J Clin Pathol 1993;46:817-821

Evaluation of Questor urine screening system forbacteriuria and pyuria

M Stevens, C J Mitchell, S A Livsey, C A MacDonald

AbstractAius-To evaluate the Questor auto-mated bacteriuria and pyuria screeningsystem; to compare its performance withthat of a reference method; and to assessits usefulness in a routine clinical labora-tory.Methods-The Questor urine screeningsystem was compared with a comprehen-sive regimen to detect urinary tractinfection, using pour-plate viable countsto determine the numbers of bacteriapresent in urine samples, a wide range ofother cultural methods, microscopicfindings and clinical information.Results-The optimal performance indetecting significant growths was a sen-sitivity of 93%, a specificity of 74%, apositive predictive value of 43% and anegative predictive value of 98%. The listprice per test is £0.17 and the capital costof the system is £39 950. Questor can test50 samples an hour and can be operatedby one member of the laboratory staff,who is not required to make interpreta-tive judgments-for example, a medicallaboratory assistant.Conclusions-The sensitivity and speci-ficity of the Questor was better than thatobtained from other screening systemsusing the same protocol. The system waseasy to use and is a useful addition tothe methods available for screening forbacteriuria.

(J Clin Pathol 1993;46:817-821)

Figure 1 Complete Questor system with computer.

Urinary infections are second only in preva-lence to those of the upper respiratory tract.But the number of requests submitted to clin-ical microbiology laboratories for the micro-biological examination of urine far exceedsthe number of upper respiratory tract speci-mens. Bacteriuria may be chronic, persistent,and possibly influence structure and functionoutside the urinary tract. It has an importantrole in disease throughout life, from prema-turity to hypertension and renal failure.'Many methods for screening for bacteriuriahave been described and reviewed,2 but thereremains a need for an accurate, rapid, costeffective system for use in routine clinical lab-oratories. Questor (Difco Laboratories Ltd,East Molesey, Surrey) is an automated sys-tem which is designed to categorise and enu-merate bacteria and other cells found in urineusing particle counting technology.An evaluation of the system was commis-

sioned by the Medical Devices Directorate ofthe Department of Health and carried out atthe Leicester Public Health Laboratory.The system's performance was compared

with a comprehensive reference methodwhich included a variety of cultural tech-niques, microscopic examination, and clinicalinformation. A full report of the evaluationhas been produced.'

MethodsThe Questor system (fig 1) is an automatedsingle-channel impedance particle counterwhich uses a direct physical method to countand categorise particles. A 100 PI aliquot ofthe urine to be tested is pipetted into one wellof an eight-well disposable plastic tray whichis then placed in the instrument. The follow-ing automatic process is then carried out. Thespecimen is diluted with a conductive elec-trolyte solution and offered to a particlecounting probe (fig 2). There is a small ori-fice (A) in the glass end of the probe whichalso contains two platinum electrodes, oneinternal (B) and one external (C). An electri-cal field is established between the electrodesvia the electrolyte in the orifice. The sample isdrawn through the orifice under vacuum. Asa particle is drawn through the orifice andenters the electrical field, it displaces its ownvolume of electrolyte, causing a change in theimpedance of the electrical circuit. Thechanges are detected as pulses, the number ofwhich indicate the quantity of particles pre-sent in the sample.The height and area of the pulse is propor-

Department ofHealthEvaluation Unit,Public HealthLaboratory, The RoyalInfirmary, LeicesterLEI 5WWM StevensC J MitchellS A IivseyMedical DevicesDirectorateDepartment ofHealth, RussellSquare, LondonC A MacDonaldCorrespondence to:M StevensAccepted for publication7 April 1993

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Figure 2 Schematicrepresentation of theQuestor probe: (A) orifice;(B) internal electrode; (C)external electrode; (D)movable insert showingdirection of movement (E).

tional to the mean spherical volume of theparticle. A histogram of the particles and theirsizes is formed and analysed by the computersoftware to give enumeration of the particlesof different size bands. In Questor, the vol-ume of the orifice is varied by inserting atapered spike (the insert) (D) into and out ofthe orifice (A). Thus the characteristics of theorifice can be optimised for the size of theparticles of interest (from 0 5 to 40 jim). Theanalyser records the volume distributions forfour different particle size ranges correspond-ing to organisms, erythrocytes, leucocytes andepithelial cells.

