Candida auris Updates: Outbreak Evaluation through Molecular Assays and Antifungal Stewardship—A Narrative Review

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The authors reviewed molecular methods detections and antifungal stewardship concerning Candida auris. The authors have addressed an important topic and their review covers the recent developments concerning C. auris. The text is good but there are a number of smaller items which have to be improved.

The conclusion is too unspecific and recommendations for the identification of C. auris infections optimal in clinical settings, dealing with C. auris infections, and future researches foci should be given.

Set all genus and species names in italics

Line 49: 28S (capital S)

Lines 69ff: this sentence needs rewording

Line 73: what does this mean? Please be more specific

Line 127: the new fungus

Line 199: wrong wording. Did you mean underlining?

There are a number of wrong wording and grammar errors, which need correction. Smoothing of the wording is desirable.

Comments on the Quality of English Language

a complete overhaul of the text is necessary

Author Response

Point-by-point response to the reviewers’ and editor’s comments.

 

Title: “Candida auris updates: molecular methods detections, antifungal stewardship”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #1:

Comments and Suggestions for Authors

 

The authors reviewed molecular methods detections and antifungal stewardship concerning Candida auris. The authors have addressed an important topic, and their review covers the recent developments concerning C. auris. The text is good but there are a number of smaller items which have to be improved.

The conclusion is too unspecific and recommendations for the identification of C. auris infections optimal in clinical settings, dealing with C. auris infections, and future research foci should begiven.

 

• Response to Reviewer: We thank the reviewer for this comment. We modified the conclusion as suggested (lines 320 – 332).

 

Candida auris was involved in more than twenty outbreaks, happening in all continents, in the last years. Given the fact that is no specific prevention available, fast detection of this pathogen in patients sample, without prior DNA extraction, can prove to be a salutary step in the management of these cases. In this way, by ensuring fast detection, safety procedures for limiting the spread of C. auris in the hospital environment could be implemented. From all the identified assays (e.g., WGS, MALDI-TOF, PCR) to diagnose C. auris outbreaks, the most appropriate for routine diagnosis is, from our point of view, Real Time PCR, due to its detection speed, possibility of also identifying resistance genes and readily-available laboratory equipment. The commercially available kits should be validated by each laboratory, before implementing them in routine diagnosis. Moreover, studies that already identified the resistance mechanisms to antifungals used for the treatment of these infections will favor stewardship strategies, contributing to prevent future resistance emergence.

 

 

 

Set all genus and species names in italics.

• Response to Reviewer: We thank the reviewer for this comment. We wrote all taxonomic names in italics.

Line 49: 28S (capital S)

• Response to Reviewer: We thank the reviewer for this comment. We modified as suggested.

Lines 69ff: this sentence needs rewording.

• Response to Reviewer: We thank the reviewer for this comment. We modified as suggested.

Line 73: what does this mean? Please be more specific.

• Response to Reviewer: We thank the reviewer for this comment. We modified as suggested.

” Worldwide guidelines elaborated to control C. auris outbreaks”

Line 127: the new fungus

• Response to Reviewer: We thank the reviewer for this comment. We modified as suggested.

Line 199: wrong wording. Did you mean underlining?

• Response to Reviewer: We thank the reviewer for this comment. We modified as suggested.

 

There are a number of wrong wording and grammar errors, which need correction. Smoothing of the wording is desirable.

 

Comments on the Quality of English Language: a complete overhaul of the text is necessary.

• Response to Reviewer: We thank the reviewer for this comment. In case of acceptance, we will ask MDPI for an official English Article Editing.

 

 

Reviewer 2 Report

Comments and Suggestions for Authors

The subject is relevant, although several reviews on Candida auris have been published in the last two years, and most of them have covered the aspects included in the manuscript. Despite this, a review manuscript gathering the latest papers dealing with species identification is valuable.

The first section reviews facts since the first report of this fungal species to the latest. I appreciate this, but including only some facts makes this part biased. There are no specific criteria to only include these relevant papers among a bunch of manuscripts that are as relevant as the included ones.

