Abortion In Small Ruminants Research Articles

Search for Possible Additional Reservoirs for Human Q Fever, the Netherlands

To the Editor: Q fever is a zoonosis caused by the bacterium Coxiella burnetii. The Q fever outbreak in the Netherlands affected ≈4,000 humans during 2007–2010 (1). In this outbreak, 1 genotype of C. burnetii appeared to be responsible for abortions in small ruminants and for clinical disease in humans (2,3). However, little is known about the outbreak genotype and the prevalence of C. burnetii in possible additional reservoirs for human Q fever (i.e., cats, dogs, horses, sheep, and cattle) in the Netherlands. We aimed to search for possible additional reservoirs for human Q fever in the Netherlands. Placentas from 15 cats, 54 dogs, and 31 horses were collected in 2011 at 5 veterinary practices. Placentas were collected by targeted sampling at breeding facilities and during parturition with veterinary assistance. In addition, 27 ovine, 11 caprine, 16 porcine, 8 equine, and 139 bovine placentas (originating from aborting animals from throughout the Netherlands that were submitted in 2011 to investigate the abortion cause) were included in the study. Samples were stored at −20°C before testing. DNA was extracted from the allantochorion of the placenta and analyzed as described (2). Samples with sufficient DNA load (cycle threshold [Ct] value <32) were typed by using 2 multilocus variable-number tandem-repeat analyses (MLVA) genotyping methods (MLVA-12 and MLVA-6), and the multispacer sequence typing method (3–5). Two C. burnetii strains from the Netherlands representing the outbreak genotype (X09003262, 3345937) and the Nine Mile RSA 493 were included as reference. For prevalence calculations, the Netherlands was divided in a southern part, comprising the Q fever hot spot area of notified cases in humans and small ruminants during the 2007–2010 epidemic (1,6), and a northern part, comprising the rest of the country. C. burnetii DNA was not detected in placentas from cats, goats, or pigs. C. burnetii DNA was detected in 4 (7% [95% CI 0.4–14.4]) of 54 canine placentas; 3 from the north and 1 from the south of the Netherlands. C. burnetii DNA was detected in 3 (8% [95% CI 0.0–16.1]) of 39 equine placentas, all from the north of the country, without known abortion history. C. burnetii DNA was detected in 7 (26% [95% CI 9.4–42.5]) of 27 ovine and in 33 (24% [95% CI 16.7–30.8]) of 139 bovine placentas. The prevalence of C. burnetii DNA–positive ovine and bovine placentas from the north and the south did not differ significantly. The C. burnetii DNA load in the placentas from dogs (Ct value 37.4–38.0) and horses (Ct value 35.4–37.4) was too low to be suitable for genotyping. Typing of 1 positive sheep sample resulted in an incomplete genotype, which is related to the outbreak genotype (sheep 192, Figure). Seven of the 33 C. burnetii DNA–positive bovine placentas were suitable for typing. One sample had a genotype similar to the outbreak genotype (2,3). Six other samples revealed a (partial) genotype related to bovine genotypes from the Netherlands (2,5,7), including a novel one. MLVA-6 and multispacer sequence typing results were consistent with the MLVA-12 results (Figure). Figure Phylogenetic tree of the genotypes of Coxiella burnetii from the samples of this study based on multilocus variable-number tandem-repeat analyses (MLVA) including 12 loci (MLVA-12). Repeats per locus are shown, and open spots indicate missing values. ... Results give no indication for major reservoirs of C. burnetii in cats, goats, and pigs in the Netherlands in 2011. However, the low numbers of placentas may have biased the results. Dogs and horses should be considered as reservoirs for C. burnetii. The detection of C. burnetii DNA–positive placentas in dogs and horses in the northern part of the country indicates the presence of a true reservoir rather than a spillover effect from the contaminated environment in the south. This observation is consistent with a reported seroprevalence of 13% in dogs in the Netherlands in 1992 (1). Until now, horses had been discussed as a risk factor in the Q fever outbreak in the Netherlands (8). Prevalence data from sheep and cattle suggest that C. burnetii is present in placentas in 25% of the abortion cases in these species. Presence of the outbreak genotype of C. burnetii in sheep has been observed (2,5), indicating sheep are a reservoir for Q fever in humans. Genotyping data show a distinct genotype in 6 of the 7 cattle samples in accordance with previous work (2,5,7). However, the outbreak genotype was detected in 1 sample from a cow. Whether this is an incidental finding or the first observation of the outbreak genotype being transferred to the cattle population is not clear. If the latter, exposure to cattle also possibly might become a risk factor for human Q fever, in addition to goats and sheep.

