Multimodal Biomarkers That Predict the Presence of Gleason Pattern 4: Potential Impact for Active Surveillance.

PURPOSE: Latent grade group ≥2 prostate cancer can impact the performance of active surveillance protocols. To date, molecular biomarkers for active surveillance have relied solely on RNA or protein. We trained and independently validated multimodal (mRNA abundance, DNA methylation, and/or DNA copy number) biomarkers that more accurately separate grade group 1 from grade group ≥2 cancers. MATERIALS AND METHODS: Low- and intermediate-risk prostate cancer patients were assigned to training (n=333) and validation (n=202) cohorts. We profiled the abundance of 342 mRNAs, 100 DNA copy number alteration loci, and 14 hypermethylation sites at 2 locations per tumor. Using the training cohort with cross-validation, we evaluated methods for training classifiers of pathological grade group ≥2 in centrally reviewed radical prostatectomies. We trained 2 distinct classifiers, PRONTO-e and PRONTO-m, and validated them in an independent radical prostatectomy cohort. RESULTS: PRONTO-e comprises 353 mRNA and copy number alteration features. PRONTO-m includes 94 clinical, mRNAs, copy number alterations, and methylation features at 14 and 12 loci, respectively. In independent validation, PRONTO-e and PRONTO-m predicted grade group ≥2 with respective true-positive rates of 0.81 and 0.76, and false-positive rates of 0.43 and 0.26. Both classifiers were resistant to sampling error and identified more upgrading cases than a well-validated presurgical risk calculator, CAPRA (Cancer of the Prostate Risk Assessment; P < .001). CONCLUSIONS: Two grade group classifiers with superior accuracy were developed by incorporating RNA and DNA features and validated in an independent cohort. Upon further validation in biopsy samples, classifiers with these performance characteristics could refine selection of men for active surveillance, extending their treatment-free survival and intervals between surveillance.

Three-dimensional tumor visualization of invasive breast carcinomas using whole-mount serial section histopathology: implications for tumor size assessment.

PURPOSE: Linear tumor size (T-size) estimated with conventional histology informs breast cancer management. Previously we demonstrated significant differences in margin and focality estimates using conventional histology versus digital whole-mount serial sections (WMSS). Using WMSS we can measure T-size or volume. Here, we compare WMSS T-size with volume, and with T-size measured conventionally. We also compare the ellipsoid model for calculating tumor volume to direct, WMSS measurement. METHODS: Two pathologists contoured regions of invasive carcinoma and measured T-size from both WMSS and (simulated) conventional sections in 55 consecutive lumpectomy specimens. Volume was measured directly from the contours. Measurements were compared using the paired t-test or Spearman's rank-order correlation. A five-point 'border index' was devised and assigned to each case to parametrize tumor shape considering 'compactness' or cellularity. Tumor volumes calculated assuming ellipsoid geometry were compared with direct, WMSS measurements. RESULTS: WMSS reported significantly larger T-size than conventional histology in the majority of cases [61.8%, 34/55; means = (2.34 cm; 1.99 cm), p < 0.001], with a 16.4% (9/55) rate of 'upstaging'. The majority of discordances were due to undersampling. T-size and volume were strongly correlated (r = 0.838, p < 0.001). Significantly lower volume was obtained with WMSS versus ellipsoid modeling [means = (1.18 cm3; 1.45 cm3), p < 0.001]. CONCLUSIONS: Significantly larger T-size is measured with WMSS than conventionally, due primarily to undersampling in the latter. Volume and linear size are highly correlated. Diffuse tumors interspersed with normal or non-invasive elements may be sampled less extensively than more localized masses. The ellipsoid model overestimates tumor volume.

Cost-effectiveness analysis of whole-mount pathology processing for patients with early breast cancer undergoing breast conservation.

