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A DNA array for rapid detection and genotyping of the pathogenic microbes of epidemic hemorrhagic fever, tsutsugamushi disease, leptospirosis, malaria, schistosomiasis, cholera, and hemorrhagic colitis was developed. The specific and relatively conserved PCR primers and DNA probes were screened from the characteristic genes of the pathogenic microbes. The PCR or RT-PCR methods were established for amplifying and labeling the DNA fragments of the pathogenic microbes. All the probes with the same Tm value were synthesized chemically and modified with an NH2 at their 5' terminus, they were printed on glass slides for fabrication of a oligonucleotide DNA array. The developed DNA array could be used for detecting and genotyping the pathogenic microbes simultaneously, and they had a high sensitivity and specificity.

Blood from patients suspected of leptospirosis 148 specimens were cultured for leptospira. Twenty two specimens were positive (15%). The isolated leptospira were tested against the 24 serovars of standard antisera by Microscopic Agglutination Test (MAT). It was found that all 22 leptospira isolates reacted strongly against L. autumnalis, except 1 isolate that also reacted against serovar djasiman. The patient's sera were collected from only 14 cases. When the sera of the 14 patients were tested with the 24 reference serovars by MAT it was found that sera reacted the most against L. australis and in decreasing order against L. bratislava, L. autumnalis, L. rachmati, L. copenhageni, L. javanica. There had some cross reactions against several serovars in a single patient. The present study showed inconsistency between culture results and serum assays. Since sera showed cross reactivities against several serovars, it was not possible to determine which serovar was etiologic. Therefore, the isolation of leptospira though time consuming is specific in the identification of the serovar.

A case of fulminant leptospirosis is presented, manifesting as rapid progression from acute undifferentiated febrile illness to refractory shock, jaundice, renal failure and massive pulmonary hemorrhage. The patient received aggressive intensive care unit support including prolonged intubation and ventilation. This case emphasizes that acute leptospirosis may well not be characterized by the classic scenario of a biphasic illness, but rather by a fulminant, monophasic illness.

Canine leptospirosis has been known as Stuttgart disease since 1898, and dogs are considered to be the second principal source of infection in man. The isolation of a pathogenic serovar from dog urine that was diagnosed clinically and laboratorial as having leptospirosis and its utilization to test serological samples of human and canine cases of leptospirosis, has demonstrated its importance to the ecosystem of the southern region of Brazil. The results of the serological microscopic agglutination test indicated that 100% of human serum samples from 12 patients from the serum bank of 2001 at the Center for Control of Zoonoses, that had titers between 25 and 3,200 with the canicola serovar, and 72% of 105 canine serum samples from the same serum bank, also reacted with the new isolate. The mean and median titers of the human serum samples tested with the battery of antigens recommended by WHO was 630 and 100 respectively, and when tested with the isolate it was 1,823 and 400. In the dog sera, the values were respectively 347 and 100 with the battery, and 1,088 and 200 with the isolate.

This study describes the isolation of a new leptospiral serovar from the Djasiman group from an Argentinean aborted fetus of a dog. The strain was isolated from a culture of mixed liver and kidney tissue from one aborted dog fetus. Bitch's serum showed a titre of 1:800 against the new serovar and 1:400 or less against other serovars tested. Microscopic Agglutination test (MAT) with a panel of 38 rabbit anti-Leptospira sera representative for all pathogenic serogroups was performed to establish the putative serogroup of the isolated strain. Serovar identification was done by Cross-Agglutination Absorption Test (CAAT). Species determination was carried out on basis of sequence identity of primer pair G1/G2 generated PCR products from the isolate with those from reference strains belonging to the various pathogenic Leptospira species. Results showed that the new strain named, Baires, belongs to species L. interrogans sensu stricto and the serogroup Djasiman. However, the isolate could not be identified as any of the known serovars within this serogroup. These results allow us to conclude that the strain Baires represents a new serovar (Buenos Aires) of Djasiman serogroup.

