Exploring the tumor genomic landscape of aggressive prostate cancer by whole-genome sequencing of tissue or liquid biopsies.

Treatment resistance remains a major issue in aggressive prostate cancer (PC), and novel genomic biomarkers may guide better treatment selection. Circulating tumor DNA (ctDNA) can provide minimally invasive information about tumor genomes, but the genomic landscape of aggressive PC based on whole-genome sequencing (WGS) of ctDNA remains incompletely characterized. Thus, we here performed WGS of tumor tissue (n = 31) or plasma ctDNA (n = 10) from a total of 41 aggressive PC patients, including 11 hormone-naïve, 15 hormone-sensitive, and 15 castration-resistant patients. Across all variant types, we found progressively more altered tumor genomic profiles in later stages of aggressive PC. The potential driver genes most frequently affected by single-nucleotide variants or insertions/deletions included the known PC-related genes TP53, CDK12, and PTEN and the novel genes COL13A1, KCNH3, and SENP3. Etiologically, aggressive PC was associated with age-related and DNA repair-related mutational signatures. Copy number variants most frequently affected 14q11.2 and 8p21.2, where no well-recognized PC-related genes are located, and also frequently affected regions near the known PC-related genes MYC, AR, TP53, PTEN, and BRCA1. Structural variants most frequently involved not only the known PC-related genes TMPRSS2 and ERG but also the less extensively studied gene in this context, PTPRD. Finally, clinically actionable variants were detected throughout all stages of aggressive PC and in both plasma and tissue samples, emphasizing the potential clinical applicability of WGS of minimally invasive plasma samples. Overall, our study highlights the feasibility of using liquid biopsies for comprehensive genomic characterization as an alternative to tissue biopsies in advanced/aggressive PC.

Perioperative management of upper tract urothelial carcinoma in the Nordic countries.

BACKGROUND: Upper tract urothelial carcinoma (UTUC) is a rare malignancy, with typically only few new cases annually per urological department. Adherence to European association of urology (EAU) guidelines on UTUC in the Nordic countries is unknown. The objective of this survey was to examine the implementation of EAU guidelines, the perioperative management and organization of the treatment of UTUC in the Nordic countries. METHODS: The electronic survey was distributed to 93 hospitals in the Nordic countries performing radical nephroureterectomy (NU). The survey consisted of 57 main questions and data was collected between December 1st, 2021 and April 23rd, 2022. RESULTS: Overall response rate was 47/93 (67%) with a completion rate of 98%. Five out of the 6 examined subjects on diagnostic practice are applied by ≥ 72% of the participating centers. NU as treatment for high-risk UTUC is performed by 37/47 (79%), and 91% include a bladder cuff excision. CONCLUSIONS: Adherence to EAU guidelines is high on diagnostic practice in the Nordic countries, whereas disease management is less coherent.

Consumer preferences for attributes in sweet beverages and market impacts of beverage innovation.

Consumption of sugar-sweetened beverages (SSB) is considered as an important risk factor for the development of overweight and obesity in populations worldwide, with a particular focus on the risks in the younger parts of the population - children and adolescents. Together with fiscal measures and information tools, innovation-based approaches such as the development of sugar-free or sugar-reduced versions of established beverages and development of new beverage products have been used to reduce this challenge, but the effects of product innovation on sugar intake are not well understood from the literature, as previous studies have largely ignored substitution effects of product innovation in the beverage domain. The objective of the present study was to investigate the potential effectiveness of product innovation as a strategy to affect consumers' intake of energy from sweetened non-alcoholic beverages. Using household panel shopping data from approximately 3000 Danish households over the years 2006-2014, we developed a hedonic pricing approach to estimate the influence of product attributes on consumers' utility, based on observed data for Danish households' purchases of sweet drinks. Overall, the study found that beverages' degree of sweetness positively affected the satiation effect of beverage consumption and in turn made the demand for these beverages less sensitive to e.g. price changes or introduction of competing products, whereas the energy density of the beverages positively affected the demand sensitivity to market changes. Findings like these can be useful for assessing market effects as well as environmental and public health impacts of changes to the market environment.

Spatial whole transcriptome profiling of primary tumor from patients with metastatic prostate cancer.

