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.     

Seroepidemiology of leptospirosis in livestock in Trinidad

A study was conducted to determine the seroprevalence of leptospirosis and infecting serovars across livestock (cattle, sheep, goats, and pigs) in Trinidad using the microscopic agglutination test with an international panel of 23 serovars. Of a total of 590 cattle tested, 21.5% were seropositive with agglutinations to 13 of the 23 antigens used in the panel. Icterohaemorrhagiae (9.3%), Sejroe (4.1%), Ballum (4.1%), and Autumnalis (1.9%) were the predominant serogroups detected in the cattle sampled (n = 590). Of 222 sheep tested, 5.0% were seropositive with agglutinations to five serovars belonging to two serogroups. These serogroups were Autumnalis at 2.7%, and Icterohaemorrhagiae at 2.3% of all sheep tested (n = 222). Of a total of 180 goats tested, 3.3% were seropositive, all agglutinating to the Icterohaemorrhagiae serogroup, 1.7% to serovar Copenhageni, 1.1% to serovar Mankarso, and 0.6% to serovar Icterohaemorrhagiae. Among pigs (n = 200), 5.0% were seropositive for five serovars belonging to three serogroups. These serogroups were Icterohaemorrhagiae at 2.5%, Australis at 2%, and Ballum at 0.5%. Overall, age and sex of animals were not significantly associated with leptospirosis with the exception of cattle where age was a significant factor for seropositivity. It was concluded that for livestock, leptospirosis may be an important zoonotic and economic disease, particularly in the case of cattle. It is imperative that the impact of leptospirosis on abortion, stillbirths, and decreased milk production in livestock in the country be assessed.     

Presence of Leptospiral DNA in Semen Suggests Venereal Transmission in Horses

The purpose of the present study was to detect leptospiral DNA by PCR in semen and urine samples of stallions to test for venereal transmission in horses. A total of 10 stallions from four herds were studied, and sampling was conducted in semen and urine for culture and PCR and serum for serology. From the 10 serum samples tested, 6 (60%) were seroreactive. No pure culture was obtained, but leptospiral DNA was detected by PCR in 50% of the semen samples and 30% of urine samples. The present study aimed to detect leptospiral DNA by PCR in semen and urine samples of stallions to test for venereal transmission in horses. Based on these findings, we suggest that there is potential transmission of leptospirosis in horses by sexual transmission.     

Serologic prevalence of toxoplasmosis in cattle, sheep, goats, pigs, bison, and elk in Montana

Serum samples from 2,539 cattle, 649 sheep, 123 goats, 413 pigs, 93 bison, and 56 elk from Montana were examined for antibody to Toxoplasma gondii in the Sabin-Feldman dye test or the modified agglutination test (MAT). Cattle, bison, and elk serum samples were treated with 0.2 M-mercaptoethanol before examination in MAT. In the dye test, 13.2% of sheep, 5.0% of pigs, and 22.7% of goats had antibody at a dilution of greater than or equal to 1:16. In the MAT, 3.2% of cattle, 3.1% of bison, and none of the elk were positive at a dilution of greater than or equal to 1:128.     

Role of Intraocular Leptospira Infections in the Pathogenesis of Equine Recurrent Uveitis in the Southern United States

Equine recurrent uveitis (ERU) has been linked to leptospirosis in Europe; however, regional differences exist in reports from the United States. The objective of this study was to investigate the role of intraocular leptospiral infections in horses with ERU in the Southern United States. Blood and ocular fluid samples were collected from horses with ERU and normal controls. Leptospira serology was performed using microscopic agglutination test (MAT). Aqueous and vitreous humor samples were obtained and submitted for aerobic and Leptospira culture, polymerase chain reaction (PCR), and MAT. Twenty-one control horses (40 eyes) and 31 ERU horses (46 eyes) were available. Serology was available for 48 of 52 horses: 16 of 21 control and 23 of 27 affected horses were positive for at least one serovar; bratislava was the most common serovar identified. Bacillus sp. and Micrococcus sp. were cultured from one control horse's eye; Streptococcus sp. (n = 1) and Leptospira (n = 6) were cultured from the eyes of six ERU horses. Leptospira isolates belonged to serogroup pomona (n = 4) and grippotyphosa (n = 2). Polymerase chain reaction results were positive in 14 of 31 (45%) horses with ERU; no control horses were positive by PCR (P = .0001). Microscopic agglutination test was positive for 17 of 24 ERU horses (71%) and one 21 (4.7%) normal horses (P < .0001). Horses with ERU had a high prevalence of Leptospira infection based on PCR and MAT results from intraocular fluids compared with control horses. The diagnosis of intraocular infections was not aided by serology and required specific invasive sampling of ocular fluid. Leptospira infection should be considered as a cause of ERU in the Southern United States.     

