Instrument and system for rapid microorganism identification and antimicrobial agent susceptibility testing

• US 10,253,355 B2
• Filed: 10/03/2016
• Issued: 04/09/2019
• Est. Priority Date: 03/30/2015
• Status: Active Grant

• Current Assignee: Accelerate Diagnostics, Inc.
• Original Assignee: Accelerate Diagnostics, Inc.
• Inventors: Richards, William L., Ashby, Austin, Ketterer, Matthew, Marshall, Kevin, Harrison, Josh, Mette, Matthew, Richards, Paul, Showalter, Wayne, Cote, Jasmin, Halbert, Phillip C., Bourgeois, Solene, Metzger, Steven W., Hance, Ken, Mensack, Meghan, Michel, Carlos, Allers, Elke, Gamage, Dulini, Prisbrey, Landon, Gusyatin, Oleg, Shamsheyeva, Alena, Turng, Ben, Ghusson, Andrew, Reinhardt, Kurt
• Primary Examiner(s): Bhat, Narayan K
• Application Number:
  US15/283,922
• Publication Number:
  US 20170023599A1
• Time in Patent Office: 918 Days
• CPC Class Codes: B01L 3/545 for laboratory containers
• CPC Class Codes: C12M 41/36 of biomass, e.g. colony counters or by turbidity measurements electrooptical investigation of individual particles G01N15/14, flow cytometers G01N15/1404
• CPC Class Codes: C12Q 1/025 for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics antimicrobial activity C12Q1/18
• CPC Class Codes: C12Q 1/6841 In situ hybridisation
• CPC Class Codes: G01N 2021/6419 Excitation at two or more wavelengths
• CPC Class Codes: G01N 2035/00158 Elements containing microarrays, i.e. "biochip"
• CPC Class Codes: G01N 2035/00346 Heating or cooling arrangements
• CPC Class Codes: G01N 2035/1032 Dilution or aliquotting
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G01N 35/00069 whereby the sample substrate is of the bio-disk type, i.e. having the format of an optical disk
• CPC Class Codes: G01N 35/025 having a carousel or turntable for reaction cells or cuvettes
• CPC Class Codes: G01N 35/1002 Reagent dispensers
• CPC Class Codes: G02B 7/28 Systems for automatic generation of focusing signals

Abstract

A system for automated microorganism identification and antibiotic susceptibility testing comprising a reagent cartridge, a reagent stage, a cassette, a cassette, stage, a pipettor assembly, an optical detection system, and a controller is disclosed. The system is designed to dynamically adjust motor idle torque to control heat load and employs a fast focus process for determining the true focus position of an individual microorganism. The system also may quantify the relative abundance of viable microorganisms in a sample using dynamic dilution, and facilitate growth of microorganisms in customized media for rapid, accurate antimicrobial susceptibility testing. Automated quality control test components and methods of their use are also disclosed.

Claims

1. An automated antibiotic susceptibility testing (AST) quality control (QC) component, comprising:

a frame including a plurality of wells;

at least one clip attached to the frame, wherein the at least one clip includes at least one clip that interacts with a reagent cartridge;

at least one microbial isolate in at least one of the plurality of wells; and

a means of identifying the component as an AST QC test by an automated system.

2. The AST QC component of claim 1, wherein the at least one clip includes at least one clip that-interacts with a microbial identification QC component.

3. The AST QC component of claim 1, wherein the plurality of wells comprises 8-12 wells.

4. The AST QC component of claim 1, wherein the microbial isolate comprises Escherichia coli, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus, or Candida albicans.

5. The AST QC component of claim 4, wherein the E. coli is E. coli ATCC 25922 and/or E. coli ATCC 35218, the Pseudomonas aeruginosa is P. aeruginosa ATCC 27853, the Enterococcus faecalis is E. faecalis ATCC 29212, the Staphylococcus aureus is S. aureus ATCC 29213, S. aureus ATCC 43300, and/or S. aureus ATCC BAA-977, and/or the Candida albicans is C. albicans ATCC 96268.

6. The AST QC component of claim 1, wherein the means of identifying the component as an AST QC test comprises a bar code or an RFID tag.

12. A kit comprising one or more automated AST QC components of claim 1.

13. The kit of claim 12, further comprising one or more automated microbial identification (ID) quality control (QC) components comprising:

a frame including a plurality of wells;

at least one clip attached to the frame;

at least one target microbial nucleic acid molecule in at least one of the plurality of wells, wherein the one or more microbial ID QC components are configured to interlock with the one or more AST QC components; and

a means of identifying the one or more microbial ID QC components as a microbial ID QC test by an automated system.

