Antibiotic Resistance: Horizontal Gene Transfer

 

Robert Lo, Ph. D, D.V.M.

 

Antimicrobial compounds, including both naturally and chemically synthesized compounds, have been one of the most important inventions to combat infections. Penicillin was the first natural antibiotic to be discovered from Penicillium fungus by Alexander Fleming in 1928. Following the discovery of penicillin, researchers started search for more antibiotics from soil microorganisms, including bacteria and fungi. Streptomycin produced by Streptomyces griseus was found in 1943. Most antibiotics in use today originated from the phylum Actinobacteria with nearly 80% of actinobacterial-derived antibiotics produced by soil-dwelling bacteria of the genus Streptomyces (Barka et al., 2016). Currently, antibiotics are classified into different groups based on their mechanism of antimicrobial activity. The main groups are: inhibit cell wall synthesis (β-lactams and glycopeptides), depolarize cell membrane (lipopeptides), inhibit protein synthesis (aminoglycosamides, chloramphenicol, lincosamides, macrolides, oxazolidinones, streptogramins, and tetracyclines), inhibit nucleic acid synthesis (Quinolones), inhibit fatty acid synthesis (platensimycin), and inhibit metabolic pathway (sulfonamides and trimethoprim).

 

With so many groups of antibiotics to treat pathogens, it seems that antibiotics would win the battles against the infection. However, the marked increase of antibiotics resistance pathogens have exceeded new drug discovery, and this resistance occurs shortly after a new drug is approved for treatment. Therefore the emergence and spread of antibiotic resistance among pathogenic bacteria has raised the concerns for public health worldwide. How did antibiotics resistance pathogens appear so quickly? More importantly, how did resistance develop?

 

Antibiotic resistance is ancient and it is expected results of the interaction of many organisms with their environment. Since most antimicrobial compounds are produced by bacteria, it is nature that those antibiotic-producing bacteria also contain self-resistance mechanisms against their own antibiotics. D’Costa et al. identified that most Streptomyces are resistant to an average of seven to eight antibiotics, including newly developed and clinically important therapeutics (D’Costa et al., 2006). In addition, it is believed that non-antibiotics producing environmental bacteria have evolved resistance mechanisms to overcome the action in order to survive via co-existence with antibiotic-producing bacteria. Researchers also identified the presence of gene-encoding resistance to β-lactams, tetracycline, and glycopeptide antibiotics in metagenome samples of 30,000-year-old permafrost (D’Costa et al., 2011). Analysis of the vancomycin resistance gene cluster in this metagenome also revealed conservation of gene sequence and synteny with modern resistance clusters in the clinic as well as protein function and structure.

 

Resistance to antibiotics occurs either by mutation or by acquisition of resistance genes via horizontal gene transfer. It is believed that horizontal gene transfer is the most important factor in the current pandemic of antimicrobial resistance. Though the mechanisms has long been existence, the rate of antibiotic resistant gene spread and the number of resistant strains has increased tremendously over the past few decades because of selective pressure through antibiotic use. The use and misuse of antibiotics in medicine, agriculture, and aquaculture has been linked to the emergence of resistant bacteria (Cabello, 2006; Economou & Gousia, 2015). In addition, the majority of consumed antibiotics are excreted unchanged, and are then introduced into the environment directly or through waste streams (Sarmah et al, 2006). As a result, waste streams become hotspots for the spread of antimicrobial resistance because of the introduction of antibiotic selection pressure into commensals and pathogens (Graham et al., 2011). These increasing environmental exposure to antibiotics has significantly increased antibiotic resistance genes of pathogenic bacteria, especially with the tetracycline resistance gene being 15 times more abundant now than in the 1970s (Knapp et al., 2010). In addition, increasing selection pressure has accelerated bacterial horizontal gene transfer process, increasing the number of resistome elements which reside mobile DNA compared to the pre-antibiotics era (Datta and Hughes, 1983).

 

Role of horizontal gene transfer in spread of antibiotic resistance genes

 

Conjugation

Conjugation is the process by which one bacterium transfers genetic material to another through direct contact. In conjugation, DNA is transferred from one bacterium to another. After the donor cell pulls itself close to the recipient via cell surface pili or adhesins, DNA is transferred between cells. Plasmid- mediated conjugation is considered the most prevalent method of horizontal gene transfer mechanisms in nature (Volkova et al., 2014). The acquisition of novel genes by plasmids through mobile genetic elements such as transposons (Babakhani and Oloomi., 2018) or integrons (Akrami et al., 2019), and their ability to replicate in a wide range of hosts, made them perfect vectors for the spread of AMR.

