Highly Pathogenic Leptospira Found in Urban Brown Rats (Rattus norvegicus) in the Largest Cities of Sweden.

Leptospirosis is an emerging zoonosis of global concern; however, its contemporary occurrence in Sweden, a European country partly located north of the Arctic Circle, is poorly known. Four out of 30 brown rats, captured within urban districts in Sweden, were found to be positive for antibodies to Leptospira interrogans serovar Icterohaemorrhagiae. This serovar causes Weil's disease in humans, a severe infection with jaundice, renal failure, and hemorrhage. Our study is the first finding of this highly pathogenic serovar in Swedish rats since the 1930s.

Waterborne Leptospirosis: Survival and Preservation of the Virulence of Pathogenic Leptospira spp. in Fresh Water.

Many studies have implicated fresh water as a source of leptospirosis outbreaks. To estimate the survival and the preservation of the virulence of pathogenic Leptospira spp. maintained in water, we selected five still waters with various pH and mineral profiles. The water samples were artificially inoculated with a culture of a pathogenic strain belonging to serovar Icterohaemorrhagiae. Samples were stored for 20 months at 4, 20 or 30 °C. The survival and preservation of virulence of this pathogenic strain was estimated by subculturing these stored samples. After 14 and 20 months of storage, the strain Icterohaemorrhagiae was re-isolated, and its virulence was determined using an animal model. In these waters, the mean survival was 130 days for storage at 4 °C, 263 days at 20 °C, and 316 days at 30 °C. Unexpectedly, the mean survival was 344 days for a final pH < 7 and 129 days for pH ≥ 7. Moreover, the pathogenic strain remained fully virulent and was able to induce a lethal disease in gerbils even when the pH of the contaminated waters decreased to <6. These data showed that despite unfavourable storage conditions such as cold, nutrient-poor acidic waters, the survival and virulence of pathogenic Leptospira spp. was fully preserved over at least 20 months.

Role of 72 kDa protein of Leptospira interrogans as a diagnostic marker in acute leptospirosis.

BACKGROUND & OBJECTIVES: Leptospirosis is a widespread zoonotic disease and a public health problem, particularly in tropical and subtropical countries. Varied clinical manifestations of the disease frequently lead to misdiagnosis resulting in life-threatening multi-organ complications. Therefore, early laboratory investigation using an appropriate diagnostic approach is crucial. In the present study, a potential protein marker was identified and evaluated for its usefulness in the serodiagnosis of acute leptospirosis. METHODS: Leptospira interrogans serovar Icterohaemorrhagiae (L44), which represents a commonly prevalent serovar in Malaysia, was cultivated for preparation of sequential protein extract (SEQ). SDS-PAGE and immunoblotting were performed with a serum panel comprising confirmed cases of leptospirosis and controls (n=42 each). Identification and characterization of the highest scoring protein from the antigenic band was performed. Subsequently based on the nucleotide coding sequence of the protein, the corresponding recombinant protein was custom-produced. It was then evaluated for sensitivity and specificity by testing against 20 serum samples from leptospirosis patients and 32 from controls. RESULTS: Among the antigenic components, a 72 kDa protein band demonstrated significant sensitivity (83.3%) and specificity (95.2%) for the detection of specific anti-leptospiral IgM antibodies. The protein was identified by mass-spectrometry analysis as heat shock protein DnaK of L. interrogans. Recombinant form of the protein (r72SEQ) showed 85 per cent sensitivity and 81 per cent specificity for the detection of specific anti-leptospiral IgM antibodies. INTERPRETATION & CONCLUSIONS: The findings of our study indicate that a protein (72 kDa) of L. interrogans has the potential utility of being used for the diagnosis of acute leptospirosis. Further studies need to be done to confirm these findings.

[Fever and jaundice... and if it was a leptospirosis. About a case of L. interrogans icterohaemorrhagiae in Northern France]. / Ictère fébrile et si c'était une leptospirose. À propos d'un cas de L. interrogans Icterohaemorrhagiae dans le Nord de la France.

