SARS-CoV-2 Variants of Concern and Clinical Severity in the Mexican Pediatric Population

Round 1

Reviewer 1 Report

This manuscript is interesting with a large sample size in Mexico. Outcomes were straightforward and consistent with the other finding in other countries or continents. However, the clinical data were lacking. Suggest adding more data, especially vaccine used to make it more information in these populations. The genetic part was clear and no doubt of it.

 

Major concerns.

1. Suggest adding the COVID-19 vaccination parameters to Table 2; vaccine types (inactivated, viral vector, mRNA...), and vaccination times (never, 1-dose, 2-dose, 3-dose, 4-dose or more).

Vaccination is an important variable that could reduce the chance of severe or life-threatening infection. This point should be added.

 

Comments.

1. Figure S2. Readers may print out a monochrome hard copy. Colour in Figures may be turned to undistinguishable greyscale. Moreover, nearly colour tones may be undifferentiated in readers with colourblindness.

Suggest changing the colour pattern following these recommendations to ensure maximum accessibility.

- http://www.cookbook-r.com/Graphs/Colors_(ggplot2)

- https://colorbrewer2.org/#type=sequential&scheme=BuGn&n=3

You may be adding a solid pattern to each section.

2. Line 54. "~6 × 10−4 nucleotides/genome/year", suggest superscript of "-4".

Author Response

Reviewer 1

Comments and Suggestions for Authors

This manuscript is interesting with a large sample size in Mexico. Outcomes were straightforward and consistent with the other finding in other countries or continents. However, the clinical data were lacking. Suggest adding more data, especially vaccine used to make it more information in these populations. The genetic part was clear and no doubt of it.

Response:

We deeply appreciate the time and effort for reviewing our manuscript, we have been taking in consideration all your suggestions. Unfortunately, regarding the vaccination data, the Epidemiological Surveillance System for COVID-19 in Mexico (SINAVE), has significant shortcomings, such as the lack of information regarding to COVID-19 vaccination. The distribution and recording of vaccines were carried out by the Mexican Government and the Ministry of Defense. As a result, this information is not available for open data assessment and could not be included in our analysis.

Major concerns.

1. Suggest adding the COVID-19 vaccination parameters to Table 2; vaccine types (inactivated, viral vector, mRNA...), and vaccination times (never, 1-dose, 2-dose, 3-dose, 4-dose or more).

Vaccination is an important variable that could reduce the chance of severe or life-threatening infection. This point should be added.

Response:

Added in the discussion section (line 391-396):

In Mexico, the first COVID-19 vaccination was received for emergency use and authorization for ages 12 to 17 was in September 2021. Vaccination for children aged 5 to 12 started in March 2022. Unfortunately, the vaccination status regarding COVID-19 in the pediatric population could not be determined because there is no proper record in the epidemiological surveillance systems (SINAVE) to be used for this analysis.

Added in the study limitations (lines 405-408):

Further studies combining genomic variability, immunization status, comorbidities, epidemiological detail information and clinical features of COVID-19 pediatric patients may be useful to characterize the physiopathological effects on the final pediatric clinical outcomes.

Comments.

1. Figure S2. Readers may print out a monochrome hard copy. Colour in Figures may be turned to undistinguishable greyscale. Moreover, nearly colour tones may be undifferentiated in readers with colourblindness.

Suggest changing the colour pattern following these recommendations to ensure maximum accessibility.

- http://www.cookbook-r.com/Graphs/Colors_(ggplot2)

- https://colorbrewer2.org/#type=sequential&scheme=BuGn&n=3

You may be adding a solid pattern to each section.

Response:

A colorblind-friendly palette used with ggplot2, was added to all the graphics as suggested. For the map (Figure 3), color brewer 2.0 was used to create a colorblind safe, blue single hue palette.

2. Line 54. "~6 × 10−4 nucleotides/genome/year", suggest superscript of "-4".

Response:

Added in the introduction section (lines 53-54):

Preliminary estimates of the evolutionary rate of SARS-CoV-2 are close to ~6 × 10⁻⁴ nucleotides/genome/year, similar to other RNA virus genomes.

Reviewer 2 Report

The  observation that the age of the patients influences the development of mutations and maybe mutationals hotspots in the viral genome is novel and important. It may be worth to investigate the molecular rmechanisms thar underly this observation (but this would go beyond the study.

 

My sole but major concern is that the authors did not comment on comorbidities or underlying chronic diseases in those pediatric patients that required mechanical ventilation/ICU and who died. This would be important, as in European countries those serious clinical courses were always associated with a serious risk factor. In this context it would also be worth to check (if possible) if any comorbidity is associated with a lineage dependent more severe outcome. If the case numbers are to slow, I rommend e.g. a Bonferroni calculation or similar to check for statistical significance.

