Diversity of Staphylococcus aureus isolated from nares of ruminants.

AIMS: To examine the diversity of Staphylococcus aureus isolated from nasal swabs of ruminants in Rwanda. METHODS AND RESULTS: A total of 454 nasal swabs from 203 cows, 170 goats, and 81 sheep were examined for the presence of S. aureus, and 30 S. aureus isolates were detected and characterized pheno- and genotypically. Resistance to penicillin and/or tetracycline was observed. The isolates were assigned to eight different spa types (t21057 (novel), t10103, t18853, t20842, t318, t355, t458, and t9432) belonging to six clonal complexes (CCs) (CC152, CC30, CC3591, CC3666, CC522, and CC97). Panton-Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83), and the human and bovine variants of the toxic shock syndrome toxin gene tst-1 variants were detected. CONCLUSION: These findings demonstrate that the nares of ruminants in Rwanda are colonized with mastitis-associated S. aureus, including lineages that are also carried by humans, underscoring the zoonotic risk, especially for livestock keepers. These results highlight the crucial importance of hygiene measures when handling livestock.

Host cell interactions of novel antigenic membrane proteins of Mycoplasma agalactiae.

BACKGROUND: Mycoplasma agalactiae is the main etiological agent of Contagious Agalactia syndrome of small ruminants notifiable to the World Organization for Animal Health. Despite serious economic losses, successful vaccines are unavailable, largely because its colonization and invasion factors are not well understood. This study evaluates the role of two recently identified antigenic proteins (MAG_1560, MAG_6130) and the cytadhesin P40 in pathogenicity related phenotypes. RESULTS: Adhesion to HeLa and sheep primary mammary stromal cells (MSC) was evaluated using ELISA, as well as in vitro adhesion assays on monolayer cell cultures. The results demonstrated MAG_6130 as a novel adhesin of M. agalactiae whose capacity to adhere to eukaryotic cells was significantly reduced by specific antiserum. Additionally, these proteins exhibited significant binding to plasminogen and extracellular matrix (ECM) proteins like lactoferrin, fibrinogen and fibronectin, a feature that could potentially support the pathogen in host colonization, tissue migration and immune evasion. Furthermore, these proteins played a detrimental role on the host cell proliferation and viability and were observed to activate pro-apoptotic genes indicating their involvement in cell death when eukaryotic cells were infected with M. agalactiae. CONCLUSIONS: To summarize, the hypothetical protein corresponding to MAG_6130 has not only been assigned novel adhesion functions but together with P40 it is demonstrated for the first time to bind to lactoferrin and ECM proteins thereby playing important roles in host colonization and pathogenicity.

Ottowia testudinis sp. nov., isolated from the cloaca of a giant Asian pond turtle (Heosemys grandis).

A bacterial strain designated 27CT isolated from the cloaca of a giant Asian pond turtle was subjected to polyphasic taxonomic characterization. The strain was Gram-stain negative and oxidase- and catalase-positive. It had highest 16S rRNA gene sequence similarity to Ottowia beijingensis GCS-AN-3T (97.6 %) and Ottowia flava GY511T 96.0% and less than 96.0 % to other established species including Ramlibacter rhizophilus YS3.2.7T, Ottowia konkukae SK3863T, Acidovorax caeni E-24608T and Ottowia thiooxydans DSM 14619T. Phylogenetically, strain 27CT formed a branch with O. beijingensis GCS-AN-3T within the Ottowia clade. The genome size was 4.32 Mbp and the G+C content was 65.7 mol%. Strain 27CT shared highest ANIb values with O. beijingensis GCS-AN-3T (82.71/82.73 %) followed by O. oryzae KADR8-3T (78.9/79.0 %) and O. caeni BD-1T (73.3/75.2 %). The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid and the quinone system was ubiquinone Q-8. Predominant compounds in the polar lipid profile were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and phosphatidylmonomethylethanolamine. Major polyamines were 2-hydroxyputrescine and putrescine. In the fatty acid profile, summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C16 : 0, summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), C14 : 0, C10 : 0 3-OH and C16 : 0 2-OH were detected. All these data identify strain 27CT as representing a novel species of the genus Ottowia and hence we propose the name Ottowia testudinis sp. nov. The type strain is 27CT (=CCM 9138T=LMG 32213T).

