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

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.

1 . (canceled) 
     
     
         2 . A method for initiating antimicrobial sensitivity testing of cells in a sample, the method comprising:
 a. determining a microbial cell concentration in the sample;   b. diluting the sample using a sample dilution factor to generate a target sample;   c. determining microbial cell concentration in the target sample;   d. generating a non-linear dilution curve using the microbial cell concentration in the sample and the sample dilution factor;   e. determine a target dilution factor using the microbial cell concentration in the target sample and the non-linear dilution curve;   f. diluting the sample to a target concentration using the target dilution factor to create a dynamically diluted sample; and   g. initiating antimicrobial sensitivity testing on the dynamically diluted sample.   
     
     
         3 . The method of  claim 2  further comprising, preselecting the sample dilution factor. 
     
     
         4 . The method of  claim 2  further comprising, prior to initiating antimicrobial sensitivity testing the dynamically diluted sample is subject to a pre-growth step. 
     
     
         5 . The method of  claim 2  further comprising, using one or more subsamples of the sample to generate the non-linear dilution curve. 
     
     
         6 . The method of  claim 5 , further comprising diluting a first subsample using a first dilution factor and diluting a second subsample using a second dilution factor wherein the first dilution factor and second dilution factor are different. 
     
     
         7 . The method of  claim 5 , further comprising creating dilution test points for a first subsample, wherein a first dilution test point comprises a first subsample microbial cell concentration and a first sample dilution factor and a second dilution test point comprises a second subsample microbial cell concentration and a second sample dilution factor. 
     
     
         8 . A method for initiating antimicrobial sensitivity testing of cells in a sample, the method comprising:
 a. splitting the sample into a first subsample and a second subsample;   b. determining a microbial cell concentration in the first subsample and second sub sample;   c. diluting the first subsample using a first sample dilution factor and diluting the second subsample using a second sample dilution factor to generate a first target sample and a second target sample wherein the first sample dilution factor and the second dilution factor are different;   d. determining microbial cell concentration in the first target sample and the second target sample;   e. generating a non-linear dilution curve using the microbial cell concentration in the first subsample and second subsample, the first dilution factor, and second dilution factor;   f. determining a first target dilution factor for the first target sample using the microbial cell concentration from the first target sample and the second target sample and the non-linear dilution curve;   g. diluting the first subsample using the first target dilution factor to create a dynamically diluted first sample; and   h. initiating antimicrobial sensitivity testing on the dynamically diluted first sample.   
     
     
         9 . The method of  claim 8  further comprising, determining the cell concentration of the dynamically diluted first sample. 
     
     
         10 . The method of  claim 8 , wherein determining a microbial cell concentration in the first subsample and second subsample does not involve plating. 
     
     
         11 . The method of  claim 8 , wherein the dynamically diluted first sample has an ideal dilution point at a midpoint of an upper limit and a lower limit of a target range for a microbe. 
     
     
         12 . The method of  claim 8 , wherein the first subsample and the second subsample have different cell concentrations. 
     
     
         13 . The method of  claim 8  further comprising, adjusting the target dilution factor with a growth factor. 
     
     
         14 . The method of  claim 8 , wherein creating a dynamically diluting first sample comprises:
 i. centering cell counts of a first microbial species and a second microbial species between the upper and lower limits of an inoculum.   
     
     
         15 . The method of  claim 8 , wherein creating a dynamically diluting first sample comprises:
 i. diluting the sample so that cell counts for a first microbial species is diluted to a midpoint of an inoculum upper and lower limits and cell counts for a second microbial species are diluted to a point below a lower limit of an inoculum.   
     
     
         16 . The method of  claim 15 , further comprising
 i. culturing the sample;   ii. determining a target dilution factor;   iii. using the target dilution factor to dynamically dilute the sample wherein dynamically diluting the sample comprises centering the cell counts of the second microbial species between the upper and lower limits of the inoculum; and   iv. initiating antimicrobial testing on the second microbial species.   
     
     
         17 . A method for initiating antimicrobial sensitivity testing of cells in a sample, the method comprising:
 a. determining a target dilution factor usable in diluting the sample to a target cell concentration, the determining comprising:
 i. identifying a growth factor associated with a microorganism species in the sample; 
 ii. dividing the sample into one or more subsamples; 
 iii. determining a cell concentration of each of the one or more subsamples; 
 iv. diluting each of the one or more subsamples using an associated sample dilution factor unique to each of the one or more subsamples to obtain one or more diluted subsamples; 
 v. determining a cell concentration of each of the one or more diluted subsamples; 
 vi. creating a model of a non-linear dilution curve; and 
 vii. determining the target dilution factor for each diluted subsample; 
   b. diluting the sample using the determined target dilution factor to produce the final cell concentration; and   c. initiating antimicrobial testing.   
     
     
         18 . The method of  claim 17 , wherein the sample is divided into 3-5 subsamples. 
     
     
         19 . The method of  claim 17 , wherein creating a model of a non-linear dilution curve comprises using the cell concentration of each of the one or more subsamples and the associated sample dilution factors or determining an average of the cell concentration of each of the one or more subsamples using a proportionality constant. 
     
     
         20 . The method of  claim 19 , wherein the proportionality constant has a value less than 1 and greater than or equal to 0.5. 
     
     
         21 . The method of  claim 17 , wherein determining the target dilution factor for each diluted subsample comprises using the model of the non-linear dilution curve, the growth factor, and the one or more diluted subsamples concentrations or identifying a first dilution test point of the one or more diluted subsamples that is less than the target cell concentration, identifying a second dilution test point of the one or more diluted subsamples that is greater than or equal to the target cell concentration, and using an interpolation between the first and second dilution test points to determine the target dilution factor.