Efficacy and Safety of an mRNA-Based RSV PreF Vaccine in Older Adults.

BACKGROUND: Respiratory syncytial virus (RSV) can cause substantial morbidity and mortality among older adults. An mRNA-based RSV vaccine, mRNA-1345, encoding the stabilized RSV prefusion F glycoprotein, is under clinical investigation. METHODS: In this ongoing, randomized, double-blind, placebo-controlled, phase 2-3 trial, we randomly assigned, in a 1:1 ratio, adults 60 years of age or older to receive one dose of mRNA-1345 (50 µg) or placebo. The two primary efficacy end points were the prevention of RSV-associated lower respiratory tract disease with at least two signs or symptoms and with at least three signs or symptoms. A key secondary efficacy end point was the prevention of RSV-associated acute respiratory disease. Safety was also assessed. RESULTS: Overall, 35,541 participants were assigned to receive the mRNA-1345 vaccine (17,793 participants) or placebo (17,748). The median follow-up was 112 days (range, 1 to 379). The primary analyses were conducted when at least 50% of the anticipated cases of RSV-associated lower respiratory tract disease had occurred. Vaccine efficacy was 83.7% (95.88% confidence interval [CI], 66.0 to 92.2) against RSV-associated lower respiratory tract disease with at least two signs or symptoms and 82.4% (96.36% CI, 34.8 to 95.3) against the disease with at least three signs or symptoms. Vaccine efficacy was 68.4% (95% CI, 50.9 to 79.7) against RSV-associated acute respiratory disease. Protection was observed against both RSV subtypes (A and B) and was generally consistent across subgroups defined according to age and coexisting conditions. Participants in the mRNA-1345 group had a higher incidence than those in the placebo group of solicited local adverse reactions (58.7% vs. 16.2%) and of systemic adverse reactions (47.7% vs. 32.9%); most reactions were mild to moderate in severity and were transient. Serious adverse events occurred in 2.8% of the participants in each trial group. CONCLUSIONS: A single dose of the mRNA-1345 vaccine resulted in no evident safety concerns and led to a lower incidence of RSV-associated lower respiratory tract disease and of RSV-associated acute respiratory disease than placebo among adults 60 years of age or older. (Funded by Moderna; ConquerRSV ClinicalTrials.gov number, NCT05127434.).

Evaluation of mRNA-1273 Vaccine in Children 6 Months to 5 Years of Age.

BACKGROUND: The safety, reactogenicity, immunogenicity, and efficacy of the mRNA-1273 coronavirus disease 2019 (Covid-19) vaccine in young children are unknown. METHODS: Part 1 of this ongoing phase 2-3 trial was open label for dose selection; part 2 was an observer-blinded, placebo-controlled evaluation of the selected dose. In part 2, we randomly assigned young children (6 months to 5 years of age) in a 3:1 ratio to receive two 25-µg injections of mRNA-1273 or placebo, administered 28 days apart. The primary objectives were to evaluate the safety and reactogenicity of the vaccine and to determine whether the immune response in these children was noninferior to that in young adults (18 to 25 years of age) in a related phase 3 trial. Secondary objectives were to determine the incidences of Covid-19 and severe acute respiratory syndrome coronavirus 2 infection after administration of mRNA-1273 or placebo. RESULTS: On the basis of safety and immunogenicity results in part 1 of the trial, the 25-µg dose was evaluated in part 2. In part 2, 3040 children 2 to 5 years of age and 1762 children 6 to 23 months of age were randomly assigned to receive two 25-µg injections of mRNA-1273; 1008 children 2 to 5 years of age and 593 children 6 to 23 months of age were randomly assigned to receive placebo. The median duration of follow-up after the second injection was 71 days in the 2-to-5-year-old cohort and 68 days in the 6-to-23-month-old cohort. Adverse events were mainly low-grade and transient, and no new safety concerns were identified. At day 57, neutralizing antibody geometric mean concentrations were 1410 (95% confidence interval [CI], 1272 to 1563) among 2-to-5-year-olds and 1781 (95% CI, 1616 to 1962) among 6-to-23-month-olds, as compared with 1391 (95% CI, 1263 to 1531) among young adults, who had received 100-µg injections of mRNA-1273, findings that met the noninferiority criteria for immune responses for both age cohorts. The estimated vaccine efficacy against Covid-19 was 36.8% (95% CI, 12.5 to 54.0) among 2-to-5-year-olds and 50.6% (95% CI, 21.4 to 68.6) among 6-to-23-month-olds, at a time when B.1.1.529 (omicron) was the predominant circulating variant. CONCLUSIONS: Two 25-µg doses of the mRNA-1273 vaccine were found to be safe in children 6 months to 5 years of age and elicited immune responses that were noninferior to those in young adults. (Funded by the Biomedical Advanced Research and Development Authority and National Institute of Allergy and Infectious Diseases; KidCOVE ClinicalTrials.gov number, NCT04796896.).

