At 0.1 M and 0.3 mM, no influence on the activity of the pSCS6-encoded enzyme. The activity of the pSCS7-encoded enzyme decreases to 10-15%. At 1.0 mM HgCl2, the pSCS6-encoded enzyme retains 50% of ist original activity, complete inactivation of the enzyme from pSCS7-encoded enzyme.
A disposable microarray was developed for detection of up to 90 antibiotic resistance genes in gram-positive bacteria by hybridization. Each antibiotic resistance gene is represented by two specific oligonucleotides chosen from consensus sequences of gene families, except for nine genes for which only one specific oligonucleotide could be developed. A total of 137 oligonucleotides (26 to 33 nucleotides in length with similar physicochemical parameters) were spotted onto the microarray. The microarrays (ArrayTubes) were hybridized with 36 strains carrying specific antibiotic resistance genes that allowed testing of the sensitivity and specificity of 125 oligonucleotides. Among these were well-characterized multidrug-resistant strains of Enterococcus faecalis, Enterococcus faecium, and Lactococcus lactis and an avirulent strain of Bacillus anthracis harboring the broad-host-range resistance plasmid pRE25. Analysis of two multidrug-resistant field strains allowed the detection of 12 different antibiotic resistance genes in a Staphylococcus haemolyticus strain isolated from mastitis milk and 6 resistance genes in a Clostridium perfringens strain isolated from a calf. In both cases, the microarray genotyping corresponded to the phenotype of the strains.
The ArrayTube platform presents the advantage of rapidly screening bacteria for the presence of antibiotic resistance genes known in gram-positive bacteria. This technology has a large potential for applications in basic research, food safety, and surveillance programs for antimicrobial resistance. The intensive use of antibiotics in both public health and animal husbandry has selected for antibiotic-resistant bacteria (). Under antibiotic selective pressure, bacteria have the ability to develop and exchange resistance genes, making them nonsusceptible to the antimicrobial substances deployed.
While antibiotic resistance has emerged in some important animal and human gram-positive pathogens, such as Staphylococcus and Streptococcus spp. And Clostridium perfringens, others, such as Bacillus anthracis, are currently still sensitive to antibiotics (, ).
Nevertheless, B. Anthracis can acquire resistance genes from other gram-positive bacteria in vitro, as previously described (, ) and as demonstrated in this study. It is therefore important to follow the evolution of antibiotic resistance in the bacterial population in order to prevent and repress the emergence of multidrug-resistant strains of those bacteria that can still be treated with antibiotics. Furthermore, commensal bacteria represent a reservoir of antibiotic resistance genes that have the potential to be transferred to human and animal pathogens. An effort has therefore been made in Europe to reduce the emergence and spread of resistant bacteria. The use of antimicrobial substances for nontherapeutic purposes in animal husbandry has been banned, and surveillance programs for antibiotic-resistant bacteria among both human and animal isolates have been implemented ().
Additionally, it has been proposed that bacteria used as probiotics in food or feed or as starter cultures for the food industry must be free of antibiotic resistance genes (). Bacteria used in food preparation are mainly gram positive and include Lactococcus, Lactobacillus, Pediococcus, Leuconostoc, Carnobacterium, Enterococcus, Micrococcus, Streptococcus, Staphylococcus, and Propionibacterium spp. Animal probiotics consist mainly of strains of Bacillus, Enterococcus faecium, Pediococcus, Lactobacillus, and Streptococcus.
A simple method which allows the rapid detection of antibiotic resistance genes would complement the standard MIC determination for pathogenic and commensal bacteria. In the clinic, this would have the advantage of detecting silent antibiotic resistance genes which might be turned on in vivo or spread to other bacteria and would help in prescribing the appropriate antibiotic. Such a method could also be applied to slow-growing bacteria, for which the MIC determination may cause problems.