Cross-border outbreak of extensively drug-resistant tuberculosis linked to a university in Romania.

Extensively drug-resistant (XDR) tuberculosis (TB) poses a threat to public health due to its complicated, expensive and often unsuccessful treatment. A cluster of three XDR TB cases was detected among foreign medical students of a Romanian university. The contact investigations included tuberculin skin testing or interferon gamma release assay, chest X-ray, sputum smear microscopy, culture, drug susceptibility testing, genotyping and whole-genome sequencing (WGS), and were addressed to students, personnel of the university, family members or other close contacts of the cases. These investigations increased the total number of cases to seven. All confirmed cases shared a very similar WGS profile. Two more cases were epidemiologically linked, but no laboratory confirmation exists. Despite all the efforts done, the source of the outbreak was not identified, but the transmission was controlled. The investigation was conducted by a team including epidemiologists and microbiologists from five countries (Finland, Israel, Romania, Sweden and the UK) and from the European Centre for Disease Prevention and Control. Our report shows how countries can collaborate to control the spread of XDR TB by exchanging information about cases and their contacts to enable identification of additional cases and transmission and to perform the source investigation.

Empirical evaluation of genetic clustering methods using multilocus genotypes from 20 chicken breeds.

We tested the utility of genetic cluster analysis in ascertaining population structure of a large data set for which population structure was previously known. Each of 600 individuals representing 20 distinct chicken breeds was genotyped for 27 microsatellite loci, and individual multilocus genotypes were used to infer genetic clusters. Individuals from each breed were inferred to belong mostly to the same cluster. The clustering success rate, measuring the fraction of individuals that were properly inferred to belong to their correct breeds, was consistently approximately 98%. When markers of highest expected heterozygosity were used, genotypes that included at least 8-10 highly variable markers from among the 27 markers genotyped also achieved >95% clustering success. When 12-15 highly variable markers and only 15-20 of the 30 individuals per breed were used, clustering success was at least 90%. We suggest that in species for which population structure is of interest, databases of multilocus genotypes at highly variable markers should be compiled. These genotypes could then be used as training samples for genetic cluster analysis and to facilitate assignments of individuals of unknown origin to populations. The clustering algorithm has potential applications in defining the within-species genetic units that are useful in problems of conservation.

Bioelectrophoresis: a rapid procedure for the identification of antimicrobial agents.

Sensitivity discs were placed on 1% Noble agar for electrophoresis in a 20 mM phosphate-buffered, pH 8.5-8.8 system. The discs were removed after electrophoresis, and the agar was overlaid with Bacillus subtilis spores in Mueller-Hinton agar. After incubation at 37 C, each individual antimicrobial agent produced a distinctive identifying pattern of B. subtilis growth inhibition. The few antimicrobial agents with similar patterns could be easily differentiated by the use of a strain of Escherichia coli containing a multiple-resistance factor. Filter discs impregnated with commercial antibiotic preparations in buffer or serum yielded growth-inhibition patterns which usually resembled those obtained from corresponding sensitivity discs.

Immunoelectrophoresis of avian viral proteins in a phosphate-buffered system.

Avian influenza and hemorrhagic enteritis viral preparations were immunoelectrophoresed in a phosphate-buffered system. Excellent separation and resolution of viral proteins were achieved. Reasons are given why this method might be preferred over the conventional method employing a veronal (barbital)-buffered system.

Molecular epidemiology and mapping of tuberculosis in Israel: do migrants transmit the disease to locals?

SETTING: Israel receives migrants from various countries, some of which have high tuberculosis (TB) prevalence. OBJECTIVE: To assess the predominant Mycobacterium tuberculosis strains in Israel isolated during 2008-2010 among Israeli-born and migrant patients, and to investigate possible transmission of TB from migrants to the local population. METHODS: Molecular characterisation employed 43-spacer spoligotyping and 16-loci mycobacterial interspersed repetitive units-variable number of tandem repeats typing. All patients were classified according to those who were members of a cluster and those who were not. RESULTS: Among 684 M. tuberculosis strains isolated from new patients genotyped and assigned to their specific cohort populations during the study period, major spoligotype families were Central Asian (CAS) (n = 140, 20%), Beijing (n = 101, 15%) and T (n = 160, 23%). Most Beijing strains (66%) were isolated from patients from the former Soviet Union (FSU), while CAS strains were mainly (74%) from Ethiopia, Eritrea and Sudan (EES). For the heterogeneous T-clade, patient countries of origin were 38% EES and 33% FSU. CONCLUSIONS: Predominant M. tuberculosis genotypes in Israel in 2008-2010 were similar to genotypes endemic to the migrants' countries of origin. Epidemiological investigations did not demonstrate transmission between migrants and Israeli-born patients sharing the same cluster.

Mycoplasma gallisepticum in culture with biosilon microcarrier beads.

Various dilutions of Mycoplasma gallisepticum were cultured in the presence of Biosilon microcarrier beads. The microcarriers did not affect the recoverability or the growth rate of M. gallisepticum. Cultures attained a higher density in the presence of microcarriers. The initiation of a culture could be accomplished by the transfer of one bead from a microcarrier culture.

Destructive effect of heparin on avian erythrocytes.