Seroprevalence of five different Leptospira interrogans serovars, Toxoplasma gondii and Leishmania infantum in stray dogs in Ankara was investigated. A total of 116 dog sera collected from apparently healthy stray dogs were tested for L. interrogans serovars by microscopic agglutination test (MAT), for T. gondii antibodies by Sabin-Feldman dye test (SFDT), and for L. infantum antibodies by indirect fluorescence antibody test (IFAT). Of the 116 dogs, 51 (43.96%) were seropositive for leptospirosis, 72 (62.06%) for T. gondii and 3 (2.58%) for L. infantum. No statistically significant difference was observed between male and female dogs in the seroprevalences of toxoplasmosis and leptospirosis (P>0.05), but statistically significant difference was observed among different age groups in the seroprevalences of toxoplasmosis and leptospirosis (P<0.05). Although the seroprevalence of L. infantum was low, asymptomatic animals should be considered as a reservoir for the spread of the disease.

Leptospirosis is globally important infectious disease affecting almost all mammals. Pathogenic Leptospira encodes immunoglobulin-like protein (Lig) that is found to express only during infection. We report the development of conventional and real time PCR assays targeting lig genes of leptospires for the early diagnosis of leptospirosis. Sensitivity of the newly designed Lig1/Lig2 primers for conventional PCR was compared with previously published primers LP1/LP2 and G1/G2. G1/G2 primers amplified the target DNA from all the serovars including non-pathogenic Leptospira biflexa whereas LP1/LP2 and Lig1/Lig2 primers amplified only pathogenic leptospires. Diagnostic PCR assay was also developed for the detection of pathogenic Leptospira interrogans in urine samples. We obtained the highest sensitivity in PCR using our Lig1/Lig2 primers with a detection of 6 leptospires. A rapid and sensitive lig-based real time PCR assay was also developed with a detection range of 10-10(7) gene copies. To evaluate the early diagnosis for leptospirosis, we compared the culture with conventional and real time PCR for the detection of spirochetes in experimentally infected hamsters during a time-course study. Culture of infected hamster tissues detected the presence of leptospires from Day 2 of infection but not on the day of infection or Day 1, whereas conventional PCR and real time PCR detected the leptospires from the day of infection. Hence, conventional and real time PCR with lig primers would be a sensitive and rapid tool for early diagnosis of leptospirosis.

A Leptospira strain was isolated from a hemoculture of a patient admitted with suspicion of leptospirosis that had been previously vaccinated. It corresponded to the Bataviae pathogenic complex. This result shows that there exists immunogenic specificity and that other serogroups should be incorporated to the current vaccine.

The effect of different alkalizing agents of urine on the isolation of leptospiras was studied. Better results were obtained on using the basic solution of NaOH 1N inoculated in urine one hour after being alkalized.

26 patients with active posterior uveitis were studied by methodologies for the confirmation of leptospirosis. ELISA-IgG was positive in 10 patients and MAT in 21. Leptospiras were observed in blood and urine. The results suggest that posterior uveitis may be caused by pathogenic leptospiras.

Serum samples were collected randomly from Mongol breed cattle in three geographically distinct provinces of Mongolia. Antibodies specific to Leptospira interrogans serovar Hardjo were detected by microscopic agglutination test (MAT) at titres of 100 or more in 80.4% (86/107) of the samples from Dornod Province, but in only 28.9% (13/45) in Arkhangai and 23.5% (12/51) in Khuvsgul Provinces, respectively. Treatment of 9 serum samples from Dornod, positive to serovar Hardjo in MAT and negative in the homologous immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA), with 2-mercaptoethanol (2-ME) indicated that those animals were recently infected.

Aims: Andaman and Nicobar Islands in India has a century long history of human leptospirosis. Several isolates have been recovered over the years from different locations. The present study was undertaken to understand the clonal relationship between all these pathogenic leptospires recovered from these islands., Methods and Results: Arbitrarily primed polymerase chain reaction (AP-PCR) was employed to genetically characterize 40 isolates recovered during 1995--2001 and their fingerprints were compared with those of 26 reference strains of known genetic and serological affinities. Sequences of PCR-amplified products from representative isolates were compared with those of different strains belonging to seven genospecies. AP-PCR fingerprints revealed that 32 of the 40 isolates were clonal in nature and fingerprints of all the isolates matched with known reference strains of pathogenic Leptospira interrogans sensu stricto. Comparison of sequence data of PCR amplified products of reference strains and isolates also corroborated these findings., Conclusions: The study revealed that 80% of the isolates recovered from these islands were clonal in nature and all the isolates taken in the study belonged to Leptospira interrogans sensu stricto., Significance and Impact of the Study: An extension of the study in animal population would help in understanding the transmission dynamics of this commonly circulating clone in these islands, which in turn might help in effective control of this public health problem.

