Erratum to "MAT cross-reactions or vaccine cross-protection: retrospective study of 863 leptospirosis canine cases" [Heliyon 4 (11) (November 2018) e00869].

[This corrects the article DOI: 10.1016/j.heliyon.2018.e00869.].

MAT cross-reactions or vaccine cross-protection: retrospective study of 863 leptospirosis canine cases.

Dogs are naturally exposed to numerous pathogenic serogroups. Leptospirosis vaccines are claimed to afford a clinical protection restricted to the serogroups of which they are composed. OBJECTIVES: Dogs exhibiting liver and kidney injury were suspected of having leptospirosis. The purpose of this study was to compare the microscopic agglutination test (MAT) results in naive and vaccinated dogs experiencing leptospirosis outcomes. Only MAT-positive animals were included in the study. METHODS: Over five years, 3 512 dogs were suspected of having leptospirosis. For each case, biochemical parameter results were recorded. Leptospirosis involvement was investigated by MAT performed against 6 major serogroups (Icterohaemorrhagiae, Canicola, Australis, Autumnalis, Grippotyphosa and Sejroë). MAT-positive results confirmed leptospirosis cases in 147 naïve dogs and in 580 fully vaccinated dogs. Serological titres of agglutinating antibodies were related to the severity of liver and kidney failure. RESULTS: The most prevalent outcome of leptospirosis in unvaccinated dogs was liver failure (57.8%) compared to 51.7% for kidney disease, but the most severe onset (90.8%) was found among the cases of acute kidney injury compared to the severe (42.3%) hepatitis cases. In dogs vaccinated by bivalent Icterohaemorrhagiae and Canicola bacterins, hepatitis decreased from 57.8 to 46.5% and acute kidney injury from 51.7 to 21.6%. The decrease was shown in leptospirosis cases induced by field strains belonging to the six most prevalent serogroups, including the 4 serogroups heterologous to the vaccine. CONCLUSION: Common vaccination was efficient in decreasing hepatitis and kidney failure induced by field Leptospira spp infection regardless of the MAT-prominent serogroup and limited the disease severity in the remaining cases.

Waterborne Leptospirosis: Survival and Preservation of the Virulence of Pathogenic Leptospira spp. in Fresh Water.

Many studies have implicated fresh water as a source of leptospirosis outbreaks. To estimate the survival and the preservation of the virulence of pathogenic Leptospira spp. maintained in water, we selected five still waters with various pH and mineral profiles. The water samples were artificially inoculated with a culture of a pathogenic strain belonging to serovar Icterohaemorrhagiae. Samples were stored for 20 months at 4, 20 or 30 °C. The survival and preservation of virulence of this pathogenic strain was estimated by subculturing these stored samples. After 14 and 20 months of storage, the strain Icterohaemorrhagiae was re-isolated, and its virulence was determined using an animal model. In these waters, the mean survival was 130 days for storage at 4 °C, 263 days at 20 °C, and 316 days at 30 °C. Unexpectedly, the mean survival was 344 days for a final pH < 7 and 129 days for pH ≥ 7. Moreover, the pathogenic strain remained fully virulent and was able to induce a lethal disease in gerbils even when the pH of the contaminated waters decreased to <6. These data showed that despite unfavourable storage conditions such as cold, nutrient-poor acidic waters, the survival and virulence of pathogenic Leptospira spp. was fully preserved over at least 20 months.

Undiagnosed leptospirosis cases in naïve and vaccinated dogs: properties of a serological test based on a synthetic peptide derived from Hap1/LipL32 (residues 154-178).

Leptospirosis is a common disease in dogs, despite having current vaccinations. However, leptospirosis diagnosis based on the routine Microscopic Agglutination Test (MAT) leads to confusing conclusions, especially for infected vaccinated dogs. Indeed, both bacterin and natural infection stimulate the production of agglutinating antibodies. In experimentally infected dogs, antibodies against the peptide PP derived from Hap1/Lipl32 were raised earlier than agglutinating antibodies. The background level of these antibodies was determined in a group of 109 healthy dogs, either vaccinated or not against leptospirosis, with a specificity for IgM of 96.4% and for IgG of 95.5%. PP ELISA was subsequently performed with 118 sera from dogs with suspected leptospirosis that was not confirmed by MAT. New leptospirosis cases based on the PP ELISA results were suspected in 14 out of 102 vaccinated dogs and in two out of 16 non-vaccinated dogs. These results highlight the importance of serological diagnosis corresponding to an interesting window when it is too late for PCR detection and too early to be confirmed by MAT.

