EVALUATION OF A HARDJO-POMONA VACCINE TO PREVENT LEPTOSPIRURIA IN CATTLE EXPOSED TO A NATURAL CHALLENGE WITH LEPTOSPIRA INTERROGANS SEROVAR HARDJO

SUMMARY The efficacy of a vaccine against Leptospira interrogans serovar pomona and Leptospira interrogans serovar hardjo was evaluated in a group of dairy heifers that were serologically negative at the time of vaccination and later subjected to natural challenge with L. interogans serovar hardjo. Thirty-nine heifers were vaccinated twice, at a one-month interval, with a commercially prepared bivalent vaccine, while 43 unvaccinated heifers were used as controls. After vaccination, microscopic agglutination (MA) titres of serums to L. interrogans serovar hardjo ranged from 32 to 512, and those to L. interrogans serovar pomona ranged from 32 to 2048. Titres resulting from vaccination were short-lived and after the first vaccination the serums of 95% of vaccinated heifers did not react in the MA test by 24 weeks. The first indication of infection in the heifers was noted at week 6, and by week 16, elevated MA titres (≥128) to L. interrogans serovar hardjo had occurred in 62% of unvaccinated heifers and had increased to 85% by week 24. At week 18, 18% of the vaccinated heifers and 56% of the unvaccinated heifers had leptospiruria (p<0.01); after 22 weeks, 13% of the vaccinated heifers and 58% of the unvaccinated heifers showed evidence of leptospiruria (p<0.01).     

The prevalence of antibodies to serovars of Leptospira interrogans in horses

SUMMARY Serum samples from 272 horses, some III, were tested by the microscopic agglutination test for the presence of antibody to 12 serovars of Leptospira interrogans. Serums from 41.5% of horses reacted to one or more of the serovars tested; the most common reactions were to L. interrogans serovar ballum (15.1%), L. interrogans serovar autumnalis (11.8%), L. interrogans serovar icterohaemorrhagiae (9.9%), L. interrogans serovar pomona (8.1%) and L. interrogans serovar hardjo (7.7%).     

Haemolytic disease associated with Leptospira Interrogans serovar pomona in red deer calves (Cervus elaphus)

Three red deer calves (Cervus elaphus) died with a haemolytic disease associated with infection by Leptospira interrogans serovar pomona.

Infection within the herd was more prevalent than disease. Sera from 16 herd mates were tested by the microscopic agglutination test (MAT) and 12 had leptospiral titres, the majority to serovar pomona. A few calves had titres to balcanica and hardjo.

Urine was obtained for culture from six of these calves and serovar pomona was isolated from five with titres to pomona, and hardjo from one with a litre to hardjo but not pomona.

A fourth calf died with severe nephritis but a diagnosis of leptospirosis was not confirmed in this case.     

Phage typing of staphylococci isolated from dairy cows in New Zealand

Four red deer calves (Cervus elaphus) died with severe nephritis apparently associated with infection by Leptospira interrogans serovar pomona. The sera of 12 in-contact red deer calves were examined for leptospiral agglutinins and nine showed titres to pomona consistent with recent infection. Two also showed titres of 1:100 to serovar hardjo. The urine of five of these in-contact calves was examined periodically over a period of nine months. All were initially leptospiruric, four being infected with pomona and one with hardjo. In four animals leptospiruria could only be detected for up to six months, but one animal infected with pomona was leptospiruric for at least eight months.

The apparent source of infection was from infected cattle, and it is suggested that deer are unlikely to act as maintenance hosts for serovar pomona.     

Clinical and biochemical changes in sheep inoculated with leptospira interrogans serotype pomona

Clinical and biochemical changes occurring in whole blood, urine, plasma and serum of sheep experimentally infected with Leptospira interrogans serotype pomona were studied.

Half-time and fractional clearance rates of sulphobromophthalein, phenosulphophthalein and chromium (^5lCr) EDTA were also measured to assess functional damage to the liverand kidney.

The results suggested that the biochemical changes occurring following pomona infection were due mainly to haemolysis of erythrocytes and to mild impairment of tubular and glomerular kidney function. No alteration in liver function was observed: Analyses of isocitrate dehydrogenase, arginase, and total bilirubin in serum and measurements of kidney glomerular and tubular function appeared to be the most useful biochemical indicators of pomona infection in sheep.     

