Marked subcutaneous mast cell and eosinophilic infiltration associated with the presence of multiple Dirofilaria repens microfilariae in 4 dogs

Dirofilaria repens is a parasitic nematode in the subcutaneous tissue of carnivores, including dogs and cats, transmitted by mosquitoes. Human beings may be accidental hosts. Infection of a dog with D repens was first reported in Palestine in 1934, and 2 additional cases were reported in dogs in Israel to date. This report describes D repens infection in 4 dogs in Israel that presented with subcutaneous masses, which were cytologically characterized by marked mast cell and eosinophil infiltration. In 3 cases, multiple microfilariae were present in the lesions; rare microfilariae were present in the 4^th case. In all 4 dogs, PCR of fine‐needle aspirates from the lesions were positive for D repens. The mast cells observed in all lesions were uniform and highly granulated, and with the presence of the microfilariae, a mast cell tumor was considered less likely. This report suggests that D repens infection‐associated subcutaneous lesions, characterized cytologically by massive mast cell and eosinophil infiltration, should be considered a differential diagnosis for mast cell tumor, especially in geographic locations endemic for this nematode. Notably, all 4 dogs were infected with D repens despite a routine prophylactic doramectin therapy administered every 3 months, probably due to the relatively long time interval between treatments.     

Dirofilaria repens—An etiological factor or an incidental finding in cytologic and histopathologic biopsies from dogs

Dirofilaria repens is an endemic, zoonotic parasite of carnivores, causing subcutaneous dirofilariasis, which is mostly asymptomatic. The aim of this study was to describe 22 cases of canine subcutaneous dirofilariasis. The cytologic and histopathologic samples were collected from dogs, which presented with various clinical signs such as cutaneous/subcutaneous nodules, hydropericardium, ascites, and lymphadenomegaly. The cytologic or histopathologic examination revealed purulent, pyogranulomatous, granulomatous or eosinophilic dermatitis/panniculitis, and the presence of D repens microfilariae or adults. The microfilariae or adults were also found incidentally in neoplastic cutaneous or subcutaneous tumors and in a sialocele. For the first time, microfilariae were also detected and described in pericardial and abdominal effusions and in enlarged reactive lymph nodes. Although it is well known that D repens can cause dermatitis and panniculitis in dogs, no previous reports of D repens microfilariae in body cavity fluids were found. The role of this parasite in the accumulation of body cavity fluid or in reactive lymphadenomegaly requires further investigation. Due to its zoonotic potential, and apparently underestimated pathogenicity, each case of canine subcutaneous dirofilariasis should be treated.     

Precision Medicine in Cats: Novel Niemann‐Pick Type C1 Diagnosed by Whole‐Genome Sequencing

State‐of‐the‐art health care includes genome sequencing of the patient to identify genetic variants that contribute to either the cause of their malady or variants that can be targeted to improve treatment. The goal was to introduce state‐of‐the‐art health care to cats using genomics and a precision medicine approach. To test the feasibility of a precision medicine approach in domestic cats, a single cat that presented to the University of Missouri, Veterinary Health Center with an undiagnosed neurologic disease was whole‐genome sequenced. The DNA variants from the cat were compared to the DNA variant database produced by the 99 Lives Cat Genome Sequencing Consortium. Approximately 25× genomic coverage was produced for the cat. A predicted p.H441P missense mutation was identified in NPC1, the gene causing Niemann‐Pick type C1 on cat chromosome D3.47456793 caused by an adenine‐to‐cytosine transversion, c.1322A>C. The cat was homozygous for the variant. The variant was not identified in any other 73 domestic and 9 wild felids in the sequence database or 190 additionally genotyped cats of various breeds. The successful effort suggested precision medicine is feasible for cats and other undiagnosed cats may benefit from a genomic analysis approach. The 99 Lives DNA variant database was sufficient but would benefit from additional cat sequences. Other cats with the mutation may be identified and could be introduced as a new biomedical model for NPC1. A genetic test could eliminate the disease variant from the population.     

