Interrogans Serovars Research Articles

Herd- and animal-level risk factors for bovine leptospirosis in Tanga region of Tanzania

Leptospirosis is the zoonosis of worldwide distribution and common cause of economic loss and ill health among animals and human populations. A cross-sectional seroprevalence study, using a microscopic agglutination test (MAT) with a threshold titre of >or=1:160, to elucidate disease magnitude, distribution and associated risk factors in cattle in Tanga, Tanzania was conducted from May 2003 to January 2004. Serum (n = 655) samples collected from randomly selected herds (n = 130) were tested for antibodies against four different Leptospira interrogans serovars (Bataviae, Tarassovi, Hardjo and Pomona) used in the agglutination test. Positive titres were detected in 30.3% [95% confidence intervals (CI) = 26.7-33.9] of cattle and 58.5% (95% CI = 49.5-67.1) of herds, respectively. Of the 198 MAT positive serum samples, 98 (49.5%) were positive against serovar Hardjo, 80 (40.4%) were positive against serovar Tarassovi, 12 (6.1%) was positive against serovar Bataviae and eight (4%) were positive against serovar Pomona. Associations found to be statistically significant in univariate analyses (at P < 0.1) were assessed by multivariable logistic regression to control for confounding factors. The results showed that risk factors for cattle were pasture grazing [odd ratio (OR) = 2.83, 95% CI = 1.57-5.12, P = 0.001], presence of goats/sheep on the farm (OR = 1.73, 95% CI = 1.17-2.56, P = 0.001) and age of the animal (OR = 2.05, 95% CI = 1.42-2.96, P = 0.001), while concrete floor housing was protective (OR = 0.47, 95% CI = 0.30-0.74, P = 0.001). Herds managed under pasture grazing system were more likely to be sero-positive than those managed under zero grazed practices (OR = 9.31; 95% CI = 3.67-23.64 for grazing herd). We concluded that bovine leptospirosis is an endemic and locally widespread disease in Tanga and suggest that it may play a role in zoonotic transmission to humans.

In silico identification of common putative drug targets in Leptospira interrogans

Infectious diseases are the leading causes of death worldwide. Hence, there is a need to develop new antimicrobial agents. Traditional method of drug discovery is time consuming and yields a few drug targets with little intracellular information for guiding target selection. Thus, focus in drug development has been shifted to computational comparative genomics for identifying novel drug targets. Leptospirosis is a worldwide zoonosis of global concern caused by Leptospira interrogans. Availability of L. interrogans serovars and human genome sequences facilitated to search for novel drug targets using bioinformatics tools. The genome sequence of L. interrogans serovar Copenhageni has 5,124 genes while that of serovar Lai has 4,727 genes. Through subtractive genomic approach 218 genes in serovar Copenhageni and 158 genes in serovar Lai have been identified as putative drug targets. Comparative genomic approach had revealed that 88 drug targets were common to both the serovars. Pathway analysis using the Kyoto Encyclopaedia of Genes and Genomes revealed that 66 targets are enzymes and 22 are non-enzymes. Sixty two common drug targets were predicted to be localized in cytoplasm and 16 were surface proteins. The identified potential drug targets form a platform for further investigation in discovery of novel therapeutic compounds against Leptospira.

High-resolution melt-curve analysis of random-amplified-polymorphic-DNA markers, for the characterisation of pathogenicLeptospira

A new test for pathogenic Leptospira isolates, based on RAPD-PCR and high-resolution melt (HRM) analysis (which measures the melting temperature of amplicons in real time, using a fluorescent DNA-binding dye), has recently been developed. A characteristic profile of the amplicons can be used to define serovars or detect genotypes. Ten serovars, of leptospires from the species Leptospira interrogans (serovars Australis, Robinsoni, Hardjo, Pomona, Zanoni, Copenhageni and Szwajizak), L. borgpetersenii (serovar Arborea), L. kirschneri (serovar Cynopteri) and L. weilii (serovar Celledoni), were typed against 13 previously published RAPD primers, using a real-time cycler (the Corbett Life Science RotorGene 6000) and the optimised reagents from a commercial kit (Quantace SensiMix). RAPD-HRM at specific temperatures generated defining amplicon melt profiles for each of the tested serovars. These profiles were evaluated as difference-curve graphs generated using the RotorGene software package, with a cut-off of at least 8 'U' (plus or minus). The results demonstrated that RAPD-HRM can be used to measure serovar diversity and establish identity, with a high degree of stability. The characterisation of Leptospira serotypes using a DNA-based methodology is now possible. As an objective and relatively inexpensive and rapid method of serovar identification, at least for cultured isolates, RAPD-HRM assays show convincing potential.

