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Myobia sp. and Demodex sp. are two skin mites that infest mice, particularly immunodeficient or transgenic lab mice. In the present study, wild house mice from five localities from the Barcelona Roberstonian system were analysed in order to detect skin mites and compare their prevalence between standard (2n = 40) and Robertsonian mice (2n > 40). We found and identified skin mites through real-time qPCR by comparing sequences from the mitochondrial 16S rRNA and the nuclear 18S rRNA genes since no sequences are available so far using the mitochondrial gene. Fourteen positive samples were identified as Myobia musculi except for a deletion of 296 bp out to 465 bp sequenced, and one sample was identified as Demodex canis. Sampling one body site, the mite prevalence in standard and Robertsonian mice was 0 and 26%, respectively. The malfunction of the immune system elicits an overgrowth of skin mites and consequently leads to diseases such as canine demodicosis in dogs or rosacea in humans. In immunosuppressed mice, the probability of developing demodicosis is higher than in healthy mice. Since six murine toll-like receptors (TLRs) are located in four chromosomes affected by Robertsonian fusions, we cannot dismiss that differences in mite prevalence could be the consequence of the interruption of TLR function. Although ecological and/or morphological factors cannot be disregarded to explain differences in mite prevalence, the detection of translocation breakpoints in TLR genes or the analysis of TLR gene expression are needed to elucidate how Robertsonian fusions affect the immune system in mice.

Dogs present almost all their skin sites covered by hair, but canine skin disorders are more common in certain skin sites and breeds. The goal of our study is to characterize the composition and variability of the skin microbiota in healthy dogs and to evaluate the effect of the breed, the skin site, and the individual. We have analyzed eight skin sites of nine healthy dogs from three different breeds by massive sequencing of 16S rRNA gene V1–V2 hypervariable regions. The main phyla inhabiting the skin microbiota in healthy dogs are Proteobacteria, Firmicutes, Fusobacteria, Actinobacteria, and Bacteroidetes. Our results suggest that skin microbiota composition pattern is individual specific, with some dogs presenting an even representation of the main phyla and other dogs with only a major phylum. The individual is the main force driving skin microbiota composition and diversity rather than the skin site or the breed. The individual is explaining 45% of the distances among samples, whereas skin site explains 19% and breed 9%. Moreover, analysis of similarities suggests a strong dissimilarity among individuals (R = 0.79, P = 0.001) that is mainly explained by low-abundant species in each dog. Skin site also plays a role: inner pinna presents the highest diversity value, whereas perianal region presents the lowest one and the most differentiated microbiota composition.

In the original article, there was a mistake in Figure 3 as published. Although Figure 3 is scientifically meaningful, it does not fit the legend and the associated text on the manuscript. The corrected Figure 3 appears below. The authors apologize for this error and state that this does not change the scientific conclusions of the article in any way.

Leishmania infantum -specific antibodies are used extensively for the diagnosis and monitoring of treatment in canine leishmaniosis. Different views have been described for the measurement of L. infantum antibody levels for the monitoring of anti-leishmanial treatment. In addition, molecular techniques using blood are frequently employed in the clinical setting. However, there are not enough studies to prove the usefulness of PCR in diagnosis, treatment monitoring and in assessing the prognosis of the disease. The objectives of this study were to evaluate L. infantum -specific antibodies and blood parasitemia at the time of diagnosis and during treatment and to correlate these with the dog's clinical status. Thirty-seven dogs were diagnosed and followed-up during treatment (days 30, 180 and 365). The treatment protocol consisted of a combination of meglumine antimoniate for one month and allopurinol for at least one year. Leishmania infantum -specific antibodies and blood parasitemia were assessed by an end point sera dilution ELISA and by real-time PCR, respectively. The majority of dogs were classified as LeishVet stage II (moderate disease) at the time of diagnosis (86 %) and the rest as stage III. Results showed variable levels of specific antibodies at the time of diagnosis [median ± interquartile range (IQR): 1372 ± 8803 ELISA units (EU)]. Twenty-three seropositive dogs (64 %) were detected as PCR-positive at the time of diagnosis. Interestingly, a rapid significant antibody level reduction was observed by day 30 of treatment (median ± IQR: 604 ± 2168 EU). A continuing significant decrease of specific antibodies was also found at days 180 (median ± IQR: 201 ± 676 EU) and 365 (median ± IQR: 133 ± 329 EU) in association with clinical improvement. A significant blood parasitemia reduction was also observed at all time points studied. Mean parasites/ml ± SD were 19.4 ± 79.1 on day 0, 2.2 ± 11.7 on day 30, 0.9 ± 2.9 on day 180, and 0.3 ± 0.7 on day 365. This study

