Noninfectious causes of abortion are similar to those known to cause embryonic loss, including nutritional imbalances, malnutrition, stress, environmental toxins, teratogenic compounds, hormone imbalances, and genetic abnormalities. An expanded list of etiologic agents is shown in Table 54.1 and in the following chapters.

Pathophysiology of injury to the conceptus

Maintenance of the first half of gestation (the first 200 days) in cattle requires a persistent corpus luteum (CL), which is maintained by the fetus.^7,13 Luteolysis may occur in pregnant cows due to excess prostaglandins (exogenous administration or secondary to heat stress) or Gram-negative bacterial septicemia.¹⁴ If the CL is destroyed at an early stage, death and rapid loss of the embryo with minimal degeneration may be seen.¹⁵ If there is fetal death prior to luteolysis, the CL may regress with eventual expulsion of autolyzed fetal tissues. However, in some cases the CL is maintained after fetal death, which can result in expulsion, resorption, or mummification.¹⁶ Early abortions are typically not recognized; most are occultly expelled, severely autolyzed, or mummified.

The maintenance of late-stage pregnancy requires both the fetus and placenta. At this stage in development, sufficient fetal stress can induce parturition through natural endocrine mechanisms. In this way, chronically diseased fetuses can initiate their own premature delivery.

Fetal death often leads to abortion within a few days rather than immediately, allowing sufficient time for autolytic changes to occur. The mechanism for expulsion of dead fetuses is unknown, but may share many similarities with normal parturition.¹⁶ Autolytic changes include generalized edema, pallor, and hemoglobin-stained fluids in the body cavities. Visceral tissues become pulpy or semi-liquid.¹⁶ In some cases, the time between fetal death and expulsion may be characteristic for a pathogen, and may therefore be useful to the clinician. Fetal infections due to Listeria monocytogenes, Trueperella pyogenes, nonseptate fungi (Absidia spp., Mucor spp., Rhizopus spp.), and bovine herpesvirus (BHV)-1 may lead to fetal death with expulsion days later. Fetuses infected by Campylobacter fetus and Aspergillus spp. may be delivered alive.¹⁷

It is important to recognize that infection of the conceptus does not necessarily lead to fetal death. The ability of a pathogen to injure the conceptus is influenced by the dam (general health, previous exposure), the stage of fetal development, and the virulence of the infectious agent. Fetal development is a continuum of organogenesis, physiological development, and immune development. Early stages of gestation are more prone to infection and severe disease, as the capabilities of the fetus are underdeveloped. The closer the fetus is to parturition, the stronger and more capable it is of defending itself from pathogens. Thus infection at different stages of development will produce different outcomes in the fetus.

Bovine viral diarrhea virus (BVDV) infection is a good example of this. Infection in the first trimester often leads to fetal death and resorption, while infection in the second trimester may lead to developmental anomalies. The character of the developmental defects depends on which cells or tissues are susceptible to the virus at the time of infection. In the second trimester, cerebellar growth is maximal and BVDV infection at this time may lead to cerebellar necrosis and cerebellar hypoplasia. Also at this time, hair growth is highly active and BVDV infection may damage follicles, resulting in hypotrichia. By the third trimester, the immunocompetent fetus may react to BVDV infection by mounting a sufficient immune response with antibody production and virus elimination. The only evidence for such an infection would be fetal specific antibody titers.¹⁸

Other viruses, such as BHV-1, cause extensive cell necrosis and hemorrhage, leading to rapid fetal death. Neospora caninum infection in the third trimester may cause only mild inflammation, with the birth of asymptomatic calves.

