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HiStrep™ Identification Kit

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KB005A

Intended Use

KB005A is a standardized, colorimetric identification system, a combination of 12 tests for identification of Streptococcus species from clinical specimen and non clinical samples using pure isolate.

Kit Contains

  1. Each kit contains 5/10/20 kits of KB005A, sufficient material to perform 5/10/20 tests. Kit contains sterile media for Voges Proskauer's, Esculin hydrolysis, PYR, ONPG, Arginine utilization tests and 7 different carbohydrates utilization tests - Glucose, Lactose, Arabinose, Sucrose, Sorbitol, Mannitol, Raffinose
  2. Baritt reagent A (R029) for VP test
  3. Baritt reagent B (R030) for VP test.
  4. PYR reagent (R043) for PYR test
  5. Technical product insert.
  6. Result Interpretation Chart and Result Entry Datasheet.

Material Required but not Supplied

  • 1.McFarland standard
  • 2.Inoculation loops, pipettes
  • 3.Enrichment medium / Isolation media

Direction

Preparation of inoculum

  • KB005A cannot be used directly for clinical specimens. The microorganisms to be identified have to be first isolated on appropriate isolation media. Only pure cultures should be used.
  • Isolate the organism to be identified on a common medium like Nutrient Agar (M001) or Tryptone Soya Agar (M290). Pick up a single isolated colony and inoculate in 5 ml BHI Broth (M210) and incubate at 35-37°C for 4-6 hours until the inoculum turbidity is 0.1 OD at 620nm or 0.5 McFarland standard. Some organisms may require more than 6 hours of incubation. In this case incubate till the inoculum turbidity reaches 0.1 OD at 620nm.
  • Alternatively, prepare the inoculum by picking 1-3 well isolated colonies and make a homogeneous suspension in 2-3 ml sterile saline. The density of the suspension should 0.1 OD at 620nm.

Inoculation of the kit

  1. Open the kit aseptically. Peel off the sealing foil.
  2. Inoculate each well with 50 µl of the above inoculum by surface inoculation method.
  3. Alternatively, the kit can also be inoculated by stabbing each individual well with a loopful of inoculum

Incubation

Temperature of incubation: 35±2°C. Duration of incubation: 18-24 hours.

Interpretation of results

Interpret results as per the standards given in the identification index. Addition of reagents in well no 1 and 3 should be done at the end of incubation period that is after 18- 24 hours.

Principle

KB005A is a standardized, colorimetric identification system utilizing twelve conventional biochemical tests. The tests are based on the principle of pH change and substrate utilization. On incubation, organisms undergo metabolic changes which are indicated as a colour change in the media that is either visible spontaneously or after addition of a reagent(1).

Type of Specimen

Pure isolate from clinical specimen and non clinical sample

Specimen Collection and Handling

Refer direction

Warning and Precautions

In Vitro diagnostic Use. For professional use only. Read the label before opening the pack. Wear protective gloves/ protective clothing/eye protection/ face protection. Follow good microbiological lab practices while handling specimens and culture. Standard precautions as per established guidelines should be followed while handling clinical specimens. Aseptic conditions should be maintained during inoculation and handling of the kits. Reagents should not come in contact with skin, eyes or clothing. Safety guidelines may be referred in individual safety data sheets.

Limitations

  1. Allow the reagents to come to room temperature after removal from the refrigerator.
  2. In case of carbohydrate fermentation test some microorganisms show weak reaction. In this case record the reaction as± and incubate further upto 48 hours. Orange colour after 48 hours of incubation should be interpreted as a negative reaction.
  3. At times organisms give conflicting result because of mutation or the media used for isolation, cultivation and maintenance.
  4. The identification index has been compiled from standard references and results of tests carried out in the laboratory.
  5. Erroneous false negative results may be obtained if the inoculum turbidity is less than 0.1 OD.
  6. Results are more prominent if an enriched culture is used instead of a suspension.
  7. It cannot be used directly for clinical specimens. The microorganisms to be identified have to be first isolated on appropriate isolation media. Only pure cultures should be used.

Performance and Evaluation

Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at recommended temperature.

Quality Control

Appearance

Sterile white opaque strip with 12 wells containing media for Voges Proskauer's, Esculin hydrolysis, PYR, ONPG, Arginine utilization tests and 7 different carbohydrates utilization tests - Glucose, Lactose, Arabinose, Sucrose, Sorbitol, Mannitol, Raffinose.

Quantity of medium

0.8 ml of medium in each well.

Sterility Check

Passes release criteria

Interpretation of results

Interpret results as per the standards given in the identification index. Addition of reagents well no 1 and 3 should be done at the end of incubation period that is after 18 - 24 hours.

