The protease inhibitor JO146 demonstrates a critical role for CtHtrA for Chlamydia trachomatis reversion from penicillin persistence

The Chlamydia trachomatis serine protease HtrA (CtHtrA) has recently been demonstrated to be essential during the replicative phase of the chlamydial developmental cycle. A chemical inhibition strategy (serine protease inhibitor JO146) was used to demonstrate this essential role and it was found that the chlamydial inclusions diminish in size and are lost from the cell after CtHtrA inhibition without formation of viable elementary bodies. The inhibitor (JO146) was used in this study to investigate the role of CtHtrA for penicillin persistence and heat stress conditions for Chlamydia trachomatis. JO146 addition during penicillin persistence resulted in only minor reductions (~1 log) in the final viable infectious yield after persistent Chlamydia were reverted from persistence. However, JO146 treatment during the reversion and recovery from penicillin persistence was completely lethal for Chlamydia trachomatis. JO146 was completely lethal when added either during heat stress conditions, or during the recovery from heat stress conditions. These data together indicate that CtHtrA has essential roles during some stress environments (heat shock), recovery from stress environments (heat shock and penicillin persistence), as well as the previously characterized essential role during the replicative phase of the chlamydial developmental cycle. Thus, CtHtrA is an essential protease with both replicative phase and stress condition functions for Chlamydia trachomatis. © 2013 Ong, Marsh, Lawrence, Allan, Timms and Huston

In vitro susceptibility of recent Chlamydia trachomatis clinical isolates to the CtHtrA inhibitor JO146

© 2015 Institut Pasteur. The present study aimed to establish if a previously identified Chlamydia trachomatis HtrA (CtHtrA) inhibitor, JO146, is effective against currently circulating clinical isolates to validate if CtHtrA is a clinically relevant target for future therapeutic development. Inhibition of CtHtrA during the middle of the chlamydial replicative cycle until the completion of the cycle resulted in loss of infectious progeny for six unique clinical isolates representing different serovars. This supports the potential for CtHtrA to be a clinically relevant target for development of new therapeutics and suggests the importance of further investigation of JO146 as a lead compound

Cervical Vascular and Upper Airway Asymmetry in Velo-Cardio-Facial Syndrome: Correlation of Nasopharyngoscopy With MRA

Purpose Velo-cardio-facial syndrome (VCFS), the most common genetic syndrome causing cleft palate, is associated with internal carotid and vertebral artery anomalies, as well as upper airway asymmetry. Medially displaced internal carotid arteries, often immediately submucosal, present a risk of vascular injury during pharyngeal flap surgery for velopharyngeal insufficiency (VPI). We evaluate the frequency and spectrum of cervical vascular anomalies in a large cohort of VCFS patients correlating MRA with nasopharyngolaryngoscopy in detecting at risk carotid arteries. Furthermore, we assess the relationship with respect to laterality between cervical vascular patterns and the asymmetric abnormalities of these subjects’ upper airways. Methods Cervical MRAs of 86 subjects with VCFS and 50 control subjects were independently reviewed by three neuroradiologists. The course of the internal carotid and vertebral arteries was identified within the pharyngeal soft tissues. Medial deviation, level of bifurcation, dominance, anomalous origin, and vessel tortuosity were recorded. Nasopharyngoscopy examinations were available for retrospective review in 43 patients and were assessed for palatal and posterior pharyngeal wall symmetry, true vocal cord motion and size, and for the presence or absence of carotid pulsations. The endoscopic findings were compared with MRA results. Results Of the 86 subjects, 80 (93%) had one or more vascular anomalies. 42 subjects (49%) were found to have medial deviation of at least one internal carotid artery. In 24 subjects (28%) the anomalous internal carotid artery was directly submucosal; four of these were bilateral (5% of the total sample, 17% of those with a submucosal internal carotid). Other carotid anomalies included low carotid bifurcation (44 subjects or 51%), anomalous origin of the right common carotid (32 cases, or 37%), and two cases of internal carotid agenesis/hypoplasia. Vertebral artery anomalies included vessel tortuosity (34 cases, or 40%), hypoplasia (10 cases, or 12%), looping (4 cases, or 5%), and one case of a double left vertebral artery. Though patients in our study showed an asymmetric distribution of vascular anomalies, no association was found between the laterality of palatal motion, pharyngeal fullness, or laryngeal movement and structure with ipsilateral vertebral or carotid artery anomalies. Of the 33 pulsatile carotid arteries visualized at nasopharyngoscopy, only nine were found to be submucosal on MRA. In contrast, 11 submucosal carotid arteries confirmed at MRA demonstrated no visible pulsations. Positive and negative predictive values of pulsative arteries seen endoscopically for MRA confirmation of a submucosal carotid course was 27% and 79% respectively. Conclusions Carotid and vertebral artery anomalies are common in VCFS including marked medial deviation of the internal carotid artery in close proximity to the donor site for pharyngeal flap surgery. Lack of correlation between laterality of vascular anomalies and upper airway structural asymmetry in VCFS does not support the hypothesis that palatal, pharyngeal, and laryngeal anomalies are due to secondary developmental sequences caused by in utero vascular insufficiency. The presence or absence of carotid pulsations seen by nasopharyngoscopy does not correlate with the carotid arterial depth identified on MRA. Furthermore, identification of the relative medial–lateral retropharyngeal position of a submucosal carotid affords the opportunity to modify the surgical approach. These findings further support the routine use of pre-operative neck MRA in VCFS patients in surgical planning

