- Email: [email protected]
157 Volumetric Laser Endomicroscopy Targeted Tissue Sampling Eliminates Unnecessary Biopsies During Planned Ablation of Barrett's Esophagus
Diagnostic Accuracy of EUS for Distinguishing Celiac Ganglia From Celiac Lymph Nodes Naveen Gara, Ferga Gleeson, Mark Topazian, Barham K. Abu Dayyeh, Suresh Chari, Michael B. Farnell, Prasad Iyer, Michael L. Kendrick, Randall K. Pearson, Bret T. Petersen, Elizabeth Rajan, Mark J. Truty, Santhi Swaroop Vege, Kenneth K. Wang, Michael J. Levy
155 Background: Endoscopic ultrasound (EUS) allows visualization of celiac axis lymph nodes (CLNs) and celiac ganglia (CG); however, the EUS accuracy in distinguishing between these anatomical structures is unknown. This distinction is important given the impact on clinical tumor staging and the targeting of celiac ganglia for blockade, neurolysis or other ablative therapies. Aim: To determine the EUS diagnostic accuracy in distinguishing CLNs from CG. Methods: Patients who underwent EUS FNA of either suspected CLNs or CG (10/200410/2015) were identified from a prospectively maintained EUS database. Results: 453 patients [58% male; age 65 years (15-93)] underwent EUS FNA of either suspected CLNs or CG. Most (376, 83%) procedures were performed to evaluate a broad spectrum of malignancies, most commonly pancreatic cancer (33%), esophageal cancer (21%), and cholangiocarcinoma (17%). EUS FNA was performed of: 1.) a presumed CLN [219 (48%) patients; median 4 passes, range 1-15)], 2.) a presumed CG [157 (35%) patients; 4 passes (1-11)], 3.) both CLN and CG [(34 (8%) patients; 3 passes (1-6) for LN and 3 passes (18) for CG)] and 4.) an indeterminate CLN or CG [43 (9%) patients; 3 passes (1-5)]. In total, there were 487 structures sampled whereby the endosonographer perceived that they represented CLNs (n=253), CG (n=191) or were indeterminate (n=43). The cytopathology report provided a definitive gold standard for 332 (73.3%) sites indicating either CG (based on the presence of neural structures without lymphocytes) or CLN (lymphocytes without neural glial cells). The cytopathology report for the remaining 155 patients simply assessed for the presence or absence of malignancy without mention of the presence of neuronal glial cells or lymphocytes. Based on those patients with a cytopathology gold standard, the accuracy of EUS, when the endosonographer was confident that they biopsied a CLN or CG, was 97% (165/171) or 91% (125/138), respectively. When including 43 patients in whom the site was indeterminate by the endosonographer, the diagnostic accuracy was reduced to 89% (165/186) and 86% (125/146) for CLNs and CG, respectively. Post procedure clinical follow-up within 4-7 days was available in 83% (378/453). The adverse event rate was 3.97% (15 of 378) and was comprised of abdominal pain (n=7), mild pancreatitis (n= 2), bleeding (n=2; hemobilia and post EMR bleed), and 1 patient each with duodenal perforation, fever, excessive moderate sedation and non-cardiac chest pain; most of which appear to relate to other procedures and interventions performed. Conclusions: EUS accurately distinguishes CLNs from CG. This information can be valuable in overall patient care to help enhance clinical tumor staging and for selectively targeting celiac ganglia pain management.
