MRI-visualized T2 hyperintense breast lesions: identifying clinical and imaging factors linked to malignant biopsy outcomes.

PURPOSE: To determine the malignancy rate for MRI-guided breast biopsies performed for T2 hyperintense breast lesions and to assess additional clinical and MRI characteristics that can predict benign and malignant outcomes. METHODS: A retrospective chart review of consecutive MRI-guided breast biopsies performed in two tertiary hospitals was conducted over two years. Biopsies performed for T2 hyperintense lesions were selected, and further lesion imaging characteristics and patient risk factors were collected. Univariate and multivariate modeling regression were used to determine additional imaging and patient factors associated with malignant outcomes for biopsies of T2 hyperintense lesions. RESULTS: Out of 369 MRI-guided breast biopsies, 100 (27%) were performed for T2 hyperintense lesions. Two biopsy-proven benign lesions were excluded as the patient was lost on follow-up. With a study cohort of 98 lesions, the final pathology results were benign for 80 (80%) of these lesions, while 18 (18%) were malignant. Using multivariate logistic modeling, patient age > 50 (OR 5.99 (1.49, 24.08 95% CI), p < 0.05) and lesion size > 3 cm (OR 5.54 (1.54-18.7), p < 0.01) were found to be important predictors of malignant outcomes for MRI biopsies performed for T2 hyperintense lesions. CONCLUSION: Our study observed a high malignancy rate, challenging the assumption that T2 hyperintensity can be considered a benign imaging characteristic for otherwise suspicious MRI-detected lesions. Decision-making regarding tissue sampling should be made based on a thorough evaluation of more reliable additional demographic and imaging factors, including patient age and lesion size.

Evaluating persistent T1-weighted lesions without concurrent abnormal enhancement on breast MRI in neoadjuvant chemotherapy patients: implications for complete pathological response.

OBJECTIVE: This study aims to determine whether persistent T1-weighted lesions signify a complete pathological response (pCR) in breast cancer patients treated with neoadjuvant chemotherapy and surgery, and to evaluate their correlation with imaging responses on MRI. MATERIALS AND METHODS: A retrospective review was conducted on data from breast cancer patients treated between January 2011 and December 2018. Patients who underwent breast MRI and pre- and post-neoadjuvant chemotherapy followed by surgery were included. Those with distant metastasis, no planned surgery, pre-surgery radiation, ineligibility for neoadjuvant chemotherapy, or unavailable surgical pathology were excluded. Groups with and without persistent T1-weighted lesions were compared using the chi-square test for categorical variables and the Student t test or Wilcox rank sum test for continuous variables. Univariate logistic regression was used to evaluate the association of the final pathological response with the presence of T1-persistent lesion and other characteristics. RESULTS: Out of 319 patients, 294 met the inclusion criteria (breast cancer patients treated with neoadjuvant chemotherapy and subsequent surgery); 157 had persistent T1 lesions on post-chemotherapy MRI and 137 did not. A persistent T1 lesion indicated reduced likelihood of complete pathological response (14% vs. 39%, p < 0.001) and imaging response (69% vs. 93%, p < 0.001). Multivariable analysis confirmed these findings: OR 0.37 (95% CI 0.18-0.76), p = 0.007. No other characteristics correlated with T1 residual lesions. CONCLUSION: Persistent T1-weighted lesions without associated abnormal enhancement on post-treatment breast MRI correlate with lower complete pathological and imaging response rates. CLINICAL RELEVANCE STATEMENT: The study underscores the importance of persistent T1-weighted lesions on breast MRI as vital clinical markers, being inversely related to a complete pathological response following neoadjuvant chemotherapy; they should be a key factor in guiding post-neoadjuvant chemotherapy treatment decisions. KEY POINTS: • Persistent T1 lesions on post-chemotherapy breast MRI indicate a reduced likelihood of achieving a complete pathological response (14% vs. 39%, p < 0.001) and imaging response (69% vs. 93%, p < 0.001). • Through multivariable analysis, it was confirmed that the presence of a persistent T1 lesion on breast MRI post-chemotherapy is linked to a decreased likelihood of complete pathological response, with an odds ratio (OR) of 0.37 (95% CI 0.18-0.76; p = 0.007). • In addition to the convention of equating the absence of residual enhancement to complete imaging response, our results suggest that the presence or absence of residual T1 lesions should also be considered.

