Multimodal surveillance of SARS-CoV-2 at a university enables development of a robust outbreak response framework.

BACKGROUND: Universities are vulnerable to infectious disease outbreaks, making them ideal environments to study transmission dynamics and evaluate mitigation and surveillance measures. Here, we analyze multimodal COVID-19-associated data collected during the 2020-2021 academic year at Colorado Mesa University and introduce a SARS-CoV-2 surveillance and response framework. METHODS: We analyzed epidemiological and sociobehavioral data (demographics, contact tracing, and WiFi-based co-location data) alongside pathogen surveillance data (wastewater and diagnostic testing, and viral genomic sequencing of wastewater and clinical specimens) to characterize outbreak dynamics and inform policy. We applied relative risk, multiple linear regression, and social network assortativity to identify attributes or behaviors associated with contracting SARS-CoV-2. To characterize SARS-CoV-2 transmission, we used viral sequencing, phylogenomic tools, and functional assays. FINDINGS: Athletes, particularly those on high-contact teams, had the highest risk of testing positive. On average, individuals who tested positive had more contacts and longer interaction durations than individuals who never tested positive. The distribution of contacts per individual was overdispersed, although not as overdispersed as the distribution of phylogenomic descendants. Corroboration via technical replicates was essential for identification of wastewater mutations. CONCLUSIONS: Based on our findings, we formulate a framework that combines tools into an integrated disease surveillance program that can be implemented in other congregate settings with limited resources. FUNDING: This work was supported by the National Science Foundation, the Hertz Foundation, the National Institutes of Health, the Centers for Disease Control and Prevention, the Massachusetts Consortium on Pathogen Readiness, the Howard Hughes Medical Institute, the Flu Lab, and the Audacious Project.

Multimodal surveillance of SARS-CoV-2 at a university enables development of a robust outbreak response framework

Universities are particularly vulnerable to infectious disease outbreaks and are also ideal environments to study transmission dynamics and evaluate mitigation and surveillance measures when outbreaks occur. Here, we introduce a SARS-CoV-2 surveillance and response framework based on high-resolution, multimodal data collected during the 2020-2021 academic year at Colorado Mesa University. We analyzed epidemiological and sociobehavioral data (demographics, contact tracing, and wifi-based co-location data) alongside pathogen surveillance data (wastewater, random, and reflexive diagnostic testing; and viral genomic sequencing of wastewater and clinical specimens) to characterize outbreak dynamics and inform policy decisions. We quantified group attributes that increased disease risk, and highlighted parallels between traditional and wifi-based contact tracing. We additionally used clinical and environmental viral sequencing to identify cryptic transmission, cluster overdispersion, and novel lineages or mutations. Ultimately, we used distinct data types to identify information that may help shape institutional policy and to develop a model of pathogen surveillance suitable for the future of outbreak preparedness.

Gravitational lensing reveals ionizing ultraviolet photons escaping from a distant galaxy.

During the epoch of reionization, neutral gas in the early Universe was ionized by hard ultraviolet radiation emitted by young stars in the first galaxies. To do so, ionizing ultraviolet photons must escape from the host galaxy. We present Hubble Space Telescope observations of the gravitationally lensed post-reionization galaxy PSZ1-ARC G311.6602-18.4624 (nicknamed the "Sunburst Arc"), revealing bright, multiply imaged ionizing photon escape from a compact star-forming region through a narrow channel in an optically thick gas. The gravitational lensing magnification shows how ionizing photons escape this galaxy, contributing to the reionization of the Universe. The multiple sight lines to the source probe absorption by intergalactic neutral hydrogen on a scale of less than a few hundred parsecs.

Clinicopathological characteristics and non-adhesive organ culture of insulinomas.

