A systematic analysis of biosynthetic gene clusters in the human microbiome reveals a common family of antibiotics.

In complex biological systems, small molecules often mediate microbe-microbe and microbe-host interactions. Using a systematic approach, we identified 3,118 small-molecule biosynthetic gene clusters (BGCs) in genomes of human-associated bacteria and studied their representation in 752 metagenomic samples from the NIH Human Microbiome Project. Remarkably, we discovered that BGCs for a class of antibiotics in clinical trials, thiopeptides, are widely distributed in genomes and metagenomes of the human microbiota. We purified and solved the structure of a thiopeptide antibiotic, lactocillin, from a prominent member of the vaginal microbiota. We demonstrate that lactocillin has potent antibacterial activity against a range of Gram-positive vaginal pathogens, and we show that lactocillin and other thiopeptide BGCs are expressed in vivo by analyzing human metatranscriptomic sequencing data. Our findings illustrate the widespread distribution of small-molecule-encoding BGCs in the human microbiome, and they demonstrate the bacterial production of drug-like molecules in humans. PAPERCLIP:

Targeting placental growth factor/neuropilin 1 pathway inhibits growth and spread of medulloblastoma.

Medulloblastoma is the most common pediatric malignant brain tumor. Although current therapies improve survival, these regimens are highly toxic and are associated with significant morbidity. Here, we report that placental growth factor (PlGF) is expressed in the majority of medulloblastomas, independent of their subtype. Moreover, high expression of PlGF receptor neuropilin 1 (Nrp1) correlates with poor overall survival in patients. We demonstrate that PlGF and Nrp1 are required for the growth and spread of medulloblastoma: PlGF/Nrp1 blockade results in direct antitumor effects in vivo, resulting in medulloblastoma regression, decreased metastasis, and increased mouse survival. We reveal that PlGF is produced in the cerebellar stroma via tumor-derived Sonic hedgehog (Shh) and show that PlGF acts through Nrp1-and not vascular endothelial growth factor receptor 1-to promote tumor cell survival. This critical tumor-stroma interaction-mediated by Shh, PlGF, and Nrp1 across medulloblastoma subtypes-supports the development of therapies targeting PlGF/Nrp1 pathway.

Utility of methylated DNA markers for the diagnosis of malignant biliary strictures.

BACKGROUND AND AIMS: Early identification of malignant biliary strictures (MBSs) is challenging, with up to 20% classified as indeterminants after preliminary testing and tissue sampling with endoscopic retrograde cholangiopancreatography. We aimed to evaluate the use of methylated DNA markers (MDMs) from biliary brushings to enhance MBS detection in a prospective cohort. APPROACH: Candidate MDMs were evaluated for their utility in MBS diagnosis through a series of discovery and validation phases. DNA was extracted from biliary brushing samples, quantified, bisulfite-converted, and then subjected to methylation-specific droplet digital polymerase chain reaction.  Patients were considered to have no malignancy if the sampling was negative and there was no evidence of malignancy after 1 year or definitive negative surgical histopathology. RESULTS: Fourteen candidate MDMs were evaluated in the discovery phase, with top-performing and new markers evaluated in the technical validation phase. The top 4 MDMs were TWIST1, HOXA1, VSTM2B, and CLEC11A, which individually achieved AUC values of 0.82, 0.81, 0.83, and 0.78, respectively, with sensitivities of 59.4%, 53.1%, 62.5%, and 50.0%, respectively, at high specificities for malignancy of 95.2%-95.3% for the final biologic validation phase. When combined as a panel, the AUC was 0.86, achieving 73.4% sensitivity and 92.9% specificity, which outperformed cytology and fluorescence in situ hybridization (FISH). CONCLUSIONS: The selected MDMs demonstrated improved performance characteristics for the detection of MBS compared to cytology and FISH. Therefore, MDMs should be considered viable candidates for inclusion in diagnostic testing algorithms.

Direct Proteomic Detection and Prioritization of 19 Onchocerciasis Biomarker Candidates in Humans.

