The aim of this retrospective study was to evaluate the contribution of fluorine-18-fluoro-deoxyglucose (FDG) positron emission tomography (PET) to the clinical management and survival outcome of patients (pts) suspected of recurrent ovarian carcinoma, with the hypothesis that early diagnosis of recurrent ovarian cancer may improve overall survival (OS).Fifty-three FDG PET/CT scans were retrospectively analyzed for 42 pts. CT and PET/CT findings were confirmed by imaging and clinical follow-up, and/or pathology, which were considered as the gold standard diagnosis. The treatment plan based on CT staging was compared with that based on PET/CT findings. Medical records were reviewed for pts characteristics, progression-free survival (PFS), and OS. PFS and OS were analyzed using the Cox proportional hazards regression model.n = 16), or by a median clinical follow-up of 6.5 years (range 0.5-7.5) after the PET/CT (n = 37). PET/CT provided a higher detection sensitivity  than CT  (p < 0.001). Globally, PET/CT modified the treatment plan in 56.6% (30/53) and in 65.2% (15/23) when the CT was negative prior to PET/CT. In 30 cases, those benefited from a modified treatment plan, these changes led to the intensification of a previous treatment procedure in 83.3% (25/30), and to a reduction in the previous treatment procedure in 16.6% of cases (5/30). The Cox regression multivariate analysis showed that the number of lesions visualized by CT and presence of lung lesions detected by PET/CT were significantly associated with PFS .The final diagnosis of recurrence was established pathologically (On account of its impact on treatment planning, and especially in predicting patient outcome, FDG PET is a valuable diagnostic tool for cases of suspected ovarian cancer recurrence. Ovarian cancer has the highest mortality of all gynecological cancers . OvarianPositron emission tomography (PET) with fluorine-18-fluoro-deoxyglucose (FDG) has been proposed as a way of overcoming these limitations. It has been found to be highly sensitive for detecting recurrent ovarian cancer, especially in patients with an unexplained elevation of serum tumor markers. It offers the combined benefits of anatomical and functional imaging, and has been used to localize areas of increased FDG with improved anatomical specificity, and to exclude disease in sites of residual structural abnormality . A recenPatients included retrospectively were of over 18 years of age, with histologically proven ovarian cancer that was suspected of recurrence, and had been treated in our institution between 2006 and 2010. All patients of the institution received written information on the possibility of use of their medical data in retrospective studies. After initial diagnosis, the patients were monitored by physical examination, CA 125 assay, and CT every 6 months over a 5-year period. Recurrent disease was suspected following abnormal examination results and/or symptoms suggestive of recurrence, equivocal results on CI, and/or the isolated elevated serum tumor marker (CA 125). PET scans were acquired only if recurrence was suspected from the different criteria described above. The International Federation of Gynecology and Obstetrics (FIGO) classification was used for clinical staging . Patients fasted for ≥4 h before CT imaging. Scans were acquired in the craniocaudal direction, between the diaphragm and the perineum. After a series of unenhanced sections, all patients received intravenous bolus injection of contrast medium  at a rate of 2.5–3 mL/s and a volume of 75–90 mL. The Hi-Speed CT scanner generated contiguous slices measuring 5 mm in thickness and reconstruction interval was 1.25 mm.Scanning, from the patient’s head to the pelvic floor with their arm held above their head, was performed using a Siemens Biograph mCT40 fitted with a 20 cm axial field of view, true of flight (TOF) feature, and in-plane resolution of 4.4 mm in full width at half maximum . Images were reconstructed with Ultra HD and TOF . Patients fasted for at least 4 h before PET acquisition. Fasting blood glucose, which had to be <7 mmol/L, was checked prior to injection of 3 MBq/kg of FDG. Intravenous injection was followed by a tracer-uptake period of approximately 60 min, during which patients remained seated in a quiet room. Low dose-unenhanced CT was performed for localization and attenuation correction. The reformatted, transverse, coronal, and sagittal views were used for interpretation.CT and PET/CT clinical reports were collected and retrospectively analyzed. All PET/CT images were interpreted independently by two nuclear medicine physicians. The observers were aware of the clinical data and findings obtained with other anatomic imaging modalities. When abnormal FDG uptake was observed, its exact anatomical location was indicated on the CT scans. On PET images, the presence of a relapse tumor was suspected when accumulation of FDG was moderately to markedly increased, in comparison with that of comparable normal contralateral structures or surrounding tissues, excluding physiological bowel and urinary activity.For both ethical and practical reasons, not every suspected involved lesion was evaluated by histology. A gold standard reference was therefore established based on histology and 6 months follow-up data . True-positive results corresponded to an abnormal image by an imaging method (CT or PET/CT) confirmed by histopathology or follow-up. For example, foci that were detected by PET/CT, which were not histologically examined, were considered to be true-positives if the disease became obvious upon clinical observation or with the imaging methods during the follow-up. A negative finding on an imaging method was considered to be false-negative if positive by histopathology or follow-up. Indeed, a negative finding on an imaging method that was detected by another imaging method and confirmed by histopathology or by a clinical or imaging progression was considered to be false-negative. True-negative results corresponded to the absence of an abnormality by an imaging method, confirmation by a negative tissue biopsy, or lack of recurrence during the follow-up. For example, when no abnormality was found by PET/CT, or when no intervention was performed, the result was considered to be a true-negative if no disease was identified by other imaging studies or by clinical observation during 6 months follow-up. Finally, an abnormal result on an imaging method was considered to be false-positive if negative by histopathology or by follow-up. Suspicious involved lymph nodes were followed by CT although their size initially was not enough RECIST 1.1 criteria. Indeed, their development after treatment or not treatment has defined their status.t-test or the Mann–Whitney U test where necessary. Survival curves were calculated by means of the Kaplan–Meier method and compared by a log rank test. Multivariate analysis was performed by means of a Cox regression model. Analyses were performed using SAS 9.1  and Stata/SE 10 . All tests were performed at a two-sided significance level of α = 0.05.Results are expressed as mean values ±SD or median (range) in the case of small group sizes. Qualitative variables were compared using the Pearson test or the Fisher exact test where necessary . Paired qualitative variables were compared using the Mac Nemar test. Quantitative variables were compared using the Student’s We retrospectively evaluated 42 patients, with a median age of 45.5 years 35–81). For these patients, 53 PET/CT were performed. Patient characteristics are described in Table . For thep < 0.001), especially with a sensitivity of 92.2% for PET/CT compared to 60.8% for CI. The four PET/CT false-negative results were local recurrences, all of which were also negative by CT.Table n = 47), the sensitivity of PET/CT was 91.3% (42/47), compared to 50% (23/46) for CI. PET/CT accurately diagnosed a recurrent disease in 91.5% of patients (43/47) compared to 48.9% (23/47) by CI (p < 0.001). The most frequent site of recurrence was in the lymph nodes ] were analyzed: age, increased CA125, the number of CT lesions, the number of PET/CT lesions, and semi-quantitative indexes, such as SUVmax, SUVmean, SUVpeak, and total lesion glycolysis (TLG).p = 0.035) but also for patients with abnormal CT than with normal CT (p = 0.002), as described in Table p = 0.049).Lung lesions detected by FDG or abnormal CT were predictive of progression free survival (PFS) Figure . The PFSp = 0.034) or with bone foci (p = 0.020), as described in Table Bone or lung lesions detected only by FDG PET were predictive of OS, whereas CI was not contributive Figure . The OS To our knowledge, this study is the first of its kind to extensively follow-up suspected recurrent ovarian carcinoma by PET/CT imaging, with the goal of determining prognostic factors for disease-free and OS, as well as the clinical impact of FDG PET in this indication. There are two major findings in this current study: (1) the absence of lung recurrence sites by PET/CT imaging was independent and prognostic of a good PFS (42% 1-year PFS) and (2) the absence of bone lesions by whole-body FDG PET was an independent and robust prognostic factor (90.2% 1-year OS).Of all the variables analyzed , lung lesions detected only by PET/CT were significantly associated with PFS, while bone or lung metastases detected only by PET/CT were associated with OS. The semi-quantitative index of PET/CT has no prognostic value in our study. One possible explanation is the heterogeneity of the population studied, with one or more relapses per patient, different types of treatments, and different disease resistance to treatment. The results of our study are consistent with that of Hebel et al. . They dip < 0.05). PET/CT has good sensitivity in detecting lymph node metastases  in the diagnosis of ovarian cancer recurrence , CA 125 n = 16/53). However, clinical follow-up (n = 37/53) is also a valid way to evaluate diagnostic accuracy and response to therapy, but it would have been unethical to investigate all PET/CT detected foci using invasive procedures. Finally, the number of patients in the study was relatively small and it warranted the necessity to gather patients for survival analysis in order to retain a valid and strong power by statistical analysis.Our study has several limitations. First, it is a retrospective study with potential inherent biases, and further prospective studies are needed to confirm the results. Second, the ideal gold standard for any analysis is histological confirmation . Nonetheless, further evaluations appear necessary.