The characteristics associated with upgrade on surgical pathology of conventional imaging occult DCIS diagnosed by MRI.

PURPOSE: To characterize the clinical, pathological, and imaging features of DCIS occult on conventional imaging diagnosed on MRI-guided biopsy associated with increased risk of invasive disease on surgical excision. MATERIALS AND METHODS: All consecutive patients with MRI-detected DCIS occult on conventional imaging between January 2009 and December 2018 were included. Women were divided into two groups based on final pathology: Pure DCIS or DCIS with invasive component. Clinical, imaging, and pathological risk factors for upgrade to invasion were evaluated. RESULTS: Of 50 patients who met the inclusion criteria, 12 (24%) were upgraded to invasive malignancy in the final pathology. The only parameters that showed statistically significant association with upgrade were related to kinetic characteristics: 53% of patients with the combination of fast early upstroke and either plateau or washout curve were upgraded, compared to 12% of women without this combination (p = 0.006). The sensitivity of combined kinetic features for predicting upgrade was 67% (95% CI 35-90%), specificity was 84% (CI 95% 68-94%), positive predictive value was 57% (CI 95% 37-75%), negative predictive value was 89% (CI 95% 77-95%), and OR was 78% (64-88%). CONCLUSION: Kinetic characteristics show the strongest association with upgrade to invasion in DCIS occult on mammogram and US. Larger studies should be encouraged to consolidate our findings, which may have implication for treatment planning.

Lobular neoplasia occult on conventional imaging and diagnosed on MRI-guided biopsy: can we estimate upgrade on surgical pathology?

PURPOSE: The goal of this study is to evaluate the frequency and imaging features of lobular neoplasia (LN) diagnosed on MRI-guided biopsy, determine the upgrade rate to malignancy, and assess for any features that may be associated with an upgrade on surgical excision. MATERIALS AND METHODS: Research ethical board approved the review of consecutive patients with MRI-detected LN between January 2009 and December 2018 with differentiation between pure LN and LN with associated other high-risk lesions. The final outcome was determined by final pathology results from surgical excision or 24 months of follow-up. Appropriate statistical tests were used. RESULTS: Out of 1250 MRI-guided biopsies performed, 76 lesions (6%) fulfilled the inclusion criteria and formed the study cohort. Of the 76 lesions, 54 (71%) were pure LN while the rest had coexistent high-risk lesion. Non-mass enhancement (NME) was the most common lesion type (62, 82%). Fifty-nine lesions (78%) were surgically excised, the other 17 had benign follow-up. Overall, 8 lesions (11%) were upgraded to malignancy on final pathology. Malignant outcome was associated with larger lesion size (5.5 versus 1.9 cm, P < 0.001) and a clumped NME pattern (75% versus 24%, P = 0.006). Lesion size and clumped NME remained significantly associated with upgrade on sub-analysis of the pure LN group. CONCLUSION: Larger lesion size and clumped NME are imaging findings associated with upgrade of LN diagnosed by MRI-guided biopsy. This may influence patient management in this clinical setting. Additional larger studies are needed to consolidate our results and to potentially detect additional factors associated with upgrade.

Can breast MRI accurately exclude malignancy in mammographic architectural distortion?

OBJECTIVES: To investigate the diagnostic accuracy of problem-solving breast magnetic resonance imaging (MRI) in excluding malignancy in a cohort of patients diagnosed with mammographic architectural distortion (MAD). METHODS: The Institutional Review Board approved the study. Imaging database with 40,245 breast MRIs done between January 2008 and September 2018 was retrospectively reviewed. The study included all exams considered problem-solving MRI for MAD. Two radiologists reviewed the imaging data. Outcome was determined by the pathology results of biopsy/surgical excision or at least 1 year of clinical and radiological follow-up. Predictors for malignancy were examined, and appropriate statistical tests were applied. RESULTS: One hundred seventy-five patients (median age 53 years) fulfilled the inclusion criteria and formed the study cohort. No cancers were diagnosed in 106 patients with a negative MRI. Out of 69 women with positive MRI findings, 48 (70%) had benign outcome defined either by pathology result or by negative follow-up, and 21 (30%) yielded malignancy. Malignancy was significantly associated with positive MRI (p < 0.001) and older age (p = 0.014). Falsely positive MRIs were frequently found in women with radial scars. The sensitivity, specificity, negative predictive value, positive predictive value, and overall accuracy of breast MRI were 100% (95% CI 84 to 100%), 68% (CI 61 to 76%), 100% (CI 95 to 100%), 30% (CI 26 to 36%), and 73% (95% CI 66-79), respectively. CONCLUSION: A negative breast MRI in patients with MAD was reliable in excluding malignancy in this cohort and may have a role as a precision medicine tool for avoiding unnecessary interventions. KEY POINTS: • MRI shows a high negative predictive value in MAD cases. • MRI displays low accuracy in differentiating malignancy from RS. • MRI is a reliable non-invasive method to exclude malignancy in women with mammographic architectural distortion, potentially avoiding unnecessary biopsies and surgeries.

