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Comparison of serology and isolates for the identification of infecting leptospiral serogroups in Hawaii, 1979-1998.

Laboratory confirmation of leptospirosis is usually accomplished serologically, without isolates, using the microscopic agglutination test (MAT). However, optimal performance of the MAT is dependent on the knowledge of enzootic serogroups and serovars so that an appropriate MAT antigen testing battery can be established. Infecting leptospiral serogroups can be identified serologically without isolates, using the MAT, or by serogrouping of isolates, but little information is available regarding the correlation between these methods. The identification of infecting serogroups for 53 culture-confirmed leptospirosis cases, diagnosed in Hawaii between 1979 and 1998, using serology and culture isolates were compared. The overall agreement between the two methods was good (kappa = 0.71, 95% CI: 0.56, 0.86). However, the agreement varied between serogroups from 0 to 100%. In establishing the prevalence of serogroups, results obtained via MAT serology (in the absence of serogrouped isolates) should be considered presumptive rather than definitive.

Agglutination Tests↗

Canine leptospirosis in New Jersey and Michigan: 17 cases (1990-1995).

The clinical, laboratory, and serological findings in 17 dogs with disease resulting from leptospiral infection were evaluated retrospectively. Acute renal failure was the most common syndrome, but cholestatic hepatic disease also was common. The most prevalent serovars identified were pomona, grippotyphosa, and autumnalis. Paired serology was available on 10 dogs. Aggressive fluid therapy in combination with ampicillin or amoxicillin resulted in a good survival rate. Canine leptospirosis may be more common than suspected, and paired serology often is necessary to confirm a diagnosis.

Amoxicillin↗

Leptospiral and hepatitis A antibodies amongst windsurfers and waterskiers in Bristol City Docks.

Previous epidemiological studies have detected a substantial risk of gastroenteritis associated with snorkel swimming events in the Bristol City Docks. The event has since been discontinued. Similar risks were not identified at that time for surface-based events. A subsequent fatality from Weil's disease (Leptospirosis) and a case of Hepatitis A associated with surface water immersion in the Bristol City Docks water prompted this study of Leptospiral and Hepatitis A immunity amongst regular windsurfers and waterskiers. Although one person showed evidence of past Leptospiral infection and several people showed evidence of previous Hepatitis A, a causal association with Docks water exposure was not established. Nevertheless, there was sufficient public concern to justify the introduction of precautions against the potential health risks associated with immersion in the City Docks. The study findings and these preventive measures are described.

Adolescent↗

Ultrastructure of renal lesions in pigs with acute leptospirosis caused by Leptospira pomona.

Twelve 4-day old, pathogen-free pigs were inoculated intraperitoneally with virulent L. pomona. Two pigs each were killed 1, 2, 3, 5, 7, and 14 days after inoculation and their kidneys examined by light and electron microscopy. Three to 5 days after inoculation there was hyperemia, edema and swelling of tubular epithelium; leptospires were detected intravascularly, in interstitium and embedded within basement membranes. At day 7, leptospires were at basal surfaces of proximal tubules and in phagosomes in tubular epithelium. Perivascular aggregates of monocytes and macrophages were prominent. At day 14, intersitial cells were chiefly plasmacytes and lymphocytes. Leptospires were only on tubular epithelial cell surface microvilli. In another experiment, all pigs receiving low-virulent strains had leptospires in the renal interstitium at 7 days after inoculation but had none at 14 days. All low-virulent strains produced degenerative changes and lymphocyte-plasmacytic infiltration. These studies indicate that non of the low-virulent isolates were acceptable for use in vaccines. Fialure of leptospires to remain attached to microvilli of proximal tubules in late stages of acute infection may be related to differences in virulence of various isolates.

Anemia↗

Detection of antibodies to leptospiral genus-specific antigen in human and animal sera by indirect hemagglutination test with a partially purified genus-specific protein antigen.

Antibodies against leptospiral genus-specific antigen were detected in the sera from clinically diagnosed human leptospirosis and suspected animal leptospirosis by indirect hemagglutination (IHA) test with a partially purified genus-specific protein antigen (GP-Ag). The reaction was positive in the infected humans and animals irrespective of the leptospiral serovars. No significant correlation was found between IHA titer against GP-Ag and microscopic agglutination (MA) titer. IHA titer did not always develop in parallel with MA titers. Sera obtained from healthy individuals were negative in both IHA and MA tests.

Animals↗

Leptospiral antigens (L. interrogans serogroup ictero-haemorrhagiae) in the kidney of experimentally infected guinea pigs and their relation to the pathogenesis of the renal injury.

