Leptospira Recombinant Proteins with conventional vaccine in experimental animals.

    PubMed

    Parthiban, M; Kumar, S Senthil; Balachandran, C; Kumanan, K; Aarthi, K S; Nireesha, G

    2015-12-01

    Leptospirosis is a bacterial disease caused by bacteria of the genus Leptospira affecting humans and animals. Untreated leptospirosis may result in severe kidney damage, meningitis, liver failure, respiratory distress, and even death. Virulent leptospirosis can rapidly enter kidney fibroblasts and induce a programmed cell death. Thus, it is a challenge for immunologists to develop an effective and safe leptospirosis vaccine. Here, we compared the commercial canine leptospira vaccine and recombinant proteins (OmpL1 and LipL41) with and without adjuvant in terms of immune response and challenge studies in hamsters and immune response studies alone in experimental dogs. The outer membrane proteins viz., lipL41 and OmpL1 of leptospira interrogans serovars icterohaemorrhagiae were amplified. The primers were designed in such a way that amplified products of OmpL1 and lipL41 were ligated and cloned simultaneously into a single vector. The cloned products were expressed in E. coli BL21 cells. The immunoprotection studies were conducted for both recombinant proteins and commercial vaccine. The challenge experiment studies revealed that combination of both rLip41 and rOmpL1 and commercial vaccine gave 83% and 87% protection, respectively. Histopathological investigation revealed mild sub lethal changes were noticed in liver and kidney in commercially vaccinated group alone. The immune responses against recombinant leptospiral proteins were also demonstrated in dogs. PMID:26742322

  19. Multiple Posttranslational Modifications of Leptospira biflexa Proteins as Revealed by Proteomic Analysis

    PubMed Central

    Carroll, James A.; Olano, L. Rennee; Sturdevant, Daniel E.; Rosa, Patricia A.

    2015-01-01

    The saprophyte Leptospira biflexa is an excellent model for studying the physiology of the medically important Leptospira genus, the pathogenic members of which are more recalcitrant to genetic manipulation and have significantly slower in vitro growth. However, relatively little is known regarding the proteome of L. biflexa, limiting its utility as a model for some studies. Therefore, we have generated a proteomic map of both soluble and membrane-associated proteins of L. biflexa during exponential growth and in stationary phase. Using these data, we identified abundantly produced proteins in each cellular fraction and quantified the transcript levels from a subset of these genes using quantitative reverse transcription-PCR (RT-PCR). These proteins should prove useful as cellular markers and as controls for gene expression studies. We also observed a significant number of L. biflexa membrane-associated proteins with multiple isoforms, each having unique isoelectric focusing points. L. biflexa cell lysates were examined for several posttranslational modifications suggested by the protein patterns. Methylation and acetylation of lysine residues were predominately observed in the proteins of the membrane-associated fraction, while phosphorylation was detected mainly among soluble proteins. These three posttranslational modification systems appear to be conserved between the free-living species L. biflexa and the pathogenic species Leptospira interrogans, suggesting an important physiological advantage despite the varied life cycles of the different species. PMID:26655756

  20. High Leptospira Diversity in Animals and Humans Complicates the Search for Common Reservoirs of Human Disease in Rural Ecuador

    PubMed Central

    Chiriboga, Jorge; Miller, Erin; Olivas, Sonora; Birdsell, Dawn; Hepp, Crystal; Hornstra, Heidie; Schupp, James M.; Morales, Melba; Gonzalez, Manuel; Reyes, Soraya; de la Cruz, Carmen; Keim, Paul; Hartskeerl, Rudy; Trueba, Gabriel; Pearson, Talima

    2016-01-01

    Background Leptospirosis is a zoonotic disease responsible for high morbidity around the world, especially in tropical and low income countries. Rats are thought to be the main vector of human leptospirosis in urban settings. However, differences between urban and low-income rural communities provide additional insights into the epidemiology of the disease. Methodology/Principal findings Our study was conducted in two low-income rural communities near the coast of Ecuador. We detected and characterized infectious leptospira DNA in a wide variety of samples using new real time quantitative PCR assays and amplicon sequencing. We detected infectious leptospira in a high percentage of febrile patients (14.7%). In contrast to previous studies on leptospirosis risk factors, higher positivity was not found in rats (3.0%) but rather in cows (35.8%) and pigs (21.1%). Six leptospira species were identified (L. borgpetersenii, L kirschnerii, L santarosai, L. interrogans, L noguchii, and an intermediate species within the L. licerasiae and L. wolffii clade) and no significant differences in the species of leptospira present in each animal species was detected (χ2 = 9.89, adj.p-value = 0.27). Conclusions/Significance A large portion of the world’s human population lives in low-income, rural communities, however, there is limited information about leptospirosis transmission dynamics in these settings. In these areas, exposure to peridomestic livestock is particularly common and high prevalence of infectious leptospira in cows and pigs suggest that they may be the most important reservoir for human transmission. Genotyping clinical samples show that multiple species of leptospira are involved in human disease. As these genotypes were also detected in samples from a variety of animals, genotype data must be used in conjunction with epidemiological data to provide evidence of transmission and the importance of different potential leptospirosis reservoirs. PMID:27622673

  1. Isolation of a Seawater Tolerant Leptospira spp. from a Southern Right Whale (Eubalaena australis).

    PubMed

    Grune Loffler, Sylvia; Rago, Virginia; Martínez, Mara; Uhart, Marcela; Florin-Christensen, Monica; Romero, Graciela; Brihuega, Bibiana

    2015-01-01

    Leptospirosis is the most widespread zoonotic disease in the world. It is caused by pathogenic spirochetes of the genus Leptospira spp. and is maintained in nature through chronic renal infection of carrier animals. Rodents and other small mammals are the main reservoirs. Information on leptospirosis in marine mammals is scarce; however, cases of leptospirosis have been documented in pinniped populations from the Pacific coast of North America from southern California to British Columbia. We report the isolation of a Leptospira spp. strain, here named Manara, from a kidney sample obtained from a Southern Right Whale (Eubalaena australis) calf, which stranded dead in Playa Manara, Península Valdés, Argentina. This strain showed motility and morphology typical of the genus Leptospira spp. under dark-field microscopy; and grew in Ellinghausen-McCullough-Johnson-Harris (EMJH) medium and Fletcher medium after 90 days of incubation at 28°C. Considering the source of this bacterium, we tested its ability to grow in Fletcher medium diluted with seawater at different percentages (1%, 3%, 5%, 7% and 10% v/v). Bacterial growth was detected 48 h after inoculation of Fletcher medium supplemented with 5% sea water, demonstrating the halophilic nature of the strain Manara. Phylogenetic analysis of 16S rRNA gene sequences placed this novel strain within the radiation of the pathogenic species of the genus Leptospira spp., with sequence similarities within the range 97-100%, and closely related to L. interrogans. Two different PCR protocols targeting genus-specific pathogenic genes (G1-G2, B64I-B64II and LigB) gave positive results, which indicates that the strain Manara is likely pathogenic. Further studies are needed to confirm this possibility as well as determine its serogroup. These results could modify our understanding of the epidemiology of this zoonosis. Until now, the resistance and ability to grow in seawater for long periods of time had been proven for the strain

  2. Isolation of a Seawater Tolerant Leptospira spp. from a Southern Right Whale (Eubalaena australis)

    PubMed Central

    Rago, Virginia; Uhart, Marcela

    2015-01-01

    Leptospirosis is the most widespread zoonotic disease in the world. It is caused by pathogenic spirochetes of the genus Leptospira spp. and is maintained in nature through chronic renal infection of carrier animals. Rodents and other small mammals are the main reservoirs. Information on leptospirosis in marine mammals is scarce; however, cases of leptospirosis have been documented in pinniped populations from the Pacific coast of North America from southern California to British Columbia. We report the isolation of a Leptospira spp. strain, here named Manara, from a kidney sample obtained from a Southern Right Whale (Eubalaena australis) calf, which stranded dead in Playa Manara, Península Valdés, Argentina. This strain showed motility and morphology typical of the genus Leptospira spp. under dark-field microscopy; and grew in Ellinghausen-McCullough-Johnson-Harris (EMJH) medium and Fletcher medium after 90 days of incubation at 28°C. Considering the source of this bacterium, we tested its ability to grow in Fletcher medium diluted with seawater at different percentages (1%, 3%, 5%, 7% and 10% v/v). Bacterial growth was detected 48 h after inoculation of Fletcher medium supplemented with 5% sea water, demonstrating the halophilic nature of the strain Manara. Phylogenetic analysis of 16S rRNA gene sequences placed this novel strain within the radiation of the pathogenic species of the genus Leptospira spp., with sequence similarities within the range 97–100%, and closely related to L. interrogans. Two different PCR protocols targeting genus-specific pathogenic genes (G1-G2, B64I-B64II and LigB) gave positive results, which indicates that the strain Manara is likely pathogenic. Further studies are needed to confirm this possibility as well as determine its serogroup. These results could modify our understanding of the epidemiology of this zoonosis. Until now, the resistance and ability to grow in seawater for long periods of time had been proven for the strain

  3. Isolation of a Seawater Tolerant Leptospira spp. from a Southern Right Whale (Eubalaena australis).

    PubMed

    Grune Loffler, Sylvia; Rago, Virginia; Martínez, Mara; Uhart, Marcela; Florin-Christensen, Monica; Romero, Graciela; Brihuega, Bibiana

    2015-01-01

    Leptospirosis is the most widespread zoonotic disease in the world. It is caused by pathogenic spirochetes of the genus Leptospira spp. and is maintained in nature through chronic renal infection of carrier animals. Rodents and other small mammals are the main reservoirs. Information on leptospirosis in marine mammals is scarce; however, cases of leptospirosis have been documented in pinniped populations from the Pacific coast of North America from southern California to British Columbia. We report the isolation of a Leptospira spp. strain, here named Manara, from a kidney sample obtained from a Southern Right Whale (Eubalaena australis) calf, which stranded dead in Playa Manara, Península Valdés, Argentina. This strain showed motility and morphology typical of the genus Leptospira spp. under dark-field microscopy; and grew in Ellinghausen-McCullough-Johnson-Harris (EMJH) medium and Fletcher medium after 90 days of incubation at 28°C. Considering the source of this bacterium, we tested its ability to grow in Fletcher medium diluted with seawater at different percentages (1%, 3%, 5%, 7% and 10% v/v). Bacterial growth was detected 48 h after inoculation of Fletcher medium supplemented with 5% sea water, demonstrating the halophilic nature of the strain Manara. Phylogenetic analysis of 16S rRNA gene sequences placed this novel strain within the radiation of the pathogenic species of the genus Leptospira spp., with sequence similarities within the range 97-100%, and closely related to L. interrogans. Two different PCR protocols targeting genus-specific pathogenic genes (G1-G2, B64I-B64II and LigB) gave positive results, which indicates that the strain Manara is likely pathogenic. Further studies are needed to confirm this possibility as well as determine its serogroup. These results could modify our understanding of the epidemiology of this zoonosis. Until now, the resistance and ability to grow in seawater for long periods of time had been proven for the strain

  4. Structural and Functional Characterization of an Orphan ATP-Binding Cassette ATPase Involved in Manganese Utilization and Tolerance in Leptospira spp.

    PubMed Central

    Saul, Frederick; Bellalou, Jacques; Miras, Isabelle; Weber, Patrick; Bondet, Vincent; Murray, Gerald L.; Adler, Ben; Ristow, Paula; Louvel, Hélène; Haouz, Ahmed

    2013-01-01

    Pathogenic Leptospira species are the etiological agents of the widespread zoonotic disease leptospirosis. Most organisms, including Leptospira, require divalent cations for proper growth, but because of their high reactivity, these metals are toxic at high concentrations. Therefore, bacteria have acquired strategies to maintain metal homeostasis, such as metal import and efflux. By screening Leptospira biflexa transposon mutants for their ability to use Mn2+, we have identified a gene encoding a putative orphan ATP-binding cassette (ABC) ATPase of unknown function. Inactivation of this gene in both L. biflexa and L. interrogans strains led to mutants unable to grow in medium in which iron was replaced by Mn2+, suggesting an involvement of this ABC ATPase in divalent cation uptake. A mutation in this ATPase-coding gene increased susceptibility to Mn2+ toxicity. Recombinant ABC ATPase of the pathogen L. interrogans exhibited Mg2+-dependent ATPase activity involving a P-loop motif. The structure of this ATPase was solved from a crystal containing two monomers in the asymmetric unit. Each monomer adopted a canonical two-subdomain organization of the ABC ATPase fold with an α/β subdomain containing the Walker motifs and an α subdomain containing the ABC signature motif (LSSGE). The two monomers were arranged in a head-to-tail orientation, forming a V-shaped particle with all the conserved ABC motifs at the dimer interface, similar to functional ABC ATPases. These results provide the first structural and functional characterization of a leptospiral ABC ATPase. PMID:24123817

  5. An Extended Multilocus Sequence Typing (MLST) Scheme for Rapid Direct Typing of Leptospira from Clinical Samples

    PubMed Central

    Menezes, Angela; Woods, Kate; Chanthongthip, Anisone; Dittrich, Sabine; Opoku-Boateng, Agatha; Kimuli, Maimuna; Chalker, Victoria

    2016-01-01

    Background Rapid typing of Leptospira is currently impaired by requiring time consuming culture of leptospires. The objective of this study was to develop an assay that provides multilocus sequence typing (MLST) data direct from patient specimens while minimising costs for subsequent sequencing. Methodology and Findings An existing PCR based MLST scheme was modified by designing nested primers including anchors for facilitated subsequent sequencing. The assay was applied to various specimen types from patients diagnosed with leptospirosis between 2014 and 2015 in the United Kingdom (UK) and the Lao Peoples Democratic Republic (Lao PDR). Of 44 clinical samples (23 serum, 6 whole blood, 3 buffy coat, 12 urine) PCR positive for pathogenic Leptospira spp. at least one allele was amplified in 22 samples (50%) and used for phylogenetic inference. Full allelic profiles were obtained from ten specimens, representing all sample types (23%). No nonspecific amplicons were observed in any of the samples. Of twelve PCR positive urine specimens three gave full allelic profiles (25%) and two a partial profile. Phylogenetic analysis allowed for species assignment. The predominant species detected was L. interrogans (10/14 and 7/8 from UK and Lao PDR, respectively). All other species were detected in samples from only one country (Lao PDR: L. borgpetersenii [1/8]; UK: L. kirschneri [1/14], L. santarosai [1/14], L. weilii [2/14]). Conclusion Typing information of pathogenic Leptospira spp. was obtained directly from a variety of clinical samples using a modified MLST assay. This assay negates the need for time-consuming culture of Leptospira prior to typing and will be of use both in surveillance, as single alleles enable species determination, and outbreaks for the rapid identification of clusters. PMID:27654037

  6. Genomic survey and expression analysis of DNA repair genes in the genus Leptospira.

    PubMed

    Martins-Pinheiro, Marinalva; Schons-Fonseca, Luciane; da Silva, Josefa B; Domingos, Renan H; Momo, Leonardo Hiroyuki Santos; Simões, Ana Carolina Quirino; Ho, Paulo Lee; da Costa, Renata M A

    2016-04-01

    Leptospirosis is an emerging zoonosis with important economic and public health consequences and is caused by pathogenic leptospires. The genus Leptospira belongs to the order Spirochaetales and comprises saprophytic (L. biflexa), pathogenic (L. interrogans) and host-dependent (L. borgpetersenii) members. Here, we present an in silico search for DNA repair pathways in Leptospira spp. The relevance of such DNA repair pathways was assessed through the identification of mRNA levels of some genes during infection in animal model and after exposition to spleen cells. The search was performed by comparison of available Leptospira spp. genomes in public databases with known DNA repair-related genes. Leptospires exhibit some distinct and unexpected characteristics, for instance the existence of a redundant mechanism for repairing a chemically diverse spectrum of alkylated nucleobases, a new mutS-like gene and a new shorter version of uvrD. Leptospira spp. shares some characteristics from Gram-positive, as the presence of PcrA, two RecQ paralogs and two SSB proteins; the latter is considered a feature shared by naturally competent bacteria. We did not find a significant reduction in the number of DNA repair-related genes in both pathogenic and host-dependent species. Pathogenic leptospires were enriched for genes dedicated to base excision repair and non-homologous end joining. Their evolutionary history reveals a remarkable importance of lateral gene transfer events for the evolution of the genus. Up-regulation of specific DNA repair genes, including components of SOS regulon, during infection in animal model validates the critical role of DNA repair mechanisms for the complex interplay between host/pathogen.

  7. Severe Leptospira interrogans serovar Icterohaemorrhagiae infection with hepato-renal-pulmonary involvement treated with corticosteroids

    PubMed Central

    Schulze, Marco H; Raschel, Heribert; Langen, Heinz-Jakob; Stich, August; Tappe, Dennis

    2014-01-01

    Key Clinical Message The traditional concept of immediate antibiotic treatment in suspected leptospirosis seems to be especially important for patients up to day 4 of clinical illness. As immune mechanisms probably play a crucial role in advanced leptospirosis with presumed pulmonary hemorrhages, patients might benefit from corticosteroids or other immunosuppressive agents beside antibiotics. PMID:25614810

  8. Geographical Dissemination of Leptospira interrogans serovar Pomona During Seasonal Migration of California Sea Lions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospirosis is one of the most widespread bacterial zoonoses in the world, and affects both terrestrial and marine mammals. Little information is available describing the geographical spread of leptospirosis, but it is logical to assume that animal migrations contribute to this process. There ha...

  9. Evaluation of the Leptospira interrogans Outer Membrane Protein OmpL37 as a Vaccine Candidate.

    PubMed

    Oliveira, Thaís Larré; Grassmann, André Alex; Schuch, Rodrigo Andrade; Seixas Neto, Amilton Clair Pinto; Mendonça, Marcelo; Hartwig, Daiane Drawanz; McBride, Alan John Alexander; Dellagostin, Odir Antônio

    2015-01-01

    The identification of potential vaccine candidates against leptospirosis remains a challenge. However, one such candidate is OmpL37, a potentially surface-exposed antigen that has the highest elastin-binding ability described to date, suggesting that it plays an important role in host colonization. In order to evaluate OmpL37's ability to induce a protective immune response, prime-boost, DNA and subunit vaccine strategies were tested in the hamster model of lethal leptospirosis. The humoral immune response was evaluated using an indirect ELISA test, and the cytokine profile in whole blood was determined by quantitative real-time PCR. Unlike the DNA vaccine, the administration of recombinant OmpL37 induced a strong IgG antibody response. When individually administrated, both formulations stimulated a TNF-α mediated inflammatory response. However, none of the OmpL37 formulations or vaccination strategies induced protective immunity. Further studies are required towards the identification of new vaccine targets against leptospirosis. PMID:26588685

  10. Titers to Leptospira species in horses in Alberta.

    PubMed Central

    Lees, V W; Gale, S P

    1994-01-01

    Sera from horses in Alberta, submitted to Agriculture and Agri-Food Canada for routine testing for equine infectious anemia from January 1987 to June 1989, were tested for antibody against 13 serovars of pathogenic Leptospira spp., using the microscopic agglutination test. The purpose of the study was to investigate the prevalence of serum titers to those serovars in horses in Alberta, and to analyze the associated risk factors. Descriptive statistics were compiled and logistic regressions were computed. Titers to L. interrogans serovars icterohaemorrhagiae, bratislava, copenhageni, and autumnalis were common (94.6%, 56.6%, 46.5%, and 43.5%, respectively). The prevalence of titers to other serovars ranged from 0.8% to 27.2%. Age was almost always significantly associated with the presence of titers. In general, the chances of being seropositive rose by approximately 10% with each year of life. Horses managed individually (eg, track horses) were approximately half as likely to be seropositive as were horses managed in groups (eg, rodeo horses). PMID:7994706

  11. Morphological alterations in the kidney of rats with natural and experimental Leptospira infection.

    PubMed

    Tucunduva de Faria, M; Athanazio, D A; Gonçalves Ramos, E A; Silva, E F; Reis, M G; Ko, A I

    2007-11-01

    Leptospirosis is a widespread anthropozoonosis, with a broad array of mammalian reservoirs, occurring as rural endemics, urban outbreaks related to floods, and emergent disease associated with water sports and recreational exposure in developed countries. Rats are the major source of human infection, particularly in urban areas; however few reports have focused on the pathology of leptospirosis in this host. This study reports pathological changes in 60 kidneys from captured wild rats and compares these with changes in the kidney of Wistar rats experimentally infected with Leptospira interrogans serovar Copenhageni strain FIOCRUZ L1-130. A broad range of morphological alterations were detected in the kidneys from captured rats but interstitial nephritis was the only feature reproduced under experimental conditions. The role of interstitial nephritis in the pathogenesis of leptospirosis is reviewed and it is suggested that rats may provide a potential tool for the study of colonization mechanisms and host resistance in acute leptospiral disease.

  12. InvA protein is a Nudix hydrolase required for infection by pathogenic Leptospira in cell lines and animals.

    PubMed

    Luo, Yihui; Liu, Yan; Sun, Dexter; Ojcius, David M; Zhao, Jinfang; Lin, Xuai; Wu, Dong; Zhang, Rongguang; Chen, Ming; Li, Lanjuan; Yan, Jie

    2011-10-21

    Leptospirosis caused by pathogenic species of the genus Leptospira is a re-emerging zoonotic disease, which affects a wide variety of host species and is transmitted by contaminated water. The genomes of several pathogenic Leptospira species contain a gene named invA, which contains a Nudix domain. However, the function of this gene has never been characterized. Here, we demonstrated that the invA gene was highly conserved in protein sequence and present in all tested pathogenic Leptospira species. The recombinant InvA protein of pathogenic L. interrogans strain Lai hydrolyzed several specific dinucleoside oligophosphate substrates, reflecting the enzymatic activity of Nudix in Leptospira species. Pathogenic leptospires did not express this protein in media but temporarily expressed it at early stages (within 60 min) of infection of macrophages and nephric epithelial cells. Comparing with the wild type, the invA-deficient mutant displayed much lower infectivity and a significantly reduced survival rate in macrophages and nephric epithelial cells. Moreover, the invA-deficient leptospires presented an attenuated virulence in hamsters, caused mild histopathological damage, and were transmitted in lower numbers in the urine, compared with the wild-type strain. The invA revertant, made by complementing the invA-deficient mutant with the invA gene, reacquired virulence similar to the wild type in vitro and in vivo. The LD(50) in hamsters was 1000-fold higher for the invA-deficient mutant than for the invA revertant and wild type. These results demonstrate that the InvA protein is a Nudix hydrolase, and the invA gene is essential for virulence in pathogenic Leptospira species.

  13. Comparison of Two Multilocus Sequence Based Genotyping Schemes for Leptospira Species

    PubMed Central

    Boonsilp, Siriphan; Wuthiekanun, Vanaporn; Nalam, Kishore; Spratt, Brian G.; Aanensen, David M.; Smythe, Lee D.; Ahmed, Niyaz; Feil, Edward J.; Hartskeerl, Rudy A.; Peacock, Sharon J.

    2011-01-01

    Background Several sequence based genotyping schemes have been developed for Leptospira spp. The objective of this study was to genotype a collection of clinical and reference isolates using the two most commonly used schemes and compare and contrast the results. Methods and Findings A total of 48 isolates consisting of L. interrogans (n = 40) and L. kirschneri (n = 8) were typed by the 7 locus MLST scheme described by Thaipadungpanit et al., and the 6 locus genotyping scheme described by Ahmed et al., (termed 7L and 6L, respectively). Two L. interrogans isolates were not typed using 6L because of a deletion of three nucleotides in lipL32. The remaining 46 isolates were resolved into 21 sequence types (STs) by 7L, and 30 genotypes by 6L. Overall nucleotide diversity (based on concatenated sequence) was 3.6% and 2.3% for 7L and 6L, respectively. The D value (discriminatory ability) of 7L and 6L were comparable, i.e. 92.0 (95% CI 87.5–96.5) vs. 93.5 (95% CI 88.6–98.4). The dN/dS ratios calculated for each locus indicated that none were under positive selection. Neighbor joining trees were reconstructed based on the concatenated sequences for each scheme. Both trees showed two distinct groups corresponding to L. interrogans and L. kirschneri, and both identified two clones containing 10 and 7 clinical isolates, respectively. There were six instances in which 6L split single STs as defined by 7L into closely related clusters. We noted two discrepancies between the trees in which the genetic relatedness between two pairs of strains were more closely related by 7L than by 6L. Conclusions This genetic analysis indicates that the two schemes are comparable. We discuss their practical advantages and disadvantages. PMID:22087342

  14. A novel flagellar sheath protein, FcpA, determines filament coiling, translational motility and virulence for the Leptospira spirochete.

    PubMed

    Wunder, Elsio A; Figueira, Cláudio P; Benaroudj, Nadia; Hu, Bo; Tong, Brian A; Trajtenberg, Felipe; Liu, Jun; Reis, Mitermayer G; Charon, Nyles W; Buschiazzo, Alejandro; Picardeau, Mathieu; Ko, Albert I

    2016-08-01

    Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends and the presence of periplasmic flagella (PF) with pronounced spontaneous supercoiling. The factors that provoke such supercoiling, as well as the role that PF coiling plays in generating the characteristic hook-end cell morphology and motility, have not been elucidated. We have now identified an abundant protein from the pathogen L. interrogans, exposed on the PF surface, and named it Flagellar-coiling protein A (FcpA). The gene encoding FcpA is highly conserved among Leptospira and was not found in other bacteria. fcpA(-) mutants, obtained from clinical isolates or by allelic exchange, had relatively straight, smaller-diameter PF, and were not able to produce translational motility. These mutants lost their ability to cause disease in the standard hamster model of leptospirosis. Complementation of fcpA restored the wild-type morphology, motility and virulence phenotypes. In summary, we identified a novel Leptospira 36-kDa protein, the main component of the spirochete's PF sheath, and a key determinant of the flagella's coiled structure. FcpA is essential for bacterial translational motility and to enable the spirochete to penetrate the host, traverse tissue barriers, disseminate to cause systemic infection and reach target organs. PMID:27113476

  15. A novel flagellar sheath protein, FcpA, determines filament coiling, translational motility and virulence for the Leptospira spirochete.

    PubMed

    Wunder, Elsio A; Figueira, Cláudio P; Benaroudj, Nadia; Hu, Bo; Tong, Brian A; Trajtenberg, Felipe; Liu, Jun; Reis, Mitermayer G; Charon, Nyles W; Buschiazzo, Alejandro; Picardeau, Mathieu; Ko, Albert I

    2016-08-01

    Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends and the presence of periplasmic flagella (PF) with pronounced spontaneous supercoiling. The factors that provoke such supercoiling, as well as the role that PF coiling plays in generating the characteristic hook-end cell morphology and motility, have not been elucidated. We have now identified an abundant protein from the pathogen L. interrogans, exposed on the PF surface, and named it Flagellar-coiling protein A (FcpA). The gene encoding FcpA is highly conserved among Leptospira and was not found in other bacteria. fcpA(-) mutants, obtained from clinical isolates or by allelic exchange, had relatively straight, smaller-diameter PF, and were not able to produce translational motility. These mutants lost their ability to cause disease in the standard hamster model of leptospirosis. Complementation of fcpA restored the wild-type morphology, motility and virulence phenotypes. In summary, we identified a novel Leptospira 36-kDa protein, the main component of the spirochete's PF sheath, and a key determinant of the flagella's coiled structure. FcpA is essential for bacterial translational motility and to enable the spirochete to penetrate the host, traverse tissue barriers, disseminate to cause systemic infection and reach target organs.

  16. Urinary shedding of leptospires and presence of Leptospira antibodies in healthy dogs from Upper Bavaria.

    PubMed

    Llewellyn, Julia-Rebecca; Krupka-Dyachenko, Inke; Rettinger, Anna Lena; Dyachenko, Viktor; Stamm, Ivonne; Kopp, Peter Andreas; Straubinger, Reinhard Konrad; Hartmann, Katrin

    2016-01-01

    Leptospirosis is classified as a re-emerging zoonotic disease with global impor- tance. The aim of this study was to determine urinary shedding of leptospires in healthy dogs and to identify the shedded leptospire species. Furthermore, antibody presence against leptospires was evaluated. In a prospective study urine samples of 200 healthy dogs from Upper Bavaria were randomly collected and evaluated by real-time polymerase chain reaction (PCR) specific for the lipL32 gene of pathogenic Leptospira (L) spp. Positive samples were further character- ized via multilocus sequence typing (MLST) to identify the Leptospira species. Microagglutination test (MAT) was performed to determine serum antibody titers. Three of 200 urine samples were found to be PCR-positive resulting in a urinary shedding prevalence of 1.5% (95% confidence interval 0.3-4.5%). All three dogs had been vaccinated before with a bivalent vaccine, covering the serogroups Canicola and lcterohaemorrhagiae. One dog shed leptospires of the species L. borgpetersenii, and two of the species L. interrogans. Of all dogs, 17.0% had antibody titers ≥ 1:100, and 3.5% titers ≥ 1:400 to serovars of non-vaccinal sero- groups. Healthy dogs that shed leptospires represent a possible risk for humans and other animals. The study emphasizes the importance of general hygiene measures in veterinary practice while handling urine of all dogs, and the use of vaccines that protect against a broader range of serogroups and that prevent urinary shedding. PMID:27344919

  17. Urinary shedding of leptospires and presence of Leptospira antibodies in healthy dogs from Upper Bavaria.

    PubMed

    Llewellyn, Julia-Rebecca; Krupka-Dyachenko, Inke; Rettinger, Anna Lena; Dyachenko, Viktor; Stamm, Ivonne; Kopp, Peter Andreas; Straubinger, Reinhard Konrad; Hartmann, Katrin

    2016-01-01

    Leptospirosis is classified as a re-emerging zoonotic disease with global impor- tance. The aim of this study was to determine urinary shedding of leptospires in healthy dogs and to identify the shedded leptospire species. Furthermore, antibody presence against leptospires was evaluated. In a prospective study urine samples of 200 healthy dogs from Upper Bavaria were randomly collected and evaluated by real-time polymerase chain reaction (PCR) specific for the lipL32 gene of pathogenic Leptospira (L) spp. Positive samples were further character- ized via multilocus sequence typing (MLST) to identify the Leptospira species. Microagglutination test (MAT) was performed to determine serum antibody titers. Three of 200 urine samples were found to be PCR-positive resulting in a urinary shedding prevalence of 1.5% (95% confidence interval 0.3-4.5%). All three dogs had been vaccinated before with a bivalent vaccine, covering the serogroups Canicola and lcterohaemorrhagiae. One dog shed leptospires of the species L. borgpetersenii, and two of the species L. interrogans. Of all dogs, 17.0% had antibody titers ≥ 1:100, and 3.5% titers ≥ 1:400 to serovars of non-vaccinal sero- groups. Healthy dogs that shed leptospires represent a possible risk for humans and other animals. The study emphasizes the importance of general hygiene measures in veterinary practice while handling urine of all dogs, and the use of vaccines that protect against a broader range of serogroups and that prevent urinary shedding.

  18. Highly Sensitive Loop-Mediated Isothermal Amplification for the Detection of Leptospira.

    PubMed

    Chen, Hua-Wei; Weissenberger, Giulia; Atkins, Erin; Chao, Chien-Chung; Suputtamongkol, Yupin; Ching, Wei-Mei

    2015-01-01

    Leptospirosis is a worldwide zoonosis caused by an infection with the pathogenic species of Leptospira. We have developed a loop-mediated isothermal amplification (LAMP) assay to detect the DNA of Leptospira spp. Six sets of primers targeting the gene of the subsurface protein, lipL32, were evaluated for their detection sensitivity. The best primer set detected less than 25 copies of lipL32 per reaction of both plasmid DNA template and purified leptospiral genomic DNA. By combining primers targeting lipL32 with the previously published primer set targeting lipL41, the sensitivity of the assay was improved to 12 copies of L. interrogans. The specificity of the LAMP assay was evaluated by using the genomic DNA from other clinically encountered bacterial species such as different strains of Orientia tsutsugamushi, Rickettsia typhi, Rickettsia conorii, Rickettsia rickettsii, Coxiella burnetii, and Bartonella bacilliformis. These genomic DNA samples were all negative in our LAMP assay. The sensitivity of the LAMP assay was very similar to that of quantitative real time PCR. Several detection methods for the amplified product of LAMP assay were performed to demonstrate the simplicity of the assay. In summary, our results have suggested that this assay is rapid, robust, and easy to perform and has the potential to be used in endemic locations.

  19. Isolation and Characterization of New Leptospira Genotypes from Patients in Mayotte (Indian Ocean)

    PubMed Central

    Bourhy, Pascale; Collet, Louis; Clément, Sabine; Huerre, Michel; Ave, Patrick; Giry, Claude; Pettinelli, François; Picardeau, Mathieu

    2010-01-01

    Background Leptospirosis has been implicated as a severe and fatal form of disease in Mayotte, a French-administrated territory located in the Comoros archipelago (southwestern Indian Ocean). To date, Leptospira isolates have never been isolated in this endemic region. Methods and Findings Leptospires were isolated from blood samples from 22 patients with febrile illness during a 17-month period after a PCR-based screening test was positive. Strains were typed using hyper-immune antisera raised against the major Leptospira serogroups: 20 of 22 clinical isolates were assigned to serogroup Mini; the other two strains belonged to serogroups Grippotyphosa and Pyrogenes, respectively. These isolates were further characterized using partial sequencing of 16S rRNA and ligB gene, Multi Locus VNTR Analysis (MLVA), and pulsed field gel electrophoresis (PFGE). Of the 22 isolates, 14 were L. borgpetersenii strains, 7 L. kirschneri strains, and 1, belonging to serogoup Pyrogenes, was L. interrogans. Results of the genotyping methods were consistent. MLVA defined five genotypes, whereas PFGE allowed the recognition of additional subgroups within the genotypes. PFGE fingerprint patterns of clinical strains did not match any of the patterns in the reference strains belonging to the same serogroup, suggesting that the strains were novel serovars. Conclusions Preliminary PCR screening of blood specimen allowed a high isolation frequency of leptospires among patients with febrile illness. Typing of leptospiral isolates showed that causative agents of leptospirosis in Mayotte have unique molecular features. PMID:20582311

  20. Neutrophil Extracellular Traps are Involved in the Innate Immune Response to Infection with Leptospira

    PubMed Central

    Scharrig, Emilia; Carestia, Agostina; Ferrer, María F.; Cédola, Maia; Pretre, Gabriela; Drut, Ricardo; Picardeau, Mathieu; Schattner, Mirta; Gómez, Ricardo M.

    2015-01-01

    NETosis is a process by which neutrophils extrude their DNA together with bactericidal proteins that trap and/or kill pathogens. In the present study, we evaluated the ability of Leptospira spp. to induce NETosis using human ex vivo and murine in vivo models. Microscopy and fluorometric studies showed that incubation of human neutrophils with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 (LIC) resulted in the release of DNA extracellular traps (NETs). The bacteria number, pathogenicity and viability were relevant factors for induction of NETs, but bacteria motility was not. Entrapment of LIC in the NETs resulted in LIC death; however, pathogenic but not saprophytic Leptospira sp. exerted nuclease activity and degraded DNA. Mice infected with LIC showed circulating NETs after 2 days post-infection (dpi). Depletion of neutrophils with mAb1A8 significantly reduced the amount of intravascular NETs in LIC-infected mice, increasing bacteremia at 3 dpi. Although there was a low bacterial burden, scarce neutrophils and an absence of inflammation in the early stages of infection in the kidney and liver, at the beginning of the leptospiruric phase, the bacterial burden was significantly higher in kidneys of neutrophil-depleted-mice compared to non-depleted and infected mice. Surprisingly, interstitial nephritis was of similar intensity in both groups of infected mice. Taken together, these data suggest that LIC triggers NETs, and that the intravascular formation of these DNA traps appears to be critical not only to prevent early leptospiral dissemination but also to preclude further bacterial burden. PMID:26161745

  1. Pathogenic Leptospira species express surface-exposed proteins belonging to the bacterial immunoglobulin superfamily.

    PubMed

    Matsunaga, James; Barocchi, Michele A; Croda, Julio; Young, Tracy A; Sanchez, Yolanda; Siqueira, Isadora; Bolin, Carole A; Reis, Mitermayer G; Riley, Lee W; Haake, David A; Ko, Albert I

    2003-08-01

    Proteins with bacterial immunoglobulin-like (Big) domains, such as the Yersinia pseudotuberculosis invasin and Escherichia coli intimin, are surface-expressed proteins that mediate host mammalian cell invasion or attachment. Here, we report the identification and characterization of a new family of Big domain proteins, referred to as Lig (leptospiral Ig-like) proteins, in pathogenic Leptospira. Screening of L. interrogans and L. kirschneri expression libraries with sera from leptospirosis patients identified 13 lambda phage clones that encode tandem repeats of the 90 amino acid Big domain. Two lig genes, designated ligA and ligB, and one pseudogene, ligC, were identified. The ligA and ligB genes encode amino-terminal lipoprotein signal peptides followed by 10 or 11 Big domain repeats and, in the case of ligB, a unique carboxy-terminal non-repeat domain. The organization of ligC is similar to that of ligB but contains mutations that disrupt the reading frame. The lig sequences are present in pathogenic but not saprophytic Leptospira species. LigA and LigB are expressed by a variety of virulent leptospiral strains. Loss of Lig protein and RNA transcript expression is correlated with the observed loss of virulence during culture attenuation of pathogenic strains. High-pressure freeze substitution followed by immunocytochemical electron microscopy confirmed that the Lig proteins were localized to the bacterial surface. Immunoblot studies with patient sera found that the Lig proteins are a major antigen recognized during the acute host infection. These observations demonstrate that the Lig proteins are a newly identified surface protein of pathogenic Leptospira, which by analogy to other bacterial immunoglobulin superfamily virulence factors, may play a role in host cell attachment and invasion during leptospiral pathogenesis. PMID:12890019

  2. Molecular and serological investigation of Leptospira and leptospirosis in dogs in Japan.

    PubMed

    Koizumi, Nobuo; Muto, Maki Mizutani; Akachi, Shigehiro; Okano, Shou; Yamamoto, Seigo; Horikawa, Kazumi; Harada, Seiya; Funatsumaru, Sadayuki; Ohnishi, Makoto

    2013-04-01

    Canine leptospirosis, which is caused by infection with pathogenic Leptospira species, occurs worldwide, but information regarding the causative Leptospira serotypes and genotypes and their effects on virulence in dogs remains limited. Monitoring acute leptospirosis in dogs as sentinels can also aid in estimating the risk of human leptospirosis, particularly when the disease is rare, as it currently is in Japan. Among 283 clinically suspected cases of leptospirosis diagnosed from August 2007 to March 2011 in Japan, 83 cases were laboratory diagnosed as leptospirosis by blood culture, a rise in antibody titres in paired sera using a microscopic agglutination test (MAT) and/or DNA detection using flaB-nested PCR. The infected dogs comprised hunting dogs (31 dogs) and companion animals (50 dogs) and two unknown; 63.4 % of the infected dogs were males. The mortality rate was 53.2 %. A rise of at least fourfold in MAT titre was detected in 30 dogs whose paired serum samples were obtained, and the predominant reactive serogroup was Hebdomadis (53.3 %), followed by Australis (16.7 %) and Autumnalis (16.7 %). Leptospira interrogans was isolated from 45 dogs of the following serogroups: Australis (16), Autumnalis (six), Canicola (one), Hebdomadis (21) and Icterohaemorrhagiae (one). All of these serogroups caused lethal infections (57.1-100 %). Genetic heterogeneity was demonstrated in serogroups Australis, Autumnalis and Hebdomadis by multilocus sequence typing (MLST) and/or RFLP analysis based on PFGE. In serogroup Hebdomadis, each genotype determined by MLST had a unique mortality rate in the infected dogs. Although classic canine leptospirosis is associated with serovars Canicola and Icterohaemorrhagiae, serogroup Hebdomadis has become the predominant serogroup causing high mortality in Japan. This study suggests that the virulence of members of serogroup Hebdomadis in dogs may be associated with the genotypes in this serogroup. PMID:23264455

  3. The usefulness of semi-solid medium in the isolation of highly virulent Leptospira strains from wild rats in an urban area of Fukuoka, Japan.

    PubMed

    Saito, Mitsumasa; Villanueva, Sharon Y A M; Masuzawa, Toshiyuki; Haraguchi, Yusuke; Ita, Shuhei; Miyahara, Satoshi; Ozuru, Ryo; Yamaguchi, Takayoshi; Yoshimura, Michinobu; Ikejiri, Mami; Aramaki, Natsumi; Amran, Muhammad Yunus; Muslich, Lisa Tenriesa; Iida, Ken-ichiro; Yanagihara, Yasutake; Gloriani, Nina G; Yoshida, Shin-ichi

    2015-06-01

    Leptospirosis is a worldwide zoonosis. The importance of urban leptospirosis is recognized in Japan: urban rats carry pathogenic leptospires and people acquire these pathogens through contact with surface water or soil contaminated by the urine of the infected animals. To determine the current Leptospira carriage rate in urban rats, 29 wild rats were trapped in the central area of Fukuoka and strains isolated from their kidneys and urine analyzed. When semi-solid Korthof's medium containing 0.1% agar was used for isolation, 72.2% and 30.8% of the kidney and urine cultures, respectively, were found to be Leptospira-positive. The isolates belonged to Leptospira interrogans, and were classified into two groups (serogroups Pomona and Icterohaemorrhagiae) based on the results of gyrB sequence analysis and microscopic agglutination testing (MAT). Strains belonging to serogroup Icterohemorrhagiae grew well in liquid medium. On the other hand, serogroup Pomona isolates multiplied very little in liquid medium, but did grow in a semi-solid medium. Although strains belonging to serogroup Pomona have not been recognized as native to Japan, this strain may be widely distributed in urban rats. Representative strains from each group were found to be highly pathogenic to hamsters. Our findings should serve as a warning that it is still possible to become infected with leptospires from wild rats living in inner cities of Japan. Furthermore, the use of semi-solid medium for culture will improve the isolation rate of leptospires from the kidneys of wild rats.

  4. Newly recognized Leptospira species ("Leptospira inadai" serovar lyme) isolated from human skin.

    PubMed Central

    Schmid, G P; Steere, A C; Kornblatt, A N; Kaufmann, A F; Moss, C W; Johnson, R C; Hovind-Hougen, K; Brenner, D J

    1986-01-01

    Leptospira strain 10, which represents a new Leptospira species, was isolated from a skin biopsy of a patient with Lyme disease. Although pathogenic for laboratory animals, the organism was not considered to have a significant role in the patient's illness. PMID:3760144

  5. 77 FR 43827 - International Workshop on Alternative Methods for Leptospira

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-26

    ... HUMAN SERVICES International Workshop on Alternative Methods for Leptospira Vaccine Potency Testing... for the Evaluation of Alternative Toxicological Methods (NICEATM) announces an ``International Workshop on Alternative Methods for Leptospira Vaccine Potency Testing: State of the Science and the...

  6. Isolation and molecular characterization of Leptospira borgpetersenii serovar Hardjo strain Hardjobovis in the urine of naturally infected cattle in Brazil.

    PubMed

    Chideroli, R T; Pereira, U P; Gonçalves, D D; Nakamura, A Y; Alfieri, A A; Alfieri, A F; Freitas, J C

    2016-02-19

    Most epidemiologic studies on bovine leptospirosis are based on serological tests that use antibodies against several serotypes, including the serovar Hardjo, which is widespread and considered to be the most adapted to bovine hosts. However, using only serological studies is not sufficient to identify and distinguish species of leptospires. The aim of this study was report the first isolation in Brazil of two strains serovar Hardjo obtained in urine samples from naturally infected cows in a small Brazilian dairy herd and find the genetic species and consequently the type strain Hardjobovis by molecular characterization. Fifteen dairy cows with a history of reproductive failure, such as abortion and infertility, were selected. Urine samples obtained from each animal were immediately seeded in tubes containing Ellinghausen-McCullough-Johnson-Harris culture medium. The identification of the isolates was performed by Multilocus variable-number tandem-repeat analysis (MLVA) technique and phylogenetic analysis of partial sequence of gene sec Y. From the 15 urine samples evaluated, two Leptospira were found and identified as the Londrina 49 and Londrina 54 strains. The MLVA profiles and sequencing of gene sec Y characterized the isolates as L. borgpetersenii serovar Hardjo strain Hadjobovis because it has different genetic pattern of Leptospira interrogans serovar Hardjo strain Hardjoprajitno. Therefore, more studies are needed including isolation and molecular characterization from regional strains to obtain a better knowledge about epidemiology of serovar Hardjo in bovine which may assist in future strategies of prevention and control of bovine leptospirosis.

  7. Leptospira immunoglobulin-like proteins as a serodiagnostic marker for acute leptospirosis.

    PubMed

    Croda, Julio; Ramos, João G R; Matsunaga, James; Queiroz, Adriano; Homma, Akira; Riley, Lee W; Haake, David A; Reis, Mitermayer G; Ko, Albert I

    2007-05-01

    There is an urgent need for improved diagnosis of leptospirosis, an emerging infectious disease which imparts a large disease burden in developing countries. We evaluated the use of Leptospira immunoglobulin (Ig)-like (Lig) proteins as a serodiagnostic marker for leptospirosis. Lig proteins have bacterial immunoglobulin-like (Big) tandem repeat domains, a moiety found in virulence factors in other pathogens. Sera from patients identified during urban outbreaks in Brazil reacted strongly with immunoblots of a recombinant fragment comprised of the second to sixth Big domains of LigB from L. interrogans serovar Copenhageni, the principal agent for transmission in this setting. Furthermore, the sera recognized an analogous LigB fragment derived from L. kirschneri serovar Grippotyphosa, a pathogenic serovar which is not endemic to the study area. The immunoblot assay detected anti-LigB IgM antibodies in sera from 92% (95% confidence interval, 85 to 96%) of patients during acute-phase leptospirosis. The assay had a sensitivity of 81% for sera from patients with less than 7 days of illness. Anti-LigB antibodies were found in sera from 57% of the patients who did not have detectable anti-whole-Leptospira responses as detected by IgM enzyme-linked immunosorbent assay and microagglutination test. The specificities of the assay were 93 to 100% and 90 to 97% among sera from healthy individuals and patients with diseases that have clinical presentations that overlap with those of leptospirosis, respectively. These findings indicate that the antibody response to this putative virulence determinant is a sensitive and specific marker for acute infection. The use of this marker may aid the prompt and timely diagnosis required to reduce the high mortality associated with severe forms of the disease. PMID:17360842

  8. Para-(benzoyl)-phenylalanine as a potential inhibitor against LpxC of Leptospira spp.: homology modeling, docking, and molecular dynamics study.

    PubMed

    Pradhan, Dibyabhaba; Priyadarshini, Vani; Munikumar, Manne; Swargam, Sandeep; Umamaheswari, Amineni; Bitla, Aparna

    2014-01-01

    Leptospira interrogans, a Gram-negative bacterial pathogen is the main cause of human leptospirosis. Lipid A is a highly immunoreactive endotoxic center of lipopolysaccharide (LPS) that anchors LPS into the outer membrane of Leptospira. Discovery of compounds inhibiting lipid-A biosynthetic pathway would be promising for dissolving the structural integrity of membrane leading to cell lysis and death of Leptospira. LpxC, a unique enzyme of lipid-A biosynthetic pathway was identified as common drug target of Leptospira. Herein, homology modeling, docking, and molecular dynamics (MD) simulations were employed to discover potential inhibitors of LpxC. A reliable tertiary structure of LpxC in complex with inhibitor BB-78485 was constructed in Modeller 9v8. A data-set of BB-78485 structural analogs were docked with LpxC in Maestro v9.2 virtual screening workflow, which implements three stage Glide docking protocol. Twelve lead molecules with better XP Gscore compared to BB-78485 were proposed as potential inhibitors of LpxC. Para-(benzoyl)-phenylalanine - that showed lowest XP Gscore (-10.35 kcal/mol) - was predicted to have best binding affinity towards LpxC. MD simulations were performed for LpxC and para-(benzoyl)-phenylalanine docking complex in Desmond v3.0. Trajectory analysis showed the docking complex and inter-molecular interactions was stable throughout the entire production part of MD simulations. The results indicate para-(benzoyl)-phenylalanine as a potent drug molecule against leptospirosis. An animated Interactive 3D Complement (I3DC) is available in Proteopedia at http://proteopedia.org/w/Journal:JBSD:10. PMID:23383626

  9. Para-(benzoyl)-phenylalanine as a potential inhibitor against LpxC of Leptospira spp.: homology modeling, docking, and molecular dynamics study.

    PubMed

    Pradhan, Dibyabhaba; Priyadarshini, Vani; Munikumar, Manne; Swargam, Sandeep; Umamaheswari, Amineni; Bitla, Aparna

    2014-01-01

    Leptospira interrogans, a Gram-negative bacterial pathogen is the main cause of human leptospirosis. Lipid A is a highly immunoreactive endotoxic center of lipopolysaccharide (LPS) that anchors LPS into the outer membrane of Leptospira. Discovery of compounds inhibiting lipid-A biosynthetic pathway would be promising for dissolving the structural integrity of membrane leading to cell lysis and death of Leptospira. LpxC, a unique enzyme of lipid-A biosynthetic pathway was identified as common drug target of Leptospira. Herein, homology modeling, docking, and molecular dynamics (MD) simulations were employed to discover potential inhibitors of LpxC. A reliable tertiary structure of LpxC in complex with inhibitor BB-78485 was constructed in Modeller 9v8. A data-set of BB-78485 structural analogs were docked with LpxC in Maestro v9.2 virtual screening workflow, which implements three stage Glide docking protocol. Twelve lead molecules with better XP Gscore compared to BB-78485 were proposed as potential inhibitors of LpxC. Para-(benzoyl)-phenylalanine - that showed lowest XP Gscore (-10.35 kcal/mol) - was predicted to have best binding affinity towards LpxC. MD simulations were performed for LpxC and para-(benzoyl)-phenylalanine docking complex in Desmond v3.0. Trajectory analysis showed the docking complex and inter-molecular interactions was stable throughout the entire production part of MD simulations. The results indicate para-(benzoyl)-phenylalanine as a potent drug molecule against leptospirosis. An animated Interactive 3D Complement (I3DC) is available in Proteopedia at http://proteopedia.org/w/Journal:JBSD:10.

  10. [Research on the recombinant plasmid pDJH2 of L. interrogans serovar lai: sequencing and alignment with other known bacterial Omp sequence].

