Structured feedback and operative video debriefing with critical view of safety annotation in training of laparoscopic cholecystectomy: a randomized controlled study.

BACKGROUND: The learning curve in minimally invasive surgery (MIS) is lengthened compared to open surgery. It has been reported that structured feedback and training in teams of two trainees improves MIS training and MIS performance. Annotation of surgical images and videos may prove beneficial for surgical training. This study investigated whether structured feedback and video debriefing, including annotation of critical view of safety (CVS), have beneficial learning effects in a predefined, multi-modal MIS training curriculum in teams of two trainees. METHODS: This randomized-controlled single-center study included medical students without MIS experience (n = 80). The participants first completed a standardized and structured multi-modal MIS training curriculum. They were then randomly divided into two groups (n = 40 each), and four laparoscopic cholecystectomies (LCs) were performed on ex-vivo porcine livers each. Students in the intervention group received structured feedback after each LC, consisting of LC performance evaluations through tutor-trainee joint video debriefing and CVS video annotation. Performance was evaluated using global and LC-specific Objective Structured Assessments of Technical Skills (OSATS) and Global Operative Assessment of Laparoscopic Skills (GOALS) scores. RESULTS: The participants in the intervention group had higher global and LC-specific OSATS as well as global and LC-specific GOALS scores than the participants in the control group (25.5 ± 7.3 vs. 23.4 ± 5.1, p = 0.003; 47.6 ± 12.9 vs. 36 ± 12.8, p < 0.001; 17.5 ± 4.4 vs. 16 ± 3.8, p < 0.001; 6.6 ± 2.3 vs. 5.9 ± 2.1, p = 0.005). The intervention group achieved CVS more often than the control group (1. LC: 20 vs. 10 participants, p = 0.037, 2. LC: 24 vs. 8, p = 0.001, 3. LC: 31 vs. 8, p < 0.001, 4. LC: 31 vs. 10, p < 0.001). CONCLUSIONS: Structured feedback and video debriefing with CVS annotation improves CVS achievement and ex-vivo porcine LC training performance based on OSATS and GOALS scores.

Technical note: A comparison of physician doses in C-Arm and CT fluoroscopy procedures.

PURPOSE: We address the misconception that the typical physician dose is higher for CT fluoroscopy (CTF) procedures compared to C-Arm procedures. METHODS: We compare physician scatter doses using two methods: a literature review of reported doses and a model based on a modified form of the dose area product (DAP). We define this modified form of DAP, "cumulative absorbed DAP," as the product of the area of the x-ray beam striking the patient, the dose rate per unit area, and the exposure time. RESULTS: The patient entrance dose rate for C-Arm fluoroscopy (0.2 mGy/s) was found to be 15 times lower than for CT fluoroscopy (3 mGy/s). A typical beam entrance area for C-Arm fluoroscopy reported in the literature was found to be 10.6 × 10.6 cm (112 cm2), whereas for CTF was 0.75 × 32 cm (24 cm2). The absorbed DAP rate for C-Arm fluoroscopy (22 mGy*cm2/s) was found to be 3.3 times lower than for CTF (72 mGy*cm2/s). The mean fluoroscopy time for C-Arm procedures (710 s) was found to be 21 times higher than for CT fluoroscopy procedures (23 s). The cumulative absorbed DAP for C-Arm procedures was found to be 9.4 times higher when compared to CT procedures (1.59 mGy*m2 vs. 0.17 mGy*m2). CONCLUSIONS: The higher fluoroscopy time in C-Arm procedures leads to a much lower cumulative DAP (i.e., physician scatter dose) in CTF procedures. This result can inform interventional physicians deciding on whether to perform inter-procedural imaging inside the room as opposed to retreating from the room.

Plastic Food Packaging from Five Countries Contains Endocrine- and Metabolism-Disrupting Chemicals.