Prostate cancer (PCa) is a highly heterogeneous disease in terms of its molecular makeup and clinical prognosis. The prostate tumor microenvironment (TME) is hypothesized to play an important role in driving disease aggressiveness, but precise mechanisms remain elusive. In our study, we used spatial transcriptomics to explore for the first time the spatial gene expression heterogeneity within primary prostate tumors from patients with metastatic disease. In total, we analyzed 5459 tissue spots from three PCa patients comprising castration-resistant (CRPC) and neuroendocrine (NEPC) disease stages. Within CRPC, we identified a T cell cluster whose activity might be impaired by nearby regulatory T cells, potentially mediating the aggressive disease phenotype. Moreover, we identified Hallmark signatures of epithelial-mesenchymal transition in a cancer-associated fibroblast (CAF) cluster, indicating the aggressive characteristic of the primary TME leading to metastatic dissemination. Within NEPC, we identified active immune-stroma cross-talk exemplified by significant ligand-receptor interactions between CAFs and M2 macrophages. Further, we identified a malignant cell population that was associated with the down-regulation of an immune-related gene signature. Lower expression of this signature was associated with higher levels of genomic instability in advanced PCa patients (SU2C cohort, n = 99) and poor recurrence free survival in early-stage PCa patients (TCGA cohort, n = 395), suggesting prognostic biomarker potential. Taken together, our study reveals the importance of whole transcriptome profiling at spatial resolution for biomarker discovery and for advancing our understanding of tumor biology.

Value of a catch-up HPV test in women aged 65 and above: A Danish population-based nonrandomized intervention study.

BACKGROUND: High-risk human papillomavirus (HPV) test is replacing cytology as the primary cervical cancer screening test due to superior sensitivity, but in most countries women ≥65 years have never had an HPV test despite they account for around 50% of cervical cancer deaths. We explored the effect of a catch-up HPV test among 65- to 69-year-old women without previous record of HPV-based screening. METHODS AND FINDINGS: This population-based nonrandomized intervention study (quasi-experimental design) included Danish women aged 65 to 69 with no record of cervical cancer screening in the last ≥5.5 years and no HPV-exit test at age 60 to 64 at the time of study inclusion. Eligible women residing in the Central Denmark Region were invited for HPV screening either by attending clinician-based sampling or requesting a vaginal self-sampling kit (intervention group, n = 11,192). Women residing in the remaining four Danish regions received standard care which was the opportunity to have a cervical cytology collected for whatever reason (reference group, n = 33,387). Main outcome measures were detection of cervical intraepithelial neoplasia (CIN) grade 2 or worse (CIN2+) per 1,000 women eligible for the screening offer and the benefit-harm ratio of the intervention and standard practice measured as the number of colposcopies needed to detect one CIN2+ case. The minimum follow-up time was 13 months for all tested women (range: 13 to 25 months). In the intervention group, 6,965 (62.2%) were screened within 12 months from the date of study inclusion and 743 (2.2%) women had a cervical cytology collected in the reference group. The CIN2+ detection was significantly higher in the intervention group (3.9, 95% confidence interval (CI): [2.9, 5.3]; p < 0.001; n = 44/11,192) as compared to the reference group (0.3, 95% CI: [0.2, 0.6]; n = 11/33,387). For the benefit-harm ratio, 11.6 (95% CI: [8.5, 15.8]; p = 0.69; n = 511/44) colposcopies were performed to detect one CIN2+ in the intervention group as compared to 10.1 (95% CI: [5.4, 18.8]; n = 111/11) colposcopies in the reference group. The study design entails a risk of confounding due to the lack of randomization. CONCLUSIONS: The higher CIN2+ detection per 1,000 eligible women in the intervention group supports that a catch-up HPV test could potentially improve cervical cancer prevention in older women. This study informs the current scientific debate as to whether women aged 65 and above should be offered a catch-up HPV test if they never had an HPV test. TRIAL REGISTRATION: ClinicalTrials.gov NCT04114968.

Prognostic Value of Low-Pass Whole Genome Sequencing of Circulating Tumor DNA in Metastatic Castration-Resistant Prostate Cancer.

