An Unconventional Oral Candidiasis in an Immunocompetent Patient

Round 1

Reviewer 1 Report

The overall structure and pacing of the manuscript was easy to follow and read.  The authors did a wonderful job presenting the data in a clear and concise manner, moving from one logical step to the next with ease and clarity.  All of the results shown and explained sufficiently supported the reported hypothesis of the authors throughout the body of the manuscript.

 

However, upon reaching the discussion section of the manuscript the authors then claim that their finding and observed drug resistance phenotypes were the result of a C. albicans biofilm infection. Not once in the manuscript did the authors state or show data indicating the presence of a biofilm. The authors did show data (gram stains showing the presence of hyphae), however this does not indicate the presence of a biofilm as Candida yeast have the ability to form hyphae/pseudohyphae in response to their environment. The stains do show the presence of hyphae/pseudohyphae but they do not have the characteristic look of a biofilm nor do they prove the existence of a biofilm in this particular patient.  

The authors did include extensive MIC testing against known azoles, polyenes, echinocandins, yet these were done against planktonic cells and not their biofilm counterparts.  It has been shown and reported that yeast cells that originate from a biofilm can exhibit MICs in the range that are susceptible to antifungals, yet upon transition to the biofilm state gain resistance upwards to 1000x the recommended antifungal dosage. By the authors stating that this particular infection was the result of a biofilm infection is highly presumptuous and should not be concluded based upon the results they are showing in this manuscript.  If this infection is believed to be the result of a biofilm infection then I think there needs to be results presented in the manuscript that are able to support this claim.

 

Author Response

The authors did include extensive MIC testing against known azoles, polyenes, echinocandins, yet these were done against planktonic cells and not their biofilm counterparts.  It has been shown and reported that yeast cells that originate from a biofilm can exhibit MICs in the range that are susceptible to antifungals, yet upon transition to the biofilm state gain resistance upwards to 1000x the recommended antifungal dosage. By the authors stating that this particular infection was the result of a biofilm infection is highly presumptuous and should not be concluded based upon the results they are showing in this manuscript.  If this infection is believed to be the result of a biofilm infection then I think there needs to be results presented in the manuscript that are able to support this claim.

 

Dear Reviewer,

thank you so much for your positive evaluation of our manuscript and for appreciating our work. We agree with you that the thesis on biofilm formation needed to be supported by real data. As you requested, we carried out a biofilm formation assay using the two isolated strains. The results have been added to the manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

Dear Authors,

The manuscript has been written well and the case was presented perfectly. However, some ambiguous issues remained to be clarify:

1.    Are you sure that you didn’t miss the multi-sensitive C. albicans isolate in the first round of identification?

2.    How the patient was cured in spite of the persistence of the C. albicans multi-resistant strain? Please discuss about this.

3.    Please add timeline for presenting the case. The progress have to be defined exactly.

 

 

 

 

 

Author Response

• Are you sure that you didn’t miss the multi-sensitive C. albicans isolate in the first round of identification?

 

Dear Reviewer,

thank you so much for your positive evaluation of our manuscript and for appreciating our work. We do not think we have missed the multi-sensitive C. albicans in the first round of identification, first of all because the swabs were carried out both in the private laboratory and in our microbiology unit and the result was superimposable in both identification rounds, on the other hand, the antimycograms were repeated more than once on different colonies and never showed the presence of two different profiles. Furthermore, the molecular profiles of the two isolates appear to be identical, therefore it is probable that they are the same strain that changed its phenotype, and which in the meantime had also reached the intestinal tract (as found in coproculture).

 

• How the patient was cured in spite of the persistence of the C. albicans multi-resistant strain? Please discuss about this.

 

Done (lines 222-226 and 434-440)

 

• Please add timeline for presenting the case. The progress have to be defined exactly.

 

The timeline has been reported along with the text, for a better presentation of the case.

Please consider that the patient followed the conventional routes as a case report and not an experimental study. Hence, the patient performed serological investigation and urine and stool analyses in an external laboratory as the ORL visit. We first aimed to eradicate the infection from the patients and acted under the guidelines for managing oral candidiasis, supported by the evidence from the literature when we chose to use probiotics and betadine instead of hospitalizing the patient, otherwise healthy.

Author Response File: Author Response.pdf

Reviewer 3 Report

 Manuscript ID: jof-2190049
Type of manuscript: Case Report

Journal. Journal of fungi. Case report. MDPI.

