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https://doi.org/10.37427/botcro-2024-001

Diversity of fungal endophytes isolated from the invasive plant Solanum rostratum

Nigora Kuchkarova ; State Key Laboratory of Desert and Oasis Ecology, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China
Caixia Han ; State Key Laboratory of Desert and Oasis Ecology, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China
Zokir Toshmatov ; State Key Laboratory of Desert and Oasis Ecology, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China; Institute of Genetics and Plant Experimental Biology, Academy of Sciences of the Republic of Uzbekistan, Kibray, Tashkent region 111208, Uzbekistan
Hongyang Chen ; State Key Laboratory of Desert and Oasis Ecology, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China
Hua Shao ; State Key Laboratory of Desert and Oasis Ecology, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China; Research Center for Ecology and Environment of Central Asia, Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Urumqi 830011, China; University of Chinese Academy of Sciences, Beijing 100049, China


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Sažetak

A culture-dependent method was used to isolate fungal endophytes from the leaves, stems, and roots of the invasive plant Solanum rostratum Dunal. growing in Xinjiang Province, China. All isolates were identified according to ITS (internal transcribed spacer) region of ribosomal DNA sequences and analyzed by Nucleotide BLAST according to NCBI GenBank and Mycobank database. Altogether 176 endophytic fungal isolates corresponding to 44 OTUs were identified, which were classified into 12 genera, with Penicillium (59.66%) and Aspergillus (23.29%) being the highly dominant genera. Ten endophytic isolates (OTU1, OTU15, OTU16, OTU21, OTU23, OTU25, OTU26, OTU30, OTU37 and OTU44) were identified as potential new species.

Ključne riječi

Culture-dependent endophyte isolation; endophytes; Penicillium; Mucor circinelloides

Hrčak ID:

309560

URI

https://hrcak.srce.hr/309560

Datum izdavanja:

1.4.2024.

Posjeta: 354 *




Introduction

Solanum rostratum Dunal. is an annual weed with a strong capacity for propagation and adaptation. A notorious invader, it also serves as the primary host of the potato leaf-roll virus and Leptinotarsa decemlineata (potato beetles), which pose substantial threats to biodiversity and the environment in China (Zhao et al. 2013, Liu et al. 2020). Additionally, S. rostratum contains abundant amounts of secondary metabolites, primarily flavonoids, alkaloids, steroids, and other compounds (Liu et al. 2020).

Endophytic fungi live in plants for all or part of their lives without harming the host (Ripa et al. 2019). They may affect a plant's ability to reproduce, grow, or resist abiotic stress or natural enemies (Rho et al. 2018). To the best of our knowledge, previous papers largely concentrated on the biological traits and phytochemical profile of S. rostratum and there is no report about the diversity of endophytic fungi of this invasive plant. The main goal of this study is to explore the community of the fungal endophytes of S. rostratum. Identification of the endophytes may help explain the invasive success of S. rostratum from the perspective of plant-microbe interaction; these endophytes are also potentially valuable resources of bioactive substances that have various biological activities.

Materials and methods

Forty-five mature S. rostratum plants at flowering stage were collected on June 28, 2018 from 3 different locations (15 plants from each location) in Urumqi and Changji city of Xinjiang province: location 1: 43º 55'60'' N, 87º 20'41'' E (Loc-1); location 2: 43º 56'0'' N, 87º 20'41'' E (Loc-2); location 3: 43º 46'14'' N, 87º 46'31'' E (Loc-3). Endophytes were isolated using a culture-dependent method within two days of collection of plants. Surface sterilization of plant parts (roots, stems, and leaves) and isolation of the endophytes were conducted following the protocol of Schulz et al. (1993). Colonization rate (CR) was counted by following Petrini et al. (1982).

DNA of the endophytic isolates was extracted by using the DNA Extraction Kit for fungi (Solarbio Life Sciences, Beijing, China), according to the manufacturer’s instructions (Abd-Elsalam et al. 2003). PCR amplification of the rDNA ITS (internal transcribe spacer) region was conducted with the use of ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') fungal primer pairs (White et al. 1990). The sequences of the fragments were identified using the basic local alignment search tool BLAST (http://www.ncbi.nlm.nih.gov/) of the NCBI and Mycobank (https://www.mycobank.org) databases. Fungal identities were generated by sequence alignment analysis with those previously submitted to GenBank.

Results and discussion

In total, 176 endophytic fungal isolates corresponding to 44 OTUs (operational taxonomic units) were isolated and were classified into 12 genera (Tab. 1). Among the isolates, 55 (31.25%) were obtained from leaves, 55 from stems (31.25%), and 66 from roots (37.50%); 34 out of 44 OTUs had between 97.14% and 100.00% sequence similarity with relevant entries in Mycobank and GenBank databases, whereas OTU1, OTU15, OTU16, OTU21, OTU23, OTU25, OTU26, OTU30, OTU37 and OTU44 had between 70.13% and 95.87% sequence similarity with species belonging to the genera Aspergillus, Penicillium, Microascus, Purpureocillium and Mucor (Tab. 1), indicating they might be potential new species. Penicillium and Aspergillus were the dominant genera of the endophytic fungal community (Cheng et al. 2018). OTU23 (closest hit Purpureocillium lilacinum CBS 284.36) was chosen for further study on its secondary metabolites due to its high plant growth regulatory activity, which resulted in the isolation and identification of 3 bioactive compounds, i.e., adenosine, cerevisterol, and thymine, which were found to possess significant plant growth regulatory activity (Kuchkarova et al. 2020).