BACKGROUND: Multiple treatments are available for metastatic castration-resistant prostate cancer (mCRPC), including androgen receptor signaling inhibitors (ARSI) enzalutamide and abiraterone, but therapy resistance remains a major clinical obstacle. We examined the clinical utility of low-pass whole-genome sequencing (LPWGS) of circulating tumor DNA (ctDNA) for prognostication in mCRPC. METHODS: A total of 200 plasma samples from 143 mCRPC patients collected at the start of first-line ARSI treatment (baseline) and at treatment termination (n = 57, matched) were analyzed by LPWGS (median: 0.50X) to access ctDNA% and copy number alteration (CNA) patterns. The best confirmed prostate specific antigen (PSA) response (≥50% decline [PSA50]), PSA progression-free survival (PFS), and overall survival (OS) were used as endpoints. For external validation, we used plasma LPWGS data from an independent cohort of 70 mCRPC patients receiving first-line ARSI. RESULTS: Baseline ctDNA% ranged from ≤3.0% to 73% (median: 6.6%) and CNA burden from 0% to 82% (median: 13.1%) in the discovery cohort. High ctDNA% and high CNA burden at baseline was associated with poor PSA50 response (P = 0.0123/0.0081), poor PFS (P < 0.0001), and poor OS (P < 0.0001). ctDNA% and CNA burden was higher at PSA progression than at baseline in 32.7% and 42.3% of the patients. High ctDNA% and high CNA burden at baseline was also associated with poor PFS and OS (P ≤ 0.0272) in the validation cohort. CONCLUSIONS: LPWGS of ctDNA provides clinically relevant information about the tumor genome in mCRPC patients. Using LPWGS data, we show that high ctDNA% and CNA burden at baseline is associated with short PFS and OS in 2 independent cohorts.

Changes in prevalence of Mycoplasma genitalium macrolide and fluoroquinolone resistance: notes from a 13-year retrospective survey in Dalarna County, Sweden.

OBJECTIVE: Macrolide and fluoroquinolone resistance in Mycoplasma genitalium (MG) is of emerging global concern. Compared with neighbouring countries such as Denmark, Sweden has had lower rates of macrolide resistance while fluoroquinolone resistance rates are less well documented. This study retrospectively examined macrolide, fluoroquinolone and multidrug resistance rates from Dalarna County, Sweden over a 13-year period. METHODS: MG-positive samples from 2006 to 2018 from patients examined at the Department of Venereology, Central Hospital, Falun, Sweden were tested by sequencing for macrolide resistance mutations (MRM) and fluoroquinolone resistance-associated mutations (QRAM) in the parC and gyrA subunit regions. A subset of these samples from 2006 to 2011 have been reported on previously, although only for MRM. RESULTS: Of 874 samples, 98 (11.2%, 95% CI 9.1% to 13.6%) had mutations associated with resistance to macrolides and 19 of 828 (2.3%, 95% CI 8.9% to 23.1%) to quinolones. Mutations associated with resistance to both drugs were detected in 5 of 828 (0.6%, 95% CI 0.1% to 1.4%) samples overall. A significant positive linear trend (p=0.004) for an increase in the rate of macrolide resistance was observed (from 0% in 2006 to 31% in 2018) while the increase in QRAM from 0% in 2006 to 12.3% in 2018 was not statistically significant. CONCLUSIONS: Despite a decrease in macrolide and fluoroquinolone consumption in Sweden, there was an overall increase in MG macrolide, fluoroquinolone and dual resistance from 2006 to 2018, although the difference in fluoroquinolone resistance rates was not statistically significant. In order to maintain comparably low resistance rates, resistance-guided therapy for MG infections will be crucial.

Low- versus standard- pneumoperitoneum in patients undergoing robot-assisted radical prostatectomy: a randomised, triple-blinded study.