Efficacy of Norway Spruce Ointments and Bacterial and Fungal Alterations in the Treatment of Castration Wounds in Piglets.

This study aimed to evaluate the efficacy of Norway spruce ointments on wound healing of castration wounds in piglets. This study included 95 male pigs randomly divided into five treatment groups: Norway spruce balm (Vulpuran), Norway spruce resin (Abilar), pork lard (ointment base of Vulpuran), no treatment (negative control) and antibiotic blue spray (Cyclo spray, positive control). Wound healing parameters (such as healing time, wound size, reddening of wound edges and surrounding, swelling, secretion and wound contamination), microbiological status and the haptoglobin level as an inflammation parameter were investigated. In the Norway spruce groups, some positive effects on wound healing parameters were found. In the first 6 days of treatment, Abilar or Vulpuran showed the smallest means of wound areas, and at the end of the study (day 15 + 17), the highest rates of completely closed wounds compared to the other groups. Vulpuran treatment led to significantly lower wound secretion (p = 0.003) and wound contamination (p = 0.015) than the untreated control did. Furthermore, the microbiological status was determined using MALDI-TOF-MS and partial 16S rRNA gene sequencing at different days of treatment. A comparison of the five treatment groups on day 3 revealed that Norway spruce led to the lowest rate of wounds colonised with fungi, mainly classified into genus Candida, (Abilar 77%, Vulpuran 70%) in comparison with blue spray (89%), lard (100%) and untreated control (100%). Fungi could only be detected in one of the 13 samples treated with Vulpuran on day 8, which nearly reached significance (p = 0.055).

Outbreak of Cronobacter turicensis in European brown hares (Lepus europaeus).

This is the first report of acute deaths in five European brown hares (Lepus europaeus) attributed to mucoid and necrotizing typhlocolitis caused by genetically different Cronobacter (C.) turicensis strains in northeastern Austria. As this opportunistic pathogen is mainly known for causing disease in immunocompromised humans and neonates, this previously unrecognized potential for a spill over from a wildlife reservoir to humans warrants further attention.

Luteolibacter ambystomatis sp. nov., isolated from the skin of an Anderson's salamander (Ambystoma andersoni).

A bacterial strain designated 32AT was isolated from the skin of an Anderson's salamander (Ambystoma andersoni) and subjected to a comprehensive taxonomic study. The strain was Gram-stain-negative, rod-shaped, non-motile, oxidase- and urease-negative, and catalase-positive. 16S rRNA gene sequence comparisons placed the strain in the genus Luteolibacter with highest sequence similarities to Luteolibacter pohnpeiensis A4T-83T (95.2%), Luteolibacter gellanilyticus CB-286403T (95.1%) and Luteolibacter cuticulihirudinis E100T (94.9%). Genomic sequence analysis revealed a size of 5.3 Mbp, a G+C-content of 62.2 mol% and highest ANI values with Luteolibacter luteus (71.2%), Luteolibacter yonseiensis (71.4%) and L. pohnpeiensis (69.5%). In the polyamine pattern, 1,3-diaminopropane and spermidine were predominant. The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. The quinone system was composed of the major menaquinones MK-9 and MK-10. Major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, the unidentified aminolipid AL2, the unidentified phospholipid PL2 and the unidentified aminophospholipid APL1. The fatty acid profile contained major amounts of iso-C14:0, iso-C16:0, C16 : 0 and C16 : 1 ω9c. In addition, C14 : 0, C15:0, anteiso-C15 : 0, summed feature 2 (C14 : 0 3OH and/or iso-C16 : 0 I), and the hydroxylated fatty acids iso-C14 : 0 3OH, iso-C16 : 0 3OH and C16 : 0 3-OH were detected. Physiologically, strain 32AT is distinguishable from its next relatives. Based on phylogenetic, genomic, physiological and chemotaxonomic data, strain 32AT represents a novel species of the genus Luteolibacter for which we propose the name Luteolibacter ambystomatis sp. nov. The type strain is 32AT (=CCM 9141T=LMG 32214T).

Paralysiella testudinis gen. nov., sp. nov., isolated from the cloaca of a toad-headed turtle (Mesoclemmys nasuta).

