Infectious disease introduced by non-native species is increasingly cited as a facilitator of native population declines, but direct evidence may be lacking due to inadequate population and disease prevalence data surrounding an outbreak. Previous indirect evidence and theoretical models support squirrelpox virus (SQPV) as being potentially involved in the decline of red squirrels (Sciurus vulgaris) following the introduction of the non-native gray squirrel (Sciurus carolinensis) to the United Kingdom. The red squirrel is a major UK conservation concern and understanding its continuing decline is important for any attempt to mitigate the decline. The red squirrel–gray squirrel system is also exemplary of the interplay between infectious disease (apparent competition) and direct competition in driving the replacement of a native by an invasive species. Time series data from Merseyside are presented on squirrel abundance and squirrelpox disease (SQPx) incidence, to determine the effect of the pathogen and the non-native species on the native red squirrel populations. Analysis indicates that SQPx in red squirrels has a significant negative impact on squirrel densities and their population growth rate (PGR). There is little evidence for a direct gray squirrel impact; only gray squirrel presence (but not density) proved to influence red squirrel density, but not red squirrel PGR. The dynamics of red SQPx cases are largely determined by previous red SQPx cases, although previous infection of local gray squirrels also feature, and thus, SQPV-infected gray squirrels are identified as potentially initiating outbreaks of SQPx in red squirrels. Retrospective serology indicates that approximately 8% of red squirrels exposed to SQPV may survive infection during an epidemic. This study further highlights the UK red squirrel – gray squirrel system as a classic example of a native species population decline strongly facilitated by infectious disease introduced by a non-native species. It is therefore paramount that disease prevention and control measures are integral in attempts to conserve red squirrels in the United Kingdom.

No abstract

Native red squirrels Sciurus vulgaris in Great Britain and Ireland are threatened by alien grey squirrels S. carolinensis through exploitation competition and spillover of squirrelpox virus (SQPV). By accelerating the replacement of red squirrels by the invader, SQPV represents a fundamental factor to consider when planning management and conservation strategies. In mainland Europe, grey squirrels introduced to Italy threaten the survival of the whole continental red squirrel population, but no extensive surveys for SQPV presence have been carried out in the region. We therefore investigated SQPV infection in north Italian grey squirrel populations through a combination of serological and molecular methods. Firstly, we analysed sera from 285 individuals through an enzyme-linked immunosorbent assay (ELISA) to detect antibodies against SQPV. Secondly, a PCR designed to amplify a segment of the G8R SQPV gene was carried out on DNA extracted from swabs and skin tissue samples from a second set of 66 grey squirrels. ELISA tests identified four reactors (1.4%), but the subsequent PCR survey did not detect any SQPV DNA. Based on the low prevalence observed and on PCR results, we believe that the four suspected positives were the result of an ELISA cross-reaction following exposure to another pox virus. Considering sample size and performances of the two methods, confidence of freedom from SQPV resulted above 99.9%. However, because of the severe impact of SQPV on red squirrels, we recommend the implementation of a passive surveillance plan for the early detection of an SQPV emergence in continental Europe.

Rapid development in polymerase chain reaction (PCR) technology has revolutionised the speed and accuracy of many diagnostic assays. However, comparatively few wildlife epidemiological studies use quantitative PCR (qPCR) for pathogen detection, even fewer employ an internal control, to ensure confidence in negative results, and PCR's ability to multiplex and therefore detect several targets in a single reaction is underutilised. Here, we describe the development of two multiplex qPCR assays for the red and grey squirrel that detect the pathogens squirrelpox virus (SQPV) and adenovirus in squirrels (SADV), both of which cause mortality in the red squirrel. Both assays use a section of the squirrel phosphoglycerate kinase gene as an endogenous internal control that identifies and compensates for both, inadequate sampling or PCR inhibition. Tests on infected squirrel tissue demonstrate that simple swab samples (particularly from distal antebrachial skin) are sufficient to detect and identify the relative quantity of SQPV DNA in both squirrel species, while rectal swabs and blood cell pellets can be used to reliably indicate SADV infection. These assays are sensitive and specific with an endogenous internal control providing confidence in negative results and allowing comparison across laboratories. Using such assays should prove advantageous in wildlife studies with limited resources while allowing the maximum data yield.