Safety and Immunogenicity of a Messenger RNA-Based Cytomegalovirus Vaccine in Healthy Adults: Results From a Phase 1, Randomized, Clinical Trial.

BACKGROUND: This phase 1 trial evaluated the safety, reactogenicity, and immunogenicity of mRNA-1647, an mRNA-based cytomegalovirus (CMV) vaccine, in CMV-seronegative and -seropositive adults. METHODS: Participants were randomly assigned to receive 30, 90, 180, or 300 µg of mRNA-1647 or placebo on a 0-, 2-, and 6-month schedule and followed for 12 months after the last dose. RESULTS: A total of 154 (80 CMV-seronegative and 74 CMV-seropositive) participants were enrolled; 118 participants were randomized to mRNA-1647 and 36 to placebo. Mean (SD) age was 32.5 (8.6) and 35.1 (8.9) years in the placebo and mRNA-1647 groups, respectively, in phase B (63% and 64% female) and 42.5 (6.2) and 33.3 (8.7) years, respectively, in phase C (2% and 16% female). No deaths, related serious adverse events, or adverse events of special interest were reported. Most adverse reactions were grade ≤2 severity. Increased neutralizing antibody, binding antibody, and antigen-specific cell-mediated responses were observed across mRNA-1647 treatment groups, regardless of CMV serostatus. CONCLUSIONS: This phase 1, first-in-human trial demonstrated mRNA-1647 has an acceptable safety profile in adults and elicits humoral and cellular immune responses. TRIALS REGISTRATION: NCT03382405; https://clinicaltrials.gov/ct2/show/NCT03382405.

Safety and Immunogenicity of an mRNA-Based RSV Vaccine Including a 12-Month Booster in a Phase I Clinical Trial in Healthy Older Adults.

BACKGROUND: An mRNA-based RSV vaccine, mRNA-1345, is under clinical investigation to address RSV disease burden in older adults. METHODS: This phase 1, randomized, observer-blind, placebo-controlled, dose-ranging study evaluated safety, reactogenicity, and immunogenicity of mRNA-1345 in adults 65-79 years (NCT04528719). Participants were randomized to receive 1-dose of mRNA-1345 (12.5, 25, 50, 100, or 200-µg) or placebo and matched mRNA-1345 booster or placebo at 12-months. RESULTS: Overall, 298 participants received the first injection; 247 received the 12-month booster injection. mRNA-1345 was generally well-tolerated after both injections, with the most frequently reported solicited adverse reactions being injection-site pain, fatigue, headache, arthralgia, and myalgia. Reactogenicity was higher after the booster injection than the first injection but similar severity, time-to-onset, and duration. A single mRNA-1345 injection boosted RSV-A and RSV-B neutralizing antibody titers (nAb) and prefusion-F-binding antibody (preF-bAb) concentrations at 1-month (geometric mean-fold rises: RSV-A, 10.2-16.5; RSV-B, 5.3-12.5; preF-bAb, 7.2-12.1). RSV antibody levels remained above baseline through 12-months, indicating immune persistence. A 12-month booster injection also increased RSV-A and RSV-B nAb titers and preF-bAb concentrations; titers post-booster injection were numerically lower compared to titers after the first-dose, with overlapping 95% CIs. CONCLUSIONS: mRNA-1345 was well-tolerated and immunogenic following a single injection and a 12-month booster. TRIAL REGISTRATION: ClinicalTrials.gov, NCT04528719.

