Leptospirosis is a zoonosis of multisystem involvement caused by pathogenic strains of the genus Leptospira.In the last few years, intensive studies aimed at the development of a vaccine have provided important knowledge about the nature of the immunological mechanisms of the host. The purpose of this study was to analyze the immune responses to two recombinant proteins, MPL17 and MPL21 (encoded by the genes LIC10765 and LIC13131, respectively) of Leptospira interrogans serovar Copenhageni in individuals during infection. The recombinant proteins were expressed in Escherichia coli as six-His tag fusion proteins and were purified from the soluble bacterial fraction by affinity chromatography with Ni 2؉ -charged resin. The recombinant proteins were used to evaluate their ability to bind to immunoglobulin G (IgG) (and IgG subclass) or IgM antibodies in serum samples from patients in the early and convalescent phases of leptospirosis (n ‫؍‬ 52) by enzyme-linked immunosorbent assays. The prevalences of total IgG antibodies against MPL17 and MPL21 were 38.5% and 21.2%, respectively. The titers achieved with MPL17 were statistically significantly higher than those obtained by the reference microscopic agglutination test. The specificity of the assay was estimated to be 95.5% for MPL17 and 80.6% for MPL21 when serum samples from individuals with unrelated febrile diseases and control healthy donors were tested. The proteins are conserved among Leptospira strains that cause human and animal diseases. MPL17 and MPL21 are most likely new surface proteins of leptospires, as revealed by liquid-phase immunofluorescence assays with living organisms. Our results demonstrate that these recombinant proteins are highly immunogenic and, when they are used together, might be useful as a means of diagnosing leptospirosis.

A collection of 101 Leptospira isolates was tested by multilocus sequence typing (MLST) and by traditional serotyping. MLST divided the isolates into 4 sequence types (STs), while serotyping classified them into 6 serogroups. Two isolates failed to generate products for some genes by MLST. MLST was less discriminatory than serotyping for uncommonly occurring isolates from humans in Brazil.Leptospirosis, caused by pathogenic leptospires, is one of the most widespread zoonotic diseases known. The genus Leptospira is divided into 20 species with more than 200 pathogenic serovars organized into 24 serogroups on the basis of antigenic relatedness (2,9,13,30). The disease occurs in wild and domesticated animals, both of which can be a source of human infection. Exposures that occur during flooding events are the main risk factors of human leptospirosis in Brazil (1,3,14,22). In São Paulo State, Brazil, 13,620 cases have been reported in the last 20 years (ftp://ftp.cve.saude.sp.gov.br/doc_tec/zoo /lepto09_perfil.pdf). However, information about circulating isolates of Leptospira spp. in Brazil is limited. Identification of isolates to the serovar level is essential for understanding the epidemiology of the disease in both humans and animals.Since certain serovars are often associated with specific mammalian hosts and with the symptoms and severity of the disease, the identification of the serovar usually permits the prediction of sources of infection, thereby enabling control of the spread of the disease (17). Monitoring the serovars and genetic profiles of strains collected over time and from different regions is also important to better understand the current circulating Leptospira population worldwide. Isolate discrimination can be performed by a microagglutination test (MAT) at the serogroup level and by a cross-agglutinin absorption test (CAAT) at the serovar level (5). However, performing those methods is tedious, as live cultures of collection strains must be maintained for use as antigens and rabbit hyperimmune sera are required. Although identification by serotyping is valuable, molecular methods with higher reproducibility and discriminatory power may be more useful in epidemiological investigations.New molecular methods such as multilocus sequence typing (MLST) have been recently developed and applied to the study of many bacterial species (4,6,7,15,18). MLST is a simple PCR-based technique that makes use of automated DNA sequencers to assign and characterize the alleles present in different target genes. The method allows one to generate sequence data on a low-to high-throughput scale that is unambiguous and suitable for epidemiological and population studies. The selected loci are generally housekeeping genes, which evolve very slowly over an evolutionary time scale (8).Our goal was to evaluate the discriminatory power of MLST compared to serotyping using a set of Brazilian human isolates. A total of 101 Leptospira clinical strains (95 from blood, 5 from cerebrospinal fluid, and 1 from urine) isolated fro...

