Correlation Between Physicochemical Properties of Soil and Presence of Leptospira.

Leptospirosis is an important global public health problem. Favourable environmental factors are influencing the survival of leptospires in soil, which is an important link in the transmission cycle. The present study was designed to understand the correlation between various soil nutrients and presence of Leptospira in soil samples of different regions of Andaman and Nicobar Islands. The study revealed a significant positive relationship between presence of Leptospira and concentration of iron, manganese and copper in soil. Presence of iron, manganese and copper in the soil may influence the survival and transmission of leptospirosis.

Changing trend in the seroprevalence and risk factors of human leptospirosis in the South Andaman Island, India.

Seroprevalence of leptospirosis among a healthy population of the South Andaman Island was assessed through random sampling. Previous studies have high seroprevalences of up to 55% in general population and 65% in agricultural labourers. The study subjects (1,181 in total, 781 rural and 400 urban) were interviewed and tested for antibodies against Leptospira. Multivariate models were developed to determine the risk factors in the rural and the urban population. The overall seroprevalence was 10.9%, with rural (12.9%) being higher than the urban subjects (7.0%). The commonest infecting serogroup was Icterohaemorrhagiae (53.5%), followed by Grippotyphosa (13.2%). Compared to the earlier observation, seroprevalence was lower and an apparent shift in the infecting serogroup was found. This shift was in concordance with the changing trend in animal population. Significant difference in risk factors, both in rural and urban areas, was also observed. Similar trends in seroprevalence are being observed around the world. Therefore, time to time prevalence studies are needed for the development of effective control measure.

Can Subminimal Inhibitory Concentrations of Antibiotics Induce the Formation of Biofilm in Leptospira?

Antibiotics at subminimal inhibitory concentrations (sub-MICs) are known to induce biofilm formation in numerous bacteria in vitro. In this report, the effect of sub-MIC levels of antibiotics (doxycycline and tetracycline) on biofilm formation by leptospiral reference strains and isolates was investigated. The sub-MIC levels of both tetracycline and doxycycline were able to induce biofilm in some of the leptospiral strains. This is the first report demonstrating the effect of sub-MIC level of antibiotics in inducing biofilm formation in Leptospira. The induction of biofilm may solely be a response to the amount of threshold stress enforced by low levels of antibiotics. The mechanism of biofilm induction by subinhibitory antibiotic concentrations needs to be explored further. Studies are required to understand the clinical relevance of the phenomenon and its contribution to biofilm formation in the host, resulting in the failure of antimicrobial therapy during the treatment of chronic leptospirosis.

In Vitro Antimicrobial Susceptibility of Pathogenic Leptospira Biofilm.

Pathogenic Leptospira spp. are the causative agent of leptospirosis. Biofilm formation in leptospires is a new area of study, and its role in pathogenesis is not fully explored. As in other biofilm-forming bacteria, Leptospira biofilm may play a significant role in antibiotic resistance. In this study, the antimicrobial susceptibility of Leptospira biofilm was investigated by 96-well plate assay using Alamar Blue. Leptospira biofilm showed five to sixfold increase in resistance in all the strains used. The range of minimal bactericidal concentrations for penicillin G, ampicillin, tetracycline, and doxycycline was 1,600 U/ml, 800-1,600 µg/ml, 800-1,600 µg/ml, and 800-1,600 µg/ml, respectively. In agar substrate, the biofilm showed six- to sevenfold increase in resistance to antibiotics compared to planktonic cell. The present study emphasizes the importance of biofilm formation and its antibiotic susceptibility patterns. This could pave the way for devising appropriate strategy to prevent the occurrence of potential chronic leptospirosis in endemic areas and also during an outbreak situation.

Detection of LipL32-specific IgM by ELISA in sera of patients with a clinical diagnosis of leptospirosis.

Successful treatment of leptospirosis is heavily dependent on early diagnosis and prompt initiation of antibiotic therapy. An ELISA test to detect specific IgM antibodies against LipL32 for early diagnosis of leptospirosis is described and evaluated here. One thousand one hundred and eighty sera from clinically suspected leptospirosis cases were enrolled together with 109 healthy volunteers selected from an endemic area between October 2007 and January 2010. Patients were categorized based on their clinical signs and symptoms. Sera were screened for leptospiral antibodies by the microscopic agglutination test (MAT) using a panel of locally circulating serovars followed by enzyme-linked immunosorbent assay (ELISA) based on recombinant LipL32 from Leptospira interrogans serovar Autumnalis strain N2. The sensitivity and specificity of the ELISA test were determined to establish its diagnostic efficiency. The cut-off value was determined to be 0·205. Overall sensitivity and specificity compared to the MAT were found to be 96·4 and 90·4%, respectively. The LipL32-specific IgM ELISA had good sensitivity and acceptable specificity and may be a candidate for the early serodiagnosis of human leptospirosis.

Seroprevalence of Leptospira borgpetersenii serovar javanica infection among dairy cattle, rats and humans in the Cauvery river valley of southern India.

