Lights, fluorescence, action-Influencing wound treatment plans including debridement of bacteria and biofilms.

High bacterial loads within chronic wounds increase the risk of infection and complication. Detection and localization of bacterial loads through point-of-care fluorescence (FL) imaging can objectively inform and support bacterial treatment decisions. This single time-point, retrospective analysis describes the treatment decisions made on 1000 chronic wounds (DFUs, VLUs, PIs, surgical wounds, burns, and others) at 211 wound-care facilities across 36 US states. Clinical assessment findings and treatment plans derived from them, as well as subsequent FL-imaging (MolecuLight®) findings and any associated treatment plan changes, were recorded for analysis. FL signals indicating elevated bacterial loads were observed in 701 wounds (70.8%), while only 293 (29.6%) showed signs/symptoms of infection. After FL-imaging, treatment plans changed in 528 wounds as follows: more extensive debridement (18.7%), more extensive hygiene (17.2%), FL-targeted debridement (17.2%), new topical therapies (10.1%), new systemic antibiotic prescriptions (9.0%), FL-guided sampling for microbiological analysis (6.2%), and changes in dressing selection (3.2%). These real-world findings of asymptomatic bacterial load/biofilm incidence, and of the frequent treatment plan changes post-imaging, are in accordance with clinical trial findings using this technology. These data, from a range of wound types, facilities, and clinician skill sets, suggest that point-of-care FL-imaging information improves bacterial infection management.

Workflow Integration of Research AI Tools into a Hospital Radiology Rapid Prototyping Environment.

The field of artificial intelligence (AI) in medical imaging is undergoing explosive growth, and Radiology is a prime target for innovation. The American College of Radiology Data Science Institute has identified more than 240 specific use cases where AI could be used to improve clinical practice. In this context, thousands of potential methods are developed by research labs and industry innovators. Deploying AI tools within a clinical enterprise, even on limited retrospective evaluation, is complicated by security and privacy concerns. Thus, innovation must be weighed against the substantive resources required for local clinical evaluation. To reduce barriers to AI validation while maintaining rigorous security and privacy standards, we developed the AI Imaging Incubator. The AI Imaging Incubator serves as a DICOM storage destination within a clinical enterprise where images can be directed for novel research evaluation under Institutional Review Board approval. AI Imaging Incubator is controlled by a secure HIPAA-compliant front end and provides access to a menu of AI procedures captured within network-isolated containers. Results are served via a secure website that supports research and clinical data formats. Deployment of new AI approaches within this system is streamlined through a standardized application programming interface. This manuscript presents case studies of the AI Imaging Incubator applied to randomizing lung biopsies on chest CT, liver fat assessment on abdomen CT, and brain volumetry on head MRI.

Advances in cocaine signature methodology: Alkaloid and isotope profiles of coca grown in Puno, Peru.

Drug Enforcement Administration's (DEA) Cocaine Signature Program previously identified 19 coca-growing regions within South America and developed methodology to geo-source cocaine using a combination of trace cocaine alkaloids, stable isotopes, and multivariate statistics. Twenty-nine coca leaf samples collected in 2016 and 2019 from a previously unanalyzed coca-growing region located in Puno, Peru, were analyzed with this methodology. Trace cocaine alkaloids and stable isotopes were compared with other Peruvian regions. Minor differences were observed in the extracted cocaine alkaloid profiles when compared with samples collected from the Ucayali-Huallaga Valley and Cusco-Apurimac regions while the stable isotopes of δ2 H (-177.1‰) and δ18 O (23.8‰) were enriched. Puno's alkaloid and stable isotope results are presented in this publication to assist forensic laboratories and enhance their cocaine geo-sourcing capabilities.

Detection of bacterial fluorescence from in vivo wound biofilms using a point-of-care fluorescence imaging device.