A bacterial strain designated 26BT, which had been isolated from the cloaca of a toad-headed turtle, was subjected to a comprehensive taxonomic study. Comparison of 16S rRNA gene sequences demonstrated that strain 26BT is a member of the family Neisseriaceae. Based on highest similarity values, Neisseria animaloris DSM 21642T (95.15 %), Alysiella filiformis ATCC 15532T (95.06 %), Uruburuella testudinis 07_OD624T (94.71 %), Uruburuella suis CCUG 47806T (94.66 %) and Alysiella crassa DSM 2578T (94.64 %) were identified as the closest relatives. Average nucleotide identity values based on the blast algorithm (ANIb) indicated that U. suis (76.10/76.17 %), Neisseria shayeganii 871T (74.34/74.51 %), Stenoxybacter acetivorans (73.30/73.41 %), N. animaloris (72.98/72.80) %, A. filiformis (71.14/71.21 %) and A. crassa (70.53/71.15 %) are the next closest relatives. Like ANIb, genome-based phylogeny did not suggest the affiliation of strain 26BT with any established genus. The polyamine pattern consisted of the major compounds putrescine, 1,3-diaminopropane and spermidine and the major quinone was ubiquinone Q-8. In the polar lipid profile, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an ornithine lipid were predominant. The fatty acid profile contained predominantly C16 : 1 ω7c, C12 : 0, C14 : 0, C16 : 0 and C12 : 0 3OH. The size of the genome was 2.91 Mbp and the genomic G+C content was 54.0 mol%. Since these data do not demonstrate an unambiguous association with any established genus, we here propose the novel genus Paralysiella with the type species Paralysiella testudinis gen. nov., sp. nov. The type strain is 26BT (=CCM 9137T=LMG 32212T).

Entomomonas asaccharolytica sp. nov., isolated from Acheta domesticus.

Strain F2AT, isolated from the cricket Acheta domesticus, was subjected to a polyphasic taxonomic characterization. Cells of the strain were rod-shaped, Gram-stain-negative and catalase- and oxidase-positive. It did not assimilate any carbohydrates. The strain's 16S rRNA gene sequence showed highest similarity to Entomomonas moraniae QZS01T (96.4 %). The next highest similarity values were found to representatives of related genera (<93 %). The genome size of strain F2AT was 3.2 Mbp and the G+C content was 36.4 mol%. Average nucleotide identity values based on blast and MUMmer and average amino acid identity values between strain F2AT and E. moraniae QZS01T were 74.29/74.43, 83.88 and 74.70 %, respectively. The quinone system predominantly contained ubiquinone Q-8. In the polar lipid profile, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and an unidentified phospholipid were detected. The polyamine pattern consisted of the major compounds putrescine and spermidine. Major fatty acids were C18 : 1 ω7c and C16 : 0 and the hydroxyl acids were C12 : 0 3-OH, C14 : 0 2-OH and C14 : 0 3-OH. The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. Due to its association with the only species of the genus Entomomonas but its distinctness from E. moraniae we here propose the novel species Entomomonas asaccharolytica sp. nov. F2AT (=CCM 9136T=LMG 32211T).

Presence of ß-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals.

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant ß-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-ß-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Is a Superior Diagnostic Tool for the Identification and Differentiation of Mycoplasmas Isolated from Animals.

In veterinary diagnostic laboratories, identification of mycoplasmas is achieved by demanding, cost-intensive, and time-consuming methods that rely on antigenic or genetic identification. Since matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) seems to represent a promising alternative to the currently practiced cumbersome diagnostics, we assessed its applicability for the identification of almost all mycoplasma species isolated from vertebrate animals so far. For generating main spectrum profiles (MSPs), the type strains of 98 Mycoplasma, 11 Acholeplasma, and 5 Ureaplasma species and, in the case of 69 species, 1 to 7 clinical isolates were used. To complete the database, 3 to 7 representatives of 23 undescribed Mycoplasma species isolated from livestock, companion animals, and wildlife were also analyzed. A large in-house library containing 530 MSPs was generated, and the diversity of spectra within a species was assessed by constructing dendrograms based on a similarity matrix. All strains of a given species formed cohesive clusters clearly distinct from all other species. In addition, phylogenetically closely related species also clustered closely but were separated accurately, indicating that the established database was highly robust, reproducible, and reliable. Further validation of the in-house mycoplasma library using 335 independent clinical isolates of 32 mycoplasma species confirmed the robustness of the established database by achieving reliable species identification with log scores of ≥1.80. In summary, MALDI-TOF MS proved to be an excellent method for the identification and differentiation of animal mycoplasmas, combining convenience, ease, speed, precision, and low running costs. Furthermore, this method is a powerful and supportive tool for the taxonomic resolution of animal mycoplasmas.

