Review for "Streaming algorithms for identification of pathogens and antibiotic resistance potential from real-time MinIONTM sequencing"

Completed on 10 May 2016 by Willem Van Schaik .

License: http://creativecommons.org/licenses/by/4.0/

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Comments to author

Author response is in blue.

Here is Willem van Schaik's review of the manuscript submitted to GigaScience.

I have previously manuscript this manuscript on academickarma.com and my report and the authors' response to my previous comments can be found here http://academickarma.org/review/j2423hkrck6w

I believe this revised version is an improvement over the previous version I reviewed, but I still have a number of concerns that remain to be addressed. I remain poorly qualified to assess the bioinformatic and computational approaches and therefore focus on the interpretation of the data.

In my opinion, the section on MLST still needs some editing. Bacterial typing is a term that describes the use of methods to identify relatedness between strains. MLST is a sequence-based typing method that was first introduced in the late 1990s (http://www.ncbi.nlm.nih.gov/pubmed/9501229) and has been widely used since. It has created a vocabulary which describes high-risk 'clones' based on their sequence type (ST). However, for MLST it is essential that STs are assigned with absolute certainty and clearly this is not the case in this dataset. Based on MinION data, the STs of two of three strains cannot be resolved. I feel the only valid conclusion from these analyses can be that the method to identify sequence types may work, but that sequence quality or sequence coverage are essential and need to be higher than in the data sets that were generated as part of this study.

In the next section ('strain typing by presence of absence of genes') is confusing. The authors frequently use the word 'strain type' where they should have used 'sequence type' (e.g. in the line 'and identified their strain types using the relevant MLST schemes'), so please check carefully when strain type should be replaced with 'sequence type' or when 'strain type' means something else than 'sequence type'. The authors set up a method to classify strains on the basis of presence/absence of genes. I have expressed my concerns about this methodology in the previous version of the manuscript and the authors have responded to these concerns as follows: 'The gene presence/absence typing approach is designed to provide preliminary strain information extremely rapidly, using both 1D and 2D reads. It is primarily designed for the situation in which an exemplar strain has already been sequenced. We argue that this does have applicability, for example in an outbreak situation where it is very useful to know if a known strain is present in a new sample.' I believe this is a valid potential application but this is not well explained in the manuscript and I believe this point should be made more clearly. The limitation of the approach (i.e. that gene content does not necessarily correlate with sequence type) should also be highlighted.

Fig 1 adds little information because they key steps of the algorithms (the arrows to 'species typing, strain typing and resistance profile') are not explained in any detail. I believe that adding a schematic overview of this part of the pipeline would make this figure considerably more informative.

Minor comments:
The discussion section is long and unfocused.
Fig 3 serves little purpose in my opinion and may be better placed in the Supplementary data.
Fig 6. Panel c) text is very difficult to read.
p. 1, l. 52 Correct 'when to when to'
p. 2, l. 57: write quasipneuminiae
p. 8, line 44. Correct 'the our real-time analysis'
p. 16, line 40. Write 'an affine gap'

Are the methods appropriate to the aims of the study, are they well described, and are necessary controls included?
If not, please specify what is required in your comments to the authors.

Yes.

Are the conclusions adequately supported by the data shown?
If not, please explain in your comments to the authors.

No.

Does the manuscript adhere to the journal’s guidelines on minimum standards of reporting?
If not, please specify what is required in your comments to the authors.

Yes.

Are you able to assess all statistics in the manuscript, including the appropriateness of statistical tests used?

Yes, and I have assessed the statistics in my report.

Quality of written English

Please indicate the quality of language in the manuscript:
Acceptable.

Declaration of competing interests
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None.

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Authors' response to reviews: (https://static-content.springer.com/openpeerreview/art%3A10.1186%2Fs13742-016-0137-2/13742_2016_137_AuthorComment_V1.pdf)