Mitogenome sequences of domestic cats demonstrate lineage expansions and dynamic mutation processes in a mitochondrial minisatellite.

BACKGROUND: As a population genetic tool, mitochondrial DNA is commonly divided into the ~ 1-kb control region (CR), in which single nucleotide variant (SNV) diversity is relatively high, and the coding region, in which selective constraint is greater and diversity lower, but which provides an informative phylogeny. In some species, the CR contains variable tandemly repeated sequences that are understudied due to heteroplasmy. Domestic cats (Felis catus) have a recent origin and therefore traditional CR-based analysis of populations yields only a small number of haplotypes. RESULTS: To increase resolution we used Nanopore sequencing to analyse 119 cat mitogenomes via a long-amplicon approach. This greatly improves discrimination (from 15 to 87 distinct haplotypes in our dataset) and defines a phylogeny showing similar starlike topologies within all major clades (haplogroups), likely reflecting post-domestication expansion. We sequenced RS2, a CR tandem array of 80-bp repeat units, placing RS2 array structures within the phylogeny and increasing overall haplotype diversity. Repeat number varies between 3 and 12 (median: 4) with over 30 different repeat unit types differing largely by SNVs. Five SNVs show evidence of independent recurrence within the phylogeny, and seven are involved in at least 11 instances of rapid spread along repeat arrays within haplogroups. CONCLUSIONS: In defining mitogenome variation our study provides key information for the forensic genetic analysis of cat hair evidence, and for the first time a phylogenetically informed picture of tandem repeat variation that reveals remarkably dynamic mutation processes at work in the mitochondrion.

Defining cat mitogenome variation and accounting for numts via multiplex amplification and Nanopore sequencing.

Hair shed by domestic cats is a potentially useful source of forensic evidence. Analysable hair DNA is predominantly mitochondrial, but the recent domestication history of cats means that mtDNA diversity is low. A 402-bp control region segment is usually sequenced, defining only a small number of distinct haplotypes in populations. Previously, we used a long-amplicon approach to sequence whole mitogenomes in a sample of blood DNAs from 119 UK cats, greatly increasing observed diversity and reducing random match probabilities. To exploit this variation for forensic analysis, we here describe a multiplex system that amplifies the cat mitogenome in 60 overlapping amplicons of mean length 360 bp, followed by Nanopore sequencing. Variants detected in multiplex sequence data from unrooted hair completely mirror those from long-amplicon data from blood from the same individuals. However, applying the multiplex to matched blood DNA reveals additional sequence variants which derive from the major feline nuclear mitochondrial insertion sequence (numt), which covers 7.9 kb of the 17-kb mitogenome and exists in multiple tandem copies. We use long-amplicon Nanopore sequencing to investigate numt variation in a set of cats, together with an analysis of published genome sequences, and show that numt arrays are variable in both structure and sequence, thus providing a potential source of uncertainty when nuclear DNA predominates in a sample. Forensic application of the multiplex was demonstrated by matching hairs from a cat with skeletal remains from its putative mother, both of which shared a globally common haplotype at the control region. The random match probability in this case with the CR 402-bp segment was 0.21 and this decreased to 0.03 when considering the whole mitogenome. The developed multiplex and sequencing approach, when applied to cat hair where nuclear DNA is scarce, can provide a reliable and highly discriminating source of forensic genetic evidence from a single hair. The confounding effect of numt co-amplification in degraded samples where mixed sequences are observed can be mitigated by variant phasing, and by comparison with numt sequence diversity data, such as those presented here.

Challenges in the diagnosis and management of skin diseases in alpacas, goats, pigs, and sheep.

The primary aim of this article is to provide an overview of several selected skin conditions in livestock species. Topics include ectoparasites in alpacas, antler velvet in reindeer, immune-mediated disease in goats, ectoparasites in pigs, Culicoides allergic dermatitis and parapox infection in sheep. When dealing with skin disease in livestock, it is important to collect a detailed history and undertake a thorough clinical examination to include the axilla, groin, limbs and feet. While the diagnosis will often be anticipated from the history and presentation, it is important to consider a differential diagnosis list and appropriate diagnostic testing before embarking on a poly-pharmacy approach to "rule out" causes of disease. This is particularly important where morbidity is high and the livestock of perceived high value to the keeper/owner, such as goats and small-breed pigs, or when the skin condition is long standing/chronic. Ideally, the management plan should sequentially clarify the role of microbial infection and then ectoparasites before considering less common allergic and autoimmune conditions. Skin cytology is an invaluable in-house diagnostic method that can support the findings of culture. Taking skin samples for histopathology and possibly culture may prove valuable once other diagnostic methods have been explored. Given the need to protect the use of parenteral antimicrobials, topical antimicrobial therapies can be deployed successfully. The repeated use of macrocyclic lactones (avermectins) must be balanced in terms of the risks of promoting anthelmintic resistance versus controlling or eradicating the ectoparasites that have, ideally, been specifically identified.

