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Current culture methods to investigate changes in  Escherichia coli  community structure are often slow and laborious. Genes such as  gnd  (6-phosphogluconate dehydrogenase) have a highly variable nucleotide sequence and may provide a target for  E .  coli  microbiome analysis using culture-independent methods. Metabarcoded PCR primers were used to generate separate libraries from calf faecal samples for high throughput sequencing. Although a total of 348 separate  gnd  sequence types (gSTs) were identified, 188 were likely to be due to sequencing errors. Of the remaining 160 gSTs, 92 did not match those in a database of 319 separate  gnd  sequences. ‘Animal’ was the main determinant of  E .  coli  diversity with limited impact of sample type or DNA extraction method on intra-host  E .  coli  community variation from faeces and recto-anal mucosal swab samples. This culture-independent study has addressed the difficulties of quantifying bacterial intra-species diversity and revealed that, whilst individual animals may harbour >50 separate  E .  coli  strains, communities are dominated by <10 strains alongside a large pool of subdominant strains present at low abundances. This method will be useful for characterising the diversity and population structure of  E .  coli  in experimental studies designed to assess the impact of interventions on the gut microbiome.

Culture independent analysis using gnd as a target gene to assess Escherichia coli diversity and community structure