Equine Abortion Research Articles

Is the horse a reservoir or an indicator of Coxiella burnetii infection? Systematic review and biomolecular investigation

The role of the horse in Coxiella burnetii infection has not been defined. Accordingly, a twofold approach was taken to further our knowledge on this topic: (1) conduct a systematic review of the literature to establish available evidence of C. burnetii infection in the horse; (2) undertake a biomolecular investigation of 122 cases of equine abortion, stillbirth and neonatal foal death, for the presence of C. burnetii using a PCR test targeting the IS1111 gene of C. burnetii.A review of the literature turned up seven studies that identified C. burnetii DNA in equine specimens, especially aborted fetuses, while an additional 34 studies sought to determine seroprevalence of the infection in the horse. A meta-analytical approach was taken to calculate a pooled mean seroprevalence in equines based on published studies. A seroprevalence of 15.8% (95% confidence interval: 9.6–23.0%) was obtained. This figure is comparable to those previously reported in other species, especially ruminants.None of the 122 cases of equine abortion, stillbirth or neonatal foal death were positive for C. burnetii DNA.C. burnetii has rarely been looked for in equine specimens in previous studies. Cases of equine abortion should be comprehensively investigated to assess the risk of abortion in a pregnant mare infected with C. burnetii. Consideration should also be given to the possible role of the horse as a source of the organism for other animal species including humans.

Equine herpesvirus type 1 infection induces procoagulant activity in equine monocytes

The alphaherpesvirus, equine herpesvirus type 1 (EHV-1), is a highly prevalent cause of equine infectious abortion and encephalomyelopathy. These syndromes have been attributed to ischemic necrosis from thrombosis in placental and neural vessels, although the mechanisms underlying thrombosis are unknown. After inhalation, EHV-1 establishes a peripheral blood mononuclear cell-associated viremia, with monocytes being a target of infection. Monocytes are also the main source of tissue factor (TF) in diseased states. Since TF is the primary activator of coagulation, increased monocyte TF expression could be involved in EHV-1-associated thrombosis. We hypothesized that EHV-1 infection would induce TF-dependent procoagulant activity in equine monocytes. Monocyte-enriched fractions of blood were infected with abortigenic (RacL11, NY03) and neuropathogenic (Ab4) EHV-1 strains. All strains induced procoagulant activity, to variable degrees, within 1 to 4 h, with maximal activity at 24 h, after infection. Virus-induced procoagulant activity was similar to that seen with lipopolysaccharide, a known stimulant of TF-mediated procoagulant responses. Virus-induced procoagulant activity was factor VIIa-dependent and temporally associated with TF gene transcription, implicating TF as the main driver of the activity. Procoagulant activity was mildly decreased (30-40%) when virus was inactivated by ultraviolet light or when infected cells were treated with aphidicolin, a virus DNA polymerase inhibitor, suggesting early events of virus infection (attachment, entry or intracellular trafficking) are the primary stimulus of procoagulant activity. Our results indicate that EHV-1 rapidly stimulates procoagulant activity in equine monocytes in vitro. The EHV-1-induced procoagulant activity in monocytes may contribute to clinical thrombosis in horses with EHV-1 infection.

Isolation and partial characterization of lytic phage against Salmonella Abortusequi

Aim: To use partially characterized phages for generation of phage lysates having potential to be developed as vaccines against Salmonella Abortusequi. Material and Methods: Phage/s isolation from several sewage samples was attempted against Salmonella Abortusequi strains procured from the National Salmonella Reference Centre, IVRI using enrichment and agar overlay technique. A concentrated stock of the phage was prepared after its purification. The lytic range, morphology and DNA size were determined. Results: Two lytic phages could be isolated and purified. The phage with wider lytic range was found to possess a hexagonal head with a very short tail. The DNA size was found to be 21kb. Conclusion: The investigation reports first isolation of a lytic phage against Salmonella Abortusequi. The phage can be used in future for development of phage lysate based immune therapeutic agent against equine abortions.

