Phage Communities Respond to Antibiotics and May Spread Resistance

Antibiotics have been, and continue to be, a cornerstone of modern medicine.  Unfortunately, their widespread use has not come without cost.  The use of antibiotics has continued to result in an increased prevalence of antibiotic resistant bacteria, a problem that was even recognized by Alexander Fleming following his discovery of Penicillin [1, 2].  Presently we do have some understanding of how antibiotic resistance is propagated throughout bacterial populations, such as through mutations that prevent antibiotic efficacy, and through antibiotic driven selection for antibiotic resistant bacteria.  In addition to these mechanisms for the propagation of antibiotic resistance, there are also less well characterized means through which antibiotic resistance is spread through bacterial populations.  One of these mechanisms is through bacteriophages, which have been shown to mediate transfer of bacterial genes, especially when their hosts are stressed by antibiotics (this often occurs when prophages are induced in response to antibiotics, a process that results gene transduction).

While phages have been shown to mediate antibiotic resistance gene transfer in response to antibiotic treatment in culture-based studies, comprehensive community-wide studies (independent of culturing) have been lacking [3].  In order to better understand the complex roles phages play in the development of bacterial antibiotic resistance, a recent study conducted by Modi, et al investigated the response of mouse gut bacteriophage communities (collectively called the gut “phageome”) to antibiotic treatment [4].  This study was performed by combining mouse treatment techniques and high throughput shotgun sequencing technologies.

The researchers in this study treated mice with two common antibiotics: ciprofloxacin (a quinolone antibiotic, which means it interferes with DNA replication) and ampicillin (a beta-lactam antibiotic, meaning it interferes with bacterial cell wall synthesis).  They collected stool samples from the mice, isolated the bacteriophage DNA (see reference [5] for a detailed explanation), and performed high-throughput shotgun sequencing, meaning they collected many DNA sequences for many pieces of phage genomes.  From here they were able to perform different types of community analyses to evaluate the differences between those mice treated with antibiotics, and those without treatment.

As the title of the paper suggests, the phageomes of the antibiotic treated mice had an increased abundance of antibiotic resistance genes.  This is an important and medically relevant finding because phages are known to mediate horizontal gene transfer, and an increase in frequency of antibiotic resistance genes in the phageome could suggest a higher frequency of antibiotic resistance transfer among bacteria.  The researchers not only showed this possibility through their sequencing analysis, but they also functionally showed an increase in antibiotic resistance transfer in untreated mice when they were orally exposed to purified phages from treated mice.  By confirming their sequencing based findings with a functional proof-of-principle, this group presents compelling evidence for roles of phageomes in mediating antibiotic resistance gene transfer.

In addition to investigating antibiotic resistance genes, the group also reported a high frequency of bacterial metabolic related genes throughout the different phageomes.  They reported increases in many different types of metabolic genes in the phageomes in response to antibiotic treatment.  This is particularly interesting because, as they point out in the paper, this could promote phage mediated gene transfer of advantageous metabolic genes among certain bacteria, which could offer those bacteria a selective advantage.  I think this could be interesting in cases such as Clostridium difficile infections, which are thought to be caused by an overgrowth of C. difficile after the antibiotic mediated destruction of the competing commensal bacteria.  We could speculate that, while the destruction of the commensal flora is most likely a key factor, antibiotics may mediate advantageous gene acquisition by the C. difficile bacteria through phages, thereby promoting their overgrowth and consequent infection.  The authors also speculate that this maintenance of metabolic genes in the phageome could preserve the gene content that would otherwise be lost upon the general bacterial destruction.  This preservation could allow the genes to be transferred back to the commensal bacteria as they grow after cessation of antibiotic treatment.

The main point I took away from this paper is that mammalian phageomes likely play important roles in antibiotic stimulated increases in antibiotic resistance (through horizontal gene transfer of antibiotic resistance genes) and increases in bacterial pathogenesis (through transfer of beneficial metabolic genes to certain, potentially pathogenic, bacteria).  I think this is particularly important because, while these implications of phages in antibiotic resistance, or pathogenic gene transfer in response to antibiotics, have been studied in culture based systems, robust community-wide studies have been lacking.  I think these findings stress the importance of understanding the roles of bacteriophages when studying infections by antibiotic resistant or opportunistically pathogenic bacteria.  Finally, it will be interesting to see how these studies influence the use and development of antibiotic drugs in the future.  Perhaps the knowledge of what drugs have a greater potential for promoting phage mediated gene transfer will change what antibiotics are prescribed, or even what types of new antibiotic drug classes are pursued by pharmaceutical companies.

Works Cited
1. Obolski U, Hadany L. Implications of stress-induced genetic variation for minimizing multidrug resistance in bacteria. BMC medicine. 2012;10:89. Epub 2012/08/15. doi: 10.1186/1741-7015-10-89. PubMed PMID: 22889082; PubMed Central PMCID: PMC3482572.
2. FLEMING A. Penicillin. Nobel Lecture; 1945.
3. Varga M, Kuntova L, Pantucek R, Maslanova I, Ruzickova V, Doskar J. Efficient transfer of antibiotic resistance plasmids by transduction within methicillin-resistant Staphylococcus aureus USA300 clone. FEMS microbiology letters. 2012;332(2):146-52. Epub 2012/05/05. doi: 10.1111/j.1574-6968.2012.02589.x. PubMed PMID: 22553940.
4. Modi SR, Lee HH, Spina CS, Collins JJ. Antibiotic treatment expands the resistance reservoir and ecological network of the phage metagenome. Nature. 2013. Epub 2013/06/12. doi: 10.1038/nature12212. PubMed PMID: 23748443.
5. Thurber RV, Haynes M, Breitbart M, Wegley L, Rohwer F. Laboratory procedures to generate viral metagenomes. Nature protocols. 2009;4(4):470-83. Epub 2009/03/21. doi: 10.1038/nprot.2009.10. PubMed PMID: 19300441.