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Feline Morbillivirus in Northern Italy Prevalence in Urine and Kidneys

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Feline Morbillivirus in Northern Italy Prevalence in Urine and Kidneys Veterinary Microbiology 233 (2019) 133–139 Contents lists available at ScienceDirect Veterinary Microbiology journal homepage: www.elsevier.com/locate/vetmic Feline morbillivirus in Northern Italy: prevalence in urine and kidneys with T and without renal disease ⁎ Angelica Stranieria,b, , Stefania Lauzia,b, Annachiara Dallaria, Maria Elena Gelainc, Federico Bonsembiantec, Silvia Ferroc, Saverio Paltrinieria,b a Department of Veterinary Medicine, University of Milan, Milan, Italy b Central Laboratory, Veterinary Teaching Hospital, University of Milan, Lodi, Italy c Department of Comparative Biomedicine and Food Science, University of Padova, Legnaro, Italy ARTICLE INFO ABSTRACT Keywords: Feline morbillivirus (FeMV) is an emerging virus that was first described in Hong Kong in 2012. Several reports Feline morbillivirus suggested the epidemiological association of FeMV infection with chronic kidney disease (CKD) in cats. The aim Chronic kidney disease of this study was to investigate the presence and the genetic diversity of FeMV as well as the relationship Clinical pathology between FeMV infection and CKD in cats from Northern Italy. Urine (n = 81) and kidney samples (n = 27) from Urinalysis 92 cats admitted to the Veterinary Teaching Hospital of the University of Milan between 2014 and 2017 were investigated for FeMV infection. FeMV RNA was detected in one urine sample (1.23%; 95% CI: 0.03–6.68%) and in two kidneys (7.40%; 95% CI: 0.91–24.28%). FeMV RNA was revealed only in urine or kidneys of cats without evidence of CKD. Phylogenetic analysis showed that the three strains clustered with FeMV strains retrieved from public database, forming a distinct sub-cluster of FeMV. The presence of distinct genotypes of FeMV found in this study is in accordance with previous studies demonstrating that FeMV strains are genetically diverse. A clear relationship between the presence of FeMV infection and CKD in the cats from Northern Italy was not observed, confirming recent reports that do not support the hypothesis that FeMV infection is associated withthedevel- opment of CKD. 1. Introduction (2015) found in cats from Germany viruses with sequences different from any other morbillivirus and with a nucleotide homology close to Morbilliviruses belong to the subfamily Paramyxovirinae, family bat and rodent paramyxoviruses, thereby named feline para- Paramoxyviridae, which is composed of enveloped, single-stranded, myxoviruses (FPaV). The same authors found a correlation between negative-sense RNA viruses (de Vries et al., 2015). Morbilliviruses in- FeMV and FPaV presence in urine and CKD (Sieg et al., 2015). Recently, fect humans (e.g. Measles morbillivirus) and animals (e.g. Canine dis- a high prevalence of FeMV was found, but without a strong correlation temper virus, Peste des petits ruminants virus) (Nambulli et al., 2016). between TIN and infection (Park et al., 2016). Moreover, Yilmaz et al Feline morbillivirus (FeMV) was firstly isolated in Hong Kong between (2017) did not find any significant difference in both histopathological 2009 and 2011, mainly in urine, and a role of this virus in the patho- and clinico-pathological findings between FeMV infected and not in- genesis of feline tubular interstitial nephritis (TIN) and chronic kidney fected cats (Yilmaz et al., 2017). In addition, no relationship between disease (CKD) was postulated (Woo et al., 2012). Two years later, FeMV the shedding of the virus in urine and laboratory alterations imputable was isolated in Japan with a prevalence of 6.1% in urine, 10% in blood to CKD were found in recent studies (Darold et al., 2017; McCallum and 40% in kidneys with nephritis (Furuya et al., 2014). Up to date, et al., 2018). FeMV has been identified also in USA, South America, Turkey and In 2015, FeMV was identified in Italy from the urine of a 15-years Europe, with sequences closely related to previously identified FeMV old stray cat with symptoms and laboratory alterations imputable to strains (Lorusso et al., 2015; Sieg et al., 2015; Sharp et al., 2016; Darold chronic kidney disease (CKD). Even if a kidney biopsy was not per- et al., 2017; Yilmaz et al., 2017; McCallum et la., 2018). Sieg et al formed, the fact that the cat continued to shed the virus in the urine for ⁎ Corresponding author at: Department of Veterinary Medicine, University of Milan, Via Celoria 10, Milan, Italy. E-mail addresses: [email protected] (A. Stranieri), [email protected] (S. Lauzi), [email protected] (A. Dallari), [email protected] (M.E. Gelain), [email protected] (F. Bonsembiante), [email protected] (S. Ferro), [email protected] (S. Paltrinieri). https://doi.org/10.1016/j.vetmic.2019.04.027 