The search in Pubmed seems to be even more biased, what were the keywords introduced? on which date? I searched for "candida auris molecular diagnosis" and I had 54 results, different from the authors (https://pubmed.ncbi.nlm.nih.gov/?term=candida+auris+molecular+diagnosis&sort=date). There are no inclusion/exclusion criteria for the selected papers, making this part of the revision confusing.

The part dealing with antifungal drugs is even more messy. It is not clear whether a similar approach for diagnosis was used here or arbitrary manuscripts were included in this part. This aspect has been thoroughly addressed in recent years and certainly, this part of the manuscript is far from being a comprehensive review on this subject. Moreover, it seems the authors are minimizing the phenomenon to the presence or expression of a small group of genes to explain antifungal drug resistance in Candida auris, and this trait is more complex than that. My suggestion is to delete this part and focus only on diagnosis.

As a minor note, gene names and taxonomical terms should be in italics.

 

Comments on the Quality of English Language

There are typos and misspellings throughout the whole manuscript.

Author Response

Point-by-point response to the reviewers’ and editor’s comments

 

Title: “Candida auris updates: molecular methods detections, antifungal stewardship”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #2:

 

Comments and Suggestions for Authors

• The subject is relevant, although several reviews on Candida auris have been published in the last two years, and most of them have covered the aspects included in the manuscript. Despite this, a review manuscript gathering the latest papers dealing with species identification is valuable.
• Response to Reviewer: We thank the reviewer for this comment.

 

• The first section reviews facts since the first report of this fungal species to the latest. I appreciate this, but including only some facts makes this part biased. There are no specific criteria to only include these relevant papers among a bunch of manuscripts that are as relevant as the included ones.

 

• Response to Reviewer: We thank the reviewer for this comment. We have presented the latest important publications (e.g., Lancet), to certify the clinical importance of our review. By presenting the guidelines released in all the continents, we just proved that Candida auris is really an important medical issue and need to be furthered kept under close observation.

 

 

• The search in Pubmed seems to be even more biased, what were the keywords introduced? on which date? I searched for "candida auris molecular diagnosis" and I had 54 results, different from the authors

(https://pubmed.ncbi.nlm.nih.gov/?term=candida+auris+molecular+diagnosis&sort=date). There are no inclusion/exclusion criteria for the selected papers, making this part of the revision confusing.

• Response to Reviewer: We thank the reviewer for this comment. We have specified the moment of our PubMed search for the last 5 years, and we also specified the inclusion and exclusion criteria. The used key words were ”Candida auris outbreak molecular detection”, which were specified in the abstract and in the manuscript also.

https://pubmed.ncbi.nlm.nih.gov/?term=Candida+auris+outbreak+molecular+detection&filter=datesearch.y_5&sort=pubdate&size=200

Since February, 2 other papers were published and we introduced them in our manuscript, in Table 1.

 

Our search initiated in 12.02.2024, in PubMed Database, using “Candida auris; outbreak; molecular detection” key words, revealed 34 results for the last 5 years. We included in our analysis (Table 1) only 25 original research papers [14-38]. We excluded review papers. Since our last search, two more papers were published, and out Table now contains 25 papers.

 

 

The final inclusion / exclusion criteria are:

 

Our search initiated in 12.02.2024, in PubMed Database, using “Candida auris; outbreak; molecular detection” key words, revealed 34 results for the last 5 years. We included in our analysis (Table 1) only 25 original research papers [14-38]. We excluded review papers. (lines 152 – 154),

 

 

• The part dealing with antifungal drugs is even more messy. It is not clear whether a similar approach for diagnosis was used here, or arbitrary manuscripts were included in this part. This aspect has been thoroughly addressed in recent years and certainly, this part of the manuscript is far from being a comprehensive review on this subject. Moreover, it seems the authors are minimizing the phenomenon to the presence or expression of a small group of genes to explain antifungal drug resistance in Candida auris, and this trait is more complex than that. My suggestion is to delete this part and focus only on diagnosis.
• Response to Reviewer: We thank the reviewer for this comment. Antifungal drug resistance was not our central aim of this review. But considering that some of the studies that we included in Table 1 describe molecular methods that are capable of detecting genes which are responsible for antifungal resistance, this led us to prepare a paragraph about this topic, without it being related with our main search about molecular methods.  Of course, this could be a separate topic for a new paper. But we find it important to include some of this information in our review. For us, it is useful to disseminate the information that the most frequent mutations responsible for antifungal resistance can be detected by Real Time PCR assays, the method which we consider the most readily accessible and with fast and accurate results, when implemented properly.