Evaluation of three alternative methods for diagnosis of equine herpesvirus abortion

use of a labeled probe, to enable detection of amplified products using a LFD. The assay to identify T. equigenitalis successfully amplified a 121bp fragment of the 23S rDNA genes. The assay detected as few as 46 gene copies in 12 minutes. The assay did not cross-hybridise with any of the 38 different bacterial organisms tested. The RPA assay was tested against a panel of 60 clinical equine genital swabs. All of the culture negative DNA extracts were correctly identified. Of the 30 culture positive DNA extracts, 24 were correctly identified. Culture positive, RPA negative DNA extracts had high real-time PCR CT values, indicating low levels of target DNA present. The introduction of a second probe into the assay enabled the simultaneous detection and differentiation of T. equigenitalis and T. asinigenitalis. The T. equigenitalis assaywas successfully adapted to enable detection of amplification products LFDs. We have successfully developed a rapid and sensitive isothermal assay to identify T. equigenitalis. Additionally, the inclusion of a second probe enables the detection of T. asinigenitalis. The amplification products can be detected in real time or via LFD. This test provides a fast (under 15 minutes) and accurate means of identification. The low energy requirements, lack of expensive equipment and LFD readout makes this test a suitable candidate for rapid diagnosis in low technology laboratories or near to the point of sampling.

Strain impact on equine herpesvirus type 1 (EHV-1) abortion models: Viral loads in fetal and placental tissues and foals

Equine herpesvirus-1 (EHV-1) continues to cause both sporadic and epidemic abortions despite extensive vaccination. Lack of progress in the development of protective vaccines may be hindered by the lack of equine abortion models that employ contemporary EHV-1 strains. The objective of our experiments was to compare a contemporary EHV-1 strain with a previously described challenge strain, and to quantify EHV-1 loads in various maternal and fetal tissues. Infection experiments were performed in two groups of 7 pregnant pony mares at 270–290 days of gestation with a contemporary EHV-1 strain (University of Findlay 2003 isolate – OH03) or an EHV-1 strain isolated over 30 years ago, and previously described in abortion models (Ab4). All mares in both groups exhibited nasal viral shedding and viremia. Infection with OH03 resulted in 1/7 abortion and infection with Ab4 resulted in 5/7 abortions. In the OH03 challenge, placentas of foals delivered at term showed little detectable virus, while the aborted fetus expressed high levels of virus infection in the spleen and liver, lower levels in the lung and thymus, and lowest levels in the chorioallantois. After Ab4 challenge, high viral loads were detected in fetal and placental tissues in abortions. In the two normal deliveries, the chorioallantois contained virus levels comparable with the chorioallantois of aborted foals and both foals shed EHV-1 starting on day 4 of life, but were clinically healthy. Our results demonstrate the continued importance of strain selection for abortion models, and this study is the first report of viral load quantification using contemporary methods. Extremely high EHV-1 loads in decidua from abortions illustrate the infection risk posed to other horses.

Causes of equine abortion, stillbirth and neonatal death in central Italy

ABORTION, stillbirths and neonatal deaths are an important source of economic loss for an equine industry. Many of the infectious disease agents that have been implicated are important from the...

A racing start in life? The hurdles of equine feto‐placental pathology

Significant progress has been made in understanding and monitoring the causes of equine abortion over past decades. However, not all in utero pathology results in abortion. It has long been recognised that some in utero pathology, such as twinning or chronic placentitis, can result in the birth of live but growth-retarded foals and there is historical evidence that birth weight may influence future athletic performance. Clinical experience (e.g. from twins) and experimental studies (pony-Thoroughbred embryo transfer) have highlighted the importance of reduced functional placental area in limiting growth in utero in horses. Many other nonfatal in utero pathologies (e.g. umbilical cord-related circulatory compromise) can potentially affect either placental function or other organ systems. Their influence on the short- and long-term health of the foal and its future athletic performance is in many cases poorly documented or understood. This review summarises the main causes of in utero pathology and reflects on how these may potentially affect the foal if born alive, highlighting the need for long-term studies on this important subject.

