Isolation and characterization of partially purified leptospiral antigens

The methanol extract of Leptospira interrogans serovar canicola was purified by precipitation with acetone or acetone and chloroform. The antigenicity of the antigen was not altered by heating or treatment with pepsin and pronase. However the antigenicity was lost when the antigen was treated with periodic acid. Chemical analysis revealed the presence of 40% carbohydrate (22% methylpentose, 28%; hexoses),4% protein, 20% lipid and 2,7% phosphate. The complement fixation test with sera from patients with leptospirosis agreed with the microscopic agglutination reaction.

Application of anti-leptospira ELISA-lgM for the etiologic elucidation of meningitis

Leptospirosis is one of the causes of meningitis, although its importance is not well known. In the present study we contributed to this knowledge by demonstrating specific IgM class anti-leptospira antibodies by the immunoenzymatic method ELISA in 14.6% of cerebrospinal fluid (CSF) samples from 171 patients with meningitis considered to be of indeterminate etiology. The frequencies of positivity were similar in cases with predominance of polymorphonuclear or lymphomononuclear leucocytes in the CSF. Age distribution showed a predominance of the 5 to 15 year age range (72%), and sex distribution showed a predominance of males (68%). The authors discuss the contribution of this method to the etiologic elucidation of meningitis.

THE PERSISTENCE OF LEPTOSPIRAL AGGLUTININS TITERS IN HUMAN SERA DIAGNOSED BY THE MICROSCOPIC AGGLUTINATION TEST

The persistence of agglutinins detected by MAT has created some problems to the interpretation of the results. The aim of this study was to examine the data of serology from 70 patients with serologically confirmed diagnosis of leptospirosis by during 3-13 months after being affected with leptospires in order to elucidate the interpretation of the persistence of agglutinins detected by MAT. Sixty-one patients sera (87.14%) had titers equal or greater than 800. Of these, two individuals maintained titers of 800 thirteen months after the onset. This study showed that only one sample of sera with high titers is not reliable to determine the time at which infection occurred.

Borrelia burgdorferi antibodies in dogs from Cotia county, São Paulo State, Brazil

Dogs sera samples collected from Cotia County, São Paulo were tested using indirect immunoenzymatic test (ELISA) in order to study Lyme disease serology in dogs. ELISA method was standardized and G39/40 North American strain of Borrelia burgdorferi was used as antigen. Positive results were confirmed employing the Western blotting technique. Because of the possibility of cross-reactions, sera were also tested for different serological strains of Leptospira interrogans and L. biflexa using microscopic sera agglutination test. Twenty-three of 237 (9.7%) serum samples were positive in the ELISA; 20 of them (86.9%) were confirmed by the Western blotting, what suggests that Cotia may be a risk area for Lyme disease. Although 4 samples (1.7%) were positive for Lyme disease and leptospirosis, no correlation was found between the results (X² = 0.725; p = 0.394) what suggests absence of serological cross reactivity.

Occurrence of antibodies against Leptospira spp. in horses of the urban area of Londrina, Paraná, Brazil

A total of 320 horses were studied in this paper, both male and female, between two and 17 years of age, which were used for traction of wagons in the urban area of the municipality of Londrina (PR). These animals were kept, after their daily work, in abandoned areas or plots, in the outskirts of the urban area of the city. When these animals were attended by the veterinarians, between 1996 and 2005, none of them presented symptoms suggesting leptospirosis. The most frequent reasons for the visit were loss of weight, unwillingness for work, parasitism, laminess, and wounds. Microscopic Seroagglutination Test (SAM), with 22 Leptospira serovars, was performed in sera sample from all these animals. The aim of this study was to investigate the occurrence of antibodies against Leptospira spp. in horses from the urban area of Londrina (PR). From the samples tested, 214 (66.88%) were considered positive, with titers between 100 and 3200, being that 49 (22.90%) presented antibodies against a single serovar of Leptospira, and 165 (77.10%) samples presented antibodies against two or more serovars simultaneously, where in 88 (53.33%) it was possible to characterize the most likely probable serovar. Antibodies against the serovar Icterohaemorrhagiae were detected in 32 (23.36%) animals.

