How to identify a bacterium using TrueBac TM ID

Bacterial taxonomy now fully embraces genomics. Bacterial species can now be objectively and precisely defined using a comparison of whole genome sequences. Bacterial identification at the species level can, in theory, be always accurate if genome sequences are used. In fact, it has been shown that the substantial portion of clinical isolates are either misidentified or belong to hitherto undescribed/unrecognized species, named genomospecies when genome sequencing is used for the identification at the species level(Roach et al., 2015). In routine microbiology laboratories where bacterial identification is carried out on a regular basis, the scheme described in the proposed minimal standard for bacterial taxonomy (Chun et al. 2018) can be applied to ensure accurate identification using a combination of 16S rRNA gene (16S) and whole genome sequences.

16S and whole genome sequences for use in bacterial identification

The use of 16S and whole genome sequences for the purpose of bacterial identification has its pros and cons:

Both 16S and whole genome sequencing are valuable tools in routine microbiology laboratories. In most cases, the genome sequence contains at least one 16S sequence, so specific 16S amplicon sequencing is not necessary. Here are three possible scenarios for routine bacterial identification by sequencing:

1. Whole genome sequencing only

Bacterial isolates are subjected to whole genome sequencing using one of available NGS platforms. Assemble raw data to generate contigs and upload the resulting genome assembly to EzBioCloud’s TrueBac ID-Genome for the definitive identification. TrueBac ID-Genome also provides the search result of the 16S sequence included in the genome assembly. If your isolate exhibits the average nucleotide identity (ANI) value of ≥95% to a known species, it is considered successfully identified. Otherwise, it is likely a member of a hitherto unknown novel species. Finding a novel species in either environmental or clinical laboratories is not a rare event.

2. 16S sequencing only

The 16S sequence is determined from an isolate using a Sanger sequencing instrument (e.g., ABI 3700). The final 16S sequence should be edited manually to ensure the highest quality possible. Upload the sequence into EzBioCloud’s TrueBac ID-16S service. If 16S of the isolate shows <98.7% sequence similarity to all known species, your isolate likely belongs to a novel species. However, the opposite is not always true. Many closely related species show 16S similarity of ≥98.7%, even identical 16S sequences in some cases. If you want to achieve an accurate and definitive identification at the species level, you should use whole genome sequencing instead of 16S sequencing.

3. 16S sequencing, and then whole genome sequencing

To reduce cost (but not time), 16S amplicon sequencing can be performed first to see if the isolate belongs to a novel species.  Otherwise, the whole genome is then sequenced. This strategy was proposed by Chun et al. (2018). However, this sequential approach does require extra time.

The EzBioCloud team / Last edited on May 5, 2018