Objectives: To evaluate the discriminatory power and usefulness of arbitrarily primed-polymerase chain reaction (AP-PCR) characterization of leptospires with M16 primer., Methods: AP-PCR fingerprints of 20 reference strains of Leptospira representing 20 different serovars belonging to seven genospecies (Leptospira interrogans, 11; L. noguchii, 2; L. borgpetersenii, 1; L. santarosai, 2; L. biflexa, 2; L. kirschneri, 1; L. weilii, 1) were generated by employing M16 primer. Fingerprints generated with this primer were compared with those generated with two other commonly used primers PB1, and L10. An attempt was also made to type 20 leptospiral isolates with the M16 primer., Results and Conclusion: Fingerprints with M16 primer could not only differentiate between strains of different genospecies, but also between strains of the same genospecies belonging to different serovars. While two commonly used primers (PB1 and L10) failed to discriminate between some of the different serovars belonging to the same genospecies, this primer was able to generate discriminatory fingerprints for all strains tested. All 20 Leptospira isolates, recovered from patients in Andaman Islands, could also be typed by fingerprints generated with the M16 primer. The discriminatory power of M16 primer adds more specificity to the rapidity of this system of characterization and can be used as an excellent tool in epidemiological studies on Leptospira.

Risk factors for leptospirosis in France were investigated to improve the vaccination program for this disease. Data from 90 hospitalized case patients and 169 matched control subjects were analyzed in a case-control study. Skin lesions, canoeing, contact with wild rodents, and country residence were independently associated with leptospirosis, emphasizing that leisure activity is a risk factor for this illness.

A bacteriological survey of kidneys from 145 abattoir horses was performed, which resulted in the isolation of two Leptospira strains. The isolates were serologically typed as belonging to serogroups Australis and Pomona, and REA identified them as L. interrogans serovar Bratislava and L. kirschneri serovar Tsaratsovo, respectively. These are the first Leptospira isolates obtained from horses in Portugal and the Bratislava strain is the first serogroup Australis strain to be isolated in this country. The 145 horses were also serologically tested for leptospiral antibodies, and 37% had MAT titres #10878;1:10.

Leptospirosis is considered an important reemerging infectious disease worldwide. The standard and most widespread method for the diagnosis of leptospirosis is the microscopic agglutination test (MAT). This test is laborious and time-consuming, and the interpretation of the results is subjective. In the present work we describe an application of flow cytometry (FCM) as a tool for the serological diagnosis of leptospirosis. The analysis is based on the sensitivity of FCM to the size and shape of the bacteria analyzed by measurement of light scatter parameters: forward scatter (FSC) and side scatter (SSC). The addition of positive serum to an infecting leptospiral serovar results in a shift of the light scatter parameter to a different location with higher FSC and SSC values, indicating the formation of leptospiral aggregates. By using immunofluorescent staining, we have shown that the large particles formed are the agglutinated leptospires. Quantification of the agglutination process has been achieved by calculating an agglutination factor (Af), based on changes in the light scatter parameters measured by FCM. Af enables us to determine the specificity of the serological reaction of the patient serum with each leptospiral serovar. In this work, 27 serum samples from 18 leptospirosis patients were tested by both the MAT and the FCM techniques, in which each serum sample was tested against 13 serovars. Twenty-six human serum samples derived from patients with a variety of other defined illnesses were used as negative controls and enabled us to define the Af threshold value as < 9.3 for negative patients, while any value higher than that would be a positive result for leptospirosis. Compared to MAT, the FCM technique was found to be more specific and sensitive, especially in identifying the serogroup in the acute phase of the disease. The whole process was found to be rapid and took less than 1.5 h. Moreover, FCM analysis is objective and can be automated for the handling of large numbers of samples.