Sinais clínicos e ocorrência de anticorpos anti-Chlamydophila abortus em ovinos de São Paulo e Minas Gerais

A Chlamydophila abortusi, anteriormente conhecida como Chlamydia psittaci sovovar 1, é uma bactéria Gram negativa, intracelular obrigatória. Esse micro-organismo é frequentemente encontrado em distúrbios reprodutivos em ovinos, bovinos e caprinos, sendo o aborto epizoótico dos bovinos e o aborto enzoótico dos ovinos e caprinos as manifestações mais importantes. Considerando-se o pouco material literário a respeito da clamidofilose no Brasil, a pesquisa teve como objetivo determinar a presença de anticorpos fixadores de complemento anti-Chlamydophila abortusi, correlacionando os resultados obtidos com achados no exame clínico e histórico dos animais, além de alterações nos índices zootécnicos, em especial na esfera reprodutiva, tais como alto índice de repetição de cio, número elevado de abortamentos, elevado número de natimortos, entre outros. Foram testadas para prova de fixação do complemento 220 amostras de soro de ovinos, de 26 propriedades, distribuídas em 19 municípios, com relato de manifestação reprodutiva, obtendo-se 19,55% (43/220) de testes positivos para Chlamydophila abortusi, com ocorrência de foco constatada de 61,53%. No geral, a titulação de anticorpos encontrada foi baixa, com título não superior a 64. A frequência de manifestação reprodutiva mais observada foi o aborto, representando 65,12% (28/43) do número total de animais soropositivos, seguido de repetição de cio juntamente com nascimento de cordeiro fraco, com frequência de 6,98% (3/43) e, por fim, morte neonatal com 4,65% (2/43), sendo que não houve associação significativa entre animais que foram positivos ao teste e a esses fatores.

Vaccination programs for reproductive disorders of small ruminants

Vaccines are available for the control of contagious epididymitis and abortion in small ruminants, although many of them have significant limitations either in efficacy or safety to both the animals vaccinated and to the people handling the vaccine or animals. Shelf-life of vaccines should be extended and improved, so that the vaccine remains effective with longer term storage and ideally without refrigeration, so that use in under-developed rural areas is not restricted (e.g., Brucella melitensis, Toxoplasma gondii). The vaccines should not be dangerous for veterinarians or producers to handle (again as examples, B. melitensis, T. gondii). The vaccines should prevent shedding of the organism, in order to prevent spread of the disease causal agent through the sale of vaccinated but shedding animals (e.g., inactivated killed Chlamydophila abortus vaccines), as well as to prevent possible exposure to people handling those vaccinated animals. Production of vaccines using zoonotic disease agents is problematic and sometime dangerous, which increases regulatory restrictions and reduces availability of those vaccines (e.g., C. abortus, Coxiella burnetii). Development of subunit recombinant DNA vaccines may offer a method to increase access to these important vaccines, as long as they are also effective, prevent shedding and remain cost effective. It is important that these vaccines are brought to international commercial production. As many of these disease agents are zoonotic and prevalent world-wide, improvement in vaccine efficacy and safety is of extreme importance.