BACKGROUND AND AIMS: Insulinoma is a very rare type of islet cell tumour, but nevertheless the most common endocrine tumour of the pancreas. We aimed at reviewing our clinical experience with this tumour type and to assess whether organ culture could be obtained from surgically resected insulinoma material. MATERIAL AND METHODS: All patients with insulinomas (6 men and 10 women) referred to Haukeland University Hospital between 1986 and 2006 were included in the study. Median age of onset was 53 years (range 21-74). Biochemical diagnosis was established during a 72 h fast test. Imaging and localization of the tumours were performed with intra-operative ultrasonography, endoscopic ultrasonography, CT-scan and/or transcutaneous ultrasonography. For six patients, organ cultures were set up from tumour tissue fragments. RESULTS: The annual incidence of insulinoma was 0.8 per million. The patients generally presented with non-specific, episodic symptoms, which often were mistaken for cardiovascular, neurological or diabetic disease and in some cases delayed the diagnosis with several years. Two patients had diabetes prior to the diagnosis of insulinoma. Patient weight gain was probably due to increased food intake, compensating for the hypoglycemia. Intra-operative ultrasonography detected all tumours correctly, whereas 73% were detected by endoscopic ultrasonography and 38% by CT scan. Five insulinomas were located in the head, eight in the body and three in the tail of the pancreas. All were removed by open-access surgery, eleven cases by resection and five by enucleation. One tumour was malignant with liver metastases and two patients had tumours defined as borderline. Insulinoma tissue fragments developed into spheroids during the first week of culturing and insulin secretion into the media was demonstrated. CONCLUSIONS: Insulinomas are rare and diagnostically challenging tumours. Intra-operative ultrasonography was superior to other imaging modalities to locate the lesion. In organ culture, insulinomas readily form spheroids which may be used to yield insight into beta-cell biology.

Saliva cortisol measurement: simple and reliable assessment of the glucocorticoid replacement therapy in Addison's disease.

No ideal parameter is available for assessment of the glucocorticoid replacement therapy in Addison's disease. Serum cortisol day-curves can be used to monitor the therapy, but this technique is cumbersome and expensive. We evaluated the potential for saliva cortisol measurement in this setting. We found excellent correlation between serum and saliva cortisol after oral intake of cortisone acetate (no. 7) or iv administration of hydrocortisone (no. 4) (Pearson's R=0.83-0.98, p<0.002). A morning dose of 12.5 mg cortisone acetate yielded wide interindividual variations in cortisol levels in saliva. Saliva cortisol measurements were successfully adopted to evaluate and adjust doses in outpatients. We conclude that cortisol measurement in saliva is practical and reliable, and is preferable to serum cortisol measurement in the assessment of the glucocorticoid replacement therapy. Our results confirm that only a minority of patients require more than 12.5 mg of cortisone acetate (equivalent to 10 mg hydrocortisone) in the morning to have sufficient cortisol levels during the first part of the day.

Predictive value of tumour cell proliferation in locally advanced breast cancer treated with neoadjuvant chemotherapy.

We previously reported that defects in apoptotic pathways (mutations in the TP53 gene) predicted resistance to doxorubicin monotherapy. The aim of this study was to evaluate whether cell proliferation, as assessed by mitotic frequency and Ki-67 levels, may provide additional predictive information in the same tumours and to assess any potential correlations between these markers and mutations in the TP53 gene and erbB-2 overexpression. Surgical specimens were obtained from ninety locally advanced breast cancers before commencing primary chemotherapy consisting of weekly doxorubicin (14 mg/m2) for 16 weeks. 38% of the patients had a partial response (PR) to therapy, 52% had stable disease (SD) while 10% had progressive disease (PD). Univariate analysis showed a significant association between a high cell proliferation rate (expressed as a high mitotic frequency) and resistance to doxorubicin (P = 0.001). Further analyses revealed this association to be limited to the subgroup of tumour expressing wild-type TP53 (P = 0.016), and TP53 mutation status was the only factor predicting drug resistance in the multivariate analyses. The finding that a high mitotic frequency, as well as a high Ki-67 staining, correlated to TP53 mutations (P = 0.001 for both), suggests TP53 mutations are the key predictor of drug resistance, although cell proliferation may play an additional role in tumours harbouring wild-type TP53. Regarding overall (OS) and relapse-free survival (RFS), multivariate analyses (Cox' proportional hazards regression) revealed a high histological grade and negative oestrogen receptor (ER) status to be the variables that were most strongly related to breast cancer death (P = 0.001 and P = 0.001, respectively). A key reason for this difference with respect to the factors predicting chemotherapy resistance could be due to the adjuvant use of tamoxifen in all patients harbouring ER-positive tumours.

