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Mechanistic investigations of host-microbe interactions in the human gut are limited by current coculture model systems. The intestinal epithelium requires oxygen for viability, while gut bacteria are facultative or obligate anaerobes. The ability to model host-commensal interactions under dynamic oxygen conditions is critical to understanding host-pathogen interactions in the human gut. Here, we demonstrate a simple, cost-effective method for co-culturing obligate anaerobic bacteria with human intestinal enteroid monolayers under variable oxygen conditions. The Enteroid-Anaerobe Co-Culture (EACC) system is able to recapitulate the steep oxygen gradient seen in vivo and induce expression of hypoxia-associated phenotypes such as increased barrier integrity and expression of antimicrobial peptide genes. Using clinical strains of the commensal anaerobes Bacteroides thetaiotaomicron and Blautia sp. on established patient-derived intestinal enteroid cell lines under physiological hypoxia, the EACC system can sustain host-anaerobe interactions for at least 24 hours. Following co-culture with anaerobic bacteria, we demonstrate patient-specific differences in epithelial response, reinforcing the potential to develop a personalized medicine approach to bacteriotherapy and host-microbe interaction investigations. Our innovative EACC system provides a robust model for investigating host-microbe interactions in complex, patient-derived intestinal tissues, that facilitates study of mechanisms underlying the role of the microbiome in health and disease.

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The most common etiologic agents for viral gastroenteritis are rotavirus, adenovirus, and members of the “small round structure virus” category (enterovirus, norwalk virus, calicivirus, astrovirus). A rapid diagnosis of viral infection can avoid unnecessary antibiotic therapy, extensive and costly medical workups and reduce the patient’s hospital stay. Pseudoreplication has been the preparation method of choice at many institutions, because of its concentrating effect on viral particles. Unfortunately, in addition to concentrating viral particles, this method also concentrates fecal debris, making the screening process quite difficult at times and time consuming.A rapid method of negative staining for viral particle identification in fecal samples is presented. The method first described by Cubitt et al has been modified, resulting in a reduction in the amount of debris and improved staining quality of the viral particles, while still concentrating the number of viral particles. This method requires less than 1 ml of stool specimen and may be prepared in less than three minutes.