A Mutation in the SUV39H2 Gene in Labrador Retrievers with Hereditary Nasal Parakeratosis (HNPK) Provides Insights into the Epigenetics of Keratinocyte Differentiation.

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Molecular population and colonisation factor analysis of the Staphylococcus intermedius group

The Gram-positive bacterium Staphylococcus intermedius is regarded as the major cause of canine pyoderma, a common skin infection of dogs. However, despite its clinical importance, the population genetic structure of S. intermedius is poorly understood. The current study examined the population genetic structure of S. intermedius using a multilocus DNA sequencing approach. A collection of 99 isolates phenotypically identified as S. intermedius and originating from a broad array of animal hosts in several different countries was investigated. Phylogenetic analysis indicated that the isolates belonged to three distinct species including S. intermedius, staphylococcus pseudintermedius, and Staphylococcus delphini, together referred to as the S. intermedius group (SIG). Importantly, it was discovered that all canine isolates investigated belonged to the S. pseudintermedius phylotype and it was concluded that S. pseudintermedius, not S. intermedius, is the common cause of canine pyoderma. Further, it was revealed that S. delphini is more clinically important than was previously thought. The allelic variation of agrD, which encodes the autoinducing peptide (AIP) of the agr quorum sensing system in staphylococci, was determined for all isolates. Four AIP variants were identified, including three which were present in all three species, suggesting that a common quorum sensing capacity has been conserved despite species differentiation in very different niches. Considerable clonal diversity was revealed within the S. pseudintermedius species, including several methicillin-resistant clones which have evolved by recent acquisition of the mecA gene. Using the sequence diversity identified, a simple diagnostic test was developed based on a PCR-RFLP approach to discriminate S. pseudintermedius from S. intermedius and S. delphini. Having established that S. pseudintermedius is the common canine pyoderma pathogen, this study aimed to investugate key host-pathogen interactions involved in colonisation of its canine host. Bioinformatic analysis of the whole genome sequence of a clinical isolate of S. pseudintermedius (strain ED99) revealed 17 genes encoding predicted LPXTG-containing cell wall-anchored (CWA) surface proteins. A diverse collection of S. pseudintermedius isolates and closely related staphylococcal species was screened for the presence of the genes encodng the novel CWA proteins. The majority of genes were widely distributed among the isolates examined, with nine genes being exclusive to S. pseudintermedius and eight being also present in other members of the SIG. In Gram-positive bacteria, a family of CWA proteins called microbial surgace components recognising adhesive matrix molecules (MSCRAMMs)mediates bacterial adherence to extracellular matrix proteins of the host. Three of the 17 predicted novel CWA proteins, designated SpsD, SpsL and SpsO, were selected for further characterisation of their role in host-pathogen interactions and were cloned and expressed on the surface of the surrogate host Lactococcus lactis. Solid phase adherence assays employing host extracellular matrix proteins and canine corneocytes were performed to identify host extracellular matrix proteins and canine corneocytes were performed to identify host receptors for the putative MSCRAMMs. L. lactis expressing SpsD demonstrated binding to fibronectin, fibrinogen and cytokeratin 10, SpsL mediated binding of L. lactis to fibronectin and canine fibrrinogen, and SpsD and SpsO both mediated L. lactis adherence to canine corneocytes. Additionally, a cell culture assay using a commercially available canine epidermal cell line was developed and the adherence of S. pseudintermedius ED99 and the L. lactis constructs to the cell line was tested. S. pseudintermedius ED99, but none of the MSCRAMM-expressing L. lactis strains, adhered to the canine epidermal cells in vitro, suggesting that receptors for S. pseudintermedius adherence which are present in ex vivo corneocytes are not present in undifferentiated canine epidermal cell line preparations. Take together, the present study provides broad new insights into the classification and evolution of the SIG, and the molecular interaction of S. pseudintermedius with its canine host

Sarcoptes infestation in two miniature pigs with zoonotic transmission – a case report

Abstract Background Scabies is a contagious skin disease rarely described in miniature pigs. To the best of the authors’ knowledge, a zoonotic transfer from infected pet pigs to humans has not been reported previously. Case presentation This case report describes the infestation with Sarcoptes scabiei mites in two miniature pigs presenting with unusual clinical signs, and disease transmission to a child. Two 7-month-old male castrated miniature pig siblings were examined. Both had developed skin lesions, one animal was presented for neurological signs and emaciation. They were housed together in an indoor- and outdoor enclosure. Dermatological examination revealed a dull, greasy coat with generalized hypotrichosis and multifocal erythema. Microscopic examination of skin scrapings, impression smears of affected skin and ear swabs revealed high numbers of Sarcoptes mites in both animals as well as bacterial overgrowth. A subcutaneous injection of ivermectin 0.3 mg/kg was administered to both animals and repeated after 2 weeks. Both miniature pigs received subcutaneous injections with butafosfan and cyanocobalamin, were washed with a 3% chlorhexidine shampoo and were fed on a well-balanced diet. Pig enclosures were cleaned. The infested child was examined by a physician and an antipruritic cream was prescribed. Both miniature pigs and the child went into clinical remission after treatment. Conclusion Sarcoptic mange is rare or even eradicated in commercial pig farming in many countries but miniature pigs may represent a niche for Sarcoptes scabiei infections. This case report indicates that miniature pigs kept as pets can efficiently transmit zoonotic disease to humans. In addition, these animals may represent a niche for Sarcoptes scabiei infestation in countries where sarcoptic mange in commercial pig farms has been eradicated and could therefore pose, a hazard for specific pathogen free farms

