Savi Scout Localization for Extrapelvic Endometriosis Resection.

BACKGROUND Endometriosis is a common cause of chronic pelvic pain among women globally. Pharmacological therapy for endometriosis includes non-steroidal anti-inflammatory drugs (NSAIDs) and hormonal contraceptives, while surgical therapy often involves either laparoscopic excision and ablation of endometriosis implants or open surgery. Surgical therapy is one of the mainstays of treatment especially for extrapelvic endometriomas. However, little guidance exists for the treatment of non-palpable or intermittently palpable lesions of this nature. CASE REPORT A 33-year-old woman with a previous cesarean section presented with complaints of intermittent discomfort in the area between her umbilicus and the surgical incision, for the previous 7 years, that worsened during her menstrual cycle. A 3×3-cm area of fullness was only intermittently palpable during various clinic visits, but was visualizable on computed tomography and magnetic resonance imaging. Given the lesion's varying palpability, a Savi Scout radar localization device was placed into the lesion pre-operatively to aid with surgical resection. The mass was excised, pathologic examination revealed endometrial tissue, and the patient had an uncomplicated postoperative course with resolution of her symptoms. CONCLUSIONS Surgical removal of extrapelvic endometrioma lesions can be made difficult by varying levels of palpability or localizability due to a patient's menstrual cycle. The Savi Scout, most commonly used in breast mass localization, is a useful tool in guiding surgical excision of non-palpable or intermittently palpable extrapelvic endometrioma lesions.

A Rare Case of Ruptured Appendicitis Secondary to Metastatic Ovarian Cancer.

BACKGROUND Patients with advanced stage ovarian cancer typically have vague non-specific abdominal symptoms related to pelvic tumor, metastasis, and ascites. When these patients present with more acute abdominal pain, appendicitis is rarely considered. Acute appendicitis due to metastatic ovarian cancer has been sparsely documented in the medical literature; only twice, to our knowledge. CASE REPORT A 61-year-old woman with a 3-week history of abdominal pain, shortness of breath, and bloating was diagnosed with ovarian cancer after computed tomography (CT) demonstrated a large pelvic cystic and solid mass. Five weeks later she underwent an omental biopsy to determine cell type and potential upstaging of the ovarian cancer to stage IV, as other aggressive cancers such as breast cancer can also involve the pelvis/omentum. Seven hours after her biopsy, she presented with increasing abdominal pain. Post-biopsy complications such as hemorrhage or bowel perforation were initially suspected to be the cause of her abdominal pain. However, CT demonstrated ruptured appendicitis. The patient underwent an appendectomy and histopathologic examination of the specimen revealed infiltration by low-grade ovarian serous carcinoma. CONCLUSIONS Given the low incidence of spontaneous acute appendicitis in this patient's age group, and the lack of any other clinical, surgical, or histopathological evidence to suggest another cause, metastatic disease was ruled to be the likely source of her acute appendicitis. Providers should be aware of appendicitis in a broad differential diagnosis and have a low threshold for ordering abdominal pelvis CT when advanced stage ovarian cancer patients present with acute abdominal pain.

Lost microbes of COVID-19: Bifidobacterium, Faecalibacterium depletion and decreased microbiome diversity associated with SARS-CoV-2 infection severity.

OBJECTIVE: The study objective was to compare gut microbiome diversity and composition in SARS-CoV-2 PCR-positive patients whose symptoms ranged from asymptomatic to severe versus PCR-negative exposed controls. DESIGN: Using a cross-sectional design, we performed shotgun next-generation sequencing on stool samples to evaluate gut microbiome composition and diversity in both patients with SARS-CoV-2 PCR-confirmed infections, which had presented to Ventura Clinical Trials for care from March 2020 through October 2021 and SARS-CoV-2 PCR-negative exposed controls. Patients were classified as being asymptomatic or having mild, moderate or severe symptoms based on National Institute of Health criteria. Exposed controls were individuals with prolonged or repeated close contact with patients with SARS-CoV-2 infection or their samples, for example, household members of patients or frontline healthcare workers. Microbiome diversity and composition were compared between patients and exposed controls at all taxonomic levels. RESULTS: Compared with controls (n=20), severely symptomatic SARS-CoV-2-infected patients (n=28) had significantly less bacterial diversity (Shannon Index, p=0.0499; Simpson Index, p=0.0581), and positive patients overall had lower relative abundances of Bifidobacterium (p<0.0001), Faecalibacterium (p=0.0077) and Roseburium (p=0.0327), while having increased Bacteroides (p=0.0075). Interestingly, there was an inverse association between disease severity and abundance of the same bacteria. CONCLUSION: We hypothesise that low bacterial diversity and depletion of Bifidobacterium genera either before or after infection led to reduced proimmune function, thereby allowing SARS-CoV-2 infection to become symptomatic. This particular dysbiosis pattern may be a susceptibility marker for symptomatic severity from SARS-CoV-2 infection and may be amenable to preinfection, intrainfection or postinfection intervention. TRIAL REGISTRATION NUMBER: NCT04031469 (PCR-) and 04359836 (PCR+).