Long-term prediction by DNA methylation of high-grade cervical intraepithelial neoplasia: Results of the ARTISTIC cohort.

Methylation markers have shown potential for triaging high-risk HPV-positive (hrHPV+) women to identify those at increased risk of invasive cervical cancer (ICC). Our aim was to assess the performance of the S5 DNA methylation classifier for predicting incident high-grade cervical intraepithelial neoplasia (CIN) and ICC among hrHPV+ women in the ARTISTIC screening trial cohort. The S5 classifier, comprising target regions of tumour suppressor gene EPB41L3 and L1 and L2 regions of HPV16, HPV18, HPV31, and HPV33, was assayed by pyrosequencing in archived hrHPV+ liquid-based samples from 343 women with high-grade disease (139 CIN2, 186 CIN3, and 18 ICC) compared to 800 hrHPV+ controls. S5 DNA methylation correlated directly with increasing severity of disease and inversely with lead time to diagnosis. S5 could discriminate between hrHPV+ women who developed CIN3 or ICC and hrHPV+ controls (p <.0001) using samples taken on average 5 years before diagnosis. This relationship was independent of cytology at baseline. The S5 test showed much higher sensitivity than HPV16/18 genotyping for identifying prevalent CIN3 (93% vs. 61%, p = .01) but lower specificity (50% vs. 66%, p <.0001). The S5 classifier identified most women at high risk of developing precancer and missed very few prevalent advanced lesions thus appearing to be an objective test for triage of hrHPV+ women. The combination of methylation of host and HPV genes enables S5 to combine the predictive power of methylation with HPV genotyping to identify hrHPV-positive women who are at highest risk of developing CIN3 and ICC in the future.

Sutimlimab in Cold Agglutinin Disease.

BACKGROUND: Cold agglutinin disease is a rare autoimmune hemolytic anemia characterized by hemolysis that is caused by activation of the classic complement pathway. Sutimlimab, a humanized monoclonal antibody, selectively targets the C1s protein, a C1 complex serine protease responsible for activating this pathway. METHODS: We conducted a 26-week multicenter, open-label, single-group study to assess the efficacy and safety of intravenous sutimlimab in patients with cold agglutinin disease and a recent history of transfusion. The composite primary end point was a normalization of the hemoglobin level to 12 g or more per deciliter or an increase in the hemoglobin level of 2 g or more per deciliter from baseline, without red-cell transfusion or medications prohibited by the protocol. RESULTS: A total of 24 patients were enrolled and received at least one dose of sutimlimab; 13 patients (54%) met the criteria for the composite primary end point. The least-squares mean increase in hemoglobin level was 2.6 g per deciliter at the time of treatment assessment (weeks 23, 25, and 26). A mean hemoglobin level of more than 11 g per deciliter was maintained in patients from week 3 through the end of the study period. The mean bilirubin levels normalized by week 3. A total of 17 patients (71%) did not receive a transfusion from week 5 through week 26. Clinically meaningful reductions in fatigue were observed by week 1 and were maintained throughout the study. Activity in the classic complement pathway was rapidly inhibited, as assessed by a functional assay. Increased hemoglobin levels, reduced bilirubin levels, and reduced fatigue coincided with inhibition of the classic complement pathway. At least one adverse event occurred during the treatment period in 22 patients (92%). Seven patients (29%) had at least one serious adverse event, none of which were determined by the investigators to be related to sutimlimab. No meningococcal infections occurred. CONCLUSIONS: In patients with cold agglutinin disease who received sutimlimab, selective upstream inhibition of activity in the classic complement pathway rapidly halted hemolysis, increased hemoglobin levels, and reduced fatigue. (Funded by Sanofi; CARDINAL ClinicalTrials.gov number, NCT03347396.).

