Molecular survey on vector-borne pathogens in clinically healthy stray cats in Zaragoza (Spain).

BACKGROUND: In Europe, feline vector-borne infections are gaining importance because of the changing climate, expanding habitats of potential vectors and expanding pathogen reservoirs. The main objective of this study was to assess the prevalence of vector-borne pathogens (VBPs) in stray cats in Zaragoza, Spain, and to investigate potential risk factors for infection, including feline leukaemia virus (FeLV) and feline immunodeficiency virus (FIV). METHODS: Blood samples from stray cats presented to the veterinary faculty in Zaragoza between February 2020 and 2022 were tested by polymerase chain reaction (PCR) for the presence of Anaplasma phagocytophilum, Anaplasma platys, Bartonella henselae, Ehrlichia canis, Rickettsia spp., haemotropic Mycoplasma spp., Hepatozoon spp., Leishmania infantum, piroplasms and microfilariae at the LABOKLIN laboratory. The cats were also tested for FeLV and FIV by PCR. RESULTS: Nearly half of the cats (158/332, 47.6%) were positive for at least one VBP. Hepatozoon spp. were detected in 25.6%, haemotropic Mycoplasma spp. in 22.9%, B. henselae in 9.3% and L. infantum in 2.1% of the cats. Male sex had a statistically significant association with test results for haemotropic Mycoplasma spp. (odds ratio 1.38 [1.21;1.57]); regionality with Hepatozoon spp., B. henseale and FIV; and seasonality with Hepatozoon spp., haemotropic Mycoplasma spp., L. infantum and FeLV (P ≤ 0.05 each). A strong positive correlation was reported for the amount of rainfall and the number of cats that tested positive for Hepatozoon spp. (ρ = 753, P = 0.05). None of the cats tested positive for A. phagocytophilum, A. platys, E. canis, Rickettsia spp., piroplasms, or microfilariae. Co-infections with multiple VBPs were detected in 56 out of 332 cats (16.9%). Thirty-one of the 332 cats included in the study (9.3%) tested positive for FeLV (6.9%) and for FIV (3.6%). In 20/31 cats (64.5%) that tested positive for FeLV/FIV, coinfections with VBP were detected (P = 0.048, OR 2.15 [0.99; 4.64]). CONCLUSIONS: VBPs were frequently detected in stray cats in Zaragoza. In particular, regionality and seasonality had a statistically significant association with PCR results for most VBPs included in the study.

White-Toothed Shrews (Genus Crocidura): Potential Reservoirs for Zoonotic Leptospira spp. and Arthropod-Borne Pathogens?

Three species of white-toothed shrews of the order Eulipotyphla are present in central Europe: the bicolored (Crocidura leucodon), greater (Crocidura russula) and lesser (Crocidura suaveolens) white-toothed shrews. Their precise distribution in Germany is ill-defined and little is known about them as reservoirs for zoonotic pathogens (Leptospira spp., Coxiella burnetii, Brucella spp., Anaplasma phagocytophilum, Babesia spp., Neoehrlichia mikurensis and Bartonella spp.). We investigated 372 Crocidura spp. from Germany (n = 341), Austria (n = 18), Luxembourg (n = 2) and Slovakia (n = 11). West European hedgehogs (Erinaceus europaeus) were added to compare the presence of pathogens in co-occurring insectivores. Crocidura russula were distributed mainly in western and C. suaveolens mainly in north-eastern Germany. Crocidura leucodon occurred in overlapping ranges with the other shrews. Leptospira spp. DNA was detected in 28/227 C. russula and 2/78 C. leucodon samples. Further characterization revealed that Leptospira kirschneri had a sequence type (ST) 100. Neoehrlichia mikurensis DNA was detected in spleen tissue from 2/213 C. russula samples. Hedgehogs carried DNA from L. kirschneri (ST 100), L. interrogans (ST 24), A. phagocytophilum and two Bartonella species. This study improves the knowledge of the current distribution of Crocidura shrews and identifies C. russula as carrier of Leptospira kirschneri. However, shrews seem to play little-to-no role in the circulation of the arthropod-borne pathogens investigated.

Feline leishmaniosis with focus on ocular manifestation: a case report.