Successful Bacterial Engraftment Identified by Next-Generation Sequencing Predicts Success of Fecal Microbiota Transplant for Clostridioides difficile.

BACKGROUND: The effectiveness of fecal microbiota transplantation (FMT), a treatment for Clostridioides difficile infection (CDI), is dependent on successful engraftment (incorporation) of donor stool. We present a method for evaluating engraftment success based on next-generation sequencing (NGS)-based profiling of bacterial strains present in donor and recipient stool, and we suggest its potential to guide treatment decisions. METHODS: Bacterial strains in stool samples from three patients from the clinic and one donor were analyzed via NGS and metagenomic sequencing, before and 1 month after FMT for CDI. The similarity of strains present was assessed via relative abundance, principal component analysis, Shannon and Simpson diversity indexes, and Bray-Curtis dissimilarity matrix. A positive outcome was successful engraftment, where the post-FMT sample closely resembled that of the donor and CDI was cured. RESULTS: Patients (Pts.) 1 and 2, but not Pt. 3's stool samples closely resembled the donor specimen post-FMT. Noteworthy, Pt. 3 pre-FMT sample was less similar to the donor than that of Pts. 1 and 2. All methods of assessing similarity and dissimilarity used yielded virtually identical conclusions. Pts. 1 and 2 which closely resembled donor specimen, eradicated CDI giving a surrogate objective measure of engraftment. CONCLUSIONS: Success of engraftment in FMT can be assessed using NGS and metagenomic analysis and parallels success in curing CDI of the microbiome. The statistical methods we present here are reliable and consistent for such purposes. The dissimilarity of Pt. 3 to the donor combined with the failure of engraftment and failure to cure CDI in Pt. 3 suggests that FMT success may be predictable by comparing pre-FMT samples to donor. There is no clinical trial registry listing this study.

Recurrent Macroprolactinoma with Malignant Conversion to Carcinoma with Spinal Metastasis.

In contrast to pituitary adenomas, pituitary carcinomas represent a rare malignant neoplasm with a remarkable high mortality. Pituitary carcinomas can arise from any pituitary tumor cell line and are determined to be carcinomas when there is distant metastasis or central nervous system dissemination. In this case vignette, we describe a rare case of malignant prolactinoma with intraspinal metastasis, and we also provide a review of relevant literature and treatment.

Detection of SARS-CoV-2 from patient fecal samples by whole genome sequencing.

BACKGROUND: SARS-CoV-2 has been detected not only in respiratory secretions, but also in stool collections. Here were sought to identify SARS-CoV-2 by enrichment next-generation sequencing (NGS) from fecal samples, and to utilize whole genome analysis to characterize SARS-CoV-2 mutational variations in COVID-19 patients. RESULTS: Study participants underwent testing for SARS-CoV-2 from fecal samples by whole genome enrichment NGS (n = 14), and RT-PCR nasopharyngeal swab analysis (n = 12). The concordance of SARS-CoV-2 detection by enrichment NGS from stools with RT-PCR nasopharyngeal analysis was 100%. Unique variants were identified in four patients, with a total of 33 different mutations among those in which SARS-CoV-2 was detected by whole genome enrichment NGS. CONCLUSION: These results highlight the potential viability of SARS-CoV-2 in feces, its ongoing mutational accumulation, and its possible role in fecal-oral transmission. This study also elucidates the advantages of SARS-CoV-2 enrichment NGS, which may be a key methodology to document complete viral eradication. Trial registration ClinicalTrials.gov, NCT04359836, Registered 24 April 2020, https://clinicaltrials.gov/ct2/show/NCT04359836?term=NCT04359836&draw=2&rank=1 ).

Surgical Management of Giant Colonic Diverticulum Presenting After Laparoscopic Lavage for Diverticular Perforation and Sepsis.

