Trichomonas vaginalis Detection in Urogenital Specimens from Symptomatic and Asymptomatic Men and Women by Use of the cobas TV/MG Test

Trichomonas vaginalis is a prevalent sexually transmitted infection (STI). Diagnosis has historically relied on either microscopic analysis or culture, the latter being the previous gold standard. However, these tests are not readily available for male diagnosis, generally only perform well for symptomatic women, and are not as sensitive as nucleic acid amplification tests (NAATs). Men are largely asymptomatic but carry the organism and transmit to their sexual partners. This multicenter, prospective study evaluated the performance of the cobas T. vaginalis/Mycoplasma genitalium (TV/MG) assay for detection of T. vaginalis DNA compared with patient infection status (PIS) defined by a combination of commercially available NAATs and culture using urogenital specimens. A total of 2,064 subjects (984 men and 1,080 women, 940 [45.5%] symptomatic, 1,124 [54.5%] asymptomatic) were evaluable. In women, sensitivity ranged from 99.4% (95% confidence interval [CI] 96.8 to 99.9%) using vaginal samples to 94.7% (95% CI 90.2 to 97.2%) in PreservCyt samples. Specificity ranged from 98.9 to 96.8% (95% CI 95.4 to 97.8%). In men, the cobas TV/MG assay was 100% sensitive for the detection of T. vaginalis in both male urine samples and meatal swabs, with specificity of 98.4% in urine samples and 92.5% in meatal swabs. The cobas TV/MG is a suitable diagnostic test for the detection of T. vaginalis, which could support public health efforts toward infection control and complement existing STI programs

Vibrio cholerae H-NS Silences Virulence Gene Expression at Multiple Steps in the ToxR Regulatory Cascade

H-NS is an abundant nucleoid-associated protein involved in the maintenance of chromosomal architecture in bacteria. H-NS also has a role in silencing the expression of a variety of environmentally regulated genes during growth under nonpermissive conditions. In this study we demonstrate a role for H-NS in the negative modulation of expression of several genes within the ToxR virulence regulon of Vibrio cholerae. Deletion of hns resulted in high, nearly constitutive levels of expression of the genes encoding cholera toxin, toxin-coregulated pilus, and the ToxT virulence gene regulatory protein. For the cholera toxin- and ToxT-encoding genes, elevated expression in an hns mutant was found to occur in the absence of the cognate activator proteins, suggesting that H-NS functions directly at these promoters to decrease gene expression. Deletion analysis of the region upstream of toxT suggests that an extensive region located far upstream of the transcriptional start site is required for complete H-NS-mediated repression of gene expression. These data indicate that H-NS negatively influences multiple levels of gene expression within the V. cholerae virulence cascade and raise the possibility that the transcriptional activator proteins in the ToxR regulon function to counteract the repressive effects of H-NS at the various promoters as well as to recruit RNA polymerase

H-NS Binding and Repression of the ctx Promoter in Vibrio cholerae▿

Expression of the ctx and tcp genes, which encode cholera toxin and the toxin coregulated pilus, the Vibrio cholerae O1 virulence determinants having the largest contribution to cholera disease, is repressed by the nucleoid-associated protein H-NS and activated by the AraC-like transcriptional regulator ToxT. To elucidate the molecular mechanism by which H-NS controls transcription of the ctxAB operon, H-NS repression and binding were characterized by using a promoter truncation series, gel mobility shift assays, and DNase I footprinting. Promoter regions found to be important for H-NS repression correlated with in vitro binding. Four main H-NS binding regions are present at ctx. One region overlaps the high-affinity ToxT binding site and extends upstream, another overlaps the ToxT low-affinity binding site around the −35 element, and the remaining two are located adjacent to one another downstream of the transcriptional start site. Competition for binding to the overlapping H-NS/ToxT binding sites was observed in gel mobility shift assays, where ToxT was found to displace H-NS from the ctx promoter region. In addition, regulatory differences between the ctx and tcpA promoters were examined. H-NS was found to have a higher relative binding affinity for the ctx promoter than for the tcpA promoter in vitro. In contrast to ToxT-dependent activation of the tcpA promoter, ToxT activation of ctx did not require the C-terminal domain of the α-subunit of RNA polymerase. These findings demonstrate that transcriptional regulation of ctx and tcpA by H-NS and ToxT is mechanistically distinct, and this may lead to important differences in the expression of these coregulated genes

Evaluation of the Performance of the Cobas CT/NG Test for Use on the Cobas 6800/8800 Systems for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae in Male and Female Urogenital Samples