Tethered Capsule OCT Endomicroscopy With Micro-Motor Scanning for Esophageal Imaging Jing Dong, Michalina J. Gora, Rohith Reddy, Wolfgang K. Trasischker, Timothy N. Ford, Weina Lu, Catriona N. Grant, Amna R. Soomro, Mireille Rosenberg, Norman S. Nishioka, Guillermo J. Tearney Introduction: Our lab has previously developed and reported a new technology termed tethered capsule endomicroscopy (TCE) that utilizes optical coherence tomography (OCT) for microscopic (10µm) imaging of internal structures of the human esophagus without the need of sedation [1]. To date, we have successfully performed ~100 procedures with the TCE device. To justify its use for population based screening for diseases in upper gastrointestinal tract, we have developed a second generation device that is cost-effective and easy to manufacture, and designed to provide a comfortable experience for the patient. Materials and Methods: The new generation TCE device incorporates a micro-motor [2] at the distal end of a 11mm x 24.4mm swallowable capsule. The capsule is connected to a 1mm diameter, 1.6m long flexible tether that houses an optical fiber and two electrical wires which power up the micro-motor. The near-infrared light from the fiber is focused by the micro-optics comprising a monolithic ball lens and deflected to the side of the capsule by the angle polished shaft of the micro-motor. Rotation of the shaft effectuates circumferential scanning of the esophageal wall. After unsedated subjects swallowed the capsule, multiple crosssectional microscopic OCT images of the esophagus were obtained and visualized in real time. The capsule descended to the stomach via peristalsis and then was gradually pulled back up through the esophagus using the tether. After imaging the esophagus twice, the capsule was removed and sterilized for reuse. After the procedure, subjects were asked if they would recommend the procedure to other patients and if any capsule vibrations were felt. Results: To date, 4 healthy subjects have been enrolled in the tethered micro-motor capsule endomicroscopy study. A total of 3/4 (75%) subjects successfully swallowed the capsule. For those who swallowed the capsule, high quality microscopic images of the entire esophagus were obtained. The entire procedure lasted an average of 3±1 minutes from capsule insertion to removal. There were no complications related to the procedure. All subjects who successfully swallowed the capsule would recommend the procedure; none of the subjects felt vibration of the capsule, 1 subject reported that the new generation of TCE was more comfortable than previous TCE device. Conclusion: Our initial experience with the tethered micro-motor capsule endomicroscopy has shown that the new generation device is able to image the esophageal wall in a rapid and safe manner while rendering the same high quality, 3-dimensional microstructural images as the previous version. Additionally, because this device is simpler to manufacture and less costly, it may be desirable for the diagnosis of esophageal diseases in screening settings. [1] Gora MJ et al. Nat Med. 2013, 19(2):238-40 [2] Liang K et al. BOE. 2015, 6(4):1146-63
157 Volumetric Laser Endomicroscopy Targeted Tissue Sampling Eliminates Unnecessary Biopsies During Planned Ablation of Barrett's Esophagus Udayakumar Navaneethan, Brooks D. Cash, Matthew McKinley, Virendra Joshi, Paul R. Tarnasky, Kenneth J. Chang, Satish K. Singh, Michael B. Wallace, Michael S. Smith Background: Endoscopic ablation of Barrett's esophagus (BE) is accomplished with a variety of modalities. Tissue sampling performed during the same endoscopy can affect visualization and potentially procedure outcome. Volumetric laser endomicroscopy (VLE) uses nextgeneration optical coherence tomography to rapidly scan a large area of the esophagus to a depth of 3 mm and a resolution of 7 microns. Our aim was to assess whether VLE-targeted tissue sampling maintains adequate sampling while decreasing unnecessary biopsies at the time of BE ablation. Methods: Interim analysis was performed for a US-based multicenter registry that prospectively collected demographic, endoscopic and histologic data when VLE was used during the same endoscopy as BE field ablation. All cases were performed using the NvisionVLE™ system (NinePoint Medical, Bedford, MA). Patients could be treatment naïve or incompletely treated. Treatments permitted for this study included radiofrequency ablation (RFA), cryoablation (Cryo) or argon plasma coagulation (APC). Patients who had VLE in the same session as endoscopic mucosal resection (EMR) were excluded, as these cases underwent separate analysis. Results: A total of 182 procedures from 140 patients met inclusion criteria. The mean age was 66.5 years (range 33-88) and 81% were male. Among these patients, 37 (26%) were BE treatment naïve, while the remaining 103 previously were treated at least once. BE was seen on white light or narrow band imaging in 145/182 (80%) procedures. VLE and endoscopic visualization identified abnormalities that led to targeted biopsies in 57/182 (31%) cases. In 12 cases VLE alone targeted the biopsies, yielding 6 cases (50%) of non-dysplastic BE, 3 cases (25%) indefinite for dysplasia and 3 cases (25%) with definite dysplasia or neoplasia. In 2 of 3 procedures where VLE-only targeted and random biopsies were obtained, targeted samples yielded a higher grade of pathology than random results. No cases had more severe pathology on random biopsies. In 3 procedures where other advanced imaging techniques were employed, along with VLE, no BE or dysplasia was seen on targeted biopsies. Conclusions: VLE aided in targeting biopsies in almost a third of cases where patients were undergoing same-session ablative treatment for BE. Of those cases where VLE alone was used for targeting, histology identified BE in 50% of cases and findings for low-grade dysplasia or more severe in 25% of cases. Random and nonVLE targeted biopsies did not find any disease missed by VLE-targeted biopsies. These findings suggest that, in order to minimize time, expense and tissue trauma from biopsies when also performing BE field ablation, it appears unnecessary to sample areas not found to be suspicious on VLE. Use of VLE during endoscopy may therefore decrease the need for and risk of extensive tissue sampling when ablation is planned. Patient Demographics
(A) Photograph of the tethered micro-motor capsule endomicroscopy device, adjacent to a penny for size comparison. (B) Schematic of the capsule including the micro-motor and micro-optics.
(C) One esophageal circumferential image obtained by the new device from a human subject in vivo.