Onchocerca volvulus, the causative agent of onchocerciasis, infects over 20 million people and can cause severe dermatitis and ocular conditions including blindness. Current treatments employed in mass drug administration programs do not kill adult female worms, and common diagnostic tests cannot reliably assess viability of adult worms. There is an urgent need for better diagnostic tests to facilitate monitoring the efficacy of new treatments and disease elimination efforts. Here, eight plasma samples collected from individuals infected with O. volvulus and seven from uninfected individuals were analyzed by MS/MS spectrometry to directly identify O. volvulus proteins present in infected but absent in uninfected control samples. This direct proteomic approach for biomarker discovery had not been previously employed for onchocerciasis. Among all detected proteins, 19 biomarker candidates were supported by two or more unique peptides, identified in the plasma of at least three O. volvulus-infected human samples and absent in all control samples. Comprehensive analysis and ranking of these candidates included detailed functional annotation and a review of RNA-seq gene expression profiles. Isotope-labeled standard peptides were run in parallel and validated MS/MS peptide identifications for 15 peptides from 11 of the 19 proteins, and two infected urine and one uninfected urine sample was used for additional validation. A major antigen/OVOC11613 was identified as the most promising candidate with eight unique peptides across five plasma samples and one urine sample. Additional strong candidates included OVOC1523/ATP synthase, OVOC247/laminin and OVOC11626/PLK5, and along with OVOC11613, and were also detected in urine samples from onchocerciasis patients. This study has identified a promising novel set of proteins that will be carried forward to develop assays that can be used for diagnosis of O. volvulus infections and for monitoring treatment efficacy.

A theory of rapid evolutionary change explaining the de novo appearance of megakaryocytes and platelets in mammals.

Platelets are found only in mammals. Uniquely, they have a log Gaussian volume distribution and are produced from megakaryocytes, large cells that have polyploid nuclei. In this Hypothesis, we propose that a possible explanation for the origin of megakaryocytes and platelets is that, ∼220 million years ago, an inheritable change occurred in a mammalian ancestor that caused the haemostatic cell line of the animal to become polyploid. This inheritable change occurred specifically in the genetic programme of the cell lineage from which the haemostatic cell originated and led, because of increase in cell size, to its fragmentation into cytoplasmic particles (platelets) in the pulmonary circulatory system, as found in modern mammals. We hypothesize that these fragments originating from the new large haemostatic polyploid cells proved to be more efficient at stopping bleeding, and, therefore, the progeny of this ancestor prospered through natural selection. We also propose experimental strategies that could provide evidence to support this hypothesis.

Torpor use in the wild by one of the world's largest bats.

Torpor is widespread among bats presumably because most species are small, and torpor greatly reduces their high mass-specific resting energy expenditure, especially in the cold. Torpor has not been recorded in any bat species larger than 50 g, yet in theory could be beneficial even in the world's largest bats (flying-foxes; Pteropus spp.) that are exposed to adverse environmental conditions causing energy bottlenecks. We used temperature telemetry to measure body temperature in wild-living adult male grey-headed flying-foxes (P. poliocephalus; 799 g) during winter in southern Australia. We found that all individuals used torpor while day-roosting, with minimum body temperature reaching 27°C. Torpor was recorded following a period of cool, wet and windy weather, and on a day with the coldest maximum air temperature, suggesting it is an adaptation to reduce energy expenditure during periods of increased thermoregulatory costs and depleted body energy stores. A capacity for torpor among flying-foxes has implications for understanding their distribution, behavioural ecology and life history. Furthermore, our discovery increases the body mass of bats known to use torpor by more than tenfold and extends the documented use of this energy-saving strategy under wild conditions to all bat superfamilies, with implications for the evolutionary maintenance of torpor among bats and other mammals.

THE ROLE OF MAGNETIC RESONANCE CHOLANGIOPANCREATOGRAPHY FOR PREVENTING PANCREATIC FLUID COLLECTION RECURRENCE AFTER EUS-GUIDED DRAINAGE OF WALLED-OFF NECROSIS.