OBJECTIVE: To investigate the effectiveness and impact of low-pressure pneumoperitoneum (Pnp) on postoperative quality of recovery (QoR) and surgical workspace (SWS) in patients with prostate cancer undergoing robot-assisted radical prostatectomy (RARP). PATIENTS AND METHODS: A randomised, triple-blinded trial was conducted in a single centre in Denmark from March 2021 to January 2022. A total of 98 patients with prostate cancer undergoing RARP were randomly assigned to either low-pressure Pnp (7 mmHg) or standard-pressure Pnp (12 mmHg). Co-primary outcomes were postoperative QoR measured via the QoR-15 questionnaire on postoperative Day 1 (POD1), POD3, POD14, and POD30, and SWS assessed intraoperatively by a blinded assessor (surgeon) via a validated SWS scale. Data analysis was performed according to the intention-to-treat principle. RESULTS: Patients who underwent RARP at low Pnp pressure demonstrated better postoperative QoR on POD1 (mean difference = 10, 95% confidence interval [CI] 4.4-15.5), but no significant differences were observed in the SWS (mean difference = 0.25, 95% CI -0.02 to 0.54). Patients allocated to low-pressure Pnp experienced statistically higher blood loss than those in the standard-pressure Pnp group (mean difference = 67 mL, P = 0.01). Domain analysis revealed significant improvements in pain (P = 0.001), physical comfort (P = 0.007), and emotional state (P = 0.006) for patients with low-pressure Pnp. This trial was registered at ClinicalTrials.gov, NCT04755452, on 16/02/2021. CONCLUSION: Performing RARP at low Pnp pressure is feasible without compromising the SWS and improves postoperative QoR, including pain, physical comfort, and emotional state, compared to the standard pressure.

Mycoplasma phocimorsus sp. nov., isolated from Scandinavian patients with seal finger or septic arthritis after contact with seals.

Seal finger (sealer's finger, spekk finger), an extremely painful hand infection contracted by individuals handling seals, has previously been associated with Mycoplasma phocacerebrale. From 2000 to 2014, six independent strains of a novel Mycoplasma species were isolated at Statens Serum Institut, Denmark, from Scandinavian patients with seal finger (M5725T, M6447, M6620, M6642 and M6879) or septic arthritis (M6921). Prior to the onset of infection, all patients had reported contact with unspeciated seals. All isolates grew within 2-5 days in Friis' modified broth and metabolized glucose and arginine but not urea. Strains M5725T, M6447, M6642 and M6921 also grew in Hayflick-type media. Colonies on agar media were large (0.5-1.0 mm) and had a typical 'fried egg' appearance, reduced tetrazolium, and were digitonin sensitive. Growth occurred at 32 °C but not at 42 °C. Strains were susceptible to doxycycline and moxifloxacin but resistant to azithromycin and erythromycin. The genomes of the six strains were sequenced and relatedness to all known Mycoplasma species was inferred. Phylogenetic analyses using 16S rRNA gene sequences and core genome single nucleotide polymorphisms showed that the isolated strains were highly similar and phylogenetically distinct from all other species within the genus Mycoplasma. The sizes of the genome sequences of the strains ranged from 744 321 to 772409 bp, with a G+C content of 25.0-25.2 mol%. Based on these analyses, we propose a novel species of the genus Mycoplasma with the name Mycoplasma phocimorsus sp. nov. with the first isolate M5725T (NCTC 14922T=DSM 116188T) as the proposed type strain and representative strains M6447, M6620, M6642, M6879 and M6921.

Treatment patterns, survival, and healthcare utilisation and costs in patients with locally advanced and metastatic bladder cancer in Denmark 2015-2020.

BACKGROUND: Treatment patterns in locally advanced and metastatic urothelial bladder cancer (La/mUBC) is changing, but little is known about current treatment patterns, survival, and costs of these patients. Our aim was to describe treatment patterns, survival, and healthcare utilisation/costs in Danish La/mUBC patients in a routine clinical care setting. METHODS: Registry-based nationwide cohort study including all bladder cancer patients aged 18 years or older with a La/mUBC tumour in the pathology register and a concomitant bladder cancer diagnosis in the Danish National Patient Registry in the period 2015-2020. We categorised the patients according to (1) La/mUBC at time of first bladder cancer diagnosis (de novo La/mUBC) and (2) non-invasive or localised muscle-invasive bladder cancer at time of diagnosis which had progressed to La/mUBC. All patients were included at date of pathology-confirmed La/mUBC. Follow-up ended 30 September 2022. RESULTS: We identified 1278 patients (69% men) with La/mUBC and no other previous cancer. Of these, 212 (17%) had de novo La/mUBC, while 1066 (83%) had progressed to La/mUBC. Median age was 72 years. Patients were followed for a median of 13.0 months (interquartile range 4.7;32.0). During follow-up, 651 (51%) patients started first-line treatment, of these, 285 progressed to second-line treatment, and 112 also started third-line treatment. Median survival was 13.0 months from La/mUBC diagnosis, 12.1 months from start of first-line treatment, 9.8 months from start of second-line treatment, and 8.6 months from start of third-line treatment. The mean number of days admitted to hospital was 3.47, 3.97, and 4.07 per month following initiation of first-line, second-line, and third-line treatment, respectively. CONCLUSION: Patients with La/mUBC have a poor prognosis, and in routine clinical care only around half of the patients received systemic anti-cancer treatment suggesting an unmet need for novel treatments. The overall costs only increased slightly from first to third-line treatment.