S-39
AGA Abstracts
AGA Abstracts
156
Figure 2: Graphic representation of overall cognitive learning curves among trainees achieving and not achieving competence by using cumulative sum analysis - crossing the lower limit threshold indicates performance within the acceptable rate. Primary analysis: success defined as score of 1 or 2 (no assistance/minimal verbal cues), Acceptable failure rate -p0=0.1 and unacceptable failure rate - p1=0.3
AGA Abstracts
on gene expression in intestinal tissues of infant mice in vivo. E. faecalis strains colonizing MAT infant mice are not vancomycin resistant, thus allowing characterization of nonpathogenic E. faecalis in vivo. Methods: We used the Mouse Innate and Adaptive Immune Responses RT2 Profiler PCR Array (Qiagen) to assess the expression of 84 genes involved in the host response to bacterial infection and sepsis. To distinguish tissue- and age-specific effects, distal ileum and proximal colon total tissue RNA from MAT infant littermates at day of life (DOL) 15 and 21 was compared gene expression relative to age matched controls (CTRL). To assess cell type specific gene and protein expression of candidate genes we further isolated intestinal epithelial cells and intestinal and splenic lymphocytes . Results: In MAT infant mice at DOL 15, 60 genes were modulated in the colon and 30 were modulated in the ileum, whereas at DOL 21, 59 genes were modulated in the colon versus 64 genes in the ileum. Based on the array data we found that IL10, TLR3, TLR9, and Traf6 were significantly regulated in MAT infant mouse intestines. We confirmed differential expression of TLRs, cytokines and signaling molecule genes in MAT and CTRL intestines by qPCR from 3 separate litters of mice. IL10, IL1a, TGFbeta, and TLR3 were down-regulated in colonic tissue from MAT infants, while Tyk2 and TLR2 were up-regulated. Conclusions: Infant mice born of mothers treated with antibiotics during the perinatal period can acquire an intestinal microbiota dominated by non-pathogenic bacteria. We show that an E. faecalis dominated microbiota in infant mice modulates TLR and cytokine expression. This result is similar to those in which the effect of E. faecalis strains from newborn infants cultured in vitro with human intestinal cell lines was examined. Our mouse model provides the opportunity for targeted experiments that further characterize how the limited diversity intestinal microbiota of infancy regulates immune homeostasis during this period.
160 Alterations in the Gut Microbiome and Correlation With Bloodstream Infection in Infants With Necrotizing Enterocolitis-Induced Short Bowel Syndrome Conrad R. Cole, Ethan A. Mezoff, Charles E. Robertson, Daniel N. Frank, Thomas Ziegler Introduction: Necrotizing enterocolitis is a major predisposing factor for surgical short bowel syndrome (SBS) in infants. Central line-associated blood stream infection (CLA-BSI) is a significant contributor to the morbidity and mortality of these children. Gut permeability defects leading to translocation of gut-associated bacteria may contribute to CLA-BSI. The aim of this study was to gain insight into the microbial diversity of stool of these infants in relationship to CLA-BSI. Methods: Ten infants with SBS caused by necrotizing enterocolitis who were dependent on parenteral nutrition for > 6 weeks were evaluated, along with 9 age-matched control subjects without intestinal failure or SBS. Stool was collected at baseline and if the child was hospitalized for CLA-BSI. Gut microbiome profiling of collected stool was accomplished by pan-bacterial 16S rRNA gene PCR and pyrosequencing. Results: An average of 53.9 genus-level bacterial groups were identified per subject. Klebsiella spp, were the most abundant microorganisms (mean = 17% of sequences) in SBS infants, but were notably less abundant in control specimens (mean = 2% of sequences; p = 0.053). Other genera, such as Enterococcus and Lactobacillus also were enriched in the stool of infants with SBS relative to controls, but the results were not statistically significant. Klebsiella pneumonia (35%) was the most common bacteria cultured in CLA-BSI. Conclusion: These results suggest the potential utility for use of culture-independent analysis to characterize microbial diversity in SBS. The preliminary data support the hypothesis that alterations in the gut luminal microbiome in pediatric SBS patients, versus non-SBS controls, is a predisposing factor in SBS-associated CLA-BSI.
