The microbiome and cancer.

Humans coexist with a vast bacterial, fungal and viral microbiome with which we have coevolved for millions of years. Several long recognized epidemiological associations between particular bacteria and cancer are now understood at the molecular level. At the same time, the arrival of next-generation sequencing technology has permitted a thorough exploration of microbiomes such as that of the human gut, enabling observation of taxonomic and metabolomic relationships between the microbiome and cancer. These studies have revealed causal mechanisms for both microbes within tumours and microbes in other host niches separated from tumours, mediated through direct and immunological mechanisms. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

COMPARATIVE BIOAVAILABILITY ANALYSIS OF ORAL ALENDRONATE SODIUM FORMULATIONS IN PAKISTAN.

Alendronate sodium, a bisphosphonate drug, it is used to treat osteoporosis and other bone diseases. The present study was designed to conduct comparative bioavailability analysis of oral formulations of aledronate sodium through an open-label, randomized, 2-sequence, 2-period crossover study. Healthy adult male Pakistani volunteers received a single 70 mg dose of the test or reference formulation of alendronate sodium followed by a 7 day washout period. Plasma drug concentrations were determined using a validated HPLC post column fluorescence derivatization method. AUC(01,) AUC(0-8,) C(max). and T(max) were determined by non-compartmental analysis and were found within the permitted range of 80% to 125% set by the US Food and Drug Administration (FDA). Results show that both in vitio and in vivo assays of all test brands were within the spec- ification of the US Pharmacopoeial limits and were statistically bioequivalent. No adverse events were reported in this study.

Characteristics and outcomes of trauma patients with ICU lengths of stay 30 days and greater: a seven-year retrospective study.

INTRODUCTION: Prolonged intensive care unit lengths of stay (ICU LOS) for critical illness can have acceptable mortality rates and quality of life despite significant costs. Only a few studies have specifically addressed prolonged ICU LOS after trauma. Our goals were to examine characteristics and outcomes of trauma patients with LOS >or= 30 days, predictors of prolonged stay and mortality. METHODS: All trauma ICU admissions over a seven-year period in a level 1 trauma center were analyzed. Admission characteristics, pre-existing conditions and acquired complications in the ICU were recorded. Logistic regression was used to identify independent predictors of prolonged LOS and predictors of mortality among those with prolonged LOS after univariate analyses. RESULTS: Of 4920 ICU admissions, 205 (4%) had ICU LOS >30 days. These patients were older and more severely injured. Age and injury severity score (ISS) were associated with prolonged LOS. After logistic regression analysis, sepsis, acute respiratory distress syndrome, and several infectious complications were important independent predictors of prolonged LOS. Within the group with ICU LOS >30 days, predictors of mortality were age, pre-existing renal disease as well as the development of renal failure requiring dialysis. Overall mortality was 12%. CONCLUSIONS: The majority of patients with ICU LOS >or= 30 days will survive their hospitalization. Infectious and pulmonary complications were predictors of prolonged stay. Further efforts targeting prevention of these complications are warranted.

Comparison of outcomes using 2 delivery models of anticoagulation care.

BACKGROUND: Studies demonstrate the effectiveness of anticoagulation management service (AMS) in providing antithrombotic therapy for eligible patients. We sought to extend this concept by determining whether an interim telephone model (IT) is comparable to our current AMS model at achieving optimal therapeutic outcomes. METHODS: The 36-month trial (24-month study plus 12-month extension) enrolled 192 eligible patients receiving long-term warfarin therapy at a Veterans Affairs hospital. Consenting participants were randomly assigned to either our current face-to-face clinic model (AMS), or our IT model. The primary outcome was the percentage of time individuals' international normalized ratios (INRs) were maintained within their target INR range (2.0-3.0 or 2.5-3.5). Secondary outcomes included the number of adverse events (eg, thromboembolism or hemorrhage) experienced during the study. RESULTS: We found no statistically significant difference between the 2 groups in the percentage of time maintained within INR target range overall (55.1% for AMS; 57.8% for IT; P = .28) nor over the course of the study. There were no statistically significant differences in the rate of thromboembolic or serious bleeding events between IT and AMS participants. Nevertheless, we did note differences related to intensity of anticoagulation. The IT group receiving treatment at a higher intensity (INR, 2.5-3.5) experienced greater anticoagulation control (P = .04) and fewer complications than the AMS group. The IT participants, however, reported a significantly higher rate of minor bleeding events, experienced mainly by those at an INR range of 2.0 to 3.0. CONCLUSION: Our IT model is a viable modification of our AMS model for the management of patients undergoing chronic anticoagulant therapy.