A 2-year-old Thoroughbred filly presented with ocular pain and epiphora of the left eye. The pupil was miotic and the cornea edematous near the ventro-temporal limbus, but did not retain any fluorescein. The topical antibiotics and atropine and diclofenac, and systemic flunixin meglumine and antibiotic therapy did not resolve the condition. A pink and fleshy infiltrate developed near the limbus indicating nonulcerative keratouveitis. The anterior uveitis deteriorated as manifested by the presence of dyscoria, hypopyon, and organized fibrin in the anterior chamber. Ocular signs were improved by topical and subconjunctival corticosteroids, but repeatedly deteriorated as the frequency of medication was reduced. The horse was seropositive to three serovars of Leptospira interrogans. The animal was diagnosed as blind on day 91 by the absence of pupillary light and menace reflexes, and donated for histopathologic diagnosis. The corneal opacity was histologically fibrotic and infiltrated predominantly by lymphocytes with Descemet's membrane partially disrupted by macrophages. The choroid was infiltrated by lymphocytes, eosinophils and basophils, and was positive to IgG and C3. There were filamentous or spiral structures positive to Warthin-Starry stain in the renal cortex. There was also polymerase chain reaction amplification of the leptospiral gene in the kidney. From these findings nonulcerative keratouveitis was believed to be caused by systemic infection with Leptospira.

Two candidate vaccines of Ballum serogroup Leptospira interrogans were microbiologically characterized as part of the work directed to the obtention of new antileptospirosis vaccine formulations for human use. The growth kynetics of both strains was evaluated in EMJH protein medium and in 3 protein free media. The virulence was estimated in hamsters by the calculation of the mean lethal dose. The cellular and extracellular antigenic profiles were analyzed by unidimensional SDS-PAGE and compared with those from strains of Canicola, Icterohaemorrhagiae and Pomona serogroups. The antigenic homology among heterologous groups was analyzed by western blotting with serun from hasterms vaccinated with vax-SPIRAL. The results obtained showed a fastidious growth of both strains of Ballum in the studied media, a high virulence in the animal model and a large antigenic homology with strains from other serogroups of Lepstospira prevailing in Cuba.

Twenty-four specific pathogen-free beagles were randomly allocated into four groups (three vaccinated groups and one control group) and inoculated at nine and 12 weeks of age with one of three commercial inactivated Leptospira vaccines: A (Vanguard 7; Pfizer Santé Animale), B (Dohyvac 7L; Fort Dodge), and C (Nobivac DHPPi + Lepto; Intervet International); the control group received Nobivac DHPPi (Intervet International). Seven weeks after the second vaccination all the dogs were challenged with Leptospira interrogans serogroup canicola. All the vaccinated dogs developed a mild serological response (microscopic agglutination titres) after the booster vaccination. A significant serological response after the challenge was observed, particularly in the controls. The challenge induced fever and clinical disorders in the control group, whereas in the vaccinated groups the clinical signs were mild. Blood cultures became positive in all control dogs, and in one of six dogs vaccinated with vaccine A and two of four dogs vaccinated with vaccine B; none of the six dogs vaccinated with vaccine C was leptospiraemic at any stage of the experiment. Urine cultures were positive in all the control dogs two weeks after the challenge. One of six dogs vaccinated with vaccine A and two of four dogs vaccinated with vaccine B shed bacteria in their urine after the challenge, but none of the dogs vaccinated with vaccine C shed bacteria in their urine at any time during the experiment.

Sera obtained from cattle in the state of Yucatan, Mexico, were screened using the microscopic agglutination test against 13 serovars of Leptospira interrogans. A total of 62.8% (461/734) cows were positive for one or more serovars. This seroprevalence probably reflects infection because vaccination against leptospirosis has not been practised in Yucatan. The most common antibodies detected were those against antigens of serovars hardjo (54.1%) and tarassovi (53.3%). Region was the only risk factor associated with the seroprevalence of leptospirosis (p < 0.05).

One kidney was collected from each of 32 fattening pigs at an abattoir in southern Vietnam in 2001 in order to demonstrate infecting Leptospira serovar and to associate renal macro- and microscopic findings with the presence of renal leptospires. Leptospires were demonstrated in 22 (69%) of the investigated kidneys by immunofluorescence. Multifocal interstitial nephritis (MFIN) and gross renal lesions (white spots) were each demonstrated in 24 (75%) kidneys. Leptospira interrogans serovar bratislava was isolated from one kidney. There was no association between presence of leptospires and MFIN (P=0.19), respectively and white spots (P=0.98), respectively. These data suggest that Leptospira infection is common among fattening pigs in the study area and that these animals may be considered as an occupational human health hazard. It is also suggested that the presence of white spots is an unreliable indicator of the presence of renal leptospires.