Vpma phase variation is important for survival and persistence of Mycoplasma agalactiae in the immunocompetent host.

Despite very small genomes, mycoplasmas retain large multigene families encoding variable antigens whose exact role in pathogenesis needs to be proven. To understand their in vivo significance, we used Mycoplasma agalactiae as a model exhibiting high-frequency variations of a family of immunodominant Vpma lipoproteins via Xer1-mediated site-specific recombinations. Phase-Locked Mutants (PLMs) expressing single stable Vpma products served as first breakthrough tools in mycoplasmology to study the role of such sophisticated antigenic variation systems. Comparing the general clinical features of sheep infected with a mixture of phase-invariable PLMs (PLMU and PLMY) and the wild type strain, it was earlier concluded that Vpma phase variation is not necessary for infection. Conversely, the current study demonstrates the in vivo indispensability of Vpma switching as inferred from the Vpma phenotypic and genotypic analyses of reisolates obtained during sheep infection and necropsy. PLMY and PLMU stably expressing VpmaY and VpmaU, respectively, for numerous in vitro generations, switched to new Vpma phenotypes inside the sheep. Molecular genetic analysis of selected 'switchover' clones confirmed xer1 disruption and revealed complex new rearrangements like chimeras, deletions and duplications in the vpma loci that were previously unknown in type strain PG2. Another novel finding is the differential infection potential of Vpma variants, as local infection sites demonstrated an almost complete dominance of PLMY over PLMU especially during early stages of both conjunctival and intramammary co-challenge infections, indicating a comparatively better in vivo fitness of VpmaY expressors. The data suggest that Vpma antigenic variation is imperative for survival and persistence inside the immunocompetent host, and although Xer1 is necessary for causing Vpma variation in vitro, it is not a virulence factor because alternative Xer1-independent mechanisms operate in vivo, likely under the selection pressure of the host-induced immune response. This singular study highlights exciting new aspects of mycoplasma antigenic variation systems, including the regulation of expression by host factors.

Classification of three corynebacterial strains isolated from the Northern Bald Ibis (Geronticus eremita): proposal of Corynebacterium choanae sp. nov., Corynebacterium pseudopelargi sp. nov., and Corynebacterium gerontici sp. nov.

Three Gram-stain-positive, rod-to-coccoid-shaped, catalase-positive and non-motile bacterial strains isolated from the choanae of a Northern bald ibis, designated strains 200CHT, W8T and 812CHT, respectively, were subjected to comprehensive taxonomic characterization. The three strains were oxidase-negative. The 16S rRNA gene sequence of 200CHT showed highest similarities to Corynebacterium epidermidicanis 410T (96.7 %) followed by Corynebacterium argentoratense DSM 44202T, Corynebacterium ulcerans NCTC 7910T and Corynebacterium pseudotuberculosis CIP 102968T (each 96.3 %). Strains W8T and 812CHT both showed highest 16S rRNA gene sequence similarities to Corynebacterium pelargi 136/3T (98.0 and 99.9 %, respectively). Comparison of the partial housekeeping gene sequence of fusA showed higher sequence similarities of 812CHT to C. pelargi (95.8 %) than W8T (90.9 %) which was also confirmed by corresponding amino acid sequences. In both, fusA gene and corresponding protein sequence strain 200CHT showed low sequence similarities to C. epidermidicanis 410T(81.6 and 87.4 %, respectively). Strains 812CHT and W8T had 76.7 % ANI similarity to each other and 88.2 and 76.4 % to C. pelargi 136/3T, respectively. In silico DNA-DNA hybridization values for 812CHT and W8T were 22.1 % among the two strains and 35.3 and 21.7 % to C. pelargi 136/3T, respectively. These data not only demonstrate that strain W8T is a representative of a novel species, but despite the high 16S rRNA gene sequence similarity to C. pelargi, strain 812CHT is also a representative of another novel species. All three strains possessed corynemycolic acids and contained meso-diaminopimelic acid as the diagnostic diamino acid of the peptidoglycan. The two strains, 200CHT and W8T, are distinguished from each other and established Corynebacterium species phylogenetically and phenotypically. In conclusion, three novel species of the genus Corynebacterium are proposed, namely Corynebacteriumpseudopelargi 812CHT (=LMG 30627T=CCM 8832T), Corynebacterium choanae 200CHT (=LMG 30628T=CCM 8831T) and Corynebacteriumgerontici W8T (=LMG 30629T=CCM 8833T), respectively.