Occurrence of Neospora caninum and Toxoplasma gondii infections in ovine and caprine abortions

Neospora caninum and Toxoplasma gondii are closely related cyst-forming apicomplexan parasites identified as important causes of reproductive failure in cattle and small ruminants, respectively. Protozoan abortion in small ruminants is traditionally associated with T. gondii, but the importance of N. caninum remains uncertain. The aim of this study was to investigate the presence of N. caninum and T. gondii infections in abortion cases in small ruminants submitted for diagnosis. For this purpose, 74 ovine and 26 caprine aborted foetuses were recovered from different areas in Spain. Foetal histopathology was used to detect the presence of protozoal-associated lesions in brain. The presence of N. caninum and T. gondii was confirmed by PCR. Protozoal infection was detected in 17 out of 100 (17%) foetuses examined by at least one of the diagnostic techniques used. Lesions suggestive of protozoal infection were observed in 10.8% (8/74) and 15.4% (4/26) of the ovine and caprine abortions respectively. N. caninum and T. gondii infection was detected by PCR in 6.8% (5/74) and 5.4% (4/74) of sheep foetuses, respectively, of which five showed protozoal-associated lesions. N. caninum DNA was detected in 11.5% (3/26) of goat foetuses, of which two showed protozoal-associated lesions, whereas T. gondii DNA was detected in one goat foetus with no lesions. The simultaneous presence of N. caninum and T. gondii DNA was detected in one sheep foetus with severe lesions. This study demonstrates that N. caninum plays a significant role in abortion in small ruminants in the studied population. In addition, our results highlight the importance of differentiating between protozoa whenever characteristic lesions are observed.

The liposoluble proteome of Mycoplasma agalactiae: an insight into the minimal protein complement of a bacterial membrane.

BackgroundMycoplasmas are the simplest bacteria capable of autonomous replication. Their evolution proceeded from gram-positive bacteria, with the loss of many biosynthetic pathways and of the cell wall. In this work, the liposoluble protein complement of Mycoplasma agalactiae, a minimal bacterial pathogen causing mastitis, polyarthritis, keratoconjunctivitis, and abortion in small ruminants, was subjected to systematic characterization in order to gain insights into its membrane proteome composition.ResultsThe selective enrichment for M. agalactiae PG2T liposoluble proteins was accomplished by means of Triton X-114 fractionation. Liposoluble proteins were subjected to 2-D PAGE-MS, leading to the identification of 40 unique proteins and to the generation of a reference 2D map of the M. agalactiae liposoluble proteome. Liposoluble proteins from the type strain PG2 and two field isolates were then compared by means of 2D DIGE, revealing reproducible differences in protein expression among isolates. An in-depth analysis was then performed by GeLC-MS/MS in order to achieve a higher coverage of the liposoluble proteome. Using this approach, a total of 194 unique proteins were identified, corresponding to 26% of all M. agalactiae PG2T genes. A gene ontology analysis and classification for localization and function was also carried out on all protein identifications. Interestingly, the 11.5% of expressed membrane proteins derived from putative horizontal gene transfer events.ConclusionsThis study led to the in-depth systematic characterization of the M. agalactiae liposoluble protein component, providing useful insights into its membrane organization.