14. A method of performing automated quality control (QC) testing, comprising:

(a) attaching the AST QC component of claim 1 to a reagent cartridge, wherein the reagent cartridge comprises;

a plurality of wells comprising reagents; and

an opening,

wherein the plurality of wells comprising reagents comprise at least one well comprising a probe and at least one well comprising an antimicrobial agent;

(b) inserting the reagent cartridge with the attached AST QC component into a system, wherein the system comprises;

a reagent stage comprising an annular shape defining an interior opening, wherein the reagent stage is configured to rotate in a first plane and wherein the reagent stage is configured to receive the reagent cartridge;

a cassette comprising a plurality of microfluidic sample channels structured to receive samples, each of the plurality of microfluidic sample channels comprising an inlet port configured to receive a pipette tip;

a cassette stage located within the interior opening of the reagent stage, wherein the cassette stage is configured to rotate and move laterally in the first plane, and wherein the cassette stage is configured to receive the cassette;

a pipettor assembly configured to move a plurality of reagents between the plurality of wells of the reagent cartridge and the inlet ports of each of the plurality of microfluidic sample channels;

an optical scanning device configured to identify a QC component;

an optical detection system configured to obtain darkfield and fluorescence photomicrographs of one or more microorganisms contained in the plurality of microfluidic sample channels; and

a controller configured to direct operation of the system and process microorganism information derived from photomicrographs obtained by the optical detection system;

(c) introducing one or more microbial isolates from the AST QC component to the plurality of microfluidic channels of the cassette of the system;

(d) growing the one or more microbial isolates in the presence or absence of one or more antimicrobial agents;

(e) monitoring the growth of the microbial isolates over time;

(f) determining susceptibility or resistance of the one or more microbial isolates to the one or more antimicrobial agents; and

(g) reporting accuracy of the microbial isolate antimicrobial susceptibility or resistance provided by the system.

15. The method of claim 14, wherein determining susceptibility or resistance of the one or more microbial isolates to the one or more antimicrobial agents further comprises qualitatively or quantitatively measuring growth or lysis of the one or more microbial isolates in the presence of the one or more antimicrobial agents compared to a control that is not exposed to the antimicrobial agents.

16. The method of claim 14, wherein the one or more microbial isolates are grown in Mueller-Hinton nutrient-depleted media or are grown in 1% phytone tryptose Mueller-Hinton agar.

17. The method of claim 14, wherein the one or more antimicrobial agents comprise one or more of amikacin, ampicillin, ampicillin-sulbactam, aztreonam, cefazolin, cefepime, ceftaroline, ceftazidime, ceftriaxone, ciprofloxacin, colistin, daptomycin, doxycycline, erythromycin, ertapenem, gentamicin, imipenem, linezolid, meropenem, minocycline, piperacillin-tazobactam, tobramycin, trimethoprim-sulfamethoxazole and vancomycin.

18. The method of claim 14, further comprising:

(a) attaching a microbial identification (ID) quality control (QC) component to the reagent cartridge prior to inserting the reagent cartridge in the system, wherein the microbial ID QC component comprises;

a frame including a plurality of wells;

at least one clip attached to the frame;

at least one target microbial nucleic acid molecule in at least one of the plurality of wells, wherein the microbial ID QC component is configured to interlock with the AST QC component; and

a means of identifying the component as a microbial ID QC test by an automated system;

(b) introducing the at least one target microbial nucleic acid molecules into the plurality of microfluidic channels of the cassette;

(c) introducing one or more labeled target probes that recognize and bind sequences of the at least one target microbial nucleic acid molecule into the at least one of the plurality of microfluidic channels;

(d) incubating the labeled probes with the at least one target microbial nucleic acid molecule for a time sufficient to enable binding of the probes to target microbial nucleic acid molecules; and

(e) detecting a signal from the one or more labeled target probes, wherein presence of a signal indicates a positive ID result and absence of a signal indicates a negative ID result; and

(f) reporting accuracy of microbial identification provided by the system.

19. The method of claim 18, wherein the at least one target microbial nucleic acid molecule is coupled to a microsphere.

20. The method of claim 18, wherein the at least one target microbial nucleic acid molecule is complementary to a nucleic acid from one or more of Staphylococcus aureus, Staphylococcus lugdunensis, coagulase-negative Staphylococcus species, Enterococcus faecalis, Enterococcus faecium, Streptococcus pneumoniae, Streptococcus agalactiae, Streptococcus spp., Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella spp., Escherichia coli, Enterobacter spp., Proteus spp., Citrobacter spp., Serratia marcescens, Candida albicans and Candida glabrata.

21. The method of claim 14, wherein the automated quality control method achieves about a 20% to about a 30% improvement in the effective pass rate over a manual quality control test procedure.

22. The method of claim 18, wherein the automated quality control method achieves about a 20% to about a 30% improvement in the effective pass rate over a manual quality control test procedure.

7. An automated microbial identification (ID) quality control (QC) component, comprising

a frame including a plurality of wells;

at least one clip attached to the frame;

at least one target microbial nucleic acid molecule in at least one of the plurality of wells, wherein the microbial ID QC component is configured to interlock with an antibiotic susceptibility QC component; and

a means of identifying the component as a microbial ID QC test by an automated system.

8. The microbial ID QC component of claim 7, wherein the at least one clip interacts with an antibiotic susceptibility testing QC component.

9. The microbial ID QC component of claim 7, wherein the plurality of wells comprises 11-13 wells.

10. The microbial ID QC component of claim 7, wherein the at least one target nucleic acid molecule is from Acinetobacter baumannii, Citrobacter spp., Enterobacter spp., Escherichia coli, Klebsiella spp., Proteus spp., Pseudomonas aeruginosa, Serratia marcescens, Coagulase-negative Staphylococcus spp., Enterococcus faecalis, Enterococcus faecium, Staphylococcus aureus, Staphylococcus lugdunensis, Streptococcus spp., Candida albicans, or Candida glabrata.

11. The microbial ID QC component of claim 7, wherein the means of identifying the component as a microbial ID QC test comprises a bar code or an RFID tag.