Immunohistochemistry and real-time PCR as diagnostic tools for detection of Borrelia burgdorferi sensu lato in ticks collected from humans

Abstract

The objective of this study was to evaluate different methods used for detection of Borrelia burgdorferi sensu lato (s.l.) in ticks: immunohistochemistry followed by focus floating microscopy (FFM) and real-time polymerase chain reaction (real-time PCR) targeting the ospA and hbb genes. Additionally, an optimized ospA real-time PCR assay was developed with an integrated internal amplification control (IAC) for the detection of inhibition in the PCR assay and was validated as an improved screening tool for B. burgdorferi. One hundred and thirty-six ticks collected from humans in a hospital from Cluj-Napoca, Romania, were investigated regarding genus, stage of development and sex, and then tested by all three assays. A poor quality of agreement was found between FFM and each of the two real-time PCR assays, as assessed by concordance analysis (Cohen's kappa), whereas the agreement between the two real-time PCR assays was moderate. The present study argues for a low sensitivity of FFM and underlines that discordant results of different assays used for detection of B. burgdorferi in ticks are frequent.

Multiple Tick-Borne Pathogens in Ixodes ricinus Ticks Collected from Humans in Romania

“…So far, studies conducted in Romania have evaluated the prevalence of Borrelia spp., N. mikurensis, Anaplasma spp., and Rickettsia spp. in I. ricinus [4][5][6][7][8]. The only study with a large dataset elaborated in Romania has evaluated the infection prevalence of B. burgdorferi s.l.…”

“…The only study with a large dataset elaborated in Romania has evaluated the infection prevalence of B. burgdorferi s.l. in ticks collected from humans and in patients bitten by Borrelia-positive ticks [4,5].…”

A Canine Leptospirosis Clinical Case Due to Leptospira interrogans (Serogroup Icterohaemorrhagiae) in a Dog Kennel in Castelvetrano (Western Sicily, South Italy)

“…Real-time PCRs were carried out on the DNA extracted from the kidney fragment of the sick dog to detect DNA from Chlamydia spp. [ 28 ], Neospora caninum [ 29 ], Coxiella burnetii [ 30 ], Borrelia burgdorferi [ 31 , 32 ], and Toxoplasma gondii [ 33 ]. A nested PCR for T. gondii was also performed [ 34 ].…”

Development and validation of a multi-target TaqMan qPCR method for detection of Borrelia burgdorferi sensu lato

“…While few published studies aim to compare or evaluate the specificity and sensitivity of TaqMan qPCR methods used to detect B. burgdorferi s.l. species and most of them for diagnostic purposes on human clinical samples (Briciu et al, 2016, de Leeuw et al, 2014, Ferdin et al, 2010, Maes et al, 2017, Nunes et al, 2018, Venczel et al, 2016, there is still a need for more standardized and characterized detection/quantification approaches.…”