BACKGROUND AND AIMS: Pancreatic fluid collections (PFC) may recur after initial successful endoscopic drainage of walled-off necrosis (WON), most commonly due to disconnected pancreatic duct syndrome (DPDS). The primary aim of this study is to assess the role of magnetic resonance cholangiopancreatography (MRCP) for identifying DPDS to guide appropriate management and prevent PFC recurrence. METHODS: Patients with WON undergoing LAMS drainage of a PFC were retrospectively identified and categorized as those with versus those without MRCP prior to removal of transmural stents. Data on patient demographics, procedural details, cross-sectional imaging, and recurrence rates were collected through chart review. RESULTS: A total of 121 patients with WON were identified, of whom 44 (36.4%) had an MRCP prior to transmural stent removal. In patients without MRCP, 13/77 (16.8%) had PFC recurrence versus 0/44 (0%; p=0.003) in those with MRCP. MRCP identified DPDS in 12 (27.2%) patients, all of whom were managed with indefinite drainage with double-pigtail plastic stents (DPPS) without recurrence. In the group without MRCP, PFCs recurred at a median interval of 284 days (IQR 182-618 days) after transmural stent removal. Among the 13 patients with PFC recurrence, 11 patients (85%) had undiagnosed DPDS detected on subsequent imaging, of whom 9 were subsequently managed with indefinite DPPS, without further PFC recurrence. CONCLUSION: Patients with WON who underwent MRCP prior to transmural stent removal had a lower rate of PFC recurrence largely due to the identification of DPD with appropriate endoscopic management.

Incidence of Post-ERCP Pancreatitis in Patients Receiving Rectal Indomethacin vs. Compounded Rectal Diclofenac Prophylaxis.

BACKGROUND AND AIMS: Endoscopic retrograde cholangiopancreatography (ERCP) carries a 3-15% risk of post-ERCP pancreatitis (PEP). Rectal indomethacin reduces the risk of PEP, but its cost has increased more than 20-fold over the past decade. Rectal diclofenac is also used to prevent PEP but is not commercially available in the United States. The aim of this study is to compare the incidence of PEP after administration of commercially available rectal indomethacin versus compounded rectal diclofenac and assess financial implications. METHODS: ERCP cases at our institution with administration of 100 mg rectal indomethacin or 100 mg compounded rectal diclofenac between May 2018 and January 2022 were retrospectively reviewed. The incidence and severity of PEP was compared between the indomethacin (n = 728) and diclofenac (n = 304) groups. Risk factors (young age, female sex, history of pancreatitis or PEP, sphincterotomy during procedure, pancreatic indication, trainee involvement) and protective factors (prior sphincterotomy, pancreatic duct stenting) for PEP were compared between groups. RESULTS: 60 patients (8.2%) in the rectal indomethacin group and 25 patients (8.2%) in the compounded rectal diclofenac group developed PEP, resulting in moderate or severe PEP in 9 (15.0%) and 2 (8.0%) patients, respectively. The compounded rectal diclofenac group had more trainee involvement (46.1% vs. 32.8%, p = 0.0001) and more prior sphincterotomy cases (15.8% vs. 10.6%, p = 0.0193) compared to the rectal indomethacin group; no statistically significant differences were observed in all other risk and protective factors. Following switch to compounded rectal diclofenac, institutional annual cost savings amounted to $441,460.62 and patient charge decreased 45-fold. CONCLUSION: This retrospective single-center real-world analysis showed similar efficacy of rectal indomethacin and compounded rectal diclofenac in preventing PEP but demonstrates substantial cost savings after switching to compounded rectal diclofenac.

Quantitative 23Na magnetic resonance imaging in the abdomen at 3 T.