    PubMed

    Jiang, N; Dai, B; Yan, Z; Yang, W; Li, S; Fang, Z; Zhao, H; Wu, W; Ye, D; Yan, R; Liu, J; Song, S; Yang, Y; Zhang, Y; Liu, F; Tu, Y; Yang, H; Huang, Z; Liang, L; Hu, L; Zhao, M

    1996-12-01

    The Leptospira whole cell vaccine (LWCV) currently used in China is safe and effective, out the immunity following vaccination with two doses of the fluid medium vaccine is of low order. The duration of immunity conferred by this vaccine is rather short, six months or at most one year. Therefore, it is necessary to develop new generation vaccines against Leptospirosis for the developing world. In this paper we report the sequencing of the insert fragment of pDJH2 from genomic DNA of L. interrogans sevovar lai strain 017 and its alignment with other bacterial omp sequences. A genomic library of Leptospira interrogaans serovar lai strain 017 was constructed with the plasmid vector pUC18. A recombinant plasmid designated pJDH2 was screened from the genomic library. Inserted fragment of pDH2 is 1.9 kb by gel electrophoresis. Immunization/protection was studied in BALB/c mice model. The results showed highly significant difference between pDJH2 and pUC18 (control). Inserted fragment of pDJH2 DNA sequencing was performed by Dr Yan Zhengxin (Max-Planck-Institut for Biology. Tubingen, Germany). Insert fragment was cloned into pBluescript II KS-(stratagene) and sequenced by using AB1 (Applied Bio Systems, Model 373A). Two open reading frames of 565 and 662 nucleotides were identified. There were identifiable initiation codons, terminators, Shine-Dalgano ribosome combining site, Pribnow boxes and Sextama boxes within the 2 sequenced regions. Nucleotide sequences were analysed using Gene Work, a suit of computer program developed by Department of Biochemistry St. Jude Children's Research Hospital Memphis. U.S.A. The results of formatted alignment showed the predicted nucleotide sequence of ORF1 of the serovar lai had significant similarity with ORF2 (49.36%). L. kirschneri ompL1 (49.26%), Borrelia burgdoferi omp (48.97%), Treponema phagedenis omp (47.3%); Salmonella typhimurium ompC(46.87%), Yersinia enterocolitica ompH (46.7%), Leptospira borgpeterseni pfap (46.3%), and

  11. Methylation and in vivo expression of the surface-exposed Leptospira interrogans outer membrane protein OmpL32

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recent studies have revealed that bacterial protein methylation is a widespread post-translational modification that is required for virulence in selected pathogenic bacteria. In particular, altered methylation of outer membrane proteins has been shown to modulate the effectiveness of the host immu...

  12. Isolation of Leptospira from a phocid: acute renal failure and mortality from Leptospirosis in rehabilitated northern elephant seals (Mirounga angustirostris), California, USA.

    PubMed

    Delaney, Martha A; Colegrove, Kathleen M; Spraker, Terry R; Zuerner, Richard L; Galloway, Renee L; Gulland, Frances M D

    2014-07-01

    During rehabilitation, acute renal failure due to leptospirosis occurred in eight male northern elephant seals (Mirounga angustirostris) that stranded along the central California coast in 2011. Characteristic histologic lesions including renal tubular degeneration, necrosis, and mineralization, and mild lymphoplasmacytic interstitial nephritis were noted in the six animals examined. Immunohistochemistry, bacterial culture, and PCR were positive in 2/3, 2/3, and 3/4 seals, respectively, and 6/8 had high serum antibody titers to Leptospira interrogans serovar pomona. Pulsed-field gel electrophoresis confirmed one isolate as serovar pomona. Variable number tandem repeat (VNTR) analysis showed both elephant seal isolates were identical to each other but distinct from those isolated from California sea lions (Zalophus californianus). The time from stranding to onset of azotemia was 1 to 38 (median=24) days, suggesting some seals were infected at the rehabilitation facility. Based on temporal and spatial incidence of infection, transmission among elephant seals likely occurred during rehabilitation. Molecular (VNTR) analysis of the two isolates indicates there is a unique L. interrogans serovar pomona genotype in elephant seals, and sea lions were not the source of infection prior to or during rehabilitation. This study confirms the susceptibility of northern elephant seals to leptospirosis, indicates intraspecies transmission during rehabilitation, and reports the first isolation and preliminary characterization of leptospires from elephant seals. PMID:24807176

  13. Isolation of Leptospira from a phocid: acute renal failure and mortality from Leptospirosis in rehabilitated northern elephant seals (Mirounga angustirostris), California, USA.

    PubMed

    Delaney, Martha A; Colegrove, Kathleen M; Spraker, Terry R; Zuerner, Richard L; Galloway, Renee L; Gulland, Frances M D

    2014-07-01

    During rehabilitation, acute renal failure due to leptospirosis occurred in eight male northern elephant seals (Mirounga angustirostris) that stranded along the central California coast in 2011. Characteristic histologic lesions including renal tubular degeneration, necrosis, and mineralization, and mild lymphoplasmacytic interstitial nephritis were noted in the six animals examined. Immunohistochemistry, bacterial culture, and PCR were positive in 2/3, 2/3, and 3/4 seals, respectively, and 6/8 had high serum antibody titers to Leptospira interrogans serovar pomona. Pulsed-field gel electrophoresis confirmed one isolate as serovar pomona. Variable number tandem repeat (VNTR) analysis showed both elephant seal isolates were identical to each other but distinct from those isolated from California sea lions (Zalophus californianus). The time from stranding to onset of azotemia was 1 to 38 (median=24) days, suggesting some seals were infected at the rehabilitation facility. Based on temporal and spatial incidence of infection, transmission among elephant seals likely occurred during rehabilitation. Molecular (VNTR) analysis of the two isolates indicates there is a unique L. interrogans serovar pomona genotype in elephant seals, and sea lions were not the source of infection prior to or during rehabilitation. This study confirms the susceptibility of northern elephant seals to leptospirosis, indicates intraspecies transmission during rehabilitation, and reports the first isolation and preliminary characterization of leptospires from elephant seals.

  14. Leptospira Exposure and Gardeners: A Case-Control Seroprevalence Study

    PubMed Central

    Alvarado-Esquivel, Cosme; Hernandez-Tinoco, Jesus; Sanchez-Anguiano, Luis Francisco; Ramos-Nevarez, Agar; Cerrillo-Soto, Sandra Margarita; Guido-Arreola, Carlos Alberto

    2016-01-01

    Background Leptospira can be found in soil. However, it is unclear whether occupational exposure to soil may represent a risk for Leptospira infection in humans. Therefore, we sought to determine the association of Leptospira IgG seroprevalence with the occupation of gardener, and to determine the epidemiological characteristics of gardeners associated with Leptospira exposure. Methods We performed a case-control study in 168 gardeners and 168 age- and gender-matched control subjects without gardening occupation in Durango City, Mexico. The seroprevalence of anti-Leptospira IgG antibodies in cases and controls was determined using an enzyme immunoassay. Bivariate and multivariate analyses were used to assess the association of Leptospira exposure and the characteristics of the gardeners. Results Anti-Leptospira IgG antibodies were found in 10 (6%) of 168 gardeners and in 15 (8.9%) of 168 control subjects (odds ratio (OR): 0.64; 95% confidence interval (CI): 0.28 - 1.48; P = 0.40). Multivariate analysis showed that Leptospira seropositivity was positively associated with female gender (OR: 5.82; 95% CI: 1.11 - 30.46; P = 0.03), and negatively associated with eating while working (OR: 0.21; 95% CI: 0.05 - 0.87; P = 0.03). In addition, multivariate analysis showed that high anti-Leptospira levels were associated with consumption of boar meat (OR: 28.00; 95% CI: 1.20 - 648.80; P = 0.03). Conclusions This is the first case-control study of Leptospira exposure in gardeners. Results do not support an association of Leptospira exposure with the occupation of gardener. However, further studies to confirm the lack of this association are needed. The potential role of consumption of boar meat in Leptospira infection deserves further investigation. PMID:26668679

  15. Molecular modeling and in-silico engineering of Cardamom mosaic virus coat protein for the presentation of immunogenic epitopes of Leptospira LipL32.

    PubMed

    Kumar, Vikram; Damodharan, S; Pandaranayaka, Eswari P J; Madathiparambil, Madanan G; Tennyson, Jebasingh

    2016-01-01

    Expression of Cardamom mosaic virus (CdMV) coat protein (CP) in E. coli forms virus-like particles. In this study, the structure of CdMV CP was predicted and used as a platform to display epitopes of the most abundant surface-associated protein, LipL32 of Leptospira at C, N, and both the termini of CdMV CP. In silico, we have mapped sequential and conformational B-cell epitopes from the crystal structure of LipL32 of Leptospira interrogans serovar Copenhageni str. Fiocruz L1-130 using IEDB Elipro, ABCpred, BCPRED, and VaxiJen servers. Our results show that the epitopes displayed at the N-terminus of CdMV CP are promising vaccine candidates as compared to those displayed at the C-terminus or at both the termini. LipL32 epitopes, EP2, EP3, EP4, and EP6 are found to be promising B-cell epitopes for vaccine development. Based on the type of amino acids, length, surface accessibility, and docking energy with CdMV CP model, the order of antigenicity of the LipL32 epitopes was found to be EP4 > EP3 > EP2 > EP6. PMID:25692534

  16. Molecular modeling and in-silico engineering of Cardamom mosaic virus coat protein for the presentation of immunogenic epitopes of Leptospira LipL32.

    PubMed

    Kumar, Vikram; Damodharan, S; Pandaranayaka, Eswari P J; Madathiparambil, Madanan G; Tennyson, Jebasingh

    2016-01-01

    Expression of Cardamom mosaic virus (CdMV) coat protein (CP) in E. coli forms virus-like particles. In this study, the structure of CdMV CP was predicted and used as a platform to display epitopes of the most abundant surface-associated protein, LipL32 of Leptospira at C, N, and both the termini of CdMV CP. In silico, we have mapped sequential and conformational B-cell epitopes from the crystal structure of LipL32 of Leptospira interrogans serovar Copenhageni str. Fiocruz L1-130 using IEDB Elipro, ABCpred, BCPRED, and VaxiJen servers. Our results show that the epitopes displayed at the N-terminus of CdMV CP are promising vaccine candidates as compared to those displayed at the C-terminus or at both the termini. LipL32 epitopes, EP2, EP3, EP4, and EP6 are found to be promising B-cell epitopes for vaccine development. Based on the type of amino acids, length, surface accessibility, and docking energy with CdMV CP model, the order of antigenicity of the LipL32 epitopes was found to be EP4 > EP3 > EP2 > EP6.

  17. 9 CFR 113.105 - Leptospira Hardjo Bacterin.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Leptospira Hardjo Bacterin. 113.105 Section 113.105 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF... produced from a culture of Leptospira hardjo which has been inactivated and is nontoxic. Each serial...

  18. 9 CFR 113.105 - Leptospira Hardjo Bacterin.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Leptospira Hardjo Bacterin. 113.105 Section 113.105 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF... produced from a culture of Leptospira hardjo which has been inactivated and is nontoxic. Each serial...

  19. Shedding and seroprevalence of pathogenic Leptospira spp. in sheep and cattle at a New Zealand Abattoir.

    PubMed

    Fang, F; Collins-Emerson, J M; Cullum, A; Heuer, C; Wilson, P R; Benschop, J

    2015-06-01

    A cross-sectional study was carried out on sheep and cattle slaughtered at a New Zealand abattoir from September to November 2010 to investigate the supplier-specific shedding rate, renal carriage rate and seroprevalence of leptospires. In the 2008/2009 season, this abattoir experienced three human leptospirosis cases from 20 staff, of which two were hospitalized. Urine, kidney and blood samples were collected from carcasses of 399 sheep (six suppliers, 17 slaughter lines) and 146 cattle (three suppliers, 22 slaughter lines). The urine and kidney samples were tested by quantitative real-time PCR (qPCR), while serum samples (from coagulated blood samples) were tested by microscopic agglutination test (MAT). In total, 27% (73/274; 95% CI: 18-37) of urine samples tested positive by qPCR. Species-specific shedding rates (prevalence of positive urine qPCR) were 31% (95% CI: 17-48) for sheep and 21% (95% CI: 14-30) for cattle. For 545 kidney samples tested, 145 were qPCR positive (27%; 95% CI: 17-39). The average prevalence of kidney qPCR positivity was 29% (95% CI: 17-45) for sheep and 21% (95% CI: 15-28) for cattle. Three hundred and thirty of 542 sampled sheep and cattle had antibodies against Leptospira borgpetersenii serovar Hardjobovis (Hardjobovis) and/or Leptospira interrogans serovar Pomona (Pomona), based on reciprocal MAT titre ≥1 : 48 (overall seroprevalence of 61%; 95% CI: 48-73). Seroprevalence was 57% (95% CI: 40-72) for sheep and 73% (95% CI: 59-83) for cattle. Among the seropositive animals, 41% (70/170; 95% CI: 30-54) were shedding (tested positive by urine qPCR) and 42% (137/330; 95% CI: 30-54) had renal carriage (tested positive by kidney qPCR). Some risk management options for abattoirs or farms to prevent human leptospirosis infections include vaccination of maintenance hosts, the use of personal protective equipment, and the application of urine qPCR to detect shedding status of stock as surveillance and as an alert.

  20. Molecular Typing of Pathogenic Leptospira Serogroup Icterohaemorrhagiae Strains Circulating in China during the Past 50 Years

    PubMed Central

    Zhang, Cuicai; Yang, Huimian; Li, Xiuwen; Cao, Zhiqiang; Zhou, Haijian; Zeng, Linzi; Xu, Jianmin; Xu, Yinghua; Chang, Yung-Fu; Guo, Xiaokui; Zhu, Yongzhang; Jiang, Xiugao

    2015-01-01

    Background Leptospirosis is one of the most important neglected tropical infectious diseases worldwide. Icterohaemorrhagiae has been throughout recent history, and still is, the predominant serogroup of this pathogen in China. However, very little in detail is known about the serovars or genotypes of this serogroup. Methodology/Principal Findings In this study, 120 epidemic strains from five geographically diverse regions in China collected over a 50 year period (1958~2008), and 8 international reference strains characterized by 16S rRNA sequencing and MLST analysis. 115, 11 and 2 strains were identified as L. interrogans, L. borgpetersenii, and L. kirschneri, respectively. 17 different STs were identified including 69 ST1 strains, 18 ST17, 18 ST128, 9 ST143 and 2 ST209. The remaining 12 strains belonged to 12 different STs. eBURST analysis demonstrated that, among the clonal complexes isolated (CCs), CC1 accounted for 73.3% (88/120) strains representing three STs: ST1, ST128 and ST98. ST1 was the most likely ancestral strain of this CC, followed by singleton CC17 (17/120) and CC143 (11/120). Further analysis of adding 116 serogroup Icterohaemorrhagiae strains in the MLST database and studies previously described using global eBURST analysis and MST dendrogram revealed relatively similar ST clustering patterns with five main CCs and 8 singletons among these 244 strains. CC17 was found to be the most prevalent clone of pathogenic Leptospira circulating worldwide. This is the first time, to our knowledge, that ST1 and ST17 strains were distributed among 4 distinct serovars, indicating a highly complicated relationship between serovars and STs. Conclusions/Significance Our studies demonstrated a high level of genetic diversity in the serogroup Icterohaemorrhagiae strains. Distinct from ST17 or ST37 circulating elsewhere, ST1 included in CC1, has over the past 50 years or so, proven to be the most prevalent ST of pathogenic leptospires isolated in China. Moreover, the

  1. Genome Sequence of the Saprophyte Leptospira Biflexa Provides Insights into the Evolution of Leptospira and the Pathogenesis of Leptospirosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospira biflexa is a free-living spirochete found broadly distributed in aquatic environments, and is the primary model for studying gene function in Leptospira spp. The L. biflexa genome has 3,590 protein-coding genes (excluding transposases) distributed across three circular replicons: two of w...

  2. Leptospira species in floodwater during the 2011 floods in the Bangkok Metropolitan Region, Thailand.

    PubMed

    Thaipadungpanit, Janjira; Wuthiekanun, Vanaporn; Chantratita, Narisara; Yimsamran, Surapon; Amornchai, Premjit; Boonsilp, Siriphan; Maneeboonyang, Wanchai; Tharnpoophasiam, Prapin; Saiprom, Natnaree; Mahakunkijcharoen, Yuvadee; Day, Nicholas P J; Singhasivanon, Pratap; Peacock, Sharon J; Limmathurotsakul, Direk

    2013-10-01

    Floodwater samples (N = 110) collected during the 2011 Bangkok floods were tested for Leptospira using culture and polymerase chain reaction (PCR); 65 samples were PCR-positive for putatively non-pathogenic Leptospira species, 1 sample contained a putatively pathogenic Leptospira, and 6 samples contained Leptospira clustering phylogenetically with the intermediate group. The low prevalence of pathogenic and intermediate Leptospira in floodwater was consistent with the low number of human leptospirosis cases reported to the Bureau of Epidemiology in Thailand. This study provides baseline information on environmental Leptospira in Bangkok together with a set of laboratory tests that could be readily deployed in the event of future flooding.

  3. Factor analysis of serogroups botanica and aurisina of Leptospira biflexa.

    PubMed

    Cinco, M

    1977-11-01

    Factor analysis is performed on serovars of Botanica and Aurisina serogroup of Leptospira biflexa. The results show the arrangement of main factors serovar and serogroup specific, as well as the antigens common with serovars of heterologous serogroups.

  4. Synanthropic Cockroaches (Blattidae: Periplaneta spp.) Harbor Pathogenic Leptospira in Colombia.

    PubMed

    Gonzalez-Astudillo, Viviana; Bustamante-Rengifo, Javier A; Bonilla, Álvaro; Lehmicke, Anna Joy J; Castillo, Andrés; Astudillo-Hernández, Miryam

    2016-01-01

    Leptospirosis cases in Colombia are typically linked to peridomestic rodents; however, empirical data suggest that Leptospira-infected patients with no apparent exposure to these reservoirs are common. Cockroaches (Periplaneta spp.) have equal or greater interaction with humans than rodents, yet their potential role as carriers of Leptospira has not been assessed. We determined if pathogenic Leptospira is harbored by Periplaneta spp. in Cali (Colombia) and the variables influencing this relationship. Fifty-nine cockroaches were captured from seven sites and DNA was extracted from the body surface and digestive tract for a multiplex polymerase chain reaction, targeting genes secY and flaB. Logistic regression models and proportion tests showed a higher likelihood for Leptospira to be isolated from body surfaces (P > 0.001) and from individuals inside houses (six times more likely). These findings are the first to demonstrate an association between Periplaneta spp. and Leptospira, suggesting the need to investigate the potential for cockroaches to serve as reservoirs or transport hosts for Leptospira.

  5. Microcinematographic analysis of tethered Leptospira illini.

    PubMed Central

    Charon, N W; Daughtry, G R; McCuskey, R S; Franz, G N

    1984-01-01

    A model of Leptospira motility was recently proposed. One element of the model states that in translating cells the anterior spiral-shaped end gyrates counterclockwise and the posterior hook-shaped end gyrates clockwise. We tested these predictions by analyzing cells tethered to a glass surface. Leptospira illini was incubated with antibody-coated latex beads (Ab-beads). These beads adhered to the cells, and subsequently some cells became attached to either the slide or the cover glass via the Ab-beads. As previously reported, these cells rapidly moved back and forth across the surface of the beads. In addition, a general trend was observed: cells tethered to the cover glass rotated clockwise around the Ab-bead; cells tethered to the slide rotated counterclockwise around the Ab-bead. A computer-aided microcinematographic analysis of tethered cells indicated that the direction of rotation of cells around the Ab-bead was a function of both the surface of attachment and the shape of the cell ends. The results can best be explained by assuming that the gyrating ends interact with the glass surface to cause rotation around the Ab-beads. The analysis obtained indicates that the hook- and spiral-shaped ends rotate in the directions predicted by the model. In addition, the tethered cell assay permitted detection of rapid, coordinated reversals of the cell ends, e.g., cells rapidly switched from a hook-spiral configuration to a spiral-hook configuration. These results suggest the existance of a mechanism which coordinates the shape of the cell ends of L. illini. Images PMID:6501226

  6. Contribution of Leptospira, Neospora caninum and bovine viral diarrhea virus to fetal loss of beef cattle in New Zealand.

    PubMed

    Sanhueza, J M; Heuer, C; West, D

    2013-10-01

    The profitability of beef breeding farms in New Zealand depends principally on optimal reproductive performance. The aim of this study was to estimate the impact of four major pathogens, bovine viral diarrhea virus (BVDV), Neospora caninum (N. caninum), Leptospira borgpetersenii serovar Hardjo (Hardjo), and Leptospira interrogans serovar Pomona (Pomona), on rates of fetal loss in commercial beef breeding herds. Farms reporting fetal loss were recruited, and a blood sample from aborting cows (cases) was collected. Controls were normally calving cows from the same farm. At least four controls were selected from each farm contributing cases. Samples were tested using ELISA for detection of antibodies against BVDV and N. caninum, and microscopic agglutination test (MAT) for detection of antibody against Hardjo and Pomona. A selection of titer cut-offs was conducted to evaluate the relationship between fetal loss and seropositivity to each pathogen using conditional logistic regression. The cut-off titer with the strongest association with fetal loss was included in the multivariate model. A significant increased risk of fetal loss was found for animals seropositive to N. caninum (odds ratio (OR)=3.36; 95% confidence interval (95% CI)=1.27-8.89), Hardjo (OR=1.84; 95% CI=1.01-3.33), and Pomona in non-vaccinated cows (OR=14.91, 95% CI=1.73-128.84) at the ELISA titer ≥ 30, and MAT titers of ≥ 1:384 and ≥ 1:768 for a positive sample, respectively. A marginally non-significant increased risk of fetal loss was found for animals exposed to BVDV (OR=2.01; 95% CI=0.99-4.11) at the ELISA titer of ≤ 1. Vaccination did not affect ORs for Hardjo or BVDV and no herd vaccinated against N. caninum. Approximately 14.0% of all fetal loss in the beef breeding cattle population in New Zealand may be attributable to BVDV (3.5%), N. caninum (3.0%), Hardjo (4.7%), and Pomona (3.6%).

  7. A unique strain of Leptospira isolated from a patient with pulmonary haemorrhages in the Andaman Islands: a proposal of serovar portblairi of serogroup Sehgali.

    PubMed Central

    Vijayacharit, P.; Hartskeerl, R. A.; Sharma, S.; Natarajaseenivasan, K.; Roy, S.; Terpstra, W. J.; Sehgal, S. C.

    2004-01-01

    Leptospirosis is endemic in the Andaman Islands, often occurring as outbreaks during the post-monsoon period. Pulmonary involvement is common and associated with high morbidity and mortality. During the investigation of an outbreak in North Andaman in 1996 an isolate was recovered from the blood of a patient with fever, headache, body aches and haemoptysis with respiratory distress as presenting symptoms. The isolate was characterized using the cross-agglutination absorption test (CAAT) and monoclonal antibodies (mAbs). The isolate showed typical morphology and characteristic motility of the genus Leptospira. Growth was inhibited at 13 degrees C and in the presence of 8-azaguanine. The isolate could not be identified with grouping sera representing 25 serogroups, CAAT and mAbs. A new serovar of a new serogroup is proposed. Genetic characterization using polymerase chain reaction (PCR) followed by sequencing of the PCR product and randomly amplified polymorphic DNA fingerprinting (RAPD) showed that the isolate was genetically similar to L. interrogans sensu stricto. PMID:15310168

  8. Seroepidemiology of Leptospira Exposure in General Population in Rural Durango, Mexico.

    PubMed

    Alvarado-Esquivel, Cosme; Sánchez-Anguiano, Luis Francisco; Hernández-Tinoco, Jesús

    2015-01-01

    The magnitude of Leptospira exposure in rural Mexico is largely unknown. We sought to determine the seroprevalence of Leptospira IgG antibodies in adults in rural Durango, Mexico, and to determine the sociodemographic, behavioral, and housing characteristics of the subjects associated with Leptospira seropositivity. We performed a cross-sectional study in 282 adults living in rural Durango, Mexico. Sera from participants were analyzed for Leptospira IgG antibodies using a commercially available enzyme immunoassay. Seroprevalence association with the characteristics of the subjects was analyzed by bivariate and multivariate analyses. Of the 282 rural subjects (42.91 ± 17.53 years old) studied, 44 (15.6%) had anti-Leptospira IgG antibodies. Seropositivity to Leptospira was not associated with gender, educational level, employment, socioeconomic status, contact with animals or soil, or type of floors at home. In contrast, multivariate analysis showed that Leptospira exposure was associated with national trips (OR = 2.09; 95% CI: 1.05-4.16; P = 0.03) and poor education of the head of the family (OR = 2.96; 95% CI: 1.51-5.78; P = 0.001). We demonstrated serological evidence of Leptospira exposure in adults in rural northern Mexico. The contributing factors associated with Leptospira exposure found in the present study may be useful for optimal planning of preventive measures against Leptospira infection.

  9. Seroepidemiology of Leptospira Exposure in General Population in Rural Durango, Mexico

    PubMed Central

    Alvarado-Esquivel, Cosme; Sánchez-Anguiano, Luis Francisco; Hernández-Tinoco, Jesús

    2015-01-01

    The magnitude of Leptospira exposure in rural Mexico is largely unknown. We sought to determine the seroprevalence of Leptospira IgG antibodies in adults in rural Durango, Mexico, and to determine the sociodemographic, behavioral, and housing characteristics of the subjects associated with Leptospira seropositivity. We performed a cross-sectional study in 282 adults living in rural Durango, Mexico. Sera from participants were analyzed for Leptospira IgG antibodies using a commercially available enzyme immunoassay. Seroprevalence association with the characteristics of the subjects was analyzed by bivariate and multivariate analyses. Of the 282 rural subjects (42.91 ± 17.53 years old) studied, 44 (15.6%) had anti-Leptospira IgG antibodies. Seropositivity to Leptospira was not associated with gender, educational level, employment, socioeconomic status, contact with animals or soil, or type of floors at home. In contrast, multivariate analysis showed that Leptospira exposure was associated with national trips (OR = 2.09; 95% CI: 1.05–4.16; P = 0.03) and poor education of the head of the family (OR = 2.96; 95% CI: 1.51–5.78; P = 0.001). We demonstrated serological evidence of Leptospira exposure in adults in rural northern Mexico. The contributing factors associated with Leptospira exposure found in the present study may be useful for optimal planning of preventive measures against Leptospira infection. PMID:26240822

  10. What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira.

    PubMed

    Fouts, Derrick E; Matthias, Michael A; Adhikarla, Haritha; Adler, Ben; Amorim-Santos, Luciane; Berg, Douglas E; Bulach, Dieter; Buschiazzo, Alejandro; Chang, Yung-Fu; Galloway, Renee L; Haake, David A; Haft, Daniel H; Hartskeerl, Rudy; Ko, Albert I; Levett, Paul N; Matsunaga, James; Mechaly, Ariel E; Monk, Jonathan M; Nascimento, Ana L T; Nelson, Karen E; Palsson, Bernhard; Peacock, Sharon J; Picardeau, Mathieu; Ricaldi, Jessica N; Thaipandungpanit, Janjira; Wunder, Elsio A; Yang, X Frank; Zhang, Jun-Jie; Vinetz, Joseph M

    2016-02-01

    Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade's refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic

  11. What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira.

    PubMed

    Fouts, Derrick E; Matthias, Michael A; Adhikarla, Haritha; Adler, Ben; Amorim-Santos, Luciane; Berg, Douglas E; Bulach, Dieter; Buschiazzo, Alejandro; Chang, Yung-Fu; Galloway, Renee L; Haake, David A; Haft, Daniel H; Hartskeerl, Rudy; Ko, Albert I; Levett, Paul N; Matsunaga, James; Mechaly, Ariel E; Monk, Jonathan M; Nascimento, Ana L T; Nelson, Karen E; Palsson, Bernhard; Peacock, Sharon J; Picardeau, Mathieu; Ricaldi, Jessica N; Thaipandungpanit, Janjira; Wunder, Elsio A; Yang, X Frank; Zhang, Jun-Jie; Vinetz, Joseph M

    2016-02-01

    Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade's refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic

  12. What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira

    PubMed Central

    Fouts, Derrick E.; Matthias, Michael A.; Adhikarla, Haritha; Adler, Ben; Amorim-Santos, Luciane; Berg, Douglas E.; Bulach, Dieter; Buschiazzo, Alejandro; Chang, Yung-Fu; Galloway, Renee L.; Haake, David A.; Haft, Daniel H.; Hartskeerl, Rudy; Ko, Albert I.; Levett, Paul N.; Matsunaga, James; Mechaly, Ariel E.; Monk, Jonathan M.; Nascimento, Ana L. T.; Nelson, Karen E.; Palsson, Bernhard; Peacock, Sharon J.; Picardeau, Mathieu; Ricaldi, Jessica N.; Thaipandungpanit, Janjira; Wunder, Elsio A.; Yang, X. Frank; Zhang, Jun-Jie; Vinetz, Joseph M.

    2016-01-01

    Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade’s refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic

  13. 9 CFR 113.105 - Leptospira Hardjo Bacterin.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... produced from a culture of Leptospira hardjo which has been inactivated and is nontoxic. Each serial of... and shall be tested for purity, safety, and potency as prescribed in this section. A serial found... completed product from each serial and each subserial shall be tested for viable bacteria and fungi...

  14. 9 CFR 113.105 - Leptospira Hardjo Bacterin.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... produced from a culture of Leptospira hardjo which has been inactivated and is nontoxic. Each serial of... and shall be tested for purity, safety, and potency as prescribed in this section. A serial found... completed product from each serial and each subserial shall be tested for viable bacteria and fungi...

  15. [Occurrance of antibodies against Leptospira in horses in Middle Germany].

    PubMed

    Pikalo, Jutta; Sattler, Tatjana; Eichinger, Michaela; Loitsch, Angelika; Sun, Hao; Schmoll, Friedrich; Schusser, Gerald Fritz

    2016-01-01

    Aim of the study was to detect antibodies and potential risk factors for an infec- tion with Leptospira in horses in Middle Germany. Serum samples of 314 horses were examined retrospectively by microscopic agglutination test for the presence of antibodies against eight Leptospira serovars. In total, 17.2% (n = 54) of the horses were positive for one or more of the serovars analyzed. The most prevalent serovar was lcterohaemorrhagiae (11.1%), followed by serovar Bratislava (9.6 %) and Grippotyphosa (1.9%). Mares showed a significantly higher occurrence of antibodies (p < 0.05) than geldings or stallions. Horses used for breeding have a significantly lower risk than horses used in sport or horses used for leisure activity. There was also a significantly higher prevalence (p < 0.05) in summer than in the other seasons. No significant influence of breed, husbandry conditions and age on the antibody occurrence was observed (p > 0.05). The clinical chemical parameters did not differ significantly between horses with positive or negative Leptospira antibody result (p > 0.05). It became apparent that horses can be infected with Leptospira without developing of clinical symptoms. PMID:27344912

  16. 9 CFR 113.105 - Leptospira Hardjo Bacterin.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Leptospira Hardjo Bacterin. 113.105 Section 113.105 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Inactivated Bacterial Products §...

  17. 9 CFR 113.101 - Leptospira Pomona Bacterin.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10-10,000 hamster...

  18. 9 CFR 113.103 - Leptospira Canicola Bacterin.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10-10,000...

  19. 9 CFR 113.103 - Leptospira Canicola Bacterin.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10-10,000...

  20. 9 CFR 113.101 - Leptospira Pomona Bacterin.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10-10,000 hamster...

  1. 9 CFR 113.101 - Leptospira Pomona Bacterin.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10-10,000 hamster...

  2. 9 CFR 113.101 - Leptospira Pomona Bacterin.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10-10,000 hamster...

  3. 9 CFR 113.103 - Leptospira Canicola Bacterin.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10-10,000...

  4. 9 CFR 113.103 - Leptospira Canicola Bacterin.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10-10,000...

  5. 9 CFR 113.103 - Leptospira Canicola Bacterin.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10-10,000...

  6. 9 CFR 113.101 - Leptospira Pomona Bacterin.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either... least 10 but not more than 12 additional hamsters from the same group as unvaccinated controls. (3... intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10-10,000 hamster...

  7. GLOBAL PATTERNS OF LEPTOSPIRA PREVALENCE IN VERTEBRATE RESERVOIR HOSTS.

    PubMed

    Andersen-Ranberg, Emilie U; Pipper, Christian; Jensen, Per M

    2016-07-01

    Leptospirosis is a widespread emerging bacterial zoonosis. As the transmission is believed to be predominantly waterborne, human incidence is expected to increase in conjunction with global climate change and associated extreme weather events. Providing more accurate predictions of human leptospirosis requires more detailed information on animal reservoirs that are the source of human infection. We evaluated the prevalence of Leptospira in vertebrates worldwide and its association with taxonomy, geographic region, host biology, ambient temperature, and precipitation patterns. A multivariate regression analysis with a meta-analysis-like approach was used to analyze compiled data extracted from 300 Leptospira-related peer reviewed papers. A fairly uniform Leptospira infection prevalence of about 15% was found in the majority of mammalian families. Higher prevalence was frequently associated with species occupying urban habitats, and this may explain why climatic factors were not significantly correlated with prevalence as consistently as expected. Across different approaches of the multiple regression analyses, the variables most frequently correlated with Leptospira infection prevalence were the host's ability to swim, minimum ambient temperature, and methodologic quality of the study. Prevalence in carnivores was not associated with any climatic variable, and the importance of environmental risk factors were indicated to be of lesser consequence in nonhuman mammals. The dataset is made available for further analysis. PMID:27187029

  8. 21 CFR 866.3350 - Leptospira spp. serological reagents.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Leptospira spp. serological reagents. 866.3350 Section 866.3350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3350...

  9. 21 CFR 866.3350 - Leptospira spp. serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Leptospira spp. serological reagents. 866.3350 Section 866.3350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3350...

  10. 21 CFR 866.3350 - Leptospira spp. serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Leptospira spp. serological reagents. 866.3350 Section 866.3350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3350...

  11. 21 CFR 866.3350 - Leptospira spp. serological reagents.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Leptospira spp. serological reagents. 866.3350 Section 866.3350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3350...

  12. 21 CFR 866.3350 - Leptospira spp. serological reagents.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Leptospira spp. serological reagents. 866.3350 Section 866.3350 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3350...

  13. [Occurrance of antibodies against Leptospira in horses in Middle Germany].

    PubMed

    Pikalo, Jutta; Sattler, Tatjana; Eichinger, Michaela; Loitsch, Angelika; Sun, Hao; Schmoll, Friedrich; Schusser, Gerald Fritz

    2016-01-01

    Aim of the study was to detect antibodies and potential risk factors for an infec- tion with Leptospira in horses in Middle Germany. Serum samples of 314 horses were examined retrospectively by microscopic agglutination test for the presence of antibodies against eight Leptospira serovars. In total, 17.2% (n = 54) of the horses were positive for one or more of the serovars analyzed. The most prevalent serovar was lcterohaemorrhagiae (11.1%), followed by serovar Bratislava (9.6 %) and Grippotyphosa (1.9%). Mares showed a significantly higher occurrence of antibodies (p < 0.05) than geldings or stallions. Horses used for breeding have a significantly lower risk than horses used in sport or horses used for leisure activity. There was also a significantly higher prevalence (p < 0.05) in summer than in the other seasons. No significant influence of breed, husbandry conditions and age on the antibody occurrence was observed (p > 0.05). The clinical chemical parameters did not differ significantly between horses with positive or negative Leptospira antibody result (p > 0.05). It became apparent that horses can be infected with Leptospira without developing of clinical symptoms.

  14. Leptospira Serovars for Diagnosis of Leptospirosis in Humans and Animals in Africa: Common Leptospira Isolates and Reservoir Hosts

    PubMed Central

    Mgode, Georgies F.; Machang’u, Robert S.; Mhamphi, Ginethon G.; Katakweba, Abdul; Mulungu, Loth S.; Durnez, Lies; Leirs, Herwig; Hartskeerl, Rudy A.; Belmain, Steven R.

    2015-01-01

    The burden of leptospirosis in humans and animals in Africa is higher than that reported from other parts of the world. However, the disease is not routinely diagnosed in the continent. One of major factors limiting diagnosis is the poor availability of live isolates of locally circulating Leptospira serovars for inclusion in the antigen panel of the gold standard microscopic agglutination test (MAT) for detecting antibodies against leptospirosis. To gain insight in Leptospira serovars and their natural hosts occurring in Tanzania, concomitantly enabling the improvement of the MAT by inclusion of fresh local isolates, a total of 52 Leptospira isolates were obtained from fresh urine and kidney homogenates, collected between 1996 and 2006 from small mammals, cattle and pigs. Isolates were identified by serogrouping, cross agglutination absorption test (CAAT), and molecular typing. Common Leptospira serovars with their respective animal hosts were: Sokoine (cattle and rodents); Kenya (rodents and shrews); Mwogolo (rodents); Lora (rodents); Qunjian (rodent); serogroup Grippotyphosa (cattle); and an unknown serogroup from pigs. Inclusion of local serovars particularly serovar Sokoine in MAT revealed a 10-fold increase in leptospirosis prevalence in Tanzania from 1.9% to 16.9% in rodents and 0.26% to 10.75% in humans. This indicates that local serovars are useful for diagnosis of human and animal leptospirosis in Tanzania and other African countries. PMID:26624890

  15. Distribution of Leptospira Serogroups in Cattle Herds and Dogs in France

    PubMed Central

    Ayral, Florence C.; Bicout, Dominique J.; Pereira, Helena; Artois, Marc; Kodjo, Angeli

    2014-01-01

    A retrospective study was conducted to identify and describe the distribution pattern of Leptospira serogroups in domestic animals in France. The population consisted of cattle herds and dogs with clinically suspected leptospirosis that were tested at the “Laboratoire des Leptospires” between 2008 and 2011. The laboratory database was queried for records of cattle and dogs in which seroreactivity in Leptospira microagglutination tests was consistent with a recent or current infection, excluding vaccine serogroups in dogs. A total of 394 cattle herds and 232 dogs were diagnosed with clinical leptospirosis, and the results suggested infection by the Leptospira serogroup Australis in 43% and 63%, respectively; by the Leptospira serogroup Grippotyphosa in 17% and 9%, respectively; and by the Leptospira serogroup Sejroe in 33% and 6%, respectively. This inventory of infecting Leptospira serogroups revealed that current vaccines in France are not fully capable of preventing the clinical form of the disease. PMID:25092816

  16. Distribution of Leptospira serogroups in cattle herds and dogs in France.

    PubMed

    Ayral, Florence C; Bicout, Dominique J; Pereira, Helena; Artois, Marc; Kodjo, Angeli

    2014-10-01

    A retrospective study was conducted to identify and describe the distribution pattern of Leptospira serogroups in domestic animals in France. The population consisted of cattle herds and dogs with clinically suspected leptospirosis that were tested at the "Laboratoire des Leptospires" between 2008 and 2011. The laboratory database was queried for records of cattle and dogs in which seroreactivity in Leptospira microagglutination tests was consistent with a recent or current infection, excluding vaccine serogroups in dogs. A total of 394 cattle herds and 232 dogs were diagnosed with clinical leptospirosis, and the results suggested infection by the Leptospira serogroup Australis in 43% and 63%, respectively; by the Leptospira serogroup Grippotyphosa in 17% and 9%, respectively; and by the Leptospira serogroup Sejroe in 33% and 6%, respectively. This inventory of infecting Leptospira serogroups revealed that current vaccines in France are not fully capable of preventing the clinical form of the disease. PMID:25092816

  17. USDA regulatory guidelines and practices for veterinary Leptospira vaccine potency testing.

    PubMed

    Srinivas, G B; Walker, A; Rippke, B

    2013-09-01

    Batch-release potency testing of leptospiral vaccines licensed by the United States Department of Agriculture (USDA) historically was conducted through animal vaccination-challenge models. The hamster vaccination-challenge assay was Codified in 1974 for bacterins containing Leptospira pomona, Leptospira icterohaemorrhagiae, and Leptospira canicola, and in 1975 for bacterins containing Leptospira grippotyphosa. In brief, 10 hamsters are vaccinated with a specified dilution of bacterin. After a holding period, the vaccinated hamsters, as well as nonvaccinated controls, are challenged with virulent Leptospira and observed for mortality. Eighty percent of vaccinated hamsters must survive in the face of a valid challenge. The high cost of the Codified tests, in terms of monetary expense and animal welfare, prompted the Center for Veterinary Biologics (CVB) to develop ELISA alternatives for them. Potency tests for other serogroups, such as Leptospira hardjo-bovis, that do not have Codified requirements for potency testing continue to be examined on a case-by-case basis. PMID:23838570

  18. Leptospira Exposure and Waste Pickers: A Case-Control Seroprevalence Study in Durango, Mexico

    PubMed Central

    Alvarado-Esquivel, Cosme; Hernandez-Tinoco, Jesus; Sanchez-Anguiano, Luis Francisco; Ramos-Nevarez, Agar; Cerrillo-Soto, Sandra Margarita; Guido-Arreola, Carlos Alberto

    2015-01-01

    Background Infection with Leptospira may occur by contact with Leptospira-infected animals. Waste pickers are in contact with rodents and dogs while picking in the garbage. Whether waste pickers are at risk for Leptospira infection is largely unknown. This study was aimed to determine the association of Leptospira IgG seroprevalence with the occupation of waste picking, and to determine the epidemiological characteristics of the waste pickers with Leptospira exposure. Methods Through a case-control study, we determined the seroprevalence of anti-Leptospira IgG antibodies in 90 waste pickers and 90 age- and gender-matched control subjects in Durango City, Mexico using an enzyme immunoassay. Data were analyzed by bivariate and multivariate analyses. Results The prevalence of anti-Leptospira IgG antibodies was similar in waste pickers (4/90: 4.4%) to that in control subjects (5/90: 5.6%) (P = 1.00). Bivariate analysis showed that Leptospira exposure in waste pickers was associated with increasing age (P = 0.009), no education (P = 0.008), and consumption of rat meat (P = 0.04). However, these associations were no longer found by multivariate analysis. Leptospira exposure in waste pickers was not associated with health status, duration in the activity, wearing hand gloves and facemasks, history of injuries with sharp material of the garbage, or contact with animals or soil. Conclusions This is the first study about Leptospira exposure in waste pickers. Results suggest that waste pickers are not at increasing risk for Leptospira exposure in Durango City, Mexico. Further research with a larger sample size to elucidate the association of Leptospira exposure with waste picking activity is needed. PMID:26124911

  19. High Seroprevalence of Leptospira Exposure in Meat Workers in Northern Mexico: A Case-Control Study

    PubMed Central

    Alvarado-Esquivel, Cosme; Hernandez-Tinoco, Jesus; Sanchez-Anguiano, Luis Francisco; Ramos-Nevarez, Agar; Cerrillo-Soto, Sandra Margarita; Saenz-Soto, Leandro; Martinez-Ramirez, Lucio

    2016-01-01

    Background The seroepidemiology of Leptospira infection in workers occupationally exposed to raw meat has been poorly studied. This work aimed to determine the association between Leptospira exposure and the occupation of meat worker, and to determine the seroprevalence association with socio-demographic, work, clinical and behavioral characteristics of the meat workers studied. Methods We performed a case-control study in 124 meat workers and 124 age- and gender-matched control subjects in Durango City, Mexico. Sera of cases and controls were analyzed for anti-Leptospira IgG antibodies using a commercially available enzyme immunoassay. Data of meat workers were obtained with the aid of a questionnaire. The association of Leptospira exposure with the characteristics of meat workers was analyzed by bivariate and multivariate analyses. Results Anti-Leptospira IgG antibodies were found in 22 (17.7%) of 124 meat workers and in eight (6.5%) of 124 controls (OR = 3.12; 95% CI: 1.33 - 7.33; P = 0.006). Seroprevalence of Leptospira infection was similar between male butchers (17.6%) and female butchers (18.2%) (P = 1.00). Multivariate analysis of socio-demographic, work and behavioral variables showed that Leptospira exposure was associated with duration in the activity, rural residence, and consumption of snake meat and unwashed raw fruits. Conclusions This is the first case-control study of the association of Leptospira exposure with the occupation of meat worker. Results indicate that meat workers represent a risk group for Leptospira exposure. Risk factors for Leptospira exposure found in this study may help in the design of optimal preventive measures against Leptospira infection. PMID:26858797

  20. Modulation of Hemostatic and Inflammatory Responses by Leptospira Spp.

    PubMed Central

    Vieira, Mônica L.; Naudin, Clément; Mörgelin, Matthias; Romero, Eliete C.; Nascimento, Ana Lucia T. O.; Herwald, Heiko

    2016-01-01

    Leptospirosis is a worldwide spread zoonotic and neglected infectious disease of human and veterinary concern that is caused by pathogenic Leptospira species. In severe infections, hemostatic impairments such as coagulation/fibrinolysis dysfunction are frequently observed. These complications often occur when the host response is controlled and/or modulated by the bacterial pathogen. In the present investigation, we aimed to analyze the modulation of the hemostatic and inflammatory host responses by the bacterial pathogen Leptospira. The effects of leptospires and their secreted products on stimulation of human intrinsic and extrinsic pathways of coagulation were investigated by means of altered clotting times, assembly and activation of contact system and induction of tissue factor. We show that both extrinsic and intrinsic coagulation cascades are modulated in response to Leptospira or leptospiral secreted proteins. We further find that the pro-inflammatory mediator bradykinin is released following contact activation at the bacterial surface and that pro-coagulant microvesicles are shed from monocytes in response to infection. Also, we show that human leptospirosis patients present higher levels of circulating pro-coagulant microvesicles than healthy individuals. Here we show that both pathways of the coagulation system are modulated by leptospires, possibly leading to altered hemostatic and inflammatory responses during the disease. Our results contribute to the understanding of the leptospirosis pathophysiological mechanisms and may open new routes for the discovery of novel treatments for the severe manifestations of the disease. PMID:27167223

  1. Modulation of Hemostatic and Inflammatory Responses by Leptospira Spp.

    PubMed

    Vieira, Mônica L; Naudin, Clément; Mörgelin, Matthias; Romero, Eliete C; Nascimento, Ana Lucia T O; Herwald, Heiko

    2016-05-01

    Leptospirosis is a worldwide spread zoonotic and neglected infectious disease of human and veterinary concern that is caused by pathogenic Leptospira species. In severe infections, hemostatic impairments such as coagulation/fibrinolysis dysfunction are frequently observed. These complications often occur when the host response is controlled and/or modulated by the bacterial pathogen. In the present investigation, we aimed to analyze the modulation of the hemostatic and inflammatory host responses by the bacterial pathogen Leptospira. The effects of leptospires and their secreted products on stimulation of human intrinsic and extrinsic pathways of coagulation were investigated by means of altered clotting times, assembly and activation of contact system and induction of tissue factor. We show that both extrinsic and intrinsic coagulation cascades are modulated in response to Leptospira or leptospiral secreted proteins. We further find that the pro-inflammatory mediator bradykinin is released following contact activation at the bacterial surface and that pro-coagulant microvesicles are shed from monocytes in response to infection. Also, we show that human leptospirosis patients present higher levels of circulating pro-coagulant microvesicles than healthy individuals. Here we show that both pathways of the coagulation system are modulated by leptospires, possibly leading to altered hemostatic and inflammatory responses during the disease. Our results contribute to the understanding of the leptospirosis pathophysiological mechanisms and may open new routes for the discovery of novel treatments for the severe manifestations of the disease.

  2. Modulation of Hemostatic and Inflammatory Responses by Leptospira Spp.

    PubMed

    Vieira, Mônica L; Naudin, Clément; Mörgelin, Matthias; Romero, Eliete C; Nascimento, Ana Lucia T O; Herwald, Heiko

    2016-05-01

    Leptospirosis is a worldwide spread zoonotic and neglected infectious disease of human and veterinary concern that is caused by pathogenic Leptospira species. In severe infections, hemostatic impairments such as coagulation/fibrinolysis dysfunction are frequently observed. These complications often occur when the host response is controlled and/or modulated by the bacterial pathogen. In the present investigation, we aimed to analyze the modulation of the hemostatic and inflammatory host responses by the bacterial pathogen Leptospira. The effects of leptospires and their secreted products on stimulation of human intrinsic and extrinsic pathways of coagulation were investigated by means of altered clotting times, assembly and activation of contact system and induction of tissue factor. We show that both extrinsic and intrinsic coagulation cascades are modulated in response to Leptospira or leptospiral secreted proteins. We further find that the pro-inflammatory mediator bradykinin is released following contact activation at the bacterial surface and that pro-coagulant microvesicles are shed from monocytes in response to infection. Also, we show that human leptospirosis patients present higher levels of circulating pro-coagulant microvesicles than healthy individuals. Here we show that both pathways of the coagulation system are modulated by leptospires, possibly leading to altered hemostatic and inflammatory responses during the disease. Our results contribute to the understanding of the leptospirosis pathophysiological mechanisms and may open new routes for the discovery of novel treatments for the severe manifestations of the disease. PMID:27167223

  3. Leptospira Exposure and Patients with Liver Diseases: A Case-Control Seroprevalence Study

    PubMed Central

    Alvarado-Esquivel, Cosme; Sánchez-Anguiano, Luis Francisco; Hernández-Tinoco, Jesús; Ramos-Nevárez, Agar; Margarita Cerrillo-Soto, Sandra; Alberto Guido-Arreola, Carlos

    2016-01-01

    The seroepidemiology of Leptospira infection in patients suffering from liver disease has been poorly studied. Information about risk factors associated with infection in liver disease patients may help in the optimal planning of preventive measures. We sought to determine the association of Leptospira IgG seroprevalence and patients with liver diseases, and to determine the characteristics of the patients with Leptospira exposure. We performed a case-control study of 75 patients suffering from liver diseases and 150 age- and gender-matched control subjects. Diagnoses of liver disease included liver cirrhosis, steatosis, chronic hepatitis, acute hepatitis, and amoebic liver abscess. Sera of participants were analyzed for the presence of anti- Leptospira IgG antibodies using a commercially available enzyme immunoassay. Anti-Leptospira IgG antibodies were found in 17 (22.7%) of 75 patients and in 15 (10.0%) of 150 control subjects (OR = 2.32; 95% CI: 1.09-4.94; P=0.03). This is the first age- and gender-matched case control study about Leptospira seroprevalence in patients with liver diseases. Results indicate that Leptospira infection is associated with chronic and acute liver diseases. Results warrants for additional studies on the role of Leptospira exposure in chronic liver disease. PMID:27493589

  4. Leptospira infections in freshwater fish in Morogoro Tanzania: a hidden public health threat.

    PubMed

    Mgode, Georgis F; Mhamphi, Ginethon G; Katkweba, Abdul S; Thomas, Michael

    2014-04-01

    Leptospirosis caused by spirochete bacterium of genus Leptospira affects humans and animals worldwide. Rodents are major reservoirs of leptospires whereas wetland and aquatic migratory birds also carry and transmit leptospires. Leptospirosis studies in fish are lacking in African countries despite favourable environment and abundant reservoirs, which can spread leptospires into aquatic habitats and infect fish. The objectives of this study were to determine presence of Leptospira in fish; the prevalent Leptospira serovars and whether are related to serovars reported in animals; and to ascertain potential public health risk. Live tilapia, catfish and eel fish (n = 48) were caught in Mindu Dam in Morogoro Municipality in eastern Tanzania. Blood sample was collected using syringes and needles to obtain serum for serological detection of leptospirosis using gold standard microagglutination test utilizing local and reference Leptospira serovars Sokoine, Kenya, Pomona and Hebdomadis. Twenty-six fish (54.2%) were positive for serovar Kenya (29.2%) and Sokoine (25%). Leptospira prevalence was high in both catfish (58.3%) and tilapia fish (47.8%). Thus, different fish types are infected with Leptospira found in animals. Fish could be source of Leptospira infection to humans since tilapia and catfish are the common fish type widely consumed in Tanzania. Further study covering lakes, rivers and dams is required to better understand the prevalence of Leptospira in fish and actual public health threats. PMID:26875305

  5. Development of Chronic and Acute Golden Syrian Hamster Infection Models with Leptospira borgpetersenii serovar Hardjo

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The golden Syrian hamster (Mesocricetus auratus) is frequently used as a model to study virulence for several species of Leptospira. Onset of an acute, lethal infection following infection with several pathogenic Leptospira species has been widely adopted for vaccine testing. An important exceptio...

