Objectives The purpose of this study was to investigate the prevalence of Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis, and Mycoplasma genitalium, in women attending a sexually transmitted disease (STD) clinic, as well as the frequency of coinfections, and relationship of each organism to cervicitis. Methods In this cross-sectional study of 324 women attending Baltimore City STD Clinics, C. trachomatis, N. gonorrhoeae, T. vaginalis, and M. genitalium were detected using nucleic acid amplification tests. Demographic characteristics and risk factors were ascertained. Results Overall prevalence of infection with C. trachomatis, N. gonorrhoeae, T. vaginalis, and M. genitalium was found to be 11.1%, 4.6%, 15.3%, and 19.2%, respectively. Prevalence in women with cervicitis was 15.8%, 6%, 18.9%, and 28.6% for C. trachomatis, N. gonorrhoeae, T. vaginalis, and M. genitalium, respectively. Percentages of coinfections were high. C. trachomatis and M. genitalium were significantly associated with cervicitis in univariate analysis, but only M. genitalium was significantly associated with cervicitis (AOR: 2.5) in multiple logistic regression models. Conclusion Knowledge of the statistical association of M. genitalium with cervicitis in this study increases the need for further confirmation of the etiologic significance of this organism with cervicitis in more diverse populations. The high prevalence merits more study and may have implications for diagnosis and treatment of cervicitis.

A rapid assay for eubacterial species identification is described using high-resolution melt analysis to characterize PCR products. Unique melt profiles generated from multiple hypervariable regions of the 16S rRNA gene for 100 clinically relevant bacterial pathogens, including category A and B biothreat agents and their surrogates, allowed highly specific species identification.Rapid and accurate diagnostic tools are critical for infectious disease surveillance and early diagnosis of disease (8,12). A simple platform which could deliver broad-based screening and specific pathogen identification would be invaluable for the timely recognition of emerging and biothreat (BT) outbreaks, as well as commonly encountered clinical infections (2,7,9,11,12).We previously reported a probe-based PCR assay, which utilizes conserved and variable 16S rRNA gene sequences for initial broad-based eubacterial detection and subsequent identification of specific bacterial agents (11). The assay demonstrated high analytical sensitivity but was limited by an inability to differentiate closely related pathogens due to decreased specificity of the TaqMan probe chemistry and high sequence homology within selected hypervariable regions of the 16S rRNA gene. Probe-based amplicon characterization accordingly limits testing to a finite number of anticipated pathogens. Alternative strategies for amplicon analysis, such as sequencing and mass spectrometry, allow broader-scale product characterization but are costly, time-consuming, and lacking in throughput (1, 6). High-resolution melt analysis (HRMA) offers a simple, low-cost, closed-tube approach to amplicon analysis with the capacity for single-nucleotide discrimination and easy integration with PCR analysis (10). We report a unique strategy for the rapid, highly specific identification of BTrelated and non-BT-related bacterial pathogens which couples eubacterial PCR with HRMA.

Objectives-This purpose of this study was to investigate prevalence of M. genitalium C. trachomatis, N. gonorrhoeae, and T. vaginalis in men, frequency of coinfections, and relationships among organisms with urethritis in men.Methods-This was a cross-sectional study of 290 men (age range 19-34 yr) attending Baltimore City STD Clinics. M. genitalium, C. trachomatis, N. gonorrhoeae, and T. vaginalis, during 2004 were detected using nucleic acid amplification tests (NAATs). (N = 153 with urethritis and 137 without urethritis). Demographic characteristics and risk factors were ascertained.Results-The overall prevalences of infection with C. trachomatis, N. gonorrhoeae, T. vaginalis, and M. genitalium, were 20.3%, 12.8%, 3.4%, and 15.2% respectively. Prevalences in men with urethritis were 32.7%, 24.2%, 5.2%, and 22.2% for C. trachomatis, N. gonorrhoeae, T. vaginalis, and M. genitalium, respectively. Percentages of coinfections were high. All men with N. gonorrhoeae had urethritis. C. trachomatis and M. genitalium were found to be significantly associated with urethritis in univariate analysis and in multiple logistic regression analysis.Conclusion-The association of M. genitalium with urethritis in this study provides confirmation of the importance of screening men for M. genitalium as a cause of non-gonococcal urethritis and supports treatment considerations for urethritis for agents other than gonococci and chlamydia. Short Summary-Men attending STD clinics were found to have high prevalences of M. genitalium (MG), C. trachomatis (CT), and N. gonorrhoeae (NG); moderate prevalence of T. vaginalis. MG was associated with urethritis in addition to NG and CT.

Septic arthritis (SA) is a rheumatologic emergency associated with significant morbidity and mortality. Delayed or inadequate treatment of SA can lead to irreversible joint destruction and disability. Current methods of diagnosing SA rely on synovial fluid analysis and culture which are known to be imprecise and time-consuming. We report a novel adaptation of a probe-based real-time PCR assay targeting the 16S rRNA gene for early and accurate diagnosis of bacterial SA. The assay algorithm consists of initial broad-range eubacterial detection, followed by Gram typing and species characterization of the pathogen. The platform demonstrated a high analytical sensitivity with a limit of detection of 10 1 CFU/ml with a panel of SA-related organisms. Gram typing and pathogen-specific probes correctly identified their respective targets in a mock test panel of 36 common clinically relevant pathogens. One hundred twenty-one clinical synovial fluid samples from patients presenting with suspected acute SA were tested. The sensitivity and specificity of the assay were 95% and 97%, respectively, versus synovial fluid culture results. Gram-typing probes correctly identified 100% of eubacterial positive samples as to gram-positive or gram-negative status, and pathogen-specific probes correctly identified the etiologic agent in 16/20 eubacterial positive samples. The total assay time from sample collection to result is 3 h. We have demonstrated that a real-time broad-based PCR assay has high analytical and clinical performance with an improved time to detection versus culture for SA. This assay may be a useful diagnostic adjunct for clinicians, particularly those practicing in the acute care setting where rapid pathogen detection and identification would assist in disposition and treatment decisions.Septic arthritis (SA) is a rheumatologic emergency associated with significant morbidity and mortality (6, 9). Delayed or inadequate treatment of SA can lead to irreversible joint destruction with subsequent disability. Accordingly, prompt diagnosis and early initiation of therapy are critical in improving the outcome (7).The diagnosis of SA in the acute care setting is challenging because of the relatively poor sensitivity and specificity of clinical examination findings and lack of a rapid reliable diagnostic assay. Further, overreliance on conventional laboratory tests for synovial fluid analysis is hindered by the relatively poor performance characteristics of these methods (11,12,16). In particular, the sensitivity of Gram staining has been reported in the range of 29% to 50% (3, 4), and the sensitivity of culture may be only 82% (9). Lack of a rapid and accurate diagnostic tool results in acute care clinicians often choosing the conservative approach of hospital admission and empirical broadspectrum antibiotics for patients with suspected SA. The benefits of this management strategy may be offset, however, by added costs and potential iatrogenic complications associated with unnecessary treatment and hospitalizations, as well ...