Endolysin Regulation in Phage Mu Lysis.

Bacteriophage Mu is a paradigm coliphage studied mainly because of its use of transposition for genome replication. However, in extensive nonsense mutant screens, only one lysis gene has been identified, the endolysin gp22. This is surprising because in Gram-negative hosts, lysis by Caudovirales phages has been shown to require proteins which disrupt all three layers of the cell envelope. Usually this involves a holin, an endolysin, and a spanin targeting the cytoplasmic membrane, peptidoglycan (PG), and outer membrane (OM), respectively, with the holin determining the timing of lysis initiation. Here, we demonstrate that gp22 is a signal-anchor-release (SAR) endolysin and identify gp23 and gp23.1 as two-component spanin subunits. However, we find that Mu lacks a holin and instead encodes a membrane-tethered cytoplasmic protein, gp25, which is required for the release of the SAR endolysin. Mutational analysis showed that this dependence on gp25 is conferred by lysine residues at positions 6 and 7 of the short cytoplasmic domain of gp22. gp25, which we designate as a releasin, also facilitates the release of SAR endolysins from other phages. Moreover, the entire length of gp25, including its N-terminal transmembrane domain, belongs to a protein family, DUF2730, found in many Mu-like phages, including those with cytoplasmic endolysins. These results are discussed in terms of models for the evolution and mechanism of releasin function and a rationale for Mu lysis without holin control. IMPORTANCE Host cell lysis is the terminal event of the bacteriophage infection cycle. In Gram-negative hosts, lysis requires proteins that disrupt each of the three cell envelope components, only one of which has been identified in Mu: the endolysin gp22. We show that gp22 can be characterized as a SAR endolysin, a muralytic enzyme that activates upon release from the membrane to degrade the cell wall. Furthermore, we identify genes 23 and 23.1 as spanin subunits used for outer membrane disruption. Significantly, we demonstrate that Mu is the first known Caudovirales phage to lack a holin, a protein that disrupts the inner membrane and is traditionally known to release endolysins. In its stead, we report the discovery of a lysis protein, termed the releasin, which Mu uses for SAR endolysin release. This is an example of a system where the dynamic membrane localization of one protein is controlled by a secondary protein.

Crowdsourcing biocuration: The Community Assessment of Community Annotation with Ontologies (CACAO).

Experimental data about gene functions curated from the primary literature have enormous value for research scientists in understanding biology. Using the Gene Ontology (GO), manual curation by experts has provided an important resource for studying gene function, especially within model organisms. Unprecedented expansion of the scientific literature and validation of the predicted proteins have increased both data value and the challenges of keeping pace. Capturing literature-based functional annotations is limited by the ability of biocurators to handle the massive and rapidly growing scientific literature. Within the community-oriented wiki framework for GO annotation called the Gene Ontology Normal Usage Tracking System (GONUTS), we describe an approach to expand biocuration through crowdsourcing with undergraduates. This multiplies the number of high-quality annotations in international databases, enriches our coverage of the literature on normal gene function, and pushes the field in new directions. From an intercollegiate competition judged by experienced biocurators, Community Assessment of Community Annotation with Ontologies (CACAO), we have contributed nearly 5,000 literature-based annotations. Many of those annotations are to organisms not currently well-represented within GO. Over a 10-year history, our community contributors have spurred changes to the ontology not traditionally covered by professional biocurators. The CACAO principle of relying on community members to participate in and shape the future of biocuration in GO is a powerful and scalable model used to promote the scientific enterprise. It also provides undergraduate students with a unique and enriching introduction to critical reading of primary literature and acquisition of marketable skills.

Insights from the reanalysis of high-throughput chemical genomics data for Escherichia coli K-12.

Despite the demonstrated success of genome-wide genetic screens and chemical genomics studies at predicting functions for genes of unknown function or predicting new functions for well-characterized genes, their potential to provide insights into gene function has not been fully explored. We systematically reanalyzed a published high-throughput phenotypic dataset for the model Gram-negative bacterium Escherichia coli K-12. The availability of high-quality annotation sets allowed us to compare the power of different metrics for measuring phenotypic profile similarity to correctly infer gene function. We conclude that there is no single best method; the three metrics tested gave comparable results for most gene pairs. We also assessed how converting quantitative phenotypes to discrete, qualitative phenotypes affected the association between phenotype and function. Our results indicate that this approach may allow phenotypic data from different studies to be combined to produce a larger dataset that may reveal functional connections between genes not detected in individual studies.

