Use of ATP-Binding Cassette Subfamily A Member 13 (ABCA13) for Sensitive Detection of Focal Pathological Forms of Subclinical Bovine Paratuberculosis.

Cristina Blanco-Vázquez,Patricia Vázquez,Rosa Casais,Natalia Iglesias,Joseba M Garrido,Marta Alonso-Hearn,María Canive,Manuel A Queipo,Tania Iglesias,Ana Balseiro,Javier Amado,Ramón A Juste

doi:10.3389/fvets.2022.816135

Abstract

Bovine paratuberculosis (PTB) is a chronic enteritis caused by Mycobacterium avium subspecies paratuberculosis (Map) that causes a heavy economic impact worldwide. Map infected animals can remain asymptomatic for years while transmitting the mycobacteria to other members of the herd. Therefore, accurate detection of subclinically infected animals is crucial for disease control. In a previous RNA-Seq study, we identified several mRNAs that were overexpressed in whole blood of cows with different PTB-associated histological lesions compared with control animals without detected lesions. The proteins encoded by two of these mRNAs, ATP binding cassette subfamily A member 13 (ABCA13) and Matrix Metallopeptidase 8 (MMP8) were significantly overexpressed in whole blood of animals with focal histological lesions, the most frequent pathological form in the subclinical stages of the disease. In the current study, the potential of sensitive early diagnostic tools of commercial ELISAs, based on the detection of these two biomarkers, was evaluated in serum samples of 704 Holstein Friesian cows (566 infected animals and 138 control animals from PTB-free farms). For this evaluation, infected animals were classified into three groups, according to the type of histological lesions present in their gut tissues: focal (n = 447), multifocal (n = 59), and diffuse (n = 60). The ELISA based on the detection of ABCA13 was successfully validated showing good discriminatory power between animals with focal lesions and control animals (sensitivity 82.99% and specificity 80.43%). Conversely, the MMP8-based ELISA showed a poor discriminatory power between the different histological groups and non-infected controls. The ABCA13-based ELISA showed a higher diagnostic value (0.822) than the IDEXX ELISA (0.517), the fecal bacterial isolation (0.523) and the real-time PCR (0.531) for the detection of animals with focal lesions. Overall, our results indicate that this ABCA13 ELISA greatly improves the identification of subclinically infected animals with focal lesions that are undetectable using current diagnostic methods.

Highlights

Bovine paratuberculosis (PTB) or Johne’s disease (JD) is a chronic contagious and debilitating enteritis of domestic and wild ruminants caused by Mycobacterium avium subspecies paratuberculosis (Map)
Receiver operator characteristic (ROC) analysis of the MMP8-based Enzyme-Linked Immunosorbent Assay (ELISA) was performed using 698 samples, 442 were from animals with focal lesions, 58 with multifocal, 60 with diffuse, and 138 from negative controls (6 sera were only analyzed by the ABCA13-based ELISA as we did not have enough volume to perform both ELISAs)
ABCA13, bovine ATP binding cassette subfamily A member 13; MMP8, bovine matrix metallopeptidase 8; STD, standard deviation; Control, refers to the PTB-free control group; The concentration values are expressed as ng/mL The asterisks indicate if the differences between each histopathological group and the control are or not significant (***p < 0.001)

Summary

Introduction

Bovine paratuberculosis (PTB) or Johne’s disease (JD) is a chronic contagious and debilitating enteritis of domestic and wild ruminants caused by Mycobacterium avium subspecies paratuberculosis (Map). PTB produces important economic losses in dairy herds worldwide due to increase in mortality, decrease in milk production, weight loss, premature culling and reduced slaughter value [1–4]. PTB has been related to reduced fertility rates [5, 6] and increased susceptibility to other diseases, mammary infections [7]. The importance of this disease would be even greater when considering its zoonotic potential and the risk of transmission of viable Map through pasteurized milk and milk products [8–10]. Access to contaminated pasture or water sources and contact with other ruminants may be involved in the spread of the disease [18–20]

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Results