Strategies of the control of an outbreak of leptospiral infection in dairy cattle in Northeastern Brazil

The aim of the present study was to describe the strategies of the control of an outbreak of leptospiral infection in dairy cattle in Maranhão State, Northeastern Brazil. In the period from January to July 2015, 18 (17%) out of 106 cows presented abortion, six (5.7%) stillbirth, and 12 (11.3%) repeated estrus, totaling 24 animals with reproductive problems. The diagnosis of leptospirosis was based on serology (microscopic agglutination test—MAT), bacteriological culture, and polymerase chain reaction (PCR). Antibiotic therapy, vaccination protocols, and changes in management practices were suggested as control measures. Of all animals on the farm (n?=?280), 136 (48.6%) were seropositive for at least one serovar of Leptospira sp. No pure leptospiral culture was obtained. Eight of the animals with reproductive problems yielded positive PCR results (vaginal fluid of seven animals and urine and vaginal fluid of one animal). Genetic sequencing of a vaginal fluid/urine PCR-positive sample revealed Leptospira borgpetersenii. One year after the adoption of control measures, no reproductive problems were observed. Thus, leptospirosis probably caused the reproductive failures in the herd, and the control and prevention measures implemented were efficient in controlling the disease.

     

Leptospira interrogans infection in domestic and wild animals in Fiji

Clinical and serological evidence has indicated that human leptospirosis in Fiji is an important disease, and the prevalence of antibody is exceptionally high. A serological survey of the rural population showed that only 12% of the people studied did not have complement fixing (CF) leptospiral antibody. As the origin of this infection could not be explained by the known distribution of leptospiral infection in domestic and wild animals, a serological survey using the complement fixation test (CFT) was undertaken as the first stage of an epidemiological investigation into human and animal leptospirosis. Sera from domestic and wild animals were tested for CF antibody to 12 leptospiral serovars, namely: pomona, copenhageni, grippotyphosa, hardjo, ballum, tarassovi, canicola, australis, bratislava, autumnalis, pyrogenes and bataviae. Antibody was detected in 27.5% of 480 cattle, 17.1% of 70 sheep, 10.3% of 252 goats, 10.0% of 480 pigs, 57.0% of 100 dogs, 55.8% of 34 rats (Rattus rattus, R. frugivorus, R. exulans and R. norvegicus), 53.1% of 32 mongooses (Herpestes auropunctatus) and 40.0% of 10 mice (species unknown.) Cross-reactivity precluded the identification of infecting serogroups with the exception of pomona in pigs and icterohaemorrhagiae, ballum and australis in dogs. Infection of dogs with a serovar of the australis serogroup may explain the predominance of serological reactions to bratislava in man. The survey revealed a significant level of leptospiral antibody in the animal populations of Fiji and indicated that cattle, dogs, rats, mongooses and mice are probably the most important maintenance hosts. Consequently, further investigation will concentrate on the attempted isolation of leptospires from these species.     

