Name: fecal microbiota transplantation via colonoscopy for recurrent c. difficile infection
Uploaded: 2014-12-08T15:00:00.000+00:00
Duration: 7 min 6 s
Description: Watch this Scientific Journal Video about Fecal Microbiota Transplantation via Colonoscopy for Recurrent C. difficile Infection at JoVE.com

The authors would like to acknowledge the Infectious Disease Department at Brigham and Women&#8217;s Hospital especially Michael S. Calderwood, MD, MPH for helping us construct this protocol. We would also like to thank the Institution&#8217;s endoscopy center and it&#8217;s leadership team for their support.

<ol>
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R., de Leon, L., Jasutkar, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fecal+microbiota+transplantation+for+relapsing+Clostridium+difficile+infection+in+26+patients:+methodology+and+results.">Fecal microbiota transplantation for relapsing Clostridium difficile infection in 26 patients: methodology and results.</a> J Clin Gastroenterol. 46 (2), 145-149 (2012).</li><li>Lund-Tonnesen, S., Berstad, A., Schreiner, A., Midtvedt, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clostridium+difficile-associated+diarrhea+treated+with+homologous+feces.">Clostridium difficile-associated diarrhea treated with homologous feces.</a> Tidsskr Nor Laegeforen. 118 (7), 1027-1030 (1998).</li><li>MacConnachie, A. A., Fox, R., Kennedy, D. R., Seaton, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Faecal+transplant+for+recurrent+Clostridium+difficile-associated+diarrhoea:+a+UK+case+series.">Faecal transplant for recurrent Clostridium difficile-associated diarrhoea: a UK case series.</a> QJM. 102 (11), 781-784 (2009).</li><li>Mellow, M. H., Kanatzar, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colonoscopic+fecal+bacteriotherapy+in+the+treatment+of+recurrent+Clostridium+difficile+infection--results+and+follow-up.">Colonoscopic fecal bacteriotherapy in the treatment of recurrent Clostridium difficile infection--results and follow-up.</a> J Okla State Med Assoc. 104 (3), 89-91 (2011).</li><li>Polak, P., Freibergerova, M., Jurankova, J., Kocourkova, H., Mikesova, L., Svacina, R., Husa, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=First+experiences+with+faecal+bacteriotherapy+in+the+treatment+of+relapsing+pseudomembranous+colitis+due+to+Clostridium+difficile.">First experiences with faecal bacteriotherapy in the treatment of relapsing pseudomembranous colitis due to Clostridium difficile.</a> Klin Mikrobiol Infekc Lek. 17 (6), 214-217 (2011).</li><li>Rohlke, F., Surawicz, C. M., Stollman, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fecal+flora+reconstitution+for+recurrent+Clostridium+difficile+infection:+results+and+methodology.">Fecal flora reconstitution for recurrent Clostridium difficile infection: results and methodology.</a> J Clin Gastroenterol. 44 (8), 567-570 (2010).</li><li>Sekirov, I., Russell, S. L., Antunes, L. C., Finlay, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gut+microbiota+in+health+and+disease.">Gut microbiota in health and disease.</a> Physiol Rev. 90 (3), 859-904 (2010).</li><li>Yoon, S. S., Brandt, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment+of+refractory/recurrent+C.+difficile-associated+disease+by+donated+stool+transplanted+via+colonoscopy:+a+case+series+of+12+patients.">Treatment of refractory/recurrent C. difficile-associated disease by donated stool transplanted via colonoscopy: a case series of 12 patients.</a> J Clin Gastroenterol. 44 (8), 562-566 (2010).</li><li>Brandt, L. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+follow-up+of+colonoscopic+fecal+microbiota+transplant+for+recurrent+Clostridium+difficile+infection.">Long-term follow-up of colonoscopic fecal microbiota transplant for recurrent Clostridium difficile infection.</a> Am J Gastroenterol. 107 (7), 1079-1087 (2012).</li><li>Kelly, C. R., Kunde, S. S., Khoruts, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Guidance+on+preparing+an+investigational+new+drug+application+for+fecal+microbiota+transplantation+studies.">Guidance on preparing an investigational new drug application for fecal microbiota transplantation studies.</a> Clin Gastroenterol Hepatol. 12 (2), 283-288 (2014).</li></ol>

The authors have nothing to disclose relevant to the material presented in this manuscript.

