Motility Tests : Principle, procedure, result interpretation
Principle of Motility tests
Organisms are motile due to flagella and he motility tests use to detect the presence of ﬂagella by bacteria, allowing them to travel in and out of the microscopic ﬁeld or beyond their initial inoculation in agar. For the wet preparation, a drop of organism in broth is suspended on a clean and grease free glass slide, a cover slip is added, and the culture is observed microscopically for motility. Occasionally the organism is incubated in the broth prior to examination where as In the tube test, semisolid motility medium like SIM (Sulfide, Indole, Motility) or MIU (Motility, Indole, Urea) or MIO (Motility, Indole, Ornithine) is inoculated in a straight line down through the center of a tube. Motile organisms will migrate out from the line of inoculation, causing visible turbidity throughout the tube. Non-motile organisms will grow only along the line of inoculation.
Requirements for Motility tests
Organisms tested-Campylobacter; Legionella; enterococci; Enterobacteriaceae; Listeria; Bacillus; other gram-positive rods; non-glucose-fermenting, gram-negative rods; and any other organism where motility is useful for identiﬁcation
At 30°C for non-fermenting, gram-negative rods and enterococci for 24 hours.
If there is a question regarding a negative result, incubate at 25°C.
For Listeria and Yersinia, incubate two tubes, one at 35°C and one at 25°C.
Result and interpretation of Motility tests
A. Wet mount preparation
Motility test positive:organisms change position with respect to one another
Motility test Negative: organisms do not change position with respect to one another
Brownian movement:It is random jiggling or shaking due to molecular bombardment, where the organisms remain in the same relative position with respect to each other, should not be mistaken for true motility. Campylobacter displays a very active motility which appears as tiny dots darting in and out of the ﬁeld.
B. Tube media
Motility Test positive:Diffuse growth outward away from stab line or turbidity of the media
Test negative or non motile:A clear tube (the same as the uninoculated media) with growth only along the line of inoculation.
Key notes of Motility tests
Enterococcus casseliﬂavus and E. gallinarum are motile.
Listeria organisms are motile at 25°C but not at 35°C, with a characteristic umbrella-shaped growth at the top of the tube. On wet mount they exhibit tumbling motility.
Bacillus spp. should be motile but lack of motility could indicate Bacillus anthracis.
Yersinia enterocolitica is motile at 25°C but non-motile at 35°C.
Acinetobacter species are non-motile.
Non-fermenting, gram-negative rods and Enterobacteriaceae vary in their motility.
Limitations of Motility tests
Motility results for enterococci in MIO have been reported to have poor sensitivity.
.Excessive heat on a microscope slide can affect the test results.
False-negative reactions may occur if bacterial ﬂagella are damaged due to heating, shaking, or other trauma and such environmental shock will render the organism non-motile.
Some microorganisms do not express ﬂagellar proteins at 35 to 37°C but do so at 22°C.
Bacillus species are best tested directly from a fresh agar plate. If a fresh plate is not available, inoculate a plate and incubate for 4 hours. Then perform wet mount.
A large number of Enterococcuscasseliﬂavus and Enterococcusgallinarum organisms have been reported as non-motile using some tube motility agar. If a vancomycin MIC is between 4 to 16 µg/ml and the isolate is ampicillin susceptible, but the enterococcus is non-motile, conﬁrm results with the 2 hours broth method or perform the MGP test
Cowan and Steel’s ,manual for the identification of medical bacteria
Lynne S. Garcia ,Clinical Microbiology Procedures Handbook