Evaluation of a validated methylation triage signature for human papillomavirus positive women in the HPV FOCAL cervical cancer screening trial.

Human papillomavirus (HPV)-based cervical cancer screening requires triage of HPV positive women to identify those at risk of cervical intraepithelial neoplasia grade 2 (CIN2) or worse. We conducted a blinded case-control study within the HPV FOCAL randomized cervical cancer screening trial of women aged 25-65 to examine whether baseline methylation testing using the S5 classifier provided triage performance similar to an algorithm relying on cytology and HPV genotyping. Groups were randomly selected from women with known HPV/cytology results and pathology outcomes. Group 1: 104 HPV positive (HPV+), abnormal cytology (54 CIN2/3; 50

A Validated Scale for Assessing the Severity of Acute Infectious Mononucleosis.

OBJECTIVES: To develop a scale for the severity of mononucleosis. STUDY DESIGN: One to 5 percent of college students develop infectious mononucleosis annually, and about 10% meet criteria for chronic fatigue syndrome (CFS) 6 months following infectious mononucleosis. We developed a severity of mononucleosis scale based on a review of the literature. College students were enrolled, generally when they were healthy. When the students developed infectious mononucleosis, an assessment was made as to the severity of their infectious mononucleosis independently by 2 physicians using the severity of mononucleosis scale. This scale was correlated with corticosteroid use and hospitalization. Six months following infectious mononucleosis, an assessment is made for recovery from infectious mononucleosis or meeting 1 or more case definitions of CFS. RESULTS: In total, 126 severity of mononucleosis scales were analyzed. The concordance between the 2 physician reviewers was 95%. All 3 hospitalized subjects had severity of mononucleosis scores ≥2. Subjects with severity of mononucleosis scores of ≥1 were 1.83 times as likely to be given corticosteroids. Students with severity of mononucleosis scores of 0 or 1 were less likely to meet more than 1 case definition of CFS 6 months following infectious mononucleosis. CONCLUSIONS: The severity of mononucleosis scale has interobserver, concurrent and predictive validity for hospitalization, corticosteroid use, and meeting criteria for CFS 6 months following infectious mononucleosis.

Molecular progression to cervical precancer, epigenetic switch or sequential model?

The evolution of precancerous cervical lesions is poorly understood. A widely held model of cervical intraepithelial neoplasia grade 3 (CIN3) development is sequential progression from normal through CIN1 and CIN2 to CIN3. Another hypothesis, the "molecular switch" model, postulates that CIN3 can evolve directly from human papillomavirus (HPV)-infected normal epithelium without progressing through CIN1 and CIN2. To shed light on this process, we compared DNA methylation of selected human biomarkers and HPV types in two groups of CIN1: CIN1 that were near or adjacent to CIN3 (adjacent-CIN1) and CIN1 that were the principal lesions with no CIN3 detected (principal-CIN1). 354 CIN (CIN1 and CIN3) and normal tissue areas were dissected and typed for HPV from 127 women who underwent loop electrosurgical excision procedures (LEEP). Methylation of genes EPB41L3 and the viral regions of HPV16-L1/L2, HPV18-L2, HPV31-L1, and HPV33-L2 were determined by a highly accurate quantitative pyrosequencing of bisulfite converted DNA. There was a significant trend of increased methylation with disease grade comparing normal to CIN1 and CIN3 (p < 0.0001). Adjacent-CIN1 predominantly shared the same HPV types as the CIN3, however, methylation differed substantially between adjacent-CIN1 and CIN3 (p = 0.008). In contrast diagnostically principal-CIN1 had an indistinguishable methylation distribution compared to adjacent-CIN1 (EPB41L3: p = 0.49; HPVme-All: p = 0.11). Our results suggest that progression from normal epithelium to CIN1 or CIN3 is usually promoted by the same HPV type but occurs via distinct DNA epigenotypes, thus favoring the "molecular switch" model.