Leptospirosa pomona Abortion Storm in a Cattle Herd in Saskatchewan

Abortions occurred in 18% of 131 beef cows and heifers during two months, on a farm in southern Saskatchewan. The losses began two weeks after acute febrile illness and agalactia in a dairy cow to which the beef herd had been exposed. A diagnosis of Leptospira interrogans serovar pomona infection was made on the basis of serology in cows and the finding of leptospires in fetal tissues by fluorescent antibody test. Tentative diagnosis of infectious bovine rhinotracheitis delayed treatment and prophylaxis until infection attained high intensity in the herd and severe losses to the farmer occurred. Abortions ceased after vaccination against pomona and oxytetracycline treatment of pregnant cows, although chronic debility followed the acute phase of the disease in some cows. Recrudescence of infection was suspected four months later, when acute agalactia occurred in one cow and debility in calves and cows was recurring. Pomona infection was not proven, but dihydrostreptomycin treatment and revaccination were applied to the whole herd. Seroconversion and IgM antibody continued to indicate a persistent source of infection and susceptibility in a minority of the population one year after onset. The source of the original infection is believed to have been a carrier beef cow, or a dairy cow which was leptospiruric at the time of contact with the beef herd. With the exception of one aborted calf, no evidence of pomona infection was found outside the farm, in cattle or wild mammals tested serologically within a radius of 30 km, during one year following the outbreak.     

Acute kidney injury due to Leptospira interrogans in 4 foals and use of renal replacement therapy with intermittent hemodiafiltration in 1 foal

Four 2-month-old foals were presented to an equine hospital with acute kidney injury caused by Leptospira interrogans infection. Clinical signs were nonspecific and included lethargy, fever, and unwillingness to nurse. The most important hematologic and clinicopathologic findings were azotemia, anemia, thrombocytopenia, hyponatremia, and hypochloremia. The diagnosis was based on urinary real-time PCR, serology using a microscopic agglutination test, or both. The most important serovars involved were L. interrogans serogroup australis serovar Bratislava and Australis. Treatment consisted of IV fluid therapy and antimicrobial treatment. Renal replacement therapy with hemodiafiltration was performed in 1 of the foals. All foals survived to discharge. This report highlights the importance of early diagnosis and treatment in foals with acute kidney injury caused by L. interrogans infection.     

An unusual serological response in cattle after use of a commercial leptospiral combined hardjo-pomona vaccine

Vaccination of four calves with Leptavoid (Wellcome New Zealand Limited) gave rise to Leptospira interrogans serovars hardjo and pomona microscopic agglutination test titres that could not be distinguished in magnitude from post-infection titres. Vaccination of four calves with Lepto-3 (ICI Tasman Limited) gave rise to much lower titres.

Revaccination of cows with Leptavoid caused a rise in hardjo titres which was significantly greater than after use of Lepto-3.

The possibility that titres were due to the simultaneous infection with serovars pomona and hardjo of only the animals vaccinated with Leptavoid must he discounted.     

Leptospirosis: I. Clinical investigation of the infection in dairy cattle in the Waikato district of New Zealand

An investigation was made into the prevalence of leptospiral infection in cattle. An area 50 km radius was selected in a region where leptospirosis was reputedly common. Farmers volunteered 250 herds with 39 500 cows for testing and 7 500 animals were selected and sampled. Twenty-nine cows (0.4%) on 14 (5.6%) of the farms had leptospiruria at the first examination. Leptospirae were cultured from the urines of nine of these animals and all were Leptospira interrogans serovar hardjo. Serologically 12.5% of cows had titres of 1:200 or greater to hardjo and 3.5% titres of 1:200 or greater to pomona. In the Spring of 1977, there was evidence of clinical leptospirosis in calves associated with only one of the herds and no clinical leptospirosis in the 250 lactating herds, although leptospiral titres were found in 88% of them. This indicated that clinical disease was much less common than infection. We concluded that leptospirosis was of minor economic importance in dairy cattle, although it could be significant in individual herds, and a health hazard to farm workers.     

Leptospira interrogans serovar pomona vaccines with different adjuvants in cattle

Three groups of four heifers were vaccinated twice, 11 weeks apart. One group received a commercial pomona vaccine with aluminium hydroxide adjuvant, the second a similar experimental vaccine, and the third a Freund's compete adjuvant (FCA) vaccine. After 47 weeks, the heifers were challenged with at least 65 × 10⁸ virulent serovar pomona organisms.

All vaccinated animals resisted the challenge, and leptospirae were only found in urine from unvaccinated controls.

The outcome of the challenge was not predictable from microscopic agglutination, cold and warm complement fixation, and growth inhibition titres.

The FCA vaccine gave rise to considerably higher antibody responses than the two aluminium hydroxide vaccines, which gave similar responses.     