  6. Leptospira Exposure and Patients with Liver Diseases: A Case-Control Seroprevalence Study.

    PubMed

    Alvarado-Esquivel, Cosme; Sánchez-Anguiano, Luis Francisco; Hernández-Tinoco, Jesús; Ramos-Nevárez, Agar; Margarita Cerrillo-Soto, Sandra; Alberto Guido-Arreola, Carlos

    2016-06-01

    The seroepidemiology of Leptospira infection in patients suffering from liver disease has been poorly studied. Information about risk factors associated with infection in liver disease patients may help in the optimal planning of preventive measures. We sought to determine the association of Leptospira IgG seroprevalence and patients with liver diseases, and to determine the characteristics of the patients with Leptospira exposure. We performed a case-control study of 75 patients suffering from liver diseases and 150 age- and gender-matched control subjects. Diagnoses of liver disease included liver cirrhosis, steatosis, chronic hepatitis, acute hepatitis, and amoebic liver abscess. Sera of participants were analyzed for the presence of anti- Leptospira IgG antibodies using a commercially available enzyme immunoassay. Anti-Leptospira IgG antibodies were found in 17 (22.7%) of 75 patients and in 15 (10.0%) of 150 control subjects (OR = 2.32; 95% CI: 1.09-4.94; P=0.03). This is the first age- and gender-matched case control study about Leptospira seroprevalence in patients with liver diseases. Results indicate that Leptospira infection is associated with chronic and acute liver diseases. Results warrants for additional studies on the role of Leptospira exposure in chronic liver disease. PMID:27493589

  7. Isolation of Leptospira hardjo from kidneys of Ontario cattle at slaughter.

    PubMed Central

    Prescott, J F; Miller, R B; Nicholson, V M

    1987-01-01

    Kidneys from 117 cattle from 110 Ontario farms were examined at slaughter for leptospires. Leptospira hardjo (hardjo-bovis A) was isolated from 11 kidneys and L. kennewicki from one. The isolations were all made (12/89, 13.5%) from beef cattle from feedlots, no isolates being obtained from dairy or beef cattle from extensive farms (0/28). Isolations were only made from cattle with antibody titers (greater than or equal to 20) against the serovars recovered. Isolation was more sensitive than immunofluorescence in identifying leptospira, particularly in animals with low antibody titers against L. hardjo. Leptospira were isolated from two kidneys with multiple gross lesions of focal nephritis, but there was no correlation between the presence of scanty kidney lesions and isolations of leptospira. Leptospira hardjo infection appears to be common in Ontario feedlot cattle. PMID:3300922

  8. Molecular Characterization of Leptospira spp. in Environmental Samples from North-Eastern Malaysia Revealed a Pathogenic Strain, Leptospira alstonii

    PubMed Central

    Azali, Muhammad Azharuddin; Yean Yean, Chan; Aminuddin Baki, Nurul Najian

    2016-01-01

    The presence of pathogenic Leptospira spp. in the environment poses threats to human health. The aim of this study was to detect and characterize Leptospira spp. from environmental samples. A total of 144 samples comprised of 72 soil and 72 water samples were collected from markets and recreational areas in a north-eastern state in Malaysia. Samples were cultured on Ellinghausen and McCullough modified by Johnson and Harris media. Leptospires were positive in 22.9% (n = 33) of the isolates. Based on partial sequences of 16S rRNA, a pathogenic leptospire, Leptospira alstonii (n = 1/33), was identified in 3% of the isolates followed by intermediate leptospire (L. wolffii, n = 1/33, and L. licerasiae, n = 7/33) and nonpathogenic leptospire, L. meyeri (n = 22/33) in 24.2% and 66.7%, respectively. This study demonstrates the presence of a clinically significant pathogenic L. alstonii in the environments which could pose health risks to the occupants and visitors. PMID:27127522

  9. Molecular Characterization of Leptospira spp. in Environmental Samples from North-Eastern Malaysia Revealed a Pathogenic Strain, Leptospira alstonii.

    PubMed

    Azali, Muhammad Azharuddin; Yean Yean, Chan; Harun, Azian; Aminuddin Baki, Nurul Najian; Ismail, Nabilah

    2016-01-01

    The presence of pathogenic Leptospira spp. in the environment poses threats to human health. The aim of this study was to detect and characterize Leptospira spp. from environmental samples. A total of 144 samples comprised of 72 soil and 72 water samples were collected from markets and recreational areas in a north-eastern state in Malaysia. Samples were cultured on Ellinghausen and McCullough modified by Johnson and Harris media. Leptospires were positive in 22.9% (n = 33) of the isolates. Based on partial sequences of 16S rRNA, a pathogenic leptospire, Leptospira alstonii (n = 1/33), was identified in 3% of the isolates followed by intermediate leptospire (L. wolffii, n = 1/33, and L. licerasiae, n = 7/33) and nonpathogenic leptospire, L. meyeri (n = 22/33) in 24.2% and 66.7%, respectively. This study demonstrates the presence of a clinically significant pathogenic L. alstonii in the environments which could pose health risks to the occupants and visitors. PMID:27127522

  10. Opportunities and strategies to further reduce animal use for Leptospira vaccine potency testing.

    PubMed

    Walker, A; Srinivas, G B

    2013-09-01

    Hamsters are routinely infected with virulent Leptospira for two purposes in the regulation of biologics: the performance of Codified potency tests and maintenance of challenge culture for the Codified potency tests. Options for reducing animal use in these processes were explored in a plenary lecture at the "International Workshop on Alternative Methods for Leptospira Vaccine Potency Testing: State of the Science and the Way Forward" held at the Center for Veterinary Biologics in September 2012. The use of validated in vitro potency assays such as those developed by the U.S. Department of Agriculture for Leptospira (L.) canicola, Leptospira grippotyphosa, Leptospira pomona, and Leptospira icterohaemorrhagiae rather than the Codified hamster vaccination-challenge assay was encouraged. Alternatives such as reduced animal numbers in the hamster vaccination-challenge testing were considered for problematic situations. Specifically, the merits of sharing challenge controls, reducing group sizes, and eliminating animals for concurrent challenge dose titration were assessed. Options for maintaining virulent, stable cultures without serial passage through hamsters or with decreased hamster use were also discussed. The maintenance of virulent Leptospira without the use of live animals is especially difficult since a reliable means to maintain virulence after multiple in vitro passages has not yet been identified.

  11. Multiple co-infections of rodents with hantaviruses, Leptospira, and Babesia in Croatia.

    PubMed

    Tadin, Ante; Turk, Nenad; Korva, Miša; Margaletić, Josip; Beck, Relja; Vucelja, Marko; Habuš, Josipa; Svoboda, Petra; Zupanc, Tatjana Avšič; Henttonen, Heikki; Markotić, Alemka

    2012-05-01

    Hantaviruses, Leptospira spp., and Babesia spp. are rodent-borne pathogens present worldwide. We studied multiple co-infections of small rodents in Croatia with all three pathogens. Twenty-eight Apodemus flavicollis and 16 Myodes glareolus were tested for the presence of hantavirus RNA by real-time RT-PCR, Leptospira strains by renoculture method and Babesia DNA by PCR. Anti-hantavirus antibodies and anti-Leptospira antibodies were detected by serological methods. Very high infection rates with each pathogen were found in A. flavicollis: 20 of 28 rodents (71%) were infected with Dobrava virus, 13 rodents (46%) were infected with Leptospira, and 5 rodents (18%) were infected with Babesia. Multiple co-infections with all three pathogens were found in 3 of 28 (11%) A. flavicollis animals, suggesting that the same rodent host can be infected with several pathogens at the same time. Dual infections with both hantaviruses and Leptospira were found in 7 of 44 rodents (16%), with hantaviruses and Babesia in 2 rodents (5%), and double infection with both Leptospira and Babesia were found in 1 rodent (2%). Since hantaviruses, Leptospira, and Babesia have similar geographical distributions, it is to be expected that in other parts of the world multiple co-infections, representing a serious threat to public health, can be found.

  12. Leptospira borgpetersenii from free-living white-eared opossum (Didelphis albiventris): first isolation in Brazil.

    PubMed

    Jorge, Sérgio; Hartleben, Cláudia P; Seixas, Fabiana K; Coimbra, Marco A A; Stark, Cledir B; Larrondo, Adriana G; Amaral, Marta G; Albano, Ana Paula N; Minello, Luiz F; Dellagostin, Odir A; Brod, Claudiomar S

    2012-11-01

    Leptospirosis is a zoonotic disease that occurs all over the world, caused by bacteria of the genus Leptospira. Marsupial and didelphidae families are considered susceptible to infection caused by a wide range of Leptospira serovars for which they serve as reservoirs. Thirty-three free-living white-eared opossums (Didelphis albiventris) were captured in Southern Brazil and bodily fluids were collected. From the urine samples it was possible to obtain an isolate identified as Leptospira borgpetersenii by rpoB gene sequencing and belonging to serovar Castellonis by Multilocus Variable-Number Tandem-Repeat Analysis. This is the first report of the isolation of Leptospira spp. from the white-eared opossum in Brazil. In addition, the new strain was also virulent in the hamster model of lethal leptospirosis. The microscopic agglutination test (MAT) was used for detecting the presence of antibodies against Leptospira spp. in white-eared opossum, human, cattle and canine sera using a panel of 59 Leptospira strains that included the new isolate. The inclusion of the new strain in the MAT battery increased the MAT sensitivity for canine sera. These findings suggest that the white-eared opossum is an important reservoir of pathogenic Leptospira spp.

  13. Opportunities and strategies to further reduce animal use for Leptospira vaccine potency testing.

    PubMed

    Walker, A; Srinivas, G B

    2013-09-01

    Hamsters are routinely infected with virulent Leptospira for two purposes in the regulation of biologics: the performance of Codified potency tests and maintenance of challenge culture for the Codified potency tests. Options for reducing animal use in these processes were explored in a plenary lecture at the "International Workshop on Alternative Methods for Leptospira Vaccine Potency Testing: State of the Science and the Way Forward" held at the Center for Veterinary Biologics in September 2012. The use of validated in vitro potency assays such as those developed by the U.S. Department of Agriculture for Leptospira (L.) canicola, Leptospira grippotyphosa, Leptospira pomona, and Leptospira icterohaemorrhagiae rather than the Codified hamster vaccination-challenge assay was encouraged. Alternatives such as reduced animal numbers in the hamster vaccination-challenge testing were considered for problematic situations. Specifically, the merits of sharing challenge controls, reducing group sizes, and eliminating animals for concurrent challenge dose titration were assessed. Options for maintaining virulent, stable cultures without serial passage through hamsters or with decreased hamster use were also discussed. The maintenance of virulent Leptospira without the use of live animals is especially difficult since a reliable means to maintain virulence after multiple in vitro passages has not yet been identified. PMID:23891496

  14. Seroprevalence of Brucella abortus and Leptospira hardjo in cattle

    PubMed Central

    Pandian, S. Jegaveera; Ray, Pradeep Kumar; Chandran, P. C.; Kumar, Manoj

    2015-01-01

    Aim: The aim was to assess the seroprevalence of B. abortus and Leptospira hardjo in the cattle population of Bihar, this work was carried out. Materials and Methods: Randomly selected 450 cattle from nine districts of Bihar were serologically screened for antibodies against L. hardjo and B. abortus. DAS-ELISA for leptospira and AB-ELISA for brucella were carried out. Based on the results prevalence in each district and the state are reported herewith. Results and Discussion: In this study, it was found that the seroprevalence of L. hardjo was 9.11% and that of B. abortus was 12.2% in Bihar. Indigenous cattle were found to be less susceptible to leptospirosis and brucellosis even though they accounted for 83.11% of the study population. Conclusion: Although there was no acute disease, antibodies detected against L. hardjo and B. abortus in the cattle population indicated the presence of chronic and subclinical infection, which could challenge the fertility of the animals. PMID:27047076

  15. Draft Genome Sequence of Brazilian Leptospira noguchii Serogroup Panama Strain U73, Isolated from Cattle.

    PubMed

    Moreno, Luisa Z; Loureiro, Ana P; Miraglia, Fabiana; Matajira, Carlos E C; Kremer, Frederico S; Eslabao, Marcos R; Dellagostin, Odir A; Lilenbaum, Walter; Moreno, Andrea M

    2015-10-15

    Leptospira noguchii is a current zoonotic pathogen in Brazil. Here, we report the draft genome sequence of the Brazilian L. noguchii serogroup Panama strain U73, isolated from asymptomatic cattle urine.

  16. Draft Genome Sequence of Brazilian Leptospira noguchii Serogroup Panama Strain U73, Isolated from Cattle

    PubMed Central

    Moreno, Luisa Z.; Loureiro, Ana P.; Miraglia, Fabiana; Matajira, Carlos E. C.; Kremer, Frederico S.; Eslabao, Marcos R.; Dellagostin, Odir A.; Lilenbaum, Walter

    2015-01-01

    Leptospira noguchii is a current zoonotic pathogen in Brazil. Here, we report the draft genome sequence of the Brazilian L. noguchii serogroup Panama strain U73, isolated from asymptomatic cattle urine. PMID:26472831

  17. Malagasy bats shelter a considerable genetic diversity of pathogenic Leptospira suggesting notable host-specificity patterns.

    PubMed

    Gomard, Yann; Dietrich, Muriel; Wieseke, Nicolas; Ramasindrazana, Beza; Lagadec, Erwan; Goodman, Steven M; Dellagi, Koussay; Tortosa, Pablo

    2016-04-01

    Pathogenic Leptospira are the causative agents of leptospirosis, a disease of global concern with major impact in tropical regions. Despite the importance of this zoonosis for human health, the evolutionary and ecological drivers shaping bacterial communities in host reservoirs remain poorly investigated. Here, we describe Leptospira communities hosted by Malagasy bats, composed of mostly endemic species, in order to characterize host-pathogen associations and investigate their evolutionary histories. We screened 947 individual bats (representing 31 species, 18 genera and seven families) for Leptospira infection and subsequently genotyped positive samples using three different bacterial loci. Molecular identification showed that these Leptospira are notably diverse and include several distinct lineages mostly belonging to Leptospira borgpetersenii and L. kirschneri. The exploration of the most probable host-pathogen evolutionary scenarios suggests that bacterial genetic diversity results from a combination of events related to the ecology and the evolutionary history of their hosts. Importantly, based on the data set presented herein, the notable host-specificity we have uncovered, together with a lack of geographical structuration of bacterial genetic diversity, indicates that the Leptospira community at a given site depends on the co-occurring bat species assemblage. The implications of such tight host-specificity on the epidemiology of leptospirosis are discussed. PMID:26902801

  18. Amino acid sequences of proteins from Leptospira serovar pomona.

    PubMed

    Alves, S F; Lefebvre, R B; Probert, W

    2000-01-01

    This report describes a partial amino acid sequences from three putative outer envelope proteins from Leptospira serovar pomona. In order to obtain internal fragments for protein sequencing, enzymatic and chemical digestion was performed. The enzyme clostripain was used to digest the proteins 32 and 45 kDa. In situ digestion of 40 kDa molecular weight protein was accomplished using cyanogen bromide. The 32 kDa protein generated two fragments, one of 21 kDa and another of 10 kDa that yielded five residues. A fragment of 24 kDa that yielded nineteen residues of amino acids was obtained from 45 kDa protein. A fragment with a molecular weight of 20 kDa, yielding a twenty amino acids sequence from the 40 kDa protein.

  19. Pathogenic and saprophytic Leptospira species in water and soils from selected urban sites in peninsular Malaysia.

    PubMed

    Benacer, Douadi; Woh, Pei Yee; Mohd Zain, Siti Nursheena; Amran, Fairuz; Thong, Kwai Lin

    2013-01-01

    Leptospira species were studied in water and soils from selected urban sites in Malaysia. A total of 151 water (n=121) and soil (n=30) samples were collected from 12 recreational lakes and wet markets. All samples were filtered and inoculated into semi-solid Ellinghausen and McCullough modified by Johnson and Harris (EMJH) media supplemented with additional 5-fluorouracil. The cultures were then incubated at 30°C and observed under a dark field microscope with intervals of 10 days. A PCR assay targeting the rrs gene was used to confirm the genus Leptospira among the isolates. Subsequently, the pathogenic status of the isolates was determined using primer sets G1/G2 and Sapro1/Sapro2, which target the secY and rrs genes, respectively. The isolates were identified at serogroup level using the microscopic agglutination test (MAT) while their genetic diversity was assessed by pulsed field gel electrophoresis (PFGE). Based on dark field microscopy, 23.1% (28/121) water and 23.3% (7/30) soil cultures were positive for Leptospira spp. Of the 35 positive cultures, only 8 were pure and confirmed as Leptospira genus by PCR assay. Two out of 8 isolates were confirmed as pathogenic, 5 were saprophytic and one was intermediate. These 8 isolates were negative for the 25 reference hyperimmune rabbit sera tested in the MAT. PFGE showed that all 8 of these environmental Leptospira spp. were genetically diverse. In conclusion, the presence of pathogenic Leptospira spp. in the urban Malaysian environment may indicate and highlight the importance of water screening, especially in recreational lakes, in order to minimize any chance of Leptospira infection. PMID:23363618

  20. Impact of Environment and Social Gradient on Leptospira Infection in Urban Slums

    PubMed Central

    Felzemburgh, Ridalva D. M.; Santana, Francisco S.; Mohr, Sharif; Melendez, Astrid X. T. O.; Queiroz, Adriano; Santos, Andréia C.; Ravines, Romy R.; Tassinari, Wagner S.; Carvalho, Marília S.; Reis, Mitermayer G.; Ko, Albert I.

    2008-01-01

    Background Leptospirosis has become an urban health problem as slum settlements have expanded worldwide. Efforts to identify interventions for urban leptospirosis have been hampered by the lack of population-based information on Leptospira transmission determinants. The aim of the study was to estimate the prevalence of Leptospira infection and identify risk factors for infection in the urban slum setting. Methods and Findings We performed a community-based survey of 3,171 slum residents from Salvador, Brazil. Leptospira agglutinating antibodies were measured as a marker for prior infection. Poisson regression models evaluated the association between the presence of Leptospira antibodies and environmental attributes obtained from Geographical Information System surveys and indicators of socioeconomic status and exposures for individuals. Overall prevalence of Leptospira antibodies was 15.4% (95% confidence interval [CI], 14.0–16.8). Households of subjects with Leptospira antibodies clustered in squatter areas at the bottom of valleys. The risk of acquiring Leptospira antibodies was associated with household environmental factors such as residence in flood-risk regions with open sewers (prevalence ratio [PR] 1.42, 95% CI 1.14–1.75) and proximity to accumulated refuse (1.43, 1.04–1.88), sighting rats (1.32, 1.10–1.58), and the presence of chickens (1.26, 1.05–1.51). Furthermore, low income and black race (1.25, 1.03–1.50) were independent risk factors. An increase of US$1 per day in per capita household income was associated with an 11% (95% CI 5%–18%) decrease in infection risk. Conclusions Deficiencies in the sanitation infrastructure where slum inhabitants reside were found to be environmental sources of Leptospira transmission. Even after controlling for environmental factors, differences in socioeconomic status contributed to the risk of Leptospira infection, indicating that effective prevention of leptospirosis may need to address the social

  1. Occurrence of Leptospira DNA in water and soil samples collected in eastern Poland.

    PubMed

    Wójcik-Fatla, Angelina; Zając, Violetta; Wasiński, Bernard; Sroka, Jacek; Cisak, Ewa; Sawczyn, Anna; Dutkiewicz, Jacek

    2014-01-01

    Leptospira is an important re-emerging zoonotic human pathogen, disseminated by sick and carrier animals, water and soil. Weather calamities, such as flooding or cyclones favour the spreading of these bacteria. To check a potential role of natural water and soil in the persistence and spread of Leptospira on the territory of eastern Poland, 40 samples of natural water and 40 samples of soil were collected from areas exposed to flooding, and 64 samples of natural water and 68 samples of soil were collected from areas not exposed to flooding. Samples of water were taken from various reservoirs (rivers, natural lakes, artificial lakes, canals, ponds, farm wells) and samples of soils were taken at the distance of 1-3 meters from the edge of the reservoirs. The samples were examined for the presence of Leptospira DNA by nested-PCR. Two out of 40 samples of water (5.0%) collected from the area exposed to flooding showed the presence of Leptospira DNA, while all 40 samples of soil from this area were negative. All samples of water and soil (64 and 68, respectively) collected from the areas not exposed to flooding were negative. No significant difference were found between the results obtained in the areas exposed and not exposed to flooding. In conclusion, these results suggest that water and soil have only limited significance in the persistence and dissemination of Leptospira in eastern Poland. PMID:25528911

  2. Characterization of Leptospira santarosai Serogroup Grippotyphosa Serovar Bananal Isolated from Capybara ( Hydrochaeris hydrochaeris ) in Brazil.

    PubMed

    Moreno, Luisa Z; Miraglia, Fabiana; Marvulo, Maria F V; Silva, Jean C R; Paula, Catia D; Costa, Barbara L P; Morais, Zenaide M; Ferreira, Fernando; Neto, José S Ferreira; Dellagostin, Odir A; Hartskeerl, Rudy A; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-07-01

    Leptospirosis is a widespread zoonosis caused by bacteria of the genus Leptospira. Rodents appear to be the most important reservoirs of infection. They contaminate the environment and food and can transmit the pathogen when they are consumed by carnivores. Capybara ( Hydrochaeris hydrochaeris ) are efficient reservoirs of Leptospira, and because they are in close contact with farm animals and are found in semiurban areas, they represent a risk to public health. We isolated five Leptospira strains from capybara kidneys in Sao Paulo State, Brazil, in 2001 and typed them using serologic and molecular techniques. These strains include the Leptospira santarosai serogroup Grippotyphosa serovar Bananal. Pulsed field gel electrophoresis resulted in a unique pattern distinct from the reference strains, and the isolates clustered with greater than 85% similarity. The isolates also presented higher growth rates than other Leptospira serovars, with high minimal inhibitory concentration values for most of the tested antibiotics, with the exception of penicillin and ampicillin. This isolation and characterization of the L. santarosai serogroup Grippotyphosa serovar Bananal from capybara, highlights the importance of wild and sinantropic rodents as carriers of pathogenic leptospires.

  3. Characterization of Leptospira santarosai Serogroup Grippotyphosa Serovar Bananal Isolated from Capybara ( Hydrochaeris hydrochaeris ) in Brazil.

    PubMed

    Moreno, Luisa Z; Miraglia, Fabiana; Marvulo, Maria F V; Silva, Jean C R; Paula, Catia D; Costa, Barbara L P; Morais, Zenaide M; Ferreira, Fernando; Neto, José S Ferreira; Dellagostin, Odir A; Hartskeerl, Rudy A; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-07-01

    Leptospirosis is a widespread zoonosis caused by bacteria of the genus Leptospira. Rodents appear to be the most important reservoirs of infection. They contaminate the environment and food and can transmit the pathogen when they are consumed by carnivores. Capybara ( Hydrochaeris hydrochaeris ) are efficient reservoirs of Leptospira, and because they are in close contact with farm animals and are found in semiurban areas, they represent a risk to public health. We isolated five Leptospira strains from capybara kidneys in Sao Paulo State, Brazil, in 2001 and typed them using serologic and molecular techniques. These strains include the Leptospira santarosai serogroup Grippotyphosa serovar Bananal. Pulsed field gel electrophoresis resulted in a unique pattern distinct from the reference strains, and the isolates clustered with greater than 85% similarity. The isolates also presented higher growth rates than other Leptospira serovars, with high minimal inhibitory concentration values for most of the tested antibiotics, with the exception of penicillin and ampicillin. This isolation and characterization of the L. santarosai serogroup Grippotyphosa serovar Bananal from capybara, highlights the importance of wild and sinantropic rodents as carriers of pathogenic leptospires. PMID:27258408

  4. Whole genome sequencing revealed host adaptation-focused genomic plasticity of pathogenic Leptospira

    PubMed Central

    Xu, Yinghua; Zhu, Yongzhang; Wang, Yuezhu; Chang, Yung-Fu; Zhang, Ying; Jiang, Xiugao; Zhuang, Xuran; Zhu, Yongqiang; Zhang, Jinlong; Zeng, Lingbing; Yang, Minjun; Li, Shijun; Wang, Shengyue; Ye, Qiang; Xin, Xiaofang; Zhao, Guoping; Zheng, Huajun; Guo, Xiaokui; Wang, Junzhi

    2016-01-01

    Leptospirosis, caused by pathogenic Leptospira spp., has recently been recognized as an emerging infectious disease worldwide. Despite its severity and global importance, knowledge about the molecular pathogenesis and virulence evolution of Leptospira spp. remains limited. Here we sequenced and analyzed 102 isolates representing global sources. A high genomic variability were observed among different Leptospira species, which was attributed to massive gene gain and loss events allowing for adaptation to specific niche conditions and changing host environments. Horizontal gene transfer and gene duplication allowed the stepwise acquisition of virulence factors in pathogenic Leptospira evolved from a recent common ancestor. More importantly, the abundant expansion of specific virulence-related protein families, such as metalloproteases-associated paralogs, were exclusively identified in pathogenic species, reflecting the importance of these protein families in the pathogenesis of leptospirosis. Our observations also indicated that positive selection played a crucial role on this bacteria adaptation to hosts. These novel findings may lead to greater understanding of the global diversity and virulence evolution of Leptospira spp. PMID:26833181

  5. Leptospira contamination in household and environmental water in rural communities in southern Chile.

    PubMed

    Muñoz-Zanzi, Claudia; Mason, Meghan R; Encina, Carolina; Astroza, Angel; Romero, Alex

    2014-07-01

    Leptospirosis is a zoonosis of global distribution that affects tropical and temperate areas. Under suitable conditions, Leptospira can survive in water and soil and contribute to human and animal infections. The objective of this study was to describe the presence of pathogenic Leptospira in peri-domestic water samples from rural households in southern Chile. Water samples, including puddles, containers, animal troughs, rivers, canals, and drinking water were collected from 236 households and tested for Leptospira using a PCR assay targeting the lipL32 gene. Evidence of Leptospira presence was detected in all sample types; overall, 13.5% (77/570) samples tested positive. A total of 10/22 (45.5%) open containers, 12/83 (14.5%) animal drinking sources, 9/47 (19.1%) human drinking sources, and 36/306 (19.3%) puddles tested positive. Lower income (OR = 4.35, p = 0.003), increased temperature (OR = 1.23, p < 0.001), and presence of dogs (OR = 15.9, p = 0.022) were positively associated with positive puddles. Increased number of rodent signs was associated with positive puddles in the household (OR = 3.22); however, only in the lower income households. There was no association between PCR positive rodents and puddles at the household level. Results revealed the ubiquity of Leptospira in the household environment and highlight the need to develop formal approaches for systematic monitoring. PMID:24972030

  6. Leptospira and Brucella antibodies in collared anteaters (Tamandua tetradactyla) in Brazilian zoos.

    PubMed

    Sales, Indiara dos Santos; Folly, Márcio Manhães; Garcia, Luize Néli Nunes; Ramos, Tatiane Mendes Varela; da Silva, Mariana Cristina; Pereira, Martha Maria

    2012-12-01

    The presence of Leptospira spp. and Brucella spp. antibodies was investigated in serum samples from 28 collared anteaters (Tamandua tetradactyla) kept in seven Brazilian zoos. Sera were tested against 19 Leptospira serovars using microscopic agglutination. Samples reacted to the following serovars: two (7.14%) to Patoc, three (10.71%) to Tarrasovi, three (10.71%) to both Patoc and Tarrasovi, two (7.14%) to Wolffi, and one (3.57%) to Australis. Two (7.14%) samples reacted to the buffered Brucella antigen test, but no confirmatory reaction occurred using the 2-mercaptoethanol slow slide agglutination test. No sample was reactive in the agar gel immunodiffusion test for rugose species of Brucella. The presence of anti-leptospira agglutinins in captive T. tetradactyla serum indicates that this species may be susceptible to infection by these bacteria.

  7. LepBank: a Leptospira sequence repository and a portal for phylogenetic studies.

    PubMed

    Eslabão, Marcus R; Dellagostin, Odir A; Cerqueira, Gustavo M

    2010-05-01

    Leptospirosis is a neglected infectious disease that constitutes a threat to both humans and animals. Comprehension about the epidemiological behavior and population dynamics of Leptospira may be helpful for the development of control measures. Thus, an effort was made to organize leptospiral sequences in a new and specific database. In addition, online bioinformatics tools were clustered in a web portal to facilitate sequences manipulation by scientists. LepBank (http://.lepbank.ufpel.edu.br) is a Leptospira sequences repository and a suite for systematics, which brings simplicity to leptospirosis research, integrating sophisticated online programs to a sequence database. We intend the database to be useful for the leptospirosis scientific community, providing standardized and high quality information and facilitating research into key aspects of the Leptospira taxonomy and phylogeny.

  8. Preliminary Investigations on the Distribution of Leptospira Serovars in Domestic Animals in North-west Morocco.

    PubMed

    Benkirane, A; Noury, S; Hartskeerl, R A; Goris, M G A; Ahmed, A; Nally, J E

    2016-04-01

    Leptospirosis is a neglected zoonosis of global importance with a complex epidemiology that affects humans, domestic and wild mammals. However, due to the diversity of clinical signs and difficulties of establishing a confirmatory laboratory diagnosis, the disease remains poorly investigated, particularly in the developing world. In Morocco, a descriptive study of the seroprevalence of Leptospira infection in animals has never been undertaken. To fill this gap, the current study was conducted on a subset of animals in north-west Morocco as a preliminary step towards understanding the epidemiological patterns of animal leptospirosis in the country. The study was conducted on 289 serum samples collected between January and April 2012 from dogs, cattle, sheep, goats and donkeys in the areas of Rabat-Temara, Sidi Kacem and Oulmes. All serum samples were tested by the MAT with 14 reference strains of the most prevalent pathogenic serovars of Leptospira and two serovars of non-pathogenic Leptospira. The overall seroprevalence of Leptospira in cattle, sheep, goats, dogs and donkeys was 15%, 18%, 20%, 21% and 20%, respectively. The most prevalent serogroups found in each species were Ballum, Sejroe, and Australis in cattle, Ballum, Australis and Sejroe in sheep, Australis and Ballum in goats, Javanica and Australis in donkey and Australis, Ballum and Canicola in dogs. Of all the serogroups tested in this study, Icterohaemorrhagiae, the only serogroup which has been previously reported in humans in Morocco, was rarely reactive. The majority of reactive sera were collected from low land areas. A large number of sera samples classified as seronegative when tested against pathogenic leptospires were positive when tested against non-pathogenic leptospires; this is suggestive of possible novel, as yet unclassified, Leptospira serovars in Morocco. Eleven of thirteen sheep urine samples were positive by real-time PCR confirming their role as Leptospira carriers in Morocco. PMID

  9. Leptospira: The Dawn of the Molecular Genetics Era for an Emerging Zoonotic Pathogen

    PubMed Central

    Ko, Albert I.; Goarant, Cyrille; Picardeau, Mathieu

    2011-01-01

    Leptospirosis is a zoonotic disease which has emerged as a major cause of morbidity and mortality among impoverished populations. One centenary after the discovery of the causative spirochaetal agent, little is understood of Leptospira pathogenesis, which in turn has hampered the identification of new intervention strategies to address this neglected disease. However the recent availability of complete genome sequences for Leptospira and discovery of genetic tools to transform the pathogen has led to major insights into the biology and pathogenesis of this pathogen. We discuss the life cycle of the bacterium and the new advances that have been made and their implications for the future prevention of this disease. PMID:19756012

  10. Epidemiology of Leptospira weilii serovar Topaz infections in Australia.

    PubMed

    Slack, Andrew T; Symonds, Meegan L; Dohnt, Michael F; Corney, Bruce G; Smythe, Lee D

    2007-06-01

    Leptospirosis is a zoonotic disease with a worldwide distribution. Leptospira weilii serovar (sv.) Topaz is a newly described serovar first isolated in the far north of Queensland, Australia. The epidemiology of L. weilii sv. Topaz infections in Australia was characterised through the use of surveillance questionnaires and molecular studies. There have been 24 human and 2 animal (bovine and bandicoot) L. weilii sv. Topaz infections diagnosed since 1991. The majority of these infections have occurred in Far North Queensland, with the remaining infections occurring in South East Queensland and in Western Australia. The majority of patients with L. weilii sv. Topaz infections presented with classical leptospirosis symptoms including; fever, headaches, sweats, chills and myalgia. The occupations of human cases of L. weilii sv. Topaz infection included banana farming, dairy and beef cattle production and tourist related activities. Fluorescent amplified fragment length polymorphism (FAFLP) was performed on 15 L. weilii sv. Topaz isolates including 2 animal isolates. Clustering analysis grouped the 15 isolates into 5 main clades with 13 unique FAFLP profiles. A high level of relatedness was demonstrated between 2 animal and 2 human isolates. PMID:17724998

  11. Epidemiology of Leptospira weilii serovar Topaz infections in Australia.

    PubMed

    Slack, Andrew T; Symonds, Meegan L; Dohnt, Michael F; Corney, Bruce G; Smythe, Lee D

    2007-06-01

    Leptospirosis is a zoonotic disease with a worldwide distribution. Leptospira weilii serovar (sv.) Topaz is a newly described serovar first isolated in the far north of Queensland, Australia. The epidemiology of L. weilii sv. Topaz infections in Australia was characterised through the use of surveillance questionnaires and molecular studies. There have been 24 human and 2 animal (bovine and bandicoot) L. weilii sv. Topaz infections diagnosed since 1991. The majority of these infections have occurred in Far North Queensland, with the remaining infections occurring in South East Queensland and in Western Australia. The majority of patients with L. weilii sv. Topaz infections presented with classical leptospirosis symptoms including; fever, headaches, sweats, chills and myalgia. The occupations of human cases of L. weilii sv. Topaz infection included banana farming, dairy and beef cattle production and tourist related activities. Fluorescent amplified fragment length polymorphism (FAFLP) was performed on 15 L. weilii sv. Topaz isolates including 2 animal isolates. Clustering analysis grouped the 15 isolates into 5 main clades with 13 unique FAFLP profiles. A high level of relatedness was demonstrated between 2 animal and 2 human isolates.

  12. Molecular epidemiology of pathogenic Leptospira spp. in the straw-colored fruit bat (Eidolon helvum) migrating to Zambia from the Democratic Republic of Congo.

    PubMed

    Ogawa, Hirohito; Koizumi, Nobuo; Ohnuma, Aiko; Mutemwa, Alisheke; Hang'ombe, Bernard M; Mweene, Aaron S; Takada, Ayato; Sugimoto, Chihiro; Suzuki, Yasuhiko; Kida, Hiroshi; Sawa, Hirofumi

    2015-06-01

    The role played by bats as a potential source of transmission of Leptospira spp. to humans is poorly understood, despite various pathogenic Leptospira spp. being identified in these mammals. Here, we investigated the prevalence and diversity of pathogenic Leptospira spp. that infect the straw-colored fruit bat (Eidolon helvum). We captured this bat species, which is widely distributed in Africa, in Zambia during 2008-2013. We detected the flagellin B gene (flaB) from pathogenic Leptospira spp. in kidney samples from 79 of 529 E. helvum (14.9%) bats. Phylogenetic analysis of 70 flaB fragments amplified from E. helvum samples and previously reported sequences, revealed that 12 of the fragments grouped with Leptospira borgpetersenii and Leptospira kirschneri; however, the remaining 58 flaB fragments appeared not to be associated with any reported species. Additionally, the 16S ribosomal RNA gene (rrs) amplified from 27 randomly chosen flaB-positive samples was compared with previously reported sequences, including bat-derived Leptospira spp. All 27 rrs fragments clustered into a pathogenic group. Eight fragments were located in unique branches, the other 19 fragments were closely related to Leptospira spp. detected in bats. These results show that rrs sequences in bats are genetically related to each other without regional variation, suggesting that Leptospira are evolutionarily well-adapted to bats and have uniquely evolved in the bat population. Our study indicates that pathogenic Leptospira spp. in E. helvum in Zambia have unique genotypes.

  13. Presence of antibodies against Leptospira serovars in Chaetophractus villosus (Mammalia, Dasypodidae), La Pampa province, Argentina.

    PubMed

    Kin, Marta S; Brihuega, Bibiana; Fort, Marcelo; Delgado, Fernando; Bedotti, Daniel; Casanave, Emma B

    2015-01-01

    Leptospirosis is a zoonosis of worldwide distribution. The aim of this study was to examine the presence of antibodies against 21 Leptospira reactive serovars in Chaetophractus villosus in La Pampa province, Argentina, using the microscopic agglutination test (MAT). Pathologic changes compatible with leptospirosis and in situ detection of the agent by immunohistochemistry were studied in 24 and 3 individuals respectively. Only 35/150 (23.3%) serum samples had antibodies against Leptospira sp. Six percent of the samples reacted with serovar Canicola, 4.7% with serovar Castellonis, 1.3% with serovar Icterohemorrhagieae and 0.7% with serovar Hardjo. Sixteen (10.6%) serum samples agglutinated with Castellonis-Icterohemorrhagiae and Canicola-Castellonis serovars, both with 4.7%, and Canicola-Hardjo and Castellonis-Canicola-Icterohemorrhagiae both with 0.6%. Fourteen animals had variable degrees of lesions, which were more severe in animals with higher serological titers (3200), and Leptospira sp. was detected in 3 animals by immunohistochemistry. These results represent the first record of the presence of Leptospira in C. villosus in La Pampa. PMID:25754485

  14. Comparison of Mucosal, Subcutaneous and Intraperitoneal Routes of Rat Leptospira Infection

    PubMed Central

    Zilber, Anne-Laure; Belli, Patrick; Grezel, Delphine; Artois, Marc; Kodjo, Angeli; Djelouadji, Zoheira

    2016-01-01

    Leptospirosis is a zoonosis found worldwide that is caused by a spirochete. The main reservoirs of Leptospira, which presents an asymptomatic infection, are wild rodents, including the brown rat (Rattus norvegicus). Experimental studies of the mechanisms of its renal colonization in rats have previously used an intraperitoneal inoculation route. However, knowledge of rat-rat transmission requires the use of a natural route of inoculation, such as a mucosal or subcutaneous route. We investigated for the first time the effects of subcutaneous and mucosal inoculation routes compared to the reference intraperitoneal route during Leptospira infection in adult rats. Infection characteristics were studied using Leptospira renal isolation, serology, and molecular and histological analyses. Leptospira infection was asymptomatic using each inoculation route, and caused similar antibody production regardless of renal colonization. The observed renal colonization rates were 8 out of 8 rats, 5 out of 8 rats and 1 out of 8 rats for the intraperitoneal, mucosal and subcutaneous inoculation routes, respectively. Thus, among the natural infection routes studied, mucosal inoculation was more efficient for renal colonization associated with urinary excretion than the subcutaneous route and induced a slower-progressing infection than the intraperitoneal route. These results can facilitate understanding of the infection modalities in rats, unlike the epidemiological studies conducted in wild rats. Future studies of other natural inoculation routes in rat models will increase our knowledge of rat-rat disease transmission and allow the investigation of infection kinetics. PMID:27031867

  15. Development of Hamster Models for Acute and Chronic Infections with Leptospira borgpetersenii serovar Hardjo

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Golden Syrian hamster is frequently used as a small animal model to study acute leptospirosis. However, use of this small animal model to study Leptospira borgpetersenii serovar Hardjo infections has not been well documented. Cattle are the normal maintenance hosts of L. borgpetersenii serovar...

  16. Comparison of Mucosal, Subcutaneous and Intraperitoneal Routes of Rat Leptospira Infection.

    PubMed

    Zilber, Anne-Laure; Belli, Patrick; Grezel, Delphine; Artois, Marc; Kodjo, Angeli; Djelouadji, Zoheira

    2016-03-01

    Leptospirosis is a zoonosis found worldwide that is caused by a spirochete. The main reservoirs of Leptospira, which presents an asymptomatic infection, are wild rodents, including the brown rat (Rattus norvegicus). Experimental studies of the mechanisms of its renal colonization in rats have previously used an intraperitoneal inoculation route. However, knowledge of rat-rat transmission requires the use of a natural route of inoculation, such as a mucosal or subcutaneous route. We investigated for the first time the effects of subcutaneous and mucosal inoculation routes compared to the reference intraperitoneal route during Leptospira infection in adult rats. Infection characteristics were studied using Leptospira renal isolation, serology, and molecular and histological analyses. Leptospira infection was asymptomatic using each inoculation route, and caused similar antibody production regardless of renal colonization. The observed renal colonization rates were 8 out of 8 rats, 5 out of 8 rats and 1 out of 8 rats for the intraperitoneal, mucosal and subcutaneous inoculation routes, respectively. Thus, among the natural infection routes studied, mucosal inoculation was more efficient for renal colonization associated with urinary excretion than the subcutaneous route and induced a slower-progressing infection than the intraperitoneal route. These results can facilitate understanding of the infection modalities in rats, unlike the epidemiological studies conducted in wild rats. Future studies of other natural inoculation routes in rat models will increase our knowledge of rat-rat disease transmission and allow the investigation of infection kinetics.

  17. Presence of antibodies against Leptospira serovars in Chaetophractus villosus (Mammalia, Dasypodidae), La Pampa province, Argentina.

    PubMed

    Kin, Marta S; Brihuega, Bibiana; Fort, Marcelo; Delgado, Fernando; Bedotti, Daniel; Casanave, Emma B

    2015-01-01

    Leptospirosis is a zoonosis of worldwide distribution. The aim of this study was to examine the presence of antibodies against 21 Leptospira reactive serovars in Chaetophractus villosus in La Pampa province, Argentina, using the microscopic agglutination test (MAT). Pathologic changes compatible with leptospirosis and in situ detection of the agent by immunohistochemistry were studied in 24 and 3 individuals respectively. Only 35/150 (23.3%) serum samples had antibodies against Leptospira sp. Six percent of the samples reacted with serovar Canicola, 4.7% with serovar Castellonis, 1.3% with serovar Icterohemorrhagieae and 0.7% with serovar Hardjo. Sixteen (10.6%) serum samples agglutinated with Castellonis-Icterohemorrhagiae and Canicola-Castellonis serovars, both with 4.7%, and Canicola-Hardjo and Castellonis-Canicola-Icterohemorrhagiae both with 0.6%. Fourteen animals had variable degrees of lesions, which were more severe in animals with higher serological titers (3200), and Leptospira sp. was detected in 3 animals by immunohistochemistry. These results represent the first record of the presence of Leptospira in C. villosus in La Pampa.

  18. Leptospira Seroprevalence and Risk Factors in Health Centre Patients in Hoima District, Western Uganda

    PubMed Central

    Pearson, Raewynne; Kankya, Clovice; Kajura, Charles; Alinaitwe, Lordrick; Kakooza, Steven; Pelican, Katharine M.; Travis, Dominic A.; Mahero, Michael; Boulware, David R.; Mugisha, Lawrence

    2016-01-01

    Background The burden of human leptospirosis in Uganda is unknown. We estimated the seroprevalence of Leptospira antibodies, probable acute/recent leptospirosis, and risk factors for seropositivity in humans in rural Western Uganda. Methodology and Principal Findings 359 non-pregnant adults visiting the Kikuube and Kigorobya Health Centers were sequentially recruited during March and April 2014. A health history survey and serum were collected from consented participants. Overall, 69% reported having fever in the past year, with 49% reporting malaria, 14% malaria relapse, 6% typhoid fever, 3% brucellosis, and 0% leptospirosis. We tested sera by microscopic agglutination test (MAT) against eight Leptospira serovars representing seven serogroups. Leptospira seroprevalence was 35% (126/359; 95%CI 30.2–40.3%) defined as MAT titer ≥ 1:100 for any serovar. The highest prevalence was against L. borgpetersenii Nigeria (serogroup Pyrogenes) at 19.8% (71/359; 95%CI 15.9–24.4%). The prevalence of probable recent leptospirosis (MAT titer ≥1:800) was 1.9% (95%CI 0.9–4.2%) and uniquely related to serovar Nigeria (serogroup Pyrogenes). Probable recent leptospirosis was associated with having self-reported malaria within the past year (p = 0.048). Higher risk activities included skinning cattle (n = 6) with 12.3 higher odds (95%CI 1.4–108.6; p = 0.024) of Leptospira seropositivity compared with those who had not. Participants living in close proximity to monkeys (n = 229) had 1.92 higher odds (95%CI 1.2–3.1; p = 0.009) of seropositivity compared with participants without monkeys nearby. Conclusions/Significance The 35% prevalence of Leptospira antibodies suggests that exposure to leptospirosis is common in rural Uganda, in particular the Nigeria serovar (Pyrogenes serogroup). Leptospirosis should be a diagnostic consideration in febrile illness and “smear-negative malaria” in rural East Africa. PMID:27487398

  19. Molecular Detection of Leptospira in Two Returned Travelers: Higher Bacterial Load in Cerebrospinal Fluid versus Serum or Plasma

    PubMed Central

    Waggoner, Jesse J.; Soda, Elizabeth A.; Seibert, Ryan; Grant, Philip; Pinsky, Benjamin A.

    2015-01-01

    Leptospirosis is a potentially severe illness in returned travelers. Patients often present with fever, headache, and neck pain, which may lead to a workup for meningitis including the acquisition of cerebrospinal fluid (CSF). Although Leptospira DNA has been detected in CSF by polymerase chain reaction (PCR), little data exist regarding the utility of testing CSF in addition to serum or plasma obtained on presentation. In this report, we present two cases of leptospirosis in returned travelers presenting with fever and headache. Our first patient had neutrophilic meningitis, and Leptospira was detectable only in CSF obtained on admission. The second patient had a normal CSF profile, but Leptospira was detected in CSF at a bacterial load 5- to 10-fold higher than that in plasma. CSF is an important specimen for the diagnosis of Leptospira by molecular methods and may yield an actionable diagnosis in the absence of leptospiremia. PMID:26033024

  20. High Prevalence of Intermediate Leptospira spp. DNA in Febrile Humans from Urban and Rural Ecuador

    PubMed Central

    Chiriboga, Jorge; Barragan, Verónica; Arroyo, Gabriela; Sosa, Andrea; Birdsell, Dawn N.; España, Karool; Mora, Ana; Espín, Emilia; Mejía, María Eugenia; Morales, Melba; Pinargote, Carmina; Gonzalez, Manuel; Hartskeerl, Rudy; Keim, Paul; Bretas, Gustavo; Eisenberg, Joseph N.S.