Wound biofilms must be identified to target disruption and bacterial eradication but are challenging to detect with standard clinical assessment. This study tested whether bacterial fluorescence imaging could detect porphyrin-producing bacteria within a biofilm using well-established in vivo models. Mouse wounds were inoculated on Day 0 with planktonic bacteria (n = 39, porphyrin-producing and non-porphyrin-producing species, 107  colony forming units (CFU)/wound) or with polymicrobial biofilms (n = 16, 3 biofilms per mouse, each with 1:1:1 parts Staphylococcus aureus/Escherichia coli/Enterobacter cloacae, 107  CFU/biofilm) that were grown in vitro. Mouse wounds inoculated with biofilm underwent fluorescence imaging up to Day 4 or 5. Wounds were then excised and sent for microbiological analysis. Bacteria-matrix interaction was assessed with scanning electron microscopy (SEM) and histopathology. A total of 48 hours after inoculation with planktonic bacteria or biofilm, red fluorescence was readily detected in wounds; red fluorescence intensified up to Day 4. Red fluorescence from biofilms persisted in excised wound tissue post-wash. SEM and histopathology confirmed bacteria-matrix interaction. This pre-clinical study is the first to demonstrate the fluorescence detection of bacterial biofilm in vivo using a point-of-care wound imaging device. These findings have implications for clinicians targeting biofilm and may facilitate improved visualisation and removal of biofilms.

An ErbB2 splice variant lacking exon 16 drives lung carcinoma.

Lung cancer causes more deaths annually than any other malignancy. A subset of non-small cell lung cancer (NSCLC) is driven by amplification and overexpression or activating mutation of the receptor tyrosine kinase (RTK) ERBB2 In some contexts, notably breast cancer, alternative splicing of ERBB2 causes skipping of exon 16, leading to the expression of an oncogenic ERBB2 isoform (ERBB2ΔEx16) that forms constitutively active homodimers. However, the broader implications of ERBB2 alternative splicing in human cancers have not been explored. Here, we have used genomic and transcriptomic analysis to identify elevated ERBB2ΔEx16 expression in a subset of NSCLC cases, as well as splicing site mutations facilitating exon 16 skipping and deletions of exon 16 in a subset of these lung tumors and in a number of other carcinomas. Supporting the potential of ERBB2ΔEx16 as a lung cancer driver, its expression transformed immortalized lung epithelial cells while a transgenic model featuring inducible ERBB2ΔEx16 specifically in the lung epithelium rapidly developed lung adenocarcinomas following transgene induction. Collectively, these observations indicate that ERBB2ΔEx16 is a lung cancer oncogene with potential clinical importance for a proportion of patients.

4E-BP-Dependent Translational Control of Irf8 Mediates Adipose Tissue Macrophage Inflammatory Response.

Deregulation of mRNA translation engenders many human disorders, including obesity, neurodegenerative diseases, and cancer, and is associated with pathogen infections. The role of eIF4E-dependent translational control in macrophage inflammatory responses in vivo is largely unexplored. In this study, we investigated the involvement of the translation inhibitors eIF4E-binding proteins (4E-BPs) in the regulation of macrophage inflammatory responses in vitro and in vivo. We show that the lack of 4E-BPs exacerbates inflammatory polarization of bone marrow-derived macrophages and that 4E-BP-null adipose tissue macrophages display enhanced inflammatory gene expression following exposure to a high-fat diet (HFD). The exaggerated inflammatory response in HFD-fed 4E-BP-null mice coincides with significantly higher weight gain, higher Irf8 mRNA translation, and increased expression of IRF8 in adipose tissue compared with wild-type mice. Thus, 4E-BP-dependent translational control limits, in part, the proinflammatory response during HFD. These data underscore the activity of the 4E-BP-IRF8 axis as a paramount regulatory mechanism of proinflammatory responses in adipose tissue macrophages.

Redefining proteostasis transcription factors in organismal stress responses, development, metabolism, and health.

Eukaryotic organisms have evolved complex and robust cellular stress response pathways to ensure maintenance of proteostasis and survival during fluctuating environmental conditions. Highly conserved stress response pathways can be triggered and coordinated at the cell-autonomous and cell-nonautonomous level by proteostasis transcription factors, including HSF1, SKN-1/NRF2, HIF1, and DAF-16/FOXO that combat proteotoxic stress caused by environmental challenges. While these transcription factors are often associated with a specific stress condition, they also direct "noncanonical" transcriptional programs that serve to integrate a multitude of physiological responses required for development, metabolism, and defense responses to pathogen infections. In this review, we outline the established function of these key proteostasis transcription factors at the cell-autonomous and cell-nonautonomous level and discuss a newly emerging stress responsive transcription factor, PQM-1, within the proteostasis network. We look beyond the canonical stress response roles of proteostasis transcription factors and highlight their function in integrating different physiological stimuli to maintain cytosolic organismal proteostasis.