Vector-borne pathogens are the subject of several investigations due to the zoonotic concern of some of them. However, limited data are available about the simultaneous presence of these pathogens in cats and their ectoparasites. The aim of the present study was to define the species of ectoparasites found on cats as well as to investigate vector-borne pathogens in cats and their ectoparasites in southern Italy. Blood from 42 cats and fleas or flea pools (n = 28) and ticks (n = 73) collected from them were investigated by quantitative PCR for the detection of vector-borne pathogens. Feline serum samples were tested by IFAT to detect IgG antibodies against Leishmania infantum, Bartonella henselae, Rickettsia conorii, Rickettsia felis, Rickettsia typhi, Babesia microti, Ehrlichia canis and Anaplasma phagocytophilum antigens. Only one flea species (Ctenocephalides felis) and four tick species belonging to the genera Rhipicephalus and Ixodes were identified on cats from southern Italy. Molecular evidence of Bartonella spp., Rickettsia spp., hemoplasmas, Babesia vogeli and L. infantum was found in ectoparasites (fleas and/or ticks) while DNA from Hepatozoon felis and Ehrlichia/Anaplasma spp. was not detected. Likewise, DNAs from Bartonella, hemoplasma and Leishmania were the only pathogens amplified from feline blood samples. Cats had also antibodies against all the investigated pathogens with the exception of Rickettsia typhi. Agreement between serological and molecular results in individual cats and their ectoparasites was not found. The only exception was for Bartonella with a fair to moderate agreement between individual cats and their ectoparasites. Bartonella clarridgeiae was the species most frequently found in cats and their fleas followed by B. henselae. In conclusion, cats harboring ticks and fleas are frequently exposed to vector-borne pathogens. Furthermore, ticks and fleas harbored by cats frequently carry pathogens of zoonotic concern therefore appropriate

A novel methodology for the isothermal amplification of Leishmania DNA using labeled primers combined with the advantages of magnetic purification/preconcentration and the use of gold nanoparticle (AuNP) tags for the sensitive electrochemical detection of such amplified DNA is developed. Primers labeled with AuNPs and magnetic beads (MBs) are used for the first time for the isothermal amplification reaction, being the amplified product ready for the electrochemical detection. The electrocatalytic activity of the AuNP tags toward the hydrogen evolution reaction allows the rapid quantification of the DNA on screen-printed carbon electrodes. Amplified products from the blood of dogs with Leishmania (positive samples) are discriminated from those of healthy dogs (blank samples). Quantitative studies demonstrate that the optimized method allows us to detect less than one parasite per microliter of blood (8 × 10 parasites in the isothermal amplification reaction). This pioneering approach is much more sensitive than traditional methods based on real-time polymerase chain reaction (PCR), and is also more rapid, cheap, and user-friendly.