Ileocolic intestinal intussusception in a dog with leptospirosis

Leptospirosis is a zoonotic infection causing clinical and subclinical disease in dogs, humans, and other mammalian species. Although more than 200 different Leptospira interrogans serovars have been identified (11), the serovars most commonly isolated from dogs with clinical leptospirosis in Southern Germany are L. grippothyphosa and L. saxkoebing (3). However, vaccines currently available in Germany only include the serovars L. canicola and L. icterohaemorrhagiae. They lack the ability to build up a sufficient protective immune response against other serovars, therefore explaining clinical disease in regularly vaccinated dogs (3, 16). The best known maintenance hosts for leptospira serovars are rodents (17), and an increase in the prevalence of cases seems to be linked to higher average rainfalls (19). This might explain why male outdoor working or herding dogs in rural areas are found to carry an increased risk for infection (19, 20). Clinical signs associated with canine leptospirosis often include non-specific symptoms such as fever, anorexia, vomiting, and lethargy, and clinical signs of hepatic and renal failure such as icterus, polyuria/polydipsia, and anuria (3, 6, 7, 17). Typical clinical features also seem to depend on the serovars involved in infection (4). Other less common clinical signs include dyspnoea, cough, diarrhoea, abdominal pain, muscle pain, and stiffness (10, 17). The patient described in this case report presented with acute hepatic icterus and renal failure. In addition to these common clinical signs of canine leptospirosis, the dog developed intestinal intussusception as a rare complication associated with the infection. Case history

Evaluation of a recombinant LigB protein of Leptospira interrogans serovar Canicola in an enzyme-linked immunosorbent assay for the serodiagnosis of bovine leptospirosis

A recombinant leptospiral lipoprotein, LigB, was evaluated for use in the diagnosis of bovine leptospirosis by enzyme-linked immunosorbent assay (rLigB IgG ELISA). The standard reference test (Microscopic agglutination test, MAT) of 200 serum samples from cattle suspected of leptospirosis showed that 95 (47.5%) samples had positive agglutination titres, which ranged from 100 to 1600. In rLigB IgG ELISA, 49% of the samples were positive. Sensitivity of IgG ELISA for 95 bovine sera, which had MAT titres of greater than or equal to 100, were 100%. ELISA showed a specificity of 97.1% with 105 bovine sera, which were negative at a 1:50 dilution in MAT for Leptospira interrogans serovars. The results of ELISA and MAT correspond very good. When analytical specificity of IgG ELISA was evaluated using bovine serum samples from animals showing the serum antibodies to other pathogens, no cross-reaction was observed. Thus the recombinant LigB IgG ELISA can be used instead of the MAT as an aid to the diagnosis of bovine leptospirosis.

Retrospective Evaluation of Idiopathic Hematuria and Associated Pathology in Grant's Gazelles (Gazella granti): 10 Cases

