Evolution and engineering of pathways for aromatic O-demethylation in Pseudomonas putida KT2440.

Biological conversion of lignin from biomass offers a promising strategy for sustainable production of fuels and chemicals. However, aromatic compounds derived from lignin commonly contain methoxy groups, and O-demethylation of these substrates is often a rate-limiting reaction that influences catabolic efficiency. Several enzyme families catalyze aromatic O-demethylation, but they are rarely compared in vivo to determine an optimal biocatalytic strategy. Here, two pathways for aromatic O-demethylation were compared in Pseudomonas putida KT2440. The native Rieske non-heme iron monooxygenase (VanAB) and, separately, a heterologous tetrahydrofolate-dependent demethylase (LigM) were constitutively expressed in P. putida, and the strains were optimized via adaptive laboratory evolution (ALE) with vanillate as a model substrate. All evolved strains displayed improved growth phenotypes, with the evolved strains harboring the native VanAB pathway exhibiting growth rates ∼1.8x faster than those harboring the heterologous LigM pathway. Enzyme kinetics and transcriptomics studies investigated the contribution of selected mutations toward enhanced utilization of vanillate. The VanAB-overexpressing strains contained the most impactful mutations, including those in VanB, the reductase for vanillate O-demethylase, PP_3494, a global regulator of vanillate catabolism, and fghA, involved in formaldehyde detoxification. These three mutations were combined into a single strain, which exhibited approximately 5x faster vanillate consumption than the wild-type strain in the first 8 h of cultivation. Overall, this study illuminates the details of vanillate catabolism in the context of two distinct enzymatic mechanisms, yielding a platform strain for efficient O-demethylation of lignin-related aromatic compounds to value-added products.

Biosensors for the detection of chorismate and cis, cis-muconic acid in Corynebacterium glutamicum.

Corynebacterium glutamicum ATCC 13032 is a promising microbial chassis for industrial production of valuable compounds, including aromatic amino acids derived from the shikimate pathway. In this work, we developed two whole-cell, transcription factor based fluorescent biosensors to track cis, cis-muconic acid (ccMA) and chorismate in C. glutamicum. Chorismate is a key intermediate in the shikimate pathway from which value-added chemicals can be produced, and a shunt from the shikimate pathway can divert carbon to ccMA, a high value chemical. We transferred a ccMA-inducible transcription factor, CatM, from Acinetobacter baylyi ADP1 into C. glutamicum and screened a promoter library to isolate variants with high sensitivity and dynamic range to ccMA by providing benzoate, which is converted to ccMA intracellularly. The biosensor also detected exogenously supplied ccMA, suggesting the presence of a putative ccMA transporter in C. glutamicum, though the external ccMA concentration threshold to elicit a response was 100-fold higher than the concentration of benzoate required to do so through intracellular ccMA production. We then developed a chorismate biosensor, in which a chorismate inducible promoter regulated by natively expressed QsuR was optimized to exhibit a dose-dependent response to exogenously supplemented quinate (a chorismate precursor). A chorismate-pyruvate lyase encoding gene, ubiC, was introduced into C. glutamicum to lower the intracellular chorismate pool, which resulted in loss of dose-dependence to quinate. Further, a knockout strain that blocked the conversion of quinate to chorismate, also resulted in absence of dose-dependence to quinate, validating that the chorismate biosensor is specific to intracellular chorismate pool. The ccMA and chorismate biosensors were dually inserted into C. glutamicum to simultaneously detect intracellularly produced chorismate and ccMA. Biosensors, such as those developed in this study, can be applied in C. glutamicum for multiplex sensing to expedite pathway design and optimization through metabolic engineering in this promising chassis organism.

Evaluation and Treatment of Primary Spinal Epidural Lymphoma Presenting as Acute Myelopathy in a Young Adult: A Case Report.

CASE: Primary spinal epidural lymphoma (PSEL) presenting as myelopathy is extremely rare, particularly within young, healthy adults. This case report describes a 26-year-old man presenting with progressive thoracic myelopathy. Magnetic resonance imaging revealed spinal epidural masses spanning T5-T10 and T12-L2 with multilevel cord compression and edema. After evaluation, the patient underwent emergent posterior decompression to prevent progressive neurological decline. Histology was consistent with diffuse large B-cell lymphoma, germinal center type. At 3 months postoperatively, the patient regained full neurologic function. CONCLUSION: Although rare, PSELs should be considered in patients presenting with myelopathy to facilitate timely diagnosis and treatment.