Threshold settings to define a positiveresult can be selected by the user. If theorganism count is above the selected thresh-old-for example, 1.0 x 105-the interpreta-tion given is "positive". If the leucocyte countis above the selected threshold- .50/pl-theinterpretation given is "pos-white cell". If theerythrocyte count is above the selectedthreshold- ,250/pl-the interpretation givenis "pos-red blood cell". The user has theoption of choosing a threshold value for anycombination of organisms, leucocytes, anderythrocytes above which results are inter-preted as positive.A previous evaluation of the Ramus urine

screening system,4 which utilised particlecounting, showed that the fixed, small sizeorifice used was prone to blocking. In theQuestor system the manufacturers claim that,"putting the insert into the orifice forms anannulus. The flow pattern sets up a standingwave at the entrance of the annulus. Thiswave sweeps the large particles away from theannulus but allows the small particlesthrough. This is called the 'filter effect',which greatly reduces the incidence of probeblockage."

Apart from the Questor system, the itemsrequired for testing urines are plastic traysand particle-free isotonic diluent (both ofwhich are supplied by the manufacturer), a

micropipette and tips, plastic waste disposalbags, and computer consumables: paper,ribbons and floppy disks.

Evaluation methodsThe urine samples examined during the eval-uation were random, unselected, consecutivespecimens and were representative of a crosssection of the types submitted to the routineclinical laboratory. A total of 1023 urinespecimens were examined by Questor and bycomprehensive comparative methods. Of the1023, 1003 were received preserved in boricacid. The remaining 20 were unpreserved butwere refrigerated and tested by Questor andthe reference methods concurrently.A pour-plate viable bacterial counting

technique was used as the basis of the refer-ence method. Urines were diluted in quarterstrength Ringer solution to produce dilutionsof 102 and 104. One millilitre of each dilutionand of the undiluted urine was mixed with20 ml of sterile, molten Columbia agar base(Oxoid CM331) held at a temperature of45°C. Plates were incubated overnight at37°C and colonies were counted. The rangeof dilutions was designed to allow bacterialnumbers in the range 0 to 107 cfu/ml to becounted. To facilitate the identification oforganisms, a calibrated 1 jul loop was used toseed urine on to one half of a 9 cm diameteragar plate containing cystine-lactose-electrolyte deficient (CLED) medium (OxoidCM301). Inoculated plates were incubated at37°C for 18 hours. Isolates were identifiedusing a variety of standard methods.5

Growths were considered to be clinicallysignificant if a count of 105 cfu/ml of nomore than two types of organisms were grownor there was a predominant organism in amixed culture. Counts between 104 and 105cfu/ml were considered to be clinically signifi-cant if they were in pure culture and thepatient was receiving antimicrobial treatment,or the urine contained > 10 leucocytes/,ul.

Cultures designed to detect and enumeratecarboxyphilic and obligately anaerobic bacte-ria were prepared as follows: each urine sam-ple was cultured, using a l/ul loop on toblood agar and heated blood agar plates.These were incubated anaerobically and in anatmosphere of 10% CO2 in air, respectively,for 48 hours at 37°C.

All urine samples were tested for the pres-ence of antibacterial agents. A multipointinoculator was used to apply urine to lawncultures of Staphylococcus aureus (NCTC6571) and Escherichia coli (NCTC 10418).These strains were chosen because of theirsusceptibility to antibiotics commonly used inthe treatment of urinary tract infections. Afterovernight incubation, the presence of anti-bacterial agents was recorded where zones ofinhibition occurred.

Statistical analysesThe results of individual tests from Questorand the reference method were categorised

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Evaluation of Questor urine screening system

Table 1 Categories of results comparing reference andQuestor methods

Questor resultReference testresult Positive Negative

Positive True positive False negativeNegative False positive True negative

(table 1). Using these categories, the sensitiv-ity, specificity, and positive and negative pre-dictive values were calculated as follows6:

Sensitivity (%)True positives

True positives + false negatives

Specificity (%)True negatives

True negatives + false positives

Positive predictive value (%)True positives

True positives + false positives

Negative predictive value (%)True negatives

True negatives + false negatives

The performance of Questor was calcu-lated by using varying threshold settings forthe organism counts between 1-0 x 1 04 to1 1 x 10 in steps of 0-1 log, together withleucocyte counts of 20, 30, 40 and 50 per,uland an erythrocyte count of 250 (table 2).From the many possible combinations ofthresholds, the optimal performance was cal-culated using the method of Koenig et al.7 Inthis method sensitivity is weighted moreheavily than specificity, because the conse-quences of reporting a false negative resultare potentially more serious than those of a

