MYCOFAST® Screening EvolutioN 3

Intended use

Mycoplasmas are the smallest and simplest of the procaryotypes capable of self-reproduction (0.15 to 0.25 μm). They differ from other bacteria in their lack of a cell wall and hence a natural resistance to ß-lactams. Since mycoplasmas are relatively fragile, they will only grow in acellular cultures in the presence of various growth factors and at a constant temperature of 35 to 37 °C.

Most human mycoplasmas are commensal. Of the 9 species that have been isolated from the urogenital tract, Ureaplasma urealyticum and Mycoplasma hominis are the most commonly found. U.u. and M.h. are sexually transmitted and can be pathogenic. Respiratory infections or meningitis can occur in the neonate as a result of contamination from the genital tract at birth. In adults, the infections caused by U.u. and M.h. are described in the table below:

Conventional diagnosis is based upon culture on A7 agar plates followed by microscopical identification of U.u. (sea urchin shaped) or M.h. (fried-egg shaped) colonies.
Since both U.u. and M.h. are commensal, infection can only be diagnosed through the determination of the pathological threshold, followed by precise enumeration.

MYCOFAST® Screening EvolutioN 3 gives the solution and makes urogenital diagnosis much easier.

Principle

MYCOFAST® Screening EvolutioN 3 identifies U.u. and M.h. growth after incubation in a liquid medium. During growth, U.u. and M.h. metabolize urea and arginine respectively resulting in a color change of the medium, which contains phenol red indicator, from yellow-orange to red. This color change is due to liberation of ammonia resulting in an alkaline pH of the medium.

Mycoplasma growth thus viewed enables:

• Screening in UMM medium; and in case of positivity:
• Enumeration of mycoplasma based on the rate of urea or arginine hydrolysis, which is proportional to the number of germs contained in the sample.
  U.u:  > 10³, 10⁴ and 10⁵ UCC/ml
  M.h: > 10⁴ UCC/ml
• Identification based on the sensitivity or otherwise of the germ to three antibiotics (lincomycin, SXT and erythromycin) = Identibiotic.
• U.u and M.h susceptibility testing to 7 antibiotics :
  Doxycycline (DO) 4-8 μg/mL ,  Pristinamycin (PT) 2 μg/mL , Roxithromycin (ROX) 1-4 μg/mL, Azithromycin (AZM) 0.5-4 μg/mL, Josamycin (JM) 1-4 μg/mL, Ciprofloxacin (CIP) 1-2 μg/mL et Ofloxacin (OFX) 1-4 μg/mL.

Simple protocol

Easy-to-read and easy-to-interpret results

The results are read by the color obtained in the different wells. Urogenital mycoplasma growth is indicated when the medium turns red (alkaline). The medium remains yellow when no growth of urogenital mycoplasma occurs.

Reagents and material

• UMMt:
  • Vial of mycoplasma transport medium (3 mL)
  • Ref. 00042 Quantity: 35
  • Ref. 00077 Quantity: 35
• UMMlyo:
  • Vial of lyophilized growth medium (+3 mL UMMt)
  • Ref. 00042 Quantity: 35
  • Ref. 00077 Quantity: 35
• MYCOFAST® EvolutioN 3 trays:
  • Tray of 20-well wrapped individually in an aluminum sachet
  • Ref. 00042 Quantity: 12
• S.M.h:
  • Mycoplama hominis growth activator (4.5 mL)
  • Quantity: 1

Stability and storage

• All the reagents are ready-to-use. The vials may be stored at 2-8 °C, in their original packaging until the expiry date shown on the kit.
• The UMMt medium may be stored temporarily at room temperature but is more stable at 2-8 °C.

Performance
1- Screening in UMM medium

A study was carried out on 332 clinical specimens (196 endocervical specimens, 10 urethral specimens, 110 sperm specimens, 5 urine specimens, 3 celioscopy specimens and 5 miscellaneous specimens) of which 85 specimens were detected as positive (78 positives for U.u and 21 positives for M.h). After 24 and 48 hours of incubation the color change of the seeded UMM medium (500μL) was compared to the results of the enumeration obtained with the direct MYCOFAST® EvolutioN 3 method. The screening results are summarized below:

2- Detection and numeration in tray

• In the same study the results obtained directly with the MYCOFAST® EvolutioN 3 method were compared with those obtained with a second MYCOFAST® EvolutioN 3 tray, inoculated after conservation of the seeded medium for 24 to 48 hours:
• For clinical interpretation the agreement of the results is 85.9% for U. urealyticum and 76.2% for M. hominis, with a total agreement of 83.8%.
• 21/78 positive specimens in U.u and 4/21 positive specimens in M.h showed a numeration lower than that obtained with the direct method.

3- Antimicrobial susceptibility testing

• Please refer to MYCOFAST EvolutioN 3 webpage.

Material required but not provided

• Sample collecting material (Swabs, cytobrushes, sterile containers for liquid samples)
• Pipettes and tips
• Waste container for contaminated waste
• Paraffin oil
• Incubator at 37 ± 1 °C