[SARS-CoV-2 infections in cats, dogs, and other animal species: Findings on infection and data from Switzerland]. / SARS-CoV-2 Infektionen bei Katzen, Hunden und anderen Tieren: Erkenntnisse zur Infektion und Daten aus der Schweiz.

INTRODUCTION: The pandemic with the novel coronavirus (SARS-CoV-2) has led to infections and deaths worldwide. Apart from humans, certain animal species are susceptible to the viral infection. Spillover between humans and animals is favored by close contact; thus, surveillance of animals is an important component to fight the pandemic from a One Health perspective. The Clinical Laboratory of the Vetsuisse Faculty Zurich has been investigating SARS-CoV-2 infections in animals since the beginning of the pandemic. In November 2020, the first SARS-CoV-2 positive Swiss cat was reported to the World Organisation for Animal Health (OIE-WAHIS). The cat showed respiratory signs and lived in a COVID-19 affected household. By now, over 500 natural SARS-CoV-2 infections have been recorded in animals worldwide. A prevalence study on SARS-CoV-2 infections in dogs and cats was carried out together with clinics from Germany and Italy during the first wave of the pandemic (March-July 2020). Among the tested 1137 animals, only one cat and one dog were positive. The prevalence of infection in dogs and cats presented to veterinary clinics was low, even in pandemic hotspot regions. However, recent studies that focused on animals in COVID-19 households found a higher prevalence of infection. A study is currently underway that specifically collects samples from pets from Swiss COVID-19 affected household and collects data on human-animal interaction.

INTRODUCTION: La pandémie à nouveau coronavirus (SARS-CoV-2) a entraîné des infections et des décès dans le monde entier. En dehors de l'homme, certaines espèces animales sont sensibles à cette infection virale. Le passage entre les humains et les animaux est favorisé par un contact étroit, la surveillance des animaux est donc un élément important pour lutter contre la pandémie dans une perspective One Health. Depuis le début de la pandémie, le laboratoire clinique de la faculté Vetsuisse de Zurich étudie les infections par le SRAS-CoV-2 chez les animaux. En novembre 2020, le premier chat suisse positif au SARS-CoV-2 a été signalé à l'Organisation mondiale de la santé animale (OIE-WAHIS). Le chat a montré des signes respiratoires et vivait dans un ménage touché par le COVID-19. À l'heure actuelle, plus de 500 infections naturelles au SRAS-CoV-2 ont été enregistrées chez des animaux dans le monde. Une étude de prévalence sur les infections par le SRAS-CoV-2 chez les chiens et les chats a été réalisée avec des cliniques d'Allemagne et d'Italie pendant la première vague de la pandémie (mars-juillet 2020). Parmi les 1137 animaux testés, seuls un chat et un chien étaient positifs. La prévalence de l'infection chez les chiens et les chats présentés aux cliniques vétérinaires était faible, même dans les régions fortement touchées par la pandémie. Cependant des études récentes, qui se sont concentrées sur les animaux dans les ménages COVID-19, ont révélé une prévalence d'infection plus élevée. Une étude est actuellement en cours qui collecte spécifiquement des échantillons d'animaux de compagnie des ménages suisses touchés par le COVID-19 et enregistre des données sur l'interaction homme-animal.

Inflammatory Mediators in the Mesenteric Lymph Nodes, Site of a Possible Intermediate Phase in the Immune Response to Feline Coronavirus and the Pathogenesis of Feline Infectious Peritonitis?

Feline infectious peritonitis (FIP) is an almost invariably fatal feline coronavirus (FCoV)-induced disease thought to arise from a combination of viral mutations and an overexuberant immune response. Natural initial enteric FCoV infection may remain subclinical, or result in mild enteric signs or the development of FIP; cats may also carry the virus systemically with no adverse effect. This study screened mesenteric lymph nodes (MLNs), the presumed first site of FCoV spread from the intestine regardless of viraemia, for changes in the transcription of a panel of innate immune response mediators in response to systemic FCoV infection and with FIP, aiming to identify key pathways triggered by FCoV. Cats with and without FIP, the latter with and without FCoV infection in the MLN, were compared. Higher expression levels in FIP were found for toll-like receptors (TLRs) 2, 4 and 8. These are part of the first line of defence and suggest a response to both viral structural proteins and viral nucleic acid. Expression of genes encoding inflammatory cytokines and chemokines, including interleukin (IL)-1ß, IL-6, IL-15, tumour necrosis factor (TNF)-α, CXCL10, CCL8, interferon (IFN)-α, IFN-ß and IFN-γ, was higher in cats with FIP, consistent with inflammatory pathway activation. Expression of genes encoding transcription factors STAT1 and 2, regulating signalling pathways, particularly of the interferons, was also higher. Among cats without FIP, there were few differences between virus-positive and virus-negative MLNs; however, TLR9 and STAT2 expression were higher with infection, suggesting a direct viral effect. The study provides evidence for TLR involvement in the response to FCoV. This could open up new avenues for therapeutic approaches.