Parasitemia due to Sarcocystis neurona‐like infection in a clinically ill domestic cat

An 8‐year‐old, 6‐kg, male neutered Domestic Shorthair cat was presented to The Ohio State University Veterinary Medical Center (OSU‐VMC) for difficulty breathing. Physical examination and thoracic radiographs indicated pneumonia, a soft‐tissue mass in the left caudal lung lobe, and diffuse pleural effusion. The effusion was classified as modified transudate. Rare extracellular elongated (~5–7 μm × 1–2 μm) zoites with a central round to oval‐shaped purple to deep purple vesicular nucleus with coarsely stippled chromatin and light blue cytoplasm were seen on a peripheral blood smear. Serum IgG and IgM were positive for Sarcocystis sp. antibodies and negative for Toxoplasma gondii antibodies, suggesting that the infection was acute rather than a recrudescence of prior infection. This organism was most consistent with either Sarcocystis neurona or Sarcocystis dasypi based on DNA sequence analysis of PCR products using COC ssRNA, ITS‐1, snSAG2, and JNB25/JD396 primer sets. This is the first report to visualize by light microscopy circulating Sarcocystis sp. merozoites in the peripheral blood of a domestic cat. Therefore, Sarcocystis should be considered as a differential diagnosis in cats with suspected systemic protozoal infection.     

Sarcocystid organisms found in bile from a dog with acute hepatitis: a case report and review of intestinal and hepatobiliary Sarcocystidae infections in dogs and cats

Sarcocystidae is a family of coccidian protozoa from the phylum Apicomplexa that includes Toxoplasma, Neospora, Sarcocystis, Hammondia, and Besnoitia spp. All species undergo a 2‐host sexual and asexual cycle. In the definitive host, replication is enteroepithelial, and infection is typically asymptomatic or less commonly causes mild diarrhea. Clinical disease is most frequently observed in the intermediate host, often as an aberrant infection, and is mostly associated with neurologic, muscular, or hepatic inflammation. Here, we review the literature regarding intestinal Sarcocystidae infections in dogs and cats, with emphasis on the life cycle stages and the available diagnostic assays and their limitations. We also report the diagnostic findings for an 11‐year‐old dog with acute neutrophilic hepatitis, biliary protozoa, and negative biliary culture. Although Toxoplasma and Neospora IgG titers were both high, PCR for these 2 organisms was negative for bile. The organisms were identified by 18S rDNA PCR as most consistent with Hammondia, either H heydorni or H triffittae. This is the first report of presumed Hammondia organisms being found in canine bile.     

Dirofilaria repens infection in a dog imported to Norway

Dirofilaria repens infection was diagnosed in a dog that had been imported to Norway from Hungary three years previously. The dog was a four-year-old castrated male mixed-breed dog and presented for examination of two masses on the right thoracic wall. Fine needle sampling from the subcutaneous nodules and subsequent cytological examination revealed a high number of microfilariae and a pyogranulomatous inflammation. At re-examination approximately 3 weeks later, both masses had apparently disappeared spontaneously, based on both inspection and palpation. However, examination of peripheral blood by a modified Knott’s test revealed a high number of unsheathed microfilariae with mean length of 360 μm and mean width of 6-7 μm, often with the classic umbrella handle appearance of D. repens. Polymerase chain reaction and sequencing confirmed the D. repens diagnosis. Subcutaneous dirofilariosis caused by D. repens is probably the most common cause of human zoonotic dirofilariosis in Europe, but currently is rarely encountered in northern countries such as Norway. However, travelling, import and relocation of dogs have increased, and thus the geographical range of these parasites is likely to increase from traditionally endemic southern regions. Increasing numbers of autochthonous cases of D. repens infections in dogs have been reported in eastern and central Europe. Although infection with D. repens often induces only mild signs or subclinical infections in dogs, they nevertheless represent a reservoir for zoonotic transmission and thus a public health concern, and, in addition, due to the long prepatent period and the high frequency of subclinical infections or infections with unspecific clinical signs, could easily be missed. Lack of experience and expectation of these parasites may mean that infection is underdiagnosed in veterinary clinics in northern countries. Also, predicted climate changes suggest that conditions in some countries where this infection is currently not endemic are likely to become more suitable for development in the intermediate host, and thus the establishment of the infection in new areas.     