Cellular fibroepithelial lesions diagnosed on core needle biopsy: Is there any role of clinical-sonography features helping to differentiate fibroadenomas and phyllodes tumor?

OBJECTIVE: The purpose of this study is to determine the role of clinico-sonographic features of breast cellular fibroepithelial lesions (CFELs) diagnosed on core needle biopsy (CNB) in the differentiation between fibroadenoma (FA) and phyllodes. MATERIALS AND METHODS: Results of consecutive women with a CNB showing CFEL from 2005 to 2010 were retrospectively reviewed. Clinical and sonographic findings were compared with surgical outcomes. Chi-square and Fisher's exact tests were used followed by a regression model for statistical analysis. RESULTS: A total of 131 women with 134 CFEL were included in the study; 89 (66%) were FAs and 45 (34%) were phyllodes (32 benign; 13 malignant). Significant predictors of increased risk of phyllodes tumor were patient age equal to or greater than 50 years (P = .021) and lesion size less than 2 cm at sonography (P = .043). No other imaging or clinical features were able to differentiate FA from phyllodes tumors. CONCLUSION: CFEL with a larger size in older women is associated with the surgical pathological result of phyllodes tumor and management should be tailored accordingly. Younger patients with small size nodules might be approached less aggressively, depending on a personalized discussion with the surgeons, taking into account the results obtained in this study.

Prevalence of pain reports in pediatric primary care and association with demographics, body mass index, and exam findings: a cross-sectional study.

BACKGROUND: Pediatric pain is associated to patient weight and demographics in specialized settings, but pain prevalence and its associated patient attributes in general pediatric outpatient care are unknown. Our objective was to determine the rate of positive pain screenings in pediatric primary care and evaluate the relationship between reported pain and obesity, demographics, and exam findings during routine pediatric encounters. METHODS: Cross-sectional observational study of 26,180 patients ages 2 to 19 seen in five urban pediatric primary care clinics between 2009 and 2016. Data were collected from systematic screening using a computerized clinical decision support system. Multivariable logistic regressions were used to analyze the association between pain reporting and obesity (body mass index), age, sex, race, season, insurance status, clinic site, prior pain reporting, pain reporting method, and exam findings. RESULTS: Pain was reported by the patient or caregiver in 14.9% of visits. In adjusted models, pain reporting was associated with obesity (Odds Ratio (OR) 1.23, 95% Confidence Intervals (CI) 1.11-1.35) and severe obesity (OR 1.32, CI 1.17-1.49); adolescents (OR 1.47, CI 1.33-1.61); and females (OR 1.21, CI 1.12-1.29). Pain reported at the preceding visit increased odds of pain reporting 2.67 times (CI 2.42-2.95). Abnormal abdominal, extremity, ear, nose, throat, and lymph node exams were associated with pain reporting. Pain reporting increased in minority races within clinics that predominantly saw a concordant race. CONCLUSIONS: Pain is common in general pediatric encounters, and occurs more frequently in obese children and those who previously reported pain. Pain reporting may be influenced by seasonal variation and clinic factors. Future pediatric pain screening may be guided by associated risk factors to improve identification and targeted healthcare interventions.

Novel messenger RNAs for body fluid identification.