158 Novel Insight Into Salmonella Typhi Pathogenesis From Ex Vivo Human Tissue Models Kourtney Nickerson, Stefania Senger, Marcelo B. Sztein, Alessio Fasano, Maria R. Fiorentino Introduction. Salmonella enterica serovar Typhi is the causative agent of Typhoid fever, from which an estimated 22 million cases occur annually resulting in 200,000 deaths. In some areas of the globe, the incidence of Typhoid fever is as high as 500 cases out of every 100,000 children. At present, little is understood about host response to Typhi infection. As such, no long-term preventive vaccine therapy is available. Current therapeutics include antibiotic treatment, however antibiotic resistant serovars are increasing worldwide. Furthermore, chronic Typhi colonization of the gall bladder is sufficient to cause gall bladder cancer therefore demonstrating significant health risks upon both long and short-term infection. To design alternative therapeutic strategies, there is immediate need to understand Typhi infection and pathogenesis. Materials and Methods. Terminal ileum biopsies were collected from donors for direct infection or generation of organoids. Whole biopsy infections were conducted using micro-snapwell mounting of tissue followed by addition of Salmonella enterica serovar Typhi 2a to the apical surface. Organoids were differentiated into epithelial compartment monolayers followed by infection with Typhi. Prior to infection, Typhi was grown in Luria Burtoni broth under static or shaking conditions. Upon infection, changes in trans-epithelial electrical resistance (TEER), cytokine release, gene expression, and cellular localization were assessed. Results and Conclusions. Terminal ileum derived organoids give rise to a diversity of epithelial cells, including goblet, paneth and M cells, which are grown as a monolayer in vitro. Use of the epithelial monolayer (EM) and whole biopsy (WB) model identified specific contributions of the epithelium in response to Typhi infection as assessed by RNA-sequencing, qPCR, and cytokine secretion. Consideration for bacterial inoculum preparation revealed that Typhi grown under static conditions express the Vi antigen, as well as, SPI-1 and SPI-2 effector proteins. Therefore, static or shaking inoculums were applied to both models. Differences in cellular association of bacteria were assessed using IF and TEM. To identify how the gut mucosa responds to infection, apical and basolateral culture supernatants were collected for ELISA analysis. ELISA analysis demonstrated differences in IL-8, IL-1b and IL-12p70 cytokine production dependent on infection model and secretion location. Interestingly, infected monolayers showed the highest levels of basolateral cytokine release. Finally, infection decreased TEER under both inoculum conditions in the EM model, but only the static inoculum decreased TEER in the WB model. All together, our data characterizes key aspects of terminal ileum response to Typhi infection addressing a critical gap in our current understanding of Typhoid fever pathogenesis.
161 Impact of Early Life Intervention With Bifidobacteria on the Structure and Function of Infant Fecal Microbiota Monika Bazanella, Tanja Maier, Thomas Clavel, Ilias Lagkouvardos, Lars Bode, Philippe Schmitt-Kopplin, Dirk Haller Background: Development of the intestinal microbiota in infants is a dynamic process and probiotic intervention was shown to be effective in preventing newborn pathologies like necrotizing enterocolitis and infantile colic. However, the impact of early-life bacterial intervention on the natural assembly of fecal communities and metabolic functions is unknown. We designed a randomized, double-blinded, placebo controlled intervention trial with healthy neonates receiving infant formula supplemented with or without a mixture of four different bifidobacteria during the first year of life. Methods: 106 infants were randomized to receive bacteria-supplemented (Bifidobacterium longum, B. infantis, B. breve, B. bifidum ) infant formula (n=48) or non-supplemented formula (n=49) after weaning. 9 infants were exclusively breast-fed. Fecal samples were collected monthly over a period of one year. High-throughput 16S rRNA amplicon sequencing by Illumina, high resolution metabolite analysis (UPLC-MS), and human milk oligosaccharide (HMO) analysis were applied. Results: Formula- and breast-fed infants harbored distinct bacterial communities, even after 1 year of age, and breast feeding was characterized by significantly reduced diversity. Metabolite profiling identified distinct clusters according to the feeding regiment and age of the infants. Supplementation of infant formula with bifidobacteria did not affect community structures or metabolite profiles in infants exclusively fed with formula (N=22). However in combination with breast feeding, bifidobacteria supplementation modulated the microbiota up to 7 months of age. All feeding groups were dominated by bifidobacteria, representing 69% (breastfed), 35% (bacteria-supplemented formula), and 32% (control formula) of total sequences. Bacteroidaceae, with Bacteroides fragilis as prevalent member, showed increased relative abundances during the first weeks of colonization in the control formula group. A core microbiota including molecular species classified as two Bifidobacterium spp., Escherichia-Shigella sp., Streptococcus sp. and Enterococcus sp., was detected in all infants at least once in the first year of life. Caesarean section was associated with increased relative abundance of Firmicutes and decrease in Actinobacteria during the first three months. Breast milk HMO analysis pointed towards a link between maternal secretor status (presence or absence of fucosyltransferase 2) and infantile microbiota composition. Conclusion: Infant formula significantly influenced assembly and metabolite profiles of the early life microbiota, particularly associated