Parachlamydia acanthamoebaeInfection and Abortion in Small Ruminants

<i>Parachlamydia acanthamoebae</i>Infection and Abortion in Small Ruminants

Detection of Chlamydophila abortus in Sheep (Ovis aries) in Mexico

Chlamydophila abortus is one of the pathogens which induce abortion in small ruminants; this pathogen has a tropism for ruminant placenta and causes the disease commonly referred to as Ovine Enzootic Abortion (OEA). In Europe are estimated economic losses of around 20 million pounds a year by OEA. In the American Continent the disease has been reported only in Canada, the United States, Colombia and Chile while in Mexico it is unknown whether OEA is common and it is causing abortions in flocks of sheep from Estado de Mexico. The objective of this study was investigating the prevalence of anti-Chlamydophila abortus IgG antibodies and detection of C. abortus DNA in sheep with clinical abort history by mean of ELISA assay (C. abortus ELISA, Institute Pourquier, Montpellier, France) and molecular identification of the principal outer membrane protein (POMP 90-91B) gene by PCR, respectively. A cross-sectional study was carried out to enroll and random sample of ewes from november 2003 until march 2005. A total of 349 sera and vaginal swabs samples were collected from 35 flocks of sheep from Xalatlaco. The results showed that the seropositive rate was 31.1% (14/45) for healthy and 21.3% (65/304) for sheep with history clinical of abort. In vaginal swabs, the PCR showed 0% (0/45) for healthy animals and 0.65% (2/304) for aborted sheep. Samples from the lungs and liver of the fetus of one of these animals were also positive for C. abortus. In conclusion, these results confirmed that infection with C. abortus is common and is affecting sheep flocks in the Mexican highlands. Therefore, is necessary that the authorities responsible for animal welfare in Mexico (SAGARPA) to set up appropriate epidemiological surveillance and control programs to eradicate this disease.

Development and production of vaccines against abortion caused by Chlamydophila abortus and Coxiella burnetii in small ruminants

Chlamydophila abortus and Coxiella burnetii are obligate intracellular parasites, small Gram-negative bacteria, which are aetiological agents of abortion in small ruminants. The organisms also have zoonotic potential, which highlights the importance of the prevention of the transmission of these diseases. Treatment of infected animals with oxytetracycline and its derivatives, can limit abortions, but does not suppress the bacterial excretion, therefore the only effective solution against the infection is the prevention by vaccination. To produce and make available such vaccines, is a long process from the emergence of a concept, through feasibility, development, production, control until obtaining marketing authorisation, and it can take 5–10 years or more. After registration, pharmacovigilance follows the vaccines on the field. C. burnetii and Cp. abortus present serious health risk to humans (Class II and III micro-organisms), thus the propagation of living bacteria is performed in confined Class III areas. Our vaccine strain against chlamydiosis is Cp. abortus 1B ts, whilst the strain of vaccine developed against coxiellosis is C. burnetii Nine Mile phase I. We present hereby the advantages of these vaccines produced with the strains mentioned above over other, existing vaccines and describe their major phases of production.

Influence of the Th2 immune response established by Nippostrongylus brasiliensis infection on the protection offered by different vaccines against Chlamydophila abortus infection

Chlamydophila abortus is the aetiological agent of enzootic abortion in small ruminants in which it infects the placenta to cause abortion during the last trimester of gestation. In a mouse model, a Th1 immune response involving IFN-gamma production and CD8+ T cells is necessary for the infection to be resolved. The authors previously demonstrated that infection with Nippostrongylus brasiliensis, a rodent gastrointestinal nematode extensively used in experimental models to induce Th2 responses, alters the specific immune response against C. abortus infection, increasing bacterial multiplication in liver and reducing specific IFN-gamma production. The aim of the present work was to clarify whether a Th2 immune response has any influence on the success of vaccination using both inactivated and attenuated vaccines. The results showed that the Th2 response established prior to vaccination did not influence the induction of protection offered by the vaccines. However, the effectiveness of this protective response can be altered, depending on the adjuvant employed in the inactivated vaccines, when the Th2 response is established after vaccination, just before challenge with C. abortus.

Application of touchdown enzyme time release (TETR)-PCR for diagnosis of Chlamydophila abortus infection

Chlamydophila abortus-DNA was detected using a touchdown enzyme time-release (TETR)-polymerase chain reaction (PCR) assay as an improved test for sensitive and rapid diagnosis of abortion in small ruminants. Two hundred and fifty two placentae, liver or spleen tissue samples from aborting ewes and goats or aborted lambs and kids in which C. abortus infection was suspected were examined by TETR-PCR and the results were compared with cell culture. Sixty-five tissue samples were found to be TETR-PCR positive while only 56 samples were cell culture-positive. After resolution of discrepant samples with a confirmatory nested PCR assay, TETR-PCR had a sensitivity of 97% and a specificity of 99.5% while culture had a sensitivity of 84.8% and a specificity of 100%. The analytical sensitivity of the TETR-PCR assay was determined with DNA extracted from 4-fold serial dilution of C. abortus B577 culture and found to be 0.25 inclusion-forming unit per PCR. No reduction in the analytical sensitivity was noted when the assay was tested with mouse liver samples spiked with 4-fold serial dilution of C. abortus B577 culture. No target product was amplified when DNA from Chlamydophila pecorum was tested. TETR-PCR used in this study is a practical, rapid, sensitive and specific assay that could be used for the detection of C. abortus in infected tissue samples. We recommend the use of this assay as a supplemental diagnostic tool for detection of C. abortus in infected tissue samples.

