Targeting Multicopy Prophage Genes for the Increased Detection of Borrelia burgdorferi Sensu Lato (s.l.), the Causative Agents of Lyme Disease, in Blood

Abstract

The successful treatment of Lyme disease (LD) is contingent on accurate diagnosis. However, current laboratory detection assays lack sensitivity in the early stages of the disease. Because delayed diagnosis of LD incurs high healthcare costs and great suffering, new highly sensitive tests are in need. To overcome these challenges, we developed an internally controlled quantitative PCR (Ter-qPCR) that targets the multicopy terminase large subunit (terL) gene encoded by prophages that are only found in LD-causing bacteria. The terL protein helps phages pack their DNA. Strikingly, the detection limit of the Ter-qPCR was analytically estimated to be 22 copies and one bacterial cell in bacteria spiked blood. Furthermore, significant quantitative differences was observed in terms of the amount of terL detected in healthy individuals and patients with either early or late disease. Together, the data suggests that the prophage-targeting PCR has significant power to improve success detection for LD. After rigorous clinical validation, this new test could deliver a step-change in the detection of LD. Prophage encoded markers are prevalent in many other pathogenic bacteria rendering this approach highly applicable to bacterial identification in general.

Induced Native Phage Therapy for the Treatment of Lyme Disease and Relapsing Fever: A Retrospective Review of First 14 Months in One Clinic

“…Due to the very high specificity of native phages to their host bacteria, Inducen-LD/RF appears to eliminate the infection based upon the understanding of the death of native phages, if there are no host bacteria present to continue replicating more phages. The sustained disappearance of Borrelia phages for as long as 26 weeks, as seen in this review, strongly suggests the complete elimination of the targeted infection [ 12 ]. With the rapid clearance of the infection by the native phages, we observed the typical clinical objectives for treatment rapidly shifted towards correcting the many consequences the infection inflicted upon the patient.…”

“…Bacteriophages are the most abundant viruses in the body [ 11 ]. It is the prevalence of phages that has led to the development of extremely sensitive laboratory phage quantitative polymerase chain reaction (qPCR) testing, the PBP test, which was utilized for our study [ 12 ].…”

“…To make matters worse, the various strains of Borrelia , such as those associated with LD and RF, are morphologically similar and clinically present with almost indistinguishable symptoms. The PBP testing brings a much higher level of diagnostic and treatment confidence with its ability to identify the exact strain of Borrelia , whether early or late in the disease progression [ 12 ]. Treatment selection can be tailored to the specific type of infection, a very important fact since LD and RF require different medications.…”

Opinion: Methodological Shortcomings in the Study on a Prophage-based PCR Test for Lyme Borreliosis

“… Alignment, individuals values of copies/PCR, and ROC analysis of copies/PCR. (A) Sequences published by Amouriaux et al ( 1993 ) and by Shan et al ( 2021 ) (termed cp32-1 to cp32-9 and lp56, lines 2–9) were aligned in MEGA 5 (Tamura et al, 2011 ). The alignment shows that there is some overlap between these sequences.…”

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We read the article by Shan et al (2021) with great interest, as new diagnostic tests for Lyme borreliosis (LB) are urgently needed (Cruickshank et al, 2018;Dessau et al, 2018). The article represents a proof of principle paper and an initial validation of an already commercially available test [Phelix Phage Borrelia-R.E.D.

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