OBJECTIVES: To assess the feasibility of sodium-23 MRI for performing quantitative and non-invasive measurements of total sodium concentration (TSC) and relaxation in a variety of abdominal organs. MATERIALS AND METHODS: Proton and sodium imaging of the abdomen was performed in 19 healthy volunteers using a 3D cones sequence and a sodium-tuned 4-rung transmit/receive body coil on a clinical 3 T system. The effects of B1 non-uniformity on TSC measurements were corrected using the double-angle method. The long-component of 23Na T2* relaxation time was measured using a series of variable echo-times. RESULTS: The mean and standard deviation of TSC and long-component 23Na T2* values were calculated across the healthy volunteer group in the kidneys, cerebrospinal fluid (CSF), liver, gallbladder, spleen, aorta, and inferior vena cava. DISCUSSION: Mean TSC values in the kidneys, liver, and spleen were similar to those reported using 23Na-MRI previously in the literature. Measurements in the CSF and gallbladder were lower, potentially due to the reduced spatial resolution achievable in a clinically acceptable scan time. Mean long-component 23Na T2* values were consistent with previous reports from the kidneys and CSF. Intra-population standard error was larger in smaller, fluid-filled structures due to fluid motion and partial volume effects.

Spinal cord representation of motor cortex plasticity reflects corticospinal tract LTP.

Although it is well known that activity-dependent motor cortex (MCX) plasticity produces long-term potentiation (LTP) of local cortical circuits, leading to enhanced muscle function, the effects on the corticospinal projection to spinal neurons has not yet been thoroughly studied. Here, we investigate a spinal locus for corticospinal tract (CST) plasticity in anesthetized rats using multichannel recording of motor-evoked, intraspinal local field potentials (LFPs) at the sixth cervical spinal cord segment. We produced LTP by intermittent theta burst electrical stimulation (iTBS) of the wrist area of MCX. Approximately 3 min of MCX iTBS potentiated the monosynaptic excitatory LFP recorded within the CST termination field in the dorsal horn and intermediate zone for at least 15 min after stimulation. Ventrolaterally, in the spinal cord gray matter, which is outside the CST termination field in rats, iTBS potentiated an oligosynaptic negative LFP that was localized to the wrist muscle motor pool. Spinal LTP remained robust, despite pharmacological blockade of iTBS-induced LTP within MCX using MK801, showing that activity-dependent spinal plasticity can be induced without concurrent MCX LTP. Pyramidal tract iTBS, which preferentially activates the CST, also produced significant spinal LTP, indicating the capacity for plasticity at the CST-spinal interneuron synapse. Our findings show CST monosynaptic LTP in spinal interneurons and demonstrate that spinal premotor circuits are capable of further modifying descending MCX control signals in an activity-dependent manner.

Ultrasound-on-chip platform for medical imaging, analysis, and collective intelligence.

Over the past half-century, ultrasound imaging has become a key technology for assessing an ever-widening range of medical conditions at all stages of life. Despite ultrasound's proven value, expensive systems that require domain expertise in image acquisition and interpretation have limited its broad adoption. The proliferation of portable and low-cost ultrasound imaging can improve global health and also enable broad clinical and academic studies with great impact on the fields of medicine. Here, we describe the design of a complete ultrasound-on-chip, the first to be cleared by the Food and Drug Administration for 13 indications, comprising a two-dimensional array of silicon-based microelectromechanical systems (MEMS) ultrasonic sensors directly integrated into complementary metal-oxide-semiconductor-based control and processing electronics to enable an inexpensive whole-body imaging probe. The fabrication and design of the transducer array with on-chip analog and digital circuits, having an operating power consumption of 3 W or less, are described, in which approximately 9,000 seven-level feedback-based pulsers are individually addressable to each MEMS element and more than 11,000 amplifiers, more than 1,100 analog-to-digital converters, and more than 1 trillion operations per second are implemented. We quantify the measured performance and the ability to image areas of the body that traditionally takes three separate probes. Additionally, two applications of this platform are described-augmented reality assistance that guides the user in the acquisition of diagnostic-quality images of the heart and algorithms that automate the measurement of cardiac ejection fraction, an indicator of heart health.