Molecular population and colonisation factor analysis of the Staphylococcus intermedius group

The Gram-positive bacterium Staphylococcus intermedius is regarded as the major cause of canine pyoderma, a common skin infection of dogs. However, despite its clinical importance, the population genetic structure of S. intermedius is poorly understood. The current study examined the population genetic structure of S. intermedius using a multilocus DNA sequencing approach. A collection of 99 isolates phenotypically identified as S. intermedius and originating from a broad array of animal hosts in several different countries was investigated. Phylogenetic analysis indicated that the isolates belonged to three distinct species including S. intermedius, staphylococcus pseudintermedius, and Staphylococcus delphini, together referred to as the S. intermedius group (SIG). Importantly, it was discovered that all canine isolates investigated belonged to the S. pseudintermedius phylotype and it was concluded that S. pseudintermedius, not S. intermedius, is the common cause of canine pyoderma. Further, it was revealed that S. delphini is more clinically important than was previously thought. The allelic variation of agrD, which encodes the autoinducing peptide (AIP) of the agr quorum sensing system in staphylococci, was determined for all isolates. Four AIP variants were identified, including three which were present in all three species, suggesting that a common quorum sensing capacity has been conserved despite species differentiation in very different niches. Considerable clonal diversity was revealed within the S. pseudintermedius species, including several methicillin-resistant clones which have evolved by recent acquisition of the mecA gene. Using the sequence diversity identified, a simple diagnostic test was developed based on a PCR-RFLP approach to discriminate S. pseudintermedius from S. intermedius and S. delphini. Having established that S. pseudintermedius is the common canine pyoderma pathogen, this study aimed to investugate key host-pathogen interactions involved in colonisation of its canine host. Bioinformatic analysis of the whole genome sequence of a clinical isolate of S. pseudintermedius (strain ED99) revealed 17 genes encoding predicted LPXTG-containing cell wall-anchored (CWA) surface proteins. A diverse collection of S. pseudintermedius isolates and closely related staphylococcal species was screened for the presence of the genes encodng the novel CWA proteins. The majority of genes were widely distributed among the isolates examined, with nine genes being exclusive to S. pseudintermedius and eight being also present in other members of the SIG. In Gram-positive bacteria, a family of CWA proteins called microbial surgace components recognising adhesive matrix molecules (MSCRAMMs)mediates bacterial adherence to extracellular matrix proteins of the host. Three of the 17 predicted novel CWA proteins, designated SpsD, SpsL and SpsO, were selected for further characterisation of their role in host-pathogen interactions and were cloned and expressed on the surface of the surrogate host Lactococcus lactis. Solid phase adherence assays employing host extracellular matrix proteins and canine corneocytes were performed to identify host extracellular matrix proteins and canine corneocytes were performed to identify host receptors for the putative MSCRAMMs. L. lactis expressing SpsD demonstrated binding to fibronectin, fibrinogen and cytokeratin 10, SpsL mediated binding of L. lactis to fibronectin and canine fibrrinogen, and SpsD and SpsO both mediated L. lactis adherence to canine corneocytes. Additionally, a cell culture assay using a commercially available canine epidermal cell line was developed and the adherence of S. pseudintermedius ED99 and the L. lactis constructs to the cell line was tested. S. pseudintermedius ED99, but none of the MSCRAMM-expressing L. lactis strains, adhered to the canine epidermal cells in vitro, suggesting that receptors for S. pseudintermedius adherence which are present in ex vivo corneocytes are not present in undifferentiated canine epidermal cell line preparations. Take together, the present study provides broad new insights into the classification and evolution of the SIG, and the molecular interaction of S. pseudintermedius with its canine host.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Molecular Diagnostic Identification of Staphylococcus pseudintermedius▿

We report the first diagnostic test for the identification of Staphylococcus pseudintermedius involving a simple PCR-restriction fragment length polymorphism approach. The method allows discrimination of S. pseudintermedius from the closely related members of the Staphylococcus intermedius group and other important staphylococcal pathogens of humans and dogs

Complete Genome Sequence of the Canine Pathogen Staphylococcus pseudintermedius▿

We report the first whole-genome sequence for a clinical isolate of Staphylococcus pseudintermedius (ED99), the major pathogen responsible for canine bacterial pyoderma. S. pseudintermedius contains numerous mobile genetic elements and encodes an array of putative virulence factors, including superantigenic, cytolytic, and exfoliative toxins and cell wall-associated surface proteins

Abnormal keratinocyte differentiation in the nasal planum of Labrador Retrievers with hereditary nasal parakeratosis (HNPK).

