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International Journal of Human Nutrition and Functional Medicine

www.IntJHumNutrFunctMed.Org

2014 Final PDF

Bacteria Used:

Cryovials containing frozen aliquots of

Shigella sonnei, Salmonella typhi, Escherichia coli

0157:H7

,

Campylobacter

jejuni,

Vibrio

parahemolyticus, Aeromonas caviae, Plesiomonas

shigelloides,

Edwardsiella

tarda,

Yersinia

enterocolitica

, and

Clostridium difficile

were utilized.

Bacterial preparations were made after aseptically

inoculating bacteria into 25 mL of Trypticase Soy

Broth.

S. sonnei, S. typhi, E. coli, V. parahemolyticus,

A. hydrophilia, P. shigelloides, E. tarda,

and

Y.

enterolytica

spiked broths were incubated overnight at

37 ± 2°C overnight.

C. jejuni

and

C. difficile

broths

were cultured in anaerobic jars with BD GasPaks™ for

2-3 days at 40 ± 2°C and for 2 days at 37 ± 2°C,

respectively.

Colony Counts:

Each overnight incubated culture was

diluted in 0.1% peptone to a concentration of

approximately 1.0 x 10

7

colony forming units/mL

(CFU/mL) using McFarland standardization. Serial

dilutions were plated in quintuplicate to confirm the

concentration of the spike-aliquots. Titer plates were

incubated for the various bacteria as described.

Results

A total of 34 stool samples were sent for Stool Testing.

The stool pool was tested extensively, using

conventional methodologies, on two separate days and

found to be free of entero pathogenic bacteria, yeast and

parasites. Thirty-one specimens were spiked with

bacterial pathogens at clinically significant levels that

are within the sensitivity of culture based methods, and

at higher levels well above the Subject Laboratory’s

reported lower limit for detection of pathogens. Three

“control” specimens were unaltered and contained no

bacterial, fungal or parasitic pathogens. All 31 stool

specimens containing bacterial pathogens were

reported negative for the indicated pathogens by the

Subject Laboratory. Seventeen samples were reported

as “Parasite present, taxonomy unavailable.” Fifteen

samples from the same stool specimen were reported as

“No Ova or Parasites.” One specimen was reported to

contain

Cryptosporidium

sp. and one specimen was

reported to contain

Enterobius vermicularis

. Two of the

samples that were reported to contain “Parasite present,

taxonomy unavailable,” were also reported to contain

Cryptosporidium

sp. Complete results are shown in

Table 1.

Table 1. Results of Stool analysis Conducted by Subject Laboratory:

(-) bacteria not present; (+) bacteria present

Sample

ID

Organism

Added to

Normal Stool

Specimen

Quantity

Normal Stool

Flora

Opportunistic

Bacteria

Pathogenic

Bacteria

Yeast/ Fungi Parasites

1

Shigella sonnei

3.4x10

2

CFU/g

+

-

-

-

Parasite

Present;

taxonomy

unavailable

2

Shigella sonnei

3.4x10

5

CFU/g

+

-

-

-

Parasite

Present;

taxonomy

unavailable

3

Salmonella typhi

4.4x102

CFU/g

+

-

-

-

Parasite

Present;

taxonomy

unavailable

4

Salmonella typhi

4.4x105

CFU/g

+

-

-

4+ =>

1000000pg

DNA/g

specimen

Geotricum sp.

No Ova or

Parasites

5

E. coli 0157:H7

2.8x102

CFU/g

+

-

-

-

Cryptosporidium

sp. Positive,

Parasite

Present;

taxonomy

unavailable

6

E. coli 0157:H7

2.8x105

CFU/g

+

-

-

-

Parasite

Present;

taxonomy

unavailable

7

Campylobacter

jejuni

2.8x102

CFU/g

+

-

-

-

Parasite

Present;

taxonomy

unavailable