Music exposure enhances resistance to Salmonella infection by promoting healthy gut microbiota

Abstract

Music intervention is gaining recognition as a cost-effective therapeutic for improving human health. Despite its growing application, the mechanisms through which music exerts beneficial health effects remain largely unexplored. Here, we show that music can exert beneficial effects in mice through modulating gut microbiome composition. Adult mice were exposed to ambient noise, Mozart's Flute Quartet in D Major, K. 285, or white noise over a three-week period. Afterward, we observed treatment-specific changes in the community of gut commensal bacteria in these animals. Upon subsequent challenge with the bacterial pathogen Salmonella typhimurium, control groups exhibited significant weight loss and increased Salmonella colonization, whereas the Mozart-treated group did not. 16S ribosomal RNA gene sequencing revealed that the Mozart group showed a significant increase in Lactobacillus salivarius, a probiotic known for its antibacterial properties. Further experiments confirmed that L. salivarius mitigated Salmonella infection in mice and that L. salivarius acidified local environments in in vitro culture, thus inhibiting Salmonella growth. Additionally, mice exposed to Mozart consumed more food but showed similar body weight compared to the control groups. Behavioral assessments, including open field and object location tests, revealed that Mozart-treated mice were more active, less anxious, and exhibited enhanced spatial memory. Finally, Mozart exposure was shown to significantly boost colonization of administered L. salivarius and alter gut metabolite profiles. These findings suggest that music exposure fosters healthier gut microbiota, enhancing resistance to bacterial infections and highlighting the potential of music therapy as a novel strategy to combat drug-resistant pathogen infections.

Importance: Music therapy is increasingly recognized as a low-cost approach to improving health, but how it works remains unclear. Our study demonstrates that music can positively influence health by altering the gut microbiome. In a mouse model, exposure to Mozart's Flute Quartet in D Major enhanced the gut microbiota, specifically increasing levels of the beneficial bacterium Lactobacillus salivarius. This probiotic protected mice from Salmonella infection by creating an acidic environment that inhibited pathogen growth. Mozart-treated mice also showed reduced anxiety, better spatial memory, and higher food intake without weight gain, suggesting the benefits of music exposure. These findings reveal a novel link between music, gut health, and disease resistance, suggesting that music therapy could be a promising strategy for enhancing gut microbiota and combating infections, including those caused by drug-resistant bacteria.

Keywords: Lactobacillus; Salmonella; metabolomics; microbiota; music therapy.

Fig 1

Sound treatments impact the composition of mouse gut microbiota. (A) Schematic diagram of procedures for animal experiments. T0: prior to the sound exposure; T1: 3 weeks after sound exposure. (B) Unweighted unifrac principal coordinates analysis (PCoA) of 16S gene V4 region amplicon sequencing. Percent variance is explained by each axis in parentheses. (C) Sound effects on the relative abundance of L. salivarius. ns, no significance; *P < 0.05 (one-way ANOVA).

Fig 2

Salmonella infection of sound-exposed mice. (A, B) Weight changes of mice prior (A) and after (B) exposure to 10⁸/mouse Salmonella were orally inoculated into mice exposed to different sound treatments. In A, weight change of the weight on Day 0 of the experiment, and in B, % change of weight on Day 0 of Salmonella infection. ***P < 0.005 (Unpaired t-test). (C) Colonization of Salmonella. Fecal pellets were collected, and viable CFU of Salmonella was determined by serial dilution and spreading them on selective LB agar plates. *P < 0.05 (unpaired t-test).