Role of quantitative p16INK4A mRNA assay and digital reading of p16INK4A immunostained sections in diagnosis of cervical intraepithelial neoplasia.

Visual interpretation of cervical biopsies is subjective and variable, generally showing fair to moderate inter-reader agreement in distinguishing high from low grade cervical intraepithelial neoplasia (CIN). We investigated the performance of two objective p16 quantitative tests in comparison with visual assessment: (i) p16-mRNA assay and (ii) digital analysis of sections stained for p16 protein. The primary analysis considered 232 high-risk human papilloma virus positive (HPV+) samples from diagnostic cervical specimens. A p16 RT-qPCR (p16-mRNA assay) was run on mRNA extracted from formalin-fixed paraffin-embedded sections. Two p16 immunohistochemistry (IHC) readings, a visual read by a histopathologist (Visual IHC) and a digital read of a high-resolution scan (Digital IHC), were done on adjacent sections. The worst reviewed CIN grade (agreed by at least two histopathologists) from up to two biopsies and a loop excision was taken, with CIN2/3 as the primary endpoint. Visual IHC attained a specificity of 70% (95%CI 61-77) for 85% (95%CI 77-91%) sensitivity. The four-point Visual IHC staining area under the curve (AUC) was 0.77 (95%CI 0.71-0.82), compared with 0.71 (95%CI 0.64-0.77) for p16-mRNA and 0.67 (95%CI 0.60-0.74) for Digital IHC. Spearman rank-order correlations were: visual to p16-mRNA 0.41, visual to digital 0.49 and p16-mRNA to digital: 0.22. The addition of p16-mRNA assay to visual reading of p16 IHC improved the AUC from 0.77 to 0.84 (p = 0.0049). p16-mRNA testing may be complementary to visual IHC p16 staining for a more accurate diagnosis of CIN, or perhaps a substitute in locations with a lack of skilled pathologists.

Central venous catheter salvage in children with Staphylococcus aureus central line-associated bloodstream infection.

BACKGROUND: Prompt central venous catheter (CVC) removal is currently recommended in children with Staphylococcus aureus central line-associated bloodstream infection (CLABSI). Our objective was to examine the outcome of attempted line salvage in children with S. aureus CLABSI and assess predictors of success. METHODS: A single-institution, retrospective cohort study was performed of all children with S. aureus CLABSI between 2012 and 2015. Patients with and without immediate CVC removal (≤ 2 days after first positive culture) were compared. The primary outcome was failed CVC salvage (removal after 3+ days). RESULTS: Seventy-seven children met criteria for S. aureus CLABSI. Immediate CVC removal was performed in 27.3% of patients. Among the 72.7% patients in whom CVC salvage was attempted, 78.6% were successful and 21.4% required delayed CVC removal. Malignancy, short gut syndrome, neutropenia, methicillin-resistant S. aureus, and line type were not associated with salvage failure. No associated morbidity or mortality occurred in patients with a failed salvage attempt. New or recurrent bacteremia occurred in five patients, but three were successfully salvaged a second time. CONCLUSIONS: CVC salvage was feasible in the majority of children with S. aureus CLABSI and was not associated with significant complications or attributable mortality as reported in adults.

Validation of a DNA methylation HPV triage classifier in a screening sample.