 Title : An unconventional Oral Candidiasis in an Immunocompetent  Patient

Authors:  Alessandra Fusco, Maria Contaldo, Vittoria Savio, Adone Baroni, Giuseppe A. Ferraro , Dario Di Stasio, 4 Alberta Lucchese, Giovanna Donnarumma and Rosario Serpico

 

More details are needed at different levels to explain the failure of the immune system concerning the particular case mentioned in the article.

At several levels, the failure of the immune systems must be evoked thanks to specific markers.

The clinical management of the pathology must begin with sanitation of the oral cavity (hygiene, scaling, root planing). Even before starting an antifungal treatment. The bacterial component is preponderant in the establishment of mycofilms (antagonism, synergy, opportunism, neutrality) hence the interest of probiotics and prebiotics. The failure of the immune system can also be measured using several markers of inflammation such as: (  Cytokines/chemokines: Interleukin 1-alpha and -beta (IL-1α, IL-1β), Interleukin-4 (IL-4); Interleukin-6 (IL-6), Interleukin-8 (IL-8), Interleukin-10 (IL-10); Interleukin-13 (IL-13); Monocyte chemoattractant protein-1 (MCP-1), Interferon-gamma (IFN-ϒ); Tumor necrosis factor-alpha (TNF-α)

 

Antifungals are not specific because they can also harm host cells. Finally, the total eradication of the candidal population is not desirable. The whole difficulty is to try to maintain this population at an acceptable level in order to restore the balance within the microbiota over the long term.

  Concerning C. lusitaniae (teleomorph Clavispora lusitaniae) to be part of the normal mycobiota of animals, though its prevalence among isolates from clinical samples is low. Acquired resistance by C. lusitaniae to the polyene antifungal  has also been noted. Serious infections by C. lusitaniae typically involve patients with hematological malignancies as well as other types of individuals being treated in intensive care units.

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mycobiota of animals, though its prevalence among isolates from clinical samples is low. Acquired resistance by C. lusitaniae to the polyene antifungal  has also been noted. Serious infections by C. lusitaniae typically involve patients with hematological malignancies as well as other types of individuals being treated in intensive care units.

 

mycobiota of animals, though its prevalence among isolates from clinical samples is low. Acquired resistance by C. lusitaniae to the polyene antifungal  has also been noted. Serious infections by C. lusitaniae typically involve patients with hematological malignancies as well as other types of individuals being treated in intensive care units.

 view notes on PDF

 

Comments for author File: Comments.pdf

Author Response

Dear Reviewer,

Thank you so much for your careful analysis of our manuscript.

 

• The clinical management of the pathology must begin with sanitation of the oral cavity (hygiene, scaling, root planing). Even before starting an antifungal treatment. The bacterial component is preponderant in the establishment of mycofilms (antagonism, synergy, opportunism, neutrality) hence the interest of probiotics and prebiotics.

 

In the routine clinical practice adopted at our facility, being a dental clinic, we constantly evaluated our patients' oral hygiene before prescribing any pharmacological or surgical treatment. In this case, the patient referred us to have undergone recent professional hygiene for dental calculus debridement. On the occasion of the first clinical examination, no calculus or evident dental plaque was found, nor signs of gingivitis. What you define as "visible inflammation on palatal gingiva", is the condition of the gums that appears to be inflamed but is instead the condition of the detached and no longer inflamed marginal gingiva, as usually occurs after a professional scaling and root planing due to reduction of gingival edema which makes the classical picture of a fake "worsening": in reality, the gingiva is simply and provisionally detached, waiting for the physiologically epithelial attachment to recover in time.

We did not report this obvious sentence previously, but you can find it now in lines 70-72.

 

• The failure of the immune system can also be measured using several markers of inflammation such as: (  Cytokines/chemokines: Interleukin 1-alpha and -beta (IL-1α, IL-1β), Interleukin-4 (IL-4); Interleukin-6 (IL-6), Interleukin-8 (IL-8), Interleukin-10 (IL-10); Interleukin-13 (IL-13); Monocyte chemoattractant protein-1 (MCP-1), Interferon-gamma (IFN-ϒ); Tumor necrosis factor-alpha (TNF-α)

 

Considering the general picture, the hematological profile, the lymphocyte populations, the erythrocyte sedimentation rate just slightly higher than the normal range, and other inflammatory markers, we did not deem it appropriate to persevere and verify these other inflammatory markers because they are out of standard clinical practice. We could have valued them in the saliva, but inevitably they would have been altered due to the local oral conditions.

Further replies to your comments are in the attached file

Author Response File: Author Response.pdf