    2015-01-01

    Leptospira spp., which comprise 3 clusters (pathogenic, saprophytic, and intermediate) that vary in pathogenicity, infect >1 million persons worldwide each year. The disease burden of the intermediate leptospires is unclear. To increase knowledge of this cluster, we used new molecular approaches to characterize Leptospira spp. in 464 samples from febrile patients in rural, semiurban, and urban communities in Ecuador; in 20 samples from nonfebrile persons in the rural community; and in 206 samples from animals in the semiurban community. We observed a higher percentage of leptospiral DNA–positive samples from febrile persons in rural (64%) versus urban (21%) and semiurban (25%) communities; no leptospires were detected in nonfebrile persons. The percentage of intermediate cluster strains in humans (96%) was higher than that of pathogenic cluster strains (4%); strains in animal samples belonged to intermediate (49%) and pathogenic (51%) clusters. Intermediate cluster strains may be causing a substantial amount of fever in coastal Ecuador. PMID:26583534

  1. Infection with Leptospira kirschneri Serovar Mozdok: First Report from the Southern Hemisphere.

    PubMed

    da Cunha, Carlos Eduardo Pouey; Felix, Samuel Rodrigues; Neto, Amilton Clair Pinto Seixas; Campello-Felix, Anelize; Kremer, Frederico Schmitt; Monte, Leonardo Garcia; Amaral, Marta Gonçalves; de Oliveira Nobre, Márcia; da Silva, Éverton Fagonde; Hartleben, Cláudia Pinho; McBride, Alan John Alexander; Dellagostin, Odir Antonio

    2016-03-01

    Leptospirosis is a global zoonosis caused by pathogenic Leptospira spp. In this study, we characterized two Leptospira kirschneri serogroup Pomona serovar Mozdok isolates, one obtained from a dog and the other from a patient with severe leptospirosis, 4 years later. Histopathological analysis showed that both isolates caused severe tissue damage when used to infect hamsters. While L. kirschneri serogroup Pomona serovar Mozdok is endemic in animals in Europe, there is only one report of human leptospirosis in the literature. Although strains belonging to L. kirschneri serogroup Pomona have been identified in cases of human leptospirosis in Europe, serovar Mozdok has not yet been implicated. The 4-year interval between isolations and the fact that this is the first report of serovar Mozdok as the causative agent of human leptospirosis in the southern hemisphere, demonstrates its epidemiological importance to public health. Moreover, the presence of serovar Mozdok in Brazil has the potential to affect vaccine and diagnostic test development. PMID:26755566

  2. Comparative genomic analysis of Brazilian Leptospira kirschneri serogroup Pomona serovar Mozdok.

    PubMed

    Moreno, Luisa Z; Kremer, Frederico S; Miraglia, Fabiana; Loureiro, Ana P; Eslabao, Marcus R; Dellagostin, Odir A; Lilenbaum, Walter; Moreno, Andrea M

    2016-08-01

    Leptospira kirschneri is one of the pathogenic species of the Leptospira genus. Human and animal infection from L. kirschneri gained further attention over the last few decades. Here we present the isolation and characterisation of Brazilian L. kirschneri serogroup Pomona serovar Mozdok strain M36/05 and the comparative genomic analysis with Brazilian human strain 61H. The M36/05 strain caused pulmonary hemorrhagic lesions in the hamster model, showing high virulence. The studied genomes presented high symmetrical identity and the in silico multilocus sequence typing analysis resulted in a new allelic profile (ST101) that so far has only been associated with the Brazilian L. kirschneri serogroup Pomona serovar Mozdok strains. Considering the environmental conditions and high genomic similarity observed between strains, we suggest the existence of a Brazilian L. kirschneri serogroup Pomona serovar Mozdok lineage that could represent a high public health risk; further studies are necessary to confirm the lineage significance and distribution. PMID:27581124

  3. Molecular typing of Leptospira spp. strains isolated from field mice confirms a link to human leptospirosis.

    PubMed

    Li, S J; Wang, D M; Zhang, C C; Li, X W; Yang, H M; Tian, K C; Wei, X Y; Liu, Y; Tang, G P; Jiang, X G; Yan, J

    2013-11-01

    In recent years, human leptospirosis has been reported in Jinping and Liping counties, Guizhou province, but the leptospires have never been isolated. To track the source of infection and understand the aetiological characteristics, we performed surveillance for field mice carriage of leptospirosis in 2011. Four strains of leptospire were isolated from Apodemus agrarius. PCR confirmed the four isolates as pathogenic. Multiple-locus variable-number tandem repeat analysis (MLVA) showed that the four strains were closely related to serovar Lai strain 56601 belonging to serogroup Icterohaemorrhagiae, which is consistent with the antibody detection results from local patients. Furthermore, the diversity of leptospiral isolates from different hosts and regions was demonstrated with MLVA. Our results suggest that A. agrarius may be the main carrier of Leptospira in Jinping and Liping counties, and the serogroup Icterohaemorrhagiae serovar may be the epidemic serogroup of Leptospira. This will contribute to the control and prevention of leptospirosis in these localities. PMID:23406882

  4. Comparative genomic analysis of Brazilian Leptospira kirschneri serogroup Pomona serovar Mozdok

    PubMed Central

    Moreno, Luisa Z; Kremer, Frederico S; Miraglia, Fabiana; Loureiro, Ana P; Eslabao, Marcus R; Dellagostin, Odir A; Lilenbaum, Walter; Moreno, Andrea M

    2016-01-01

    Leptospira kirschneri is one of the pathogenic species of the Leptospira genus. Human and animal infection from L. kirschneri gained further attention over the last few decades. Here we present the isolation and characterisation of Brazilian L. kirschneri serogroup Pomona serovar Mozdok strain M36/05 and the comparative genomic analysis with Brazilian human strain 61H. The M36/05 strain caused pulmonary hemorrhagic lesions in the hamster model, showing high virulence. The studied genomes presented high symmetrical identity and the in silico multilocus sequence typing analysis resulted in a new allelic profile (ST101) that so far has only been associated with the Brazilian L. kirschneri serogroup Pomona serovar Mozdok strains. Considering the environmental conditions and high genomic similarity observed between strains, we suggest the existence of a Brazilian L. kirschneri serogroup Pomona serovar Mozdok lineage that could represent a high public health risk; further studies are necessary to confirm the lineage significance and distribution. PMID:27581124

  5. Comparative genomic analysis of Brazilian Leptospira kirschneri serogroup Pomona serovar Mozdok.

    PubMed

    Moreno, Luisa Z; Kremer, Frederico S; Miraglia, Fabiana; Loureiro, Ana P; Eslabao, Marcus R; Dellagostin, Odir A; Lilenbaum, Walter; Moreno, Andrea M

    2016-08-01

    Leptospira kirschneri is one of the pathogenic species of the Leptospira genus. Human and animal infection from L. kirschneri gained further attention over the last few decades. Here we present the isolation and characterisation of Brazilian L. kirschneri serogroup Pomona serovar Mozdok strain M36/05 and the comparative genomic analysis with Brazilian human strain 61H. The M36/05 strain caused pulmonary hemorrhagic lesions in the hamster model, showing high virulence. The studied genomes presented high symmetrical identity and the in silico multilocus sequence typing analysis resulted in a new allelic profile (ST101) that so far has only been associated with the Brazilian L. kirschneri serogroup Pomona serovar Mozdok strains. Considering the environmental conditions and high genomic similarity observed between strains, we suggest the existence of a Brazilian L. kirschneri serogroup Pomona serovar Mozdok lineage that could represent a high public health risk; further studies are necessary to confirm the lineage significance and distribution.

  6. Prevalence of leptospira in acute hepatitis syndrome and assessment of IL-8 and TNF-alpha level in leptospiral hepatitis.

    PubMed

    Rizvi, M; Azam, M; Ajmal, M R; Shukla, I; Malik, A

    2011-10-01

    To study the prevalence of leptospira in acute hepatitis syndrome and to assess interleukin (IL)-8 and tumour necrosis factor (TNF)-alpha levels in the pathogenesis of hepatitis due to leptospiral infection. Two hundred and forty-seven consecutive cases with symptoms of acute hepatitis and 30 healthy controls were enrolled in the study and detailed clinical history was elicited from them. Enzyme-linked immunosorbent assays (ELISAs) for HAV, HBV, HCV and HEV were performed to rule out common viral aetiology of hepatitis. IgM antibodies to leptospira were detected by ELISA. IL-8 and TNF-alpha levels were estimated in leptospira-positive cases and healthy controls by ELISA. Out of 247 cases of acute hepatitis, 46 (18.62%) were observed to be positive for IgM antibodies for leptospira. The mean age of these patients was 31.99 ± 0.28 years (25 males and 21 females; M/F ratio: 1.19:1). The mean ALT, AST and ASP were raised in the majority of patients. IL-8 was found to be elevated (130.81 pg/ml) in a large majority of cases 41/46, 89.1% (P < 0.001). Patients with more severe symptoms were associated with higher levels of IL-8. One mortality was observed due to leptospira. Unpredictably, TNF-alpha level was largely suppressed (45.63 pg/ml) in most of the leptospira-positive patients in comparison with healthy controls. Leptospira-induced hepatitis should be actively looked for in patients negative for A-E viral hepatitis. IL-8 appears to play an important role in the pathogenesis of leptospiral hepatitis. High TNF-alpha should alert clinicians for aggressive in hospital management of patients. PMID:22185944

  7. In vitro susceptibilities of Leptospira spp. and Borrelia burgdorferi isolates to amoxicillin, tilmicosin, and enrofloxacin.

    PubMed

    Kim, Doo; Kordick, Dorsey; Divers, Thomas; Chang, Yung Fu

    2006-12-01

    Antimicrobial susceptibility testing was conducted with 6 different spirochetal strains (4 strains of Leptospira spp. and 2 strains of Borrelia burgdorferi) against 3 antimicrobial agents, commonly used in equine and bovine practice. The ranges of MIC and MBC of amoxicillin against Leptospira spp. were 0.05 - 6.25 microgram/ml and 6.25 - 25.0 microgram/ml, respectively. And the ranges of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of amoxicillin against B. burgdorferi were 0.05 - 0.39 microgram/ml and 0.20 - 0.78 microgram/ml, respectively. The ranges of MIC and MBC of enrofloxacin against Leptospira spp. were 0.05 - 0.39 microgram/ml and 0.05 - 0.39 microgram/ml, respectively. Two strains of B. burgdorferi were resistant to enrofloxacin at the highest concentration tested for MBC (>or=100 microgram/ml). Therefore, the potential role of tilmicosin in the treatment of leptospirosis and borreliosis should be further evaluated in animal models to understand whether the in vivo studies will confirm in vitro results. All spirochetal isolates were inhibited (MIC) and were killed (MBC) by tilmicosin at concentrations below the limit of testing (

  8. In vitro susceptibilities of Leptospira spp. and Borrelia burgdorferi isolates to amoxicillin, tilmicosin, and enrofloxacin

    PubMed Central

    Kim, Doo; Kordick, Dorsey; Divers, Thomas

    2006-01-01

    Antimicrobial susceptibility testing was conducted with 6 different spirochetal strains (4 strains of Leptospira spp. and 2 strains of Borrelia burgdorferi) against 3 antimicrobial agents, commonly used in equine and bovine practice. The ranges of MIC and MBC of amoxicillin against Leptospira spp. were 0.05-6.25 µg/ml and 6.25-25.0 µg/ml, respectively. And the ranges of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of amoxicillin against B. burgdorferi were 0.05-0.39 µg/ml and 0.20-0.78 µg/ml, respectively. The ranges of MIC and MBC of enrofloxacin against Leptospira spp. were 0.05-0.39 µg/ml and 0.05-0.39 µg/ml, respectively. Two strains of B. burgdorferi were resistant to enrofloxacin at the highest concentration tested for MBC (≥100 µg/ml). Therefore, the potential role of tilmicosin in the treatment of leptospirosis and borreliosis should be further evaluated in animal models to understand whether the in vivo studies will confirm in vitro results. All spirochetal isolates were inhibited (MIC) and were killed (MBC) by tilmicosin at concentrations below the limit of testing (≤0.01 µg/ml). PMID:17106227

  9. Seroprevalence of Leptospira spp. and Borrelia burgdorferi sensu lato in Italian horses.

    PubMed

    Ebani, Valentina V; Bertelloni, Fabrizio; Pinzauti, Paolo; Cerri, Domenico

    2012-01-01

    The aim of the study was to determine the seroprevalence of Leptospira spp. and Borrelia burgdorferi sensu lato in healthy horses living in 7 provinces of central Italy. In the period 2007-2009, sera from 386 horses were tested by microagglutination test (MAT) to detect antibodies to Leptospira spp., employing the following serovars as antigens: Bratislava, Ballum, Canicola, Icterohaemorrhagiae, Grippotyphosa, Hardjo, Pomona, Tarassovi. 3 animals were positive for the serovars Icterohaemorrhagiae, 2 to Bratislava, and 1 to Pomona, for a total 1.5% seroprevalence. All sera were examined by immunofluorence antibody test (IFAT) to reveal anti-B. burgdorferi s.l. antibodies. 94 (24.3%) horses were positive with antibody titres ranging from 1:64 to 1:1,024. The seroprevalence was significantly higher in >10 year-old horses compared to younger subjects. No significant differences in the mean seroprevalence were observed in the respective years. The total mean seroprevalence were strictly related to the environmental conditions of the areas in which the horses lived. No cross-reactions between Leptospira and Borrelia were observed. This is the first serological survey on antibodies to B. burgdorferi s.l. in Italian horses. PMID:22742794

  10. Leptospira spp. infection in wild ruminants: a survey in Central Italian Alps.

    PubMed

    Andreoli, Elena; Radaelli, Enrico; Bertoletti, Irene; Bianchi, Alessandro; Scanziani, Eugenio; Tagliabue, Silvia; Mattiello, Silvana

    2014-12-29

    Leptospirosis is an important zoonotic disease diffused worldwide, and wildlife species are commonly considered to be important epidemiological carriers. Four-hundred and forty-one serological and 198 renal samples from red deer, roe deer and chamois collected in the Province of Sondrio were analysed using the microscopic agglutination test and histopathologic examination. Positive serological findings were found only in 15 red deer and 19 positive serologic reactions were recorded. The most frequent serovars were Bratislava and Grippotyphosa, followed by Pomona, Hardjo and Copenhagheni. Twenty-two per cent of renal samples from seropositive red deer were affected by mild to moderate multifocal chronic lymphoplasmacytic and fibrosing tubulo-interstitial nephritis, mainly involving the cortical parenchyma. In this study, antibodies to Leptospira spp. were infrequent in wild ruminants, and only red deer seemed to be sensitive to the infection. Given the low presence and the fact that there was no record of Leptospira spp. infections in cattle, sheep, goats and also hunters in area during the study period, wild ruminants in Alpine environments cannot be considered as reservoirs or important sources of Leptospira spp. infection for humans or domestic animals.

  11. The Utility of Blood Culture Fluid for the Molecular Diagnosis of Leptospira: A Prospective Evaluation.

    PubMed

    Dittrich, Sabine; Rudgard, William E; Woods, Kate L; Silisouk, Joy; Phuklia, Weerawat; Davong, Viengmon; Vongsouvath, Manivanh; Phommasone, Koukeo; Rattanavong, Sayaphet; Knappik, Michael; Craig, Scott B; Weier, Steven L; Tulsiani, Suhella M; Dance, David A B; Newton, Paul N

    2016-04-01

    Leptospirosis is an important zoonosis worldwide, with infections occurring after exposure to contaminated water. Despite being a global problem, laboratory diagnosis remains difficult with culture results taking up to 3 months, serology being retrospective by nature, and polymerase chain reaction showing limited sensitivity. Leptospira have been shown to survive and multiply in blood culture media, and we hypothesized that extracting DNA from incubated blood culture fluid (BCF), followed by quantitative real-time polymerase chain reaction (qPCR) could improve the accuracy and speed of leptospira diagnosis. We assessed this retrospectively, using preincubated BCF of Leptospira spp. positive (N= 109) and negative (N= 63) febrile patients in Vientiane, Lao PDR. The final method showed promising sensitivities of 66% (95% confidence interval [CI]: 55-76) and 59% (95% CI: 49-68) compared with direct or direct and indirect testing combined, as the respective reference standards (specificities > 95%). Despite these promising diagnostic parameters, a subsequent prospective evaluation in a Lao hospital population (N= 352) showed that the sensitivity was very low (∼30%) compared with qPCR on venous blood samples. The disappointingly low sensitivity does suggest that venous blood samples are preferable for the clinical microbiology laboratory, although BCF might be an alternative if leptospirosis is only suspected postadmission after antibiotics have been used. PMID:26880775

  12. Development of Lyophilized Loop-Mediated Isothermal Amplification Reagents for the Detection of Leptospira.

    PubMed

    Chen, Hua-Wei; Weissenberger, Giulia; Ching, Wei-Mei

    2016-05-01

    Leptospirosis is considered to be the most widespread zoonosis. This worldwide emerging infectious disease is caused by the pathogenic species belonging to the genus Leptospira. Polymerase chain reaction (PCR)-based diagnostic assays have been developed for detecting Leptospira DNA in cell cultures and clinical samples. Because PCR requires specialized equipment and extensive end-user training, it is not suitable for routine work in resource-limited areas. We have developed a loop-mediated isothermal amplification (LAMP) assay to detect the presence of Leptospira in patient, animal and environmental samples using lyophilized reagents at a single temperature of around 63°C with a heating block. The sensitivity of this LAMP assay is very similar to the PCR method. The amplified DNA products can be visualized with the naked eyes using hydroxy naphthol blue or detected by the fluorescence signal of SYBR green dye in the reaction when an ultraviolet lamp or compact fluorescence tube scanner is used. This LAMP assay is simple, rapid, and can be performed with a water bath or heating block. The lyophilized LAMP reagents are stable for 3 months when stored at 4°C and 1 month when stored at 25°C, respectively. It is ideal for resource-limited settings where leptospirosis is endemic. PMID:27168577

  13. Identification of Leptospira serovars by RFLP of the RNA polymerase beta subunit gene (rpoB)

    PubMed Central

    Jung, Lenice Roteia Cardoso; Bomfim, Maria Rosa Quaresma; Kroon, Erna Geessien; Nunes, Álvaro Cantini

    2015-01-01

    Leptospires are usually classified by methods based on DNA-DNA hybridization and the conventional cross-agglutination absorption test, which uses polyclonal antibodies against lipopolysaccharides. In this study, the amplification of the rpoB gene, which encodes the beta-subunit of RNA polymerase, was used as an alternative tool to identify Leptospira. DNA extracts from sixty-eight serovars were obtained, and the hypervariable region located between 1990 and 2500-bp in the rpoB gene was amplified by polymerase chain reaction (PCR). The 600-bp amplicons of the rpoB gene were digested with the restriction endonucleases TaqI, Tru1I, Sau3AI and MslI, and the restriction fragments were separated by 6% polyacrylamide gel electrophoresis. Thirty-five fragment patters were obtained from the combined data of restriction fragment length polymorphism (PCR-RFLP) analysis and used to infer the phylogenetic relationships among the Leptospira species and serovars. The species assignments obtained were in full agreement with the established taxonomic classifications. Twenty-two serovars were effectively identified based on differences in their molecular profiles. However, the other 46 serovars remained clustered in groups that included more than one serovar of different species. This study demonstrates the value of RFLP analysis of PCR-amplified rpoB as an initial method for identifying Leptospira species and serovars. PMID:26273261

  14. In vitro susceptibilities of Leptospira spp. and Borrelia burgdorferi isolates to amoxicillin, tilmicosin, and enrofloxacin.

    PubMed

    Kim, Doo; Kordick, Dorsey; Divers, Thomas; Chang, Yung Fu

    2006-12-01

    Antimicrobial susceptibility testing was conducted with 6 different spirochetal strains (4 strains of Leptospira spp. and 2 strains of Borrelia burgdorferi) against 3 antimicrobial agents, commonly used in equine and bovine practice. The ranges of MIC and MBC of amoxicillin against Leptospira spp. were 0.05 - 6.25 microgram/ml and 6.25 - 25.0 microgram/ml, respectively. And the ranges of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of amoxicillin against B. burgdorferi were 0.05 - 0.39 microgram/ml and 0.20 - 0.78 microgram/ml, respectively. The ranges of MIC and MBC of enrofloxacin against Leptospira spp. were 0.05 - 0.39 microgram/ml and 0.05 - 0.39 microgram/ml, respectively. Two strains of B. burgdorferi were resistant to enrofloxacin at the highest concentration tested for MBC (>or=100 microgram/ml). Therefore, the potential role of tilmicosin in the treatment of leptospirosis and borreliosis should be further evaluated in animal models to understand whether the in vivo studies will confirm in vitro results. All spirochetal isolates were inhibited (MIC) and were killed (MBC) by tilmicosin at concentrations below the limit of testing (

  15. The C-terminal variable domain of LigB from Leptospira mediates binding to fibronectin

    PubMed Central

    Lin, Yi-Pin

    2008-01-01

    Adhesion through microbial surface components that recognize adhesive matrix molecules is an essential step in infection for most pathogenic bacteria. In this study, we report that LigB interacts with fibronectin (Fn) through its variable region. A possible role for LigB in bacterial attachment to host cells during the course of infection is supported by the following observations: (i) binding of the variable region of LigB to Madin-Darby canine kidney (MDCK) cells in a dose-dependent manner reduces the adhesion of Leptospira, (ii) inhibition of leptospiral attachment to Fn by the variable region of LigB, and (iii) decrease in binding of the variable region of LigB to the MDCK cells in the presence of Fn. Furthermore, we found a significant reduction in binding of the variable region of LigB to Fn using small interfering RNA (siRNA). Finally, the isothermal titration calorimetric results confirmed the interaction between the variable region of LigB and Fn. This is the first report to demonstrate that LigB binds to MDCK cells. In addition, the reduction of Fn expression in the MDCK cells, by siRNA, reduced the binding of LigB. Taken together, the data from the present study showed that LigB is a Fn-binding protein of pathogenic Leptospira spp. and may play a pivotal role in Leptospira-host interaction during the initial stage of infection. PMID:18487934

  16. [Construction of genomic library of L. interrogans serovar lai using lambda gt11 as the vector and a study of recombiant plasmid pDL121].

    PubMed

    Liu, H; Dai, B; Jing, B; Wu, W; Li, S; Fang, Z; Zhao, H; Ye, D; Yan, R; Liu, J; Song, S; Yang, Y; Zhang, Y; Liu, F; Tu, Y; Yang, H; Huang, Z; Liang, L; Hu, L; Zhao, M

    1997-03-01

    A genomic library of L. interrogans serovar lai strain 017 has been constructed using lambda gt11 as the vector. DNA was partially digested by two blunt-end restriction enzymes, then methylated with EcoR I methylase; after EcoR I linker was added to the DNA, the linker-ended DNA was ligated to the dephosphorylated EcoR I digested lambda gt11 arms. The recombined DNA was packaged in vitro, and used to transduct E. coli Y1090 for amplification. There were 2.1 x 10(6) recombinant bacteriophages as recognized by their ability to form white plaques plated on Lac host in the presence of both IPTG and X-Ga1. A positive clone, designated lambda DL12, was screened with a rabbit anti-serum against L. interrogans serovar lai from the genomic library. The DNA from lambda DL12 was subcloned into plasmid pUC18. A recombinant (designated as pDL121) was obtained. SDS-PAGE analysis indicated that a 23 kd was expressed in E. coli JM 103 harboring pDL121. Western blotting analysis showed that a specific protein band molecular weight of 23 kd could be recognized by the rabbit antiserum against L. interrogans serovar lai strain 017. PMID:10684055

  17. Contrasting Patterns in Mammal–Bacteria Coevolution: Bartonella and Leptospira in Bats and Rodents

    PubMed Central

    Lei, Bonnie R.; Olival, Kevin J.

    2014-01-01

    Background Emerging bacterial zoonoses in bats and rodents remain relatively understudied. We conduct the first comparative host–pathogen coevolutionary analyses of bacterial pathogens in these hosts, using Bartonella spp. and Leptospira spp. as a model. Methodology/Principal Findings We used published genetic data for 51 Bartonella genotypes from 24 bat species, 129 Bartonella from 38 rodents, and 26 Leptospira from 20 bats. We generated maximum likelihood and Bayesian phylogenies for hosts and bacteria, and tested for coevoutionary congruence using programs ParaFit, PACO, and Jane. Bartonella spp. and their bat hosts had a significant coevolutionary fit (ParaFitGlobal = 1.9703, P≤0.001; m2 global value = 7.3320, P≤0.0001). Bartonella spp. and rodent hosts also indicated strong overall patterns of cospeciation (ParaFitGlobal = 102.4409, P≤0.001; m2 global value = 86.532, P≤0.0001). In contrast, we were unable to reject independence of speciation events in Leptospira and bats (ParaFitGlobal = 0.0042, P = 0.84; m2 global value = 4.6310, P = 0.5629). Separate analyses of New World and Old World data subsets yielded results congruent with analysis from entire datasets. We also conducted event-based cophylogeny analyses to reconstruct likely evolutionary histories for each group of pathogens and hosts. Leptospira and bats had the greatest number of host switches per parasite (0.731), while Bartonella and rodents had the fewest (0.264). Conclusions/Significance In both bat and rodent hosts, Bartonella exhibits significant coevolution with minimal host switching, while Leptospira in bats lacks evolutionary congruence with its host and has high number of host switches. Reasons underlying these variable coevolutionary patterns in host range are likely due to differences in disease-specific transmission and host ecology. Understanding the coevolutionary patterns and frequency of host-switching events between bacterial pathogens and

  18. Frequency and type of renal lesions in dogs naturally infected with leptospira species.

    PubMed

    Ortega-Pacheco, A; Colin-Flores, R F; Gutiérrez-Blanco, E; Jiménez-Coello, M

    2008-12-01

    The aim of this study was to determine the frequency and type of renal lesions associated with positive titers against Leptospira sp. in a stray dog population. Three hundred fifty pairs of kidneys and an equal number of serum samples were collected from dogs captured by the dog pound of Merida, Yucatan, Mexico. Euthanasia of dogs was performed following the regulations of the Official Mexican Health Ministry (NOM-033-ZOO-1995). Serum samples were evaluated with the microscopic agglutination test, and tissue samples were processed and fixed in paraffin. After staining with hematoxylin and eosin, the frequency of renal lesions was determined and classified. As an additional evaluation, samples with interstitial nephritis were stained by the Warthin-Starry method in order to observe the presence of spirochete forms that could be morphologically compatible with Leptospira spp. We found that 98% of cases presented at least one type of lesion. The main histological lesions found were mesangial proliferative glomerulonephritis (MPGN) in 63.7% (n= 223), mesangial proliferative glomerulonephritis and interstitial nephritis (MPGN+IN) in 34% (n= 119), nephrosclerosis in 0.57% (n= 2), mesangial glomerulonephritis in 0.28% (n= 1), and interstitial nephritis (IN) in 0.28% (n= 1). Thirty-four percent (n= 122) of the dogs were seropositive to Leptospira sp., mainly against serovar canicola. Among dogs with IN (alone or associated with MPGN) (n= 120), 49.1% were seropositive to Leptospira sp., but only 17% of them showed spirochete forms compatible with the bacteria. A statistical association between seropositive dogs and the presence of MPGN+IN was determined (P < 0.0001; odds ratio 2.7, confidence interval 1.7-4.5). We concluded that the frequency of renal lesions found in this study is high and L. canicola is probably the most common circulating serovar in dogs from this area. Dogs that have been in contact with Leptospira spp. have a higher risk of developing renal lesions of

  19. Asymptomatic Renal Colonization of Humans in the Peruvian Amazon by Leptospira

    PubMed Central

    Ganoza, Christian A.; Matthias, Michael A.; Saito, Mayuko; Cespedes, Manuel; Gotuzzo, Eduardo; Vinetz, Joseph M.

    2010-01-01

    Background Renal carriage and shedding of leptospires is characteristic of carrier or maintenance animal hosts. Sporadic reports indicate that after infection, humans may excrete leptospires for extended periods. We hypothesized that, like mammalian reservoir hosts, humans develop asymptomatic leptospiruria in settings of high disease transmission such as the Peruvian Amazon. Methodology/Principal Findings Using a cross-sectional study design, we used a combination of epidemiological data, serology and molecular detection of the leptospiral 16S rRNA gene to identify asymptomatic urinary shedders of Leptospira. Approximately one-third of the 314 asymptomatic participants had circulating anti-leptospiral antibodies. Among enrolled participants, 189/314 (59%) had evidence of recent infection (microscopic agglutination test (MAT0 ≥1∶800 or ELISA IgM-positive or both). The proportion of MAT-positive and high MAT-titer (≥1∶800) persons was higher in men than women (p = 0.006). Among these people, 13/314 (4.1%) had Leptospira DNA-positive urine samples. Of these, the 16S rRNA gene from 10 samples was able to be sequenced. The urine-derived species clustered within both pathogenic (n = 6) and intermediate clades of Leptospira (n = 4). All of the thirteen participants with leptospiral DNA in urine were women. The median age of the DNA-positive group was older compared to the negative group (p≤0.05). A group of asymptomatic participants (“long-term asymptomatic individuals,” 102/341 (32.5%) of enrolled individuals) without serological evidence of recent infection was identified; within this group, 6/102 (5.9%) excreted pathogenic and intermediate-pathogenic Leptospira (75–229 bacteria/mL of urine). Conclusions/Significance Asymptomatic renal colonization of leptospires in a region of high disease transmission is common, including among people without serological or clinical evidence of recent infection. Both pathogenic and intermediate Leptospira

  20. Duration of immunity of a multivalent (DHPPi/L4R) canine vaccine against four Leptospira serovars.

    PubMed

    Wilson, Stephen; Stirling, Catrina; Thomas, Anne; King, Vickie; Plevová, Edita; Chromá, Ludmila; Siedek, Elisabeth; Illambas, Joanna; Salt, Jeremy; Sture, Gordon

    2013-06-28

    Despite effective vaccines against common Leptospira serovars, the development of new products with long duration of immunity is still important to protect dogs against leptospirosis. The results from four challenge studies performed one year after vaccination of dogs with a multivalent vaccine containing four Leptospira antigens are reported. Six week old dogs received two vaccinations, three weeks apart, and were challenged 367 days later. Clinical observations were recorded, while blood (culture, biochemistry and haematology), urine (culture) and liver and kidney (culture) samples were collected throughout the study or at necropsy. All control dogs remained seronegative until challenge, when they seroconverted. Antibody titres to Leptospira antigens were seen in vaccinated dogs 21 days after first vaccination and peaked three to six weeks after the second vaccination. Titres decreased in all studies over the following 12 months, until challenge when anamnestic responses were observed. In all studies control dogs demonstrated various abnormal clinical signs, while no vaccinated dogs were affected; differences between groups were only significant following L. bratislava challenge. Analysis of blood cultures showed all control and five of the 24 vaccinated dogs were Leptospira positive after challenge; all studies showed significant differences between treatment groups in mean number of days with positive cultures. Significant differences between vaccinated and control groups in mean number of days with positive urine cultures were also observed, with all non-vaccinated and one vaccinated dog Leptospira positive. The urine culture positive vaccinated dog also gave positive culture from kidney and liver samples. All except one control dog also showed positive Leptospira isolation from kidney or liver, with significant differences between vaccinated and control groups observed. The results demonstrate that administration of a new vaccine to six week old puppies

  1. Comparative study of four saprophytic leptospira strains as screening antigens in the serodiagnosis of leptospirosis in water buffaloes (Bubalus bubalis).

    PubMed

    Girio, R J; Yanaguita, R M; Mathias, L A

    1985-03-01

    The profitability of four saprophytic leptospira strains (Buenos Aires, Patoc 1, Rufino and São Paulo) as polyvalent antigen in the serodiagnosis of leptospirosis in water buffaloes was studied by the microscopic agglutination test. From 104 examined sera tested against 16 pathogenic leptospira serotypes, 52 were positives and 52 were negatives. The results led to the conclusion that from four studied strains, Buenos Aires strain showed the best results when utilized in screening tests for the serodiagnosis of leptospirosis in water buffaloes. The other three strains showed a good specificity but their sensibilities were poor and therefore should not be recommended for diagnosing water buffalo leptospirosis.

  2. A cross sectional observational study to estimate herd level risk factors for Leptospira spp. serovars in small holder dairy cattle farms in southern Chile

    PubMed Central

    2014-01-01

    Background The south of Chile constitutes the main cattle milk producing area of the country. Regarding leptospirosis control in Chile, there is neither an official program nor an epidemiological characterization of smallholder dairy farms. This study was carried out to determine Leptospira seroprevalence and to evaluate risk factors associated with seropositivity at herd level in smallholder bovine dairy herds in southern Chile. A cross-sectional study was conducted, and a convenient sample of 1,537 apparently healthy dairy cows was included in the study. Individual blood samples were taken and examined for six selected reference Leptospira serovars by the Microscopic Agglutination Test (MAT). Results Of the included herds 75% (52/69) showed serological titers against one or more Leptospira serovar. Leptospira borgpetersenii serovar Hardjo was the serovar most frequently (81%) reported from animals with positive results. The variables considered risk factors for Leptospira seropositivity were calve natural breeding system, using a specific calving area and vaccination against Leptospira. Adult cows in contact with calves weaned, proved to be a protective factor against infection. Conclusions Herds neglecting the management practices mentioned in this study could represent an important source of Leptospira infection for other herds in the same geographic area, as well as for other animal species. PMID:24906684

  3. OmpL1 Is an Extracellular Matrix- and Plasminogen-Interacting Protein of Leptospira spp.

    PubMed Central

    Fernandes, Luis G. V.; Vieira, Monica L.; Kirchgatter, Karin; Alves, Ivy J.; de Morais, Zenaide M.; Vasconcellos, Silvio A.; Romero, Eliete C.

    2012-01-01

    Leptospirosis is a zoonosis with multisystem involvement caused by pathogenic strains of the genus Leptospira. OmpL1 is an outer membrane protein of Leptospira spp. that is expressed during infection. In this work, we investigated novel features of this protein. We describe that OmpL1 is a novel leptospiral extracellular matrix (ECM)-binding protein and a plasminogen (PLG) receptor. The recombinant protein was expressed in Escherichia coli BL21(DE3) Star/pLysS as inclusion bodies, refolded, and purified by metal-chelating chromatography. The protein presented a typical β-strand secondary structure, as evaluated by circular dichroism spectroscopy. The recombinant protein reacted with antibodies in serum samples from convalescent leptospirosis patients with a high specificity compared to serum samples from individuals with unrelated diseases. These data strengthen the usefulness of OmpL1 as a diagnostic marker of leptospirosis. The characterization of the immunogenicity of recombinant OmpL1 in inoculated BALB/c mice showed that the protein has the capacity to elicit humoral and cellular immune responses, as denoted by high antibody titers and the proliferation of lymphocytes. We demonstrate that OmpL1 has the ability to mediate attachment to laminin and plasma fibronectin, with KD (equilibrium dissociation constant) values of 2,099.93 ± 871.03 nM and 1,239.23 ± 506.85 nM, respectively. OmpL1 is also a PLG receptor, with a KD of 368.63 ± 121.23 nM, capable of generating enzymatically active plasmin. This is the first report that shows and characterizes OmpL1 as an ECM-interacting and a PLG-binding protein of Leptospira spp. that may play a role in bacterial pathogenesis when expressed during infection. PMID:22802342

  4. An Optimized Method for Quantification of Pathogenic Leptospira in Environmental Water Samples.

    PubMed

    Riediger, Irina N; Hoffmaster, Alex R; Casanovas-Massana, Arnau; Biondo, Alexander W; Ko, Albert I; Stoddard, Robyn A

    2016-01-01

    Leptospirosis is a zoonotic disease usually acquired by contact with water contaminated with urine of infected animals. However, few molecular methods have been used to monitor or quantify pathogenic Leptospira in environmental water samples. Here we optimized a DNA extraction method for the quantification of leptospires using a previously described Taqman-based qPCR method targeting lipL32, a gene unique to and highly conserved in pathogenic Leptospira. QIAamp DNA mini, MO BIO PowerWater DNA and PowerSoil DNA Isolation kits were evaluated to extract DNA from sewage, pond, river and ultrapure water samples spiked with leptospires. Performance of each kit varied with sample type. Sample processing methods were further evaluated and optimized using the PowerSoil DNA kit due to its performance on turbid water samples and reproducibility. Centrifugation speeds, water volumes and use of Escherichia coli as a carrier were compared to improve DNA recovery. All matrices showed a strong linearity in a range of concentrations from 106 to 10° leptospires/mL and lower limits of detection ranging from <1 cell /ml for river water to 36 cells/mL for ultrapure water with E. coli as a carrier. In conclusion, we optimized a method to quantify pathogenic Leptospira in environmental waters (river, pond and sewage) which consists of the concentration of 40 mL samples by centrifugation at 15,000×g for 20 minutes at 4°C, followed by DNA extraction with the PowerSoil DNA Isolation kit. Although the method described herein needs to be validated in environmental studies, it potentially provides the opportunity for effective, timely and sensitive assessment of environmental leptospiral burden. PMID:27487084

  5. An Optimized Method for Quantification of Pathogenic Leptospira in Environmental Water Samples

    PubMed Central

    Riediger, Irina N.; Hoffmaster, Alex R.; Biondo, Alexander W.; Ko, Albert I.; Stoddard, Robyn A.

    2016-01-01

    Leptospirosis is a zoonotic disease usually acquired by contact with water contaminated with urine of infected animals. However, few molecular methods have been used to monitor or quantify pathogenic Leptospira in environmental water samples. Here we optimized a DNA extraction method for the quantification of leptospires using a previously described Taqman-based qPCR method targeting lipL32, a gene unique to and highly conserved in pathogenic Leptospira. QIAamp DNA mini, MO BIO PowerWater DNA and PowerSoil DNA Isolation kits were evaluated to extract DNA from sewage, pond, river and ultrapure water samples spiked with leptospires. Performance of each kit varied with sample type. Sample processing methods were further evaluated and optimized using the PowerSoil DNA kit due to its performance on turbid water samples and reproducibility. Centrifugation speeds, water volumes and use of Escherichia coli as a carrier were compared to improve DNA recovery. All matrices showed a strong linearity in a range of concentrations from 106 to 10° leptospires/mL and lower limits of detection ranging from <1 cell /ml for river water to 36 cells/mL for ultrapure water with E. coli as a carrier. In conclusion, we optimized a method to quantify pathogenic Leptospira in environmental waters (river, pond and sewage) which consists of the concentration of 40 mL samples by centrifugation at 15,000×g for 20 minutes at 4°C, followed by DNA extraction with the PowerSoil DNA Isolation kit. Although the method described herein needs to be validated in environmental studies, it potentially provides the opportunity for effective, timely and sensitive assessment of environmental leptospiral burden. PMID:27487084

  6. Orientia, rickettsia, and leptospira pathogens as causes of CNS infections in Laos: a prospective study

    PubMed Central

    Dittrich, Sabine; Rattanavong, Sayaphet; Lee, Sue J; Panyanivong, Phonepasith; Craig, Scott B; Tulsiani, Suhella M; Blacksell, Stuart D; Dance, David A B; Dubot-Pérès, Audrey; Sengduangphachanh, Amphone; Phoumin, Phonelavanh; Paris, Daniel H; Newton, Paul N

    2015-01-01

    Summary Background Scrub typhus (caused by Orientia tsutsugamushi), murine typhus (caused by Rickettsia typhi), and leptospirosis are common causes of febrile illness in Asia; meningitis and meningoencephalitis are severe complications. However, scarce data exist for the burden of these pathogens in patients with CNS disease in endemic countries. Laos is representative of vast economically poor rural areas in Asia with little medical information to guide public health policy. We assessed whether these pathogens are important causes of CNS infections in Laos. Methods Between Jan 10, 2003, and Nov 25, 2011, we enrolled 1112 consecutive patients of all ages admitted with CNS symptoms or signs requiring a lumbar puncture at Mahosot Hospital, Vientiane, Laos. Microbiological examinations (culture, PCR, and serology) targeted so-called conventional bacterial infections (Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae, S suis) and O tsutsugamushi, Rickettsia typhi/Rickettsia spp, and Leptospira spp infections in blood or cerebrospinal fluid (CSF). We analysed and compared causes and clinical and CSF characteristics between patient groups. Findings 1051 (95%) of 1112 patients who presented had CSF available for analysis, of whom 254 (24%) had a CNS infection attributable to a bacterial or fungal pathogen. 90 (35%) of these 254 infections were caused by O tsutsugamushi, R typhi/Rickettsia spp, or Leptospira spp. These pathogens were significantly more frequent than conventional bacterial infections (90/1051 [9%] vs 42/1051 [4%]; p<0·0001) by use of conservative diagnostic definitions. CNS infections had a high mortality (236/876 [27%]), with 18% (13/71) for R typhi/Rickettsia spp, O tsutsugamushi, and Leptospira spp combined, and 33% (13/39) for conventional bacterial infections (p=0·076). Interpretation Our data suggest that R typhi/Rickettsia spp, O tsutsugamushi, and Leptospira spp infections are important causes of CNS infections in Laos

  7. Transcriptome Sequencing Reveals Wide Expression Reprogramming of Basal and Unknown Genes in Leptospira biflexa Biofilms.

    PubMed

    Iraola, Gregorio; Spangenberg, Lucía; Lopes Bastos, Bruno; Graña, Martín; Vasconcelos, Larissa; Almeida, Áurea; Greif, Gonzalo; Robello, Carlos; Ristow, Paula; Naya, Hugo

    2016-01-01

    The genus Leptospira is composed of pathogenic and saprophytic spirochetes. Pathogenic Leptospira is the etiological agent of leptospirosis, a globally spread neglected disease. A key ecological feature of some pathogenic species is their ability to survive both within and outside the host. For most leptospires, the ability to persist outside the host is associated with biofilm formation, a most important bacterial strategy to face and overcome hostile environmental conditions. The architecture and biochemistry of leptospiral biofilms are rather well understood; however, the genetic program underpinning biofilm formation remains mostly unknown. In this work, we used the saprophyte Leptospira biflexa as a model organism to assess over- and underrepresented transcripts during the biofilm state, using transcriptome sequencing (RNA-seq) technology. Our results showed that some basal biological processes like DNA replication and cell division are downregulated in the mature biofilm. Additionally, we identified significant expression reprogramming for genes involved in motility, sugar/lipid metabolism, and iron scavenging, as well as for outer membrane-encoding genes. A careful manual annotation process allowed us to assign molecular functions to many previously uncharacterized genes that are probably involved in biofilm metabolism. We also provided evidence for the presence of small regulatory RNAs in this species. Finally, coexpression networks were reconstructed to pinpoint functionally related gene clusters that may explain how biofilm maintenance is regulated. Beyond elucidating some genetic aspects of biofilm formation, this work reveals a number of pathways whose functional dissection may impact our understanding of leptospiral biology, in particular how these organisms adapt to environmental changes. IMPORTANCE In this work, we describe the first transcriptome based on RNA-seq technology focused on studying transcriptional changes associated with biofilm growth

  8. Transcriptome Sequencing Reveals Wide Expression Reprogramming of Basal and Unknown Genes in Leptospira biflexa Biofilms

    PubMed Central

    Spangenberg, Lucía; Lopes Bastos, Bruno; Graña, Martín; Vasconcelos, Larissa; Almeida, Áurea; Greif, Gonzalo; Robello, Carlos; Ristow, Paula

    2016-01-01

    ABSTRACT The genus Leptospira is composed of pathogenic and saprophytic spirochetes. Pathogenic Leptospira is the etiological agent of leptospirosis, a globally spread neglected disease. A key ecological feature of some pathogenic species is their ability to survive both within and outside the host. For most leptospires, the ability to persist outside the host is associated with biofilm formation, a most important bacterial strategy to face and overcome hostile environmental conditions. The architecture and biochemistry of leptospiral biofilms are rather well understood; however, the genetic program underpinning biofilm formation remains mostly unknown. In this work, we used the saprophyte Leptospira biflexa as a model organism to assess over- and underrepresented transcripts during the biofilm state, using transcriptome sequencing (RNA-seq) technology. Our results showed that some basal biological processes like DNA replication and cell division are downregulated in the mature biofilm. Additionally, we identified significant expression reprogramming for genes involved in motility, sugar/lipid metabolism, and iron scavenging, as well as for outer membrane-encoding genes. A careful manual annotation process allowed us to assign molecular functions to many previously uncharacterized genes that are probably involved in biofilm metabolism. We also provided evidence for the presence of small regulatory RNAs in this species. Finally, coexpression networks were reconstructed to pinpoint functionally related gene clusters that may explain how biofilm maintenance is regulated. Beyond elucidating some genetic aspects of biofilm formation, this work reveals a number of pathways whose functional dissection may impact our understanding of leptospiral biology, in particular how these organisms adapt to environmental changes. IMPORTANCE In this work, we describe the first transcriptome based on RNA-seq technology focused on studying transcriptional changes associated with biofilm

  9. Transcriptome Sequencing Reveals Wide Expression Reprogramming of Basal and Unknown Genes in Leptospira biflexa Biofilms.

    PubMed

    Iraola, Gregorio; Spangenberg, Lucía; Lopes Bastos, Bruno; Graña, Martín; Vasconcelos, Larissa; Almeida, Áurea; Greif, Gonzalo; Robello, Carlos; Ristow, Paula; Naya, Hugo

    2016-01-01

    The genus Leptospira is composed of pathogenic and saprophytic spirochetes. Pathogenic Leptospira is the etiological agent of leptospirosis, a globally spread neglected disease. A key ecological feature of some pathogenic species is their ability to survive both within and outside the host. For most leptospires, the ability to persist outside the host is associated with biofilm formation, a most important bacterial strategy to face and overcome hostile environmental conditions. The architecture and biochemistry of leptospiral biofilms are rather well understood; however, the genetic program underpinning biofilm formation remains mostly unknown. In this work, we used the saprophyte Leptospira biflexa as a model organism to assess over- and underrepresented transcripts during the biofilm state, using transcriptome sequencing (RNA-seq) technology. Our results showed that some basal biological processes like DNA replication and cell division are downregulated in the mature biofilm. Additionally, we identified significant expression reprogramming for genes involved in motility, sugar/lipid metabolism, and iron scavenging, as well as for outer membrane-encoding genes. A careful manual annotation process allowed us to assign molecular functions to many previously uncharacterized genes that are probably involved in biofilm metabolism. We also provided evidence for the presence of small regulatory RNAs in this species. Finally, coexpression networks were reconstructed to pinpoint functionally related gene clusters that may explain how biofilm maintenance is regulated. Beyond elucidating some genetic aspects of biofilm formation, this work reveals a number of pathways whose functional dissection may impact our understanding of leptospiral biology, in particular how these organisms adapt to environmental changes. IMPORTANCE In this work, we describe the first transcriptome based on RNA-seq technology focused on studying transcriptional changes associated with biofilm growth

  10. Household Characteristics Associated with Rodent Presence and Leptospira Infection in Rural and Urban Communities from Southern Chile

    PubMed Central

    Muñoz-Zanzi, Claudia; Mason, Meghan; Encina, Carolina; Gonzalez, Marcelo; Berg, Sergey

    2014-01-01

    Rodents are well-recognized reservoirs of Leptospira, contributing to its maintenance in endemic areas and playing a role in the public health risk associated with the infection. This study sought to provide some insights into rodent populations from Chile and their Leptospira carriage. In total, 393 rodents were trapped in 177 households. Higher rodent counts were associated with year 2 of the study, rainfall, and number of rodent signs. There was an inverse correlation with the number of cats. The number of rodents was higher in villages compared with slums (rate ratio = 3.23) but modified by average household age. Eighty rodents (20.4%) tested positive for Leptospira: 19.7% on the farms, 25.9% in villages, and 12.3% in the slums. Prevalence was 22.5% in Mus musculus, 20.7% in Rattus rattus, 21.1% in wild rodents, and 10.3% in R. norvegicus. Seasonal and temporal effects were the major determinants of Leptospira infection in rodent populations. PMID:24445209

  11. Spatial characterization of Leptospira spp. infection in equids from the Brejo Paraibano micro-region in Brazil.

    PubMed

    Oliveira Filho, Ruy Brayner; Campos Malta, Karla; Assis Santana, Vania Lucia; Vance Harrop, Mabel Hanna; Tancler Stipp, Danilo; Friguglietti Brandespim, Daniel; Aparecido Mota, Rinaldo; Wilton Pinheiro Júnior, José

    2014-05-01

    The present study, the first to spatially characterize Leptospira spp. infection among equids in the Brejo Paraibano micro-region of the Paraiba state in the northeast of Brazil, investigated 257 animals in 26 farms properties. Serum samples from 204 horses, 46 mules and seven donkeys were serologically diagnosed using the microscopic agglutination test (MAT). The distribution of Leptospira spp. was studied by employing specific antigens from 24 different Leptospira serovars. All farms were georeferenced and their distribution visualised on a map of the Brejo Paraibano micro-region. In addition, rainfall data were obtained from the same year, in which the sampling was performed. Among the 20 farms found to harbour animals with leptospirosis, 14 (70%) exhibited low prevalence, five (25%) medium prevalence and one (5%), high prevalence. Certain areas had a higher density of infected farms and required intervention to control the infection. Many serovars were widely distributed, while others were more common in particular areas. There was no significant association between the prevalence of Leptospira spp. infection and rainfall.

  12. Exposure of free-ranging wild carnivores, horses and domestic dogs to Leptospira spp in the northern Pantanal, Brazil.

    PubMed

    Jorge, Rodrigo Silva Pinto; Ferreira, Fernando; Ferreira Neto, José Soares; Vasconcellos, Silvio de Arruda; Lima, Edson de Souza; Morais, Zenaide Maria de; Souza, Gisele Oliveira de

    2011-06-01

    Leptospirosis is a zoonotic disease affecting most mammals and is distributed throughout the world. Several species of domestic and wild animals may act as reservoirs for this disease. The purpose of this study was to assess the exposure of free-ranging wild carnivores, horses and domestic dogs on a private reserve located in the northern Pantanal (Brazil) and the surrounding areas to Leptospira spp from 2002-2006, 75 free-ranging wild carnivores were captured in the Pantanal and serum samples were collected. In addition, samples from 103 domestic dogs and 23 horses in the region were collected. Serum samples were tested for the presence of Leptospira antibodies using the microscopic agglutination test. Thirty-two wild carnivores (42.7%) were considered positive with titres ≥ 100, and 18 domestic dogs (17.5%) and 20 horses (74.1%) were also found to be positive. Our study showed that horses, dogs and several species of free-ranging wild carnivores have been exposed to Leptospira spp in the Pantanal, suggesting that the peculiar characteristics of this biome, such as high temperatures and an extended period of flooding, may favour bacterial persistence and transmission. In this region, wild carnivores and horses seem to be important hosts for the epidemiology of Leptospira species.

  13. Plasminogen Binding Proteins and Plasmin Generation on the Surface of Leptospira spp.: The Contribution to the Bacteria-Host Interactions

    PubMed Central

    Vieira, Monica L.; Atzingen, Marina V.; Oliveira, Rosane; Mendes, Renata S.; Domingos, Renan F.; Vasconcellos, Silvio A.; Nascimento, Ana L. T. O.

    2012-01-01

    Leptospirosis is considered a neglected infectious disease of human and veterinary concern. Although extensive investigations on host-pathogen interactions have been pursued by several research groups, mechanisms of infection, invasion and persistence of pathogenic Leptospira spp. remain to be elucidated. We have reported the ability of leptospires to bind human plasminogen (PLG) and to generate enzimatically active plasmin (PLA) on the bacteria surface. PLA-coated Leptospira can degrade immobilized ECM molecules, an activity with implications in host tissue penetration. Moreover, we have identified and characterized several proteins that may act as PLG-binding receptors, each of them competent to generate active plasmin. The PLA activity associated to the outer surface of Leptospira could hamper the host immune attack by conferring the bacteria some benefit during infection. The PLA-coated leptospires obstruct complement C3b and IgG depositions on the bacterial surface, most probably through degradation. The decrease of leptospiral opsonization might be an important aspect of the immune evasion strategy. We believe that the presence of PLA on the leptospiral surface may (i) facilitate host tissue penetration, (ii) help the bacteria to evade the immune system and, as a consequence, (iii) permit Leptospira to reach secondary sites of infection. PMID:23118516

  14. Prevalence of Leptospira antibodies in wild boars (Sus scrofa) from Northern Portugal: risk factor analysis.

    PubMed

    Vale-Gonçalves, H M; Cabral, J A; Faria, M C; Nunes-Pereira, M; Faria, A S; Veloso, O; Vieira, M L; Paiva-Cardoso, Md N

    2015-07-01

    Leptospirosis is a zoonosis of worldwide distribution, caused by infection with pathogenic spirochaetes of the genus Leptospira. The wild boar (Sus scrofa), an important hunting species in Europe, seems to play a significant role in the epidemiological cycle of leptospirosis. A total of 101 serum samples from wild boar hunted in Northern Portugal were analysed for leptospiral antibodies detection by microscopic agglutination test. Sera were collected during hunting seasons (2011-2013) and tested with 17 different pathogenic serovars of Leptospira. Antibodies against nine serovars were detected in 66 (65·4%) of these sera. Serovars Tarassovi and Altodouro exhibited the highest seroreactivity rates (23·8% and 16·8%, respectively), followed by Autumnalis (7·9%) and Bratislava (6·9%). Age and district of origin were found to be risk factors for the presence of leptospiral antibodies in contrast to gender. From a One Health perspective, this study revealed that wild boar should be considered as a potential source of leptospirosis dissemination for humans and animal species (domestic and wild) in shared environments, particularly in the Trás-os-Montes region.

  15. [Exposure to Leptospira in stray dogs in the city of Cali].