RENAL LEPTOSPIROSIS IN VACCINATED PIGS

SUMMARY The ability of 2 commercial leptospirosis vaccines to prevent renal colonisation with Leptospira interrogans serovar pomona was studied in pigs. Based on bacteriological, pathological and serological findings some degree of protection was found in vaccinates when compared with non-vaccinates after challenge 4, 7 and 12 months after vaccination. However, the degree of protection was insufficient to prevent colonisation and renal leptospirosis in 6 of 10 pigs at 4 months, 7 of 10 pigs at 7 months and 8 of 10 pigs at 12 months.     

Serological patterns,antibody half-life and shedding in urine of Leptospira spp. in naturally exposed sheep

Abstract

AIMS: To determine within-farm prevalence, longitudinal pattern of exposure measured by serology, antibody titre longevity and point prevalence of shedding in urine of Leptospira borgpetersenii serovar Hardjo and L. interrogans serovar Pomona in naturally infected sheep on a sample of commercial farms in New Zealand.

METHODS: On eight commercial sheep farms, between September 2011 and January 2014, blood samples were collected from 115–217 ewe lambs on each farm, at intervals of 2–11 months. They were analysed by microscopic agglutination test (MAT) for antibodies to L. borgpetersenii serovar Hardjo and L. interrogans serovar Pomona, using a titre cut-point of 48. Urine from 98 animals was tested by quantitative PCR (qPCR). The half-life of antibodies was estimated in 185 sheep for serovar Hardjo and 21 for Pomona, and the seroprevalence and mean titre of animals lost to follow-up was compared with those remaining in the study.

RESULTS: Within-flock seroprevalence for serovar Hardjo reached a maximum at 17–22 months of age, ranging from 79 to 100%. Seroprevalence for serovar Pomona rose above 10% on three farms and increased to 21–54% by 4–14 months. Seroconversions occurred mainly from late autumn to early summer at 7–15 months of age. Seroprevalences ranging from 3 to 76% for serovar Hardjo and 0.5 to 15% for serovar Pomona were observed up to 3 months of age, likely due to maternally derived immunity. The half-life of antibody in response to infection was estimated to be 6.7 (95% CI=5.8–7.9) months for serovar Hardjo and 6.3 (95% CI=4.8–9.0) months for Pomona. The prevalence of sheep with urine positive for leptospires on qPCR on each farm ranged from 11 to 88%. All but one of the qPCR-positive animals were seropositive for serovar Hardjo. On two farms where Pomona exposure was observed, animals that were lost to follow-up had a higher geometric mean titre for serovar Pomona than those remaining in the study.

CONCLUSIONS: This study demonstrated seasonal exposure from autumn to early summer in young sheep, a wide range of within-flock serological and shedding prevalence, and gives an estimation of the half-life of MAT titres in sheep. More extensive data are needed to fully understand the epidemiology of leptospirosis in sheep flocks across New Zealand and, along with economic analysis, to justify and design cost-effective and efficient control measures to protect human and animal health.     

Protection of cattle against natural challenge with Leptospira interrogans serovar hardjo using a hardjo-pomona vaccine

The hardjo component of a bivalent hardjo/pomona vaccine prepared from cultures grown in a protein-free medium was tested in calves which experienced a natural challenge about 7 months after vaccination. A significant degree of protection was obtained using either seroconversion or positive urine cultures as in-dicators of infection. Seroconversion occurred in 10/10 non-vaccinates and 2/9 vaccinates (p <0.01), Leptospires of serovar hardjo were isolated from the urine of 6/10 non-vaccinated calves and 0/9 vaccinated animals (p <0.05).     

Leptospirosis in Colombia: Isolation ofLeptospira SPP. from the kidneys of brown rats (Rattus Norvegicus) trapped on infected piggeries

Summary Leptospira interrogansserotypes pomonaand icterohaemorrhagiaewere isolated from the kidneys of Rattus norvegicustrapped on pomonainfected piggeries.The isolation of pomonafrom one of 111cultured kidneys, although of interest, strongly suggests that this rat does not play an important part in the epidemiology of infection by this serotype. Kidney tubular degeneration, focal mononuclear infiltrations and fibroplasia are morphological traces which remain after an attack of leptospirosis. The high percentage (48.6)of these lesions found in the rat kidneys examined does not seem to be related to serotype pomonabut to icterohaemorrhagiae.The importance of this last serotype in swine in the Cauca Valley has yet to be assessed.