Leptospirosis is a zoonotic infection caused by pathogenic members of the genus Leptospira spp. Knowledge of the prevalent serovars and their maintenance hosts is essential to understand the disease. The aim of this study was to evaluate the ability of serology by the microscopic agglutination test (MAT) to predict the serogroups compared with results of identification of leptospires in São Paulo, Brazil. MAT correctly assigned the serogroup of the infecting isolate in 49/52 cases (94.23%). The serogroup Icterohaemorrhagiae was the predominant serogroup (88.46%). This study showed the usefulness of the MAT to correctly identify the infecting serogroup with a good overall agreement between the serologically-identified infecting serogroup and by identification of the isolate and can be used in epidemiological surveys in São Paulo. However, it should be complemented by the identification of Leptospira isolates.

Aims:  Leptospirosis is a public health problem worldwide. Traditionally, microscopic agglutination test (MAT) and cross‐agglutinin absorption test (CAAT) are used to identify leptospires. However, these techniques are laborious and time‐consuming, requiring the maintenance of a collection of more than 200 reference strains and correspondent rabbit antisera. The purpose of this study was to evaluate the pulsed‐field gel electrophoresis (PFGE) method for discrimination of Leptospira serovars. Methods and Results:  Fourteen clinical isolates of Leptospira spp. were analysed by MAT before being characterized by PFGE. The isolates were compared with a library of 206 different reference Leptospira serovars. All the isolates gave clear profiles with high resolution. PFGE and MAT results were in agreement for all clinical isolates evaluated. Twelve isolates were classified as serovar Icterohaemorrhagiae/Copenhageni by PFGE. By MAT, these isolates were classified as serogroup Icterohaemorrhagiae with titres ranging from 3200 to 25 600. Two isolates were classified as serovar Canicola by PFGE, and as serogroup Canicola by MAT with titres higher than 3200. Conclusions:  PFGE offers the advantages of simple, reliable and reproducible results. Significance and Impact of the Study:  PFGE provides a convenient tool for the identification of clinical isolates.

Objective: Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira spp. The aim of this study was to report the incidence of leptospirosis from 1998 to 2012 in the state of São Paulo, Brazil to show the importance of human leptospirosis and to describe some epidemiological characteristics. Methods: From January 1998 to December 2012, sera from patients with suspected leptospirosis were analyzed. The microscopic agglutination test (MAT) was used for serological investigations and MLST, serotyping and PFGE methods for the identification of leptospires. The descriptive seasonal analysis was performed with Excel Microsoft version 2007. Pearson's correlation was used to assess the association between rainfall and the number of cases. Results: Among 22,795 serum samples, 2,430 cases of leptospirosis were laboratory confirmed, giving an average incidence rate of 1.35/100,000 inhabitants. Of these patients, 2,032 (83.62%) were male with a predominance in the age groups of 21-50 years. The highest incidence and rainfall were from December to April. There was correlation between the rainfall and the number of cases. Icterohaemorrhagiae was the predominant serogroup. Conclusions: This study shows that leptospirosis is a seasonal disease in São Paulo with most cases occurring during the rainy season, and thus, will continue to be a disease of public health importance.

Aims:  Development of a simple, specific, rapid and inexpensive Dot‐ELISA test for early diagnosis of human leptospirosis. Methods and Results:  Serum samples from 90 patients diagnosed with leptospirosis were analysed by Dot‐ELISA test incorporating Glycolipoprotein (GLP) antigen from serovars Copenhageni and Patoc. Results were compared with those obtained with microscopic agglutination test, currently, the gold standard reference serological method. Serum samples from healthy blood bank donors and patients diagnosed with diseases other than leptospirosis were used as negative controls. The specificities of both GLP‐based assays were 97·1% and 100% with serum samples from patients with other diseases and with serum samples from healthy control group, respectively. With serum samples from patients with acute leptospirosis, sensitivity was 76·6% with Dot‐ELISA Copenhageni and 90·0% with Dot‐ELISA Patoc. With serum samples from patients in convalescence, sensitivity was 100% with both GLP‐based assays. Conclusions:  This Dot‐ELISA provides a candidate antigen for serodiagnosis of leptospirosis during all phases of illness and could be a good alternative method for the early diagnosis of leptospirosis. Significance and Impact of the Study:  The Dot‐ELISA test is simple, specific, rapid and inexpensive. It is suitable for identifying a large number of samples and, hence, reducing the death rate of patients with leptospirosis.

Leptospirosis is a zoonotic disease caused by the pathogenic

Household contacts are important sources of Bordetella pertussis in infants. A total of 353 household contacts of 97 index cases were evaluated for pertussis by culture and polymerase chain reaction. Twenty eight contacts were positive (8.0%). The presence of symptoms did not influence the rate of diagnosed bacteriologic pertussis in communicants. We conclude that contacts with an index case can be positive for B. pertussis independently of the presence of symptoms.