Effects of maternal disease

Infectious pathogens may cause abortion either by directly infecting the conceptus, or by affecting the dam such that severe physiological disturbance leads to abortion. The dam is predisposed to infection through nonspecific immunosuppression that occurs during pregnancy.¹⁹ Fever (mastitis, pneumonia), circulatory disease (myocarditis, severe anemia), hypoxia, and endotoxemia (Gram-negative sepsis) are possible conditions that may cause abortion indirectly.¹⁸ Prostaglandins may be elevated in febrile states, which can lead to luteolysis and abortion.²⁰ The likelihood of abortion probably increases with the number of pregnancies or previous abortions by the dam.⁶

Routes of infection to the conceptus

In cases where infectious organisms invade the conceptus, four routes are likely.¹⁸ Hematogenous spread from the dam to the placenta is commonly suspected in cases of infectious abortion, particularly due to Listeria monocytogenes, Leptospira interrogans, Salmonella enterica, Brucella abortus, fungus, BHV-1, or BVDV infection. Ascending infection from the vagina through the cervix can occur from primary vaginitis (characteristic of Tritrichomonas foetus) or contamination at insemination. The presence of pathogens within the uterus (endometritis due to Campylobacter fetus, Trueperella pyogenes) and descent from the abdomen via the uterine tubes (mycobacteriosis) are also possible pathways for organisms to reach the conceptus.

Route from the placenta to the fetus

Once a pathogen reaches the placenta, it may proceed to the fetus through the blood, via the umbilical veins, or by contamination of the amniotic fluid¹⁸ (Figure 54.1). For example, BVDV primarily infects the fetus hematogenously whereas BHV-1 may first infect the placenta, and then proceed to the fetus. Primary placental infection with subsequent amniotic fluid contamination is characteristic of fungal infections. In some cases a pathogen need not infect the fetus, such as with Salmonella enterica infection, where severe infection of the placenta may lead to generalized fetal hypoxia and subsequent death.

Lesions in placenta

Placentitis most commonly develops by one of three ways: hematogenous spread, extension from a diseased uterus, or an ascending vaginal infection. Gross placental lesions are characteristic of chronic bacterial and fungal disease, while viral infections generally produce no lesions.²¹ Changes of significance may occur in both the cotyledons and/or intercotyledonary spaces. The presence of fibrin (yellow, stringy, friable material) on cotyledons is an indication of inflammation (Figure 54.2). Fibrin must be distinguished from necrotic inflammatory exudate or entrapped caruncle fragments (necrotic uninfected endometrial tissue). The presence of ecchymosis and extension of the changes to the intercotyledonary space are suggestive of an infectious inflammatory process. With chronicity, stereotypical changes of placentitis include (i) placental fibrosis and edema; (ii) cupping of cotyledons; (iii) exudate on chorionic surfaces; and (iv) necrosis of the cotyledons.¹⁰

Fungal and bacterial inflammation of the cotyledons is often associated with necrosis, which imparts a firm to friable, tan, irregular or nodular consistency to the tissues with interspersed ecchymoses. Fungal organisms prefer to grow along and within blood vessels; this often leads to thrombosis, hemorrhage, and necrosis (Figure 54.3). Fungal organisms may be particularly abundant at the periphery of lesions rather than in the centers. A useful feature for interpretation is the extension of the pathologic process into the pericotyledonary chorioallantois. Such infiltrates may bridge cotyledons and often impart a leathery thickened texture. Another differential for such a change would be adventitial placentation (discussed in the section on pathology). Leptospirosis and BHV-1 infections may also produce a placentitis and appear similar to one another.²²

Ureaplasma diversum is known to produce a rather characteristic hemorrhagic amnionitis, where the membranes are thickened, opaque, and ecchymotic with fibrin, necrosis, and fibrosis.²³

Lesions in fetus

Gross lesions of the fetus, aside from malformations, are generally uncommonly noticed. They may be inapparent for four reasons: (i) the fetal immune system is not of sufficient robustness to mount a response that is easily observed at the gross level; (ii) fetuses often die before lesions appear; (iii) lesions are masked by autolysis, and (iv) fetal pathology may manifest itself in ways that are difficult to recognize.²⁴ Table 54.2 lists some causes for commonly encountered lesions in aborted fetuses.