Premorbid body weight and its relations to primary tumour diameter in breast cancer patients; its dependence on estrogen and progesteron receptor status.

Hormonal mechanisms have been offered as an explanation for the higher frequency of large tumours, lymph node metastases and poorer prognosis in obese breast cancer patients than in lean ones. If hormonal mechanisms are important for these relations, they should probably act more strongly in patients with hormonal receptor positive tumours than in those with negative ones. We have examined if the relations between premorbid body weight or Quetelet's index (weight/height2) and tumour diameter are modified by estrogen receptor alpha (ER) and progesteron receptor (PgR) status. The analyses were based on 1,241 women with unilateral disease treated with modified radical mastectomy living in the geografic area of Haukeland Hospital. Their body weight and height have been measured as a mean 12.5 years before presentation of the disease. Body weight and Quetelet's index have been adjusted for age. The relations were studied using linear regression analyses adjusting the effect of body weight with height and mean nuclear area of the tumour cells and adjusting the effect of Quetelet's index for mean nuclear area. The main findings showed that patients with high body weight or Quetelet's index presented more often with PgR positive tumours than lean ones. Quetelet's index was also positively related to ER. These relations were present in patients older than 50 years of age (older). Patients with large tumours (>2.0 cm) had significantly higher body weight and Quetelet's index than those with small ones. These differences were significantly present in older patients and in patients with PgR negative and ER negative-PgR negative tumours. Linear regression analyses confirmed that tumour diameter increases with body weight and Quetelet's index. These relations were present in both lymph node groups and in older patients. Stratification according to hormonal receptor status showed these relations to be significant in patients with ER negative, with PgR negative and those with ER negative-PgR negative tumours only. Taking age and hormonal receptor status into consideration simultaneously, both body weight and Quetelet's index were significantly related to tumour diameter in older patients with hormone receptor negative tumours. In conclusion body size was positively related to hormone receptor status and to diameter of the primary tumour. The relation to tumour diameter was present in older patients with hormone receptor negative tumours. Although hormonal mechanisms able to act on the tumour can not be excluded, mechanisms acting independent of hormonal receptors must be considered. Different mechanisms related to body fat cytokines are discussed.

Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications.

The purpose of this study was to classify breast carcinomas based on variations in gene expression patterns derived from cDNA microarrays and to correlate tumor characteristics to clinical outcome. A total of 85 cDNA microarray experiments representing 78 cancers, three fibroadenomas, and four normal breast tissues were analyzed by hierarchical clustering. As reported previously, the cancers could be classified into a basal epithelial-like group, an ERBB2-overexpressing group and a normal breast-like group based on variations in gene expression. A novel finding was that the previously characterized luminal epithelial/estrogen receptor-positive group could be divided into at least two subgroups, each with a distinctive expression profile. These subtypes proved to be reasonably robust by clustering using two different gene sets: first, a set of 456 cDNA clones previously selected to reflect intrinsic properties of the tumors and, second, a gene set that highly correlated with patient outcome. Survival analyses on a subcohort of patients with locally advanced breast cancer uniformly treated in a prospective study showed significantly different outcomes for the patients belonging to the various groups, including a poor prognosis for the basal-like subtype and a significant difference in outcome for the two estrogen receptor-positive groups.

Dynamic pattern formation in a vesicle-generating microfluidic device.

Spatiotemporal pattern formation occurs in a variety of nonequilibrium physical and chemical systems. Here we show that a microfluidic device designed to produce reverse micelles can generate complex, ordered patterns as it is continuously operated far from thermodynamic equilibrium. Flow in a microfluidic system is usually simple-viscous effects dominate and the low Reynolds number leads to laminar flow. Self-assembly of the vesicles into patterns depends on channel geometry and relative fluid pressures, enabling the production of motifs ranging from monodisperse droplets to helices and ribbons.

