Widespread heteroresistance to antibiotics in Lactobacillus species

Abstract

Lactobacilli are prevalent members of the intestinal and reproductive tract microbiota of humans and other species. They are commonly used in probiotics and various food products due to their beneficial effects on human health. For example, these beneficial microbes are used to treat diarrhea caused by antibiotic therapy and are commonly given during antibiotic treatment. Despite the many studies conducted to understand the beneficial effects of Lactobacilli, less is known about their resistance and heteroresistance to antibiotics. In this study, we evaluated the resistance heterogeneity in eight Lactobacillus species. Our results demonstrate that several Lactobacilli species, including Lactobacillus rhamnosus, are heteroresistant to antibiotics, a recently discovered phenotype commonly seen in multidrug-resistant organisms that cause clinical failures but understudied in commensals and probiotics.

Keywords: Lactobacillus; antibiotic resistance; heteroresistance; microbiota; probiotics.

Figure 1:. Lactobacilli have varied resistance to different antibiotics.

Population analysis profile assays (PAPs) of seven Lactobacillus isolates plated on A) Ciprofloxacin (MIC: 4μg/ml), B) Amoxicillin (MIC: 16μg/ml), C) Cefepime (MIC: 16μg/ml), D) Vancomycin (MIC: 32μg/ml), E) Streptomycin (MIC: 16μg/ml), F) Azithromycin (MIC: 8μg/ml), G) Tetracycline (MIC: 16μg/ml) in aerobic conditions. Gray-filled dots indicate samples below the level of detection. Symbols represent bacterial species. Graphs are representative of 3 independent experiments (n=3).

Figure 2:. Lactobacillus murinus is heteroresistant to ciprofloxacin.

Lactobacillus murinus was subcultured in MRS with ciprofloxacin (MIC: 4μg/ml) for 48 hours followed by subcultures in ciprofloxacin-free MRS media for 14 days. Data represents the proportion of ciprofloxacin resistant bacteria in the culture each day. Symbols represent the mean of triplicates and is representative of three independent experiments (n=9).

Figure 3:. L. rhamnosus is heteroresistant to Fluoroquinolones Ciprofloxacin and Levofloxacin

Population analysis profiles assay (PAPs) of Lactobacillus rhamnosus plated on A) Ciprofloxacin (MIC: 4μg/ml), B) Amoxicillin (MIC: 16μg/ml), C) Cefepime (MIC: 16μg/ml), D) Vancomycin (MIC: 32μg/ml), E) Streptomycin (MIC: 16μg/ml), F) Azithromycin (MIC: 8μg/ml), G) Tetracycline (MIC: 16μg/ml) in aerobic conditions. Gray-filled dots indicate samples below the level of detection. PAPs were performed in triplicates Graphs are represewntative of 3 independent experiments. H) L. rhamnosus was subcultured in MRS with ciprofloxacin (MIC: 4μg/ml) for 48 hours followed by subcultures in ciprofloxacin-free MRS media for 14 days. Data represents the proportion of ciprofloxacin resistant bacteria in the culture each day. Symbols represent the mean of triplicates and are representative of three independent experiments (n=9).