Enno Luge

Resurgence of Field Fever in a Temperate Country: An Epidemic of Leptospirosis among Seasonal Strawberry Harvesters in Germany in 2007

BACKGROUND Although leptospirosis is a reemerging zoonosis of global importance, outbreaks related to agricultural exposures are primarily situated in tropical countries. In July 2007, a suspected leptospirosis outbreak was recognized among strawberry harvesters from Eastern Europe who were working in Germany. An investigation was initiated to identify the outbreak source and the risk factors for infection. METHODS We conducted a retrospective cohort study with use of a questionnaire administered to harvesters by health authorities in Romania, Slovakia, and Poland. Collected serum samples were tested by microscopic agglutination test and immunoglobulin M enzyme-linked immunosorbent assay. A case patient was defined as a person who worked in the strawberry field during the period 5 June-8 September 2007 and had leptospirosis-compatible symptoms and either an antibody titer 1:800 and a positive immunoglobulin M enzyme-linked immunosorbent assay result (for a confirmed case) or no serological confirmation (for a suspected case). Local rodents were examined for leptospirosis. RESULTS Among 153 strawberry harvesters, we detected 13 confirmed case patients who had test results positive for antibodies against Leptospira species serogroup Grippotyphosa and 11 suspected case patients (attack rate, 16%). Risk of disease increased with each day that an individual worked in the rain with hand wounds (odds ratio, 1.1; 95% confidence interval, 1.04-1.14) and accidental rodent contact (odds ratio, 4.8; 95% confidence interval, 1.5-15.9). Leptospires of the serogroup Grippotyphosa were isolated from the kidneys of 7 (64%) of 11 voles. CONCLUSIONS This is, to our knowledge, the largest leptospirosis epidemic to occur in Germany since the 1960s. Contact between hand lesions and contaminated water or soil and infected voles was the most likely outbreak source. The unusually warm winter of 2006-2007 supported vole population growth and contributed to this resurgence of leptospirosis in Germany. Because of ongoing climate change, heightened awareness of leptospirosis in temperate regions is warranted.

Leptospirosis in Urban Wild Boars, Berlin, Germany

We found antibodies to leptospires in 25 (18%) of 141 wild boars from Berlin (95% confidence interval 12–25). Seropositivity was associated with chronic interstitial nephritis (odds ratio 10.5; p = 0.01), and leptospires were detected in kidney tissues. Wild boars represent a potential source for human leptospirosis in urban environments.

Outbreak of leptospirosis among triathlon participants in Germany, 2006

BackgroundIn August 2006, a case of leptospirosis occurred in an athlete after a triathlon held around Heidelberg and in the Neckar river. In order to study a possible outbreak and to determine risk factors for infection an epidemiological investigation was performed.MethodsParticipants of the triathlon were contacted by e-mail and were asked to fill out a standardized questionnaire. In addition, they were asked to supply a serum sample for laboratory diagnosis of leptospirosis. A confirmed case patient was defined as a clinical case (i.e. fever and at least one additional symptom suggestive for leptospirosis) with at least two of the following tests positive: ELISA IgM, latex agglutination testing, or microscopic agglutination testing. Rainfall and temperature records were obtained.ResultsA total of 142 of 507 triathletes were contacted; among these, five confirmed leptospirosis cases were found. Open wounds were identified as the only significant risk factor for illness (p = 0.02). Heavy rains that preceded the swimming event likely increased leptospiral contamination of the Neckar River.DiscussionThis is the first outbreak of leptospirosis related to a competitive sports event in Germany. Among people with contact to freshwater, the risk of contracting leptospirosis should be considered by health care providers also in temperate countries, particularly in the summer after heavy rains.

Pulmonary Abnormalities in Dogs with Leptospirosis

BACKGROUND Leptospirosis in dogs is a multiorgan disease affecting mostly kidneys and liver. OBJECTIVES The objective was to characterize prevalence, clinical, and radiological features and outcome of dogs with leptospirosis and pulmonary abnormalities. ANIMALS Fifty dogs with leptospirosis. METHODS Medical records of dogs diagnosed with leptospirosis at the Small Animal Clinic, Berlin, were reviewed. Diagnosis was based on microscopic agglutination test, blood or urine polymerase chain reaction, and histopathology. Based on clinical and/or radiological signs, patients were grouped into dogs with lung abnormalities (group 1) or without (group 2). Severity of respiratory distress was scored as mild to moderate (grade 1) or severe (grade 2). Thoracic radiographs were scored based on pulmonary changes and location as grade 1 (caudal interstitial pattern), 2 (generalized mild to moderate reticulonodular interstitial pattern), or 3 (generalized severe reticulonodular interstitial pattern with patchy alveolar consolidations). Results of CBC and biochemistry were compared between groups. RESULTS Thirty-five dogs had radiological pulmonary changes (grade 1: 5; grade 2: 14; grade 3: 16); 31 of them had pulmonary distress (grade 1: 13, grade 2: 18). Sixty-seven percent of the dogs with dyspnea grade 2 were mainly euthanized because of respiratory distress. Fifteen percent of the dogs with dyspnea grade 1 and 21% without clinical respiratory signs were euthanized because of acute renal failure or sepsis. CONCLUSIONS AND CLINICAL IMPORTANCE In 70% of dogs with leptospirosis pulmonary changes were detected. Lung involvement represented a severe complication causing increased case fatality depending on the severity of respiratory distress.