وقد ثبت زرع البراز ليكون علاجا فعالا وآمنا للCDI المتكررة. وقد أوجزنا بروتوكول قياسي لFMT عن طريق تنظير القولون وشملت نتائجنا لمدة 22 المرضى الذين خضعوا لإجراءات CDI المتكررة. نتائجنا قابلة للمقارنة مع البيانات المنشورة لمرض المتكرر حتى الآن مع مريض واحد فقط تتكرر ش?...

وقد وصفت زرع البراز لعلاج التهاب القولون pseumembranous للمرة الأولى في عام 1958 1. ومع ذلك، تقارير عن تناول البراز لعلاج التسمم الغذائي وتاريخ الإسهال الشديد إلى الوراء في وقت مبكر من القرن الرابع الصين 2. ويطلق البراز الجراثيم زرع (FMT) أيضا في الأدب &quot;المعالجة بالجراثيم البراز&quot;، &quot;ضخ بروبيوتيك البشري&quot;، &quot;زرع البراز&quot;، &quot;المعوية استعادة microbiome&quot;، و &quot;نقل البراز&quot;. أنها تنطوي على جمع البراز من الأصحاء، فحص ما قبل الجهات المانحة وتقديم الطين على استعداد في القناة الهضمية للمتلقي عن طريق أنبوب أنفي معدي، esophagogastricduodenoscopy، تنظير القولون، أو حقنة شرجية 3. وأفادت العديد من المسلسلات حالة فعالية الإدارة عبر NGT والحقنة الشرجية، وقد أظهرت لكن أكثر حداثة الدراسات أن إدارة البراز المانحة عن طريق فحص القولون بالمنظار آمنة وفعالة للغاية 4.في حين أساليب أخرى مثل الإدارة أنبوب أنفي معدي تم الإبلاغ، والتسليم من البراز المانحة عن طريق تنظير القولون له آثار جانبية أقل الفورية (مثل الانتفاخ في البطن والغثيان) (4) ولقد وجدنا أن المرضى هم أكثر تقبلا لمفهوم FMT عندما يتم تنفيذ ذلك من قبل تنظير القولون. واحدة من المزايا العلاجية لFMT عن طريق تنظير القولون هو انخفاض في الكتلة الحيوية القولون نتيجة لغسيل الأمعاء التي يتم تنفيذها قبل إجراء 4. ودراسات متعددة دور FMT لعلاج التهاب القولون والإسهال الناجمة عن العوامل المسببة للأمراض الانتهازية C. صعب (CDI). استمر حدوث CDI في الارتفاع وسبب رئيسي للمراضة والوفيات مع عبء اقتصادي كبير في جميع أنحاء العالم. علاج الخط الأول لCDI تتكون من العلاج بالمضادات الحيوية، ولكن تم الإبلاغ عن معدلات تكرار بين 15-35٪ 5. عدة سلسلة من الحالات والتقارير ديك القيام بهcumented سلامة وفعالية FMT لCDI صهر للعلاج الطبي القياسية مع المضادات الحيوية 4،6-16. أفادت دراسة تبحث في المدى الطويل متابعة المرضى بعد FMT عن طريق تنظير القولون لCDI نسبة شفاء الأساسي 91٪ في 77 مريضا 17. في مركزنا (بريجهام ومستشفى النساء وقسم أمراض الجهاز الهضمي والكبد والمناظير)، ونحن برنامجا زرع البراز لمرضى المتكررة أو CDI الحرارية. يتم تعريف المرشحين المناسبين من المرضى الذين لديهم CDI المتكررة (تاريخ من 3 أو أكثر من حلقة، أو 2 حلقة التي تتطلب دخول المستشفى)، أو المرضى الذين يعانون من مرض الحرارية التي لا تستجيب للمضادات الحيوية التقليدية. ونحن نعتقد نهج منتظم لجميع مراحل هذا الإجراء يزيد فعالية. في هذه المخطوطة والفيديو المصاحب، ونحن بالتفصيل بروتوكول لقد استخدمنا في مركز لدينا، والتي تشمل المريض والجهات المانحة الفرز، وإعداد البراز، وإيصال البراز لر وقت تنظير القولون. هذه الطريقة قد أسفرت عن نتائج إيجابية مماثلة لالكتابات المنشورة. حالة عرض تقديمي: ويمثل هذا المريض المريض نموذجي المشار لزرع البراز. كان المريض امرأة تبلغ من العمر 69 مع تاريخ من سرطان الدم الليمفاوي المزمن الذي كان انتكاسة للمرض تحتاج إلى مزيد من العلاج مع العلاج الكيميائي. عانت من ثلاث حلقات من CDI في العام الماضي، وبالتالي أحيل إلى العيادة للنظر في FMT مسبق العلاج الكيميائي لإعادة بدء. الحلقة الأولى لها من CDI وقعت في أكتوبر 2012. وقالت إنها لم يكن لديه أي المضادات الحيوية السابقة. وكانت مريضة جدا في وحدة العناية المركزة بعد تقديم مع الصدمة الإنتانية وخضع لتحويل حلقة فغر اللفائفي مع تقدمي فانكومايسين الحقن الشرجية لدورة 6 أسابيع في تركيبة مع فانكومايسين عن طريق الفم نظرا لشدة مرضها. فعلت بشكل جيد جدا مع قرار من الإسهال لها وانها استطاعت أن تأتيالمضادات الحيوية خارج. وقد ظهرت عليها فتق حول ستوما لها لذلك كان عكس ذلك. بعد فترة وجيزة من انعكاس الستوما أنها وضعت مرة أخرى الإسهال وجدت مرة أخرى ليكون C. السم صعب إيجابية. أكملت دورة أخرى 6 أسابيع من فانكومايسين عن طريق الفم وتمكنت من الاضمحلال منه بنجاح. في وقت لاحق ولكن عدة أشهر، وقالت انها وضعت مرة أخرى الإسهال الذي كان C. السم صعب إيجابية وبدأت في الدورة الثالثة لها من فانكومايسين عن طريق الفم. كانت تفعله جيدا على المضادات الحيوية عندما لوحظ أن لها العد الأبيض ارتفع إلى&gt; 100 وأنها قد وجدت لديهم تكرار CLL لها على خزعة نخاع العظام. كانت الأورام لها عن قلقها إزاء بدء العلاج الكيميائي دون حل كامل من CDI لها وذلك طلب منا أن أداء FMT.

