PurposeWe wanted to improve the diagnosis of implant-related infection using molecular biological techniques after sonication.MethodsWe studied 258 retrieved implant components (185 prosthetic implants and 73 osteosynthesis implants) from 126 patients. 47 patients had a clinical diagnosis of infection (108 components) and 79 patients did not (150 components). The fluids from sonication of retrieved implants were tested in culture and were also analyzed using a modified commercial PCR kit for detection of Gram-positive and Gram-negative bacteria (GenoType BC; Hain Lifescience) after extraction of the DNA.Results38 of 47 patients with a clinical diagnosis of infection were also diagnosed as being infected using culture and/or PCR (35 by culture alone). Also, 24 patients of the 79 cases with no clinical diagnosis of infection were identified microbiologically as being infected (4 by culture, 16 by PCR, and 4 by both culture and PCR). Comparing culture and PCR, positive culture results were obtained in 28 of the 79 patients and positive PCR results were obtained in 35. There were 21 discordant results in patients who were originally clinically diagnosed as being infected and 28 discordant results in patients who had no clinical diagnosis of infection.InterpretationFor prosthetic joint infections and relative to culture, molecular detection can increase (by one tenth) the number of patients diagnosed as having an infection. Positive results from patients who have no clinical diagnosis of infection must be interpreted carefully.

Background: During the last few years, PCR-based methods have been developed to simplify and reduce the time required for genotyping Mycobacterium tuberculosis (MTB) by standard approaches based on IS6110-Restriction Fragment Length Polymorphism (RFLP). Of these, MIRU-12-VNTR (Mycobacterial interspersed repetitive units-variable number of tandem repeats) (MIRU-12) has been considered a good alternative. Nevertheless, some limitations and discrepancies with RFLP, which are minimized if the technique is complemented with spoligotyping, have been found. Recently, a new version of MIRU-VNTR targeting 15 loci (MIRU-15) has been proposed to improve the MIRU-12 format.

Recent transmission among immigrants and transmission permeability between the immigrant and autochthonous populations were found. Epidemiologic strategies that combine universal genotyping and refined surveys of the clustered patients are needed to investigate transmission patterns in complex scenarios.

In recent years, the application of molecular tools has shown us that clonal complexity in infection byMycobacterium tuberculosis is not anecdotal. Exogenous reinfections, mixed infections, compartmentalization, and microevolution are different aspects of this issue. The detection and characterization of clonal variants of M. tuberculosis by standard genotyping methods is laborious and frequently requires expertise. Our aim was to evaluate a new genotyping PCR-based method for M. tuberculosis, mycobacterial interspersed repetitive unitvariable-number tandem repeat typing (MIRU-VNTR), as a potential tool to simplify and optimize the clonal analysis of tuberculosis. MIRU-VNTR was able to detect mixed clonal variants in vitro, even for clones at low ratios (1:99). This technique was prospectively applied to search for cases infected by more than one clone. Clonal variants within the same host were detected in 3 out of 115 cases (2.6%), including cases with clones which were indistinguishable by restriction fragment length polymorphism or spoligotyping. In one case, coinfecting clonal variants differed in antibiotic susceptibilities. MIRU-VNTR was applied to cases with proven polyclonal infection, and it succeeded in detecting the coinfecting strains and proved useful in confirming cases of compartmentalized infection. MIRU-VNTR is a simple, rapid, and sensitive method which could facilitate and optimize the identification and characterization of clonal complexity in M. tuberculosis infection.In recent years, clonal analysis of Mycobacterium tuberculosis infection has shown us that, in certain circumstances, tuberculosis is a much more complex situation than the schematic vision of one strain infecting one host. The complex situations found in tuberculosis include recurrences caused by exogenous reinfections (2,4,5,7,19); simultaneous coinfections by more than one M. tuberculosis strain (3,5,8,13,20); compartmentalization of the infection; with different strains infecting different tissues (3, 9, 12); and microevolution phenomena leading to the appearance of clonal variants within a host (6, 10). Unfortunately, the methodological approach to a search for clonal heterogenity or polyclonality is complex and/or laborious because it requires (i) a refined analysis of genotypic patterns to prove the existence of different clones or (ii) the analysis of multiple independent colonies from each specimen in order to detect the presence of clonal variants. These methodological exigencies have been responsible for the small number of reports examining this issue. Mycobacterial interspersed repetitive unit-variable-number tandem repeat typing (MIRU-VNTR) (17) is a novel PCR-based typing method for M. tuberculosis which is based on the amplification of 12 independent loci and the assignation of the number of tandem repeats found in each locus. The purpose of this study is to test the usefulness of this novel method for simplifying and optimizing the exploration and characterization of clonal complexity in tuberculosis. MATERIALS...