Syndromic surveillance: etiologic study of acute febrile illness in dengue suspicious cases with negative serology. Brazil, Federal District, 2008

With the aim of identifying the etiology of acute febrile illness in patients suspected of having dengue, yet with non reagent serum, a descriptive study was conducted with 144 people using secondary serum samples collected during convalescence. The study was conducted between January and May of 2008. All the exams were re-tested for dengue, which was confirmed in 11.8% (n = 17); the samples that remained negative for dengue (n = 127) were tested for rubella, with 3.9% (n = 5) positive results. Among those non reactive for rubella (n = 122), tests were made for leptospirosis and hantavirus. Positive tests for leptospirosis were 13.9% (n = 17) and none for hantavirus. Non reactive results (70.8%) were considered as Indefinite Febrile Illness (IFI). Low schooling was statistically associated with dengue, rubella and leptospirosis (p = 0.009), dyspnea was statistically associated with dengue and leptospirosis (p = 0.012), and exanthem/petechia with dengue and rubella (p = 0.001). Among those with leptospirosis, activities in empty or vacant lots showed statistical association with the disease (p = 0.013). Syndromic surveillance was shown to be an important tool in the etiologic identification of IFI in the Federal District of Brazil.

HIGH SENSITIVE PCR METHOD FOR DETECTION OF PATHOGENIC Leptospira spp. IN PARAFFIN-EMBEDDED TISSUES

This study describes the development and application of a new PCR assay for the specific detection of pathogenic leptospires and its comparison with a previously reported PCR protocol. New primers were designed for PCR optimization and evaluation in artificially-infected paraffin-embedded tissues. PCR was then applied to post-mortem, paraffin-embedded samples, followed by amplicon sequencing. The PCR was more efficient than the reported protocol, allowing the amplification of expected DNA fragment from the artificially infected samples and from 44% of the post-mortem samples. The sequences of PCR amplicons from different patients showed >99% homology with pathogenic leptospires DNA sequences. The applicability of a highly sensitive and specific tool to screen histological specimens for the detection of pathogenic Leptospira spp. would facilitate a better assessment of the prevalence and epidemiology of leptospirosis, which constitutes a health problem in many countries.

DETECTION OF Leptospira spp. AND Brucella abortus ANTIBODIES IN FREE-LIVING JAGUARS (Panthera onca) IN TWO PROTECTED AREAS OF NORTHERN PANTANAL, BRAZIL

 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.

NEW WILDLIFE HOSTS OF Leptospira interrogans IN CAMPECHE, MEXICO

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.

Factors associated with Leptospira sp infection in a large urban center in northeastern Brazil

Leptospirosis is a zoonotic disease that has emerged to cause epidemics in urban communities in developing countries. However, little is known about the infection in the general population. A seroprevalence survey was performed on a random sample of 1,390 subjects in Salvador, Brazil. Data on environmental and socioeconomic factors were collected. The microagglutination test of serum samples was used to show any prior Leptospira infection. The overall seroprevalence was 12.4%. Among the seropositive individuals, 111 (61%) had high titers for serovars of the Icterohaemorrhagiae serogroup. Seroprevalence increased with age and was similar for males and females. A positive correlation between Leptospira infection and low educational level was found. These findings indicate that a significant proportion of this urban population is exposed to pathogenic Leptospira. Public health actions for leptospirosis control will need to target not only the traditional groups at risk of infection with severe forms of this disease, but also the general population that is at risk.