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Staphylococcus aureus infection model that served as a surrogate for predicting efficacious exposures for the treatment of Neisseria gonorrhoeae infections. A wide safety margin to the efficacious exposure in toxicological evaluations supported progression to Phase 1. Dosing ETX0914 in human volunteers showed sufficient exposure and minimal adverse effects to expect a highly efficacious anti-gonorrhea therapy.With the diminishing effectiveness of current antibacterial therapies, it is critically important to discover agents that operate by a mechanism that circumvents existing resistance. ETX0914, the first of a new class of antibacterial agent targeted for the treatment of gonorrhea, operates by a novel mode-of-inhibition against bacterial type II topoisomerases. Incorporating an oxazolidinone on the scaffold mitigated toxicological issues often seen with topoisomerase inhibitors. Organisms resistant to other topoisomerase inhibitors were not cross-resistant with ETX0914 nor were spontaneous resistant mutants to ETX0914 cross-resistant with other topoisomerase inhibitor classes, including the widely used fluoroquinolone class. Preclinical evaluation of ETX0914 pharmacokinetics and pharmacodynamics showed distribution into vascular tissues and efficacy in a murine  Such infections include bone, chronic osteomyelitis and infections caused by prosthetic joints, valves and implantable cathetersThere have been repeated calls over the last few decades for the discovery and development of novel antibacterial agents to address widespread resistance to current treatment regimens145N. gonorrhoeae as an “Urgent Threat” requiring aggressive action as treatment failures against uncomplicated gonorrhea accumulate11N. gonorrhoeae resistant to the current first-line treatment options for gonorrhea including azithromycin and the extended spectrum cephalosporins, cefixime and ceftriaxone, raising concerns that gonorrhea will become untreatable1314N. gonorrhoeae.In 2013 the United States Centers for Disease Control and Prevention (CDC) released a report classifying drug-resistant 2B2 enzymes consisting of two units (A = GyrA or ParC) responsible for cleavage/re-ligation of the leading DNA strand, and two units (B = GyrB or ParE) responsible for ATP hydrolysis needed for passing the lagging DNA strand through the cleaved leading DNA strand. Fluoroquinolones, including ciprofloxacin 2, have been the most widely used class of antibacterials inhibiting topoisomerases. However, decades of fluoroquinolone use has led to extensive target site mediated resistance limiting the utility against S. aureus and N. gonorrhoeae among other bacteria163 of 18194 is the only member of this class that has been commercialized despite a diverse array of GyrB/ParE ATP competitive chemotypes that have been investigated21ETX0914 follows three other classes of antibacterial agents  that tarTherefore, in addition to a low potential for cross-resistance to current regimens, an agent advancing to clinical trials should demonstrate a sufficient margin between adverse effects in animal models and predicted efficacious doses. ETX0914 shows key attributes to support its development as a new antibacterial treatment option with a novel mode-of-inhibition relative to fluoroquinolones, NBTIs and novobiocin. The extensive preclinical and early clinical evaluations performed to support its progression to Phase 2 clinical trials are discussed in this manuscript.5  to plasma protein binding (PPB) of 5 were sufficiently high to warrant an extensive analog program. Pharmacokinetic (PK) properties of 5 were also favorable as seen by the low plasma clearance (CLp) and good bioavailability in dog. The subsequent analog program around 5 established a rather restricted structural latitude for antibacterial activity. The spirocyclic pyrimidinetrione of the scaffold could not be altered by substitution, isosteric replacement or truncation. The specific -stereochemistry around the fused morpholine ring was required for activity. The morpholine ring oxygen can be replaced with carbon, but at the expense of solubility, and the two methyl substituents proved to be optimal. The benzisoxazole fluorine atom was also optimal; though replacement with a chlorine atom afforded similar activity, it increased lipophilicity and decreased solubility and fu24Spiropyrimidinetriones were assembled via a key tertiary amino effect reaction affording a spirocyclic architecture . Previou5  was descin vitro toxicological data associated with compounds 6–8 and ETX0914 show a general progression to compounds of greater safety (in vitro through a mouse micronucleus aberration assay (MMA) and a mouse lymphoma assay as well as in vivo through a rat MMA, has been of concern for some fluoroquinolones including gemifloxacin5 showed a moderate, but measurable genotoxicity signal at 100 μM in the in vitro MMA; for comparison, the values for gemifloxacin and ciprofloxacin were 6 and 100 μM, respectively. Compound 6 with an N-linked triazole ring showed higher antibacterial activity than 1, but it induced micronucleation similar to gemifloxacin. Furthermore, dosing 6 at 100 mg/kg/day for 14 days in the rat led to haematologic effects and a potential for a reduction in bone marrow function as manifested by decreases in reticulocytes, neutrophils, lymphocytes and monocytes . With such favorable in vitro toxicological properties and DMPK characteristics, ETX0914 was thereby selected for clinical development.The broad tolerance for substitution at the benzisoxazole 3-position for antibacterial activity enabled diversification and optimization toward compounds that might mitigate undesirable mammalian toxicological effects. The r safety . Genotoxonocytes . Hence, in vitro . Bone mao assays . The lacrhythmia . MarginsS. aureus and Escherichia coli topoisomerases demonstrated that ciprofloxacin inhibited the re-ligation reaction of DNA gyrase by stabilizing a complex with DNA through a non-catalytic Mg2+ that interacts with an aspartate/glutamate  and by decatenating DNA (primarily by Topo IV). The complex course of the catalytic mechanism encompasses multiple conformational states and offers multiple possibilities for inhibition35ate E88,  and a sed assays . To difftrations . A similtrations . Hence, S. aureus (ARC2381), S. pneumoniae (ARC2480) and N. gonorrhoeae  that are resistant to fluoroquinolones due to three resistance determinants, S84, S85 and E88 (S. aureus numbering), but are fully susceptible to ETX0914. This follows previous studies where ciprofloxacin resistant S. aureus and S. pneumoniae, manifested by mutations in GyrA and ParC, were fully susceptible to spiropyrimidinetriones52+ chelated by fluoroquinolones; S85P is thought to alter the conformational integrity of the α-helix that spans S84 and E88, thereby altering ciprofloxacin binding  with reduced novobiocin susceptibility due to changes in the ATP binding region were generated by incubating the bacteria with ATPase inhibitors on agar plates. These mutants were fully susceptible to ETX0914, consistent with the compound’s lack of inhibition of ATPase activity as measured in Malachite Green based assays of DNA gyrase or Topo IVS. aureus mutant ARC2796 was derived from ARC516 and was not cross-resistant to ETX0914, nor was it cross-resistant to ciprofloxacin or novobiocin  and N. gonorrhoeae (1.5 × 10−8 to <5.2 × 10−9 at 4X MIC) relative to those seen for fluoroquinolones79437N and D429N mutations7To further investigate the mode-of-inhibition among the topoisomerase inhibitors, spontaneous resistant mutants to ETX0914 were assessed for cross-susceptibility in a int) of ETX0914 indicated that metabolism mediated clearance was low in human (1.0 μL/min/106 cells) relative to dog, rat and mouse (int in hepatocytes was predictive of the observed in vivo plasma clearance (CLp) values of 22 and 4 mL/min/kg observed, whereas the mouse Clint predicted a significantly lower in vivo clearance than that observed (CLp = 70 mL/min/kg). Pre-dosing with 1-aminobenzotriazole p nearly 5-fold to 15 mL/min/kg indicating that P450 metabolism contributed significantly to the high mouse CLp. The reason for the low turnover in mouse hepatocytes relative to the in vivo CLp could not be determined. These results led to murine efficacy experiments being carried out in the presence of ABT. The observed low apparent clearance in human from the healthy volunteer study (see below) confirmed that the higher clearance observed in mice was specific to mice. In addition, higher bioavailability (F) indicating low first pass effects was recorded in dog (71%) and monkey (58%) relative to mouse (46%) and rat (34%). The half-life after oral and intravenous administration in both dog and monkey was similar, indicating that absorption was not a rate limiting process in these species. PPB of 14C-ETX0914 was independent of concentration between expected  clinically relevant concentrations of 1–50 μM and was similar across species, ranging from 17.0% free (human) to 22.1% free (mouse). Free plasma levels were used for the assessment of pharmacodynamic (PD) and toxicological effects (below).In hepatocytes, the intrinsic clearance  and %T > MIC (R2 = 0.85) (The PK/PD index best-associated with ETX0914 efficacy was defined in a hollow fiber infection model (HFIM) against  = 0.85) .N. gonorrhoeae in animal studies due to spontaneous eradication of the bacterium; the existing animal model only provides a qualitative response negating a quantitative translation to clinical outcomeN. gonorrhoeae, a surrogate pathogen approach was undertaken utilizing PK/PD determinations from a mouse S. aureus neutropenic thigh model that has been shown to correlate with human clinical efficacyS. aureus and N. gonorrhoeae infections to assess the translatability of PK/PD indices  and the human fu of 17%, translated to a predicted efficacious mean AUC in humans of 49 μg*h/mL.The translatability of the PK/PD targets from animal models to clinical utility was established by comparing the free plasma PK/PD magnitudes obtained for the reference compounds in the in vitro characteristics observed in ETX0914 related to drug toxicity were followed by extensive in vivo toxicology investigations, which is particularly important for a drug with a new chemotype and inhibitory mechanism. Data collected included clinical observations, body weight changes, food consumption, clinical pathology and histopathology on dosing rats and dogs. In the rat, ETX0914 was tolerated up to 1000 mg/kg/day  for 28 days  were tolerated for 28 days. At this highest dose tested, tachycardia was observed, consistent with an IV telemetry dog study wherein decreases in blood pressure, contractility and PR interval and an increase in the heart rate were observed at 100 mg/kg. These effects were transient and fully reversible and, importantly, can be monitored in the clinical setting. There were no changes to other