The most common finding in abortion is autolysis, the degree of which depends on the cause of death, the time from death to abortion, and the time from abortion to examination.²² In experimentally induced, sterile, dead ovine fetuses, changes associated strictly with autolysis included (i) lack of odor, (ii) subcutaneous blood-tinged gelatinous edema, (iii) blood-tinged fluids in body cavities, (iv) renal cortex softening, (v) liver softening, (vi) abomasal content that was cloudy yellow to red, and (vii) uniform color (pink/red) of tissues²⁵ (Figures 54.4 and 54.5). By 12 hours after death, the fetal corneas were cloudy, the liver and kidneys were friable, and abomasal content became cloudy with brown flecks. After 36 hours, the subcutis contained edema and the skin would slough. By 144 hours, progressive dehydration was obvious (mummification).²⁵ Signs of fetal infection include fibrinous exudates in body cavities, white to tan foci in the liver or lungs, and evidence of abnormal development²² (Figure 54.6).

Pathogens that invade the fetus via umbilical veins may produce lesions in the liver, as it is the first organ encountered (Figure 54.7). Listeria monocytogenes, BHV-1, Yersinia pseudotuberculosis, and Salmonella enterica infections are often associated with liver necrosis. For pathogens that infect the amniotic fluid, exposure of the skin, lung, and intestines may occur. Skin lesions are particularly well known in cases of fungal infection, where pale patches of thickened skin are evident (Figure 54.8). Lung and pleural disease may be manifestations of hematogenous systemic spread or inhalation of infected amniotic fluid during fetal distress.¹⁸

Epizootic bovine abortion causes distinctive lesions in the fetal liver, lymph nodes, and spleen. The liver is irregular and nodular; however this change may also be seen in cases of congenital heart disease where chronic passive congestion occurs (Figure 54.9). The spleen and lymph nodes are characteristically enlarged due to lymphoid and mononuclear cell hyperplasia.¹⁶

Outcome of exposure to abortifacient agents

Fetuses exposed to abortifacient agents or conditions may die (resorption/abortion/stillbirth), they may be infected (with or without disease), they may be malformed (live/dead), or they may be normal.¹⁹

Death in the embryonic stage leads to resorption. In the fetal stage the presence of skin and musculoskeletal structures, in addition to placental endocrine activity, prevents resorption. Instead fetal death leads to autolysis and expulsion, or retention with mummification/maceration. In late gestation, a sick or stressed fetus may precipitate its own premature delivery with one of three results: the fetus may be born and then die, it may be born and fail to thrive, or it may be born and thrive even if infected.¹⁸

Disease may impair fetal motion, a necessary requirement for proper musculoskeletal development. In such cases, the calves may exhibit severe changes, including arthrogryposis, ankylosis, scoliosis, torticollis, and similar defects that lead to dystocia, stillbirth, or death in the perinatal period.

History/background investigation

The importance of a thorough history cannot be overstated; essential information is found in a systematic query of farm details. Abortion is both an individual and potential herd problem, and lack of investigation in both will severely hamper efforts to alleviate unacceptably high losses. It is important to keep in mind that more than one agent may be involved in abortion. Similarly, although more than one infectious agent may be present in an aborted fetus, only one may be the cause of death. Organisms regarded as low virulence may be the cause of abortion, but are also commonly present as contaminants.¹⁷

Sporadic abortions may be a manifestation of an ongoing herd problem or they may be the beginning of an outbreak; in either case investigation is warranted. Standard forms are used to reliably collect important cow and herd information (see Appendix); systematic evaluation of management practices will help direct investigations. Pertinent questions are listed below.³⁹ These questions and other information are incorporated into the investigation form provided. An expanded questionnaire has been published.⁴⁰

Examination of the cow

In some cases, the health condition of the cow has direct implications as to the cause of abortion (plane of nutrition, respiratory/intestinal disease, Anaplasma sp. infection). Any vaginal discharge or cervical mucus should be collected and sent to the diagnostic laboratory for microbiological analysis. Blood samples for serology should be taken from the dam as well as unaffected cows and those that have recently aborted. Serum should be acquired 2–3 weeks later in order to provide paired samples. Collecting serum from 10 other herdmates (or 10% of the herd) will make serological assessment more meaningful.⁶ Whole blood in EDTA is useful for blood smear and serum chemistry analysis of the dam. Gloves are to be worn during all examinations, as zoonotic agents may be present.

