Journal of Veterinary Diagnostic Investigation最新文献

African swine fever (ASF) is a high-consequence transboundary animal disease caused by African swine fever virus (ASFV). Given that vaccines are not widely available, ASFV detection, including by molecular and serologic assays, is paramount to efficacious control and mitigation of ASF. ASFV-specific antibodies can be detected as early as 7-10 d postinfection in infected animals and may persist for several months or longer. Accurate detection of ASFV-specific antibody is critical for the identification of chronically infected, subclinically infected, or recovered animals. ELISAs are commonly used for the rapid screening of large numbers of animals for ASFV antibodies. The World Organisation for Animal Health recommends that ELISA-positive results should be confirmed with a second serologic method, such as an indirect immunofluorescent assay, indirect immunoperoxidase test (IPT), or immunoblot test. Commercial kits are not available for those tests. We developed and validated an in-house IPT by using a currently circulating genotype II ASFV strain as antigen. The sensitivity and specificity of the in-house IPT are comparable to the reference IPT developed by an international ASFV reference laboratory and superior to a commercial blocking ELISA.

Piglet lethality is one of the major concerns in pig breeding programs. Deletion of a 212-kb region within the Bardet-Biedl syndrome 9 (BBS9) gene has been linked to a reduction in the number of piglets born alive per litter. The BBS9 mutant gene carrier-by-carrier mating scheme could result in mummification of piglets carrying 2 copies of the BBS9 mutant allele, which ultimately affects the reproductive performance of the sow. Our aim was to develop a simple, rapid, and cost-efficient method that could be applied in a BBS9 mutant gene carrier screening program in low- and middle-income countries within basic laboratory settings. Here, we report an optimized multiplex PCR assay that we have established successfully for detection of a 212-kb deletion within the BBS9 genomic sequence. We genotyped 420 animals from Yorkshire, Duroc, and Landrace purebred populations in Vietnam. We found that while the BBS9 mutant allele was not identified in Duroc pigs, the frequency of BBS9 carriers was 10% in both Yorkshire and Landrace populations. We subsequently validated our results using Sanger sequencing. Our multiplex PCR method could be utilized as a BBS9 screening test in pig breeding programs.

Mass spectrometry (MS) has long been considered a cornerstone technique in analytical chemistry. However, the use of MS in animal health laboratories (AHLs) has been limited, however, largely because of the expense involved in purchasing and maintaining these systems. Nevertheless, since ~2020, the use of MS techniques has increased significantly in AHLs. As expected, developments in new instrumentation have shown significant benefits in veterinary analytical toxicology as well as bacteriology. Creative researchers continue to push the boundaries of MS analysis, and MS now promises to impact disciplines other than toxicology and bacteriology. I include a short discussion of MS instrumentation, more detailed discussions of the MS techniques introduced since ~2020, and a variety of new techniques that promise to bring the benefits of MS to disciplines such as virology and pathology.

Congenital structural anomalies of the lower airways of the respiratory tract are uncommon in cats. We describe here a case of cystic pulmonary lesions in a 6-wk-old domestic shorthair cat consistent with congenital pulmonary airway malformation (CPAM; formerly referred to as cystic adenomatoid malformation of the lung, or congenital pulmonary adenomatoid malformation; Stocker type II). CPAM is rarely reported in veterinary species and, to our knowledge, has not been reported in cats. In humans and veterinary species, individuals with CPAM (Stocker types I-IV) can be asymptomatic at birth but are predisposed to developing respiratory abnormalities that typically manifest clinically in the early years of life. We review the pathologic features of CPAM.

A free-ranging, adult female two-toed sloth (Choloepus hoffmanni) was brought to a wildlife rescue center in Costa Rica with ocular and auricular myiasis and numerous skin lesions. After one month of unsuccessful systemic and topical antimicrobial treatment, the patient died. A postmortem examination was performed, and tissues were examined histologically, confirming disseminated amebic infection with intralesional trophozoites and cysts in the lungs, liver, eye, heart, spleen, and stomach. Immunohistochemistry identified the ameba as Acanthamoeba sp. A multiplex real-time PCR assay, 18S ribosomal DNA PCR, and sequencing performed on formalin-fixed, paraffin-embedded lung tissue confirmed the Acanthamoeba T17 genotype. The Acanthamoeba genus is in the group of free-living amebas that cause infection in humans and animals, and it is ubiquitous in the environment. Acanthamoeba T17 has been isolated from water and soil, but to our knowledge, this genotype has not been implicated in infections of animals previously and has not been reported from Costa Rica. Systemic Acanthamoeba infection has not been described in sloths previously. We provide a comprehensive literature review describing infections by free-living amebas of the genus Acanthamoeba spp., Balamuthia spp., and Naegleria spp. in domestic, zoo, and wild mammals.

The incidence of enzootic bovine leukosis (EBL), a type of B-cell lymphoma, is increasing in Japan. EBL is caused by bovine leukemia virus (BLV; Retroviridae, Deltaretrovirus bovleu) infection and is diagnosed by detecting antibodies against BLV in milk and blood or BLV DNA in blood. We assessed the feasibility of using stable flies (Stomoxys calcitrans) as a sampling tool to assess BLV infection status in cattle herds. First, we collected blood from 3 cattle herds and, based on the measurement of BLV-proviral load (PVL) by quantitative real-time PCR (qPCR), identified 1) a BLV-free herd, 2) a herd with a low prevalence of BLV-infected cattle and low PVL, and 3) a herd wherein half of the cattle were BLV-infected with low-to-high PVLs. Next, we collected stable flies from the 3 herds, extracted DNA from their blood meals, analyzed it for BLV DNA, and measured the BLV PVL. Cattle DNA and BLV DNA, but not other mammalian DNA, were successfully detected by digestion of the flies. Based on fly blood meal qPCR, we identified one herd as BLV-free and the other 2 herds as having <50% prevalence of BLV-infected cattle with low PVLs. Our fly results were not consistent with preliminary BLV-PVL measurements on cattle blood. Our pilot study indicated that, to assess the feasibility of a stable fly blood meal test as an alternative technique for evaluating BLV infection status in dairy and beef cattle, additional investigations involving more cattle herds and stable flies are needed.