Leptospirosis is a major problem of dairy farms in Tamilnadu, India, resulting in abortions, stillbirths and infertility. Serologic and genetic analyses of samples from cattle, humans and rodents were performed in order to estimate infection prevalence and identify leptospiral species. Five hundred and fifteen sera and 76 urine samples were collected from dairy cattle on 25 farms including a farm that practiced rat control. Sera and kidney samples were also collected from field rats (Rattus norvegicus) in the vicinity of these farms. In addition, sera were collected from farm workers. Serum antibody was measured by the microscopic agglutination test. Leptospires isolated from blood, kidney, and urine were characterized as to serovar. Genomospecies were predicted using random amplified polymorphic DNA (RAPD) profiling. SecY gene sequencing was performed as a tool for tracing of source. Seroprevalence of 87.%, 51.% and 76.5% for cattle, rats and humans, respectively, was observed on endemic farms. Prevalences on a non-endemic farm were lower. Antibodies to Autumnalis, Javanica, Icterohaemorrhagiae and Pomona predominated in both cattle and rats. Thirteen isolates from rat kidneys were identified as serogroup Javanica, serovar Javanica. RAPD comparisons and secY gene sequencing identified these isolates as Leptospira borgpetersenii. These results altogether indicated that L. borgpetersenii was the dominant species in these areas with serovar Javanica apparently derived from rats which provided an important source of infection in cattle resulting a high incidence of infertility, abortion and.still-birth in the Cauvery river valley, Tiruchirappalli, Tamilnadu.

FlaB PCR-based identification of pathogenic leptospiral isolates.

BACKGROUND/PURPOSE: The genus Leptospira comprises pathogenic and saprophytic strains. Conventional methods for the identification of pathogenic leptospiral isolates are cumbersome and laborious. In view of these limitations, the search for alternative methods have been focused on DNA based techniques. In this study, we have developed an effective method for the rapid identification of pathogenic and saprophytic leptospiral isolates based on DNA-based techniques. METHODS: A polymerase chain reaction(PCR)-based approach was developed using specific primer sets (flaB, G1-G2, B64I-II, and A-B) to differentiate pathogenic and saprophytic leptospiral strains. Fifty-five leptospiral isolates were used for this study. The pathogenic status of the isolates was compared with the results obtained using conventional techniques, which included growth in the presence of 8-azaguanine and growth at 13 degrees C. RESULTS: In this analysis, 46 leptospiral isolates were confirmed as pathogenic and nine were confirmed as saprophytic. PCR with the A-B primer set yielded an amplified product of 331 bp in all of the pathogenic and saprophytic isolates. The other primer sets, G1-G2, B64I-II and flaB, yielded products of 258 bp, 568 bp, and 793 bp, respectively, exclusively for the pathogenic leptospiral strains. None of the saprophytic strains yielded products with these primer sets. CONCLUSION: The flaB-specific primers consistently yielded an amplification product for all of the pathogenic leptospiral isolates, indicating the presence of the flaB gene only among pathogenic leptospires, and making this a useful tool for distinguishing between pathogenic and saprophytic leptospires. The efficiency of PCR-based identification corroborates the implementation of these techniques for the identification of pathogenic and saprophytic leptospiral strains.

Characterization of Leptospira borgpetersenii isolates from field rats (Rattus norvegicus) by 16S rRNA and lipL32 gene sequencing

The main goal of this study was to evaluate the prevalence of leptospirosis among field rodents of Tiruchirappalli district, Tamil Nadu, India. In total 35 field rats were trapped and tested for seroprevalence by the microscopic agglutination test (MAT). Isolation of leptospires was performed from blood and kidney tissues and characterized to serovar level. Genomospecies identification was carried out using 16S rRNA and lipL32 gene sequencing. The molecular phylogeny was constructed to find out species segregation. Seroprevalence was about 51.4 percent, and the predominant serovars were Autumnalis, Javanica, Icterohaemorrhagiae and Pomona. Two isolates from the kidneys were identified as serovar Javanica of Serogroup Javanica, and sequence based molecular phylogeny indicated these two isolates were Leptospira borgpetersenii.

Characterization of leptospira borgpetersenii isolates from field rats (rattus norvegicus) by 16s rrna and lipl32 gene sequencing.

The main goal of this study was to evaluate the prevalence of leptospirosis among field rodents of Tiruchirappalli district, Tamil Nadu, India. In total 35 field rats were trapped and tested for seroprevalence by the microscopic agglutination test (MAT). Isolation of leptospires was performed from blood and kidney tissues and characterized to serovar level. Genomospecies identification was carried out using 16S rRNA and lipL32 gene sequencing. The molecular phylogeny was constructed to find out species segregation. Seroprevalence was about 51.4 %, and the predominant serovars were Autumnalis, Javanica, Icterohaemorrhagiae and Pomona. Two isolates from the kidneys were identified as serovar Javanica of Serogroup Javanica, and sequence based molecular phylogeny indicated these two isolates were Leptospira borgpetersenii.