    PubMed

    Rodríguez, Ana Lucía; Ferro, Beatriz Eugenia; Varona, María Ximena; Santafé, Mauricio

    2004-09-01

    In Colombia, little information is available concerning the epidemiology of leptospirosis in urban environments. Furthermore, the role of dogs in the transmission cycle of leptospirosis in the urban setting is unclear. To explore the potential role of canines in the transmission of leptospirosis in Cali, a serological study was conducted with 197 serum samples collected from stray dogs during 2001 and 2003. Serum specimens were screened with the Microscopic Agglutination Test (MAT) and 7 serovars--Icterohaemorrhagiae, Canicola, Gryppotyphosa, Hardjo strain Hardjobovis, Pomona, Hardjo strain Hardjoprajitno and Bratislava. All serovars were provided by the Instituto Colombiano Agropecuario (ICA), Tuluá, Colombia. The MAT was considered positive when 50% or more leptospiras were agglutinated with one or more serovars in a serum dilution of 1:100. At least one serovar showed evidence of infection in 41.1% of the dogs. The most prevalent serovar was Icterohaemorrhagiae, found in 55.6% of the seropositive dogs. 48.1% were co-agglutinations. No reactions against the serovars Pomona, Hardjo strain Hardjoprajitno and Bratislava were observed. These findings suggested that stray dogs are potential reservoirs of Leptospira in Cali and underscored the need to study the epidemiology of this disease in Colombia.

  16. First evidence for a restriction-modification system in Leptospira sp.

    PubMed

    Brenot, A; Werts, C; Ottone, C; Sertour, N; Charon, N W; Postic, D; Baranton, G; Saint Girons, I

    2001-07-24

    The LE1 leptophage exhibited a host range restricted to the saprophytic Leptospira biflexa [Saint Girons et al., Res. Microbiol. 141 (1990) 1131-1133] and mainly to the Patoc 1 strain (hereafter called PFRA) kept in the Paris, France collection. Results of titration of LE1 lysates indicated the presence of a host-controlled modification and restriction system within PUSA (Patoc 1 strain maintained in the Morgantown, WV, USA collection) that was absent in PFRA. Because genomic DNA of PITAL (Patoc 1 strain maintained in Trieste, Italy) appeared smeared in pulsed field gel electrophoresis (PFGE), this strain is likely to contain nucleases that are activated upon DNA isolation. Moreover, comparative NotI digestions of PUSA and PFRA DNAs, as visualized by PFGE, indicated that PUSA belonged to a different serovar than PFRA. Finally, 16S ribosomal sequence analysis indicated that PUSA belonged to the saprophytic Leptospira meyeri species, while PITAL and PFRA appertained to L. biflexa. The evolutionary significance and the importance of the restriction and modification enzymes or non-specific nucleases within strains for genetic experiments are discussed.

  17. Detection of wild animals as carriers of Leptospira by PCR in the Pantanal biome, Brazil.

    PubMed

    Vieira, Anahi S; Narduche, Lorena; Martins, Gabriel; Schabib Péres, Igor A H F; Zimmermann, Namor P; Juliano, Raquel S; Pellegrin, Aiesca O; Lilenbaum, Walter

    2016-11-01

    Leptospiral infection is widespread in wildlife. In this context, wild ecosystems in tropical countries hold a vast biodiversity, including several species that may act as potential reservoirs of leptospires. The Pantanal biome presents highly favorable environmental conditions for the occurrence of leptospirosis, such as high temperatures, constant flooding, and high biodiversity. The purpose of this study was to detect wild animals as carriers of Leptospira sp. using direct methods (PCR and culture) in the Pantanal biome, Brazil. A total of 35 animals were studied, namely Cerdocyon thous, Nasua nasua, Ozotoceros bezoarticus, and Sus scrofa species. Blood for serology (MAT) and urine for bacteriological culturing and PCR was sampled. The most prevalent serogroups were Javanica and Djasiman. Additionally, 40.6% of these animals presented PCR positive reactions. Seroreactivity associated with the high frequency of leptospiral carriers among the different studied species suggests a high level of exposure of the studied animals to pathogenic Leptospira strains. Our results are still limited and the actual role of the studied animals in the epidemiology of leptospirosis in the Pantanal region remains to be elucidated. PMID:27496621

  18. Antimicrobial susceptibility of Leptospira isolates from dogs and rats to 12 antimicrobial agents.

    PubMed

    Suepaul, S M; Carrington, C; Campbell, M; Borde, G; Adesiyun, A A

    2015-03-01

    This study determined the antimicrobial susceptibilities of 67 isolates of Leptospira from dogs (suspect canine cases: n=7 and stray dogs: n=6) and rodents (n=54) in Trinidad to 12 antimicrobial agents using broth microdilution and macrodilution techniques. Commonly used antimicrobial agents such as the penicillin G and ceftriaxone had relatively low minimal inhibitory concentrations (MICs) while doxycycline displayed a relatively higher value but was still considered to be effective. While imipenem was the most effective with low MIC values in vitro, sulphamethoxazole-trimethoprim had the highest i.e. least effective. Based on these results, the drugs commonly used in the treatment of leptospirosis (penicillin G, penicillin-streptomycin, doxycycline and ceftriaxone) in both humans and animals in Trinidad appear to have similar MICs and MBCs in vitro when compared with published reports. The serovar of Leptospira spp. and in most cases the origin of the isolates did not significantly (P>0.05) influence their susceptibilities to the antimicrobial agents tested. PMID:25801249

  19. Detection of anti-Leptospira antibodies in captive nonhuman primates from Salvador, Brazil.

    PubMed

    Pinna, Melissa H; Martins, Gabriel; Pinheiro, Ana Carla O; Almeida, Daniela S; Oriá, Arianne P; Lilenbaum, Walter

    2012-01-01

    Leptospirosis is a widely distributed zoonosis that affects several species of domestic and wild animals. Under captive conditions, Leptospirosis is a potential problem because the physical conditions in most zoos and research centers cannot prevent the captive animals from being exposed to rodents, raccoons, opossums, and other local wildlife that are known carriers. Yet, despite the potential risk, animals that are destined for reintroduction into the wild are not routinely tested for anti-Leptospira antibodies before their release. The purpose of this study was to determine the occurrence of anti-Leptospira antibodies in captive New World monkeys that were housed in the Wild Animals Screening Center in Salvador, Brazil. Blood samples were collected from 44 monkeys (28 Callithrix jacchus, eight Callithrix pennicilata, and eight Cebus sp.). The animals were screened for antibodies with the microscopic agglutination test. Twenty-five (56.8%) primates were seroreactive, with Icterohaemorrhagiae being the most frequent serogroup. None of the monkeys, however, presented clinical signs of leptospirosis. Thus, seroreactivity with low titers in asymptomatic animals, as observed in this study, suggests exposure to the agent. The unexpected predominance of the serogroup Icterohaemorrhagiae further suggests that exposure to this serogroup occurred in captivity. Therefore, the dangerous possibility cannot be ignored that reintroduced monkeys will carry the leptospiral serovars into wild populations. In conclusion, primates exposed to urban serovars before their release from captivity represent a potentially significant health risk to wild populations.

  20. Detection of wild animals as carriers of Leptospira by PCR in the Pantanal biome, Brazil.

    PubMed

    Vieira, Anahi S; Narduche, Lorena; Martins, Gabriel; Schabib Péres, Igor A H F; Zimmermann, Namor P; Juliano, Raquel S; Pellegrin, Aiesca O; Lilenbaum, Walter

    2016-11-01

    Leptospiral infection is widespread in wildlife. In this context, wild ecosystems in tropical countries hold a vast biodiversity, including several species that may act as potential reservoirs of leptospires. The Pantanal biome presents highly favorable environmental conditions for the occurrence of leptospirosis, such as high temperatures, constant flooding, and high biodiversity. The purpose of this study was to detect wild animals as carriers of Leptospira sp. using direct methods (PCR and culture) in the Pantanal biome, Brazil. A total of 35 animals were studied, namely Cerdocyon thous, Nasua nasua, Ozotoceros bezoarticus, and Sus scrofa species. Blood for serology (MAT) and urine for bacteriological culturing and PCR was sampled. The most prevalent serogroups were Javanica and Djasiman. Additionally, 40.6% of these animals presented PCR positive reactions. Seroreactivity associated with the high frequency of leptospiral carriers among the different studied species suggests a high level of exposure of the studied animals to pathogenic Leptospira strains. Our results are still limited and the actual role of the studied animals in the epidemiology of leptospirosis in the Pantanal region remains to be elucidated.

  1. Development of a recombinase polymerase amplification assay for the detection of pathogenic Leptospira.

    PubMed

    Ahmed, Ahmed; van der Linden, Hans; Hartskeerl, Rudy A

    2014-05-08

    Detection of leptospires based on DNA amplification techniques is essential for the early diagnosis of leptospirosis when anti-Leptospira antibodies are below the detection limit of most serological tests. In middle and low income countries where leptospirosis is endemic, routine implementation of real-time PCR is financially and technically challenging due to the requirement of expensive thermocycler equipment. In this study we report the development and evaluation of a novel isothermal recombinase polymerase amplification assay (RPA) for detection of pathogenic Leptospira based on TwistAmp chemistry. RPA enabled the detection of less than two genome copies per reaction. Retrospective evaluation revealed a high diagnostic accuracy (sensitivity and specificity of 94.7% and 97.7%, respectively) compared to culturing as the reference standard. RPA presents a powerful tool for the early diagnosis of leptospirosis in humans and in animals. Furthermore, it enables the detection of the causative agent in reservoirs and environment, and as such is a valuable adjunct to current tools for surveillance and early outbreak warning.

  2. Serological investigation of Leptospira infection and its circulation in one intensive-type water buffalo farm in the Philippines.

    PubMed

    Villanueva, Marvin A; Mingala, Claro N; Gloriani, Nina G; Yanagihara, Yasutake; Isoda, Norikazu; Nakajima, Chie; Suzuki, Yasuhiko; Koizumi, Nobuo

    2016-02-01

    Water buffalo is an indispensable livestock in the Philippines. Leptospirosis is a serious zoonosis that can be fatal to humans and cause reproductive problems in livestock. Leptospirosis has been reported in some countries where water buffaloes are commercially raised, highlighting the Leptospira prevalence in this farming system, but information on leptospirosis in water buffalo farms in the Philippines is limited. In this study, we collected blood samples from rats (n = 21), and water buffaloes (n = 170) from different groups and locations in one intensive-type buffalo farm in the Philippines. Serum was analyzed by microscopic agglutination test (MAT). Anti-Leptospira antibodies reacting with serogroups Canicola, Icterohaemorrhagiae and Pomona were found in sera of 30% tested rats, and 48% of water buffalo sera tested positive for at least one Leptospira strain, in which serogroups Mini, Hebdomadis, Tarassovi and Pyrogenes were predominantly agglutinated. The number of seropositive young water buffaloes (< 1 year-old) was lower than that of older seropositive ones. Furthermore, sera from younger water buffaloes were reactive with single serotypes with low MAT titers, but older animals were reactive with multiple Leptospira strains with variable MAT titers. In addition, antibodies against serogroups Icterohaemorrhagiae and Pomona were detected in both animals. Finally, Leptospira infection was found associated with age and animal grouping, highlighting the impact of management in the persistence of leptospirosis at intensive-type buffalo farm settings in the Philippines. Further investigation and appropriate control strategies are required to prevent leptospirosis from causing risks to public health and economic losses to the water buffalo farming industry. PMID:27348885

  3. Serological investigation of Leptospira infection and its circulation in one intensive-type water buffalo farm in the Philippines.

    PubMed

    Villanueva, Marvin A; Mingala, Claro N; Gloriani, Nina G; Yanagihara, Yasutake; Isoda, Norikazu; Nakajima, Chie; Suzuki, Yasuhiko; Koizumi, Nobuo

    2016-02-01

    Water buffalo is an indispensable livestock in the Philippines. Leptospirosis is a serious zoonosis that can be fatal to humans and cause reproductive problems in livestock. Leptospirosis has been reported in some countries where water buffaloes are commercially raised, highlighting the Leptospira prevalence in this farming system, but information on leptospirosis in water buffalo farms in the Philippines is limited. In this study, we collected blood samples from rats (n = 21), and water buffaloes (n = 170) from different groups and locations in one intensive-type buffalo farm in the Philippines. Serum was analyzed by microscopic agglutination test (MAT). Anti-Leptospira antibodies reacting with serogroups Canicola, Icterohaemorrhagiae and Pomona were found in sera of 30% tested rats, and 48% of water buffalo sera tested positive for at least one Leptospira strain, in which serogroups Mini, Hebdomadis, Tarassovi and Pyrogenes were predominantly agglutinated. The number of seropositive young water buffaloes (< 1 year-old) was lower than that of older seropositive ones. Furthermore, sera from younger water buffaloes were reactive with single serotypes with low MAT titers, but older animals were reactive with multiple Leptospira strains with variable MAT titers. In addition, antibodies against serogroups Icterohaemorrhagiae and Pomona were detected in both animals. Finally, Leptospira infection was found associated with age and animal grouping, highlighting the impact of management in the persistence of leptospirosis at intensive-type buffalo farm settings in the Philippines. Further investigation and appropriate control strategies are required to prevent leptospirosis from causing risks to public health and economic losses to the water buffalo farming industry.

  4. Identification of Tenrec ecaudatus, a Wild Mammal Introduced to Mayotte Island, as a Reservoir of the Newly Identified Human Pathogenic Leptospira mayottensis.

    PubMed

    Lagadec, Erwan; Gomard, Yann; Le Minter, Gildas; Cordonin, Colette; Cardinale, Eric; Ramasindrazana, Beza; Dietrich, Muriel; Goodman, Steven M; Tortosa, Pablo; Dellagi, Koussay

    2016-08-01

    Leptospirosis is a bacterial zoonosis of major concern on tropical islands. Human populations on western Indian Ocean islands are strongly affected by the disease although each archipelago shows contrasting epidemiology. For instance, Mayotte, part of the Comoros Archipelago, differs from the other neighbouring islands by a high diversity of Leptospira species infecting humans that includes Leptospira mayottensis, a species thought to be unique to this island. Using bacterial culture, molecular detection and typing, the present study explored the wild and domestic local mammalian fauna for renal carriage of leptospires and addressed the genetic relationships of the infecting strains with local isolates obtained from acute human cases and with Leptospira strains hosted by mammal species endemic to nearby Madagascar. Tenrec (Tenrec ecaudatus, Family Tenrecidae), a terrestrial mammal introduced from Madagascar, is identified as a reservoir of L. mayottensis. All isolated L. mayottensis sequence types form a monophyletic clade that includes Leptospira strains infecting humans and tenrecs on Mayotte, as well as two other Malagasy endemic tenrecid species of the genus Microgale. The lower diversity of L. mayottensis in tenrecs from Mayotte, compared to that occurring in Madagascar, suggests that L. mayottensis has indeed a Malagasy origin. This study also showed that introduced rats (Rattus rattus) and dogs are probably the main reservoirs of Leptospira borgpetersenii and Leptospira kirschneri, both bacteria being prevalent in local clinical cases. Data emphasize the epidemiological link between the two neighbouring islands and the role of introduced small mammals in shaping the local epidemiology of leptospirosis.

  5. Identification of Tenrec ecaudatus, a Wild Mammal Introduced to Mayotte Island, as a Reservoir of the Newly Identified Human Pathogenic Leptospira mayottensis

    PubMed Central

    Lagadec, Erwan; Gomard, Yann; Le Minter, Gildas; Cordonin, Colette; Cardinale, Eric; Ramasindrazana, Beza; Dietrich, Muriel; Goodman, Steven M; Tortosa, Pablo; Dellagi, Koussay

    2016-01-01

    Leptospirosis is a bacterial zoonosis of major concern on tropical islands. Human populations on western Indian Ocean islands are strongly affected by the disease although each archipelago shows contrasting epidemiology. For instance, Mayotte, part of the Comoros Archipelago, differs from the other neighbouring islands by a high diversity of Leptospira species infecting humans that includes Leptospira mayottensis, a species thought to be unique to this island. Using bacterial culture, molecular detection and typing, the present study explored the wild and domestic local mammalian fauna for renal carriage of leptospires and addressed the genetic relationships of the infecting strains with local isolates obtained from acute human cases and with Leptospira strains hosted by mammal species endemic to nearby Madagascar. Tenrec (Tenrec ecaudatus, Family Tenrecidae), a terrestrial mammal introduced from Madagascar, is identified as a reservoir of L. mayottensis. All isolated L. mayottensis sequence types form a monophyletic clade that includes Leptospira strains infecting humans and tenrecs on Mayotte, as well as two other Malagasy endemic tenrecid species of the genus Microgale. The lower diversity of L. mayottensis in tenrecs from Mayotte, compared to that occurring in Madagascar, suggests that L. mayottensis has indeed a Malagasy origin. This study also showed that introduced rats (Rattus rattus) and dogs are probably the main reservoirs of Leptospira borgpetersenii and Leptospira kirschneri, both bacteria being prevalent in local clinical cases. Data emphasize the epidemiological link between the two neighbouring islands and the role of introduced small mammals in shaping the local epidemiology of leptospirosis. PMID:27574792

  6. Identification of Tenrec ecaudatus, a Wild Mammal Introduced to Mayotte Island, as a Reservoir of the Newly Identified Human Pathogenic Leptospira mayottensis.

    PubMed

    Lagadec, Erwan; Gomard, Yann; Le Minter, Gildas; Cordonin, Colette; Cardinale, Eric; Ramasindrazana, Beza; Dietrich, Muriel; Goodman, Steven M; Tortosa, Pablo; Dellagi, Koussay

    2016-08-01

    Leptospirosis is a bacterial zoonosis of major concern on tropical islands. Human populations on western Indian Ocean islands are strongly affected by the disease although each archipelago shows contrasting epidemiology. For instance, Mayotte, part of the Comoros Archipelago, differs from the other neighbouring islands by a high diversity of Leptospira species infecting humans that includes Leptospira mayottensis, a species thought to be unique to this island. Using bacterial culture, molecular detection and typing, the present study explored the wild and domestic local mammalian fauna for renal carriage of leptospires and addressed the genetic relationships of the infecting strains with local isolates obtained from acute human cases and with Leptospira strains hosted by mammal species endemic to nearby Madagascar. Tenrec (Tenrec ecaudatus, Family Tenrecidae), a terrestrial mammal introduced from Madagascar, is identified as a reservoir of L. mayottensis. All isolated L. mayottensis sequence types form a monophyletic clade that includes Leptospira strains infecting humans and tenrecs on Mayotte, as well as two other Malagasy endemic tenrecid species of the genus Microgale. The lower diversity of L. mayottensis in tenrecs from Mayotte, compared to that occurring in Madagascar, suggests that L. mayottensis has indeed a Malagasy origin. This study also showed that introduced rats (Rattus rattus) and dogs are probably the main reservoirs of Leptospira borgpetersenii and Leptospira kirschneri, both bacteria being prevalent in local clinical cases. Data emphasize the epidemiological link between the two neighbouring islands and the role of introduced small mammals in shaping the local epidemiology of leptospirosis. PMID:27574792

  7. Circulating serovars of Leptospira in cart horses of central and southern Ethiopia and associated risk factors.

    PubMed

    Tsegay, K; Potts, A D; Aklilu, N; Lötter, C; Gummow, B

    2016-03-01

    Little work has been done on diseases of horses in Ethiopia or tropical regions of the world. Yet, Ethiopia has the largest horse population in Africa and their horses play a pivotal role in their economy as traction animals. A serological and questionnaire survey was therefore conducted to determine the circulating serovars of Leptospira and their association with potential risk factors in the cart horse population of Central and Southern Ethiopia. A total of 184 out of 418 cart horses from 13 districts had antibody titres of 1:100 or greater to at least one of 16 serovars of Leptospira species in Central and Southern Ethiopian horses. A significantly higher seropositivity (62.1%) was noted in horses from the highland agroecology followed by midland (44.4%) and lowland (39.8%). Serovar Bratislava (34.5%) was the predominant serovar followed by serovars Djasiman (9.8%), Topaz (5.98%) and Pomona (5.3%). Age and location proved to be associated with seropositive horses with older horses being more commonly affected and the districts of Ziway (Batu) (Apparent Prevalence (AP)=65.5%), Shashemene (AP=48.3%) and Sebeta (AP=41.4%) having the highest prevalence. Multivariable logistic regression found risk factors significantly associated with Leptospira seropositive horses were drinking river water (OR=2.8) and horses 7-12 years old (OR=5) and risk factors specifically associated with serovar Bratislava seropositive horses were drinking river water (OR=2.5), horses ≥13 years (OR=3.5) and the presence of dogs in adjacent neighbouring properties (OR=0.3). Dogs had a protective effect against seropositivity to serovars Bratislava and Djasiman, which may be due to their ability to control rodents. The high seroprevalence confirm that leptospirosis is endemic among horses of Central and Southern Ethiopia. The predominance of serovar Bratislava supports the idea that serovar Bratislava may be adapted to and maintained by the horse population of Central and Southern Ethiopia

  8. Leptospira spp. in Domestic Cats from Different Environments: Prevalence of Antibodies and Risk Factors Associated with the Seropositivity

    PubMed Central

    Azócar-Aedo, Lucía; Monti, Gustavo; Jara, Ronald

    2014-01-01

    Simple Summary Although Leptospira infection occurs in domestic cat populations, studies on leptospirosis are very limited in felines and the role of cats in the epidemiology of this zoonosis has not received much attention. The present work is an epidemiologic study intended to determine the prevalence of anti-Leptospira antibodies and risk factors related with the seropositivity in cats from urban and rural environments. A higher prevalence in rural cats was detected (25.2%) compared with urban animals (1.8%). Characteristics of the habitat of the animals and some agricultural activities performed by cat’s owners were found to be risk factors associated with the seropositivity. Abstract Leptospirosis is an emerging zoonotic disease of worldwide distribution. A cross-sectional study was conducted in urban and rural environments in southern Chile (1) to detect domestic cats with serologic evidence of exposure to Leptospira spp.; (2) to determine the prevalence of anti-Leptospira antibodies; (3) to describe seroprevalences according to different characteristics of the animals, and (4) to identify risk factors associated with the seropositivity in the Microscopic Agglutination Test (MAT). Blood samples were taken from 124 owned cats. A frequentist and Bayesian approach were applied for prevalence estimation. The overall apparent prevalence of anti-Leptospira antibodies was 8.1% (95% Confident Interval = 3.9–4.3). With the Bayesian approach, the overall True Prevalence (TP) was 5.2% (95% Credibility Interval (CrI) = 0.6–12.4). The TP for urban cats was 1.8% (95% CrI = 0.1–7.2) and the TP for rural felines was 25.2% (95% CrI = 9.3–46.6). Cats that live in a place where agricultural activities are performed with water that flows in streams or backwater and cats that live in places near flooded areas had a higher risk of seropositivity in MAT. The exposure to Leptospira spp. in domestic cats of urban and rural origin in Southern Chile is a public health concern

  9. Reverse-Transcriptase PCR Detection of Leptospira: Absence of Agreement with Single-Specimen Microscopic Agglutination Testing

    PubMed Central

    Waggoner, Jesse J.; Balassiano, Ilana; Mohamed-Hadley, Alisha; Vital-Brazil, Juliana Magalhães; Sahoo, Malaya K.; Pinsky, Benjamin A.

    2015-01-01

    Background Reference diagnostic tests for leptospirosis include nucleic acid amplification tests, bacterial culture, and microscopic agglutination testing (MAT) of acute and convalescent serum. However, clinical laboratories often do not receive paired specimens. In the current study, we tested serum samples using a highly sensitive real-time nucleic acid amplification test for Leptospira and compared results to MAT performed on the same specimens. Methods/Principal Findings 478 serum samples from suspected leptospirosis cases in Rio de Janeiro were tested using a real-time RT-PCR for the diagnosis of leptospirosis, malaria and dengue (the Lepto-MD assay). The Lepto-MD assay detects all species of Leptospira (saprophytic, intermediate, and pathogenic), and in the current study, we demonstrate that this assay amplifies both Leptospira RNA and DNA. Dengue virus RNA was identified in 10 patients, and no cases of malaria were detected. A total of 65 samples (13.6%) were positive for Leptospira: 35 samples (7.3%) in the Lepto-MD assay, 33 samples (6.9%) by MAT, and 3 samples tested positive by both (kappa statistic 0.02). Poor agreement between methods was consistent regardless of the titer used to define positive MAT results or the day of disease at sample collection. Leptospira nucleic acids were detected in the Lepto-MD assay as late as day 22, and cycle threshold values did not differ based on the day of disease. When Lepto-MD assay results were added to the MAT results for all patients in 2008 (n=818), the number of detected leptospirosis cases increased by 30.4%, from 102 (12.5%) to 133 (16.3%). Conclusions/Significance This study demonstrates a lack of agreement between nucleic acid detection of Leptospira and single-specimen MAT, which may result from the clearance of bacteremia coinciding with the appearance of agglutinating antibodies. A combined testing strategy for acute leptospirosis, including molecular and serologic testing, appears necessary to maximize

  10. Anti-Leptospira spp. antibodies in Crotalus durissus collilineatus kept in captivity and its zoonotic relevance.

    PubMed

    Rodrigues, T C S; Santos, A L Q; Lima, A M C; Gomes, D O; Brites, V L C

    2016-06-01

    Leptospirosis is a worldwide spread zoonosis that can affect all groups of vertebrates, including reptiles. Because it has been little studied in snakes, this study focused on determining the occurrence of anti-Leptospira spp. antibodies in 64 Crotalus durissus collilineatus kept in captivity and on identifying the most common serovars in these animals, using the microscopic agglutination test. Of these, almost 90% were positive and there were reactions to the 22 serovars used in the study. The most common serovar in these snakes was Javanica, Andamana and Patoc. Most frequent titers were 25 and 50, although high titers (such as 1600) were also recorded, despite the absence of clinical symptoms. The possibility should be considered of captive snakes serving as a serious source of leptospiral infection in humans, which is why it is essential to study, prevent and control the disease in breeding centers and serpentariums. PMID:26875762

  11. Anti-Leptospira spp. antibodies in Crotalus durissus collilineatus kept in captivity and its zoonotic relevance.

    PubMed

    Rodrigues, T C S; Santos, A L Q; Lima, A M C; Gomes, D O; Brites, V L C

    2016-06-01

    Leptospirosis is a worldwide spread zoonosis that can affect all groups of vertebrates, including reptiles. Because it has been little studied in snakes, this study focused on determining the occurrence of anti-Leptospira spp. antibodies in 64 Crotalus durissus collilineatus kept in captivity and on identifying the most common serovars in these animals, using the microscopic agglutination test. Of these, almost 90% were positive and there were reactions to the 22 serovars used in the study. The most common serovar in these snakes was Javanica, Andamana and Patoc. Most frequent titers were 25 and 50, although high titers (such as 1600) were also recorded, despite the absence of clinical symptoms. The possibility should be considered of captive snakes serving as a serious source of leptospiral infection in humans, which is why it is essential to study, prevent and control the disease in breeding centers and serpentariums.

  12. Aseptic meningitis caused by Leptospira spp diagnosed by polymerase chain reaction.

    PubMed

    Romero, Eliete Caló; Blanco, Roberta Morozetti; Yasuda, Paulo Hideki

    2010-12-01

    Leptospirosis is a zoonotic disease caused by the pathogenic Leptospira spp. The clinical presentations are diverse, ranging from undifferentiated fever to fulminant disease including meningeal forms. The neurological leptospirosis forms are usually neglected. The aim of this study was to investigate leptospirosis as the cause of aseptic meningitis using different diagnostic techniques including the polymerase chain reaction (PCR). Thirty-nine cerebrospinal fluid (CSF) samples from patients presenting with meningeal abnormalities, predominance of lymphocytes and negative results by traditional microbiological tests were processed by leptospiral culture, anti-leptospiral antibody response and PCR. Leptospira spp DNA was detected in 23 (58.97%) of the CSF samples. Anti-leptospiral antibodies were found in 13 (33.33%) CSF samples. Twelve CSF samples were positive by PCR assay and negative by microscopic agglutination test (MAT) assay. Two CSF samples were positive by MAT and negative by PCR. The positive and negative agreement between both tests was 11 and 14, respectively. CSF samples from six cases of unknown diagnosis were positive by PCR assay. Eight cases showed positive results using PCR and MAT. Leptospirosis could be detected by PCR assay from the 3rd-26th day after illness onset. The sensitivity of the PCR was assessed with confirmed cases of leptospirosis (by MAT) and found to be 89.5%. All CSFs were negative by culture. PCR was found to be a powerful tool for diagnosing meningitis cases of leptospirosis. We recommend that it may be used as a supplementary diagnostic tool, especially in the early stages of the disease, when other diagnostic techniques such as serology are not sensitive.

  13. Detection of Leptospira spp. in wildlife reservoir hosts in Ontario through comparison of immunohistochemical and polymerase chain reaction genotyping methods.

    PubMed

    Shearer, Karen E; Harte, Michael J; Ojkic, Davor; Delay, Josepha; Campbell, Douglas

    2014-03-01

    A total of 460 kidney samples from wildlife (beavers, coyotes, deer, foxes, opossums, otters, raccoons, skunks) were obtained from road-kill and hunter/trapper donations in Ontario between January 2010 and November 2012. The objectives of the study were to detect Leptospira spp. by immunohistochemistry and polymerase chain reaction (PCR), to map presence of leptospires in wildlife relative to livestock and human populations, and to characterize positive samples by sequencing and comparison to leptospires known to affect domestic animals and humans. The proportion of samples that tested positive ranged from 0% to 42%, with the highest rates in skunks and raccoons. Leptospira spp. were present in kidneys of wildlife across Ontario, particularly in areas of high human density, and areas in which livestock populations are abundant. The PCR was too weak in most samples to permit genotyping and examination of the relationship between the leptospires found in this study and those affecting domestic animals and humans.

  14. Product-specific validation of a serological potency test for release of Leptospira vaccines in the European Union.

    PubMed

    Stirling, Catrina; Novokova, Viera

    2013-09-01

    Historically in the European Union, all Leptospira vaccines were released using the European Pharmacopoeia (Ph. Eur.) hamster potency assay. Recently, there has been a shift toward alternatives that offer either refinement of testing or replacement of animals for product release. This is being driven by animal welfare concerns but also by a drive to have more consistent, cheaper, and faster batch release tests. This publication discusses one such example of a multicomponent canine vaccine that includes three Leptospira serovars and has recently been registered in the European Union. The potency release test is a refinement because it uses rabbit serology rather than hamster challenge. This publication covers the principles of the test method, challenges faced during its development and registration, and discussion about benefits and limitations of this method. It concludes with a view of how the use of serology testing could fit into an overall strategy to move to fully in vitro testing by adopting a consistency approach.

  15. Product-specific validation of a serological potency test for release of Leptospira vaccines in the European Union.

    PubMed

    Stirling, Catrina; Novokova, Viera

    2013-09-01

    Historically in the European Union, all Leptospira vaccines were released using the European Pharmacopoeia (Ph. Eur.) hamster potency assay. Recently, there has been a shift toward alternatives that offer either refinement of testing or replacement of animals for product release. This is being driven by animal welfare concerns but also by a drive to have more consistent, cheaper, and faster batch release tests. This publication discusses one such example of a multicomponent canine vaccine that includes three Leptospira serovars and has recently been registered in the European Union. The potency release test is a refinement because it uses rabbit serology rather than hamster challenge. This publication covers the principles of the test method, challenges faced during its development and registration, and discussion about benefits and limitations of this method. It concludes with a view of how the use of serology testing could fit into an overall strategy to move to fully in vitro testing by adopting a consistency approach. PMID:23849308

  16. Detection of Leptospira spp. in wildlife reservoir hosts in Ontario through comparison of immunohistochemical and polymerase chain reaction genotyping methods

    PubMed Central

    Shearer, Karen E.; Harte, Michael J.; Ojkic, Davor; DeLay, Josepha; Campbell, Douglas

    2014-01-01

    A total of 460 kidney samples from wildlife (beavers, coyotes, deer, foxes, opossums, otters, raccoons, skunks) were obtained from road-kill and hunter/trapper donations in Ontario between January 2010 and November 2012. The objectives of the study were to detect Leptospira spp. by immunohistochemistry and polymerase chain reaction (PCR), to map presence of leptospires in wildlife relative to livestock and human populations, and to characterize positive samples by sequencing and comparison to leptospires known to affect domestic animals and humans. The proportion of samples that tested positive ranged from 0% to 42%, with the highest rates in skunks and raccoons. Leptospira spp. were present in kidneys of wildlife across Ontario, particularly in areas of high human density, and areas in which livestock populations are abundant. The PCR was too weak in most samples to permit genotyping and examination of the relationship between the leptospires found in this study and those affecting domestic animals and humans. PMID:24587507

  17. Detection of Leptospira-Specific Antibodies Using a Recombinant Antigen-Based Enzyme-Linked Immunosorbent Assay

    PubMed Central

    Chen, Hua-Wei; Zhang, Zhiwen; Halsey, Eric S.; Guevara, Carolina; Canal, Enrique; Hall, Eric; Maves, Ryan; Tilley, Drake H.; Kochel, Tadeusz J.; Ching, Wei-Mei

    2013-01-01

    We produced three highly purified recombinant antigens rLipL32, rLipL41, and rLigA-Rep (leptospiral immunoglobulin-like A repeat region) for the detection of Leptospira-specific antibodies in an enzyme-linked immunosorbent assay (ELISA). The performance of these recombinant antigens was evaluated using 121 human sera. Among them, 63 sera were microscopic agglutination test (MAT)-confirmed positive sera from febrile patients in Peru, 22 sera were indigenous MAT-negative febrile patient sera, and 36 sera were from patients with other febrile diseases from Southeast Asia, where leptospirosis is also endemic. Combining the results of immunoglobulin M (IgM) and IgG detection from these three antigens, the overall sensitivity is close to 90% based on the MAT. These results suggest that an ELISA using multiple recombinant antigens may be used as an alternative method for the detection of Leptospira-specific antibodies. PMID:24166046

  18. PATHOGENIC LEPTOSPIRA SEROVARS IN FREE-LIVING SEA LIONS IN THE GULF OF CALIFORNIA AND ALONG THE BAJA CALIFORNIA COAST OF MEXICO.

    PubMed

    Avalos-Téllez, Rosalía; Carrillo-Casas, Erika M; Atilano-López, Daniel; Godínez-Reyes, Carlos R; Díaz-Aparicio, Efrén; Ramírez-Delgado, David; Ramírez-Echenique, María F; Leyva-Leyva, Margarita; Suzán, Gerardo; Suárez-Güemes, Francisco

    2016-04-28

    The California sea lion ( Zalophus californianus ), a permanent inhabitant of the Gulf of California in Mexico, is susceptible to pathogenic Leptospira spp. infection, which can result in hepatic and renal damage and may lead to renal failure and death. During summer 2013, we used the microscopic agglutination test (MAT) to investigate the prevalence of anti-Leptospira antibodies in blood of clinically healthy sea lion pups from seven rookery islands on the Pacific Coast of Baja California (Pacific Ocean) and in the Gulf of California. We also used PCR to examine blood for Leptospira DNA. Isolation of Leptospira in liquid media was unsuccessful. We found higher antibody prevalence in sea lions from the rookery islands in the gulf than in those from the Pacific Coast. Antibodies against 11 serovars were identified in the Gulf of California population; the most frequent reactions were against serovars Bataviae (90%), Pyrogenes (86%), Wolffi (86%), Celledoni (71%), and Pomona (65%). In the Pacific Ocean population, MAT was positive against eight serovars, where Wolffi (88%), Pomona (75%), and Bataviae (70%) were the most frequent. Serum samples agglutinated with more than one Leptospira serovar. The maximum titer was 3,200. Each island had a different serology profile, and islands combined showed a distinct profile for each region. We detected pathogenic Leptospira DNA in 63% of blood samples, but we found no saprophytic Leptospira. Positive PCR results were obtained in blood samples with high and low MAT titers. Together, these two methods enhance the diagnosis and interpretation of sea lion leptospirosis. Our results may be related to human activities or the presence of other reservoirs with which sea lions interact, and they may also be related to sea lion stranding. PMID:26967136

  19. PATHOGENIC LEPTOSPIRA SEROVARS IN FREE-LIVING SEA LIONS IN THE GULF OF CALIFORNIA AND ALONG THE BAJA CALIFORNIA COAST OF MEXICO.

    PubMed

    Avalos-Téllez, Rosalía; Carrillo-Casas, Erika M; Atilano-López, Daniel; Godínez-Reyes, Carlos R; Díaz-Aparicio, Efrén; Ramírez-Delgado, David; Ramírez-Echenique, María F; Leyva-Leyva, Margarita; Suzán, Gerardo; Suárez-Güemes, Francisco

    2016-04-28

    The California sea lion ( Zalophus californianus ), a permanent inhabitant of the Gulf of California in Mexico, is susceptible to pathogenic Leptospira spp. infection, which can result in hepatic and renal damage and may lead to renal failure and death. During summer 2013, we used the microscopic agglutination test (MAT) to investigate the prevalence of anti-Leptospira antibodies in blood of clinically healthy sea lion pups from seven rookery islands on the Pacific Coast of Baja California (Pacific Ocean) and in the Gulf of California. We also used PCR to examine blood for Leptospira DNA. Isolation of Leptospira in liquid media was unsuccessful. We found higher antibody prevalence in sea lions from the rookery islands in the gulf than in those from the Pacific Coast. Antibodies against 11 serovars were identified in the Gulf of California population; the most frequent reactions were against serovars Bataviae (90%), Pyrogenes (86%), Wolffi (86%), Celledoni (71%), and Pomona (65%). In the Pacific Ocean population, MAT was positive against eight serovars, where Wolffi (88%), Pomona (75%), and Bataviae (70%) were the most frequent. Serum samples agglutinated with more than one Leptospira serovar. The maximum titer was 3,200. Each island had a different serology profile, and islands combined showed a distinct profile for each region. We detected pathogenic Leptospira DNA in 63% of blood samples, but we found no saprophytic Leptospira. Positive PCR results were obtained in blood samples with high and low MAT titers. Together, these two methods enhance the diagnosis and interpretation of sea lion leptospirosis. Our results may be related to human activities or the presence of other reservoirs with which sea lions interact, and they may also be related to sea lion stranding.

  20. IMMUNOLOGICAL STUDIES WITH A STRAIN OF LEPTOSPIRA ISOLATED FROM A CASE OF YELLOW FEVER IN MERIDA, YUCATAN

    PubMed Central

    Noguchi, Hideyo; Kligler, I. J.

    1920-01-01

    Identification of the leptospira isolated from a case of yellow fever in Merida was accomplished by means of an anti-icteroides immune serum prepared in a horse with several Guayaquil strains of Leptospira icteroides. The immune serum showed a protective action of high titer against the Merida strain, thus establishing its efficacy as a therapeutic agent against this strain. Polyvalent anti-icteroides immune serum prepared in the horse or monovalent anti-icteroides immune serums prepared in the rabbit had a definite devitalizing action upon the Merida strain, while immune serums similarly prepared with strains of icterohœmorrhagiœ had no perceptible effect upon the Merida strain. Serums from yellow fever convalescents in Merida gave a positive Pfeiffer reaction with the Merida strain of Leptospira icteroides. The reactions between the Guayaquil strains (Nos. 1 and 5) and two of these serums from convalescents varied from definitely positive to doubtful, owing probably to the diminution of active immune principles in the serums during the prolonged and unfavorable conditions of their transportation. PMID:19868465

  1. Detection of antibodies against Brucella abortus, Leptospira spp., and Apicomplexa protozoa in water buffaloes in the Northeast of Argentina.

    PubMed

    Konrad, José L; Campero, Lucía M; Caspe, Gastón S; Brihuega, Bibiana; Draghi, Graciela; Moore, Dadin P; Crudeli, Gustavo A; Venturini, María C; Campero, Carlos M

    2013-11-01

    Water buffalo industry has become a profitable activity worldwide, including the Northeast of Argentina (NEA). However, research on diseases affecting this species is scarce. The aim of the present study was to detect antibodies against Brucella abortus, Leptospira spp., Neospora caninum, Toxoplasma gondii, and Sarcocystis spp. in 500 water buffalo cows from five ranches (100 animals each) in the NEA. Serum samples were tested for B. abortus by fluorescence polarization assay, Leptospira spp. by microagglutination test, and N. caninum, T. gondii, and Sarcocystis spp. by indirect fluorescent antibody tests. Overall, the proportion of seropositive animals was 6.4, 22.2, 42.2, 25.4, and 50.8 % for brucellosis, leptospirosis, neosporosis, toxoplasmosis, and sarcocystosis, respectively. The proportion of seropositive animals for all diseases was statistically different among herds (p < 0.05). Statistical differences were also detected among age groups for brucellosis and neosporosis (p < 0.05). The detection of specific antibodies to B. abortus, Leptospira spp., and several Apicomplexa protozoans in water buffaloes in the NEA is reported in this study. PMID:23765549

  2. Detection of antibodies against Brucella abortus, Leptospira spp., and Apicomplexa protozoa in water buffaloes in the Northeast of Argentina.

    PubMed

    Konrad, José L; Campero, Lucía M; Caspe, Gastón S; Brihuega, Bibiana; Draghi, Graciela; Moore, Dadin P; Crudeli, Gustavo A; Venturini, María C; Campero, Carlos M

    2013-11-01

    Water buffalo industry has become a profitable activity worldwide, including the Northeast of Argentina (NEA). However, research on diseases affecting this species is scarce. The aim of the present study was to detect antibodies against Brucella abortus, Leptospira spp., Neospora caninum, Toxoplasma gondii, and Sarcocystis spp. in 500 water buffalo cows from five ranches (100 animals each) in the NEA. Serum samples were tested for B. abortus by fluorescence polarization assay, Leptospira spp. by microagglutination test, and N. caninum, T. gondii, and Sarcocystis spp. by indirect fluorescent antibody tests. Overall, the proportion of seropositive animals was 6.4, 22.2, 42.2, 25.4, and 50.8 % for brucellosis, leptospirosis, neosporosis, toxoplasmosis, and sarcocystosis, respectively. The proportion of seropositive animals for all diseases was statistically different among herds (p < 0.05). Statistical differences were also detected among age groups for brucellosis and neosporosis (p < 0.05). The detection of specific antibodies to B. abortus, Leptospira spp., and several Apicomplexa protozoans in water buffaloes in the NEA is reported in this study.

  3. Leptospira weilii serovar Topaz, a new member of the Tarassovi serogroup isolated from a bovine source in Queensland, Australia.

    PubMed

    Corney, B G; Slack, A T; Symonds, M L; Dohnt, M F; McClintock, C S; McGowan, M R; Smythe, L D

    2008-10-01

    This paper reports on a Leptospira isolate of bovine origin and its identification as belonging to a previously unknown serovar, for which the name Topaz is proposed. The isolate (94-79970/3) was cultured from bovine urine from a north Queensland dairy farm in Australia. Strain 94-79970/3 grew at 30 degrees C in Ellinghausen McCullough Johnson Harris (EMJH) medium but failed to grow at 13 degrees C in EMJH medium or in the presence of 8-azaguanine. Serologically, strain 94-79970/3 produced titres against the Leptospira borgpetersenii serovar Tarassovi, the reference strain for the Tarassovi serogroup; however, no significant titres to any other serovars within the serogroup were obtained. Using 16S rRNA and DNA gyrase subunit B gene analysis, strain 94-79970/3 was identified as a member of the species Leptospira weilii. We propose that the serovar be named Topaz, after the location where the original isolate was obtained. The reference strain for this serovar is 94-79970/3 (=KIT 94-79970/3=LT722).

  4. Sensitive Real-Time PCR Detection of Pathogenic Leptospira spp. and a Comparison of Nucleic Acid Amplification Methods for the Diagnosis of Leptospirosis

    PubMed Central

    Waggoner, Jesse J.; Balassiano, Ilana; Abeynayake, Janaki; Sahoo, Malaya K.; Mohamed-Hadley, Alisha; Liu, Yuanyuan; Vital-Brazil, Juliana Magalhães; Pinsky, Benjamin A.

    2014-01-01

    Background Bacteria of the genus Leptospira, the causative agents of leptospirosis, are categorized into pathogenic and non-pathogenic species. However, the benefit of using a clinical diagnostic that is specific for pathogenic species remains unclear. In this study, we present the development of a real-time PCR (rtPCR) for the detection of pathogenic Leptospira (the pathogenic rtPCR), and we perform a comparison of the pathogenic rtPCR with a published assay that detects all Leptospira species [the undifferentiated febrile illness (UFI) assay] and a reference 16S Leptospira rtPCR, which was originally designed to detect pathogenic species. Methodology/Principal Findings For the pathogenic rtPCR, a new hydrolysis probe was designed for use with primers from the UFI assay, which targets the 16S gene. The pathogenic rtPCR detected Leptospira DNA in 37/37 cultured isolates from 5 pathogenic and one intermediate species. Two strains of the non-pathogenic L. biflexa produced no signal. Clinical samples from 65 patients with suspected leptospirosis were then tested using the pathogenic rtPCR and a reference Leptospira 16S rtPCR. All 65 samples had tested positive for Leptospira using the UFI assay; 62 (95.4%) samples tested positive using the pathogenic rtPCR (p = 0.24). Only 24 (36.9%) samples tested positive in the reference 16S rtPCR (p<0.0001 for comparison with the pathogenic rtPCR and UFI assays). Amplicon sequencing confirmed the detection of pathogenic Leptospira species in 49/50 cases, including 3 cases that were only detected using the UFI assay. Conclusions/Significance The pathogenic rtPCR displayed similar sensitivity to the UFI assay when testing clinical specimens with no difference in specificity. Both assays proved significantly more sensitive than a real-time molecular test used for comparison. Future studies are needed to investigate the clinical and epidemiologic significance of more sensitive Leptospira detection using these tests. PMID:25379890

  5. Detection of Leptospira spp. and Brucella abortus antibodies in free-living jaguars (Panthera onca) in two protected areas of northern Pantanal, Brazil.

    PubMed

    Onuma, Selma Samiko Miyazaki; Kantek, Daniel Luis Zanella; Crawshaw Júnior, Peter Gransden; Morato, Ronaldo Gonçalves; May-Júnior, Joares Adenilson; Morais, Zenaide Maria de; Ferreira Neto, José Soares; Aguiar, Daniel Moura de

    2015-01-01

    This study aimed to assess the exposure of free-living jaguars (Panthera onca) to Leptospira spp. and Brucella abortus in two conservation units in the Pantanal of Mato Grosso, Brazil. The presence of antibodies in blood samples of eleven jaguars was investigated using autochthonous antigens isolated in Brazil added to reference antigen collection applied to diagnosis of leptospirosis by Microscopic Agglutination Test (MAT). The Rose Bengal test was applied for B. abortus antibodies. Two (18.2%) jaguars were seroreactive for the Leptospira spp. antigen and the serovar considered as most infective in both animals was a Brazilian isolate of serovar Canicola (L01). All jaguars were seronegative for B. abortus. These data indicate that the inclusion of autochthonous antigens in serological studies can significantly increase the number of reactive animals, as well as modify the epidemiological profile of Leptospira spp. infection.

  6. Detection of Leptospira spp. and Brucella abortus antibodies in free-living jaguars (Panthera onca) in two protected areas of northern Pantanal, Brazil.

    PubMed

    Onuma, Selma Samiko Miyazaki; Kantek, Daniel Luis Zanella; Crawshaw Júnior, Peter Gransden; Morato, Ronaldo Gonçalves; May-Júnior, Joares Adenilson; Morais, Zenaide Maria de; Ferreira Neto, José Soares; Aguiar, Daniel Moura de

    2015-01-01

    This study aimed to assess the exposure of free-living jaguars (Panthera onca) to Leptospira spp. and Brucella abortus in two conservation units in the Pantanal of Mato Grosso, Brazil. The presence of antibodies in blood samples of eleven jaguars was investigated using autochthonous antigens isolated in Brazil added to reference antigen collection applied to diagnosis of leptospirosis by Microscopic Agglutination Test (MAT). The Rose Bengal test was applied for B. abortus antibodies. Two (18.2%) jaguars were seroreactive for the Leptospira spp. antigen and the serovar considered as most infective in both animals was a Brazilian isolate of serovar Canicola (L01). All jaguars were seronegative for B. abortus. These data indicate that the inclusion of autochthonous antigens in serological studies can significantly increase the number of reactive animals, as well as modify the epidemiological profile of Leptospira spp. infection. PMID:25923900

  7. DETECTION OF Leptospira spp. AND Brucella abortus ANTIBODIES IN FREE-LIVING JAGUARS (Panthera onca) IN TWO PROTECTED AREAS OF NORTHERN PANTANAL, BRAZIL

    PubMed Central

    ONUMA, Selma Samiko Miyazaki; KANTEK, Daniel Luis Zanella; CRAWSHAW, Peter Gransden; MORATO, Ronaldo Gonçalves; MAY-JÚNIOR, Joares Adenilson; de MORAIS, Zenaide Maria; FERREIRA, José Soares; de AGUIAR, Daniel Moura

    2015-01-01

     This study aimed to assess the exposure of free-living jaguars (Panthera onca) to Leptospira spp. and Brucella abortus in two conservation units in the Pantanal of Mato Grosso, Brazil. The presence of antibodies in blood samples of eleven jaguars was investigated using autochthonous antigens isolated in Brazil added to reference antigen collection applied to diagnosis of leptospirosis by Microscopic Agglutination Test (MAT). The Rose Bengal test was applied for B. abortus antibodies. Two (18.2%) jaguars were seroreactive for the Leptospira spp. antigen and the serovar considered as most infective in both animals was a Brazilian isolate of serovar Canicola (L01). All jaguars were seronegative for B. abortus. These data indicate that the inclusion of autochthonous antigens in serological studies can significantly increase the number of reactive animals, as well as modify the epidemiological profile of Leptospira spp. infection. PMID:25923900

  8. Comparison of fluorescent antibody and microscopic agglutination testing for Leptospira in pregnant and nonpregnant cows.

    PubMed

    Rajeev, Sreekumari; Berghaus, Roy D; Overton, Michael W; Pence, Mel E; Baldwin, Charles A

    2010-01-01

    Serum and urine samples from 30 cows (15 pregnant and 15 nonpregnant) from each of 10 Georgia dairy herds (total cows = 300) were examined by microscopic agglutination testing (MAT) and direct fluorescent antibody testing (FAT), respectively. Seven of the 10 herds had at least 1 cow with a positive FAT, and all of the herds had at least 1 cow with a reciprocal MAT titer > or =100 for 1 or more serovars. Serological testing was not helpful in identifying the infecting serovar for cows with a positive FAT result. The MAT titers for all 7 of the serovars evaluated were significantly correlated with one another, with 17 (81%) of the 21 Spearman rank correlation coefficients > or =0.4 in magnitude. Twenty (56%) of 36 FAT-positive cows did not have a titer that was highest for any particular serovar. Four of the 7 herds that reported using a Leptospira borgpetersenii serovar Hardjo-bovis vaccine had one or more FAT-positive cows compared with 3 out of 3 herds that reported they were not using this type of vaccine, although this difference was not statistically significant.

