key: cord-1049294-6l5m0jp8
authors: Ruiz‐Arrondo, Ignacio; Portillo, Aránzazu; Palomar, Ana M.; Santibáñez, Sonia; Santibáñez, Paula; Cervera, Cristina; Oteo, José A.
title: Detection of SARS‐CoV‐2 in pets living with COVID‐19 owners diagnosed during the COVID‐19 lockdown in Spain: A case of an asymptomatic cat with SARS‐CoV‐2 in Europe
date: 2020-08-18
journal: Transbound Emerg Dis
DOI: 10.1111/tbed.13803
sha: 15923169a2874d40af0fdcb3ddf0666491e23793
doc_id: 1049294
cord_uid: 6l5m0jp8

Pets from COVID‐19 owners were screened for SARS‐CoV‐2 (April‐May 2020). From 23 pets, an asymptomatic cat showed positive RT‐qPCRs results from oropharyngeal swab (negative rectal swab). Remaining pets were negative. This suggests that cats can contract the virus from their infected owners and may act as potential hosts for SARS‐CoV‐2. Their role in carrying live or infectious viruses and disseminating them needs more investigation.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for the current coronavirus disease 2019 (COVID-19) pandemic, is the seventh coronaviruses that cause human respiratory infections. It belongs to the genus Betacoronavirus, such as the two other zoonotic coronaviruses (SARS-1 and MERS) that recently caused outbreaks. The natural origin of SARS-CoV-2 seems derived from bats (spillover). Pangolins, snakes, turtles, hamsters or yaks have been considered as potential intermediate hosts before spreading to humans, but this still remains unclear (Luan et al., 2020) . According to World Health Organization (WHO) (May 8, 2020), there is no evidence that dogs and/or cats can disseminate SARS-CoV-2 and act as source of human infection (https://www.who.int/es/emergencies/diseases/novelcoronavirus-2019/advice-for-public/q-a-coronaviruses). Nevertheless, in any new or emerging disease, especially if there are 'knowledge gaps' in the epidemiology, to assess the potential for domestic transmission through pets (mainly, dogs and cats) is relevant (https://www.oie.int/fileadmin/Home/eng/Our_scientific_expertise/docs/pdf/COV-19/COVID19_21Feb.pdf). Our aim was to evaluate, at an early stage of COVID-19 pandemic, the state of infection and, in consequence, the potential role of SARS-CoV-2 transmission from humans to companion animals (especially dogs and cats) under a one health approach in La Rioja (northern Spain).

A total of 23 asymptomatic mammalian pets under quarantine (8 cats, 1 guinea pig, 2 rabbits and 12 dogs) from 17 households with confirmed human cases of COVID-19 infection diagnosed at the Hospital Universitario San Pedro (Logroño, Spain) were included in our study (April 8-May 4, 2020) ( Table 1) 

This article is protected by copyright. All rights reserved extract, with 500 nM each primer and 125 nM probe. Thermal cycler was performed at 42 ºC for 5 min for reverse transcription, followed by 95ºC for 10 s and 45 cycles of 95 ºC for 5s and 55º C for 30s. All specimens were also analysed using the commercial kit GPS™ CoVID-19 dtec-RT-qPCR Test (Genetic PCR Solutions™, Alicante, Spain). Positive cases were confirmed by RT-qPCR for the envelope (E) protein-encoding gene (Corman et al., 2020), as described above for the N1 gene, with these modifications:

final concentrations of each primer and probe were 400 and 200 nM, respectively, and the annealing took place at 58ºC for 30s. Synthetic plasmid controls with the complete SARS-CoV-2 N gene (Integrated DNA Technologies, Leuven, Belgium) and the E gene (Eurofins Genomics, Ebersberg, Germany) were used to generate standard curves based on ten-fold serial dilutions for quantification. The GPS™ CoVID-19 dtec-RT-qPCR Test kit also included a positive control. Positive and negative (extraction and amplification) controls were included in all the RT-qPCR assays. Samples and controls were tested in triplicate.

