Faculty Mentor: Todd Primm, Associate Professor,
Department of Biological Sciences
Student Team Members:
Jeannette Carlson, John Pinard, & Dayna Bishoff
Faculty Report
Antibiotics are powerful tools to fight infectious bacteria, but they can also disrupt the normal microbiome (bacteria associated with the host normally). While this disruption has been observed in many model systems, including our own, it has not been clearly demonstrated that the change in microbiome results in a clear negative effect for the host. Below are our results on how the host can be affected.
Method:Our fish host is exposed to the antibiotic rifampicin for three days, which changes the community composition of the skin and gut microbiome, but the overall number of bacteria is similar.
Part 1: How much and how long is the microbiome affected? By culture the percent of skin bacteria that are drug resistant is 90% after treatment, while after one week of recovery drops to 17% yet persists at that level after a second week. So the rate of resistance drops after 14 days, but resistance is not lost. Our metagenomic DNA sequencing data shows that before antibiotic the top five (58% of all sequences) genera on the skin are Massilia, Captivus, Rickettsia, Leptothrix, and Ottowia. After treatment those five are only 5.1% of sequences, and the most abundant (68% of total) genera are Variovorax, Hydrogenophaga, Mitsuaria, Pelomonas, and Ottowia. So only one of the original dominating organisms, Ottowia, persisted. Thus treatment caused a dramatic change in the microbiome. After two weeks of recovery, the five most abundant (67%) bacterial genera were Flavobacterium, Pseudomonas, Mitsuaria, Comamonas, and Variovorax. The microbiome does not recover in two weeks to the original composition, and in fact, it changes even more.
Part 2: Does the altered microbiome affect digestion and nutrient uptake? Fish were exposed to antibiotic, then fed two food pellets daily and weight measured weekly. Of nine untreated control fish, 8 of those gained weight, but with the nine treated fish, 4 gained and 5 lost weight, yet the average weight gain in those four treated fish was higher than in the eight increasing untreated fish. Our current best explanation is that the antibiotic-disrupted gut microbiome may be worse or even slightly better than the original microbiome. This suggests that there are many different possible compositions for the host, which span a broad range of negative to positive effects, and the original composition may not necessarily be the most optimal one.
Repeat experiments are underway to explore these effects.
Part 3: Does the altered microbiome protect the fish against infection as well as the original? The treated fish were given 12 hours of rest in order to eliminate the antibiotic from their tissues. Both groups of fish were infected by E. ict pathogen for 24 hours and then moved into clean APW. The average time to death for the untreated group was 102+/-56 hrs and the average for the untreated group was 98.5+/-15 hrs. The data collected in this run demonstrates that the untreated fish died at a faster rate than the untreated. This evidence suggests that a disrupted microbiome does not protect as well versus a pathogen.
Part 4: Does the altered microbiome prevent the skin from balancing salt in the water? When challenged with half of the salinity of seawater, 3 of 7 untreated fish died at an average of 22.2 hrs, while 7 of 8 treated fish died at an average of 18.9 hrs. This suggests that the disrupted microbiome has a negative effect on the fish’s ability to adjust to high sodium chloride. Perhaps osmotic balance is a newly discovered function of the microbiome.
Part 5: Are fish with an altered microbiome more sensitive to harm from a contaminated environment? The first challenge examined was adding organic-rich soil to the water, thus loading it with many different soil bacterial species. All fish, treated and untreated, survived this trial. It may be that this stress is so mild that no effects are observed with a few days. The second challenge was more severe, being suspension of feces in the water. This is a higher number of bacteria, and the bacteria are most likely to be invasive. Again, no differences between treated and untreated fish were found. So while the altered microbiome is less protective against a specific pathogen, it is similar to a large number of non-pathogenic bacteria.
Conclusion: In our system, antibiotic may affect nutrition, resistance to infection, and osmotic balance of fish. This could relate to some side effects of antibiotics in humans, such as diarrhea and repeated infections. Future studies will focus on the mechanisms of how these host effects occur.