Molecular and Serological Studies on Potential SARS-CoV-2 Infection among 43 Lemurs under Human Care-Evidence for Past Infection in at Least One Individual

Abstract

In the setting of the recent COVID-19 pandemic, transmission of SARS-CoV-2 to animals has been reported in both domestic and wild animals and is a matter of concern. Given the genetic and functional similarities to humans, non-human primates merit particular attention. In the case of lemurs, generally considered endangered, they are believed to be susceptible to SARS-CoV-2 infection. We have conducted a study for evidence of SARS-CoV-2 infection among the 43 lemurs of Mundomar, a zoological park in Benidorm, Spain. They belong to two endangered lemur species, 23 black-and-white ruffed lemurs (Varecia variegata) and 20 ring-tailed lemurs (Lemur catta). Health assessments conducted in 2022 and 2023 included molecular analyses for SARS-CoV-2 RNA of oral and rectal swabs using two different RT-qPCR assays, always with negative results for SARS-CoV-2 in all animals. The assessment also included serological testing for antibodies against the receptor-binding domain (RBD) of the spike protein (S) of SARS-CoV-2, which again yielded negative results in all animals except one black-and-white ruffed lemur, supporting prior infection of that animal with SARS-CoV-2. Our data, while not indicating a high susceptibility of lemurs to SARS-CoV-2 infection, show that they can be infected, adding to the existing information body on potential ways for SARS-CoV-2 virus spreading in zoos, highlighting the need for animal surveillance for the virus.

Keywords: COVID-19; Lemur catta; SARS-CoV-2; Varecia variegata; animal COVID-19; lemurs, captive; one health.

Figure 1

Weights of the lemurs of the two species determined in the present assessments of the lemurs’ health, plotted against the age of the animals on examination. The curves show the Lowess fit (carried out by GraphPad Prism, for 10 points in the smoothing window) for the pooled data for males (blue circles) and females (red squares). In the left panel, the solid red squares correspond to weight determinations in animal #6, the only animal that was seropositive for SARS-CoV-2.

Figure 2

Illustrative example of RT-PCR, for animal 6, the lemur that tested positive by serology, on the date of assessment that gave seropositivity. The sample was RNA from the oropharynx. For details on the procedure and the positive and negative controls, see Section 2.3 Thresholds for positivity were drawn by the program on the basis of the negative controls.

Figure 3

Serology results for anti-RBD of SARS-CoV-2 in the lemurs of the present study. For details, see Materials and Methods. (A) Serological titers determined in this study (see Table 1 for the times and animals sampled for serum) grouped per lemur species as indicated. The results for lemur #6 (identified in Table 1; it is a V. variegata individual) are singled out in the center as filled circles, indicating the date of the blood extractions. The horizontal lines represent means and standard deviations for the points in the cohort. (B) Frequency of serological titers found in our entire animal cohort (pooling of both species) in the indicated intervals of titer values. Results for lemur #6 are in black filling. (C) Serology titers for all the animals for which two serum samples were obtained with a 7–11 month interval between the two blood extractions (as indicated). Colors and symbols identify each individual as shown. The two results for each animal are connected with a straight line, except in the three cases, in which the second value was smaller than the first one, in which the lines are the exponentials for a t_1/2 for the decay of 1 month.