Impact of tebipenem pivoxil on the intestinal microbiota and on establishment of colonization with carbapenem-resistant Klebsiella pneumoniae in mice

Abstract

Tebipenem pivoxil has potent in vitro activity against Enterobacterales pathogens, but requires combination with β-lactamase inhibitor to achieve activity against carbapenemase producers, including metallo-β-lactamases (MBLs). Herein, we evaluate the potential of tebipenem pivoxil, alone and in combination with the prodrug of the experimental MBL inhibitor CS319 (CS319-piv-SAc), to disrupt the indigenous mice microbiota of the colon and promote colonization by pathogens. The effect of antibiotic treatment (daily for 3 days with subcutaneous saline [control], subcutaneous clindamycin, oral tebipenem pivoxil alone and in combination with CS319-piv-Sac, or oral CS319-piv-Sac) on the intestinal microbiota was assessed by culture for enterococci and facultative Gram-negative bacilli and by 16S rRNA amplicon sequencing. Mice were also challenged with 10,000 colony-forming units (CFU) of multidrug-resistant (MDR) strain Klebsiella pneumoniae bla_NDM-1, 6 h after the second dose. The concentrations of the MDR K. pneumoniae in stool were measured on days 1, 3, and 6 after challenge. In comparison to saline controls, clindamycin (P = 0.001) and tebipenem pivoxil plus CS319-piv-SAc (P = 0.02) treatment resulted in significant changes in the alpha diversity patterns, whereas tebipenem pivoxil and CS319-piv-SAc individual treatments did not (P > 0.05). Moreover, clindamycin treatment resulted in substantial overgrowth of MDR K. pneumoniae (mean concentration after 6 days of infection, 6.1 vs 2.9 log₁₀ CFU/g stool), whereas the other treatments did not (≤3.6 log10 CFU/g). Although tebipenem pivoxil alone or in combination with an MBL inhibitor, CS319, caused alteration of the mice intestinal microbiota, neither treatment promoted overgrowth of carbapenem-resistant K. pneumoniae.IMPORTANCEIn this work, we used a mouse model to determine the impact of tebipenem pivoxil alone and in combination with a prodrug of an experimental metallo-β-lactamase inhibitor, CS319, on the intestinal microbiota and on the establishment of colonization with carbapenem-resistant Klebsiella pneumoniae. We found that while treatment with tebipenem pivoxil plus the prodrug of CS319 caused alteration of the intestinal microbiota, it did not promote the overgrowth of carbapenem-resistant K. pneumoniae. Although additional studies are needed to examine the impact of tebipenem pivoxil treatment on other multidrug-resistant Gram-negative bacilli, Clostridioides difficile, and Candida spp., our study is a step forward in the understanding of the potential effect of this oral carbapenem on the indigenous microbiota of the colon and on the promotion of colonization by pathogens.

Keywords: carbapenem-resistant Enterobacterales; gut microbiome; intestinal colonization; metallo-β-lactamase inhibitor; tebipenem.

Fig 1

Chemical structures of (A) tebipenem and its oral formulations, and (B) metallo-β-lactamase inhibitor CS319 and its prodrugs CS319-piv-SAc and CS319-SAc.

Fig 2

Effect of treatment on the concentrations of facultative gram-negative bacilli and enterococci in stool. Mice (three per group) received daily doses of the treatments for 3 days and stool specimens were collected ~4 h after the second daily dose. Error bars represent standard error.

Fig 3

Impact of antibiotic treatment on the total bacterial diversity in the stool of mice as indicated by the Shannon diversity index.

Fig 4

Impact of different antibiotic treatment on the relative abundance of different bacterial genera (A). Differentially abundance analysis (P < 0.05, ANOVA, Benjamini-Hochberg [BH]) highlighting 19 taxa at genus level whose abundance was differentially altered in the antibiotic treatment groups versus the saline control group (B).

Fig 5

Effect of treatment on establishment of colonization by Klebsiella pneumoniae. Mice (eight per group) received 10,000 colony-forming units (CFU) of K. pneumoniae 141 by oral gavage on days 2 of 3 of daily antibiotic treatment. The concentration of K. pneumoniae in stool was measured at baseline and on days 1, 3, and 6 after gavage. Error bars represent standard error.