Recommended rejection of the names Malacoplasma gen. nov., Mesomycoplasma gen. nov., Metamycoplasma gen. nov., Metamycoplasmataceae fam. nov., Mycoplasmoidaceae fam. nov., Mycoplasmoidales ord. nov., Mycoplasmoides gen. nov., Mycoplasmopsis gen. nov. [Gupta, Sawnani, Adeolu, Alnajar and Oren 2018] and all proposed species comb. nov. placed therein.

The consensus of the members of the International Committee on Systematics of Prokaryotes' Subcommittee on the taxonomy of Mollicutes is that recently proposed sweeping changes to nomenclature of members of the Mycoplasmatales, specifically involving introduction of the names Malacoplasma gen. nov., Mesomycoplasma gen. nov., Metamycoplasma gen. nov., Metamycoplasmataceaefam. nov., Mycoplasmoidaceaefam. nov., Mycoplasmoidalesord. nov., Mycoplasmoides gen. nov., Mycoplasmopsis gen. nov., and all proposed species or subspecies comb. nov. placed therein, should be rejected because they violate one or more essential points of the International Code of Nomenclature of Prokaryotes.

Urban brown rats (Rattus norvegicus) as possible source of multidrug-resistant Enterobacteriaceae and meticillin-resistant Staphylococcus spp., Vienna, Austria, 2016 and 2017.

BackgroundBrown rats (Rattus norvegicus) are an important wildlife species in cities, where they live in close proximity to humans. However, few studies have investigated their role as reservoir of antimicrobial-resistant bacteria.AimWe intended to determine whether urban rats at two highly frequented sites in Vienna, Austria, carry extended-spectrum ß-lactamase-producing Enterobacteriaceae, fluoroquinolone-resistant Enterobacteriaceae and meticillin-resistant (MR) Staphylococcus spp. (MRS).MethodsWe surveyed the presence of antimicrobial resistance in 62 urban brown rats captured in 2016 and 2017 in Vienna, Austria. Intestinal and nasopharyngeal samples were cultured on selective media. We characterised the isolates and their antimicrobial properties using microbiological and genetic methods including disk diffusion, microarray analysis, sequencing, and detection and characterisation of plasmids.ResultsEight multidrug-resistant Escherichia coli and two extensively drug-resistant New Delhi metallo-ß-lactamases-1 (NDM-1)-producing Enterobacter xiangfangensis ST114 (En. cloacae complex) were isolated from nine of 62 rats. Nine Enterobacteriaceae isolates harboured the bla CTX-M gene and one carried a plasmid-encoded ampC gene (bla CMY-2). Forty-four MRS were isolated from 37 rats; they belonged to seven different staphylococcal species: S. fleurettii, S. sciuri, S. aureus, S. pseudintermedius, S. epidermidis, S. haemolyticus (all mecA-positive) and mecC-positive S. xylosus.ConclusionOur findings suggest that brown rats in cities are a potential source of multidrug-resistant bacteria, including carbapenem-resistant En. xiangfangensis ST114. Considering the increasing worldwide urbanisation, rodent control remains an important priority for health in modern cities.

Effect of presence or absence of antibiotics and use of modified single layer centrifugation on bacteria in pony stallion semen.