Subconjunctival Dirofilaria repens infection in a dog resident in the UK

Dirofilaria repens infection was diagnosed in a 5‐year‐old female German shepherd crossbreed, originally from Romania but brought into the UK in February 2014. The dog presented with conjunctivitis in March 2014 and then again 2 months later with additional ocular and nasal mucopurulent discharge. Bacterial cultures from the nasolacrimal duct were negative for bacterial growth. The case was referred in August 2014 for ophthalmic examination, which revealed abnormalities in both eyes, especially the left. They included mild palpebral conjunctival hyperaemia and marked follicular conjunctivitis, as well as a dorsonasal bulbar conjunctival mass. Serum biochemistry was unremarkable and a conjunctival biopsy taken from the dorsonasal bulbar conjunctival mass revealed eosinophilic/lymphoplasmacytic conjunctivitis. At re‐examination, nematodes were found in the area of the previous biopsy site and in the ventral palpebral conjunctival fornix. Polymerase chain reaction and sequencing confirmed these to be D. repens. Treatment with 10% imidacloprid and 2·5% moxidectin (Advocate Spot‐On) was successful, and clinical signs resolved over a 6‐week period. This case report indicates that D. repens infection should be considered as a possible aetiological cause of ocular lesions in dogs in the UK, especially those with a history of foreign travel. Implications for establishment and spread of D. repens in the UK are discussed.     

Atypical actinobacillosis affecting hind limbs and lungs in a single beef cattle herd

Actinobacillosis usually is a sporadic infection that affects the tongue in cattle (“wooden tongue”) with possible spread to the digestive tract. Two 4‐year‐old Rouge‐des‐Prés cows from a single French beef herd were referred for chronic (2‐6 months) swelling and cutaneous nodules in the distal hind limbs. In addition to cutaneous signs, physical examination disclosed cachexia, lameness, lymphadenitis of the hind limbs, and pneumonia in both cows. Cytologic examination of direct skin smears was inconclusive, and no parasites were observed in examination of multiple skin scrapings. Histopathological examination of skin and lung biopsy specimens identified chronic, diffuse, severe pyogranulomatous dermatitis, associated with Splendore‐Hoeppli phenomenon and intralesional Gram‐negative bacteria. Cultures from skin, lymph nodes, and lungs (both cows were euthanized for welfare reasons) identified a Pasteurellaceae organism, confirmed as Actinobacillus lignieresii by partial sequencing of the rpoB gene. This report emphasizes that actinobacillosis can appear as a small outbreak in cattle with cutaneous and respiratory signs.     

Isolation of a Campylobacter lanienae‐like Bacterium from Laboratory Chinchillas (Chinchilla laniger)

Routine necropsies of 27 asymptomatic juvenile chinchillas revealed a high prevalence of gastric ulcers with microscopic lymphoplasmacytic gastroenteritis and typhlocolitis. Polymerase chain reaction (PCR) analysis using Campylobacter genus‐specific partial 16S rRNA primers revealed the presence of Campylobacter spp. DNA in the faeces of 12 of 27 animals (44.4%). Species‐specific partial 16S rRNA PCR and sequencing confirmed that these animals were colonized with Campylobacter lanienae, a gram‐negative, microaerophilic bacterium that was first identified on routine faecal screening of slaughterhouse employees and subsequently isolated from faeces of livestock. Campylobacter lanienae was isolated from the faeces of six PCR‐positive animals and identified with species‐specific PCR and full 16S rRNA sequencing. Phylogenetic analysis showed that these isolates clustered with C. lanienae strain NCTC 13004. PCR analysis of DNA extracted from gastrointestinal tissues revealed the presence of C. lanienae DNA in the caecum and colon of these chinchillas. Gastrointestinal lesions were scored and compared between C. lanienae‐positive and C. lanienae‐negative animals. There was no correlation between colonization status and lesion severity in the stomach, liver, duodenum, or colon. Possible routes of C. lanienae infection in chinchillas could include waterborne transmission and faecal–oral transmission from wild mice and rats or livestock. Based on these findings, the authors conclude that C. lanienae colonizes the lower bowel of chinchillas in the absence of clinical disease. This is the first report of C. lanienae in any rodent species. Campylobacter lanienae isolates from different mammalian species demonstrate heterogeneity by 16S rRNA sequence comparison. Analysis using rpoB suggests that isolates and clones currently identified as C. lanienae may represent multiple species or subspecies.     