The search for leptospiral antigens (L. interrogans serogroup icterohaemorrhagiae) was carried out in 24 guinea pigs experimentally inoculated with 1 ml of culture containing 10(7)-10(8) leprospires and sequentially sacrificed from the first until the 6th day of infection. Semiquantitative analysis of histopathological variables comprising kidney interstitium, tubules and glomeruli was done in 1 micron sections of tissue embedded in glycolmetacrylate. Leptospiral antigen (LAg) and its glycolipoprotein (GLP) expression were detected through PAP in paraffin embedded tissue. The mild interstitial involvement of the kidney, manifested chiefly by oedema and focal interstitial nephritis seen at the 4th day, progressed to tubular damage at the 6th day, characterized by either swelling or cytoplasmic acidophilia of epithelial cells with loss of cell cohesion and sloughing of cells into the tubular lumina. Brush border alterations and mitochondrial changes were observed. Endothelial cell injury was noted in the interstitial vessels. LAg expression was parallel to the kidney changes: small deposits of elongated forms of LAg were detected at the 4th day either within the vascular lumen or free in the interstitium. A rise in the antigen expression was observed at the 5th day when it was seen either around tubules or in their walls. LAg was detected inside the tubular lumina at the 6th day of infection when granular LAg and GLP were abundant. This sequence reproduces the pathway of leptospires in the kidney and the crescent amounts of antigens detected toward the end of the experiment, with antigen concentration in cases of major tissue damage suggesting a direct action of the microorganisms and/or their products in the pathogesis of the lesions.

Animals↗

Direct detection of leptospiral material in human postmortem samples.

Leptospiral culture, direct immunofluorescence, and the polymerase chain reaction (PCR) were used to detect leptospiral material in postmortem specimens collected from eight patients who died of leptospirosis. Diagnosis of leptospiral infection was based on clinical summary (premortem) and confirmed by serological analysis and/or culture of leptospires. Leptospiral culture was the least sensitive technique, yielding two isolates (3%) from 65 samples. Both isolates were from the aqueous humour and cerebrospinal fluid of the same patient. Direct immunofluorescence was of intermediate sensitivity for detection of leptospires, confirming the presence of leptospires in 11% (2 of 18) of tissue samples from three patients. PCR analysis was the most sensitive technique for detection of leptospiral material in tissue samples, being positive in 20% (11 of 56) of samples from eight patients. Both samples (cerebellum and liver) positive by immunofluorescence were also positive by PCR. The sensitivity of the PCR assay was 1-10 leptospires ml(-1) sample, and the assay was specific for Leptospira pathogenic species. Multi-system involvement was indicated based on successful amplification of leptospiral DNA from more than one tissue sample, which corroborated with the clinical and pathologic findings. The results suggest that in acute and/or fatal leptospirosis, the pathogenesis of the pathologic features are related to the presence of the organisms in the tissues. In conclusion, PCR combined with serology appears to be a useful tool for diagnosis of leptospirosis and may be invaluable in epidemiological studies.

Antigens, Bacterial↗

Serologic survey for selected microbial pathogens in bison from Kansas.

A serologic survey was conducted on an American bison (Bison bison) herd in Kansas for antibodies against Brucella spp., Leptospira interrogans serovar canicola, pomona, grippotyphosa, icterohaemorrhagiae, and hardjo, Anaplasma spp., bluetongue virus, infectious bovine rhinotracheitis virus and bovine viral diarrhea virus. There was an increase in prevalence of bluetongue antibodies from 38% in 1987 to 100% in 1989 in animals greater than or equal to 24-mo-old. Prevalences of antibodies against the other livestock pathogens were either negative or at levels associated with previous vaccination.

Anaplasmosis↗

Clinical, bacteriologic and histopathologic studies on induced leptospirosis in stray dog pups.

Eighteen 8-12 days old stray dog (canis familiaris mongrel) pups of either sex; 6 pups each in test groups and control group were infected with lepotspiral serovars autumnalis and canicola. The experimental animals, clinical, bacteriologic and histopathologic kinetics were observed. Both the serovars had evoked typical clinical manifestations. Leptospiraemia could be demonstratedin between the post inoculation (PI) days 1 & 5. Leptospiruria commensed in between the PI days 5 & 7 and lasted throughout the study period. Histopathologic study did not reveal any marked pathologic changes except hydropic changes in the liver of both the test groups.

Animals↗

Diseases of cattle in the Solomon Islands.