Determinants of vitamin D levels in nursing home residents.

OBJECTIVE: To determine the effect of various doses of vitamin D(2) and D(3), as well as ambulatory status (a surrogate for sun exposure), on 25-OH-D levels. DESIGN: Cross-sectional study with multiple regression analysis. SETTING: A state veterans home for veterans and their spouses. PARTICIPANTS: Three hundred two of 609 eligible residents. MEASUREMENTS: Serum 25-OH-D and parathyroid hormone (PTH) level, supplemental dose of vitamins D(2) and D(3) per kilogram of body weight, and 3 levels of ambulatory status. RESULTS: The mean 25-OH-D level was 28.6 + 9.2 ng/mL; 6.6% of subjects had values of 16 ng/mL or below. Thirty-two percent of participants had 25-OH-D levels below 30 ng/mL and PTH elevation based on stage of kidney disease, evidence that the suboptimal 25-OH-D level had physiologic consequences. Residents unable to transfer independently had 25-OH-D levels 1.6 ng/mL lower than those able to transfer independently. A regression analysis performed in residents unable to transfer independently (less likely to be exposed to the sun) demonstrated that the average increase in 25-OH-D level per 100 IU of D(3) in a 70-kg resident was 2.1 ng/mL versus 1.8 ng/mL for vitamin D(2). CONCLUSION: Nursing home residents should receive at least 800-1000 IU of D(3) per day in an effort to maintain optimal vitamin D levels.

Clustering of multiple endemic strains of methicillin-resistant Staphylococcus aureus in a nursing home: an 8-year study.

PFGE was performed on residents' first clinical MRSA isolate (n=94) during 8 years. Sixty-one percent of the isolates were clustered in time (P < .05) and space (P < .05) (i.e., 2 separate statistically significant tests). Isolates from individual units were genetically related, with only the occasional unrelated isolate.

Updated patterns of failure after multimodality therapy for malignant pleural mesothelioma.

OBJECTIVE: We have previously described patterns of failure after extrapleural pneumonectomy and multimodality therapy for malignant pleural mesothelioma and sought to update our results with a larger cohort of recent patients. METHODS: A total of 169 patients underwent extrapleural pneumonectomy without preoperative chemotherapy between 2001 and 2010. Data for treatment, recurrence, and survival were determined from medical records. A thoracic radiologist reviewed postoperative computed tomography or positron emission tomography computed tomography scans to determine sites of recurrence. Time to recurrence was estimated by the Kaplan-Meier method. Rates were compared using the Fisher exact test. RESULTS: The median age of patients was 62 years. Histology on final pathology was epithelial for 104 patients (62%) and nonepithelial for 65 patients (38%). A total of 132 patients (78%) received heated intraoperative chemotherapy; 77 patients (45%) received adjuvant chemotherapy, and 71 patients (42%) received adjuvant radiation therapy. Most chemotherapy regimens included platinum or pemetrexed. Median radiation therapy dose was 54 Gy. Among 158 evaluable patients, a recurrence developed in 118 (75%). Median follow-up was 83 months, median time to recurrence was 13.1 months, and median survival was 15 months. Sites of first recurrence were in the ipsilateral hemithorax or mediastinum for 54% of patients, in the abdomen for 39% of patients, in the contralateral hemithorax for 28% of patients, and in other distant sites for 5% of patients. Some patients had simultaneous recurrences in multiple sites. CONCLUSIONS: The most common site of recurrence after extrapleural pneumonectomy and planned multimodality therapy remains the ipsilateral hemithorax (including mediastinum), and true distant failure (other than the abdomen or contralateral hemithorax) remains unusual. The distribution of recurrences is strikingly similar to our prior report.

Anatomic determinants of sleep-disordered breathing across the spectrum of clinical and nonclinical male subjects.