High-risk human papillomavirus (hrHPV) DNA tests have excellent sensitivity for detection of cervical intraepithelial neoplasia 2 or higher (CIN2+). A drawback of hrHPV screening, however, is modest specificity. Therefore, hrHPV-positive women might need triage to reduce adverse events and costs associated with unnecessary colposcopy. We compared the performance of HPV16/18 genotyping with a predefined DNA methylation triage test (S5) based on target regions of the human gene EPB41L3, and viral late gene regions of HPV16, HPV18, HPV31 and HPV33. Assays were run using exfoliated cervical specimens from 710 women attending routine screening, of whom 38 were diagnosed with CIN2+ within a year after triage to colposcopy based on cytology and 341 were hrHPV positive. Sensitivity and specificity of the investigated triage methods were compared by McNemar's test. At the predefined cutoff, S5 showed better sensitivity than HPV16/18 genotyping (74% vs 54%, P = 0.04) in identifying CIN2+ in hrHPV-positive women, and similar specificity (65% vs 71%, P = 0.07). When the S5 cutoff was altered to allow equal sensitivity to that of genotyping, a significantly higher specificity of 91% was reached (P < 0.0001). Thus, a DNA methylation test for the triage of hrHPV-positive women on original screening specimens might be a valid approach with better performance than genotyping.

Relationship of ZNF423 and CTSO with breast cancer risk in two randomised tamoxifen prevention trials.

A case-control study from two randomised breast cancer prevention trials of tamoxifen and raloxifene (P-1 and P-2) identified single-nucleotide polymorphisms (SNPs) in or near genes ZNF423 and CTSO as factors which predict which women will derive most anti-cancer benefit from selective oestrogen receptor modulator (SERM) therapy. In this article, we further examine this question using blood samples from two randomised tamoxifen prevention trials: the International Breast Cancer Intervention Study I (IBIS-I) and the Royal Marsden trial (Marsden). A nested case-control study was designed with 2:1 matching in IBIS-I and 1:1 matching in Marsden. The OncoArray was used for genotyping and included two SNPs previously identified (rs8060157 in ZNF423 and rs10030044 near CTSO), and 102 further SNPs within the same regions. Overall, there were 369 cases and 662 controls, with 148 cases and 268 controls from the tamoxifen arms. Odds ratios were estimated by conditional logistic regression, with Wald 95 % confidence intervals. In the tamoxifen arms, the per-allele odds ratio for rs8060157 was 0.99 (95 %CI 0.73-1.34) and 1.00 (95 %CI 0.76-1.33) for rs10030044. In the placebo arm, the odds ratio was 1.10 (95 %CI 0.87-1.40) for rs8060157 and 1.01 (95 %CI 0.79-1.29) for rs10030044. There was no evidence to suggest that other SNPs in the surrounding regions of these SNPs might predict response to tamoxifen. Results from these two prevention trials do not support the earlier findings. rs8060157 in ZNF423 and rs10030044 near CTSO do not appear to predict response to tamoxifen.

Polymorphism in the neurofibromin gene, Nf1, is associated with antagonistic selection on wing size and development time in Drosophila melanogaster.

In many invertebrates, body size shows genetically based clines, with size increasing in colder climates. Large body size is typically associated with prolonged development times. We consider variation in the CNS-specific gene neurofibromin 1 (Nf1) and its association with body size and development time. We identified two major Nf1 haplotypes in natural populations, Nf1-insertion-A and Nf1-deletion-G. These haplotypes are characterized by a 45-base insertion/deletion (INDEL) in Nf1 intron 2 and an A/G synonymous substitution (locus L17277). Linkage disequilibrium (LD) between the INDEL and adjacent sites is high but appears to be restricted within the Nf1 gene interval. In Australia, the frequency of the Nf1-insertion-A haplotype increases with latitude where wing size is larger, independent of the chromosomal inversion In(3R)Payne. Unexpectedly, the Nf1-insertion-A haplotype is negatively associated with wing size. We found that the Nf1-insertion-A haplotype is enriched in females with shorter development time. This suggests that the Nf1 haplotype cline may be driven by selection for development time rather than size; females from southern (higher latitude) D. melanogaster populations maintain a rapid development time despite being relatively larger, and the higher incidence of Nf1-insertion-A in Southern Australia may contribute to this pattern, whereas the effects of the Nf1 haplotypes on size may be countered by other loci with antagonistic effects on size and development time. Our results point to the potential complexity involved in identifying selection on genetic variants exhibiting pleiotropic effects when studies are based on spatial patterns or association studies.

