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Evaluation of Accuracy and Efficiency for Novel Automatic Colony Counting System for Ready-to-Use Culture Media, Easy Plate
Kentaro Takenaka1, Shinichiro Sugiura2(1) Kikkoman Corporation, (2) Kikkoman Biochemifa Company

Continuous Scanning

Automated Counting

Contamination

Onion salad
Bean sprout

pH adjustment

No need to adjust parameters according to the sample-CCS only needs a general-purpose
document scanner, thus reducing
the initial investment-All 5 types of Easy Plate supported

Tortilla roll
Raw salmon

Escherichia coli
NBRC 15034
was inoculated

Frozen pizza
Kikkoman soymilk
Vegetable juice

pH 7 using 1N NaOH

Stomached and diluted in phosphate-buffered saline (PBS)

Inoculation & Incubation
CCCorrelation coefficients between manual counting and the CCS
method were greater than 0.98 for both AC and CC (Figures 2aand 2b). However, in some cases, such as in areas X and Y, nocorrelation was obtained.
It is assumed that the differences in area X due to the false
detection of bubbles in CCS method (blue arrow in Figure 2c) andthe missing of small colonies in manual counting (orange arrow).
The differences in area Y may be due to the presence of verythin colonies that did not show up on the scanner in CCS method(blue arrows in Figure 2d) .
(a)
300

Medium, type of Easy Plate

Incubation temperature & timePlates

For aerobic countFor coliform count

35±1℃, 48±2hr

35±1℃, 24±1hr

15 plates

Contamination level(CFU/plate)

Low: 10-50, Middle: 50-150, High: 150-300

CCS method

Manual counting

Category
PC

OptiPlex 3080 SFF(Dell), Windows10, Corei3-10105(4C/3.74.4GHz) /16GB/256GBSSD+1TBHDDScanner

ADS-4300N (Brother Industry, LTD)

Scanner software

Brother ScanEssentials Ver. 1.1.0.2(Brother Industry, LTD)Counting software

CCS for Easy Plate Ver. 1.0.2150100x3 technicians (T1~3) 5 plates

150Y
100
5000150200

250300050

100150200

250300Manual counting (CFU/plate)(d)x1 CCS

40

Faster
18.9

19.2

Faster

90

70.0
60.0
50.0
40.0
30.0
20.0
10.0
0.0

10

5.6

5.7

0
T1T2
Technicians

T3

CCSCCS

T1T2T3

Technicians

CCS

7060
50
4030
201050

100

150

200

250300050

Number of colonies (CFU/plate)

100150200250300

Number of colonies (CFU/plate)
CCS

T1

T2

T3

Figure 3. Comparison of counting speeds between counting methods

ACCC
8.00

7.40

7.00

5.91

6.00

6.03

Faster
5.30

5.27

5.24

5.00
4.00
3.00
2.00
1.00

7.40

Faster

7.00
5.70

6.00

5.27

5.00

4.99

5.02

4.97

4.00
3.00
2.00
1.00
0.00

515

30

4560

90

Counting

2.60

1.42

1.07

0.86

0.88

0.80

4.16

Scanning

4.80

4.28

4.20

4.13

4.13

4.17

5.24

Sum

7.40

5.70

5.27

4.99

5.02

4.97

51530

45

60

90

Counting

2.70

1.70

1.73

1.18

1.13

1.09

Scanning

4.704.21

4.30

4.12

4.13

Sum

7.40

5.91

6.03

5.30

5.27

Number of plates processed

CCS

Figure 4. Comparison of average counting speeds between methods at (a)lowand (b)high contamination

80000.00

Number of plates processed

Figure 5. Comparison of number of plates processed and CCS method speed

References
1. OKOCHI, Norihiko, et al. Dai Nippon Printing Co., Ltd, Medi· Ca AC for Enumeration of

15 plates

Aerobic Bacteria. Journal of AOAC International, 2014, 97.3: 837-842.

Figure 2. Correlation coefficients of (a) AC and (b) CC for bacterial countsbetween counting methods. (c) Portion of AC in area X and (d) CC in area Y.

Link to CCS info

• The counting speed is faster when more plates are processed at a batch.
• The CCS method uses a general-purpose scanner, which makes it a low-cost
option for users.
• The CCS method is an effective way to reduce counting time while
maintaining accuracy.

2. SAITO, Fumihiko, et al. Dai Nippon Printing Co., Ltd Medi· Ca CC for Enumeration ofAccuracy and efficiency analysis

80.0

Conclusion

20050

100

50

17.2100

8.00

60

20

90.0

64.4R² = 0.986

2502005065.1CC

CC

Manual counting (CFU/plate)(c)

×Detail

R² = 0.986062.830

ACHigh contamination level (150-300 CFU)300

X250Counting
System configuration of CCS

(b)

AC

CCS method (CFU/plate)-Simple and easy-to-use UI

No adjustment(b)

Low contamination level (10-50 CFU)
70

CCS method (CFU/plate)AI-based image recognition algorithm-

Raw shrimp

Naturallycontaminated(35℃ 0-5 hours)

AC-(a)

Figure 1. Marked or not marked AC and CC plate image

Raw ground chicken

Colony counting system for Easy Plate (CCS), jointly developedby Kikkoman Biochemifa Company and NTT DATA BUSINESSSYSTEMS Corporation, is an automated counting software
dedicated for Easy Plate and has the following features.

Since the CCS method automatically counts all images at once
after scanning, it was found that the more plates processed in abatch, the more efficient the method could be, as the counting
time per plate was reduced (Figure 5).

10-50
CFU

Raw ground beef

Colonies on Easy Plate are easy to count because they formbright, high-contrast colonies, but manual counting is timeconsuming and causes artificial errors.

Compared to manual counting, the CCS method was 3.3 times
faster for low contamination levels (10-50 CFU/plate) and 11.1 times
faster for high contamination levels (150-300 CFU/plate) (Figure 4).

50-150CFU

Sample preparation

10 food samples

CC

Counting time per plates(sec/plate)

Cultured Easy Plates

Conventional manual counting took time in proportion to thenumber of colonies. On the other hand, the CCS method showed
an average time of 5.8 seconds/plate, regardless of thecontamination level or the type of Easy Plate (Figure 3).

150-300
CFU

CCS method workflow

Materials and Methods

How to use Easy Plate

ACRaw ground beef

Easy Plate AC for aerobic bacteria count and Easy Plate CCfor coliform bacteria count are AOAC PTM certified1, 2) andMicroVal certified3) RTU dried medium that spreads sample
suspension evenly over the plate surface by simply closing thecover film. It also has advantages over conventional agar plates,
such as reduced plastic usage and reduced GHG emissions.

The count results of CCS method both AC and CC

Counting time per plate (sec/plate)In this study, we evaluated the accuracy of conventional manual
counting and automatic counting with CCS (CCS method) for
Easy Plate AC (AC) and Easy Plate CC (CC) and compared theirefficiency.

counting time(sec/plate)

Ready-to-use (RTU) culture media enhances food safety andproductivity because of its quickness, compactness, simplicity,and visibility of colonies compared to conventional agar plates.

Results – Efficiency

Results – AccuracyCounting time per plate (sec/plate)

Objectives

counting time(sec/plate)

Introduction

Coliform Bacteria. Journal of AOAC International, 2015, 98.1: 62-70.
3. https://microval.org/en/issued-certificates/

SCAN ME

Contact information
Kentaro Takenaka✉ktakenaka@mail.Kikkoman.co.jp
Kikkoman Biochemifa Company
https://biochemifa.kikkoman.com/e/kit/
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