Inter- and Intra-Pathogen Interactions Emanating from Coinfection with Different Fungal and Viral strains in Canola Cultivars with Differing Host Resistances.

Few recent investigations examine coinfection interactions between fungal and viral plant pathogens. Here, we investigated coinfections between Leptosphaeria maculans and turnip mosaic virus (TuMV) in canola (Brassica napus). Different combinations of L. maculans isolate P11 and resistance breaking isolates L. maculans UWA192 and TuMV 12.1, were inoculated to three cultivars with differing pathogen resistances/susceptibilities. They were inoculated first to entire or half cotyledons 10-12 days after emergence, and second to opposite entire or half cotyledons on the same day (day 0) or 3 or 7 days afterwards. The parameters measured were L. maculans cotyledon disease index (%CDI), and TuMV systemically infected leaf symptom intensity (SI) and virus concentration (VC). Except when both day 0 inoculations were with isolate UWA192, %CDI values were supressed strongly or only weakly when isolates P11 and/or UWA192 were inoculated to plants with L. maculans single gene resistance (SGR) or polygenic resistance, respectively. However, except when isolate P11 was inoculated first and UWA192 second, these values declined after inoculation day 0 when SGR was absent. TuMV infection suppressed %CDI values, although this decrease was usually smaller following day 0 half cotyledon inoculations. When TuMV temperature sensitive extreme resistance was present and both inoculations were with TuMV, SI and VC values diminished greatly. However, the extent of this decrease was reduced when second inoculations were with L. maculans. SI and VC values were also smaller when SGR was present and second inoculations were with L. maculans. When L. maculans resistance was lacking, SI and VC values were smaller when second inoculations to entire cotyledons were with L. maculans rather than TuMV. This also occurred after second half cotyledon inoculations with isolate P11 but not isolate UWA192. Therefore, diverse inter- or intra-pathogen interactions developed depending upon host resistance, isolate combination, cotyledon inoculation approach and second inoculation timing.

Not so Fast: Extended Oral Antibiotic Prophylaxis Does Not Reduce 90-Day Infection Rate Following Joint Arthroplasty.

BACKGROUND: Periprosthetic joint infection (PJI) is a devastating complication following both total hip (THA) and knee (TKA) arthroplasty. Extended oral antibiotic (EOA) prophylaxis has been reported to reduce PJI following TJA in high-risk patients. The purpose of this study was to determine if EOA reduces PJI in all-comers and high-risk THA and TKA populations. METHODS: This is a retrospective cohort study, including 4,576 patients undergoing primary THA or TKA at a single institution from 2018 to 2022. Beginning in 2020, EOA prophylaxis was administered for 10 days following THA or TKA at our institution. Patients were separated into 2 cohorts (1,769 EOA, 2,807 no EOA) based on whether they received postoperative EOA. The 90-day and 1-year outcomes, with a focus on PJI, were then compared between groups. A subgroup analysis of high-risk patients was also performed. RESULTS: There was no difference in 90-day PJI rates between cohorts (EOA 1 versus no EOA 0.8%; P = .6). The difference in the rate of PJI remained insignificant at 1 year (EOA 1 versus no EOA 1%; P = .9). Similarly, our subgroup analysis of high-risk patients demonstrated no difference in postoperative PJI between EOA (n = 254) and no EOA (n = 396) (0.8 versus 2.3%, respectively; P = .2). Reassuringly, we also found no differences in the incidence of Clostridium difficile infection (EOA 0.1 versus no EOA 0.1%; P > .9) or in antibiotic resistance among those who developed PJI within 90 days (EOA 59 versus no EOA 83%; P = .2). CONCLUSIONS: With the numbers available for analysis, EOA prophylaxis was not associated with PJI risk reduction following primary TJA when universally deployed. Furthermore, among high-risk patients, there was no statistically significant difference. While we did not identify increased antibiotic resistance or Clostridium difficile infection, we cannot recommend wide-spread adoption of EOA prophylaxis, and clarification regarding the role of EOA, even in high-risk patients, is needed.