 

 

• As a minor note, gene names and taxonomical terms should be in italics.
• Response to Reviewer: We thank the reviewer for this comment. We have modified as suggested.

 

 

Comments on the Quality of English Language

There are typos and misspellings throughout the whole manuscript.

• Response to Reviewer: We thank the reviewer for this comment. In case of acceptance, we will kindly ask MDPI for an official English Article Editing.

 

 

 

 

 

Reviewer 3 Report

Comments and Suggestions for Authors

This manuscript describes the latest knowledge on detection methods and antifungal stewardship of C. auris.  This seems to be interesting and provide useful information to readers, but contains several concerns as follows.

1. Title should be reconsidered. Is "molecular methods detection" correct? 

2. There have been many review papers on C. auris to date. In this manuscript, what point did authors focus? Authors should write objectives of this review in Introduction.

3. In Abstract, there is explanation of new nomenclature of some fungal species. However, in main text, there is no such description. Information in Abstract should appear in main text with more contents. 

4. Throughout the manuscript, C. auris and other species names should be italicized.

5.line 119: How did authors select 23 research articles? The reason should be written.

6. Chapter 3 has a title, "Molecular assay....". This is a main portion of this review. However, descriptions are just arrangement of published findings and not written systematically. Information of mutation related to antifungal resistance are also mixed up. Review article should summarize these information systematically.  For example, molecular methods that have been used are listed as "1)PCR, 2)sequencing of  ITS region, 3)MALDI-TOF, 4)whole genome analysis, ...., and describe their sensitivity, feasibility in clinical laboratory, cost, frequency of usage, good points, short point, etc. 

7. Table 1 has a title of "Molecular detection.....". However, In the column of "Assay used for C. auris detection", description is not in uniformity, and "sensitivity test" is also shown, without information of its method. For example, in first article by Naeimi, assay shows "Pan-fungal primers...". This should be written as "conventional PCR", and primers or target gene should be added as supplementary information. Table 1 should be substantially revised.

8. line 166, 168: authors wrote "clade" and its specificity or common features. But it is very simply written. It is better to add explanation of C. auris clades in introduction section, and specificity of mutation or resistance mechanism with clade should be described in more detail in text, or in a newly created Table.      

Comments on the Quality of English Language

This manuscript is generally acceptable, but contains some poor phrases.

Author Response

Point-by-point response to the reviewers’ and editor’s comments

 

Title: “Candida auris updates: molecular methods detections, antifungal stewardship”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #3:

 

Comments and Suggestions for Authors

 

This manuscript describes the latest knowledge on detection methods and antifungal stewardship of C. auris.  This seems to be interesting and provide useful information to readers but contains several concerns as follows.

 

1. Title should be reconsidered. Is "molecular methods detection" correct?

• Response to Reviewer: We thank the reviewer for this comment. We have modified the title, as suggested.

 

Candida auris Updates: Outbreaks Evaluation Through Molecular Assays and Antifungal Stewardship – a Narrative Review

2. There have been many review papers on C. auris to date. In this manuscript, what point did authors focus? Authors should write objectives of this review in Introduction.

• Response to Reviewer: We thank the reviewer for this comment. We have added the suggested paragraph in the abstract and introduction (lines 24-25, 101-106).

 

We aimed to assess which molecular assays have been used to diagnose Candida auris outbreaks in the last five years. (lines 24-25)

 

The aim of this review was to summarize the recent findings regarding Candida auris molecular diagnosis in the worldwide outbreaks, registered in the last five years. So far, there is no FDA-approved assay for this fungus, and therefore hospital microbiology laboratories should be properly informed, from a scientific and logistic point of view, regarding which assay is most suitable to be implemented for C. auris fast and accurate diagnosis. (lines 101-106)

 

3. In Abstract, there is explanation of new nomenclature of some fungal species. However, in main text, there is no such description. Information in Abstract should appear in main text with more contents.

