European Journal of Parenteral & Pharmaceutical Sciences 2017; 22(1): 13-19

© 2017 Pharmaceutical and Healthcare Sciences Society

Use of contact plates to perform environmental

settle plate testing

Aleshia Samson, Andrew Sage, David Jones*

Rapid Micro Biosystems, Lowell, Boston, Massachusetts, USA

An evaluation was performed to investigate the use of contact plates in place of Petri plates to
perform environmental monitoring settle plates. A Petri plate has approximately 2.5 times the
surface area of a contact plate and a different agar profile for exposure to air. Petri plates also have
plastic walls that extend above the agar surface and may affect air flows. To evaluate the possible use
of contact plates, analysis of capture and recovery of bioburden was performed in a number of
different environments, with differing numbers of exposed plates. Overall, a good correlation was
shown when two contact plates were used as a substitute for one Petri plate with a 4-hour exposure
time. The ability to use contact plates for a settle plate test allows the use of a single consumable, the
contact plate, to be stocked for environmental monitoring testing.

Key words: Contact plate, settle plate, environmental monitoring.

Introduction
As part of an environmental monitoring programme, a
pharmaceutical company often performs air monitoring using
an active air sampler and settle plates. Each method has its
attributes for the tracking of environmental contaminants as
part of a trending programme for environmental quality
control. The current requirements call for a 90 mm Petri plate
to be exposed in the test area for up to 4 hours prior to
incubation and colony counting1,2. In some cases, the
substitution of the 90 mm Petri plate to a 60 mm contact plate
may be required either due to a technological requirement
(new rapid microbiological method (RMM)), or with a view
to rationalise all environmental monitoring testing to one
plate type (less growth promotion testing and stock control).
One drawback to this change concerns the fact that the
surface of a standard contact plate is smaller than a Petri plate.
The contact plates used in this study have a surface contact
diameter of 57 mm which, compared to a 90 mm diameter
Petri plate, gives 25.5 cm2 compared to 63.6 cm2. The contact
plate has approximately 2.5 times less surface area. Due to
this smaller sampling area3, a strategy is required to replace
the Petri plate for routine use with the smaller contact plate.
A number of options could be envisioned to replace a Petri
plate with a contact plate.
1. Use of a single contact plate as a trend monitor for
excursions and accepting that the smaller surface area may
lead to fewer detections.
*Corresponding author: David Jones, Director of Technical Services,
Rapid Micro Biosystems, 1001 Pawtucket Blvd West, Lowell 01854, Boston,
Massachusetts, USA; Email: DJones@rapidmicrobio.com
2. Using two contact plates exposed at the same time and site
to equate the surface area with the Petri plate.
3. Using a single contact plate and doubling the time of
exposure to 8 hours to equate to the total collection time
for the Petri plate and the surface area difference.
Data for each of these options have been collected and will be
described below.
Materials and methods
Materials
1. Environmental monitoring contact plates filled with
tryptone soya agar (TSA)-LP80 media and prepared by
Rapid Micro Biosystems (Boston, MA, USA) using BD
media powder (Becton Dickinson, Franklin Lakes, NJ,
USA).
2. TSA Petri plates: standard 90 mm filled with TSA
manufactured by Becton Dickinson.
3. TSA-LP80 Petri plates: standard 90 mm filled with TSA-
LP80 manufactured by Merck Millipore (Billerica, MA,
USA).
Methods
To compare settle plate capture of one, two or three contact
plates versus one Petri plate, 50, 100 or 150 contact plates
and 50 Petri plates were placed in different room grades
from Grade A to unqualified as per the diagram (Figure 1).
The contact plates and Petri plates were labelled in order to
track relative counts in each group. One or two of the
contact plates and the Petri plate were exposed for 4 hours at

13
14 ALESHIA SAMSON, ANDREW SAGE, DAVID JONES

Figure 1. Sampling layout: one, two or three contact plates (CPs) are laid around each Petri plate (PP).

room temperature. Also, depending on the study, the Results

third contact plate where required was exposed for 8
hours.
The sets of plates were placed side-by-side in small
groups of five to ten sets on flat surfaces with as little
intervention as possible from the operator.
After the 4-hour exposure time was completed, the
two contact plates and the single Petri plate per test set
were covered and removed. The remaining contact
plates were exposed for a total of 8 hours before being
covered and removed as required. The contact plates
and the Petri plates were incubated at 32.5°C for 72
hours with counts at day 2 and 3 in case spreading
colonies were present.
To verify the amount of dehydration that occurs from
the Petri plate and contact plate media during the
exposure, each plate was weighed pre- and post-test.
Comparison of single and double contact plates
at 4 hours and single contact plate at 8 hours of
exposure to a non-controlled environment
The non-controlled environment (office space) was used to
ensure high CFU counts to show equivalence for the data
in Figures 2 and 3. The results from the comparison of the
single contact plate experiment are shown in Figure 2. The
individual result for each contact plate was compared to
the corresponding TSA Petri plate in its set. The individual
contact plates have a lower capture than the larger Petri
plate in the 4-hour exposure time.
The comparison of summation of CFU counts for the
two test contact plates compared to the TSA Petri plate
counts in each set are shown in Figure 3. The summed CFU
count is generally higher than seen with the Petri plate.