Bacteria contaminate semen during collection and handling. The objective of this study was to identify the bacteria in pony stallion semen, the effects of antibiotics included in commercial semen extenders (lincomycin and spectinomycin) and the effect of modified single layer centrifugation (MSLC), on bacterial load. Ejaculates from six pony stallions, 3 ejaculates per animal, were extended in EquiPlus extender either with or without antibiotics. Aliquots were processed by MSLC to form four treatment groups: control and MSLC with antibiotics (CA and SA, respectively) and control and MSLC without antibiotics (CW and SW, respectively). Bacteriological examinations were carried out within 2 hr. Thirty-one species of bacteria were isolated from one or more ejaculates, with Corynebacterium spp. being the most frequently detected. Corynebacterium spp. were present in all ejaculates. The MSLC resulted in a significantly lower total bacterial count than controls (CA vs. SA, p < 0.001; CW vs. SW, p < 0.0001).

The effects of two different types of bandage contact lenses on the healthy canine eye.

OBJECTIVE: To compare two types of bandage contact lenses on the healthy canine eye. ANIMALS STUDIED: Six healthy Beagles. PROCEDURES: Two different types of bandage contact lenses (single sized human silicone contact lens 'PureVision 2' (Bausch & Lomb Incorporated, Rochester, NY, USA) and specially designed veterinary hydrogel contact lens 'AcriVet Pat D' (Bausch & Lomb Incorporated) were placed in 12 eyes of healthy Beagle dogs. Retention times and the effects of the lenses regarding irritation of the eye, changes in tear production, impact of contact lenses on tonometric readings, and cytologic and microbiological alterations of the canine eye were investigated. RESULTS: Mean retention times for veterinary hydrogel lenses with special dimensions were significantly shorter (2 days) than for one size human silicon lenses (8.8 days). Irritation scores were overall low for both types of lenses apart from one human lens causing severe irritation and keratoconjunctivitis as a sequel to folding and displacement. Tear production remained stable in human contact lenses. Intraocular pressure readings with a contact lens in place were only slightly altered; the most accurate readings were obtained through a human lens with an applanation tonometer. Cytology revealed a slight, nonsignificant increase in neutrophilic granulocytes with both types of lenses; the microflora did not change significantly. DISCUSSION: Human silicone lenses have significantly longer retention times and are less expensive than veterinary hydrogel lenses. In regard to irritation, bacterial growth and inflammation, both types of lenses can be recommended for use in canine eyes.

Frequency of Th17 cells correlates with the presence of lung lesions in pigs chronically infected with Actinobacillus pleuropneumoniae.

Porcine contagious pleuropneumonia caused by Actinobacillus pleuropneumoniae (APP) remains one of the major causes of poor growth performance and respiratory disease in pig herds. While the role of antibodies against APP has been intensely studied, the porcine T cell response remains poorly characterized. To address this, pigs were intranasally infected with APP serotype 2 and euthanized during the acute phase [6-10 days post-infection (dpi)] or the chronic phase of APP infection (27-31 dpi). Lymphocytes isolated from blood, tonsils, lung tissue and tracheobronchial lymph nodes were analyzed by intracellular cytokine staining (ICS) for IL-17A, IL-10 and TNF-α production after in vitro stimulation with crude capsular extract (CCE) of the APP inoculation strain. This was combined with cell surface staining for the expression of CD4, CD8α and TCR-γδ. Clinical records, microbiological investigations and pathological findings confirmed the induction of a subclinical APP infection. ICS-assays revealed the presence of APP-CCE specific CD4+CD8αdim IL-17A-producing T cells in blood and lung tissue in most infected animals during the acute and chronic phase of infection and a minor fraction of these cells co-produced TNF-α. APP-CCE specific IL-17A-producing γδ T cells could not be found and APP-CCE specific IL-10-producing CD4+ T cells were present in various organs but only in a few infected animals. The frequency of identified putative Th17 cells (CD4+CD8αdimIL-17A+) in lung and blood correlated positively with lung lesion scores and APP-specific antibody titers during the chronic phase. These results suggest a potential role of Th17 cells in the immune pathogenesis of APP infection.

Host-pathogen interplay at primary infection sites in pigs challenged with Actinobacillus pleuropneumoniae.