Serological survey of canine leptospirosis in three countries of tropical Africa: Sudan, Gabon and Ivory Coast.

A cross-sectional survey was conducted to estimate the presence of leptospiral antibodies among 475 dogs from three countries of tropical Africa: Sudan (n=62), Gabon (n=255) and Ivory Coast (n=158). Sixteen reference strains belonging to seven serogroups were used as antigen in the microscopic agglutination test. Overall, considering titres ≥1:40, 453 samples were positive towards one or several serovars of pathogenic leptospires. Focusing on high titres, i.e. ≥1:320, the seroprevalence was 40.8%. In Gabon, the seroprevalence was higher in rural areas than in an urban environment (p<0.001). In Ivory Coast, the seroprevalence for serogroups Icterohaemorrhagiae and Canicola was not statistically different according to the vaccinal status. Predominant serogroups varied according to the countries but Grippotyphosa and Sejroë were the most common, while Icterohaemorragiae and Canicola were dominant in Sudan. In these three countries, dogs are heavily exposed to pathogenic Leptospira and humans living in the same environment are also at risk of infection.

Development and validation of an enzyme-linked immunosorbent assay for the detection of circulating antibodies raised against growth hormone as a consequence of rbST treatment in cows.

The recombinant bovine growth hormone (rbST) is used to increase lactating performances of dairy cows. Administration of rbST is banned in the European Union; nevertheless, its use is probable. Until now, efficient analytical strategies to detect such practice are based on the direct detection by mass spectrometry of the presence of rbST in biological fluids, which suits for confirmatory purposes. Current screening strategies do not offer satisfactory performances; therefore, alternative screening strategies are required. The aim of the present work is to develop and validate an ELISA to measure the production of specific antibodies upon rbST in bovine sera. In this immunoassay, rbST is absorbed onto microtiter plate. After specific purification of the antibodies in serum, samples are analysed and the presence of antibodies anti-rbST is detected by Protein G peroxidase conjugate and 2-2'-azino di-ethyl benz-thiazoline-6-sulphonic acid (ABTS). The mean reproducibility of the OD (λ=405 nm) measurement was calculated with a CV of 13%. The intra- and inter-assay CVs ranged from 0.79% to 7.91% and from 2.69% to 20% respectively. The test presents cross-reaction with other growth hormones such as the recombinant equine (reST) and porcine (pST) (100% and 80% respectively). The specificity of the test toward rbST anabolic treatment was confirmed through the analysis of sera samples collected on animals administered with other anabolic compounds (steroids). The performances of the present anti-rbST ELISA proves its efficiency as a new screening tool to highlight illegal administration of rbST in cattle up to at least 3 weeks after treatment.

Leptospirosis in free-ranging endangered European mink (Mustela lutreola) and other small carnivores (Mustelidae, Viverridae) from southwestern France.

To study the possible role of disease in the decline of endangered European mink (Mustela lutreola), we conducted a survey of antibody prevalence and renal carriage of pathogenic leptospira (Leptospira interrogans sensu lato) using serum and kidney samples collected from 1990 to 2007 from several free-ranging small carnivores and farmed American mink (Mustela vison) in southwestern France. An indirect microscopic agglutination test using a panel of 16 serovars belonging to 6 serogroups (Australis, Autumnalis, Icterohæmorrhagiæ, Grippotyphosa, Panama, Sejroe) revealed antibodies in all species, with significant differences in antibody prevalences: 74% in European mink (n=99), 65.4% in European polecats (Mustela putorius, n=133), 86% in American mink (n=74), 89% in stone martens (Martes foina, n=19), 74% in pine martens (Martes martes, n=19), 35% in common genets (Genetta genetta, n=79), and 31% in farmed American mink (n=51). Serogroups Australis and Icterohæmorragiæ were dominant in most free-ranging species; serogroup Grippotyphosa had high prevalences in European mink. Such high antibody prevalences have never been reported. They are probably related to the large number of known reservoirs, rats (Rattus spp.), muskrat (Ondatra zibethicus), and coypu (Myocastor coypu), in the study area. The polymerase chain reaction test specific for pathogenic leptospiral DNA detected renal carriage in 23% of 34 European mink, 22% of 18 polecats, and 15% of 33 free-ranging American mink, with no significant differences. Renal carriage shows that mustelids may shed leptospira for short periods, but their epidemiologic role is probably limited. High antibody prevalences suggest that the disease is unlikely to be highly pathogenic for these species; however, chronic forms of the disease (abortions, renal lesions) could reduce the reproductive success or life span of infected animals. Further studies on the pathogenicity of leptospirosis in these populations are needed to measure its impact on the population dynamics of these rodent predators.