Molecular investigations of cat fleas (Ctenocephalides felis) provide the first evidence of Rickettsia felis in Malta and Candidatus Rickettsia senegalensis in Israel.

Rickettsia felis, the causative agent of flea-borne spotted fever, occurs on all continents except Antarctica, owing to the cosmopolitan distribution of its cat flea vector. In this study, cat fleas were collected in two countries where the occurrence of R. felis was either unknown (Malta) or where accurate prevalence data were lacking (Israel). Altogether 129 fleas were molecularly analysed for the presence of rickettsial DNA. On the basis of three genetic markers, R. felis was identified in 39.5% (15/38) of the cat fleas from Malta. Sequences showed 100% identity to each other and to relevant sequences in GenBank. Among the 91 cat fleas from Israel, two (2.2%) contained the DNA of Candidatus Rickettsia senegalensis. Phylogenetically, the R. felis and Candidatus R. senegalensis identified here clustered separately (with high support) but within one clade, which was a sister group to that formed by the typhus group and spotted fever group rickettsiae. This is the first record of R. felis in Malta and of Candidatus R. senegalensis outside its formerly reported geographical range including Africa, Asia and North America.

Detection of 'Candidatus Neoehrlichia mikurensis' and other Anaplasmataceae and Rickettsiaceae in Canidae in Switzerland and Mediterranean countries.

INTRODUCTION: 'Candidatus Neoehrlichia mikurensis' is an emerging tick-borne zoonotic agent that primarily affects immunocompromised human patients. Dogs and foxes are frequently exposed to ticks, and both species are in close proximity to humans. This is the first study to systematically investigate the occurrence of 'Candidatus Neoehrlichia mikurensis' in Canidae in Europa. We analyzed 1'739 blood samples from dogs in Switzerland, Italy, Spain and Portugal and 162 blood samples from free-ranging red foxes (Vulpes vulpes) in Switzerland. All samples were tested using a previously described multiplex real-time PCR for the Anaplasmataceae family, the 'Candidatus Neoehrlichia' genus and the 'Candidatus Neoehrlichia mikurensis' species. All Anaplasmataceae positive samples were subsequently tested using specific real-time PCRs for Anaplasma phagocytophilum, Anaplasma platys, Ehrlichia canis and Rickettsia helvetica. Among the tested animals, one dog from Zurich tested positive for 'Candidatus Neoehrlichia mikurensis'. The 12-year old West Highland white terrier had been splenectomized 3 months prior to the blood collection and presented with polyuria/polydipsia. Fanconi syndrome was diagnosed based on glucosuria with normoglycemia and hyperaminoaciduria. A. platys and E. canis were detected in 14/249 dogs from Sicily and Portugal; two of the dogs were coinfected with both agents. Four Swiss foxes tested positive for A. phagocytophilium. R. helvetica was detected for the first time in a red fox. In conclusion, 'Candidatus Neoehrlichia mikurensis' infection should be considered in sick dogs, particularly when immunocompromised. The pathogen seems not to be widespread in Canidae in the investigated countries. Conversely, other Anaplasmataceae were more readily detected in dogs and foxes.