In forensic investigations, the identification of the cellular or body fluid source of biological evidence can provide crucial probative information for the court. Messenger RNA (mRNA) profiling has become a valuable tool for body fluid and cell type identification due to its high sensitivity and compatibility with DNA analysis. However, using a single marker to determine the somatic origin of a sample can lead to misinterpretation as a result of cross-reactions. While false positives may be avoided through the simultaneous detection of multiple markers per body fluid, this approach is currently limited by the small number of known differentially expressed mRNAs. Here we characterise six novel mRNAs, partly identified from RNA-Seq, which can supplement existing markers for the detection of circulatory blood, semen (with and without spermatozoa), and menstrual fluid: HBD and SLC4A1 for blood, TNP1 for spermatozoa, KLK2 for seminal fluid, and MMP3 and STC1 for menstrual fluid. Respective expression profiles were evaluated by singleplex endpoint reverse transcription polymerase chain reaction (RT-PCR). HBD, SLC4A1, and KLK2 were specific to their target body fluids. TNP1, MMP3, and STC1 each cross-reacted with two non-target samples; however, these signals were below 350RFU, not reproducible, and likely resulted from large body fluid inputs. All candidates were more sensitive for the detection of their target body fluids than corresponding well-known mRNAs, in particular those for menstrual fluid. The increased sensitivities were statistically significant, except for KLK2. Thus, the new mRNAs introduced here are promising new targets for improved body fluid profiling.

Evaluation of the Utility of Screening Mammography for High-Risk Women Undergoing Screening Breast MR Imaging.

Purpose To evaluate the value of mammography in detecting breast cancer in high-risk women undergoing screening breast magnetic resonance (MR) imaging. Materials and Methods An ethics-approved, retrospective review of prospective databases was performed to identify outcomes of 3934 screening studies (1977 screening MR imaging examinations and 1957 screening mammograms) performed between January 2012 and July 2014 in 1249 high-risk women. Performance measures including recall and cancer detection rates, sensitivity, specificity, and positive predictive values were calculated for both mammography and MR imaging. Results A total of 45 cancers (33 invasive and 12 ductal carcinomas in situ) were diagnosed, 43 were seen with MR imaging and 14 with both mammography and MR imaging. Additional tests (further imaging and/or biopsy) were recommended in 461 screening MR imaging studies (recall rate, 23.3%; 95% confidence interval [CI]: 21.5%, 25.2%), and mammography recalled 217 (recall rate, 11.1%; 95% CI: 9.7%, 12.6%). The cancer detection rate for MR imaging was 21.8 cancers per 1000 examinations (95% CI: 15.78, 29.19) and that for mammography was 7.2 cancers per 1000 examinations (95% CI: 3.92, 11.97; P < .001). Sensitivity and specificity of MR imaging were 96% and 78% respectively, and those of mammography were 31% and 89%, respectively (P < .001). Positive predictive value for MR imaging recalls was 9.3% (95% CI: 6.83%, 12.36%) and that for mammography recalls was 6.5% (95% CI: 3.57%, 10.59%). Conclusion Contemporaneous screening mammography did not have added value in detection of breast cancer for women who undergo screening MR imaging. Routine use of screening mammography in women undergoing screening breast MR imaging warrants reconsideration. © RSNA, 2017 Online supplemental material is available for this article.

A time-course analysis of mRNA expression during injury healing in human dermal injuries.

The determination of dermal injury age is important in forensic practice. It helps answer questions that are important to an investigation such as the timing of the injury and incident, the order of infliction (where there is more than one injury), the survival time after injury (post-infliction interval) and the relation of the injury to the incident. Despite the importance of injury age determination, there currently exists no reliable method to estimate dermal injury age. In this study, the expression of the following 14 mRNAs was studied in human dermal injuries and their usefulness in the estimation of human dermal injury age was evaluated: dual specificity phosphate 1 (DUSP1), interleukin 7 (IL7), vascular cell adhesion molecule 1, tenascin C, cluster of differentiation 14, interleukin 6, tumour necrosis factor alpha (TNFα), interleukin 1beta (IL1ß), chymase 1 (CMA1), collagen type III alpha I, interleukin 2, collagen type I alpha I, collagen type I alpha II and vascular endothelial growth factor A (VEGFA). DUSP1, IL7, TNFα and VEGFA showed an initial decrease in expression during the early stages followed by an increase in expression towards the middle and late phases. IL1ß and CMA1 expression was limited to specific time points. The remaining markers either showed inconsistent expression or were undetected in our samples. The expression patterns of the detected markers suggest they have potential to predict injury age, especially during the initial stages of injury healing, if used in combination with one another.

Early cow's milk introduction is associated with failed personal-social milestones after 1 year of age.