Structural contributions of different manganese oxide minerals on the redox transformations and proliferation of iodine.

The redox capabilities of birnessite minerals are contingent upon the physical characteristics of the solid, indicating that different allotropes have various reactivities. Here, the role of these structural differences on the oxidation of iodine, a risk driving environmental contaminant in several federal complexes, was investigated. The mechanism of which can be seen here, with one of the minerals of study, acid birnessite. The pH range chosen for this study was pH 5-6. Throughout the experiments it was seen that the average oxidation state (AOS) had the greatest contribution to the differences in redox capabilities of the various birnessite minerals. Several trends were observed throughout this study: as AOS decreased, oxidation of iodide (I-) increased; as specific surface area (SSA) increased, the sorption of iodate (IO3-) increased. Additional experiments were conducted at trace levels of iodine, to better model environmental conditions. In that case, a one-step conversion of I- to IO3- occurred, to a greater extent than under artificially elevated concentrations.

Complex regulation in a Comamonas platform for diverse aromatic carbon metabolism.

Critical to a sustainable energy future are microbial platforms that can process aromatic carbons from the largely untapped reservoir of lignin and plastic feedstocks. Comamonas species present promising bacterial candidates for such platforms because they can use a range of natural and xenobiotic aromatic compounds and often possess innate genetic constraints that avoid competition with sugars. However, the metabolic reactions of these species are underexplored, and the regulatory mechanisms are unknown. Here we identify multilevel regulation in the conversion of lignin-related natural aromatic compounds, 4-hydroxybenzoate and vanillate, and the plastics-related xenobiotic aromatic compound, terephthalate, in Comamonas testosteroni KF-1. Transcription-level regulation controls initial catabolism and cleavage, but metabolite-level thermodynamic regulation governs fluxes in central carbon metabolism. Quantitative 13C mapping of tricarboxylic acid cycle and cataplerotic reactions elucidates key carbon routing not evident from enzyme abundance changes. This scheme of transcriptional activation coupled with metabolic fine-tuning challenges outcome predictions during metabolic manipulations.

Impact of radiocapitellar interposition arthroplasty on ulnohumeral joint biomechanics.

BACKGROUND: Radial head excision (RHE) has been shown to increase contact pressures within the ulnohumeral joint. Radiocapitellar interposition arthroplasty (RCIA) with the use of a soft tissue graft is an alternative for the treatment of isolated radiocapitellar arthritis or with failure of radial head replacement. We investigated contact pressures and contact area within the ulnohumeral joint after RHE compared to RCIA with dermal autograft. METHODS: Six fresh-frozen cadaver elbows were tested on a custom dynamic elbow frame. A pressure sensor was inserted into the intact elbow joint, and mean contact pressure, peak contact pressure, contact area, and force within the ulnohumeral joint were recorded at 0°, 30°, 60°, 90°, and 120° of flexion as a valgus load was applied to the elbow. The radial head was then excised and specimens were retested. Finally, a dermal graft matched to the size of the resected radial head was inserted in the radiocapitellar space and the specimens were tested a third time. RESULTS: At 90° of flexion, contact pressure within the ulnohumeral joint was significantly lower with RCIA compared with RHE (110.8 kPa vs 216.8 kPa; P = .013). The mean peak contact pressure was also significantly lower with RCIA compared with RHE at 90° (279.4 vs 626.7 kPa; P = .025). No statistically significant differences were seen in mean contact area or force between the 3 testing conditions at any flexion position. CONCLUSION: RCIA with a dermal graft reduced contact pressures within the ulnohumeral joint compared to RHE at 90° of flexion without a significant change in contact area or contact force.

Rimegepant for the treatment of migraine.

