CleanRoom Spore Size Study

8/2/2019 CleanRoom Spore Size Study

http://slidepdf.com/reader/full/cleanroom-spore-size-study 1/32





• GOALSGOALS -- Better understand properties of simulantsBetter understand properties of simulantsvs. agents. Provide data to help choose appropriatevs. agents. Provide data to help choose appropriatesimulants and surrogates for specific tasks.simulants and surrogates for specific tasks.

•• METHODMETHOD -- Compare properties of simulants to thoseCompare properties of simulants to thoseof the agents.of the agents.

IntroductionIntroduction

•• FACILITYFACILITYMicrobiologyMicrobiologyTeamTeam

BSL3BSL3LaboratoryLaboratory



Properties (NonProperties (Non --Medical)Medical)•• Spore SizeSpore Size –– length, width, aspect ratiolength, width, aspect ratio

•• Spore DensitySpore Density

•• Effect of decon agentsEffect of decon agents

•• Fluorescence spectraFluorescence spectra

•• Effect of UV radiationEffect of UV radiation



Spore SizeSpore Size•• Important to design and development of samplersImportant to design and development of samplersand detectorsand detectors

•• Published spore sizesPublished spore sizes -- spores produced underspores produced underdifferent conditions for each species withoutdifferent conditions for each species withoutextensive comparisons, size distributions or ranges.extensive comparisons, size distributions or ranges.

•• Systematic comparison of the size of Systematic comparison of the size of B. B. anthracis anthracisspores to size of other Bacillus sporesspores to size of other Bacillus spores

(simulants/surrogates)(simulants/surrogates) -- all spores produced underall spores produced underthe same conditions.the same conditions.





B. B. globigii globigii SB512SB512 (B.(B. atrophaeus atrophaeus ) )

•• LengthLength

Mean 1.22Mean 1.22 ±±0.120.12 µmm

Range 0.91Range 0.91 --1.901.90 µmm•• WidthWidth

Mean 0.65Mean 0.65 ±±0.050.05 µmm

Range 0.56Range 0.56 --0.800.80 µmm

B. globigii Size Disribution

0

5

10

15

20

2530

35

40

45

900 1000110012001300140015001600170018001900More

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%

FrequencyCumulative %

B. globigii Size Distribution

0

5

10

15

20

25

500 550 600 650 700 750 800

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. subtilis subtilis 10311031

•• LengthLength

Mean 1.07Mean 1.07 ±±0.090.09 µmm


Mean 0.48Mean 0.48 ±±0.030.03 µmm

Range 0.41Range 0.41 --0.580.58 µmm

B. subtilis Size Distribution

0

5

10

15

20

25

30

35

8 5 0 9 0 0 9 5 0 1 0 0 0

1 0 5 0

1 1 0 0

1 1 5 0

1 2 0 0

1 2 5 0

1 3 0 0

1 3 5 0

1 4 0 0

1 4 5 0 M o r e

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%


B. subtilis Size Distribution

0

5

10

15

20

25

30

400 425 450 475 500 525 550 575 600 MoreWidth (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%FrequencyCumulative %



B. cereus B. cereus ATCC 10702ATCC 10702

•• LengthLengthMean 1.55Mean 1.55 ±±0.160.16 µmm


Mean 0.90Mean 0.90 ±±0.070.07 µmmRange 0.76Range 0.76 --1.141.14 µmm

B. cereus

0

5

10

15

20

25

30

1200 1300 1400 1500 1600 1700 1800 1900 2000 More

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%


B. cereus

0

5

10

15

20

25

30

750 800 850 900 950 1000 1050 1100 1150 More

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%




B. B. megaterium megaterium WWWW --1515 --49004900

•• LengthLength

Mean 1.60Mean 1.60 ±±0.160.16 µmm


Mean 0.84Mean 0.84 ±±0.070.07 µmm

Range 0.70Range 0.70 --1.051.05 µmm

B. megaterium

0

5

10

15

20

25

30

1200 13001400 1500 1600 1700 1800 1900 2000 2100 More

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%


B. megaterium

0

5

10

15

20

25

30

700 750 800 850 900 950 1000 10501100More

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. mycoides mycoides ATCC 10206ATCC 10206