BACKGROUND: Obtaining accurate histopathologic detail for breast lumpectomy specimens is challenging because of sampling and loss of three-dimensional conformational features with conventional processing. The whole-mount (wm) technique is a novel method of serial pathologic sectioning designed to optimize cross-sectional visualization of resected specimens and determination of margin status. METHODS: Using a Markov chain cohort simulation cost-effectiveness model, we compared conventional processing with wm technique for breast lumpectomies. Cost-effectiveness was evaluated from the perspective of the Canadian health care system and compared using incremental cost-effectiveness ratios (icers) for cost per quality-adjusted life-year (qaly) over a 10-year time horizon. Deterministic and probabilistic sensitivity analyses were performed to test the robustness of the model with willingness-to-pay (wtp) thresholds of $0-$100,000. Costs are reported in adjusted 2014 Canadian dollars, discounted at a rate of 3%. RESULTS: Compared with conventional processing, wm processing is more costly ($19,989 vs. $18,427) but generates 0.03 more qalys over 10 years. The icer is $45,414, indicating that this additional amount is required for each additional qaly obtained. The model was robust to all variance in parameters, with the prevalence of positive margins accounting for most of the model's variability. CONCLUSIONS: After a wtp threshold of $45,414, wm processing becomes cost-effective and ultimately generates fewer recurrences and marginally more qalys over time. Excellent baseline outcomes for the current treatment of breast cancer mean that incremental differences in survival are small. However, the overall benefit of the wm technique should be considered in the context of achieving improved accuracy and not just enhancements in clinical effectiveness.

Diazinon resistance, fluctuating asymmetry and fitness in the Australian sheep blowfly, lucilia cuprina.

Genetic evidence suggests that the evolution of resistance to the insecticide diazinon in Lucilia cuprina initially produced an increase in asymmetry. At that time resistant flies were presumed to be at a selective disadvantage in the absence of diazinon. Subsequent evolution in natural populations selected modifiers to ameliorate these effects. The fitness and fluctuating asymmetry levels of resistant flies are currently similar to those of susceptibles. Previous genetic analyses have shown the fitness modifier to co-segregate with the region of chromosome III marked by the white eyes, w, locus, unlinked to the diazinon resistance locus, Rop-1, on chromosome IV. This study maps the asymmetry modifier to the same region, shows, as in the case of the fitness modifier, its effect to be dominant and presents data consistent with the fitness/asymmetry modifier being the same gene (gene complex). These results suggest changes in fluctuating asymmetry reflect changes in fitness.

Wings and bristles: character specificity of the asymmetry phenotype in insecticide-resistant strains of Lucilia cuprina.

We investigated the hypothesis that observed higher levels of asymmetry displayed by insecticide-resistance genotypes of Lucilia cuprina are restricted to bristle characters, due to the action of resistance genes in bristle cell development, rather than through the disruption of genomic coadaptation. We compared the level of asymmetry of three bristle characters and three wing characters in non-modified and modified-resistance genotypes. Consistent with previous studies, resistance genotypes displayed greater levels of bristle asymmetry than either susceptible or modified genotypes. However, there were no differences among genotypes for any of the wing characters. To confirm that this result is attributable to the action of the resistance and modifier genes themselves, we also examined the responses of both bristle and wing characters to the more general developmental stress of extreme temperature. Sub-optimal temperature was shown to increase both bristle and wing asymmetry, suggesting that there are no underlying differences between the two character types which could, of themselves, explain the differential response observed in the resistance genotypes.

Randomized controlled trial of vasodilator therapy after myocardial infarction.

Hemodynamic changes and mortality and morbidity were compared in a randomized controlled trial of sodium nitroprusside after acute myocardial infarction. Fifty patients with a mean pulmonary capillary wedge pressure of more than 20 mm Hg within 24 hours of acute infarction were randomly assigned to one of two groups: 25 patients treated with nitroprusside and 25 treated with furosemide. Nitroprusside rapidly produced a sustained decrease in systemic vascular resistance and increase in cardiac index (thermodilution). After 1 hour the cardiac index had increased 16 +/- 3 (mean +/- standard error of the mean) percent (p less than 0.001) compared with a decrease of 7 +/- 3 percent with administration of furosemide (p less than 0.01). Differences in systemic vascular resistance and cardiac index in the two groups persisted throughout the 48 hour treatment period (p less than 0.001). Pulmonary capillary wedge pressure decreased rapidly with nitroprusside and slowly with furosemide so that, although it was significantly lower in the former group overall (p less than 0.001), by 48 hours the values were not different. Although beneficial acute hemodynamic effects of nitroprusside were demonstrated, there was no difference in mortality or in morbidity assessed clinically, by chest X-ray film, echocardiogram or graded treadmill stress testing after 6 months or 1 year.