Plastics are complex chemical mixtures of polymers and various intentionally and nonintentionally added substances. Despite the well-established links between certain plastic chemicals (bisphenols and phthalates) and adverse health effects, the composition and toxicity of real-world mixtures of plastic chemicals are not well understood. To assess both, we analyzed the chemicals from 36 plastic food contact articles from five countries using nontarget high-resolution mass spectrometry and reporter-gene assays for four nuclear receptors that represent key components of the endocrine and metabolic system. We found that chemicals activating the pregnane X receptor (PXR), peroxisome proliferator receptor γ (PPARγ), estrogen receptor α (ERα), and inhibiting the androgen receptor (AR) are prevalent in plastic packaging. We detected up to 9936 chemical features in a single product and found that each product had a rather unique chemical fingerprint. To tackle this chemical complexity, we used stepwise partial least-squares regressions and prioritized and tentatively identified the chemical features associated with receptor activity. Our findings demonstrate that most plastic food packaging contains endocrine- and metabolism-disrupting chemicals. Since samples with fewer chemical features induce less toxicity, chemical simplification is key to producing safer plastic packaging.

Beyond the Nucleus: Plastic Chemicals Activate G Protein-Coupled Receptors.

G protein-coupled receptors (GPCRs) are central mediators of cell signaling and physiological function. Despite their biological significance, GPCRs have not been widely studied in the field of toxicology. Herein, we investigated these receptors as novel targets of plastic chemicals using a high-throughput drug screening assay with 126 human non-olfactory GPCRs. In a first-pass screen, we tested the activity of triphenol phosphate, bisphenol A, and diethyl phthalate, as well as three real-world mixtures of chemicals extracted from plastic food packaging covering all major polymer types. We found 11 GPCR-chemical interactions, of which the chemical mixtures exhibited the most robust activity at adenosine receptor 1 (ADORA1) and melatonin receptor 1 (MTNR1A). We further confirm that polyvinyl chloride and polyurethane products contain ADORA1 or MTNRA1 agonists using a confirmatory secondary screen and pharmacological knockdown experiments. Finally, an analysis of the associated gene ontology terms suggests that ADORA1 and MTNR1A activation may be linked to downstream effects on circadian and metabolic processes. This work highlights that signaling disruption caused by plastic chemicals is broader than that previously believed and demonstrates the relevance of nongenomic pathways, which have, thus far, remained unexplored.

A target containing phantom for accuracy assessment of cone-beam CT-guided histotripsy.

PURPOSE: Histotripsy is a nonionizing, noninvasive, and nonthermal focal tumor therapy. Cone-beam computed tomography (CBCT) guidance was developed for targeting tumors not visible on ultrasound. This approach assumes cavitation is formed at the geometrical focal point of the therapy transducer. In practice, the exact location might vary slightly between transducers. In this study, we present a phantom with an embedded target to evaluate CBCT-guided histotripsy accuracy and assess the completeness of treatments. METHODS: Spherical (2.8 cm) targets with alternating layers of agar and radiopaque barium were embedded in larger phantoms with similar layers. The layer geometry was designed so that targets were visible on pre-treatment CBCT scans. The actual histotripsy treatment zone was visualized via the mixing of adjacent barium and agar layers in post-treatment CBCT images. CBCT-guided histotripsy treatments of the targets were performed in six phantoms. Offsets between planned and actual treatment zones were measured and used for calibration refinement. To measure targeting accuracy after calibration refinement, six additional phantoms were treated. In a separate investigation, two groups (N = 3) of phantoms were treated to assess visualization of incomplete treatments ("undertreatment" group: 2 cm treatment within 2.8 cm tumor, "mistarget" group: 2.8 cm treatment intentionally shifted laterally). Treatment zones were segmented (3D Slicer 5.0.3), and the centroid distance between the prescribed target and actual treatment zones was quantified. RESULTS: In the calibration refinement group, a 2 mm offset in the direction of ultrasound propagation (Z) was measured. After calibration refinement, the centroid-to-centroid distance between prescribed and actual treatment volumes was 0.5 ± 0.2 mm. Average difference between the prescribed and measured treatment sizes in the incomplete treatment groups was 0.5 ± 0.7 mm. In the mistarget group, the distance between prescribed and measured shifts was 0.2 ± 0.1 mm. CONCLUSION: The proposed prototype phantom allowed for accurate measurement of treatment size and location, and the CBCT visible target provided a simple way to detect misalignments for preliminary quality assurance of CBCT-guided histotripsy.