159 Modulation of TLR and Cytokine Gene Expression in Commensal Enterococcus Faecalis Dominated Infant Mouse Intestine Tessa M. Tekieli, Esi Lamouse-Smith Background: Enterococcus faecalis is normal member of the intestinal microbiota and a founding member of commensal bacteria colonizing the intestine of humans and mice at birth. While well known as a human pathogen its role and function as a commensal symbiont in the intestine of infants is less well characterized. In our mouse model of maternal antibiotic treatment (MAT), infant MAT mice maintain an intestinal flora almost entirely dominated by E. faecalis. These mice gain weight and grow appropriately, but are unable to control a systemic vaccinia virus infection. We hypothesized that the E. faecalis dominated microbiota modulates the expression of genes that can regulate mucosal and systemic immunity. Our aim in this study was to characterize the selective impact of commensal E. faecalis colonization
AGA Abstracts
S-40
155 Background: Endoscopic ultrasound (EUS) allows visualization of celiac axis lymph nodes (CLNs) and celiac ganglia (CG); however, the EUS accuracy in distinguishing between these anatomical structures is unknown. This distinction is important given the impact on clinical tumor staging and the targeting of celiac ganglia for blockade, neurolysis or other ablative therapies. Aim: To determine the EUS diagnostic accuracy in distinguishing CLNs from CG. Methods: Patients who underwent EUS FNA of either suspected CLNs or CG (10/200410/2015) were identified from a prospectively maintained EUS database. Results: 453 patients [58% male; age 65 years (15-93)] underwent EUS FNA of either suspected CLNs or CG. Most (376, 83%) procedures were performed to evaluate a broad spectrum of malignancies, most commonly pancreatic cancer (33%), esophageal cancer (21%), and cholangiocarcinoma (17%). EUS FNA was performed of: 1.) a presumed CLN [219 (48%) patients; median 4 passes, range 1-15)], 2.) a presumed CG [157 (35%) patients; 4 passes (1-11)], 3.) both CLN and CG [(34 (8%) patients; 3 passes (1-6) for LN and 3 passes (18) for CG)] and 4.) an indeterminate CLN or CG [43 (9%) patients; 3 passes (1-5)]. In total, there were 487 structures sampled whereby the endosonographer perceived that they represented CLNs (n=253), CG (n=191) or were indeterminate (n=43). The cytopathology report provided a definitive gold standard for 332 (73.3%) sites indicating either CG (based on the presence of neural structures without lymphocytes) or CLN (lymphocytes without neural glial cells). The cytopathology report for the remaining 155 patients simply assessed for the presence or absence of malignancy without mention of the presence of neuronal glial cells or lymphocytes. Based on those patients with a cytopathology gold standard, the accuracy of EUS, when the endosonographer was confident that they biopsied a CLN or CG, was 97% (165/171) or 91% (125/138), respectively. When including 43 patients in whom the site was indeterminate by the endosonographer, the diagnostic accuracy was reduced to 89% (165/186) and 86% (125/146) for CLNs and CG, respectively. Post procedure clinical follow-up within 4-7 days was available in 83% (378/453). The adverse event rate was 3.97% (15 of 378) and was comprised of abdominal pain (n=7), mild pancreatitis (n= 2), bleeding (n=2; hemobilia and post EMR bleed), and 1 patient each with duodenal perforation, fever, excessive moderate sedation and non-cardiac chest pain; most of which appear to relate to other procedures and interventions performed. Conclusions: EUS accurately distinguishes CLNs from CG. This information can be valuable in overall patient care to help enhance clinical tumor staging and for selectively targeting celiac ganglia pain management.
Tethered Capsule OCT Endomicroscopy With Micro-Motor Scanning for Esophageal Imaging Jing Dong, Michalina J. Gora, Rohith Reddy, Wolfgang K. Trasischker, Timothy N. Ford, Weina Lu, Catriona N. Grant, Amna R. Soomro, Mireille Rosenberg, Norman S. Nishioka, Guillermo J. Tearney Introduction: Our lab has previously developed and reported a new technology termed tethered capsule endomicroscopy (TCE) that utilizes optical coherence tomography (OCT) for microscopic (10µm) imaging of internal structures of the human esophagus without the need of sedation [1]. To date, we have successfully performed ~100 procedures with the TCE device. To justify its use for population based screening for diseases in upper gastrointestinal tract, we have developed a second generation device that is cost-effective and easy to manufacture, and designed to provide a comfortable experience for the patient. Materials and Methods: The new generation TCE device incorporates a micro-motor [2] at the distal end of a 11mm x 24.4mm swallowable capsule. The capsule is connected to a 1mm diameter, 1.6m long flexible tether that houses an optical fiber and two electrical wires which power up the micro-motor. The near-infrared light from the