In vitro detection of porphyrin-producing wound bacteria with real-time fluorescence imaging.

Aim: Fluorescence imaging can visualize polymicrobial populations in chronic and acute wounds based on porphyrin fluorescence. We investigated the fluorescent properties of specific wound pathogens and the fluorescence detected from bacteria in biofilm. Methods: Utilizing Remel Porphyrin Test Agar, 32 bacterial and four yeast species were examined for red fluorescence under 405 nm violet light illumination. Polymicrobial biofilms, supplemented with δ-aminolevulinic acid, were investigated similarly. Results: A total of 28/32 bacteria, 1/4 yeast species and polymicrobial biofilms produced red fluorescence, in agreement with their known porphyrin production abilities. Conclusion: These results identify common wound pathogens capable of producing porphyrin-specific fluorescence and support clinical observations using fluorescence imaging to detect pathogenic bacteria in chronic wounds.

Use of a bacterial fluorescence imaging device: wound measurement, bacterial detection and targeted debridement.

OBJECTIVE: Diagnostics which provide objective information to facilitate evidence-based treatment decisions could improve the chance of wound healing. Accurate wound measurements, objective bacterial assessment, and the regular, consistent tracking of these parameters are important aspects of wound care. This study aimed to assess the accuracy, clinical incorporation and documentation capabilities of a handheld bacterial fluorescence imaging device (MolecuLight i:X). METHOD: Benchtop wound models with known dimensions and clinical wound images were repeatedly measured by trained clinicians to quantify accuracy and intra/inter-user coefficients of variation (COV) of the imaging device measurement software. In a clinical trial of 50 wounds, wound dimensions were digitally measured and fluorescence images were acquired to assess for the presence of bacteria at moderate-to-heavy loads. Finally, fluorescence imaging was implemented into the routine assessment of 22 routine diabetic foot ulcers (DFU) to determine appropriate debridement level and location based on bacterial fluorescence signals. RESULTS: Wound measurement accuracy was >95% (COV <3%). In the clinical trial of 50 wounds, 72% of study wounds demonstrated positive bacterial fluorescence signals. Levine sampling of wounds was found to under-report bacterial loads relative to fluorescence-guided curettage samples. Furthermore, fluorescence documentation of bacterial presence and location(s) resulted in more aggressive, fluorescence-targeted debridement in 17/20 DFUs after standard of care debridement failed to eliminate bacterial fluorescence in 100% of DFU debridements. CONCLUSION: The bacterial fluorescence imaging device can be readily implemented for objective, evidenced-based wound assessment and documentation at the bedside. Bedside localisation of regions with moderate-to-heavy bacterial loads facilitated improved sampling, debridement targeting and improved wound bed preparation.

Opinions and Practices of Lung Cancer Screening by Physician Specialty.

BACKGROUND In response to the National Lung Screening Trial, numerous professional organizations published guidelines recommending annual lung cancer screening with low-dose computed tomography (LDCT) for high-risk patients. Prior studies found that physician attitudes and knowledge about lung cancer screening directly impacts the number of screening exams ordered.METHODS In 2015, we surveyed 34 pulmonologists and 186 primary care providers (PCPs) to evaluate opinions and practices of lung cancer screening in a large academic medical center. We compared PCP and pulmonologist responses using t-tests and χ2 tests.RESULTS The overall survey response rate was 40% (39% for PCPs and 50% for pulmonologists). Pulmonologists were more likely than PCPs to report lung cancer screening as beneficial for patients (88.2% versus 37.7%, P < .0001) and as being cost-effective (47.1% versus 14.3%, P = .02). More pulmonologists (76%) reported ordering a LDCT for screening in the past 12 months compared to PCPs (41%, P = .012). Pulmonologists and PCPs reported similar barriers to referring patients for lung cancer screening, including patient costs (82.4% versus 77.8%), potential for emotional harm (58.8% versus 58.3%), high false positive rate (47.1% versus 69.4%), and likelihood for medical complications (47.1% versus 59.7%).LIMITATIONS Our results are generalizable to academic medical centers and responses may be susceptible to recall bias, non-response bias, and social desirability bias.CONCLUSION We found significant differences in opinions and practices between PCPs and pulmonologists regarding lung cancer screening referrals and perceived benefits. As lung cancer screening continues to emerge in clinical practice, it is important to understand these differences across provider specialty to ensure screening is implemented and offered to patients appropriately.