  9. Pathogenic Leptospira Species Acquire Factor H and Vitronectin via the Surface Protein LcpA

    PubMed Central

    da Silva, Ludmila Bezerra; Miragaia, Lidia dos Santos; Breda, Leandro Carvalho Dantas; Abe, Cecilia Mari; Schmidt, Mariana Costa Braga; Moro, Ana Maria; Monaris, Denize; Conde, Jonas Nascimento; Józsi, Mihály; Isaac, Lourdes; Abreu, Patrícia Antônia Estima

    2014-01-01

    Upon infection, pathogenic Leptospira species bind several complement regulators in order to overcome host innate immunity. We previously characterized a 20-kDa leptospiral surface protein which interacts with C4b binding protein (C4BP): leptospiral complement regulator-acquiring protein A (LcpA). Here we show that LcpA also interacts with human factor H (FH), which remains functionally active once bound to the protein. Antibodies directed against short consensus repeat 20 (SCR20) inhibited binding of FH to LcpA by approximately 90%, thus confirming that this particular domain is involved in the interaction. We have also shown for the first time that leptospires bind human vitronectin and that the interaction is mediated by LcpA. Coincubation with heparin blocked LcpA-vitronectin interaction in a dose-dependent manner, strongly suggesting that binding may occur through the heparin binding domains of vitronectin. LcpA also bound to the terminal pathway component C9 and inhibited Zn2+-induced polymerization and membrane attack complex (MAC) formation. Competitive binding assays indicated that LcpA interacts with C4BP, FH, and vitronectin through distinct sites. Taken together, our findings indicate that LcpA may play a role in leptospiral immune evasion. PMID:25534939

  10. Variables Associated with Infections of Cattle by Brucella abortus., Leptospira spp. and Neospora spp. in Amazon Region in Brazil.

    PubMed

    Chiebao, D P; Valadas, S Y O B; Minervino, A H H; Castro, V; Romaldini, A H C N; Calhau, A S; De Souza, R A B; Gennari, S M; Keid, L B; Soares, R M

    2015-10-01

    The frequency of Neospora spp., Leptospira spp. and Brucella abortus infections in adult cattle was determined in herds of the State of Pará, Brazil, which is an important region for cattle production located in the Amazon region. A total of 3466 adult female cattle from 176 herds were tested, leading to a frequency of seropositive animals of 14.7%, 3.7% and 65.5% and a herd positivity of 87.4%, 41.3% and 98.8% for infections caused by Neospora spp., B. abortus and Leptospira spp., respectively. The five most frequently diagnosed serologic responses to Leptospira spp. were those against serovars hardjo, wolfii, grippotyphosa, hebdomadis and shermani. The following associations were found: practice of artificial insemination, large farm size, large herd size, large number of dogs and high number of total abortions per year with the presence of antibodies against serovar hardjo; positive results to serovar grippotyphosa with the presence of dogs; inappropriate disposal of aborted foetuses with positivity to serovar hebdomadis. Serovar grippotyphosa was also associated with number of episodes of abortions. Neospora spp. positive herds were associated with episodes of abortion and B. abortus infection with the disposal of dead animals and aborted foetuses on pastures and with the use of artificial insemination. In conclusion, the high frequency of brucellosis, leptospirosis and neosporosis in the region may be a consequence of social, natural and raising conditions as: (i) climate conditions that favour the survival and spread of pathogens in the environment; (ii) farms located in regions bordering forest areas; (iii) farms in areas of difficult access to the veterinary service; (iv) extensive beef herds raised at pastures with different age and productive groups inter-mingled; and (v) minimal concerns regarding hygiene practices and disease prevention measures. PMID:26302373

  11. Variables Associated with Infections of Cattle by Brucella abortus., Leptospira spp. and Neospora spp. in Amazon Region in Brazil.

    PubMed

    Chiebao, D P; Valadas, S Y O B; Minervino, A H H; Castro, V; Romaldini, A H C N; Calhau, A S; De Souza, R A B; Gennari, S M; Keid, L B; Soares, R M

    2015-10-01

    The frequency of Neospora spp., Leptospira spp. and Brucella abortus infections in adult cattle was determined in herds of the State of Pará, Brazil, which is an important region for cattle production located in the Amazon region. A total of 3466 adult female cattle from 176 herds were tested, leading to a frequency of seropositive animals of 14.7%, 3.7% and 65.5% and a herd positivity of 87.4%, 41.3% and 98.8% for infections caused by Neospora spp., B. abortus and Leptospira spp., respectively. The five most frequently diagnosed serologic responses to Leptospira spp. were those against serovars hardjo, wolfii, grippotyphosa, hebdomadis and shermani. The following associations were found: practice of artificial insemination, large farm size, large herd size, large number of dogs and high number of total abortions per year with the presence of antibodies against serovar hardjo; positive results to serovar grippotyphosa with the presence of dogs; inappropriate disposal of aborted foetuses with positivity to serovar hebdomadis. Serovar grippotyphosa was also associated with number of episodes of abortions. Neospora spp. positive herds were associated with episodes of abortion and B. abortus infection with the disposal of dead animals and aborted foetuses on pastures and with the use of artificial insemination. In conclusion, the high frequency of brucellosis, leptospirosis and neosporosis in the region may be a consequence of social, natural and raising conditions as: (i) climate conditions that favour the survival and spread of pathogens in the environment; (ii) farms located in regions bordering forest areas; (iii) farms in areas of difficult access to the veterinary service; (iv) extensive beef herds raised at pastures with different age and productive groups inter-mingled; and (v) minimal concerns regarding hygiene practices and disease prevention measures.

  12. [Detection of Leptospira organisms in Rattus rattus of two islands in the Mozambique Channel: Europa and Juan-de-Nova].

    PubMed

    Freulon, M; Aboubaker, M; Marié, J-L; Drancourt, M; Davoust, B

    2010-02-01

    Europa and Juan-de-Nova are two little coral islands in the Mozambique Channel. They are only occupied by a troop detachment of 15 men, who exercise the French sovereignty and maintain the island. During these activities, the men work in dampness and they can encounter rats. The aim of this survey is to show presence of leptospirosis in these islands by testing rat kidney by specific PCR. The results found a positive specimen on each island (2/52) indicating a previously unknown presence of Leptospira organisms in these islands.

  13. Detection of Pathogenic Leptospira Bacteria in Pinniped Populations via PCR and Identification of a Source of Transmission for Zoonotic Leptospirosis in the Marine Environment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leptospirosis, caused by the spirochete bacterium Leptospira, is a geographically widespread disease that affects a broad range of mammals, including marine mammals. During 2004 an outbreak of leptospirosis occurred among select pinniped populations along the West Coast of North America, with cases...

  14. Serosurvey of Smooth Brucella, Leptospira spp. and Toxoplasma gondii in Free-Ranging Jaguars (Panthera onca) and Domestic Animals from Brazil.

    PubMed

    Furtado, Mariana Malzoni; Gennari, Solange Maria; Ikuta, Cassia Yumi; Jácomo, Anah Tereza de Almeida; de Morais, Zenaide Maria; Pena, Hilda Fátima de Jesus; Porfírio, Grasiela Edith de Oliveira; Silveira, Leandro; Sollmann, Rahel; de Souza, Gisele Oliveira; Tôrres, Natália Mundim; Ferreira Neto, José Soares

    2015-01-01

    This study investigated the exposure of jaguar populations and domestic animals to smooth Brucella, Leptospira spp. and Toxoplasma gondii in the Cerrado, Pantanal and Amazon biomes of Brazil. Between February 2000 and January 2010, serum samples from 31 jaguars (Panthera onca), 1,245 cattle (Bos taurus), 168 domestic dogs (Canis lupus familiaris) and 29 domestic cats (Felis catus) were collected and analysed by rose bengal test for smooth Brucella, microscopic agglutination test for Leptospira spp. and modified agglutination test for T. gondii. Cattle populations from all sites (9.88%) were exposed to smooth Brucella, but only one jaguar from Cerrado was exposed to this agent. Jaguars captured in the Cerrado (60.0%) and in the Pantanal (45.5%) were seropositive for different serovars of Leptospira spp., cattle (72.18%) and domestic dogs (13.1%) from the three sites and one domestic cat from Pantanal were also seropositive for the agent. The most prevalent serotype of Leptospira spp. identified in jaguars from the Cerrado (Grippotyphosa) and the Pantanal (Pomona) biomes were distinct from those found in the domestic animals sampled. Jaguars (100%), domestic dogs (38.28%) and domestic cats (82.76%) from the three areas were exposed to T. gondii. Our results show that brucellosis and leptospirosis could have been transmitted to jaguars by domestic animals; and jaguars probably play an important role in the maintenance of T. gondii in nature.

  15. Serosurvey of Smooth Brucella, Leptospira spp. and Toxoplasma gondii in Free-Ranging Jaguars (Panthera onca) and Domestic Animals from Brazil

    PubMed Central

    Furtado, Mariana Malzoni; Gennari, Solange Maria; Ikuta, Cassia Yumi; Jácomo, Anah Tereza de Almeida; de Morais, Zenaide Maria; Pena, Hilda Fátima de Jesus; Porfírio, Grasiela Edith de Oliveira; Silveira, Leandro; Sollmann, Rahel; de Souza, Gisele Oliveira; Tôrres, Natália Mundim; Ferreira Neto, José Soares

    2015-01-01

    This study investigated the exposure of jaguar populations and domestic animals to smooth Brucella, Leptospira spp. and Toxoplasma gondii in the Cerrado, Pantanal and Amazon biomes of Brazil. Between February 2000 and January 2010, serum samples from 31 jaguars (Panthera onca), 1,245 cattle (Bos taurus), 168 domestic dogs (Canis lupus familiaris) and 29 domestic cats (Felis catus) were collected and analysed by rose bengal test for smooth Brucella, microscopic agglutination test for Leptospira spp. and modified agglutination test for T. gondii. Cattle populations from all sites (9.88%) were exposed to smooth Brucella, but only one jaguar from Cerrado was exposed to this agent. Jaguars captured in the Cerrado (60.0%) and in the Pantanal (45.5%) were seropositive for different serovars of Leptospira spp., cattle (72.18%) and domestic dogs (13.1%) from the three sites and one domestic cat from Pantanal were also seropositive for the agent. The most prevalent serotype of Leptospira spp. identified in jaguars from the Cerrado (Grippotyphosa) and the Pantanal (Pomona) biomes were distinct from those found in the domestic animals sampled. Jaguars (100%), domestic dogs (38.28%) and domestic cats (82.76%) from the three areas were exposed to T. gondii. Our results show that brucellosis and leptospirosis could have been transmitted to jaguars by domestic animals; and jaguars probably play an important role in the maintenance of T. gondii in nature. PMID:26605787

  16. Serosurvey of Smooth Brucella, Leptospira spp. and Toxoplasma gondii in Free-Ranging Jaguars (Panthera onca) and Domestic Animals from Brazil.

    PubMed

    Furtado, Mariana Malzoni; Gennari, Solange Maria; Ikuta, Cassia Yumi; Jácomo, Anah Tereza de Almeida; de Morais, Zenaide Maria; Pena, Hilda Fátima de Jesus; Porfírio, Grasiela Edith de Oliveira; Silveira, Leandro; Sollmann, Rahel; de Souza, Gisele Oliveira; Tôrres, Natália Mundim; Ferreira Neto, José Soares

    2015-01-01

    This study investigated the exposure of jaguar populations and domestic animals to smooth Brucella, Leptospira spp. and Toxoplasma gondii in the Cerrado, Pantanal and Amazon biomes of Brazil. Between February 2000 and January 2010, serum samples from 31 jaguars (Panthera onca), 1,245 cattle (Bos taurus), 168 domestic dogs (Canis lupus familiaris) and 29 domestic cats (Felis catus) were collected and analysed by rose bengal test for smooth Brucella, microscopic agglutination test for Leptospira spp. and modified agglutination test for T. gondii. Cattle populations from all sites (9.88%) were exposed to smooth Brucella, but only one jaguar from Cerrado was exposed to this agent. Jaguars captured in the Cerrado (60.0%) and in the Pantanal (45.5%) were seropositive for different serovars of Leptospira spp., cattle (72.18%) and domestic dogs (13.1%) from the three sites and one domestic cat from Pantanal were also seropositive for the agent. The most prevalent serotype of Leptospira spp. identified in jaguars from the Cerrado (Grippotyphosa) and the Pantanal (Pomona) biomes were distinct from those found in the domestic animals sampled. Jaguars (100%), domestic dogs (38.28%) and domestic cats (82.76%) from the three areas were exposed to T. gondii. Our results show that brucellosis and leptospirosis could have been transmitted to jaguars by domestic animals; and jaguars probably play an important role in the maintenance of T. gondii in nature. PMID:26605787

  17. PCR and Culture Identification of Pathogenic Leptospira spp. from Coastal Soil in Leyte, Philippines, after a Storm Surge during Super Typhoon Haiyan (Yolanda)

    PubMed Central

    Miyahara, Satoshi; Villanueva, Sharon Y. A. M.; Aramaki, Natsumi; Ikejiri, Mami; Kobayashi, Yoshie; Guevarra, Jonathan P.; Masuzawa, Toshiyuki; Gloriani, Nina G.; Yanagihara, Yasutake; Yoshida, Shin-ichi

    2014-01-01

    Leptospirosis is a zoonosis caused by pathogenic Leptospira spp. Most of the outbreaks of leptospirosis occur after floods caused by heavy rain in countries where Leptospira spp. are endemic. It has been believed that the overflow of seawater rarely causes outbreaks of leptospirosis because the leptospires are killed by salt water. On 8 November 2013, a storm surge caused by Super Typhoon Haiyan (Yolanda) inundated the entire coastal areas of Tacloban and Palo in Leyte, Philippines. The present study was carried out in order to determine whether the environmental leptospires in soil were able to survive after the storm surge in the affected areas. We collected 23 wet soil samples along the coastal areas of Tacloban and Palo 2 months after the storm surge. The samples were suspended in HEPES buffer, and the supernatants were cultured in liquid or semisolid Korthof's medium supplemented with five antimicrobial agents to inhibit the growth of contaminants. Leptospires were isolated from primary cultures of 22 out of 23 samples. The DNA of pathogenic Leptospira species was detected in 11 samples (47.8%) by analysis of flaB by nested PCR. Eventually, two pathogenic Leptospira strains were isolated and showed the highest 16S rRNA gene sequence similarity to Leptospira kmetyi. When these isolates were experimentally mixed with soil, they were found to survive in seawater for 4 days. These results show the possibility that leptospires living in soil survived after the storm surge. Our findings may serve as a warning that when seawater inundates the land during a storm surge or a tsunami, an outbreak of leptospirosis could occur in the disaster-stricken area. PMID:25172869

  18. Evaluation of three 5' exonuclease-based real-time polymerase chain reaction assays for detection of pathogenic Leptospira species in canine urine.

    PubMed

    Fink, Jamie M; Moore, George E; Landau, Ruth; Vemulapalli, Ramesh

    2015-03-01

    Leptospirosis is caused by several pathogenic Leptospira species, and is an important infectious disease of dogs. Early detection of infection is crucial for an effective antibiotic treatment of the disease. Though different polymerase chain reaction (PCR) assays have been developed for detection of pathogenic Leptospira spp., thorough evaluation of the performance of these assays using dog urine samples has not been carried out. In the current study, the performance of 3 real-time PCR (qPCR) assays was assessed, 1 targeting the 16S ribosomal RNA (rRNA) gene and the other 2 targeting the lipL32 gene, a gene for the LipL32 outer membrane protein. With DNA extracted from laboratory-cultured pathogenic Leptospira spp., all 3 qPCR assays showed 100% specificity and had identical lower limits of detection. Compared to a conventional, gel-based PCR assay, all 3 qPCR assays were 100-fold more sensitive. There was a 100% agreement in the results of the 3 assays when tested on urine samples collected aseptically from 30 dogs suspected for leptospirosis. However, when tested on 30 urine samples that were collected by the free-catch method, the 16S rRNA-based assay falsely detected 13.3% of the samples as positive for pathogenic Leptospira spp. Nucleotide sequence analysis of the amplified DNA fragments showed that the assay resulted in false positives because of unrelated bacteria. All urine samples collected from 100 apparently healthy dogs at a local animal shelter tested negative for pathogenic Leptospira spp. These results highlight the importance of sample-specific validation of PCR-based diagnostic assays and the application of appropriately validated assays for more reliable pathogen detection. PMID:25776541

  19. A new multivalent (DHPPi/L4R) canine combination vaccine prevents infection, shedding and clinical signs following experimental challenge with four Leptospira serovars.

    PubMed

    Wilson, Stephen; Stirling, Catrina; Thomas, Anne; King, Vickie; Plevová, Edita; Chromá, Ludmila; Siedek, Elisabeth; Illambas, Joanna; Salt, Jeremy; Sture, Gordon

    2013-06-28

    Although effective vaccines have been developed against the common Leptospira serovars, they are still reported in clinical cases, while others are increasingly prevalent. The results from four challenge studies following vaccination of dogs with a new combination vaccine (DHPPi/L4R) containing inactivated L. serovars, L. canicola, L. icterohaemorrhagiae, L. bratislava and L. grippotyphosa conducted to satisfy the requirements of the European Pharmacopoeia monograph (01/2008:0447), are reported. Six week old dogs received two vaccinations, three weeks apart, and were challenged 25 days later with different isolates of the L. serovars. Clinical observations were recorded, and blood, urine and tissue samples were collected for analysis. Following challenge, non-vaccinated dogs demonstrated various clinical signs, while no vaccinated dogs were affected; significant differences in mean clinical scores were observed. Measurable antibody titres to each Leptospira antigen were seen in vaccinated dogs 21 days following the first vaccination, with further increases in antibody titres observed following challenge with the respective Leptospira strain. Non-vaccinated dogs remained seronegative until challenge. Leptospira were re-isolated from the blood, urine, kidney and liver of all non-vaccinated dogs following challenge. In contrast no vaccinated dogs had Leptospira re-isolated from the same tissues. Significant differences were seen in number of days with positive isolation (blood and urine) and in number of dogs with positive samples (kidney and liver). In conclusion, vaccination of dogs with the new vaccine induces protective immunity 25 days after second vaccination with protection against infection, renal infection and clinical signs following challenge.

  20. PCR and culture identification of pathogenic Leptospira spp. from coastal soil in Leyte, Philippines, after a storm surge during Super Typhoon Haiyan (Yolanda).

    PubMed

    Saito, Mitsumasa; Miyahara, Satoshi; Villanueva, Sharon Y A M; Aramaki, Natsumi; Ikejiri, Mami; Kobayashi, Yoshie; Guevarra, Jonathan P; Masuzawa, Toshiyuki; Gloriani, Nina G; Yanagihara, Yasutake; Yoshida, Shin-ichi

    2014-11-01

    Leptospirosis is a zoonosis caused by pathogenic Leptospira spp. Most of the outbreaks of leptospirosis occur after floods caused by heavy rain in countries where Leptospira spp. are endemic. It has been believed that the overflow of seawater rarely causes outbreaks of leptospirosis because the leptospires are killed by salt water. On 8 November 2013, a storm surge caused by Super Typhoon Haiyan (Yolanda) inundated the entire coastal areas of Tacloban and Palo in Leyte, Philippines. The present study was carried out in order to determine whether the environmental leptospires in soil were able to survive after the storm surge in the affected areas. We collected 23 wet soil samples along the coastal areas of Tacloban and Palo 2 months after the storm surge. The samples were suspended in HEPES buffer, and the supernatants were cultured in liquid or semisolid Korthof's medium supplemented with five antimicrobial agents to inhibit the growth of contaminants. Leptospires were isolated from primary cultures of 22 out of 23 samples. The DNA of pathogenic Leptospira species was detected in 11 samples (47.8%) by analysis of flaB by nested PCR. Eventually, two pathogenic Leptospira strains were isolated and showed the highest 16S rRNA gene sequence similarity to Leptospira kmetyi. When these isolates were experimentally mixed with soil, they were found to survive in seawater for 4 days. These results show the possibility that leptospires living in soil survived after the storm surge. Our findings may serve as a warning that when seawater inundates the land during a storm surge or a tsunami, an outbreak of leptospirosis could occur in the disaster-stricken area. PMID:25172869

  1. PCR and culture identification of pathogenic Leptospira spp. from coastal soil in Leyte, Philippines, after a storm surge during Super Typhoon Haiyan (Yolanda).

    PubMed

    Saito, Mitsumasa; Miyahara, Satoshi; Villanueva, Sharon Y A M; Aramaki, Natsumi; Ikejiri, Mami; Kobayashi, Yoshie; Guevarra, Jonathan P; Masuzawa, Toshiyuki; Gloriani, Nina G; Yanagihara, Yasutake; Yoshida, Shin-ichi

    2014-11-01

    Leptospirosis is a zoonosis caused by pathogenic Leptospira spp. Most of the outbreaks of leptospirosis occur after floods caused by heavy rain in countries where Leptospira spp. are endemic. It has been believed that the overflow of seawater rarely causes outbreaks of leptospirosis because the leptospires are killed by salt water. On 8 November 2013, a storm surge caused by Super Typhoon Haiyan (Yolanda) inundated the entire coastal areas of Tacloban and Palo in Leyte, Philippines. The present study was carried out in order to determine whether the environmental leptospires in soil were able to survive after the storm surge in the affected areas. We collected 23 wet soil samples along the coastal areas of Tacloban and Palo 2 months after the storm surge. The samples were suspended in HEPES buffer, and the supernatants were cultured in liquid or semisolid Korthof's medium supplemented with five antimicrobial agents to inhibit the growth of contaminants. Leptospires were isolated from primary cultures of 22 out of 23 samples. The DNA of pathogenic Leptospira species was detected in 11 samples (47.8%) by analysis of flaB by nested PCR. Eventually, two pathogenic Leptospira strains were isolated and showed the highest 16S rRNA gene sequence similarity to Leptospira kmetyi. When these isolates were experimentally mixed with soil, they were found to survive in seawater for 4 days. These results show the possibility that leptospires living in soil survived after the storm surge. Our findings may serve as a warning that when seawater inundates the land during a storm surge or a tsunami, an outbreak of leptospirosis could occur in the disaster-stricken area.

  2. The adaptor molecule Trif contributes to murine host defense during Leptospiral infection.

    PubMed

    Jayaraman, Priya A; Devlin, Amy A; Miller, Jennifer C; Scholle, Frank

    2016-09-01

    Leptospirosis is a zoonotic disease and is caused by pathogenic species of the Leptospira genus, including Leptospira interrogans (L. interrogans). Humans, domestic and wild animals are susceptible to acute or chronic infection. The innate immune response is a critical defense mechanism against Leptospira interrogans, and has been investigated in mouse models. Murine Toll-like receptors (TLRs) have been shown to be key factors in sensing and responding to L. interrogans infection. Specifically, TLR2, TLR4 and the TLR adaptor molecule MyD88 are essential for host defense against L. interrogans; however, the role of the TLR adaptor molecule TIR-domain-containing adaptor-inducing interferon β (TRIF) in the response to L. interrogans has not been previously determined. In the present study, TRIF was found to play an important role during leptospiral infection. Following challenge with L. interrogans, Trif(-/-) mice exhibited delayed weight gain compared to wild-type mice. Moreover, Trif(-/-) mice exhibited an increase in L. interrogans burden in the kidneys, lungs, and blood at early time points (less than 7days post infection). Multiple components of the innate immune responses were dampened in response to leptospiral infection including transcription and production of cytokines, and the humoral response, which suggested that TRIF contributes to expression and production of cytokines important for the host defense against L. interrogans. PMID:27259371

  3. Isolation and clinical sample typing of human leptospirosis cases in Argentina.

    PubMed

    Chiani, Yosena; Jacob, Paulina; Varni, Vanina; Landolt, Noelia; Schmeling, María Fernanda; Pujato, Nazarena; Caimi, Karina; Vanasco, Bibiana

    2016-01-01

    Leptospira typing is carried out using isolated strains. Because of difficulties in obtaining them, direct identification of infective Leptospira in clinical samples is a high priority. Multilocus sequence typing (MLST) proved highly discriminatory for seven pathogenic species of Leptospira, allowing isolate characterization and robust assignment to species, in addition to phylogenetic evidence for the relatedness between species. In this study we characterized Leptospira strains circulating in Argentina, using typing methods applied to human clinical samples and isolates. Phylogenetic studies based on 16S ribosomal RNA gene sequences enabled typing of 8 isolates (6 Leptospira interrogans, one Leptospira wolffii and one Leptospira broomii) and 58 out of 85 (68.2%) clinical samples (55 L. interrogans, 2 Leptospira meyeri, and one Leptospira kirschneri). MLST results for the L. interrogans isolates indicated that five were probably Canicola serogroup (ST37) and one was probably Icterohaemorrhagiae serogroup (ST17). Eleven clinical samples (21.6%), provided MLST interpretable data: five were probably Pyrogenes serogroup (ST13), four Sejroe (ST20), one Autumnalis (ST22) and one Canicola (ST37). To the best of our knowledge this study is the first report of the use of an MLST typing scheme with seven loci to identify Leptospira directly from clinical samples in Argentina. The use of clinical samples presents the advantage of the possibility of knowing the infecting strain without resorting to isolates. This study also allowed, for the first time, the characterization of isolates of intermediate pathogenicity species (L. wolffii and L. broomii) from symptomatic patients. PMID:26658064

  4. Isolation and clinical sample typing of human leptospirosis cases in Argentina.

    PubMed

    Chiani, Yosena; Jacob, Paulina; Varni, Vanina; Landolt, Noelia; Schmeling, María Fernanda; Pujato, Nazarena; Caimi, Karina; Vanasco, Bibiana

    2016-01-01

    Leptospira typing is carried out using isolated strains. Because of difficulties in obtaining them, direct identification of infective Leptospira in clinical samples is a high priority. Multilocus sequence typing (MLST) proved highly discriminatory for seven pathogenic species of Leptospira, allowing isolate characterization and robust assignment to species, in addition to phylogenetic evidence for the relatedness between species. In this study we characterized Leptospira strains circulating in Argentina, using typing methods applied to human clinical samples and isolates. Phylogenetic studies based on 16S ribosomal RNA gene sequences enabled typing of 8 isolates (6 Leptospira interrogans, one Leptospira wolffii and one Leptospira broomii) and 58 out of 85 (68.2%) clinical samples (55 L. interrogans, 2 Leptospira meyeri, and one Leptospira kirschneri). MLST results for the L. interrogans isolates indicated that five were probably Canicola serogroup (ST37) and one was probably Icterohaemorrhagiae serogroup (ST17). Eleven clinical samples (21.6%), provided MLST interpretable data: five were probably Pyrogenes serogroup (ST13), four Sejroe (ST20), one Autumnalis (ST22) and one Canicola (ST37). To the best of our knowledge this study is the first report of the use of an MLST typing scheme with seven loci to identify Leptospira directly from clinical samples in Argentina. The use of clinical samples presents the advantage of the possibility of knowing the infecting strain without resorting to isolates. This study also allowed, for the first time, the characterization of isolates of intermediate pathogenicity species (L. wolffii and L. broomii) from symptomatic patients.

  5. Case Series of Fatal Leptospira spp./Dengue Virus Co-Infections—Puerto Rico, 2010–2012

    PubMed Central

    Pérez Rodríguez, Nicole M.; Galloway, Renee; Blau, Dianna M.; Traxler, Rita; Bhatnagar, Julu; Zaki, Sherif R.; Rivera, Aidsa; Torres, Jose V.; Noyd, David; Santiago-Albizu, Xavier E.; García, Brenda Rivera; Tomashek, Kay M.; Bower, William A.; Sharp, Tyler M.

    2014-01-01

    Co-infection with pathogens that cause acute febrile illness creates a diagnostic challenge as a result of overlapping clinical manifestations. Here, we describe four fatal cases of Leptospira species/dengue virus co-infection in Puerto Rico. Although all patients sought care early, antibiotic administration was delayed for most. Steroids were administered to all patients, in most cases before antibiotics. These cases show the need for clinicians evaluating patients in or recently returned from the tropics with acute febrile illness to consider both dengue and leptospirosis. Furthermore, they illustrate the need for nucleic acid- or antigen-based rapid diagnostic tests to enable timely patient diagnosis and management. In particular, antibiotic therapy should be initiated early for patients with suspected leptospirosis, and steroids should not be administered to patients with suspected dengue. PMID:25092820

  6. Serologic survey for Leptospira spp. in captive neotropical felids in Foz do Iguaçu, Paraná, Brazil.

    PubMed

    Ullmann, Leila Sabrina; Hoffmann, Juliano L; de Moraes, Wanderlei; Cubas, Zalmir S; dos Santos, Leonilda Correia; da Silva, Rodrigo Costa; Moreira, Nei; Guimaraes, Ana Marcia Sa; Camossi, Lucilene Granuzzio; Langoni, Helio; Biondo, Alexander W

    2012-06-01

    Leptospirosis is a bacterial zoonosis of worldwide distribution and is endemic in tropical countries, where rodents and other wild mammals are abundant and may act as reservoirs. Leptospirosis has become a concern in captive wild animals, due mostly to their exposure to contaminated urine or environment. Although domestic cats (Felis catus) have been reported refractory to leptospirosis, serology and disease in captive wild felids is still unclear. In this study 57 adult, clinically healthy felids, including 1 Geoffroy's cat (Leopardus geoffroyi), 3 jaguarundis (Puma yagouaroundi), 17 margays (Leopardus wiedii), 22 little spotted cats (Leopardus tigrinus), and 14 ocelots (Leopardus pardalis) kept in captivity at the Sanctuary at the Itaipu Binacional hydroelectric power plant (Bela Vista Biological Sanctuary), Foz do Iguacu City, Paraná State, Brazil, were serologically surveyed for the presence of antibodies against 28 serovars of Leptospira spp. by microagglutination test (MAT). Two animals (3.5%) were seropositive: one male ocelot to the serovar Cynopteri (titer 100) and one female margay to Autumnalis (100) and Butembo (200). The captive-born, 5-yr-old ocelot had been solitary housed in an individual cage. The approximately 21-yr-old wild-caught margay was also kept individually. None of the tested animals showed signs ofleptospirosis. During a study conducted 4 yr previously in the same facility, this particular margay also tested positive for the same two serovars, among others. The present study indicates that the felids tested for Leptospira spp. by MAT were exposed to serovars, but did not demonstrate clinical signs of disease. Comparison with a previous study suggests that serovar titers may vary over time and that leptospirosis dynamics remains unclear in wild felids.

  7. Urban Market Gardening and Rodent-Borne Pathogenic Leptospira in Arid Zones: A Case Study in Niamey, Niger.

    PubMed

    Dobigny, Gauthier; Garba, Madougou; Tatard, Caroline; Loiseau, Anne; Galan, Max; Kadaouré, Ibrahima; Rossi, Jean-Pierre; Picardeau, Mathieu; Bertherat, Eric

    2015-01-01

    Leptospirosis essentially affects human following contact with rodent urine-contaminated water. As such, it was mainly found associated with rice culture, recreational activities and flooding. This is also the reason why it has mainly been investigated in temperate as well as warm and humid regions, while arid zones have been only very occasionally monitored for this disease. In particular, data for West African countries are extremely scarce. Here, we took advantage of an extensive survey of urban rodents in Niamey, Niger, in order to look for rodent-borne pathogenic Leptospira species presence and distribution across the city. To do so, we used high throughput bacterial 16S-based metabarcoding, lipL32 gene-targeting RT-PCR, rrs gene sequencing and VNTR typing as well as GIS-based multivariate spatial analysis. Our results show that leptospires seem absent from the core city where usual Leptospira reservoir rodent species (namely R. rattus and M. natalensis) are yet abundant. On the contrary, L. kirschneri was detected in Arvicanthis niloticus and Cricetomys gambianus, two rodent species that are restricted to irrigated cultures within the city. Moreover, the VNTR profiles showed that rodent-borne leptospires in Niamey belong to previously undescribed serovars. Altogether, our study points towards the importance of market gardening in maintain and circulation of leptospirosis within Sahelian cities. In Africa, irrigated urban agriculture constitutes a pivotal source of food supply, especially in the context of the ongoing extensive urbanization of the continent. With this in mind, we speculate that leptospirosis may represent a zoonotic disease of concern also in arid regions that would deserve to be more rigorously surveyed, especially in urban agricultural settings. PMID:26437456

  8. Serologic survey for Leptospira spp. in captive neotropical felids in Foz do Iguaçu, Paraná, Brazil.

    PubMed

    Ullmann, Leila Sabrina; Hoffmann, Juliano L; de Moraes, Wanderlei; Cubas, Zalmir S; dos Santos, Leonilda Correia; da Silva, Rodrigo Costa; Moreira, Nei; Guimaraes, Ana Marcia Sa; Camossi, Lucilene Granuzzio; Langoni, Helio; Biondo, Alexander W

    2012-06-01

    Leptospirosis is a bacterial zoonosis of worldwide distribution and is endemic in tropical countries, where rodents and other wild mammals are abundant and may act as reservoirs. Leptospirosis has become a concern in captive wild animals, due mostly to their exposure to contaminated urine or environment. Although domestic cats (Felis catus) have been reported refractory to leptospirosis, serology and disease in captive wild felids is still unclear. In this study 57 adult, clinically healthy felids, including 1 Geoffroy's cat (Leopardus geoffroyi), 3 jaguarundis (Puma yagouaroundi), 17 margays (Leopardus wiedii), 22 little spotted cats (Leopardus tigrinus), and 14 ocelots (Leopardus pardalis) kept in captivity at the Sanctuary at the Itaipu Binacional hydroelectric power plant (Bela Vista Biological Sanctuary), Foz do Iguacu City, Paraná State, Brazil, were serologically surveyed for the presence of antibodies against 28 serovars of Leptospira spp. by microagglutination test (MAT). Two animals (3.5%) were seropositive: one male ocelot to the serovar Cynopteri (titer 100) and one female margay to Autumnalis (100) and Butembo (200). The captive-born, 5-yr-old ocelot had been solitary housed in an individual cage. The approximately 21-yr-old wild-caught margay was also kept individually. None of the tested animals showed signs ofleptospirosis. During a study conducted 4 yr previously in the same facility, this particular margay also tested positive for the same two serovars, among others. The present study indicates that the felids tested for Leptospira spp. by MAT were exposed to serovars, but did not demonstrate clinical signs of disease. Comparison with a previous study suggests that serovar titers may vary over time and that leptospirosis dynamics remains unclear in wild felids. PMID:22779223

  9. Urban Market Gardening and Rodent-Borne Pathogenic Leptospira in Arid Zones: A Case Study in Niamey, Niger

    PubMed Central

    Dobigny, Gauthier; Garba, Madougou; Tatard, Caroline; Loiseau, Anne; Galan, Max; Kadaouré, Ibrahima; Rossi, Jean-Pierre; Picardeau, Mathieu; Bertherat, Eric

    2015-01-01

    Leptospirosis essentially affects human following contact with rodent urine-contaminated water. As such, it was mainly found associated with rice culture, recreational activities and flooding. This is also the reason why it has mainly been investigated in temperate as well as warm and humid regions, while arid zones have been only very occasionally monitored for this disease. In particular, data for West African countries are extremely scarce. Here, we took advantage of an extensive survey of urban rodents in Niamey, Niger, in order to look for rodent-borne pathogenic Leptospira species presence and distribution across the city. To do so, we used high throughput bacterial 16S-based metabarcoding, lipL32 gene-targeting RT-PCR, rrs gene sequencing and VNTR typing as well as GIS-based multivariate spatial analysis. Our results show that leptospires seem absent from the core city where usual Leptospira reservoir rodent species (namely R. rattus and M. natalensis) are yet abundant. On the contrary, L. kirschneri was detected in Arvicanthis niloticus and Cricetomys gambianus, two rodent species that are restricted to irrigated cultures within the city. Moreover, the VNTR profiles showed that rodent-borne leptospires in Niamey belong to previously undescribed serovars. Altogether, our study points towards the importance of market gardening in maintain and circulation of leptospirosis within Sahelian cities. In Africa, irrigated urban agriculture constitutes a pivotal source of food supply, especially in the context of the ongoing extensive urbanization of the continent. With this in mind, we speculate that leptospirosis may represent a zoonotic disease of concern also in arid regions that would deserve to be more rigorously surveyed, especially in urban agricultural settings. PMID:26437456

  10. Urban Market Gardening and Rodent-Borne Pathogenic Leptospira in Arid Zones: A Case Study in Niamey, Niger.

    PubMed

    Dobigny, Gauthier; Garba, Madougou; Tatard, Caroline; Loiseau, Anne; Galan, Max; Kadaouré, Ibrahima; Rossi, Jean-Pierre; Picardeau, Mathieu; Bertherat, Eric

    2015-01-01

    Leptospirosis essentially affects human following contact with rodent urine-contaminated water. As such, it was mainly found associated with rice culture, recreational activities and flooding. This is also the reason why it has mainly been investigated in temperate as well as warm and humid regions, while arid zones have been only very occasionally monitored for this disease. In particular, data for West African countries are extremely scarce. Here, we took advantage of an extensive survey of urban rodents in Niamey, Niger, in order to look for rodent-borne pathogenic Leptospira species presence and distribution across the city. To do so, we used high throughput bacterial 16S-based metabarcoding, lipL32 gene-targeting RT-PCR, rrs gene sequencing and VNTR typing as well as GIS-based multivariate spatial analysis. Our results show that leptospires seem absent from the core city where usual Leptospira reservoir rodent species (namely R. rattus and M. natalensis) are yet abundant. On the contrary, L. kirschneri was detected in Arvicanthis niloticus and Cricetomys gambianus, two rodent species that are restricted to irrigated cultures within the city. Moreover, the VNTR profiles showed that rodent-borne leptospires in Niamey belong to previously undescribed serovars. Altogether, our study points towards the importance of market gardening in maintain and circulation of leptospirosis within Sahelian cities. In Africa, irrigated urban agriculture constitutes a pivotal source of food supply, especially in the context of the ongoing extensive urbanization of the continent. With this in mind, we speculate that leptospirosis may represent a zoonotic disease of concern also in arid regions that would deserve to be more rigorously surveyed, especially in urban agricultural settings.

  11. Serology, isolation, and molecular detection of Leptospira spp. from the tissues and blood of rats captured in a wild animal preservation centre in Brazil.

    PubMed

    Paixão, Mirian dos Santos; Alves-Martin, Maria Fernanda; Tenório, Michely da S; Starke-Buzetti, Wilma A; Alves, Maria Luana; da Silva, Diogo Tiago; Ferreira, Andréa Gonçalves; Floró e Silva, Marina; Sousa, Lúcio O; Lucheis, Simone B

    2014-07-01

    To elucidate the occurrence and epidemiology of leptospirosis in rats cohabitating with forest animals, 13 rats were captured at seven locations of the Centre for the Conservation of Wild Fauna (CCWF) in Sao Paulo state, Brazil, and samples of their blood, liver, and kidneys were collected. The diagnostic techniques utilized were the microscopic agglutination test (MAT), polymerase chain reaction (PCR) for Leptospira spp., and cultures of rat kidneys and liver in Fletcher's medium. The MAT results showed that 13 (100%) of the samples were reactive to 12 serovars among the 29 Leptospira spp. tested, and the Australis and Tarassovi serovars were the most frequently identified serovars. To research the agent in fragments of the liver and kidney, 13 samples from each tissue were cultured in Fletcher's medium, and the results revealed seven positive samples (53.8%; three from the kidneys and four from the livers). The analysis of the blood samples by PCR for Leptospira spp. showed that six animals (46.1%) were positive, whereas the analysis of the organs (kidneys and liver) by PCR revealed that nine animals (69.2%) were positive, and the culture of the organs revealed four positive animals (30.8%). These results suggest that the presence of Leptospira spp. infection in rats at the study site and the knowledge of the serovars that exist in this environment are important for the epidemiological comprehension of the disease and for the identification of control measures that should be considered to reduce the risk of transmission of the disease through this animal reservoir.

  12. Serologic-based investigation of leptospirosis in a population of free-ranging eastern grey kangaroos (Macropus giganteus) indicating the presence of Leptospira weilii serovar Topaz.

    PubMed

    Roberts, Michael W; Smythe, Lee; Dohnt, Michael; Symonds, Meegan; Slack, Andrew

    2010-04-01

    Eastern grey kangaroos (Macropus giganteus) are one of the most abundant large macropodids sharing the landscape with humans. Despite this, little is known about the prevalence of Leptospira carriage within this species and the role that they may partake in the transmission of this disease in Australia. The sera of 87 free-ranging eastern grey kangaroos, captured in the Warragamba Catchment Area, Sydney, Australia, from June 2004 to November 2006, were screened against a reference panel of 22 Leptospira serovars using the microscopic agglutination test (MAT). Leptospiral antibodies were detected in 47% (41 of 87) of serum samples collected. Leptospira weilii Topaz, a newly emergent serovar in Australia, was detected in all seropositive kangaroos (41 of 41; 100%). The sex and tail-fat body condition index of kangaroos appeared to have no significant effect on the exposure to the disease. This serologic-based study is the first reported for L. weilii serovar Topaz in New South Wales, to our knowledge, having previously been isolated only in humans and two other animal species (bovine and long-nosed bandicoot [Perameles nasuta]) in Western Australia and Queensland. The potential role of eastern grey kangaroos in the maintenance and zoonotic spread of the disease to livestock and humans is discussed.

  13. Seroprevalence and risk factors of antibodies against Leptospira spp. in ovines from Uberlândia municipality, Minas Gerais state, Brazil

    PubMed Central

    Salaberry, Sandra R.S.; Castro, Vanessa; Nassar, Alessandra F.C.; Castro, Jacqueline R.; Guimarães, Ednaldo C.; Lima-Ribeiro, Anna M.C.

    2011-01-01

    The objectives of the present study were to verify the seroprevalence of anti-Leptospira spp. antibodies, identify the most frequent serovars and the risk factors associated with the infection in Uberlândia, Minas Gerais, Brazil. A total of 334 ovines blood samples were collected in 12 farms from Uberlândia municipality to be evaluated by means the Microscopic Agglutination Test (MAT) against 22 serovars of Leptospira spp. and an epidemiologic questionnaire was applied for each farm in order to correlate with risk factors of leptospirosis: sex, age and breed as well as contact with cattle, contact with dogs and presence of rodents. The prevalence of seropositive to MAT was found in seventy four ovines (22.2%; CI 95% 17.6–26.4%), with titers ranging from 100 to 3200. The most frequent serovars identified were: Hardjo, Autumnalis, Hardjo and Wolffi association and Grippotyphosa. Statistically significant differences were found in males, pure breeds and presence of rodents (p<0.05). The prevalence of anti-Leptospira spp. antibodies found in the present study demonstrated that this bacterium occurs in ovines of Uberlândia municipality, MG, Brazil. The need for the adoption of efficient management for the control of rodents and infection in ovines in order to avoid leptospirosis in the local flocks and future transmission to humans. PMID:24031773

  14. An in vitro growth inhibition test for measuring the potency of Leptospira spp. Sejroe group vaccine in buffaloes.

    PubMed

    de Nardi, Geraldo; Genovez, Margareth Elide; Ribeiro, Marcio Garcia; Castro, Vanessa; Jorge, André Mendes

    2010-07-01

    Leptospira spp. serovars Hardjo and Wollfi from Sejroe serogroup have been detected in livestock in Brazil, where the main control procedures rely on vaccination. The potency of two commercial vaccines available in this country was monitored by microagglutination test-MAT and in vitro growth inhibition test-GIT in serum samples from 33 female buffaloes divided into: G1-unvaccinated control; G2-vaccinated with Leptobac-6 containing serovars Hardjo and Wolffi and G3-vaccinated with Triangle-9 containing serovar Hardjo. G2 and G3 animals were vaccinated on day zero, and received a booster and two revaccinations on days 30, 210 and 390 and G1 animals received phosphate buffered saline. Serum samples were collected at 15-day intervals between days 0 and 60; and at 30-day intervals between days 60 and 540 and were tested by MAT and GIT with serovars Hardjo and Wolffi. G1 remained negative throughout the experiment. Both vaccines were able to induce agglutinating and growth inhibition antibodies. Six months after the last revaccination, all animals tested negative by MAT, but still were positive by GIT until the end of experimental period. GIT could be a good tool to evaluate the potency and to monitor antibodies responses of vaccines of Sejroe group serovars. PMID:20332068

  15. Antibodies to Leptospira among blood donors in higher-risk areas of Australia: possible implications for transfusion safety

    PubMed Central

    Faddy, Helen; Seed, Clive; Lau, Colleen; Racloz, Vanessa; Flower, Robert; Smythe, Lee; Burns, Mary-Anne; Dohnt, Michael; Craig, Scott; Harley, Robert; Weinstein, Philip

    2015-01-01

    Background Leptospirosis is one of the most common bacterial zoonoses worldwide, and clinical manifestations range from asymptomatic infection to acute febrile illness, multi-organ failure and death. Asymptomatic, acute bacteraemia in a blood donor provides a potential for transfusion-transmission, although only a single such case from India has been recorded. Human leptospirosis is uncommon in developed countries; however, the state of Queensland in Australia has one of the highest rates among developed countries, especially after increased rainfall. This study examined the prevalence of antibodies to Leptospira spp. in blood donors residing in higher-risk areas of Australia, to evaluate the appropriateness of current blood safety guidelines. Materials and methods Plasma samples collected from blood donors residing in higher-risk areas of Australia during 2009 and 2011 were included in the study. All samples were tested for the presence of antibodies to 22 leptospiral serovars using the microscopic agglutination test. Result No sample had antibody titres suggestive of a current or recent infection, however, seven samples (1.44%, 95% CI: 0.38–2.50%) had titres suggestive of a past infection. Discussion This study provides data that may support the appropriateness of current relevant donor selection policies in Australia. Given that the risk profile for leptospirosis is expanding and that the infection is likely to become more prevalent with climate change, this disease may become more of a concern for transfusion safety in the future. PMID:24960651

  16. A Leptospira borgpetersenii Serovar Hardjo Vaccine Induces a Th1 Response, Activates NK Cells, and Reduces Renal Colonization ▿

    PubMed Central

    Zuerner, Richard L.; Alt, David P.; Palmer, Mitchell V.; Thacker, Tyler C.; Olsen, Steven C.

    2011-01-01

    Chronic infection of cattle with Leptospira borgpetersenii serovar Hardjo reduces animal production through reproductive failure and presents a persistent health threat to workers in the animal industry. Cattle are maintenance hosts for serovar Hardjo, and development of vaccines that establish long-term protective immunity has been problematic; induction of high titers of anti-serovar Hardjo antibody does not appear to be protective. Rather, development of an antigen-specific Th1 response appears to be critical for limiting renal colonization and urinary shedding of bacteria. In this study we compared two monovalent killed bacterial cell vaccines to assess long-term (12 months) protection against live serovar Hardjo challenge. Although neither vaccine prevented infection, renal colonization and urinary shedding of bacteria were reduced compared to those of control animals. Increased proliferation of CD4+, CD8+, and γδ T cells from vaccinated, but not control, animals was detected. In addition, NK cells from vaccinated animals and from all animals following infection, when exposed to antigen ex vivo, demonstrated a gamma interferon (IFN-γ) recall response. We propose that programming NK cells to respond quickly to L. borgpetersenii serovar Hardjo infection may be an important step toward developing protective immunity. PMID:21288995

  17. Seroprevalence of bovine leptospiral antibodies by microscopic agglutination test in Southeast of Iran

    PubMed Central

    Khalili, Mohammad; Sakhaee, Ehsanollah; Aflatoonian, Mohammad Reza; Abdollahpour, Gholamreza; Tabrizi, Saeed Sattari; Damaneh, Elham Mohammadi; Hossini-nasab, Sajad

    2014-01-01

    Objective To evaluate serological findings of bovine leptospirosis which is a zoonotic disease with worldwide distribution caused by Leptospira interrogans. Methods One hundred and sixty seven sera were collected from 9 commercial dairy herds in jiroft suburbs, from July to October 2011. Microscopic agglutination test (MAT) was used to evaluates serological findings of bovine leptospirosis in Jiroft suburb dairy farms, Kerman province, Iran. Results Antibodies were found by MAT at least against one serovar of Leptospira interrogans in 29 samples (17.36%) among 167 sera at a dilution 1:100 or higher, and Leptospira pomona was the most prevalent serovar. Positive titers against more than one serovar were detected in 6 sera of the positive samples. Conclusion This study is the first report of leptospirosis in Southeast Iran and showed that Leptospira pomona was the most and Leptospira icterohaemorrhagiae the least prevalent serovars in Southeast Iran. PMID:25182718

  18. Molecular cloning and sequence analysis of the gene encoding OmpL1, a transmembrane outer membrane protein of pathogenic Leptospira spp.

    PubMed Central

    Haake, D A; Champion, C I; Martinich, C; Shang, E S; Blanco, D R; Miller, J N; Lovett, M A

    1993-01-01

    Pathogenic Leptospira spp. are spirochetes that have a low transmembrane outer membrane protein content relative to that of enteric gram-negative bacteria. In a previous study we identified a 31-kDa surface protein that was present in strains of Leptospira alstoni in amounts which correlated with the outer membrane particle density observed by freeze fracture electron microscopy (D. A. Haake, E. M. Walker, D. R. Blanco, C. A. Bolin, J. N. Miller, and M. A. Lovett, Infect. Immun. 59:1131-1140, 1991). The N-terminal amino acid sequence was used to design a pair of oligonucleotides which were utilized to screen a lambda ZAP II library containing EcoRI fragments of L. alstoni DNA. A 2.5-kb DNA fragment which contained the entire structural ompL1 gene was identified. The structural gene deduced from the sequence of this DNA fragment would encode a 320-amino-acid polypeptide with a 24-amino-acid leader peptide and a leader peptidase I cleavage site. Processing of OmpL1 results in a mature protein with a predicted molecular mass of 31,113 Da. Secondary-structure prediction identified repeated stretches of amphipathic beta-sheets typical of outer membrane protein membrane-spanning sequences. A topological model of OmpL1 containing 10 transmembrane segments is suggested. A recombinant OmpL1 fusion protein was expressed in Escherichia coli in order to immunize rabbits with the purified protein. Upon Triton X-114 extraction of L. alstoni and phase separation, anti-OmpL1 antiserum recognized a single band on immunoblots of the hydrophobic detergent fraction which was not present in the hydrophilic aqueous fraction. Immunoelectron microscopy with anti-OmpL1 antiserum demonstrates binding to the surface of intact L. alstoni. DNA hybridization studies indicate that the ompL1 gene is present in a single copy in all pathogenic Leptospira species that have been tested and is absent in nonpathogenic Leptospira species. OmpL1 may be the first spirochetal transmembrane outer membrane

  19. Separating Putative Pathogens from Background Contamination with Principal Orthogonal Decomposition: Evidence for Leptospira in the Ugandan Neonatal Septisome.