Examination of tissues

A necropsy is an examination with a definite purpose in order to yield the maximum amount of information. As such, it is vitally important to be systematic so that a full complement of tissues is taken during each examination. Optimally one should send the entire fetus and membranes to a diagnostic laboratory for evaluation. It is of primary importance to send fetal membranes as these may be diagnostic. If this is not convenient, then a standard approach should be adopted, as described here.

Examinations should be directed toward suspected etiologies, but keep in mind that more than one pathogen may be present. Typical findings in aborted calves that died in utero include edematous red tissues that lack normal tinctorial distinction, clear red fluids in body cavities, pale soft livers, and friable autolytic kidneys. Digital photographs are an excellent way to document lesions and may be useful in communications with diagnostic laboratories.

Examination of the fetus

It is important to verify the approximate time of death, which can be done based on gross findings. In antepartum fetal death, tissues are autolyzed or mummified and include the findings described above. Partum death is characterized by signs of life (localized head or limb edema, partial lung aeration, limb/rib fractures, subcutaneous shoulder hemorrhage, liver fractures with hemorrhage) in addition to proper degrees of fetal maturation/formation. Neonatal deaths are characterized by aeration of the lungs, umbilical artery thrombosis (swollen purple/red nodules along the arteries that run lateral to the urinary bladder), loss of the eponychium (soft fetal hoof keratin), and milk in the stomachs.⁴¹

The method of necropsy for aborted fetuses is basically the same as for any animal. The degree of autolysis and weight of the fetus and placenta are noted. Even if severely autolyzed, fetal tissues may be of use for polymerase chain reaction (PCR) screening for pathogens. Estimation of gestational age can be accomplished by the following formula: x = 2.5(y + 21), where y is crown–rump length in centimeters and x is gestation age in days.⁷ The crown is a point midway between the orbits on a line traversing the frontal eminence; the rump is the base of the tail or first coccygeal vertebra.⁴² In embryos, the greatest total length may be used instead of crown to rump. For stillbirths, fetal weight is useful in the diagnosis of dystocia due to fetal–maternal disproportion. Also, comparison of calf weight to thyroid weight in cases of suspect hyperthyroidism (goiter) may be useful. Reports of normal thyroid weight in calves vary from 6.5 to over 18 g.⁴³ Further discussion on this subject can be found in the section on noninfectious causes of abortion.

Most aborted calves have no gross lesions. Small white/tan foci in the lungs, liver, and kidneys may be present and indicate necrosis or inflammation. The presence of fibrin on organ surfaces is helpful for confirming inflammation. Meconium on the perineum or in airways/stomach indicates fetal stress and anoxia, typically associated with placentitis or dystocia (Figure 54.12). Hemorrhage and fractures of shoulder and hip joints are most often seen as a result of dystocia.

Slightly raised, white skin plaques that are firmly adherent, particularly around the eyes and face indicate fungal infection. Sometimes calves will have scattered white mineral on the hair, which is easily removed; this finding is not significant but must be distinguished from adherent fungal plaques (Figure 54.13). In calves born dead, the lungs are uninflated; they are uniformly dull red/plum, heavy, and sink in formalin (see Figure 54.4). In cases of dystocia where fetuses may have taken a partial breath, there may be incompletely inflated lungs; assess lung aeration in sections of cranial and caudal lobes by degree of the flotation in fixative/fluid. Make note of any milk in the stomachs, as this indicates colostrum consumption, which will interfere with bacteriological and serological findings.⁷

Necropsy method

It is best to have an abortion kit prepared beforehand; Table 54.4 enumerates materials to have in a kit. Zoonotic pathogens may be present, so always wear gloves, protective clothing, eye/face protection, and use careful technique when examining dams and fetal tissues. In addition, materials used in standard field necropsies are needed: clean water, bucket, brush, and disinfectant. Make sure to disinfect contaminated sites and equipment. If there are questions as to which samples are best to take, contact the diagnostic laboratory to which samples will be submitted for advice. Taking photographs of lesions is a relatively simple and excellent practice which will aid diagnosis.