The clinical performance of the Cobas CT/NG assay on the Cobas 6800/8800 systems (Cobas) for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae was established in a multisite, prospective collection study using male and female urogenital specimens; supportive data from archived specimens were also included. The results obtained with the Cobas assay were compared with the patient infected status derived from a combination of U.S. Food and Drug Administration-approved nucleic acid amplification tests to determine the sensitivity and specificity of detection from each sample type. The sensitivity of Cobas for the detection of C. trachomatis in female specimens was 95.6% (95% confidence interval [CI], 92.4% to 97.4%) for urine; 98.6% (95% CI, 95.2% to 99.6%) and 99.2% (95% CI, 95.4% to 99.9%) for clinician- and self-collected vaginal swab specimens, respectively; 93.3% (95% CI, 89.6% to 95.7%) for endocervical swabs; and 92.5% (95% CI, 88.7% to 95.1%) for cervical swab samples in PreservCyt. The specificity for the detection of C. trachomatis was ≥98.8% for all female sample types. Sensitivity and specificity estimates of Cobas for the detection of C. trachomatis in male urine samples were 100% (96.8% to 100.0%) and 99.7% (95% CI, 99.2% to 99.9%), respectively. The sensitivity of Cobas for the detection of N. gonorrhoeae in female specimens was 94.8% (95% CI, 89.6% to 97.4%) for urine; 100.0% (95% CI, 87.9% to 100.0%) and 100.0% (95% CI, 87.9% to 100.0%) for clinician- and self-collected vaginal swab specimens, respectively; 97.0% (95% CI, 91.5% to 99.0%) for endocervical swabs; and 96.6% (95% CI, 90.6% to 98.8%) for cervical samples in PreservCyt; the specificity for all female sample types was >99.0%. The sensitivity and specificity of Cobas for detecting N. gonorrhoeae in male urine were 100.0% (95% CI, 95.8% to 100.0%) and 99.5% (95% CI, 98.8% to 99.8%), respectively. Fully automated assays help fill the clinical need for a sensitive, high-throughput screening tool to aid public health efforts to control C. trachomatis and N. gonorrhoeae infections

Detection of Chlamydia trachomatis and Neisseria gonorrhoeae with the cobas CT/NG v2.0 test: performance compared with the BD ProbeTec CT Qx and GC Qx amplified DNA and Aptima AC2 assays

ObjectivesInfections due to Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are among the most common bacterial sexually transmitted infections worldwide, most of which are asymptomatic. Detection of infection using a variety of specimen types in symptomatic and asymptomatic subjects is important to effectively combat CT/NG infections. The performance of the cobas CT/NG v2.0 test was assessed for urogenital swabs, urine and cervical cytology samples collected in PreservCyt Solution from 5266 symptomatic and asymptomatic women (including 202 who were pregnant), and urine from 738 men.MethodsSensitivity and specificity were estimated compared with a patient infected status determined using two US Food and Drug Administration-cleared nucleic acid amplification tests.ResultsAmong 6004 participants, 487 CT (8.1%) and 159 NG (2.6%) infections were identified. Sensitivity estimates for CT for women ranged from 91.2% to 97.6% depending on specimen type, and the estimate for male urine specimens was 98.4%. Specificity for CT ranged from 99.2% to 99.7%. Sensitivity estimates for NG ranged from 95.6% to 100.0% for women, and the estimate for men was 100.0%. Specificity for NG ranged from 99.3% to 100.0%.ConclusionsThe cobas CT/NG v2.0 test performs well using urogenital swabs, urine and cervical samples collected in PreservCyt solution

Mycoplasma genitalium Detection in Urogenital Specimens from Symptomatic and Asymptomatic Men and Women by Use of the cobas TV/MG Test

Mycoplasma genitalium (MG) infections are a growing concern within the field of sexually transmitted infections. However, diagnostic assays for M. genitalium have been limited in the United States. As most infections are asymptomatic, individuals can unknowingly pass the infection on, and the prevalence is likely to be underestimated. Diagnosis of M. genitalium infection is recommended using a nucleic acid test. This multicenter study assessed the performance of the cobas Trichomonas vaginalis (TV)/MG assay (cobas) for the detection of M. genitalium, using 22,150 urogenital specimens from both symptomatic and asymptomatic men and women collected at geographically diverse sites across the United States. The performance was compared to a reference standard of three laboratory-developed tests (LDTs). The specificity of the cobas assay for M. genitalium ranged from 96.0% to 99.8% across symptomatic and asymptomatic men and women. The sensitivities in female vaginal swabs and urine samples were 96.6% (95% confidence interval [CI], 88.5 to 99.1%) and 86.4% (95% CI, 75.5 to 93.0%), respectively. The sensitivities in male urine and meatal swab samples were 100% (95% CI, 94.0 to 100%) and 85.0% (95% CI, 73.9 to 91.9%), respectively. This study demonstrated that the cobas assay was highly sensitive and specific in all relevant clinical samples for the detection of M. genitalium