    PubMed

    Schiff, Steven J; Kiwanuka, Julius; Riggio, Gina; Nguyen, Lan; Mu, Kevin; Sproul, Emily; Bazira, Joel; Mwanga-Amumpaire, Juliet; Tumusiime, Dickson; Nyesigire, Eunice; Lwanga, Nkangi; Bogale, Kaleb T; Kapur, Vivek; Broach, James R; Morton, Sarah U; Warf, Benjamin C; Poss, Mary

    2016-01-01

    Neonatal sepsis (NS) is responsible for over 1 million yearly deaths worldwide. In the developing world, NS is often treated without an identified microbial pathogen. Amplicon sequencing of the bacterial 16S rRNA gene can be used to identify organisms that are difficult to detect by routine microbiological methods. However, contaminating bacteria are ubiquitous in both hospital settings and research reagents and must be accounted for to make effective use of these data. In this study, we sequenced the bacterial 16S rRNA gene obtained from blood and cerebrospinal fluid (CSF) of 80 neonates presenting with NS to the Mbarara Regional Hospital in Uganda. Assuming that patterns of background contamination would be independent of pathogenic microorganism DNA, we applied a novel quantitative approach using principal orthogonal decomposition to separate background contamination from potential pathogens in sequencing data. We designed our quantitative approach contrasting blood, CSF, and control specimens and employed a variety of statistical random matrix bootstrap hypotheses to estimate statistical significance. These analyses demonstrate that Leptospira appears present in some infants presenting within 48 h of birth, indicative of infection in utero, and up to 28 days of age, suggesting environmental exposure. This organism cannot be cultured in routine bacteriological settings and is enzootic in the cattle that often live in close proximity to the rural peoples of western Uganda. Our findings demonstrate that statistical approaches to remove background organisms common in 16S sequence data can reveal putative pathogens in small volume biological samples from newborns. This computational analysis thus reveals an important medical finding that has the potential to alter therapy and prevention efforts in a critically ill population. PMID:27379237

  20. Separating Putative Pathogens from Background Contamination with Principal Orthogonal Decomposition: Evidence for Leptospira in the Ugandan Neonatal Septisome

    PubMed Central

    Schiff, Steven J.; Kiwanuka, Julius; Riggio, Gina; Nguyen, Lan; Mu, Kevin; Sproul, Emily; Bazira, Joel; Mwanga-Amumpaire, Juliet; Tumusiime, Dickson; Nyesigire, Eunice; Lwanga, Nkangi; Bogale, Kaleb T.; Kapur, Vivek; Broach, James R.; Morton, Sarah U.; Warf, Benjamin C.; Poss, Mary

    2016-01-01

    Neonatal sepsis (NS) is responsible for over 1 million yearly deaths worldwide. In the developing world, NS is often treated without an identified microbial pathogen. Amplicon sequencing of the bacterial 16S rRNA gene can be used to identify organisms that are difficult to detect by routine microbiological methods. However, contaminating bacteria are ubiquitous in both hospital settings and research reagents and must be accounted for to make effective use of these data. In this study, we sequenced the bacterial 16S rRNA gene obtained from blood and cerebrospinal fluid (CSF) of 80 neonates presenting with NS to the Mbarara Regional Hospital in Uganda. Assuming that patterns of background contamination would be independent of pathogenic microorganism DNA, we applied a novel quantitative approach using principal orthogonal decomposition to separate background contamination from potential pathogens in sequencing data. We designed our quantitative approach contrasting blood, CSF, and control specimens and employed a variety of statistical random matrix bootstrap hypotheses to estimate statistical significance. These analyses demonstrate that Leptospira appears present in some infants presenting within 48 h of birth, indicative of infection in utero, and up to 28 days of age, suggesting environmental exposure. This organism cannot be cultured in routine bacteriological settings and is enzootic in the cattle that often live in close proximity to the rural peoples of western Uganda. Our findings demonstrate that statistical approaches to remove background organisms common in 16S sequence data can reveal putative pathogens in small volume biological samples from newborns. This computational analysis thus reveals an important medical finding that has the potential to alter therapy and prevention efforts in a critically ill population. PMID:27379237

  1. Evaluation of loop-mediated isothermal amplification method (LAMP) for pathogenic Leptospira spp. detection with leptospires isolation and real-time PCR.

    PubMed

    Suwancharoen, Duangjai; Sittiwicheanwong, Busara; Wiratsudakul, Anuwat

    2016-09-01

    Leptospirosis has been one of the worldwide zoonotic diseases caused by pathogenic Leptospira spp. Many molecular techniques have consecutively been developed to detect such pathogen including loop-mediated isothermal amplification method (LAMP). The objectives of this study were to evaluate the diagnostic accuracy of LAMP assay and real-time PCR using bacterial culture as the gold standard and to assess the agreement among these three tests using Cohen's kappa statistics. In total, 533 urine samples were collected from 266 beef and 267 dairy cattle reared in central region of Thailand. Sensitivity and specificity of LAMP were 96.8% (95% CI 81.5-99.8) and 97.0% (95% CI 94.9-98.2), respectively. The accuracy of LAMP (97.0%) was significantly higher than that of real-time PCR (91.9%) at 95% CI. With Cohen's kappa statistics, culture method and LAMP were substantially agreed with each other (77.4%), whereas real-time PCR only moderately agreed with culture (47.7%) and LAMP (45.3%), respectively. Consequently, LAMP was more effective than real-time PCR in detecting Leptospira spp. in the urine of cattle. Besides, LAMP had less cost and was simpler than real-time PCR. Thus, LAMP was an excellent alternative for routine surveillance of leptospirosis in cattle. PMID:27150208

  2. Evaluation of loop-mediated isothermal amplification method (LAMP) for pathogenic Leptospira spp. detection with leptospires isolation and real-time PCR.

    PubMed

    Suwancharoen, Duangjai; Sittiwicheanwong, Busara; Wiratsudakul, Anuwat

    2016-09-01

    Leptospirosis has been one of the worldwide zoonotic diseases caused by pathogenic Leptospira spp. Many molecular techniques have consecutively been developed to detect such pathogen including loop-mediated isothermal amplification method (LAMP). The objectives of this study were to evaluate the diagnostic accuracy of LAMP assay and real-time PCR using bacterial culture as the gold standard and to assess the agreement among these three tests using Cohen's kappa statistics. In total, 533 urine samples were collected from 266 beef and 267 dairy cattle reared in central region of Thailand. Sensitivity and specificity of LAMP were 96.8% (95% CI 81.5-99.8) and 97.0% (95% CI 94.9-98.2), respectively. The accuracy of LAMP (97.0%) was significantly higher than that of real-time PCR (91.9%) at 95% CI. With Cohen's kappa statistics, culture method and LAMP were substantially agreed with each other (77.4%), whereas real-time PCR only moderately agreed with culture (47.7%) and LAMP (45.3%), respectively. Consequently, LAMP was more effective than real-time PCR in detecting Leptospira spp. in the urine of cattle. Besides, LAMP had less cost and was simpler than real-time PCR. Thus, LAMP was an excellent alternative for routine surveillance of leptospirosis in cattle.

  3. Human Leptospirosis on Reunion Island, Indian Ocean: Are Rodents the (Only) Ones to Blame?

    PubMed Central

    Cordonin, Colette; Le Minter, Gildas; Gomard, Yann; Pagès, Frédéric; Jaffar-Bandjee, Marie-Christine; Tortosa, Pablo; Dellagi, Koussay

    2016-01-01

    Background Although leptospirosis is a zoonosis of major concern on tropical islands, the molecular epidemiology of the disease aiming at linking human cases to specific animal reservoirs has been rarely explored within these peculiar ecosystems. Methodology/Principal Findings Five species of wild small mammals (n = 995) as well as domestic animals (n = 101) were screened for Leptospira infection on Reunion Island; positive samples were subsequently genotyped and compared to Leptospira from clinical cases diagnosed in 2012–2013 (n = 66), using MLST analysis. We identified two pathogenic species in human cases, namely Leptospira interrogans and Leptospira borgpetersenii. Leptospira interrogans was by far dominant both in clinical samples (96.6%) and in infected animal samples (95.8%), with Rattus spp and dogs being its exclusive carriers. The genetic diversity within L. interrogans was apparently limited to two sequence types (STs): ST02, identified among most clinical samples and in all rats with complete MLST, and ST34, identified in six humans, but not in rats. Noteworthy, L. interrogans detected in two stray dogs partially matched with ST02 and ST34. Leptospira borgpetersenii was identified in two clinical samples only (3.4%), as well as in cows and mice; four haplotypes were identified, of which two seemingly identical in clinical and animal samples. Leptospira borgpetersenii haplotypes detected in human cases were clearly distinct from the lineage detected so far in the endemic bat species Mormopterus francoismoutoui, thus excluding a role for this volant mammal in the local human epidemiology of the disease. Conclusions/Significance Our data confirm rats as a major reservoir of Leptospira on Reunion Island, but also pinpoint a possible role of dogs, cows and mice in the local epidemiology of human leptospirosis. This study shows that a comprehensive molecular characterization of pathogenic Leptospira in both clinical and animal samples helps to gaining

  4. Serologic, trace element, and fecal parasite survey of free-ranging, female mule deer (Odocoileus hemionus) in eastern Washington, USA.

    PubMed

    Myers, Woodrow L; Foreyt, William J; Talcott, Patricia A; Evermann, James F; Chang, Wan-Ying

    2015-01-01

    Blood and fecal samples collected from 97 free-ranging mule deer (Odocoileus hemionus), from four distinct herds during the spring of 2000 or 2001 in eastern Washington, US, were tested for exposure to selected pathogens, concentrations of trace elements, and presence of parasites in feces. Antibodies were detected to the following: Leptospira interrogans serovar Bratislava (4%), Leptospira interrogans serovar Canicola (1%), Leptospira interrogans serovar Grippotyphosa (13%), Bovine viral diarrhea virus 1 (57%), Bovine respiratory syncytial virus (71%), Infectious bovine rhinotracheitis virus (51%), Bovine parainfluenza virus 3 (61%), Bluetongue virus (25%), and Epizootic hemorrhagic disease virus (25%); 3 of 63 (5%) samples had antibody to Neospora spp. All samples tested for antibody to Brucella abortus and L. interrogans serovar Icterohaemorrhagiae, L. interrogans serovar Pomona, and L. interrogans serovar Hardjo samples were negative. Trace element concentrations from 97 sera were deficient for selenium (17%), copper (19%), iron (34%), calcium (3%), and phosphorus (2%) compared with thresholds established for domestic livestock. Parasites detected in 97 fecal samples included dorsal-spined larvae (probably Parelaphostrongylus sp.) (40%), abomasal nematode eggs (1%), Capillaria sp. eggs (1%), Nematodirus sp. eggs (26%), Moniezia sp. eggs (1%), and Eimeria sp. (2%).

  5. The emergence of Leptospira borgpetersenii serovar Arborea as the dominant infecting serovar following the summer of natural disasters in Queensland, Australia 2011.

    PubMed

    Wynwood, S J; Craig, S B; Graham, G C; Blair, B R; Burns, M A; Weier, S L; Collet, T A; McKay, D B

    2014-06-01

    The following research reports the emergence of Leptospira borgpetersenii serovar Arborea as the dominant infecting serovar following the summer of disasters and the ensuing clean up in Queensland, Australia during 2011. For the 12 month period (1 January to 31 December) L. borgpetersenii serovar Arborea accounted for over 49% of infections. In response to a flooding event public health officials need to issue community wide announcements warning the population about the dangers of leptospirosis and other water borne diseases. Communication with physicians working in the affected community should also be increased to update physicians with information such as clinical presentation of leptospirosis and other waterborne diseases. These recommendations will furnish public health officials with considerations for disease management when dealing with future disaster management programs.

  6. EMJH medium with 5-fluorouracil and nalidixic acid associated with serial dilution technique used to recover Leptospira spp from experimentally contaminated bovine semen.

    PubMed

    Miraglia, Fabiana; de Moraes, Zenaide Maria; Melville, Priscilla Anne; Dias, Ricardo Augusto; Vasconcellos, Silvio Arruda

    2009-01-01

    Bovine semen experimentally contaminated with Leptospira santarosai serovar Guaricura was submitted to the modified EMJH medium with 5-fluorouracil (300mg/L) and nalidixic acid (20mg/L), named as "selective medium" and using the serial dilution technique, in order to evaluate the percentage of recovery of the added microorganism. The selective EMJH medium was found with higher percentage of recovery of leptospiras and minor losses of samples due to contamination with opportunistic microorganisms than the non-selective EMJH medium: 151/376 (40.0%) of positive growth; and 38/376 (10.0%) contamination and 58/376 (15%) and 129/376 (34.0%), respectively. These results were statistically significant (p<0. 0001; Fisher). Differences were found when the frequencies of positive leptospires recovery have been compared in the serial dilution technique (10(-1) to 10(-4)) between the selective and non-selective media at different dilution factors. At 1/10(th) dilution the percentages found were (0%, 0/80) and (38%, 30/80), at 1/100(th) dilution, (3%, 2/80) and (49%, 39/ 80) and at 1/1,000(th) dilution, (25%, 20/80) and (50%, 40/80), respectively. The percentage of recovery of leptospires was found to be directly proportional to the dilution used. The methodology of the serial dilution technique (setting at least three dilutions) and the use of selective EMJH medium have been found to be efficient for the isolation of leptospires from the bovine semen samples.

  7. Monoclonal antibodies against LipL32, the major outer membrane protein of pathogenic Leptospira: production, characterization, and testing in diagnostic applications.

    PubMed

    Fernandes, Cláudia P H; Seixas, Fabiana K; Coutinho, Mariana L; Vasconcellos, Flávia A; Seyffert, Núbia; Croda, Julio; McBride, Alan J; Ko, Albert I; Dellagostin, Odir A; Aleixo, José A G

    2007-02-01

    Pathogenic serovars of Leptospira have a wide antigenic diversity attributed mainly to the lipopolysaccharide present in the outer membrane. In contrast, antigens conserved among pathogenic serovars are mainly represented by outer membrane proteins. Surface exposure of a major and highly conserved outer membrane lipoprotein (LipL32) was recently demonstrated on pathogenic Leptospira. LipL32 in its recombinant form (rLipL32) was used to immunize BALB/c mice to develop murine monoclonal antibodies (MAbs). Three MAbs against rLipL32 were produced, isotyped, and evaluated for further use in diagnostic tests of leptospirosis using different approaches. MAbs were conjugated to peroxidase and evaluated in a native protein enzyme-linked immunosorbent assay (ELISA) with intact and heat-treated leptospiral cells, conjugated to fluorescein isothiocyanate (FITC) for indirect immunofluorescence with intact and methanol fixed cells and were used for LipL32 immunoprecipitation from leptospiral cells. rLipL32 MAbs conjugated to peroxidase or used as primary antibody bound to intact and heat-treated cells in ELISA, proving that they could be used in enzyme immunoassays for detection of the native protein. In immunofluorescence assay, MAbs labeled bacterial cells either intact or methanol fixed. Two MAbs were able to immunoprecipitate the native protein from live and motile leptospiral cells and, adsorbed onto magnetic beads, captured intact bacteria from artificially contaminated human sera for detection by polymerase chain reaction (PCR) amplification. Results of this study suggest that the MAbs produced can be useful for the development of diagnostic tests based on detection of LipL32 leptospiral antigen in biological fluids. PMID:17316084

  8. Evaluation of truncated LipL32 expressed by Escherichia coli and Pichia pastoris for serodiagnosis of Leptospira infection in rodents

    PubMed Central

    SHIOKAWA, Kanae; GAMAGE, Chandika D.; KOIZUMI, Nobuo; SAKODA, Yoshihiro; SHIMIZU, Kenta; TSUDA, Yoshimi; YOSHIMATSU, Kumiko; ARIKAWA, Jiro

    2015-01-01

    The applicability of the recombinant LipL32 for serodiagnosis of leptospiral infection in field rodents was assessed in this study. An immunodominant region of LipL32 was determined by monoclonal antibodies, and then, truncated LipL32 (tLipL32) was designed to contain the region (87–188th amino acid). The tLipL32 was compared between two recombinant expression hosts Escherichia coli and Pichia pastoris in ELISA. With field rat sera, tLipL32 expressed by P. pastoris (tLipL32p) had high antigenicity without background reactions, while tLipL32 expressed by E. coli (tLipL32e) showed high background reactions, which were reduced by pre-adsorption of sera with E. coli. To evaluate tLipL32-ELISA, field rat sera were tentatively divided into a Leptospira infection positive (12 sera) and a negative group (12 sera) based on the results from flaB gene PCR of kidney samples and WB with whole Leptospira cell. Consequently, the sensitivity of tLipL32p-ELISA for field rat sera was 83% . A similar result was obtained from tLipL32e-ELISA with adsorbed sera, (92%). However, sensitivity of tLipL32e-ELISA using sera without an adsorption treatment was 50%. Regardless of the expression host, tLipL32-ELISA had 100% specificity and sensitivity in experimentally infected laboratory rats. These results suggest that recombinant LipL32 expressed by P. pastoris is more applicable for serodiagnosis in field rats due to a lack of background reaction. PMID:26412049

  9. EMJH medium with 5-fluorouracil and nalidixic acid associated with serial dilution technique used to recover Leptospira spp from experimentally contaminated bovine semen

    PubMed Central

    Miraglia, Fabiana; de Moraes, Zenaide Maria; Melville, Priscilla Anne; Dias, Ricardo Augusto; Vasconcellos, Silvio Arruda

    2009-01-01

    Bovine semen experimentally contaminated with Leptospira santarosai serovar Guaricura was submitted to the modified EMJH medium with 5-fluorouracil (300mg/L) and nalidixic acid (20mg/L), named as “selective medium” and using the serial dilution technique, in order to evaluate the percentage of recovery of the added microorganism. The selective EMJH medium was found with higher percentage of recovery of leptospiras and minor losses of samples due to contamination with opportunistic microorganisms than the non-selective EMJH medium: 151/376 (40.0%) of positive growth; and 38/376 (10.0%) contamination and 58/376 (15%) and 129/376 (34.0%), respectively. These results were statistically significant (p<0. 0001; Fisher). Differences were found when the frequencies of positive leptospires recovery have been compared in the serial dilution technique (10-1 to 10-4) between the selective and non-selective media at different dilution factors. At 1/10th dilution the percentages found were (0%, 0/80) and (38%, 30/80), at 1/100th dilution, (3%, 2/80) and (49%, 39/ 80) and at 1/1,000th dilution, (25%, 20/80) and (50%, 40/80), respectively. The percentage of recovery of leptospires was found to be directly proportional to the dilution used. The methodology of the serial dilution technique (setting at least three dilutions) and the use of selective EMJH medium have been found to be efficient for the isolation of leptospires from the bovine semen samples. PMID:24031342

  10. Seroepidemiologic study of three zoonoses (leptospirosis, Q fever, and tularemia) among trappers in Québec, Canada.

    PubMed Central

    Lévesque, B; De Serres, G; Higgins, R; D'Halewyn, M A; Artsob, H; Grondin, J; Major, M; Garvie, M; Duval, B

    1995-01-01

    This study was undertaken to evaluate the prevalence of antibodies against Francisella tularensis, Coxiella burnetii, and certain serovars of Leptospira interrogans among trappers in Québec, Canada. Muskrat trapping was identified as a risk factor for F. tularensis infection, whereas having a cat at home apparently protected trappers against infection by L. interrogans. High percentages of control sera were positive for antibodies against C. burnetii (15%) and L. interrogans (5%), most frequently serovar bratislava. This is the first report of human infection by serovar bratislava in North America. PMID:7583933

  11. Production of reactive oxygen (H2O2) and nitrogen (NO) intermediates and tnf-α in mice genetically selected for high (H) and low (L) antibody response and experimentally infected with Leptospira serovar pomona

    PubMed Central

    Haanwinckel, Maria Cristina Santos; de Oliveira, Silvio Luis

    2011-01-01

    The aim of the present study was to evaluate the activity of macrophages, and the production of TNF-α and antibodies against experimental infection by Leptospira serovar Pomona in mice genetically selected for High (H) or Low (L) humoral immune response. To evaluate macrophagic activity, peritoneal and splenic lavages were performed for determination of oxygen (H2O2) and nitrogen (NO) intermediates. The production of the tumor necrosis factor (TNF-α) was investigated through bioassays in serum and homogenates of splenic and hepatic cells of control and infected animals, as was as specific antibodies production. The immune response against serovar Pomona in those lines, was characterized by high antibody production, especially in later periods of the infectious process, whereas values of bacterial recovery in culture medium were lower. The production of reactives oxygen and nitrogen intermediate, also helped to eliminate Leptospira Pomona in both lines; H2O2 production an important factor in HIV-A, as well as NO production in LIV-A, especially in later post-inoculation periods. The same was detected for TNF-α. Results suggest that such lines could be an important model to investigate the pathogenesis and the immune response of animals against the several Leptospira serovars. PMID:24031688

  12. Leptospirosis in raccoons in Quebec: 2 case reports and seroprevalence in a recreational area.

    PubMed Central

    Mikaelian, I; Higgins, R; Lequient, M; Major, M; Lefebvre, F; Martineau, D

    1997-01-01

    Raccoons may represent a source of leptospires for humans and domestic animals. We describe a case of severe interstitial nephritis associated with the serovar bratislava of Leptospira interrogans (1st report in wildlife), and the seroprevalence to 4 leptospire serovars in a recreational area in Quebec. Images Figure 1. p442-a PMID:9220134

  13. NMR Solution Structure of the Terminal Immunoglobulin-like Domain from the Leptospira Host-Interacting Outer Membrane Protein, LigB

    PubMed Central

    2015-01-01

    A number of surface proteins specific to pathogenic strains of Leptospira have been identified. The Lig protein family has shown promise as a marker in typing leptospiral isolates for pathogenesis and as an antigen in vaccines. We used NMR spectroscopy to solve the solution structure of the twelfth immunoglobulin-like (Ig-like) repeat domain from LigB (LigB-12). The fold is similar to that of other bacterial Ig-like domains and comprised mainly of β-strands that form a β-sandwich based on a Greek-key folding arrangement. Based on sequence analysis and conservation of structurally important residues, homology models for the other LigB Ig-like domains were generated. The set of LigB models illustrates the electrostatic differences between the domains as well as the possible interactions between neighboring domains. Understanding the structure of the extracellular portion of LigB and related proteins is important for developing diagnostic methods and new therapeutics directed toward leptospirosis. PMID:25068811

  14. A cross sectional study of leptospirosis and fetal death in Yucatan, Mexico

    PubMed Central

    Vado-Solis, Ignacio; Pérez-Osorio, Carlos; Peniche-Lara, Gaspar; Segura-Correa, José

    2016-01-01

    Introduction: Leptospirosis is a zoonotic disease affecting mainly to low income human population. Acute leptospiral infection during pregnancy has been associated with spontaneous abortion and fetal death during the first trimester and the abortion may occur as consequence of systemic failure. Objective: To estimate the frequency of Leptospira interrogans infection in women with spontaneous abortion in the state of Yucatan, Mexico. Methods: A cross sectional study on women with spontaneous abortion was conducted. Serum samples were tested for Leptospirosis by the microaglutination test, to estimate the frequency of the infecting serovar. The indirect ELISA IgM was used to detect recent infection by L. interrogans. DNA was extracted from paraffin-embedded tissue of placenta for PCR detection of L. interrogans. Results: Overall frequency of infection with L. interrogans in the 81 women with abortion was 13.6%. Five of the 12 serovars evaluated were found and included. Two of the 11 women with abortion and positive to microaglutination test were also positive to the ELISA IgM test. None samples were positive for PCR Leptospira diagnosis. Conclusion: two women could be associated with spontaneous abortion due to leptospirosis, because they showed antibodies against L. interrogans in the microaglutination test and ELISA IgM assays. Differences between regions were found with respect to the prevalences of lesptospirosis. PMID:27226658

  15. Leptospirosis: clinical presentation and correlation with serovars.

    PubMed

    Chidambaram, N; Ramanathan, M; Anandi, V; Sasikala, S; Innocent, D J P; Sarayu, L

    2007-06-01

    A total of 2400 patients with pyrexia of unknown origin and or suspected leptospirosis were included in this study. Dark field microscopy detected Leptospira in 690 cases, Leptospira serological Investigations proved positive in 570 out of these 690 patients. Among them 212 had the classical icteric and the other 358 had anicteric type of presentation. Notably eptospira interrogans serovar ictero haemorrhagiae infection was encountered in 212 patients. In 30 patients, who had multi organ dysfunction which included renal failure, hepatic dysfunction or meningitis was due to Leptospira interrogans Serovar cannicola. Coexsistense of leptospirosis and hepatitis B virus infection were noted in 15 patients. Antibody to Leptospira interrogans was demonstrated by Micro agglutination test (MAT) in addition to dark field microscopy positivity in these cases. Similarly HIV antibody was demonstrated in 30 of the 330 anicteric patients. 554 out of 570 cases responded to intra venous penicillin (216), and oral Doxycycline (182) and Augmentin (156), and the remaining 16 patients succumbed to death. PMID:18338689

  16. An outbreak of severe leptospirosis in capuchin (Cebus) monkeys

    PubMed Central

    Szonyi, Barbara; Agudelo-Flórez, Piedad; Ramírez, Marcela; Moreno, Natali; Ko, Albert I.

    2013-01-01

    Naturally acquired acute leptospirosis in monkeys is uncommon. This study reports an outbreak of severe leptospirosis among 52 capuchin (Cebus) monkeys that had been rescued from homes and housed in a wildlife rehabilitation center in Colombia in 2007. Case confirmation consisted of Leptospira isolation followed by a polymerase chain reaction targeting the LipL32 gene. The attack and mortality rates were 71% and 27%, respectively. Sixteen cases were confirmed. Necropsy revealed diffuse jaundice and pulmonary hemorrhage. Multi-locus sequence typing identified the agent to be Leptospira interrogans sequence type 17, indicating rats as the source of infection. An environmental survey confirmed rodent infestation as the cause of the outbreak. The extent of Leptospira transmission between humans and monkeys is unknown. Improper husbandry of non-human primates could create new reservoirs and transmission routes for Leptospira threatening conservation efforts and public health. PMID:20554228

  17. Serological survey of Bartonella spp., Borrelia burgdorferi, Brucella spp., Coxiella burnetii, Francisella tularensis, Leptospira spp., Echinococcus, Hanta-, TBE- and XMR-virus infection in employees of two forestry enterprises in North Rhine-Westphalia, Germany, 2011-2013.

    PubMed

    Jurke, Annette; Bannert, N; Brehm, K; Fingerle, V; Kempf, V A J; Kömpf, D; Lunemann, M; Mayer-Scholl, A; Niedrig, M; Nöckler, K; Scholz, H; Splettstoesser, W; Tappe, D; Fischer, Silke F

    2015-10-01

    We initiated a survey to collect basic data on the frequency and regional distribution of various zoonoses in 722 employees of forestry enterprises in the German state of North Rhine-Westphalia (NRW) from 2011 to 2013. Exposures associated with seropositivity were identified to give insight into the possible risk factors for infection with each pathogen. 41.2% of participants were found to be seropositive for anti-Bartonella IgG, 30.6% for anti-Borrelia burgdorferi IgG, 14.2% for anti-Leptospira IgG, 6.5% for anti-Coxiella burnetii IgG, 6.0% for anti-Hantavirus IgG, 4.0% for anti-Francisella tularensis IgG, 3.4% for anti-TBE-virus IgG, 1.7% for anti-Echinococcus IgG, 0.0% for anti-Brucella IgG and anti-XMRV IgG. Participants seropositive for B. burgdorferi were 3.96 times more likely to be professional forestry workers (univariable analysis: OR 3.96; 95% CI 2.60-6.04; p<0.001); and participants seropositive for Hantavirus 3.72 times more likely (univariable analysis: OR 3.72; 95% CI 1.44-9.57; p=0.007). This study found a surprisingly high percentage of participants seropositive for anti-B. henselae IgG and for anti-F. tularensis IgG. The relatively high seroprevalence for anti-Leptospira IgG seen in this study could be related to living conditions rather than to exposure at work. No specific risk for exposure to C. burnetii and Echinococcus was identified, indicating that neither forestry workers nor office workers represent a risk population and that NRW is not a typical endemic area. Forestry workers appear to have higher risk for contact with B. burgdorferi-infected ticks and a regionally diverse risk for acquiring Hantavirus-infection. The regional epidemiology of zoonoses is without question of great importance for public health. Knowledge of the regional risk factors facilitates the development of efficient prevention strategies and the implementation of such prevention measures in a sustainable manner.

  18. Six new leptospiral serovars isolated from wild animals in Peru.

    PubMed Central

    Liceras de Hidalgo, J L; Sulzer, K R

    1984-01-01

    Six new serovars of Leptospira interrogans were isolated from opossums (Didelphis marsupialis and Philander opossum) trapped in the Peruvian jungle. The proposed names, type strain designation, and serogroup of the serovars, respectively, were: huallaga, strain M-7, Djasiman serogroup; luis, strain M-6, Tarassovi serogroup; machiguenga, strain MMD-3, Icterohaemorrhagiae serogroup; rioja, strain MR-12, Bataviae serogroup; rupa rupa, strain M-3, Sejroe serogroup; and tingomaria, strain M-13, Cynopteri serogroup. PMID:6470106

  19. Serologic evidence for selected infectious diseases in Marsican brown bears (Ursus arctos marsicanus) in Italy (2004-09).

    PubMed

    Di Francesco, Cristina Esmeralda; Gentile, Leonardo; Di Pirro, Vincenza; Ladiana, Lara; Tagliabue, Silvia; Marsilio, Fulvio

    2015-01-01

    We tested 30 serum samples collected during 2004-09 from 22 free-ranging Marsican brown bears (Ursus arctos marsicanus) in the National Park of Abruzzo, Lazio, and Molise, Italy, for antibodies against canine distemper virus (CDV), canine adenovirus type 2 (CAV-2), canine parvovirus type 2 (CPV-2), Brucella spp., and eight Leptospira interrogans sensu lato serovars. Antibody to CDV was detected in 11 samples (37%); only two bears (10%) had detectable CAV-2 and Brucella spp. antibodies; three bears were positive for L. interrogans serovar Bratislava; and one sample had antibody against L. interrogans serovar Copenhageni. All samples were positive for CPV-2 antibody. The CPV-2 antibody titers varied from 1∶640 to 1∶10,240, suggesting that transmission was still active. Fifty percent of bears were positive for antibody to two or more pathogens. Our results highlight the need to consider infectious diseases as a potential risk for Marsican brown bear conservation.

  20. A survey of rodent-borne pathogens carried by wild Rattus spp. in Northern Vietnam.

    PubMed

    Koma, T; Yoshimatsu, K; Yasuda, S P; Li, T; Amada, T; Shimizu, K; Isozumi, R; Mai, L T Q; Hoa, N T; Nguyen, V; Yamashiro, T; Hasebe, F; Arikawa, J

    2013-09-01

    To examine the prevalence of human pathogens carried by rats in urban areas in Hanoi and Hai Phong, Vietnam, we live-trapped 100 rats in January 2011 and screened them for a panel of bacteria and viruses. Antibodies against Leptospira interrogans (22·0%), Seoul virus (14·0%) and rat hepatitis E virus (23·0%) were detected in rats, but antibodies against Yersinia pestis were not detected. Antibodies against L. interrogans and Seoul virus were found only in adult rats. In contrast, antibodies to rat hepatitis E virus were also found in juvenile and sub-adult rats, indicating that the transmission mode of rat hepatitis E virus is different from that of L. interrogans and Seoul virus. Moreover, phylogenetic analyses of the S and M segments of Seoul viruses found in Rattus norvegicus showed that Seoul viruses from Hai Phong and Hanoi formed different clades. Human exposure to these pathogens has become a significant public health concern.

  1. Serologic survey for cross-species pathogens in urban coyotes (Canis latrans), Colorado, USA.

    PubMed

    Malmlov, Ashley; Breck, Stewart; Fry, Tricia; Duncan, Colleen

    2014-10-01

    Abstract As coyotes (Canis latrans) adapt to living in urban environments, the opportunity for cross-species transmission of pathogens may increase. We investigated the prevalence of antibodies to pathogens that are either zoonotic or affect multiple animal species in urban coyotes in the Denver metropolitan area, Colorado, USA, in 2012. We assayed for antibodies to canine parvovirus-2, canine distemper virus, rabies virus, Toxoplasma gondii, Yersinia pestis, and serotypes of Leptospira interrogans. Overall, 84% of the animals had antibodies to canine parvovirus-2, 44% for canine distemper virus, 20% for T. gondii (IgG), 28% for Y. pestis, and 4% for L. interrogans serotype Grippotyphosa. No neutralizing antibodies were detected to rabies virus, T. gondii (IgM), or L. interrogans serotypes other than Grippotyphosa. With 88% of animals exposed to at least one pathogen, our results suggest that coyotes may serve as important reservoirs and sentinels for etiologic agents.

  2. Seroepidemiology of leptospirosis in Minnesota wolves

    USGS Publications Warehouse

    Khan, M.A.; Goyal, S.M.; Diesch, S.L.; Mech, L.D.; Fritts, S.H.

    1991-01-01

    Serum samples (n = 457) from wolves (Canis lupus) in northern Minnesota were collected from 1972 through 1986 and were tested for antibodies against Leptospira interrogans using a microtiter agglutination test. Twelve serovars included in the study were: australis, autumnalis, ballum, bataviae, bratislava, canicola, copenhageni, grippotyphosa, hardjo, pomona, pyrogenes, and tarassovi. Fifty-two (11%) sera had antibody titers of greater than or equal to 1:50 against one or more serovars of L. interrogans. The seroprevalence of different serovars in decreasing order was: grippotyphosa, bratislava, autumnalis, canicola, pomona, ballum, pyrogenes, hardjo, and copenhageni. No antibodies were found against australis, bataviae, and tarassovi. These results indicate that L. interrogans infection may occur in wolves of Minnesota.

  3. Serologic survey for cross-species pathogens in urban coyotes (Canis latrans), Colorado, USA.

    PubMed

    Malmlov, Ashley; Breck, Stewart; Fry, Tricia; Duncan, Colleen

    2014-10-01

    Abstract As coyotes (Canis latrans) adapt to living in urban environments, the opportunity for cross-species transmission of pathogens may increase. We investigated the prevalence of antibodies to pathogens that are either zoonotic or affect multiple animal species in urban coyotes in the Denver metropolitan area, Colorado, USA, in 2012. We assayed for antibodies to canine parvovirus-2, canine distemper virus, rabies virus, Toxoplasma gondii, Yersinia pestis, and serotypes of Leptospira interrogans. Overall, 84% of the animals had antibodies to canine parvovirus-2, 44% for canine distemper virus, 20% for T. gondii (IgG), 28% for Y. pestis, and 4% for L. interrogans serotype Grippotyphosa. No neutralizing antibodies were detected to rabies virus, T. gondii (IgM), or L. interrogans serotypes other than Grippotyphosa. With 88% of animals exposed to at least one pathogen, our results suggest that coyotes may serve as important reservoirs and sentinels for etiologic agents. PMID:25121408

  4. Pulsed-field gel electrophoretic analysis of leptospiral DNA.

    PubMed Central

    Taylor, K A; Barbour, A G; Thomas, D D

    1991-01-01

    The genomic structures of spirochete species are not well characterized, and genetic studies on these organisms have been hampered by lack of a genetic exchange mechanism in these bacteria. In view of these observations, pulsed-field gel electrophoresis was used to examine the genomes of Leptospira species. Live cells, prepared in agarose plugs, were lysed in situ, and the DNA was analyzed under different electrophoretic conditions. Pulsed-field gel electrophoresis of DNA digested with infrequently cutting restriction enzymes showed that the genome of Leptospira interrogans serovar canicola is approximately 3.1 Mb, while that of the saprophytic L. biflexa serovar patoc I is 3.5 Mb. DNA forms of approximately 2,000 and 350 kb which were present in samples from L. interrogans serovars were not readily detected in nonpathogenic serovars. Three distinct populations, designated type alpha, beta, and gamma, of L. interrogans DNA molecules were further analyzed with two-dimensional gel electrophoresis. Evidence suggested that two of these DNA forms, type alpha and gamma, were linear structures. Pulsed-field gel electrophoresis has proven to be a valuable tool with which to size bacterial genomes and to take the first steps toward characterization of a form of leptospiral DNA which behaves as a linear molecule and which may be related to the virulence of L. interrogans. Images PMID:1987046

  5. SEROLOGIC SURVEY AND RESULTS OF URINARY PCR TESTING FOR LEPTOSPIROSIS IN CAPTIVE BLACK-TAILED PRAIRIE DOGS (CYNOMYS LUDOVICIANUS).

    PubMed

    Olds, June E; Sun, Yaxuan; Baum, David H; Gauger, Phillip

    2015-12-01

    Leptospirosis is an important zoonotic disease occurring clinically and subclinically in humans and a wide variety of mammal species worldwide. Often, rodents and wild animals are identified as important reservoirs for the disease. Twenty-two captive black-tailed prairie dogs (Cynomys ludovicianus) housed within a zoo were examined as part of a routine census and preventive medicine program. During examinations, blood and urine were collected to screen for exposure to, or infection with, leptospirosis. All animals were apparently healthy at the time of examination. Leptospira microscopic agglutination test identified 12 of 22 (54.5%) prairie dogs with antibody titers ≥1 : 100 against Leptospira interrogans serovar bratislava on initial serologic examination. All prairie dogs within this collection were serologically negative for L. interrogans serovars canicola, hardjo, icterohaemorrhagiae, and pomona and Leptospira kirschneri serovar grippotyphosa. Leptospira polymerase chain reaction (PCR) testing of urine was negative in all animals tested. This report describes evidence that captive prairie dogs may be exposed to leptospirosis, most likely from wild rodent reservoirs; however, serum titers are low, and lack of leptospiral DNA detected by PCR indicates that these captive animals are unlikely to be important reservoirs for the disease. PMID:26667541

  6. Antileptospiral activity of xanthones from Garcinia mangostana and synergy of gamma-mangostin with penicillin G

    PubMed Central

    2013-01-01

    Background Leptospirosis, one of the most widespread zoonotic infectious diseases worldwide, is caused by spirochetes bacteria of the genus Leptospira. The present study examined inhibitory activity of purified xanthones and crude extracts from Garcinia mangostana against both non-pathogenic and pathogenic leptospira. Synergy between γ-mangostin and penicillin G against leptospires was also determined. Methods Minimal inhibitory concentrations (MIC) of crude extracts and purified xanthones from G. mangostana and penicillin G for a non-pathogenic (L. biflexa serovar Patoc) and pathogenic (L. interrogans serovar Bataviae, Autumnalis, Javanica and Saigon) leptospires were determined by using broth microdilution method and alamar blue. The synergy was evaluated by calculating the fractional inhibitory concentration (FIC) index. Results The results of broth microdilution test demonstrated that the crude extract and purified xanthones from mangosteen possessed antileptospiral activities. The crude extracts were active against all five serovars of test leptospira with MICs ranging from 200 to ≥ 800 μg/ml. Among the crude extracts and purified xanthones, garcinone C was the most active compound against both of pathogenic (MIC =100 μg/ml) and non-pathogenic leptospira (MIC = 200 μg/ml). However, these MIC values were higher than those of traditional antibiotics. Combinations of γ-mangostin with penicillin G generated synergistic effect against L. interrogans serovars Bataviae, Autumnalis and Javanica (FIC = 0.52, 0.50, and 0.04, respectively) and no interaction against L. biflexa serovar Patoc (FIC =0.75). However, antagonistic activity (FIC = 4.03) was observed in L. interrogans serovar Saigon. Conclusions Crude extracts and purified xanthones from fruit pericarp of G. mangostana with significant antibacterial activity may be used to control leptospirosis. The combination of xanthone with antibiotic enhances the antileptospiral efficacy. PMID

  7. Leptospira Immunoglobulin-Like Protein B (LigB) Binds to Both the C-Terminal 23 Amino Acids of Fibrinogen αC Domain and Factor XIII: Insight into the Mechanism of LigB-Mediated Blockage of Fibrinogen α Chain Cross-Linking

    PubMed Central

    Hsieh, Ching-Lin; Chang, Eric; Tseng, Andrew; Ptak, Christopher; Wu, Li-Chen; Su, Chun-Li; McDonough, Sean P.; Lin, Yi-Pin; Chang, Yung-Fu

    2016-01-01

    The coagulation system provides a primitive but effective defense against hemorrhage. Soluble fibrinogen (Fg) monomers, composed of α, β and γ chains, are recruited to provide structural support for the formation of a hemostatic plug. Fg binds to platelets and is processed into a cross-linked fibrin polymer by the enzymatic clotting factors, thrombin and Factor XIII (FXIII). The newly formed fibrin-platelet clot can act as barrier to protect against pathogens from entering the bloodstream. Further, injuries caused by bacterial infections can be confined to the initial wound site. Many pathogenic bacteria have Fg-binding adhesins that can circumvent the coagulation pathway and allow the bacteria to sidestep containment. Fg expression is upregulated during lung infection providing an attachment surface for bacteria with the ability to produce Fg-binding adhesins. Fg binding by leptospira might play a crucial factor in Leptospira-associated pulmonary hemorrhage, the main factor contributing to lethality in severe cases of leptospirosis. The 12th domain of Leptospira immunoglobulin-like protein B (LigB12), a leptospiral adhesin, interacts with the C-terminus of FgαC (FgαCC). In this study, the binding site for LigB12 was mapped to the final 23 amino acids at the C-terminal end of FgαCC (FgαCC8). The association of FgαCC8 with LigB12 (ELISA, KD = 0.76 μM; SPR, KD = 0.96 μM) was reduced by mutations of both charged residues (R608, R611 and H614 from FgαCC8; D1061 from LigB12) and hydrophobic residues (I613 from FgαCC8; F1054 and A1065 from LigB12). Additionally, LigB12 bound strongly to FXIII and also inhibited fibrin formation, suggesting that LigB can disrupt coagulation by suppressing FXIII activity. Here, the detailed binding mechanism of a leptospiral adhesin to a host hemostatic factor is characterized for the first time and should provide better insight into the pathogenesis of leptospirosis. PMID:27622634

  8. Leptospira Immunoglobulin-Like Protein B (LigB) Binds to Both the C-Terminal 23 Amino Acids of Fibrinogen αC Domain and Factor XIII: Insight into the Mechanism of LigB-Mediated Blockage of Fibrinogen α Chain Cross-Linking.

    PubMed

    Hsieh, Ching-Lin; Chang, Eric; Tseng, Andrew; Ptak, Christopher; Wu, Li-Chen; Su, Chun-Li; McDonough, Sean P; Lin, Yi-Pin; Chang, Yung-Fu

    2016-09-01

    The coagulation system provides a primitive but effective defense against hemorrhage. Soluble fibrinogen (Fg) monomers, composed of α, β and γ chains, are recruited to provide structural support for the formation of a hemostatic plug. Fg binds to platelets and is processed into a cross-linked fibrin polymer by the enzymatic clotting factors, thrombin and Factor XIII (FXIII). The newly formed fibrin-platelet clot can act as barrier to protect against pathogens from entering the bloodstream. Further, injuries caused by bacterial infections can be confined to the initial wound site. Many pathogenic bacteria have Fg-binding adhesins that can circumvent the coagulation pathway and allow the bacteria to sidestep containment. Fg expression is upregulated during lung infection providing an attachment surface for bacteria with the ability to produce Fg-binding adhesins. Fg binding by leptospira might play a crucial factor in Leptospira-associated pulmonary hemorrhage, the main factor contributing to lethality in severe cases of leptospirosis. The 12th domain of Leptospira immunoglobulin-like protein B (LigB12), a leptospiral adhesin, interacts with the C-terminus of FgαC (FgαCC). In this study, the binding site for LigB12 was mapped to the final 23 amino acids at the C-terminal end of FgαCC (FgαCC8). The association of FgαCC8 with LigB12 (ELISA, KD = 0.76 μM; SPR, KD = 0.96 μM) was reduced by mutations of both charged residues (R608, R611 and H614 from FgαCC8; D1061 from LigB12) and hydrophobic residues (I613 from FgαCC8; F1054 and A1065 from LigB12). Additionally, LigB12 bound strongly to FXIII and also inhibited fibrin formation, suggesting that LigB can disrupt coagulation by suppressing FXIII activity. Here, the detailed binding mechanism of a leptospiral adhesin to a host hemostatic factor is characterized for the first time and should provide better insight into the pathogenesis of leptospirosis. PMID:27622634

  9. Leptospirosis and tularaemia in raccoons (Procyon lotor) of Larimer County, [corrected] Colorado.

    PubMed

    Duncan, C; Krafsur, G; Podell, B; Baeten, L A; LeVan, I; Charles, B; Ehrhart, E J

    2012-02-01

    Raccoons (Procyon lotor) are commonly implicated as carriers of many zoonotic pathogens. The purpose of this cross-sectional study was to look for Leptospira interrogans and Francisella tularensis in opportunistically sampled, free-ranging raccoons of Larimer Country, Colorado, USA. Sixty-five animals were included in the study and testing consisted of gross post-mortem examination, histopathology, and both immunohistochemistry and PCR for L. interrogans and F. tularensis. No significant gross lesions were identified and the most common histological lesions were lymphoplasmacytic interstitial nephritis and pulmonary silicosis; rare periportal hepatitis, splenic lymphoid hyperplasia and small pulmonary granulomas were also identified. Of 65 animals, 20 (30%) were positive for Leptospira on IHC but only one by PCR. Animals with inflammation in their kidneys were seven times more likely to be positive for Leptospira than animals without inflammation. The severity of inflammation was variable but often mild with minimal associated renal pathology. One animal was positive for Francisella on both IHC and PCR; IHC staining was localized to histiocytic cells within a pulmonary granuloma. In Colorado the significance and epidemiology of Leptospira is poorly understood. The high prevalence of infection in raccoons in this study population suggests that this species may be important in the regional epidemiology or could be used to estimate risk to domestic animals and humans. Identification of a single Francisella positive animal is significant as this is an uncommon disease in terrestrial animals within the state; the apparently higher prevalence in this peridomestic species implies that raccoons may be good indicators of the pathogen in the region. The results of this study suggest that raccoons may serve as effective sentinels for both Leptospira and Francisella in the state of Colorado. Further studies are needed to better characterize the prevalence and epidemiology of

  10. Detección y estudio mediante Fluorescencia Inducida por Láser de radicales libres formados por Disociación Multifotónica Infrarroja

    NASA Astrophysics Data System (ADS)

    Santos, M.; Díaz, L.; Torresano, J. A.; Rubio, L.; Samoudi, B.

    Una de las principales aplicaciones actuales de los procesos de disociación multifotónica inducidos por radiación láser infrarroja (DMI) es la producción de radiales libres, con el fin de estudiar sus propiedades cinéticas y espectroscópicas. La disociación de moléculas poliatómicas en el IR con láseres de CO2 tiene lugar desde la superficie de energía molecular mas baja y conduce generalmente a la formación de fragmentos en el estado electrónico fundamental, con diversos grados de excitación vibracional. En el Grupo de Procesos Multifotónicos del Instituto de Estructura de la Materia del C.S.I.C. hemos puesto a punto la técnica de Fluorescencia Inducida por Láser (LIF) para la detección y análisis en tiempo real de los fragmentos producidos en la DMI inducida mediante uno o dos campos láseres de diferentes longitudes de onda. Objetivos de nuestro trabajo han sido el estudio de los canales de disociación mayoritarios y de las especies transitoria producidas, así como de la distribución de energía interna con que éstas son generadas. En particular hemos detectado mediante LIF las especies: C2, CF, CH, SiH2, CF2, CH2, SiHCl, y CF3 a partir de la disociación de, entre otras, las siguientes moléculas: C2H3Br, C3F6, C4H8Si, C2H5ClSi y CH5ClSi. En este trabajo presentamos algunos de los resultados obtenidos mediante el estudio por LIF de estos radicales: estudio temporal de la señal LIF obtenida con determinación de tiempos de vida, espectros de excitación y fluorescencia, temperaturas vibracionales de formación, variación de la intensidad LIF con el tiempo de retraso entre los láseres de disociación y prueba, etc.

  11. Serological prevalence to six leptospiral serovars in cattle in Asturias (Northern Spain).

    PubMed Central

    Espi, A.; Prieto, J. M.; Fernandez, M.; Alvarez, M.

    2000-01-01

    The prevalence of antibody to six serovars of Leptospira interrogans in cattle in Asturias (Northern Spain) was determined by the microscopic agglutination test (MAT). Using 50% agglutination or lysis at a dilution of 1:10 or more as the criteria for seropositivity, 371 of 3578 (10.36%) animals were found to react with one serovar. The most commonly detected serovars were pomona (5.59%) and grippotyphosa (2.37%), whilst serovar hardjo (0.75%), icterohaemorrhagiae (0.64%), poi (0.64%) and autumnalis (0.36%) were found at lower frequencies. PMID:10982084

  12. A 23-year-old Man with Leptospirosis and Acute Abdominal Pain

    PubMed Central

    Mazhar, Momal; Kao, Janet J

    2016-01-01

    Leptospirosis is a zoonosis caused by the spirochete Leptospira interrogans. Most cases of leptospirosis are mild to moderate, and self-limited. The course of disease, however, may be complicated by multiorgan dysfunction such as in Weil's disease. We present a case of Weil's disease with pancreatitis in a young Caucasian man residing in Hawai‘i. Although leptospirosis is common in Hawai‘i, few patients present with pancreatitis. This report of leptospirosis-induced pancreatitis should help raise awareness of clinicians to assess for pancreatitis when evaluating a patient with leptospirosis and acute abdominal pain. PMID:27738562

  13. [Detection of leptospirosis reservoirs in Madagascar using the polymerase chain reaction technique].

    PubMed

    Ralaiarijaona, R L; Bellenger, E; Chanteau, S; Roger, F; Pérolat, P; Rasolofo Razanamparany, V

    2001-01-01

    A polymerase chain reaction (PCR) technique was used for detection of the Leptospira interrogans rrs gene in kidney tissue from 115 rats, 50 zebu cattles and 13 pigs in an attempt to identify a possible animal reservoir of leptospirosis in Madagascar. In addition, serological testing of 105 individuals in close contact with animals was carried out. The PCR analysis was negative for all the samples tested and only one person was found seropositive at a low titer. The findings suggest that leptospirosis, if prevalent in Madagascar, is likely rare.

  14. Decay-Accelerating Factor 1 Deficiency Exacerbates Leptospiral-Induced Murine Chronic Nephritis and Renal Fibrosis

    PubMed Central

    Ferrer, María F.; Scharrig, Emilia; Alberdi, Lucrecia; Cedola, Maia; Pretre, Gabriela; Drut, Ricardo; Song, Wen-Chao; Gomez, Ricardo M.

