Jo Handelsman likes to follow the bugs—especially those bacteria that might hide new forms of antibiotic resistance within. While the antibiotic resistance that shows up in clinical setting gets scads of attention from researchers, Handelsman’s team at Yale University in New Haven, Connecticut wanted to investigate how new antibiotic resistance (AR) genes might make their way into human pathogens. In particular, in a study published this week in mBio®, they investigated an altogether different form of the farm-to-table movement.
“We were interested in the role of manure,” commonly used as a soil fertilizer in agriculture, says Handelsman, who is also an HHMI Professor. She paints a scenario where manure used on some vegetable crops contains bacteria from the cow’s gut, which could cling to root crops especially, such as carrots. The manure bacteria might then move from the produce to people, or they might transfer AR genes to bacteria that colonize humans.
“Is this a route for movement of these genes from the barn to the table?” she asks. “This was the first step, looking at the presence of antibiotic resistance genes in the manure.”
Using a powerful functional metagenomics approach, her team surveyed five manure samples from four dairy cows housed at the University of Connecticut. From these samples, the researchers extracted all of the bacterial DNA present, chopped it into bits, and put each bit back into a laboratory strain of Escherichia coli bacteria.
The E. coli were then grown on plates containing four different classes of antibiotics: beta-lactams (like penicillin), aminoglycosides (like kanamycin), tetracycline, and chloramphenicol. Any E. coli that could grow on the antibiotics likely contained an AR gene isolated from the manure bacteria. The team then used third-generation sequencing to pull out the gene and its flanking DNA to identify both the gene and the likely species of gut bacteria that originally hosted it.
They identified 80 distinct and functional AR genes, 75% of which were novel—genes so divergent in sequence identity from known AR genes that some might not have been found through sequencing efforts alone.
“That was exactly the motivation behind the functional process, to deepen the pool of known genes with AR function,” says Handelsman. Also, by expressing the genes in E. coli, the team selected for genes that truly confer resistance in a species that can be pathogenic and associated with farm products.
The deep divergence from known AR genes surprised Handelsman. “From an evolutionary perspective, these are distant from the ones we see in the database, which largely represent the AR genes seen in the clinic.” That signals that cow manure bacteria may not be a major source of resistance that currently causes problems for human patients. However, as lead author Fabienne Wichmann points out, it could also mean that “cow manure harbors an unprecedented reservoir of AR genes that we should know about.”
In other words, these new AR genes could be the next ones to show up in bacteria infecting humans. There are two ways that could happen—bacteria containing these AR genes colonize humans, or through horizontal gene transfer, the genes jump to other bacteria that already colonize humans. Benign manure bacteria might transfer AR genes to pathogens at any point—in manure, soil, food, or humans.
Expanding our repertoire of known resistance-conferring genes, will help clinicians better predict which kinds of antibiotic resistance they might face in the future, says Wichmann. Studies like this will also give drug developers better information for designing pre-emptive strike inhibitors to AR genes.
This is just the first in a sequence of studies, says Handelsman. “We’re hoping this study will open up a larger field of surveillance, to start looking at new types of resistance before they show up in the clinic.”