CHAPTER 24 Bacterial Diseases of the Fetus and Placenta Associated with Fetal Loss in the Mare
Fetal lesions and placentitis associated with abortions and stillbirths are major causes of economic loss in mares in many countries throughout the world.1–4 Before the mid-1980s the majority of pathogenic bacteria associated with equine placentitis and abortions were reportedly due to a group of bacterial microorganisms including Streptococcus zooepidemicus, S. equisimilis and other streptococci including non-beta-hemolytic Streptococcus spp., Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, and Actinobacillus equuli.1,2,4–6 However, since the late 1980s and early 1990s other microorganisms have dramatically emerged as very significant causes of fetal loss in mares.7–9 These agents include leptospires, nocardioform actinomycete group of organisms,10 and microorganisms associated with early and late fetal loss in mares.11 This chapter will review the emerging bacterial diseases currently causing equine fetal loss in mares.
The most common bacterial group currently being diagnosed is categorized as a group of organisms belonging to the nocardioform actinomycetes, of which there are several,12 followed by the streptococcal group of bacteria including Streptococcus zooepidemicus, Streptococcus spp., and Streptococcus equisimilis. Other commonly isolated bacteria include E. coli, Pantoea spp., Pseudomonas spp., and Leptospira spp. In addition, several less commonly isolated bacteria are associated with fetal septicemia. The largest number of fetal losses are free of known pathogens, but are infected with a group of bacteria referred to as nonpathogens and saphrophytes.11
Most gross and microscopic lesions affecting allantochorion fall into three categories: diffuse placentitis associated with a hematogenous origin, an ascending placentitis originating from vaginal tract and cervix, and a focal mucoid placentitis located in the anterior body of unknown origin.13 It is not uncommon to find overlapping of placental lesions when multiple etiologic syndromes simultaneously affect the fetus. In the umbilical cord and amnion, the primary lesions are edema and hemorrhage, and secondary inflammation apparently is associated with secondary opportunistic and pathogenic bacteria. Seemingly, lesions in umbilical cord and amnion are emerging as a significant cause of fetal death associated with midterm and late-term fetal losses.
Leptospirosis is a zoonotic bacterial disease of worldwide distribution that infects many species, including humans. In horses as well as other species, clinical responses range from inapparent to severe infections. Pregnant mares exposed to infected animals may abort.14–16 Most infections are caused by leptospires in the Pomona serogroup, and less frequently by the servor grippotyphosa, and rarely other strains.1,7,18–20 The horse is seemingly an incidental host for these and other serovars, with the possible exception of bratislava.15,16
The most dramatic pathologic changes in the fetus are seen in the liver, which is enlarged, mottled, and pale to yellow. The microscopic changes consist of hepatocellular dissociation, hepatocellular vacuolar degeneration, and focal to multifocal fibrosis and necrosis of individual hepatocytes and Kupffer cells. There is intrahepatic bile stasis. Some of the hepatocytes develop into giant hepatocytes with abundant cytoplasm and binucleated to multinucleated nuclei. There is a mixed leukocytic infiltration in the portal trials.21
The kidneys are grossly enlarged and edematous with pale white radiating streaks in the cortex and medulla. The microscopic lesions consist of multiple focally extensive microabscesses with few to several giant cells, and multifocal nonsuppurative interstitial nephritis. The tubules may be dilated.21
The changes in the lungs and heart are occasionally present and include varying degrees of bronchopneumonia and subpleural, interlobular, and alveolar hemorrhages. The pneumonic changes are usually minimal to mild and consist of focal neutrophilic infiltrations. The microscopic changes in the heart are occasionally found and consist of multifocal infiltrations of mixed leukocytes in the myocardium. The lymphoid organs may have a mild lymphoid necrosis with neutrophilic infiltration and the reticuloendothelial cells proliferate in the sinuses.21
The placental changes occur in the allantochorion where there are patchy areas of necrotic mucoid exudates on the chorionic epithelium. There may be lesions of nodular cystic hyperplasia in the allantois. Microscopic changes in the chorion consist of thrombosis, vasculitis, mixed inflammatory cell infiltration of the chorion, and villous necrosis and calcification. There is cystic adenomatous hyperplasia of allantoic epithelium.9,21 The spirochetes can be demonstrated in the majority of cases with immunofluorescent microscopic examination and in histologic sections stained by the Warthin-Starry method for spirochetes.9,21,22