UNLABELLED: Both the American Academy of Pediatrics (AAP) and the Institute of Medicine (IOM) recommend delaying the introduction of cow's milk until after 1 year of age due to its low absorbable iron content. We used a novel computerized decision support system to gather data from multiple general pediatrics offices. We asked families whether their child received cow's milk before 1 year of age, had a low-iron diet, or used low-iron formula. Then, at subsequent visits, we performed a modified developmental assessment using the Denver II. We assessed the effect of early cow's milk or a low-iron diet on the later failure of achieving developmental milestones. We controlled for covariates using logistic regression. Early cow's milk introduction (odds ratio (OR) 1.30, p = 0.012), as well as a low-iron diet or low-iron formula (OR 1.42, p < 0.001), was associated with increased rates of milestone failure. Only personal-social milestones (OR 1.44, p = 0.002) showed a significantly higher rate of milestone failure. Both personal-social (OR 1.42, p < 0.001) and language (OR 1.22, p = 0.009) showed higher rates of failure in children with a low-iron diet. CONCLUSIONS: There is an association between the introduction of cow's milk before 1 year of age and the rate of delayed developmental milestones after 1 year of age. This adds strength to the recommendations from the AAP and IOM to delay cow's milk introduction until after 1 year of age.

Developing an interpretation model for body fluid identification.

Criminal investigations, particularly sexual assaults, frequently require the identification of body fluid type in addition to body fluid donor to provide context. In most cases this can be achieved by conventional methods, however, in certain scenarios, alternative molecular methods are required. An example of this is the detection of menstrual fluid and vaginal material, which are not able to be identified using conventional techniques. Endpoint reverse-transcription PCR (RT-PCR) is currently used for this purpose to amplify body fluid specific messenger RNA (mRNA) transcripts in forensic casework. Real-time quantitative reverse-transcription PCR (RT-qPCR) is a similar method but utilises fluorescent markers to generate quantitative results in the form of threshold cycle (Cq) values. Despite the uncertainty surrounding body fluid identification, most interpretation guidelines utilise categorical statements. Probabilistic modelling is more realistic as it reflects biological variation as well as the known performance of the method. This research describes the application of various machine learning models to single-source mRNA profiles obtained by RT-qPCR and assesses their performance. Multinomial logistic regression (MLR), Naïve Bayes (NB), and linear discriminant analysis (LDA) were used to discriminate between the following body fluid categories: saliva, circulatory blood, menstrual fluid, vaginal material, and semen. We identified that the performance of MLR was somewhat improved when the quantitative dataset of the original Cq values was used (overall accuracy of approximately 0.95) rather than presence/absence coded data (overall accuracy of approximately 0.94). This indicates that the quantitative information obtained by RT-qPCR amplification is useful in assigning body fluid class. Of the three classification methods, MLR performed the best. When we utilised receiver operating characteristic curves to observe performance by body fluid class, it was clear that all methods found difficulty in classifying menstrual blood samples. Future work will involve the modelling of body fluid mixtures, which are common in samples analysed as part of sexual assault investigations.

Families Benefit After Utilization of a Clinic-Based Food Pantry Irrespective of Food Insecurity Experiences in a Pediatric Obesity Treatment Program.

PURPOSE: To evaluate the congruence between food insecurity screening outcome and clinic-based food pantry utilization and to examine caregiver reported comfort, motivation, and benefits of utilization. DESIGN: Mixed-methods study. SETTING: Academic pediatric obesity treatment clinic. SUBJECTS: Convenience sample of caregivers. INTERVENTION: Clinic-based food pantry offered irrespective of food insecurity screening outcome. MEASURES: Food insecurity screening (Hunger Vital Sign) and severity, self-rated caregiver health, willingness to disclose food insecurity and receive food, and food-related stress. ANALYSIS: Chi-square and t-tests were utilized to examine associations and descriptive analysis explored benefits. Rapid qualitative analysis was utilized to identify themes. RESULTS: Caregivers of 120 children were included (child mean age 11.8; 56.7% female, 67.6% Non-Hispanic Black), with 47 of 59 eligible completing follow-up surveys and 14 completing in-depth interviews. Approximately half (N = 30/59, 50.8%) of families utilizing the food pantry screened negative for food insecurity. Families utilizing the food pantry were more likely to report severe food insecurity (N = 23/59; 38.9%) compared to those declining (N = 3/61; 4.9%, P < .001). Caregivers accepting food were able to meet a child health goal (N = 30/47, 63.8%). Caregivers reported feeling comfortable receiving food (N = 13/14) and felt utilizing the food pantry led to consumption of healthier foods (N = 7/14). CONCLUSIONS: Families who screened both positive and negative for food insecurity utilized and benefited from a clinic-based food pantry. Clinics should consider strategies offering food resources to all families irrespective of screening outcome.