Multilocus sequence analysis for typing Leptospira interrogans and Leptospira kirschneri.

Fifty-three strains belonging to the pathogenic species Leptospira interrogans and Leptospira kirschneri were analyzed by multilocus sequence analysis. The species formed two distinct branches. In the L. interrogans branch, the phylogenetic tree clustered the strains into three subgroups. Genogroups and serogroups were superimposed but not strictly.

Detection of secondary biomarker of met-eGH as a strategy to screen for somatotropin misuse in horseracing.

Since the Australian commercialisation of the recombinant equine growth hormone (reGH) in 1998 (EquiGen-5), Bresagen), this reGH, which differs only from eGH by an additional methionine at the N-terminal end (met-eGH), is worldwide suspected to be administered to racehorses as a doping agent. Indeed, the use of this biological drug is considered as a threat to horseracing since it acts both on growth, development or reproductive functions, and on the improvement of performances. In this work, we describe two reliable techniques based on surface plasmon resonance biosensor immunoassay (SPR-BIA) and solid-phase enzyme-linked immunosorbent assay (ELISA) as new, rapid and efficient long-term screening methods applicable to horseracing antidoping analysis. The ELISA and SPR-BIA tests were applied to octanoic acid purified IgGs from serum/plasma samples collected on two thoroughbreds treated with recombinant equine growth hormone for a period of two weeks. The first kinetic study of serum/plasma antibodies raised as a consequence of recombinant equine growth hormone administrations, which allows the detection from eight days up to 200 days after the beginning of the treatment, was performed. In order to trace the occurrence of anti-reGH antibodies in routine analysis and to monitor the animal level exposure to this forbidden molecule, a random population study was conducted on 233 post-race horses.

Canine leptospirosis--do we have a problem?

Acute canine leptospirosis is well known to vet surgeons. To protect dogs against this lethal disease, vaccination is widely used. However, chronic forms of leptospirosis have been noticed even in vaccinated animals, generally induced by bacteria from serogroups other than Icterohaemorrhagiae and Canicola such as Sejroë, Australis or Grippotyphosa. In a survey on 98 ill cats, 48% were positive in microagglutination test (MAT) to Leptospira spp., showing that this infection is also frequent in the feline species.

Identification of pathogenic Leptospira strains in tissues of a premature foal by use of polymerase chain reaction analysis.

Studies were carried out to determine the cause of death in a prematurely born Thoroughbred foal that died 24 hours after birth. Necropsy revealed gross lesions suggestive of septicemia. A commercial Leptospira polymerase chain reaction (PCR) assay designed to specifically amplify the hemolysis-associated protein 1 (hap1) gene present only in pathogenic Leptospira strains detected the presence of Leptospira DNA in various tissues of the foal. Histologic examination of lung, liver, kidney, and myocardium revealed numerous spirochetes in Warthin-Starry-stained tissue sections. Results of PCR analysis and histologic examination suggested a leptospiral infection in the newborn foal. At the moment of death, the infection coexisted with a streptococcal-associated aspiration bronchopneumonia and postpartum septicemia. These findings indicate that the PCR assay based on the amplification of the hap1 gene represents a useful tool for specific detection of pathogenic leptospira in field samples taken from horses.

Polymerase chain reaction assay specific for pathogenic Leptospira based on the gene hap1 encoding the hemolysis-associated protein-1.

In this study, we used Southern hybridization of genomic DNA with the integral hap1 gene as a probe to show that this gene is only present in pathogenic Leptospira strains. We then selected PCR primers based on the hap1 gene, and tested them on several Leptospira strains and biological samples. Specific amplification was obtained for all pathogenic strains tested. Negative PCR results were observed with all saprophytic leptospire strains used as well as with other spirochetes and bacteria commonly found in biological samples. The results of direct PCR performed on biological samples, such as blood, urine or kidneys correlated with the results obtained with the classical Leptospira tests (culture and MAT). A PCR assay based on this gene would be a very useful tool for the rapid, sensitive and specific identification of pathogenic leptospires in samples for diagnosis or epidemiological survey.