Acute kidney injury in a tropical country: a cohort study of 253 patients in an infectious diseases intensive care unit

Introduction: Acute kidney injury (AKI) is a frequent and potentially fatal complication in infectious diseases. The aim of this study was to investigate the clinical aspects of AKI associated with infectious diseases and the factors associated with mortality. Methods: This retrospective study was conducted in patients with AKI who were admitted to the intensive care unit (ICU) of a tertiary infectious diseases hospital from January 2003 to January 2012. The major underlying diseases and clinical and laboratory findings were evaluated. Results: A total of 253 cases were included. The mean age was 46±16 years, and 72% of the patients were male. The main diseases were human immunodeficiency virus (HIV) infection, HIV/acquired immunodeficiency syndrome (AIDS) (30%), tuberculosis (12%), leptospirosis (11%) and dengue (4%). Dialysis was performed in 70 cases (27.6%). The patients were classified as risk (4.4%), injury (63.6%) or failure (32%). The time between AKI diagnosis and dialysis was 3.6±4.7 days. Oliguria was observed in 112 cases (45.7%). The Acute Physiology and Chronic Health Evaluation (APACHE) II scores were higher in patients with HIV/AIDS (57±20, p-value=0.01) and dengue (68±11, p-value=0.01). Death occurred in 159 cases (62.8%). Mortality was higher in patients with HIV/AIDS (76.6%, p-value=0.02). A multivariate analysis identified the following independent risk factors for death: oliguria, metabolic acidosis, sepsis, hypovolemia, the need for vasoactive drugs, the need for mechanical ventilation and the APACHE II score. Conclusions: AKI is a common complication in infectious diseases, with high mortality. Mortality was higher in patients with HIV/AIDS, most likely due to the severity of immunosuppression and opportunistic diseases.

Seroprevalence and seroincidence of Leptospira infection in dogs during a one-year period in an endemic urban area in Southern Brazil

INTRODUCTION: Leptospirosis is a zoonosis that affects both humans and animals. Dogs may serve as sentinels and indicators of environmental contamination as well as potential carriers for Leptospira. This study aimed to evaluate the seroprevalence and seroincidence of leptospirosis infection in dogs in an urban low-income community in southern Brazil where human leptospirosis is endemic. METHODS: A prospective cohort study was designed that consisted of sampling at recruitment and four consecutive trimestral follow-up sampling trials. All households in the area were visited, and those that owned dogs were invited to participate in the study. The seroprevalence (MAT titers ≥100) of Leptospira infection in dogs was calculated for each visit, the seroincidence (seroconversion or four-fold increase in serogroup-specific MAT titer) density rate was calculated for each follow-up, and a global seroincidence density rate was calculated for the overall period. RESULTS: A total of 378 dogs and 902.7 dog-trimesters were recruited and followed, respectively. The seroprevalence of infection ranged from 9.3% (95% CI; 6.7 - 12.6) to 19% (14.1 - 25.2), the seroincidence density rate of infection ranged from 6% (3.3 - 10.6) to 15.3% (10.8 - 21.2), and the global seroincidence density rate of infection was 11% (9.1 - 13.2) per dog-trimester. Canicola and Icterohaemorraghiae were the most frequent incident serogroups observed in all follow-ups. CONCLUSIONS: Follow-ups with mean trimester intervals were incapable of detecting any increase in seroprevalence due to seroincident cases of canine leptospirosis, suggesting that antibody titers may fall within three months. Further studies on incident infections, disease burden or risk factors for incident Leptospira cases should take into account the detectable lifespan of the antibody.

