BIODIVERSITAS ISSN: 1412-033X Volume 21, Number 4, April 2020 E-ISSN: 2085-4722 Pages: 1678-1685 DOI: 10.13057/biodiv/d210449 Short Communication: New record of Stenotrophomonas sp. as endosymbiont bacteria in Rhizopus microsporus DEWI PETI VIRGIANTI 1,3,♥ , DESSY NATALIA 2 , I NYOMAN PUGEG ARYANTHA 1 1 Doctoral Program of Biology, School of Life Sciences and Technology, Institut Teknologi Bandung, Jl. Ganesa 10, Bandung 40132, West Java, Indonesia 2 Biochemistry Research Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesa 10, Bandung 40132, West Java, Indonesia 3 Program of Medical Laboratory Technique, Sekolah Tinggi Ilmu Kesehatan Bakti Tunas Husada, Jl. Cilolohan 36, Tasikmalaya 46115, West Java, Indonesia. Tel.: +62-265-334740,  email: dewivirgianti@gmail.com Manuscript received: 9 December 2019. Revision accepted: 27 March 2020. Abstract. Virgianti DP, Natalia D, Aryantha INP. 2020. Short Communication: New record of Stenotrophomonas sp. as endosymbiont bacteria in Rhizopus microsporus. Biodiversitas 21: 1678-1685. Rhizopus sp. are closely related to humankind in many aspects of human life. Several species of Rhizopus are important in food, agriculture, and health industries. Rhizopus microsporus has endosymbiont bacteria that has been identified as Burkholderia sp. This study aimed to identify the endosymbiont bacteria in R. microsporus isolated from Moringa oleifera Lam leaves. Ring technique, simple method of agar heap technique and antibiotic technique, was conducted to purify the hyphae and to eliminate ectosymbiont and contaminant bacteria outside the hyphae. The presence of endosymbiont bacteria in R. microsporus was determined by using LIVE/DEAD® BacLight Bacterial Viability Kits L13152 and the Fluorescent in Situ Hybridization (FISH) method using EUB338 probes. It was suggested that the culturable endosymbiont bacteria was identified as Stenotrophomonas sp. based on the phylogenetic tree using the 16S rDNA Keywords: Bacterial-fungal interaction, BFI, EHB, endo hyphal bacteria, endosymbiont, FISH, Rhizopus INTRODUCTION Many species of Rhizopus are used in human life, as bioprocess agents in the fermented food industry (Nout and Aidoo 2010) and enzyme industries, and contain various secondary metabolites (Ghosh and Ray 2011; Freitas et al. 2014). However, some species of Rhizopus can cause diseases in plants (Ghosh et al. 2015), post-harvest damage especially in fruits (Kwon et al. 2011) and produce harmful toxins to human (Rohm et al. 2010). Some species of Rhizopus play a role in mucormycosis, especially in immunosuppressed patients (Dolatabadi et al. 2016). Mondo et al. (2017) reported that Burkholderia sp. is mutualistic symbiotic bacteria controls sexual reproduction in Rhizopus sp. However, endosymbiont Burkholderia rhizoxina in R. microspores produced rhizoxin ten times in pure culture compared to that in symbiotic forms (Scherlach et al. 2006). The ability of the fungus to produce rhizoxin depends on the presence of endosymbiont bacteria (Partida-Martinez and Hertweck 2005). The first mycotoxin rhizonin is a hepatotoxic cyclopeptide produced in the symbiotic form of low-level fungus R. microsporus van Tieghem with Burkholderia bacteria (Partida-Martinez et al. 2007). The transmission process of Burkholderia sp. to R. microsporus occur horizontally and vertically (Partida-Martinez et al. 2007; Lackner et al. 2009). Chitinase encoded by the type 2 secretion system (T2SS) gene cluster is used to enter hyphae (Moebius et al. 2014). The symbiosis between Rhizopus sp. and Burkholderia sp. did not occur in every isolate of Rhizopus. Of the six samples of R. microsporus originating from the soil, rice seedlings, and human body tissues, only four species had symbiosis with Burkholderia, namely R. microsporus var microsporus and R. microsporus van Tieghem var Chinensis (Partida-Martinez and Hertweck 2005). Scherlack et al. (2006) succeeded in finding six different symbionts from a total of 15 R. microsporus isolates. The study by Dolatabadi et al. (2016) showed only seven strains (11%) were positive containing endosymbiont of a total of 64 Rhizopus sp. strains originating from various clinical, environmental, and food samples. Data showed that 85% of strains containing endosymbiont bacteria were non-clinical samples. The endosymbiotic occurs in Rhizopus sp. originating from various habitats and could not be linked to specific habitats. Data obtained from various studies indicated that the species of R. microsporus found symbiosis with B. rhizoxina and B. endofungarium were derived from soil, rice seedlings, peanuts, Vietnamese Sufu starter, and tempeh (Lacker et al. 2009; Rohm et al. 2010; Dolatabadi et al. 2016). A different genus of endosymbiont bacteria has been isolated from the clinical isolate of R. microsporus was identified as Ralstonia pickettii contributing to fungal stress resistance and immune cell evasion as part of fungal pathogenesis in animal models (Itabangi et al. 2019). The aim of this study was to determine and identify endosymbiont bacteria in Rhizopus sp. isolated from different sources, namely Moringa oleifera leaves.