Safety and efficacy of classical complement pathway inhibition with sutimlimab in chronic immune thrombocytopenia.

Chronic/refractory immune thrombocytopenia (ITP) is a rare and pathophysiologically heterogeneous disorder with variable responsiveness to available treatments. Sutimlimab, a first-in-class humanized monoclonal anti-C1s IgG4 antibody, selectively inhibits the classical pathway. This phase 1 study (NCT03275454) assessed the safety, efficacy, pharmacokinetics, and pharmacodynamics of biweekly sutimlimab in patients with chronic/refractory ITP with an inadequate response to ≥2 therapies (platelet count ≤ 30 × 109/L). Twelve patients (median age 42 years) received sutimlimab for a median of 20.5 weeks followed by a median 2-week washout period (part A). In part B, 7 of the 12 eligible patients received sutimlimab retreatment for a median of 113 weeks. In part A, the mean (standard deviation) platelet count increased from 25 × 109/L (17) to 54 × 109/L (60) 24 hours after starting sutimlimab, maintaining ≥50 × 109/L throughout part A. Five patients (42%) achieved durable platelet count responses (≥50 × 109/L in ≥50% of follow-up visits) and 4 achieved complete response (platelet count ≥100 × 109/L). The mean platelet count returned to baseline during washout and increased upon retreatment in part B. The mean platelet count improvements accompanied the rapid inhibition of the classical pathway. There were 74 treatment-emergent adverse events in part A (n = 10) and 70 in part B (n = 6). Five serious adverse events were observed; 1 event (migraine) was assessed by the investigator as related to sutimlimab. These results demonstrated that in some patients with ITP, autoantibodies activate the classical complement pathway, accelerating platelet destruction or impairing platelet production and contributing to treatment failure. Thus, C1s inhibition may be a safe and beneficial therapeutic approach for patients with chronic/refractory ITP.

Molecular basis of adaptive shift in body size in Drosophila melanogaster: functional and sequence analyses of the Dca gene.

Latitudinal body size clines in animals conforming to Bergmann's rule occur on many continents but isolating their underlying genetic basis remains a challenge. In Drosophila melanogaster, the gene Dca accounts for approximately 5-10% of the natural wing size variation (McKechnie SW, Blacket MJ, Song SV, Rako L, Carroll X, Johnson TK, Jensen LT, Lee SF, Wee CW, Hoffmann AA. 2010. A clinally varying promoter polymorphism associated with adaptive variation in wing size in Drosophila. Mol Ecol. 19:775-784). We present here functional evidence that Dca is a negative regulator of wing size. A significant negative latitudinal cline of Dca gene expression was detected in synchronized third instar larvae. In addition, we clarified the evolutionary history of the three most common Dca promoter alleles (Dca237-1, Dca237-2, and Dca247) and showed that the insertion allele (Dca247), whose frequency increases with latitude, is associated with larger wing centroid size and higher average cell number in male flies. Finally, we showed that the overall linkage disequilibrium (LD) was low in the Dca promoter and that the insertion/deletion polymorphism that defines the Dca alleles was in strong LD with two other upstream sites. Our results provide strong support that Dca is a candidate for climatic adaptation in D. melanogaster.

Documentation of Indications: Agreement Between Order Entry and Clinical Notes and Effect on Time to Antibiotic Administration.