Ten cases of hematuria in Grant's gazelle (Gazella granti) (two male and eight female) from five institutions were examined and the clinicopathologic data summarized. Five gazelles died spontaneously and five were euthanized. All gazelles had marked hematuria without pyuria. Mean age at the onset of clinical signs and time of euthanasia or death was 5.0 +/- 1.4 yr and 8.2 +/- 3.7 yr, respectively. The severity of clinical signs with hematuria ranged from episodes of chronic intermittent hematuria to marked dysuria, with urinary bladder rupture secondary to obstructive blood clots in one case. Submandibular edema was the most common associated clinical sign (five of 10 cases). Serum chemistries from eight gazelles obtained during hematuria episodes revealed hypocalcemia (8/8), hypoproteinemia (7/8), hypoalbuminemia (7/8), and hyperphosphatemia (6/8). Fifty percent of the gazelles (4/8) developed anemia over the course of hematuria episodes. Prothrombin times and partial thromboplastin times were presumed increased in two of four animals evaluated. The predominant histologic lesions in seven of 10 gazelles reviewed were vascular necrosis, vasculitis, and perivasculitis in the urinary tract. Lesions in necropsied gazelles were identified in the urinary bladder (7/10 gazelles), kidney (3/10), and ureter (3/10). Additional urinary tract lesions included tubulointerstitial nephritis (5/10 gazelles), hemorrhagic cystitis (4/10), renal tubular necrosis (4/10), and subacute renal infarcts (2/10). Polymerase chain reaction testing on paraffin-embedded urinary tract tissue for alcelaphine herpesvirus-1 and -2, ovine herpesvirus-2, bluetongue virus, and epizootic hemorrhagic disease virus was negative for the six cases tested. One gazelle that had been vaccinated for Leptospira interrogans had a titer to serovar icterohaemorrhagiae, but serum from the six other gazelles tested was negative for all L. interrogans serovars. No exposure to any toxic agent was identified. An underlying cause for vascular lesions associated with episodic hematuria in Grant's gazelles remains to be determined.

Risk factors associated with the seroprevalence of leptospirosis, amongst at-risk groups in and around Tanga city, Tanzania

The epidemiological status of human leptospirosis in Tanzania has not been well defined because of a general lack of awareness of the disease and difficulties in its diagnosis. A cross-sectional seroprevalence study, using the microscopic agglutination test with a threshold titre of 1 : 160, was therefore conducted, among various occupational groups from Tanga city, in November 2005. Of the 199 subjects investigated, 30 (15.1%) were found to have antibodies that reacted with antigens from at least one of the six Leptospira interrogans serovars used in the agglutination test. Of the seropositive samples, 36.6% were positive for serovar Icterohaemorrhagiae, 30.0% for Bataviae, 20.0% for Hardjo, 6.6% for Tarrassovi, 3.3% for Ballum and 3.3% for Pomona. Amongst the occupational groups examined, livestock farmers, veterinary/meat inspectors and abattoir workers appeared to be most at risk, with seroprevalences of 19.4%, 18.1% and 17.1%, respectively. Subjects who reported that they milked cows were significantly more likely to be found seropositive for Leptospira infection than the other subjects (odds ratio=3.44; P<0.001). These findings indicate that a significant proportion of the population of Tanga city is being exposed to pathogenic Leptospira. Public-health interventions against leptospirosis will need to target not only the occupational groups considered to be at high risk of infection but also the general population at large.

Analysis of differential proteomes in pathogenic and non‐pathogenic Leptospira: Potential pathogenic and virulence factors

Leptospirosis is a bacterial zoonotic disease caused by spirochetes in the genus Leptospira. To date, factors determining the pathogenicity and virulence of leptospires remain unclear. We performed a gel-based proteomic analysis to evaluate differential leptospiral proteomes in the pathogenic L. interrogans (serovars Australis, Bratislava, Autumnalis, and Icterohaemorrhagiae) and the non-pathogenic L. biflexa (serovar Patoc). Quantitative proteome analysis and MS protein identification revealed 42 forms of 33 unique proteins whose levels were significantly greater in the pathogenic serovars compared with the non-pathogenic serovar. Among the four pathogenic serovars, the more virulent serovar Icterohaemorrhagiae (which is most commonly associated with severe leptospirosis in patients) had significantly greater levels of 14 forms of 12 unique proteins, when compared with the other three pathogenic serovars. Some of these identified proteins may serve as the pathogenic and/or virulence factors of leptospirosis.

