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Bactogram : Spatial Analysis of Bacterial Colonization in Epidermal Wounds

Wallblom, Karl LU orcid ; Forsberg, Fredrik LU ; Lundgren, Sigrid LU ; Fisher, Jane LU ; Cardoso, José LU ; Petruk, Ganna LU orcid ; Strömdahl, Ann-Charlotte LU ; Saleh, Karim LU ; Puthia, Manoj LU and Schmidtchen, Artur LU (2024)
Abstract
Skin barrier damage and subsequent development of harmful microbiota contribute to conditions such as wound infections, atopic dermatitis, and chronic wounds, which impact millions of people globally and pose a significant economic burden on healthcare systems. Established microbial sampling methods, such as swabs and tissue biopsies, provide limited information on the spatial distribution of bacteria. We here describe a new method that produces a visual map of the distribution of cultivable bacteria, denoted “Bactogram”, across the whole wound and surrounding skin, suitable for image-based quantification. As part of an exploratory endpoint in a clinical trial (NCT05378997) we applied the Bactogram method to 48 suction blister wounds in 24... (More)
Skin barrier damage and subsequent development of harmful microbiota contribute to conditions such as wound infections, atopic dermatitis, and chronic wounds, which impact millions of people globally and pose a significant economic burden on healthcare systems. Established microbial sampling methods, such as swabs and tissue biopsies, provide limited information on the spatial distribution of bacteria. We here describe a new method that produces a visual map of the distribution of cultivable bacteria, denoted “Bactogram”, across the whole wound and surrounding skin, suitable for image-based quantification. As part of an exploratory endpoint in a clinical trial (NCT05378997) we applied the Bactogram method to 48 suction blister wounds in 24 healthy volunteers. Bacteria developed in all wounds, predominantly on the skin under the dressing and near wound edges. Two quantification methods, based on visual scoring and image analysis, demonstrated high inter-, and intra-rater agreement and were used to characterize bacterial re-colonization during epidermal wound healing. We also demonstrated proof of concept that the method can be used with chromogenic agar to enable spatial identification of pathogenic bacterial species, such as Staphylococcus aureus. In conclusion, this study introduces a simple method for sampling bacteria over large areas and generating a bacterial map that can identify spatial variations in bacterial composition and abundance in skin and wound conditions.Competing Interest StatementA.S. is a founder of in2cure AB, a parent company of Xinnate AB, companies that are developing therapies based on thrombin-derived peptides and variants. G.P. is employed part-time (20%) by Xinnate AB. The other authors have declared that no conflict of interest exists.Clinical TrialNCT05378997Clinical Protocols https://doi.org/10.1136/bmjopen-2022-064866 Funding StatementThe exploratory data presented here was supported by grants from the Swedish Research Council (project 2017-02341, 2020-02016), Edvard Welanders Stiftelse and Finsenstiftelsen (Hudfonden), the Royal Physiographic Society, the Crafoord and Österlund Foundations, and the Swedish Government Funds for Clinical Research (ALF). Xinnate AB provided the project management resources and expertise for the regulatory development enabling the clinical parts of the Safety study that generated the control samples used in this work.Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Etikprövningsmyndigheten (Swedish Ethics committee) gave ethical approval for this work.I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).YesI have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable.YesThe generated pictures of the Bactograms will be available on reasonable request, after the publication of the treated wound data in a separate publication, since the original images inevitably include both treated and untreated wounds. The code used for the ImageJ macros can be found in Supplementary Data 3. The Python code for creating spatial heat maps can be found in Supplementary Data 4. The complete score data from both the visual scoring and the computer-assisted method is provided in Supplementary Data 5. Further information or instructions required to reanalyze the data reported in this paper are available from the lead contact upon reasonable request. (Less)
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; ; ; ; ; ; ; ; and
organization
publishing date
type
Working paper/Preprint
publication status
published
subject
keywords
Wound healing, Clinical study, Wound infection, Staphylococcus aureus, Microbiology
pages
33 pages
publisher
medRxiv
DOI
10.1101/2024.04.18.24305961
language
English
LU publication?