<table><tbody><tr><td>Blender</td><td>Hamilton Beach</td><td>51101</td><td>&lt;a href=&quot;http://www.amazon.com/Hamilton-Beach-51101-Personal-Blender/dp/B00065L6CU/ref=pd_sim_k_2?ie=UTF8&amp;amp;refRID=1Q06MXAE9XKTB4QG0AEN&quot; target=&quot;_blank&quot;&gt;Link to Amazon.com&lt;/a&gt;</td></tr><tr><td>Strainer</td><td>Winco</td><td>MS3A-8S</td><td>&lt;a href=&quot;http://www.amazon.com/Winco-MS3A-8S-Strainer-Single-Wooden/dp/B001VZ5E40/ref=sr_1_3?s=home-garden&amp;amp;ie=UTF8&amp;amp;qid=1395001295&amp;amp;sr=1-3&amp;amp;keywords=kitchen+strainer+wooden+handle&quot; target=&quot;_blank&quot;&gt;Link to Amazon.com&lt;/a&gt;</td></tr><tr><td>Disposable pads (chux), 17 x 24 in.</td><td>hospital standard</td><td></td><td>one package</td></tr><tr><td>Pink rectangular hospital standard wash basin, 8 quart</td><td>hospital standard</td><td></td><td>1 or 2</td></tr><tr><td>500 cc bottle of sterile normal saline</td><td>hospital standard</td><td></td><td>2</td></tr><tr><td>60 cc luer lock syringes</td><td>hospital standard</td><td></td><td>8 or 9</td></tr><tr><td>Biopsy channel cap with extension</td><td>hospital standard</td><td></td><td></td></tr></tbody></table>

fecal microbiota transplantation via colonoscopy for recurrent c. difficile infection