Energy expenditure and intensity of ritual jumping-dancing in male Maasai.

OBJECTIVES: Traditional jumping-dance rituals performed by Maasai men involve prolonged physical exertion that may contribute significantly to overall physical activity level. We aimed to objectively quantify the metabolic intensity of jumping-dance activity and assess associations with habitual physical activity and cardiorespiratory fitness (CRF). METHODS: Twenty Maasai men (18-37 years) from rural Tanzania volunteered to participate in the study. Habitual physical activity was monitored using combined heart rate (HR) and movement sensing over 3 days, and jumping-dance engagement was self-reported. A 1-h jumping-dance session resembling a traditional ritual was organized, during which participants' vertical acceleration and HR were monitored. An incremental, submaximal 8-min step test was performed to calibrate HR to physical activity energy expenditure (PAEE) and assess CRF. RESULTS: Mean (range) habitual PAEE was 60 (37-116) kJ day-1 kg-1 , and CRF was 43 (32-54) mL O2 min-1 kg-1 . The jumping-dance activity was performed at an absolute HR of 122 (83-169) beats·min-1 , and PAEE of 283 (84-484) J min-1 kg-1 or 42 (18-75)% when expressed relative to CRF. The total PAEE for the session was 17 (range 5-29) kJ kg-1 , ~28% of the daily total. Self-reported engagement in habitual jumping-dance frequency was 3.8 (1-7) sessions/week, with a total duration of 2.1 (0.5-6.0) h/session. CONCLUSIONS: Intensity during traditional jumping-dance activity was moderate, but on average sevenfold higher than habitual physical activity. These rituals are common, and can make a substantial contribution to overall physical activity in Maasai men, and thus be promoted as a culture-specific activity to increase energy expenditure and maintain good health in this population.

Similar performance after intake of carbohydrate plus whey protein and carbohydrate only in the early phase after non-exhaustive cycling.

AIM: The aim of the present study was to compare performance 5 h after a 90-min endurance training session when either carbohydrate only or carbohydrate with added whey hydrolysate or whey isolate was ingested during the first 2 h of the recovery period. METHODS: Thirteen highly trained competitive male cyclists completed three exercise and diet interventions (double-blinded, randomized, crossover design) separated by 1 week. The 90-min morning session (EX1) included a 60 min time-trial (TT60 ). Immediately and 1 h after exercise, participants ingested either (1) 1.2 g carbohydrate∙kg-1 ∙h-1 (CHO), (2) 0.8 g carbohydrate∙kg-1 ∙h-1 + 0.4 g isolate whey protein∙kg-1 ∙h-1 (ISO) or (3) 0.8 g carbohydrate∙kg-1 ∙h-1 + 0.4 g hydrolysate whey protein∙kg-1 ∙h-1 (HYD). Additional intakes were identical between interventions. After 5 h of recovery, participants completed a time-trial performance (TTP ) during which a specific amount of work was performed. Blood and urine were collected throughout the day. RESULTS: TTP did not differ significantly between dietary interventions (CHO: 43:54 ± 1:36, ISO: 46:55 ± 2:32, HYD: 44:31 ± 2:01 min). Nitrogen balance during CHO was lower than ISO (p < 0.0001) and HYD (p < 0.0001), with no difference between ISO and HYD (p = 0.317). In recovery, the area under the curve for blood glucose was higher in CHO compared to ISO and HYD. HR, VO2 , RER, glucose, and lactate during EX2 were similar between interventions. CONCLUSION: Performance did not differ after 5 h of recovery whether carbohydrate only or isocaloric carbohydrate plus protein was ingested during the first 2 h. Correspondingly, participants were not in negative nitrogen balance in any dietary intervention.