fiber is focused by the micro-optics comprising a monolithic ball lens and deflected to the side of the capsule by the angle polished shaft of the micro-motor. Rotation of the shaft effectuates circumferential scanning of the esophageal wall. After unsedated subjects swallowed the capsule, multiple crosssectional microscopic OCT images of the esophagus were obtained and visualized in real time. The capsule descended to the stomach via peristalsis and then was gradually pulled back up through the esophagus using the tether. After imaging the esophagus twice, the capsule was removed and sterilized for reuse. After the procedure, subjects were asked if they would recommend the procedure to other patients and if any capsule vibrations were felt. Results: To date, 4 healthy subjects have been enrolled in the tethered micro-motor capsule endomicroscopy study. A total of 3/4 (75%) subjects successfully swallowed the capsule. For those who swallowed the capsule, high quality microscopic images of the entire esophagus were obtained. The entire procedure lasted an average of 3±1 minutes from capsule insertion to removal. There were no complications related to the procedure. All subjects who successfully swallowed the capsule would recommend the procedure; none of the subjects felt vibration of the capsule, 1 subject reported that the new generation of TCE was more comfortable than previous TCE device. Conclusion: Our initial experience with the tethered micro-motor capsule endomicroscopy has shown that the new generation device is able to image the esophageal wall in a rapid and safe manner while rendering the same high quality, 3-dimensional microstructural images as the previous version. Additionally, because this device is simpler to manufacture and less costly, it may be desirable for the diagnosis of esophageal diseases in screening settings. [1] Gora MJ et al. Nat Med. 2013, 19(2):238-40 [2] Liang K et al. BOE. 2015, 6(4):1146-63
157 Volumetric Laser Endomicroscopy Targeted Tissue Sampling Eliminates Unnecessary Biopsies During Planned Ablation of Barrett's Esophagus Udayakumar Navaneethan, Brooks D. Cash, Matthew McKinley, Virendra Joshi, Paul R. Tarnasky, Kenneth J. Chang, Satish K. Singh, Michael B. Wallace, Michael S. Smith Background: Endoscopic ablation of Barrett's esophagus (BE) is accomplished with a variety of modalities. Tissue sampling performed during the same endoscopy can affect visualization and potentially procedure outcome. Volumetric laser endomicroscopy (VLE) uses nextgeneration optical coherence tomography to rapidly scan a large area of the esophagus to a depth of 3 mm and a resolution of 7 microns. Our aim was to assess whether VLE-targeted tissue sampling maintains adequate sampling while decreasing unnecessary biopsies at the time of BE ablation. Methods: Interim analysis was performed for a US-based multicenter registry that prospectively collected demographic, endoscopic and histologic data when VLE was used during the same endoscopy as BE field ablation. All cases were performed using the NvisionVLE™ system (NinePoint Medical, Bedford, MA). Patients could be treatment naïve or incompletely treated. Treatments permitted for this study included radiofrequency ablation (RFA), cryoablation (Cryo) or argon plasma coagulation (APC). Patients who had VLE in the same session as endoscopic mucosal resection (EMR) were excluded, as these cases underwent separate analysis. Results: A total of 182 procedures from 140 patients met inclusion criteria. The mean age was 66.5 years (range 33-88) and 81% were male. Among these patients, 37 (26%) were BE treatment naïve, while the remaining 103 previously were treated at least once. BE was seen on white light or narrow band imaging in 145/182 (80%) procedures. VLE and endoscopic visualization identified abnormalities that led to targeted biopsies in 57/182 (31%) cases. In 12 cases VLE alone targeted the biopsies, yielding 6 cases (50%) of non-dysplastic BE, 3 cases (25%) indefinite for dysplasia and 3 cases (25%) with definite dysplasia or neoplasia. In 2 of 3 procedures where VLE-only targeted and random biopsies were obtained, targeted samples yielded a higher grade of pathology than random results. No cases had more severe pathology on random biopsies. In 3 procedures where other advanced imaging techniques were employed, along with VLE, no BE or dysplasia was seen on targeted biopsies. Conclusions: VLE aided in targeting biopsies in almost a third of cases where patients were undergoing same-session ablative treatment for BE. Of those cases where VLE alone was used for targeting, histology identified BE in 50% of cases and findings for low-grade dysplasia or more severe in 25% of cases. Random and nonVLE targeted biopsies did not find any disease missed by VLE-targeted biopsies. These findings suggest that, in order to minimize time, expense and tissue trauma from biopsies when also performing BE field ablation, it appears unnecessary to sample areas not found to be suspicious on VLE. Use of VLE during endoscopy may therefore decrease the need for and risk of extensive tissue sampling when ablation is planned. Patient Demographics
(A) Photograph of the tethered micro-motor capsule endomicroscopy device, adjacent to a penny for size comparison. (B) Schematic of the capsule including the micro-motor and micro-optics.
(C) One esophageal circumferential image obtained by the new device from a human subject in vivo.