 

Some of known plasmids-mediated antibiotic resistance gene transfer events are the spread of carbapenemase blaCTX-M extended-spectrum β-lactamases (ESBLs), which can hydrolyze β-lactams (Ramos et al., 2013), and spread of quinolone resistance gene (Robicsek et al., 2006). Transposons are found in both gram negative bacteria, including Tn1, Tn3, Tn5, Tn9, Tn10, Tn21, Tn501, Tn903, Tn1525, Tn1721, Tn2350 and Tn3926, and gram positive bacteria, including Tn551, Tn917, Tn4001, Tn4451, and Tn4003. Resistance gene cassettes most frequently associated with class 1 integrons are streptomycin-spectinomycin resistance genes and trimethoprim resistance genes. Class 1 integrins has been found in gram negative bacteria, including Escherichia, Klebsiella, Aeromonas, Enterobacter, Providencia, Mycobacterium, Burkholderia, Alcaligenes, Campylobacter, Citrobacter, Stenotrophomonas, Acinetobacter, Pseudomonas, Salmonella, Serratia, Vibrio, and Shigella (Partridge et al., 2009; Xu et al., 2011 ), and in gram positive bacteria, including Corynebacterium, Aeromonas, Staphylococcus, Streptococcus, Enterococcus, and Brevibacterium (Nandi et al., 2004; Xu et al., 2010). Class 2 integrons have been found in Salmonella, Enterobacteriaceae, Enterococcus, Acinetobacter, and Pseudomonas (Machado et al., 2008; Xu et al., 2010; Xu et al., 2011). Class 3 integrons have been reported in Escherichia, Citrobacter, Klebsiella, Salmonella, Pseudomonas, Acinetobacter, Alcaligenes, and Serratia (Rowe-Magnus et al., 2001; Ploy et al., 2003). Class 4 integrons have been linked to Pseudomonas, Vibrionaceae, Xanthomonas, Shewanella, and other proteobacteria (Clark et al., 2000; Rowe-Magnus et al., 2001).

 

Transformation

In transformation, a bacterium takes in DNA from its environment, often DNA that has been shed by other bacteria. If the DNA is in the form of a circular DNA called a plasmid, it can be copied in the receiving cell and passed on to its descendants.

 

This direct genetic exchange was first identified between different strains of Streptococcus pneumonia in1928 (Griffith F., 1928). In 1951 Hotchkiss demonstrated penicillin and streptomycin resistance genes were induced into previously sensitive strains of S. pneumonia from resistant strains via transformation (Hotchkiss, 1951).Other antibiotic resistance genes, such as streptomycin, rifampicin, erythromycin, nalidixic acid, and kanamycin have been in vitro transformed into different bacteria (Harford and Mergeay., 1973; M Kristensen et al., 2012; Prudhomme et al., 2006).

 

 

Transduction

In transduction, bacteriophages, viruses that infect bacteria, move short pieces of chromosomal DNA from one bacterium to another “by accident.”

Transduction is believed to play a major role in evolution of resistance in S. aureus (Haaber et al., 2017). The transfer of antibiotic resistance genes by bacteriophages has also been reported in various bacterial species: transfer of erythromycin between strains of Staphylococcus pyogenes (Hyder and Streitfeld., 1978), the transfer of tetracycline and gentamicin resistance between enterococci (Mazaheri Nezhad Fard et al., 2011), or the transfer of antibiotics resistance plasmids in Methicillin-resistant Staphylococcus aureus (Varga et al., 2012)

 

Conclusion

Antibiotic resistance genes have been existed in antibiotic producing bacteria and non-pathogenic environmental bacteria for a long time (Barlow and Hall., 2002). These resistance genes may be transferred into clinical bacteria via horizontal gene transfer mechanisms, and the transfer rate of these events have been rapidly increased since the widespread use of antibiotics in clinical treatment (Barlow M., 2009). Horizontal gene transfer mechanisms, including conjugation, transformation and transduction, are the main mechanisms for transfer of antibiotic resistance genes between bacterial populations (Fig 1).

Figure 1. Involved mechanisms in horizontal gene transfer. Transduction, conjugation, and transformation are the main mechanisms by which bacterial species can mobilize and share genetic material with both related and non-related species. These mechanisms imply a pathway for the evolution of bacteria in different environments, allowing them to survive in their niches. A clear example of this is the acquisition of antibiotic resistance mechanisms, virulent traits, and other resources used by the microorganism to guarantee its survival. (Bello-López et al.; 2019).

 

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