Canine ocular thelaziosis caused by Thelazia callipaeda in Portugal

Ocular thelaziosis caused by Thelazia callipaeda is a vector‐borne disease affecting dogs and humans. We report a case of thelaziosis in a 10‐year‐old German Shepherd dog from Vila Real city (Portugal). Ophthalmological examination revealed bulbar and nictitating membrane conjunctival hyperemia with serous discharge noted at the left medial canthus and blepharitis. Schirmer tear test value and intraocular pressure were within the reference ranges in both eyes, and biomicroscopy showed a transparent cornea without lesions or edema and no inflammatory reaction in the anterior chamber. No funduscopic alterations were detected by direct and indirect ophthalmoscopic examination. When testing the nasolacrimal patency, two white worms were observed on the caruncle conjunctival surface with undulating movements that increased with light intensity. In total, eight worms were collected and morphologically identified as T. callipaeda (seven mature females and one male). PCR amplification of a 689 sequence of partial cytochrome c oxidase subunit 1 target gene confirmed the nematodes were T. callipaeda, haplotype 1. The dog was treated with a single subcutaneous injection of ivermectin combined with additional topical application of ophthalmic fusidic acid drops and oral milbemycin oxime. One week after treatment, no worms were detected and the ocular clinical signs resolved. The most recent reports of canine thelaziosis in the Iberian Peninsula should alert local health authorities to the zoonotic potential of infestation with T. callipaeda, which should be included in the differential diagnosis of conjunctivitis in dogs and humans.     

First molecular detection of Dirofilaria immitis (Spirurida: Onchocercidae) infection in dogs from Northern Algeria

Dirofilaria immitis and Dirofilaria repens are mosquito-borne filarioid nematodes that affect dogs and other domestic and wild carnivores, causing heartworm disease and subcutaneous dirofilariosis, respectively. In Algeria, the data about the epidemiology of these infections is largely unknown. The present study was designed to establish the occurrence of D. immitis and D. repens in dogs in Algeria using molecular tools.In 2014 and 2015, a total of 209 dogs over one year of age of different breed and sex, living in Northern Algeria, were examined and blood samples were collected from each dog. The presence of D. immitis and D. repens in these samples was detected by real-time PCR followed by standard PCR and sequencing. Overall, the blood of 209 dogs from two departments was collected and only 3 (1.4%) of the blood samples were found positive for D. immitis DNA. Sequencing of the corresponding amplicon displayed a 99.8% identity to D. immitis, confirming the presence of this mosquito-borne nematode in Algeria. Furthermore, all tested samples were negative for D. repens.     

Novel Borrelia genotypes in bats from the Macaregua Cave,Colombia

Bats have been implicated as reservoirs of relapsing fever group spirochaetes since the beginning of the last century. Recently, bat‐associated spirochaetes have been reported as human pathogens. In 1968, a spirochaete was detected in blood of the bat Natalus tumidirostris captured inside the Macaregua cave, Colombia. Data on this microorganism were never published again. The aim of this study was to evaluate the presence of Borrelia DNA in blood from bats of Macaregua cave. We performed molecular analyses using a genus‐specific real‐time PCR targeting the 16S rRNA to detect DNA of Borrelia in blood samples from 46 bats captured in the Macaregua cave. Positive samples were submitted to a battery of PCRs aiming to amply Borrelia 16S rRNA, flaB, glpQ, p66, ospC, clpA, clpX, nifS, pepX, pyrG, recG, rplB and uvrA genes. Seventeen samples were positive for Borrelia after real‐time PCR. With the exception of flaB gene, attempts to amplify further loci were unsuccessful. Nucleotide and amino acid divergences of four flaB haplotypes characterized from blood of Carollia perspicillata showed Borrelia burgdorferi sensu lato (Bbsl) as the most closely related group. A phylogenetic tree including 74 sequences of the genus confirmed this trend, since Borrelia genotypes detected in bats from Macaregua formed a monophyletic group basally positioned to Bbsl. Our results suggest that Borrelia genotypes characterized from bats roosting in the Macaregua cave might constitute a new taxon within the genus. This is the first molecular characterization of a Borrelia sp. in Colombia.     