ECG intervals including QTc or QRS with ETX0914 administration. The dog NOAEL was established at 100 mg/kg PO for the 28 day study with a Cmax of 85 μg/mL and AUC24h of 870 μmol*h/mL measured on day 27. There were no significant changes in haematology or platelet count indicating anaemia or thrombocytopenia at the NOAEL in neither the rat nor dog,. Histopathology in both rat and dog at higher doses showed a variety of findings  was seen if the drug was preceded by a high fat meal. The Cmax increased from 22.9 to 24.0 μg/mL and the Tmax from 2.2 to 4.0 hours with food in line with solubility/absorption being rate limiting at the higher dose.A single center, multipart Phase 1 trial with an oral suspension of ETX0914 versus placebo was conducted in 48 healthy men and women of non-child bearing potential at doses ranging from 200–4000 mg. ETX0914 was absorbed relatively quickly under fasting conditions with a median T fashion . Followimax values for ETX0914 at the various doses also point to a margin of safety as seen by the multiples to the rat and dog NOAEL values (No serious adverse events (AEs) were encountered during the Phase 1 study. Mild and non-serious AEs entailed transient dysgeusia and headache, which were reversible in all subjects. There were no significant changes in haematological, clinical chemistry, urinalysis, vital signs or ECG. Doses of 2000 mg and 3000 mg were selected for a Phase 2 study in patients with uncomplicated gonococcal infections to maximize the probability of achieving the target ratio of plasma AUC to MIC in more than 90% of the patient population determined via a Monte Carlo simulation for the AUC distribution of ETX0914 (to be published elsewhere). The AUC and CL values . Hence, Mice, rats, dogs and monkeys were maintained under protocols in accordance with the Institutional Animal Care and Use Committee at AstraZeneca and IUCAC guidelines and were approved by in accordance with the American Association for Accreditation of Laboratory Animal Care . Rat and dog toxicology studies were performed in compliance with UK GLP regulations and OECD GLP principles and were approved by the United Kingdom Department of Health.S. aureus ARC2381, S. pneumoniae ARC548 and ARC 2800, and N. gonorrhoeae FA1090, ARC4672, and ARC4680 are contemporary clinical isolates. S. aureus ARC3445, ATCC33591-D1 and ATCC33591-D2, S. pneumoniae ARC2800, and N. gonorrhoeae 49226-TF, ARC4676-D1, ARC4676-D3 and ARC4676-D3-2 were described previously79S. aureus and S. pneumoniae isolates was performed using the broth microdilution method and against N. gonorrhoeae isolates using the standard agar dilution method. The quality control isolates obtained from the American Type Culture Collection and used during testing were N. gonorrhoeae ATCC49226, S. aureus ATCC29213, and S. pneumoniae ATCC49619. Reference antimicrobials  were obtained from the US Pharmacopeia Convention and MP Biomedicals , and were tested in accordance with CLSI recommendations. For isolates evaluated in animal efficacy studies, at least ten MIC replicates were performed to obtain the modal MIC. Modal MIC values were used for reporting and in PK/PD analyses of ETX0914 efficacy.The bacterial strains included in these studies are maintained in the AstraZeneca Research Collection (ARC). S. aureus (ARC516) were transferred onto plates of Mueller-Hinton agar containing the NBTI cis-4--1-[2-ethyl]cyclohexanolgyrA, gyrB, parC, and parE) were amplified by PCR and sequenced using standard protocols described previouslygyrA gene: Met121Lys. This same mutation was isolated by selection in S. aureus against NXL101Suspensions of cultures of methicillin-susceptible S. aureus ARC516 (inoculum 106 CFU/mL) grown in cellulosic cartridges were exposed to simulated steady state free plasma drug concentrations of ETX0914. The doses were administered once daily (q24h), every 12 h (q12h), or every 6 h (q6h) to vary the AUC/MIC, Cmax/MIC, and %T > MIC achieved across the cartridges. The system simulated a single-compartment model with mono-exponential elimination. A projected human half-life for both 6 h and 1 h infusion duration was applied in these studies. Serial samples were collected to ascertain simulated drug exposure and bacterial burden over 24 h. A one-compartment linear PK model was fit to the observed concentration-time profiles using WinNonlin 6.3. The change in bacterial burden at 24 h relative to the burden at time zero was correlated to drug exposures determined by the PK model, and a sigmoid Emax model was used to analyze the data and simulate the PK/PD indices.Bacterial response over 24 h to various concentrations of ETX0914 was determined using HFIM methods described elsewhere63S. aureus  prepared from a fresh subculture, to achieve a target inoculum of 5 × 105 CFU/thigh. ETX0914 was dosed subcutaneously 2 h post-inoculation. Two separate dose response studies were run for each isolate. PK parameters of ETX0914 in infected mice were evaluated following single doses. Blood samples were collected from 3 mice per time point, and a total of 9 time points were collected per dose over 24 h. Plasma was obtained after centrifugation of blood samples in the presence of EDTA and stored at −20 °C until bioanalysis. Concentrations of ETX0914 in mouse plasma were measured by liquid chromatography-mass spectrometry (LC/MS/MS), for which the assay was linear over 0.005–10 μg/mL (r2 = 0.995) and quality control accuracy was 100 ± 15%. To reflect free drug, simulated plasma concentration-time data were corrected for ETX0914 protein binding in mouse plasma, 22.1%. PK parameters were determined from single dose data by compartmental analysis . Plasma exposures following multiple dose administration in efficacy studies were simulated by mean parameters and verified. The correlation between efficacy and the PK/PD parameter of AUC/MIC was determined using a sigmoidal Emax model, and the AUC/MIC magnitudes for different efficacy end points were calculated.Female CD-1 mice were rendered neutropenic by intraperitoneal injections of cyclophosphamide 150 mg/kg on Day -4 and 100 mg/kg on Day -1 prior to infection. Two hours prior to infection, mice received 50 mg/kg ABT orally. Mice received a second 50 mg/kg ABT administration 12 h later. Thigh infections were produced by intramuscular injection of a mid-log culture of 2. The following were assessed: clinical observations, body weight, food consumption, body temperature, hematology, plasma chemistry, toxicokinetics and gross and microscopic pathology. All necropsies consisted of an external and internal examination including body orifices  and cranial, thoracic, abdominal organs and tissues.The following dosing formulation for rat and dog 28 day PO studies was used: a 100 mg/mL stock of ETX0914 in 5% dextrose and 2.5M meglumine adjusted to pH 9 using 5M HCl or 2.5M meglumine. The dosing formulations were diluted as required with phosphate buffered saline and dispensed daily. Formulated ETX0914 was administered once daily by oral gavage for 28 days, at dose levels of 200, 500 or 1000 mg/kg/day to groups of 10 male and 10 female Han Wistar rats. A control group of 10 male and 10 females were given the vehicle only. An additional 3 animals per sex were allocated to each group for toxicokinetics wherein blood samples (1 mL) from the tail vein pre-trial and on Day 27 were collected. An additional 5 animals per sex were assigned to the control and high dose groups to demonstrate the reversibility of any findings over a 3 month (91 day) recovery period. Animals were terminated by exposure to COFormulated ETX0914 was administered once daily by oral gavage for 28 days, at 100, 200 or 500 mg/kg/day to groups of 3 male and 3 female beagle dogs. A control group of 3 male and 3 females were given the vehicle only. Three animals per sex were assigned to the control and high dose groups to demonstrate the reversibility of any findings over a 92 day recovery period. The electrocardiogram (ECG) of each animal was recorded using a DSI jacketed external telemetry system. Recordings were obtained from all animals once pretrial and on Days 2, 23 and 27 of the dosing period and then in the final week of the recovery period. The following parameters were recorded: PR, QRS, QT and heart rate. An additional recording was obtained from 50 days after the completion of dosing for the females or 51 days after the completion of dosing from the males. The animals were fasted overnight before collecting blood samples (1 mL) from the jugular vein pre-trial and on Day 27. The blood samples were then transferred into tubes containing anticoagulant (0.9 mL 3.8% trisodium citrate). All study animals were terminated by intravenous injection of sodium pentobarbital for necropsy on the day after the final dose was given (day 29). or on day 120 for reversibility studies. All necropsies consisted of an external and internal examination including body orifices  and cranial, thoracic, abdominal organs and tissues.Samples of the following tissues from both rats and dogs taken at necropsy from all main and recovery study animals were recorded and tissues were weighed, fixed and preserved: Adrenal glands, aortic artery, rib bone marrow smear , brain, epididymides, eyes, femoral bone , gross lesions/masses, gall bladder, gut associated lymphoid tissues, heart, intestinal duodenum, intestinal ileum, intestinal jejunum, intestinal colon, intestinal caecum, intestinal rectum, kidneys, liver, lacrimal gland, lungs, mandibular lymph node, mandibular, mesenteric lymph node, axiliary lymph node (for dogs), skeletal muscle (from thigh), sciatic nerve, oesophagus, optic nerves, ovaries, pancreas, pituitary gland, prostate gland, mandibular salivary glands, parotid salivary glands, skin and mammary gland, spinal cord , spleen, sternum (including bone marrow), stomach, testes, thymus, thyroid glands with parathyroid glands, tongue, trachea, urinary bladder, ureter, uterus with cervix and oviducts, vagina. Bone marrow smears were stained using May-Grunwald-Giemsa as the Romanowsky stain. Modified Davidson’s fixative was used for testes tissues, Davidson’s fixative for eyes and optic nerve tissues and 10% neutral buffered formalin for all other tissues. Wax blocked tissues from all animals were cut into 4–6 μm thick sections and stained with haematoxylin and eosin, and examined by light microscopy. Blood (0.5 mL) for rat haematology analyses was taken via the lateral tail vein and analyzed using a Siemens, ADVIA 120 haematology analyzer.A Phase 1 multipart studyHow to cite this article: Basarab, G. S. et al. Responding to the challenge of untreatable gonorrhea: ETX0914, a first-in-class agent with a distinct mechanism-of-action against bacterial Type II topoisomerases. Sci. Rep.5, 11827; doi: 10.1038/srep11827 (2015).