Assessment of owner perceptions of caregiver burden, veterinarian-client relationship and satisfaction with the provider in canine pruritus: An experimental vignette study.

BACKGROUND: Complicated treatments for skin disease are linked to owner-caregiver burden and poorer perception of the veterinarian-client relationship, regardless of disease severity. HYPOTHESES/OBJECTIVES: Using experimental vignettes, we explored the impact on owner perception of the interaction of treatment complexity and skin disease outcomes. We hypothesised that: (i) vignette conditions involving injection therapy would result in lower burden, better veterinary-client relationship and greater satisfaction relative to multimodal therapy conditions; (ii) the vignette condition of injection therapy with a completely effective outcome would be superior to all other conditions; (iii) ineffective vignette conditions would be inferior to all other conditions; and (iv) the vignette condition injection with a mostly effective outcome would be similar or superior to the multimodal therapy with a completely effective outcome condition. PARTICIPANTS: Three hundred and nine owners of pruritic dogs recruited from a general veterinary practice, pet-related podcast, or social media dog allergy group. MATERIALS AND METHODS: Participants were presented with six counterbalanced online vignettes representing three levels of treatment effectiveness (Completely Effective, Mostly Effective, Ineffective) and two treatment regimens (Multimodal, Injection). Measurements of participant perceptions of caregiver burden, veterinarian-client relationship and satisfaction were recorded. RESULTS: Injection with perfect outcome was superior to other conditions (p < 0.001). Conditions with poor effectiveness were inferior (p < 0.001). Comparison of Injection with a mostly effective outcome to Multimodal treatment with perfect outcome yielded small-to-medium effects of preference for the latter in veterinarian-client relationship and satisfaction (p < 0.01); no difference was observed for caregiver burden. When good effectiveness was assured, injection was preferred (p < 0.001). CONCLUSIONS AND CLINICAL RELEVANCE: Owners preferred a Completely Effective outcome and were prepared to select the Injection regimen or Multimodal therapy to achieve this; Injection was preferred when effectiveness was assured.

Relationships between physical and immunological tumor microenvironment in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is physically palpated as a hard tumor with an unfavorable prognosis. Assessing physical features and their association with pathological features could help to elucidate the mechanism of physical abnormalities in cancer tissues. A total of 93 patients who underwent radical surgery for pancreatic and bile duct cancers at a single center hospital during a 28-month period were recruited for this study that aimed to estimate the stiffness of PDAC tissues compared to the other neoplasms and assess relationships between tumor stiffness and pathological features. Physical alterations and pathological features of PDAC, with or without preoperative therapy, were analyzed. The immunological tumor microenvironment was evaluated using multiplexed fluorescent immunohistochemistry. The stiffness of PDAC correlated with the ratio of Azan-Mallory staining, α-smooth muscle actin, and collagen I-positive areas of the tumors. Densities of CD8+ T cells and CD204+ macrophages were associated with tumor stiffness in cases without preoperative therapy. Pancreatic ductal adenocarcinoma treated with preoperative therapy was softer than that without, and the association between tumor stiffness and immune cell infiltration was not shown after preoperative therapy. We observed the relationship between tumor stiffness and immunological features in human PDAC for the first time. Immune cell densities in the tumor center were smaller in hard tumors than in soft tumors without preoperative therapies. Preoperative therapy could alter physical and immunological aspects, warranting further study. Understanding of the correlations between physical and immunological aspects could lead to the development of new therapies.

Combined Drainage and Protocolized Necrosectomy Through a Coaxial Lumen-apposing Metal Stent for Pancreatic Walled-off Necrosis: A Prospective Multicenter Trial.

