An apparent case of brodifacoum toxicosis in a whelping dog

Abstract

A 7-y-old Weimaraner bitch was presented to emergency service after 3 h of active labor with no puppies produced. Hemoabdomen and hemothorax were present at the time of surgery; prothrombin time (PT) and activated partial thromboplastin time (aPTT) were both found to be within normal ranges. Surgical cesarean section was performed; 4 dead puppies and 5 live puppies were delivered. Because hemostasis was difficult to achieve, a hysterectomy was performed; however, the dog died as the operation was being completed. At autopsy, the pleural cavity contained 1.5 L of unclotted blood; the peritoneal cavity was relatively normal, and no obvious hemorrhage was associated with the surgical sites. All 4 dead fetuses were opened, and their pleural cavities were filled with unclotted blood. An anticoagulant screen was performed, and brodifacoum was identified in the liver of the bitch. This case is unusual in that the PT and aPTT were within reference intervals, but brodifacoum was present in sufficient amounts to potentially result in this dog bleeding to death, and also is suspected to have crossed the placenta and caused hemothorax and death in 4 of 9 puppies in utero.

A 7-y-old Weimaraner bitch was presented to the Emergency Service, Small Animal Clinic, Veterinary Teaching Hospital, Michigan State University on June 11, 2016. The dog was in distress after 3 h of active unproductive labor at home. On presentation, the patient was markedly tachypneic with a restrictive breathing pattern. Thoracic radiographs were performed, which revealed suspected diaphragmatic hernia with a moderate amount of pleural effusion present. Abdominal radiographs confirmed questionable integrity of the diaphragm and the presence of 9 fetal skeletons.

Additional testing was performed to assess for signs of fetal distress and maternal factors precipitating dystocia. Blood gas analysis, packed cell volume and total solids (PCV/TS), prothrombin time and activated partial thromboplastin time (PT/aPTT; Automate 5 system, Diagnostics Stago, Parsippany, NJ), blood typing, and an abdominal focused assessment using sonography for trauma (AFAST) scan were performed. Blood gas levels were unremarkable. The patient was anemic with a PCV of 0.29 L/L and total solids of 55 g/L. PT was 13 s (reference interval [RI]: 11–17 s), and aPTT was 80 s (RI: 72–102 s). AFAST was concerning for fetal distress, with multiple fetal heart rates noted to be <180 beats per min. Cesarean section was recommended.

Upon entering the peritoneal cavity, a moderate amount of peritoneal hemorrhage was discovered. Cesarean section was performed; however, difficulty in stopping the bleeding from the uterine incision resulted in the decision to perform an ovariohysterectomy. The surgery resulted in the delivery of 5 live and 4 dead puppies. Shortly thereafter, the dam went into cardiopulmonary arrest. The diaphragm was found to be intact so an incision was made and a moderate amount of unclotted blood was discovered in the thoracic cavity. Trans-diaphragmatic cardiac massage and manual ventilation were performed. Rescue drugs were administered, and 2 units of dog erythrocyte antigen 1.1 negative packed red blood cells were transfused. However, cardiopulmonary resuscitation was unsuccessful, with acute, massive hemorrhage as the suspected cause of death.

At autopsy, the dog had ~1.5 L of unclotted blood filling the pleural cavity; the peritoneal cavity contained no obvious hemorrhage, and no bleeding was associated with the ovariohysterectomy site and ligatures. The lungs were atelectatic, dark red, edematous, and heavy. No other gross anatomic lesions were noted. Liver was frozen for possible toxicology; placenta from several pups was collected for bacterial culture; and representative sections of a variety of tissues from the bitch were collected in formalin for histopathology. All 4 dead puppies had their body cavities opened, and all 4 thoraxes were filled with unclotted blood.

Lung sections had random multifocal alveolar septal fibrosis, and obliterative bronchiolitis affected low numbers of scattered bronchioles associated with fibrotic areas. The pleura of the lungs had diffuse mild reactive mesothelial cells, likely in response to the pleural hemorrhage. No other diagnostically significant lesions were found in the other tissues examined; specifically, the liver, spleen, and bone marrow had no lesions suggestive of degeneration, inflammation, or response to anemia.

Based on the gross and histologic findings, specifically the unclotted blood filling the pleural spaces of the bitch and all 4 dead pups, the normal active bone marrow in the bitch, and the lack of liver disease or obvious disseminated intravascular coagulation (DIC) histologically, a provisional diagnosis of anticoagulant poisoning was suggested. An anticoagulant screen was conducted on the frozen liver. The anticoagulant rodenticide screen performed was a modified high-performance liquid chromatography method.² The screen was positive for brodifacoum (0.024 mg/kg [0.011 mg/lb]). Other rodenticides included in this screen were negative at the detection limits (0.002 mg/kg [0.0009 mg/lb] for brodifacoum; 0.02 mg/kg [0.009 mg/lb] for bromadiolone, difenacoum, and warfarin; 0.07 mg/kg [0.03 mg/lb] for difethialone; 0.20 mg/kg [0.091 mg/lb] for chlorophacinone and diphacinone) of the assay. These results support exposure to brodifacoum in the bitch, and are suggestive of potential anticoagulant poisoning. The owner was contacted by the State Veterinarian’s office after toxicology results became available. One of the 5 living puppies had died of unrelated causes but it was recommended that the 4 remaining puppies receive plasma transfusions. The owner was unaware of any possible sources of rodenticide exposure. The puppies remained nonclinical for rodenticide poisoning before and after this treatment. They did not receive vitamin K. At the time of manuscript submission, the puppies were ~6-mo-old and reportedly healthy.