Contrast-enhanced ultrasound imaging using capacitive micromachined ultrasonic transducers.

Capacitive micromachined ultrasonic transducers (CMUTs) have a nonlinear relationship between the applied voltage and the emitted signal, which is detrimental to conventional contrast enhanced ultrasound (CEUS) techniques. Instead, a three-pulse amplitude modulation (AM) sequence has been proposed, which is not adversely affected by the nonlinearly emitted harmonics. In this paper, this is shown theoretically, and the performance of the sequence is verified using a 4.8 MHz linear capacitive micromachined ultrasonic transducer (CMUT) array, and a comparable lead zirconate titanate (PZT) array, across 6-60 V applied alternating current (AC) voltage. CEUS images of the contrast agent SonoVue flowing through a 3D printed hydrogel phantom showed an average enhancement in contrast-to-tissue ratio (CTR) between B-mode and CEUS images of 49.9 and 37.4 dB for the PZT array and CMUT, respectively. Furthermore, hydrophone recordings of the emitted signals showed that the nonlinear emissions from the CMUT did not significantly degrade the cancellation in the compounded AM signal, leaving an average of 2% of the emitted power between 26 and 60 V of AC. Thus, it is demonstrated that CMUTs are capable of CEUS imaging independent of the applied excitation voltage when using a three-pulse AM sequence.

The use of digital technologies to support home-based prehabilitation prior to major surgery: A systematic review.

BACKGROUND: Prehabilitation is a multidimensional and multidisciplinary approach, to prepare patients for surgery, with the aim of improving postoperative outcomes. Worldwide, healthcare systems are facing challenges with a gap between demand and supply of healthcare services. Telehealth is seen as a solution for delivering sustainable and efficient treatments. The effect of providing prehabilitation using telehealth solutions is unclear. OBJECTIVE: This systematic review investigated the existing literature regarding the effects and feasibility of technologies for remotely supporting home-based prehabilitation compared to standard care on pre- and postoperative outcomes in adults undergoing elective major surgery. METHODS: A literature search identified relevant studies published between 1 January 2012 and December 2022 in PubMed, PsychInfo, Cinahl and the Cochrane Library. We included RCTs, feasibility and pilot studies. The quality of studies was evaluated using Cochrane's Risk of bias assessment and by narratively rating the certainty of evidence. RESULTS: Six randomized controlled trials and 20 pilot/feasibility studies were included. The number and content of the interventions varied, depending on context and resources. Multiple approaches in the use of digital healthcare solutions were applied and the results highlight the potential of providing health services remotely. CONCLUSION: The use of technologies to support remote home-based prehabilitation in patients undergoing elective major surgery is feasible and has high acceptability, though telehealth is a broad term and wide-ranging strategies are used. Digital technologies for supporting home-based prehabilitation are likely to play an essential role in future health care as resources are scarce and innovative solutions are needed.

The retinacular ligaments of the digital extensor expansion revisited: An anatomical and biomechanical study.

Our aim was to clarify the anatomy and function of the retinacular ligaments. Forty-eight fingers were dissected and the thickness of the oblique retinacular ligament (ORL) was graded. In four fingers, the motion in the proximal interphalangeal (PIP) and distal interphalangeal (DIP) joints was subjected to an in-depth analysis before and after sectioning of either the lateral bands (LB) or the ORLs. The function of the ORLs in restricting flexion of the DIP joint with full extension of the PIP joint was measured in 10 fingers. An ORL could be identified on both sides in all 48 fingers. The distribution of the insertion on the proximal phalanx was 7/96 (7%) in the distal third, 70/96 (73%) in the middle third and 19/96 (20%) in the proximal third. Among the specimens, 29% were graded as having a strong bundle of fibers, 51% as having well-defined and regular fibers and 20% as having thin and sparse fibers. With the PIP joint extended, there was a statistically significant increase in flexion of the DIP joint after the ORL was cut. After the LB was sectioned, the ORL was able to extend the DIP joint with an extension lag of 10-22°. Cutting the ORL did not affect the ability to extend that joint. We conclude that the retinacular ligaments are consistent. Their major role is not connected with finger movement, but to provide stabilizing links between the PIP and DIP joints. They are active in not only specific, uncommon finger positions but also in ordinary use of the fingers.