Novel COVID-19 phenotype definitions reveal phenotypically distinct patterns of genetic association and protective effects

INTRODUCTION PARAGRAPHMultiple large COVID-19 genome-wide association studies (GWAS) have identified reproducible genetic associations indicating that some infection susceptibility and severity risk is heritable.1-5 Most of these studies ascertained COVID-19 cases in medical clinics and hospitals, which can lead to an overrepresentation of cases with severe outcomes, such as hospitalization, intensive care unit admission, or ventilation. Here, we demonstrate the utility and validity of deep phenotyping with self-reported outcomes in a population with a large proportion of mild and subclinical cases. Using these data, we defined eight different phenotypes related to COVID-19 outcomes: four that align with previously studied COVID-19 definitions and four novel definitions that focus on susceptibility given exposure, mild clinical manifestations, and an aggregate score of symptom severity. We assessed replication of 13 previously identified COVID-19 genetic associations with all eight phenotypes and found distinct patterns of association, most notably related to the chr3/SLC6A20/LZTFL1 and chr9/ABO regions. We then performed a discovery GWAS, which suggested some novel phenotypes may better capture protective associations and also identified a novel association in chr11/GALNT18 that reproduced in two fully independent populations.

The Diagnostic and Treatment Patterns of Urologists in the United States for Interstitial Cystitis/Painful Bladder Syndrome.

INTRODUCTION: Interstitial cystitis/bladder pain syndrome is not completely understood, making it challenging to diagnose and treat. The current literature elucidating this disease process is inconsistent. Without a clear consensus regarding management it is important to evaluate how urologists are treating these patients. METHODS: Urologists across the United States completed a 19-item survey addressing diagnostic and treatment methods for interstitial cystitis. Participation was voluntary and no compensation was provided to complete the survey. RESULTS: A total of 95 surveys were completed and returned between December 2012 and January 2013. Of the respondents 92% considered themselves general urologists and most preferred to manage interstitial cystitis/bladder pain syndrome themselves with only 33% referring these patients. Of the respondents 47% believed that the etiology of interstitial cystitis is still unknown. Cystoscopy with hydrodistention was the most common approach to diagnosis (70% of respondents) followed closely by validated symptoms scores (65%). Oral medication was the most commonly used treatment (92% of respondents), of which pentosan polysulfate was the most commonly used agent. Oral medication was followed by intravesical treatment and bladder hydrodistention at 77% and 74% of respondents, respectively. Most urologists ultimately used multimodal therapy. AUA (American Urological Association) guidelines were followed by only 15% of respondent urologists. CONCLUSIONS: The treatment of patients with interstitial cystitis/bladder pain syndrome is variable and many urologists use multiple modalities for diagnosis and treatment. This variability in diagnosis and/or treatment reflects the deficiency of our current understanding of this disease process. Until the pathophysiology is better delineated diagnosis and treatment will remain without consensus.

Successful Surgical Management of Giant Condyloma Acuminatum (Buschke Lowenstein Tumor) in the Urethra of a Female Patient: A Case Report.

The Buschke-Lowenstein tumor (BLT) is a slow-growing, locally destructive verrucous plaque that typically appears on the penis but may occur elsewhere in the anogenital region. It most commonly is considered to be a regional variant of verrucous carcinoma. It is rare but accounts for 5-24% of all penile cancers. It can also affect the perineum and other portions of the genitalia. It was first described by Buschke and Löwenstein in 1925, and is also known as giant condyloma acuminatum (GCA). Regardless of the treatment modality, careful follow-up is recommended because of the high risk of recurrence and the possibility for malignant transformation in 30-56% of patients. We present a case of a 47-year-old Hispanic female that presented to the urology clinic for dysuria and upon work-up was found to have a GCA. GCA typically affects the penis, although the perianal region, vulva, vagina, rectum, scrotum, perineum and bladder may be involved. To date, we believe this is the only reported case of GCA in the urethra of a female patient with sparing of the bladder. This lesion was successfully removed with wide local excision. We believe that further studies are needed to define this disease, identify its pathogenesis, and the most successful treatment protocol.

A Case Series of Spontaneous Rupture of the Urinary Bladder.

We report 2 cases of spontaneous bladder rupture related to chronic outlet obstruction and urinary retention. In both cases, focal perforation was identified within diverticula. Bladder rupture in the absence of trauma is a rare and serious event with a mortality rate approaching 50%. These injuries are often initially misdiagnosed and it is our goal to provide insight to the presentation, management and treatment of this rare event.