In rural parts of Africa, dogs live in close association with humans and livestock, roam freely, and usually do not receive prophylactic measures. Thus, they are a source of infectious disease for humans and for wildlife such as protected carnivores. In 2011, an epidemiological study was carried out around three conservation areas in Uganda to detect the presence and determine the prevalence of vector-borne pathogens in rural dogs and associated ticks to evaluate the risk that these pathogens pose to humans and wildlife. Serum samples (n = 105), blood smears (n = 43) and blood preserved on FTA cards (n = 38) and ticks (58 monospecific pools of Haemaphysalis leachi and Rhipicephalus praetextatus including 312 ticks from 52 dogs) were collected from dogs. Dog sera were tested by indirect immunofluorescence to detect the presence of antibodies against Rickettsia conorii and Ehrlichia canis. Antibodies against R. conorii were also examined by indirect enzyme immunoassay. Real time PCR for the detection of Rickettsia spp., Anaplasmataceae, Bartonella spp. and Babesia spp. was performed in DNA extracted from FTA cards and ticks. 99 % of the dogs were seropositive to Rickettsia spp. and 29.5 % to Ehrlichia spp. Molecular analyses revealed that 7.8 % of the blood samples were infected with Babesia rossi, and all were negative for Rickettsia spp. and Ehrlichia spp. Ticks were infected with Rickettsia sp. (18.9 %), including R. conorii and R. massiliae ; Ehrlichia sp. (18.9 %), including E. chaffeensis and Anaplasma platys ; and B. rossi (1.7 %). Bartonella spp. was not detected in any of the blood or tick samples. This study confirms the presence of previously undetected vector-borne pathogens of humans and animals in East Africa. We recommend that dog owners in rural Uganda be advised to protect their animals against ectoparasites to prevent the transmission of pathogens to humans and wildlife.

Limited information is available about the species of ticks infesting the cat and the pathogens that they harbor. The aims of the present study were to identify the species of ticks removed from cats living in Sicily and Calabria (Italy) and to detect DNA of vector-borne pathogens in the same ticks. Morphological identification of 132 adult ticks collected throughout the year from cats was carried out. Real-time PCRs for Hepatozoon felis, Piroplasmid, Ehrlichia/Anaplasma spp., Rickettsia spp., Bartonella spp., Mycoplasma spp. and Leishmania infantum were performed from each individual tick. Ticks belonging to Rhipicephalus (R. sanguineus sensu lato, R. pusillus) and Ixodes (I. ricinus, I. ventalloi) genera were identified. Ixodes ventalloi was the most frequently found tick species (47 %). The positivity rate to at least one pathogen was 14.4 % (19/132 ticks). Leishmania infantum, Rickettsia spp. (R. monacensis and R. helvetica), Bartonella spp. (B. clarridgeiae), Piroplasmid (Babesia vogeli), and Ehrlichia/Anaplasma spp. (E. canis) DNAs were amplified in 8.3, 5.3, 1.5, 0.75 and 0.75 % of ticks, respectively. Hepatozoon felis, Anaplasma spp. and hemotropic Mycoplasma spp. DNAs were not detected. Four (21.1 %) out of nineteen positive ticks were co-infected. This study provides novel data about ticks infesting cats and the DNA of pathogens that they harbor. In Southern Italy, anti-tick prophylaxis should be implemented throughout the year in cats without neglecting winter time.

The diagnosis of feline haemoplasmosis has improved over the years, with several techniques enabling a clear and specific diagnosis, and where polymerase chain reaction (PCR) is considered as the ‘gold standard’. The aim of this study was to survey the prevalence of feline haemoplasmas in 320 cats from the north-central region of Portugal by the use of real-time PCR, as well as to evaluate any associations between infection, clinical presentation and risk factors. The overall prevalence of infection by feline haemoplasmas was 43.43% (139/320), where 41.56% (133/320) corresponded to Candidatus Mycoplasma haemominutum (CMhm), 12.81% (41/320) to Mycoplasma haemofelis (Mhf), 4.38% (14/320) to Candidatus Mycoplasma haematoparvum and 1.25% (4/320) to Candidatus Mycoplasma turicensis. Almost 13% (47/320) of the samples were co-infected, with the most common co-infection being CMhm and Mhf (23.74%). Infection was found statistically significant with feline immunodeficiency/feline leukaemia virus status (P = 0.034), but no significant association was found for breed, sex, fertility status (neutered/spayed/entire), age, clinical status, living conditions (in/outdoor), anaemia status, or the presence/absence of ticks or fleas. Cats from north-central Portugal are infected with all the known feline haemoplasma species, with CMhm being the most common one. Prevalence of all feline haemoplasmas was higher than that reported previously in cats from other European countries, but similar to that described in Portugal for dogs. These data provide a better perspective regarding Mycoplasma species infection in Europe, and new information that helps us better understand feline haemoplasmosis.

