Comparison of Sampling Methods for Detecting Protein in Gastrointestinal Endoscopes.

Background: Persistent microbial contamination of flexible endoscopes has been linked to infections and outbreaks. Valid and reliable sampling methods are critical for monitoring processing effectiveness in flexible endoscopes. In this study, the effectiveness of protein extraction via turbulent fluid flow (TFF) sampling was compared with flush-only sampling in manually cleaned gastrointestinal endoscopes. Methods: A crossover study design, in which both sampling methods were used in alternating order during each endoscope encounter, was utilized to assess protein levels after colonoscopes and gastroscopes underwent manual cleaning. Endoscope channels were sampled with 20 mL sterile water using TFF and flush-only methods. Protein levels were quantified using a spectrophotometer. Results: Protein samples were collected during a total of 40 encounters with 20 unique endoscopes (19 colonoscopes and 21 gastroscopes) following procedural use. More effluent was captured following TFF (20-30 mL) compared with flush-only (19-21 mL) sampling. Zero samples had detectable protein after flush-only sampling, and nine samples (22.5%; two gastroscopes and seven colonoscopes) had detectable protein following TFF sampling (range 1-4 μg/mL). Of those, four exceeded the 2 μg/mL study threshold for recleaning after the first cleaning and three of four dropped to 2 μg/mL or less after recleaning. Conclusion: TFF sampling of the entire suction-biopsy channel allowed the detection of residual protein in nine gastrointestinal endoscopes, whereas no protein was detected in samples obtained by manually flushing the instrument channel. More research is needed to characterize the real-world utility of using the TFF system to verify whether soil and bioburden have been effectively removed during processing. Numerous studies have documented that a majority of fully processed, patient-ready endoscopes harbor microbes.^1-8 Microbes found in endoscopes include high-concern organisms (e.g., multidrug-resistant microbes and pathogens) that have been linked to endoscopy-associated outbreaks.^9-12 In these outbreaks, visible residual soil was discovered during the outbreak investigation. Current guidelines and standards note that effectively cleaning endoscopes is critical to the success of high-level disinfection (HLD) and sterilization.^13,14 Several studies by Ofstead and colleagues^6,15,16 have documented high protein levels on endoscopes. A study involving colonoscopes and gastroscopes detected protein on 100% of manually cleaned endoscopes (range 3-11 μg/mL).⁶ Other studies also found protein in 100% of manually cleaned bronchoscopes (range 2-30 μg/mL) and sterilized ureteroscopes (range 9-32 μg/mL).^15,16 These contamination levels were higher than positive controls, which were dirty gastroscopes that had not been manually cleaned. Microbes were found on 12.5% to 60% of fully processed endoscopes, including potential pathogens such as Pseudomonas aeruginosa, Escherichia coli, and Micrococcus luteus.^6,15,16 This reinforces the need to verify that endoscopes are clean prior to undergoing HLD or sterilization. Evidence shows that protein can persist through multiple rounds of cleaning.^17,18 Despite efforts to clean the endoscope, harvesting samples from surfaces that remain contaminated with soil can be challenging because sampling commonly uses the same tools as cleaning (e.g., brushes or swabs and flushing). Residual soil or bioburden may also be encased in a biofilm matrix that has been hardened through exposure to harsh chemicals used during HLD and/or sterilization and repeated cycles of drying,^19,20 thereby increasing the difficulty of capturing a sample. Hervé et al.²¹ noted that protein deposits in endoscopes were able to resist brushing and flushing, especially in the presence of wear and damage. Historically, flush-only ("flush") sampling was used,^22,23 but this method often was limited to the instrument channel and captured lower yields compared with more robust methods.^4,24,25 As the effectiveness of sampling affects the validity of results of tests for organic soil and microbial cultures, more robust sampling methods may be required.²⁶ Guidance on sampling for microbial cultures provided by the Food and Drug Administration (FDA) and Centers for Disease Control and Prevention (CDC) involves incorporating a brushing step and an additional flushing step ("flush-brush-flush") to dislodge and flush out microbes.²⁷ This method has been found to be more effective than flush sampling,^3,24 but brushes cannot access every endoscope channel and may leave behind bristles. Researchers have reported that the FDA/CDC sampling method is cumbersome, time consuming,²⁸ and prone to contamination.⁷ Even when using recommended sampling methods, investigators have reported needing to rely on external experts and destructive sampling to effectively harvest samples that ultimately revealed the outbreak pathogen.^10,11 This underscores the importance of robust sampling methods, both to avoid false negatives from failing to capture soil or bioburden that is present and to avoid false positives from environmental contamination.^7,29 Given the challenges associated with current sampling techniques for organic soil testing and microbial cultures, this study was conducted to evaluate a method that could potentially improve sample validity and reduce the influence of human factors on sampling. The automated turbulent fluid flow (TFF) system pumps a mixture of air and water through the suction and instrument channels from the suction connector to the distal end and into a sterile collection cup that is sealed during sampling to maintain a closed system. The turbulent flow provides friction to endoscope interior surfaces without needing to use a brush.³⁰ In this study, protein extraction via TFF sampling was compared with flush sampling in manually cleaned gastrointestinal endoscopes.