Seroprevalence and risk factors for leptospirosis in cattle,sheep, and goats at consorted rearing from the State of Piauí, northeastern Brazil

Leptospirosis is an endemic disease in Latin America, caused by pathogenic bacteria of the genus Leptospira. It is considered one of the main causes responsible for the negative economic impact on global livestock by causing reproductive problems. The research aimed to determine the prevalence of leptospirosis in cattle, sheep, and goats at consorted rearing in the micro-region of Teresina, Piauí state, northeastern Brazil, as well as to identify prevalent serovars and risk factors associated with seroprevalence. Serum samples were analyzed in 336 sheep, 292 goats, and 253 cattle using microscopic agglutination test (MAT). Overall, 378 samples were positive to MAT, with seroprevalence of 42.9%. The prevalences in cattle, sheep, and goats were 50.5, 40.5, and 34.6%, respectively. All herds presented at least one seropositive animal; the Hardjo/Wolffi serovar association was the most common in cattle and Icterohaemorrhagiae in goats and sheep. Beef production (OR?=?4.9), cattle herd over 35 animals (OR?=?4.0), feeding on pasture (OR?=?6.4), weir and/or stream as water source (OR?=?2.1), and no veterinary services (OR?=?2.9) were risk factors for cattle infection. For sheep, intensive management system (OR?=?5.3), suspended slatted facilities (OR?=?2.2), more than 20 sheep in reproductive age (OR?=?1.9), and absence of deworming (OR?=?3.5) were the risk factors, while for goats, the identified risk factors were sheep herd over 52 animals (OR?=?1.9) and no veterinary services (OR?=?1.8). We conclude that the infection was spreading in consorted herds in this region. Thus, it would be interesting and important to conduct educative activities to farmers on the economic impacts of this disease and the need of preventive and control strategies mainly focused on sanitary measures and animal handling.     

Evaluation of a recombinant LipL41 antigen of Leptospira interrogans serovar canicola in ELISA for serodiagnosis of bovine leptospirosis

The efficacy of a recombinant leptospiral lipoprotein LipL41 as an antigen for conducting enzyme-linked immunosorbent assay (ELISA) for diagnosis of bovine leptospirosis was evaluated. Using known positive and known negative cattle sera the recombinant antigen was found to be highly reactive in the concentration of 100 ng/well. Using a total of 321 field cattle sera the sensitivity of ELISA as compared to microscopic agglutination test (MAT) was calculated to be 100% whereas the specificity was 85.3%. The seropositivity of leptospirosis among bovine population was found to be 21.18% having the predominance of serovars Sejroe and Pomona. It was concluded that rLipL41 protein could be a putative diagnostic candidate for serodiagnosis of bovine leptospirosis.     

Identification of Leptospira spp. carriers among seroreactive goats and sheep by polymerase chain reaction

Few studies were conducted on the diagnosis and control of small ruminants’ leptospirosis. Thirteen goat herds and seven sheep flocks located in the state of Rio de Janeiro, Brazil, were screened for leptospirosis. From the three herds and three flocks with greatest seroreactivity by MAT (Microscopic Agglutination Test), 19 and 40 seropositive goats and sheep, respectively, were selected, and urine samples were collected for bacteriology and PCR. For both species of animals, the most prevalent reactions were due to serogroups Sejroe and Shermani. Although leptospires were observed by darkfield microscopy in eight samples, pure isolates were obtained by bacteriological culture from only two samples. However, twelve urine samples (six goats and six sheep) were positive by PCR. Based on these findings, we consider that the combined use of MAT as a screening test followed by urine PCR for the direct detection of Leptospira spp. DNA was adequate for the identification of carrier animals among goats and sheep. These are valuable tools for the control of leptospirosis in small ruminants.     

Neutralizing antibody to bovine rotavirus in various animal species

A total of 288 serum samples were collected from 12 species of animals in various localities of Japan from 1975 to 1977. Neutralizing antibody to bovine rotavirus was found in serum samples from all the species, viz., horses, cattle, sheep, goats, pigs, dogs, rabbits, guinea pigs, rats, mice, chickens and human beings. The incidence of neutralizing antibody in titers of 1 : 2 or higher ranged from 31 to 100%, and high incidences exceeding 70% were obtained in horses, cattle, sheep, pigs, dogs, rabbits and human beings. High titers were most common in horses, cattle, sheep and pigs. These serological results suggest that rotaviruses occur commonly in these species of animals.     