Forty-two urine and 96 blood and serum samples were obtained from California sea lion (Zalophus californianus) pups from the Gulf of California during the 2000 reproductive season. Antibody prevalence to 13 serovars of Leptospira interrogans was determined by microagglutination tests (MAT); presence of pathogenic leptospires was detected by polymerase chain reaction (PCR). Samples with antibody titers > or = 1:25 or 115 bp fragments on ethidium bromidestained 1.5% agarose gels were considered positive. Antibody prevalence was 54% overall with highest prevalence against serovar cynopteri (50% of all positive reactions). Highest antibody titers (1:50) were detected against serovars cynopteri and pomona. Polymerase chain reaction products were observed in two of 42 urine samples, six of 96 blood samples, and one of 96 serum samples. Presence of PCR products in blood and serum was demonstrated in pups that were seronegative. Kruskall-Wallis tests and corresponding post hoc Tukey tests (alpha = 0.05) showed that prevalence of leptospirosis was significantly different among all rookeries. The high seroprevalence (54%), low antibody titers (maximum 1:50), absence of pups showing clinical signs indicative of the disease, and lack of recent reports of increased mortality of sea lions in the Gulf of California are suggestive of the presence of enzootic host-adapted serovars. Crowding in rookeries as well as the presence of bats and rodents on some of the islands may explain infection by L. interrogans (sensu lato) and some of the differences in seroprevalence among reproductive rookeries.

Serum samples from 77 animals belonging to 38 species and 19 families in Rio de Janeiro Zoo, Brazil were tested for antibodies against serovars of Leptospira interrogans by microscopic agglutination test. Antibodies were detected in serum samples of 37.7% of all animals belonging to 10 families. Seropositivity was more common in the Carnivora Canidae (7/9), Procyonidae (5/9) and the Edentata Myrmecophagidae (5/9). Icterohaemorrhagiae was the most common serogroup. Seropositivity was less common in primates.

A 2-year cohort study was conducted to investigate the probability of disease introduction into Dutch dairy farms. The farms were tested regularly for diseases and were visited biannually to collect management data. Ninety-five specific pathogen-free (SPF) dairy farms were selected from a database of bovine herpesvirus type 1 (BHV1)-free farms to study the probability of, and risk factors for, introduction of BHV1, bovine viral diarrhoea virus (BVDV), Salmonella enterica subsp. enterica serotype Dublin (S. dublin), and Leptospira interrogans serovar hardjo (L. hardjo). Although most of the 95 SPF farms had a low risk on introduction of infectious diseases, one disease was introduced into 12 farms and two diseases were introduced into one farm. Three farms experienced an outbreak of BHV1, one farm an outbreak of L. hardjo, two farms BVDV, six farms S. dublin, and one farm both BHV1 and S. dublin. The total incidence rate was 0.09 (0.06-0.12) per herd-year at risk. The results suggest that the "non-outbreak" farms were significantly more closed than the "outbreak" farms. Direct animal contacts with other cattle should be avoided and professional visitors should be instructed to wear protective clothing before handling cattle.

The presence of Salmonella enterica and serologic evidence of infection by Leptospira interrogans, were detected in the opossum Didelphis virginiana in a semi-urban locality of the Yucatán State, México. Ninety-one opossums were captured during the period April 1996 and May 1998. From a total of 17 feces samples, four Salmonella enterica subsp. enterica serotypes (Sandiego, Newport, Anatum, and Minnesota), and one Salmonella enterica subsp. arizonae serovar O44:Z4,Z23:- were isolated. Some opossums presented mixed infections. From 81 sera samples, four (4.9%) were positive to antibodies to Leptospira serovars pomona and wolfii. Both animals infected with Salmonella enterica and those serologically positive to Leptospira interrogans were captured in peridomestic habitat. Opossums infected with Salmonella enterica, were captured in dry season, and those seropositive to Leptospira interrogans during the rainy season. The implications of infection and reactivity of these zoonotic pathogens in D. virginiana in the Yucatan state are briefly discussed.

