Characterization of the outer membrane proteome of Francisella noatunensis subsp. orientalis.

AIMS: The aims of the current study were to characterize the outer membrane proteins (OMPs) of Francisella noatunensis subsp. orientalis (Fno) STIR-GUS-F2f7, and identify proteins recognized by sera from tilapia, Oreochromis niloticus, (L) that survived experimental challenge with Fno. METHODS AND RESULTS: The composition of the OMPs of a virulent strain of Fno (STIR-GUS-F2f7), isolated from diseased red Nile tilapia in the United Kingdom, was examined. The sarcosine-insoluble OMPs fraction was screened with tilapia hyperimmune sera by western blot analysis following separation of the proteins by 1D SDS-PAGE. Liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was used to identify the various proteins present in the OMP profile. Two hundred and thirty-nine proteins were identified, of which 44 were found in the immunogenic band recognized by the tilapia hyperimmune serum. In silico analysis was performed to predict the function and location of the OMPs identified by MS. CONCLUSIONS: Using a powerful proteomic-based approach in conjugation with western immunoblotting, proteins comprising the outer membrane fraction of Fno STIR-GUS-F2f7 were identified, catalogued and screened for immune recognition by tilapia sera. SIGNIFICANCE AND IMPACT OF THE STUDY: The current study is the first report on the characterization of Fno-OMPs. The findings here provide preliminary data on bacterial surface proteins that exist in direct contact with the host's immune defences during infection and offer an insight into the pathogenesis of Fno.

Role of breast density measurement in screening for breast cancer.

Mammographic breast density is an established risk factor for breast cancer and significantly reduces the sensitivity of mammography, yet its use within breast cancer screening programs in Australia is limited. We provide a contemporary snapshot of the role of breast density measurement in screening for breast cancer and review the recent evidence for an increasing role of breast density measurement as: a predictor of breast cancer risk, a predictor of 'masking', and a biomarker to monitor effectiveness of intervention strategies for primary prevention or improved breast cancer outcomes.

The Influence of Clinical and Patient-Reported Outcomes on Post-surgery Satisfaction in Cholecystectomy Patients.

BACKGROUND: Patient-reported outcomes (PROMs) and post-operative satisfaction have become a growing focus of surgical outcome evaluation and are considered key components of the movement towards patient-centred care. The aim was to compare the association of traditional clinical outcome measures and PROMs with post-surgery satisfaction in cholecystectomy patients. METHODS: Patients who had undergone elective or emergency cholecystectomy for gallstone disease were sent validated PROM questionnaires, and telephone follow-up was performed in all cases. Categorical data were compared with the Chi-square and Fisher's exact tests. Satisfaction was investigated using a "top-box" approach, and multivariable logistic regression was performed for factors significantly (p < 0.05) associated with satisfaction in univariable analyses. RESULTS: A total of 234 patients underwent cholecystectomy between 1 March 2014 and 1 May 2014, and 147 patients (63%) completed the questionnaire. 104/147 (71%) reported being "very satisfied" with their surgical outcome. In univariable analyses, satisfaction showed significant association with an absence of hospital-recorded 30-day complications (OR = 4.11, 95% CI 1.29-13.84), but not re-attendance, readmission, or length of stay. In a multiple regression analysis, no traditional clinical outcome measures were associated with satisfaction. By contrast, self-perceived health (OR = 4.04, 95% CI 1.44-11.86), the absence of patient-reported wound pain (OR = 6.11, 95% CI 1.83-21.74), and a return to normal leisure activities (OR = 11.14, 95% CI 2.61-55.26) were associated with satisfaction. CONCLUSION: PROMs are the major determinants of patient satisfaction following cholecystectomy. When assessing outcomes following cholecystectomy, the measurement of clinical outcomes alone is inadequate and should be supplemented by the use of PROMs.

Patient experience and overall satisfaction after emergency abdominal surgery.