Table 2 Performance of Questor at different positivethresholds

Questor Sensitivity Specificity Performanceorganisms ('o) (00) index

Leucocytes 50 pl5-0 x 104 93 2 590 83-060 x 104 932 65-3 84-870 x 104 927 713 86-38-0 x 104 910 75-8 8649.0 X 104 89-8 77-7 86-21-0 x 10' 89-3 78-4 86-01*1 x 10' 88-7 79-3 86-9Leucocytes 40 p15 0 x 104 93-2 58-0 82-760 x 104 932 64-2 84-57-0 x 104 93-2 700 86-38-0 x 104 92-7 74 0 87-19-0 X 104 91-5 75-9 86-91-0 x 10' 91-5 76-5 87-01-1 X 105 91.0 772 868Leucocytes 30 ul5 0 x 104 93-8 57-1 82 86-0 x 104 93-8 63-0 84-67 0 x 104 93-8 68 3 86-28-0 x 104 93-2 71-8 86-89-0 x 104 92-1 74-0 86-71-0 X 105 92 1 74-0 86-71-1 X 105 915 745 864Leucocytes 20 pl5-0 x 104 94 9 54-6 82-86-0 x 104 94-9 59-7 84-470 x 104 949 64-1 85780 x 104 944 67-3 86-29-0 X 104 93-8 68-4 86-21.0 X 105 93-8 68-9 86-31*1 X 105 93-8 69-2 86-4

false positive result. The optimal performancewas determined by calculating an index thatemphasised sensitivity more than specificityin a ratio of 70:30-70% of the sensitivity wasadded to 30% of the specificity at eachthreshold combination (table 2). The 70:30ratio is arbitrary; small changes in the ratio donot alter the optimal thresholds.

ResultsOf the 1023 urine samples tested, 225(22%)contained > 105 cfu/ml of bacteria as deter-mined by the pour-plate counts. Of these225, 167 were considered to be clinically sig-nificant growths. An additional 10 specimenscontained between 104 and 105 cfu/ml andwere considered to be clinically significant.Because the growth was pure, it was associ-ated with the presence of 10 or more leuco-cytes and the clinical information indicatedan infection. There were therefore a total of177 significant growths.Using the method of Koenig et al7; the

optimal performance of Questor was with thefollowing thresholds: bacteria 8-0 x 104, or

leucocytes 40/pl, or erythrocytes 250/pul. Theperformance achieved at these thresholds isshown in table 3.Of the 13 false negative results 10 grew

pure cultures of E coli, all of which had lessthan 10 leucocytes/,ul. One specimen grewEnterococcus faecalis and another Proteusmirabilis, associated with leucocyte counts ofl/ul and 6/,ul, respectively. The clinical signif-icance of these growths may be questioned.The remaining false negative grew E coli andhad a leucocyte count of 22/,ul.

Questor assigned the following interpreta-tions to the false positive results: (n = numberof samples in category): "positive" 109, "poswhite cell" 84, and "pos-red blood cell" 27.The leucocyte counts of the false positive

results determined by microscopy is shown intable 4. Questor was thus able to detect a

large number of leucocytes (,>50) in almost50% of the false positive results.Of the 220 false positive results, 86 (39%)

contained anaerobic or carboxyphilic organ-isms and a further 74 (34%) contained a

mixed bacterial population. Thus in 73% of

Table 3 Optimal performance of Questor

True positives 164True negatives 626False positives 220False negatives 13Sensitivity 92-7%Specificity 74-0%Positive predictive value 42-7%Negative predictive value 98 0%

Table 4 False positive samples containing leucocytes

Number of leucocytes Number ofsamples

0-9 7210-19 1720-29 530-39 840-49 950-99 21> 100 88

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the false positive results, organisms were pre-sent in numbers greater than 105 cfu/ml.Antibiotic substances were detected in asignificantly higher percentage (47 of 220, or21%) of the false positive samples comparedwith 147/1023 (14%) of all the samplestested (p = 0-001).

Questor assigned the following interpreta-tion to the true positive results: (n = numberof samples in category): "positive" 105, "pos-white cells" 48, and "pos-red blood cells" 10.Comparisons between the Questor leucocyte,erythrocyte, and epithelial cells counts andmicroscopy are given in tables 5, 6, and 7respectively.

DiscussionThe Questor system was more sensitive andspecific than other bacteriuria screening sys-tems evaluated using the same protocol.489The results showed that the system was

able selectively to detect bacteria, leucocytes,erythrocytes and epithelial cells. The bacterialcounts given were accurate, except below1.0 x 104 and above 1 1 x 105, and theevaluators suggest that as part of the installa-tion process for individual laboratories, thesoftware should be configured so that resultsoutside these figures should read <1 0 x 104and >1 0 x 105 , respectively.