Bovine brucellosis and leptospirosis are important causes of bovine abortion around the world. Both diseases can be serologically diagnosed, but many factors may cause false positive and negative results. Direct methods based on bacteriological isolation are usually employed, but they are difficult, time consuming and dangerous. Monoplex polymerase chain reaction (PCR) have been successfully described for the detection of Brucella spp. and Leptospira spp. Aiming at improvement in the direct diagnosis, a multiplex PCR (mPCR) for the detection of these agents in aborted bovine fetuses is described. The detection threshold of the mPCR was evaluated in experimentally contaminated bovine clinical samples using a conventional proteinase K/SDS or a boiling-based extraction protocols. The mPCR was applied to two groups of clinical samples: 63 episodes of bovine abortion and eight hamsters experimentally infected with Leptospira interrogans serovar pomona. Adopting microbiological isolation as reference, the test showed a sensitivity of 100% in both groups of clinical samples. Seven samples collected from bovine fetuses were Brucella spp. culture negative but showed positive results in mPCR. Regarding Leptospira spp. detection, similar results were observed in three bovine clinical samples. All hamsters infected with Leptospira were positive in both microbiological culture and mPCR. The boiling extraction protocol showed better results in some clinical samples, probably by the removal of PCR inhibitors by heat treatment. The high sensitivity, simplicity and the possibility of detection of both bacteria in a single tube reaction support the use of the mPCR described in the routine diagnosis.

History and Clinical Findings: A 65 year-old man was transferred to our department from a neighbouring hospital with anuria and epistaxis. A few days prior to hospitalization, he had experienced severe muscular and joint pain accompanied by chills. A careful history revealed that, in recent weeks, the patient had frequently collected wild walnuts growing, for the most part, on the banks of a small stream, known to have an infestation of rats. The physical examination revealed pronounced jaundice of the skin and sclerae, and petechia on the lower legs., Investigations: Laboratory results showed marked thrombocytopenia, hyperbilirubinaemia, appreciably elevated urine retention parameters and increased C-reactive protein. During the subsequent course of his illness, serum leptospiral antibody titres were elevated, indicating an acute leptospiral infection manifesting as Weil's syndrome. Silver staining (>>Warthin-Starry<<) revealed rod-shaped bacteria, presumably representing leptospires, in some bone marrow macrophages., Treatment and Course: Treatment with i. v. penicillin was immediately initiated, and urine output established by intravenous fluid resuscitation in the intensive care unit, so that haemodialysis was not necessary. The platelet count returned to normal and bilirubin began to decrease again. The patient was discharged home after 2 weeks in the hospital., Conclusion: When a patient presents with the triad of renal failure, jaundice and thrombocytpenia in the setting of a possible infection, then the severe form of leptospirosis known as Weil inverted question marks syndrome must be considered, and antibiotic treatment initiated without delay. Of importance for the definitive diagnosis is the repeated determination of the titres of antibodies to leptospires in the serum and urine, which usually become positive only in the second week of the illness. In our case, we detected bacteria directly in some bone marrow macrophages as well.

Objective: To measure antibody titers against Leptospira interrogans in serum and vitreous humor and determine the prevalence of L interrogans in vitreous humor of horses with recurrent uveitis., Design: Cross-sectional study., Animals: 242 horses (270 eyes) with recurrent uveitis undergoing vitrectomy and 39 control horses (54 eyes) without any history or clinical signs of recurrent uveitis undergoing euthanasia or enucleation for unrelated reasons., Procedure: Serum and vitreous humor were tested for antibodies against 13 serovars of L interrogans. Vitreous humor was submitted for leptospiral culture; isolates were typed to the serogroup level., Results: Leptospira interrogans was isolated from vitreous humor from 120/229 (52%) horses (126/252 [50%] eyes) with recurrent uveitis but was not isolated from vitreous humor from 36 eyes of 21 control horses. Duration of recurrent uveitis was > or = 1 year for 45 of the 120 (38%) horses from which the organism was isolated. Geometric mean antibody titers against L interrogans in the vitreous humor and serum of horses with recurrent uveitis were 1:1,332 and 1:186, respectively. Only 91 of 120 (76%) horses from which the organism was isolated had a 4-fold or greater difference between serum and vitreous humor antibody titers., Conclusions and Clinical Relevance: Results suggest that persistent ocular infection with L interrogans is common in horses with recurrent uveitis. A 4-fold increase in vitreous humor versus serum antibody titers may not be a sensitive test for the diagnosis of L interrogans-induced recurrent uveitis. We hypothesize that the immune component of recurrent uveitis can be directly induced and maintained by persistent infection of the eye with L interrogans.