1. Measure crown–rump length along the vertebral ridge (see above) and weigh fetus and placenta separately. Examine skin for exudate and abnormalities, around the eyes and face in particular for signs of fungal infection.
   
2. Examine the body for congenital abnormalities, umbilical hernias, cleft palates, facial anomalies.
   
3. Wash the surface of fetus with water to remove gross contaminants.
   
4. With the left side down, reflect right legs with a knife and continue the incision from cranial to caudal, reflecting the skin to the dorsal and ventral midlines. Collect any pooled blood between incised tissue planes with a syringe if available (for serology).
   
5. Open abdomen at the highest point along the caudal aspect of the ribs, being careful not to contaminate any peritoneal fluid or tissues. Aspirate any abdominal fluids with a sterile syringe, place in sterile tube, label, refrigerate. Continue the cut along the caudal aspect of the ribs, along the lumbar area, and cranial to the pelvis to expose the abdominal viscera. Note any lesions, and volume, character and color of fluids.
   
6. Incise the diaphragm (noting if negative pressure is present) and cut the ribs at the costal angles and along the sternum; remove the chest wall. Aspirate any thoracic fluids with a sterile syringe, place in sterile tube, label, refrigerate. Note any lesions, and volume, character and color of fluids. The pleural surface of the removed chest wall may be used as a makeshift cutting board/sterile field for dissection.
   
7. Aspirate fluids from the unopened abomasum with sterile syringe, place in sterile tube, label, refrigerate. Note the character of the abomasal fluid. Take samples of any other suspicious fluids at this time.
   
8. Examine the body cavities and organs in situ, making note of fibrin, hemorrhages, or abnormally situated organs.
   
9. Remove thoracic viscera from tongue to lung (pluck). Examine the mouth; one may split the mandibular symphysis with a knife. Incise the esophagus along the entire length; incise the dorsal tracheal membrane from the larynx to the bronchi. Examine the chest for any fractures. Slice multiple lung lobes, examining the cut sections for changes in the lobules and airways. Examine the thyroid gland. Collect tissues, especially lesions (see numbers 16 and 17).
   
10. Examine the heart, making sure to inspect all four main valves and four chambers. Look for ventricular septal defects or other anomalies.
    
11. Collect visceral organs, examining gastrointestinal tract last. Identify the kidneys, ureters, and urinary bladder. Remove the gastrointestinal tract by incising the root of the mesentery and continuing the cut to the diaphragm along the dorsum, avoiding the urinary tract. The tract may then be lifted out of the abdomen, leaving the colon attached and uncut. Collect tissues, especially lesions (see numbers 16 and 17).
    
12. Remove the adrenal glands.
    
13. Remove the urinary tract, making multiple slices through the kidneys. Identify genital organs.
    
14. Serially section the liver and spleen.
    
15. Open the forestomachs and abomasum, examine for content. Continue into the intestine, making several incisions along each major section (duodenum, jejunum, ileum, cecum, colon). Examine content and the mucosa.
    
16. Collect organs (kidney, liver, lung, spleen, thymus, adrenal) in sterile bags (placenta kept in separate bag), refrigerate.
    
17. Collect in formalin for histopathology: lung, liver, kidney, spleen, heart, adrenal glands, skeletal muscle (three to four sections including tongue, diaphragm), thyroid, ileum, skin (including eyelid), thymus, and mesenteric lymph node in 5-mm sections.
    