Duplication of hsp-110 Is Implicated in Differential Success of Globodera Species under Climate Change.

Managing the emergence and spread of crop pests and pathogens is essential for global food security. Understanding how organisms have adapted to their native climate is key to predicting the impact of climate change. The potato cyst nematodes Globodera pallida and G. rostochiensis are economically important plant pathogens that cause yield losses of up to 50% in potato. The two species have different thermal optima that may relate to differences in the altitude of their regions of origin in the Andes. Here, we demonstrate that juveniles of G. pallida are less able to recover from heat stress than those of G. rostochiensis. Genome-wide analysis revealed that while both Globodera species respond to heat stress by induction of various protective heat-inducible genes, G. pallida experiences heat stress at lower temperatures. We use C. elegans as a model to demonstrate the dependence of the heat stress response on expression of Heat Shock Factor-1 (HSF-1). Moreover, we show that hsp-110 is induced by heat stress in G. rostochiensis, but not in the less thermotolerant G. pallida. Sequence analysis revealed that this gene and its promoter was duplicated in G. rostochiensis and acquired thermoregulatory properties. We show that hsp-110 is required for recovery from acute thermal stress in both C. elegans and in G. rostochiensis. Our findings point towards an underlying molecular mechanism that allows the differential expansion of one species relative to another closely related species under current climate change scenarios. Similar mechanisms may be true of other invertebrate species with pest status.

A PQM-1-Mediated Response Triggers Transcellular Chaperone Signaling and Regulates Organismal Proteostasis.

In metazoans, tissues experiencing proteotoxic stress induce "transcellular chaperone signaling" (TCS) that activates molecular chaperones, such as hsp-90, in distal tissues. How this form of inter-tissue communication is mediated to upregulate systemic chaperone expression and whether it can be utilized to protect against protein misfolding diseases remain open questions. Using C. elegans, we identified key components of a systemic stress signaling pathway that links the innate immune response with proteostasis maintenance. We show that mild perturbation of proteostasis in the neurons or the intestine activates TCS via the GATA zinc-finger transcription factor PQM-1. PQM-1 coordinates neuron-activated TCS via the innate immunity-associated transmembrane protein CLEC-41, whereas intestine-activated TCS depends on the aspartic protease ASP-12. Both TCS pathways can induce hsp-90 in muscle cells and facilitate amelioration of Aß3-42-associated toxicity. This may have powerful implications for the treatment of diseases related to proteostasis dysfunction.

Effector gene birth in plant parasitic nematodes: Neofunctionalization of a housekeeping glutathione synthetase gene.

Plant pathogens and parasites are a major threat to global food security. Plant parasitism has arisen four times independently within the phylum Nematoda, resulting in at least one parasite of every major food crop in the world. Some species within the most economically important order (Tylenchida) secrete proteins termed effectors into their host during infection to re-programme host development and immunity. The precise detail of how nematodes evolve new effectors is not clear. Here we reconstruct the evolutionary history of a novel effector gene family. We show that during the evolution of plant parasitism in the Tylenchida, the housekeeping glutathione synthetase (GS) gene was extensively replicated. New GS paralogues acquired multiple dorsal gland promoter elements, altered spatial expression to the secretory dorsal gland, altered temporal expression to primarily parasitic stages, and gained a signal peptide for secretion. The gene products are delivered into the host plant cell during infection, giving rise to "GS-like effectors". Remarkably, by solving the structure of GS-like effectors we show that during this process they have also diversified in biochemical activity, and likely represent the founding members of a novel class of GS-like enzyme. Our results demonstrate the re-purposing of an endogenous housekeeping gene to form a family of effectors with modified functions. We anticipate that our discovery will be a blueprint to understand the evolution of other plant-parasitic nematode effectors, and the foundation to uncover a novel enzymatic function.