Leptospirosis is an anthropozoonose, an animal disease transmissible to humans, caused by a spirochete of the genus Leptospira that lives mainly among rodents but also in wetlands. It occurs worldwide, particularly in Asia, Latin America and Africa. In Europe, the incidence is small (except in France and Great Britain, where its frequency has increased in recent years) but the frequency may be underestimated. Some areas overseas are particularly affected. In France, the potential epidemic of leptospirosis is subject to climatic variations, justifying a constant monitoring of the disease provided by the National Reference Centre (CNR) of leptospires. Transmission to humans primarily occurs through contact with environments contaminated by the urine of infected animals. The disease can affect the liver and kidneys (hepatonephritis) as cytolysis, cholestasis and renal failure associated with fever. A coagulopathy usually accompanies the clinical table. Its diagnosis is difficult because of the clinical polymorphism. Early diagnosis of leptospirosis allows effective medical care, improving patient outcomes. This is currently based on gene amplification (PCR) or serology positive by the microscopic agglutination test (MAT), which is the reference method. Its evolution is usually favorable with appropriate antibiotic treatment (aminopenicillin). However 5-10% of symptomatic patients have a severe multisystem defaillance. Nearly a century after the discovery of the causative agent, this zoonosis remains a public health problem, zoonosis priority in terms of virulence, its reporting is mandatory in our country. We report the case of a severe form of hepatonephritis due to water contaminated with Leptospira observed in Northern France.

Molecular characterization of virulent Leptospira interrogans serogroup icterohaemorrhagiae isolated from Cavia aperea.

Leptospirosis is a worldwide zoonotic infection caused by pathogenic Leptospira. Synanthropic rodents are recognized carriers of leptospires; however, the role of wild rodents in the epidemiology of the disease is still incipient. In this work, we describe Leptospira strain isolated from Cavia aperea (Brazilian guinea pig). The isolated strain was characterized by partial rpoB gene sequencing, variable-number tandem-repeats and histopathological analysis. The strain was identified as Leptospira interrogans, serogroup Icterohaemorrhagiae and caused clinical signs of leptospirosis in the hamster model, attesting to its virulence. In conclusion, these findings could be useful for elucidating the epidemiological role of C. aperea in leptospirosis.

Cholinesterases as markers of the inflammatory process in rats infected with Leptospira interrogans serovar Icterohaemorrhagiae.

The aim of this study was evaluate changes in the cholinesterase activity in blood, lymphocytes and serum of rats infected with Leptospira interrogans serovar Icterohaemorrhagiae ('L. icterohaemorrhagiae'). Sixty adult Wistar rats were divided into six groups of 10 animals: three control groups and three test groups. The animals from the test groups were intraperitoneally inoculated with 1 ml medium containing 1 × 10(8) leptospires. The activity of acetylcholinesterase in blood and butyrylcholinesterase in serum increased on days 5 (P<0.05) and 30 (P<0.021) post-infection, respectively. A decrease in lymphocyte count was observed on days 15 (P<0.01) and 30 post-infection (P<0.05). On day 15 post-infection, acetylcholinesterase activity (P<0.001) in lymphocytes decreased in infected rats. However, on day 30 post-infection there was an increase in acetylcholinesterase activity in lymphocytes. In conclusion, our results showed that the activity of enzymes of the cholinergic system in the total blood, lymphocytes and serum is altered as a result of inflammation caused by infection with L. icterohaemorrhagiae. The possible causes of these alterations will be discussed in this paper.

Toll-like receptor 4 protects against lethal Leptospira interrogans serovar icterohaemorrhagiae infection and contributes to in vivo control of leptospiral burden.

