Abstract
Mass spectrometry enhanced by nanotechnology can achieve previously unattainable sensitivity for characterizing urinary pathogen-derived peptides. We utilized mass spectrometry enhanced by affinity hydrogel particles (analytical sensitivity = 2.5 pg/mL) to study tick pathogen-specific proteins shed in the urine of patients with (1) erythema migrans rash and acute symptoms, (2) post treatment Lyme disease syndrome (PTLDS), and (3) clinical suspicion of tick-borne illnesses (TBI). Targeted pathogens were Borrelia, Babesia, Anaplasma, Rickettsia, Ehrlichia, Bartonella, Francisella, Powassan virus, tick-borne encephalitis virus, and Colorado tick fever virus. Specificity was defined by 100% amino acid sequence identity with tick-borne pathogen proteins, evolutionary taxonomic verification for related pathogens, and no identity with human or other organisms. Using a cut off of two pathogen peptides, 9/10 acute Lyme Borreliosis patients resulted positive, while we identified zero false positive in 250 controls. Two or more pathogen peptides were identified in 40% of samples from PTLDS and TBI patients (categories 2 and 3 above, n = 59/148). Collectively, 279 distinct unique tick-borne pathogen derived peptides were identified. The number of pathogen specific peptides was directly correlated with presence or absence of symptoms reported by patients (ordinal regression pseudo-R2 = 0.392, p = 0.010). Enhanced mass spectrometry is a new tool for studying tick-borne pathogen infections.
Highlights
Mass spectrometry enhanced by nanotechnology can achieve previously unattainable sensitivity for characterizing urinary pathogen-derived peptides
Patients enrolled in this study fall into five categories: (1) patients with acute stage Borrelia infection defined by a two-tier serology criteria; (2) symptomatic patients with a clinical diagnosis of PTLDS4,13; (3) patients treated in community centers and private practices with clinical suspicion of tick-borne illnesses but in the absence of complete clinical information regarding previous symptoms and treatments; (4) diseased controls, which include patients harboring non-tick-borne infections, who are hospitalized in Peru, a geographic region where ticks are very rare, and U.S patients with a diagnosis of traumatic brain injury and acute respiratory distress syndrome; and (5) healthy controls
In the training phase of the study, we analyzed urinary pathogen peptides in N = 10 patients suffering from early stage Lyme borreliosis and diag
Summary
Mass spectrometry enhanced by nanotechnology can achieve previously unattainable sensitivity for characterizing urinary pathogen-derived peptides. We utilized mass spectrometry enhanced by affinity hydrogel particles (analytical sensitivity = 2.5 pg/mL) to study tick pathogen-specific proteins shed in the urine of patients with (1) erythema migrans rash and acute symptoms, (2) post treatment Lyme disease syndrome (PTLDS), and (3) clinical suspicion of tick-borne illnesses (TBI). Recent molecular evidence suggested that post treatment persistence of Lyme arthritis symptoms maybe influenced by the persistence of Borrelia peptidoglycans in synovial fluid[17] Whether these biomolecules are derived from viable pathogens or persist in the body long after the infection has resolved remains to be determined. Specificity of the mass spectrometry analytic method is ensured by a three-tier authentication algorithm which requires stringent filters for peptide identification, 100% amino acid sequence identity with tick-borne pathogen proteins, evolutionary taxonomic verification for related pathogens, and lack of identity with human or non-tick-borne pathogenic organisms (Fig. S1). The correlation between the number of pathogen-specific urinary peptides and the presence or absence of symptoms as assessed by health care professionals is investigated
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