A novel isolation method of Brucella species and molecular tracking of Brucella suis biovar 2 in domestic and wild animals

Brucella suis biovar 2 is the most common aetiological agent of porcine brucellosis in Europe. B. suis biovar 2 is considered to have low zoonotic potential, but is a causative agent of reproductive losses in pigs, and it is thus economically important. The multilocus variable-number of tandem repeats genotyping analysis of 16 loci (MLVA-16) has proven to be highly discriminatory and is the most suitable assay for simultaneously identifying B. suis and tracking infections. The aim of this study was to investigate the relatedness between isolates of B. suis biovar 2 obtained during a brucellosis outbreak in domestic pigs and isolates from wild boars and hares collected from proximal or remote geographical areas by MLVA-16. A cluster analysis of the MLVA-16 data revealed that most of the isolates obtained from Switzerland clustered together, with the exception of one isolate. The outbreak isolates constituted a unique subcluster (with a genetic similarity >93.8%) distinct from that of the isolates obtained from wild animals, suggesting that direct transmission of the bacterium from wild boars to domestic pigs did not occur in this outbreak. To obtain a representative number of isolates for MLVA-16, alternative methods of Brucella spp. isolation from tissue samples were compared with conventional direct cultivation on a Brucella-selective agar. We observed an enhanced sensitivity when mechanical homogenisation was followed by host cell lysis prior to cultivation on the Brucella-selective agar. This work demonstrates that MLVA-16 is an excellent tool for both monitoring brucellosis and investigating outbreaks. Additionally, we present efficient alternatives for the isolation of Brucella spp.

Acinetobacter baumannii isolates from pets and horses in Switzerland: molecular characterization and clinical data

OBJECTIVES: We investigated whether Acinetobacter baumannii isolates of veterinary origin shared common molecular characteristics with those described in humans. METHODS: Nineteen A. baumannii isolates collected in pets and horses were analysed. Clonality was studied using repetitive extragenic palindromic PCR (rep-PCR) and multilocus sequence typing (MLST). PCR and DNA sequencing for various beta-lactamase, aminoglycoside-modifying enzyme, gyrA and parC, ISAba1 and IS1133, adeR and adeS of the AdeABC efflux pump, carO porin and class 1/2/3 integron genes were performed. RESULTS: Two main clones [A (n = 8) and B (n = 9)] were observed by rep-PCR. MLST indicated that clone A contained isolates of sequence type (ST) ST12 (international clone II) and clone B contained isolates of ST15 (international clone I). Two isolates of ST10 and ST20 were also noted. Seventeen isolates were resistant to gentamicin, 12 to ciprofloxacin and 3 to carbapenems. Isolates of ST12 carried bla(OXA-66), bla(ADC-25), bla(TEM-1), aacC2 and IS1133. Strains of ST15 possessed bla(OXA-69), bla(ADC-11), bla(TEM-1) and a class 1 integron carrying aacC1 and aadA1. ISAba1 was found upstream of bla(ADC) (one ST10 and one ST12) and/or bla(OXA-66) (seven ST12). Twelve isolates of different STs contained the substitutions Ser83Leu in GyrA and Ser80Leu or Glu84Lys in ParC. Significant disruptions of CarO porin and overexpressed efflux pumps were not observed. The majority of infections were hospital acquired and in animals with predisposing conditions for infection. CONCLUSIONS: STs and the molecular background of resistance observed in our collection have been frequently described in A. baumannii detected in human patients. Animals should be considered as a potential reservoir of multidrug-resistant A. baumannii.

Increased IL-4 and decreased regulatory cytokine production following relocation of Icelandic horses from a high to low endoparasite environment

Insect bite hypersensitivity (IBH) is an IgE-mediated dermatitis of horses caused by bites of Culicoides spp. IBH does not occur in Iceland where Culicoides are absent. However, following importation into continental Europe where Culicoides are present, >or=50% of Icelandic horses (1st generation) develop IBH but

Distinct reactivities of interleukin-4-specific antibodies with recombinant and native canine interleukin-4 in various assays

Interleukin 4 (IL-4) plays a central role in immune responses to parasites and allergens. IL-4 drives the differentiation of naive T cells into Th2 cells and regulates immunoglobulin class switching to IgE.Little is known about the role of IL-4 in canine allergies and parasite infections. Most of the information derives from measurement of IL-4 mRNA expression in dog tissues, but detection of IL-4 protein has been difficult so far, probably due to low sensitivity of available methods. Antibodies (Ab) specific for canine IL-4 are available from various sources, but these Ab have been produced against recombinant Escherichia coli-expressed canine IL-4 and there is only limited information on their reactivities with native canine IL-4. Therefore, in the present study, we tested six available canine IL-4-specific Ab for their reactivities with recombinant canine IL-4 expressed in E. coli (rec.IL-4) or in mammalian cells (mam.IL-4), and with supernatants from stimulated canine peripheral blood mononuclear cells (PBMCs) using several detection methods, including Western blotting, ELISA, cytokine bead assay, and intracellular IL-4 staining. Additionally, we tested a bovine IL-4-specific antibody that has been previously shown to cross-react with canine IL-4. All tested Ab except anti-bovine IL-4 reacted with rec.IL-4, and most of them reacted with mam.IL-4. However, only the cytokine bead assay was sensitive enough to allow the detection of IL-4 in supernatants of canine PBMCs.

Molecular cloning and characterization of equine thymic stromal lymphopoietin

Thymic stromal lymphopoietin (TSLP) is a novel cytokine that plays a central role in T helper 2 (Th2) cell differentiation and allergic inflammation. It is predominantly expressed by epithelial cells, and its expression is increased in patients with atopic dermatitis and asthma. Mice overexpressing TSLP in the skin develop allergic dermatitis and mice overexpressing TSLP in lungs develop asthma-like disease. However, it is not known whether TSLP plays an important role in equine allergies. Therefore, we cloned and sequenced the complete translated region of equine TSLP gene and measured its expression in various tissues. The equine TSLP gene is organized in 4 exons and encodes a protein of 143 amino acids, which has 62% amino acid identity with human TSLP.