INTRODUCTION: 'Candidatus Neoehrlichia mikurensis' est un agent de zoonose transmis par les tiques qui gagne en importance et concerne principalement les patients immunosupprimés. Les chiens comme les renards sont souvent concernés par des morsures de tiques et vivent en contact étroit avec les êtres humains. Dans le présent travail, nous étudions pour la première fois systématiquement la présence de 'Candidatus Neoehrlichia mikurensis' chez les canidés en Europe. Les échantillons sanguins analysés provenaient de 1'739 chiens de Suisse, d'Italie, d'Espagne et du Portugal ainsi que de 162 renards (Vulpes vulpes) de Suisse. Tous les échantillons ont été examinés avec un test de PCR multiplex en temps réel déjà publié quant à la présence d'agents de la famille des Anaplasmataceae, du genre 'Candidatus Neoehrlichia' et de l'espèce 'Candidatus Neoehrlichia mikurensis'. Les échantillons positifs aux Anaplasmataceae ont ensuite été testés avec un test PCR en temps réel spécifique quant à Anaplasma phagocytophilum, Anaplasma platys, Ehrlichia canis und Rickettsia helvetica. Parmi les échantillons examinés se trouvait celui d'un chien de Zürich qui était infecté par 'Candidatus Neoehrlichia mikurensis'. Ce West Highland White Terrier de 12 ans avait été présenté pour polyurie/polydipsie; il avait été splénectomisé trois mois avant la prise de l'échantillon. Au vu d'une glycosurie et d'une hyperaminoacidurie accompagnées d'une glycémie normale, on a posé le diagnostic de syndrome de Fanconi. A. platys et E. canis ont été mis en évidence chez 14/249 chiens provenant de Sicile et du Portugal; deux chiens étaient infectés par les deux agents pathogènes. Quatre renards suisses étaient positifs à A. phagocytophilium et R. helvetica a été trouvé pour la première fois chez un renard. En résumé, on peut dire qu'une infection à 'Candidatus Neoehrlichia mikurensis' chez un chien malade doit être prise en considération comme diagnostic différentiel, particulièrement chez les anomaux immunosupprimés. Toutefois cet agent n'est pas très répandu chez les canidés des pays examinés, contrairement aux autres Anaplasmataceae spp. qui ont été trouvées plus souvent chez les chiens et les renards.

Feline infectious peritonitis: still an enigma?

Feline infectious peritonitis (FIP) is one of the most important fatal infectious diseases of cats, the pathogenesis of which has not yet been fully revealed. The present review focuses on the biology of feline coronavirus (FCoV) infection and the pathogenesis and pathological features of FIP. Recent studies have revealed functions of many viral proteins, differing receptor specificity for type I and type II FCoV, and genomic differences between feline enteric coronaviruses (FECVs) and FIP viruses (FIPVs). FECV and FIP also exhibit functional differences, since FECVs replicate mainly in intestinal epithelium and are shed in feces, and FIPVs replicate efficiently in monocytes and induce systemic disease. Thus, key events in the pathogenesis of FIP are systemic infection with FIPV, effective and sustainable viral replication in monocytes, and activation of infected monocytes. The host's genetics and immune system also play important roles. It is the activation of monocytes and macrophages that directly leads to the pathologic features of FIP, including vasculitis, body cavity effusions, and fibrinous and granulomatous inflammatory lesions. Advances have been made in the clinical diagnosis of FIP, based on the clinical pathologic findings, serologic testing, and detection of virus using molecular (polymerase chain reaction) or antibody-based methods. Nevertheless, the clinical diagnosis remains challenging in particular in the dry form of FIP, which is partly due to the incomplete understanding of infection biology and pathogenesis in FIP. So, while much progress has been made, many aspects of FIP pathogenesis still remain an enigma.

[Haemoplasma infection in a dairy cow]. / Hämoplasmeninfektion bei einer Milchkuh.

The present work describes the clinical and laboratory examination as well as the treatment of a 7-year-old local dairy breed cow presented with reduced appetite, decreasing milk yield and striking yellowish discoloured skin and mucosa. The laboratory examination revealed a high degree regenerative anaemia and hyperbilirubinaemia. The bovine haemotrophic mycoplasma species Mycoplasma wenyonii and 'Candidatus Mycoplasma haemobos' were detected in the blood by PCR. Treatment with oxytetracycline rapidly improved the general condition, and milk production was increased. In a follow-up study, blood samples of all 23 animals from the same herd were examined. Fifteen cows were found to be infected with both haemoplasma species, three animals were only infected with 'Candidatus Mycoplasma haemobos' and one animal only with Mycoplasma wenyonii. Two out of three tested calves were positive for 'Candidatus Mycoplasma haemobos'. Except for the above described anaemic cow, all other animals were clinically healthy.