For serologic testing, heart blood and pericardial fluid are ideal for testing fetuses and stillborn foals, and blood from the aborting mares can be tested for leptospiral antibodies by the microscopic agglutination test (MAT). The most common serovar found in affected fetuses and aborting mares are Leptospira interrogans serovars pomona, grippotyphosa, copenhageni, hardjo, canicola, and bratislava. The majority of cases react to only one serovar, but a few cases show cross-reactivity to more than one serovar.9,21,22
Horses appear to serve as maintenance host for the serovar bratislava due to the high seroprevalence of bratislava antibodies in various horse populations throughout the world.15 If in fact serovar bratislava is host adapted, transmission would be by direct contact with infected urine or by the venereal route from infected horses. Bratislava may be localized in the kidneys of horses and may be shed in the urine for the duration of the horse’s life.15
In North America most horses seemingly are infected with serovars maintained in other animal species, and mares are considered incidental hosts. In the central Kentucky area and likely other areas in North America, kennewicki, grippotyphosa, and hardjo, found in skunks, raccoons, and cattle, respectively, are the nonhost-adaped serovars associated with equine abortions. 10 These species are commonly found on or around horse farms. Once horses are infected on a farm, horse-to-horse transmission is likely to occur as infected mares may shed high numbers of the nonhost-adaped leptospires for several weeks.10
Attempts should be made to prevent pregnant mares from coming into direct contact with the urine of maintenance hosts as well as indirect contact with contaminated environmental drinking water, feed, and bedding contaminated with urine from maintenance hosts. Various antibiotics have been used to treat infected and carrier mares with mixed results with apparent success in some mares, but not in others.10
Bacteria that induce lesions of mucoid placentitis are categorized as a group of organisms belonging to the nocardioform actinomycete strains of bacteria, of which there are several 12 The causative organisms associated with these lesions are gram-positive, non-acid-fast, filame ntous bacteria. The bacteria were previously deemed to be nonpathogenic when they were found in lesions of mucoid placentitis. However, when the numbers of cases became more numerous in the early 1980s23,24 and subsequently increased dramatically in the late 1990s, these previously called nonpathogenic bacteria are now considered emerging opportunistic pathogens and a significant cause of abortions and premature foals.10
Mucoid placentitis is a chronic form of placentitis that is focally extensive, affecting the junction of the horns and body of the allantochorion. Typically, the exudate on the surface of the chorion is mucoid and mimics a mycotic placentitis. Unlike mycotic infections of the placenta, the cervical area of the allantochorion is not affected, which suggests a hematogenous route of infection. The causal organisms induce extensive lesions on the chorionic surface that result in a mild to marked placental insufficiency, depending on the extent of the placental lesions. Affected fetuses either abort, or are born prematurely and typically resemble a twin due to the placental insufficiency. Affected fetuses are usually alive when aborted and lesions are restricted to the liver, which is enlarged and discolored due to the placental insufficiency.
Unlike mycotic organisms,25 the causative bacteria are not easily demonstrated upon histopathologic examination with hematoxylin and eosin (H&E) staining, but can be demonstrated with the Gram stain.24 The organisms are easily cultured on blood agar using routine bacteriologic techniques but may require more time to grow than other bacteria or contaminating organisms.10,26
Using molecular techniques and comparing the sequence of the 16S rDNA gene against the public databases indicated a relationship to members of the suborder Pseudonocardineae. One of the causative organisms has been further classified as Crossiella equi.27 Other nocardioform bacteria, recently classified as Amycolatopis spp., are less commonly found in affected placentas.28,29 It is likely that other similar nocardioform bacteria are also associated with mucoid placentitis lesions.