BACKGROUND: There is a growing recognition of the importance of patient experience in healthcare, however little is known in the context of emergency abdominal surgery. This study sought to quantify the association between patient experience and overall satisfaction. METHODS: Patient demographics, operation details and 30-day clinical outcome data of consecutive patients undergoing emergency abdominal surgery were collected. Data was collected using validated Patient Reported Experience Measures (PREMs) questionnaires. Categorical data were tested using Mann Whitney U test. Multivariable regression was used to determine independent factors associated with satisfaction. RESULTS: In a well-fitting multivariable analysis (R2 = 0.71), variables significantly associated with a higher global satisfaction score were "sufficient information given about treatment" (ß = 0.86, 95% CI 0.01-1.70, p = 0.047), "sufficient explanation of risks and benefits of surgery" (ß = 1.26, 95% CI 0.18-2.34, p = 0.020), "absence of night-time noise" (ß = 1.35, 95% CI 0.56-2.14, p = 0.001) and "confidence and trust in nurses" (ß = 1.51, 95% CI 0.54-2.49, p = 0.003). CONCLUSIONS: Overall patient satisfaction was strongly associated with perceptions of good communication and transfer of information. Confidence and trust in the clinical team is an important determinant of patient experience. Improving the ward environment by reducing noise at night may also improve the overall experience and satisfaction in emergency surgery.

Role of the placenta in developmental programming: Observations from models using large animals.

Developmental programming, which proposes that "insults" or "stressors" during intrauterine or postnatal development can have not only immediate but also long-term consequences for healthy and productivity, has emerged as a major biological principle, and based on studies in many animal species also seems to be a universal phenomenon. In eutherians, the placenta appears to be programmed during its development, which has consequences for fetal growth and development throughout pregnancy, and likewise has long-term consequences for postnatal development, leading to programming of organ function of the offspring even into adulthood. This review summarizes our current understanding of the placenta's role in developmental programming, the mechanisms involved, and the challenges remaining.

Identification of N-terminal regions of wheat leaf ferredoxin NADP+ oxidoreductase important for interactions with ferredoxin.

Wheat leaves contain two isoproteins of the photosynthetic ferredoxin:NADP(+) reductase (pFNRI and pFNRII). Truncated forms of both enzymes have been detected in vivo, but only pFNRII displays N-terminal length-dependent changes in activity. To investigate the impact of N-terminal truncation on interaction with ferredoxin (Fd), recombinant pFNRII proteins, differing by deletions of up to 25 amino acids, were generated. During purification of the isoproteins found in vivo, the longer forms of pFNRII bound more strongly to a Fd affinity column than did the shorter forms, pFNRII(ISKK) and pFNRII[N-2](KKQD). Further truncation of the N-termini resulted in a pFNRII protein which failed to bind to a Fd column. Similar k(cat) values (104-140 s(-1)) for cytochrome c reduction were measured for all but the most truncated pFNRII[N-5](DEGV), which had a k(cat) of 38 s(-1). Stopped-flow kinetic studies, examining the impact of truncation on electron flow between mutant pFNRII proteins and Fd, showed there was a variation in k(obs) from 76 to 265 s(-1) dependent on the pFNRII partner. To analyze the sites which contribute to Fd binding at the pFNRII N-terminal, three mutants were generated, in which a single or double lysine residue was changed to glutamine within the in vivo N-terminal truncation region. The mutations affected binding of pFNRII to the Fd column. Based on activity measurements, the double lysine residue change resulted in a pFNRII enzyme with decreased Fd affinity. The results highlight the importance of this flexible N-terminal region of the pFNRII protein in binding the Fd partner.

Topical aminolaevulinic acid-photodynamic therapy produces an inflammatory infiltrate but reduces Langerhans cells in healthy human skin in vivo.

BACKGROUND: Topical photodynamic therapy (PDT) elicits a therapeutic response in both skin cancer and immune-mediated skin disorders. While PDT induces direct cell death, host inflammatory and immune responses to PDT may contribute to the therapeutic effects. OBJECTIVES: To examine the impact of topical PDT on leucocyte trafficking and mediators of chemotaxis in healthy human skin. METHODS: Aminolaevulinic acid (ALA)-PDT was performed on the buttock skin of seven healthy volunteers. Biopsies for immunohistochemical assessment were taken 1, 4 and 24 h post-PDT and from untreated contralateral buttock skin (baseline). RESULTS: A significant dermal neutrophilic infiltrate appeared early, peaking at 4 h (P < 0·01) and returning to near baseline by 24 h. Expression of E-selectin was significantly higher at 4 h (P < 0·05) and correlated strongly with neutrophil numbers (r = 0·93). Expression of intercellular adhesion molecule 1 was significantly elevated after 24 h (P < 0·05) with an apparent gradual increase in CD4+ T cells up to this time point. Notably, epidermal Langerhans cells were significantly reduced 24 h post-PDT compared with baseline (P < 0·01) and comprised a significantly larger proportion of cells with migratory rather than dendritic morphology (P < 0·05). The number of epidermal cells expressing tumour necrosis factor-α significantly increased at 4 h (P < 0·05) and remained elevated 24 h post-PDT, whereas no significant change in expression of interleukin (IL)-1ß or IL-8 was seen. CONCLUSIONS: Reduction of Langerhans cells by topical PDT of human skin may play a significant role in PDT-induced local immunosuppression, potentially benefiting the treatment of immune-mediated skin disorders but negatively impacting on antitumour responses. Further exploration according to disease indication/treatment protocol is warranted.