Table 1. Data for the 2 x 50 contact plates for 4 hours, 1 x 50 contact plates for 8 hours versus 1 x 50 90 mm Petri plates for 4 hours.
Contact A Contact B Contacts A+B Contact Petri plate
4-hour 4-hour 4-hour 8-hour 4-hour
exposure exposure exposure exposure exposure

Mean CFU 6.78 7.34 14.12 10.96 10.44

Minimum CFU 2 3 7 5 4

Maximum CFU 12 13 21 19 18

% Petri plate 64.9 70.3 135.2 105 100

USE OF CONTACT PLATES TO PERFORM ENVIRONMENTAL SETTLE PLATE TESTING 15

Figure 2. Two replicates of contact plates (CP Replicates A and B) compared to a single Petri plate from each set

Figure 3. Summation of the two contact plates compared to the Petri plate CFU counts.
16 ALESHIA SAMSON, ANDREW SAGE, DAVID JONES

Table 2. 50 Petri plates and 100 contact plates exposed for 4 hours with the sample pattern shown in Figure 1.
Contact A Contact B Contact A + B 90 mm Petri plate

Mean CFU 6.98 3.92 5.45 8.94

Minimum CFU 1 0 0 0

Maximum CFU 19 12 19 19

% Petri plate 78.1 43.8 61.0 100.0

Table 3. Comparison of single, double and 8-hour exposure compared to the 90 mm TSA Petri plate.
Contact A Contact B Contacts A+B Contact 90 mm Petri plate
4-hour 4-hour 4-hour 8-hour 4-hour
exposure exposure exposure exposure exposure

Test with 0 CFU 49 50 99 46 45

Test with 1 CFU 1 0 1 4 4

Test with 2 CFU 0 0 0 0 1

Table 4. Comparison of one, two and combined contact plate count to the Petri plate with 4-hour exposure.
Sum of the CFU Tests with 0 CFU Tests with 1 CFU Tests with >1 CFU
on 50 replicates n n n

TSA Petri plate 42 40 1 9

Contact replicate 1 18 40 4 6

Contact replicate 2 7 46 1 3

Combined contacts 25 84 5 9

Table 5. Comparison of one contact plate replicate to the Petri plate with a 4-hour exposure time.
Sum of the CFU Tests with 0 CFU Tests with 1 CFU Tests with >1 CFU
on 49 replicates n n n

TSA Petri plate 7 42 7 0

Combined plate 1 48 1 0

Using a single contact plate and exposing it for 8 hours,
gives a very good agreement with the counts obtained
with the Petri plates (see Table 1 for data summary). The
one caveat with this approach is the high dehydration seen
on the contact plate compared to the 4-hour exposure.
Growth promotion was confirmed at the more extreme
loss of water, see later data set.
Comparison of contact plate to TSA Petri plate in
low bioburden environments
A second experiment was performed in the Class C
environment. Fifty TSA Petri plates were laid in a similar
pattern as described in Figure 1 with two contact plates for
each Petri plate. Exposure was for 4 hours. The data is
shown in Table 2. For this run, lower counts were observed
for both the contact and the Petri plates. The mean recovery
for the combined contact plate count being approximately
50% of the recovery seen in the first experiment at 61%.
Comparison of contact plate to TSA Petri plate in
Class A/B environments
In both the previous experiments, a higher CFU count was
seen than would be expected in a Class A/B environment.
To determine the equivalency in the Class A/B
environment, an experimental setup was made that used
the laminar airflow and a biological safety cabinet in the
Class B rooms pre-cleaning. As expected, much lower
counts were seen with the majority being 0 CFU counts.
The frequency of hits was recorded and summarised in
Tables 3, 4 and 5.
Table 3 shows the results for the first study. With a low
environmental count, the detection of organisms is less
efficient using the single contact option, both for the
single and combined counts. The 8-hour exposure gives a
good equivalent hit rate.
Further testing was performed to investigate the
equivalence of single and combined plate counts
compared to the control TSA Petri plate. The summaries
are shown in Tables 4 and 5. As it can be seen, the single
contact plate used in isolation is not equivalent to the Petri
plate for frequency of hits or total CFU captured. With the
combined values, the sum of CFU is 25 which is 61% of
the Petri plate with a total of 14 positive hits compared to
10 with the Petri plate.
USE OF CONTACT PLATES TO PERFORM ENVIRONMENTAL SETTLE PLATE TESTING 17

Table 6. Comparison of one, two and combined TSA-LP80 contact plate counts to the Petri plate with 4-hour exposure.
Sum of the CFU Tests with 0 CFU Tests with 1 CFU Tests with >1 CFU
on 50 replicates n n n

TSA-LP80 Petri plate 69 19 12 19

Contact replicate 1 28 28 18 4

Contact replicate 2 23 33 11 6

Combined contacts 51 20 25 5

Table 7. Contact and Petri plate dehydration as determined by weight loss after 4 hours of exposure.
Sample Weight (g) pre-exposure Weight (g) post-exposure Weight loss (g)

