Genotyping of Francisella tularensis strains by pulsed-field gel electrophoresis, amplified fragment length polymorphism fingerprinting, and 16S rRNA gene sequencing.

We evaluated three molecular methods for identification of Francisella strains: pulsed-field gel electrophoresis (PFGE), amplified fragment length polymorphism (AFLP) analysis, and 16S rRNA gene sequencing. The analysis was performed with 54 Francisella tularensis subsp. holarctica, 5 F. tularensis subsp. tularensis, 2 F. tularensis subsp. novicida, and 1 F. philomiragia strains. On the basis of the combination of results obtained by PFGE with the restriction enzymes XhoI and BamHI, PFGE revealed seven pulsotypes, which allowed us to discriminate the strains to the subspecies level and which even allowed us to discriminate among some isolates of F. tularensis subsp. holarctica. The AFLP analysis technique produced some degree of discrimination among F. tularensis subsp. holarctica strains (one primary cluster with three major subclusters and minor variations within subclusters) when EcoRI-C and MseI-A, EcoRI-T and MseI-T, EcoRI-A and MseI-C, and EcoRI-0 and MseI-CA were used as primers. The degree of similarity among the strains was about 94%. The percent similarities of the AFLP profiles of this subspecies compared to those of F. tularensis subsp. tularensis, F. tularensis subsp. novicida, and F. philomiragia were less than 90%, about 72%, and less than 24%, respectively, thus permitting easy differentiation of this subspecies. 16S rRNA gene sequencing revealed 100% similarity for all F. tularensis subsp. holarctica isolates compared in this study. These results suggest that although limited genetic heterogeneity among F. tularensis subsp. holarctica isolates was observed, PFGE and AFLP analysis appear to be promising tools for the diagnosis of infections caused by different subspecies of F. tularensis and suitable techniques for the differentiation of individual strains.

In vitro efficacy of N-duopropenide, a recently developed disinfectant containing quaternary ammonium compounds, against selected gram-positive and gram-negative organisms.

OBJECTIVE: To study the efficacy of N-duopropenide against various gram-positive and gram-negative organisms. SAMPLE POPULATION: One field strain each of Pasteurella multocida subsp multocida, Staphylococcus aureus, Listeria monocytogenes, and L ivanovii, and 2 field strains of Escherichia coli. PROCEDURE: Strains were tested with and without serum as organic matter, using quantitative suspension and carrier tests. Six concentrations of active ingredient (0.005, 0.01, 0.05, 0.11, 0.27, and 0.55%) and 6 contact times (15 and 30 seconds and 1, 3, 5, and 10 minutes) were studied for each. RESULTS: Globally, N-duopropenide was more effective in suspension tests than in carrier tests, and when organisms were suspended in saline solution rather than serum. Under the most disadvantageous conditions (carrier test with serum), a concentration of 0.55% N-duopropenide acting for only 15 seconds was effective in inactivating P multocida subsp multocida and was even more effective against the 2 Listeria species tested. For E coli strains, the same concentration also was effective, but after 10 minutes of contact. On the other hand, N-duopropenide was unable to inactivate the S aureus strain in the carrier test with serum, a concentration of 0.55% for 10 minutes was necessary to inactivate it without organic matter; however, N-duopropenide was highly effective against this organism in the suspension test, even with serum. CONCLUSION: N-duopropenide was highly effective in vitro against 5 of the most commonly encountered organisms in clinical veterinary medicine and, consequently, might be a good choice in control measures against common pathogenic organisms in modern production systems.