S-39
AGA Abstracts
AGA Abstracts
156
Figure 2: Graphic representation of overall cognitive learning curves among trainees achieving and not achieving competence by using cumulative sum analysis - crossing the lower limit threshold indicates performance within the acceptable rate. Primary analysis: success defined as score of 1 or 2 (no assistance/minimal verbal cues), Acceptable failure rate -p0=0.1 and unacceptable failure rate - p1=0.3
AGA Abstracts
on gene expression in intestinal tissues of infant mice in vivo. E. faecalis strains colonizing MAT infant mice are not vancomycin resistant, thus allowing characterization of nonpathogenic E. faecalis in vivo. Methods: We used the Mouse Innate and Adaptive Immune Responses RT2 Profiler PCR Array (Qiagen) to assess the expression of 84 genes involved in the host response to bacterial infection and sepsis. To distinguish tissue- and age-specific effects, distal ileum and proximal colon total tissue RNA from MAT infant littermates at day of life (DOL) 15 and 21 was compared gene expression relative to age matched controls (CTRL). To assess cell type specific gene and protein expression of candidate genes we further isolated intestinal epithelial cells and intestinal and splenic lymphocytes . Results: In MAT infant mice at DOL 15, 60 genes were modulated in the colon and 30 were modulated in the ileum, whereas at DOL 21, 59 genes were modulated in the colon versus 64 genes in the ileum. Based on the array data we found that IL10, TLR3, TLR9, and Traf6 were significantly regulated in MAT infant mouse intestines. We confirmed differential expression of TLRs, cytokines and signaling molecule genes in MAT and CTRL intestines by qPCR from 3 separate litters of mice. IL10, IL1a, TGFbeta, and TLR3 were down-regulated in colonic tissue from MAT infants, while Tyk2 and TLR2 were up-regulated. Conclusions: Infant mice born of mothers treated with antibiotics during the perinatal period can acquire an intestinal microbiota dominated by non-pathogenic bacteria. We show that an E. faecalis dominated microbiota in infant mice modulates TLR and cytokine expression. This result is similar to those in which the effect of E. faecalis strains from newborn infants cultured in vitro with human intestinal cell lines was examined. Our mouse model provides the opportunity for targeted experiments that further characterize how the limited diversity intestinal microbiota of infancy regulates immune homeostasis during this period.
160 Alterations in the Gut Microbiome and Correlation With Bloodstream Infection in Infants With Necrotizing Enterocolitis-Induced Short Bowel Syndrome Conrad R. Cole, Ethan A. Mezoff, Charles E. Robertson, Daniel N. Frank, Thomas Ziegler Introduction: Necrotizing enterocolitis is a major predisposing factor for surgical short bowel syndrome (SBS) in infants. Central line-associated blood stream infection (CLA-BSI) is a significant contributor to the morbidity and mortality of these children. Gut permeability defects leading to translocation of gut-associated bacteria may contribute to CLA-BSI. The aim of this study was to gain insight into the microbial diversity of stool of these infants in relationship to CLA-BSI. Methods: Ten infants with SBS caused by necrotizing enterocolitis who were dependent on parenteral nutrition for > 6 weeks were evaluated, along with 9 age-matched control subjects without intestinal failure or SBS. Stool was collected at baseline and if the child was hospitalized for CLA-BSI. Gut microbiome profiling of collected stool was accomplished by pan-bacterial 16S rRNA gene PCR and pyrosequencing. Results: An average of 53.9 genus-level bacterial groups were identified per subject. Klebsiella spp, were the most abundant microorganisms (mean = 17% of sequences) in SBS infants, but were notably less abundant in control specimens (mean = 2% of sequences; p = 0.053). Other genera, such as Enterococcus and Lactobacillus also were enriched in the stool of infants with SBS relative to controls, but the results were not statistically significant. Klebsiella pneumonia (35%) was the most common bacteria cultured in CLA-BSI. Conclusion: These results suggest the potential utility for use of culture-independent analysis to characterize microbial diversity in SBS. The preliminary data support the hypothesis that alterations in the gut luminal microbiome in pediatric SBS patients, versus non-SBS controls, is a predisposing factor in SBS-associated CLA-BSI.