Serologic Survey for Selected Infectious Diseases in Raccoons (Procyon lotor) in Indiana, USA

The objective of this study was to characterize the antibody prevalence of important livestock and domestic animal pathogens in raccoons (Procyon lotor) trapped and sampled in 39 forest patches in north-central Indiana, USA, between 2004 and 2005. A total of 459 serum samples were tested for antibodies to Leptospira serovars, 512 for Canine distemper virus (CDV) antibodies, and 340 for antibodies to Porcine reproductive and respiratory syndrome virus (PRRSV). In total, 47, 16, and 0% of the samples were seropositive for at least one Leptospira serovar, CDV, and PRRSV, respectively. Most of the positive Leptospira results were to serovars grippotyphosa (36%), autumnalis (22%), and hardjo (22%). No statistically significant correlation was found between antibody prevalence estimates for different Leptospira serovars. A significant association was found between body weight and antibodies for Leptospira serovars and CDV. In addition, age (adult vs. juvenile) was significantly associated with the presence of CDV antibody, with adults exhibiting a higher prevalence than juveniles. This study confirmed that raccoons in Indiana, USA, are exposed to different Leptospira interrogans serovars and CDV and that age and weight are associated with the presence of antibodies for both pathogens.

Nationwide Survey of Leptospira Antibodies in Dogs in Japan: Results from Microscopic Agglutination Test and Enzyme-Linked Immunosorbent Assay

Leptospirosis is an infectious disease caused by Leptospira interrogans sensu lato and is common in both humans and animals. In the present study, serum samples were collected from 801 dogs across all 47 prefectures in Japan, and evaluated with a microscopic agglutination test (MAT), using 5 major L. interrogans serovars (Icterohaemorrhagiae, Canicola, Autumnalis, Hebdomadis, and Australis) as antigens, and an enzyme-linked immunosorbent assay (ELISA) using recombinant OmpL1 protein as the antigen. Across all dogs tested, 217 (27.0%) and 29 (3.6%) were MAT- and ELISA-positive, respectively. However, evidence strongly suggests that MAT also detected antibodies produced by vaccination. Of 243 dogs never inoculated with any canine vaccine, 41 (16.9%) from 23 prefectures were MAT and/or ELISA positive. The most commonly detected serovar was Icterohaemorrhagiae (22 dogs, 19 prefectures). Our results suggest that there are dogs with subclinical Leptospira infection throughout Japan. To the best of our knowledge, the present study is the first nationwide survey of Leptospira infection in dogs, and the findings are relevant not only for clinical veterinary medicine but also for public health.

Viral and bacterial serology of six free-ranging bearded seals Erignathus barbatus

Serum or heparinized plasma samples were obtained from 3 male (2 adult and 1 weaned calf) and 3 adult female free-ranging bearded seals Erignathus barbatus in May of 1994, 1995, or 1996. Blood samples were obtained from animals taken in subsistence hunts near St. Lawrence Island, Alaska and screened for antibodies to a suite of bacteria and viruses potentially pathogenic for pinnipeds and/or humans. No samples had detectable antibodies to Brucella spp., Phocine distemper virus, influenza A virus or caliciviruses (San Miguel sea lion virus strains 1, 2, and 4 to 13, vesicular exanthema of swine serotypes A48, B51, C52, D53, E54, F55, G55, H54, 155, J56, K54, 1934B, and Tillamook and Walrus calicivirus). One seal had a low titer of 100 to Leptospira interrogans serovar

Household Transmission of Leptospira Infection in Urban Slum Communities

BackgroundLeptospirosis, a spirochaetal zoonotic disease, is the cause of epidemics associated with high mortality in urban slum communities. Infection with pathogenic Leptospira occurs during environmental exposures and is traditionally associated with occupational risk activities. However, slum inhabitants reside in close proximity to environmental sources of contamination, suggesting that transmission during urban epidemics occurs in the household environment.Methods and FindingsA survey was performed to determine whether Leptospira infection clustered within households located in slum communities in the city of Salvador, Brazil. Hospital-based surveillance identified 89 confirmed cases of leptospirosis during an outbreak. Serum samples were obtained from members of 22 households with index cases of leptospirosis and 52 control households located in the same slum communities. The presence of anti-Leptospira agglutinating antibodies was used as a marker for previous infection. In households with index cases, 22 (30%) of 74 members had anti-Leptospira antibodies, whereas 16 (8%) of 195 members from control households had anti-Leptospira antibodies. Highest titres were directed against L. interrogans serovars of the Icterohaemorrhagiae serogroup in 95% and 100% of the subjects with agglutinating antibodies from case and control households, respectively. Residence in a household with an index case of leptospirosis was associated with increased risk (OR 5.29, 95% CI 2.13–13.12) of having had a Leptospira infection. Increased infection risk was found for all age groups who resided in a household with an index case, including children <15 years of age (P = 0.008).ConclusionsThis study identified significant household clustering of Leptospira infection in slum communities where recurrent epidemics of leptospirosis occur. The findings support the hypothesis that the household environment is an important transmission determinant in the urban slum setting. Prevention therefore needs to target sources of contamination and risk activities which occur in the places where slum inhabitants reside.