Migraine is a common form of primary headache, affecting up to 1 in every 6 Americans. The pathophysiology is an intricate interplay of genetic factors and environmental influence and is still being elucidated in ongoing studies. The trigeminovascular system is now known to have a significant role in the initiation of migraines, including the release of pain mediators such as CGRP and substance P. Traditional treatment of migraine is usually divided into acute and preventive treatment. Acute therapy includes non-specific therapy, such as NSAIDs and other analgesics, which may provide relief in mild to moderate migraines. 5-HT1 agonists may provide relief in severe migraine, but are not universally effective and carry a significant side-effect profile with frequent redosing requirement. Prophylactic therapy may reduce the occurrence of acute migraine attacks in selected patients, but does not completely eliminate it. More recently, CGRP antagonism has been studied and shown to be effective in both abortion and prevention of migraine. Novel medications, targeting CGRP, divide into CGRP antibodies and receptor antagonists (gepants). Rimegepant, a second-generation gepant, has shown efficacy in several clinical trials in treating acute migraine. Ongoing trials are also evaluating its role in migraine prophylaxis, and results are promising. It is also generally safer for use than existing options, does not appear to increase the chance of developing chronic migraines, and carries a very tolerable side effects profile. It is a part of a growing arsenal in migraine treatment, and may present the silver bullet for treatment of this disease.

The Ramifications of Opioid Utilization and Outcomes of Alternative Pain Control Strategies for Total Knee Arthroplasties.

Morbidity and mortality related to opioid use has generated a public health crisis in the United States. Total knee arthroplasty (TKA) is an increasingly common procedure and is often accompanied by post-operative opioid utilization. Unfortunately, post-operative opioid usage after TKA has been shown to lead to higher rates of complications, longer hospital stays, increased costs, and more frequent need for revision surgery. Pre-operative opioid utilization has been shown to be one of the most important predictors of post-operative opioid usage. Additional risk factors for continued post-operative opioid utilization after TKA include pre-operative substance and tobacco use as well as higher post-operative prescription dosages, younger age, female gender, and Medicaid insurance. One method for mitigating excessive post-operative opioid utilization are Enhanced Recovery After Surgery (ERAS) protocols, which include a multidisciplinary approach that focuses on perioperative factors to optimize patient recovery and function after surgery. Additional strategies include multimodal pain regimens with epidural anesthetics, extended duration local anesthetics and adjuvants, and ultrasound guided peripheral nerve blocks. In recent years, opioid prescribing duration limitations have also been put into place by state and federal government, hospital systems, and ambulatory surgery centers making effective acute pain management imperative for all stakeholders. In this regard, as rates of TKA continue to increase across the United States, multidisciplinary efforts by all stakeholders are needed to ensure adequate pain control while preventing the negative sequalae of opioid medications.

Opioid Use Consequences, Governmental Strategies, and Alternative Pain Control Techniques Following Total Hip Arthroplasties.

Over the last several decades, rates of opioid use and associated problems have dramatically increased in the United States leading to laws limiting prescription duration for acute pain management. As a result, orthopedic surgeons who perform total hip arthroplasty (THA), a procedure that often leads to significant postoperative pain, have been faced with substantial challenges to adequately mitigate patient pain while also reducing opioid intake. Current strategies include identifying and correcting modifiable risk factors associated with postoperative opioid use such as preoperative opioid use, alcohol and tobacco abuse, and untreated psychiatric illness. Additionally, recent evidence has emerged in the form of Enhanced Recovery After Surgery (ERAS) protocols suggesting that a multidisciplinary focus on patient factors perioperatively can lead to reduced postoperative opioid administration and decreased hospital stays. A cornerstone of ERAS protocols includes multimodal pain regimens with opioid rescue only as needed, which often includes multiple systemic pain therapies such as acetaminophen, gabapentin, non-steroidal anti-inflammatory drugs, as well as targeted pain therapies that include epidural catheters and ultrasound-guided nerve blocks. Many hospital systems and states have also implemented opioid prescribing limitations with mixed success. As the opioid epidemic continues in the United States, while contributing to poor outcomes following elective surgeries, further research is warranted to identify multidisciplinary strategies that mitigate opioid use while also allowing for adequate pain control and rehabilitation.

Can Clinical Response Predict Pathologic Response Following Neoadjuvant Chemoradiation for Esophageal Cancer?

