An improved peptide-ELISA protocol for serological identification of Borrelia burgdorferi sensu stricto, B. garinii and B. afzelii

The performance of 21 synthetic peptides for serological identification of Borrelia burgdoferi sensu stricto, B. garinii and B. afzelii was assessed in this study using two ELISA protocols based on (1) conventional passive binding onto 96-well Greiner Microloon ® 600 High-binding microplates and (2) covalent binding onto 96-well surfaced-activated Nunc™ Immobilizer Amino Plates. Sensitivity, specificity and accuracy was initially assessed testing 13 follow-up positive sera from seroconverted patients bitten by ticks that had tested positive for Borrelia burgdorferi sensu stricto ( N = 2), B. garinii (N = 2), B. afzelii ( N = 9). The optimized protocol was then applied to two cohorts of samples including plasma from Danish patients with Lyme borreliosis (LB) ( N = 32) and positive samples from Danish healthy blood donors (HBD) ( N = 44). Use of covalent binding resulted in higher OD values and significantly increased the accuracy of the test. Results of the seroprevalence study applied to LB and HBD samples showed different distribution of the three Bb sl. Borrelia burgdorferi sensu stricto ( Bb ss), B. garinii and B. afzelii were identified in 3 out 24 (12.5%), 13 out of 24 (54.2%) and 7 out of 24 (29.2%) LB samples, respectively. B. garinii was confirmed to be to the most prevalent species also in the second cohort of HBD samples as it was identified in 20 out 32 samples (62.5%) followed by Borrelia burgdorferi sensu stricto and B. afzelii and which were identified in 5 out of 32 (15.6%) and 4 out of 32 (12.5%) samples, respectively.