Between 1967 and 1977 a study was made of diseases of cattle in the Solomon Islands. Tuberculosis was found in only 3 herds and was eradicated by 1975. Brucellosis serology revealed very few reactors and by 1977 the herds involved were considered free of the disease. Significant serological reactions were found to Leptospira interrogans serovars pomona, hardjo, autumnalis and jez-bratislava. There was evidence that infectious bovine rhinotracheitis and mucosal disease were present. Seventeen parasites were identified of which Haematobia irritans exigua, Haemonchus placei, Oesophagostomum radiatum and Ceylonocotyle streptocoelium were widely distributed. Nutritional stress occurred under some forms of husbandry but environmental stress was minimal. The Solomon Islands are therefore in a most favourable situation with regard to diseases of cattle.

Animals↗

Detection of leptospires in pig kidney using DNA hybridisation.

DNA hybridisation detected leptospiral organisms in homogenised kidneys from experimentally infected pigs, and in homogenates of pig kidneys collected at abattoirs. The technique is easy to perform and had some advantages over cultural and histological methods, in permitting the rapid survey of many kidneys simultaneously. Leptospires added to a homogenate of uninfected kidney could be detected at 10(2) organisms ml-1 by DNA hybridisation, but the technique appeared to be less sensitive than culture.

Abattoirs↗

Patterns of acute renal failure in leptospirosis.

The renal profile was studied in 61 patients having leptospirosis. Leptospira-induced acute renal failure conformed to one of two distinct clinical patterns, either oliguric on non-oliguric renal failure. The prognosis was excellent in the latter variety of renal failure and mortality was confined to patients with oliguric renal failure, where severe glomerular and widespread tubulo-interstitial lesions were invariably present. These patients had unfavourable prognostic features during life, such as prolonged oliguria and anuria, absence of a diruetic phase, persistent elevations in BUN and persistently low urea excretion. The prognostic significance of the age of the patients, hypotension and jaundice in relation to the occurrence and type of acute renal failure have been discussed. Retrospective observations indicate that both a vasculotoxic or haemorrhagic state and oliguric acute renal failure are important causes for mortality in human leptospiral infections.

Acute Kidney Injury↗

[A case of leptospirosis infected in Borneo Island, Malaysia].

We report a case of leptospirosis infected in Sabah, Borneo island, Malaysia. The case is 25-year-old male who had participated in the EcoChallenge Sabah 2000 Expedition Race, a multisport event held during August 20 to September 3, 2000 at various sites in Sabah in Malaysian Borneo. He developed a high fever and headache on September 7, and he was admitted to our hospital on September 9. On admission he also had conjunctivitis and myalgias. Laboratory findings on admission revealed leukocytosis with left shift, slightly elevated transaminase levels, high CRP levels and proteinuria. Plasmodium spp. were negative on blood smears, and no bacteria were isolated from blood and feces cultures. We performed the laboratory tests for leptospirosis, based on the information about the probable leptospirosis outbreak among athletes who participated in the EcoChallenge Race, however both Leptospira antigens and antibodies were negative at that time. We diagnosed leptospirosis clinically because he manifested persistent symptoms, and minocycline 100 mg b.i.d. was administered intravenously resulting in excellent efficacy. Serum antibody tests by microscopic agglutination test (MAT) at convalescent stage revealed significant increased antibodies against Leptospira interrogans serovar hebdomadis, and the diagnosis of leptospirosis was confirmed. Infectious diseases have been global and it is important to have information concerning worldwide infectious disease situations as much as possible for accurate diagnosis.

Adult↗

Detection of specific anti-leptospiral immunoglobulins M and G in human serum by solid-phase enzyme-linked immunosorbent assay.

The enzyme-linked immunosorbent assay (ELISA) was used to detect leptospire-specific immunoglobulin M (IgM) and IgG in the sera of patients infected with leptospiral serovars hardjo, pomona, or copenhageni. All patients produced specific IgM and IgG detectable by ELISA. In contrast, only a few patients produced IgG agglutinins whereas all produced IgM agglutinins. The specificity and sensitivity of the test suggest that the ELISA anti-IgM technique is a suitable method for detecting leptospiral antibodies in human sera for diagnostic and epidemiological purposes.

Agglutination Tests↗

[A comparative evaluation of the sensitivity of different methods for the serological diagnosis of leptospirosis].