OBJECTIVES: Approximately 20-40% of patients with locally advanced esophageal cancer will achieve a pathologic complete response (ypCR) following neoadjuvant chemoradiotherapy (nCRT). Predicting ypCR based on a clinical complete response (ycCR) has been a challenge. This study assessed the correlation between ycCR and ypCR, as determined from esophagectomy specimens. METHODS: Patients undergoing esophagectomy following nCRT at three major institutions between 2005 and 2018 were reviewed. Restaging, including PET/CT, endoscopy with biopsy, and esophageal ultrasound (EUS), was performed to determine ycCR. RESULTS: Six hundred sixty patients were included, with 93.3% with esophageal adenocarcinoma histology. Six hundred fifty-eight of these patients underwent PET, 304 EUS, and 584 underwent a biopsy. Following nCRT, 148 (22.4%) were found to have a ypCR. Only 12/32 (37.5%) determined to have a ycCR were found to have a ypCR, while 136/628 (21.6%) with a non-ycCR were found to have a ypCR (p 0.075). Individual modality PPV was 28% for PET, 54% for EUS, and 26% for biopsy. When PET was combined with EUS, 168 reports were concordant and the PPV of ypCR was 50%, though the number of patients was low (1/2). With all 3 re-staging modalities combined, the PPV and NPV both rose to 100%. CONCLUSIONS: Current restaging tools cannot reliably predict ypCR after nCRT. While multimodal restaging appears to be a more accurate predictor of ypCR than any testing modality alone, patients cannot reliably be advised to avoid an esophagectomy on the assumption that ycCR predicts ypCR at this time.

Fragility of randomized controlled trials on treatment of proximal humeral fracture.

BACKGROUND: Proximal humeral fracture represents an increasingly common pathology with evaluation and treatment often guided by evidence from randomized controlled trials (RCTs), but the strength of an RCT must be considered in this process. The purpose of this study was to evaluate the strength of outcomes in RCTs on the management of proximal humeral fractures using the fragility index (FI), a method used with statistically significant dichotomous outcomes to assess the number of patients that would change an outcome measure from significant (P ≤ .05) to nonsignificant if the patient outcome changed. We also aimed to correlate the FI with other measures of study strength. METHODS: A systematic review was performed using Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines to evaluate RCTs on the management of proximal humeral fractures. The PubMed, Ovid MEDLINE, Web of Science, and Embase databases were searched from database inception to May 2021. RCTs with at least 1 statistically significant (P ≤ .05) dichotomous outcome were included. The FI was calculated for each included trial using the Fisher exact test. The FI was correlated with the study sample size and journal impact factor. RESULTS: Ten RCTs reporting on 656 patients and published between 2011 and 2020 were included. The median patient sample size was 67 (mean, 65.6; range, 40-86). Complications were the most commonly reported dichotomous statistically significant outcome. The median FI was 1 (mean, 2.6; range, 0-18), with 4 studies having an FI of 0. A median FI of 1 indicates that 1 patient experiencing an alternative outcome or having not been lost to follow-up could have changed the pertinent conclusions of the trial for a given outcome. The median number of patients lost to follow-up was 3 (mean, 4.9; range, 0-16) and exceeded the FI in 50% of studies. There was no correlation between the FI and sample size (Spearman coefficient = 0.0592, P = .865) or between the FI and journal impact factor (Spearman coefficient = -0.0229, P = .522). CONCLUSION: In most studies of proximal humeral fractures, only 1 or 2 patients experiencing an alternative outcome or lost to follow-up would change the conclusions for the dichotomous outcome studied. Although the FI cannot be used to assess continuous variables, which are often the primary outcome variables of RCTs, it does offer an additional unique measure of study strength that surgeons should consider when evaluating RCTs.

A Scoping Review of the Health Impact of the COVID-19 Pandemic on Persons Experiencing Homelessness in North America and Europe.

Persons experiencing homelessness (PEH) are at heightened risk for infection, morbidity, and mortality from COVID-19. However, health consequences of the pandemic extend far beyond those directly caused by the virus. This scoping review aimed to explore the impacts of the COVID-19 pandemic on the health and well-being of PEH in North America and Europe. A systematic search of academic and grey literature was conducted in September 2021. To be included, studies had to include primary data related to the impact of the pandemic on health or well-being of PEH and be written in English. All potentially relevant references were independently screened by two reviewers, and minor conflicts were settled with input of a third reviewer. A total of 96 articles met criteria for inclusion. Data extraction was completed for all included studies, and findings synthesised and presented thematically. Numerous health impacts of the pandemic on PEH were identified, including SARS-CoV-2 infection, morbidity, mortality, and hospitalisation, fear of infection, access to housing, hygiene, PPE, food, as well as mental health, substance use, other health-related outcomes and treatment services. Gaps in the literature relating to persons using alcohol, access to mental health support, and violence were also identified. Implications for future research are discussed.