Needle artifact reduction during interventional CT procedures using a silver filter.

BACKGROUND: MAR algorithms have not been productized in interventional imaging because they are too time-consuming. Application of a beam hardening filter can mitigate metal artifacts and doesn't increase computational burden. We evaluate the ability to reduce metal artifacts of a 0.5 mm silver (Ag) additional filter in a Multidetector Computed Tomography (MDCT) scanner during CT-guided biopsy procedures. METHODS: A biopsy needle was positioned inside the lung field of an anthropomorphic phantom (Lungman, Kyoto Kagaku, Kyoto, Japan). CT acquisitions were performed with beam energies of 100 kV, 120 kV, 135 kV, and 120 kV with the Ag filter and reconstructed using a filtered back projection algorithm. For each measurement, the CTDIvol was kept constant at 1 mGy. Quantitative profiles placed in three regions of the artifact (needle, needle tip, and trajectory artifacts) were used to obtain metrics (FWHM, FWTM, width at - 100 HU, and absolute error in HU) to evaluate the blooming artifact, artifact width, change in CT number, and artifact range. An image quality analysis was carried out through image noise measurement. A one-way analysis of variance (ANOVA) test was used to find significant differences between the conventional CT beam energies and the Ag filtered 120 kV beam. RESULTS: The 120 kV-Ag is shown to have the shortest range of artifacts compared to the other beam energies. For needle tip and trajectory artifacts, a significant reduction of - 53.6% (p < 0.001) and - 48.7% (p < 0.001) in the drop of the CT number was found, respectively, in comparison with the reference beam of 120 kV as well as a significant decrease of up to - 34.7% in the artifact width (width at - 100 HU, p < 0.001). Also, a significant reduction in the blooming artifact of - 14.2% (FWHM, p < 0.001) and - 53.3% (FWTM, p < 0.001) was found in the needle artifact. No significant changes (p > 0.05) in image noise between the conventional energies and the 120 kV-Ag were found. CONCLUSIONS: A 0.5 mm Ag additional MDCT filter demonstrated consistent metal artifact reduction generated by the biopsy needle. This reduction may lead to a better depiction of the target and surrounding structures while maintaining image quality.

Use of an uncrewed surface vehicle and near infrared hyperspectral imaging for sampling and analysis of aquatic microplastics.

Data on MP in aquatic environments have low resolution in space and time. Scaling up sampling and increasing analysis throughput are the main bottlenecks. We combined two approaches: an uncrewed surface vehicle (USV) and near infrared hyperspectral imaging (NIR-HSI) for sampling and analysis of MP > 300 µm. We collected 35 water samples over 4 d in a coastal area. Samples were analyzed using NIR-HSI and Fourier transform infrared spectroscopy (FTIR). Spiked samples were used to determine recovery. We conclude that using a USV can mitigate issues of traditional trawls like scalability, repeatability, and contamination. NIR-HSI detects more polyethylene but less polypropylene than FTIR analysis and reduces analysis time significantly. Highly variable concentrations were found at both sampling locations, with mean MP concentration of 0.28 and 0.01 MP m-3 for location A and B respectively. USV sampling in tandem with NIR-HSI is an effective analytical pipeline for MP monitoring.

[The digital operating room : Chances and risks of artificial intelligence]. / Der digitale Operationssaal : Chancen und Risiken künstlicher Intelligenz.

At the central workplace of the surgeon the digitalization of the operating room has particular consequences for the surgical work. Starting with intraoperative cross-sectional imaging and sonography, through functional imaging, minimally invasive and robot-assisted surgery up to digital surgical and anesthesiological documentation, the vast majority of operating rooms are now at least partially digitalized. The increasing digitalization of the whole process chain enables not only for the collection but also the analysis of big data. Current research focuses on artificial intelligence for the analysis of intraoperative data as the prerequisite for assistance systems that support surgical decision making or warn of risks; however, these technologies raise new ethical questions for the surgical community that affect the core of surgical work.

Comparing a virtual reality head-mounted display to on-screen three-dimensional visualization and two-dimensional computed tomography data for training in decision making in hepatic surgery: a randomized controlled study.