yes
id
d7eb3681-dd78-484a-836c-be3c337d8e6f
date added to LUP
2024-05-13 10:49:07
date last changed
2024-05-31 15:49:37
@misc{d7eb3681-dd78-484a-836c-be3c337d8e6f,
  abstract     = {{Skin barrier damage and subsequent development of harmful microbiota contribute to conditions such as wound infections, atopic dermatitis, and chronic wounds, which impact millions of people globally and pose a significant economic burden on healthcare systems. Established microbial sampling methods, such as swabs and tissue biopsies, provide limited information on the spatial distribution of bacteria. We here describe a new method that produces a visual map of the distribution of cultivable bacteria, denoted “Bactogram”, across the whole wound and surrounding skin, suitable for image-based quantification. As part of an exploratory endpoint in a clinical trial (NCT05378997) we applied the Bactogram method to 48 suction blister wounds in 24 healthy volunteers. Bacteria developed in all wounds, predominantly on the skin under the dressing and near wound edges. Two quantification methods, based on visual scoring and image analysis, demonstrated high inter-, and intra-rater agreement and were used to characterize bacterial re-colonization during epidermal wound healing. We also demonstrated proof of concept that the method can be used with chromogenic agar to enable spatial identification of pathogenic bacterial species, such as Staphylococcus aureus. In conclusion, this study introduces a simple method for sampling bacteria over large areas and generating a bacterial map that can identify spatial variations in bacterial composition and abundance in skin and wound conditions.Competing Interest StatementA.S. is a founder of in2cure AB, a parent company of Xinnate AB, companies that are developing therapies based on thrombin-derived peptides and variants. G.P. is employed part-time (20%) by Xinnate AB. The other authors have declared that no conflict of interest exists.Clinical TrialNCT05378997Clinical Protocols https://doi.org/10.1136/bmjopen-2022-064866 Funding StatementThe exploratory data presented here was supported by grants from the Swedish Research Council (project 2017-02341, 2020-02016), Edvard Welanders Stiftelse and Finsenstiftelsen (Hudfonden), the Royal Physiographic Society, the Crafoord and Österlund Foundations, and the Swedish Government Funds for Clinical Research (ALF). Xinnate AB provided the project management resources and expertise for the regulatory development enabling the clinical parts of the Safety study that generated the control samples used in this work.Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Etikprövningsmyndigheten (Swedish Ethics committee) gave ethical approval for this work.I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).YesI have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable.YesThe generated pictures of the Bactograms will be available on reasonable request, after the publication of the treated wound data in a separate publication, since the original images inevitably include both treated and untreated wounds. The code used for the ImageJ macros can be found in Supplementary Data 3. The Python code for creating spatial heat maps can be found in Supplementary Data 4. The complete score data from both the visual scoring and the computer-assisted method is provided in Supplementary Data 5. Further information or instructions required to reanalyze the data reported in this paper are available from the lead contact upon reasonable request.}},
  author       = {{Wallblom, Karl and Forsberg, Fredrik and Lundgren, Sigrid and Fisher, Jane and Cardoso, José and Petruk, Ganna and Strömdahl, Ann-Charlotte and Saleh, Karim and Puthia, Manoj and Schmidtchen, Artur}},
  keywords     = {{Wound healing; Clinical study; Wound infection; Staphylococcus aureus; Microbiology}},
  language     = {{eng}},
  month        = {{01}},
  note         = {{Preprint}},
  publisher    = {{medRxiv}},
  title        = {{Bactogram : Spatial Analysis of Bacterial Colonization in Epidermal Wounds}},
  url          = {{http://dx.doi.org/10.1101/2024.04.18.24305961}},
  doi          = {{10.1101/2024.04.18.24305961}},
  year         = {{2024}},
}