A link between prompt optical and prompt gamma-ray emission in gamma-ray bursts.

The prompt optical emission that arrives with the gamma-rays from a cosmic gamma-ray burst (GRB) is a signature of the engine powering the burst, the properties of the ultra-relativistic ejecta of the explosion, and the ejecta's interactions with the surroundings. Until now, only GRB 990123 had been detected at optical wavelengths during the burst phase. Its prompt optical emission was variable and uncorrelated with the prompt gamma-ray emission, suggesting that the optical emission was generated by a reverse shock arising from the ejecta's collision with surrounding material. Here we report prompt optical emission from GRB 041219a. It is variable and correlated with the prompt gamma-rays, indicating a common origin for the optical light and the gamma-rays. Within the context of the standard fireball model of GRBs, we attribute this new optical component to internal shocks driven into the burst ejecta by variations of the inner engine. The correlated optical emission is a direct probe of the jet isolated from the medium. The timing of the uncorrelated optical emission is strongly dependent on the nature of the medium.

An infrared flash contemporaneous with the gamma-rays of GRB 041219a.

The explosion that results in a cosmic gamma-ray burst (GRB) is thought to produce emission from two physical processes: the central engine gives rise to the high-energy emission of the burst through internal shocking, and the subsequent interaction of the flow with the external environment produces long-wavelength afterglows. Although observations of afterglows continue to refine our understanding of GRB progenitors and relativistic shocks, gamma-ray observations alone have not yielded a clear picture of the origin of the prompt emission nor details of the central engine. Only one concurrent visible-light transient has been found and it was associated with emission from an external shock. Here we report the discovery of infrared emission contemporaneous with a GRB, beginning 7.2 minutes after the onset of GRB 041219a (ref. 8). We acquired 21 images during the active phase of the burst, yielding early multi-colour observations. Our analysis of the initial infrared pulse suggests an origin consistent with internal shocks.

Whole-genome sequencing of clinical Clostridioides difficile isolates reveals molecular epidemiology and discrepancies with conventional laboratory diagnostic testing.

BACKGROUND: The high clinical burden of Clostridioides difficile infections merits rapid and sensitive identification of affected individuals. However, effective diagnosis remains challenging. Current best practice guidelines recommend molecular and/or direct toxin detection-based screening for symptomatic individuals, but previous work has called into question the concordance and performance of extant clinical assays. AIM: To better correlate the genomic and phenotypic properties of clinical C. difficile isolates with laboratory testing outcomes in both C. difficile-infected patients and asymptomatic carriers. METHODS: Whole-genome sequencing of clinical C. difficile isolates collected from an inpatient population at a single healthcare institution was performed, enabling examination of their molecular epidemiology and toxigenic gene content. Genomic findings were compared with clinical testing outcomes, identifying multiple diagnostic discrepancies. FINDINGS: Toxigenic culture, considered a 'reference standard', provided perfect sensitivity and specificity in predicting toxigenic gene content, whereas reduced performance was observed for Simplexa C. difficile Direct Assay (100% specificity, 88% sensitivity), Gene Xpert CD/Epi Assay (86% specificity, 83% sensitivity), and Quick Check Complete Tox A/B (100% specificity, 30% sensitivity). Genomic analysis additionally revealed variability in toxin gene sequences among C. difficile strains, phylogenomic equivalency between isolates from affected patients and carriers, and patient carriage with uncommon environmentally derived C. difficile lineages, as well as presenting opportunities for tracing pathogen transmission events. CONCLUSION: These results highlight the variable performance of clinical stool-based testing approaches as well as the potential diagnostic utility of whole-genome sequencing as an alternative to conventional testing algorithms.