•• LengthLength

Mean 1.85Mean 1.85 ±±0.210.21 µmm


Mean 0.91Mean 0.91 ±±0.070.07 µmm

Range 0.65Range 0.65 --1.041.04 µmm

B. mycoides

0

5

10

15

20

25

30

1 3 0 0

1 4 0 0

1 5 0 0

1 6 0 0

1 7 0 0

1 8 0 0

1 9 0 0

2 0 0 0

2 1 0 0

2 2 0 0

2 3 0 0

2 4 0 0

2 5 0 0

M o r e

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%


B. mycoides

0

5

10

15

20

25

30

35

650 700 750 800 850 900 950 1000 1050 More

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%






B. B. anthracis anthracis ((∆--SterneSterne ))

•• LengthLength

Mean 1.56Mean 1.56 ±±0.160.16 µmm


Mean 0.82Mean 0.82 ±±0.060.06 µmm

Range 0.68Range 0.68 --0.980.98 µmm

B. anthracis ∆ -Sterne

0

5

10

15

20

25

30

35

650 700 750 800 850 900 950 1000 1050 1100 More

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%


B. anthracis ∆ -Sterne

0

5

10

15

20

25

30

1100 1200 1300 1400 1500 1600 1700 1800 1900 2000 2100 More

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%

Frequency

Cumulative %



B. B. anthracis anthracis (Albia)(Albia)

•• LengthLength

Mean 1.43Mean 1.43 ±±0.140.14 µmm


Mean 0.87Mean 0.87 ±±0.070.07 µmm

Range 0.71Range 0.71 --1.031.03 µmm

B. anthracis Albia

0

2

4

6

8

10

12

14

16

18

1200 1250 1300 1350 1400 1450 1500 1550 1600 1650 1700 1750 1800 More

Length

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%


B. anthracis Albia

0

5

10

15

20

25

30

35

40

700 750 800 850 900 950 1000 1050 1100 More

Width

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. anthracis anthracis (NCTC 1087)(NCTC 1087)

•• LengthLength

Mean 1.68Mean 1.68 ±±0.190.19 µmm


Mean 0.89Mean 0.89 ±±0.060.06 µmm

Range 0.76Range 0.76 --1.091.09 µmm

B. anthracis 1087

0

5

10

15

20

25

1 2 0 0

1 3 0 0

1 4 0 0

1 5 0 0

1 6 0 0

1 7 0 0

1 8 0 0

1 9 0 0

2 0 0 0

2 1 0 0

M o r e

Length (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%80.00%

100.00%

120.00%


B. anthracis 1087

0

5

10

15

2025

30

35

40

750 800 850 900 950 1000 1050 1100 More

Width (nm)

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. anthracis anthracis (NCTC 1928)(NCTC 1928)

B. B. anthracis anthracis (LA1)(LA1)



Aspect Ratio (L/W) of Bacillus sporesAspect Ratio (L/W) of Bacillus sporesB anthracis Sterne

600

700

800

900

1000

1100

1200

1000 1200 1400 1600 1800 2000

Length

W i d t h

B globigii

500

550600650700750800850900

1000 1100 1200 1300 1400 1500 1600 1700

Length

W i d t h

B. subtilis

400

450500

550

600

650

700

800 900 1000 1100 1200 1300 1400 1500 1600

Length

W i d t h

B. a B. a . NCTC 1087. NCTC 1087

MeanMean 1.91 +/ 1.91 +/ -- 0.230.23RangeRange 1.371.37 --2.452.45

B. subtilis B. subtilis

MeanMean 2.23 +/ 2.23 +/ -- 0.250.25RangeRange 1.631.63 --2.952.95

B. globigii B. globigiiMeanMean 1.85 +/ 1.85 +/ -- 0.190.19

RangeRange 1.431.43 --2.392.39

B. a B. a .. SterneSterneMeanMean 1.75 +/ 1.75 +/ -- 0.200.20

RangeRange 1.331.33 --2.202.20B. anthracis 1087

600

700

800

900

1000

1100

1200

1000 1200 1400 1600 1800 2000 2200

Length

W i d t h



Comparison of Comparison of B. B. anthracis anthracis with otherwith other

Bacillus Bacillus speciesspeciesLength ( µ m) Width ( µ m)

B. anthracis Albia 1.43 +/- 0.14 0.87 +/- 0.07 B. anthracis 1087 1.68 +/- 0.19 0.89 +/- 0.06 B. anthracis Sterne 1.53 +/- 0.15 0.88 +/- 0.06 B. anthracis D-Sterne 1.56 +/- 0.16 0.82 +/- 0.06

B. globigii 1.22 +/- 0.12 0.65 +/- 0.05

B. subtilis 1.07 +/- 0.09 0.48 +/- 0.03

B. cereus 1.55 +/- 0.16 0.90 +/- 0.07

B. megaterium 1.60 +/- 0.16 0.84 +/- 0.07

B. mycoides 1.85 +/- 0.21 0.91 +/- 0.07



B.a. B.a. 1087 vs. B. g. (Length)1087 vs. B. g. (Length)

9 0 0

1 1 0 0

1 3 0 0

1 5 0 0

1 7 0 0

1 9 0 0

2 1 0 0

0

10

20

30

40

50

F r e q u e n c y

Length (nm)

1087BG



B.a. B.a. 1087 vs. B. g. (Width)1087 vs. B. g. (Width)

5 5 0

6 5 0

7 5 0

8 5 0

9 5 0

1 0 5 0

M o r e

05

10

1520253035

40

F r e q u e n c y

Width (nm)

B. a. 1087BG





B.a. Albia vs. B. g. (Width)B.a. Albia vs. B. g. (Width)

5 5 0

6 5 0

7 5 0

8 5 0

9 5 0

1 0 5 0

M

o r e

05

10152025303540

F r e q u e n c y

Width (nm)

B.a. AlbiaBG



B.a. B.a. 1087 vs.1087 vs. B. subtilis B. subtilis(Length)(Length)