BACKGROUND Diverticulosis and its complication of diverticulitis is a common condition that can be found in up to 35% of the population. Giant colonic diverticulum is a rare entity with fewer than 200 cases reported in the scientific literature. Development of a giant diverticulum as a sequelae of laparoscopic washout is an unreported event in current literature. CASE REPORT The patient was a 74-year-old female who had a well-known history of diverticulosis and diverticulitis. She developed perforated sigmoid diverticulitis, underwent laparoscopic washout and recovered without colon resection. Within a year after washout, she developed abdominal distention and bloating, and computed tomography (CT) imaging revealed a giant diverticulum. She went on to undergo surgery for resection of her sigmoid colon, which contained the giant diverticulum. Her recovery was otherwise uneventful. CONCLUSIONS To our knowledge, this is the first case report of giant diverticulum presenting as a complication of abdominal washout for management of acute diverticulitis. Initial CT scan performed at the time of perforation did not demonstrate this diverticulum, indicating that it developed within the year after abdominal washout for sepsis and acute rupture, likely due to weakening of the colonic wall secondary to ongoing inflammation. The very rare presentation of giant diverticulum makes it difficult to establish a clear link to washout, however, this case establishes a groundwork for further investigation as our fund of knowledge on the subject continues to grow.

Low-Grade Appendiceal Mucinous Neoplasms: A Single Institution Experience of 64 Cases With Clinical Follow-up and Correlation With the Current (Eighth Edition) AJCC Staging.

Background. In this single-institution study, we applied the current (eighth edition) American Joint Committee on Cancer pathologic staging criteria to 64 low-grade mucinous neoplasms of the appendix (LAMNs), examined their histopathologic features, and studied the patients' clinical outcomes. Design. Sixty-four LAMNs, with a median follow-up of 52 months, were reviewed. Results. The distribution of pathologic stages was pTis (n = 39), pT3 (n = 1), pT4a (n = 5), pT4aM1a (n = 8), and pT4aM1b (n = 11). Recurrence was observed in only 2 patients (both with pT4aM1b disease), one of whom died of disease. All remaining patients were disease-free after a median clinical follow-up of 60 months. Conclusions. Our study confirms that pTis LAMNs have an excellent prognosis and suggests that pT4a and pT4aM1a LAMNs may also have a low risk of developing progressive disease.

Resolution of Diffuse Intrahepatic Biliary Strictures after Chemotherapy for Metastatic Ovarian Cancer.

Sclerosing cholangitis and cholestatic jaundice secondary to metastatic disease is a rare complication. We report a rare case of secondary sclerosing cholangitis (SSC) due to lymphatic spread from ovarian cancer with complete resolution after chemotherapy. The diagnosis of SSC from metastatic ovarian cancer was clinically challenging, as endoscopic retrograde cholangiopancreatography revealed irregular hepatic ducts consistent with sclerosing cholangitis, but it did not identify any malignant cells. The final diagnosis was made with liver biopsy revealing high-grade metastatic Mullerian carcinoma. The patient responded well to chemotherapy and is in remission. A timely diagnosis is important and can lead to complete resolution of the disease.

Use of the ADAMTS13 activity assay improved the accuracy and efficiency of the diagnosis and treatment of suspected acquired thrombotic thrombocytopenic purpura.

CONTEXT: Acquired thrombotic thrombocytopenic purpura (A-TTP) is a rare but significant disease requiring rapid diagnosis and treatment. The diagnosis is often difficult because of variability in the presence of specific clinical criteria. The primary etiology of A-TTP involves inhibitors directed against ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13). Literature has shown that the ADAMTS13 activity assay is sensitive and specific for identifying cases of A-TTP, and application of this test as an on-site screening method has not been fully explored. OBJECTIVE: Our objective is to determine if the ADAMTS13 activity assay can be used as a successful, on-site diagnostic modality to rapidly identify cases of A-TTP and prevent unnecessary use of prophylactic therapeutic plasma exchange. DESIGN: A retrospective analysis was performed including 152 patients with clinically suspected A-TTP, screened using the ADAMTS13 activity assay. Results were correlated with potential therapeutic plasma exchange treatment for all cases highly suspicious for A-TTP and evaluated for unnecessary patient morbidity and financial cost. RESULTS: The ADAMTS13 activity assay had an overall sensitivity and specificity of 100% and 99%, respectively. The positive predictive value was 91% and the negative predictive value was 100%. In 95% of the studies ordered, A-TTP was ruled out, leading to decreased patient morbidity and $1.7 million of potential treatment costs avoided. CONCLUSION: Implementation of the fluorescence energy transfer-based ADAMTS13 activity assay as a point-of-care laboratory study decreased patient morbidity while also directing more efficient employment of therapeutic plasma exchange in cases of suspected A-TTP.

Evidence of ambiguous differentiation and mTOR pathway dysregulation in subependymal giant cell astrocytoma.