Serodiagnosis of leptospirosis in pigs using an axial filament enzyme-linked immunosorbent assay.

The axial filament (AF) from Leptospira interrogans serovar canicola was isolated by cesium chloride density gradient centrifugation of 2% sarcosyl treated whole cells. Isolation of AF was confirmed by electron microscopic examination, by protein-A immunogold labelling, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), and immunoblotting. Analysis by SDS-PAGE of the purified preparation showed relatively weak bands of molecular size 41 kDa and 21 kDa, and strong bands of 35 kDa and 34.5 kDa. Immunoblot analysis using antiserum to the AF against sonicated leptospires of a variety of serovars showed prominent reaction against the 41, 35, and 34.5 kDa protein bands, as well as against minor bands of molecular weight 43, 39, and 37 kDa. Antisera prepared against leptospiral serovars also identified minor bands at 33 and 32 kDa. Immunoblots with antiserum to whole cells of serovar bratislava detected the 35 and 34.5 kDa AF bands of Borrelia burgdorferi moderately and of Treponema hyodysenteriae only slightly in comparison to leptospiral AF. Antibody to B. burgdorferi did not detect the leptospiral AF antigen. Immunoblots with antiserum to T. hyodysenteriae showed a marked reaction with a 41 kDa band of B. burgdorferi but only a very minor reaction with leptospiral AF. The AF was tested in an AF-ELISA against sera from 260 pigs, many of which reacted in the microscopic agglutination test (MAT) against one or more leptospiral serovars. A sensitivity of 97.1% and a specificity of 93.1% was determined in comparison to the MAT. Only moderate correlation was observed between titres detected in the AF-ELISA and the MAT (r = 0.4). When sonicated whole cells (WC) of serovar canicola were used in an ELISA (WC-ELISA), high correlation was observed between AF-ELISA and WC-ELISA (r = 0.97). These findings show that the AF-ELISA can be used effectively as a species-specific antigen for the serological diagnosis of leptospirosis in swine and that sonicated whole cells can substitute excellently for purified AF as the antigen source. These findings may be extrapolated to the use of AF in immunodiagnosis of leptospirosis in other species.     

Serological prevalence of bovine leptospirosis in Plateau State, Nigeria

Serum samples obtained from 1.537 cattle in the 14 local government areas (LGAs) of Plateau State of Nigeria were screened for the presence of leptospiral antibodies using 13 serovars in a modified microscopic agglutination test (MAT). Two hundred and twenty-two (14.4 p.100) of the cattle tested had leptospiral antibody titres of 1:100 or higher to one or more of the test antigens. The prevalence rates of antibodies to individual serovars were: hardjo (35.6 p.100), pomona (11.7 p.100), pyrogenes (11.7 p.100), canicola (9.5 p.100), grippotyphosa (7.7 p.100), bratislava (5.9 p.100), icterohaemorrhagiae (5.9 p.100), ballum (4.5 p.100), autumnalis (3.6 p.100), bataviae (2.3 p.100) and tarassovi (1.8 p.100). The serological prevalence of bovine leptospirosis in the various local government areas of Plateau State of Nigeria differed significantly (P less than 0.05; X2).     

Prevalence and risk factors associated with Cryptosporidium spp. infection in young domestic livestock in India