These preliminary data suggest that a DUS-based assay could bypass sociocultural barriers and sampling issues and therefore could be a suitable, effective tool for epidemiological surveillance and screening programs, especially in low-income countries.Nowadays, several screening strategies are available to prevent cervical cancer, but inadequate resources, sociocultural barriers, and sampling issues impede their success in low-income countries. To overcome these issues, this study aimed to evaluate the performance of human papillomavirus (HPV) testing from dried urine spots (DUS). Eighty-eight urine samples (including 56 HPV DNA positive specimens) were spotted on filter paper, dried, and stored in paper-bags. HPV DNA was detected from the DUS after 1 week and 4 weeks of storage using a polymerase chain reaction (PCR) assay. The sensitivity, specificity, and concordance of the DUS-based HPV test were evaluated by comparing the results with those of HPV testing on fresh urine samples as the gold standard. The sensitivity of the test was 98.21% (95% CI: 90.56–99.68) for DUS stored for 1 week and 96.42% (95% CI: 87.88–99.01) for DUS stored for 4 weeks. The specificity was 100% (95% CI: 89.28–100) at both time points. The concordance between DUS and fresh urine HPV testing was “almost perfect” using the  Cervical cancer is a relevant public health problem for women worldwide, being the third most frequent cancer and the fourth most common cause of death from cancer in women , 2. OverHPVs are DNA viruses that are grouped into cutaneous and mucosal types according to their infection site and further subdivided into high-risk (HR) and low-risk (LR) types, depending on their association with disease malignancy. The International Agency for Research on Cancer (IARC) has included 25 types of HPV in the high-risk clade (HR-clade) by subdividing them into three groups . HPV typDifferent types of screening for cervical cancer are now available, such as the conventional cytology test (Pap test) and liquid based cytology (LBC), visual inspection with acetic acid (VIA), and HPV testing for HR-HPV from cervical brush. These types of screening require adequate financial resources, a developed infrastructure, trained labour, and surveillance mechanisms for screening, investigating, treating, and following up on targeted women , 6. MoreAlternative tools that can overcome these problems could improve screening coverage and reach the female population at risk in developing countries.The use of HPV testing on urine, a noninvasive and easy-to-collect sample, could be more attractive to women because it bypasses medical examination, as well as sociocultural and religious implications. The correlation between the detection of HPV in urine and cervical samples has been reported in several studies in the literature –12. In aHowever, urine samples require restrictive conditions for storage and transportation, especially when samples must be transported over large distances in a warm climate. This factor is particularly relevant in low-income countries where analysis laboratories can be located far away from the rural areas where women live. An alternative method of sample collection is the use of dried urine spots (DUS), that is, a urine sample spotted on blotting paper, which allows for stabilization by drying. This approach has logistical benefits because DUS are small and easily transported and the specimens can be stored at room temperature.DUS sample collection solves several problems associated with the sampling and storage of fresh urine samples. Therefore, the DUS approach is particularly interesting, especially in developing countries, even considering that dried spots on filter paper can be successfully used for the detection of various infectious agents, as well as metabolic and genetic diseases –16.Few studies have evaluated the use of dried samples on filter paper for the detection of HPV infection, and these investigations examined exclusively cervical brush samples –19. No dThus, the aim of this study was to evaluate the performance of HPV testing from DUS and to compare the results obtained with those from paired fresh urine samples. Chlamydia trachomatis infections [Urine samples were obtained from 88 immigrant women : 28–43 years) who attended NAGA Onlus in Milan, Italy, between June 2012 and December 2013 and were included in a large epidemiological study on HPV andOf the urine samples, 56/88 (63.6%) were HPV DNA positive and 32 (36.3%) were HPV DNA negative. Of the HPV DNA positive samples, 40/56 (71.4%) were sustained by single infections (24 belonging to HR-clade genotypes and 16 to LR genotypes) and 16/56 (28.6%) were caused by multiple infections (4 were caused by LR genotypes and 12 by at least 1 genotype of the HR-clade).μL urine sample was subdivided into eight 50 μL aliquots that were each spotted on preprinted circles on a piece of filter paper . The DUS filter papers were dried for 3 h and then stored in paper bags in a dry location at room temperature  for either 1 week or 4 weeks until the analyses took place. The analyses were carried out at the Laboratories of the Department of Biomedical Sciences for Health, University of Milan, Italy.Each 400 μL, was extracted using the commercial NucliSENS EasyMAG method , according to the manufacturer's instructions. The nucleic acids were eluted in 100 μL of the NucliSENS elution buffer.Four preprinted circles were punched or cut out from each piece of DUS filter paper using a sterile single-cut paper-punching machine or a new sterile scalpel blade. The circles were transferred into a 1.5 mL tube containing 1 mL of NucliSENS Lysis Buffer  and incubated on a roller mixer for 30 minutes at RT. Then the tube was centrifuged for 15 s at 1500 ×g. The lysate, with a volume of approximately 750 The concentration of the extracted DNA was evaluated using a spectrophotometer . The quality of DNA was validated by detection of a 268 bp fragment of the housekeeping beta-globin gene using an in-house polymerase chain reaction (PCR) assay .HPV DNA was detected using an in-house nested-PCR assay based on the amplification of a 150 bp open reading frame late gene 1 (ORF L1) fragment. The nested-PCR assay was performed using a two-step amplification to either increase sensitivity or mitigate the inhibitory effect of substances potentially present in the sample. Every PCR reaction included positive  and negative (water) controls. Strict laboratory precautions and quality assurance/quality control measures were followed to avoid cross contamination and carry over PCR. ELSI_F/ELSI_R primers were used for the first cycle of amplification and GP5+/GP6+ primers were used for the nested reaction, as previously described , 22.The amplification products were visualized by means of electrophoresis analysis on 2% agarose gels containing ethidium bromide (0.5 mg/mL). The amplified products were compared with molecular weight standards .HPV DNA positive DUS isolates were genotyped using INNO-LiPA HPV Genotyping Extra , a line probe assay based on the principle of reverse hybridization, according to the manufacturer's instructions. The resulting line patterns allowed for identification of 28 different HPV genotypes: 6, 11, 16, 18, 26, 31, 33, 35, 39, 40, 43, 44, 45, 51, 52, 53, 54, 56, 58, 59, 66, 68, 69, 70, 71, 73, 74, and 82.The sensitivity and specificity of HPV testing from DUS were evaluated in comparison to HPV testing from fresh urine samples as the gold standard. These results were presented as percentages with 95% confidence intervals (95% CIs). Proportions (95% CI) were calculated using the Wilson score model by the OpenEpi statistical program (version 3.01), which is available online .κ) statistic was calculated by dividing the difference between the observed proportion of agreement and the expected proportion of agreement by 1 minus the expected proportion of agreement. The concordance was defined as “poor” (k = 0), “slight” (0.01 ≤ k ≤ 0.20), “fair” (0.21 ≤ k ≤ 0.40), “moderate” (0.41 ≤ k ≤ 0.60), “substantial” (0.61 ≤ k ≤ 0.80), “almost perfect” (0.81 ≤ k < 1.00), or “perfect” (k = 1.00).To determine the proportion of agreement between DUS and fresh urine testing, Cohen's unweighted kappa  tested HPV DNA positive after 1 week. In contrast, HPV DNA was detected in 54/56  of the DUS samples after storage for 4 weeks.The DUS sample that tested HPV DNA negative after both 1 and 4 weeks of storage was prepared from an HPV-72 (LR HPV) infected urine sample. The other DUS that tested HPV DNA negative only after 4 weeks of storage was prepared from an HPV-56 (HR HPV) infected urine sample.All DUS prepared using HPV DNA negative urine samples tested negative for HPV DNA.κ statistic ≥ 0.81) (The sensitivity of the HPV DNA test in DUS at 1 and 4 weeks was 98.21% (95% CI: 90.56–99.68) and 96.42% (95% CI: 87.88–99.01), respectively. The specificity for the test was 100% (95% CI: 89.28–100) after both 1 week and 4 weeks of storage. The proportion of agreement between the DUS and fresh urine tests was “almost perfect” ( ≥ 0.81) .Of the 40 DUS prepared from fresh urine samples of women infected by a single HPV genotype, 38 (95%) tested HPV DNA positive after four weeks of storage. All amplified fragments from the DUS were properly genotyped, and the distribution of HPV genotypes matched across the two sample types . The disTo our knowledge, this is the first report of HPV detection from urine samples stored on filter paper. The quantity and the quality of DNA extracted from DUS were comparable to those obtained by standard collection from fresh urine, as estimated by the spectrophotometric readings and the detection of the housekeeping beta-globin gene using an in-house PCR assay. Accordingly, the high molecular weight DNA extracted from DUS samples is sufficient to perform molecular assays, either traditional or high throughput.The data obtained showed an elevated concordance between HPV DNA detection in DUS and fresh urine samples. Cohen's unweighted kappa values were very high, indicating an “almost perfect” agreement.These preliminary data support the use of DUS as simple, rapid, and safe sampling for HPV DNA detection and genotyping by using molecular tests, such as PCR and line probe assay based on the principle of reverse hybridization. Furthermore, the use of DUS strengthens the already recognized advantages of urine samples. Collection of these noninvasive specimens is more acceptable and can bypass the ethical, social, and religious barriers of speculum exams for the collection of conventional cervical brush , 8, 11.Moreover, drying urine samples on filter paper allows the DNA to be protected from degradation for a long period, as shown by the high percentage (96.4%) of HPV DNA positive samples detected after four weeks of storage at room temperature.Due to the introduction and increasing availability of novel and powerful high-throughput molecular technologies, the optimization of methods for biological samples collection and storage has become a critical issue. These results highlight the potential use of DUS samples as an alternative means of biobanking, avoiding the high costs and logistical problems associated with the storage and transportation, especially where a cold chain is absent , 24. FinHPV testing from DUS showed an elevated sensitivity and specificity and a high concordance rate compared to HPV testing from fresh urine samples. These preliminary data suggest that a DUS-based assay could bypass sociocultural barriers and sampling issues. This approach could be a suitable and effective tool for epidemiological surveillance and screening programs, especially in low-income countries.