Seroprevalence of leptospirosis in domesticated Asian elephants (Elephas maximus) in north and west Thailand in 2004

Serum samples from Asian elephants (Elephas maximus) in the Kanchanaburi, Chiang Mai and Lampang provinces of Thailand were tested using the microscopic agglutination test against 22 serovars of Leptospira interrogans....

Evaluation of the recombinant LipL32 in enzyme-linked immunosorbent assay for the serodiagnosis of bovine leptospirosis

The recombinant leptospiral protein LipL32 was evaluated for use in the diagnosis of bovine leptospirosis by enzyme-linked immunosorbent assay (rLipL32 IgG ELISA). The microscopic agglutination test (MAT) of 150 serum samples from cattle suspected of leptospirosis showed that 125 (83.3%) samples had positive reciprocal agglutination titres, which ranged from 100 to 1600. The highest titres were observed for the serovars Hardjoprajitno and Bratislava. In the rLipL32 IgG ELISA, 83.3% of the samples were positive. The sensitivity of IgG ELISA for 125 bovine sera, which had MAT titres of greater than or equal to 100, was 100%. ELISA showed a specificity of 100% with 58 bovine sera, which were negative at a 1:50 dilution in the MAT for Leptospira interrogans serovars. When analytical specificity of the IgG ELISA was evaluted using 60 bovine serum samples from animals showing serum antibodies to other pathogens that cause abortion in cattle, such as Babesia sp., Anaplasma sp. and Brucella sp. and no cross-reaction was observed. The recombinant LipL32 IgG ELISA can be an alternative to the MAT for diagnosis of leptospiral infection in cattle.

Seroepidemiology of leptospirosis, toxoplasmosis, and leishmaniosis among dogs in Ankara, Turkey

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 fluoresence 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.

Disease survey of free-ranging grey brocket deer (Mazama gouazoubira) in the Gran Chaco, Bolivia.

Samples from 17 free-ranging hunter-killed grey brocket deer (Mazama gouazoubira) in the Gran Chaco, Bolivia, were collected during June-August 1999. All 17 deer appeared to be in good condition at the time of death. Gross necropsies were performed, serum was collected for serologic evaluation of selected infectious disease agents, and feces and ectoparasites were collected for evaluation of internal and external parasites. Serologic tests were positive for antibodies against bovine respiratory syncytial virus and four Leptospira interrogans serovars, with questionable results for epizootic hemorrhagic disease virus serotypes 1 and 2. No antibodies were detected to Anaplasma marginale, Babesia bigemina, Babesia bovis, Babesia odocoilei, bluetongue virus (serotypes 2, 10, 11, 13, and 17), bovine viral diarrhea virus, Brucella abortus, foot-and-mouth disease virus, infectious bovine rhinotracheitis virus, Mycobacterium avium subsp. paratuberculosis, and parainfluenza-3 virus. Sixty-four percent (7/11) of the deer had endoparasites. Amblyomma spp. ticks were found on seven deer, flies of the family Hippoboscidae on six deer, and lice on six deer.

Serological evidence of Brucella species and Leptospira interrogans serovars in Greek swine herds

In a serological survey in intensively managed swine herds in northern Greece (280 samples) and southern Greece (120 samples), 28.2% of samples were positive for Leptospira interrogans serovars and 3% for Brucella species. These pathogens of public health significance should be systematically investigated in Greece, a swine brucellosis-free country.