This study was designed to investigate the role of leptospirae in interstitial nephritis. Sixty-eight white-spotted kidneys and 30 grossly normal kidneys from slaughtered cattle were examined histologically and immunohistochemically for the presence of Leptospira interrogans antigens. The presence of L. interrogans antigens was found in 21 of 68 white-spotted kidneys and in four of 30 normal kidneys. In conclusion, the detected incidence of infection with L. interrogans was not high, but there was a relationship between the presence of interstitial nephritis and leptospiral antigens.

Between March 1996 and September 1997, 15 farms located in Galicia (NW Spain) and experiencing suboptimal reproductive efficiency were visited and blood samples were obtained from all the lactating cows (n=442). Additionally, 1060 samples were obtained monthly from a cohort of 219 lactating cows belonging to nine of the farms between March 1996 and April 1997. All the samples were tested by microscopic agglutination (MAT) using live antigens representing the following Leptospira interrogans serogroups: Australis, Autumnalis, Ballum, Canicola, Grippotyphosa, Icterohaemorrhagiae, Louisiana, Pomona, Sejroe, Shermani and Tarassovi. Eighty-one of the 442 cows were positive against one or more serogroups (P=18.33%). Serologically, L. bratislava and L. grippotyphosa were detected as the most prevalent serovars (P=7.92 and 7.69%, respectively) and as the serovars against which the probability of seroconversion was highest (P=0.27 and 0.25, for a 12-month period, respectively) among those studied. The proportional hazards regression method was used for evaluating the seasonal trend of seroconversions against these two serovars. The risk of seroconversion against L. grippotyphosa was significantly higher during spring. The risk of seroconversion against L. bratislava did not differ significantly among seasons. Our results suggest that infections by L. bratislava did not follow, among the study animals and during the study period, the pattern typically described for non-adapted serovars, pointing out the possibility that some strains of this serovar could behave as adapted serovars.

Leptospirosis is a zoonosis caused by Leptospira interrogans. This disease is diagnosed by quantification of specific immunoglobulins in serum by the microagglutination test (MAT). The aims of this research were: a) to compare the protein profiles of 3 clinical isolates of bovine leptospirosis with the reference strain used for the MAT, and b) to identify the immunodomain antigens of the regional isolates through PAGE and immunoblotting techniques of bovine sera from infected, vaccinated and MAT-negative animals. Coomassie-blue stained gels revealed extensive protein similarities between pathogenic and reference strain. Most infected (8/10) and vaccinated animal sera (4/7) showed by immunoblotting a similar reactivity against the proteins from pathogenic leptospires, with a strong band of 25-30 kDa which was not detected in the reference strain. The lack of correlation between MAT and immunoblotting techniques for infected animals could be due either to the infection stage at which the diagnosis was made or to the immunoglobulin isotype involved in the response. Results obtained would confirm the antigenic differences between the 3 isolates and the reference strain.

On January 20, 1915, Inada and Ido announced the discovery of the causative agent of Weil's disease. Subsequently, on February 13, 1915, they published the first paper on the discovery of the causative organism (a new species of Spirochaeta) of Weil's disease. Besides discovering the causative organism of the disease, Inada and colleagues clarified the pure culture in medium, and determined the source and route of the infection, its pathology and morbid anatomy; the distribution of the organism in various organs and tissues; the excretion of the spirochete, and its division, filterability, and morphological characteristics; and the clinical picture, laboratory findings, diagnosis, prophylaxis, and treatment of the disease. These studies were conducted by Inada, Ido, Kaneko, Hoki, and Ito, in the years 1914 to 1915. In the early investigation of leptospirosis, Inada and colleagues played a prominent part. We would like to remember these remarkably complete and definitive original achievements on leptospirosis made by Inada and colleagues.