Pilot study of quantitative analysis of background enhancement on breast MR images: association with menstrual cycle and mammographic breast density.

PURPOSE: To perform semiautomated quantitative analysis of the background enhancement (BE) in a cohort of patients with newly diagnosed breast cancer and to correlate it with mammographic breast density and menstrual cycle. MATERIALS AND METHODS: Informed consent was waived after the research ethics board approved this study. Results of 177 consecutive preoperative breast magnetic resonance (MR) examinations performed from February to December 2009 were reviewed; 147 female patients (median age, 48 years; range, 26-86 years) were included. Ordinal values of BE and breast density were described by two independent readers by using the Breast Imaging Reporting and Data System lexicon. The BE coefficient (BEC) was calculated thus: (SI2 · 100/SI1) - 100, where SI is signal intensity, SI2 is the SI enhancement measured in the largest anteroposterior dimension in the axial plane 1 minute after the contrast agent injection, and SI1is the SI before contrast agent injection. BEC was used for the quantitative analysis of BE. Menstrual cycle status was based on the last menstrual period. The Wilcoxon rank-sum or Kruskal-Wallis test was used to compare quantitative assessment groups. Cohen weighted κ was used to evaluate agreement. RESULTS: Of 147 patients, 68 (46%) were premenopausal and 79 (54%) were postmenopausal. The quantitative BEC was associated with the menstrual status (BEC in premenopausal women, 31.48 ± 20.68 [standard deviation]; BEC in postmenopausal women, 25.65 ± 16.74; P = .02). The percentage of overall BE was higher when the MR imaging was performed in women in the inadequate phase of the cycle (<35 days, not 7-14 days; mean BEC, 35.7) compared with women in the postmenopausal group (P = .001). Premenopausal women had significantly higher BEC when compared with postmenopausal women (P = .03). There was no significant difference in the percentage of BE between breast density groups. CONCLUSION: Premenopausal women with breast cancer, and specifically women in the inadequate phase of the cycle, presented with higher quantitative BE than postmenopausal women. No association was found between BE and breast density.

One-step endpoint RT-PCR assays for confirmatory body fluid identification.

Messenger RNA (mRNA) expression analysis is increasingly used in casework, in the form of multiplex two-step reverse transcriptase PCR (RT-PCR) assays such as CellTyper 2 (CT2), developed by the Institute of Environmental Science and Research (ESR). This paper presents the development of a one-step endpoint RT-PCR workflow to improve the efficiency and precision of confirmatory body fluid identification. A comparative study of commercial one-step RT-PCR kits was undertaken, with the highest performing kit (RNA to CT) retained for further development. Sensitivity, specificity across body fluids, and precision was assessed simultaneously using receiver operating characteristic (ROC) curves. An optimal RFU cut-off value which maximised sensitivity and specificity was determined for each marker. All assays performed significantly better when compared to the equivalent of a completely uninformative test (area under the curve of 0.5) for their target body fluid. Sensitivity varied between different donors, but the limit of detectionss were estimated as follows; saliva markers HTN3: 1 in 100 dilution of a whole buccal swab and FDCSP: 1 in 10 dilution of a whole buccal swab, circulatory blood marker SLC4A1: 0.1 µL blood, menstrual fluid markers STC1, MMP10: 1 in 10 dilution of a whole menstrual swab, spermatozoa markers PRM1, TNP1: 0.1 µL semen, seminal fluid markers KLK2: 0.1 µL semen and MSMB: 0.01 µL semen, and vaginal material marker CYP2B7P: 1 in 1000 dilution of a whole vaginal swab. The method successfully detected most body fluids in a range of simple mixtures with 77 out of 80 markers observed when expected. The developed one-step endpoint RT-PCR assays lack the sensitivity and precision required for forensic casework and provide little benefit when compared with standard two-step endpoint RT-PCR, other than minimal time and cost savings, similar sensitivity, and improved precision for some markers. As both methods utilise endpoint RT-PCR, they have the same narrow linear dynamic range. The novel method is therefore similarly susceptible to varied RNA input, a major disadvantage of this approach. The limited sensitivity and precision consistently encountered with endpoint RT-PCR - regardless of cDNA synthesis strategy - could be addressed by a real-time PCR approach.

