Orchids and its mycorrhiza have a symbiotic relationship particularly during seed germination and can continue into the adult stage. Studies have shown that compatible orchid mycorrhiza may enhance plant growth when associating symbiotically in the roots of orchids such as the Malaysian endemic orchid, Paphiopedilum barbatum. Thus, study aims to identify the mycorrhizal fungi that form a symbiosis with the terrestrial tropical orchid, Paphiopedilum barbatum and determine root mycorrhization and growth in several in vitro Paphiopedilum plantlets. Mycorrhizal fungi were isolated from the roots of P. barbatum collected from five different locations in Peninsular Malaysia. Isolation of fungi was carried out according to the Currah method for initial morphological identification and then through molecular identification using the internal transcribed spacer (ITS) region of nuclear ribosomal DNA of the isolates. The identified mycorrhiza was then inoculated on in vitro selected Paphiopedilum plantletson a double slanting media to observe mycorrhization and the occurrence of enhancement of plant growth. Isolation of mycorrhizal fungi from the roots of P. barbatum in this study identified only one mycorrhiza; namely Tulasnella calospora (anamorph: Rhizoctonia repens) which is a ubiquitous orchid mycorrhiza. In vitro inoculation of T. calospora on seedlings from the Paphiopedilum genus after incubation for 24 weeks showed a significant 1.1% increase in fresh weight in P. rothschildianum with 68% root mycorrhization. However, no significant growth was observed in the seedlings of P. sanderianum, P. gigantifolium x P. rothschildianum and P. esquirolei x P. rothschildianum when inoculated with T. calospora after incubation for 24 weeks even though each had 32%, 24% and 13% root mycorrhization. This preliminary finding present an insight into the orchid mycorrhiza of the Paphiopedilum but a more extensive study is needed to further explore this symbiotic relationship and furthermore contribute in developing a beneficial application whether commercially or for the conservation of this genus.