Combined cone-beam CT imaging and microsurgical dissection of cadaver specimens to study cerebral venous anatomy: a technical note.

PURPOSE: Cadaver dissections and X-ray based 3D angiography are considered gold standards for studying neurovascular anatomy. We sought to develop a model that utilize the combination of both these techniques to improve current tools for anatomical research, teaching and preoperative surgical planning, particularly addressing the venous system of the brain. MATERIALS AND METHODS: Seven ethanol-fixed human cadaveric heads and one arm were injected with a latex-barium mixture into the internal jugular veins and the brachial artery. After the ethanol-based fixation, specimens were scanned by high-resolution cone-beam CT and images were post-processed on a 3D-workstation. Subsequent, microsurgical dissections were performed by an experienced neurosurgeon and venous anatomy was compared with relevant 3D venograms. RESULTS: Latex-barium mixtures resulted in a homogenous cast with filling of the cerebral venous structures down to 150 µm in diameter. The ethanol-based preparation of the cadaveric brains allowed for near-realistic microsurgical maneuverability during dissection. The model improves assessment of the venous system for anatomical education and hands-on surgical training. CONCLUSION: To our knowledge we describe the first preparation method which combines near-realistic microsurgical dissection of human heads with high-resolution 3D imaging of the cerebral venous system in the same specimens.

Combination Therapy for Mycoplasma genitalium, and New Insights Into the Utility of parC Mutant Detection to Improve Cure.

BACKGROUND: Mycoplasma genitalium (MG) infection is challenging to cure because of rising antimicrobial resistance and limited treatment options. METHODS: This was a prospective evaluation of the efficacy and tolerability of resistance-guided combination antimicrobial therapy for MG treatment at Melbourne Sexual Health Centre (August 2019-December 2020). All patients received 7 days of doxycycline before combination therapy based on the macrolide-resistant profile. Macrolide-susceptible infections received combination doxycycline + azithromycin (1 g, day 1; 500 mg, days 2-4) and macrolide-resistant infections combination doxycycline + moxifloxacin (400 mg daily for 7 days). Adherence and adverse effects were recorded at test-of-cure, recommended 14-28 days after antimicrobial completion. Sequencing was performed to determine the prevalence of single nucleotide polymorphisms (SNPs) in the parC gene and their association with moxifloxacin treatment outcomes in macrolide-resistant infections. RESULTS: Of 100 patients with macrolide-susceptible MG treated with doxycycline + azithromycin, 93 were cured (93.0%; 95% confidence interval [CI], 86.1-97.1). Of 247 patients with macrolide-resistant MG receiving doxycycline + moxifloxacin, 210 were cured (85.0%; 95% CI, 80.0-89.2). parC sequencing was available for 164 (66%) macrolide-resistant infections; 29% had SNPs at parC S83 or D87 (23% S83I). The absence of SNPs at parC S83/D87 was associated with 98.3% cure (95% CI, 93.9-99.8) following doxycycline + moxifloxacin. The presence of the parC S83I-SNP was associated with failure in 62.5% (95% CI, 45.8-77.3). Side effects were common (40%-46%) and predominantly mild and gastrointestinal. CONCLUSIONS: Combination doxycycline + azithromycin achieved high cure for macrolide-susceptible infections. However, in the context of a high prevalence of the parC S83I mutation (23%) in macrolide-resistant infections, doxycycline + moxifloxacin cured only 85%. Infections that were wild-type for S83/D87 experienced high cure following doxycycline + moxifloxacin, supporting the use of a parC-resistance/susceptibility testing strategy in clinical care.

Clinical Importance of Superior Sensitivity of the Aptima TMA-Based Assays for Mycoplasma genitalium Detection.