Efficacy and Toxicity of Stereotactic Body Radiotherapy for Early to Advanced Stage Hepatocellular Carcinoma - Initial Experience From an Australian Liver Cancer Service.

AIMS: Intrahepatic progression remains the predominant mode of cancer-related death in hepatocellular carcinoma (HCC) underscoring the need for effective local therapies. We report our initial experience with liver stereotactic body radiotherapy (SBRT) in the management of early to advanced stage HCC at an Australian tertiary liver cancer service. MATERIALS AND METHODS: Patients with liver-confined HCC unsuitable for surgical resection or thermal ablation treated with SBRT between October 2013 and December 2018 were retrospectively evaluated. The primary end point was freedom from local progression. Secondary end points were progression-free survival, disease-specific survival, overall survival and toxicity. RESULTS: Ninety-six patients were treated for 112 lesions (median size 3.8 cm, range 1.5-17 cm). The median follow-up was 13 months (range 3-65). Forty-six patients had received prior local therapies (median 1, range 1-5), 83 (86%) patients had cirrhosis with baseline Child-Pugh scores of A (88%) and B7-8 (12%). Fifty-nine (61%) patients had Barcelona Clinic Liver Cancer (BCLC) stage 0/A disease and 37 (39%) had stage B/C. Macrovascular invasion was present in 20 (21%). The median biologically effective dose (BED10) was 86 and 60 Gy for the BCLC 0/A and B/C cohorts, respectively. Freedom from local progression at 18 months was 94% for BCLC 0/A and 74% for BCLC B/C. Progression-free survival and overall survival at 12 months were 80 and 95% for BCLC 0/A and 40 and 71% for BCLC B/C, respectively. Five patients (7%) with cirrhosis and without disease progression had an increase in Child-Pugh score >1 within 3 months of SBRT, four of whom had intercurrent infections. Clinical toxicities grade ≥2 were reported in 20% of patients. CONCLUSION: SBRT is an effective ablative modality for early stage HCC with low rates of significant toxicity. Lower dose SBRT can provide durable local control for advanced stage HCC. However, out-of-field relapse remains common, providing a rationale to investigate SBRT in combination with other therapies.

Sexual health of adults working in pornographic films.

We report the frequency of sexually transmitted infections (STIs) diagnosed in performers in the adult pornographic film industry. Over a 13 month period, 445 STI screens were performed in 115 patients, 56 women and 59 men. All reported unprotected sex during filming. Seventy-five percent (86) had at least one sexual partner outside work, and 90% used condoms inconsistently with them. Women worked exclusively with women (23%), men only (38%) or both genders (39%). Almost all men (97%) worked exclusively heterosexually. Thirty-eight percent (44/115) were diagnosed with 77 STIs, including non-specific urethritis (51), gonorrhoea (10), chlamydia (6) and genital warts (6). Gonorrhoea was found exclusively at the pharynx in three heterosexual men. There were no cases of HIV, syphilis, hepatitis B or hepatitis C. Monthly screening and certification is a working requirement for this population but STIs are common in an industry where unprotected sex is the norm.

Protocol for geolocating rural villages of women in Liberia utilizing a maternity waiting home.

OBJECTIVE: Geospatial data are used by health systems and researchers to understand disease burdens, trace outbreaks, and allocate resources, however, there are few well-documented protocols for collecting and analyzing geographic information systems data in rural areas of low- and middle-income countries. Even with the proliferation of spatial technologies such as Open Street Map and Google Maps, basic geographic data-such as village locations-are not widely available in many countries in sub-Saharan Africa. The purpose of this paper is to report a step-wise protocol, using geographic information system techniques and tools, developed to collect and analyze the type of spatial data necessary to calculate the distance between rural villages and maternity waiting homes located near rural primary healthcare facilities in Bong County, Liberia. RESULTS: Using a step-wise approach incorporating local healthcare provider knowledge, intensive field work, and spatial technologies such as Open Street Map and Google Maps for village geospatial data collection and verification, we identified village locations of 93.7% of the women who accessed the five maternity waiting homes in our study from 2012 to 2016.

