Researchers map antimicrobial resistance genes in sewage

Researchers from Denmark and elsewhere have been sifting through the world’s sewers for genetic material that can explain how bacteria transfer antimicrobial resistance.

They have identified a large quantity of resistance-carrying circular pieces of DNA in sewage that have the potential to transfer antimicrobial resistance from one bacterium to the next.

The discovery paves the way for greater understanding of how antimicrobial resistance is transferred between bacteria and where it can potentially occur. The information from mapping the total genetic material in sewage from 22 countries on five continents can also be applied in biotechnology and the other life sciences in general.

“This provides insight into the antimicrobial resistance genes with the potential to be transferred between bacteria. This is basic biology that can have various long-term practical implications for monitoring antimicrobial resistance but also for using the bacteria’s genetic material biotechnologically or in other ways,” explains a researcher behind the new study, Sünje Johanna Pamp, Associate Professor, Novo Nordisk Foundation Center for Biosustainability and National Food Institute, Technical University of Denmark, Kongens Lyngby.

The research, published in mSystems, was carried out by Sünje Johanna Pamp and her colleagues at the National Food Institute, Technical University of Denmark.

Plasmids can transfer antimicrobial resistance between bacteria

Antimicrobial resistance is a growing problem that threatens global health because antibiotic-resistant bacteria can make otherwise relatively harmless infections deadly.

Bacteria themselves can become antibiotic resistant when they encounter antibiotics, but bacteria can also transfer antimicrobial resistance to each other by using small circular pieces of DNA called plasmids.

Plasmids may contain genes for antimicrobial resistance, so if one bacterium has become resistant to an antibiotic and has the gene for resistance in a plasmid, the resistance can be transferred to another bacterium, which picks up the plasmid directly from the environment or through contact with the resistant bacteria. The recipient bacterium can then incorporate it into its own genetic material or just keep and use the plasmid it has taken up.

Virulence factors are proteins that can make people sick, and these and the bacteria’s own types of antibiotics used to combat other bacteria are also present in the genetic codes of plasmids.

Sünje Johanna Pamp and colleagues investigated the presence of antibiotic-resistant plasmids in domestic sewage, since this may offer researchers greater insight into the potential for resistance developing in humans.

“We wanted to answer the very basic question of where to find resistance genes. From a public health perspective, this is a very important question for slowing down the galloping development of antimicrobial resistance,” says Sünje Johanna Pamp, who explains that there have been several attempts to map where most of the world’s problematic genes for antimicrobial resistance are located, but this has not yet succeeded.

“In a previous study, we looked for resistance genes in samples from sewers, including in New York, Copenhagen and 77 other cities in 60 countries, but we did not examine whether the resistance genes were in plasmids or in other types of genetic material. We just identified that they existed,” explains Sünje Johanna Pamp.

Exploring sewage worldwide

In the new study, the researchers examined untreated sewage samples in 22 countries on five continents.

Since sewage contains very few bacteria and a lot of water, the researchers filtered out all the solid material and then started looking for the plasmids.

The researchers used nanopore sequencing, which enabled them to sequence entire plasmids at once. Researchers usually sequence DNA in small chunks at a time, but for plasmids, this is a problem because they have many repeating DNA sequences that can be difficult to assemble as a jigsaw puzzle afterwards.

“This has never been done before, and the technique is quite groundbreaking. We isolated the plasmids, sequenced them and then used bioinformatics to examine their DNA and determine what they encoded and whether we could identify genes for antimicrobial resistance,” says Sünje Johanna Pamp.

Resistance genes in plasmids differ in Asia and Europe

The study resulted in 150,000 gene sequences, 60,000 of which had known plasmid-related genes. The rest are pieces of DNA that are unidentified so far.

In the 60,000 plasmids, the researchers found many genes for antimicrobial resistance but also identified large geographical differences in the samples.

For example, the samples from Asia had more similar genetic patterns than the samples from Europe.

The researchers also examined whether the resistance genes in the plasmids matched those from previous studies in all the genetic material in the samples – that is, how many of the resistance genes were present in the plasmids and in the bacteria’s chromosomes.

“We identified resistance genes from chromosomes and from plasmids. This is interesting because resistance genes from plasmids more easily transfer between bacteria. We should pay extra attention to these genes in the future,” explains Sünje Johanna Pamp.

Biotechnologists hunt for plasmids

Sünje Johanna Pamp explains that the study has practical implications in monitoring the development of antimicrobial resistance but also contains considerable basic scientific knowledge that may become valuable in the future.

Researchers can search the study material for novel plasmids and study their functions in more detail.

Plasmids enable bacteria to transfer antimicrobial resistance genes but are also used in biotechnology, including in cloning.

“This is basic biology, and we can learn from this and ultimately find more practical implications in biotechnology and the life sciences,” concludes Sünje Johanna Pamp.