Distribution of Leptospira Serogroups in Dogs from Berlin, Germany

Leptospirosis is a bacterial zoonosis in which dogs can act as a reservoir for human infection. The annual vaccination of dogs can prevent leptospirosis caused by serovars included in the vaccine. To date, all available vaccines in Germany include only the serovars Icterohaemorrhagiae and Canicola, the most commonly found serovars prior to the introduction of the leptospirosis vaccines. Yet, the involvement of additional serovars in the clinical presentation of leptospirosis in dogs has been described. The objective of this sero-epidemiological study was to examine the different Leptospira serovars currently circulating in a population of dogs suspicious for leptospirosis from Berlin. In 329 dogs presenting at the Small Animal Clinic in Berlin, the predominant serogroup was Australis (24%), followed by Grippotyphosa (20%) and Pomona (9%). A total of 18% of the dogs were diagnosed with clinical leptospirosis; here the most prevalent serogroups were also Australis (28%), Grippotyphosa (18%), and Pomona (14%). The serovar prevalence data presented here confirm that a change of pattern of infecting Leptospira serovars in dogs has taken place in Berlin. This data corresponds to further sero-epidemiological studies from other regions in Germany. To ensure human and canine health, available vaccines should be adapted to include the most important circulating serovars.

Comparison of Two PCR Systems for the Rapid Detection of Leptospira spp. from Kidney Tissue

In this study we compared two routine PCR systems for the detection of Leptospira spp. and assessed their performance when directly applied to kidney samples from small mammals. Although the kappa value of 0.9 indicated a high level of agreement between the tests, the outer membrane lipoprotein gene lipl32 based PCR was more robust and showed a higher number of positive kidney samples.

Wild boars as possible source of hemorrhagic leptospirosis in Berlin, Germany

Leptospirosis is a reemerging anthropozoonosis in developing and developed countries [1, 2], and Leptospira spp are endemic to a multitude of domestic and wild animals that shed the infectious pathogen in their urine [1]. Humans usually become infected through contact with urinecontaminated water or soil. Wild boars (Sus scrofa) are well known as animal hosts for Leptospira spp. Leptospires of the Pomona-serogroup, which are predominantly recognized as the infecting serogroup in domestic swine, have frequently been detected in wild boars in European and other countries [3, 4]. The population of this game species has significantly increased throughout Europe over the past decades [5]. As a consequence, migration to urban areas and close contact between wild boars and humans are more frequently observed. In September 2003, a 39-year-old male presented to the emergency unit with a 2-day history of watery diarrhea, fever and abdominal pain. There was no history of traveling abroad, but the patient remembered he had eaten raw eggs about 12 h prior to the onset of diarrhea. Physical examination on hospitalization revealed an acutely ill male with a temperature of 39.8°C, severe tachycardia (120 beats/min), and hypotension (90/60 mmHg). An initial chest radiograph was normal. Infectious gastroenteritis was suspected and the patient received intravenous fluids. On hospital day 3, his condition rapidly deteriorated. In a state of protracted shock, he developed clinical and laboratory signs of pancreatitis, cholecystitis, and hepatitis. Extensive microbiological, parasitological, and serological examinations revealed no causative pathogen and several blood cultures remained sterile. Despite an empiric antibiotic treatment with ciprofloxacin and clarithromycin and infusion therapy, the patient became hemodynamically unstable and had to be transferred to the intensive care unit. The following day, the patient became increasingly breathless and hypoxemic and required intubation. His chest radiograph revealed an adult respiratory distress syndrome pattern, while bronchoscopy showed a sanguineous lavage fluid, suggesting alveolar hemorrhage (Fig. 1). Under intubation, forced diuresis, and an antibiotic regimen including imipenem, ciprofloxacin and clarithromycin, the patient’s condition stabilized. Upon further questioning, his relatives recalled he had cleaned a pond in his garden 2 weeks before the onset of disease. During this work, he waded through the muddy water for several hours and contracted a number of small wounds on his calves. At that time, the pond was frequently used by wild boars as a drinking trough and for wallowing. Contact with other animals was negated. Leptospirosis was suspected, and three consecutive serum samples were submitted to the Eur J Clin Microbiol Infect Dis (2006) 25:544–546 DOI 10.1007/s10096-006-0174-3

Estimation of the sensitivity and specificity of a Leptospira spp. in-house ELISA through Bayesian modelling