OBJECTIVE: The exact cell of origin of subependymal giant cell astrocytoma is debated but most currently consider the tumor in the astrocytic category. Mutations and subsequent biallelic inactivation of TSC1 encoding hamartin, or TSC2 encoding tuberin appear to be the underlying genetic aberrations. Inactivation leads to loss of proteins that inhibit mammalian target of rapamycin (mTOR ) disrupting tightly regulated cell functions. MATERIAL AND METHOD: We analyzed the expression of tuberin and hamartin along with an array of neuroepithelial markers in 9 subependymal giant cell astrocytomas. In addition, RPS6 and 4EBP1 regulatory proteins that are downstream in the mTOR pathway were also evaluated. RESULTS: While hamartin and tuberin expression levels were relatively decreased compared to control tissue, this was not of particular practical use to detect the mutated gene since low levels of positivity could be detected throughout the central nervous system. As expected, the levels of RPS6 and 4EBP1 were increased, further confirming the activation of the mTOR pathway. GFAP was positive in 5 cases, while Synaptophysin positivity was found in all tumors. CD34 (a marker often observed in well differentiated glio-neuronal tumors), Olig2 (a nuclear marker present in most gliomas), IDH1 and IDH2 were entirely negative in all tumor cells. Ki67 (MIB-1) showed a low proliferation rate ranging from 2% to 8%. CONCLUSION: Staining with neuroepithelial markers supports the suggestion of ambiguous differentiation. Subependymal giant cell astrocytomas do not appear to have the typical expression profiles of astrocytic tumors, under which they have been classified.

Extracellular potassium dependency of block of HERG by quinidine and cisapride is primarily determined by the permeant ion and not by inactivation.

Drug induced Long QT syndrome results primarily from block of the cardiac potassium channel HERG (human-ether-a-go-go related gene). In some cases prolongation of the QT interval can result in the lethal arrhythmia torsade de pointes, an arrhythmia characterized by a rapid heart rate and severely compromised cardiac output. Many patients requiring medication present with abnormal serum electrolyte levels due to a variety of conditions including gastrointestinal dysfunction, renal and endocrine disorders, diuretic use, alcoholism and aging. Extracellular cations have significant influence on HERG channel gating and in some instances they have been shown to alter drug block of HERG. However, the mechanisms by which drug block is altered in different extracellular cation solutions are not well understood. In this study, HERG block by quinidine and cisapride was assessed in extracellular solutions of calcium, potassium, rubidium, cesium and tetraethylammonium (TEA) using two-electrode voltage clamping of Xenopus oocytes. Consistent with previous reports we show that increases in extracellular potassium reduce HERG block by quinidine and cisapride. We also show that increasing extracellular rubidium and cesium reduced HERG block by quinidine and cisapride whereas increasing extracellular calcium and extracellular TEA did not alter HERG block by quinidine and cisapride. These results demonstrate that at lower extracellular potassium concentrations, the permeant ion is almost exclusively responsible for the reduction in quinidine and cisapride block of HERG due to increases in extracellular potassium.

Resistance is non-futile: resistance to Cry5B in the nematode Caenorhabditis elegans.

The nematode, Caenorhabditis elegans, can be mutated to resistance to the Cry5B toxin of Bacillus thuringiensis. By cloning and characterization of these C. elegans resistance genes, we have determined that a major mechanism by which C. elegans resists Cry5B is by loss of function mutations in any one of four gylcosyltransferase genes that glycosylate glycolipids specific to arthropods. Without correct gylcosylation, binding of Cry5B is greatly impaired in C. elegans. That these specific arthroseries glycolipids do not occur in vertebrates potentially helps explain why Cry toxins are specific for arthropods.

Resistance to Bacillus thuringiensis toxin in Caenorhabditis elegans from loss of fucose.

A mutation in the Caenorhabditis elegans bre-1 gene was isolated in a screen for Bacillus thuringiensis toxin-resistant (bre) mutants to the Cry5B crystal toxin made by B. thuringiensis. bre-1 mutant animals are different from the four other cloned bre mutants in that their level of resistance is noticeably lower. bre-1 animals also display a significantly reduced brood size at 25 degrees C. Here we cloned the bre-1 gene and characterized the bre-1 mutant phenotype. bre-1 encodes a protein with significant homology to a GDP-mannose 4,6-dehydratase, which catalyzes the first step in the biosynthesis of GDP-fucose from GDP-mannose. Injection of GDP-fucose but not fucose into C. elegans intestinal cells rescues bre-1 mutant phenotypes. Thus, C. elegans lacks a functional fucose salvage pathway. Furthermore, we demonstrate that bre-1 mutant animals are defective in production of fucosylated glycolipids and that bre-1 mutant animals make quantitatively reduced levels of glycolipid receptors for Cry5B. We finally show that bre-1 mutant animals, although viable, show a lack of fucosylated N- and O-glycans, based on mass spectrometric evidence. Thus, C. elegans can survive with little fucose and can develop resistance to crystal toxin by loss of a monosaccharide biosynthetic pathway.

