Germ tube test

Introduction

The germ tube test for yeast identification is perhaps the most fundamental and simplest to perform. The germ tube test is one of the quickest methods for presumptive identification of Candida albicans and Candida dubliniensis. The yeasts isolated from clinical specimens are around 75% C. albicans, and germ tube tests usually enable reliable identification of this organism in a few hours.

Germ tubes are young hyphal-like extensions of yeast cells that develop without constriction at the place of origin from the yeast cell. C. dubliniensis, another Candida species, has been shown to produce true germ tubes. Although C. dubliniensis is relatively rare, it would require further biochemical or morphologic analysis to distinguish it from C. albicans. On the other hand C. tropicalis develops “pseudo-germ tubes,” which are constricted from the yeast cell at the base or tip of the germ tube. C. tropicalis isolates will be misidentified as C. albicans until laboratory finding distinguishs between actual germ tubes and pseudogerm tubes.

Serum or plasma, such as fetal bovine serum, is required for the standard test procedure. Leftover fresh-frozen plasma from a blood bank is suitable in this test and may be kept at 4 ° C can be used for longer duration too.. As an alternative, several additional liquid media (for example, BHI, soy broth trypticase, and nutritional broth) also can be utilised. The medium is inoculated with yeast and incubated at 37 ° C for 3 hours. It is critical not to incubate the test for over than 3 hours, as other species can develop germ tubes with prolonged incubation.

Procedure

• Using a wooden applicator stick, gently touch a yeast colony.
• Suspend the yeast cells in a fetal bovine serum tube.
• Incubate the tubes at 35° to 37°C for no longer than 3 hours.
• Place a drop of the suspension on a microscope slide.
• Place a coverslip over the suspension.
• Examine under high power for the presence or absence of germ tubes.

Result

A germ line originates from the yeast cell as a tiny lateral branch with no constriction (septum) near the yeast cell. Pseudohyphae or budding cells develop a constriction where the lateral extension meets the yeast cell. A minimum of 5 germ tubes should be examined before declaring the isolate positive.

True germ tubes lack constriction at their bases, where they adhere to the mother cell. If there is a constriction at the base of a germ tunnel, the yeast is other than candida. Such constricted germ tubes, known as pseudo-germ tubes, are more common in C. tropicalis. The inability of C. dubliniensis to grow at 42°C distinguishes it from C. albicans.

References:

• Linne & Ringsurd Clinical Laboratory Sience 6^th Edition
• Textbook of Diagnostic Microbiology 5^th Edition, Saunders Elsevier
• Clinical Microbiology Procedures Handbook 2^nd Edition, ASM Press
• Bailey & Scott’s Diagnostic Microbiology 13^th Edition, Elsevier