Plasmodium infections were common at baseline, with 41·1% of children positive for Plasmodium falciparum/Plasmodium malariae, 9·2% for P. falciparum/ Plasmodium ovale spp. and 8·0% for all three species. Moreover, on day 17, 39·9% of children infected with falciparum malaria at baseline were again positive for the same species, and 9·2% of those infected with P. malariae at baseline were positive for P. malariae. Here, chronic multi-species malaria infections persisted in children after AL treatment(s). Better point-of-care diagnostics for non-falciparum infections are needed, as well as further investigation of AL performance in asymptomatic individuals.During a longitudinal study investigating the dynamics of malaria in Ugandan lakeshore communities, a consistently high malaria prevalence was observed in young children despite regular treatment. To explore the short-term performance of artemether-lumefantrine (AL), a pilot investigation into parasite carriage after treatment(s) was conducted in Bukoba village. A total of 163 children (aged 2–7 years) with a positive blood film and rapid antigen test were treated with AL; only 8·7% of these had elevated axillary temperatures. On day 7 and then on day 17, 40 children (26·3%) and 33 (22·3%) were positive by microscopy, respectively. Real-time PCR analysis demonstrated that multi-species  Challenges to effective malaria control in this country include an extremely high transmission intensity with entomological inoculation rates greater than 100 per year in many areas, a weak health system with a fragile supply chain for front-line antimalarial treatments, inadequate case management with poor diagnostic tools in rural health centres and ineffective monitoring and evaluation of control interventions  and lactate dehydrogenase , display marked differences in their sensitivity, stability and dynamics of clearance from the blood after antimalarial treatment  promotes a ‘test-and-treat’ strategy for malaria involving parasitological diagnosis rather than presumptive treatment based on clinical indicators WHO, a. The goP. falciparum remains the major cause of morbidity and mortality in sub-Saharan Africa, non-falciparum infections, such as Plasmodium vivax, Plasmodium malariae and Plasmodium ovale spp., are also present in humans and their clinical significance should not be overlooked   or First Response RDTs , OptiMAL RDTs  and microscopy on Giemsa-stained blood films as described  community-based longitudinal study was conducted in six villages on the shores of Lake Albert and Lake Victoria in Uganda, commencing in May 2009  within the SIMI cohort (N = 188) were recruited to the study on the basis of a positive First Response RDT result and microscopy-confirmed malaria. Children were subsequently followed up 7 and 17 days after day 0 (day of initiation of treatment). Due to financial and logistical constraints, only individuals who were RDT-positive on day 17 (see below), who had been given oral quinine sulphate as an alternative exit treatment, were followed up on day 24. A blood smear archive was made on days 0, 7 and 17, and confirmatory microscopy was carried out 2 or 3 days after each sample was taken . In addition, haemoglobin (Hb) levels were recorded for each child using a HemoCue spectrometer  on days 0, 7 and 17.Bukoba village, Mayuge District, showed the highest prevalence of falciparum and non-falciparum malaria in young children see  and so wP. falciparum infections  and albendazole (400 mg) according to WHO guidelines  and glutamate-rich protein (glurp) was carried out according to published methods  and R v2.10.1 . Anaemia was categorized based on haemoglobin levels as follows: mild 10–11 g dLL−1 WHO, . To deteL−1 WHO, .Ethical approval was provided by The London School of Hygiene and Tropical Medicine and the Ugandan National Council of Science and Technology. Informed consent was given by mothers on behalf of their children and documented in writing or by thumbprint (in cases of illiteracy).S. mansoni infection was very low, 9·0% of children had egg-patent hookworm infections.μL−1 blood. On day 7, 40 children (26·3%) remained malaria positive by microscopy and on day 17, 33 children (21·3%) had microscopically detectable malaria infections. After genotyping using msp1, msp2 and glurp loci, 33 out of 40 and 17 out of 33 children were shown to have recrudescent P. falciparum infections on days 7 and 17, respectively. By real-time PCR 96·9% of children were positive for P. falciparum on day 0 and multi-species Plasmodium infections were common, with 41·1% positive for P. falciparum/P. malariae, 9·2% positive for P. falciparum/P. ovale spp. and 8·0% for all three species . Thus strong evidence for clustering of malaria infections after AL treatment was not found, although this may reflect a lack of power due to the small numbers of infected individuals on days 7 and 17.To determine whether there were differing risks of infection after treatment for children living in different parts of the village, their households and infection status on days 7 and 17 were mapped . SpatialP. falciparum parasitaemia 7 days after treatment and that 22·3% (11·7% recrudescent) had parasites in their blood after 17 days, despite the fact that 33% of these positive children had already been retreated with AL at day 7. A substantial number of additional recurrent infections were identified by real-time PCR, including P. malariae and P. ovale spp. infections. Although none of these infections could be considered ‘complicated’ clinical malaria, their significance needs to be better quantified.In light of the challenges in diagnosis of falciparum and non-falciparum malaria and the chronically high prevalence of infections in Bukoba village, Uganda, we document the short-term effect of AL treatment on multiple malaria species in a cohort of young children living in this area. The study was designed to gain insights into treatment responses when AL was distributed to parasitaemic children in a community-based setting (unsupervised treatment), rather than as a carefully controlled standard efficacy trial. It is outside the remit of this paper to explore how well the results of standard efficacy trials predict the effectiveness of home-based management strategies in the same population. However, we found by microscopy that 26·3% of treated children (21·9% recrudescent by PCR genotyping) showed et al.et al.et al.et al.et al.et al.et al.et al.et al.et al.et al. reported a small decline in parasitological response rates to ACTs (including AL) in coastal Kenya  in recurrent infections soon after AL treatment , there was negligible risk of parasitaemia before day 21 and that risks of treatment failure by day 28 ranged from 1–9%  and those with lower parasitaemia (data not shown). Similar responses to AL were also observed when children with fever or moderate/severe anaemia on day 0 were compared with those with no fever or anaemia. However, the number of symptomatic individuals in our study may be too small to detect subtle differences in responses to AL treatment between symptomatic and asymptomatic individuals.This study was based on assessment of parasitaemia rather than clinical illness and most children had asymptomatic infections on day 0. The 21-day efficacy of the ACT dihydroartemisinin piperiquine (DP) for treatment of et al.et al.et al.et al.et al.et al. in Kenya, who showed that residual sub-microscopic parasitaemia on day 3 after commencement of AL or DP treatment was associated with longer gametocyte carriage, increased transmission to mosquitoes and recurrence of microscopically detectable malaria infections on day 24 or day 48 (Beshir et al.There has been little work assessing the dynamics of parasite clearance after AL treatment using PCR, although a median clearance time of 2 days has been reported (Aydin-Schmidt P. malariae and P. ovale infections detected by real-time PCR at baseline were mixed infections with P. falciparum, which were difficult to detect on thick films despite cross-checking. Plasmodium malariae was not fully sensitive to AL in this study as around 9% of infected individuals were still positive on day 17, and two children were P. malariae positive at day 24 after three antimalarial treatments. The fact that 50% of study participants were infected with P. malariae and 18% with P. ovale indicates that the number of mixed infections receiving ACTs is substantially larger than previously thought. Mombo-Ngoma et al. reported, in 38 Gabonese patients, a 28-day cure rate by microscopy of 100% for AL in treatment of P. malariae, P. ovale and mixed infections in Gabon, although post hoc PCR revealed that only 19 of the patients in the study had been correctly diagnosed with non-falciparum malaria (Mombo-Ngoma et al.et al. detected persistent P. malariae and P. ovale spp. infections in Ghanaian schoolchildren 21 days after ACT treatment using qPCR as the endpoint measure (Dinko et al.P. malariae and P. ovale spp. would be of great assistance in these endeavours.The vast majority of et al.It must be borne in mind that our study did not formally determine how well the participants complied with the approved antimalarial treatment regime, i.e. two doses a day for 3 days after consumption of fatty food WHO, a. Measurin vivo study protocols, with sensitive molecular endpoints, must now be developed that are specifically designed to measure the efficacy of antimalarial regimens against asymptomatic and sub-patent Plasmodium spp. infections.The burden of malaria remains high in Uganda. Whilst the Ugandan malaria policy promotes AL as the first-line antimalarial treatment and is doubtless saving many lives, other challenges in its implementation remain. Our results suggest that even though ACTs remain highly efficacious in treatment of clinical disease in sub-Saharan Africa, asymptomatic and submicroscopic falciparum and non-falciparum infections often persist after treatment. These infections may be significant in terms of later recurrence of clinical disease and parasite transmission. Improved diagnostics are needed for field-based detection of non-falciparum malaria infections, gametocytes and drug-resistance markers. Increased community-based surveillance using a range of diagnostic tests is required to provide further insights into the extent of persistent parasitaemia after ACT treatment and for early detection of ACT-tolerant parasites in sub-Saharan Africa. Finally, new


Polycystic ovary syndrome (PCOS) is a common endocrinopathy, associated with metabolic abnormalities. Metabolic features of various phenotypes of this syndrome are still debatable. The aim of present study hence was to evaluate the metabolic and hormonal features of PCOS phenotypes in comparison to a group of healthy control.A total of 646 reproductive-aged women were randomly selected using the stratified, multistage probability cluster sampling method. The subjects were divided into five phenotypes: A (oligo/anovulation + hyperandrogenism + polycystic ovaries), B (oligo/anovulation + hyperandrogenism), C (hyperandrogenism + polycystic ovaries) and D (oligo/anovulation + polycystic ovaries). Hormonal and metabolic profiles and the prevalence of metabolic syndrome among these groups were compared using ANCOVA adjusted for age and body mass index.insulin%20resistance (HOMA-IR), compared to PCOS women of group D. Serum concentrations of cholesterol, low density lipoprotein, triglycerides and glucose in group A were higher than in other phenotypes, whereas the metabolic syndrome was more prevalent among group B.Among women with PCOS (n = 85), those of groups A and C had higher serum levels of insulin and homeostatic model assessment for Women who had all three components of the syndrome showed the highest level of metabolic disturbances indicating that metabolic screening of the severest phenotype of PCOS may be necessary.The online version of this article (doi:10.1186/1477-7827-12-89) contains supplementary material, which is available to authorized users. Polycystic ovary syndrome (PCOS) is one of the most common, complex and heterogeneous endocrine disorders –4 affectinsulin%20resistance (HOMA-IR) among groups B and D [IR is strongly linked to PCOS  and hype B and D ; hence i B and D , 23.Due to a lack of adequate population based studies on the metabolic aspects of the various PCOS phenotypes, we aimed to compare the metabolic parameters of four PCOS phenotypes to a group of non-PCOS