# US 2024/0200151 A1 · Metagenomic next-generation sequencing of microbial cell-free nucleic acids in subjects with lyme disease > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2024/0200151 A1 - Document ID: US20240200151A1 - Status: Published application - Kind: A1 - Application number: 18/365,876 - Filed: 2023-08-04 - Published: 2024-06-20 - Earliest claimed: 2021-02-12 - Assignees: Karius, Inc.; The Research Foundation for The State University of New York - Applicants: Karius, Inc.; The Research Foundation for The State University of New York - Inventors: Asim A. Ahmed; Carine Ho; Desiree D. Hollemon; Andrew Handel; Christy Beneri - Web page: https://patentgrep.com/patent/US20240200151A1 ## CPC - C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) - C12Q 1/689: for bacteria - C12N 15/1034: {Isolating an individual clone by screening libraries} - C12Q 2600/112: Disease subtyping, staging or classification - Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change - C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE ## Abstract What is disclosed herein is a method of detecting and treating Lyme disease, particularly Lyme arthritis. The method includes metagenomic next-generation sequencing (mNGS) of plasma microbial cell-free DNA (mcfDNA) to detect and quantify Borrelia spp. in the circulation. Also disclosed is a method of treating Lyme arthritis by treating the patient with an anti-microbial treatment until Borrelia mcfDNA is lowered to below detectable levels, such as less than 1 molecule per microliter of plasma. ## Claims ### Claim 1 A method of treating a Borrelia spp. infection in a subject that has received a diagnosis of the Borrelia spp. infection, wherein the diagnosis of the Borrelia spp. infection was based at least in part on [[the]]a method comprising: a. collecting one or more blood samples from the subject at a time when the subject does not have an erythema migrans (EM) rash and wherein the one or more blood samples comprise microbial cell-free nucleic acids (mcfNA); b. detecting mcfNA from Borrelia spp. in the one or more blood samples; and c. diagnosing the subject with the Borrelia spp. infection based at least in part on the detecting the mcfNA from the Borrelia spp. ### Claim 2 (depends on 1) The method of claim 1, further comprising quantifying the mcfNA from Borrelia spp. ### Claim 3 (canceled) ### Claim 4 (depends on 1) The method of claim 1, wherein the one or more blood samples are one or more plasma samples. ### Claim 5 (depends on 1) The method of claim 1, further comprising attaching nucleic acid adapters to cell-free nucleic acids in the one or more blood samples to prepare a sequencing library comprising the mcfNA. ### Claim 6 (depends on 5) The method of claim 5, further comprising generating sequence reads from the sequencing library comprising the mcfNA, aligning the sequence reads to Borrelia spp. genomic sequences in a reference data set to obtain aligned sequence reads, and identifying the Borrelia spp. based on the aligned sequence reads. ### Claim 7 (depends on 1) The method of claim 1, the method further comprising wherein the treating comprises administering a therapeutic treatment to the subject to treat a Borrelia spp. infection. ### Claim 8 (depends on 7) The method of claim 7, wherein the therapeutic treatment is a Borrelia-directed therapy. ### Claim 9 (depends on 8) The method of claim 8, wherein the Borrelia -directed therapy is at least one therapy selected from the group consisting of: doxycycline, amoxicillin, cefuroxime axetil, ceftriaxone, and cefotaxime. ### Claim 10 (depends on 1) The method of claim 1, further comprising spiking the one or more blood samples with a known concentration of synthetic DNA. ### Claim 11 (depends on 4) The method of claim 4, further comprising spiking the one or more plasma samples with a known concentration of synthetic DNA. ### Claim 12 (canceled) ### Claim 13 (canceled) ### Claim 14 (canceled) ### Claim 15 (depends on 1) The method of claim 1, wherein the subject has arthritis of a joint. ### Claim 16 (depends on 15) The method of claim 15, wherein the joint comprises at least one joint selected from the group consisting of knee, elbow, temporomandibular joint, and hip. ### Claim 17 (depends on 1) The method of claim 1, wherein the subject is blood culture negative for Borrelia at the time of the collecting of the one or more blood samples. ### Claim 18 (depends on 1) The method of claim 1, wherein the subject is negative for Borrelia when measured by a polymerase chain reaction (PCR) test of a blood sample or synovial fluid sample of blood from the subject. ### Claim 19 (canceled) ### Claim 20 (depends on 1) The method of claim 1, wherein the subject was bitten by a tick carrying Borrelia bacteria at least 6 months prior to the collecting of the one or more blood samples. ### Claim 21 (canceled) ### Claim 22 (canceled) ### Claim 23 (depends on 1) The method of claim 1, wherein the subject is serologically positive for Borrelia antibodies. ### Claim 24 (canceled) ### Claim 25 (depends on 1) The method of claim 1, wherein the concentration of Borrelia mcfDNA is 1-1,000 molecules per microliter (MPM) of plasma. ### Claim 26 (depends on 1) The method of claim 1, wherein the subject has disseminated late-stage Lyme disease. ### Claim 27 (depends on 1) The method of claim 1, wherein a sensitivity of detecting the Borrelia mcfDNA is at least 60%. ### Claim 28 (depends on 1) The method of claim 1, wherein the Borrelia mcfNA comprises mcfNA derived from B. burgdorferi or B. maynoii bacteria. ### Claim at position 29 29-36. (canceled)