Spread of Multidrug-Resistant Bacteria by Moth Flies from Hospital Waste Water System

We documented and analyzed moth fly occurrence and spread of multidrug-resistant bacteria in a tertiary care hospital in Germany. The moth flies (Clogmia albipunctata) bred in the sewage system, then moved into the hospital, carrying biofilm and multidrug-resistant bacteria on their feet. Subsequently, the hospital developed a pest control protocol.

Fluorescence in situ hybridization and fluorescence microscopy on sections of embedded moth flies showed the presence of biofilms and bacteria on their feet ( Figure 2). Not every moth fly caught was tested; however, representative moth flies from each identified location were tested. Furthermore, biofilm in sewage pipes revealed a kind of ecosystem consisting of moth fly larvae, vermicular, fungi, and bacteria (Appendix Figure 1). Subsequently we collected and microbiologically analyzed C. albipunctata and biofilm from different parts of the hospital (Tables 1, 2). Of the moth flies we analyzed microbiologically, we found that 41.1% carried MDR or even XDR bacteria (Appendix). Overall, 43.9% of specimens were MDR or XDR.
We subsequently examined pest control options. Our first approach, removing biofilm from accessible pipes in the sewage system, did not successfully reduce or eliminate moth flies. Our second approach, mechanically and chemically cleaning all sinks and proximal sewage lines, also did not prevent periodic reoccurrence of moth flies. Our third approach was more successful. We flushed all sinks in the OR at the same time with 60°C hot water for 15 min/wk, daily during summer, which suppressed C. albipunctata moth flies in the OR but not in the rest of the hospital.

Conclusions
Infection with and colonization by MDR bacteria is an increasing challenge in health care (6). Psychodidae (moth flies) are small, 1-4 mm in size, and have been Magnified Clogmia albipunctata moth fly. The length of the corpus is 2.5 mm. B) Fluorescence in situ hybridization (FISH) from biofilm of a sewage pipe with a blind end in the operating room (OR) using the pan-bacterial FISH-probe EUB338 labeled with fluorescein isothiocyanate (green), Pseudomonas aeruginosa specific probe labeled with Cy3 (orange), and nucleic acid stain DAPI (Thermo Fisher Scientific, https://www.thermofisher.com/) (blue). The oval structure seen is consistent with an eggshell of Psychodidae, which is colonized by bacteria. C) Schematic map of floor -3 (topographically representative of the hospital building). Blue lines represent the main sewage pipe; heat map shows the frequency of C. albipunctata occurrence merged from all floors of the building; blue arrow indicates sewage system discharge from the hospital; black arrows indicate where moth flies were captured on level -3. Arrow numbers correspond to the numbers in Tables 1, 2  Overview showing an overlay of the fluorescent images with a phase contrast to visualize the limbs of the legs. B) Higher magnification of the inset from panel A shows the anatomy of the tarsus and claws with an adjacent biofilm, which is stained by the bacterial probe (green) and DAPI (blue). C) Higher magnification of the inset from panel B shows the biofilm. Blue represents DAPI staining of DNA; bacteria were stained green with pan-bacterial FISH probe EUB338-FITC, Enterobacterales stained orange with an Escherichia coli-specific FISH probe (data not shown), and NONEUB (nonsense EUB) probe labeled with Cy5 was used to exclude unspecific probe binding. D) Overlay of the DAPI and fluorescein isothiocyanate channel shows the biofilm with different bacterial morphotypes. Different planes of the z-stack in the green channel (pan-bacterial probe) of the identical microscopic field depicts the different claws embracing the biofilm (D1 and D2). D3 shows the DAPI filterset only with the DNA of the bacteria, whereas D4 shows the autofluorescence in the Cy5 filter-set NONEUB probe.
regarded as unharmful vermin except in highly sterile areas. Therefore, they have often been overlooked or ignored and not considered a high-consequence problem.
The results of our study suggest a change in this point of view is needed. If generalized to other hospitals, our findings indicate that C. albipunctata moth flies in hospitals, combined with MDR, XDR, or PDR (pandrug-resistant) bacteria in biofilms, pose an underestimated threat. The danger from this symbiotic system between moth flies and these bacteria results from moth fly eggs and larvae living in biofilm that is contaminated by a patient's bacterial flora. Furthermore, biofilms can rapidly grow and spread over distances kilometers in length (7) and are almost impossible to eradicate. In the third and fourth stages of development, larvae living in the biofilm can begin to move, thus overcoming the water barriers in showers, bathtubs, toilets, and other washing units. At this point, adult moth flies can enter the hospital (Video 1) and transport drug-resistant bacteria from the microbial flora of the biofilm into the hospital.
We frequently found Stenotrophomonas maltophilia on C. albipunctata moth flies and also in clinical samples from deep respiratory material, wounds, blood culture, urine, and bile. In 1 patient, for example, we found hospital-acquired S. maltophilia and a genetically identical strain in drains in a ward ≈250 m away (data not shown). Even though this evidence is scant, it does support our hypothesis.
In addition, low doses of antimicrobials excreted by patients can result in the quick development and spread of plasmids (resistance genes) and virulence factors in biofilms (8). This process might result in resistance developing not only in a patient's microbiota but also in hospital biofilm. Our observations suggest that the adult C. albipunctata moth flies can move freely throughout sewage systems and that they carry bacterial biofilm on their feet. Many authors have suggested the existence of missing links in polyclonal outbreaks and in other hard-to-explain observations (9,10). We hypothesize that moth flies in symbiotic combination with biofilms could, in part, explain one such observed Patient numbers correspond to the numbers in Table 2 and Figure 1 ¶In OR 12 only 1 moth fly was found even after an intense search. From all immediately reported occasions 1-2 moth flies were caught and analyzed microbiologically (see Table 2 for results).
transmission. However, the findings of this study are limited by the moderate number of moth flies, which should be addressed in future investigations. Currently, there are no proven strategies, including chemical methods, to prevent or eradicate moth flies in sewage systems. However, weekly or, during summer, daily flushing with hot water (60°C) for 15 min was sufficient to suppress the moth flies in our study. We propose a prevention protocol including flushing weekly or daily with hot water (60°C), mechanical removal of biofilms; deconstruction of unused siphons or replacement by heatable siphons; and checking for unexpected outlets, such as drill holes, from drains into hospital rooms. These measures will not eliminate but might substantially suppress the problem. Once moth flies leave the drains, among the few available biofilms are patient wounds. Research has reported that adult moth flies are attracted to them, and C. albipunctata larvae have been found in wounds (11,12). Searching for moth flies and determining their microbial load might be advisable, especially if an unexpected bacterial outbreak occurs. Finally, our observations should be taken into account in the planning of hospital sewage systems in the future. Moth fly ICU floor C Sterile NA ND NA *Numbers at left are patient numbers, which correspond to the numbers in Table 1 and Figure 1, panel C. CS, cardiosurgery; C, cardiology; ICU, intensive care unit; MDR, multidrug resistant; OR, operating room; NA, not applicable; ND, not detected; XDR, extensively drug resistant. †Microbiology, histology, and fluorescence in situ hybridization findings in the investigated specimens. Note that not every moth fly from the Table 1 count column was analyzed. ‡Resistance levels defined in Magiorakos et al. (13). §Potential XDR means that the international standards have not been applied to the bacterium S. maltophilia. Additional details are available in the Appendix (https://wwwnc.cdc.gov/EID/article/26/8/19-0750-App1.pdf). ¶Possible XDR means that it could be XDR but not all antimicrobial categories were tested. Additional details are available in the Appendix.