برازي الجراثيم زرع (FMT) هو علاج آمن وفعال للغاية لالمتكرر وصهر C. عدوى صعب (CDI). وقد تم الإبلاغ عن طرق مختلفة لإدارة FMT في الأدب بما في ذلك أنبوب أنفي معدي، التنظير العلوي، والحقنة الشرجية وتنظير القولون. FMT عبر عائدات قولون معدلات الشفاء ممتازة وأيضا هو جيد التحمل. لقد وجدنا أن المرضى يجدون هذا في وضع مقبول ومقبول من الولادة. في مركزنا، شرعنا في برنامج زرع البراز لمرضى المتكررة أو CDI الحرارية. لقد قمنا بتطوير بروتوكول باستخدام عملية تكرارية المراجعة وأدوا 24 زرع البراز في 22 المرضى الذين يعانون من معدلات نجاح مماثلة لالكتابات المنشورة الحالي. نهج منتظم لالمريض والجهات المانحة الفرز، وإعداد البراز، والتسليم من البراز يزيد النجاح العلاجي. نحن هنا بالتفصيل كل خطوة من بروتوكول FMT التي يمكن القيام بها في أي التنظيرالمركز مع درجة عالية من السلامة والنجاح.

وقد أجرينا 24 FMTS على 22 مريضا في مركز لدينا (الجدول 1) باستخدام بروتوكول أعلاه. وكان تسعة مرضى IBD المتزامنة (ستة يعانون من مرض كرون والتهاب القولون التقرحي مع ثلاثة). كان كل تسعة حل CDI. كان واحدا من المرضى الذين يعانون من مرض كرون الوضع مرحلة ما بعد استئصال القولون ...

Watch this Scientific Journal Video about Fecal Microbiota Transplantation via Colonoscopy for Recurrent C. difficile Infection at JoVE.com

Fecal Microbiota Transplantation via Colonoscopy for Recurrent C. difficile Infection

برازي الجراثيم زرع عن طريق تنظير القولون هو علاج آمن وفعال للعدوى C.difficile المتكرر وصهر. وهناك أسلوب منهجي لالمريض وفحص المانحة، وإعداد البراز، والتسليم من البراز خلال تنظير القولون تعظيم النجاح العلاجي.

برازي الجراثيم زرع عن طريق تنظير القولون لالمتكررة<em&gt; C. صعب</em&gt; العدوى

Fecal Microbiota Transplantation (FMT) is a safe and highly effective treatment for recurrent and refractory C. difficile infection (CDI). Various methods ...

fecal-microbiota-transplantation-via-colonoscopy-for-recurrent-c

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

Fecal Microbiota Transplantation via Colonoscopy for Recurrent C. difficile Infection

27.0K Views.  Brigham and Women‘s Hospital.  برازي الجراثيم زرع عن طريق تنظير القولون هو علاج آمن وفعال للعدوى C.difficile المتكرر وصهر. وهناك أسلوب منهجي لالمريض وفحص المانحة، وإعداد البراز، والتسليم من البراز خلال تنظير القولون تعظيم النجاح العلاجي. 