Progression of the load of waterborne and intestinal parasitic diseases in the State of Amazonas

In the State of Amazonas, Brazil, urban expansion together with precarious basic sanitation conditions and human settlement on river banks has contributed to the persistence of waterborne and intestinal parasitic diseases. Time series of the recorded cases of cholera, typhoid fever, hepatitis A and leptospirosis are described, using data from different levels of the surveillance systems. The sources for intestinal parasitosis prevalence data (non-compulsory reporting in Brazil) were Medical Literature Analysis and Retrieval System Online (MEDLINE), Literatura Latino-Americana (LILACS) and the annals of major scientific meetings. Relevant papers and abstracts in all languages were accessed by two independent reviewers. The references cited by each relevant paper were scrutinized to locate additional papers. Despite its initial dissemination across the entire State of Amazonas, cholera was controlled in 1998. The magnitude of typhoid fever has decreased; however, a pattern characterized by eventual outbreaks still remains. Leptospirosis is an increasing cause of concern in association with the annual floods. The overall prevalence of intestinal parasites is high regardless of the municipality and the characteristics of areas and populations. The incidence of hepatitis A has decreased over the past decade. A comparison of older and recent surveys shows that the prevalence of intestinal parasitic diseases has remained constant. The load of waterborne and intestinal parasitic diseases ranks high among the health problems present in the State of Amazonas. Interventions aiming at basic sanitation and vaccination for hepatitis A were formulated and implemented, but assessment of their effectiveness in the targeted populations is still needed.

Comparison of a Recombinant-antigen Enzyme Immunoassay with Treponema pallidum Hemagglutination Test for Serological Confirmation of Syphilis

A recombinant-antigen enzyme immunoassay (EIA), BioSCREEN TM anti-Treponema pallidum, was compared favorably with the T. pallidum hemagglutination test, in the detection of specific antibodies in different groups of sera from patients with primary (n = 38), secondary (n = 10), early latent (n = 28) and congenital syphilis (n = 2), patients with leptospirosis ( n= 8), infectious mononucleosis (n = 7), hepatitis (n = 9), diabetes mellitus (n = 11), rheumatoid arthritis (n = 13), leprosy (n = 11), tuberculosis (n = 9), HIV/Aids ( n= 12), systemic lupus erythematosus (n = 4), rheumatic fever (n = 3), old-persons (n = 9), pregnant women (n = 29) and blood donors (n = 164). The coincidence between them was 95.1%. The sensitivity and specificity of the EIA were 93.3% and 95.5%, respectively. Fifteen serum specimens belonging to old-persons, pregnant women, blood donors, and patients with human leptospirosis, hepatitis, diabetes mellitus, tuberculosis and rheumatic fever gave false-positive results by Venereal Disease Research Laboratory and/or Rapid Plasma Reagin. The EIA can be used as alternative method for the serological confirmation of syphilis.

Molecular characterization of Leptospira sp. strains isolated from human subjects in São Paulo, Brazil using a polymerase chain reaction-based assay: a public health tool

A polymerase chain reaction (PCR)-based assay which amplifies repetitive DNA elements present within bacterial genomes was used to characterize and differentiate Leptospira sp. Thirty-five strains from a reference culture collection and 18 clinical isolates which had been previously analyzed by cross agglutinin absorption test (CAAT) were evaluated by this technique. PCR results from analysis of the reference culture collection showed no bands corresponding to serogroups Australis, Autumnalis, Bataviae, Celledoni, Cynopteri, Djasiman, Panama, Pomona, Pyrogenes, and Tarassovi. However, the PCR method was able to clearly discriminate the serogroups Andamana, Ballum, Canicola, Grippotyphosa, Hebdomadis, Icterohaemorrhagiae, Javanica, Sejroe, Semaranga, and Shermani. Clinical isolates previously characterized by CAAT as serovar Copenhageni, serovar Castellonis, and as serovar Canicola were in agreement with PCR results. The clinical isolate previously characterized as serovar Pomona was not differentiated by PCR. Forty additional clinical isolates from patients with leptospirosis obtained in São Paulo, Brazil were also evaluated by this PCR method. Thirty-nine of these were determined to belong to serogroup Icterohaemorrhagiae (97.5%) and one to serogroup Sejroe (2.5%). These results demonstrate that the PCR method described in this study has utility for rapid typing of Leptospira sp. at the serogroup level and can be used in epidemiological survey.