Received 6 September 2018; Received in revised form 23 April 2019; Accepted 23 April 2019 0378-1135/ © 2019 Elsevier B.V. All rights reserved. A. Stranieri, et al. Veterinary Microbiology 233 (2019) 133–139 14 days, led the Authors to suggest a possible role of this virus in the of relevant laboratory changes, and as diseased if history, clinical signs, development of renal failure (Lorusso et al., 2015). FeMV is now cited diagnostic imaging, or laboratory changes were consistent with a spe- as a possible cause of CKD, which is a very common feline disease, with cific disease. Sick cats were further classified in specific disease groups a multifactorial and still unclear pathogenesis (Brown et al., 2016). according to their diagnosis: CKD alone; CKD along with other diseases; However, only few studies on FeMV provided information about cats urinary tract disorders without CKD; neoplastic, infectious, endocrine signalment and anamnesis, as well as on clinicopathological changes or miscellaneous diseases. Based on the International Renal Interest related to FeMV infection. Moreover, data regarding the correlation Society (IRIS) guidelines (http://www.iris-kidney.com), the con- between CKD and FeMV infection are often discordant and they have centration of serum creatinine and the UPC values were used to stage not been investigated in Italy yet. Thus, the aims of this study were to and substage cats for CKD, respectively. investigate the presence of feline paramyxoviruses in Italy with mole- cular methods, regardless of the presence of clinical signs consistent 2.3. Kidney samples with CKD, to perform molecular characterization and phylogenetic analysis and to evaluate the existence of a correlation between para- Kidneys were collected during necropsies performed either within myxoviruses infection and clinicopathological as well as histopatholo- studies on feline infectious peritonitis (FIP) or during routine diagnostic gical changes imputable to renal damage. activities of the Pathology unit of the Veterinary teaching hospital. During necropsies, two sections approximately 1 cm thick each in- 2. Material and methods cluding, when present, macroscopically apparent lesions were col- lected. One section was immediately frozen at −80 °C and subjected to 2.1. Sample selection and processing RNA extraction within two weeks from sampling, and one section was put in 10% neutral-buffered formalin for histology. Sections used for Cats routinely subjected to urine and hematological analyses for molecular biology were always collected adjacent to the sections used diagnostic purposes at the Veterinary Teaching Hospital of Milan in for histology. Histology was performed on formalin-fixed paraffin-em- 2017 were involved. Signalment (e.g. age, breed, sex, living environ- bedded (FFPE) samples using microtomic sections (3 μm) stained with ment), clinical history and health status, including results of Feline haematoxylin-eosin. Immunodeficiency virus (FIV) and Feline Leukemia virus (FeLV) ser- ology, were recorded for each cat. Either cats with a diagnosis or a 2.4. PCR for FeMV and phylogenetic analysis clinical suspicion of CKD, clinically healthy cats, and cats affected by other diseases were enrolled, regardless of age and breed. Cats were Frozen-thawed urine samples were centrifuged (3000 rpm × 5 min) examined and received additional diagnostic investigations (e.g. X- to remove debris. Then, 150 μL of the obtained supernatant were used Rays, ultrasound or other instrumental diagnostic approaches) de- for RNA extraction using the NucleoSpin RNA Virus commercial kit pending on the clinical presentation. Urine and blood samples were (Macherey-Nagle, Bethlehem, PA), following manufacturer’s instruc- collected in the context of clinicopathological investigations requested tions. RNA extraction from renal tissues was performed using the from the referring veterinarians. Additionally, 27 RNA samples ob- NucleoSpin RNA kit (Macherey-Nagel, Bethlehem, PA) according to the tained from kidneys of euthanized or spontaneously deceased cats were manufacturer’s instruction. RNA samples obtained from urine and also collected during necropsies performed for diagnostic purposes kidneys were then frozen at −80 °C or immediately investigated for the between 2014 and 2017. presence of FeMV by molecular analysis. All of the above procedures were performed within routine diag- A FeMV specific nested RT-PCR (nRT-PCR) was performed using nostic workouts and therefore, according to the decisions of the Ethical primers and methods amplifying a 401 bp fragment of the L gene, ac- Committee of the university of Milan, residual aliquots of samples or cording to Furuya et al (2014). PCR product were visualized under UV tissues collected under informed consent of the owners can be used for transilluminator
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