Adaptive laboratory evolution for improved tolerance of isobutyl acetate in Escherichia coli.

Previously, Escherichia coli was engineered to produce isobutyl acetate (IBA). Titers greater than the toxicity threshold (3 g/L) were achieved by using layer-assisted production. To avoid this costly and complex method, adaptive laboratory evolution (ALE) was applied to E. coli for improved IBA tolerance. Over 37 rounds of selective pressure, 22 IBA-tolerant mutants were isolated. Remarkably, these mutants not only tolerate high IBA concentrations, they also produce higher IBA titers. Using whole-genome sequencing followed by CRISPR/Cas9 mediated genome editing, the mutations (SNPs in metH, rho and deletion of arcA) that confer improved tolerance and higher titers were elucidated. The improved IBA titers in the evolved mutants were a result of an increased supply of acetyl-CoA and altered transcriptional machinery. Without the use of phase separation, a strain capable of 3.2-fold greater IBA production than the parent strain was constructed by combing select beneficial mutations. These results highlight the impact improved tolerance has on the production capability of a biosynthetic system.

Diagnostic Evaluation and Cervical Spine Surgery in the Setting of a Cardiac Left Ventricular Assist Device: Challenges and a Case Illustration.

Due to incompatibility with magnetic resonance imaging, patients with left ventricular assist devices (LVADs) presenting with pathologies of the spinal soft tissues or neural elements represent diagnostically complex cases. We present a case of a patient undergoing a CT (computed tomography) myelogram and subsequent successful cervical posterior laminectomy. A C1-C2 lateral puncture approach CT myelogram revealed nearly a complete block of contrast movement at the level of the C2-C3 vertebrae concerning a compressive etiology. The cervical lateral approach was chosen based on patient symptomology and concern that contrast dye injected in the lumbar spine would not travel to the region of interest due to altered CSF pulsatility caused by the LVAD device. A C3-C7 posterior laminectomy was then successfully performed. Intra-operatively, however, there was no sign of a compressive lesion, and ultrasound confirmed a decompressed spinal cord. This case highlights the diagnostic challenges of pre-operative evaluation in patients with LVADs in which the efficacy of performing CT myelograms is also questionable due to potential alterations in cerebrospinal fluid movement due to variations in arterial pulsatility due to LVAD physiology.

An Update on Postoperative Opioid Use and Alternative Pain Control Following Spine Surgery.

Opioids are commonly prescribed postoperatively for pain control, especially in spine surgery. Not only does this pose concerns for potential abuse, but it also has been shown to worsen certain outcomes. Risk factors for increased use include preoperative opioid use, female sex, psychiatric diagnoses, and drug and alcohol use. Over the past few decades, there have been increasing efforts mostly spearheaded by governmental agencies to decrease postoperative opioid use via opioid prescription limitation laws regulating the number of days and amounts of analgesics prescribed and promotion of the use of enhanced recovery after surgery (ERAS) protocols, multimodal pain regimens, epidural catheters, and ultrasound-guided peripheral nerve blocks. These strategies collectively have been efficacious in decreasing overall opioid use and better controlling patients' postoperative pain while simultaneously improving other outcomes such as postoperative nausea, vomiting, and length of stay. With an aging population undergoing an increasing number of spinal surgeries each year, it is now more important than ever to continue these efforts to improve the quality and safety of pain control methods after spinal surgery and limit the transition of acute management to the development of opioid dependence and addiction long-term.

Does minimally invasive spine surgery improve outcomes in the obese population? A retrospective review of 1442 degenerative lumbar spine surgeries.