158 Novel Insight Into Salmonella Typhi Pathogenesis From Ex Vivo Human Tissue Models Kourtney Nickerson, Stefania Senger, Marcelo B. Sztein, Alessio Fasano, Maria R. Fiorentino Introduction. Salmonella enterica serovar Typhi is the causative agent of Typhoid fever, from which an estimated 22 million cases occur annually resulting in 200,000 deaths. In some areas of the globe, the incidence of Typhoid fever is as high as 500 cases out of every 100,000 children. At present, little is understood about host response to Typhi infection. As such, no long-term preventive vaccine therapy is available. Current therapeutics include antibiotic treatment, however antibiotic resistant serovars are increasing worldwide. Furthermore, chronic Typhi colonization of the gall bladder is sufficient to cause gall bladder cancer therefore demonstrating significant health risks upon both long and short-term infection. To design alternative therapeutic strategies, there is immediate need to understand Typhi infection and pathogenesis. Materials and Methods. Terminal ileum biopsies were collected from donors for direct infection or generation of organoids. Whole biopsy infections were conducted using micro-snapwell mounting of tissue followed by addition of Salmonella enterica serovar Typhi 2a to the apical surface. Organoids were differentiated into epithelial compartment monolayers followed by infection with Typhi. Prior to infection, Typhi was grown in Luria Burtoni broth under static or shaking conditions. Upon infection, changes in trans-epithelial electrical resistance (TEER), cytokine release, gene expression, and cellular localization were assessed. Results and Conclusions. Terminal ileum derived organoids give rise to a diversity of epithelial cells, including goblet, paneth and M cells, which are grown as a monolayer in vitro. Use of the epithelial monolayer (EM) and whole biopsy (WB) model identified specific contributions of the epithelium in response to Typhi infection as assessed by RNA-sequencing, qPCR, and cytokine secretion. Consideration for bacterial inoculum preparation revealed that Typhi grown under static conditions express the Vi antigen, as well as, SPI-1 and SPI-2 effector proteins. Therefore, static or shaking inoculums were applied to both models. Differences in cellular association of bacteria were assessed using IF and TEM. To identify how the gut mucosa responds to infection, apical and basolateral culture supernatants were collected for ELISA analysis. ELISA analysis demonstrated differences in IL-8, IL-1b and IL-12p70 cytokine production dependent on infection model and secretion location. Interestingly, infected monolayers showed the highest levels of basolateral cytokine release. Finally, infection decreased TEER under both inoculum conditions in the EM model, but only the static inoculum decreased TEER in the WB model. All together, our data characterizes key aspects of terminal ileum response to Typhi infection addressing a critical gap in our current understanding of Typhoid fever pathogenesis.
161 Impact of Early Life Intervention With Bifidobacteria on the Structure and Function of Infant Fecal Microbiota Monika Bazanella, Tanja Maier, Thomas Clavel, Ilias Lagkouvardos, Lars Bode, Philippe Schmitt-Kopplin, Dirk Haller Background: Development of the intestinal microbiota in infants is a dynamic process and probiotic intervention was shown to be effective in preventing newborn pathologies like necrotizing enterocolitis and infantile colic. However, the impact of early-life bacterial intervention on the natural assembly of fecal communities and metabolic functions is unknown. We designed a randomized, double-blinded, placebo controlled intervention trial with healthy neonates receiving infant formula supplemented with or without a mixture of four different bifidobacteria during the first year of life. Methods: 106 infants were randomized to receive bacteria-supplemented (Bifidobacterium longum, B. infantis, B. breve, B. bifidum ) infant formula (n=48) or non-supplemented formula (n=49) after weaning. 9 infants were exclusively breast-fed. Fecal samples were collected monthly over a period of one year. High-throughput 16S rRNA amplicon sequencing by Illumina, high resolution metabolite analysis (UPLC-MS), and human milk oligosaccharide (HMO) analysis were applied. Results: Formula- and breast-fed infants harbored distinct bacterial communities, even after 1 year of age, and breast feeding was characterized by significantly reduced diversity. Metabolite profiling identified distinct clusters according to the feeding regiment and age of the infants. Supplementation of infant formula with bifidobacteria did not affect community structures or metabolite profiles in infants exclusively fed with formula (N=22). However in combination with breast feeding, bifidobacteria supplementation modulated the microbiota up to 7 months of age. All feeding groups were dominated by bifidobacteria, representing 69% (breastfed), 35% (bacteria-supplemented formula), and 32% (control formula) of total sequences. Bacteroidaceae, with Bacteroides fragilis as prevalent member, showed increased relative abundances during the first weeks of colonization in the control formula group. A core microbiota including molecular species classified as two Bifidobacterium spp., Escherichia-Shigella sp., Streptococcus sp. and Enterococcus sp., was detected in all infants at least once in the first year of life. Caesarean section was associated with increased relative abundance of Firmicutes and decrease in Actinobacteria during the first three months. Breast milk HMO analysis pointed towards a link between maternal secretor status (presence or absence of fucosyltransferase 2) and infantile microbiota composition. Conclusion: Infant formula significantly influenced assembly and metabolite profiles of the early life microbiota, particularly associated
159 Modulation of TLR and Cytokine Gene Expression in Commensal Enterococcus Faecalis Dominated Infant Mouse Intestine Tessa M. Tekieli, Esi Lamouse-Smith Background: Enterococcus faecalis is normal member of the intestinal microbiota and a founding member of commensal bacteria colonizing the intestine of humans and mice at birth. While well known as a human pathogen its role and function as a commensal symbiont in the intestine of infants is less well characterized. In our mouse model of maternal antibiotic treatment (MAT), infant MAT mice maintain an intestinal flora almost entirely dominated by E. faecalis. These mice gain weight and grow appropriately, but are unable to control a systemic vaccinia virus infection. We hypothesized that the E. faecalis dominated microbiota modulates the expression of genes that can regulate mucosal and systemic immunity. Our aim in this study was to characterize the selective impact of commensal E. faecalis colonization
AGA Abstracts
S-40