Bacterial Genetics and Genomics book Discussion Topic: Chapter 17, question 13
The publications of the first bacterial genome sequences were 25 years ago. The technology has come a long way since then, both in the lab and computationally. One of the first bacterial genome sequencing projects started was one undertaken to sequence the complete Escherichia coli genome. Ultimately, it was completed and published in 1997. To-date, this publication has been cited by 2,469 other articles, including a recent investigation into E. coli that are present on the surface of the human eye (Ranjith et al., 2020).
E. coli are predominantly found in the intestinal tract of humans and animals, but they are also present elsewhere, including on the ocular surface. There are, in fact, many bacterial species that reside as commensal, non-pathogenic, bacteria on the surface of the eye. These bacteria can cause opportunistic ocular infections when there has been trauma to the eye or due to other issues that compromise the immune system. To understand more about this type of E. coli, 10 eye isolates were genome sequenced. The genome sequences were analyzed for SNP variation to find single nucleotide polymorphisms between the sequences. The isolates were sorted into the nearest E. coli phylogenetic group and pathotype, as well as being assessed for antimicrobial resistance genes, prophages, and other factors that might be involved in pathogenicity.
The E. coli isolates came from two cases of conjunctivitis, two cases of bacterial keratitis, five cases of endophthalmitis, and one from orbital cellulitis. DNA was extracted from overnight cultures of the E. coli using a QIAGEN DNA isolation kit and genome sequenced using an Illumina HiSeq. The data was mapped against the reference genome sequence E. coli strain K-12 substrain MG1655 using BWA. The de novo assemblies used Velvet.
From this study, antimicrobial resistance genes were identified, which correlated with antimicrobial investigations in the laboratory. Five out of the 10 isolates were resistant to more than three classes of antibiotics. Presence of plasmids, prophages, and virulence genes were also identified, including some that may be characteristic of ocular isolates.
The different presence and absence of virulence genes, resistance genes, prophages, and plasmids were examined using BRIG, the BLAST Ring Image Generator, an example of which is described in Chapter 17 of Bacterial Genetics and Genomics and shown in Chapter 17 Figure 12 (shown below).
The 10 isolates were also able to be categorized into three of the seven known phylogenetic groups: A (2 isolates); B2 (7 isolates): and C (1 isolate). SNP analysis agreed with these relationships. Additional analysis associated the ocular isolates with four of the eight pathotypes, grouping six isolates with ExPEC (extra-intenstinal pathogenic E. coli), two with EPEC (enteropathogenic E. coli), on wit ETEC (enterotoxigenic E. coli), and one with UPEC (uropathogenic E. coli)strains. There was therefore a lack of concordance between the phylogenetic groups (A, B2, and C) and the pathotypes (ExPEC, EPEC, ETEC, and UPEC).
This study, and the over 2,000 others that have cited the original E. coli genome sequence paper, as well as many others, have shown the power of genome sequence data in revealing both differences and similarities between bacterial strains. There is tremendous diversity in the microbial world. The more we sequence, the more that is apparent. We have come a long way in 25 years. Not only are many genome sequence papers published currently that include more than one genome, but such studies are able to comparatively analyze the data, identify features that we did not know were present when investigations first started, generate the data and analyze it in a fraction of the time, and do so using much less starting material than previously required, including generating sequence data without first culturing bacteria in the lab.