controls in a community based study, conducted in the southwest of Iran.Ethical review board of the Research Institute for Endocrine Sciences see Additional file A stratified, multistage cluster with a probability in proportion to size procedure was used for the sampling method. The study design, recruitment process and data collection have been described previously in detail elsewhere . In brieA standard questionnaire was completed for eligible women n = 625). All participants underwent clinical examinations by trained staff of local medical universities/schools and their body weight, height, waist (WC), hip circumferences (HC) and blood pressure were measured and documented. BMI was calculated as weight in kilograms divided by height in meters squared (kg/m2). Acne was scored based on its number, type and distribution [. All parInsulin (Ins) was measured by immunoenzymometric assay (IEMA), . Glucose (Glu), Triglycerides (TG), Total Cholesterol (TC), Low Density Lipoprotein (LDL) and High Density Lipoprotein (HDL) were measured by enzymatic colorimetery, . HOMA-IR was calculated as fasting insulin (mIU/L) * fasting glucose (mg/dl) /405.The intra- and inter-assay coefficients of variation for Ins were 0.9% and 1.1%, respectively; for Glu they were 1.3% and 2.9%, for TG 1.8% and 2.7%, for TC 0.8% and 2.8%, and for LDL 0.7% and 2.9%; and for HDL, these values were 0.9% and 3.3%, respectively.We defined PCOS in our study using the Rotterdam (Rott) criteria, by which PCOS was defined as presence of two or more of the following: oligo/amenorrhea, clinical and/or biochemical hyperandrogenism, and polycystic ovaries . Women wHyperandrogenism (HA) was considered as clinical Metabolic syndrome was defined by the clinical diagnostic criteria used for Iranian adult metabolic syndrome as the presence of any three of five of the following characteristics: (1) Waist circumference ≥ 95 cm; (2) Fasting TG ≥ 150 mg/dl or drug treatment for elevated triglycerides; (3) HDL cholesterol < 50 mg/dl or drug treatment for reduced HDL-C; (4) Systolic blood pressure ≥130 or diastolic blood pressure ≥ 85 mmHg or treatment with antihypertensive medication, and (5) Fasting glucose ≥ 100 mg/dl or treatment with diabetes medication .th and 75th percentages, following testing for normality, and categorical variables, are expressed as percentages. Demographic and anthropometric features between phenotypes are compared using one way ANOVA and/or the Kruskal-Wallis Hand or Pearson’s χ2 test, as appropriate. The association between PCOS manifestations with hormonal and metabolic parameters are analyzed using ANCOVA, adjusted for age and BMI. The association between MetS and phenotypical groups adjusted for age and BMI are analyzed using logistic regression model. Data analysis was performed using the SPSS 15.0 PC package and statistical significance was set at P < 0.05.Continuous variables are presented as mean and standard deviation or median and the 25Of the 602 participants who completed the study, 85 women met the PCOS criteria; the most common phenotype among them was C (49.4%) i.e. they met two criteria of PCOS including hyperandrogenism and polycystic ovaries; group B (22.4%), D 15.3%), and A (12.9%) followed in that order. Non-PCOS women were significantly older than those with PCOS ; however there were no significant differences in mean weight, height, BMI, WC, HC, waist to hip ratio (WHR) between PCOS and non-PCOS participants. Table 5.3%, andPCOS women showed significantly higher serum levels of total testosterone (TT) , androstenedione (A4) , dehydroepiandrosterone sulfate (DHEAS) , free androgen index (FAI)  and prolactin (PRL)  versus non-PCOS ones. Table χ2 test revealed that non-PCOS participants indicated a significantly higher number of women with MetS, compared to PCOS ones (P = 0.005). Regarding the phenotype groups, Pearson’s χ2 test showed that group C included fewer number of women with MetS compared to those without PCOS (P = 0.01). Multivariate logistic regression showed that after adjustment for age and BMI, the prevalence of MetS in group C was significantly lower than in the non-PCOS women .Pearson’s The present community-based study revealed that PCOS phenotypes with hyperandrogenism  had the worst metabolic presentations in terms of insulin resistance and metabolic syndrome. Furthermore the lipid and glucose profiles of women with mild phenotype (D) were better than the other PCOS groups, even non-PCOS ones. Our findings also indicate that concurrency of the three PCOS symptoms leads to increased severity of metabolic disorders, especially hyperinsulinemia which may be related to more severe hyperanrogenemia.Similar to our findings, a study conducted on 93 Polish women with PCOS, showed elevated serum concentrations of TT, Chol and LDL in the classic phenotype of PCOS i.e. group A ; other rIt is noteworthy that although PCOS women have shown higher risk of cardiovascular diseases, yet hyperandrogenism has not been proven to be the main contributor to future cardiovascular diseases ; howeverOur findings of lower lipid- and Glu- profile-related disorders in normoandrogenic patients (group D) were consistent with the androgen excess and the PCOS society’s claim that PCOS should initially be considered an androgen excess disorder . Group DTwo studies documented an association between AnOvu and metabolic disorders , 32, altThe slight differences between results of studies are probably due to genetic and environmental factors, in addition to heterogeneity of PCOS phenotypes. Similar to our findings, in another study conducted in Iran, the prevalence of MetS in phenotype B was reported the highest. A study conducted by Mehrabian et al has shown that PCO has negative association with MetS  and the This study does have its strengths, such as using a community-based sample instead of recruitment from a referral centre. In addition, we used national cut points of WC and HOMA-IR for the definition of abdominal obesity and IR, respectively. To mention the limitations, since aging decreases serum androgen levels and increases the prevalence of IR and MetS , a limitIn conclusion, our results indicate that, women with hyperandrogenism exhibit the worst metabolic features. As a result screening of phenotypes with hyperandrogenic symptoms for detection of metabolic abnormalities is highly recommended in the scope of PCOS. Further longitudinal population based studies, with a prospective risk assessment approach in larger sample sizes are needed to analyze the long-term effects of metabolic profiles of each phenotype.Ethical approval.(DOCX 973 KB)Additional file 1:Consent.(DOCX 16 KB)Additional file 2:


Staphylococcus haemolyticus, with S. hominis, S. warneri, S. cohnii, and Staphylococcus epidermidis commonly detected. Antimicrobial susceptibilities and carriage of the mecA gene were determined for all of these isolates. Most (85.9%) staphylococci were resistant to multiple antibiotics with all displaying increased susceptibility toward penicillin, fusidic acid, erythromycin, and cefepime. Twenty-one (29.5%) of the isolates were mecA positive, however MIC values to oxacillin, normally associated with the carriage of mecA, varied widely in this group (from 0.06 to 256 mg/L). Fifteen of the twenty-one mecA positive isolates carried SCCmec of these seven were type V, one type I, one type II, and one type IV. Additionally, five of these 15 isolates carried a previously unreported type, 1A, which involves an association between class A mec complex and ccr type 1. The remaining six of the 21 isolates were non-typeable and carried a combination of class A mec complex and ccrC. In addition to this, we also report on new MLST types which were assigned for five S. epidermidis isolates. Four out of these five isolates had MICs between 0.06 and 256 mg/L to oxacillin and would be regarded as clinically susceptible but one isolate had a high oxacillin MIC of 256 mg/L. We demonstrated widespread multiple drug resistance among different staphylococcal species isolated from non-healthcare environments highlighting the potential for these species to act as a reservoir for methicillin and other forms of drug resistance.Antibiotic resistance in bacteria isolated from non-healthcare environments, is a potential problem to public health. In our survey a total of 71 coagulase negative staphylococci (CNS) belonging to 11 different species were isolated from three large hotels in London, UK. The most prevalent species was  S. epidermidis, S. haemolyticus, and S. hominis are amongst the CNS which can be responsible for a wide range of illnesses from minor skin infections to life threatening diseases   and ccr types   are opportunistic pathogens that have emerged as a major cause of nosocomial infections often associated with healthcare settings  outside of hospital settings, has been reported by several authors and their studies demonstrated that non-healthcare settings, including fire stations, student housing sites/common areas, public buses can be reservoir for MRSA .In addition to these samples, 12 air samples were also collected from 12 randomly selected rooms using a high volume air sampler . The airflow of sample collection was between 200 and 1000 l per min.All specimens were inoculated onto Nutrient Agar  and Mannitol Salt Agar plates . These cultures were incubated aerobically at 37°C for 24–72 h.S. aureus. All potential staphylococcal isolates were then fully identified using a Matrix-assisted laser desorption ionization time flight mass-spectroscopy  in a positive linear mode . Samples were prepared as described previously  was used as a standard for calibration and quality control.All isolates were initially screened using conventional methods including microscopy (Gram film), catalase and coagulase testing. Prolex™ staph latex kits  were used to identify CNS and The susceptibility of 12 antibiotics was tested using standard disk diffusion method as previously described  for seven housekeeping genes, as has been described previously .http://www.ncbi.nlm.nih.gov/BLAST).Amplicons were sequenced by Eurofins MWG GmBH  using ABI 3730XL DNA analyser. Sequence similarity searches were carried out using BLAST tool  as recommended by the manufacturers . Hybridization was carried out with a DNA probe for mecA at 68°C, and using the DIG luminescence detection kit according to the manufacturer's instructions .A total of 71 staphylococcal isolates were recovered in this study, this included 62 from hotel surfaces and nine collected from hotel air samples.S. haemolyticus (n = 34), S. hominis (n = 12), S. warneri (n = 5), S. cohnii (n = 5), S. epidermidis (n = 5), S. lugdunensis (n = 3), S. pettenkoferi (n = 2), S. capitis (n = 2), S. sciuri (n = 1), S. pasteuri (n = 1), and S. equorum (n = 1).We identified 71 staphylococcal isolates as belonging to 11 different staphylococcal species. These included S. haemolyticus was predominantly resistant to erythromycin (59%), followed by S. epidermidis to amoxicillin 60% and tetracycline 60% respectively and S. cohnii to erythromycin (80%), whereas S. haemolyticus (ID 54) and S. pasteuri (ID 68) were susceptible to almost all antibiotics tested except for streptomycin  out of 71 staphylococcal isolates were resistant to two or more of the 12 antibiotics evaluated , and one isolate each harbored SCCmec type I (n = 1), type II (n = 1), and type IV (n = 1). Five isolates harbored a new SCCmec type 1A, which carried combination of class A mec complex and ccr type 1. The six isolates that were non-typeable, carried a combination of class A mec complex and ccrC  were resistant to two or more antibiotics, including one isolate was resistant to 10 antibiotics; 1–9 antibiotics, four isolates were resistant to eight antibiotics; 4–7 antibiotics; 5–6 antibiotics; 7–5 antibiotics; 13–4 antibiotics; 14–3 antibiotics and 12–2 antibiotics (Tables mec types were assigned for the 21 isolates mentioned above. Type V was the most common (7 isolates). This is consistent with some previously published works carried out with clinical isolates  (Hussain et al., S. epidermidis have been focused on clinical isolates (Li et al., S. epidermidis isolated from non-healthcare sources.Finally to emphasis the wide range of genetic variability that exists among these isolates, we also report new MLST types were assigned for five S. epidermidis isolate type with an MIC to oxacillin of 256 mg/L is of particular interest and requires further investigation with regard to its classification within the existing SCCmec typing system. We also aim in the future to look into the genetics of these isolates for other 11 antibiotics used in this study. Moreover, we have several other studies on-going which are looking at the bacterial populations in non-healthcare environments that will allow us to compare these samples with those from the current as well as previous studies. The dissemination of multidrug resistance in non-healthcare environments is evidence that infection control measures in the hospitals and in public places are ineffective in limiting the spread of such clones and that these environments are a source of antibiotic resistant pathogens.In conclusion, public environments are potential reservoirs of multidrug resistant staphylococci. The characterization of an The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


