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  1. This work has been published in GigaByte Journal under a CC-BY 4.0 license (, and has published the reviews under the same license. These are as follows.

    **Reviewer 1. Zhizhi Wang **

    Is there sufficient data validation and statistical analyses of data quality? No.

    Comments and Suggestions for Authors: In this manuscript, Schultz et al reported the sex-biased transcriptomes of E. suzannae, whose reproductive system was manipulated by the bacterial endosymbiont, Cardinium. Despite de novo assembling, this paper also aimed to annotate sex determination genes and venom proteins to better understand the biology of E. suzannae. However, there are several issues with this manuscript:

    General comments It seems that the theme of the background is about bacterial endosymbiont and its function in host reproduction while the data annotation is not so relevant. Of importance, there is also no further information, e.g. gene lists, about the annotated sex determination genes and venom proteins in the manuscript. The sex determination systems in hymenopteran insects are diverse and complex, while the mechanism of two species, Apis mellifera and Nasonia vitripennis, are well characterized. Authors should also point out if there are sex determination homologies shared between E. suzannae and these two model species, such as csd and wom. For the annotation of venom protein, authors should note that the predicted venom protein is not reliable without venom gland transcriptome or venom proteome data.

    As stated, the transcriptome data have been published elsewhere focusing on the expression profile of Cardinium, it would be interesting to show potential endosymbiont response genes or pathways in E. suzannae.

    My other concern is that the number of E. suzannae coding sequences is twice as many as that of E. formosa, which leads me to doubt the purity of the assembled transcriptome. The authors use a mapping-and-removal approach to filter contaminant reads from several endosymbionts of E. suzannae and its host Bemisia tabaci, while one could not exclude the possibility that other foreign contaminants could present in the raw data. Instead, foreign contaminants can be detected—and optionally removed—using a short-read taxonomic classifier by software.

    Minor: Line 41 their use in biological control of... Line 50 B. tabaci Line 51 …or other aphelinid parasitoids… Line 54 ... they can directly damage plants by feeding… Line 84 Line 122 …mapped to the male… Line 187….the public version….

    Major Revisions.

    Re-review: The authors have addressed most of my concerns, so I would like to recommend the paper for publication.

    Reviewer 2. Shaoli Wang

    Is there sufficient data validation and statistical analyses of data quality?

    Schultz et al., used previous available RNA-seq data from E. suzannae cultures as data source. They filter possible sequence reads of symbionts and host whitefly by the method of mapping-and-removal approach to eliminate known contaminants, and assembly the rest reads to unigenes as insect E. Suzannae transcriptome data source, further annotate these unigenes. I have no more constructive comments to this paper, just some suggestions below to improve this MS, even though the authors wrote the MS very carefully.

    1. Line 33, it would be better to add some words about other results from annotation and transcriptome comparison parts, after the sentence “Benchmarking Single-Copy Orthologs (BUSCO) results indicate both assemblies are highly complete”.
    2. Line 123, “Reads that did not map to any of these bacterial genomes with greater than 94% identity……”~ After gradient screening to get this 94% identity? If yes, add some words to explain why select this value. Same question as the 97% in line 124.
    3. Line 194, Summarize a table to show annotation results about sex determination and venom proteins related genes.
    4. Line 246, “Quality control and data validation” should be moved into the front of the annotation part.
    5. The format of references cited in the MS are different, which should be revised according to the requirement of the journal.

    Even though the data presented are not very informative, there is no any obvious flaw throughout the MS.