 

Minor comments:

 

Line 263: To the very best of my knowledge, the term gender should be replaced by sex, as gender is a social term while sex defines the biological status (male or female).

 

Author Response

Reviewer 2

Comments and Suggestions for Authors.

The observation that the age of the patients influences the development of mutations and maybe mutational hotspots in the viral genome is novel and important. It may be worth to investigate the molecular mechanisms that underly this observation (but this would go beyond the study).

Response:

We deeply appreciate the time and effort for reviewing our manuscript, we have been taking in consideration all your suggestions. Understanding the clinical severity associated with SARS-CoV-2 variants of concern (VOC) is important for public health implications, both nationally and internationally. Some mutations associated to VOCs identified in this work have been target of previous studies to understand their impact in viral transmission, disease severity, reinfection rates, and vaccine effectiveness. For this reason, Table S1 outlining the key effects on viral activity that could impact the pediatric population has been included in the supplementary materials.

Added to the results section (Line 187-190):

The substantial presence of these high-frequency mutations in the pediatric population holds significant implications, particularly concerning immunological and transmission features (Table S1).

Added to the supplementary section: 

Table S1. Viral activity effects reported in high-frequency mutations associated with pediatric sequences in Mexico.

Gen / Mutation	VOCs1		Effects in viral activity
			Transmission		Immunity		Severity
Envelope
T9I	ο						Virulence decreased.[1]
Nucleocapsid
P13L	ο				Decreased T cell recognition.[2]		Decreased pathogenicity.[3,4]
Spike
T478K	δ ο		Increase ACE2 affinity.[5] Increased potential transmission.[6]		Reduced affinity and antibody response.[7,8] Increased antibody escape.[5,9]		Increase virulence.[5] Increased severity.[10]
P681H	α ο		Increase ACE2 binding.[11]  Increase infectivity and transmission.[12]		Reduced affinity and antibody response. [13,14]		Increased virulence.[15] Increase replication, viral load. .[12]
G142D	ο				Decrease antibody binding.[11]
H655Y	ɣ ο		Increase ACE2 binding.[11]
L452R	δ ο				Reduced affinity and antibody response.[7]
N679K	ο		Increase ACE2 binding.[11]
N969K	ο		Decrease infectivity.[16]
Q954H	ο		Decrease infectivity.[16]
N764K	ο						Increase virulence.[17]
G339D	ο				Increase antibody escape.[18]
S375F	ο		Increased potential transmission.[6]		Increase antibody escape.[18]
S373P	ο		Increased potential transmission.[6]		Increase antibody escape.[18]
N501Y	α β ɣ ο		Increase infectivity and transmission.[6,18]		Reduced affinity and antibody response.[7,18]

¹Variants of Concern (α) Alpha, (β)Beta, (ɣ) Gamma, (δ) Delta, and (ο) Omicron.

My sole but major concern is that the authors did not comment on comorbidities or underlying chronic diseases in those pediatric patients that required mechanical ventilation/ICU and who died. This would be important, as in European countries those serious clinical courses were always associated with a serious risk factor. In this context it would also be worth to check (if possible) if any comorbidity is associated with a lineage dependent more severe outcome. If the case numbers are to slow, I rommend e.g. a Bonferroni calculation or similar to check for statistical significance.

Response:

Pediatric comorbidities are a main feature influence in clinical severity outcomes. Unfortunately, the dataset recollected in Mexico with the Epidemiological Surveillance System for COVID-19 (SINAVE), has significant shortcomings and does not include information regarding comorbidities in pediatric patients. For this reason, we were not able to use for comorbidity parameters in our analysis.

Added to the study limitations (lines 405-407):

Further studies combining genomic variability, immunization status, comorbidities, epidemiological detail information and clinical features of COVID-19 pediatric patients may be useful to characterize the physiopathological effects on the final pediatric clinical outcomes.

Minor comments:

Line 263: To the very best of my knowledge, the term gender should be replaced by sex, as gender is a social term while sex defines the biological status (male or female).

Response:

Added to the results section (lines 282-287):

A comparison of the principal pediatric severity outcomes across the VOCs groups is presented in Table 2. Sociodemographic factors such as age and sex were included. The age means distributions across the VOCs groups were similar. Only the Omicron lineage presented a decreased age mean of 10.33±4.88 years, compared to other VOCs groups (p<0.001). The male patients resulted in a slightly increased proportion versus the female COVID-19 patients.

Added to the Table 2:

Sex

Round 2

Reviewer 2 Report

My previous comments have been thoroughly addressed, thank you. Very interesting study.