Rapid Detection of Chlamydia trachomatis and Typing of the Lymphogranuloma venereum associated L-Serovars by TaqMan PCR

<p>Abstract</p> <p>Background</p> <p>Infection due to <it>Chlamydia trachomatis </it>is the most common sexually transmitted bacterial disease of global health significance, and especially the L-serovars causing lymphogranuloma venereum are increasingly being found in Europe in men who have sex with men.</p> <p>Results</p> <p>The design and evaluation of a rapid, multiplex, real-time PCR targeting the major outer membrane protein (<it>omp-1</it>) -gene and a L-serovar-specific region of the polymorphic protein H (<it>pmp-H</it>) -gene for the detection of <it>Chlamydia trachomatis </it>is reported here. The PCR takes place as a single reaction with an internal control. For L1-, L2- and L3-serovar differentiation a second set of real-time PCRs was evaluated based on the amplification of serovar-specific <it>omp-1</it>-regions. The detection limit of each real-time PCR, multiplexed or not, was 50 genome copies per reaction with an efficiency ranging from 90,5–95,2%.</p> <p>In a retrospective analysis of 50 ocular, rectal and urogenital specimens formerly tested to be positive for <it>C. trachomatis </it>we identified six L2-serovars in rectal specimens of HIV-positive men, one in a double-infection with L3, and one L2 in a urethral specimen of an HIV-negative male.</p> <p>Conclusion</p> <p>This unique real-time PCR is specific and convenient for the rapid routine-diagnostic detection of lymphogranuloma venereum-associated L-serovars and enables the subsequent differentiation of L1, L2 and L3 for epidemiologic studies.</p

Diagnostic challenges within the Bacillus cereus-group: finding the beast without teeth

The Bacillus cereus-group (B. cereus sensu lato) includes common, usually avirulent species, often considered contaminants of patient samples in routine microbiological diagnostics, as well as the highly virulent B. anthracis. Here we describe 16 isolates from 15 patients, identified as B. cereus-group using a MALDI-TOF MS standard database. Whole genome sequencing (WGS) analysis identified five of the isolates as B. anthracis species not carrying the typical virulence plasmids pXO1 and pXO2, four isolates as B. paranthracis, three as B. cereus sensu stricto, two as B. thuringiensis, one as B. mobilis, and one isolate represents a previously undefined species of Bacillus (B. basilensis sp. nov.). More detailed analysis using alternative MALDI-TOF MS databases, biochemical phenotyping, and diagnostic PCRs, gave further conflicting species results. These cases highlight the difficulties in identifying avirulent B. anthracis within the B. cereus-group using standard methods. WGS and alternative MALDI-TOF MS databases offer more accurate species identification, but so far are not routinely applied. We discuss the diagnostic resolution and discrepancies of various identification methods

Cutibacterium modestum and 'Propionibacterium humerusii' represent the same species that is commonly misidentified as Cutibacterium acnes

Cutibacterium spp. play an increasing role in soft tissue and implant-associated infections. We isolated a novel Cutibacterium spp. from an implant and investigated this isolate using multiple identification approaches. Correct identification was hampered by inconsistent reference data. The isolate was characterised using conventional methods such as Gram stain, MALDI-TOF MS, and antimicrobial susceptibility testing against multiple antimicrobials. Partial 16S rRNA gene sequencing and whole genome sequencing were also performed. In addition, we summarised the available published sequence data and compared prior data to our strain. Conventional phenotypic identification of our isolate resulted in Cutibacterium spp. After analysis of 16S rRNA gene and genome sequences, our isolate was identified as C. modestum, a very recently described species. The 16S rRNA gene analysis was hampered by three incorrect nucleotides within the 16S rRNA gene reference sequence of C. modestum M12(T) (accession no. LC466959). We also clearly demonstrate that this novel species is identical to tentatively named "Propionibacterium humerusii". Retrospective data analysis indicates that C. modestum is a clinically important Cutibacterium species often misidentified as C. acnes. The isolation and identification of Cutibacterium spp. is still a challenge. The correct description of very recently named C. modestum and the availability of a correct 16S rRNA sequence of the type strain may help to clarify the taxonomical uncertainty concerning "P. humerusii". The novel C. modestum is an additional, clinically important species within the genus Cutibacterium and may represent a new member of the human skin microbiome