The nuclear and cytoplasmic membranes of avian erythrocytes exposed to heparin became permeable to basichromatin. The permeability was dosage dependent, which suggests that the results were not an artifact of technique. The results are in agreement with previous findings that heparin causes nuclear membranes of isolated nuclei to become permeable to chromatin.

Polymyxin B: pharmacokinetics of single doses given intravenously and intramuscularly to turkeys, and minimal inhibitory concentrations for Escherichia coli and Pasteurella multocida.

The 50% and 90% minimal inhibitory concentrations (MIC50 and MIC90) of polymyxin B for avian Escherichia coli and Pasteurella multocida isolates were determined by the agar plate dilution method. Polymyxin B at approximate MIC level in serum was bactericidal for E. coli in 2 to 4 hours. Aqueous polymyxin B sulfate was administered by a single bolus intravenous injection into turkeys at 10,000 IU/kg, and by a single bolus intramuscular injection at 5,000, 10,000 or 20,000 IU/kg. Effective serum drug concentrations after intramuscular injection (MIC50 levels or greater) were maintained for E. coli for 7.0 hr (10,000 IU/kg) and 11.5 hr (20,000 IU/kg), and for P. multocida for 3.0 hr (10,000 IU/kg) and 4.1 hr (20,000 IU/kg). Pharmacokinetic parameters were calculated by non-compartmental methods. Elimination time half-lives, mean residence time, clearance, and apparent volume of distribution at steady state (Vdss) were all much higher for i.m. injection of 20,000 IU/kg than for i.m. injection of 10,000 IU/kg. We postulate that there exists a minimal tissue-interaction threshold concentration (MTC) at which polymyxin B can enter previously unavailable compartments or bind to previously refractory tissue components. Bioavailability of polymyxin B injected i.m. was 0.904 for the 10,000 IU/kg dose and 0.675 for the 20,000 IU/kg dose. Dosage intervals necessary to produce minimal steady state concentrations (Cssmin) equal to the MIC were calculated. Certain aspects of the use of the parameter Vdss, and limitations on the use of dosage interval calculations for polymyxin B, are discussed. One week after i.m. injection of polymyxin B at 10,000 IU/kg, high tissue drug levels were present, especially in bound form in liver. Following single injections, no toxic effects on turkeys were observed.

Experimental vaccination of young chickens with a live, non-pathogenic strain of Escherichia coli.

A non-pathogenic, piliated strain of Escherichia coli (BT-7; Frommer et al., 1990), isolated from a meat-type chicken flock, was studied as a candidate for a live vaccine to protect chickens from E. coli infection. Active immunization provided substantial protection of chicks vaccinated at 14 or 21 days of age, resulting in better resistance to challenge than in those vaccinated at 1 or 7 days. Chicks vaccinated at 21 days of age and challenged 1 week later with pathogenic E. coli strains 01-.K1, 02:K1 or 078:K80, exhibited good protection for at least 2 weeks against all strains. Three vaccination routes were found to give the highest resistance to challenge with pathogenic E. coli strain 078:K80. Intramuscular (i.m.) at 7 and 21 days of age, i.m. at 21 days of age and spray at 7, followed by per os at 21 days of age. Vaccination per os once at 7 or twice at 7 and 21 days resulted in good protection. Chicks exhibiting high antibody titres by ELISA were well-protected against challenge.

Adherence-associated characteristics and pathogenicity of Escherichia coli from avian colibacillosis.

Escherichia coli isolates from avian colibacillosis were examined for adherence-associated characteristics and post-colonisation pathogenicity. The adherence-associated characteristics studied included expression of pili, ability to partition with a hydrophobic phase, and ability to bind to Biosilon plastic polar-surfaced microcarriers. None of the adherence-associated characteristics correlated absolutely with post-colonisation pathogenicity, and thus in vitro tests for these adherence-associated characteristics cannot be used to predict whether any given E. coli avian colibacillosis isolate will exhibit post-colonisation pathogenicity. The isolation of strain BT7 from avian colibacillosis, a piliated E. coli strain which binds to Biosilon polar-surfaced plastic microcarriers, but is not pathogenic, is reported.

Apramycin: minimal inhibitory concentrations for avian Escherichia coli and serum levels after intramuscular injection in turkeys.

The minimal inhibitory concentrations (MIC) of apramycin, a unique aminocyclitol antibiotic, for 100 Escherichia coli isolates recovered from clinical cases of avian colibacillosis were determined using the agar dilution method. All isolates were inhibited at apramycin concentration of 8.0 micrograms/ml; 90 and 50% of the isolates were inhibited at 6.6 and 3.4 micrograms/ml, respectively. A commercial injectable product containing 200 mg apramycin/ml was administered intramuscularly (i.m.) to groups of 6- and 12-week-old turkeys at 10, 15 and 20 mg/kg. Apramycin was quickly absorbed from the i.m. injection site. Mean peak serum drug concentrations were reached 1 h after treatment and were 19.5, 27.5 and 36.0 micrograms/ml, respectively. The serum elimination half-life (t 1/2) of the drug ranged between 1.75 h for the 10 mg/kg dose and 2.5 h for the 20 mg/kg dose. Very low concentrations of the drug were found 24 h after treatment. Duration of serum apramycin concentrations in relation to the MIC, dose, and age of birds was determined.