18. Remove the head from the body at the atlanto-occipital joint. Harvest entire brain, even if severely autolyzed, and place whole in formalin. If the fetus is severely autolyzed, take a swab culture of the brain tissue prior to fixation. The skull is relatively soft and can be easily opened with a saw, making two cuts along the inner aspects of the occipital condyles, cranially (and somewhat laterally along the curve of the calvaria) toward the inner aspects of the orbital rims. Make a third incision across the orbital ridge, connecting the first two cuts. Pry the cranium from the skull in one piece. Place the brain in the palm of your hand and with a scalpel gently cut the nervous attachments of the base of the brain, allowing the brain to fall into your hand.
    
19. Collect ocular fluid with a needle/syringe for nitrate/nitrite levels if needed, freeze.
    
20. Open several joints, looking for purulent exudate, fibrin, and hemorrhage.
    
21. Examine fetal membranes for exudates, lesions, abnormalities (see next section for detailed instruction). Remove dirt/debris from the membranes prior to sampling.
    
22. Place two to three sections with cotyledons in a sterile bag, refrigerate; make impression smears of any lesions.
    
23. Harvest the placenta for histopathology (at least three to four sections including cotyledons and intercotyledonary areas).
    
24. Collect amniotic fluid if available, refrigerate.
    
25. Collect dam blood in red top tube (herdmate blood samples may be collected as well; 10 cows or 10% of the herd has been suggested).

Examination of the membranes

Collect as much of the membranes as possible and carefully examine the chorioallantois and amnion, as lesions may be subtle, focal, and easily overlooked. In many cases, little if any placenta can be examined. It is important to try to verify if all membranes have been expelled. Because of the attachments of the amnion to the chorion, as well as the short umbilicus, calves will rupture and escape both the amnion and chorioallantois at delivery. Typically membranes are found with cotyledons on the outside.

The fetal membranes may be laid out lengthwise to identify the umbilicus and with it the internal aspect of the amnion. Examination of amniotic membranes and fluids is best done at this time. The umbilicus is composed of four muscular turgid vessels (arteries and veins) loosely enmeshed in slippery membranes. The gravid horn is easily identified as the larger half of the tissue, and should have larger placentomes. Avascular chorionic tips should be identified and inspected carefully, as true placental lesions may be present in the gravid horn near the tip.¹⁶ The umbilicus is continuous with the umbilical arteries and veins, which form a useful longitudinal axis by which to orient the placenta. From there the internal aspect of the allantoic cavity may be examined, with attention to the nature of any fluids present, thickness and color of membranes, and peculiar odors. Allantoic calculi are typically found at this time.

By placing the umbilical vessels at one side of the tissue, a lengthwise incision along the opposite side of the placenta will allow the placenta to be opened and laid flat, with the umbilical vessels forming the central axis (Figure 54.14). The horns are unequal in size, the gravid horn being much larger. In this way, vessels and cotyledon rows (normally four) may be examined thoroughly. Fresh cotyledons have a red velvety texture with a mildly irregular surface. The size variation, shape, color, consistency, and degree of cotyledon mottling should be noted, as well as degree of autolysis. Occasional small tags of attached caruncular tissue may be seen. Cotyledon margins are typically sharply defined, and the intercotyledonary chorion is white, translucent, and smooth. Adventitial placentation is a frequent finding that appears as small cotyledons or villous tuft formations adjacent to the placentomes that may cover the entire placenta.

Sample collection

A complete set of tissues should be collected in every case.⁴⁴ Biological specimens must be properly collected, prepared, stored, and transported otherwise diagnostic accuracy will suffer. It is important to maintain aseptic technique despite apparent tissue contamination; this is necessary to maximize chances for diagnosis as well as for prevention of zoonotic infection. Microbiology samples are to be kept chilled and sent to the diagnostic laboratory as soon as possible in a Styrofoam cooler with ice packs for overnight courier shipping. If fresh tissue samples cannot be sent within 2–3 days, then freezing at –70 °C and delivering on dry ice is ideal.⁴⁴ If necessary, specimens can be placed at –20 °C until dry ice can be obtained. Conventionally frozen tissues (–20 °C) are often acceptable for PCR testing of many pathogens. Abomasal and thoracic/pericardial fluid samples should be sent in sterile tubes without additives. Frozen fetal liver, lung, and kidney (2.5–5 cm chunks, –20 °C) should be kept for testing in case nutritional or toxicologic disease is suspected after initial examinations.³⁹