Efficacy of flecainide for the reversion of acute onset atrial fibrillation.

The efficacy and safety of intravenous flecainide to convert recent-onset atrial fibrillation (AF) (present for greater than or equal to 30 minutes and less than or equal to 72 hours and a ventricular response greater than or equal to 120 beats/min) was investigated. A total of 102 patients without severe heart or circulatory failure were randomized to receive either intravenous flecainide (2 mg/kg, maximum dose 150 mg; 51 patients) or placebo (51 patients) in a double-blind trial. Digoxin (500 micrograms intravenously) was administered to all patients who had not previously been receiving digoxin. The electrocardiogram was monitored continuously during the study. In 29 (57%) patients stable sinus rhythm was restored within 1 hour after flecainide and in only 7 (14%) given placebo (chi square 18.9; p = 0.000013; odds ratio 8.3; 95% confidence interval 2.9-24.8). Reversion to sinus rhythm within 1 hour after starting the trial medication was considered a pretrial end point and likely to be due to a drug effect. At the end of the 6-hour monitoring period, 34 patients (67%) in the flecainide group were in sinus rhythm whereas only 18 (35%) in the placebo group had reverted (chi square 8.83, p = 0.003; odds ratio 3.67; 95% confidence interval 1.5-9.1). Significant hypotension, although short lived, was more common in the flecainide group. One patient given flecainide developed torsades de pointes and was successfully electrically cardioverted. Flecainide is useful for the management of recent-onset AF both for control of the ventricular response and conversion to sinus rhythm.(ABSTRACT TRUNCATED AT 250 WORDS)

Reversion of recent-onset atrial fibrillation to sinus rhythm by intravenous flecainide.

Spontaneous reversion to sinus rhythm is a frequent occurrence in recent-onset atrial fibrillation (AF). In a randomized, double-blind, controlled study, intravenous flecainide (2 mg/kg, maximum dose 150 mg) was compared with placebo in the treatment of recent-onset AF (present for greater than or equal to 30 minutes and less than or equal to 72 hours' duration and a ventricular response greater than or equal to 120 beats/min). Intravenous digoxin (500 micrograms) was administered concurrently to all patients in both groups who had not previously taken digoxin. The trial medication was administered over 30 minutes. Exclusion criteria included hemodynamic instability, severe heart failure, recent antiarrhythmic therapy, hypokalemia and pacemaker dependence. One hundred two consecutive patients with recent-onset AF were enrolled in the study. All patients underwent continuous electrocardiographic monitoring in the intensive care or coronary care unit. Twenty-nine (57%) patients given flecainide and digoxin, but only 7 (14%) given placebo and digoxin, reverted to sinus rhythm in less than or equal to 1 hour after starting the trial medication infusion and remained in stable sinus rhythm (chi-square 18.9, p = 0.000013; odds ratio 8.3, 95% confidence interval 2.9 to 24.8). At the end of the 6-hour monitoring period, 34 patients (67%) in the flecainide-digoxin group were in stable sinus rhythm, whereas only 18 patients (35%) in the placebo-digoxin group had reverted (chi-square 8.83, p = 0.003; odds ratio 3.67, 95% confidence interval 1.5 to 9.1).(ABSTRACT TRUNCATED AT 250 WORDS)

Proximally delivered dilute heparin does not improve circuit life in continuous venovenous haemodiafiltration.