BACKGROUND: Actinobacillus (A.) pleuropneumoniae is the causative agent of porcine pleuropneumonia and causes significant losses in the pig industry worldwide. Early host immune response is crucial for further progression of the disease. A. pleuropneumoniae is either rapidly eliminated by the immune system or switches to a long-term persistent form. To gain insight into the host-pathogen interaction during the early stages of infection, pigs were inoculated intratracheally with A. pleuropneumoniae serotype 2 and humanely euthanized eight hours after infection. Gene expression studies of inflammatory cytokines and the acute phase proteins haptoglobin, serum amyloid A and C-reactive protein were carried out by RT-qPCR from the lung, liver, tonsils and salivary gland. In addition, the concentration of cytokines and acute phase proteins were measured by quantitative immunoassays in bronchoalveolar lavage fluid, serum and saliva. In parallel to the analyses of host response, the impact of the host on the bacterial pathogen was assessed on a metabolic level. For the latter, Fourier-Transform Infrared (FTIR-) spectroscopy was employed. RESULTS: Significant cytokine and acute phase protein gene expression was detected in the lung and the salivary gland however this was not observed in the tonsils. In parallel to the analyses of host response, the impact of the host on the bacterial pathogen was assessed on a metabolic level. For the latter investigations, Fourier-Transform Infrared (FTIR-) spectroscopy was employed. The bacteria isolated from the upper and lower respiratory tract showed distinct IR spectral patterns reflecting the organ-specific acute phase response of the host. CONCLUSIONS: In summary, this study implies a metabolic adaptation of A. pleuropneumoniae to the porcine upper respiratory tract already during early infection, which might indicate a first step towards the persistence of A. pleuropneumoniae. Not only in lung, but also in the salivary gland an increased inflammatory gene expression was detectable during the acute stage of infection.

Genetic loci of Mycoplasma agalactiae involved in systemic spreading during experimental intramammary infection of sheep.

Mycoplasmas are amongst the most successful pathogens of both humans and animals yet the molecular basis of mycoplasma pathogenesis is poorly understood. This is partly due to the lack of classical virulence factors and little similarity to common bacterial pathogenic determinants. Using Mycoplasma agalactiae as a model we initiated research in this direction by screening a transposon mutant library in the natural sheep host using a negative selection method. Having successfully identified putative factors involved in the colonization of local infection and lymphogenic sites, the current study assessed mutants unable to spread systemically in sheep after experimental intramammary infection. Analysis of distant body sites for complete absence of mutants via SSM PCR revealed that additional set of genes, such as pdhB, oppC, oppB, gtsB, MAG1890, MAG5520 and MAG3650 are required for systemic spreading apart from those that were necessary for initial colonization. Additional in vitro studies with the mutants absent at these systemic sites confirmed the potential role of some of the respective gene products concerning their interaction with host cells. Mutants of pdhB, oppC and MAG4460 exhibited significantly slower growth in the presence of HeLa cells in MEM medium. This first attempt to identify genes exclusively required for systemic spreading provides a basis for further in-depth research to understand the exact mechanism of chronicity and persistence of M. agalactiae.

Carriage of meticillin-resistant staphylococci between humans and animals on a small farm.

BACKGROUND: Meticillin-resistant staphylococci (MRS) are pathogens of increasing importance to human and animal health worldwide. Transmission of meticillin-resistant Staphylococcus aureus (MRSA) between animals and humans has been well documented. By contrast, information about transmission of other Staphylococcus spp. is limited. HYPOTHESIS/OBJECTIVES: The aim of this study was to screen animals and humans on a small farm for nasal carriage of MRS and to assess interspecies exchange. METHODS: After detection of MRSA in a lung sample of a deceased cat, which lived on a small mixed farm, nasal swabs were taken within two weeks, four and 16 months from other animals of various species and humans living on the farm. Swabs were cultured for MRS which were then characterized molecularly. RESULTS: MRSA and meticillin-resistant coagulase negative staphylococci (MRCoNS), including Staphylococcus haemolyticus, S. epidermidis and S. fleurettii, were isolated from humans and different animal species. Typing of the MRS revealed isolates with the same characteristics in different human and animal hosts. CONCLUSIONS AND CLINICAL IMPORTANCE: To the best of the authors' knowledge, this is the first report of carriage of both MRSA and MRCoNS among humans and various animals within a shared environment. The detection of strains with indistinguishable molecular characteristics strongly suggested transmission of these MRS between the various animal species and humans.