OBJECTIVE: We evaluated a protocolized endoscopic necrosectomy approach with a lumen-apposing metal stent (LAMS) in patients with large symptomatic walled-off pancreatic necrosis (WON) comprising significant necrotic content, with or without infection. SUMMARY BACKGROUND DATA: Randomized trials have shown similar efficacy of endoscopic treatment compared with surgery for infected WON. DESIGN: We conducted a regulatory, prospective, multicenter single-arm clinical trial examining the efficacy and safety of endoscopic ultrasound -guided LAMS with protocolized necrosectomy to treat symptomatic WON ≥6 cm in diameter with >30% solid necrosis. After LAMS placement, protocolized WON assessment was conducted and endoscopic necrosectomy was performed for insufficient WON size reduction and persistent symptoms. Patients with radiographic WON resolution to ≤ 3 cm and/or 60-day LAMS indwell had LAMS removal, then 6-month follow-up. Primary endpoints were probability of radiographic resolution by 60 days and procedure-related serious adverse events. RESULTS: Forty consecutive patients were enrolled September 2018 to March 2020, of whom 27 (67.5%) were inpatients and 19 (47.5%) had clinical evidence of infection at their index procedure. Mean WON size was 15.0 ± 5.6 cm with mean 53.2% ± 16.7% solid necrosis. Radiographic WON resolution was seen in 97.5% (95% CI, 86.8%, 99.9%) by 60 days, without recurrence in 34 patients with 6-month follow-up data. Mean time to radiographic WON resolution was 34.1 ± 16.8 days. Serious adverse events occurred in 3 patients (7.5%), including sepsis, vancomycin-resistant enterococcal bacteremia and shock, and upper gastrointestinal bleeding. There were no procedure-related deaths. CONCLUSIONS: Endoscopic ultrasound-guided drainage with protocolized endoscopic necrosectomy to treat large symptomatic or infected walled-off necrotic pancreatic collections was highly effective and safe. Clinicaltrials.-gov no: NCT03525808.

Turgor-dependent and coronin-mediated F-actin dynamics drive septin disc-to-ring remodeling in the blast fungus Magnaporthe oryzae.

The fungus Magnaporthe oryzae uses a specialized pressure-generating infection cell called an appressorium to break into rice leaves and initiate disease. Appressorium functionality is dependent on the formation of a cortical septin ring during its morphogenesis, but precisely how this structure assembles is unclear. Here, we show that F-actin rings are recruited to the circumference of incipient septin disc-like structures in a pressure-dependent manner, and that this is necessary for their contraction and remodeling into rings. We demonstrate that the structural integrity of these incipient septin discs requires both an intact F-actin and microtubule cytoskeleton and provide fundamental new insight into their functional organization within the appressorium. Lastly, using proximity-dependent labeling, we identify the actin modulator coronin as a septin-proximal protein and show that F-actin-mediated septin disc-to-ring remodeling is perturbed in the genetic absence of coronin. Taken together, our findings provide new insight into the dynamic remodeling of infection-specific higher-order septin structures in a globally significant fungal plant pathogen.

A Protocolized Management of Walled-Off Necrosis (WON) Reduces Time to WON Resolution and Improves Outcomes.

BACKGROUND AND AIMS: Patients with infected or symptomatic walled-off necrosis (WON) have high morbidity and health care utilization. Despite the recent adoption of nonsurgical treatment approaches, WON management remains nonalgorithmic. We investigated the impact of a protocolized early necrosectomy approach compared with a nonprotocolized, clinician-driven approach on important clinical outcomes. METHODS: Records were reviewed for consecutive patients with WON who underwent a protocolized endoscopic drainage with a lumen-apposing metal stent (cases), and for patients with WON treated with a lumen-apposing metal stent at the same tertiary referral center who were not managed according to the protocol (control subjects). The protocol required repeat cross-sectional imaging within 14 days after lumen-apposing metal stent placement, with regularly scheduled endoscopic necrosectomy if WON diameter reduction was <50%. Control patients were treated according to their clinician's preference without an a priori strategy. Inverse probability of treatment weighting-adjusted analysis was used to evaluate the influence of being in the protocolized group on time to resolution. RESULTS: A total of 24 cases and 47 control subjects were included. There were no significant differences in baseline characteristics. Although numbers of endoscopies and necrosectomies were similar, cases had lower adverse event rates, shorter intensive care unit stay, and required nutritional support for fewer days. On matched multivariate Cox regression, cases had earlier WON resolution (hazard ratio, 5.73; 95% confidence interval, 2.62-12.5). This was confirmed in the inverse probability of treatment weighting-adjusted analysis (hazard ratio, 3.4; 95% confidence interval, 1.92-6.01). CONCLUSIONS: A protocolized strategy resulted in faster WON resolution compared with a discretionary approach without the need for additional therapeutic interventions, and with a better safety profile and decreased health care utilization.