The impact of Leptospira seropositivity on reproductive performance in sows in southern Viet Nam

A serologic survey was conducted among sows in the Mekong delta in southern Viet Nam to investigate associations between leptospiral seropositivity and reproductive performance. Data were collected from a total of 339 sows in lactation or gestation, from four large-scale state farms on three occasions. The seroprevalence for Leptospira interrogans serovar (sv) autumnalis was 32%, for L. interrogans sv bratislava 29%, for L. kirschneri sv grippotyphosa 13%, for L. interrogans sv icterohaemorrhagiae 27%, for L. interrogans sv pomona 5%, and for L. borgpetersenii sv tarassovi 13%. The reproductive parameters number of days from weaning to service (WSI), number of piglets born, number of piglets born dead, and number of piglets born weak, were evaluated. Seropositivity for sv tarassovi was associated with 0.8 more dead piglets per litter ( P=0.06), and sv grippotyphosa with a 1 day longer WSI ( P=0.06). There were no significant associations between reproductive performance and sv autumnalis, sv bratislava, sv pomona, and sv icterohaemorrhagiae. It is concluded that seropositivity for Leptospira can be associated with impaired reproductive performance even in areas where a high degree of immunity among sows is expected.

Viral and bacterial serology of free-ranging Pacific walrus.

Serum or heparinized plasma samples were obtained between 1994 and 1996 from 20 male and 20 female adult free-ranging Pacific walrus (Odobenus rosmarus divergens) from St. Lawrence Island and Round Island, Alaska. Samples were screened for antibodies to some potentially pathogenic bacteria and viruses. No sample had detectable antibody to Brucella spp. Three of 40 (8%) had low antibody titers to Leptospira interrogans serovars. Phocine distemper virus antibodies were not detected. Serologic responses to one or more caliciviruses (San Miguel sea lion virus 12 or vesicular exanthema of swine serotypes E54, F55, G55, 1934B) were detected in 18% (seven of 40) walrus. Antibodies to one or more subtypes of influenza A virus (H10, N2, N3, N5, N6, N7) were detected in 21% (eight of 38). Periodic screening of free-ranging populations for exposure to infectious diseases has become an important component of bio-monitoring programs to facilitate understanding and detecting trends in marine mammal populations.

Animal- and herd-level risk factors for leptospiral seropositivity among sows in the Mekong delta, Vietnam

In 1998, a total of 424 sows had sera collected in the Mekong delta in Vietnam. Of these, 283 sows were from 151 small-scale family farms in 19 villages, and 141 from seven large-scale state farms. The sera were subjected to the microscopic agglutination test (MAT) for antibodies to 13 Leptospira serovars. The overall leptospiral seroprevalence for titres ≥1:100 and ≥1:400, was 73 and 29%, respectively, and was higher ( P=0.001) at small- than at large-scale farms. The highest seroprevalence was recorded for Leptospira interrogans serovar (sv) bratislava (52%). At small-scale farms, higher prevalences were found to serovars L. interrogans sv icterohaemorrhagiae ( P=0.04) and L. interrogans sv pomona ( P=0.02). Epidemiological information (at the individual-animal and herd-levels) was collected with a questionnaire. The data were analysed using logistic multiple regression. At the animal-level, sows seropositive for L. interrogans sv australis and sv autumnalis had less direct contact with sows in neighbouring pens (odds ratio (OR)=0.3 and 0.4, respectively) and sows seronegative for L. interrogans sv bratislava were of lower age (OR=0.1 for seropositivity). Also, sows seropositive for L. interrogans sv icterohaemorrhagiae had higher odds (OR=5.8) if they had not been born on the farm (had been introduced to it as gilts). Herds seropositive for sv javanica showed association with farms not taking measures to control the local rodent population (OR=7.8). Serovar pomona was also linked to the use of artificial insemination (AI), as opposed to natural-breeding services (OR=11.2). These results indicate that housing and management could affect the seroprevalence of Leptospira infection in pigs.