BACKGROUND: In Europe, feline leishmaniosis is commonly caused by Leishmania infantum. There is little knowledge regarding pathogenesis, ocular manifestations and long-term follow-ups in cats with leishmaniosis. FINDINGS: A 6-year-old female, spayed European Shorthair cat was imported from Spain to Germany 2 years prior to its first clinical presentation. The cat showed lethargy, weight loss, ulcerative lesions on the front limbs and high-grade chronic uveitis. The diagnosis of L. infantum infection was based on the cytological finding of amastigotes in skin lesions, positive qPCR of EDTA-blood and positive PCR of a cyto-brush sample from the conjunctiva. Supportive findings included positive serology by IFAT, serum protein capillary electrophoresis with peaks in alpha2- and gamma-globulin sections and marked elevation of SAA. Enucleation had to be performed on day 288 on both eyes because of blindness, glaucoma and high-grade uveitis. Histologically, high numbers of Leishmania spp. amastigotes were found in histiocytes. IFAT and PCR were positive in the aqueous humor in both eyes, respectively. Feline leukemia virus antigen and feline immunodeficiency virus antibody testings were positive. Hematological and biochemical results revealed mild leukocytosis with lymphocytosis, monocytosis and eosinopenia as well as marked elevation of SAA and hyperglobulinemia. The cat was treated with allopurinol, responded well and was still alive at follow-up on day 288 after first presentation. However, enucleation was necessary because of refractory glaucoma and uveitis.  CONCLUSION: For the first time, ocular evidence of Leishmania IgG antibodies was demonstrated in the aqueous humor of both eyes in cats. There is limited knowledge about the pathogenesis, treatment options and outcomes in cats infected with L. infantum. This case report supports the hypothesis that immunosuppression increases the risk of clinical signs of leishmaniasis in cats. Alpha2- and gamma-globulin peaks in serum protein capillary electrophoresis are supportive criteria for the diagnosis of L. infantum infection. SAA is valuable for monitoring. Regarding ophthalmology, uveitis and glaucoma may have a poor prognosis.

Establishment of a Real-Time PCR Assay for the Detection of Devriesea agamarum in Lizards.

(1) Background: Devriesea (D.) agamarum is a potential cause of dermatitis and cheilitis in lizards. The aim of this study was to establish a real-time PCR assay for the detection of D. agamarum. (2) Methods: Primers and probe were selected targeting the 16S rRNA gene, using sequences of 16S rRNA genes of D. agamarum as well as of other bacterial species derived from GenBank. The PCR assay was tested with 14 positive controls of different D. agamarum cultures as well as with 34 negative controls of various non-D. agamarum bacterial cultures. Additionally, samples of 38 lizards, mostly Uromastyx spp. and Pogona spp., submitted to a commercial veterinary laboratory were tested for the presence of D. agamarum using the established protocol. (3) Results: Concentrations of as low as 2 × 104 colonies per mL were detectable using dilutions of bacterial cell culture (corresponding to approximately 200 CFU per PCR). The assay resulted in an intraassay percent of coefficient of variation (CV) of 1.31% and an interassay CV of 1.80%. (4) Conclusions: The presented assay is able to detect D. agamarum in clinical samples, decreasing laboratory turn-around time in comparison to conventional culture-based detection methods.

First Description of Sarcoptic Mange in a Free-Ranging European Wildcat (Felis silvestris silvestris) from Spain.

Sarcoptic mange caused by the mite Sarcoptes scabiei is a worldwide-distributed skin infestation with a wide range of hosts, among them several species within the Felidae family. Sarcoptes scabiei was diagnosed in a dead adult female European wildcat (Felis silvestris silvestris) from Spain, based on histological evaluation of skin biopsies and identification of the arthropod from skin scrapings and molecular methods. This is the first description of Sarcoptes scabiei in a European wildcat. Due to its critical demography in the southernmost population of the Iberian Peninsula, the impacts of infectious diseases, including sarcoptic mange, as a new potential threat should be considered during disease surveillance programs of the species' populations.

Humoral responses to SARS-CoV-2 by healthy and sick dogs during the COVID-19 pandemic in Spain.