Partial validation of multiplexed real-time quantitative PCR assays for forensic body fluid identification.

Confirmatory body fluid identification using messenger RNA (mRNA) is a well-established technique to address issues encountered with conventional testing - such as poor sensitivity, specificity, and a lack of available tests for all body fluids of interest. For over a decade, endpoint reverse-transcription polymerase chain reaction (RT-PCR) assays have been used in forensic casework for such purposes. However, in comparison with real-time quantitative RT-PCR (RT-qPCR), endpoint RT-PCR has lower sensitivity, precision, and linear dynamic range. This research details the multiplexing and partial validation of confirmatory RT-qPCR assays. We have previously described novel assays for a range of body fluid targets and identified an optimal commercial kit for their amplification. Here, multiplexing was undertaken to form three assays: circulatory blood (SLC4A1) and menstrual fluid (STC1), saliva (HTN3) and vaginal material (CYP2B7P), and spermatozoa (PRM1) and seminal fluid (KLK2), all including a synthetic internal control RNA. Partial validation of the multiplexed assays incorporated the MIQE guidelines, ISO requirements, and SWGDAM guidelines. Using receiver operating characteristic (ROC) curves, each marker was significantly different from an uninformative assay and optimal cut-offs were all above 35 cycles. All assays showed a wide LDR (ranging from 3 to 5 logs with most R2 > 0.99), and high precision (most mean CV < 1 %). STC1 showed some instances of sporadic expression in blood, semen, and vaginal material at high CT values. CYP2B7P showed off-target expression in semen and blood. The sensitivities were approximated as; saliva: 1 in 1,000 dilution of a whole buccal swab, circulatory blood: 0.01-0.1 µL blood, menstrual fluid: 1 in 10,000 dilution of a whole menstrual swab, spermatozoa: 0.001 µL semen, seminal fluid: 0.01 µL semen, and vaginal material: 1 in 1,000 dilution of a whole vaginal swab. A total of 16 mock body fluid extract mixtures and 18 swab mixtures were tested and had 100% and 99% detection of target markers below each specific cut-off, respectively. Some mixtures containing high volumes of blood and semen showed off-target CYP2B7P expression. The successful application of a probabilistic model to the RT-qPCR data was also demonstrated. Further work will involve full developmental validation.

Effects of delayed callback from screening mammography due to the COVID-19 pandemic.

OBJECTIVE: To determine the frequency and distinguishing imaging characteristics of breast cancers detected on screening mammography which was initially evaluated as a probably benign lesion and the workup was delayed due to the COVID-19 pandemic. MATERIALS AND METHODS: REB-approved multicenter retrospective screening mammography studies and patient's chart review carried out between February 2020 and March 2020. According to an institutional decision, the frequency and imaging findings deemed probably benign on screening mammography after review by a breast fellowship-trained radiologist with workup deferred until after the first pandemic wave plateau in late July 2020 were recorded. Results were correlated with histopathology if tissue sample performed or an uneventful 2 years follow-up. Descriptive statistical analysis was used to describe the retrieved data set. RESULTS: Out of 1816 mammography screening between February 2020 and March 2020, 99 women, median age 58 years (range 35-84), 99 mammography had possibly benign findings with workup delayed, and two patients, age 49 and 56, had cancers (2.02%), misinterpreted as benign findings. Both malignant cases were focal asymmetries, with pathology of invasive ductal carcinoma, 12 mm and 9 mm in size. No in-situ carcinoma was detected. CONCLUSION: The low rate of cancer detected suggests that a delay callback may be a reasonable option for some likely benign findings when immediate callback is not an option, such as during a pandemic. Larger studies would be helpful to support our findings and may allow us to translate the adoption of such a model during potential future pandemic. CLINICAL RELEVANCE: The results of this study may be helpful for a future situation when delaying a call back from screening mammography is again required.

Rapid Microchip Electrophoretic Separation of Novel Transcriptomic Body Fluid Markers for Forensic Fluid Profiling.