Hereditary nasal parakeratosis (HNPK) is an inherited disorder described in Labrador Retrievers and Greyhounds. It has been associated with breed-specific variants in the SUV39H2 gene encoding a histone 3 methyltransferase involved in epigenetic silencing. Formalin-fixed biopsies of the nasal planum of Labrador Retrievers were screened by immunofluorescence microscopy for the presence and distribution of epidermal proliferation and differentiation markers. Gene expression of these markers was further analysed using RNA sequencing (RNA-seq) and ultrastructural epidermal differences were investigated by electron microscopy. Differentiation of the nasal planum in the basal and suprabasal epidermal layers of HNPK-affected dogs (n = 6) was similar compared to control dogs (n = 6). In the upper epidermal layers, clear modifications were noticed. Loricrin protein was absent in HNPK-affected nasal planum sections in contrast to sections of the same location of control dogs. However, loricrin was present in the epidermis of paw pads and abdominal skin from HNPK dogs and healthy control dogs. The patterns of keratins K1, K10 and K14, were not markedly altered in the nasal planum of HNPK-affected dogs while the expression of the terminal differentiation marker involucrin appeared less regular. Based on RNA-seq, LOR and IVL expression levels were significantly decreased, while KRT1, KRT10 and KRT14 levels were up-regulated (log2fold-changes of 2.67, 3.19 and 1.71, respectively) in HNPK-affected nasal planum (n = 3) compared to control dogs (n = 3). Electron microscopical analysis revealed structural alterations in keratinocytes and stratum corneum, and disrupted keratinocyte adhesions and distended intercellular spaces in lesional samples (n = 3) compared to a sample of a healthy control dog (n = 1). Our findings demonstrate aberrant keratinocyte terminal differentiation of the nasal planum of HNPK-affected Labrador Retrievers and provide insights into biological consequences of this inactive SUV39H2 gene variant

Population Genetic Structure of the Staphylococcus intermedius Group: Insights into agr Diversification and the Emergence of Methicillin-Resistant Strains▿ †

The population genetic structure of the animal pathogen Staphylococcus intermedius is poorly understood. We carried out a multilocus sequence phylogenetic analysis of isolates from broad host and geographic origins to investigate inter- and intraspecies diversity. We found that isolates phenotypically identified as S. intermedius are differentiated into three closely related species, S. intermedius, Staphylococcus pseudintermedius, and Staphylococcus delphini. S. pseudintermedius, not S. intermedius, is the common cause of canine pyoderma and occasionally causes zoonotic infections of humans. Over 60 extant STs were identified among the S. pseudintermedius isolates examined, including several that were distributed on different continents. As the agr quorum-sensing system of staphylococci is thought to have evolved along lines of speciation within the genus, we examined the allelic variation of agrD, which encodes the autoinducing peptide (AIP). Four AIP variants were encoded by S. pseudintermedius isolates, and identical AIP variants were shared among the three species, suggesting that a common quorum-sensing capacity has been conserved in spite of species differentiation in largely distinct ecological niches. A lack of clonal association of agr alleles suggests that assortive recombination may have contributed to the distribution of agr diversity. Finally, we discovered that the recent emergence of methicillin-resistant strains was due to multiple acquisitions of the mecA gene by different S. pseudintermedius clones found on different continents. Taken together, these data have resolved the population genetic structure of the S. intermedius group, resulting in new insights into its ancient and recent evolution

Evolutionary conservation of the asparagine residue at position 324 in the SUV39H2 protein.

<p>Position 324 within the catalytically active SET domain is perfectly conserved across vertebrates in all known SUV39H2 orthologs. In the lower part of the alignment the sequences of closely related paralogous histone methyltransferases with similar substrate specificity also demonstrate conservation of the asparagine at this position. The sequences were derived from the following database accessions: <i>C. lupus</i> SUV39H2 XP_535179.2, <i>H. sapiens</i> SUV39H2 NP_001180353.1, <i>B. taurus</i> SUV39H2 NP_001032556.1, <i>M. musculus</i> Suv39h2 NP_073561.2, <i>G. gallus</i> SUV39H2 NP_001026541.1, <i>X. laevis</i> suv39h2 NP_001091337.1, <i>H. sapiens</i> SUV39H1 NP_003164.1, <i>H. sapiens</i> EHMT1 NP_079033.4, <i>H. sapiens</i> EHMT2 NP_006700.3, <i>H. sapiens</i> SETDB1 NP_001138887.1, <i>H. sapiens</i> SETDB2 NP_114121.2.</p

Mapping of HNPK in Labrador Retrievers.

<p>(A) A genome-wide association study using 13 cases and 23 controls indicates a strong signal with multiple associated SNPs on CFA 2. (B) The detailed view of CFA 2 delineates an associated interval of ∼4 Mb. (C) Homozygosity mapping. Each horizontal bar corresponds to one of the 13 analyzed cases. Homozygous regions with shared alleles are shown in black. A shared homozygous interval of ∼1.6 Mb delineates the exact boundaries of the critical interval from 20,818,258–22,414,948 bp (CanFam 3.1 assembly). (D) Gene content of the corresponding human interval on HSA 10 (NCBI annotation, genome build 37.5).</p