Predicting Acute Urinary Retention in Patients with Elevated Post-Void Residuals.

OBJECTIVES: To perform a retrospective analysis evaluating factors that may predict which men with elevated post-void residuals (PVRs) that were at increased risk to develop acute urinary retention (AUR). METHODS: We retrospectively analyzed the records of 44 male patients who had 2 consecutive PVRs greater than 100 ml over a 6-month period. Using regression analysis, we evaluated patient's age, PVR volume, prostate specific antigen (PSA) and transrectal ultrasound prostate volume with respect to development of AUR over 24 months. RESULTS: Of the 44 patients, 4 developed AUR. When all factors were considered, prostate volume was determined to be the only that was statistically significant (p = 0.003). A 1-SD increase in prostate volume (12 ml) led to a 19.6% increased risk of developing AUR. There was a strong correlation between PSA and prostate volume (0.787). A regression analysis was then repeated excluding prostate volume. PSA then became a statistically significant predictor of AUR (p = 0.007). A 1-SD increase in PSA (1.377 ng/ml) increased the patients' risk of developing AUR by 12.3%. CONCLUSION: In men with an elevated PVR, increased transrectal ultrasound prostate volume or PSA may help predict which patients have an elevated risk of developing AUR within the next 24 months. This information may influence which patients need early surgical intervention versus medical therapy.

Procidentia as a Cause of Obstructive Uropathy and Acute Kidney Injury.

Pelvic organ prolapse can affect urinary tract function by reducing flow rates and increasing post void residual urine volumes secondary to outlet obstruction. If the diagnosis is missed or left untreated, pelvic organ prolapse can lead to acute renal injury, chronic renal failure or even end stage renal disease. Herein, we present a case of a patient who presented to Albert Einstein Medical Center in Philadelphia, PA with urinary retention and acute kidney injury secondary to complete uterine prolapse, also referred to as procidentia.

A 52-year-old male with bilaterally duplicated collecting systems with obstructing ureteral stones: a case report.

Collecting system duplication is marked by a variety of clinical syndromes. Bilateral and obstructed duplicated systems, particularly with asymmetric levels of duplication, are rare and typically due to ureteric bud development anomalies. The infrequency with which this condition exists makes it a formidable challenge for physicians and patients. To our knowledge, we present the first case report of bilateral obstruction of bilaterally duplicated collecting systems. In our case, a 52-year-old male complaining of low back pain, constipation, urinary urgency and hematuria was found to have bilateral obstructing stones as well as asymmetrical bilateral collecting system duplication. We discuss the natural history of this condition, its consequences and identification.

Primary clear cell adenocarcinoma of a urethral diverticulum treated with multidisciplinary robotic anterior pelvic exenteration.

Primary urethral carcinoma is extremely rare and is marked by a variety of clinical symptoms. Primary carcinoma of a urethral diverticulum is still rarer and clear cell adenocarcinoma of the urethra is particularly uncommon (Swartz et al., 2006). Such infrequency has led to inadequate management guidance in the literature for a disease that is often late in presentation and carries substantial morbidity and mortality. This treatable but grave disease deserves definitive curative treatment. We present the first published instance in which it was treated with robotic anterior exenteration. In our case, a 47-year-old female was referred to the urology service for investigation of recurring urinary tract infections. During the workup, the patient was found to have an advanced clear cell urethral adenocarcinoma originating in a urethral diverticulum. We discuss the natural history of this condition, its consequences, and the first instance of its treatment using robotic anterior pelvic exenteration.

How does acute urolithiasis present after renal denervation?

PURPOSE: To present our experience with the acute presentation of urolithiasis after laparoscopic renal denervation and nephropexy for intractable renal pain in the pediatric patient. PATIENTS AND METHODS: Five patients aged 8 to 16 years (mean age 11.4 years) with a history of laparoscopic renal denervation presented with acute abdominal symptoms; obstructing urolithiasis was diagnosed. RESULTS: There were a total of 12 episodes of acute urolithiasis in this cohort. All five patients presented with periumbilical and epigastric discomfort when the stone was on the denervated side. During the episodes, of the five patients, three (60%) also presented with vomiting; four (80%) had at least one episode of gross hematuria; and five (100%) had microscopic hematuria during episodes on the denervated side. None of the patients presented with flank discomfort on the denervated side. CONCLUSIONS: Stone-related pain in patients undergoing laparoscopic renal denervation and nephropexy is not a typical presentation, and one needs to be wary of vague abdominal pain in this population.