Fatal bovine anaplasmosis in a herd with new genotypes of Anaplasma marginale, Anaplasma ovis and concurrent haemoplasmosis.

Haematological and molecular analysis of blood samples was carried out during an outbreak of bovine anaplasmosis in Hungary. Acute disease was observed in five animals, two of which died. Anaplasma-carrier state was diagnosed in 69 (92%) of cattle. Further evaluation of 24 blood samples revealed concurrent infections with Mycoplasma wenyonii and 'CandidatusM. haemobos' in 22 and 21 animals, respectively. In addition, two cows were identified with rickettsaemia. Regarding molecular investigation of potential hard tick vectors, Haemaphysalis inermis and Dermacentor marginatus males collected from the animals were PCR-negative. However, in one pool (out of 18) of Ixodesricinus males, and in six pools (out of 18) of D. reticulatus males the msp4 gene of Anaplasma marginale was detected. In the same I. ricinus pool Anaplasma ovis was also identified. All ticks were negative for haemoplasmas. Anaplasma sequences yielded 97-99% homology to sequences deposited in the Genbank. This is the first report of fatal bovine anaplasmosis associated with divergent A. marginale genotypes and concurrent 'CandidatusM. haemobos' infection, as well as of an A. ovis strain in ticks collected from cattle.

In vivo induction of interferon gamma expression in grey horses with metastatic melanoma resulting from direct injection of plasmid DNA coding for equine interleukin 12.

Whole blood pharmacokinetics of intratumourally injected naked plasmid DNA coding for equine Interleukin 12 (IL-12) was assessed as a means of in vivo gene transfer in the treatment of melanoma in grey horses. The expression of induced interferon gamma (IFN-g) was evaluated in order to determine the pharmacodynamic properties of in vivo gene transduction. Seven grey horses bearing melanoma were injected intratumourally with 250 µg naked plasmid DNA coding for IL-12. Peripheral blood and biopsies from the injection site were taken at 13 time points until day 14 post injection (p.i.). Samples were analysed using quantitative real-time PCR. Plasmid DNA was quantified in blood samples and mRNA expression for IFN-g in tissue samples. Plasmid DNA showed fast elimination kinetics with more than 99 % of the plasmid disappearing within 36 hours. IFN-g expression increased quickly after IL-12 plasmid injection, but varied between individual horses. Intratumoural injection of plasmid DNA is a feasible method for inducing transgene expression in vivo. Biological activity of the transgene IL-12 was confirmed by measuring expression of IFN-g.

Molecular investigation of transplacental and vector-borne transmission of bovine haemoplasmas.

The present study was carried out in a herd with concurrent infections of Mycoplasma wenyonii and 'Candidatus M. haemobos', to investigate if transplacental and/or vector-borne transmission is possible for one or both bovine haemoplasma species. For this purpose blood samples were collected from 38 mother animals and their newborn calves; as well as from 17 uninseminated cows twice three months apart. In addition, 311 mosquitoes and blood-sucking flies (Diptera: Culicidae, Tabanidae, Muscidae) were cought near the animals. DNA was extracted from all samples, followed by real-time PCR analysis. In 10.5% of neonate calves, that were born to cows harbouring both haemoplasmas, M. wenyonii and/or 'Candidatus M. haemobos' positivity was detected. Copy numbers in positive samples from cows and their calves indicated that - in comparison with M. wenyonii - 'Candidatus M. haemobos'-bacteraemia had usually lower levels. In samples of uninseminated cows the rate of infection with the latter species decreased. These findings may explain why M. wenyonii was significantly more frequently detected in blood-sucking flies, than 'Candidatus M. haemobos'. In conclusion, molecular evidence is provided for the first time on the transplacental transmission of bovine haemoplasmas. Regarding their spread by blood-sucking arthropods, new potential vectors were identified, i.e. the horn fly (Haematobia irritans), the stable fly (Stomoxys calcitrans) and two species of horse flies (Tabanus bovinus, T. bromius).