Climate change is predicted to alter the current pest status of Globodera pallida and G. rostochiensis in the United Kingdom.

The potato cyst nematodes Globodera pallida and G. rostochiensis are economically important plant pathogens causing losses to UK potato harvests estimated at £50 m/ year. Implications of climate change on their future pest status have not been fully considered. Here, we report growth of female G. pallida and G. rostochiensis over the range 15 to 25°C. Females per plant and their fecundity declined progressively with temperatures above 17.5°C for G. pallida, whilst females per plant were optimal between 17.5 and 22.5°C for G. rostochiensis. Relative reproductive success with temperature was confirmed on two potato cultivars infected with either species at 15, 22.5 and 25°C. The reduced reproductive success of G. pallida at 22.5°C relative to 15°C was also recorded for a further seven host cultivars studied. The differences in optimal temperatures for reproductive success may relate to known differences in the altitude of their regions of origin in the Andes. Exposure of G. pallida to a diurnal temperature stress for one week during female growth significantly suppressed subsequent growth for one week at 17.5°C but had no effect on G. rostochiensis. However, after two weeks of recovery, female size was not significantly different from that for the control treatment. Future soil temperatures were simulated for medium- and high-emission scenarios and combined with nematode growth data to project future implications of climate change for the two species. Increased soil temperatures associated with climate change may reduce the pest status of G. pallida but benefit G. rostochiensis especially in the southern United Kingdom. We conclude that plant breeders may be able to exploit the thermal limits of G. pallida by developing potato cultivars able to grow under future warm summer conditions. Existing widely deployed resistance to G. rostochiensis is an important characteristic to retain for new potato cultivars.

Lung Cancer Screening Practices in North Carolina CT Facilities.

BACKGROUND: The burden of lung cancer in the United States is staggering, with more Americans dying from lung cancer than the next four most common cancers combined. With endorsement of lung cancer screening by the United States Preventive Services Task Force and reimbursement by CMS, the number screened for lung cancer with low-dose CT (LDCT) is anticipated to rise in the near future. METHODS: In 2015, we conducted a cross-sectional study of mailed surveys to 112 ACR CT facilities across North Carolina to examine recommended guidelines for lung cancer screening, referral patterns, and patient tracking methods. We describe the survey results and compare findings with surveys of academic medical centers. RESULTS: Among 48 respondents (response rate = 42.9%), 54.2% offer lung cancer screening with LDCT (93.1% community and 6.9% academic settings); of these, 70.8% use ACR/CMS guidelines. In lung cancer screening facilities, reported patient volumes are low (average eight patients screened per month, range: 0-30) and patient tracking occurs in 72% of facilities. CONCLUSIONS: Among our predominately community-based sample of facilities, we found variation in lung cancer screening guideline usage, number of patients screened, and tracking methods.

The isotopic fractionation of carbon, nitrogen, hydrogen, and oxygen during illicit production of cocaine base in South America.

Stable isotope measurements have become a key component in sourcing the origin of illicit cocaine seized within the United States. Therefore, it is imperative to understand the process by which isotopes may be fractionated during illicit cocaine processing. In a controlled observational study, there was apparent isotopic fractionation of carbon, nitrogen, hydrogen, and oxygen. To investigate the potential source of the fractionation, cocaine base was fractionally precipitated from a dilute sulfuric acid solution with dilute ammonium hydroxide. The values of δ13C, δ15N, δ2H, and δ18O for each fraction were measured by isotope ratio mass spectrometry (IRMS). There was an equilibrium fractionation observed in all measured stable isotopes. Early fractions were depleted, and later fractions were enriched, with 15N and 2H being the most affected. The described trend is opposite of the Rayleigh distillation observed for cocaine hydrochloride precipitation.

Opinions, practice patterns, and perceived barriers to lung cancer screening among attending and resident primary care physicians.

