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Rapid antibiotic susceptibility testing

一种抗生素、药敏的技术，应用在抗生素药敏性的快速测试领域

Active Publication Date: 2015-01-14
PRESIDENT & FELLOWS OF HARVARD COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, current practice (i.e. blood cultures) takes more than two days to get an answer, which often proves to be excessive

Method used

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  • Rapid antibiotic susceptibility testing
  • Rapid antibiotic susceptibility testing

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0438] Example 1 - Rapid Antibiotic Susceptibility Testing Based on Magnetic Sorting and Microscopy

[0439] Bacteremia, a condition also known as bacterial sepsis or blood poisoning, which describes a bacterial infection of a patient's blood, is a major killer in the United States and worldwide. Worldwide, there are 18 million cases of sepsis each year, resulting in more than 6 million deaths; in the United States alone, there are 750,000 cases a year, resulting in more than 200,000 deaths. When doctors suspect a patient has bacteremia, they must act quickly: Because bacteria can divide so rapidly, every hour lost before the right treatment is given can make a substantial difference in patient outcomes. So doctors must quickly answer two questions: whether the patient does have bacteremia, and if so, what antibiotics to prescribe. Unfortunately, existing methods for answering these questions (blood culture) take two days or more to get an answer, which often proves to be too...

Embodiment 2

[0450] Example 2 - Rapid Antibiotic Susceptibility Testing Based on Magnetic Sorting and ELISA

[0451] The inventors developed methods for the rapid isolation, detection and determination of antibiotic susceptibility for bacteremia or other microbial infections using magnetic sorting and ELISA / metabolic readout. This provides a technique for detecting bacteremia and determining the antibiotic resistance profile of causative agents in hours or less.

[0452] In general, the method involves: (i) extracting and concentrating pathogens from blood using functionalized magnetic beads; (ii) dividing pathogens into subsamples and incubating with antibiotic-supplemented medium (not for ELISA). Yeast extract-containing media, or other media for luciferase-based assays); and (iii) luciferase detection assays using ELISA assays (enzyme-linked immunosorbent assays) or ATP production method (e.g. BACTITER-GLO from Promega (Cat No. G8230) TM microbial cell viability assay) to detect patho...

Embodiment 3

[0464] Example 3 - Plate Capture / Natural Growing Bacteria Can Produce Artificially Low Counts

[0465] Determine capture efficiency of Akt-FcMBL beads by plate count:

[0466] -5 μl Akt-FcMBL beads + 10 μl S. aureus or E. coli (1 mL TBST-Ca 2+ middle);

[0467] -Hula 10min;

[0468] - in TBST-Ca when on magnet 2+ 1 wash in medium.

[0469] count:

[0470] - Number of bacteria added (input), washed bead supernatant (uncaptured) and bead fraction (captured).

[0471] like Figure 8 As shown, plating bead fractions yields an artificially low readout of bacterial counts. Therefore, non-plated readouts (eg Fc-MBL ELISA, metabolic assays - ATP luminescence) may be more useful for measuring growth of bacteria isolated with Akt-FcMBL beads.

[0472] Without wishing to be bound by theory, the FcMBL beads bound all bacteria. However, they clump together so that standard techniques of plating and colony counting may not be feasible for quantification of captured pathogens, with ...

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Abstract

Embodiments of various aspects described herein are directed to methods, compositions, kits and systems for rapid determination of antibiotic susceptibility of a microbe within hours after a sample is collected. In some embodiments, the methods, compositions, kits and systems described herein can allow determination of antibiotic susceptibility of a microbe based on a small number of microbes, e.g., as few as 5-10 microbes bound to a microbe-targeting substrate described herein.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS [0002] Pursuant to 35 U.S.C. §119(e), this application claims priority to US Provisional Application No. 61 / 604,878, filed February 29, 2012, and US Provisional Application No. 61 / 647,860, filed May 16, 2012, by reference is incorporated in its entirety into this article. [0003] governmental support [0004] This invention was made with government support under Grant No. N66001-11-1-4180 awarded by DARPA. The United States Government has certain rights in this invention. technical field [0005] The present disclosure relates to methods, compositions and kits for the rapid determination of antibiotic susceptibility of microbes within hours of sample collection. Background technique [0006] Each year, more than 18 million patients suffer from sepsis caused by systemic blood-borne infections, and more than 6 million of these people die. Mortality from sepsis in intensive care units worldwide ranges from 20% to 60%, and on...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/18C12Q1/04C12Q1/02
CPCC12Q1/18G01N33/56911G01N2333/245G01N2333/31Y02A90/10G01N33/56916G01N33/56938G01N2500/10
Inventor 米歇尔·舒普尔唐纳德·E·英格贝尔马克·J·卡特赖特亚历山大·沃特斯约翰·塞缪尔·沃克曼丹尼尔·勒文尼尔马丁·洛特曼
Owner PRESIDENT & FELLOWS OF HARVARD COLLEGE
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