• Response to Reviewer: We thank the reviewer for this comment. We have added the suggested text in the introduction (lines 52-58).

 

Although taxonomical changes may prove difficult to adapt to, recent fungal reclassification is another step in underlining the importance of fungal resistance and possible emergence of novel pathogens. Recently, a change in the nomenclature of fungi has been underway, reclassifying fungal species into new genera based on antifungal resistance and thermotolerance [2]. The WHO priority fungi list mentions two such reclassified species: Nakaseomyces glabrata (Candida glabrata) and Pichia kudriavzevii (Candida krusei), in the high and medium priority groups, respectively [1].

 

 

4. Throughout the manuscript, C. auris and other species names should be italicized.

• Response to Reviewer: We thank the reviewer for this comment. We have modified as suggested.

 

 

 

 

5. line 119: How did authors select 23 research articles? The reason should be written.

• Response to Reviewer: We thank the reviewer for this question. At moment of our PubMed search for the last 5 years, we found 23 papers, by using the following inclusion and exclusion criteria: The used key words were ”Candida auris outbreak molecular detection”, which were specified in the abstract and in the manuscript also.

https://pubmed.ncbi.nlm.nih.gov/?term=Candida+auris+outbreak+molecular+detection&filter=datesearch.y_5&sort=pubdate&size=200

Since February, 2 other papers were published and we introduced them in our manuscript, in Table 1.

 

Our search initiated in 12.02.2024, in PubMed Database, using “Candida auris; outbreak; molecular detection” key words, revealed 34 results for the last 5 years. We included in our analysis (Table 1) only 25 original research papers [14-38]. We excluded review papers. Since our last search, two more papers were published, and out Table now contains 25 papers.

 

 

The final inclusion / exclusion criteria are:

 

Our search initiated in 12.02.2024, in PubMed Database, using “Candida auris; outbreak; molecular detection” key words, revealed 34 results for the last 5 years. We included in our analysis (Table 1) only 25 original research papers [14-38]. We excluded review papers. (lines 153 – 156).

 

6. Chapter 3 has a title, "Molecular assay....". This is a main portion of this review. However, descriptions are just arrangement of published findings and not written systematically. Information of mutation related to antifungal resistance are also mixed up. Review article should summarize these information systematically. For example, molecular methods that have been used are listed as "1)PCR, 2)sequencing of ITS region, 3)MALDI-TOF, 4)whole genome analysis, ...., and describe their sensitivity, feasibility in clinical laboratory, cost, frequency of usage, good points, short point, etc.

• Response to Reviewer: We thank the reviewer for this comment. We reorganized the table as suggested and inserted the required comments (lines 180 – 191).

 

 

 

In our analysis, we identified 25 papers, from which 6 used WGS + MALDI-TOF, 3 used MALDI-TOF + PCR / cultivation, 11 used PCR and 5 used other methods for C. auris detection.

The most comprehensive assay is WGS, of course, being a method that offers detailed genetic information, but has the disadvantage that it requires special and expensive laboratory equipment to be performed. MALDI-TOF is a very fast and accurate identification assay, and it needs to be followed by antifungal susceptibility testing. Real Time PCR allows for fast detection, and it was the most frequently used assay, probably because many laboratories, after the emergence of the SARS-CoV-2 pandemic, acquired the necessary equipment. More than simple detection, Real Time PCR which is able to detect antifungal resistance genes seems to be even more suitable for fast diagnosis in Candida auris outbreaks.

 

 

 

7. Table 1 has a title of "Molecular detection.....". However, In the column of "Assay used for C. auris detection", description is not in uniformity, and "sensitivity test" is also shown, without information of its method. For example, in first article by Naeimi, assay shows "Pan-fungal primers...". This should be written as "conventional PCR", and primers or target gene should be added as supplementary information. Table 1 should be substantially revised.

• Response to Reviewer: We thank the reviewer for this comment. We modified the table as suggested.