COVID-19 is a zoonotic disease caused by SARS-CoV-2. Infections of animals with SARS-CoV-2 have recently been reported, and an increase of severe lung pathologies in domestic dogs has also been detected by veterinarians in Spain. Therefore, further descriptions of the pathological processes in those animals that show symptoms similar to those described in humans affected by COVID-19 would be highly valuable. The potential for companion animals to contribute to the continued transmission and community spread of this known human-to-human disease is an urgent issue to be considered. Forty animals with pulmonary pathologies were studied by chest X-ray, ultrasound analysis, and computed tomography. Nasopharyngeal and rectal swabs were analyzed to detect canine pathogens, including SARS-CoV-2. An additional twenty healthy dogs living in SARS-CoV-2-positive households were included. Immunoglobulin detection by several immunoassays was performed. Our findings show that sick dogs presented severe alveolar or interstitial patterns with pulmonary opacity, parenchymal abnormalities, and bilateral lesions. The forty sick dogs were negative for SARS-CoV-2 but Mycoplasma spp. was detected in 26 of 33 dogs. Five healthy and one pathological dog presented IgG against SARS-CoV-2. Here we report that despite detecting dogs with α-SARS-CoV-2 IgG, we never obtained a positive RT-qPCR for SARS-SoV-2, not even in dogs with severe pulmonary disease; suggesting that even in the case of canine infection, transmission would be unlikely. Moreover, dogs living in COVID-19-positive households could have been more highly exposed to infection with SARS-CoV-2.

Challenges in microbiological identification of aerobic bacteria isolated from the skin of reptiles.

INTRODUCTION: Bacterial pathogens are often involved in dermatitis in reptiles. Exact identification of reptile-specific but otherwise uncommon bacterial species may be challenging. However, identification is crucial to evaluate the importance of the detected bacterial species. OBJECTIVE: The aim of this study was to assess the number of aerobic bacterial isolates cultured from skin-derived samples of reptiles which were not reliably identified by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS), and to determine their identity. MATERIAL AND METHODS: Routine bacterial diagnostics were performed on 235 skin samples, and 417 bacterial isolates were analysed by MALDI-TOF MS. The isolates were grouped into categories based on their first score: category I (≥ 2.00), category II (≥ 1.70 and < 2.00), and category III (< 1.70). Isolates from category III were further investigated by 16S rRNA gene sequencing and the following criteria were applied: query cover 100%, e-value rounded to 0.0 and sequence identity (%) > 98.00% for genus identification, and > 99.00% for species identification. RESULTS: The majority of bacterial isolates were in category I (85.1%) or category II (8.4%). In category III (6.5%) results achieved at first by MALDI-TOF MS corresponded to the results of the molecular analysis in 8.0% of isolates at the species level and in 24.0% at the genus level. Bacterial isolates classified as category III were heterogenic in genus (e.g. Chryseobacterium, Devriesea, Pseudomonas, Staphylococcus, Uruburuella), and some have only been described in reptiles so far. CONCLUSIONS: Most of the aerobic bacterial isolates cultured from reptile skin achieved high scores by MALDI-TOF MS. However, in the majority of category III isolates MALDI-TOF MS results were different from those of the molecular analysis. This strengthens the need to carefully examine low-scored results for plausibility and to be familiar with the occurrence and morphology of relevant reptile-specific bacterial species (e.g. Devriesea agamarum) as well as with the limits of the database used.

[Prevalence of Dog circovirus in healthy and diarrhoeic dogs]. / Nachweishäufigkeit von Dog circovirus bei gesunden und an Durchfall erkrankten Hunden.

OBJECTIVE: In 2012, a Dog circovirus (DogCV) was discovered in the USA, which was followed by further descriptions of the virus in the USA, Italy and Germany. The present study is the first to examine the prevalence of DogCV in faeces of dogs from Germany and other European countries. MATERIAL AND METHODS: Faecal samples from 184 dogs with diarrhoea and from 82 clinically healthy dogs (control group) were analysed for the presence of DogCV by PCR. Furthermore, the detection of parvovirus, coronavirus, Giardia and Cryptosporidium was performed in all samples. RESULTS: In the group of dogs with diarrhoea the prevalence of DogCV was 20.1% (37/184), in the healthy control group it was 7.3% (6/82). Therefore, the virus could be detected significantly more frequently in dogs with diarrhoea. The detection frequency of DogCV is comparable with those of the other tested pathogens. In approximately 50% of the DogCV-positive dogs, infections with other enteropathogenic organisms were diagnosed. CONCLUSION AND CLINICAL RELEVANCE: The role of co-infection in the pathogenesis of the disease remains unclear, but there appears to be an association between co-infection and disease severity. Evidence of DogCV in clinically healthy dogs appears important for the epidemiology and raises questions about its pathogenicity. Further studies are needed to clarify questions regarding the pathogenesis, causal relevance and possible interference by other diarrhoeal pathogens. Nevertheless, the results of this study are an important indication that DogCV should be considered as a differential diagnosis in dogs with diarrhoea.