As the average age of the HIV-positive population increases, there is increasing need to monitor patients for the development of comorbidities as well as for drug toxicities.We examined factors associated with the frequency of measurement of liver enzymes, renal function tests, and lipid levels among participants of the Canadian Observational Cohort (CANOC) collaboration which follows people who initiated HIV antiretroviral therapy in 2000 or later. We used zero-inflated negative binomial regression models to examine the associations of demographic and clinical characteristics with the rates of measurement during follow-up. Generalized estimating equations with a logit link were used to examine factors associated with gaps of 12 months or more between measurements.Electronic laboratory data were available for 3940 of 7718 CANOC participants. The median duration of electronic follow-up was 3.5 years. The median (interquartile) rates of tests per year were 2.76 , 2.55  and 1.42  for liver, renal and lipid parameters, respectively. In multivariable zero-inflated negative binomial regression models, individuals infected through injection drug use (IDU) were significantly less likely to have any measurements. Among participants with at least one measurement, rates of measurement of liver, renal and lipid tests were significantly lower for younger individuals and Aboriginal Peoples. Hepatitis C co-infected individuals with a history of IDU had lower rates of measurement and were at greater risk of having 12 month gaps between measurements.Hepatitis C co-infected participants infected through IDU were at increased risk of gaps in testing, despite publicly funded health care and increased risk of comorbid conditions. This should be taken into consideration in analyses examining factors associated with outcomes based on laboratory parameters. Comorbidities are of particular concern in HIV-positive populations as successful treatment of HIV with combination antiretroviral therapy (cART) and increased incident HIV infection in older individuals have led to an increase in the average age of HIV-positive populations , 2. LaboThe recommended frequency of laboratory assessment after cART initiation varies among HIV treatment guidelines –7. To moRisk factors and clinical features also influence the recommended frequency of testing within the guidelines. For instance, BHIVA recommends more frequent measurement of lipid levels for patients at high risk of cardiovascular disease , 8. SimiWe conducted this analysis to determine whether demographic and clinical characteristics were associated with rates of laboratory testing and clinically significant gaps in measurement of liver enzymes, renal function, and lipid tests among individuals who initiated cART since January 1, 2000 in Canada. The motivation for this work was primarily to determine if HIV-positive individuals were accessing services equitably within a publicly funded health care system and, secondly, to determine whether there was evidence of measurement bias that could aid in interpretation of analyses of HIV co-morbidities that rely upon routinely collected laboratory marker data.The Canadian Observational Cohort (CANOC) is a national collaboration of eight cohorts of antiretroviral-naive HIV-positive individuals initiating cART after January 1, 2000 from three Canadian provinces (British Columbia (BC), Quebec, and Ontario). The study design has been reported previously . BrieflyThe human subjects activities of CANOC have been approved by the Simon Fraser University Research Ethics Board (REB) and the University of British Columbia REB. Additionally, approval from local institutional review boards (IRBs) was granted at each participating cohort site, as follows: Providence Health Care Research Institute Office of Research Services, the Ottawa Hospital REB, University Health Network (UHN) REB, Véritas IRB, Biomedical C REB of the McGill University Heath Centre (MUHC), University of Toronto HIV REB, and Women’s College Hospital REB.Local cohorts have obtained written consent from participants with the following exceptions: HAART Observational Medical Evaluation and Research (HOMER) Cohort ; Ottawa Hospital Cohort ; UHN ; MUHC ; Maple Leaf Medical Clinic ; Electronic Antiretroviral Therapy (EARTH) Cohort .The three sets of laboratory measurements studied were (a) the liver enzymes alanine transaminase (ALT) or aspartate transaminase (AST), (b) creatinine, as a measure of renal function, and (c) at least one of the following measures of lipid levels: LDL or HDL cholesterol, triglycerides or total cholesterol. For each set of measurements, we determined the rate of measurement per year  and whether the gaps between pairs of consecutive measurements exceeded 12 months, which was considered to be a clinically important gap between measurements. To avoid inflated rates of measurement from testing during apparent hospitalization or intensive monitoring during changes in cART or other therapies (e.g. for hepatitis C virus), repeat measurements within 30 days were excluded from determination of the rates of measurement. = 200 U/L), creatinine , LDL (5.0 mmol/L), triglyceride (8.48 mmol/L), total cholesterol (7.78 mmol/L) and total cholesterol/HDL ratio (7.0).We selected potential prognostic factors for laboratory testing based on a priori knowledge of associations with frequency of clinical follow-up. We examined demographic and clinical characteristics including age, sex, province, race, HIV risk factors such as injection drug use (IDU) and men having sex with men (MSM), year of first HIV positive test, co-infection with hepatitis B or C, CD4 count and HIV viral load at cART initiation, and class of ARV in the initial cART regimen. Grade 3 or 4 elevations for each laboratory measure were defined as follows: AST  = 170 units per litre (U/L)), ALT  models with a logit link were used to examine factors associated with a 12 month gap between sequential measurements. An unstructured correlation matrix was used to account for correlation among repeat observations within individuals. Time-updated variables were used to model the associations of CD4 counts, HIV viral load and grade 3 or 4 elevations of laboratory markers with a 12 month gap between measurements. Sensitivity analyses were conducted with an 18 month gap between measurements. For variables with large amounts of missing data, separate categories were created for missing values. All analyses were conducted with SAS 9.4 .A total of 7718 participants were enrolled into CANOC as of September 2011, of whom 3940 were followed during calendar time periods when electronic laboratory data were available at their site. Clinical and demographic characteristics are described for all patients and by availability of electronic laboratory data in Table versus 2.89,3.31, and 2.90 for HCV- non-IDU participants, HCV+ non-IDUs and HCV- IDUs respectively, p < 0.0001, Table p < 0.0001). Among participants with at least one measure of ALT/AST, Aboriginal ancestry was associated with lower rates of measurement (rate ratio (RR) =0.87, 95 % CI = , p < 0.01), as was hepatitis C co-infection among participants with a history of IDU , p < 0.0001) relative to HIV mono-infected non-IDUs. Increasing age , p = 0.03) was associated with a higher rate of measurement of ALT/AST.Ninety-three percent of participants for whom electronic laboratory data were available had at least one ALT/AST measurement; the median rate of measurement of ALT/AST per year of follow-up was 2.8 . The rate of measurement differed by province, gender, age, race, HIV risk factor, baseline regimen, and IDU history and hepatitis C co-infection .Ninety percent and 84 % of participants had at least one measurement of creatinine and lipids, respectively. The median rates of measurement of creatinine and lipids per year of follow-up were 2.6 IQR 1.4, 3.4) and 1.4 , respectively. Associations of demographic and clinical variables with rates of creatinine and lipid measurements were similar to those for liver enzymes (Table .4 and 1.p < 0.0001) and older age , p < 0.0001) were associated with decreased risk of a 12 month gap between ALT/AST measurements while hepatitis C co-infected participants with a history of IDU were at an increased risk of a 12 month gap , p < 0.0001) (Table p = 0.01), and a grade 3 or 4 elevation in lipid measurements , p < 0.01) were associated with a decreased risk of gap between lipid measures, and a grade 3 or 4 elevation in ALT/AST measurement was associated with a decreased risk of a gap between ALT/AST measurements , p = 0.02). Results were similar for gaps in excess of 18 months; with the exception that Black race and Grade 3 or 4 levels were no longer significantly associated with gaps in lipid measurements (data not shown).Twenty-two percent, 18 % and 33 % of participants had at least one 12 month gap between measurements of ALT/AST, creatinine and lipid measurements, respectively. In a multivariable GEE model with a logit link, viral load suppression (<50 copies/mL) at the start of an inter-test gap , 0.70, 0.8, p < 0.00.70, 0.8, p < 0.0In this cohort of people infected with HIV from across Canada who had initiated cART since 2000, rates of laboratory test measurement differed by age, race and HIV risk factor despite access to publicly funded health care even after adjusting for rate of CD4 measurement as a surrogate of engagement in care. Persons with HIV-hepatitis C co-infection and with a history of IDU had significantly lower rates of measurement of these tests and were more likely to have a 12 month gap between measurements than participants without history of IDU whether or not they were HIV mono- or HIV-hepatitis C co-infected. However, hepatitis C co-infected participants without a history of IDU were not monitored less frequently than HIV mono-infected individuals, in concordance with clinical guidelines , 9.et al observed higher rates of liver enzyme and renal function measurement than our study , but with lower rates of lipid testing [et al [Our results are similar to those of a multi-site study from the United States of HIV-positive individuals initiating cART, where older age, lower CD4 count, an AIDS diagnosis, later year of cART initiation, boosted PI-based cART regimens and Abacavir use were associated with shorter times to both the first laboratory test and repeated laboratory tests . As in o testing . The difg [et al  includedIn our previous work examining factors associated with rates of viral load (VL) measurement among CANOC participants, geographic region, HIV risk factor, age, year of cART initiation, type of cART regimen, being in the first year of cART, AIDS defining illness and whether or not the previous VL was below the limit of detection were associated with lower rates of VL measurement and gaps in VL measurement of more than 9 months . We haveet al [et al also found that hepatitis C co-infection was associated with less treatment access for IDU, consistent with our findings that hepatitis C co-infected IDU had the lowest rates of laboratory marker measurement and were significantly more likely to have clinically important gaps in measurement than both HIV mono-infected and hepatitis C co-infected non-IDU, and even HIV mono-infected IDU.Our observations within the current study suggest that challenges in HIV care engagement among people with a history of IDU may considerably limit the ability to follow clinical guidelines for laboratory testing in this population. Previously we have shown that participants with a history of IDU in CANOC were more likely to be suboptimally engaged in HIV care , consistet al  found baThe EACS and BC CfE guidelines specifically suggest increased frequency of monitoring for liver and renal function abnormalities for those co-infected with hepatitis C , 9. As sWhen interpreting studies of liver or renal toxicity or the development of comorbid conditions, differential monitoring needs to be considered, as abnormalities are likely to be detected sooner in individuals monitored more frequently . If moniStrengths of this analysis include the size and diversity of the CANOC cohort, which captures approximately half of the HIV-positive individuals who have initiated cART since 2000 in Canada . The pubDespite a publicly funded health care setting, there were disparities in the frequency of liver, renal, and metabolic function laboratory monitoring among subpopulations of HIV-positive individuals. Liver enzymes were more commonly and routinely measured than renal function and lipid tests, and frequency of monitoring differed by age, race, HIV risk factor, and history of hepatitis C co-infection. In particular, people with hepatitis C co-infection and a history of IDU had the lowest rates of laboratory marker measurement despite being at higher risk for cirrhosis and end-stage kidney disease. Further research should evaluate the clinical impact of delayed detection of laboratory abnormalities on the development of comorbid conditions.rd Canadian Conference on HIV/AIDS Research, St. John’s, Canada, 1-4 May 2014 (Abstract P041).The data summarized in this paper were presented in part at the 23


