Oxytetracycline reduces inflammation and treponeme burden whereas vitamin D3 promotes β-defensin expression in bovine infectious digital dermatitis

  • Kaitlyn M. Watts
  • Priyoshi Lahiri
  • Rakel Arrazuria
  • Jeroen De Buck
  • Cameron G. Knight
  • Karin Orsel
  • Herman W. Barkema
  • Eduardo R. CoboEmail author
Regular Article


Digital dermatitis (DD), a common ulcerative disease of the bovine foot causing lameness and reducing productivity and animal welfare, is associated with infection by spirochete Treponema bacteria. Topical tetracycline, the most common treatment, has inconsistent cure rates; therefore, new therapeutic options are needed. We compared effects of topical oxytetracycline and vitamin D3 on innate immunity in DD-affected skin. Cows with active DD lesions were treated topically with oxytetracycline or vitamin D3 and skin biopsies were collected from lesions. Tissue samples were examined histologically, transcriptional expression of pro-inflammatory cytokines, Toll-like receptors (TLRs), and host defense peptides assessed, and the presence of specific treponeme species determined. Effects of treatments at a mechanistic level were studied in a human keratinocyte model of treponeme infection. Oxytetracycline promoted hyperplastic scab formation in ulcerated DD lesions and decreased transcriptional expression of Cxcl-8 (neutrophil chemoattractant). Oxytetracycline also reduced numbers of Treponema phagedenis and T. pedis and enhanced Tlr2 mRNA expression. Vitamin D3 did not modify expression of cytokines or Tlrs, or bacterial loads, but enhanced transcription of tracheal antimicrobial peptide (Tap), a key bovine β-defensin. Combing oxytetracycline and vitamin D3 provides complementary clinical benefits in controlling DD through a combination of antimicrobial, immunomodulatory, and pro-healing activities.


Treponema spp. Digital dermatitis Cattle Oxytetracycline Vitamin D3 Cathelicidins β-Defensins Interleukin 8 



We thank certified hoof trimmers Rob Geier (Casper Trimming, Calgary, AB, Canada) and Elbert Koster (No-tilt Hoof Trimming, Calgary, AB, Canada) for facilitating collection of biopsies. We also thank Dr. Brielle Rosa for her critical review and editing the manuscript.

Author contributions

KW and EC conceived and designed the experiments. KW and KO conducted animal sampling. KW and PL conducted in vitro experiments and analyzed data. RA performed Treponema spp. identification by culture and qPCR. RA and PL conducted immunohistochemistry and CK conducted histopathological examination. KO, CK, RA, JDB, HB, and EC contributed to data analysis and interpretation as well as manuscript editing. KW, PL, CK, and EC drafted and wrote the manuscript.


This work was supported by the Natural Sciences and Engineering Research Council (NSERC) Discovery Grant (RGPAS-2017- 507827) and the Alberta Agriculture and Forestry (2019F040R) to ERC. KW was supported by Queen Elizabeth II Graduate Scholarship.

Compliance with ethical standards

The authors declare no conflicts of interest. Studies involving cattle were reviewed and approved by the University of Calgary Animal Care Guidelines (AC16-0070) for the care and use of animals. The owner’s written informed consent was obtained prior to the use of any cattle.

Supplementary material

441_2019_3082_Fig6_ESM.png (257 kb)
Supplementary Figure S1

Transcriptional gene expression of cytokines, Toll-like receptors (Tlr), host defence peptides, and vitamin D enzymes in foot lesions of cows with active (M2) DD treated with vitamin D3 (VitD3) or oxytetracycline (OTC) (5-d intervals). Relative mRNA expression of Tlr4 (a), Ifn-γ (b), cathelicidin bovine myeloid antimicrobial peptide (Bmap)-28 (c), β-defensin lingual antimicrobial peptide (Lap) (d), and vitamin D3 metabolite Cyp24A1(e) was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. Only significant comparisons (P < 0.05) are noted (one-way ANOVA using Tukey’s post hoc test). (PNG 257 kb).

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High resolution image (TIFF 9236 kb).
441_2019_3082_Fig7_ESM.png (304 kb)
Supplementary Figure S2

Transcriptional gene expressions of CXCL-8 (a, b), CYP24A1 (c, d) and CYP27B1 (e, f) in human keratinocytes (HaCaT cells) treated with tetracycline for 6 h and then challenged by T. phagedenis isolated from active (M2) DD lesions for 2 h (top row) and 24 h (bottom row) under aerobic conditions. Relative mRNA expression was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. P < 0.05 was considered significant (Student’s T test). (PNG 303 kb).

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High resolution image (TIFF 9148 kb).
441_2019_3082_Fig8_ESM.png (279 kb)
Supplementary Figure S3

Transcriptional gene expressions of IL-10 (a, b), CAMP (c, d) and IFN-γ (e, f) in human keratinocytes (HaCaT cells) treated with tetracycline for 6 h and then challenged by T. phagedenis isolated from active (M2) DD lesions for 2 h (top row) and 24 h (bottom row) under aerobic conditions. Relative mRNA expression was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. P < 0.05 was considered significant (Student’s T test). (PNG 279 kb).

441_2019_3082_MOESM3_ESM.tiff (8.9 mb)
High resolution image (TIFF 9164 kb).
441_2019_3082_Fig9_ESM.png (206 kb)
Supplementary Figure S4

Transcriptional gene expressions of TLR-2 (a, b) and TLR-4 (c, d) in human keratinocytes (HaCaT cells) treated with tetracycline for 6 h and then challenged by T. phagedenis isolated from active (M2) DD lesions for 2 h (top row) and 24 h (bottom row) under aerobic conditions. Relative mRNA expression was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. P < 0.05 was considered significant (T-test). (PNG 205 kb).

441_2019_3082_MOESM4_ESM.tiff (9.3 mb)
High resolution image (TIFF 9533 kb).
441_2019_3082_Fig10_ESM.png (260 kb)
Supplementary Figure S5

Transcriptional gene expressions of IL-10 (a, b), CAMP(c, d) and IFN-γ(e, f) in human keratinocytes (HaCaT cells) treated with vitamin D3 (VitD3) for 6 h and then challenged by T. phagedenis isolated from active (M2) DD lesions for 2 h (top row) and 24 h (bottom row) under aerobic conditions. Relative mRNA expression was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. P < 0.05 was considered significant (Student’s T test). (PNG 260 kb).

441_2019_3082_MOESM5_ESM.tiff (9.1 mb)
High resolution image (TIFF 9316 kb).
441_2019_3082_Fig11_ESM.png (196 kb)
Supplementary Figure S6

Transcriptional gene expressions of TLR-2 (a, b) and TLR-4 (c, d) in human keratinocytes (HaCaT cells) treated with vitamin D3 (VitD3) for 6 h and then challenged by T. phagedenis isolated from active (M2) DD lesions for 2 h (top row) and 24 h (bottom row) under aerobic conditions. Relative mRNA expression was determined by RT-qPCR. Means + the standard error of the mean (SEM) are presented. P < 0.05 was considered significant (Student’s T test). (PNG 195 kb).

441_2019_3082_MOESM6_ESM.tiff (9 mb)
High resolution image (TIFF 9180 kb).


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Production Animal Health, Faculty of Veterinary MedicineUniversity of CalgaryCalgaryCanada
  2. 2.Department of Veterinary Clinical and Diagnostic Science, Faculty of Veterinary MedicineUniversity of CalgaryCalgaryCanada

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