Monolithic microfabricated valves and pumps by multilayer soft lithography.

Soft lithography is an alternative to silicon-based micromachining that uses replica molding of nontraditional elastomeric materials to fabricate stamps and microfluidic channels. We describe here an extension to the soft lithography paradigm, multilayer soft lithography, with which devices consisting of multiple layers may be fabricated from soft materials. We used this technique to build active microfluidic systems containing on-off valves, switching valves, and pumps entirely out of elastomer. The softness of these materials allows the device areas to be reduced by more than two orders of magnitude compared with silicon-based devices. The other advantages of soft lithography, such as rapid prototyping, ease of fabrication, and biocompatibility, are retained.

[Student education in a laboratory for general practice skills]. / Studentundervisning i allmennmedisinsk ferdighetslaboratorium.

Clinical skills, like questioning and examining a patient, are developed gradually throughout the years of medical training. Basic skills should be mastered on graduation, but the teaching of skills in most medical schools is not systematic. We evaluated a pilot teaching project in a general practice (GP) skills laboratory. Students were randomised to an intervention group and a control group. Teaching was performed in small groups of one GP instructor and six students and consisted of four weekly three-hour sessions. For the control group, teaching was delayed until after evaluation. Evaluation of practical skills and communication skills was made with the students in random order by GP evaluators who did not know about the randomisation procedure. Each skill was scored as satisfactory or not satisfactory for each of five procedural elements. 56 fourth-year medical students volunteered. Evaluation was performed for 19 (68%) students in the training group and 18 (64%) in the control group. The training group performed better on practical skills, average score 9.7 (95% confidence interval 7.4 to 12.1) vs. 5.5 (3.2 to 7.8), (p = 0.01). There was no significant difference for communication skills, 7.7 (5.6 to 9.8) vs. 6.7 (4.6 to 8.7). Teaching clinical skills in a general practice skills laboratory is feasible. Practical skills may improve rapidly, at least for short-time performance, while our brief teaching in communication skills was not proved to be effective.

Changing doctor prescribing behaviour.

The aim of this overview was to identify interventions that change doctor prescribing behaviour and to derive conclusions for practice and further research. Relevant studies (indicating prescribing as a behaviour change) were located from a database of studies maintained by the Cochrane Collaboration on Effective Professional Practice. This register is kept up to date by searching the following databases for reports of relevant research: DHSS-DATA; EMBASE; MEDLINE; SIGLE; Resource Database in Continuing Medical Education (1975-1994), along with bibliographies of related topics, hand searching of key journals and personal contact with content area experts. Randomised controlled trials and non-equivalent group designs with pre- and post-intervention measures were included. Outcome measures were those used by the study authors. For each study we determined whether these were positive, negative or inconclusive. Positive studies (+) were those that demonstrated a statistically significant change in the majority of outcomes measured at level of p < or = 0.05 between the intervention and control groups. Negative studies (-) showed a significant change in the opposite direction and inconclusive studies (approximately) showed no significant change compared to control or no overall positive findings. We identified 79 eligible studies which described 96 separate interventions to change prescribing behaviour. Of these interventions, 49 (51%, 41%-61%) showed a positive significant change compared to the control group but interpretation of specific interventions is limited due to wide and overlapping confidence intervals.

Functional expression of horseradish peroxidase in E. coli by directed evolution.