Subcutaneous and cardiopulmonary dirofilariasis in a dog

Canine subcutaneous dirofilariasis is rare and typically caused by Dirofilaria repens. An adult male, intact, mixed breed dog from South Carolina, USA, was examined because of physical trauma to the right maxilla. The dog died during hospitalization and was submitted for necropsy. Gross post-mortem changes included approximately 150 adult nematodes morphologically consistent with Dirofilaria spp. in the pulmonary artery, right atrium, and right ventricle. Histologically there was widespread proliferative pulmonary endarteritis with intraluminal nematodes morphologically consistent with Dirofilaria spp. Four similar nematodes were present in the subcutaneous tissue of the left medial thigh and tibial area. These nematodes were located within the fascia and skeletal muscles. They were surrounded by epithelioid macrophages and multinucleated giant cells, with some lymphocytes and plasma cells, and areas of mineralization (interpreted as mineralized cross sections of nematodes). Nematodes were morphologically identified as D. immitis. Subcutaneous dirofilariasis caused by D. immitis occurs when migrating nematode larvae develop into adults in the subcutaneous and skeletal muscle tissues.Key clinical message:Subcutaneous dirofilariasis in dogs is rare and most often associated with Dirofilaria repens. This case highlights the pathology findings of subcutaneous D. immitis in a dog.     

Cytologic identification of fungal arthritis in a Labrador Retriever with disseminated Talaromyces helicus infection

An 8‐year‐old, neutered male Labrador Retriever presented with acute forelimb lameness. Clinical signs progressed over one week. On physical examination, right cubital joint effusion and bilateral axillary lymphadenomegaly were noted, and severe internal lymphadenomegaly was observed ultrasonographically. Granulomatous lymphadenitis with intralesional fungi was noted cytologically, and the dog was ultimately diagnosed with disseminated Talaromyces helicus infection via PCR of a pure isolate. Extensive medical therapy was pursued, and months later, an arthrocentesis was performed due to continued lameness and severe cubital joint effusion. The synovial fluid contained increased numbers of neutrophils, macrophages, and multinucleated giant cells. Frequent fungal hyphae were found both intracellularly and extracellularly. These basophilic organisms were 2‐4 µm in width with internal eosinophilic granules, roughly parallel walls, and occasional to frequent septa. Round to oval yeast‐like forms with thin, clear halos were also occasionally identified. Due to the severity of clinical signs, the right thoracic limb was amputated. Histologic examination of the cubital joint revealed marked granulomatous synovitis, fasciitis, panniculitis, and osteomyelitis, all with intralesional fungi. Talaromyces helicus is a very rare cause of disease, reported only in one other dog. Granulomatous lymphadenitis appears to be a feature of this disease, but this report is the first to describe a significant synovial component.     

Use of real‐time quantitative PCR to document successful treatment of Mycoplasma haemocanis infection with doxycycline in a dog

An 8‐year‐old Jack Russell Terrier was diagnosed with hemolytic anemia caused by hemoplasmosis 4 years following splenectomy. Quantitative real‐time PCR (qPCR) analysis was used initially to confirm infection with Mycoplasma haemocanis and subsequently to monitor and direct medical therapy. Doxycycline was administered beyond resolution of clinical signs until hemoplasma DNA could no longer be detected by qPCR. The dog remained clinically healthy and hemoplasma‐negative 8 months following cessation of therapy. Canine hemoplasmosis should remain as a differential diagnosis for hemolytic anemia in dogs, particularly those that are splenectomized or immunocompromised, even in geographic regions where prevalence of infection is low. Prolonged doxycycline administration has been shown by qPCR to lead to sustained absence of detectable infection and should be considered as a first line treatment for canine hemoplasmosis.     