In this work the comparative evaluation of the sensitivity and serological specificity of the microcapsular agglutination (MCA) test, the passive hemagglutination (PHA) test and the microagglutination (MA) test are presented. In the MCA test leptospiral antigens, adsorbed on synthetic carrier capsules produced by Japan Lyophilization Laboratory, were used and the PHA test was made with the use of polyvalent erythrocyte diagnosticum. The study of blood serum samples from 46 leptospirosis patients revealed that the values of antibody titers in the PHA and MCA tests were 5.5-8.1 times higher than the traditional MA test. In the MCA and PHA tests antileptospiral antibodies could be detected as early as on days 1-3 of the disease when the results of the MA test were negative or very low. The maximum values of antibody titers in the MCA and PHA tests were detected on days 11-15 of the disease and in the MA test, on days 21-25. The MCA and PHA tests are genus-specific and permit the detection of antileptospiral antibodies irrespective of the serogroup of the infective agent. In the study of the blood sera of 40 patients with diseases of nonleptospiral etiology the MCA and MA tests yielded false positive results in 7.5% and the PHA test, in 12.5% of cases in titers below the diagnostic level. These data are indicative of high sensitivity and specificity of the serological tests used in this study.

Agglutination Tests↗

Immunofluorescent staining of leptospires in pepsin treated histologic sections.

A standard immunofluorescent method was modified for the staining of leptospires in formalin fixed, paraffin embedded tissues. Routine histologic sections were deparaffinized and treated with pepsin prior to staining. Pepsin treatment greatly enhanced subsequent staining of leptospires in naturally infected bovine and porcine tissues as well as in artificially infected tissues. Leptospires in naturally infected bovine tissues were usually undetectable in untreated sections but clearly visible in stained pepsin-treated sections. Naturally infected porcine kidney usually contained high levels of leptospiral antigen which could be stained without prior pepsin treatment. However, pepsin treatment of porcine tissues greatly increased the amount of leptospiral antigen detectable and made individual leptospires more conspicuous. The staining method could employ a single antiserum for the staining of leptospires from 13 serogroups. Also, leptospires could be stained in tissues stored in formalin for more than 14 months and in 26-year-old paraffin embedded tissues.

Animals↗

Some observations on the diagnosis and epidemiology of leptospirosis in swine.

The results of combined epidemiological, clinical, serological, bacteriological and histopathological studies following an outbreak of disease caused by L. pomona on a farm stocked with cattle, sheep, pigs, goats and horses maintained for experimental purposes, are reported.The incidence of infection was high in horses, cattle and pigs. A few low titres were seen in sheep. The goats were not infected. Apart from a single bovine abortion all the clinical symptoms observed occurred in pregnant sows. Seven of these aborted or gave birth to stillborn pigs within a six week period. Fifteen species of wildlife were trapped or shot on the farm during the year following the outbreak. L. pomona was isolated from four skunks and a porcupine. Epidemiological studies indicated that wildlife reservoir hosts were the primary source of infection for the domestic livestock. Leptospiruria and the serological response were studied in a group of eight infected sows. Microscopic agglutination titres of 10(2) or less could not be associated with leptospiruria and the duration of leptospiruria was found to range from a few weeks to over two years in individual sows. Direct dark-field examination of urine proved superior to guinea-pig inoculation as a method of detecting leptospiruria and it is suggested that the former technique could be adopted with advantage as a routine aid to diagnosis.

Agglutination Tests↗

Enzyme-linked immunosorbent assay for leptospirosis immunoglobulin M specific antibody using surface antigen from a pathogenic Leptospira: a comparison with indirect hemagglutination and microagglutination tests.

A search for a sensitive and specific test for human leptospirosis was made by enzyme-linked immunosorbent assay for immunoglobulin M specific antibody (IgM ELISA) using a surface antigen from L.interrogans serovar bataviae, L. interrogans serovar pyrogenes and L.interogans serovar icterohaemorrhagiae. The IgM ELISA tests using each of the three antigens were evaluated in 103 sera primarily positive by microagglutination test (MA). Optical density of these IgM ELISA tests showed good correlation. The IgM ELISA using antigen from serovar bataviae was compared with MA and indirect hemagglutination (IHA) in 20 sera primarily positive by IHA, and 103 sera primarily positive by MA. IgM ELISA and IHA using antigen prepared from serovar bataviae in 103 sera positive for MA had a sensitivity of 98.06 and 92.23 per cent respectively. In 20 sera primarily positive by IHA, IgM ELISA and MA showed sensitivity of 80 and 45 per cent respectively. The surface antigen used in IgM ELISA is broadly specific making IgM ELISA a sensitive and specific test for human leptospirosis. IHA agreed more with IgM ELISA in comparison to MA. As MA is not sensitive for early infection, IHA and IgM ELISA should be in routine use in general laboratories.

Agglutination Tests↗