176


Table 1. Identification of Potential Nosocomial Pathogens on Practitioner Stethoscopesa Set A (n=20)


V1V2 Region Genus


Staphylococcus (g) S. aureus (s)


Pseudomonas (g) Acinetobacter (g) Clostridium (g) Enterococcus (g)


Stenotrophomonas (g) Burkholderia (g)


Frequency, No. (%)


20 (100) 11 (55) 16 (80) 13 (65) 8 (40) 8 (40) 7 (35) 3 (15)


Relative Abundance (mean ± SD)


6.82 ± 5.00


0.23 ± 0.27 0.14 ± 0.14 0.06 ± 0.13 0.04 ± 0.06 0.03 ± 0.05 0.01 ± 0.01


Frequency, No. (%)


20 (100) 13 (65) 20 (100) 20 (100) 12 (60) 18 (90) 18 (90) 3 (15)


V1V2 Region


Relative Abundance (mean ± SD)


8.06 ± 4.35


0.37 ± 1.00 0.17 ± 0.10 0.02 ± 0.02 0.14 ± 0.28 0.06 ± 0.10 0.01 ± 0.02


Frequency, No. (%)


20 (100) 13 (65) 20 (100) 20 (100) 18 (90) 18 (90) 6 (30) 8 (40)


Vincent R. Knecht et al


Set C (n=20) V4 Region


Relative Abundance (mean ± SD)


14.01 ± 6.41


1.87 ± 3.22 1.36 ± 1.05 0.12 ± 0.17 0.08 ± 0.12 0.01 ± 0.01 0.01 ± 0.01


Note. SD, standard deviation. aSequence data from the practitioner stethoscopes in sets A and C were investigated for the presence of selected bacterial genera (g) that commonly cause hospital-acquired infections


(HAIs), based on 16S rRNA gene V1V2 or V4 region sequences as described in the Methods section. Frequency indicates the proportion of practitioner stethoscopes with that genus. Relative abundance indicates the percentage of all bacterial sequences on a stethoscope that genus represents, across the practitioner stethoscope sets. For Staphylococcus, some sequences could be assigned at the species level (s), and those identified as S. aureus are indicated.


2 different sets clearly establish that these taxa are widely present on stethoscopes, albeit at low relative abundances. Other noso- comial infection-related genera were also identified on practi- tioner stethoscopes at high frequency but lower relative abundances. Because this is the first study to analyze stethoscope contamination at the molecular level, it remains to be determined what amount of contamination is clinically relevant for potential nosocomial transmission. We also applied these molecular methods to determine the


impact of cleaning the stethoscope diaphragm, testing both a standardized approach and practitioners’ usual methods. Both methods significantly reduced the 16S DNA bacterial biomass but failed to bring contamination to the level of clean stethoscopes. The standardized cleaning method reduced more stethoscopes to the clean level (5 of 10) than the practitioner-preferred method (2 of 20). PCoA analysis of bacterial communities pre- and post-cleaning revealed a shift toward background communities detected by the analysis of community membership only, but not when bacterial relative abundance was incorporated (unweighted vs weighted Uni- Frac; Fig. 5). This results implicates low-abundance taxa as con- tributing the most to differences, whereas themore abundant taxa are not substantially altered by cleaning. Notably, although the CDC offers recommendations on stethoscope decontamination,23 studies suggest that it is infrequently practiced.1,2 Our study has several limitations. We were not able to identify


bacterial taxa at the species level for most OTUs in our data set, and thus most HAI-associated bacteria could only be defined at the genus level. Future studies might complement comprehensive unbiased 16S rRNA gene analysis with species-targeted amplifi- cation. We were not able to identify drug-resistant species using this method. Because this is a DNA-based approach, it cannot distinguish between bacteria that are dead versus alive.24 Finally, we did not analyze fungal or viral sequences to understand the complete scope of organisms that stethoscopes carry in the ICU. In summary, this study is the first comprehensive molecular


analysis of bacterial contamination of stethoscopes in an ICU, the presence of potential nosocomial pathogens, and the impact of


cleaning methods. Practitioner stethoscopes are contaminated by a plethora of bacteria, including organisms that may be associated with nosocomial infections. Cleaning reduces contamination but does not bring the bacterial biomass down to the level of clean stethoscopes nor does it significantly change the overall com- munity composition. Thus, stethoscopes are a potential vector of HAI transfer. Useful future directions would be to use these molecular approaches to identify improved cleaning methods, enhance species-level identification of pathogens, quantify live versus dead bacteria, and define fungal and viral contaminants as well. In addition, shotgun metagenomic sequencing would be useful to analyze drug-resistance genes that might be carried between patients on practitioner stethoscopes.


Supplementary materials. To view supplementary material for this article, please visit https://doi.org/10.1017/ice.2018.319


Acknowledgments. We thank members of the ICU staff who assisted with this project and members of the Collman and Bushman labs who provided input and advice. We also thank D. Pegues for critical review of the manuscript.


Financial support. This work was supported by the National Institutes of Health (grant nos. R01-HL113252 and R61-HL137063). We received assistance from the Penn-CHOP Microbiome Program and the Penn Center for AIDS Research (grant no. P30-AI045008).


Conflicts of interest. The authors have no financial or other conflicts of interest relevant to this article.


References


1. Holleck JL, Merchant N, Lin S, Gupta S. Can education influence stethoscope hygiene? Am J Infect Control 2017;45:811–812.


2. Jenkins IH, Monash B,WuJ, Amin A. The third hand: low rates of stethoscope hygiene on general medical services. JHospMed 2015;10:457–458.


3. Whittington AM, Whitlow G, Hewson D, Thomas C, Brett SJ. Bacterial contamination of stethoscopes on the intensive care unit. Anaesthesia 2009;64:620–624.


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