OBJECTIVE: The effect of obesity on outcomes in minimally invasive surgery (MIS) approaches to posterior lumbar surgery is not well characterized. The authors aimed to determine if there was a difference in operative variables and complication rates in obese patients who underwent MIS versus open approaches in posterior spinal surgery, as well as between obese and nonobese patients undergoing MIS approaches. METHODS: A retrospective review of all consecutive patients who underwent posterior lumbar surgery from 2013 to 2016 at a single institution was performed. The primary outcome measure was postoperative complications. Secondary outcome measures included estimated blood loss (EBL), operative time, the need for revision, and hospital length of stay (LOS); readmission and disposition were also reviewed. Obese patients who underwent MIS were compared with those who underwent an open approach. Additionally, obese patients who underwent an MIS approach were compared with nonobese patients. Bivariate and multivariate analyses were carried out between the groups. RESULTS: In total, 423 obese patients (57.0% decompression and 43.0% fusion) underwent posterior lumbar MIS. When compared with 229 obese patients (56.8% decompression and 43.2% fusion) who underwent an open approach, patients in both the obese and nonobese groups who underwent MIS experienced significantly decreased EBL, LOS, operative time, and surgical site infections (SSIs). Of the nonobese patients, 538 (58.4% decompression and 41.6% fusion) underwent MIS procedures. When compared with nonobese patients, obese patients who underwent MIS procedures had significantly increased LOS, EBL, operative time, revision rates, complications, and readmissions in the decompression group. In the fusion group, only LOS and disposition were significantly different. CONCLUSIONS: Obese patients have poorer outcomes after posterior lumbar MIS when compared with nonobese patients. The use of an MIS technique can be of benefit, as it decreased EBL, operative time, LOS, and SSIs for posterior decompression with or without instrumented fusion in obese patients.

Minimally invasive posterior lumbar surgery in the morbidly obese, obese and non-obese populations: A single institution retrospective review.

BACKGROUND: There is a paucity of information regarding outcomes in minimally invasive surgical (MIS) approaches to posterior lumbar surgery in morbidly obese patients. We seek to determine if there are differences in operative variables and early complication rates in morbidly obese patients undergoing MIS posterior lumbar surgery compared to obese and non-obese patients. METHODS: A single institution retrospective review of patients undergoing MIS posterior lumbar surgery (decompression and/or fusion) between 2013 and 2016 was performed. Morbidly obese patients (BMI ≥ 40) were compared to obese (BMI 30-39.9) and non-obese (BMI < 30) cohorts. Postoperative complication rates and perioperative variables including estimated blood loss, operative time, and outcome measures including length of stay (LOS), in-hospital complications, readmission, and disposition were assessed. RESULTS: 47 morbidly obese, 135 obese and 224 non-obese patients underwent posterior MIS instrumented fusion. 59 morbidly obese, 182 obese and 314 non-obese patients underwent posterior MIS decompression. The morbidly obese group experienced a greater rate of deep vein thrombosis and had an increased hospital LOS (p < 0.05). Morbidly obese patients who underwent MIS decompression experienced increased postoperative complications (p < 0.01), and increased LOS (p < 0.0001) compared to obese and non-obese patients. There were no differences in revision rates, readmissions, and other complications including surgical site infection. Morbid obesity was an independent predictor of overall complications and increased LOS on multivariate analysis. CONCLUSION: Morbidly obese patients undergoing posterior MIS fusion had a higher rate of complications and increased LOS. While weight loss should be encouraged, complication rates remains acceptably low in morbidly obese patients and MIS posterior lumbar surgery should still be offered.

Doctor-patient interactions that exclude patients experiencing homelessness from health services: an ethnographic exploration.

BACKGROUND: People experiencing homelessness have poor health indices and poor access to health care. Their health service utilisation (HSU) is typified by: late illness presentations; poor attendance rates at appointments; low usage of primary care services and outpatient departments; and high utilisation of emergency departments and inpatient services. Why people experiencing homelessness have these particular HSU patterns is poorly understood. AIM: This research sought to explore barriers to health service usage for people experiencing homelessness. DESIGN & SETTING: The authors conducted critical realist ethnography over 13 months in Dublin with people experiencing homelessness at four purposively chosen sites (a food hall, a drop-in centre, an emergency department, and an outreach service for rough sleepers). METHOD: Ethnographic research was supplemented with focus groups of hospital doctors and people experiencing homelessness, and with 50 semi-structured interviews with people experiencing homelessness. The epistemological framework was critical realism. RESULTS: One of the factors identified in the research as contributing to the HSU pattern of people experiencing homelessness was recurrent interactions between health professionals and patients, whereby patients were either excluded or discouraged from attending health services, or self-excluded themselves from services. These interactions were described as 'conversations of exclusion'. Four such conversations were described: 'the benzodiazepine conversation'; 'the mistrustful conversation'; 'the blaming conversation'; and 'the assertive conversation'. CONCLUSION: There are certain recurrent interactions between people experiencing homelessness and doctors that result in the exclusion of people experiencing homelessness from health services.