OBJECTIVE: Evaluation of the benefits of a virtual reality (VR) environment with a head-mounted display (HMD) for decision-making in liver surgery. BACKGROUND: Training in liver surgery involves appraising radiologic images and considering the patient's clinical information. Accurate assessment of 2D-tomography images is complex and requires considerable experience, and often the images are divorced from the clinical information. We present a comprehensive and interactive tool for visualizing operation planning data in a VR environment using a head-mounted-display and compare it to 3D visualization and 2D-tomography. METHODS: Ninety medical students were randomized into three groups (1:1:1 ratio). All participants analyzed three liver surgery patient cases with increasing difficulty. The cases were analyzed using 2D-tomography data (group "2D"), a 3D visualization on a 2D display (group "3D") or within a VR environment (group "VR"). The VR environment was displayed using the "Oculus Rift ™" HMD technology. Participants answered 11 questions on anatomy, tumor involvement and surgical decision-making and 18 evaluative questions (Likert scale). RESULTS: Sum of correct answers were significantly higher in the 3D (7.1 ± 1.4, p < 0.001) and VR (7.1 ± 1.4, p < 0.001) groups than the 2D group (5.4 ± 1.4) while there was no difference between 3D and VR (p = 0.987). Times to answer in the 3D (6:44 ± 02:22 min, p < 0.001) and VR (6:24 ± 02:43 min, p < 0.001) groups were significantly faster than the 2D group (09:13 ± 03:10 min) while there was no difference between 3D and VR (p = 0.419). The VR environment was evaluated as most useful for identification of anatomic anomalies, risk and target structures and for the transfer of anatomical and pathological information to the intraoperative situation in the questionnaire. CONCLUSIONS: A VR environment with 3D visualization using a HMD is useful as a surgical training tool to accurately and quickly determine liver anatomy and tumor involvement in surgery.

Topology observing 3D device reconstruction from continuous-sweep limited angle fluoroscopy.

BACKGROUND: Minimally invasive procedures usually require navigating a microcatheter and guidewire through endoluminal structures such as blood vessels and airways to sites of the disease. For numerous clinical applications, two-dimensional (2D) fluoroscopy is the primary modality used for real-time image guidance during navigation. However, 2D imaging can pose challenges for navigation in complex structures. Real-time 3D visualization of devices within the anatomic context could provide considerable benefits for these procedures. Continuous-sweep limited angle (CLA) fluoroscopy has recently been proposed to provide a compromise between conventional rotational 3D acquisitions and real-time fluoroscopy. PURPOSE: The purpose of this work was to develop and evaluate a noniterative 3D device reconstruction approach for CLA fluoroscopy acquisitions, which takes into account endoluminal topology to avoid impossible paths between disconnected branches. METHODS: The algorithm relies on a static 3D roadmap (RM) of vessels or airways, which may be generated from conventional cone beam CT (CBCT) acquisitions prior to navigation. The RM is converted to a graph representation describing its topology. During catheter navigation, the device is segmented from the live 2D projection images using a deep learning approach from which the centerlines are extracted. Rays from the focal spot to detector pixels representing 2D device points are identified and intersections with the RM are computed. Based on the RM graph, a subset of line segments is selected as candidates to exclude device paths through disconnected branches of the RM. Depth localization for each point along the device is then performed by finding the point closest to the previous 3D reconstruction along the candidate segments. This process is repeated as the projection angle changes for each CLA image frame. The approach was evaluated in a phantom study in which a catheter and guidewire were navigated along five pathways within a complex vessel phantom. The result was compared to static cCBCT acquisitions of the device in the final position. RESULTS: The average root mean squared 3D distance between CLA reconstruction and reference centerline was 1.87 ± 0.30 $1.87 \pm 0.30$ mm. The Euclidean distance at the device tip was 2.92 ± 2.35 $2.92 \pm 2.35$ mm. The correct pathway was identified during reconstruction in 100 % $100\%$ of frames ( n = 1475 $n=1475$ ). The percentage of 3D device points reconstructed inside the 3D roadmap was 91.83 ± 2.52 % $91.83 \pm 2.52\%$ with an average distance of 0.62 ± 0.30 $0.62 \pm 0.30$ mm between the device points outside the roadmap and the nearest point within the roadmap. CONCLUSIONS: This study demonstrates the feasibility of reconstructing curvilinear devices such as catheters and guidewires during endoluminal procedures including intravascular and transbronchial interventions using a noniterative reconstruction approach for CLA fluoroscopy. This approach could improve device navigation in cases where the structure of vessels or airways is complex and includes overlapping branches.