Both gravitational loading and the forces generated by muscle contraction have direct effects on serum markers of bone metabolism. The object of this study was to examine the direct effects of a single session of resistance exercise or walking on biochemical markers of bone metabolism in participants with low bone mass.n = 50), walking group , or control group . Changes in bone-specific alkaline phosphatase (BALP), carboxy-terminal cross-linked telopeptide of type I collagen (CTX), and serum sclerostin concentrations were measured before and immediately after a single exercise intervention.A total of 150 otherwise healthy female subjects (mean age = 59.1 ± 7.1 years) diagnosed with osteoporosis or osteopenia were randomly allocated to either a resistance exercise group . In contrast, the changes in CTX concentrations from baseline were significant in the RG (P < 0.01) but not in the WG (P = 0.11), and there was a significant difference between resistance exercise and walking (P < 0.01).There was no significant change in BALP values in any of the groups. Sclerostin levels increased in the RG and WG, and there was significant difference between the WG and CG after the exercise intervention (In participants with low bone mass, resistance exercise influenced the serum concentrations of CTX, a marker of bone resorption, but walking did not.ISRCTN16329455; retrospectively registered on 05/05/2016.Current Controlled Trials  Biochemical markers of bone metabolism that reflect the cellular activity related to the formation and resorption of bone are useful in monitoring physical activity, and can help us understand the effects of exercise on bone . The speThe anabolic effect of physical exercise on osseous tissue is related to mechanical effort, although the osteogenic response may also be influenced by other factors . PhysicaBone metabolism is influenced by several local factors including the receptor activator of nuclear factor ĸ B (RANK) ligand (RANKL) and osteoprotegerin (OPG) cytokine system, inflammatory cytokines, growth factors, bone morphogenetic proteins, transforming growth factor, prostaglandins, colony stimulating factors, and interleukins , 7. RANKBone mass can be measured well by densitometry; however, it is more difficult to accurately examine bone structure and strength in live tissue. Some substances produced during bone remodeling are specific biochemical markers of bone metabolism. Products of active osteoblasts can serve as markers of bone formation; serum concentrations of these markers reflect osteoblast function during specific phases of bone formation . AlkalinWeight-bearing and resistance exercises that improve muscle strength are essential for preserving the health of the musculoskeletal system . Coordind = 0.3) [n = 50), walking group , or control group . Exclusion criteria included (1) any condition influencing calcium and bone metabolism , (2) ongoing hormone replacement therapy, (3) any known endocrine/metabolic, renal, or hepatic disease , (4) any physical injury  hindering the performance of physical activity, (5) osseous fracture of any origin during the previous 6 months, (6) a diagnosis of cardiovascular disease or uncontrolled hypertension, (7) any non-antibiotic medication within the past year, including steroids of any type, thyroid hormones, diuretics, or anticoagulants, and (8) any antibiotic use within the last 6 months.A total of 150 female subjects diagnosed with osteoporosis/osteopenia (mean age: 58.5 ± 7.5 years) were included in this study. Untreated subjects with recently diagnosed osteoporosis/osteopenia were recruited through a large-scale screening program promoted by the Department of Rheumatology at the Zala County Hospital, Hungary, and implemented through a network of family doctors. The needed sample size was calculated based on expected effect size and study power. There are previous studies in similar contexts with similar interventions which enable exact estimations (d = 0.3) , 28. ThuEthical approval for this study was issued by the Regional Committee of Science and Research Ethics and the Policy Administration Service of Public Health of Zala County. All participants received written and verbal information regarding the purpose and procedures of the study, and all gave their written informed consent before participation. After the study, appropriate antiporotic medication was provided to the participants as required.The mean age of the participants in the RG was 60.2 ± 6.9 years. Their exercise session included 8 min of dynamic warm-up consisting of exercises requiring movements of large muscle groups and major joints that incorporated small impacts with the ground  . The maiThe mean age of participants in the WG was 58.7 ± 6.3 years. Their exercise session consisted of moderate intensity brisk walking  to a rhythm (100 steps/min) provided by a metronome. In this rhythm, they walked continuously for 46 min outdoors on even ground (wearing training shoes). The subjects familiarized themselves with the walking style prior to participating in the study session.The mean age of participants in the control group was 57.8 ± 8.4 years. These women did not receive any intervention.Serum samples were collected and concentrations of markers of bone metabolism were measured at baseline and immediately after the interventions. Several assistants performed sample collection simultaneously to ensure rapid collection within 0–5 min after the exercise sessions. All baseline blood samples were collected between 8:00 and 8:30 AM to minimize circadian effects. Prior to the interventions the participants refrained from exercising for 24 h and fasted overnight. BALP was measured after lectin precipitation using a photometric assay. Test was performed on an Olympus AU 480 Chemistry Analyzer . CTX was measured by an electrochemiluminescence immunoassay , 30. TheParticipants completed a physiotherapist-administered questionnaire specifically designed to collect demographic data and medical history for this study. Previous and existing diseases , medications and supplements (such as Vitamin D), lifestyle factors , and current physical activity level were determined using the international physical activity questionnaire (IPAQ) . The heiAll statistical analyses were performed using SPSS . Results are expressed as means and standard deviations. Data were checked for normality of distribution by the Kolmogorov-Smirnov test. Levene’s test was used to test homoscedasticity. For the clinical parameters, paired t-tests were used to determine the significance of differences in continuous variables. A two (time) by three (intervention) analysis of variance (ANOVA) with repeated measures was conducted to explore between- and within-group differences and Scheffe post hoc analysis was used to determine the significance of all possible comparisons. The α-level was set at 0.05 for all analyses.P = 0.31).Participant characteristics are presented in Table P = 0.21). The RG and CG both experienced only a non-significant rise from baseline in serum sclerostin concentrations, while the WG experienced a significant increase in serum sclerostin. Repeated measures analysis revealed significant group x time interactions (P < 0.001). Paired sample t-tests revealed a significant change in CTX concentrations from baseline only in the RG, and repeated measures ANOVA revealed a significant main effect of the type of the physical activity (P < 0.01). We compared the osteoporotic and osteopenic participants to determine if there were any differences in the CTX changes from baseline after our interventions; RG participants who had osteoporosis experienced a significant decrease in CTX concentrations post-intervention (P < 0.01), but CTX concentrations did not change in participants in the WG or CG (P > 0.05).Baseline and post-intervention serum BALP, CTX, and sclerostin concentrations for the RG, WG, and CG are shown in Table Elevated levels of CTX indicate increased bone resorption and excessive bone turnover . In our Previous studies have reported the short-term effects of different physical activities, including walking , 38, outPrevious studies have reported that serum BALP concentrations increased , 28, 47,Normal sclerostin concentrations vary with age and gender. In premenopausal and postmenopausal women normal serum sclerostin concentrations are 24.6 ± 5.7 pmol/L and 30.3 ± 8.8 pmol/L, respectively. In this study, the mean overall sclerostin concentration before exercise was 24.8 ± 11.5 pmol/L. Serum sclerostin levels increase with age in both genders, probably contributing to the age-dependent decrease in bone formation .Situations associated with bone resorption are accompanied by increases in serum sclerostin concentrations. Ardawi et al. demonstrated that even a small increase in physical loading induced a significant decrease in serum sclerostin concentrations and a concomitant increase in markers of bone formation . Spatz eThe strength of the present study was that it examined the direct effects of a single resistance exercise or walking session on sclerostin and biochemical markers of bone metabolism. Its further positive features include the homogeneity of the groups, as well as the careful assessment and exclusion of underlying diseases, medications, hormone replacement therapies, and other confounding circumstances. However, our study also had a number of limitations. It analyzed only the short-term effects of the exercises. Participants had wide age and T-score ranges, both of which have been reported to influence bone turnover . The intOur findings provide insight into the direct effects of resistance exercises and walking on biochemical markers of bone metabolism in older adults with low bone mass. Physical activity of appropriate type, duration, and intensity can influence bone turnover, causing a detectable change in serum concentrations of biochemical markers of bone metabolism and in serum sclerostin levels. The results of the study are consistent with previous reports in the literature indicating that the forces generated by muscle contraction play an important role in stimulating bone resorption, which is evident in the change in CTX concentration after exercise in the RG group. Further studies may uncover the long-term effect of resistance training on sclerostin and Wnt/β-catenin pathway.25-OH-D, 25-hydroxy vitamin D; ANOVA, analysis of variance; BALP, bone-specific alkaline phosphatase; BMI, body mass index; BMU, bone multicellular unit; CG, control group; CTX, carboxy-terminal cross-linked telopeptide of type I collagen; DEXA, dual energy x-ray absorptiometry; IPAQ, international physical activity questionnaire; NTX, amino-terminal cross-linked telopeptide of type I collagen; OPG, osteoprotegerin; PTH, parathyroid hormone; RANK, receptor activator of nuclear factor ĸ B; RG, resistance exercise group; WG, walking group