Contact plate 1 40.58 38.76 1.82

Contact plate 2 40.80 39.09 1.71

Contact plate 3 40.85 39.32 1.53

Contact plate 4 40.88 39.18 1.71

Contact plate 5 40.57 38.92 1.65

Petri plate 1 32.50 29.84 2.66

Petri plate 2 32.51 29.84 2.67

Petri plate 3 32.43 30.15 2.28

Petri plate 4 32.31 29.58 2.74

Petri plate 5 31.86 29.38 2.48

Comparison of contact plate to TSA-LP80 Petri
plate in Class A/B environments
The previous studies showed the comparison of contact
plates filled with TSA-LP80 versus Petri plates with TSA
media. The media formulation may have had an influence
on the data obtained. To investigate this variable, a study
was performed using 50 Petri plates filled with TSA-LP80
compared to the 100 contact plates. The layout of the
contact plates was as described in Figure 1 and the testing
performed in a Class B environment during engineering
shutdown with a 4-hour exposure time.
From the data there are 19 Petri plates and 20 pairs of
contact plates with zero CFU showing good correlation.
With those plates with a contamination the Petri plate
appears to recover slightly higher number per site with 19
Petri plates having more than 1 CFU while only 5 of the
combined show >1 CFU. The difference may indicate a
lower capture, however, there is also the possibility that the

Figure 4. Single contact plate exposed for 8 hours compared to 4-hour Petri plate CFU counts for the 50 test sites.
18
Figure 5. Weight loss profile during 8 hours of contact plate exposure as settle plate sampling in a laminar air flow hood. The control
samples did not have their lids removed through the sample period.
Petri plate media may have a better recovery of organisms
compared to the contact plate media as variations between
supplier media for growth properties is well known.
Media dehydration
To determine the degree of media dehydration throughout
the exposure process, a parallel experiment was conducted
exposing the contact plates and 90 mm Petri plates to the
controlled environment in a cleanroom. The plates were
weighed, exposed for 4 hours and then capped and
reweighed. Data is shown in Table 7.
The mean weight loss was 1.68 g for the contact plates
and 2.57 g for the Petri plates. Taking the weight of the
media into account rather than the total unit weight, the
loss of water was 7.6% for the contact plates and 14.3%
for the Petri plates.
With an 8-hour exposure, the water loss would be
expected to be more severe. To mimic this worst-case
scenario, ten contact plates were distributed around the
interior of a laminar airflow hood and exposed for 8 hours.
The linear dehydration is shown in Figure 5. Over the
course of the 8 hours, 3.25 g of water was lost, which is
15.1% of the agar moisture.
To verify that the nutritive properties of the media were
not lost, six contact plates were placed in duplicate in three
different International Organization for Standardization 5
hoods. One pair of contacts were kept as a non-exposed
control. Following 8 hours of sampling, the contacts were
taken and 50 µL of a preparation of Staphylococcus aureus
was spread plated onto the media surface for both the test
and control plates. The contact plates were incubated for
ALESHIA SAMSON, ANDREW SAGE, DAVID JONES
48 hours at 32.5°C and then the colonies enumerated. From
the data, it can be seen that all plates recovered >70% of
the spiked dose of S. aureus confirming growth promotion
after exposure (Table 8). Colony size was equivalent
between the test and control plates.
Discussion
The results reported here indicate that the use of contact
plates in place of a Petri plate would be technically
possible. Currently, in the Pharmacopoeias, there are no
acceptance criteria quoted for comparison of methods
other than to say “not significantly different”. The
Pharmacopoeias do give an acceptance range for
bioburden recovery so that value has been used as a
simple acceptance test. Using an acceptance criteria of
50–200% recovery as defined in the US Pharmacopeia
Chapter <61>4 and the European Pharmacopoeia 2.6.125,
the microbial recovery is acceptable for both the double
contact plate format and the single contact plate with an
8-hour exposure. The single contact plate format passes
the >50% recovery when the counts are relatively high
(>5 CFU), however, the detection efficiency does not
work as well in environments wherein the counts are
below 5 CFU. Why there should be a difference between
the two efficiencies when bioburden load is low is not
certain. One cause could be the statistical uncertainty of
detecting low numbers, therefore larger experiments are
planned.
The 8-hour exposure gives good recovery compared to
the Petri plate, however, the contact plates are more
USE OF CONTACT PLATES TO PERFORM ENVIRONMENTAL SETTLE PLATE TESTING 19

Table 8. Recovery of S. aureus spread on the dehydrated contact plates following 8 hours of exposure.
Sampling location CFU count Mean CFU % Recovery

Control 1 45 57.5
Control 2 70
Laminar airflow hood 1 54 65 113.0
Laminar airflow hood 2 76
Rear hood 1 50 54 93.9
Rear hood 2 58
Demo 1 40 44.5 77.4
Demo 2 49

dehydrated and there may be some risk for microorganism References

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The use of two contact plates in the same location and
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