Video: Fecal Microbiota Transplantation via Colonoscopy for Recurrent C. difficile Infection

A Genetically Engineered Mouse Model of Sporadic Colorectal Cancer

Despite the advantages of easy applicability and cost-effectiveness, colorectal cancer mouse models based on tumor cell injection have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Genetically engineered mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in large organs such as the colon in which only a single tumor is desired.
As a result, an immunocompetent, genetically engineered mouse model of colorectal cancer was developed which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor development is initiated by surgical, segmental infection of the distal colon with adeno-cre virus in compound conditionally mutant mice. The tumors can be easily detected and monitored via colonoscopy. We here describe the surgical technique of segmental adeno-cre infection of the colon, the surveillance of the tumor via high-resolution colonoscopy and present the resulting colorectal tumors.

A protocol for the establishment of a genetically engineered mouse model of colorectal cancer by segmental adeno-cre infection and its surveillance via high-resolution colonoscopy is presented.

نموذج الفأر المهندسة وراثيا لسرطان القولون والمستقيم المتفرقة

Despite the advantages of easy applicability and cost-effectiveness, colorectal cancer mouse models based on tumor cell injection have severe limitations ...

Cancer Research

Culturing and Maintaining Clostridium difficile in an Anaerobic Environment

Clostridium difficile is a Gram-positive, anaerobic, sporogenic bacterium that is primarily responsible for antibiotic associated diarrhea (AAD) and is a significant nosocomial pathogen. C. difficile is notoriously difficult to isolate and cultivate and is extremely sensitive to even low levels of oxygen in the environment. Here, methods for isolating C. difficile from fecal samples and subsequently culturing C. difficile for preparation of glycerol stocks for long-term storage are presented. Techniques for preparing and enumerating spore stocks in the laboratory for a variety of downstream applications including microscopy and animal studies are also described. These techniques necessitate an anaerobic chamber, which maintains a consistent anaerobic environment to ensure proper conditions for optimal C. difficile growth. We provide protocols for transferring materials in and out of the chamber without causing significant oxygen contamination along with suggestions for regular maintenance required to sustain the appropriate anaerobic environment for efficient and consistent C. difficile cultivation.

Culturing and Maintaining Clostridium difficile in an Anaerobic Environment

Clostridium difficile is a pathogenic bacterium that is a strict anaerobe and causes antibiotic associated diarrhea (AAD). Here, methods for isolating, culturing and maintaining C. difficile vegetative cells and spores are described. These techniques necessitate an anaerobic chamber, which requires regular maintenance to ensure proper conditions for optimal C. difficile cultivation.

زراعة وصيانة<em&gt; المطثية العسيرة</em&gt; في بيئة لاهوائية

Clostridium  difficile is a Gram-positive, anaerobic, sporogenic bacterium  that is primarily responsible for antibiotic associated diarrhea (AAD) and is ...

Immunology and Infection

Murine Ileocolic Bowel Resection with Primary Anastomosis

Intestinal resections are frequently required for treatment of diseases involving the gastrointestinal tract, with Crohn&#8217;s disease and colon cancer being two common examples. Despite the frequency of these procedures, a significant knowledge gap remains in describing the inherent effects of intestinal resection on host physiology and disease pathophysiology. This article provides detailed instructions for an ileocolic resection with primary end-to-end anastomosis in mice, as well as essential aspects of peri-operative care to maximize post-operative success. When followed closely, this procedure yields a 95% long-term survival rate, no failure to thrive, and minimizes post-operative complications of bowel obstruction and anastomotic leak. The technical challenges of performing the procedure in mice are a barrier to its wide spread use in research. The skills described in this article can be acquired without previous surgical experience. Once mastered, the murine ileocolic resection procedure will provide a reproducible tool for studying the effects of intestinal resection in models of human disease.

Ileocolic resection is commonly performed in several human diseases; however, little is known regarding the impact of intestinal resection on surgical illnesses. This article provides instruction on executing the procedure in mice with high success, providing a means to study the effects of ileocolic resection in models of disease.

الفئران اللفائفي القولوني استئصال الأمعاء مع التحام الابتدائي

Intestinal resections are frequently required for treatment of diseases involving the gastrointestinal tract, with Crohn&#8217;s disease and colon cancer ...