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Leptospirosis En Colombia: Aislamiento DeLeptospira SPP. De Riñones De Ratas Pardas (Rattus Norvegicus) Atrapadas En Porquerizas Infectadas

Resumen Se aislóLeptospira interrogans serotipospomona eicterohaemorrhagiae de riñones deRattus norvegicus atrapadas en porquerizas infectadas porpomona. El aislamiento depomona de 1/111 riñones cultivados, aunque de interés, sugiere que esta rata no desempeña parte importante en la epidemiología de las infecciones por este serotipo.Degeneración tubular renal, infiltración mononuclear focal y fibroplasia son huellas morfológicas que quedan después de un ataque de leptospirosis. El alto porcentaje (48.6) de estas lesiones encontradas en los riñones de las ratas examinadas no parece estar relacionado con el serotipopomona sino con elicterohaemorrhagiae. La importancia de este último seroptipo en cerdos en el Valle del Cauca no se ha evaluado todavía.

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Leptospiroses En Colombie: Isolement DeLeptospira SPP. De Reins De Rats Bruns (Rattus Norvegicus) Captures Dans Des Porcheries

Résumé Les sérotypespomona eticterohaemorrhagiae deLeptospira interrogans ont été isolés de reins deRattus norvegicus capturés dans des porcheries infectées de (pomona?). L'isolement depomona sur une seule des 111 cultures de rein effectuées, bien qu'interessant, suggère fortement que ce rat ne joue pas un rôle important dans l'épidémiologie de l'infection par ce sérotype.La dégénerescence tubulaire des reins, des infiltrations ponctuelles par des monocucléaires et l'aspect fibreux constituent les traces morphologiques qui demeurent après une attaque de leptospirose. Le pourcentage élevé (48,6) de ces lésions trouvées dans les les reins des rats examinés semble être plus en rapport avec le sérotypeicterohaemorrhagiae qu'avec le sérotypepomona. L'importance du sérotypeicterohaemorrhagiae chez le porc de la Cauca Valley est encore à vérifier.

     

Leptospirosis outbreak in a piggery in southern alberta

Fourteen abortions, stillbirths and neonatal deaths occurred over an interval of one month in crated sows in a farrow-to-finish swine operation. Leptospira interrogans serovar pomona (kennewicki by DNA analysis) was demonstrated by culture and fluorescent antibody test to be present in sows, piglets, boars, feeders, drinking water and skunks on the premises.

Antibody was found in all clinically affected sows, at serum dilutions ranging from 1/800 to 1/25,600, and in all breeding boars at titers from 1/50 to 1/1600. Pomona antibody was present in 118 sow sera collected nine months before the outbreak, at a prevalence of 21.3%. Parvovirus infection in fetuses was intercurrent with the leptospirosis epizootic, despite vaccination for the former. Environmental contamination, feedback through skunks via drinking water, and dissemination through the piggery are discussed.

     

SEROLOGICAL TITRES OF DAIRY COWS OVER A 63-WEEK PERIOD FOLLOWING NATURAL INFECTION WITH LEPTOSPIRA INTERROGANS SEROVAR HARDJO

SUMMARY Leptospirosis due to Leptospira interrogans serovar hardjo, occurred in a dairy herd of 104 cows and produced signs of mastitis, characterised by a sudden decrease in milk production and uniformly flaccid udders, in 11 cows. The microscopic agglutination (MA) test was used to determine antibody titres to serovar hardjo in all cows in the herd at 6, 33 and 63 weeks after the initial outbreak. The prevalence of MA antibodies to hardjo at week 6 was highest in cows in the youngest age groups and lowest in cows in the oldest age group. Over the 63-week period MA antibody titres to hardjo declined in 54 out of 62 seropositive cows, remained unchanged in 6 cows, and increased slightly in 2 cows. Fourteen of 59 cows (23.7%) with MA titres greater than 100 at week 6 had titres greater than 100 by week 63. Included in this group of 14 were 5 of 7 cows that had been affected with atypical mastitis. The distribution of serological titres to hardjo showed bimodal configurations at weeks 6 and 33 with the second peaks occurring at titres greater than 100, while the configuration was linear at week 63. MA titres to hardjo were also determined for 22 heifers 1, 16 and 36 weeks after they were added to the herd. No clinical signs of leptospirosis were observed in the heifers; however, the distribution of MA titres showed bimodal configurations at weeks 16 and 33 with second peaks occurring at titres greater than 100. Leptospiras were isolated from the urine of 2 seropositive heifers 16 weeks after their introduction to the herd, and cross-agglutinin absorption tests performed on one of the isolates indicated that it was identical to serovar hardjo.     