Septum-associated microtubule organizing centers within conidia support infectious development by the blast fungus Magnaporthe oryzae.

Cytoplasmic microtubule arrays play important and diverse roles within fungal cells, including serving as molecular highways for motor-driven organelle motility. While the dynamic plus ends of cytoplasmic microtubules are free to explore the cytoplasm through their stochastic growth and shrinkage, their minus ends are nucleated at discrete organizing centers, composed of large multi-subunit protein complexes. The location and composition of these microtubule organizing centers varies depending on genus, cell type, and in some instances cell-cycle stage. Despite their obvious importance, our understanding of the nature, diversity, and regulation of microtubule organizing centers in fungi remains incomplete. Here, using three-color fluorescence microscopy based live-cell imaging, we investigate the organization and dynamic behavior of the microtubule cytoskeleton within infection-related cell types of the filamentous fungus,Magnaporthe oryzae, a highly destructive pathogen of rice and wheat. We provide data to support the idea that cytoplasmic microtubules are nucleated at septa, rather than at nuclear spindle pole bodies, within the three-celled blast conidium, and provide new insight into remodeling of the microtubule cytoskeleton during nuclear division and inheritance. Lastly, we provide a more complete picture of the architecture and subcellular organization of the prototypical blast appressorium, a specialized pressure-generating cell type used to invade host tissue. Taken together, our study provides new insight into microtubule nucleation, organization, and dynamics in specialized and differentiated fungal cell types.

Risk Adjustment of ICD-10-CM Coded Potential Inpatient Complications Using Administrative Data.

OBJECTIVE: To risk-adjust the Potential Inpatient Complication (PIC) measure set and propose a method to identify large deviations between observed and expected PIC counts. DATA SOURCES: Acute inpatient stays from the Premier Healthcare Database from January 1, 2019 to December 31, 2021. STUDY DESIGN: In 2014, the PIC list was developed to identify a broader set of potential complications that can occur as a result of care decisions. Risk adjustment for 111 PIC measures is performed across 3 age-based strata. Using patient-level risk factors and PIC occurrences, PIC-specific probabilities of occurrence are estimated through multivariate logistic regression models. Poisson Binomial cumulative mass function estimates identify deviations between observed and expected PIC counts across levels of patient-visit aggregation. Area under the curve (AUC) estimates are used to demonstrate PIC predictive performance in an 80:20 derivation-validation split framework. DATA COLLECTION/EXTRACTION METHODS: We used N=3,363,149 administrative hospitalizations between 2019 and 2021 from the Premier Healthcare Database. PRINCIPAL FINDINGS: PIC-specific model predictive performance was strong across PICs and age strata. Average area under the curve estimates across PICs were 0.95 (95% CI: 0.93-0.96), 0.91 (95% CI: 0.90-0.93), and 0.90 (95% CI: 0.89-0.91) for the neonate and infant, pediatric, and adult strata, respectively. CONCLUSIONS: The proposed method provides a consistent quality metric that adjusts for the population's case mix. Age-specific risk stratification further addresses currently ignored heterogeneity in PIC prevalence across age groups. Finally, the proposed aggregation method identifies large PIC-specific deviations between observed and expected counts, flagging areas with a potential need for quality improvements.