Caenorhabditis elegans carbohydrates in bacterial toxin resistance.

The major virulence factor produced by the bacterium Bacillus thuringiensis (Bt) is a pore-forming toxin called crystal (Cry) toxin, which targets and kills insects and nematodes. To understand how this bacterial toxin interacts with its invertebrate hosts, a genetic screen in C. elegans for nematodes resistant to Bt toxin was carried out. Four of the five genes that mutated to toxin resistance encode glycosyltransferases. These genes were found to participate in the biosynthesis of C. elegans glycosphingolipids. These glycolipids in turn were shown to directly bind Bt toxin. Thus, resistance to Bt toxin in C. elegans can develop as a result of loss of glycolipid receptors for the toxin. Here we describe the isolation of Bt toxin resistance mutants in C. elegans, isolation of C. elegans glycolipids, and their separation by thin-layer chromatography, overlay assays to demonstrate direct binding of Bt toxin to glycolipids, and the purification of specific C. elegans glycolipid species.

A purified Bacillus thuringiensis crystal protein with therapeutic activity against the hookworm parasite Ancylostoma ceylanicum.

Crystal (Cry) proteins produced by the soil bacterium Bacillus thuringiensis (Bt) are harmless to vertebrates, but they are highly toxic to insects and nematodes. Their value in controlling insects that destroy crops and transmit human diseases is well established. Although it has recently been demonstrated that a few individual Bt Cry proteins, such as Cry5B, are toxic to a wide range of free-living nematodes, the potential activity of purified Cry proteins against parasitic nematodes remains largely unknown. We report here studies aimed at characterizing in vitro and in vivo anthelminthic activities of purified recombinant Cry5B against the hookworm parasite Ancylostoma ceylanicum, a bloodfeeding gastrointestinal nematode for which humans are permissive hosts. By using in vitro larval development assays, Cry5B was found to be highly toxic to early stage hookworm larvae. Exposure of adult A. ceylanicum to Cry5B was also associated with significant toxicity, including a substantial reduction in egg excretion by adult female worms. To demonstrate therapeutic efficacy in vivo, hamsters infected with A. ceylanicum were treated with three daily oral doses of purified Cry5B, the benzimidazole anthelminthic mebendazole, or buffer. Compared with control (buffer-treated) animals, infected hamsters that received Cry5B showed statistically significant improvements in growth and blood hemoglobin levels as well as reduced worm burdens that were comparable to the mebendazole-treated animals. These data demonstrate that Cry5B is highly active in vitro and in vivo against a globally significant nematode parasite and that Cry5B warrants further clinical development for human and veterinary use.

Resistance to a bacterial toxin is mediated by removal of a conserved glycosylation pathway required for toxin-host interactions.

Crystal (Cry) proteins made by the bacterium Bacillus thuringiensis are pore-forming toxins that specifically target insects and nematodes and are used around the world to kill insect pests. To better understand how pore-forming toxins interact with their host, we have screened for Caenorhabditis elegans mutants that resist Cry protein intoxication. We find that Cry toxin resistance involves the loss of two glycosyltransferase genes, bre-2 and bre-4. These glycosyltransferases function in the intestine to confer susceptibility to toxin. Furthermore, they are required for the interaction of active toxin with intestinal cells, suggesting they make an oligosaccharide receptor for toxin. Similarly, the bre-3 resistance gene is also required for toxin interaction with intestinal cells. Cloning of the bre-3 gene indicates it is the C. elegans homologue of the Drosophila egghead (egh) gene. This identification is striking given that the previously identified bre-5 has homology to Drosophila brainiac (brn) and that egh-brn likely function as consecutive glycosyltransferases in Drosophila epithelial cells. We find that, like in Drosophila, bre-3 and bre-5 act in a single pathway in C. elegans. bre-2 and bre-4 are also part of this pathway, thereby extending it. Consistent with its homology to brn, we demonstrate that C. elegans bre-5 rescues the Drosophila brn mutant and that BRE-5 encodes the dominant UDP-GlcNAc:Man GlcNAc transferase activity in C. elegans. Resistance to Cry toxins has uncovered a four component glycosylation pathway that is functionally conserved between nematodes and insects and that provides the basis of the dominant mechanism of resistance in C. elegans.