Health and welfare of deer.

Reviewed by Aiden Foster, president of the British Deer Veterinary Association.

A survey of enteric disease agents in UK deer populations.

Wild deer products have been linked to outbreaks of Escherichia coli 0157 in humans and bovine tuberculosis (bTB) in domestic cats, raising concerns around foodborne diseases from venison. This study investigated the prevalence of several enteric disease agents in UK deer populations, including foodborne pathogens and other diseases of import to livestock health. Intestinal samples were collected from slaughtered farmed deer (N = 211, eight farms), shot wild deer (N = 136) and ground faecal samples were collected from two farms (N = 90), six parks (N = 228) and five zoos (N = 67). DNA was extracted and multiplex qPCR assays were run to amplify targets of, C. perfringens toxins, Campylobacter spp., E. coli toxins, Mycobacterium avium subsp. Paratuberculosis (MAP), Salmonella spp. and Yersinia spp. The most commonly amplified targets were E. coli astA (61.2%) and Campylobacter spp. (43.3%), but the prevalence of C. coli and C. jejuni were ≤ 3.0% and Salmonella spp., MAP and Yersinia pseudotuberculosis also had low overall prevalences of 1.6%, 3.3% and 2.6%, respectively. However, notable targets included C. perfringens toxins α (20.4%) and ß2 (16.9%); E. coli stx1 (14.6% overall, 28.0% in abattoir samples) and stx2 (17.8% overall, 31.6% in wild samples); and Yersinia enterocolitica (10.8% overall, 25% in wild samples). The low prevalence of some foodborne pathogens is reassuring for food safety; further investigations into the commonly found targets are needed to quantify the risk to public health and the possibility of wild deer acting as reservoirs of disease.

A survey of handling and transportation of UK farmed deer.

Studies on the transport of deer (Cervidae), in the UK, were published > 15 years ago. A more recent study of deer transport is required to allow for assessments and improvements to the transport of farmed deer. Sixteen deer farmers participated in a survey describing their management practices related to transport. Their responses showed that most vehicles used to transport deer were designed for other livestock. Participating farmers estimated journey times to slaughter as 1-8 h, with an arithmetic mean of 4.8 (± 2.38) h. Specific concerns raised by the respondents, relating to the transport of deer, included a need for deer-specific vehicles, stop-off areas for long journeys, market locations and haulier experience. Furthermore, data were collected from two abattoirs between July 2019 and June 2020 comprising journey times, slaughter times, bruising, location of origin, vehicle type and the number of animals. In total, 4,922 deer were transported across 133 journeys (from farm to abattoir) from 61 farms. Median and range for journey length were 3.2 (0.4-9.8) h and 154.2 (7.1-462.2) km, whereas group size and time spent in the lairage were 24 (1-121) and 17.8 (10.2-68.9) h, respectively. Group size was found to be significantly associated with both the presence of bruising in a group and the amount of bruising per deer. This study provides a much-needed update on the transport of farmed deer in the UK and highlights key areas for future research including the welfare impact of transport in larger groups and for longer durations.

Reindeer health and welfare.

This focus article has been prepared by Aiden Foster of the University of Bristol (which offers postmortem examination services as part of the APHA surveillance network) and Alan Wight, veterinary lead of the APHA Miscellaneous and Exotic Farmed Species Expert Group.

Congenital heart defects in calves.

This focus article describes some of the cardiac anomalies detected in calves submitted to APHA Veterinary Investigation Centres and other postmortem examination providers.

Otoscopy and aural cytological findings in a population of rescue cats and cases in a referral small animal hospital in England and Wales.

