Rapid, accurate, simple, convenient and economic.

Advanced System for efficient identification of 27 Enterobacteriaceae.

Indole production ONPG (hydrolysis) degradation for the detection of -D-glucosidase activity Glucose fermentation (acidity without gas)* Glucose fermentation with CO2 gas formation* Hydrogen Sulfide production Urea (hydrolysis) degradation for the ureasa activity Motility

ENTEROTESTID
Enterobacteriaceae is a large family of Gram-negative catalase positive and oxidase negative1 (mostly, as Pleisomonas family has been included in oxidase positive1) bacteria, that includes, along with many harmless symbionts, many of the more familiar pathogens.They are distributed worldwide and may be found in soil, water, plants and animals. Most species grow well at 37C, although some species grow better at 25-30C. There is a large number of species included in the family, more than 170, but the clinical and more important specimens which play a major role as infectious agents, belong to 20 to 25 species that are well known for many years. 2-6 The identification of the Enterobacteriaceae is complicated and has been traditionally based on biochemical and antigenic characteristics 2-6. With Enterotest ID, using a method well known by microbiologists worldwide, stabbing the butt and smearing a slant, six biochemical reactions are performed in a simple tube as well as bacteria growth motility for further investigations. The media tube contains two layers and a paper cap for indol determination. Upper layer(3 ml): a butt and a slant with dextrose, ONPG, enrichments, ferrous sulphate, sodium thiosulfate, agar and phenol-red Lower layer (2 ml): semi-solid media, containing urea, enrichments, sodium chloride, agar and phenol-red Paper cap: soaked with a solution of 4-dimethyl-aminobenzaldehyde for the indol test.
Warning: The prepared medium is red-peachy colour, slightly opalescent in the upper layer and light peachy in the lower layer. If there are any physical changes, discard the medium. Dehydration or liquefaction conditions may also interfere with the accuracy of Enterotest ID: * If the glucose is not fermented (the medium remains red-orange after inoculation and incubation), the strain does not belong to the Enterobacteriacea. 2-3

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Indole cap

3
1,5 cm 0,6

18 - 24 h at 35 2C

5
Citrobacter Edwardsiella Enterobacter

6
Enterotest ID Record Chart
DATE Reference Motility H2S Indol Urease ONPG Glucose Gas CO2 RESULT

Slant for bacterial growth ONPG Glucose with or without gas H2S

agg lom erans

aeroge nes

freu ndii

dive rsus

cloa cae
+ AG + +/-

tard a

STOPPER Slant

Indol ONPG Glucose H2S Motility Urea

+ AG + + -

+ + AG + -

+ AG + + -

+ AG + -

+ + AG

D A AG +/+

UPPER LAYER

Butt

LOWER LAYER

coli
+ -

+ positive in one day + positive weekly

- negative D different

[+] [+] H2S

LOT NUMBER

EXPIRY DATE

Store at room temperature in a dark place.

Urea, Motility

Pick a typical suspected ISOLATED colony from an appropriate plate culture. Appropiated medium could be MacConkey agar (cat.1052), Hektoen Enteric agar (cat.1030), Eosin Methylene Blue (EMB) (cat. 1039), Salmonella Shigella (SS) (cat.1064) or nonselective blood agar media. Only an isolated colony must be chosen. Jab the tube to the bottom and seed in the slant. Place the indol test cap. Incubate for 18-24 hours at 352C. Read the results and compare with differentiation chart included. Results can be interpreted by using the differentiation chart, comparing with known species or using the digital interpretation guide (codebook) provided. When further tests are needed (two or more organisms are listed with the same result), the codebook will inform you as well as the results will be obtained in them. Use slant growth for further serological test if needed.

REACTION INDOLE ONPG GLUCOSE CO2 SH2 MOTILITY UREASE

POSITIVE Purple-pink Orange Yellow Bubbles Black * Fuzzy Pink

NEGATIVE Colorless Cherry-Pink Yellow Clear Yellow

2 3 4 5

(*) If too much bacterium is taken in the seed, the ferric sulfide may mask other results. The black precipitate may fade or revert to negative if the enterotest ID is read after 24 h.

ADDITIONAL INFORMATION With Enterotest ID Yersinia is not confused with Proteus. When Yersiniae infection is suspected, motility will be observed at 25C and not at 352C. This characteristic is sufficient to differentiate this group from all others.7 Since there is no lactose in the media, no direct lactose fermentation is observed but, as the original colony was transferred from media containing lactose (SS agar, MacConkey, EMB, etc), this characteristic is already known before introduction in tube.8 For final identification and differentiation of E.coli and Citrobacter, a spot test on Simmons Citrate Agar is recommended. Citrobacter is positive in this test and E. Coli is negative.6,8 For differentiation between Enterobacter species and Serratia species,a few supplementary test are needed: Lysine; Ornithine, Arabinose, Sorbitol, Gelatin.6,8 For final identification and differentiation of Yersinia enterocolitica and Yersinia pseudotuberculosis a lysine decarboxylase test is recommended. Yersinia enterocolitica will give a positive result and Yersinia pseudotuberculosis will be negative.6-8
PRICE NUMBER OF TUBES /PLATES SIMPLICITY MOTILITY FLEXIBILITY FOR ADDITIONAL TEST TIME TO RESULT (LABOR TIME) INCUBATOR SPACE NEED OF EXTRA MATERIAL

ADVANTAGES OF ENTEROTEST VS OTHER METHODS

Conventional methods $$ 3* API $$$ Enterotest ID $ 6 reactions per tube easy yes yes fast small no

Cat N 20 reactions
complex no no medium small yes

hard-complex no no slow high yes

LIMITATIONS OF THE PROCEDURE Enterotest ID is designed specifically to identify Gram negative rods which are oxidase negative and should only be used to differentiate members of the Enterobacteriaceae family. Identification of Gram negative bacteria should be made with the consideration of additional characteristics such as source of specimen, history of the patient, colonial and microscopic morphology, serology and antimicrobial susceotibility aptterns.

*Kliger/Triple Sugar iron; urease broth, SIM

1. Steel, K. J. 1961. The oxidase reaction as a taxonomic tool. J. Gen. Microbiol. 25:297-306. 2. Farmer, J.J. III. 2003. Enterobacteriaceae: introduction and identification. In: Murray, P. R., E. J. Baron, J.H. Jorgensen, M. A. Pfaller, and R. H. Yolken (ed.). Manual of clinical microbiology, 8 th ed. American Society for Microbiology, Washington, D.C 3. Edwards and Ewing's identification of Enterobacteriaceae. 3rd ed. Burgess Publishing Co.,Minneapolis, Minnesota.. 1986 pp. ix + 536pp. 4. Kauffmann, f.; m. d. ]. The bacteriology of enterobacteriaceae. Collected studies of the author and his co-workers. 1966 pp. 400 pp. 5. R. Podschun, U. Ullmann. Klebsiella spp. as Nosocomial Pathogens: Epidemiology, Taxonomy, Typing Methods, and Pathogenicity Factors. Clin Microbiol Rev. 1998 October; 11(4): 589603. 6. Brenner, D,J, et al. Bergeys Manual of Systematic Bacteriology. Family I, Enterobacteriacae P595-602 7. Brubaker RR. The genus Yersinia: biochemistry and genetics of virulence. Curr Top Microbiol Immunol 1972;57:111-58 8. Ewing, W. H. 1962. Enterobacteriaceae. Biochemical tests for group differentiation. Public Health Service Publication no. 734 (revised). U.S. Department of Health, Education and Welfare, Washington, D.C.