    2014-01-01

    Leptospirosis is a global zoonosis caused by pathogenic Leptospira, which can colonize the proximal renal tubules and persist for long periods in the kidneys of infected hosts. Here, we characterized the infection of C57BL/6J wild-type and Daf1−/− mice, which have an enhanced host response, with a virulent Leptospira interrogans strain at 14 days post-infection, its persistence in the kidney, and its link to kidney fibrosis at 90 days post-infection. We found that Leptospira interrogans can induce acute moderate nephritis in wild-type mice and is able to persist in some animals, inducing fibrosis in the absence of mortality. In contrast, Daf1−/− mice showed acute mortality, with a higher bacterial burden. At the chronic stage, Daf1−/− mice showed greater inflammation and fibrosis than at 14 days post-infection and higher levels at all times than the wild-type counterpart. Compared with uninfected mice, infected wild-type mice showed higher levels of IL-4, IL-10 and IL-13, with similar levels of α-smooth muscle actin, galectin-3, TGF-β1, IL-17, IFN-γ, and lower IL-12 levels at 90 days post-infection. In contrast, fibrosis in Daf1−/− mice was accompanied by high expression of α-smooth muscle actin, galectin-3, IL-10, IL-13, and IFN-γ, similar levels of TGF-β1, IL-12, and IL-17 and lower IL-4 levels. This study demonstrates the link between Leptospira-induced murine chronic nephritis with renal fibrosis and shows a protective role of Daf1. PMID:25032961

  15. Decay-accelerating factor 1 deficiency exacerbates leptospiral-induced murine chronic nephritis and renal fibrosis.

    PubMed

    Ferrer, María F; Scharrig, Emilia; Alberdi, Lucrecia; Cedola, Maia; Pretre, Gabriela; Drut, Ricardo; Song, Wen-Chao; Gomez, Ricardo M

    2014-01-01

    Leptospirosis is a global zoonosis caused by pathogenic Leptospira, which can colonize the proximal renal tubules and persist for long periods in the kidneys of infected hosts. Here, we characterized the infection of C57BL/6J wild-type and Daf1-/- mice, which have an enhanced host response, with a virulent Leptospira interrogans strain at 14 days post-infection, its persistence in the kidney, and its link to kidney fibrosis at 90 days post-infection. We found that Leptospira interrogans can induce acute moderate nephritis in wild-type mice and is able to persist in some animals, inducing fibrosis in the absence of mortality. In contrast, Daf1-/- mice showed acute mortality, with a higher bacterial burden. At the chronic stage, Daf1-/- mice showed greater inflammation and fibrosis than at 14 days post-infection and higher levels at all times than the wild-type counterpart. Compared with uninfected mice, infected wild-type mice showed higher levels of IL-4, IL-10 and IL-13, with similar levels of α-smooth muscle actin, galectin-3, TGF-β1, IL-17, IFN-γ, and lower IL-12 levels at 90 days post-infection. In contrast, fibrosis in Daf1-/- mice was accompanied by high expression of α-smooth muscle actin, galectin-3, IL-10, IL-13, and IFN-γ, similar levels of TGF-β1, IL-12, and IL-17 and lower IL-4 levels. This study demonstrates the link between Leptospira-induced murine chronic nephritis with renal fibrosis and shows a protective role of Daf1.

  16. Coexistence and survival of pathogenic leptospires by formation of biofilm with Azospirillum.

    PubMed

    Kumar, K Vinod; Lall, Chandan; Raj, R Vimal; Vedhagiri, K; Vijayachari, P

    2015-06-01

    Pathogenic Leptospira spp. represent one cause of leptospirosis worldwide and have long been regarded as solitary organisms in soil and aquatic environments. However, in the present study, Leptospira interrogans was observed to be associated with environmental biofilms with 21 bacterial isolates belonging to 10 genera. All 21 isolates were examined for their coaggregation and biofilm-forming ability with leptospires in vitro. Among these, Azospirillum brasilense RMRCPB showed maximum interspecies coaggregation with leptospiral strains (>75%, visual score of +4). Other significant coaggregating isolates belonged to the genera Sphingomonas, Micrococcus, Brevundimonas, Acinetobacter and Paracoccus. Biofilms of leptospires in combination with A. brasilense RMRCPB showed high resistance to penicillin G, ampicillin and tetracycline (minimum bactericidal concentration ≥800 μg/mL) and tolerance to UV radiation and high temperature (up to 49°C). This study hypothesized that biofilm formation with A. brasilense protects the pathogenic Leptospira from adverse environmental conditions/stress. This coexistence of pathogenic Leptospira with other bacteria may be the key factor for its persistence and survival. However, the mechanism of biofilm formation by leptospires needs to be explored to help devise an appropriate control strategy and reduce transmission of leptospires. PMID:25962762

  17. Epidemiology of Leptospirosis in Africa: A Systematic Review of a Neglected Zoonosis and a Paradigm for ‘One Health’ in Africa

    PubMed Central

    Allan, Kathryn J.; Biggs, Holly M.; Halliday, Jo E. B.; Kazwala, Rudovick R.; Maro, Venance P.; Cleaveland, Sarah; Crump, John A.

    2015-01-01

    Background Leptospirosis is an important but neglected bacterial zoonosis that has been largely overlooked in Africa. In this systematic review, we aimed to summarise and compare current knowledge of: (1) the geographic distribution, prevalence, incidence and diversity of acute human leptospirosis in Africa; and (2) the geographic distribution, host range, prevalence and diversity of Leptospira spp. infection in animal hosts in Africa. Methods Following Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines, we searched for studies that described (1) acute human leptospirosis and (2) pathogenic Leptospira spp. infection in animals. We performed a literature search using eight international and regional databases for English and non-English articles published between January 1930 to October 2014 that met out pre-defined inclusion criteria and strict case definitions. Results and Discussion We identified 97 studies that described acute human leptospirosis (n = 46) or animal Leptospira infection (n = 51) in 26 African countries. The prevalence of acute human leptospirosis ranged from 2 3% to 19 8% (n = 11) in hospital patients with febrile illness. Incidence estimates were largely restricted to the Indian Ocean islands (3 to 101 cases per 100,000 per year (n = 6)). Data from Tanzania indicate that human disease incidence is also high in mainland Africa (75 to 102 cases per 100,000 per year). Three major species (Leptospira borgpetersenii, L. interrogans and L. kirschneri) are predominant in reports from Africa and isolates from a diverse range of serogroups have been reported in human and animal infections. Cattle appear to be important hosts of a large number of Leptospira serogroups in Africa, but few data are available to allow comparison of Leptospira infection in linked human and animal populations. We advocate a ‘One Health’ approach to promote multidisciplinary research efforts to improve understanding of the animal to human

  18. Leptospirosis in pigs, dogs, rodents, humans, and water in an area of the Colombian tropics.

    PubMed

    Calderón, Alfonso; Rodríguez, Virginia; Máttar, Salim; Arrieta, Germán

    2014-02-01

    Leptospirosis is a reemerging zoonosis of global distribution and is one of the causes of hemorrhagic fevers in the tropics. We sought to determine seroprevalence in humans and animals and isolate Leptospira interrogans sensu lato in domestic animals, rodents, and water sources. The study was conducted in a tropical area of the middle Sinú in Cordoba, Colombia. In a prospective descriptive study, we collected blood and urine from pigs and dogs, sera from rural human workers, sera and kidney macerates of rodents, and water samples from environmental sources. We used microagglutination to screen for antibodies to 13 serovars. Strains were cultured on the Ellinghausen-McCullough-Johnson-Harris medium and confirmed by PCR amplifying lipL32 gene. Seroprevalence was 55.9% in pigs, 35.2% in dogs, and 75.8% in humans; no antibody was detected, and no Leptospira were isolated from kidney macerates of rodents. Seven L. interrogans sensu lato strains were isolated: three from pigs, two from dogs, and two from water. High seroprevalence in pigs, dogs, and humans, concomitant to isolation of strains, demonstrates that in Cordoba, transmission exists among animals, the environment, and humans, which warrants the implementation of public health intervention measures to reduce the epidemiological impact of leptospirosis in the region. PMID:24254419

  19. [A Case of Leptospirosis in which the Causative Pathogen was Detected Using Cerebrospinal Fluid PCR Eight Days after Onset].

    PubMed

    Arita, Yuki; Tono, Toshihiro; Hosoda, Tomohiro; Taguchi, Hiroaki; Sakamoto, Mitsuo; Osone, Yasuo; Nozaki, Hiroyuki

    2016-05-01

    We report a patient with leptospirosis caused by infection with Leptospira interrogans serovar Rachmati. A 30-year-old Japanese man took part in a survival camp on Iriomote Island, Okinawa, from July 9 to July 15, 2014. During the camp, he swam in the river and kayaked. He developed a high fever and fatigue 7 days after completing his trip and was admitted to our hospital on July 22. On admission, he complained of a posterior cervical pain and a loss of appetite. Laboratory findings revealed granulocytosis, mildly elevated AST and ALT levels, elevated BUN and Cr levels, and a significantly elevated CRP level. No pathogenic bacteria were isolated from blood, urine, or cerebrospinal fluid cultures. We included leptospirosis in the differential diagnosis because of the patient's history of participating in a survival camp on Iriomote Island. Minocycline 200 mg, p.o. showed an excellent efficacy. The Leptospira flagellar gene FlaB was detected using a cerebrospinal fluid PCR. A microscopic agglutination test (MAT) during the convalescent stage demonstrated significant increases in antibodies against L. interrogans serovar Rachmati, confirming the diagnosis of leptospirosis. A medical history including occupation and recent travel history, and an adequate specimen sampling are crucial for the accurate and early diagnosis of leptospirosis. PMID:27529969

  20. Environmental Factors Influencing White-Tailed Deer (Odocoileus virginianus) Exposure to Livestock Pathogens in Wisconsin.

    PubMed

    Dubay, Shelli; Jacques, Christopher; Golden, Nigel; Kern, Bryant; Mahoney, Kathleen; Norton, Andrew; Patnayak, Devi; Van Deelen, Timothy

    2015-01-01

    White-tailed deer (Odocoileus virginianus) are commonly exposed to disease agents that affect livestock but environmental factors that predispose deer to exposure are unknown for many pathogens. We trapped deer during winter months on two study areas (Northern Forest and Eastern Farmland) in Wisconsin from 2010 to 2013. Deer were tested for exposure to six serovars of Leptospira interrogans (grippotyphosa, icterohaemorrhagiae, canicola, bratislava, pomona, and hardjo), bovine viral diarrhea virus (BVDV-1 and BVDV-2), infectious bovine rhinotracheitis virus (IBR), and parainfluenza 3 virus (PI3). We used logistic regression to model potential intrinsic (e.g., age, sex) and extrinsic (e.g., land type, study site, year, exposure to multiple pathogens) variables we considered biologically meaningful to exposure of deer to livestock pathogens. Deer sampled in 2010-2011 did not demonstrate exposure to BVDV, so we did not test for BVDV in subsequent years. Deer had evidence of exposure to PI3 (24.7%), IBR (7.9%), Leptospira interrogans serovar pomona (11.7%), L. i. bratislava (1.0%), L. i. grippotyphosa (2.5%) and L. i. hardjo (0.3%). Deer did not demonstrate exposure to L. interrogans serovars canicola and icterohaemorrhagiae. For PI3, we found that capture site and year influenced exposure. Fawns (n = 119) were not exposed to L. i. pomona, but land type was an important predictor of exposure to L. i. pomona for older deer. Our results serve as baseline exposure levels of Wisconsin white-tailed deer to livestock pathogens, and helped to identify important factors that explain deer exposure to livestock pathogens.

  1. Environmental Factors Influencing White-Tailed Deer (Odocoileus virginianus) Exposure to Livestock Pathogens in Wisconsin

    PubMed Central

    Kern, Bryant; Mahoney, Kathleen; Norton, Andrew; Patnayak, Devi; Van Deelen, Timothy

    2015-01-01

    White-tailed deer (Odocoileus virginianus) are commonly exposed to disease agents that affect livestock but environmental factors that predispose deer to exposure are unknown for many pathogens. We trapped deer during winter months on two study areas (Northern Forest and Eastern Farmland) in Wisconsin from 2010 to 2013. Deer were tested for exposure to six serovars of Leptospira interrogans (grippotyphosa, icterohaemorrhagiae, canicola, bratislava, pomona, and hardjo), bovine viral diarrhea virus (BVDV-1 and BVDV-2), infectious bovine rhinotracheitis virus (IBR), and parainfluenza 3 virus (PI3). We used logistic regression to model potential intrinsic (e.g., age, sex) and extrinsic (e.g., land type, study site, year, exposure to multiple pathogens) variables we considered biologically meaningful to exposure of deer to livestock pathogens. Deer sampled in 2010–2011 did not demonstrate exposure to BVDV, so we did not test for BVDV in subsequent years. Deer had evidence of exposure to PI3 (24.7%), IBR (7.9%), Leptospira interrogans serovar pomona (11.7%), L. i. bratislava (1.0%), L. i. grippotyphosa (2.5%) and L. i. hardjo (0.3%). Deer did not demonstrate exposure to L. interrogans serovars canicola and icterohaemorrhagiae. For PI3, we found that capture site and year influenced exposure. Fawns (n = 119) were not exposed to L. i. pomona, but land type was an important predictor of exposure to L. i. pomona for older deer. Our results serve as baseline exposure levels of Wisconsin white-tailed deer to livestock pathogens, and helped to identify important factors that explain deer exposure to livestock pathogens. PMID:26030150

  2. Environmental Factors Influencing White-Tailed Deer (Odocoileus virginianus) Exposure to Livestock Pathogens in Wisconsin.

    PubMed

    Dubay, Shelli; Jacques, Christopher; Golden, Nigel; Kern, Bryant; Mahoney, Kathleen; Norton, Andrew; Patnayak, Devi; Van Deelen, Timothy

    2015-01-01

    White-tailed deer (Odocoileus virginianus) are commonly exposed to disease agents that affect livestock but environmental factors that predispose deer to exposure are unknown for many pathogens. We trapped deer during winter months on two study areas (Northern Forest and Eastern Farmland) in Wisconsin from 2010 to 2013. Deer were tested for exposure to six serovars of Leptospira interrogans (grippotyphosa, icterohaemorrhagiae, canicola, bratislava, pomona, and hardjo), bovine viral diarrhea virus (BVDV-1 and BVDV-2), infectious bovine rhinotracheitis virus (IBR), and parainfluenza 3 virus (PI3). We used logistic regression to model potential intrinsic (e.g., age, sex) and extrinsic (e.g., land type, study site, year, exposure to multiple pathogens) variables we considered biologically meaningful to exposure of deer to livestock pathogens. Deer sampled in 2010-2011 did not demonstrate exposure to BVDV, so we did not test for BVDV in subsequent years. Deer had evidence of exposure to PI3 (24.7%), IBR (7.9%), Leptospira interrogans serovar pomona (11.7%), L. i. bratislava (1.0%), L. i. grippotyphosa (2.5%) and L. i. hardjo (0.3%). Deer did not demonstrate exposure to L. interrogans serovars canicola and icterohaemorrhagiae. For PI3, we found that capture site and year influenced exposure. Fawns (n = 119) were not exposed to L. i. pomona, but land type was an important predictor of exposure to L. i. pomona for older deer. Our results serve as baseline exposure levels of Wisconsin white-tailed deer to livestock pathogens, and helped to identify important factors that explain deer exposure to livestock pathogens. PMID:26030150

  3. Decrease in antithrombin III and prothrombin serum levels contribute to coagulation disorders during leptospirosis.

    PubMed

    Fernandes, Luis G V; Filho, Antonio F S; Souza, Gisele O; Vasconcellos, Silvio A; Romero, Eliete C; Nascimento, Ana L T O

    2016-08-01

    Pathogenic bacteria of the genus Leptospira are the causative agent of leptospirosis, an emergent infectious disease that affects humans and animals worldwide. Severe forms of the disease in humans include jaundice, multiple organ failure and intense haemorrhage. Up to now, mechanisms associated with the haemorrhage foci are poorly understood. We report in this work that, despite the low levels of antithrombin III in convalescent human serum samples, virulent, culture-attenuated and saprophyte strains of Leptospira are unable to bind and/or degrade this thrombin inhibitor, suggesting an indirect mechanism of pathogenesis. Lower levels of prothrombin were found in serum samples at the onset and convalescent phase of the disease when compared to normal human sera. The concomitant decreased levels of antithrombin III and prothrombin suggest a process of stimulated coagulation, which is corroborated by the increase of prothrombin fragment F1+2 in the serum samples. Data obtained with hamsters experimentally infected with virulent Leptospira interrogans serovars Kennewicki and Canicola strongly point out that haemorrhage is correlated with decreased levels of thrombin inhibitors and prothrombin. Activated coagulation might lead to an overconsumption of coagulation factors ultimately leading to bleeding and organ failure. PMID:27260249

  4. Evaluation of two novel leptospiral proteins for their interaction with human host components.

    PubMed

    Silva, Lucas P; Fernandes, Luis G V; Vieira, Monica L; de Souza, Gisele O; Heinemann, Marcos B; Vasconcellos, Silvio A; Romero, Eliete C; Nascimento, Ana L T O

    2016-07-01

    Pathogenic species of the genus Leptospira are the etiological agents of leptospirosis, the most widespread zoonosis. Mechanisms involved in leptospiral pathogenesis are not well understood. By data mining the genome sequences of Leptospira interrogans we have identified two proteins predicted to be surface exposed, LIC10821 and LIC10064. Immunofluorescence and proteinase K assays confirmed that the proteins are exposed. Reactivity of the recombinant proteins with human sera has shown that rLIC10821, but not rLIC10064, is recognized by antibodies in confirmed leptospirosis serum samples, suggesting its expression during infection. The rLIC10821 was able to bind laminin, in a dose-dependent fashion, and was called Lsa37 (leptospiral surface adhesin of 37 kDa). Studies with human plasma components demonstrated that rLIC10821 interacts with plasminogen (PLG) and fibrinogen (Fg). The binding of Lsa37 with PLG generates plasmin when PLG activator was added. Fibrin clotting reduction was observed in a thrombin-catalyzed reaction, when Fg was incubated with Lsa37, suggesting that this protein may interfere in the coagulation cascade during the disease. Although LIC10064 protein is more abundant than the corresponding Lsa37, binding activity with all the components tested was not detected. Thus, Lsa37 is a novel versatile adhesin that may mediate Leptospira-host interactions. PMID:27129366

  5. Detection and analysis of leptospiral DNA in early Leptospirosis by polymerase chain reaction and DNA hybridization with Digoxingenin-AMPPD

    NASA Astrophysics Data System (ADS)

    Bao, Lang; Yu, Ye-Rong; Terpstra, W. J.

    1994-07-01

    Fourteen serum specimens from patients with early Leptospirosis proven by blood culture and serological test were detected by PCR with the oligonucleotide primers obtained from a genomic library of leptospira interrogans. The amplified DNA were hybridized with the homologous DNA probe labeling with Digoxingenin-AMPPD. All of the samples revealed the presence of leptospira and the strong signals were visualized with homologous DNA probes hybridization. Negative and positive controls appeared correctly. The DNA fragment generated from PCR amplification homologically hybridized with the DNA of 16 strains of leptospira. The single recognized band (about 400 bps) from 6 out of the 16 strains has come out which are representative of the principal strains in Sichuan, China. The results demonstrated that PCR is an advanced diagnostic technique for early Leptospirosis. The treatment of samples is easy and has little risk of DNA loss and contamination. This is a considerable advantage over other detective techniques and can be available especially in China and other developing countries.

  6. Decrease in antithrombin III and prothrombin serum levels contribute to coagulation disorders during leptospirosis.

    PubMed

    Fernandes, Luis G V; Filho, Antonio F S; Souza, Gisele O; Vasconcellos, Silvio A; Romero, Eliete C; Nascimento, Ana L T O

    2016-08-01

    Pathogenic bacteria of the genus Leptospira are the causative agent of leptospirosis, an emergent infectious disease that affects humans and animals worldwide. Severe forms of the disease in humans include jaundice, multiple organ failure and intense haemorrhage. Up to now, mechanisms associated with the haemorrhage foci are poorly understood. We report in this work that, despite the low levels of antithrombin III in convalescent human serum samples, virulent, culture-attenuated and saprophyte strains of Leptospira are unable to bind and/or degrade this thrombin inhibitor, suggesting an indirect mechanism of pathogenesis. Lower levels of prothrombin were found in serum samples at the onset and convalescent phase of the disease when compared to normal human sera. The concomitant decreased levels of antithrombin III and prothrombin suggest a process of stimulated coagulation, which is corroborated by the increase of prothrombin fragment F1+2 in the serum samples. Data obtained with hamsters experimentally infected with virulent Leptospira interrogans serovars Kennewicki and Canicola strongly point out that haemorrhage is correlated with decreased levels of thrombin inhibitors and prothrombin. Activated coagulation might lead to an overconsumption of coagulation factors ultimately leading to bleeding and organ failure.

  7. Evaluation of two novel leptospiral proteins for their interaction with human host components.

    PubMed

    Silva, Lucas P; Fernandes, Luis G V; Vieira, Monica L; de Souza, Gisele O; Heinemann, Marcos B; Vasconcellos, Silvio A; Romero, Eliete C; Nascimento, Ana L T O

    2016-07-01

    Pathogenic species of the genus Leptospira are the etiological agents of leptospirosis, the most widespread zoonosis. Mechanisms involved in leptospiral pathogenesis are not well understood. By data mining the genome sequences of Leptospira interrogans we have identified two proteins predicted to be surface exposed, LIC10821 and LIC10064. Immunofluorescence and proteinase K assays confirmed that the proteins are exposed. Reactivity of the recombinant proteins with human sera has shown that rLIC10821, but not rLIC10064, is recognized by antibodies in confirmed leptospirosis serum samples, suggesting its expression during infection. The rLIC10821 was able to bind laminin, in a dose-dependent fashion, and was called Lsa37 (leptospiral surface adhesin of 37 kDa). Studies with human plasma components demonstrated that rLIC10821 interacts with plasminogen (PLG) and fibrinogen (Fg). The binding of Lsa37 with PLG generates plasmin when PLG activator was added. Fibrin clotting reduction was observed in a thrombin-catalyzed reaction, when Fg was incubated with Lsa37, suggesting that this protein may interfere in the coagulation cascade during the disease. Although LIC10064 protein is more abundant than the corresponding Lsa37, binding activity with all the components tested was not detected. Thus, Lsa37 is a novel versatile adhesin that may mediate Leptospira-host interactions.

  8. The Leptospiral Antigen Lp49 is a Two-Domain Protein with Putative Protein Binding Function

    SciTech Connect

    Oliveira Giuseppe,P.; Oliveira Neves, F.; Nascimento, A.; Gomes Guimaraes, B.

    2008-01-01

    Pathogenic Leptospira is the etiological agent of leptospirosis, a life-threatening disease that affects populations worldwide. Currently available vaccines have limited effectiveness and therapeutic interventions are complicated by the difficulty in making an early diagnosis of leptospirosis. The genome of Leptospira interrogans was recently sequenced and comparative genomic analysis contributed to the identification of surface antigens, potential candidates for development of new vaccines and serodiagnosis. Lp49 is a membrane-associated protein recognized by antibodies present in sera from early and convalescent phases of leptospirosis patients. Its crystal structure was determined by single-wavelength anomalous diffraction using selenomethionine-labelled crystals and refined at 2.0 Angstroms resolution. Lp49 is composed of two domains and belongs to the all-beta-proteins class. The N-terminal domain folds in an immunoglobulin-like beta-sandwich structure, whereas the C-terminal domain presents a seven-bladed beta-propeller fold. Structural analysis of Lp49 indicates putative protein-protein binding sites, suggesting a role in Leptospira-host interaction. This is the first crystal structure of a leptospiral antigen described to date.

  9. 9 CFR 113.102 - Leptospira Icterohaemorrhagiae Bacterin.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... icterohaemorrhagiae organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4)...

  10. 9 CFR 113.102 - Leptospira Icterohaemorrhagiae Bacterin.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... icterohaemorrhagiae organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4)...

  11. 9 CFR 113.102 - Leptospira Icterohaemorrhagiae Bacterin.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... icterohaemorrhagiae organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4)...

  12. 9 CFR 113.104 - Leptospira Grippotyphosa Bacterin.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4) Post-challenge...

  13. 9 CFR 113.102 - Leptospira Icterohaemorrhagiae Bacterin.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... icterohaemorrhagiae organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4)...

  14. 9 CFR 113.104 - Leptospira Grippotyphosa Bacterin.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4) Post-challenge...

  15. 9 CFR 113.102 - Leptospira Icterohaemorrhagiae Bacterin.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... icterohaemorrhagiae organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4)...

  16. 9 CFR 113.104 - Leptospira Grippotyphosa Bacterin.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4) Post-challenge...

  17. 9 CFR 113.104 - Leptospira Grippotyphosa Bacterin.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4) Post-challenge...

  18. 9 CFR 113.104 - Leptospira Grippotyphosa Bacterin.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25... recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same... organisms, using a dose of 10-10,000 hamster LD50 as determined by titration. (4) Post-challenge...

  19. Studies on the susceptibility of water buffaloes to Leptospira.

    PubMed

    Ryu, E; Liu, C K

    1968-04-01

    Eight mature farming type, Taiwan, water buffaloes were inoculate with L. australis A while six received L. canicola. Before inoculation all animals were negative to the microscopic-agglutination test (agglutinationlysis test) using the above species as antigen. No sign of clinical leptospirosis was observed although four animals developed temperatures. Cultures made from buffalo blood, kidneys and urine and from blood of guinea pigs inoculated with kidney emulsion and urine from the inoculated buffalo were all negative for leptospiral organisms. Blood samples drawn from the water buffalo 2, 3 and 4 weeks post inoculation were negative to the microscopic-agglutination test except for one animal. Blood from the animal taken two weeks post-inoculation was positive at 1:100 dilution with L. australis A antigen but that taken at 3 and 4 weeks was negative.

  20. Regional Differences of Leptospirosis in Sri Lanka: Observations from a Flood-Associated Outbreak in 2011

    PubMed Central

    Agampodi, Suneth B.; Dahanayaka, Niroshan J.; Bandaranayaka, Anoma K.; Perera, Manoj; Priyankara, Sumudu; Weerawansa, Prasanna; Matthias, Michael A.; Vinetz, Joseph M.

    2014-01-01

    Leptospirosis is known to be an important cause of weather disaster-related infectious disease epidemics. In 2011, an outbreak of leptospirosis occurred in the relatively dry district of Anuradhapura, Sri Lanka where diagnosis was resisted by local practitioners because leptospirosis was not known in the area and the clinical presentation was considered atypical. To identify the causative Leptospira associated with this outbreak, we carried out a cross-sectional study. Consecutive clinically suspected cases in this district were studied during a two-and-a-half-month period. Of 96 clinically suspected cases, 32 (33.3%) were confirmed by qPCR, of which the etiological cause in 26 cases was identified using 16S rDNA sequencing to the species level. Median bacterial load was 4.1×102/mL (inter-quartile range 3.1–6.1×102/mL). In contrast to a 2008 Sri Lankan leptospirosis outbreak in the districts of Kegalle, Kandy, and Matale, in which a predominance of Leptospira interrogans serovars Lai and Geyaweera was found, most cases in the 2011 outbreak were caused by Leptospira kirschneri. Seven (21.9%) confirmed cases had acute renal failure; five (15.6%) had myocarditis; severe thrombocytopenia (<20,000/uL) was seen in five (15.6%) cases. This outbreak of leptospirosis in the relatively dry zone of Sri Lanka due primarily to L. kirschneri was characterized by markedly different clinical presentations and low leptospiremia. These observations and data demonstrate the public health relevance of molecular diagnostics in such settings, possibly related to the microgeographic variations of different Leptospira species, but of particular value to public health intervention in what appears to have been a regionally neglected tropical disease. PMID:24454971

  1. Leptospiral LruA Is Required for Virulence and Modulates an Interaction with Mammalian Apolipoprotein AI

    PubMed Central

    Zhang, Kunkun; Murray, Gerald L.; Seemann, Torsten; Srikram, Amporn; Bartpho, Thanatchaporn; Sermswan, Rasana W.; Hoke, David E.

    2013-01-01

    Leptospirosis is a worldwide zoonosis caused by spirochetes of the genus Leptospira. While understanding of pathogenesis remains limited, the development of mutagenesis in Leptospira has provided a powerful tool for identifying novel virulence factors. LruA is a lipoprotein that has been implicated in leptospiral uveitis as a target of the immune response. In this study, two lruA mutants, M754 and M765, generated by transposon mutagenesis from Leptospira interrogans serovar Manilae, were characterized. In M754, the transposon inserted in the middle of lruA, resulting in no detectable expression of LruA. In M765, the transposon inserted toward the 3′ end of the gene, resulting in expression of a truncated protein. LruA was demonstrated to be on the cell surface in M765 and the wild type (WT). M754, but not M765, was attenuated in a hamster model of acute infection. A search for differential binding to human serum proteins identified a serum protein of around 30 kDa bound to the wild type and the LruA deletion mutant (M754), but not to the LruA truncation mutant (M765). Two-dimensional separation of proteins from leptospiral cells incubated with guinea pig serum identified the 28-kDa apolipoprotein A-I (ApoA-I) as a major mammalian serum protein that binds Leptospira in vitro. Interestingly, M754 (with no detectable LruA) bound more ApoA-I than did the LruA-expressing strains Manilae wild type and M765. Our data thus identify LruA as a surface-exposed leptospiral virulence factor that contributes to leptospiral pathogenesis, possibly by modulating cellular interactions with serum protein ApoA-I. PMID:23918777

  2. Viral and bacterial serology of six free-ranging bearded seals Erignathus barbatus.

    PubMed

    Calle, Paul P; Seagars, Dana J; McClave, Catherine; Senne, Dennis; House, Carol; House, James A

    2008-08-19

    Serum or heparinized plasma samples were obtained from 3 male (2 adult and 1 weaned calf) and 3 adult female free-ranging bearded seals Erignathus barbatus in May of 1994, 1995, or 1996. Blood samples were obtained from animals taken in subsistence hunts near St. Lawrence Island, Alaska and screened for antibodies to a suite of bacteria and viruses potentially pathogenic for pinnipeds and/or humans. No samples had detectable antibodies to Brucella spp., Phocine distemper virus, influenza A virus or caliciviruses (San Miguel sea lion virus strains 1, 2, and 4 to 13, vesicular exanthema of swine serotypes A48, B51, C52, D53, E54, F55, G55, H54, 155, J56, K54, 1934B, and Tillamook and Walrus calicivirus). One seal had a low titer of 100 to Leptospira interrogans serovar PMID:18828565

  3. Coiled to diffuse: Brownian motion of a helical bacterium.

    PubMed

    Butenko, Alexander V; Mogilko, Emma; Amitai, Lee; Pokroy, Boaz; Sloutskin, Eli

    2012-09-11

    We employ real-time three-dimensional confocal microscopy to follow the Brownian motion of a fixed helically shaped Leptospira interrogans (LI) bacterium. We extract from our measurements the translational and the rotational diffusion coefficients of this bacterium. A simple theoretical model is suggested, perfectly reproducing the experimental diffusion coefficients, with no tunable parameters. An older theoretical model, where edge effects are neglected, dramatically underestimates the observed rates of translation. Interestingly, the coiling of LI increases its rotational diffusion coefficient by a factor of 5, compared to a (hypothetical) rectified bacterium of the same contour length. Moreover, the translational diffusion coefficients would have decreased by a factor of ~1.5, if LI were rectified. This suggests that the spiral shape of the spirochaete bacteria, in addition to being employed for their active twisting motion, may also increase the ability of these bacteria to explore the surrounding fluid by passive Brownian diffusion.

  4. The use of halloysite clay and carboxyl-functionalised multi-walled carbon nanotubes for recombinant LipL32 antigen delivery enhanced the IgG response

    PubMed Central

    Hartwig, Daiane D; Bacelo, Kátia L; Oliveira, Thaís L; Schuch, Rodrigo; Seixas, Fabiana K; Collares, Tiago; Rodrigues, Oscar; Hartleben, Cláudia P; Dellagostin, Odir A

    2015-01-01

    We studied the feasibility of using halloysite clay nanotubes (HNTs) and carboxyl-functionalised multi-walled carbon nanotubes (COOH-MWCNTs) as antigen carriers to improve immune responses against a recombinant LipL32 protein (rLipL32). Immunisation using the HNTs or COOH-MWCNTs significantly increased the rLipL32-specific IgG antibody titres (p < 0.05) of Golden Syrian hamsters. None of the vaccines tested conferred protection against a challenge using a virulent Leptospira interrogans strain. These results demonstrated that nanotubes can be used as antigen carriers for delivery in hosts and the induction of a humoral immune response against purified leptospiral antigens used in subunit vaccine preparations. PMID:25742273

  5. Rats, cities, people, and pathogens: a systematic review and narrative synthesis of literature regarding the ecology of rat-associated zoonoses in urban centers.

    PubMed

    Himsworth, Chelsea G; Parsons, Kirbee L; Jardine, Claire; Patrick, David M

    2013-06-01

    Urban Norway and black rats (Rattus norvegicus and Rattus rattus) are the source of a number of pathogens responsible for significant human morbidity and mortality in cities around the world. These pathogens include zoonotic bacteria (Leptospira interrogans, Yersina pestis, Rickettsia typhi, Bartonella spp., Streptobacillus moniliformis), viruses (Seoul hantavirus), and parasites (Angiostrongylus cantonensis). A more complete understanding of the ecology of these pathogens in people and rats is critical for determining the public health risks associated with urban rats and for developing strategies to monitor and mitigate those risks. Although the ecology of rat-associated zoonoses is complex, due to the multiple ways in which rats, people, pathogens, vectors, and the environment may interact, common determinants of human disease can still be identified. This review summarizes the ecology of zoonoses associated with urban rats with a view to identifying similarities, critical differences, and avenues for further study.

  6. The use of halloysite clay and carboxyl-functionalised multi-walled carbon nanotubes for recombinant LipL32 antigen delivery enhanced the IgG response.

    PubMed

    Hartwig, Daiane D; Bacelo, Kátia L; Oliveira, Thaís L; Schuch, Rodrigo; Seixas, Fabiana K; Collares, Tiago; Rodrigues, Oscar; Hartleben, Cláudia P; Dellagostin, Odir A

    2015-02-01

    We studied the feasibility of using halloysite clay nanotubes (HNTs) and carboxyl-functionalised multi-walled carbon nanotubes (COOH-MWCNTs) as antigen carriers to improve immune responses against a recombinant LipL32 protein (rLipL32). Immunisation using the HNTs or COOH-MWCNTs significantly increased the rLipL32-specific IgG antibody titres (p < 0.05) of Golden Syrian hamsters. None of the vaccines tested conferred protection against a challenge using a virulent Leptospira interrogans strain. These results demonstrated that nanotubes can be used as antigen carriers for delivery in hosts and the induction of a humoral immune response against purified leptospiral antigens used in subunit vaccine preparations.

  7. The etiology of acute pyrexia of unknown origin in children after a flood.

    PubMed

    Pradutkanchana, Jintana; Pradutkanchana, Sukone; Kemapanmanus, Marisa; Wuthipum, Nathakul; Silpapojakul, Kamkarn

    2003-03-01

    Acute pyrexia of unknown origin (PUO) is a major public health problem in Thailand. We studied the etiology of 180 cases of acute PUO in children after a sudden severe flood in Hat Yai city in 2000. Dengue infection and leptospirosis accounted for more than half of the total cases. Dengue hemorrhagic fever was the most common (29.4%) followed by leptospirosis (27.2%) and scrub typhus infection (1.1%). Five serovars of leptospires were involved in this study. Leptospira interrogans bataviae was the most common (86.5%). Acute serum antibody testing could detect only 52.8% and 40.8% of dengue and leptospirosis cases, respectively. This study showed both should be included in the presumptive diagnosis of acute PUO in patients after flooding.

  8. Proteomic Features Predict Seroreactivity against Leptospiral Antigens in Leptospirosis Patients

    PubMed Central

    2015-01-01

    With increasing efficiency, accuracy, and speed we can access complete genome sequences from thousands of infectious microorganisms; however, the ability to predict antigenic targets of the immune system based on amino acid sequence alone is still needed. Here we use a Leptospira interrogans microarray expressing 91% (3359) of all leptospiral predicted ORFs (3667) and make an empirical accounting of all antibody reactive antigens recognized in sera from naturally infected humans; 191 antigens elicited an IgM or IgG response, representing 5% of the whole proteome. We classified the reactive antigens into 26 annotated COGs (clusters of orthologous groups), 26 JCVI Mainrole annotations, and 11 computationally predicted proteomic features. Altogether, 14 significantly enriched categories were identified, which are associated with immune recognition including mass spectrometry evidence of in vitro expression and in vivo mRNA up-regulation. Together, this group of 14 enriched categories accounts for just 25% of the leptospiral proteome but contains 50% of the immunoreactive antigens. These findings are consistent with our previous studies of other Gram-negative bacteria. This genome-wide approach provides an empirical basis to predict and classify antibody reactive antigens based on structural, physical–chemical, and functional proteomic features and a framework for understanding the breadth and specificity of the immune response to L. interrogans. PMID:25358092

  9. The terminal portion of leptospiral immunoglobulin-like protein LigA confers protective immunity against lethal infection in the hamster model of leptospirosis.

    PubMed

    Silva, Everton F; Medeiros, Marco A; McBride, Alan J A; Matsunaga, Jim; Esteves, Gabriela S; Ramos, João G R; Santos, Cleiton S; Croda, Júlio; Homma, Akira; Dellagostin, Odir A; Haake, David A; Reis, Mitermayer G; Ko, Albert I

    2007-08-14

    Subunit vaccines are a potential intervention strategy against leptospirosis, which is a major public health problem in developing countries and a veterinary disease in livestock and companion animals worldwide. Leptospiral immunoglobulin-like (Lig) proteins are a family of surface-exposed determinants that have Ig-like repeat domains found in virulence factors such as intimin and invasin. We expressed fragments of the repeat domain regions of LigA and LigB from Leptospira interrogans serovar Copenhageni. Immunization of Golden Syrian hamsters with Lig fragments in Freund's adjuvant induced robust antibody responses against recombinant protein and native protein, as detected by ELISA and immunoblot, respectively. A single fragment, LigANI, which corresponds to the six carboxy-terminal Ig-like repeat domains of the LigA molecule, conferred immunoprotection against mortality (67-100%, P<0.05) in hamsters which received a lethal inoculum of L. interrogans serovar Copenhageni. However, immunization with this fragment did not confer sterilizing immunity. These findings indicate that the carboxy-terminal portion of LigA is an immunoprotective domain and may serve as a vaccine candidate for human and veterinary leptospirosis. PMID:17629368

  10. Leptospirosis in wild and domestic carnivores in natural areas in Andalusia, Spain.

    PubMed

    Millán, Javier; Candela, Mónica G; López-Bao, José Vicente; Pereira, Marian; Jiménez, María Angeles; León-Vizcaíno, Luis

    2009-10-01

    Leptospirosis is a zoonosis that affects humans, domestic animals, and wildlife. Carnivores are at the top of the feeding chain, thus being exposed to pathogens through their preys. From June 2004 to April 2007, we analyzed for evidences of contact with 14 serovars of Leptospira interrogans Sensu Lato serum (analyzed by indirect Microscopic Agglutination Test) and urine or kidney samples (analyzed by microscopic observation, immunostaining and culture) collected from 201 wild and domestic carnivores, including 26 free-living Iberian lynx (Lynx pardinus), 33 red foxes (Vulpes vulpes), 33 Egyptian mongooses (Herpestes ichneumon), 25 common genets (Genetta genetta), two Eurasian badgers (Meles meles) and one Eurasian otter (Lutra lutra), and 53 free-roaming cats and 28 rural dogs in protected areas in Andalusia (southern Spain). Twenty-three percent of the animals presented evidences of contact, being the prevalence similar among wild (23.5%) and domestic species (22.2%). Contact with Lesptospira was detected in all the species but the otter. Prevalence was: lynx (11% by bacteriological detection, 32% by serology), fox (0%, 47%), mongoose (5%, 20%), genet (0%, 12%), badger (0%, 50%), cat (20%, 14%), dog (only serology: 36%). Serovar Icterohemorragiae accounted for 2/3 of the cases. Serovar Canicola was detected in half of the positive dogs and one lynx. Other serovars detected were Ballum, Sejroë, and Australis. No macroscopic lesions were observed in necropsied animals that showed evidence of contact with the agent, although histopathologic lesions (chiefly chronic interstitial nephritis) were observed in 7 out of the 11 microscopically analyzed individuals. Thus, L. interrogans may cause previously unrecorded disease in wild carnivores in Spain. Wild and free-roaming carnivores may not act as reservoir of L. interrogans but as a dead-end hosts, though the dog may act as reservoir of serovar Canicola. Carnivores are apparently good sentinels for the epidemiological

  11. COMPARISON OF 16S rRNA-PCR-RFLP, LipL32-PCR AND OmpL1-PCR METHODS IN THE DIAGNOSIS OF LEPTOSPIROSIS

    PubMed Central

    GÖKMEN, Tülin GÜVEN; SOYAL, Ayben; KALAYCI, Yıldız; ÖNLEN, Cansu; KÖKSAL, Fatih

    2016-01-01

    SUMMARY Leptospirosis is still one of the most important health problems in developing countries located in humid tropical and subtropical regions. Human infections are generally caused by exposure to water, soil or food contaminated with the urine of infected wild and domestic animals such as rodents and dogs. The clinical course of leptospirosis is variable and may be difficult to distinguish from many other infectious diseases. The dark-field microscopy (DFM), serology and nucleic acid amplification techniques are used to diagnose leptospirosis, however, a distinctive standard reference method is still lacking. Therefore, in this study, we aimed to determine the presence of Leptospira spp., to differentiate the pathogenic L. interrogans and the non-pathogenic L. biflexa, and also to determine the sensitivity and specificity values of molecular methods as an alternative to conventional ones. A total of 133 serum samples, from 47 humans and 86 cattle were evaluated by two conventional tests: the Microagglutination Test (MAT) and the DFM, as well as three molecular methods, the 16S rRNA-PCR followed by Restriction Fragment Lenght Polymorphism (RFLP) of the amplification products 16S rRNA-PCR-RFLP, LipL32-PCR and OmpL1-PCR. In this study, for L. interrogans, the specificity and sensitivity rates of the 16S rRNA-PCR and the LipL32-PCR were considered similar (100% versus 98.25% and 100% versus 98.68%, respectively). The OmpL1-PCR was able to classify L. interrogans into two intergroups, but this PCR was less sensitive (87.01%) than the other two PCR methods. The 16S rRNA-PCR-RFLP could detect L. biflexa DNA, but LipL32-PCR and OmpL1-PCR could not. The 16S rRNA-PCR-RFLP provided an early and accurate diagnosis and was able to distinguish pathogenic and non-pathogenic Leptospira species, hence it may be used as an alternative method to the conventional gold standard techniques for the rapid disgnosis of leptospirosis. PMID:27680169

  12. Multilocus sequence typing (MLST) of leptospiral strains isolated from two geographic locations of Tamil Nadu, India.

    PubMed

    Kanagavel, Murugesan; Princy Margreat, Alphonse Asirvatham; Arunkumar, Manivel; Prabhakaran, Shanmugarajan Gnanasekaran; Shanmughapriya, Santhanam; Natarajaseenivasan, Kalimuthusamy

    2016-01-01

    Here the rodent carrier status for the transmission of human leptospirosis in Tiruchirappalli, district, Tamil Nadu, India was assessed. The predominantly circulating leptospiral STs were recognized by multilocus sequence typing (MLST). A total of 113 rodents were trapped from different provinces of the Tiruchirappalli district. The most prevalent rodent was Bandicota bengalensis (37.2%), and of the total, 52.2% (n=59) rodents were found to be positive for leptospiral 16S rRNA. These results were validated with a leptospiral culture positivity of 45.8% (n=27). Three isolates from Chennai (2 rodents and 1 human) and 1 human isolate from Tiruchirappalli were included to understand the spatial variations and to track the source of human leptospirosis. The serogroup, serovar, and species level identification of all 31 isolates identified 28 to be Leptospira borgpetersenii serovar Javanica and three as Leptospira interrogans serovar Autumnalis. MLST analysis defined all isolates to the existing ST profiles (ST145 and ST27) with the exception of 6 L. borgpetersenii (ST DR) isolates that showed variations in the sucA and pfkB loci. The DR ST was locally confined to Chatram province of Tiruchirappalli suggesting an epidemiological link. The predominant STs, ST145 and ST-DR form a group, indicating the presence of original strain that subsequently diverged evolutionarily into two STs. The variations between L. borgpetersenii in sucA and pfkB loci may be an indication that evolutionary changes transpired in Tiruchirappalli.

  13. Efficient Detection of Pathogenic Leptospires Using 16S Ribosomal RNA

    PubMed Central

    Lindow, Janet; Wunder, Elsio A.; Reis, Mitermayer G.; Usmani-Brown, Sahar; Ledizet, Michel; Ko, Albert; Pal, Utpal

    2015-01-01

    Pathogenic Leptospira species cause a prevalent yet neglected zoonotic disease with mild to life-threatening complications in a variety of susceptible animals and humans. Diagnosis of leptospirosis, which primarily relies on antiquated serotyping methods, is particularly challenging due to presentation of non-specific symptoms shared by other febrile illnesses, often leading to misdiagnosis. Initiation of antimicrobial therapy during early infection to prevent more serious complications of disseminated infection is often not performed because of a lack of efficient diagnostic tests. Here we report that specific regions of leptospiral 16S ribosomal RNA molecules constitute a novel and efficient diagnostic target for PCR-based detection of pathogenic Leptospira serovars. Our diagnostic test using spiked human blood was at least 100-fold more sensitive than corresponding leptospiral DNA-based quantitative PCR assays, targeting the same 16S nucleotide sequence in the RNA and DNA molecules. The sensitivity and specificity of our RNA assay against laboratory-confirmed human leptospirosis clinical samples were 64% and 100%, respectively, which was superior then an established parallel DNA detection assay. Remarkably, we discovered that 16S transcripts remain appreciably stable ex vivo, including untreated and stored human blood samples, further highlighting their use for clinical detection of L. interrogans. Together, these studies underscore a novel utility of RNA targets, specifically 16S rRNA, for development of PCR-based modalities for diagnosis of human leptospirosis, and also may serve as paradigm for detection of additional bacterial pathogens for which early diagnosis is warranted. PMID:26091292

  14. Serosurvey of leptospirosis in feral hogs (Sus scrofa) in Florida.

    PubMed

    Chatfield, Jenifer; Milleson, Michael; Stoddard, Robyn; Bui, Duy M; Galloway, Renee

    2013-06-01

    Leptospira is a global pathogen of emerging public health importance in both developing and industrialized nations and can infect almost all mammalian species, including humans. As suburbanization and the popularity of outdoor recreational activities increases, so do human-wildlife and companion animal-wildlife interfaces. Florida offers a tropical climate favorable for outdoor activities and a semirural landscape that sustains an abundant feral hog population. Because no survey ofleptospirosis in feral hogs (Sus scrofa) in Florida has been published to our knowledge, we sought to establish preliminary seroprevalence ofleptospirosis exposure in feral hogs in Florida. Blood samples were collected opportunistically from 158 male and 166 female feral hogs taken at managed hunts and by permitted trappers in the northern, central, and southern regions of Florida. Samples were then analyzed using the microscopic agglutination test (MAT) for antibody titers to 20 Leptospira serovars representing 17 serogroups. A titer of > 1:100 was considered positive; 33% (107/324 total samples) were positive to at least one serovar, and 46% of those were positive to multiple serovars. Antibodies to L. interrogans serovar Bratislava strain Jez Bratislava (serogroup Australis) was the most common, with 18% (58/324) testing positive for antibodies. These initial data indicate that there is a significant possibility of feral hogs having a larger role in the complex etiology of leptospirosis in Florida than historically estimated and that further investigation is warranted.

  15. Leptospiral outer membrane protein LipL41 is not essential for acute leptospirosis but requires a small chaperone protein, lep, for stable expression.

    PubMed

    King, Amy M; Bartpho, Thanatchaporn; Sermswan, Rasana W; Bulach, Dieter M; Eshghi, Azad; Picardeau, Mathieu; Adler, Ben; Murray, Gerald L

    2013-08-01

    Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira spp., but knowledge of leptospiral pathogenesis remains limited. However, the development of mutagenesis systems has allowed the investigation of putative virulence factors and their involvement in leptospirosis. LipL41 is the third most abundant lipoprotein found in the outer membranes of pathogenic leptospires and has been considered a putative virulence factor. LipL41 is encoded on the large chromosome 28 bp upstream of a small open reading frame encoding a hypothetical protein of unknown function. This gene was named lep, for LipL41 expression partner. In this study, lipL41 was found to be cotranscribed with lep. Two transposon mutants were characterized: a lipL41 mutant and a lep mutant. In the lep mutant, LipL41 protein levels were reduced by approximately 90%. Lep was shown through cross-linking and coexpression experiments to bind to LipL41. Lep is proposed to be a molecular chaperone essential for the stable expression of LipL41. The roles of LipL41 and Lep in the pathogenesis of Leptospira interrogans were investigated; surprisingly, neither of these two unique proteins was essential for acute leptospirosis.

  16. Outbreak of leptospirosis among triathlon participants in Langau, Austria, 2010.

    PubMed

    Radl, Christoph; Müller, Maria; Revilla-Fernandez, Sandra; Karner-Zuser, Stefanie; de Martin, Alfred; Schauer, Ulrike; Karner, Franz; Stanek, Gerold; Balcke, Peter; Hallas, Andreas; Frank, Herbert; Fürnschlief, Albert; Erhart, Friedrich; Allerberger, Franz

    2011-12-01

    We report on the first documented outbreak of leptospirosis in Austria. In July 2010, four cases of serologically confirmed leptospirosis occurred in athletes after a triathlon held in Langau. Heavy rains preceded the triathlon (rainfall: 22 mm). The index case (Patient A) was a 41-year-old previously healthy male, who was admitted to hospital A on July 8 with a four-day history of fever up to 40°C that began 14 days after attending the triathlon event. On July 7, patient B, a 42-year-old male, was admitted to the same hospital, with signs and symptoms of kidney failure. Hemodialysis was performed every other day for 3 weeks. While the serum drawn on the day of admission was negative for antibodies against Leptospira, a specimen from July 28 tested positive with Leptospira interrogans. On July 11, patient C, a 40-year-old male, was admitted to hospital B for nephritis. On July 14, patient D, a 44-year-old male, was admitted to hospital C with a ten days history of intermittent fever, mild dry cough and headache. Our report underlines that in Austria recreational users of bodies of freshwater must be aware of an existing risk of contracting leptospirosis, particularly after heavy rains. The suppressive influence of a triathlon on the immune system is well documented and therefore an outbreak in this population group can be seen as a sensitive indicator concerning possible risk for the general population.

  17. Recombinant Antigens rLipL21, rLoa22, rLipL32 and rLigACon4-8 for Serological Diagnosis of Leptospirosis by Enzyme-Linked Immunosorbent Assays in Dogs

    PubMed Central

    Ye, Cuilian; Yan, Weiwei; Xiang, Hua; He, Hongxuan; Yang, Maosheng; Ijaz, Muhammad; Useh, Nicodemus; Hsieh, Ching-Lin; McDonough, Patrick L.; McDonough, Sean P.; Mohamed, Hussni; Yang, Zhibang; Chang, Yung-Fu

    2014-01-01

    Animal leptospirosis is one of the most common zoonotic diseases in the United States and around the world.