To expand surveillance testing capacity through sample pooling, a thorough understanding is needed of how sample dilution through pooling affects the sensitivity of candidate assays. We validated a robust and representative framework for assessing the dilution effect of sample pooling using duplex rtPCR surveillance of Theileria orientalis and Anaplasma marginale, both of which are causative agents of severe anemia in cattle and a serious threat to the cattle industry in Virginia and many other states. We used 200 known-positive samples with Ct values representative of typical surveillance results in a series of pools in which we re-tested each sample individually, followed by each sample diluted in equal volumes with negative samples to make pools of 2, 4, 6, 8, and 10 total samples. We compared the Ct values of the individual positives with the Ct values of each pool size to determine if Ct values increase past the limit of detection in the 45-cycle assay. We observed a maximum of 2% sensitivity loss (no more than 2 of 100 samples returned a false-negative result) for both T. orientalis and A. marginale during the pooling series, with lower-than-expected average Ct increase and sensitivity loss. We conclude that pooling up to 10 samples would be acceptable for regional surveillance of T. orientalis and A. marginale using our rtPCR assay. The described strategy is applicable to validate pooling for a wide range of single and duplex rtPCR assays, which could expand efficient disease surveillance.

Natural oak toxicity, a phenomenon sporadically reported in the United States, is due to consumption of any part of most oak trees (Quercus spp.). Ruminants, mainly cattle, are disproportionately susceptible to oak toxicity. Toxicity is attributed to degradation of the oak plant hydrolysable tannins by rumen microbes and enzymes into absorbable low-molecular-weight metabolites, which are postulated to bind and damage endothelial cells by unknown mechanisms. The clinical manifestations of acute toxicosis are nonspecific or broadly suggestive of renal disease due to acute tubular injury. Here we document the clinical, gross, histopathologic, and novel ultrastructural features of natural acute oak nephrotoxicity in 3 beef calves on 2 farms in Colorado, USA. Gross postmortem findings included perirenal edema with renomegaly and hemorrhagic gastroenteritis. Histologically, renal tubular epithelial necrosis was severe, with hemorrhage and intratubular hyaline casts. Transmission electron microscopy revealed extensive involvement of proximal and distal convoluted tubules, with predominantly intact basement membranes, and glomerular and interstitial endothelial injury and necrosis. The ultrastructural details of toxic nephropathy and vasculopathy induced by oak metabolites in natural cases of bovine oak toxicosis have not been described previously, to our knowledge.

A 2-y-old, intact female, mixed-breed dog was presented to the veterinary hospital with abdominal distension, anemia, and lethargy following a chronic history of nonspecific gastrointestinal signs. CBC and serum biochemistry revealed moderate nonregenerative anemia with neutrophilia, hypoalbuminemia, hyperglobulinemia, hypoglycemia, decreased urea and creatinine, and hypercholesterolemia. Abdominal radiographs and ultrasound revealed a large heterogeneous mesenteric mass and ascites. Abdominocentesis confirmed septic peritonitis with filamentous bacteria. Fine-needle aspiration of the mass yielded pyogranulomatous inflammation and hyphae. An exploratory laparotomy revealed a large cranial abdominal mass with granulomas present throughout the abdominal cavity. Due to the poor prognosis and disseminated disease, the owner elected euthanasia. Postmortem and histologic examinations detected intralesional mycetomas and bacterial colonies within the mesenteric masses. 16S ribosomal RNA gene PCR and sequencing using formalin-fixed, paraffin-embedded sections identified Nocardia yamanashiensis, Nocardioides cavernae, and Nocardioides zeicaulis. Fungal culture, PCR, and sequencing confirmed Scedosporium apiospermum. Our report highlights the importance of molecular methods in conjunction with culture and histologic findings for diagnosing coinfections caused by infrequent etiologic agents. Additionally, we provide a comprehensive literature review of Scedosporium apiospermum infections in dogs.

To prevent significant economic losses, some countries have successfully eradicated enzootic bovine leukosis (EBL), which is caused by bovine leukemia virus (BLV) infection. In Serbia, efforts to eliminate EBL commenced in the late 1990s. Recognizing the disparities in test selection among laboratories and variations in quality, we evaluated the diagnostic sensitivity and specificity of commercial ELISAs using field samples in Serbia. Using 5 commercial ELISA kits, we tested 138 cattle serum samples, submitted for confirmatory testing between 2020 and 2023, along with 100 serum samples from BLV-negative herds. We found 100% agreement of the ID Screen BLV Competition (IDvet), Svanovir BLV gp51-Ab (Svanova), and INgezim BLV Compac 2.0 (Ingenasa) ELISAs. We observed 93% agreement comparing these 3 kits to the Bovine Leukemia Virus Antibody test kit (VMRD). Agreements of 92% and 88.4% were determined between Idexx and IDvet, Svanova, and Ingenasa kits, and between Idexx and VMRD kits, respectively.