Motion-compensation approach for quantitative digital subtraction angiography and its effect on in-vivo blood velocity measurement.

Purpose: Quantitative monitoring of flow-altering interventions has been proposed using algorithms that quantify blood velocity from time-resolved two-dimensional angiograms. These algorithms track the movement of contrast oscillations along a vessel centerline. Vessel motion may occur relative to a statically defined vessel centerline, corrupting the blood velocity measurement. We provide a method for motion-compensated blood velocity quantification. Approach: The motion-compensation approach utilizes a vessel segmentation algorithm to perform frame-by-frame vessel registration and creates a dynamic vessel centerline that moves with the vasculature. Performance was evaluated in-vivo through comparison with manually annotated centerlines. The method was also compared to a previous uncompensated method using best- and worst-case static centerlines chosen to minimize and maximize centerline placement accuracy. Blood velocities determined through quantitative DSA (qDSA) analysis for each centerline type were compared through linear regression analysis. Results: Centerline distance errors were 0.3±0.1 mm relative to gold standard manual annotations. For the uncompensated approach, the best- and worst-case static centerlines had distance errors of 1.1±0.6 and 2.9±1.2 mm, respectively. Linear regression analysis found a high R-squared between qDSA-derived blood velocities using gold standard centerlines and motion-compensated centerlines (R2=0.97) with a slope of 1.15 and a small offset of -0.6 cm/s. The use of static centerlines resulted in low coefficients of determination for the best case (R2=0.35) and worst-case (R2=0.20) scenarios, with slopes close to zero. Conclusions: In-vivo validation of motion-compensated qDSA analysis demonstrated improved velocity quantification accuracy in vessels with motion, addressing an important clinical limitation of the current qDSA algorithm.

Improved sampling and DNA extraction procedures for microbiome analysis in food-processing environments.

Deep investigation of the microbiome of food-production and food-processing environments through whole-metagenome sequencing (WMS) can provide detailed information on the taxonomic composition and functional potential of the microbial communities that inhabit them, with huge potential benefits for environmental monitoring programs. However, certain technical challenges jeopardize the application of WMS technologies with this aim, with the most relevant one being the recovery of a sufficient amount of DNA from the frequently low-biomass samples collected from the equipment, tools and surfaces of food-processing plants. Here, we present the first complete workflow, with optimized DNA-purification methodology, to obtain high-quality WMS sequencing results from samples taken from food-production and food-processing environments and reconstruct metagenome assembled genomes (MAGs). The protocol can yield DNA loads >10 ng in >98% of samples and >500 ng in 57.1% of samples and allows the collection of, on average, 12.2 MAGs per sample (with up to 62 MAGs in a single sample) in ~1 week, including both laboratory and computational work. This markedly improves on results previously obtained in studies performing WMS of processing environments and using other protocols not specifically developed to sequence these types of sample, in which <2 MAGs per sample were obtained. The full protocol has been developed and applied in the framework of the European Union project MASTER (Microbiome applications for sustainable food systems through technologies and enterprise) in 114 food-processing facilities from different production sectors.

Bile acids regulate SF-1 to alter cholesterol balance in adrenocortical cells via S1PR2.