BACKGROUND/OBJECTIVES: Antibiotic indication documentation at the time of order entry is mandated by the Joint Commission. Inclusion of indication at order entry may have an impact on the time to administration. Our primary objective was to evaluate agreement between indication selected during order entry and clinical notes. Our secondary objective was to observe if there was a change in time to administration after indications were required during order entry. METHODS: Patients ≤18 years old who received ≥1 dose of vancomycin or ceftriaxone during a preintervention period and 3 postintervention periods were included. Indication for use, agreement between order and clinical note, and timing of antibiotic administration were collected. RESULTS: Most common indication for vancomycin (total: 789) was sepsis (26%, n = 204). Common indications for ceftriaxone (total: 1071) were sepsis (12%, n = 127), perforated appendicitis (12%, n = 125), and urinary tract infection (10%, n = 107). Postintervention, agreement between the indication selected during order entry and indication documented in clinical note for ceftriaxone and vancomycin orders were 41% and 46%, respectively. Median time to administration decreased among patients who received ceftriaxone (P < .01) but had no significant impact on time to administration of vancomycin (P = .49). CONCLUSIONS: Indication for ceftriaxone and vancomycin selected during order entry and reported in clinical notes inconsistently matched. Inclusion of antibiotic indication may impact time to administration.

Consistency of the S5 DNA methylation classifier in formalin-fixed biopsies versus corresponding exfoliated cells for the detection of pre-cancerous cervical lesions.

Methylation biomarkers are promising tools for diagnosis and disease prevention. The S5 classifier is aimed at the prevention of cervical cancer by the early detection of cervical intraepithelial neoplasia (CIN). S5 is based on pyrosequencing a promoter region of EPB41L3 and five late regions of HPV types 16, 18, 31, and 33 following bisulfite conversion of DNA. Good biomarkers should perform well in a variety of sample types such as exfoliated cells, fresh frozen or formalin-fixed paraffin-embedded (FFPE) materials. Here, we tested the performance of S5 on 315 FFPE biopsies with paired exfoliated cervical samples using four different conversion kits (Epitect Bisulfite, Epitect Fast Bisulfite, EZ DNA Methylation, and EZ DNA Methylation-Lightning). The S5 values from FFPE biopsies for all kits were significantly correlated with those obtained from their paired exfoliated cells. For the EZ DNA Methylation kit, we observed an average increased methylation of 4.4% in FFPE. This was due to incomplete conversion of DNA (73% for FFPE vs. 95% for cells). The other kits had a DNA conversion rate in FFPE similar to the cells (95%-97%). S5 performed well at discriminating

A Randomized Comparison of Different Vaginal Self-sampling Devices and Urine for Human Papillomavirus Testing-Predictors 5.1.

BACKGROUND: Human papillomavirus (HPV)-based screening is rapidly replacing cytology as the cervical screening modality of choice. In addition to being more sensitive than cytology, it can be done on self-collected vaginal or urine samples. This study will compare the high-risk HPV positivity rates and sensitivity of self-collected vaginal samples using four different collection devices and a urine sample. METHODS: A total of 620 women referred for colposcopy were invited to provide an initial stream urine sample collected with the Colli-Pee device and take two vaginal self-samples, using either a dry flocked swab (DF) and a wet dacron swab (WD), or a HerSwab (HS) and Qvintip (QT) device. HPV testing was performed by the BD Onclarity HPV Assay. RESULTS: A total of 600 vaginal sample pairs were suitable for analysis, and 505 were accompanied by a urine sample. Similar positivity rates and sensitivities for CIN2+ and CIN3+ were seen for DF, WD, and urine, but lower values were seen for QT and HS. No clear user preferences were seen between devices, but women found urine easiest to collect, and were more confident they had taken the sample correctly. The lowest confidence in collection was reported for HS. CONCLUSIONS: Urine, a DF swab, and WD swab all performed well and were well received by the women, whereas the Qvintip and HerSwab devices were less satisfactory. IMPACT: This is the first study to compare five self-sampling methods in the same women taken at the same time. It supports wider use of urine or vaginal self-sampling for cervical screening.