This material intends to present three clinical observations regarding the uveitis etiology, that is represented by the association of some etiological factors less known: Toxoplasma gondii, Listeria monocytogenes, Leptospira icterohaemorrhagiae. The uveitis described here have been severe in the fast evolution and with trends of reappearence. This work demonstrates that the strict etiological antibacteria and antiparasite therapy is not enough and only corticotherapy is able to fight against the accentuated inflammatory phenomena that go along with type of uveitis.

Leptospirosis, a common form of zoonosis, especially in rainy countries, is caused by Leptospira interrogans. In our region of Turkey this type of disease has often been encountered in connection with rice harvesting and we therefore attempted to evaluate the prevalence of L. interrogans in wild rats in our region. Fifty-nine Rattus norvegicus rats were trapped alive in different areas of an approximately 100 km stretch of seashore in the Middle Black Sea region of Turkey. L. interrogans was determined by PCR in sera, kidney and brain tissue. Sixteen (27.1%) kidney samples and 10 brain tissue samples (16.9%) were positive for L. interrogans. No PCR positivity was seen in sera samples. Five sera were positive by microagglutination test. A large proportion of wild rats in our region were found to be carriers of L. interrogans. We conclude that people who are exposed to rat urine in their daily life are at risk of acquiring L. interrogans.

A competitive enzyme-linked immunosorbent assay (ELISA) using a specific monoclonal antibody (M898) was developed for detection of bovine antibodies to Leptospira interrogans serovar pomona. This assay was evaluated using field sera (n = 190) with serovar pomona microscopic agglutination test (MAT) titers of > or =100 as the positive population (group A); field sera (n = 1,445) which were negative in the MAT (1:100 dilution) for serovar pomona (group B); and sera (from a specific-pathogen-free cattle herd [n = 210]) which were negative in the MAT (1:100 dilution) for serovars canicola, copenhageni, grippotyphosa, hardjo, pomona, and sejroe (group C). At the cutoff point recommended by receiver operating characteristic (ROC) curve analysis of the combined ELISA results of serum groups A, B, and C, the sensitivity and specificity values were 93.7 and 96.3%, respectively. The value for the area under this ROC curve was 0.977, indicating a high level of accuracy for the ELISA. Similar results were obtained from the analysis of the combined results of serum groups A and B and from the analysis of the combined results of serum groups A and C.

A new repetitive DNA element was identified in an isolate of Leptospira interrogans serovar copenhageni from a patient in Salvador, Brazil. A Sau3A genomic library from this strain was constructed and screened for repetitive DNA elements. An insert of 438 bp (Rep1) from one library clone hybridized to multiple chromosomal DNA fragments resolved electrophoretically after digestion with BamHI, HindIII, and MfeI. A single oligonucleotide primer, designated iRepl, was designed to generate multiple PCR amplicons of various electrophoretic mobilities in a PCR typing method. The method distinguished strains belonging to the eight pathogenic and three saprophytic species of the genus Leptospira. Clinical isolates obtained during urban epidemics between 1996 and 1998 in Salvador, Brazil, were analyzed by this PCR method. Although the iRep1 primer was unable to discriminate strains among L. interrogans serovar copenhageni isolates, it was able to differentiate strains belonging to different species and serogroups of Leptospira identified in Salvador. This PCR-based method may provide a faster and less expensive alternative to serologic tests used in reference laboratories.

The results of clinical trials carried out in different foci have demonstrated high diagnostic value of analysis made with the use of the polymerase chain reaction (PCR) at early stages of Leptospira infection caused by infective agents of the serogroups Grippotyphosa, Canicola and Icterohaemorrhagiae. The possibility of leptospiremia lasting considerably longer than heretofore believed to be possible, as well as the persistence of leptospires in the liquor of patients after the acute phase of the disease is over, i.e. during the early and late convalescence periods, has been shown. This is indicative of good prospects of using the PCR analysis not only for early rapid diagnostics of Leptospira infections, but also for controlling the course of the infection, for prognosticating early and late complications of the disease, as well the mechanisms of pathogenesis.