The oropharyngeal swab sample from a female cat tested positive for SARS-CoV-2 by the three RT-qPCR assays performed ( Table 1) . The specimen showed a viral load of 1.7x10 3 RNA copies/µL for the N1 fragment gene, and 1.1x10 3 RNA copies/µL for the E gene. Viral RNA was not detected in the rectal swab sample from this animal. It was an 8-year-old female domestic European cat without clinical signs related to coronavirus disease, although it had chronic feline gingivostomatitis, feline idiopathic cystitis (treated with glucosamine and chondroitin sulphate), chronic kidney disease (treated with special feeding, ranitidine and benazepril hydrochloride) and feline asthmatic bronchitis (treated with fluticasone propionate). It cannot be ruled out that the virus was found in the swab just because the cat came into contact with a surface/fomites heavily contaminated by the infected owner (e.g. due to the owner's sneezing or coughing), Our study reports the detection of an asymptomatic cat with SARS-CoV-2 in Europe, probably associated with close contact with its owner who was diagnosed with active COVID-19 infection. According to our data, a high prevalence of SARS-CoV-2 RT-qPCR positive cats was observed (1/8; 12.5%). Up to our

This article is protected by copyright. All rights reserved knowledge, the known prevalence data for cats are those from Zhang et al. (Zhang et al., 2020) . They found a 14.7% seroprevalence against SARS-CoV-2 in cats in China. To our knowledge, no previous SARS-CoV2 RNA detection in asymptomatic cats has been reported in Spain. Another case of SARS-CoV-2 infection in a cat without signs of respiratory disease has been reported in Germany (https://www.oie.int/en/scientific-expertise/specific-information-and-recommendations/questions-andanswers-on-2019novel-coronavirus/events-in-animals/). Three cats remained asymptomatic after cohousing and SARS-CoV-2 transmission by infected cats (Halfmann et al., 2020) . The remaining SARS-CoV-2 infections in felines (Belgium, USA, France and Spain) were symptomatic, and mainly showed clinical signs of respiratory and/or digestive disease. A more recent case has been detected in a cat in Russia but no clinical data are available (https://www.oie.int/en/scientific-expertise/specific-information-andrecommendations/questions-and-answers-on-2019novel-coronavirus/events-in-animals/). Probably, all these cases are also the result of contagions from their owners. Based on these results, it is possible that the number of affected cats living with COVID-19 owners is greater than that published to date, since these animals may be asymptomatic and not detected.

We consider that the limited number of animals included in our study can be a bias for the results.

Nevertheless, the exceptional circumstances lived in Spain and in our region (La Rioja) during the sampling period (lockdown, confinement, quarantine, healthcare staff's work overload, high mortality rates and health system on the brink of collapse) make these data relevant. Our finding indicates that cats can contract the virus from infected humans, although the transmission of SARS-CoV-2 from animals to humans seems unlikely (https://www.who.int/es/emergencies/diseases/novel-coronavirus-2019/advice-forpublic/q-a-coronaviruses). To date, all cases seem to be isolated ones related with human transmission through COVID-19 infected people taking care of the animals, without any epidemiological significance and considering cats as dead-end hosts (https://www.cdc.gov/coronavirus/2019-ncov/daily-lifecoping/animals.html). 

This article is protected by copyright. All rights reserved (https://www.cdc.gov/coronavirus/2019-ncov/daily-life-coping/animals.html). The role of cats in these outbreaks related to minks is unknown. Further research from a one health perspective is needed to clarify these aspects. 

Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR

Transmission of SARS-CoV-2 in Domestic Cats

SARS-CoV-2 spike protein favors ACE2 from Bovidae and Cricetidae

Susceptibility of ferrets, cats, dogs, and other domesticated animals to SARS-coronavirus 2. Science, eabb7015

SARS-CoV-2 neutralizing serum antibodies in cats: a serological investigation

We would like to acknowledge to the pets' owners for their consent to include their pets in the study.

The authors declare no conflict of interest.

The manuscript has been sent to the preprint server medRxiv (https://doi.org/10.1101/2020.05.14.20101444).

This article is protected by copyright. All rights reserved covid23 Dog Spanish water dog 3 Female Negative Negative Mild 26 †Two confirmed human cases in the same household. Mild: upperway respiratory clinic without radiological data from pulmonary involvement; Moderate: respiratory involvement with radiological alteration without the need for oxygen; Severe: respiratory involvement with radiological alteration and need of oxygen.