INTRODUCTION: The US Preventive Services Task Force recommended annual lung cancer screening with low-dose computed tomography (LDCT) for high-risk patients in December 2013. We compared lung cancer screening-related opinions and practices among attending and resident primary care physicians (PCPs). METHODS: In 2015, we conducted a 23-item survey among physicians at a large academic medical center. We surveyed 100 resident PCPs (30% response rate) and 86 attending PCPs (49% response rate) in Family Medicine and Internal Medicine. The questions focused on physicians' opinions, knowledge of recommendations, self-reported practice patterns, and barriers to lung cancer screening. In 2015 and 2016, we compared responses among attending versus resident PCPs using chi-square/Fisher's exact tests and 2-samples t-tests. RESULTS: Compared with resident PCPs, attending PCPs were older (mean age =47 vs 30 years) and more likely to be male (54% vs 37%). Over half of both groups concurred that inconsistent recommendations make deciding whether or not to screen difficult. A substantial proportion in both groups indicated that they were undecided about the benefit of lung cancer screening for patients (43% attending PCPs and 55% resident PCPs). The majority of attending and resident PCPs agreed that barriers to screening included limited time during patient visits (62% and 78%, respectively), cost to patients (74% and 83%, respectively), potential for complications (53% and 70%, respectively), and a high false-positive rate (67% and 73%, respectively). CONCLUSION: There was no evidence to suggest that attending and resident PCPs had differing opinions about lung cancer screening. For population-based implementation of lung cancer screening, physicians and trainees will need resources and time to address the benefits and harms with their patients.

The genome of the yellow potato cyst nematode, Globodera rostochiensis, reveals insights into the basis of parasitism and virulence.

BACKGROUND: The yellow potato cyst nematode, Globodera rostochiensis, is a devastating plant pathogen of global economic importance. This biotrophic parasite secretes effectors from pharyngeal glands, some of which were acquired by horizontal gene transfer, to manipulate host processes and promote parasitism. G. rostochiensis is classified into pathotypes with different plant resistance-breaking phenotypes. RESULTS: We generate a high quality genome assembly for G. rostochiensis pathotype Ro1, identify putative effectors and horizontal gene transfer events, map gene expression through the life cycle focusing on key parasitic transitions and sequence the genomes of eight populations including four additional pathotypes to identify variation. Horizontal gene transfer contributes 3.5 % of the predicted genes, of which approximately 8.5 % are deployed as effectors. Over one-third of all effector genes are clustered in 21 putative 'effector islands' in the genome. We identify a dorsal gland promoter element motif (termed DOG Box) present upstream in representatives from 26 out of 28 dorsal gland effector families, and predict a putative effector superset associated with this motif. We validate gland cell expression in two novel genes by in situ hybridisation and catalogue dorsal gland promoter element-containing effectors from available cyst nematode genomes. Comparison of effector diversity between pathotypes highlights correlation with plant resistance-breaking. CONCLUSIONS: These G. rostochiensis genome resources will facilitate major advances in understanding nematode plant-parasitism. Dorsal gland promoter element-containing effectors are at the front line of the evolutionary arms race between plant and parasite and the ability to predict gland cell expression a priori promises rapid advances in understanding their roles and mechanisms of action.

STAT3 Establishes an Immunosuppressive Microenvironment during the Early Stages of Breast Carcinogenesis to Promote Tumor Growth and Metastasis.

Immunosurveillance constitutes the first step of cancer immunoediting in which developing malignant lesions are eliminated by antitumorigenic immune cells. However, the mechanisms by which neoplastic cells induce an immunosuppressive state to evade the immune response are still unclear. The transcription factor STAT3 has been implicated in breast carcinogenesis and tumor immunosuppression in advanced disease, but its involvement in early disease development has not been established. Here, we genetically ablated Stat3 in the tumor epithelia of the inducible PyVmT mammary tumor model and found that Stat3-deficient mice recapitulated the three phases of immunoediting: elimination, equilibrium, and escape. Pathologic analyses revealed that Stat3-deficient mice initially formed hyperplastic and early adenoma-like lesions that later completely regressed, thereby preventing the emergence of mammary tumors in the majority of animals. Furthermore, tumor regression was correlated with massive immune infiltration into the Stat3-deficient lesions, leading to their elimination. In a minority of animals, focal, nonmetastatic Stat3-deficient mammary tumors escaped immune surveillance after a long latency or equilibrium period. Taken together, our findings suggest that tumor epithelial expression of Stat3 plays a critical role in promoting an immunosuppressive tumor microenvironment during breast tumor initiation and progression, and prompt further investigation of Stat3-inhibitory strategies that may reactivate the immunosurveillance program.