8 0 0

1 0 0 0

1 2 0 0

1 4 0 0

1 6 0 0

1 8 0

0

2 0 0 0

M o r e

0

1020

30

40

50

60

F r e q u e n c y

Length (nm)

B. a. 1087B. sub.



B.a. B.a. 1087 vs.1087 vs. B. subtilis B. subtilis (Width)(Width)

4 0 0

5 0 0

6 0 0

7 0 0

8 0 0

9 0 0

1 0 0 0

1 1 0 0

0

10

20

30

40

50

60

F r e q u e n c y

Width (nm)

1087

B. sub.



B.a. B.a. 1087 vs.1087 vs. B. cereus B. cereus (Length)(Length)

1 2 0 0

1 3 0 0

1 4 0 0

1 5 0 0

1 6 0 0

1 7 0 0

1 8 0 0

1 9 0 0

2

0 0 0

2 1 0 0

M o r e

0

5

10

15

20

25

30

F r e q u e n c y

Length (nm)

B. a. 1087B. cer.



B.a. B.a. 1087 vs.1087 vs. B. cereus B. cereus (Width)(Width)

6 5 0

7 5 0

8 5 0

9 5 0

1

0 5 0

1 1 5 0

05

10

1520253035

40

F r e q u e n c y

Width (nm)

1087B. cer.



B. B. globigii globigii SB512SB512 (B.(B. atrophaeus atrophaeus ) )

•• LengthLength

Mean 1.27Mean 1.27 ±±0.100.10 µmm


Mean 0.67Mean 0.67 ±±0.070.07 µmm

Range 0.50Range 0.50 --0.940.94 µmm

B. globigii

05

101520253035404550

1 1.1 1.2 1.3 1.4 1.5 1.6 1.7 More

Length

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%


B. globigii

01020304050607080

0.4 0.5 0.6 0.7 0.8 0.9 1 More

Width

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. subtilis subtilis 10311031

•• LengthLength

Mean 1.14Mean 1.14 ±±0.070.07 µmm


Mean 0.54Mean 0.54 ±±0.040.04 µmm

Range 0.46Range 0.46 --0.690.69 µmm

B. subtilis

0

5

10

15

2025

30

35

0.9 1.05 1.1 1.15 1.2 1.25 1.3 More

Length

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%


B. subtilis

0

10

20

3040

50

60

70

0.4 0.45 0.5 0.55 0.6 0.65 0.7 More

Width

F r e q u e n

c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




B. B. anthracis anthracis ((∆--SterneSterne ))

•• LengthLength

Mean 1.48Mean 1.48 ±±0.070.07 µmm


Mean 0.89Mean 0.89 ±±0.050.05 µmm

Range 0.75Range 0.75 --1.001.00 µmm

B. anthracis d-Sterne

0

5

10

15

20

25

30

35

40

1.3 1.35 1.4 1.45 1.5 1.55 1.6 1.65 1.7 1.75More

Length

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%


B. anthracis d-Sterne

0

10

20

30

40

50

60

0.7 0. 75 0. 8 0.85 0.9 0.95 1 1.05 More

Width

F r e q u e n c y

.00%

20.00%

40.00%

60.00%

80.00%

100.00%

120.00%




Comparison of MeasurementsComparison of Measurementsby TEM vs. OMby TEM vs. OM

Length WidthB. globigii TEM 1.22 +/- 0.12 0.65 +/- 0.05

OM 1.27 +/- 0.10 0.67 +/- 0.07

B. subtilis TEM 1.07 +/- 0.09 0.48 +/- 0.03OM 1.14 +/- 0.07 0.54 +/- 0.04

B. anthracis (d-Sterne) TEM 1.56 +/- 0.16 0.82 +/- 0.06OM 1.48 +/- 0.07 0.89 +/- 0.05



CONCLUSIONSCONCLUSIONS•• Systematic comparison of spore properties of severalSystematic comparison of spore properties of severalBacillus species.Bacillus species.

••

Compared spore size properties of 6 species of BacillusCompared spore size properties of 6 species of Bacillus

by TEM.by TEM.

•• B. B. anthracis anthracis spores are significantly larger than sporesspores are significantly larger than sporesof the most common simulants,of the most common simulants, B. g. B. g. andand B. B. subtilis subtilis ..

•• B. B. anthracis anthracis spores are similar in size tospores are similar in size to B. cereus B. cereus andand B. B. megaterium megaterium ..

•• Spores of pathogenic and nonSpores of pathogenic and non --pathogenicpathogenic B. B. anthracis anthracisare similar in size.are similar in size.



AcknowledgementsAcknowledgements

MICROBIOLOGY TEAMMICROBIOLOGY TEAMLeslie WilliamsLeslie WilliamsWarren GardnerWarren Gardner

UNIVERSITY OF PENNSYLVANIAUNIVERSITY OF PENNSYLVANIA ––BIOMEDICAL IMAGING CORE LABORATORYBIOMEDICAL IMAGING CORE LABORATORY

Neelima ShahNeelima ShahRay MeadeRay Meade

Documents

CleanRoom Spore Size Study