A Single Intranasal Dose of Bacterial Therapeutics to Calves Confers Longitudinal Modulation of the Nasopharyngeal Microbiota: a Pilot Study

Abstract

To address the emergence of antimicrobial-resistant pathogens in livestock, microbiome-based strategies are increasingly being sought to reduce antimicrobial use. Here, we describe the effects of intranasal application of bacterial therapeutics (BTs) on the bovine respiratory microbiota and used structural equation modeling to investigate the causal networks after BT application. Beef cattle received (i) an intranasal cocktail of previously characterized BT strains, (ii) an injection of metaphylactic antimicrobial (tulathromycin), or (iii) intranasal saline. Despite being transient colonizers, inoculated BT strains induced longitudinal modulation of the nasopharyngeal bacterial microbiota while showing no adverse effect on animal health. The BT-mediated changes in bacteria included reduced diversity and richness and strengthened cooperative and competitive interactions. In contrast, tulathromycin increased bacterial diversity and antibiotic resistance and disrupted bacterial interactions. Overall, a single intranasal dose of BTs can modulate the bovine respiratory microbiota, highlighting that microbiome-based strategies have potential in being utilized to mitigate bovine respiratory disease in feedlot cattle. IMPORTANCE Bovine respiratory disease (BRD) remains the most significant health challenge affecting the North American beef cattle industry and results in $3 billion in economic losses yearly. Current BRD control strategies mainly rely on antibiotics, with metaphylaxis commonly employed to mitigate BRD incidence in commercial feedlots. However, the emergence of multidrug-resistant BRD pathogens threatens to reduce the efficacy of antimicrobials. Here, we investigated the potential use of novel bacterial therapeutics (BTs) to modulate the nasopharyngeal microbiota in beef calves, which are commonly administered metaphylactic antibiotics to mitigate BRD when sourced from auction markets. By direct comparison of the BTs with an antibiotic commonly used for BRD metaphylaxis in feedlots, this study conveyed the potential use of the BTs to modulate respiratory microbiome and thereby improve resistance against BRD in feedlot cattle.

Keywords: bacterial therapeutics; beef cattle; bovine respiratory pathogens; cattle; intranasal; microbial interactions; respiratory microbiota; structural equation modeling; tulathromycin.

FIG 1

Prevalence of the BRD-associated pathogens in the nasopharynx of cattle over the course of 28 days, determined by culturing nasopharyngeal swabs. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group). *, significant difference between treatments (P < 0.05).

FIG 2

Estimated abundances of total bacteria (top) and Lactobacillus (bottom) in nasopharyngeal swab samples obtained from calves that received either intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group) on day 0. Gene copy numbers were quantified by qPCR. The results are presented as estimated mean ± SEM. Asterisks represent significant difference between treatments (*, P < 0.05; **, P < 0.01; ****, P < 0.0001).

FIG 3

Detrended correspondence analysis (DCA) plots of the Bray-Curtis metric for bacteria in nasopharyngeal samples collected from cattle and visualized by time or treatment. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group). The percentages of variation explained by the DCA are indicated on the axes.

FIG 4

Alpha diversity of bacteria in nasopharyngeal samples collected from cattle over 43 days. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group). The top plot shows the mean richness estimate, and the bottom panel shows the mean estimated Shannon diversity. Error bars indicate standard error of the mean.

FIG 5

The five most relatively abundant phyla in the nasopharyngeal microbiota of calves. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group).

FIG 6

Heatmap comparing the bacterial microbiota in the nasopharynx of cattle. Taxa (n = 28) that showed a significant change (P < 0.05) from baseline (day −1) in BT and MP groups above and beyond any changes in the control (CTRL) group over the course of 42 days. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group).

FIG 7

Ecological network of observed bacterial genera in nasopharyngeal samples of calves. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group).

FIG 8

Proportion (%) of the resistance determinants msr(E) and tet(H) to 16S rRNA gene copies in nasopharyngeal samples obtained from cattle, quantified by qPCR. On day 0, cattle were treated with intranasal bacterial therapeutics (BT), intranasal PBS (CTRL), or subcutaneous tulathromycin (MP) (n = 20 per group). The results are presented as estimated mean ± standard error of the mean.