Initial screening of criminal evidence often involves serological testing of stains of unknown composition and/or origin discovered at a crime scene to determine the tissue of origin. This testing is presumptive but critical for contextualizing the scene. Here, we describe a microfluidic approach for body fluid profiling via fluorescent electrophoretic separation of a published mRNA panel that provides unparalleled specificity and sensitivity. This centrifugal microfluidic approach expedites and automates the electrophoresis process by allowing for simple, rotationally driven flow and polymer loading through a 5 cm separation channel; with each disc containing three identical domains, multi-sample analysis is possible with a single disc and multi-sample detection per disc. The centrifugal platform enables a series of sequential unit operations (metering, mixing, aliquoting, heating, storage) to execute automated electrophoretic separation. Results show on-disc fluorescent detection and sizing of amplicons to perform comparably with a commercial 'gold standard' benchtop instrument and permitted sensitive, empirical discrimination between five distinct body fluids in less than 10 min. Notably, our microfluidic platform represents a faster, simpler method for separation of a transcriptomic panel to be used for forensically relevant body fluid identification.

A Mixed-Methods Examination of Referral Processes to Clinic-Community Partnership Programs for the Treatment of Childhood Obesity.

Background: Partnerships with community programs have been used to improve access to obesity care and address clinical barriers to childhood obesity management; however, little is known about the program referral process. The objective of this study was to identify factors that affect the referral from clinics to community-based programs. Methods: Active Recreation through Community-Healthcare Engagement Study (ARCHES) is a mixed-method, implementation study designed to test the feasibility of establishing clinic-community partnerships to treat childhood obesity. We collected clinical referral and program attendance data from the six ARCHES clinic-community partnerships and conducted semistructured interviews (n = 19) with key stakeholders. Logistic regression models were used to identify referral characteristics associated with ever attending a community program. We used deductive thematic analysis to examine contextual factors affecting the clinical referral and subsequent attendance at the community programs. Results: Patients referred from individual providers [odds ratio (OR): 3.20, 95% confidence interval (CI): 1.08-9.48], specialty clinics (OR: 2.73, 95% CI: 1.48-5.05), and community wellness clinics (OR: 3.42, 95% CI: 1.05-11.13), had greater odds of ever attending the programs compared with patients from primary care clinics. Patients referred to cohort-based programs compared with open enrollment programs had greater odds of ever attending the programs. Stakeholders emphasized the value of communication within the partnership and with patients in clinical settings. Effective provider communication with patients involved engaging and program endorsing conversations to explain the value of the program. Conclusions: We identified factors that may improve the referral process in clinic-community partnerships to provide resources to primary care providers looking to address childhood obesity. Clinical Trial Registration number: NCT03246763.

Detection of five specified menstrual fluid mRNA markers over the uterine cycle.

Matrix metallopeptidases (MMPs) 7, 10, and 11 are currently the most commonly employed messenger RNAs (mRNAs) for the identification of menstrual fluid (MF) in forensic analysis. However, no comprehensive study has been carried out to date to explore their time-dependent detection in vaginal samples. This research investigated the detection of MMPs 7, 10, and 11, as well as MMP3 and stanniocalcin 1 (STC1) over the uterine cycle. The aim was to associate relative transcript levels with cycle stages and thus determine which of these transcripts is most suitable for MF identification in a forensic context. Additionally, the effect of hormonal contraceptives (HCs) on their abundance was explored. A total of 300 vaginal swab samples were collected from eight female donors, including a pregnant woman, naturally cycling women, and contraceptive users. Differences among individuals were observed, but these were not consistent within the groups. Only MMP10 and STC1 mRNA abundance appeared to be unaffected by the use of HCs. MMP3, MMP7, and MMP11 transcripts were less abundant in MF samples of some HC users. Overall, MMP3 was most specific to MF, although this transcript was still detected in one of four vaginal material (VM) samples. STC1 was less specific than MMP3 (detected in 39.6 % of VM samples). However, over the days of menstruation, STC1 was more consistently detectable than the MMPs. MMP10 was least specific, with a 78.3 % detection rate in VM samples, but the presence/absence in VM was individual-specific and consistent. MMP10 may therefore be more useful as a VM marker with elevated abundance during menstruation in some individuals. MMP7 and MMP11 were the least reliable mRNAs for MF identification, despite an increased specificity compared to MMP10. Detection rates in MF were lower than those of MMP3 and STC1, whereas detection rates in VM were higher. MMP7 abundance additionally increased approximately 2-5 days after the end of menstruation in all donors except one naturally cycling individual. In view of these results, MMP3 and STC1 were identified as the most useful MF markers for forensic use. Nevertheless, mRNA typing results need to be interpreted with utmost caution.