Abortion in small ruminants in Switzerland: investigations during two lambing seasons (1996-1998) with special regard to chlamydial abortions

Abortion cases of 144 goats und 86 sheep were investigated etiologically during 2 lambing seasons (1996/1997, 1997/1998). Macroscopic inspection of fetus and placenta was completed by histopathology and bacteriological isolation of agents. In addition, immunohistologically the following antigens were labeled in formalin-fixed and paraffin-embedded tissue sections: Toxoplasma gondii, Neospora caninum, Chlamydophila abortus (formerly Chlamydia psittaci serovar 1) and Border Disease Virus. From farms with abortions caused by Chlamydophila abortus specific data were recorded. In 75% of abortion cases in sheep and in 59% of cases in goats an etiologic diagnosis could be substantiated. Chlamydophila abortus is the most commonly involved agent in the etiology of caprine and ovine abortion (sheep 39%, goats 23%), followed by Toxoplasma gondii (sheep 19%, goats 15%) and Coxiella burnetti (sheep 1%, goats 10%). All other agents are of minor importance. An infectious cause of abortion based on histopathologic findings without isolation of agents was observed in sheep (10%) and goats (21%). Malformation occurred in sheep (2%) and goats (3%) and lesions suggestive for Vitamin E/Selenium deficiency were seen in goats only (2%).

Chlamydiose abortive des petits ruminants au Maroc : valeur épidémiologique d’un kit Elisar de &lt;em&gt;Chlamydophila abortus&lt;/em&gt; (&lt;em&gt;Chlamydia psittaci&lt;/em&gt; sérotype 1)

Le test Elisa recombinant (Elisar) utilisant la protéine de 80 à 90 kDa spécifique de Chlamydophila abortus, développé à l’Inra de Nouzilly en France, a été évalué au Maroc. Trois cent sept sérums appartenant à cinq groupes d’ovins et de caprins, provenant d’élevages aux taux d’avortements élevés ou bien présentés pour contrôle vétérinaire, ont été testés. La comparaison des résultats obtenus avec le test de fixation du complément (Tfc) a montré une grande discordance entre les deux techniques. En effet, seulement 16 sérums ont été positifs avec Elisar contre 130 positifs avec Tfc. En particulier, parmi les 55 sérums provenant d’animaux avortés, 48 ont été positifs avec Tfc et 16 avec Elisar. Ceci soulève avec acuité la problématique du rôle de Chlamydophila pecorum dans les avortements des petits ruminants au Maroc. La recherche différentielle des pathologies à Chlamydophila abortus et Chlamydophila pecorum est nécessaire pour préciser l’origine des divergences observées entre Elisar et Tfc.

Preparation of a high titre Salmonella abortusovis H-antigen and antiserum and improvement of the microagglutination test

Serological diagnosis of small ruminant abortions due to Salmonella abortusovis (S.a.o) was improved by using modified procedures to produce high titre H-antigens and anti-H serum. While the titres of the two standard antigens were 1:8 and 1:20, both modified antigens had a titre of 1:50. Similarly, the use of adjuvanted and non adjuvanted antigens for the production of anti-H hyperimmune serum resulted in a titre of 1:160 000 as compared to 1:20 000 when standard procedures were used. In addition, the slow micro-agglutination test has been modified and results can now be read within the same day.