Stimulated Immune Response by TruCulture® Whole Blood Assay in Patients With European Lyme Neuroborreliosis: A Prospective Cohort Study

Abstract

Introduction: Borrelia burgdorferi sensu lato complex (B. burgdorferi) can cause a variety of clinical manifestations including Lyme neuroborreliosis. Following the tick-borne transmission, B. burgdorferi initially evade immune responses, later symptomatic infection is associated with occurrence of specific antibody responses. We hypothesized that B. burgdorferi induce immune hyporesponsiveness or immune suppression and aimed to investigate patients with Lyme neuroborreliosis ability to respond to immune stimulation.

Methods: An observational cohort study investigating the stimulated immune response by standardized whole blood assay (TruCulture^®) in adult patients with Lyme neuroborreliosis included at time of diagnosis from 01.09.2018-31.07.2020. Reference intervals were based on a 5-95% range of cytokine concentrations from healthy individuals (n = 32). Patients with Lyme neuroborreliosis and references were compared using Mann-Whitney U test. Heatmaps of cytokine responses were generated using the webtool Clustvis.

Results: In total, 22 patients with Lyme neuroborreliosis (19 definite, 3 probable) were included. In the unstimulated samples, the concentrations of cytokines in patients with Lyme neuroborreliosis were comparable with references, except interferon (IFN)-α, interleukin (IL)-17A, IL-1β and IL-8, which were all significantly below the references. Patients with Lyme neuroborreliosis had similar concentrations of most cytokines in all stimulations compared with references. IFN-α, IFN-γ, IL-12 and IL-17A were lower than references in multiple stimulations.

Conclusion: In this exploratory cohort study, we found lower or similar concentrations of circulating cytokines in blood from patients with Lyme neuroborreliosis at time of diagnosis compared with references. The stimulated cytokine release in blood from patients with Lyme neuroborreliosis was in general slightly lower than in the references. Specific patterns of low IL-12 and IFN-γ indicated low Th1-response and low concentrations of IL-17A did not support a strong Th17 response. Our results suggest that patients with Lyme neuroborreliosis elicit a slightly suppressed or impaired immune response for the investigated stimulations, however, whether the response normalizes remains unanswered.

Keywords: Borrelia burgdorferi sensu lato complex; Lyme neuroborreliosis; TruCulture; innate immune system; stimulated immune response; whole blood assay.

Figure 1

Heatmap of individual cytokine response signatures in patients with LNB (red) and references (blue). The stimulations are divided and shown horizontally, HKCA (red), LPS (blue), PolyI:C (purple), R848 (orange) and Null (green). The individual response is shown in vertical columns and every square represent the cytokine response for the given stimulation. Column (diagnosis) dendrograms were drawn based on hierarchical clustering analysis using the complete agglomeration method on Euclidian distance matrices and Ward linkage.

Figure 2

Cytokine concentration in unstimulated (A) and stimulated blood samples (B–E) from patients with LNB. The gray area represents the reference interval and every concentration is shown individually as color coded dots. Comparison between cytokine concentration in blood from patients with LNB and the reference. *p < 0.05. **p < 0.001. (A) NULL (B) HCKA (C) LPS (D) POLY I:C (E) R848.

Figure 3

Heatmap of individual cytokine response signatures in blood from patients with LNB (blue), bacterial CNS infections (red), viral meningitis (purple) and references (green). The stimulations are divided and shown horizontally, HKCA (red), LPS (blue), PolyI:C (purple), R848 (orange) and Null (green). The individual response is shown in vertical columns and every square represent the cytokine response for the given stimulation. Column (diagnosis) dendrograms were drawn based on hierarchical clustering analysis using the complete agglomeration method on Euclidian distance matrices and Ward linkage.

Figure 4

Cytokine concentration in unstimulated blood samples from patients with other bacterial CNS infections (light blue), viral CNS infections (green), patients with LNB (dark blue) and references (grey). Bacterial CNS-infections was compared with LNB (grey stars) and bacterial CNS-infections was compared with references (black stars). *p < 0.05.