A retrospective pilot study to determine whether the reproductive tract microbiota differs between women with a history of infertility and fertile women

© 2017 The Royal Australian and New Zealand College of Obstetricians and Gynaecologists Background: We know very little about the microbiota inhabiting the upper female reproductive tract and how it impacts on fertility. Aims: This pilot study aimed to examine the vaginal, cervical and endometrial microbiota for women with a history of infertility compared to women with a history of fertility. Materials and methods: Using a retrospective case–control study design, women were recruited for collection of vaginal, cervical and endometrial samples. The microbiota composition was analysed by 16S ribosomal RNA (rRNA) gene amplification and endometrial expression of selected human genes by quantitative reverse transcription polymerase chain reaction. Results: Sixty-five specimens from the reproductive tract of 31 women were successfully analysed using 16S rRNA gene amplicon sequencing (16 controls and 15 cases). The dominant microbial community members were consistent in the vagina and cervix, and generally consistent with the endometrium although the relative proportions varied. We detected three major microbiota clusters that did not group by tissue location or case–control status. There was a trend that infertile women more often had Ureaplasma in the vagina and Gardnerella in the cervix. Testing for the expression of selected genes in the endometrium did not show evidence of correlation with case–control status, or with microbial community composition, although Tenascin-C expression correlated with a history of miscarriage. Conclusions: There is a need for further exploration of the endometrial microbiota, and how the microbiota members or profile interplays with fertility or assisted reproductive technologies

Growth of nano-domains in Gd-CeO₂ mixtures:Hybrid Monte Carlo simulations

Hybrid Monte Carlo (HMC) simulations are used to study the growth of Gd-rich domains in Gd doped CeO2, and we probe the conductivity of the resulting and other configurations by molecular dynamics.</p

Assay strategies for the discovery and validation of therapeutics targeting <i>Brugia pahangi</i> Hsp90

The chemotherapy of lymphatic filariasis relies upon drugs such as diethylcarbamazine and ivermectin that largely target the microfilarial stages of the parasite, necessitating continued treatment over the long reproductive life span of the adult worm. The identification of compounds that target adult worms has been a long-term goal of WHO. Here we describe a fluorescence polarization assay for the identification of compounds that target Hsp90 in adult filarial worms. The assay was originally developed to identify inhibitors of Hsp90 in tumor cells, and relies upon the ability of small molecules to inhibit the binding of fluorescently labelled geldanamycin to Hsp90. We demonstrate that the assay works well with soluble extracts of Brugia, while extracts of the free-living nematode C. elegans fail to bind the probe, in agreement with data from other experiments. The assay was validated using known inhibitors of Hsp90 that compete with geldanamycin for binding to Hsp90, including members of the synthetic purine-scaffold series of compounds. The efficacy of some of these compounds against adult worms was confirmed in vitro. Moreover, the assay is sufficiently sensitive to differentiate between binding of purine-scaffold compounds to human and Brugia Hsp90. The assay is suitable for high-throughput screening and provides the first example of a format with the potential to identify novel inhibitors of Hsp90 in filarial worms and in other parasitic species where Hsp90 may be a target

IL-4-secreting CD4+ T cells are crucial to the development of CD8+ T-cell responses against malaria liver stages.

CD4+ T cells are crucial to the development of CD8+ T cell responses against hepatocytes infected with malaria parasites. In the absence of CD4+ T cells, CD8+ T cells initiate a seemingly normal differentiation and proliferation during the first few days after immunization. However, this response fails to develop further and is reduced by more than 90%, compared to that observed in the presence of CD4+ T cells. We report here that interleukin-4 (IL-4) secreted by CD4+ T cells is essential to the full development of this CD8+ T cell response. This is the first demonstration that IL-4 is a mediator of CD4/CD8 cross-talk leading to the development of immunity against an infectious pathogen