Feline leukemia virus outbreak in the critically endangered Iberian lynx (Lynx pardinus): high-throughput sequencing of envelope variable region A and experimental transmission.

The Iberian lynx is the most endangered felid species. During winter/spring 2006/7, a feline leukemia virus (FeLV) outbreak of unexpected virulence killed about 2/3 of the infected Iberian lynxes. All FeLV-positive animals were co-infected with feline hemoplasmas. To further characterize the Iberian lynx FeLV strain and evaluate its potential virulence, the FeLV envelope gene variable region A (VRA) mutant spectrum was analyzed using the Roche 454 sequencing technology, and an in vivo transmission study of lynx blood to specified-pathogen-free cats was performed. VRA mutations indicated weak apolipoprotein B mRNA editing enzyme and catalytic polypeptide-like cytidine deaminase (APOBEC) restriction of FeLV replication, and variants characteristic of aggressive FeLV strains, such as FeLV-C or FeLV-A/61C, were not detected. Cats exposed to FeLV/Candidatus Mycoplasma haemominutum-positive lynx blood did not show a particularly severe outcome of infection. The results underscore the special susceptibility of Iberian lynxes to infectious diseases.

Survey on blood-sucking lice (Phthiraptera: Anoplura) of ruminants and pigs with molecular detection of Anaplasma and Rickettsia spp.

Lice may serve as biological or mechanical vectors for various infectious agents. To investigate louse infestation of ruminants and pigs, and pathogens potentially transmitted by them, anopluran lice (n=1182) were collected in Hungary, and evaluated for the presence of anaplasma, rickettsia and haemotropic mycoplasma DNA. On cattle the following species were found: Linognathus vituli (57%), Haematopinus eurysternus (38%) and Solenopotes capillatus (5%). L. vituli had a lower mean individual count/host when compared to H. eurysternus. On calves only L. vituli was observed, with a higher louse burden than on full-grown cattle. H. eurysternus and S. capillatus were more likely to occur simultaneously with another species on the same host, than L. vituli. Goats infested with Linognathus stenopsis had the overall highest prevalence (68%), while pigs harbouring Haematopinus suis showed the lowest (<1%). Anaplasma DNA was detected in 50% of pools analysed. In L. vituli Anaplasma ovis (or a closely related novel Anaplasma marginale genotype) was identified. Anaplasma-positivity of H. suis suggests that pigs may extend the reservoir and/or host spectrum of relevant species. Anaplasma-infected L. stenopsis pools show for the first time that caprine anaplasmosis is endemic in Hungary. Rickettsia spp. were demonstrated from Linognathus spp. and H. eurysternus. No haemotropic mycoplasmas were detected in any samples. In conclusion, this is the first molecularly confirmed report of bovine and ovine Anaplasma spp. in L. vituli, L. stenopsis and H. suis. The present results suggest that phthirapterosis of domestic animals deserves more attention, and lice should be evaluated among the broad range of potential vectors of arthropod-borne pathogens.

[Bovine fasciolosis in Switzerland: relevance and control]. / Bovine Fasciolose in der Schweiz: Bedeutung und Bekämpfung.

In Switzerland, bovine fasciolosis is an economically important but often overlooked disease of dairy cows. The intermediate host of Fasciola hepatica in Switzerland is Galba truncatula, an amphibious snail living in humid habitats which are infected by miracidia from recently hatched Fasciola eggs. The definitive hosts include cattle, sheep, goats, horses, and free-living ruminants. Infection of these hosts occur from metacercariae, usually encysted on vegetation. Infection risk depends on the location of the habitat on the farm. There is a lower risk for the intermediate host to become infected on pastures for young stock and dry cows than on pastures for dairy cows. This in turn results in a lower infection risk for young stock and dry cows than for dairy cows. When controlling the disease, epidemiologic factors such as treatment and pasture management strategies should be taken into account. If individual control measures are followed, infection pressure and prevalence in a herd can be significantly reduced. To support veterinarians and farmers in the control of fasciolosis, an interactive map showing potential risk areas for fasciolosis was created on the basis of geographical, meteorological, and biological data of the intermediate host and the free-living parasite stages.

Prevalence of Fasciola hepatica in the intermediate host Lymnaea truncatula detected by real time TaqMan PCR in populations from 70 Swiss farms with cattle husbandry.

