Assessment of Bilge Water Degradation by Isolated Citrobacter sp. and Two Indigenous Strains and Identification of Organic Content by GC-MS

Round 1

Reviewer 1 Report

The main issue of the reviewed manuscript was isolation of the specified bacterial strains and their characterization in terms of organic contamination from the bilge water. The topic over bilge water management obviously is not novel, by Scopus search with phrases bilage AND water AND treatment 185 outputs have been obtained. However, by expanding search with biological treatment the number of outputs was reduced to 16 publications, basically from the same scientific group, which prepared this reviewed article. Nevertheless, manuscript is well prepared, organized and written.

In general, I am satisfied with the overall manuscript quality but I got some major comments and suggestions that have to be considered before final acceptance.

Abstract

1). Last sentence of the abstract is not clear. Mentioned compounds are metabolites from the decomposition of the organics from the raw bilge water or, they couldn’t be detected in raw sample due to their coverage by other more abundant organic compounds? The same issue has to be addressed in discussion section.

Introduction

2) In general, introduction section is in general well written and provide all basic information. However, most commonly find bacterial genera in the system of bilge water treatment or similar have to be presented in this section. As well biochemical mechanisms responsible for the organics degradation in such systems need to be introduced – whether these substrates are used as an energy source or just detoxification of the environment is performed to reduce harmful effects on microbial growth? This issue is also missing in the discussion section.

Materials & methods

3) In the section about real bilge water the authors mentioned that samples were collected. Thus, please provide information about how many samples were indeed analyzed during this study. If few what concentrations are presented in Table S1 and S2- averages? Where are the standard deviations?

4) Please provide more information about samples of soil and BW applied for the microorganisms isolations, preferably in the supplementary data. How many samples, localizations on map etc.

5) Please be precise using microbial terminology, for instance line 135 - resulting alignment of the 16S rRNA was compared for homology in the NCBI database by BLASTn nucleotide tool analysis

You are not analysing 16S rRNA but you are analyzing DNA sequences of the gene encoding 16S rRNA

6) 16S rRNA comprehension of homology between experimental samples and DNA sequences deposited in NCBI database is just preliminary step. In fact experimental DNA sequences and marker DNA sequences collected from the database, should be aligned together and construction of the genetic distance tree has to be done. Without this step bacteria identification is not reliable. You can check following paper for methodology in that aspect: https://doi.org/10.3390/en14175225.

Results and Discussion

This chapters are logically organized and supported by graphs and tables.

7)However, in particular Table 2 requires uniformed in terms of the editorial aspects with Table 1, as well figure 5 requires improvements of the axis’s units and descriptions readability.

Please also add information to the discussion about issues pointed to the comments for the abstract and introduction.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

Several studies reported the isolation of indigenous species from bilge water and the degradation of petroleum hydrocarbons by species isolated from oil-contaminated substrates. The manuscript lacks the novelty and can not be recommended for publication in water.

Author Response

We would like to thank the Reviewer for his/her time and reply. However, we would like to point out that in the present study (as already reported in the manuscript) we have emphasized the following points that were not reported by other studies:

• There is scarce information regarding the biodegradation of raw real BW by indigenous and non-indigenous species, as well as the specific organic compounds that are removed due to their action.

•  This is the first study that pointed out that Citrobacter species could biodegrade bilge water. In addition, it shows the biodegradation potential of two indigenous species (Halomonas and Exiguobacterium sp). Furthermore, the performance of those strains in bilge water was investigated using GC-MS analysis before and after biodegradation.

Reviewer 3 Report

The manuscript entitled “Assessment of bilge water degradation by isolated Citrobacter sp. and two indigenous strains and identification of organic content by GC-MS” presents an interesting research related to the use of the isolating microorganisms to cleaning the bilge water accumulating on on the hull at the bottom of a vessel. Effective treatment of these waters will reduce the level of pollution of sea waters by sewage from ships.

In my opinion, the abstract should be more condensed and should present the most important outcomes and highlight the novelty. Classical test methods were used for chemical analyzes, please indicate the novelty in this regard. In my opinion the Abstract should be revised and modified.

The introduction should be enhanced by referring to recent studies covering the impact of bilge water on sea degradation. It is not clear what is the hypothesis of performed study.

The “Materials and Methods” section should be clarified by adding the precise description and analysis of the test results.

The Discussion is very weak. Hence, I suggest to enhance this chapter and refer to some more articles to compare main findings of the study and to formulate conclusions valid in a broader scale. The novelty and originality of performed research should be also strengthened.

The conclusions should also include general statements regarding the practical use of research results and the importance of reducing the degradation of the marine environment. The literature review seems to be limited. I suggest authors to elaborate more on the topic and refer to many more scientific research related to the issue.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The authors have addressed all of my comments with appropriate way, thus I fully support current manuscript version for publication in Water.

Reviewer 2 Report

I found substantial improvements in manuscript. Authors revised the introduction to highlight the novelty and significance. Sec. 2 and 3 are also improved. I'll recommend this article for publication in Water.

Reviewer 3 Report

The Authors introduced significant corrections and additions to the article, which improved its quality and content. The literature review and References have also been supplemented. Minor additions may still be made to the chapter 3 Results and Discussion, but I recommend that the article can be accepted for publication in its current form. Good luck!