Abortos em equinos na região sul do Rio Grande do Sul: estudo de 72 casos

Foi realizado um estudo retrospectivo das causas de aborto ocorridas em equinos na área de influência do Laboratório Regional de Diagnóstico (LRD), da Faculdade de Veterinária da Universidade Federal de Pelotas (UFPel), no período entre janeiro de 2000 e junho de 2011. Foram revisados os protocolos de necropsia e de 1.154 equinos ou materiais de equinos recebidos 72 (6,2%) eram casos de abortos. A infecção bacteriana foi a principal causa de aborto neste estudo com 36,1% dos casos. As causas não infecciosas corresponderam a 8,3% dos casos, os abortos virais a 4,2%, os parasitários a 1,4% e os inflamatórios a 2,8%. Em 47,2% dos casos não foi possível determinar a causa/etiologia do aborto. Lesões macroscópicas e histológicas características foram observadas em casos de aborto por Leptospira sp. e por herpesvirus equino-1 sendo que nos demais casos as lesões foram inespecíficas. Ficou demonstrado que o envio do feto inteiro junto com a placenta, sob refrigeração, aumenta consideravelmente a eficiência do diagnóstico e o elevado número de abortos de causa não determinada foi atribuído em parte ao envio de material não adequado.

Molecular detection of Coxiella burnetii and Neospora caninum in equine aborted foetuses and neonates

Abortion, stillbirth and neonatal death are major causes of equine mortality and cause severe economic loss to the equine industry. The present study was based on a complete necropsy protocol associated with classical microbiological examinations and molecular biology on 407 cases of abortion, stillbirths and neonate death. Based on this retrospective survey, “less common” abortive infectious agents were characterised by molecular tools in nine independent cases of abortion or neonate mortality. Among others, Chlamydophila abortus (1 case), Coxiella burnetii (6 cases) and Neospora caninum (3 cases) were detected by real-time PCR; one of these samples being co-infected by N. caninum and C. burnetii. DNA detection of this latter bacterium is reported here for the first time in equine abortion samples. C. burnetii should, along with other common pathogens, probably be taken into account in equine abortion.

SEROREVELATION OF ANTIBODIES OF EQUINE HERPESVIRUS (EHV1/EHV4) IN SYRIA

The objective of this study is, serological investigation of Epizootic Equine Abortion (EEA) or Equine Rhinopneumonitis (ER) Equine HerpesvirusMyeloencephalopathy(EHM) that know worldwide Equid herpesvirus (EHV type 1 and 4) in Syria. Equid herpesvirus 1 and 4 are widespread in the domestic horses population worldwide. EHV1 is a major cause of abortion, respiratory and neurologicaldisorders in horses. EHV4 is responsible of respiratory disease.In this study, Antibodies of EHV1 and 4 were investigated serologically in horses in Syria. A total of 203 unvaccinated horses in five regions in Syria were sampled and tested using indirect ELISA. EHV1-specific antibodies were found to be in fifth regions as 7(3.5%). EHV4- specific antibodies were found to be in fifth regions as 195(96%). This is the first serological investigation for EHV1 and EHV4 in Syria.