A total of 938 faecal samples (461 cattle calves, 264 buffalo calves, 55 lambs, 116 kids and 42 piglets) from different livestock farms and individual small holdings in six targeted states of India were collected and screened by modified Ziehl–Neelsen staining technique to determine the prevalence of Cryptosporidium spp. and its association with age, sex, season and faecal consistency in domesticated animals. Overall, 16.2 % of the animals were positive for Cryptosporidium infection with prevalence of 16.3, 24.2, 1.8, 3.5 and 19.1 % in cattle calves, buffalo calves, lambs, kids and piglets, respectively. The prevalence of infection was significantly higher (p?<?0.05) in bovines (19.3 % cattle and 33.7 % buffalo) below 1 month of age than in animals between 1 and 3 months of age. But in piglets, it was higher in the age group of 1 to 3 months (22.6 %) than in younger animals (9.1 %). Also, higher prevalence (p?>?0.05) was recorded in females than in males. Seasons had a significant effect (p?<?0.05) on the prevalence of infection in large ruminants, with the highest prevalence in monsoon (cattle 28.8 % and buffalo 36.6 %) followed by pre-monsoon and post-monsoon season. However, in case of sheep and goats, the prevalence was higher (p?>?0.05) in post-monsoon than in monsoon season. A high degree of association was noticed between Cryptosporidium infection and diarrhoea in ruminants screened during the present study. But, in case of pigs, the prevalence was higher in non-diarrhoeic than in diarrhoeic animals. Genotyping of Cryptosporidium spp. based on nested PCR amplification of partial 18S rRNA and its subsequent digestion with SspI, VspI and MboII restriction enzymes revealed prevalence of Cryptosporidium parvum in representative number of positive samples of cattle, buffalo and goats.     

The comparative role of cattle, goats and pigs in the epidemiology of livestock trypanosomiasis on the plateau of eastern Zambia

To determine and compare the prevalence of trypanosome infections in different livestock species (cattle, pigs and goats) in areas where game animals are scarce and livestock constitute the main food source of tsetse, a survey was conducted on the plateau of the Eastern Province of Zambia in Katete and Petauke districts where Glossina morsitans morsitans is the only tsetse species present. Blood was collected from a total of 734 cattle, 333 goats and 324 pigs originating from 59 villages in both districts and was examined using the buffy coat method and the PCR-RFLP as diagnostic tools. The prevalence of trypanosome infections differed substantially between livestock species. Using microscopic diagnostic methods, trypanosome infections were detected in 13.5% of the cattle and 0.9% of the pigs. All goats were parasitologically negative. The PCR-RFLP analyses increased the trypanosomiasis prevalence to 33.5, 6.5 and 3.3% in cattle, pigs and goats respectively. The majority of the infections (91.2%) were due to Trypanosoma congolense. The presence of a trypanosome infection in cattle and pigs resulted in a significant decline in the packed cell volume. The outcome of the study clearly shows that despite the availability of goats and pigs, cattle seem to be the major livestock species affected by the disease in trypanosomiasis endemic areas. The high proportion of infections in cattle could be partly attributed to their higher availability and attractiveness to tsetse.     

New enzyme-linked immunoassay for the detection of specific antibodies against multiple Leptospira serogroups in bovine sera

Leptospirosis is a zoonotic disease with worldwide endemicity in Argentina, it is a significant public health problem in low-income populations. Bovine leptospirosis is a serious economic problem for cattle production, causing abortions, reduced milk yield, mortality in calves and decreased daily weight gain. We developed an enzyme-linked immunosorbent assay (ELISA) with sonicated Leptospira interrogans serogroup Icterohaemorrhagiae serovar Copenhageni M 20. We evaluated its performance for the detection of specific antibodies against multiple Leptospira serogroups in bovine. The microscopic agglutination test (MAT) was used as the gold standard. The performance of this ELISA was evaluated with a panel of sera (118 MAT confirmed positive and 97 MAT negative). The overall sensitivity was close to 85.6 % and the specificity was 83.5 %, according to the MAT reference method. Analytical specificity of the IgG-ELISA was evaluated using 50 bovine serum samples from animals showing serum antibodies against other pathogens that cause abortion in bovine, such as Brucella sp., Neospora sp. and Bovine Viral Diarrhea (BVD), and no cross-reaction was observed. This IgG-ELISA can be an alternative to the MAT for diagnosis of leptospiral infection in bovine.     