Fig 3

L. salivarius inhibits Salmonella growth in vivo and in vitro. (A) L. salivarius inhibits Salmonella growth in vivo. Six-week-old female CD-1 mice were given drinking water containing 1 mg/mL streptomycin for 1 day. PBS buffer (empty circles) or 10⁹ CFU/mouse L. salivarius (green squares) were then intragastrically inoculated into mice. Two days later, 10⁸ CFU/mouse Salmonella were orally inoculated into mice. Fecal pellets were collected after 1 day of infection and colonized Salmonella were enumerated. Horizontal lines: Geometric mean of 5 mice. **P < 0.01 (t-test). (B) Salmonella survival rate in the presence of L. salivarius. 10⁶ CFU/mL of Salmonella and different amounts of L. salivarius were inoculated into MRS/LB (50:50) media and incubated at 37°C without shaking for 16 h. The cultures were then serially diluted and spotted onto LB agar plates. The plates were then incubated at 37°C for 16 h before being photographed. L. salivarius was unable to grow on these plates. (C) 10⁷/mL Salmonella without or with 10⁷/ml L. gasseri and L. salivarius were inoculated into MRS/LB and incubated at 37°C stationarily. Medium pH was measured by a pH meter at the time point indicated. N = 3. ****P < 0.0001 (t-test) (compared to the medium pH of Salmonella only at the corresponding time points). (D) Salmonella was seeded in the MRS/LB media containing 0.3% agar without (top) and with (bottom) the pH indicator bromocresol purple (0.1%). Ten microliters of overnight cultures of L. salivarius and L. gasseri was then spotted. The plates were incubated at 37°C for 16 h before photographed. (E) The effects of acidified media on Salmonella growth. 10⁷/mL Salmonella were inoculated into LB containing 50% fresh MRS, L. salivarius-grown spent media (SM), and spent media adjusted to pH 6.5 using NaOH. The cultures were incubated at 37°C stationarily. OD₆₀₀ was measured. N = 12. ****P < 0.0001 (t-test) (compared to OD₆₀₀ of fresh MRS at the corresponding time points).

Fig 4

The sound effects on L. salivarius gut colonization. (A) Schematic diagram of procedures for animal experiments. Six-week-old CD-1 mice were treated with streptomycin (1 mg/mL) in their drinking water for 3 days and continued during L. salivarius inoculation. The L. salivarius used was naturally resistant to streptomycin. Mice were inoculated intragastrically with 10⁹ CFU/mouse L. salivarius daily for 3 days. Mice were then exposed to the music or white noise, as described in the Materials and Methods. From days 30 to 37, the sound treatment was removed. (B) L. salivarius colonization. At the time points indicated, fecal pellets were collected and L. salivarius colonization was determined by serial diluting the fecal homogenates and plating on the MRS plates containing 100 µg/mL streptomycin. N = 5. *P < 0.05 (unpaired t-test).

Fig 5

Music’s effects on mouse behavior. (A) Schematic diagram of procedures for animal experiments. After acclimatization, the sound treatment was from day 0 to day 30 and then removed from day 30 to day 37. (B) Percentage of weight change. The weight of each mouse was compared with the weight of the same mouse 1 day prior. (C) Food consumption, representing the weight of food placed in each cage (5 mice) minus the weight of remaining food after 24 h. *P < 0.05, **P < 0.005 (unpaired two-sample t-test for Mozart vs White Noise). (D) Open Field Tests. Thigmotaxis plot (proportion of time spent in the outer peripheral of the arena). Horizontal lines: mean of at least 9 mice. ns, no significance; **P < 0.01 (unpaired t-test). (E) Task New Object Exploration Task, showing the percentage of time spent on exploration of the moved object over the total time spent on exploring all the objects. Horizontal lines: mean of 7 mice. *P < 0.05 (unpaired t-test).

Fig 6

The sound effects on host metabolomics. The procedures for animal experiments were the same as in Fig. 5A. Fecal samples were collected prior to the sound treatment (A), 3 weeks after the treatment (B), and 1 week after the treatment stopped (C). PCoA of Euclidean distances between metabolite peak area across samples showed a clear separation and grouping of Mozart and WN samples, suggesting a clear difference in metabolites and metabolite abundance between these groups. *P < 0.05 (PERMANOVA).