OBJECTIVE: To assess the effect on circuit life in continuous venovenous haemodiafiltration (CVVHD) by manipulating heparin dilution and point of administration. DESIGN: Repeated crossover design. Cases were randomised for first circuit and heparin dilution, after which crossovers occurred until treatment was stopped. SETTING: A 24-bed combined general and surgical intensive care unit admitting 1900 patients a year. On average, 54 cases a year receive CVVHD. PATIENTS: 26 critically ill adult patients requiring CVVHD were enrolled, 18 of whom used at least one standard circuit and one modified circuit. INTERVENTIONS: Two circuit configurations and heparin dilutions were compared. In combination A, standard CVVHD blood lines and heparin concentration (100 units/ml) were used. In combination B, heparin was delivered in a more dilute volume (10 units/ml) via a modified circuit design with an administration port immediately adjacent to the venous access. MEASUREMENTS AND RESULTS: 18 randomised crossovers of circuits A and B occurred. Mean/median circuit life for the standard heparin/circuit combination A was 20.1/17.5 (SD 14.6) and for the modified combination B 21.4/15.4 (SD 19.2). There was no significant difference between circuits (paired t-test, p = 0.8175). To identify other factors which could have influenced circuit life (platelet count, heparin dose and pre- and post-filter activated partial thromboplastin time, APTT) all circuits terminated for the reasons identified (n = 105) were analysed using linear modelling. Survival analysis was used to determine the survival function of the circuit. Pre-heparin APTT was the only factor associated with an increase in filter life (p = 0.035). The hazard rate for filter failure was 0.049/h (95% confidence interval 0.04 to 0.06), the range of time until filters failed was 1.8 to 78.5 h. CONCLUSIONS: Proximally administered dilute heparin is not associated with a significant increase in circuit life.

Fluctuating asymmetry of invertebrate populations as a biological indicator of environmental quality.

Fluctuating asymmetry (FA) of populations of the shrimp, Palaemon elegans, and the blood worm, Chironomus salinarius, was used to assess the environmental impact of a fertilizer manufacturing facility on the surrounding marine and freshwater ecosystems, respectively. Populations of both species were found to have elevated levels of asymmetry compared with control populations, indicating that the facility was having a significant impact on organism development. The use of FA of invertebrate populations as a simple, sensitive and cost-effective biomonitoring system for the detection of environmental stress is discussed.

The use of irradiated embryos of Chrysomya bezziana for the detection of the Old World screw-worm fly.

Irradiation of Chrysomya bezziana embryos 1 h before hatching with doses less than or equal to 7 kilorad (kr) had a significant effect on percentage egg hatch, weights and survival of larvae. Doses greater than or equal to 1 kr allowed larval development to the end of the 3rd instar stage in vitro, but prevented normal pupal development. Cattle with wounds infested with 1st instar larvae derived from irradiated embryos had 3rd instar larvae present after 3 d but these failed to pupate. Thus it would be feasible to use such larvae for wound infestation for the enhanced detection of screw-worm fly in areas where the release of fertile flies is undesirable.

3D Pathology Volumetric Technique: A Method for Calculating Breast Tumour Volume from Whole-Mount Serial Section Images.

Tumour size, most commonly measured by maximum linear extent, remains a strong predictor of survival in breast cancer. Tumour volume, proportional to the number of tumour cells, may be a more accurate surrogate for size. We describe a novel "3D pathology volumetric technique" for lumpectomies and compare it with 2D measurements. Volume renderings and total tumour volume are computed from digitized whole-mount serial sections using custom software tools. Results are presented for two lumpectomy specimens selected for tumour features which may challenge accurate measurement of tumour burden with conventional, sampling-based pathology: (1) an infiltrative pattern admixed with normal breast elements; (2) a localized invasive mass separated from the in situ component by benign tissue. Spatial relationships between key features (tumour foci, close or involved margins) are clearly visualized in volume renderings. Invasive tumour burden can be underestimated using conventional pathology, compared to the volumetric technique (infiltrative pattern: 30% underestimation; localized mass: 3% underestimation for invasive tumour, 44% for in situ component). Tumour volume approximated from 2D measurements (i.e., maximum linear extent), assuming elliptical geometry, was seen to overestimate volume compared to the 3D volumetric calculation (by a factor of 7x for the infiltrative pattern; 1.5x for the localized invasive mass).