Mycoplasma genitalium (MG) is a common cause of nongonococcal cervicitis and urethritis. We investigated the demographic and clinical characteristics of patients tested in Denmark with the Conformité Européenne (CE)/in vitro diagnostics (IVD) Aptima Mycoplasma genitalium assay (CE/IVD AMG; Hologic) and examined the clinical significance of the higher sensitivity of the TMA-based MG assays. From March to June 2016, urogenital and extragenital specimens from consecutive attendees at a sexually transmitted infection clinic in Copenhagen, Denmark were tested with the CE/IVD AMG assay (TMA-based), the research-use-only MG Alt TMA-1 assay (Hologic), a laboratory-developed TaqMan mgpB quantitative real-time PCR (qPCR), and the Aptima Combo 2 (CT/NG; Hologic). Demographic characteristics and clinical symptoms were collected from the patient records. There were 1,245 patients included in the study. The MG prevalence among female subjects was 9.4%, and the MG prevalence among male subjects was 8.7%. Compared to the TMA-based assays, the sensitivity of the PCR-based MG assay was 64.52%, and 55 specimens from 48 individuals were missed in the mgpB qPCR. Of these, 26 individuals (54.2%) were symptomatic, whereas, among 64 individuals with concordant results, 30 individuals (46.9%) were symptomatic; no statistically significant difference was found between the groups (P = 0.567). The improved sensitivity of the TMA-based assays resulted in diagnoses of more patients with clinically relevant symptoms for which antibiotic treatment is indicated. However, approximately half of the MG-infected patients reported no symptoms, and future research is needed to investigate the pros and cons of diagnosing and treating MG in asymptomatic subjects.

Challenges of in vitro propagation and antimicrobial susceptibility testing of Mycoplasma genitalium.

The sexually transmitted bacterial pathogen Mycoplasma genitalium has proved a complex organism to work with in the laboratory setting. Exhibiting an extremely fastidious nature, successful in vitro propagation of M. genitalium has remained elusive for many researchers. Antimicrobial resistance to both first- and second-line recommended therapies (macrolides and fluoroquinolones, respectively) is commonly reported. However, phenotypic susceptibility testing is not routinely performed, due to the difficulties of in vitro growth. Instead, molecular detection of known resistance determinants is used to infer susceptibility/resistance. However, associations between determinant detection and clinical treatment failure are not always clear. Furthermore, molecular assays have limited use for detection of emerging resistance mechanisms. The present review collates and discusses the development of successful culture systems for initial isolation of this organism and current methodologies employed for phenotypic susceptibility testing to aid researchers in this field. As with Neisseria gonorrhoeae, future treatment options are extremely limited for M. genitalium and, if this sexually transmitted infection is to remain treatable, phenotypic susceptibility testing will play an invaluable role in evaluation of potential therapeutics. As such, retainment of these techniques is imperative.

Phenotypic and genotypic antimicrobial susceptibility patterns of the emerging human respiratory pathogen Mycoplasma amphoriforme isolated from the UK and Denmark.

OBJECTIVES: To determine the phenotypic and genotypic antibiotic susceptibility of Mycoplasma amphoriforme isolates recovered from patients in the UK and Denmark. METHODS: Seven isolates of M. amphoriforme were examined for antimicrobial susceptibility to seven antibiotics using the microbroth dilution assay in line with the CLSI guidelines for mycoplasmas. Each isolate was additionally subjected to WGS to identify resistance-associated mutations. Based on the consensus sequences from the genomic data, PCR primers were designed, and tested, for the amplification of the QRDR within the parC gene. RESULTS: Of the seven isolates investigated, four (57%) were resistant to moxifloxacin (0.5-1 mg/L) and levofloxacin (1-2 mg/L), compared with those that were susceptible (0.03-0.06 and 0.006 mg/L, respectively). Isolate H29 was resistant to five of the seven antibiotics tested: moxifloxacin, 0.5 mg/L; levofloxacin, 2 mg/L; azithromycin, 64 mg/L; erythromycin, 128 mg/L; and clindamycin, 64 mg/L. All isolates were susceptible to tetracycline (0.06 mg/L) and lefamulin (0.001-0.004 mg/L). Mutations from genomic data confirmed the presence of an S89F mutation within the ParC protein among all fluoroquinolone-resistant isolates and an A2059G mutation in the 23S rRNA gene in the macrolide- and lincosamide-resistant isolate H29. CONCLUSIONS: To the best of our knowledge, this is the first time where phenotypic and genotypic resistance data have been paired for M. amphoriforme confirming a correlation between the two. These data suggest the need for focused testing and resistance determination of isolates from high-risk patients given the backdrop of a high prevalence of antimicrobial resistance.