Bovine fasciolosis is an economically important parasitic disease. Quantitative real time PCR was utilized to determine the prevalence of Fasciola hepatica in the snail intermediate host Lymnaea truncatula from 70 selected, infected Swiss cattle farms, and to gain information on the infection risk to the definitive host. Snails from 130 habitats (36 streams, 21 wells, 24 drainage ditches, 33 spring swamps, 14 reeds, 1 drainage shaft and 1 pond) originating from 71 dairy cow pastures, 39 pastures for young stock, 14 hay fields and 6 dry cow pastures were collected. Of these, 51 populations were found to be infected with F. hepatica. A total of 4733 snails were examined of which 331 were infected (7.0%). The numbers of snails collected from different sites ranged from 1 to 159 snails. Clustering of infection in snails was found on the farm of origin with a mixed logistic model with random effects. The risk of infection of L. truncatula with F. hepatica was significantly higher in populations originating from spring swamps, wells and reeds compared to populations from streams. In addition the risk of snail infection was significantly lower in populations collected in young stock and dry cow pastures compared to dairy cow pastures. The greater the population size collected from a habitat also increased the risk of an individual snail being infected.

Quantitative real-time PCR for the measurement of 11beta-HSD1 and 11beta-HSD2 mRNA levels in tissues of healthy dogs.

The 11beta-hydroxysteroid dehydrogenase (11beta-HSD) exists in two isoforms, 11beta-HSD1 and 11beta-HSD2. 11beta-HSD1 generates active cortisol from cortisone and appears to be involved in insulin resistant states. 11beta-HSD2 protects the mineralocorticoid receptor from inappropriate activation by glucocorticoids and is important to prevent sodium retention and hypertension. The purposes of the present study were to develop two real-time PCR assays to assess 11beta-HSD1 and 11beta-HSD2 mRNA expression and to evaluate the tissue distribution of the two isoforms in dogs. Thirteen different tissues of 10 healthy dogs were evaluated. Both real-time PCR assays were highly specific, sensitive and reproducible. Highest 11beta-HSD1 mRNA expression was seen in liver, lung, and renal medulla; highest 11beta-HSD2 mRNA expression in renal cortex, adrenal gland, and renal medulla. Higher 11beta-HSD1 than 11beta-HSD2 mRNA levels were found in all tissues except adrenal gland, colon, and rectum. Our results demonstrate that the basic tissue distribution of 11beta-HSD1 and 11beta-HSD2 in dogs corresponds to that in humans and rodents. In a next step 11beta-HSD1 and 11beta-HSD2 expression should be assessed in diseases like obesity, hypercortisolism, and hypertension to improve our knowledge about 11beta-HSD activity, to evaluate the dog as a model for humans and to potentially find new therapeutic options.

Genotyping of Babesia bigemina from cattle from a non-endemic area (Switzerland).

In August 2002, bovine anaplasmosis and concurrent infections with Mycoplasma sp. and piroplasms were reported in a cattle herd in an alpine region of Switzerland. The piroplasms were identified by PCR/sequencing of part of the 18S rRNA gene as Babesia bigemina and Theileria of the buffeli/sergenti/orientalis-complex, which have never been diagnosed in Switzerland before. The B. bigemina isolate was genetically characterised at two loci and compared with isolates from Italy, Spain, Turkey, Kenya and Mexico. Analysis of the internal transcribed spacer 2 (ITS2) of the rRNA genes revealed high polymorphism not only among the isolates but even within the isolates, and the presence of two types of the ITS2 in every isolate was confirmed. A dendrogram based on ITS2 sequences showed that the Swiss isolate was most closely related to a Spanish isolate but no sequences of the isolate from Switzerland were identical to any of the other isolates. The isolate from Italy was not positioned in the same cluster as the Swiss and the Spanish isolate. This had been anticipated as the nearest known endemic area of B. bigemina in Central Italy. Sequence analysis of the rhoptry-associated protein-1c gene (rap1c) confirmed the similarity of the Swiss and Spanish isolate. Hence, our molecular analyses of the Swiss B. bigemina isolate did not unequivocally track its geographical origin and the way of introduction remains obscure.