Parasites observed in the proximal alimentary tract of a Przewalski's horse in China

This article reports a Przewalski's horse from the Kalamaili Nature Reserve (KNR) in China transported to our institute to determine the cause of death. The animal had been subjected to necropsy before being transported to our institute; the entire digestive tract was removed but not opened to reduce the possible loss of parasites. During examination, ascarids in the intestines and bots in the oral cavity and stomach were observed. The parasites were identified according to an identification key and enumerated. The ascarids were identified as Parascaris spp. and were associated with a volvulus, resulting in intestinal obstruction. A total of two (stomach), one (duodenum) and 106 (intestines) Parascaris spp. were counted. Bots in the digestive tract were identified as third stage larvae of Gasterophilus pecorum (G. pecorum), Gasterophilus nigricornis (G. nigricornis) and Gasterophilus nasalis (G. nasalis), which may have been associated with stomach ulcers suffered by the horse. Attached to the lining of the oral cavity and stomach were 331 and 2014, respectively, larvae of G. pecorum, while in the duodenum there were 23 third stage larvae of G. nigricornis and 20 third stage larvae of G. nasalis. Therefore, the possibility of the presence of these parasites should not be overlooked in Przewalski horses as these parasites may be responsible for severe clinical signs in infected animals.     

Guttural pouch empyema caused by Corynebacterium pseudotuberculosis in a pregnant mare

During medical management of mild colic in a 12‐year‐old Quarter Horse, mid‐gestation mare, unilateral purulent nasal discharge from the right nostril was noted. Endoscopic examination revealed guttural pouch empyema. Culture was positive for Corynebacterium pseudotuberculosis and negative for Streptococcus equi ssp. equi. A synergistic haemolysis inhibition titre of 1024 was consistent with C. pseudotuberculosis infection. Treatment included serial lavages and local infusion of antibiotics into the guttural pouches along with a 6‐week course of oral trimethoprim–sulfamethoxazole and rifampicin. Overall, no additional sites of infection were identified and the mare responded well to treatment, delivering a healthy, full‐term foal. This case emphasises that C. pseudotuberculosis, although uncommon, should be considered as a differential for guttural pouch empyema.     

Administration of L. salivarius expressing 3D8 scFv as a feed additive improved the growth performance,immune homeostasis,and gut microbiota of chickens

Probiotics have been defined as live microorganisms that are administered in an appropriate amount to provide health benefits to the host animal. In this study, we investigated the effect of L. salivarius DJ‐sa‐01 secreting the 3D8 single‐chain variable fragment (3D8 scFv) on the growth performance, cytokine secretion, and intestinal microbial flora of chickens. The experiment was divided into the control group and L. salivarius expressing 3D8 scFv experimental group. Chicken was fed 10⁹ colony‐forming units (CFUs) of wild‐type (WT) L. salivarius or 3D8 scFv‐secreting L. salivarius daily for 35 days. The administration of L. salivarius expressing 3D8 scFv significantly improved the body weight of chickens compared with the administration of WT L. salivarius. A 16S ribosomal RNA metagenomic analysis showed that Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes were the dominant phyla in both experimental groups. At the genus level, Lactobacillus was more abundant (22.82%) in the L. salivarius/3D8 group compared with the WT L. salivarius group. The serum levels of cytokines, such as IL‐8, TNF‐α, IL‐1β, IFN‐γ, IL‐4, and IGF1, were significantly reduced in the L. salivarius/3D8‐treated chickens. In summary, our results suggest that L. salivarius expressing 3D8 scFv could be considered a feed additive for improving the growth performance, immune function, and disease resistance of poultry.     

Nodular granulomatous glossitis as the sole clinical sign in canine leishmaniosis

A 5.5‐year‐old, intact male Rottweiler dog was admitted with a history of multifocal nodular tongue lesions which progressively deteriorated during the previous year. Physical examination revealed several reddish nodules with central depression on the surface of the tongue in an otherwise healthy dog. Clinicopathologic abnormalities included eosinophilia and hyperproteinemia. Lingual nodule cytopathology, histopathology, and immunohistochemistry revealed Leishmania spp. amastigotes and a severe granulomatous glossitis. The dog was also seroreactive to L infantum antigens by an indirect immunofluorescence assay. Clinical reevaluation 3 months after the institution of treatment with allopurinol and miltefosine indicated that the nodular lesions had completely regressed. In endemic areas, lingual nodular lesions may rarely be the sole clinical sign of canine leishmaniosis. Standard medical treatment may provide an excellent prognosis.