Sub-Genotyping of Acanthamoeba T4 Complex: Experience from North India

Round 1

Reviewer 1 Report

This manuscript reports the use of 16S ribosomal mitochondrial sequences to classify Acanthamoeba within the T4 genotype with respect to the tendency to produce AK. The main finding was that two strains that grouped with the previously identified subtype Acanthamoeba mauritaniensis were from AK patients. This would be the first indication that A. mauritaniensis has a pathogenic potential for humans. The paper is generally clear and well written.

The last sentence of the abstract is a little unclear and maybe better rewritten as “16S sequences from two isolated from patients of Acanthamoeba keratitis were found to group with A. mauritaniensis 22 (T4d). This group was believed to be a non-pathogenic environmental Acanthamoeba.”  Caution should be exercised here as it is not yet safe to assume that these two cases (AC1 and AC25) are actually equivalent to A. mauritaniensis as only one gene has been investigated. Also, although the authors described the morphological details of their isolated they did not tabulate them in detail sufficiently for them to be compared with known A. mauritaniensis isolates. Because of this the authors should make it clear that these two new isolates may not be equivalent to A. mauritaniensis despite the similarity apparent in this single gene. The conclusions paragraph should also reflect this point.

Author Response

We are grateful to the reviewer for this insight. The last sentence of abstract has been revised as suggested (lines 22-25). The fact that only single gene was used and whole genome sequencing should be used ideally is already acknowledged in the limitations of the study (line 210-221), however as per the reviewer’s suggestion and for better understanding of the reader, we have added this caution in lines 202-203 of text and lines 227-228 of conclusion as well.

Reviewer 2 Report

Reviewer comments

The manuscript pathogens-2134056 represents an interesting starting point to improve the T4 comprehension. After minor changes I recommend its publication.

Table 1: It would be interesting to include the GenBank sequence homologous to each sample.

Lines 81-82: what do the authors mean with “improvised”?

Line 88 and Figure 1: the authors should include the details of the microscope

Line 137: Acanthamoeba should be in italics

Author Response

Please see the attachment.

Author Response File: Author Response.pdf