[Feline hemoplasmas in Switzerland: identification of a novel species, diagnosis, prevalence, and clinical importance]. / Feline Haemoplasmen in der Schweiz: Identifikation einer neuen Spezies, Diagnose, Prävalenz und klinische Bedeutung.

Two feline hemotropic mycoplasma spp. (aka hemoplasma) have previously been recognized. We recently discovered a third novel species in a cat with hemolytic anemia, designated 'Candidatus Mycoplasma turicensis', which is closely related to rodent haemoplasmas. This novel species induced anemia after experimental transmission to two SPF cats. Three quantitative real-time PCR assays were newly designed and applied to an epidemiological study surveying the Swiss pet cat population. Blood samples from 713 healthy and ill cats were analyzed. Up to 104 parameters per cat (detailed questionnaire, case history, laboratory parameters and retroviral infections) were evaluated. 'Candidatus Mycoplasma haemominutum' infection was more prevalent (8.5%) than Mycoplasma haemofelis (0.5%) and 'Candidatus Mycoplasma turicensis' (1%). Hemoplasma infections were associated with male gender, outdoor access, and old age, but not with disease or anemia. Infections were more frequently found in the South and West of Switzerland. Several hemoplasma infected cats, some acutely infected, others co-infected with FIV or FeLV, showed hemolytic anemia indicating that additional factors might play a role in the pathogenesis of the disease.

Detection of feline leukemia virus RNA in saliva from naturally infected cats and correlation of PCR results with those of current diagnostic methods.

A novel diagnostic test for feline leukemia virus (FeLV) RNA in saliva from naturally infected cats is described in this study. We evaluated different diagnostic tests and compared them with the widely used enzyme-linked immunosorbent assay (ELISA) for the detection of p27 in the diagnosis of FeLV. Blood samples from 445 cats were tested for the presence of provirus by real-time PCR and plasma and saliva specimens from those cats were tested for the presence of viral RNA by real-time reverse transcription (RT)-PCR and for the presence of p27 by ELISA. In comparison to conventional ELISA, the diagnostic sensitivity and specificity of the detection of salivary FeLV RNA by real-time RT-PCR were found to be 98.1 and 99.2%, respectively. Detection of viral RNA in saliva had a positive predictive value of 94.6% and a negative predictive value of 99.7%. The kappa value was 0.96, demonstrating an almost perfect agreement between both tests. Furthermore, we confirmed previous results showing that a number of cats which tested negative for the presence of p27 in plasma were in fact positive for the presence of DNA provirus in blood specimens (5.4%). However, 96.4% of these latently infected cats did not shed viral RNA in saliva; therefore, we assume that these cats are of relatively low clinical importance at the time of testing. This study shows considerable diagnostic value of the detection of saliva FeLV RNA in naturally infected cats. This new diagnostic method has advantages over the conventional ELISA, such as less invasive sample collection and no requirement for trained personnel.

Shedding of feline leukemia virus RNA in saliva is a consistent feature in viremic cats.

The purpose of this investigation was to characterize the shedding pattern of feline leukemia virus (FeLV) RNA in saliva, and to correlate it with the proviral load in whole blood, viral load in plasma, levels of p27 in saliva and plasma, the isolation of infectious FeLV from saliva, and the titers of FeLV-specific antibodies of the IgG and IgA isotypes. We evaluated 24 experimentally FeLV-infected cats for these parameters using real-time RT-PCR and PCR, cell culture assay and sandwich ELISA. We observed that shedding of viral RNA in saliva was a consistent feature in viremic cats. Latently FeLV-infected cats, displaying a very low proviral load, did not shed infectious virus in saliva, but occasionally shed viral RNA. Consequently, salivary shedding of FeLV RNA may not necessarily indicate a transmission potential for susceptible cats. This study also confirmed previous results from our laboratory, showing that a negative result for p27 in plasma, or for viral RNA in plasma or saliva does not exclude FeLV infection, considering that blood cells from those cats contained provirus. We also showed that FeLV RNA and DNA were stable for more than 64 days in saliva samples stored at room temperature. We conclude that the detection of FeLV RNA in saliva may be a useful indicator of viremia, and that the detection of salivary viral RNA by RT-PCR could become a reliable tool for the diagnosis of FeLV infection, which is facilitated by the low invasive method of collection of the samples.