Also inducing lesions of mucoid placentitis is a bacterium identified as Cellulosimicrobium (Cellumonas) cellulans (formerly Oerskovia xanthineolytica).30,31 This bacterium was first reported as causing abortion in Australia31 and later in Kentucky.30 Fetuses affected with C. cellulans have lesions in the chorion similar to those caused by Crossiella equi, but unlike C. equi, in some cases the lesions affect the cervical star area of the chorion, although in other cases the cervical star is not affected.30 The chorionic epithelium is necrotic, thickened, and tan, and sharply demarcates the normal adjacent chorionic epithelium. The lesions in the body of the placenta are focally extensive, irregular, and very demarcated from adjacent normal chorionic epithelium. The villi in the center of the lesion are absent and covered by a brown, mucoid exudate. The amnion may be discolored yellow.30,31
The microscopic changes in the chorionic epithelium include short, blunted villi lined by hypertrophic trophoblastic epithelial cells. There is an infiltration of large numbers of lymphocytes and plasma cells, and fewer numbers of macrophages and neutrophils. There is an increase in vascularity and fibroplasia in the affected area of the chorion. There may be a squamous metaplasia of the trophoblastic epithelial cells. The chorionic surface is frequently covered with an amorphous eosinophic material that contains neutrophils and bacteria. Also, lesions may be present in the allantois and amnionic portion of the umbilical cord, but are less severe than the chorionic lesions and consistent with a nonsuppurative inflammation.30
The placental lesions in the chorion resemble those found with Crossiella equi, except that the cervical star area may be affected. In addition, fetuses with C. equi infections generally have no lesions, other than enlarged pale livers.10 Fetuses infected with C. cellulans are usually emaciated and have gross lesions in the lungs and occasionally the liver. The lungs are firm and mottled red-tan. The liver is enlarged and mottled and may contain multiple 1- to 5-mm pale foci visible on the capsular surface and in some cases distributed throughout the hepatic parenchyma. The microscopic lesions in the liver are multifocal and there are mixed inflammatory cells in the portal triads.30,31
The microscopic lesions in the lungs include cellular debris, granulocytes, macrophages, and multinucleated histiocytic giant cells in the lumens of the alveoli and bronchioles. The pneumocytes are frequently hypertrophied, and the alveolar walls may be infiltrated with mononuclear inflammatory cells. Low numbers of gram-positive bacillary to filamentous bacteria can be seen in the pulmonary cells and in the cytoplasm of the multinucleated giant cells.30,31
The syndrome is not new, as it was first reported in 1982 in Australia.31 Earlier, fetuses with identical lesions were seen in an outbreak of abortions on a Thoroughbred farm in central Kentucky in December 1970 (Swerczek, TW, unpublished data). Multiple abortions occurred in a herd of 25 pregnant mares. Two 10-month-old fetuses were submitted for necropsy examination, and the gross and microscopic lesions in the chorion, amnion, umbilical cord, lung, and liver were consistent with those subsequently described in 1982 in Australia31 and in 2004 in Kentucky.30 Multiple mixed bacteria were isolated from the placentas and fetuses. Gram stains of lungs with fetal pneumonia revealed bacteria in the alveolar macrophages and multinucleated giant cells that were morphologically consistent with those reported in fetuses in 198231 and 2004.30
There were no other fetuses with similar lesions submitted for necropsy examination during the 1970 and 1971 foaling season from other area farms (Swerczek, TW, unpublished data). An unconventional feeding program was given to the affected pregnant mares that aborted. The affected mares were being fed haylage starting in November 1970, and approximately 3 weeks later mares aborted. It was suspected that the causative bacterium may have been present in the haylage that was being fed to cattle as well as the pregnant mares on the farm. The farm did not experience similar abortions in subsequent years, but because haylage was the suspected as harboring the bacterium at that time, it was no longer fed after the mares aborted in the fall of 1970.
The source of infection of Cellulosimicrobium (Cellumonas) cellulans is most likely the environment. In light of the cases in which haylage was suspected as early as 1970, these bacteria that are commonly found in soils may be also associated with the oral consumption of the causative bacteria in forages.
Until more is known on the distribution of the group of causative organisms in the environment, predisposing factors, and how these bacteria enter the reproductive tract of pregnant mares no recommendations can be made.