IP-10 Is a Sensitive Biomarker of Antigen Recognition in Whole-Blood Stimulation Assays Used for the Diagnosis of Mycobacterium bovis Infection in African Buffaloes (Syncerus caffer)

Abstract

African buffaloes (Syncerus caffer) are maintenance hosts of Mycobacterium bovis, the causative agent of bovine tuberculosis. They act as reservoirs of this infection for a wide range of wildlife and domestic species, and the detection of infected animals is important to control the geographic spread and transmission of the disease. Interferon gamma (IFN-␥) release assays (IGRAs) utilizing pathogen-derived peptide antigens are highly specific tests of M. bovis infection; however, the diagnostic sensitivities of these assays are suboptimal. We evaluated the diagnostic utility of measuring antigen-dependent interferon gamma-induced protein 10 (IP-10) release as an alternative to measuring IFN-␥ levels. M. bovis-exposed buffaloes were tested using the Bovigam PC-EC and Bovigam PC-HP assays and a modified QuantiFERON TB-Gold (mQFT) assay. IP-10 was measured in the harvested plasma and was produced in significantly greater abundance in response to M. bovis antigens in Bovigam-positive than in Bovigam-negative animals. For each assay, using the Bovigam results as a reference, receiver operating characteristic curve analysis was done to determine diagnostically relevant cutoff values for IP-10. Thereafter, mQFT test results derived from measurement of IP-10 and IFN-␥ were compared and a larger number of Bovigam-positive animals were detected using IP-10 as a diagnostic marker. Moreover, using IP-10, agreement between the mQFT assay and the Bovigam assays was increased, while the excellent agreement between the Bovigam assays was retained. We conclude that IP-10 is a sensitive marker of antigen recognition and that measurement of this cytokine in antigen-stimulated whole blood might increase the sensitivity of conventional IGRAs in African buffaloes. Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB) in a wide range of domestic animals and wildlife (1). BTB in cattle populations is intensively controlled in many countries as it can result in reduced productivity or death of infected animals and poses a serious zoonotic risk. In South Africa, the African buffalo (Syncerus caffer) is a maintenance host of M. bovis, and the early detection of infected animals is important to control the transmission of the pathogen to other wildlife and domestic species and to prevent the geographic spread of this disease by translocation (2, 3).The most sensitive method for diagnosing M. bovis infection is by detection of the host's cell-mediated immune response to pathogen-specific antigens (4). Examples of such tests are the in vivo tuberculin skin test (TST) and in vitro interferon gamma (IFN-␥) release assays (IGRAs). The latter detect the release of interferon gamma in whole blood or from isolated peripheral blood mononuclear cells (PBMCs) in response to M. bovis purified protein derivative (PPD) (5) or to more specific antigens such as the 6-kDa early secreted antigenic target (ESAT-6) and the 10-kDa culture filtrate protein (CFP-10) (6, 7). Recently, IGRAs utilizing the latter antigens, i.e., the modified Quant...

In vitro responses of multiple cytokines to purified protein derivative in healthy and naturally Mycobacterium tuberculosis‐infected rhesus monkeys (Macaca mulatta)

“…14,[17][18][19] This study aimed to evaluate more cytokines as alternative biomarkers of whole-blood stimulation assays in diagnosing nonhuman Panel 1 6 TNF RII, IL19, RANTES, MIF, MPO, Galectin-3 Panel 2 37 IFN-γ, TNF-α, TNF RI, GM-CSF, IL1β, IL1RA, IL1 RII, IL2, IL2 Rα, IL4, IL5, IL6, IL6 Rα, IL8, IL10, IL15, IL17A, IL22, MCP-1, MIP-1β, MCP-3, MCP-4, CCL11, CCL18, CCL22, CCL27, CXCL1, CXCL4, IP-10, CXCL12, CXCL13, CD14, CD27, CD30 been proven to be robust biomarkers of whole-blood stimulation assays. [22][23][24] In our study, IFN-γ and IP-10 were also identified as sensitive markers for the PPD-based whole-blood stimulation assay in nonhuman primates, and IP-10 appeared to be more sensitive than…”

Interleukin-17A as a Biomarker for Bovine Tuberculosis

“…While TSTs are the mainstays for bovine TB antemortem testing, blood-based tests are attractive, as they require only a single visit to the farm, provide the opportunity for immediate repeat testing, and yield results that are less subjective than those of TSTs. Additional biomarkers have recently emerged as potential candidates for use in blood-based TB tests for humans (reviewed by Walzl et al [5] and Salgame et al [6]) and cattle, including interleukin-1␤ (IL-1␤), IL-2, tumor necrosis factor alpha (TNF-␣), nitric oxide, IL-17, and IFN-␥-induced protein 10 (IP-10) (7)(8)(9)(10)(11)(12)(13). Of these, IL-17 is particularly appealing as a biomarker for bovine TB, as IL-17A mRNA responses determined using real-time quantitative PCR (RT-qPCR) may be predictive of both vaccine efficacy (11,14,15) and lesion severity (7) when measured after vaccination and infection, respectively.…”

Development and evaluation of a diagnostic cytokine-release assay for Mycobacterium suricattae infection in meerkats (Suricata suricatta)

“…In both these species, IP-10 is produced in greater amounts than IFN-γ in antigen-stimulated whole blood and has been shown to be a more sensitive marker of mycobacterial infection [8, 9, 13]. In the present study, despite the use of ELISA antibodies produced against distantly related species, measurement of IP-10 proved to be a useful marker of immune activation.…”

“…In other species, such as cattle, in vitro tests of cell-mediated immunity (CMI) offer the most sensitive methods for TB diagnosis by detecting the release of interferon-gamma (IFN-γ) in response to mycobacterial antigens [6, 7]. Moreover, an alternative diagnostic marker of CMI, IFN-γ-inducible protein 10 (IP-10), has been shown to improve diagnostic sensitivity in humans and buffaloes ( Syncerus caffer ) [8, 9]. The specificity of such assays can be optimised by using highly specific antigens such as 6 kDa early secretory antigenic target (ESAT-6) and 10 kDa culture filtrate protein (CFP-10) [10, 11].…”