 

8. line 166, 168: authors wrote "clade" and its specificity or common features. But it is very simply written. It is better to add explanation of C. auris clades in introduction section, and specificity of mutation or resistance mechanism with clade should be described in more detail in text, or in a newly created Table.

• Response to Reviewer: We thank the reviewer for this comment. We have added the suggested in the introduction and section about antifungal stewardship (lines 58-66 and 305-310).

 

Candida auris, a critical priority fungal species, according to the WHO, emerged as a yeast that is multidrug-resistant in the past decade. This species inhabits the surface of human skin and hospital settings, leading to the occurrence of healthcare-associated epidemics and invasive candidiasis. Initial genetic studies identified four primary clades, which were designated after their corresponding geographical locations: South-Asia, East-Asia, Africa, and South America. These clades are referred to as clade I, clade II, clade III, and clade IV, respectively and are differentiated by at least of ten thousand single nucleotide polymorphisms (SNPs) between any two clades. In 2022, a fifth clade, from Iran, was confirmed by whole-genome sequencing [3]. (lines 58-66)

 

Several papers report on the relationship between C. auris clade and antifungal re-sistance, which were included in a recent review by Sharma and Kadosh, who found that isolates belonging to clades I, III, IV and V show resistance to fluconazole, cross-resistance to echinocandins and amphotericin B, with even the possibility of pan-drug resistant strains. Clade II, found predominantly in Korea and Japan, was found to have a lower de-gree of antifungal resistance [57]. (lines 305-310)

 

 

Comments on the Quality of English Language

This manuscript is generally acceptable but contains some poor phrases.

• Response to Reviewer: We thank the reviewer for this comment. In case of acceptance, we will ask MDPI for an official English Article Editing.

 

 

Submission Date

27 April 2024

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

I thank the authors for the manuscript improvement and for considering the suggestions. The revised version reads much better. My only pending concern is related to the antifungal drugs. As mentioned, the issue is oversimplified and section 4 is not at the same level as the rest of the manuscript. The authors agreed that this was not part of the original goal, so why do you think the inclusion of partial information increases the manuscript quality? On the contrary, I think it has a negative impact on it. Once again, I suggest the authors to delete section 4.

Author Response

 

Point-by-point response to the reviewers’ and editor’s comments

 

Title: “Candida auris Updates: Outbreaks Evaluation Through Molecular Assays and Antifungal Stewardship – a Narrative Review”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #2:

 

Comments and Suggestions for Authors

I thank the authors for the manuscript improvement and for considering the suggestions. The revised version reads much better. My only pending concern is related to the antifungal drugs. As mentioned, the issue is oversimplified, and section 4 is not at the same level as the rest of the manuscript. The authors agreed that this was not part of the original goal, so why do you think the inclusion of partial information increases the manuscript quality? On the contrary, I think it has a negative impact on it. Once again, I suggest the authors to delete section 4.

• Response to Reviewer: We thank the reviewer for this comment. We have removed the table, as suggested.

 

Reviewer 3 Report

Comments and Suggestions for Authors

The revised version has been well written with high quality.

Author Response

Point-by-point response to the reviewers’ and editor’s comments

 

Title: “Candida auris Updates: Outbreaks Evaluation Through Molecular Assays and Antifungal Stewardship – a Narrative Review”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #3:

 

Comments and Suggestions for Authors

The revised version has been well written with high quality.

• Response to Reviewer: We thank the reviewer for this comment and for helping us in improving the quality of our manuscript.

Round 3

Reviewer 2 Report

Comments and Suggestions for Authors

Tha manuscript is suitable for publication.

Author Response

 

Point-by-point response to the reviewers’ and editor’s comments

 

Title: “Candida auris Updates: Outbreaks Evaluation Through Molec-ular Assays and Antifungal Stewardship – a Narrative Review”

 

We thank the reviewer for giving us the opportunity to improve the quality of our manuscript.

 

Reviewer #2:

 

 

Comments and Suggestions for Authors

The manuscript is suitable for publication.

• Response to Reviewer: We thank the reviewer for helping us in improving the quality of our manuscript.

 

 

Submission Date         27 April 2024

Date of this review     24 May 2024 16:52:17

Author Response File: Author Response.docx