A unique genotype of Leptospira interrogans serovar Pomona type kennewicki is associated with equine abortion

Although serologic data indicate horses in N. America are exposed to a variety of leptospiral serovars, abortion is almost always associated with Leptospira interrogans serovar Pomona type kennewicki. A variety of wildlife including raccoons, white tailed deer, striped skunks, opossums, and red and grey foxes have been shown to host serovar Pomona and have therefore been suspect as sources of infection for pregnant mares. The aim of the present study was to examine genetic diversity in serovar Pomona type kennewicki in wildlife and in aborting mares. Our approach utilized PCR that targeted tandem repeats at the VNTR – 4 locus and a 1235bp 5′-sequence of the lk73.5 (sph2) and adjacent upstream sequence unique to serovar Pomona. All isolates/specimens of equine origin in 1992 and 2008 yielded amplicons of 1235 and 595bp, whereas 14 isolates/specimens from wildlife yielding a 1235bp amplicon characteristic of serovar Pomona produced amplicons of 1300, 550bp (3), 1300bp (10), or 595bp (6) with the VNTR – 4 primer set. Wildlife therefore hosted at least three different genetic variants of type kennewicki including the genetic variant that predominated in aborting mares. The data are consistent with other studies indicating specific genetic variants of type kennewicki show a strong tendency to be associated with a specific host. Levels of antibody in wildlife sera reactive with rLk73.5, rLig130 and sonicate of type kennewicki were poorly correlated with PCR data, although rLk73.5 was superior to rLig130 in detection of antibody responses. PCR is therefore a more reliable tool for studies of wildlife reservoirs of Leptospira sp. than serologic surveillance that targets host induced proteins or LPS-rich sonicate.

A 24-Year Retrospective Study of Equine Abortion in Normandy (France)

The main causes of abortion in mares in France were studied from 1,822 cases submitted for necropsy. The cause of abortion was established in 74.9% of cases (n = 1,365). Fetoplacental infections (n = 869) represented 63.7% of diagnosed abortions. A noninfectious cause was found in 27.2% of cases (n = 496). Of the infectious causes of abortion, the vast majority were caused by bacteria (n = 695; 79.9%), followed by viruses (15.1%), and then fungi (1.8%). In 27 cases (3.1%), no specific pathogenic agent could be identified despite the presence of lesions. Of the noninfectious causes of abortion, umbilical cord abnormalities were the most frequent (n = 300; 60.5%). Placental villous hypoplasia represented the second most frequent cause (17.3%). This was followed by lethal congenital malformations (6.9%). The causes of placental insufficiency other than placental villous hypoplasia—twins, placental edema, placental premature separation, and body pregnancy—were less frequent. The diagnosis of equine abortion cases examined in Normandy seemed to be somewhat in agreement with the findings reported in Kentucky and the United Kingdom. In this study, about 60% of the cases were associated with a condition involving the allantochorion or the umbilical cord. Thus, to enhance diagnostic success, it is of prime importance to submit the fetal membranes along with the fetus for necropsy.

Outbreak of Abortions and Infertility in Thoroughbred Mares Associated with Waterborne Aeromonas hydrophila.

At a thoroughbred equine breeding farm near Hissar (Haryana), three mares aborted in their seventh month of pregnancy. The vaginal swabs of all aborted mares, and stomach contents, heart blood, liver, spleen and placenta of aborted fetuses yielded pure culture of Aeromonas hydrophila. In addition, A. hydrophila was also isolated from the vaginal swabs of three repeat breeding mares and faecal sample of a diarrheic foal. The source of infection was possibly water supply as all the water samples collected from taps, mother tank and storage tank were found to be positive for A. hydrophila. The antibiogram of all the isolates was similar showing resistance to ampicillin, carbenicillin, gentamicin, kanamycin and amikacin but sensitive to chloramphenicol, ciprofloxacin, cefuroxime, ceftriaxone, cotrimoxazole, cotrimazine, nitrofurantoin, streptomycin and tetracycline. All the 20 sera samples collected from three aborted and three repeat breeding, and eight in-contact mares, a diarrheic foal, three cows and two male buffaloes maintained at the same farm contained antibodies against A. hydrophila with titres ranging from 80 to 640. The water supply was instantly chlorinated using 0.05% sodium hypochlorite for three consecutive days and all the culturally positive mares were treated with intravaginal administration of 1g ciprofloxacin, while the foal was given nitrofurantoin for three days. After one month, A. hydrophila could not be isolated either from mares or from their environment and antibody titre in all the seropositive animals showed a declining trend. Later, all the aborted and repeat breeding mares were confirmed to be pregnant. Thus, the present study indicated that water-borne A. hydrophila might be associated with equine abortions and infertility, and diarrhea in newborn foals.