Epidemiological Analysis of Leptospira spp. Infection in Equids from the Brejo Paraibano Microregion of Brazil

Leptospirosis is a zoonotic disease that occurs in humans and several animal species, including cattle, pigs, and horses, and it has a worldwide distribution. In equids, leptospirosis manifests through recurrent uveitis and abortion and other reproductive disorders. The objective of the present study was to investigate the epidemiological situation of Leptospira spp. infection in equids from the Brejo Paraibano microregion in the northeast of Brazil, that is, to assess the prevalence of and identify the major risk factors for infection. For this purpose, blood samples were taken from 257 equids (204 horses, 46 mules, and 7 donkeys), and a microscopic agglutination test (MAT) was used as serological analysis, with 24 serovars of Leptospira spp. as antigens. Antibodies were observed in 16.2% of horses, 13.0% of mules, and 28.6% of donkeys. Animals between the ages of 2.5 and 11 years had a risk of infection 0.4 times lower than those younger than 2.5 years old (odds ratio, 0.4; 95% confidence interval, 0.18-0.90). Further studies are needed to identify potential wild reservoirs and the main routes of infections.     

A systematic review of leptospirosis on dogs,pigs, and horses in Latin America

Leptospirosis is a worldwide zoonosis which can affect many species. Control programs need accurate diagnosis to be successful, and currently, diagnosis relies on serology. It presents three main issues: the sampling, the antigen panel, and the cutoff point. Herein, we propose a systematic review on leptospirosis among dogs, pigs, and horses in Latin America in order to improve the understanding of the seroepidemiology of leptospirosis in these species in the region as well as the temporal development of the research on this topic and, consequently, improve the chances of success on control programs. Internet databases were consulted over 2015. Inclusion criteria included serosurvey using MAT; a relevant number of animals; the presence in the antigen panel of at least one representative of serogroups Icterohaemorrhagiae and Canicola for dogs, Icterohaemorrhagiae, Australis, and Pomona for pigs, and Icterohaemorrhagiae and Australis for horses; and a cutoff point of ≥100. Overall, 240 papers were studied, of which 87 referred to dogs, 66 to pigs, 39 to horses, and 48 to more than one of the studied species. In relation to those that met all the inclusion criteria, it was 45 (66.2%) in dogs, 23 (41.8%) in pigs, and 23 (63.9%) in horses. Leptospirosis is widespread in Latin America. Predominant serogroups are Canicola to dogs and Icterohaemorrhagiae to pigs and horses. Therefore, research on animal leptospirosis should be encouraged in Latin America, in order to reach a greater standardization in studies and then achieve better results on control programs.     

35 leptospira isolated from the vitreous body of 32 horses with recurrent uveitis (ERU)

130 vitreous samples, systematically collected in 1998 from 117 horses during vitrectomy, were cultured for the presence of leptospires. All horses suffered from equine recurrent uveitis (ERU), also known as periodic ophthalmia or moon blindness, and were treated surgically to combat painful attacks, and to preserve vision. In 35 out of 130 vitreous samples (35/130 = 26.9%), leptospires could be isolated. These isolates belong to the grippotyphosa serogroup (n = 31) and to the australis serogroup (n = 4). So, for the first time, leptospires were recovered from eyes in vivo in a large number of horses with ERU. Vitreous samples and one serum sample from each horse were also tested for leptospiral antibodies using the microscopic agglutination test (MAT). In 92 vitreous samples (92/130 = 70.7%) and 96 serum samples (96/117 = 82.0%) leptospiral antibodies were detected at a dilution of > 1:100. The presence of intact leptospires and specific antibodies in eyes affected with ERU demonstrates a local antibody production to leptospiral antigen. These results indicate an important etiological role of leptospires in equine recurrent uveitis.     

Evaluation and comparison of native and recombinant LipL21 protein-based ELISAs for diagnosis of bovine leptospirosis

A 21-kDa leptospiral lipoprotein (LipL21) was evaluated for its diagnostic potential to detect bovine leptospirosis by ELISA. Both native LipL21 (nLipL21) and recombinant LipL21 (rLipL21) proteins were tested and compared regarding diagnostic efficiency, and no statistically significant difference was observed. The sensitivity of rLipL21 ELISA for 62 microscopic agglutination test (MAT) positive sera was 100% and the specificity with 378 MAT negative sera was 97.09%. Thus, rLipL21 protein-based ELISA could be used as an alternative to MAT for the diagnosis of bovine leptospirosis.