OBJECTIVES: We wished to determine the independent contribution of craniofacial dimensions of the upper airway to sleep-disordered breathing (SDB) in subjects who spanned the entire continuum of SDB. We also determined the interactive effects of body mass index (BMI) and age on the relationship between airway dimensions and SDB. DESIGN AND SUBJECTS: We studied 142 nonclinical male subjects in a working community population (average age, 47 years; average BMI, 29; average +/- SD apnea/hypopnea index [AHI], 20 +/- 20/h), and 62 patients with obstructive sleep apnea (average age, 47 years; average BMI, 32; average +/- SD AHI, 48 +/- 35/h. We determined the AHI from overnight polysomnography and the number of oxygen desaturations (> or = 2%) per hour of sleep. We used lateral facial cephalometric radiographs to measure 41 anatomic landmarks and 55 dimensions in the upper airway. SETTING: A university hospital and a sleep-disorders clinic. DATA ANALYSIS: We used stepwise regression analysis to determine the independent contributions of measured variables to SDB. MEASUREMENTS AND RESULTS: In the entire study population (n = 204), variations in BMI and six measures of craniofacial morphology accounted equally for one half of the total variance in AHI, and their interactive effects accounted for an additional 15%. Membership in the clinical or nonclinical group per se had no significant influence on these relationships. The single most important cephalometric variable in predicting AHI severity was the horizontal dimension of the maxilla (ie, porion vertical to supradentale [PV-A] distance). When the PV-A distance was relatively narrow (< 97 mm) the probability of having mild (AHI, 15 to 30/h) to severe (AHI > 30/h) SDB increased fivefold to sevenfold in nonobese subjects and threefold in obese subjects. Thus, in nonobese subjects (average BMI, 25 +/- 2) and in subjects with narrow upper airway dimensions, four cephalometric dimensions were the dominant predictors of AHI, accounting for 50% of the variance. However, in subjects with a large anteroposterior facial dimension, BMI was the major predictor of AHI and a BMI > 28 increased the probability of moderate-to-severe sleep apnea by approximately fivefold. Finally, the combination of cephalometric dimensions and BMI accounted for an increasing amount of the variance in AHI as the severity of AHI increased. CONCLUSIONS: Across the population spectrum of SDB, four cephalometric dimensions of the upper airway in combination with BMI accounted independently for up to two thirds of the variation in AHI; and the relative contribution of these two sets of determinants of AHI varied depending on airway size, obesity, and the amount of SDB.

Risk of acquiring influenza A in a nursing home from a culture-positive roommate.

Influenza A was cultured in 62 double rooms. The roommate was infected in 12 (19.4%). During 3,294 resident-seasons, influenza was cultured in 208 single rooms (6.3%). Those who lived in double rooms with a culture-positive roommate had a 3.07 relative risk (CI95, 1.61-5.78) of acquiring influenza.

The effect of culture-positive influenza type A on resource use and adverse events in nursing home residents.

OBJECTIVES: To determine adverse clinical events and resource utilization associated with culture-positive influenza A in nursing home residents. DESIGN: A retrospective cohort study with cases and controls. SETTING: Seven hundred twenty-one-bed skilled nursing facility. PARTICIPANTS: One hundred fifty-four residents (21% of all residents) from whom influenza A was isolated during the 1997/98 season and matched controls. MEASUREMENTS: Baseline parameters, staff interventions, diagnostic tests, and adverse events were recorded from 60 days before to 60 days after specimen collection. The difference between each individual's before and after measurements determined excess utilization secondary to influenza. Controls were studied to determine time series effects. RESULTS: Baseline Minimum Data Set and nutritional parameters demonstrated significantly greater (P <.05) feeding dependency and lower serum albumin in the control group. Time series effects in the control group were negligible. Among cases, there were nine deaths within 30 days; among controls, there were four (chi2 P =.26). Within 30 days of onset, an average excess of 18 notations by nursing staff, one phone call to the physician, and one to family was noted per case. In half of cases, a nonscheduled physician visit was required. There was a 20% excess in physician orders for oxygen and bronchodilators. Chest x-rays were performed in half of the cases, and antibiotics were prescribed to half. Sixteen percent of cases had radiographic pneumonia, and 2% had congestive heart failure. The average cost for excess chest x-rays, laboratory services, antimicrobials, ambulance calls, hospital days, and emergency room and physician visits was $943.44. This does not include efforts by nursing home staff who accommodate functional decline on-site. CONCLUSION: An unexpected finding was that there were more impaired individuals who were less likely to have influenza detected or less likely to acquire influenza in the control group than in the influenza group. The morbidity, mortality, excess staff effort, and measured expenditure justify efforts to prevent influenza.