Glucocorticoid synthesis typically occurs in adrenal cortex and is influenced by cholesterol balance, since cholesterol is the sole precursor of steroids. Bile acids as the signaling molecules, have been shown to promote steroidogenesis in steroidogenic cells. However, whether bile acids directly regulate cholesterol balance remains elusive. In this study, we prepared cholestatic mouse models and cultured human adrenocortical cells (H295R) treated with taurochenodeoxycholic acid (TCDCA) to determine transcription levels of cholesterol metabolism associated genes and cholesterol concentrations in adrenocortical cells. Results showed that common bile duct ligation (CBDL) and chenodeoxycholic acid (CDCA) feeding elevated the mRNA levels of Abca1, Cyp51, Hmgcr, Srb1, and Mc2r in adrenals of mice. Meanwhile, the concentrations of total cholesterol and cholesteryl ester in adrenals of CBDL and CDCA-fed mice were dramatically lowered. The total and phosphorylation levels of HSL in adrenal glands of CBDL mice were also enhanced. Similarly, TCDCA treatment in H295R cells decreased intracellular concentrations of total cholesterol and cholesteryl ester and increased transcription levels of SRB1, MC2R, and HSL as well. Inhibition of bile acids' receptor sphingosine 1-phosphate receptor 2 (S1PR2), extracellular signal-regulated kinase (ERK) phosphorylation, and steroidogenic factor 1 (SF-1) respectively successfully abolished effect of TCDCA on H295R cells. SF-1s was found to be phosphorylated at Thr75 in TCDCA-treated H295R cells. While a mild increase of intracellular cAMP concentration was detected upon TCDCA treatment, inhibition of PKA activity with Rp-Isomer in H295R cells failed to decrease the expression of SF-1 and its target genes. Our findings suggest that conjugated bile acids affect cholesterol balance through regulation of SF-1 in adrenocortical cells so as to provide an adequate cholesterol supply for glucocorticoid synthesis, which improves and enriches our understanding of the mechanism whereby bile acids regulate cholesterol balance to affect adrenal function.

Exposure to microplastics affects fatty acid composition in the Japanese quail depending on sex and particle size.

Plastic ingestion in birds is a widespread phenomenon of increasing concern. However, little is known about how exposure to microplastics (MP) affects the health of birds. In other organisms, MP exposure alters lipid metabolism and composition. If also true in birds, this could affect their fitness, especially since birds heavily rely on lipids during migration and egg production. Therefore, the aim of this study was to determine if ingestion of MP (polypropylene and polyethylene collected in nature) in two size ranges, large (3 mm) and small (<125 µm), affects lipid metabolism in the Japanese quail (Coturnix japonica). We orally exposed 55 one-week-old quail during 5 weeks to a total of 600 mg of MP in sizes of either large, small, or a mixture of both. After the exposure period, females fed small MP had higher liver masses compared to control females (on average ± SD, 8.95 ± 2.3 g vs. 6.34 ± 1.0 g), while liver lipid content did not differ in either males or females. The levels of monounsaturated fatty acids were lower in females exposed to large MP and the mixture of both MP sizes compared to controls. Females exposed to MP also had different levels of oleic- (18:1) and palmitoleic (16:1) acid compared to controls dependent on MP size. Exposure to small MP increased levels of palmitic- (16:0) and palmitoleic (16:1) acid in both males and females suggesting a possible increase in de novo fatty acid synthesis. Taken together, our results point towards a sex-specific sensitivity to MP as well as size-dependent MP effects on lipid metabolism in birds. Disruption of fatty acid composition could affect important life stages in female birds, such as migration and egg-laying. We stress the importance of further research focused on determining the mechanisms of action of MP on lipid metabolism.

Toward an interoperable, intraoperative situation recognition system via process modeling, execution, and control using the standards BPMN and CMMN.

PURPOSE: For the modeling, execution, and control of complex, non-standardized intraoperative processes, a modeling language is needed that reflects the variability of interventions. As the established Business Process Model and Notation (BPMN) reaches its limits in terms of flexibility, the Case Management Model and Notation (CMMN) was considered as it addresses weakly structured processes. METHODS: To analyze the suitability of the modeling languages, BPMN and CMMN models of a Robot-Assisted Minimally Invasive Esophagectomy and Cochlea Implantation were derived and integrated into a situation recognition workflow. Test cases were used to contrast the differences and compare the advantages and disadvantages of the models concerning modeling, execution, and control. Furthermore, the impact on transferability was investigated. RESULTS: Compared to BPMN, CMMN allows flexibility for modeling intraoperative processes while remaining understandable. Although more effort and process knowledge are needed for execution and control within a situation recognition system, CMMN enables better transferability of the models and therefore the system. Concluding, CMMN should be chosen as a supplement to BPMN for flexible process parts that can only be covered insufficiently by BPMN, or otherwise as a replacement for the entire process. CONCLUSION: CMMN offers the flexibility for variable, weakly structured process parts, and is thus suitable for surgical interventions. A combination of both notations could allow optimal use of their advantages and support the transferability of the situation recognition system.