Chilled or frozen tissues should be placed in sealed bags (whirlpack-type bags often leak), preferably with a secondary bag around them. Fluids shipped in glass tubes require ample padding in tight-fitting containers. Absorbent materials should also be included in the package to prevent soaking of the exterior, which complicates and may delay the shipping process. Disposable diapers are an effective option.³⁹ Contact your diagnostic laboratory for proper packaging and shipping requirements for abortion samples as they may contain infectious and/or zoonotic agents.

Viruses may have very specific tissue tropisms and therefore require certain tissues for isolation. Specimens need to be collected as soon as possible as autolysis can be very damaging to virions. Virology samples for abortions are typically pooled, although placing tissues in separate bags is ideal. Placental tissues should be bagged separately. Do not pool samples from multiple abortions. Tissues should be kept cool but not frozen until arrival at the diagnostic laboratory. If tissues will not be immediately used, freezing at –70 °C is better than at –20 °C.³⁹ If live viruses are sent packed in dry ice, be sure to place samples in airtight containers as the gaseous phase of dry ice is carbon dioxide, which can lower sample pH and inactivate some viruses. Certain laboratories may have preferences for virological samples, particularly if a certain pathogen is highly suspected.

Bacteriology tissue samples should be 2.5–5 cm per side in order that surfaces may be heat seared for proper culture. External and gut samples should not be bagged with internal organs. If samples are contaminated, sending a swab culture (Culturette) in addition to tissues may be helpful. For severely decomposed specimens, aseptically collected brain tissue may be useful for isolating pertinent organisms.²²

Fungal sampling should at least include affected cotyledons; however it is best to send the entire placenta. Fetal infection is inconsistent, but lung, skin, and abomasal content are useful.

For histopathology, sections approximately 5 mm wide in 10% neutral buffered formalin are best. The brain, however, should be immersed whole in formalin, without dissection. If the brain is liquefied, fix separately. For small lesions, several foci should be submitted. For large lesions, a few sections including the margin with normal and abnormal tissues should be collected. Identify clearly any lesions seen in order to alert the diagnostician.

At least 10 volumes of formalin per volume of tissue is needed for proper fixation. Tissues should be allowed to fix for at least 24 hours, preferably 3 days. The brain in particular requires ample fixative over 3–4 days. Addition of 1 part ethanol to 9 parts formalin (10% ethanol, 90% formalin) may be used to prevent freezing in cold climates.⁴⁵ If formalin is not available, 70% ethanol may be used. Tissues may be sent in formalin, otherwise fix tissues for 24 hours, drain, and send in bags with formalin-moistened gauze. Be sure to double-bag samples as containers may rupture in transit. Contact your diagnostic laboratory for instructions.

For blood samples, blood from the dam as well as the fetus is ideal. Fresh blood smears are particularly useful if anaplasmosis is suspected. Blood in EDTA is preferred for PCR testing. For serum samples, draw 8–10 mL of serum into tubes without additives, allow to sit for 1–2 hours until the clot begins to retract, then place in the refrigerator overnight. Centrifuge at 1000 × g for 10 min and decant serum; send in a separate sterile tube. One may freeze the serum (do not freeze whole blood) until needed.

Table 54.5 lists typical sample storage conditions.

Diagnosis

Arrival at an accurate diagnosis is the result of the cooperative efforts of the attending veterinarian and the diagnosticians at the laboratory. Even with the submission of proper samples, the establishment of a definitive diagnosis is problematic. Expulsion of a dead fetus often occurs hours to days after death; the resulting autolysis makes the difficult task of fetal lesion identification even more so. Fetal membranes, which may have the only significant lesions to be found, are often not present for examination. Also, in many cases examined tissues are severely contaminated, whether due to incomplete expulsion or from the environment. Finally, diagnostic laboratories run routine tests to identify well-known causes of abortion in a timely manner. As such, investigation of unusual or poorly understood etiologies is beyond their purview.¹⁷