Cefoperazone-treated Mouse Model of Clinically-relevant Clostridium difficile Strain R20291

Clostridium difficile is an anaerobic, gram-positive, spore-forming enteric pathogen that is associated with increasing morbidity and mortality and consequently poses an urgent threat to public health. Recurrence of a C. difficile infection (CDI) after successful treatment with antibiotics is high, occurring in 20-30% of patients, thus necessitating the discovery of novel therapeutics against this pathogen. Current animal models of CDI result in high mortality rates and thus do not approximate the chronic, insidious disease manifestations seen in humans with CDI. To evaluate therapeutics against C. difficile, a mouse model approximating human disease utilizing a clinically-relevant strain is needed. This protocol outlines the cefoperazone mouse model of CDI using a clinically-relevant and genetically-tractable strain, R20291. Techniques for clinical disease monitoring, C. difficile bacterial enumeration, toxin cytotoxicity, and histopathological changes throughout CDI in a mouse model are detailed in the protocol. Compared to other mouse models of CDI, this model is not uniformly lethal at the dose administered, allowing for the observation of a prolonged clinical course of infection concordant with the human disease. Therefore, this cefoperazone mouse model of CDI proves a valuable experimental platform to assess the effects of novel therapeutics on the amelioration of clinical disease and on the restoration of colonization resistance against C. difficile.

Cefoperazone-treated Mouse Model of Clinically-relevant Clostridium difficile Strain R20291

This protocol outlines the cefoperazone mouse model of Clostridium difficile infection (CDI) using a clinically-relevant and genetically-tractable strain, R20291. Emphasis on clinical disease monitoring, C. difficile bacterial enumeration, toxin cytotoxicity, and histopathological changes throughout CDI in a mouse model are detailed in the protocol.

المعالجة سيفوبيرازون ماوس نموذج سريريا ذات صلة<em&gt; المطثية العسيرة</em&gt; سلالة R20291

Clostridium difficile is an anaerobic, gram-positive, spore-forming enteric pathogen that is associated with increasing morbidity and mortality and ...

Therapeutic Evaluation of Fecal Microbiota Transplantation in an Interleukin 10-Deficient Mouse Model

With the development of microecology in recent years, the relationship between intestinal bacteria and inflammatory bowel disease (IBD) has attracted considerable attention. Accumulating evidence suggests that dysbiotic microbiota plays an active role in triggering or worsening the inflammatory process in IBD and that fecal microbiota transplantation (FMT) is an attractive therapeutic strategy since transferring a healthy microbiota to IBD patient could restore the appropriate host-microbiota communication. However, the molecular mechanisms are unclear, and the efficacy of FMT has not been very well established. Thus, further studies in animal models of IBD are necessary. In this method, we applied FMT from wild-type C57BL/6J mice to IL-10 deficient mice, a widely used mouse model of colitis. The study elaborates on collecting fecal pellets from the donor mice, making the fecal solution/suspension, administering the fecal solution, and monitoring the disease. We found that FMT significantly mitigated the cardiac impairment in IL-10 knockout mice, underlining its therapeutic potential for IBD management.

The interaction of genetic susceptibility, mucosal immunity, and intestinal microecological environment is involved in the pathogenesis of inflammatory bowel disease (IBD). In this study, we applied fecal microbiota transplantation to IL-10 deficient mice and investigated its impact on colonic inflammation and heart function.

التقييم العلاجي لزرع الميكروبات البرازية في نموذج فأر ناقص إنترلوكين 10

With the development of microecology in recent years, the relationship between intestinal bacteria and inflammatory bowel disease (IBD) has attracted ...

