Effects of danofloxacin dosing regimen on gastrointestinal pharmacokinetics and fecal microbiome in steers

Abstract

Fluoroquinolones are a class of antimicrobial commonly used in human medicine, and deemed critical by the World Health Organization. Nonetheless, two formulations are approved for the treatment of respiratory disease in beef cattle. The objective of this study was to determine the gastrointestinal pharmacokinetics and impact on enteric bacteria of cattle when receiving one of the two dosing regimens (high: 40 mg/kg SC once or low: 20 mg/kg IM q48hr) of danofloxacin, a commonly utilized synthetic fluoroquinolone in veterinary medicine. Danofloxacin was administered to 12 steers (age 7 months) fitted with intestinal ultrafiltration devices at two different dosing regimens to assess the gastrointestinal pharmacokinetics, the shifts in the gastrointestinal microbiome and the development of resistant bacterial isolates. Our results demonstrated high intestinal penetration of danofloxacin for both dosing groups, as well as, significant differences in MIC values for E. coli and Enterococcus between dosing groups at selected time points over a 38 day period. Danofloxacin treatment consistently resulted in the Euryarchaeota phyla decreasing over time, specifically due to a decrease in Methanobrevibacter. Although microbiome differences were minor between dosing groups, the low dose group had a higher number of isolates with MIC values high enough to cause clinically relevant resistance. This information would help guide veterinarians as to appropriate dosing schemes to minimize the spread of antimicrobial resistance.

Figure 1

Total concentration of danofloxacin in plasma, interstitial fluid (ISF), ileum and colon ultrafiltrate dosed at 6 mg/kg, once every 48 h. Each point represents the mean and the error bars represent the standard deviation. The circle indicates plasma concentrations, the square indicates ISF concentrations, the upside down triangle indicates ileum ultrafiltrate concentrations and the right side up triangle indicates colon ultrafiltrate concentrations. The data is presented on a semi-logarithmic axis. The solid lines represent the predicted concentration based on the model fit. The actual values are the solid points.

Figure 2

Total concentration of danofloxacin in plasma, interstitial fluid (ISF), ileum and colon ultrafiltrate dosed at 8 mg/kg once. Each point represents the mean and the error bars represent the standard deviation. The circle indicates plasma concentrations, the square indicates ISF concentrations, the upside down triangle indicates ileum ultrafiltrate concentrations and the right side up triangle indicates colon ultrafiltrate concentrations. The data is presented on a semi-logarithmic axis. The solid lines represent the predicted concentration based on the model fit. The actual values are the solid points.

Figure 3

Mean Log CFU/g of E. coli and Enterococcus per dosing group over time with standard error. (A) Shows the Mean log CFU/g of E. coli while (B) shows the mean log CFU/g of Enterococcus. The black line is the high dose group and the red line is the low dose group. Individual t-tests with a Bonferonni correction (p < 0.0125) were performed for overall log CFU/g at predetermined time points. The only significant difference seen for E. coli was comparing Day 0 to Day 6 for the low dose group only (p = 0.0006). There was no significant differences noted for Enterococcus.

Figure 4

Heat Map demonstrating the number of E. coli isolates per each MIC value over time for both the low dose (A) and high dose group (B).

Figure 5

Heat Map demonstrating the number of Enterococcus isolates per each MIC value over time for both the low dose (A) and high dose group (B).

Figure 6

The Log2 MIC value for the identified Enterococcus species over time. The Enterococcus species were determined via MALDI-TOF. The data is presented as a boxplots with the upper and low quartiles present. Outliers are shown as individual black circles. Present species include E. casseliflavus, E. durans, E. faecalis, E. faecium, E. gallinarum, E. hirae.

Figure 7

Log2 MIC over time of Enterococcus hirae per dosing group. The high dose group is shown in black and the low dose group is shown in red. The data is presented as boxplots with the upper and low quartiles present. Outliers are shown as individual black circles.

Figure 8

Shannon Diversity Index and Bray–Curtis plot assessing beta diversity. This is a Shannon Index diversity plot comparing the diversity of the samples between the high (orange) and low dose (blue) group. The higher the value on the y axis indicates there is more diversity in the microbial community. This is a Bray–Curtis plot to assess the beta diversity, or look at the variance across all the samples. The high dose group samples are shown in orange and the low dose group samples are shown in blue. The Bray–Curtis plot measures how dissimilar the samples are.

Figure 9

Relative abundance plotting of Phylum over time per dosing group. A relative abundance plot demonstrating the change in microbial composition (at the Phylum level) over time for each individual steer per high and low dose group.