Whole-specimen histopathology: a method to produce whole-mount breast serial sections for 3-D digital histopathology imaging.

AIMS: To develop a method for preparing diagnostic-quality, whole-mount serial sections of breast specimens while preserving 3-D conformation. This required supporting the fresh specimen prior to breadloafing and refining the conventional tissue processing method. The overall goal is to use digital images of whole-specimen histopathology to improve the estimation of extent of disease. METHODS AND RESULTS: To maintain a 3-D conformation, the specimen is suspended in 3.5% agar at 55 degrees C. The block is sliced at 5-mm intervals. Sectioning is performed after extended fixation in 4% formaldehyde from paraformaldehyde in 0.1 m Millonig's buffer, followed by paraffin processing using a non-routine schedule and extended paraffin infiltration. Whole-mount serial breast sections are produced with features of equal or superior quality to that which can be achieved using conventional methods. The method is compatible with some immunohistochemical stains but requires further optimization for others. CONCLUSIONS: The technique is currently suitable for research applications. With the reduction in processing time achievable with microwave-assisted processing, there is the potential for its use as a routine clinical method. This tool may improve the accuracy of margin estimates and identification of multifocality in breast cancer; further evaluation is necessary.

Tracing the geographic origin of the cosmopolitan parthenogenetic insect pest Liposcelis bostrychophila (Psocoptera: Liposcelididae).

The randomly amplified polymorphic DNA technique was used to trace the geographic origin of Liposcelis bostrychophila Badonnel populations in Australia from unknown geographic sources internationally. Haplotype (or clonal) diversity was high, with 474 unique haplotypes found from 616 individuals genotyped. Gene diversity estimates (0.10-0.28) and percent polymorphic loci (38.1-88.1%) were moderate to high for most populations. This resulted in genetic distance estimates that ranged from 0.04 to 0.26 and were significantly different for most pairwise population combinations. G ST values for all populations were also moderate (0.04-0.54) and again were significantly different for most pairwise population comparisons. Analysis of molecular variance revealed that the majority of variation was apportioned among individuals within populations regardless of the level at which they were grouped. Gene flow (Nm) was mostly low for all pairwise populations comparisons with an average Nm=1.8. A non-significant negative correlation between genetic distance and geographic distance was found for worldwide populations. In contrast, within Australian populations a significant positive correlation between genetic distance and geographic distance was detected. Genetic relationships explored using unweighted pair group method analysis and non-metric multidimensional scaling indicated a mixed pattern of genetic similarities among all populations. Multiple introductions, from a wide range of international source populations, have obscured the ability to accurately determine the geographic origin of L. bostrychophila in Australia.

Autonomic manifestations of tetanus.

Two patients with tetanus are reported to illustrate the involvement of the sympathetic and parasympathetic nervous system respectively. Aspects of the autonomic manifestations of tetanus are illustrated and discussed.

The Training/Examination Programme in Intensive Care, Australian and New Zealand College of Anaesthetists. 1. Training.

Unlike training for programmes in other countries which have published details of training programmes for Intensive Care Medicine, the training programme of the Australian and New Zealand College of Anaesthetists does not require certification in a primary specialty, although it is possible to combine training in Intensive Care and Anaesthetics. The lynchpin of the programme is the requirement that training can be undertaken in recognised posts in intensive care units which are approved by the College. Approval of the Unit requires evidence of appropriate supervision and teaching of trainees, sufficient number of admissions with a casemix suitable for the trainees' learning needs and an adequate level of staff and equipment. The Units are assessed by physical inspection by assessors appointed by the College. The programme includes a Final Examination in Intensive Care.