OBJECTIVES: Otitis externa is seen clinically in cats, although studies investigating this condition within the UK are lacking. The objective of this study was to investigate the prevalence of Otodectes cynotis mites and microbial infection in the ear canals of cats in various rescue centres and a referral hospital. METHODS: Otoscopy was performed in 332 cats. Otoscopic findings were noted, including the gross visualisation of Otodectes species. A sample of cerumen was collected for cytological evaluation and a cerumen smear for detection of Otodectes mites if there was a large amount of aural exudate present. RESULTS: O cynotis infestation was noted in 3/341 cats (0.9%, 95% confidence interval [CI] 0.3-2.6). A total of 129/341 (37.8%; 95% CI 32.7-43.0) cats were found to have Malassezia species within one or both ears. Bacteria were found unilaterally in 9/341 (2.6%; 95% CI 1.4-4.9) cats. Analysis of the cytological findings showed an increased likelihood for Malassezia species to be present as age increased (n = 293; Pearson r = 0.204, P <0.001). There was also an increased likelihood of finding Malassezia species in both ears if found within one ear (n = 327; r = 0.499, P <0.001). There was a positive correlation between the number of Malassezia organisms and the quantity of aural exudate (n = 338; r = 0.778, P <0.001). Cats in which Otodectes species infestation were noted (n = 3) had moderate or large quantities of cerumen. CONCLUSIONS AND RELEVANCE: This study shows that there was a low prevalence of O cynotis in this cohort of cats. In normal cats it was not unusual to find Malassezia microorganisms upon aural cytology, bacteria were noted far less frequently and in two cats this was associated with underlying anatomical pathology.

Owner-reported flea treatment measures and skin disease in cats.

OBJECTIVES: The aim of this study was to obtain information regarding the use of flea treatments and owner-reported flea infestation and skin disease. METHODS: Owners of 1150 cats were recruited into a long-term longitudinal study ('Bristol Cats' study) and asked to complete questionnaires at set time points. Questionnaires 1 and 5 were used and included data collection for kittens aged 2-4 months (T1) and at 2.5 years of age (T2). Owners were asked which brand of flea treatment, if any, was used at each time point. At T2 owners were asked if they had noted signs of skin disease or fleas on their cat within the previous 12 months and if they had sought veterinary attention for the skin disease or fleas. IBM SPSS version 23 was used for statistical analyses. RESULTS: Skin disease at T2 was reported by owners of 55/1150 (4.8%; 95% confidence interval [CI] 3.7-6.2) cats. Many owners who reported skin disease (25/55 [45.5%]; 95% CI 32.3-58.7) had not sought veterinary attention. There were 81/1150 (7.0%; 95% CI 5.6-8.5) cats with reported head shaking and/or ear scratching within the past 12 months at T2. The majority of cats (204/286; 71.3%) received prescription flea treatments at both points. Use of non-prescription flea treatment at T2 was more likely to be associated with owner-reported skin disease/flea infestation than those who used prescription flea treatment ( P <0.001). There was a significant association between the reported presence of fleas and reported presence of skin disease at T2 ( P = 0.03). CONCLUSIONS AND RELEVANCE: Despite the potential for reverse causality, the association between owner-reported skin disease and/or flea infestation in their cats and the use of non-prescription flea treatment was as expected. Use of flea treatments and, in particular, the type of ectoparasite control used (prescription or non-prescription) should be carefully assessed when managing cats with skin disease.

Retrospective study of the presentation, diagnosis and management of 16 cats with otitis media not due to nasopharyngeal polyp.

OBJECTIVES: The aim of this study was to analyse retrospectively cats diagnosed with otitis media (OM) not due to nasopharyngeal polyp, and to review the clinical outcome with surgical and medical management. METHODS: Patient records were searched for cats diagnosed with OM. The diagnosis of OM was based on the presence of clinical signs, including neurological signs, respiratory signs and signs of otitis externa, and on the basis of evidence of thickened or irregular bullae walls, or the presence of fluid within the tympanic cavity in those that had diagnostic imaging. In those that did not have imaging, the diagnosis was made on the basis of the presence of fluid in the bulla or organisms cultured using myringotomy. These records were analysed retrospectively. RESULTS: Of 16 cats, one had a total ear canal ablation, five had ventral bulla osteotomy surgery and 11 were medically managed. Of the cats that were medically managed, using either topical products, systemic antimicrobials or a combination of both, eight had complete resolution of clinical signs. CONCLUSIONS AND RELEVANCE: This small cohort shows that some cats with OM can be successfully managed medically. Surgery is invasive and may not necessarily be required if appropriate medical management is undertaken. This is the first study of OM treatment in cats and provides the basis for further studies, which should aim to establish specific infectious causes of OM and how they can potentially be managed with medical therapies.