Delays in the application of outbreak control prophylaxis for influenza A in a nursing home.

OBJECTIVE: To identify delayed prophylaxis from a pre-existing database and strategies to improve performance. SETTING: A skilled nursing facility with 14 floors (4 buildings). The "outbreak unit" was a 50- to 60-bed floor. METHODS: We performed surveillance during six seasons using one protocol. Prophylaxis was started when influenza was cultured in the building and 10% of residents on the floor had a new respiratory illness within 7 days. We defined delayed prophylaxis as four or more residents on a floor with positive cultures whose specimens had been collected within 5 days before the application of prophylaxis. RESULTS: We identified 14 examples of delayed prophylaxis. In three, delayed prophylaxis was related to the 3.9-day delay between culture collection and culture report There was a high degree of commonality among building attack rates within a season. During six seasons, the first case in the last building occurred 27 to 64 days after the first case in the facility. The two seasons with the greatest activity (68 and 154 cases, respectively) began with explosive, multi-floor outbreaks in a single building. The match between the circulating strain and the vaccine was good, except in 1997-1998 when there were seven examples of delayed prophylaxis. CONCLUSIONS: Influenza may involve buildings sequentially with a commonality of building attack rates. Explosive, multi-floor outbreaks early in the season could lead to a lower threshold for prophylaxis within a larger area when initial cases are encountered later in the season. This strategy could have prevented five examples of delayed prophylaxis. Rapid testing of multiple specimens while waiting for culture confirmation could have prevented three examples of delayed prophylaxis.

A comparison of previous antibiotic therapy following isolation of MRSA versus MSSA in nursing home residents: a preliminary investigation.

OBJECTIVE: Antibiotic exposure is a risk factor for isolation of methicillin-resistant Staphylococcus aureus (MRSA). We reasoned that this relationship might be especially strong if the comparison was targeted to antibiotics with activity against methicillin-sensitive Staphylococcus aureus (MSSA), but not MRSA. SUBJECTS: Forty-eight cases with initial MRSA clinical isolates (15 urinary, 19 wound, 11 sputum, 3 blood) were matched to 48 with MSSA clinical isolates from the same location. In addition, 10 whose MRSA was noted screening around an index case were matched to 10 with negative screening cultures sampled around the same index case. DESIGN: MSSA was always/usually sensitive to dicloxacillin, amoxicillin/clavulanate, cephalosporins, imipenem, quinolones, and erythromycin, whereas MRSA was always/usually resistant. These antibiotics were defined as "targeted" and would be expected to facilitate the proliferation of MRSA by suppressing competing bacteria. We compared the use of all antibiotics over 60 days in cases and controls. RESULTS: The comparison of antibiotic exposure in the 48 clinical isolates revealed that 29 (60%) with MRSA isolates versus 16 (33%) with MSSA isolates had received a targeted antibiotic (RR 1.81, CI 1.33-3.34, P = 0.006). Twelve (25%) with MRSA clinical isolates versus 3 with MSSA isolates (6%) had received a nontargeted antibiotic (RR 4.00, CI 2.10-14.18, P = 0.009). The comparison of antibiotic exposure in MRSA screening isolates versus negative controls revealed that 7 (70%) had received a targeted antibiotic versus 2 (20%) of controls (RR 3.50, CI 1.57-16.85, P = 0.009)). One in the screening group versus 2 controls had received a nontargeted antibiotic. Twenty-one (62%) of 34 with MRSA (urine or wound) isolates versus 4 MSSA controls (12%) had received a quinolone (RR 5.25, CI 1.80-15.30, P < 0.000). CONCLUSION: There was a significant association between antibiotic exposure and MRSA isolates. The association was especially strong for quinolones with urinary or wound isolation of MRSA. Our data do not support the hypothesis that targeted antibiotic use was more likely to be associated with MRSA isolation than nontargeted antibiotic use. The use of nontargeted antibiotics was low, with greater use in the MRSA clinical group.