Differential diagnoses for the intrathoracic hemorrhage and difficult-to-control bleeding from the dam’s uterus, as well as blood-filled thoraxes in the 4 dead pups, would include genetic and inherited bleeding disorders such as von Willebrand disease (vWD), hemophilia A, hemophilia B, and various clotting factor disorders (II, VII, X, XI, XII). Based on the normal PT and aPTT on the bitch, all of these except vWD, which is generally associated with normal clotting times, should have been ruled out in the dam. Other differentials include acute blood loss caused by trauma (ruled out by AFAST and autopsy) or gastrointestinal ulcers, which were not present at autopsy; thrombocytopenia, which was not recognized in bone marrow samples and should also not have affected the puppies; blood parasites, which were not seen on peripheral blood smears; advanced liver disease resulting in decreased blood clotting factors, which was not present in histologic liver sections and ruled out by normal PT and aPTT times; and DIC, which was not recognized on blood work by the absence of increased fibrin degradation products, the lack of widespread intravascular fibrin thrombi in multiple tissues on histologic section, and the absence of prolonged PT and aPTT times.

The work-up on this bitch, both clinically and on postmortem examination pointed strongly toward an anticoagulant exposure, which was subsequently confirmed by the anticoagulant screen as having detectable concentration of brodifacoum. The other primary rule-out in this case is vWD, which is generally associated with normal PT and aPTT values unless it is accompanied by a concurrent deficiency of factor VIII.⁹ Unfortunately, the buccal mucosal bleeding time (BMBT) or a direct assay for von Willebrand factor were not run on the bitch while she was still alive to confirm or deny the presence of this disease. However, a thorough review of the literature on canine vWD did not reveal any reports of multiple affected puppies being aborted or dying at delivery as a result of hemothorax associated with the disease, leading us to suggest that brodifacoum toxicosis was the most likely cause of the dam and its pups’ deaths. Furthermore, Weimaraner is not one of the canine breeds that has a reported predisposition to vWD.

Anticoagulant rodenticides comprise the most common pesticide exposure for pets and are the most commonly inquired about toxins in United States veterinary practices.⁸ Anticoagulant rodenticides exert their poisonous effects by interfering with vitamin K metabolism, thus inhibiting the synthesis of clotting factors II, VII, IX, and X.³ Brodifacoum is a second-generation, coumarin-type, anticoagulant rodenticide that is commercially available for rodent control in and around public, commercial, industrial, and residential structures. Brodifacoum is 200 times more potent than the first-generation warfarin and has a markedly longer duration of action.¹ The estimated half-life of brodifacoum in dogs is ~4 mo,¹ whereas the elimination half-life for brodifacoum has been approximated to 6 ± 4 d.¹¹

A limited number of cases were identified in the literature regarding brodifacoum poisoning during pregnancy in humans and dogs.^5,6,7,13 Cases had mixed outcomes ranging from successful treatment of the mother or dam with vitamin K,^5,13 to abortion and fetal intoxication.^6,7 There is clear evidence for the transplacental movement of brodifacoum from the literature that helps explain the hemothorax lesions found in the dead puppies in our report.

PT and aPTT assays are often used as screening tests and surrogate markers of coagulation factor levels in trauma settings.¹² PT assesses the function of the extrinsic system, whereas aPTT measures the activity of the intrinsic pathway. In our case, both PT and aPTT were normal, in spite of clear gross evidence for a coagulopathy consistent with anticoagulant rodenticide exposure. False-negative results may occur for both PT and aPTT. The aPTT screen is very sensitive to the levels of factor VIII, an acute-phase protein. If factor VIII levels are raised, then aPTT may be misleadingly short and not accurately reflect the degree of anticoagulation.¹⁰ Moreover, any difficulties in the collection of samples that leads to activation of coagulation within the collection tube may result in false-negative parameters for both aPTT and PT. This may occur when blood is slow to fill the collection container, there is considerable manipulation of a vein by the needle, samples are incompletely mixed following collection, or the tube is underfilled.⁴

Our case serves as a warning for clinicians to strongly consider anticoagulant rodenticide exposure, especially caused by brodifacoum, in pregnant dogs and their unborn or newly born puppies, when other common causes of bleeding have been ruled out. It further points out that PT and aPTT assays will not always be able to detect anticoagulant rodenticide exposure. Additionally, this case serves as a reminder that modern generation anticoagulants may have the ability to cross the placenta and affect both the pregnant adult as well as the puppies in utero.