The increased prevalence of neuropathogenic strains of EHV-1 in equine abortions

A panel of 426 archived EHV-1 isolates collected (1951-2006) from equine abortions was analyzed using a real-time Taq-Man((R)) allelic discrimination PCR assay. Based on previous findings, isolates possessing adenine at nucleotide position 2254 (A(2254)) in ORF30 were classified as having a non-neuropathogenic genotype and those with guanine at 2254 (G(2254)) were designated as the neuropathogenic genotype. The resultant data demonstrated that viruses with the neuropathogenic genotype existed in the 1950s and isolates with this genotype increased from 3.3% in the 1960s to 14.4% in the 1990s. The incidence of EHV-1 isolates from 2000 to 2006 with G at position 2254 is 19.4%, suggesting that viruses with the neuropathogenic genotype are continuing to increase in prevalence within the latent reservoir of the virus, leading to greater risks for costly outbreaks of equine herpesvirus neurologic disease. Another highly significant finding was two isolates failed to react with either probe in the allelic discrimination assay. These isolates were found to possess an adenine to cytosine substitution at position 2258 (A(2258)-->C(2258)) in ORF30, in addition to A(2254)-->G(2254). Interestingly, the non-neuropathogenic RAC-H modified live vaccine strain of EHV-1 also contains both A(2254)-->G(2254) and A(2258)-->C(2258) substitutions. This finding clearly suggests that additional research is required before the genetic basis of the neuropathogenic phenotype in EHV-1 is fully understood.

Identification of Bartonella henselae in an Aborted Equine Fetus

This report describes the characterization of a Bartonella henselae abortion in an equine fetus by gross, histologic, immunohistochemical, ultrastructural, and molecular methods. Bartonella henselae can cause cat scratch disease, bacillary angiomatosis, bacillary peliosis, and endocarditis in humans and other animals. The bacterium has been isolated from several mammalian species but only recently from equids; however, it has not been linked to abortion in equids. An aborted equine fetus exhibited necrosis and vasculitis in multiple tissues, with intralesional Gram-negative short-to-spirillar bacteria. Nucleotide sequence analysis of the bacterial 16S rRNA gene amplified from the DNA extracted from fetal tissues revealed 99.9% homology to that of B. henselae. The presence of B. henselae in the fetal tissues was further confirmed by polymerase chain reaction amplification and nucleotide sequence analysis of other Bartonella species-specific genes. Microorganisms were immunohistochemically labeled with a monoclonal antibody to B. henselae and were ultrastructurally characterized. Attempts to detect known causative agents of equine abortion were unsuccessful. Given the severity of vasculitis and the presence of intralesional bacteria, we concluded that B. henselae infection caused the abortion of this foal.

Review Paper: Mare Reproductive Loss Syndrome

An epidemic of early fetal loss (EFL), late fetal loss (LFL), fibrinous pericarditis, and unilateral uveitis which occurred during the spring of 2001, are together now known as the mare reproductive loss syndrome (MRLS). A similar epidemic with less intensity was reported during the same period of time from southern Ohio, West Virginia, and Tennessee. The same syndrome with lesser intensity recurred in 2002. The estimated economic loss from the syndrome in 2001 and 2002 together was approximately $500 million. Both EFL and LFL were characterized by the absence of specific clinical signs in aborting mares. Nonhemolytic Streptococcus spp. and Actinobacillus spp. accounted for 65% of the organisms isolated from fetuses submitted for a postmortem during the MRLS period in 2001 and 2002. The pathologic findings in fetoplacental units of LFL included bronchopneumonia and funisitis, and there were no findings in EFL. Epidemiologic studies conducted in 2001 suggested an association between the presences of eastern tent caterpillars (ETC) in pastures with MRLS. Experimental studies in pregnant mares by exposure to ETC, or administration by stomach tube or with feed material, reproduced EFL and LFL. Similar experimental studies in mouse, rats, and goats with ETC were unsuccessful. Currently, 2 hypotheses are proposed for MRLS. One hypothesis proposes that an ETC-related toxin with secondary opportunistic bacterial invasion of the fetus leads to MRLS. The second hypothesis suggests that a breach of gastrointestinal mucosal integrity by hairs of ETC leads to a bacteremia and results in MRLS. In 2004, a similar equine abortion storm was reported from Australia and caterpillar exposure was identified as a risk factor for the abortion. In 2006, the syndrome was observed in Florida and New Jersey.