In an effort to develop a bacterial expression system for horseradish peroxidase (HRP), we inserted the gene encoding HRP into the pET-22b(+) vector (Novagen) as a fusion to the signal peptide PelB. A similar construct for cytochrome c peroxidase (CcP) leads to high CcP activity in the supernatant. Expression of the wild-type HRP gene in the presence of isopropyl-beta-D-thiogalactopyranoside (IPTG) yielded no detectable activity against ABTS (azinobis(ethylbenzthiazoline sulfonate)). However, weak peroxidase activity was detected in the supernatant in the absence of IPTG. The HRP gene was subjected to directed evolution: random mutagenesis and gene recombination followed by screening in a 96-well microplate format. From 12 000 clones screened in the first generation, one was found that showed 14-fold higher HRP activity than wild-type, amounting to approximately 110 microg of HRP/L, which is similar to that reported from laborious in vitro refolding. No further improvement was obtained in subsequent generations of directed evolution. This level of expression has nonetheless enabled us to carry out further directed evolution to render the enzyme more thermostable and more resistant toward inactivation by H2O2. These results show that directed evolution can identify mutations that assist proteins to fold more efficiently in Escherichia coli. This approach will greatly facilitate efforts to "fine-tune" those many enzymes that are promising industrial biocatalysts, but for which suitable bacterial or yeast expression systems are currently lacking.

Puncture technique and postural postdural puncture headache. A randomised, double-blind study comparing transverse and parallel puncture.

BACKGROUND: This clinical study was conducted in order to investigate the effect of two different orientations of the bevel during dural puncture on development of postural postdural puncture headache (PPDPH). METHODS: Two hundred and eighteen patients aged 18 to 50 years scheduled for minor non-obstetric surgery using spinal anaesthesia (SA) were included in this randomised, double-blind study. Dural puncture was performed using a 0.42 mm O.D. (27-g) Quincke spinal needle with the orientation of the bevel parallel or transverse relative to the longitudinal axis of the dural cylinder. All patients were blinded with regard to the puncture technique, and so was the anaesthesiologist performing a telephone interview 5 to 7 days postoperatively. The occurrence and duration of headache, backache and other complaints were recorded. Headache was classified as PPDPH or non-PPDPH, and intensity of the headache was registered using a numerical rating scale (NRS) from 0 to 10. RESULTS: Two hundred and twelve patients with a mean age of 35.3 years completed the study, 106 in each group. The two groups were comparable with regard to mean age, sex, local anaesthetics used and surgical procedure performed. Headache occurred in 44 patients postoperatively. PPDPH was diagnosed in 4/106 patients (3.8%) in the parallel group and 24/106 (22.6%) in the transverse group (P < 0.0002). Postoperative backache occurred in 31 and 20 patients (parallel compared to transverse) (NS). CONCLUSIONS: Dural puncture with the bevel of the needle transverse to the longitudinal axis of the dural cylinder gave significantly more cases of PPDPH than puncture with the bevel parallel to this axis even when using a 27-g Quincke needle. When using Quincke bevelled needles care must be taken to assure that the orientation of the bevel is parallel to the longitudinal axis of the dural sac.

Ultrasound biomicroscopy. II. Intraobserver and interobserver reproducibility of measurements.

PURPOSE: To evaluate intraobserver and interobserver reproducibility of measurements in images obtained by the 50 MHz Humphrey Ultrasound Biomicroscope. METHODS: The first stored image of central cornea, central anterior chamber, and angle structures obtained at 50 examinations were measured twice by three observers. Nine different parameters were measured in angle images and two in images of central cornea and anterior chamber. Intraobserver reproducibility was assessed by calculating the coefficient of variation and interobserver reproducibility by a two-ways ANOVA. RESULTS: Intraobserver reproducibility was high for all measurements of central cornea thickness and anterior chamber depth with a coefficient of variation (%) < or = 3.8%. In angle images the intraobserver reproducibility of measurements was often lower (coefficient of variation (%) 1.3-52.6%). The lower reproducibility was most noticeable in measurements involving the less defined structures. Interobserver reproducibility was poor. CONCLUSIONS: Comparison of measurements are best done of well defined structures and by only one observer.

Modulation of mouse estrogen receptor transcription activity by protein kinase C delta.