Although Questor recorded a false positiveresult for 220 specimens, many of these couldbe accounted for by the presence of anaerobicor carboxyphilic organisms, or the presence ofantimicrobial substances and therefore thepossibility of the presence of non-viablebacteria.A total of 109 of the 220 false positive

results contained high numbers of leucocytes

Table 5 Leucocyte counts, Questor compared withmicroscopy Questor white cell count (per p1)Microscopy 0-9 10-29 30-49 >50 Totals

0-9 486 129 29 14 65810-29 14 44 9 16 8330-49 0 13 18 13 44>50 1 8 8 221 238Totals 501 194 64 264 1023

Table 6 Erythrocyte counts, Questor compared withmicroscopy Questor red blood cell count (per,l)

Microscopy 0-9 10-29 30-49 > 50 Totals

0-9 524 141 30 37 73210-29 17 43 13 5 7830-49 1 10 9 11 31>50 6 23 28 125 182Totals 548 217 80 155 1023

Table 7 Epithelial cells, Questor compared withmicroscopy Questor epithelial cell count (per 1,d)Microscopy 0-9 10-29 30-49 >50 Totals

0-9 687 34 1 1 72310-29 84 57 3 1 14530-49 9 34 1 0 44>50 3 16 14 22 55Totals 783 141 19 24 967

*Epithelial cells were not recorded by Questor for the remain-ing 56 specimens, due to the presence of an excess number ofleucocytes.

and although no organisms were detected inthese samples, these high leucocyte countshave diagnostic value.

Questor has a tendency to give higher leu-cocyte and erythrocyte counts than is foundby microscopy (tables 5 and 6).

Individual laboratories using Questor havethe option of choosing any combination ofdifferent positive threshold levels for organ-isms, leucocytes, and erythrocytes. Changingthese thresholds will alter the performance,increasing the sensitivity and lowering thespecificity or vice versa. This may be impor-tant when testing urine samples from particu-lar groups of patients. In a patient populationwith a low incidence of urinary tract infectionthe thresholds could be changed to increasethe specificity; in a symptomatic group, ahigh sensitivity is required. It is recom-mended that each user laboratory should cali-brate the system by comparing the resultsobtained with those of the best possible com-parative method.

Because the operator involvement in theanalytical process consists only of pipetting10041l of urine sample into a tray and enter-ing the sample number into the computersystem, with no interpretive steps, a medicallaboratory assistant can be used to operatethe system.

In this evaluation it was possible for amedical laboratory assistant to operate theQuestor system and to process 230 urinesamples a day. As Questor takes about 1minute to process one sample, in theory itwould be possible for the system to test over400 samples in a normal working day. This,however, would rely on specimens beingreceived at regular intervals and thereforeavailable for testing as a continuous workloadthroughout the day. In practice this did notoccur, with most samples being received inmid-afternoon. Questor is a system designedto screen urines, allowing a negative result tobe sent from the laboratory in most cases onthe same day as specimen receipt.

This could have some beneficial impact onpatient care, although this would dependgreatly on both the speed at which the speci-men is sent to the laboratory after voidingand how quickly the report is sent to therequesting physician. A benefit to the labora-tory would be that the negative specimenswould not be cultured and therefore notrequire the preparation and interpretation ofculture plates by laboratory staff. Specimensrecorded as positive by Questor wouldrequire culture, to identify causative organ-isms, to provide cultures for antimicrobialsusceptibility tests, and to detect mixed,possibly insignificant growths.The list price capital cost of the Questor

system is £39 950. The list price costs of theconsumables required, diluent, and trays total,C0-17 a sample. The WELCAN workloadsystem, when applied to all procedures asso-ciated with urine screening by Questor and aroutine microscopical and cultural method,showed a saving of staff time of 31% whenusing the automated method.

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The system was simple to use, was liked bylaboratory staff, and had excellent, user-

friendly software. The ability of the system toprovide an accurate rapid, result, the savingsin staff time taken to process samples makethe Questor system an important and usefuladdition to the methods available for screen-ing for bacteriuria.This evaluation was supported financially by the MedicalDevices Directorate, Department of Health.

1 Kass EH. Pyelonephritis and bacteriuria: a major problemin preventative medicine. Ann Intern Med 1962;56:46-53.

2 Stevens M. Screening urines for bacteriuria. Med Lab Sci

1989;46: 194-206.3 Medical Devices Directorate. An evaluation of the Questor

urine screening system. Report MDD/92/48. London:Department of Health, 1992.

4 Medical Devices Directorate. An evaluation of the OrbecRamus 256. Report STD/88/13. London: Department ofHealth, 1988.

5 Cowan & Steel's Manual for the Identification of MedicalBacteria. 2nd Edn, Cambridge: Cambridge UniversityPress, 1975.

6 Galen RS, Gambino SR. Beyond Normality: The PredictiveValue and Efficiency of Medical Diagnoses. New York:John Wiley and Sons, 1975.

7 Koenig C, Tick U, Hanna BA. Analyses of theFlashTrack DNA probe and UTIscreen biolumines-cence tests for bacteriuria. Clin Microbiol 1992;30:342-5.

8 Medical Devices Directorate. An evaluation of the Autotrakurine screening system. Report STD/88/28. London:Department of Health, 1988.

9 Medical Devices Directorate. An evaluation of the SensititreLeukoBact urine screen. Report MDD/92/03. London:Department of Health, 1992.

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