Prediction of intramuscular fat content and shear force in Texel lamb loins using combinations of different X-ray computed tomography (CT) scanning techniques.

Computed tomography (CT) parameters, including spiral computed tomography scanning (SCTS) parameters, intramuscular fat (IMF) and mechanically measured shear force were derived from two previously published studies. Purebred Texel (n = 377) of both sexes, females (n = 206) and intact males (n = 171) were used to investigate the prediction of IMF and shear force in the loin. Two and three dimensional CT density information was available. Accuracies in the prediction of shear force and IMF ranged from R2 0.02 to R2 0.13 and R2 0.51 to R2 0.71 respectively, using combinations of SCTS and CT scan information. The prediction of mechanical shear force could not be achieved at an acceptable level of accuracy employing SCTS information. However, the prediction of IMF in the loin employing information from SCTS and additional information from standard CT scans was successful, providing evidence that the prediction of IMF and related meat eating quality (MEQ) traits for Texel lambs in vivo can be achieved.

Oxidation of a specific methionine in thrombomodulin by activated neutrophil products blocks cofactor activity. A potential rapid mechanism for modulation of coagulation.

Endothelial thrombomodulin (TM) plays a critical role in hemostasis as a cofactor for thrombin-dependent formation of activated protein C, a potent anticoagulant. Chloramine T, H2O2, or hypochlorous acid generated from H2O2 by myeloperoxidase rapidly destroy 75-90% of TM cofactor activity. Activated PMN, the primary in vivo source of biological oxidants, also rapidly inactivate TM. Oxidation of TM by PMN is inhibited by diphenylene iodonium, an inhibitor of NADPH oxidase. Both Met291 and Met388 in the six epidermal growth factor-like repeat domain are oxidized; however, only substitutions of Met388 lead to TM analogues that resist oxidative inactivation. We suggest that in inflamed tissues activated PMN may inactivate TM and demonstrate further evidence of the interaction between the inflammatory process and induction of thrombotic potential.

Is it time for the virtual chaperone in genitourinary medicine clinics?

The General Medical Council recommends that chaperones must be offered during intimate examinations. Implementation of these guidelines may serve to protect both patients and staff within the genitourinary (GU) medicine clinic, but is likely to have major ramifications from a practical, financial and staffing perspective. The Virtual Chaperone (VC) is an objective device making audio and visual recordings of patients' consultations and examinations. Information cannot be manipulated and is encrypted for security. It would not replace the human chaperone. This study investigated the attitudes of GU medicine staff and patients to the VC in an inner city sexual health clinic. Voluntary, anonymized questionnaires were distributed to all members of the multidisciplinary team and 200 patients. The patient response rate was 90% (n = 180). Only 40% of respondents felt the VC was acceptable in GU medicine clinics. Two-thirds felt unsure or were against the VC recording during consultations, and fewer wished the examination to be recorded. Most opted to switch off the video entirely. Almost 50% of respondents felt that the VC was designed to protect staff, whereas only 41% thought it would protect patients. The staff response rate was 69% (n = 35). Almost three-quarters of respondents were unsure or felt the device was unacceptable. Less than half would feel comfortable with the VC recording during consults. Overall, three-quarters of respondents were either unsure or did not support the introduction of the VC. There was a clear feeling (>80%) that the VC would protect staff and patients, 71% indicating that the trust would also benefit. Despite its success in other outpatient specialties, GU medicine staff and patients do not favour the introduction of the VC.

Accuracy of in vivo muscularity indices measured by computed tomography and their association with carcass quality in lambs.

Assessments of muscle mass and skeletal dimensions by Computed Tomography (CT) enable the development of new muscularity indices for the hind leg (HL) and lumbar region (LR) in lambs. Compared to previous CT muscularity indices, the accuracy was much higher with the new index in the HL (correlations between CT and dissection indices of 0.89 vs 0.51). The accurate measurement of femur length by CT used in the calculation of the new HL index made an important contribution to the higher accuracy of the index. The improvement in accuracy was smaller for the LR (0.55 vs 0.44). The association of CT muscularity indices and carcass quality in Texel and Scottish Blackface lambs showed that improved muscularity is not phenotypically correlated with detrimental effects on carcass composition. CT muscularity indices provide an alternative method to improve carcass conformation and leanness, using measurements that at a constant weight are independent of fatness.