Exploration of Mycobiota in Cypripedium japonicum, an Endangered Species.

Orchids live with mycorrhizal fungi in mutualism. This symbiotic relationship plays an essential role in the overall life cycle of orchids from germination, growth, settlement, and reproduction. Among the 1000 species of the orchid, the Korean lady's slipper, Cypripedium japonicum, is known as an endangered species. Currently, only five natural habitats of the Korean lady's slipper remain in South Korea, and the population of Korean lady's slipper in their natural habitat is not increasing. To prevent extinction, this study was designed to understand the fungal community interacting in the rhizosphere of the Korean lady's slipper living in the native and artificial habitats. In-depth analyses were performed to discover the vital mycorrhizal fungi contributing to habitat expansion and cultivation of the endangered orchid species. Our results suggested that Lycoperdon nigrescens contributed most to the increase in natural habitats and Russula violeipes as a characteristic of successful cultivation. And the fungi that helped L. nigrescens and R. violeipes to fit into the rhizosphere community in Korean lady's slipper native place were Paraboeremia selaginellae and Metarhizium anisopliae, respectively. The findings will contribute to restoring and maintaining the endangered orchid population in natural habitats.

Cytological Study of Cypripedium japonicum Thunb. (Orchidaceae Juss.): An Endangered Species from Korea.

Changes in chromosome number and karyotype evolution are important to plant diversification, as they are both major drivers of speciation processes. Herein, chromosome number, karyotype, and genome size of the Korean lady's slipper orchid Cypripedium japonicum Thunb., an endangered species, were investigated in natural populations. Furthermore, all cytological data from this species are reported herein for the first time. The chromosome number of all investigated C. japonicum plants was diploid (2n = 2x = 22), with x = 11 as base chromosome number, whereby the species can now be clearly distinguished from the Japanese lady's slipper orchid. The karyotypes of all studied individuals were of similar length, symmetrical, and rather unimodal. Flow cytometry of the C. japonicum revealed that the genome size ranged from 28.38 to 30.14 pg/1C. Data on chromosome number and karyotypes were largely consistent with previous results indicating that Korean (x = 11) populations of C. japonicum are more closely related to Chinese populations (x = 11) compared to Japanese (x = 10) populations. These comprehensive cytological results will benefit the efforts to discriminate the geographically isolated and endangered Eastern Asian (China, Japan, and Korea) lady's slipper orchid species.

First Report of Gray Mold Disease on Endangered Species Cypripedium japonicum.

Cypripedium japonicum is known to be the indigenous plant to Korea, Japan, and China. However, C. japonicum represents the most critically endangered plant species in South Korea. The plant is esthetically pleasing due to its flower, which is larger than any other orchidaceous species. Disease symptoms relating to gray mold were observed on C. japonicum in May 2019. The suspected pathogen was successfully isolated from the symptomatic leaf tissue and conducted a pure culture of the fungi. The conidia formed consisted of a colorless or light brown single cell, which was either egg or oval-shaped with a size of 7.1 to 13.4 × 5.2 to 8.6 µm. Molecular phylogenetic relationship analysis was also confirmed that the pathogen concerned belonging to the family of Botrytis cinerea. Therefore, the findings confirmed that the pathogen isolated from C. japonicum was consistent with the unique properties of B. cinerea.

Distribution of Rhizosphere and Endosphere Fungi on the First-Class Endangered Plant Cypripedium japonicum.

Endangered native plant habitats and populations are rapidly disappearing because of climate and environmental changes. As a representative, the abundance of the first-class endangered wild plant, Cypripedium japonicum, has been rapidly decreasing in Korea. The purpose of this study was to evaluate the distribution of rhizosphere and endophytic fungi on C. japonicum in its native habitat. A total of 440 rhizosphere and 79 endosphere fungi isolates were isolated and identified on the basis of their molecular characteristics. Sixty-five genera and 119 fungi species were identified in this study. The genus Trichoderma showed the highest abundance among both rhizosphere and endosphere fungi. Mortierella, Hypocrea, and Penicillium spp. were also relatively dominant species on C. japonicum. The community structures of rhizosphere and endosphere fungi were similar, but endosphere fungi showed greater diversity.

Analysis of Fungicide Sensitivity and Genetic Diversity among Colletotrichum Species in Sweet Persimmon.

Anthracnose, caused by Colletotrichum gloeosporioides (C. gloeosporioides; Teleomorph: Glomerella cingulata), is the most destructive disease that affects sweet persimmon production worldwide. However, the biology, ecology, and genetic variations of C. gloeosporioides remain largely unknown. Therefore, in this study, the development of fungicide resistance and genetic diversity among an anthracnose pathogen population with different geographical origins and the exposure of this population to different cultivation strategies were investigated. A total of 150 pathogen isolates were tested in fungicide sensitivity assays. Five of the tested fungicides suppressed mycelial pathogen growth effectively. However, there were significant differences in the sensitivities exhibited by the pathogen isolates examined. Interestingly, the isolates obtained from practical management orchards versus organic cultivation orchards showed no differences in sensitivity to the same fungicide. PCR-restriction fragment length polymorphism (RFLP) analyses were performed to detect internal transcribed spacer regions and the ß-tubulin and glutamine synthetase genes of the pathogens examined. Both the glutamine synthetase and ß-tubulin genes contained a complex set of polymorphisms. Based on these results, the pathogens isolated from organic cultivation orchards were found to have more diversity than the isolates obtained from the practical management orchards.