Tab. 1. List of identified endophytic fungi isolated from Solanum rostratum plant parts. aBLASTN max score; blevel of identification for pairwise alignments by calculating using the Martinez-Needleman-Wunsch algorithm; clevel of similarity for pairwise alignments with the closest match, using the NCBI and Mycobank database; Accession number of the closest database match; daccession number of the closest database match. OUT - operational taxonomic unit.

Best Blast hitNumber of isolates
OTUAccession no.Closest taxa matchScore a

Query

coverage (%) b

Ident

(%) c

Accession no d By tissue typeBy location Total observed
LeafStemRoot123
1ON149677 Aspergillus lentulus 5569485.21PWQ23950011001
2ON149678 Penicillium oxalicum 84395100.00FMR 142616418184628
3ON149679 Pichia kudriavzevii 7119699.36CNRMA6.981001001
4ON149680 Aspergillus quadrilineatus 7869299.40IHEM 227051101012
5ON149681 Aspergillus rugulosus 7629299.20UOA/HCPF 100202035005
6ON149682 Emericella nidulans 7759498.22WM 06.1004206006
7ON149683 Aspergillus creber 6379497.42FMR 143641001001
8ON149684 Fusarium verticillioides 77594100.00IHEM 98350101001
9ON149708 Penicillium citrinum 7959699.60NRRL 18410021102
10ON149685 Aspergillus niger 85794100.00WM 10.767017018
11ON149686 Aspergillus nidulans 8059599.42WM 11.600021102
12ON149687 Penicillium brasilianum 84394100.00FMR 142960101001
13ON149688 Aspergillus oryzae 8529599.81WM 10.1200011001
14ON149689 Aspergillus tubingensis 84894100.00IHEM 174403324048
15ON149690 Penicillium rolfsii 6959595.10FMR 143070011001
16OM698374 Microascus cirrosus 1666270.13FMR 122560101001
17ON149691 Penicillium chrysogenum 83594100.00FMR 140081419155152848
18ON149692 Fusarium pseudonygami 73589100.00U345630011001
19ON149693 Fusarium oxysporum 7329398.77UOA/HCPF AB820011001
20ON149694 Aspergillus terreus 84892100.00WM 03.2180022002
21ON149695 Aspergillus calidoustus 6679595.87UOA/HCPF 92361001001
22ON149696 Aspergillus fumigatus 86395100.00ATCC 10221001001
23ON149697 Purpureocillium lilacinum 6247695.59CBS 284.3645243411
24ON149698 Penicillium coprophilum 8179999.24FMR 139981101102
25ON149699 Penicillium glabrum 5167594.12FMR 142920202002
26ON149700 Penicillium frequentans 5137594.09FMR 143180101001
27ON149701 Talaromyces pinophilus 8169499.42FMR 140170011001
28ON149702 Aspergillus aculeatus 79592100.00CBS 172.660041034
29ON149703 Paecilomyces lilacinus 7918398.85WM 04.4570110202
30ON149704 Aspergillus brasiliensis 7329595.48ATCC MY-A45531000101
31ON149705 Aspergillus flavus 83693100.00PWQ 23354000404
32ON149706 Penicillium echinulatum 7709498.48FMR 139450010101
33ON149707 Penicillium rubens 8279399.81FMR 138740010101
34ON149709 Penicillium crustosum 7268599.16FMR 14300610167
35ON149710 Penicillium allii 8089399.04FMR 142510010101
36ON149711 Penicillium commune 7649397.14CBS 311.480010011
37ON149712

Mucor circinelloides

f. circinelloides

7549192.19IHEM 241291000011
38ON149713 Geotrichum candidum 4829099.03WM 07.3041000011
39ON149714 Geotrichum bryndzae 4368298.93PMM09-440L1000011
40ON149715 Fusarium keratoplasticum 79893100.00FRC S-24650100011
41ON149716 Penicillium brevicompactum 7389397.67WM 06.3400520077
42ON149717 Alternaria alternata 81795100.00WM 04.4860100011
43ON149718 Penicillium griseofulvum 8139499.44CBS 185.270010011
44OM698376 Penicillium palitans 5077291.09FMR 142681000011
Total 555566723668176

The percentage of endophytic isolates belonging to Penicillium (59.66%; 105/176) was much higher than those identified as Aspergillus (23.29%; 41/176), Purpureocillium (6.25%; 11/176), Emericella (3.41%; 6/176), Fusarium (2.27%; 4/176), Paecilomyces (1.14%; 2/176), Geotrichum (1.14%; 2/176) as well as Altenaria, Microascus, Mucor, Pichia and Talaromyces, which were detected only sporadically (< 1%). The CR of the roots of the plant was higher (43.33%) than that of the stems (30.00%) and leaves (26.67%) of the identical plants. Furthermore, the CR of fungal endophytes of plants acquired from Loc-1 was much higher than that from Loc-2 and Loc-3.

To the best of our knowledge, this is the first report on the diversity of the endophytic fungi isolated from the invasive plant S. rostratum. This study demonstrated the comparatively high multiplicity of the endophytic fungi of S. rostratum from three locations in Xinjiang. Our work revealed that the invasive plant S. rostratum harbours a variety of fungal endophytes in its leaves, stems, and roots. Given the fact that endophytes are able to produce biologically active secondary metabolites that affect the growth of their hosts, we speculate that the endophytic fungi might contribute to the invasive success of S. rostratum.

Acknowledgments

This research work is financially supported by the National Foreign Experts Project granted to Nigora Kuchkarova (QN2022045006L), the CAS President’s International Fellowship Initiative (PIFI) granted to Zokir Toshmatov (2020PB0010), and the Natural Science Foundation of Xinjiang Uygur Autonomous Region (2022D01D02).

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