A survey of equine abortion and perinatal foal losses in Hungary during a three-year period (1998–2000)

Cases of equine abortion and perinatal foal losses were investigated in Hungary during a three-year period (1998-2000). Samples from aborted equine fetuses and newborn foals (total n = 96) were examined using bacteriological, virological, pathological, immunohistochemical (IHC), molecular biological and serological methods. The cause of abortion and perinatal foal loss was identified in 67/96 cases (70%); viral infection was found in 22 (23%), viral and bacterial coinfection in 1 (1%), bacterial infection in 23 (24%), protozoan infection in 1 (1%) and fungal infection in 2 cases (2%). Morphological lesions suggestive of infection were recorded in 2 (2%) and non-infectious causes in 16 cases (17%).

Neospora spp. infection associated with equine abortion and/or stillbirth rate

Neospora spp. infection associated with equine abortion and/or stillbirth rate F. Veronesi & M. Diaferia & M. T. Mandara & M. L. Marenzoni & F. Cittadini & D. Piergili Fioretti Published online: 12 August 2008 # Springer Science + Business Media B.V. 2008

Klebsiella oxytoca as a cause of equine abortion—Short communication

Klebsiella (K.) oxytoca infection induced the abortion of a female equine fetus in the 10th month of pregnancy. Bacteria were cultured from the liver, lung and stomach content. They were labelled with an anti-Mycobacterium bovis antibody in the thymus, liver and lungs and were stained with Giemsa and Brown-Brenn staining in the thymus and lung. The diffusely consolidated lungs contained numerous grey-whitish foci 2-4 mm in diameter, which corresponded to severe pyogranulomatous pneumonia characterised by numerous intraalveolar neutrophils and macrophages and multinucleated Langhans' giant cells. K. oxytoca was located in the cytoplasm of these cells, and extracellularly in the lumen of alveoli, bronchioles and bronchi, in the capsule of thymus and in the sinusoids of the liver. The results indicate that K. oxytoca can cause sporadic equine abortion.

A case of equine abortion caused by Encephalitozoon sp.

A Lippizan mare aborted a male fetus a few days before the expected foaling date without showing any clinical sings. Focal lympho-histiocytic hepatitis in the foal and multiplex focal lympho-histiocytic villitis accompanied by villus necroses and marked hypertrophy of chorionic epithelial cells in the arcades were observed. Elongated nucleated organisms were seen in groups in vacuoles or solitarily located in the cytoplasm of the chorionic epithelial cells. The organisms were in large numbers and often extracellularly in areas of villitis and villus necroses. They were Gram-positive, stained with haematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Giemsa, weakly with Warthin-Starry silver stain but not with Gömöri's methenamine-silver stain. By ultrastructural and immunohistochemical examinations, the organisms were identified as microsporidia belonging to the genus Encephalitozoon. No Encephalitozoon organisms were detected in the fetal organs. This is the first reported case of equine abortion induced by Encephalitozoon sp. in Europe. Although abortion induced by Encephalitozoon is rare, microsporidia should be considered a differential diagnosis for intracellular organisms observed in the chorionic epithelial cells of horses.