Diverse Listeria monocytogenes in-house clones are present in a dynamic frozen vegetable processing environment.

Listeria (L.) monocytogenes is of global concern for food safety as the listeriosis-causing pathogen is widely distributed in the food processing environments, where it can survive for a long time. Frozen vegetables contaminated with L. monocytogenes were recently identified as the source of two large listeriosis outbreaks in the EU and US. So far, only a few studies have investigated the occurrence and behavior of Listeria in frozen vegetables and the associated processing environment. This study investigates the occurrence of L. monocytogenes and other Listeria spp. in a frozen vegetable processing environment and in frozen vegetable products. Using whole genome sequencing (WGS), the distribution of sequence types (MLST-STs) and core genome sequence types (cgMLST-CT) of L. monocytogenes were assessed, and in-house clones were identified. Comparative genomic analyses and phenotypical characterization of the different MLST-STs and isolates were performed, including growth ability under low temperatures, as well as survival of freeze-thaw cycles. Listeria were widely disseminated in the processing environment and five in-house clones namely ST451-CT4117, ST20-CT3737, ST8-CT1349, ST8-CT6243, ST224-CT5623 were identified among L. monocytogenes isolates present in environmental swab samples. Subsequently, the identified in-house clones were also detected in product samples. Conveyor belts were a major source of contamination in the processing environment. A wide repertoire of stress resistance markers supported the colonization and survival of L. monocytogenes in the frozen vegetable processing facility. The presence of ArgB was significantly associated with in-house clones. Significant differences were also observed in the growth rate between different MLST-STs at low temperatures (4 °C and 10 °C), but not between in-house and non-in-house isolates. All isolates harbored major virulence genes such as full length InlA and InlB and LIPI-1, yet there were differences between MLST-STs in the genomic content. The results of this study demonstrate that WGS is a strong tool for tracing contamination sources and transmission routes, and for identifying in-house clones. Further research targeting the co-occurring microbiota and the presence of biofilms is needed to fully understand the mechanism of colonization and persistence in a food processing environment.

Assessment of microbial quality in poultry drinking water on farms in Austria.

The quality of poultry drinking water has a significant effect on broiler health and performance. This study conducted an analysis of aerobic mesophilic counts (AMC), Enterobacteriaceae (EB), Pseudomonadaceae (PS), and screened for the presence of Campylobacter spp. in water samples collected from a total of 14 farms in Austria, with either a public or private water source. The efficacy of two water line treatment methods was evaluated: a chemical treatment of the water lines with 4.0 ppm ClO2 (T1) and a combined chemical (4.0 ppm active ClO2 and 3.0% peracetic acid) and mechanical treatment (purging of the water lines with a high-pressure air pump; T2). However, both the T1 and T2 treatments failed to reduce the AMC counts below the maximum acceptable microbial limit of 4.0 log10 CFU/ml in water samples. In addition, no significant reduction in EB and PS counts was observed in water samples after either T1 or T2 water line treatment. The water samples showed a high level of microbial diversity with 18 to 26 different genera. The genus Pseudomonas was most frequently isolated across all poultry farms, while Campylobacter jejuni was identified in a single sample collected before water line treatment. Isolate analysis revealed the presence of opportunistic pathogens in water samples both before (T1 43.1%, T2 30.9%) and after (T1 36.3%, T2 33.3%) water line treatment. Opportunistic pathogens belonging to genera including Pseudomonas spp., Stenotrophomonas spp., and Ochrobactrum spp., were most frequently isolated from poultry drinking water. These isolates exhibited multidrug resistance and resistance phenotypes to antimicrobials commonly used in Austrian poultry farms. The findings of this study emphasize the potential risk of exposure to opportunistic pathogens for poultry and personnel, underscoring the importance of efficient water line management.