Bacteriophage treatment of carbapenemase-producing <em>Klebsiella pneumoniae</em> in a multispecies biofilm: a potential biocontrol strategy for healthcare facilities

• Main Authors: Ariel J. Santiago, Maria L. Burgos-Garay, Leila Kartforosh, Mustafa Mazher, Rodney M. Donlan
• Format: Article
• Language: English
• Published: AIMS Press 2020-03-01
• Series: AIMS Microbiology
• Subjects: biofilms; bacteriophage; carbapenemase-producing klebsiella pneumoniae; healthcare-associated infections
• Online Access: https://www.aimspress.com/article/10.3934/microbiol.2020003/fulltext.html

The p-traps of hospital handwashing sinks represent a potential reservoir for antimicrobial-resistant organisms of major public health concern, such as carbapenemase-producing KPC+ <em>Klebsiella pneumoniae</em> (CPKP). Bacteriophages have reemerged as potential biocontrol agents, particularly against biofilm-associated, drug-resistant microorganisms. The primary objective of our study was to formulate a phage cocktail capable of targeting a CPKP strain (CAV1016) at different stages of colonization within polymicrobial drinking water biofilms using a CDC biofilm reactor (CBR) p-trap model. A cocktail of four CAV1016 phages, all exhibiting depolymerase activity, were isolated from untreated wastewater using standard methods. Biofilms containing <em>Pseudomonas aeruginosa, Micrococcus luteus, Stenotrophomonas maltophilia, Elizabethkingia anophelis, Cupriavidus metallidurans,</em> and <em>Methylobacterium fujisawaense</em> were established in the CBR p-trap model for a period of 28 d. Subsequently, CAV1016 was inoculated into the p-trap model and monitored over a period of 21 d. Biofilms were treated for 2 h at either 25 °C or 37 °C with the phage cocktail (10 9 PFU/ml) at 7, 14, and 21 d post-inoculation. The effect of phage treatment on the viability of biofilm-associated CAV1016 was determined by plate count on m-Endo LES agar. Biofilm heterotrophic plate counts (HPC) were determined using R2A agar. Phage titers were determined by plaque assay. Phage treatment reduced biofilm-associated CAV1016 viability by 1 log 10 CFU/cm 2 (p &lt; 0.05) at 7 and 14 d (37 ℃) and 1.4 log 10 and 1.6 log 10 CFU/cm 2 (p &lt; 0.05) at 7 and 14 d, respectively (25 ℃). No significant reduction was observed at 21 d post-inoculation. Phage treatment had no significant effect on the biofilm HPCs (p &gt; 0.05) at any time point or temperature. Supplementation with a non-ionic surfactant appears to enhance phage association within biofilms. The results of this study suggest the potential of phages to control CPKP and other carbapenemase-producing organisms associated with microbial biofilms in the healthcare environment.