The effect of protein kinase C (PKC) delta on the transcriptional activity of the mouse estrogen receptor was investigated. The receptor was expressed transiently in Cos-1 and NIH3T3 cells in the presence of wild-type, dominant negative or constitutively active forms of PKC delta. Transfection experiments demonstrated that PKC delta stimulated both unliganded and liganded estrogen receptor transcriptional activity. This stimulatory effect was not observed using PKC alpha or PKC epsilon. 4-Hydroxytamoxifen and the pure anti-estrogen ICI 164,384 reduced receptor transcriptional activity in the presence of PKC delta. The stimulatory effect of PKC delta on estrogen receptor transcriptional activity was mediated by the N-terminal activation function 1 (AF-1) domain. The reduced stimulatory effect of PKC delta on transcriptional activity of the phosphorylation defective mutant of estrogen receptor suggests that phosphorylation of serine 122 in the AF-1 region may mediate the modulatory effect of PKC delta. Wild-type PKC delta caused a twofold increase in estrogen receptor phosphorylation, while a dominant negative mutant of PKC delta reduced the receptor phosphorylation to five percent of that caused by wild-type PKC delta. Our results suggest that PKC delta participates in the signaling pathways that lead to estrogen receptor phosphorylation and its effect on estrogen receptor transcriptional activation is both cell type and promoter specific.

High-throughput genetic analysis using microfabricated 96-sample capillary array electrophoresis microplates.

Capillary array electrophoresis (CAE) microplates that can analyze 96 samples in less than 8 min have been produced by bonding 10-cm-diameter micromachined glass wafers to form a glass sandwich structure. The microplate has 96 sample wells and 48 separation channels with an injection unit that permits the serial analysis of two different samples on each capillary. An elastomer sheet with an 8 by 12 array of holes is placed on top of the glass sandwich structure to define the sample wells. Samples are addressed with an electrode array that makes up the third layer of the assembly. Detection of all lanes with high temporal resolution was achieved by using a laser-excited confocal fluorescence scanner. To demonstrate the functionality of these microplates, electrophoretic separation and fluorescence detection of a restriction fragment marker for the diagnosis of hereditary hemochromatosis were performed. CAE microplates will facilitate all types of high-throughput genetic analysis because their high assay speed provides a throughput that is 50 to 100 times greater than that of conventional slab gels.

Eicosapentaenoic acid and sulphur substituted fatty acid analogues inhibit the proliferation of human breast cancer cells in culture.

Numerous studies have shown dietary fatty acids to influence the progression of several types of cancers. The purpose of the present investigation was to examine the influence of various types of fatty acids, including omega-3 fatty acids and a new class of hypolipidemic peroxisome proliferating fatty acid analogues, namely the 3-thia fatty acids, on MCF-7 human breast cancer cell growth. 3-thia fatty acids represent non-beta-oxidizable fatty acid analogues in which a sulphur atom substitutes for the beta-methylene group (3-position) in the saturated and unsaturated fatty acids. The effects of increasing concentrations of palmitic acid, tetradecylthioacetic acid (a 3-thia fatty acid), eicosapentaenoic acid, docosahexaenoic acid, and two 3-thia polyunsaturated fatty acids on the proliferation of MCF-7 cells, maintained in serum-free culture, were studied. At the highest concentration of fatty acid used (64 microM) tetradecylthioacetic acid was found to be the most effective of all fatty acids tested in inhibiting cell growth, whilst palmitic acid and docosahexaenoic acid had no significant effect on cell growth. Thus, of the two dietary polyunsaturated omega-3 fatty acids eicosapentaenoic acid and docosahexaenoic acid, only eicosapentaenoic acid possesses an inhibitory effect on the proliferation of MCF-7 cells. In all cases the inhibitory effect of the fatty acid was found to be reversible. Tetradecylthioacetic acid has been shown to be a potent peroxisome proliferator. It was, therefore, hypothesized that tetradecylthioacetic acid may inhibit the human MCF-7 cell growth by increasing the level of oxidative stress within the cell. However, use of agents which modify the cell's protective apparatus against oxidative stress had no influence on the inhibitory effect of tetradecylthioacetic acid. These experiments indicate that tetradecylthioacetic acid inhibits cell growth by mechanisms which may be independent of oxidative status.