Murine Fecal Isolation and Microbiota Transplantation

Gut microbiota dysbiosis plays a role in the pathophysiology of cardiovascular and metabolic disorders, but the mechanisms are not well understood. Fecal microbiota transplantation (FMT) is a valuable approach to delineating a direct role of the total microbiota or isolated species in disease pathophysiology. It is a safe treatment option for patients with recurrent Clostridium difficile infection. Preclinical studies demonstrate that manipulating the gut microbiota is a useful tool to study the mechanistic link between dysbiosis and disease. Fecal microbiota transplantation may help elucidate novel gut microbiota-targeted therapeutics for the management and treatment of cardiometabolic disease. Despite a high success rate in rodents, there remains translational changes associated with the transplantation. The goal here is to provide guidance in studying the effects of gut microbiome in experimental cardiovascular disease. In this study, a detailed protocol for the collection, handling, processing, and transplantation of fecal microbiota in murine studies is described. The collection and processing steps are described for both human and rodent donors. Lastly, we describe using a combination of the Swiss-rolling and immunostaining techniques to assess gut-specific morphology and integrity changes in cardiovascular disease and related gut microbiota mechanisms.

The goal here is to outline a protocol to investigate the mechanisms of dysbiosis in cardiovascular disease. This paper discusses how to aseptically collect and transplant murine fecal samples, isolate intestines, and use the "Swiss-roll" method, followed by immunostaining techniques to interrogate changes in the gastrointestinal tract.

عزل براز الفئران وزرع الجراثيم

Gut microbiota dysbiosis plays a role in the pathophysiology of cardiovascular and metabolic disorders, but the mechanisms are not well understood. Fecal ...

Preparation and Maintenance of Bioexclusion IsoPositive Cage Experiment for Human Fecal Transplantation into Germ-Free Mice

Germ-free mice are an important investigation tool for understanding the contribution of microorganisms in host health and disease, enabling assessment of the specific role of individuals, defined or complex groups of microorganisms in host response. Traditionally bred and reared in flexible-film or semi-rigid isolators, germ-free mouse husbandry and experimental manipulation are costly and require numerous trained staff and a large space footprint in animal housing facilities. The IsoPositive caging system allows for experimental manipulation of germ-free mice in individual, hermetically-sealed, positive-pressure isolator cages (isocages), reducing cost and enabling greater flexibility in experimental manipulations. 
Here, a protocol is described for transferring germ-free mice from breeding isolators to isocages and subsequent fecal transfer from human donor stool into mice to create stable long-term gut "humanized" mice for future studies. The materials and preparation needed for the utilization of the isocage system are described, including the use of chlorine-dioxide sterilant chemical sterilant to clean cages, supplies, equipment, and personal protective equipment. The methods for confirming the germ-free status of transferred mice and how to determine contamination in the caging system are discussed. The procedure for husbandry, including bedding, food, and water supply, is further discussed. The protocol for human fecal slurry preparation and gavage into germ-free mice to create gut "humanized" mice, along with stool collection to monitor the microbial community composition of these mice, are described. An experiment illustrates that two weeks post-human fecal transplant allows for stable colonization of donor microbiota in the murine hosts, enabling subsequent experimental usage. Furthermore, the collection of humanized mouse feces in viability preservation media, enabling use in further functional experiments, is described. Overall, these methods allow for the safe and effective establishment of humanized mouse communities in experimental gnotobiotic cages for further manipulation.

This protocol describes the best practices for germ-free mouse transfer to and housing in experimental single-cage isolators (isocages) while maintaining sterile conditions. Methods for fecal transplant into germ-free mice and the collection of viable bacteria from these gut "humanized" mice for further applications are discussed.

تحضير وصيانة تجربة القفص المتساوي الإيجابي للاستبعاد الحيوي لزرع البراز البشري في الفئران الخالية من الجراثيم

Germ-free mice are an important investigation tool for understanding the contribution of microorganisms in host health and disease, enabling assessment of ...

Fecal Microbiota Transplantation and Detection of Prevalence of IgA-Coated Bacteria in the Gut

Gut microbiota exert pleiotropic roles in human health and disease. Fecal microbiota transplantation (FMT) is an effective method to investigate the biological function of intestinal bacteria as a whole or at the species level. Several different FMT methods have been published. Here, we present an FMT protocol that successfully depletes gut microbiota in a matter of days, followed by transplantation of fecal microbiota from fresh or frozen donor intestinal contents to conventional mice. Real time-PCR is applied to test the efficacy of bacterial depletion. Sequencing of the 16S ribosomal RNA (rRNA) is then applied to test the relative abundance and identity of gut microbiota in recipient mice. We also present a flow cytometry-based detection method of immunoglobulin A (IgA)-coated bacteria in the gut.

