Performance evaluation of the PelvoCheck CT/NG test kit for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae.

OBJECTIVE: Assessment of the performance of the PelvoCheck CT/NG test (Greiner-Bio-One GmbH) to detect Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) in first-void urine (FVU) of females. DESIGN: A cross-sectional study to compare the PelvoCheck CT/NG with COBAS TaqMan CT Test V.2.0 (Roche) for the detection of CT and with an in-house porA-based PCR for the detection of NG in FVU specimens. In addition, pools of 5 FVU specimens containing only CT-negative or 1 CT-positive and 4 CT-negative samples were tested. Abbott RealTime CT/NG was used as an additional test to resolve discordant results. SETTING: Samples sent from six laboratories were tested at the University Medical Center Hamburg. PARTICIPANTS: Urine samples were from 1622 female patients attending gynaecological practices for chlamydia screening, another 120 urine samples were from patients pretested for NG at Synlab, Medical Service Center, Weiden GmbH. In addition, 50 urine samples spiked with various concentrations of reference material were used. RESULTS: For the detection of CT and NG, the sensitivity and specificity of the PelvoCheck CT/NG test were 98.8% and 100%, and 98.3% and 98.2%, respectively. The data obtained with the PelvoCheck CT/NG for pooled urine specimens resulted in a positive agreement of 90.9% and a negative agreement of 100%. CONCLUSIONS: The PelvoCheck CT/NG assay is a suitable test method for the detection of CT and NG in female FVU samples, with sensitivity and specificity comparable with other Food and Drug Administration approved CT/NG nucleic acid amplification tests. To the best of our knowledge, this is the first commercial test system validated for the analysis of pooled urine specimens. No false-positive or invalid result was observed in 55 analysed pools. Nevertheless, 5 samples were false negative due to a target concentration below the limit of detection of the PelvoCheck CT/NG test as a consequence of pooling-associated dilution.

Yeast Ist2 recruits the endoplasmic reticulum to the plasma membrane and creates a ribosome-free membrane microcompartment.

The endoplasmic reticulum (ER) forms contacts with the plasma membrane. These contacts are known to function in non-vesicular lipid transport and signaling. Ist2 resides in specific domains of the ER in Saccharomyces cerevisiae where it binds phosphoinositide lipids at the cytosolic face of the plasma membrane. Here, we report that Ist2 recruits domains of the yeast ER to the plasma membrane. Ist2 determines the amount of cortical ER present and the distance between the ER and the plasma membrane. Deletion of IST2 resulted in an increased distance between ER and plasma membrane and allowed access of ribosomes to the space between the two membranes. Cells that overexpress Ist2 showed an association of the nucleus with the plasma membrane. The morphology of the ER and yeast growth were sensitive to the abundance of Ist2. Moreover, Ist2-dependent effects on cytosolic pH and genetic interactions link Ist2 to the activity of the H(+) pump Pma1 in the plasma membrane during cellular adaptation to the growth phase of the culture. Consistently we found a partial colocalization of Ist2-containing cortical ER and Pma1-containing domains of the plasma membrane. Hence Ist2 may be critically positioned in domains that couple functions of the ER and the plasma membrane.

The Drosophila Crumbs signal peptide is unusually long and is a substrate for signal peptide peptidase.

N-terminal signal sequences mediate nascent protein targeting to and protein insertion into the membrane of the endoplasmic reticulum. They are typically 15-30 amino acid residues long with a core hydrophobic region flanked by an N-terminal (n-) and a C-terminal region. Following cleavage by signal peptidase, some of the resulting signal peptides are further processed by signal peptide peptidase (SPP) and fragments are liberated into the cytosol. Such fragments can have independent, post-targeting functions affecting diverse cellular processes. We show that Drosophila melanogaster Crumbs, a transmembrane protein controlling cell polarity and morphogenesis, is synthesized with an 83 residues-long signal sequence. To our knowledge, this is currently the longest signal sequence described for an eukaryotic protein. The unusual length is caused by an extended n-region, but the extension does neither affect protein targeting nor signal sequence cleavage. The signal sequence is cleaved off and the resulting signal peptide, SP(Crb), is proteolytically processed by SPP, thus representing the first substrate described for the Drosophila enzyme. We further show that signal peptide fragments can be degraded by the proteasome. Expression of transgenes encoding tagged variants of Crumbs in Drosophila embryos suggests that the signal peptide is short-lived in vivo. Our findings support a model suggesting that besides generating fragments with post-targeting functions, SPP-mediated processing is the first step in the degradation of signal peptides.