Engineering defined motor ensembles with DNA origami.

Many cytoskeletal motors function in groups to coordinate the spatial and temporal positioning of cellular cargo. While methods to study the biophysical properties of single motors are well established, methods to understand how multiple motors work synergistically or antagonistically are less well developed. Here, we describe a three-dimensional synthetic cargo structure made using DNA origami, which can be used to template defined numbers and types of cytoskeletal motors with programmable geometries and spacing. We describe methods for building the DNA origami structure, covalently attaching motors to DNA, forming the motor-DNA origami structure complex, and single-molecule assays to examine the motile properties of motor ensembles.

Reconstitution of dynein transport to the microtubule plus end by kinesin.

Cytoplasmic dynein powers intracellular movement of cargo toward the microtubule minus end. The first step in a variety of dynein transport events is the targeting of dynein to the dynamic microtubule plus end, but the molecular mechanism underlying this spatial regulation is not understood. Here, we reconstitute dynein plus-end transport using purified proteins from S. cerevisiae and dissect the mechanism using single-molecule microscopy. We find that two proteins-homologs of Lis1 and Clip170-are sufficient to couple dynein to Kip2, a plus-end-directed kinesin. Dynein is transported to the plus end by Kip2, but is not a passive passenger, resisting its own plus-end-directed motion. Two microtubule-associated proteins, homologs of Clip170 and EB1, act as processivity factors for Kip2, helping it overcome dynein's intrinsic minus-end-directed motility. This reveals how a minimal system of proteins transports a molecular motor to the start of its track.DOI: http://dx.doi.org/10.7554/eLife.02641.001.

Lis1 regulates dynein by sterically blocking its mechanochemical cycle.

Regulation of cytoplasmic dynein's motor activity is essential for diverse eukaryotic functions, including cell division, intracellular transport, and brain development. The dynein regulator Lis1 is known to keep dynein bound to microtubules; however, how this is accomplished mechanistically remains unknown. We have used three-dimensional electron microscopy, single-molecule imaging, biochemistry, and in vivo assays to help establish this mechanism. The three-dimensional structure of the dynein-Lis1 complex shows that binding of Lis1 to dynein's AAA+ ring sterically prevents dynein's main mechanical element, the 'linker', from completing its normal conformational cycle. Single-molecule experiments show that eliminating this block by shortening the linker to a point where it can physically bypass Lis1 renders single dynein motors insensitive to regulation by Lis1. Our data reveal that Lis1 keeps dynein in a persistent microtubule-bound state by directly blocking the progression of its mechanochemical cycle.

Engineered, harnessed, and hijacked: synthetic uses for cytoskeletal systems.

Synthetic biology re-imagines existing biological systems by designing and constructing new biological parts, devices, and systems. In the arena of cytoskeleton-based transport, synthetic approaches are currently used in two broad ways. First, molecular motors are harnessed for non-physiological functions in cells. Second, transport systems are engineered in vitro to determine the biophysical rules that govern motility. These rules are then applied to synthetic nanotechnological systems. We review recent advances in both of these areas and conclude by discussing future directions in engineering the cytoskeleton and its motors for transport.

Dynein achieves processive motion using both stochastic and coordinated stepping.

Processivity, the ability of single molecules to move continuously along a track, is a fundamental requirement of cargo-transporting molecular motors. Here, we investigate how cytoplasmic dynein, a homodimeric, microtubule-based motor, achieves processive motion. To do this, we developed a versatile method for assembling Saccharomyces cerevisiae dynein heterodimers, using complementary DNA oligonucleotides covalently linked to dynein monomers labeled with different organic fluorophores. Using two-color, single-molecule microscopy and high-precision, two-dimensional tracking, we find that dynein has a highly variable stepping pattern that is distinct from all other processive cytoskeletal motors, which use 'hand-over-hand' mechanisms. Uniquely, dynein stepping is stochastic when its two motor domains are close together. However, coordination emerges as the distance between motor domains increases, implying that a tension-based mechanism governs these steps. This plasticity may allow tuning of dynein for its diverse cellular functions.