We established an efficient way to deplete intestinal bacteria in three days, and subsequently transplant fecal microbiota via gavage of fecal fluid prepared from fresh or frozen intestinal contents in mice. We also present an optimized method to detect the frequency of IgA-coated bacteria in the gut.

Gut microbiota exert pleiotropic roles in human health and disease. Fecal microbiota transplantation (FMT) is an effective method to investigate the ...

Biology

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation

Gut microbiota dysbiosis is thought to exert a role in the progression of colitis. However, the precise standards for probiotic administration in alleviating colitis remain undefined. Most analysis methods rely on limited diversity and abundance of gut microorganisms. Therefore, observational studies cannot establish causation. In this study, we applied antibiotic-induced pseudo-germ-free mice to investigate the role of gut microbiota in regulating the probiotic effects of Bacillus&#160;cereus (B. cereus)&#160;on dextran sulfate sodium (DSS)-induced colitis in mice. This process allows for evaluating the bidirectional regulating effect of B. cereus&#160;supplementation on health and provides stable and reproducible results. Here, the detailed protocols for B. cereus cultivation, gavage operation, stool collection, and antibiotic clearance treatment on colitis mice are provided. The optimization methods are also applicable for other chronic inflammatory-associated disorders. The results showed that B. cereus administration decreased body weight loss, colon length shortening, disease activity index, and histopathological scores. However, treatment with antibiotics suppressed the positive effect of B. cereus on colitis. These results indicate that gut microbiota are needed for the alleviating effects of B. cereus on colitis. Therefore, exploring the beneficial effects of probiotics in this research is a promising approach for developing novel treatment strategies for alleviating the symptoms of chronic inflammatory-associated disorders.

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation

Here, we present a protocol to establish the antibiotic-induced pseudo-germ-free dextran sulfate sodium-induced colitis mouse model to investigate the role of gut microbiota in regulating the positive effects of Bacillus cereus on colitis.

Gut microbiota dysbiosis is thought to exert a role in the progression of colitis. However, the precise standards for probiotic administration in ...

Generating a Double Humanized BLT (Bone-Marrow, Liver, Thymus) Mouse Model

Source: Daharsh, L. et al., <a href="https://appjove.unicartagenaproxy.elogim.com/v/59773">A Double Humanized BLT-mice Model Featuring a Stable Human-Like Gut Microbiome and Human Immune System.</a>&#160;J. Vis. Exp. (2019)
This video demonstrates the method for generating a double humanized bone BLT (marrow, liver, and thymus) mouse with a functional human immune system and a stable engrafted human-like gut microbiome.

توليد نموذج فأر BLT مزدوج (نخاع العظام والكبد والغدة الصعترية)

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Mice ...

برازي الجراثيم زرع عن طريق تنظير القولون لالمتكررة

Summary

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Chapters in this video

نموذج الفأر المهندسة وراثيا لسرطان القولون والمستقيم المتفرقة

زراعة وصيانة

الفئران اللفائفي القولوني استئصال الأمعاء مع التحام الابتدائي

المعالجة سيفوبيرازون ماوس نموذج سريريا ذات صلة

التقييم العلاجي لزرع الميكروبات البرازية في نموذج فأر ناقص إنترلوكين 10

عزل براز الفئران وزرع الجراثيم

تحضير وصيانة تجربة القفص المتساوي الإيجابي للاستبعاد الحيوي لزرع البراز البشري في الفئران الخالية من الجراثيم

Fecal Microbiota Transplantation and Detection of Prevalence of IgA-Coated Bacteria in the Gut

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation

توليد نموذج فأر BLT مزدوج (نخاع العظام والكبد والغدة الصعترية)