Keep in mind that the majority of pregnancy wastage occurs at earlier stages of development, when expelled tissues are least likely to be noticed. Early gestation abortions are often detected only weeks after the fact, making investigation and diagnosis all but impossible. The majority of tissues examined by clinicians and diagnostic laboratories are of large fetuses, which may bias the interpretation of a herd problem where losses occur at various gestation stages. Late-stage abortions may be the result of injury many weeks prior to expulsion; they may not exhibit important or telling changes that could be present in earlier aborted fetuses, preventing accurate diagnosis. Worse yet, in an effort to find a cause, other coincident agents may be erroneously blamed, depending on what is found in the submitted fetus.⁶

It is important to realize that more than one disease or management problem may be at work in a herd, complicating the resolution of endemic problems. Multiple pathogens may be present; some may not be obvious while those that are readily evident may not be directly contributing to abortion. In these cases, determining the actual cause of abortion is challenging.

Serology results must be cautiously interpreted. A positive sample from a cow indicates exposure, which may or may not be relevant in light of vaccination status. Twofold to fourfold increases in titer over 2 weeks may be significant, particularly with herdmate samples for comparison.⁶ However, it is likely that dam serum antibody levels have reached their peak by the time abortion occurs, so it is not unexpected to see a lack of rise in titer 2–3 weeks later.²⁴ Serology may be particularly useful in diagnosing BVDV, leptospirosis, anaplasmosis, and abortions where the agent is difficult to demonstrate by other methods.¹⁷ The presence of a specific antibody in fetal serum does not mean that particular agent is the cause of abortion; BVDV and Neospora caninum are two examples. Serology results must be carefully weighed with microbiology, histopathology, and herd history data. Table 54.6 lists common diagnostic tests for specific pathogens.

Mummification

Mummification is an uncommon event in which there is often no certain cause. It is the process of progressive dehydration and compaction of a sufficiently mature dead fetus in utero. In order for this to occur, bacterial infection (tissue lytic organisms) of the dead fetus must be absent, no air can be present in the uterus (closed cervix), and a functional CL must be present. The time course, which may take months, depends on the size of the fetus. This process generally occurs in the second and third trimesters, when fetal bones are sufficiently developed to resist resorption. As fetal fluids are absorbed, tissues become compressed, shrunken, red/brown, and sticky to dry, without odor. As the membranes atrophy and the uterus involutes, hemorrhage occurs between the uterus and fetal tissues, imparting the red “hematic” sticky material onto the tissues (Figures 54.15 and 54.16). This hematic type of mummification apparently is distinctly bovine.^11,27

The uterus tightly wraps around the fetal materials, compacting the mummy. The longer this proceeds, the drier and firmer the fetus becomes, remaining in utero for as much as 2 years. In the cow, mummification is most common at the end of the first and beginning of the second trimesters.⁴⁶ Various infectious and noninfectious etiologies have been implicated in their occurrence (BVDV, tritrichomoniasis), but the cause is usually unknown.^10,16 Mummification may be associated with certain breeds (Guernsey, Jersey) or certain breeding/genetic combinations.¹¹ In cases of uncomplicated mummification, prognosis is good for return to fertility.²⁷

Maceration

Maceration describes the effect of dead fetal tissue softening due to fluid soaking and saprophytic digestion in utero (Figure 54.17). In cattle this is due to exposure of the retained fetal tissues to bacteria. This occurs when there is failure of fetal expulsion after death with retention of the carcass partially or entirely within the uterus. The bacteria may be those that caused fetal death or those that arrive through an open cervix.¹⁰ Maceration is distinct from autolysis, which is digestion of tissues by endogenous enzymes and requires no bacteria. When bacteria gain access to a dead fetus in the reproductive tract, they are able to multiply rapidly at body temperature and are relatively insulated from the dam’s immune system. Bacterial digestion of tissues and gas formation (emphysema) ensues. The uterus surrounds the fetus and there is an intense metritis.