The roles of innate immune responses in protection from or pathogenesis of severe leptospirosis remain unclear. We examined the role of Toll-like receptors (TLRs) in mouse infection and macrophage responses to Leptospira. C3H/HeJ mice (TLR4 deficient) and C3H/HeJ-SCID mice, but not C3H/OuJ mice (TLR4 intact), died after intraperitoneal infection with Leptospira interrogans serovar Icterohaemorrhagiae. Death in both C3H/HeJ mouse strains was associated with jaundice and pulmonary hemorrhage, similar to the patient from whom the isolate was obtained. In chronic sublethal infection, TLR4-deficient mice harbored more leptospires in liver, lung, and kidney than control mice. Heat-killed Leptospira stimulated macrophages to secrete proinflammatory cytokines, tumor necrosis factor alpha, interleukin-6, and macrophage inflammatory protein 2 not inhibited by polymyxin B, suggesting that leptospiral lipopolysaccharide (LPS) did not drive these responses. Anti-TLR4 and anti-MD-2 but not anti-CD14 monoclonal antibodies inhibited cytokine production. Peritoneal macrophages from CD14-/- and TLR2-/- mice exhibited no defect in cytokine responses to Leptospira compared to controls. Macrophages from C3H/HeJ, TLR4-/-, and MyD88-/- mice secreted far-lower levels of cytokines than wild-type macrophages in response to Leptospira. TLR4 plays a crucial role in protection from acute lethal infection and control of leptospiral burden during sublethal chronic infection. Cytokine responses in macrophages correlated with leptospiral clearance. These TLR4-dependent but CD14/TLR2-independent responses are likely mediated by a leptospiral ligand(s) other than LPS.

Regulation of complement activation at the C3-level by serum resistant leptospires.

Leptospires are spirochetes that are transmitted to humans through contacts with wild or domestic animals or via an exposure to contaminated soil or water. In this study we have compared the serum-sensitivity of five pathogenic strains of leptospires (L. interrogans) to an environmental isolate (strain Patoc). Different levels of sensitivities to human serum were seen. Interestingly, the most sensitive strain was the non-pathogenic Patoc strain. The fully and intermediately resistant strains have been isolated from human patients. Testing was performed in the absence of specific antibodies, and killing was found to be dependent on the complement system. The serum sensitive Patoc strain was killed in human serum within minutes, whereas the most resistant strains tolerated serum up to 4h. We also tested the deposition of the complement components C3, C5, C6, C8 and C5b-9 to the surfaces of the sensitive and resistant strains of Leptospira by immunofluorescence microscopy and ELISA. C3 was deposited on both the sensitive and resistant strains, but the terminal complement components were detected only on the surface of the complement-sensitive strain. The complement resistant and intermediate strains were found to bind more factor H from human serum than the complement sensitive strain. Thus, binding of this major alternative complement pathway inhibitor is related to serum resistance in Leptospira spirochetes.

[Efficacy of Spirolept vaccine against human leptospirosis as estimated by passive protection of laboratory rodents]. / Etude de l'efficacité du vaccin Spirolept contre la leptospirose par la protection passive de rongeurs de laboratoire.

OBJECTIVE: This study had for aim to assess the serological response induced by the Spirolept vaccine against human leptospirosis. METHOD: A serological follow-up was made on 31 patients at a risk of occupational exposure. The antibody titers of vaccinated patients were assessed by MAT and ELISA. In a second step, vaccinal protection was studied in vivo by checking the seroprotective effect of the human sera injected in an animal model (Meriones unguiculatus) naturally susceptible to the disease. The passive protection was studied by comparing the death rate on five batches of animals to which the bacterium was inoculated. Thus, four batches of animals were injected subcutaneously with a pooled sera of vaccinated people sampled at D0, D15, D135, and D320 after Spirolept vaccination. One control batch was given PBS. One day after injection, the latter batch was inoculated with the homologous strain Verdun of Leptospira interrogans ss icterohemorrhagiae (serogroup Icterohemorrhagiae) used to make the vaccine. RESULTS: The death rate was significantly decreased as soon as D15 after the first injection, even with pooled sera of vaccinated people negative for the MAT. COMMENTS: The Spirolept vaccine induces a protective response against icterohemorrhagiae, which can be transmitted to the animal model and thus is linked to a humoral response.