Voges-Proskaeur's Test: Well No.1

  • Add 2-3 drops of Baritt reagent A (R029) and 1 drop of Baritt reagent B (R030)
  • Positive test is indicated by a development of pinkish red colour in 5 - 10 minutes
  • No colour change or a copper colour (due to reaction of Reagent A and Reagent B) indicates a negative reaction

Esculin hydrolysis: Well No. 2

  • Cream colour changes to black colour indicates a positive reaction.
  • No colour change indicates negative reaction.

PYR test: Well No. 3

  • Add 1-2 drops of PYR reagent (R043).
  • Positive test is indicated by development and retention of cherry red colour.
  • Development of pink, orange or yellow colour indicates a negative reaction.

ONPG: Well No. 4

  • Change of colour from colourless to yellow indicates positive reaction.
  • No colour change indicates negative reaction.

Arginine utilization: Well No. 5

  • Olive green to light purple colour changes to dark purple colour indicates positive reaction.
  • No change in colour or yellow colour indicates negative reaction.

Carbohydrate fermentation Test: Well No. 6-12

  • Positive test is indicated by a colour change to yellow colour.
  • Red or no colour change indicates a negative reaction.

Result Interpretation Chart

No. Test Reagents to be added after incubation Principle Original colour of the medium Positive reaction Negative reaction
1. Voges Proskauer's 1-2 drops of Baritt reagent A and 1-2 drops of Baritt reagent B Detects acetoin production Colourless / light yellow Pinkish red Colourless/ slight copper
2 Esculin hydrolysis — Detects Esculin hydrolysis Cream Black Cream
3. PYR 1-2 drops of PYR reagent Detects PYR enzyme activity Cream Cherry Red Cream
4 ONPG — Detects -βgalactosidase activity Colourless Yellow Colourless
5 Arginine utilization — Detects Arginine decarboxylation Olive green to Light purple Purple / Dark purple No Change in colour or yellow
6 Glucose — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
7 Lactose — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
8 Arabinose — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
9 Sucrose — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
10 Sorbitol — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
11 Mannitol — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink
12 Raffinose — Carbohydrate utilization Pinkish Red / Red Yellow Red / Pink

KB005A: HiStrep™ Identification Kit (Used for Differentiation of Streptococcus Species) Identification Index of Various Streptococcus Species

Tests
Voges Proskauer's Esculin hydrolysis PYR ONPG (β-galactosidase) Arginine Utilization Glucose Lactose Arabinose Sucrose Sorbitol Mannitol Raffinose
S. anginosus + + nd d + + d + — — — v
S. mitis — d d d — + + + — v d d
S. oralis d (-) nd d (-) + + nd nd — — d
S. pneumoniae — d d d (+) + + + slow — — — +
S. porcinus + (+) — — + + d nd nd + + +
S. pyogenes — (-) + — + + — — — — — —
S. salivarius (+) + — — — + (+) — + (-) + —
S. sanguis — d — d + + — + — — — —
S. suis nd + — nd + + d — + — + —
S. equii spp. zeoepidemicus — (-) — — — + + (+) + — — —
S. agalacti — + — — — + + d + — — —
S. adjacens nd nd + — — + — nd nd — — —
S. acidominimus — + — (-) (-) + + — — — — v
S. bovis nd + — (+) — + + d + — d nd
S. defectivus nd nd + — — + d nd nd — — v
S. dysgalactiae — + — — — + + + + — — —
S. equinus + + — — — + — — — — — —
S. constellatus — + nd d (+) + + nd nd + + (-)
S. canis — + + — — + + — + — + (-)
S. equi spp. equi — (-) — — — + + — + — — —
S. mutans + + + v — + + — + + nd +
S. uberis nd + — — — + + + + (+) + —
S. faecalis + + + + + + + — + + + +

Note: Based on % strains showing reactions following symbols have been assigned from laboratory results and standard references.
0-10% = -
11-25% = [-]
26-75% = d
76-89% = [+]
90-100% = +
ND = not detected v = variable reaction

Storage and Shelf Life

On receipt store between 2-8°C. Use before expiry date on the label. Product performance is best if used within stated expiry period.

Disposal

After use, kits and the instruments used for isolation and inoculation (pipettes, loops etc.) must be disinfected using a suitable disinfectant and then discarded by incineration or autoclaving in a disposal bag. Follow established laboratory procedures in disposing of infectious materials and material that comes into contact with clinical sample must be decontaminated and disposed of in accordance with current laboratory techniques (2,3).

More Information
Product Name HiStrep™ Identification Kit
SKU KB005A
Customized Product Available No
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