[Background]: In recent years, several clinical cases and epidemiological studies of feline vector-borne diseases (FVBD) have been reported worldwide. Nonetheless, information on FVBD agents and their prevalence in Portugal is scarce., [Methods]: Three-hundred and twenty domestic cats presented to 30 veterinary medical centres in the north and centre regions of Portugal were randomly sampled. Blood was assayed by real-time polymerase chain reaction (PCR) for genera Anaplasma/Ehrlichia, genus Babesia, Hepatozoon canis, Hepatozoon felis, Leishmania infantum and the genus Rickettsia. Babesia-positive samples were further tested for Babesia canis and Babesia vogeli., [Results]: Eighty (25.0%) out of the 320 cats were positive to at least one vector-borne agent, including seven (2.2%) cats co-infected with two agents. Two cats (0.6%) were infected with Anaplasma/Ehrlichia spp., four (1.3%) with B. canis, 26 (8.1%) with B. vogeli, 50 (15.6%) with H. felis, one (0.3%) with L. infantum and four (1.3%) with Rickettsia spp. No cat tested positive for H. canis. One cat (0.3%) was co-infected with B. canis and B. vogeli, three (0.9%) with B. vogeli and H. felis, one (0.3%) with H. felis and L. infantum, and two (0.6%) with H. felis and Rickettsia spp., [Conclusions]: A considerable prevalence of infection with vector-borne pathogens among the domestic feline population of the north and centre of Portugal has been revealed by the present study. Additionally, this is the first detection of B. vogeli in cats from Europe and of H. felis in cats from Portugal.

[Background]: The current disease model for leishmaniasis suggests that only a proportion of infected individuals develop clinical disease, while others are asymptomatically infected due to immune control of infection. The factors that determine whether individuals progress to clinical disease following Leishmania infection are unclear, although previous studies suggest a role for host genetics. Our hypothesis was that canine leishmaniasis is a complex disease with multiple loci responsible for the progression of the disease from Leishmania infection., [Methodology/Principal Findings]: Genome-wide association and genomic selection approaches were applied to a population-based case-control dataset of 219 dogs from a single breed (Boxer) genotyped for ∼170,000 SNPs. Firstly, we aimed to identify individual disease loci; secondly, we quantified the genetic component of the observed phenotypic variance; and thirdly, we tested whether genome-wide SNP data could accurately predict the disease., [Conclusions/Significance]: We estimated that a substantial proportion of the genome is affecting the trait and that its heritability could be as high as 60%. Using the genome-wide association approach, the strongest associations were on chromosomes 1, 4 and 20, although none of these were statistically significant at a genome-wide level and after correcting for genetic stratification and lifestyle. Amongst these associations, chromosome 4: 61.2–76.9 Mb maps to a locus that has previously been associated with host susceptibility to human and murine leishmaniasis, and genomic selection estimated markers in this region to have the greatest effect on the phenotype. We therefore propose these regions as candidates for replication studies. An important finding of this study was the significant predictive value from using the genomic information. We found that the phenotype could be predicted with an accuracy of ∼0.29 in new samples and that the affection status was correctly predicted in 60% of dogs, significantly higher than expected by chance, and with satisfactory sensitivity-specificity values (AUC = 0.63).