A Serological Survey of Leptospira interrogans serotype pomona in Alberta and Saskatchewan Striped Skunks and Possible Transmission Between Cattle and Skunks

The range of known occurrence of Leptospira interrogans serotype pomona is extended to Alberta in striped skunks (Mephitis mephitis); no evidence of L. sejroe was found. Reacting sera from skunks were confined to the southern portion of Alberta and adjacent Saskatchewan, although a number of reactors were found sufficiently further north in Saskatchewan suggesting that a different mode of infection may be functioning there. Of 95 skunk sera from near a dairy farm infected with serotype pomona 40% were reactors. Of 438 skunk sera from other areas only 5.7% were reactors; that difference was suggestive of transmission from cattle to skunks on the dairy farm. Of 22 skunk sera collected near the dairy farm in summer none were reactors, whereas 52% of skunk sera taken the previous and following winters were. That seasonal difference was not evident among sera from other locations.     

Experimental myrotheciotoxicosis in sheep and calves

Hamster inoculation, medium inoculation and microscopical methods were used for the detection of Leptospira interrogans serovar pomona organisms in bovine urine. Urine samples were collected from both naturally and experimentally infected animals during the period when leptospiruria could be expected.

Inoculation into hamsters of 0.5 ml of urine from each of 14 samples resulted in 5 positives. The inoculation of 0.025 ml of the same sample into semi-solid EMJH medium gave 10 positives. From a cumulative total of 46 urine samples, 28 were positive using dark-ground microscopy of centrifuged urine deposits and 35 were positive using media inoculation. A cumulative total of 126 urine samples were examined, after centrifugation, under dark-ground microscopy and leptospirae were detected in 60.

Fluorouracil (100 mg/ml) proved to be beneficial for successful isolations in media, whereas neomycin (5 mg/ml) did not.     

LEPTOSPIRA INTERROGANS SEROVAR HARDJO VACCINATION OF PREGNANT BEEF COWS AND SUBSEQUENT GROWTH RATE OF PROGENY

SUMMARY Five experiments with Leptospira interrogans serovar hardjo vaccine were carried out over a 6-year period in pregnant Brahman-cross and Sahiwal-cross cows in the dry tropics of northern Queensland. The numbers ranged from 127 breeders aged 2 to 5 years in 1972 to 344 breeders aged 2 to 9 years in 1977. Half of the cows were vaccinated twice in midpregnancy except for 1977, when they were vaccinated once. In 1975–1977 inclusive, half of the heifers were given an additional dose of vaccine at commencement of mating. Vaccination caused a significant (P<0.01) reduction of prenatal loss but not of perinatal or postnatal losses. Growth rates of calves from vaccinated and unvaccinated dams were similar.     

Serological survey of pre-weaned New Zealand fur seals (Arctocephalus forsteri) for brucellosis and leptospirosis

AIM: To conduct a longitudinal serological survey for evidence of Brucella spp and Leptospira spp infection of pre-weaned New Zealand fur seals in a colony on the Otago Peninsula.

METHODS: Seal pups were repeatedly captured on a monthly basis from February through to July 2001. Pups were tagged at first capture and a blood sample was taken at each capture event. A total of 163 sera were collected from 118 seal pups. Where sufficient volume was collected, the sera were tested for leptospirosis using the microscopic agglutination test (MAT), and for brucellosis using the competitive enzyme-linked immunosorbent assay (ELISA) for Brucella abortus.

RESULTS: None of 128 sera from 101 seals tested positive to the ELISA for B. abortus. All tests for Leptospira interrogans serovars Grippotyphosa, Copenhageni, Bratislava and Leptospira borgpetersenii serovar Ballum were negative at a cut-off of <1/100 dilution. Positive or suspicious titres were found to L. interrogans serovars Canicola and Pomona and L. borgpetersenii serovar Hardjo. The highest titres (12,800) were found to serovar Pomona. The titre to serovar Pomona in one seal rose from <1/50 in March to 12,800 in April and was <1/50 when re-sampled in July. The titre to serovar Pomona in another seal dropped from 12,800 in May to <1/50 in June. These seals also had titres to serovar Hardjo, which rose or fell in the same manner. All suspicious or positive titres occurred in late April and early May, when the pups were approximately 4–5 months old. In June and July, all seals tested were negative.

CONCLUSIONS: There was no serological evidence of Brucella infection in the pre-weaned fur seals at the colony. Positive titres to serovars Pomona, Hardjo, or Canicola suggest that a Leptospira species was present at the colony, however isolation or visualisation of the organism is required to confirm this. Care should be exercised when handling New Zealand fur seals to prevent human infection or inadvertent transfer of leptospirosis to another marine mammal species.