The microscopic agglutination test (MAT) is still considered the gold standard for the diagnosis of leptospirosis, although studies have shown that the test is an imperfect gold standard for clinical samples and unsuitable for epidemiological studies. Here, test characteristics of an in-house ELISA were identified for both subclinical and clinical populations by Bayesian latent class models. A conditional dependence model for two diagnostic tests and two populations was adapted to analyse a clinical and a subclinical scenario, respectively. These Bayesian models were used to estimate the sensitivity and specificity of the in-house ELISA and the MAT as well as the prevalences. The Bayesian estimates of the in-house ELISA were: clinical sensitivity=83.0%, clinical specificity=98.5%, subclinical sensitivity=85.7% and subclinical specificity=99.1%. In contrast, the estimates of the MAT were: clinical sensitivity=65.6%, clinical specificity=97.7%, subclinical sensitivity 54.9% and subclinical specificity=97.3%. The results show the suitability of the in-house ELISA for both clinical investigations and epidemiological studies in mildly endemic areas.

Specific CD4+ T-Cell Reactivity and Cytokine Release in Different Clinical Presentations of Leptospirosis

ABSTRACT Clinical manifestations of leptospirosis are highly variable: from asymptomatic to severe and potentially fatal. The outcome of the disease is usually determined in the immunological phase, beginning in the second week of symptoms. The underlying mechanisms, predictive factors, and individual immune responses that contribute to clinical variations are not well understood. The aim of this study was to determine the specifics of CD4+ T-cell reactivity and cytokine release after stimulation with leptospiral antigens in patients with leptospirosis of different disease severities (patients with mild and severe symptoms) and in control subjects (with and without proven exposure to Leptospira). Whole-blood specimens were stimulated with Leptospira antigens in vitro. Subsequently, intracellular staining of cytokines was performed, and flow cytometry was used to assess the expression of CD40 ligand (CD40L) and the production of gamma interferon (IFN-γ), interleukin-10 (IL-10), IL-2, and tumor necrosis factor alpha (TNF-α) by CD4+ T cells. The production of inflammatory cytokines such as TNF-α by CD4+ T cells after stimulation with leptospiral antigens was highest in patients with severe disease. In contrast, the ratio of IL-10 production to TNF-α production was higher in exposed subjects than in patients with mild and severe disease. Levels of proinflammatory cytokines such as TNF-α may be useful markers of the severity of the immunological phase of leptospirosis. IL-10 production by T cells after antigen-specific stimulation may indicate a more successful downregulation of the inflammatory response and may contribute to an asymptomatic course of the disease.

Severe leptospirosis complicated by Epstein-Barr Virus reactivation.

IntroductionWeil’s disease is a severe, potentially fatal illness following Leptospira interrogans infection. The reported case of a patient suffering from acute renal failure, jaundice, thrombocytopenia, rhabdomyolysis and encephalitis syndrome highlights the clinical challenge in reference to Weil syndrome complicated by Epstein–Barr Virus (EBV) reactivation.Materials and methodsThe diagnosis of leptospirosis was performed using four different diagnostic methods. Sera were analyzed with an in-house IgM and IgG enzyme-linked immunosorbent assay (ELISA) and indirect haemagglutination assay (IHA). Microscopic agglutination test (MAT) was done using 17 reference strains comprising 14 serogroups and 17 serovars. Polyvalent EBV-IgG analysis, EBV-IgG/IgM/IgA western blot analysis as well as quantitative EBV polymerase chain reaction (PCR) were performed.ResultsLeptospira IHA showed an initial titer of 1:640 (cut-off 1:320), leptospiral IgG was negative, but IgM was positive. MAT was negative at that time for all 17 strains analyzed. One week later, leptospirosis IHA titer increased to 1:20,480. Leptospiral IgG was now positive, −IgM remained positive and urine was tested negative for leptospiral DNA. The MAT showed positive results for L. interrogans serovar Bataviae, serovar Copenhageni, serovar Pyrogenes and L. borgpetersenii serovar Serjoe. During follow-up examinations, both the leptospiral IgM and IgG remained positive and MAT showed positive results for L. interrogans of different serovars. EBV IgA immunoblot taken at admission was positive for VCA-p18, quantitative EBV-PCR showed an EBV viral load of 2.8E3 copies/ml indicating acute EBV-reactivation.ConclusionLeptospirosis represents a neglected and re-emerging disease which is difficult to diagnose since Leptospira-PCR from whole blood or urine is frequently negative in the case of early empiric antibiotic treatment. EBV-reactivation might represent a severe complication in Weil’s disease which potentially aggravates clinical manifestations of leptospirosis including hepatitis, nephritis, and rhabdomyolysis. Thus, there might be a need for peripheral blood EBV-PCR and EBV blotting in patients suffering from complicated Weil syndrome, also in terms of the choice of antibiotic treatment.