Abstract
Two Diaporthe species isolated from fruit of Citrus sinensis in China were characterized based on morphology and multilocus phylogeny of ITS, tef1, and tub2 gene sequences. The phylogeny indicated that the two species match Diaporthe taoicola and D. siamensis. A critical examination of phenotypic characteristics confirmed the phylogenetic results. Diaporthe taoicola was morphologically characterized by producing Alpha conidia with tapering toward both ends. Meanwhile, D. siamensis produced cylindrical or ellipsoidal Alpha conidia with two oil drops. Pathogenicity tests revealed that both species were pathogenic to fruit of C. sinensis. To our knowledge, the two species were firstly reported on Citrus sinensis in China.
Keywords: Diaporthe, phylogeny, morphology, pathogenicity, Citrus sinensis
Citrus (Rutaceae) is worldwide cultivated because of its nutritional values and the medicinal benefits (e.g., anti-hypertensive) [1]. In 2018, citrus production in Zigui county, Hubei province, the larger Citrus sinensis production area in China, has reached 23.3 thousand ha. As citrus has diversified as a commercial crop, it became a host for various pathogens from nursery to the storage stage.
Diaporthe species are present as pathogens, endophytes, or saprophytes on a wide range of cultivated or wild trees and ornamentals [2–5]. Species in this genus have been reported as the pathogens of blight, canker, decay, dieback, wilt, leaf spot, fruit rot and root rot across a diverse range of plant species [6–9]. Members of Diaporthe are also frequently associated with citrus diseases worldwide [10,11]. Diaporthe citri (anamorph=Phomopsis citri) caused melanose and stem-end rot of fruit, which are important in most citrus-growing areas with high humidity. Besides, D. foeniculina has also been found from New Zealand, Spain and USA associated with stem end rot on fruit [11]. In China, Diaporthe citri, D. eres and D. unshiuensis have been reported on fruit of Citrus spp. [12].
Preliminarily, species in Diaporthe were identified mainly based on morphological characters and host associations. However, morphology has been conferred to be inconsistent for identification due to inter- and intra-species variability [13]. Molecular analyses inferred that Diaporthe species are not highly host-specific [14]. More than one species is often present on one host, or one species may occur on more than one host [15]. Studies converging on the diversity of Diaporthe have been progressed in recent years in China. Huang et al. (2015) studied Diaporthe on Citrus in China found eight known species and seven novel species based on morphological comparison and multi-gene analyses. Moreover, Diaporthe associated with peach trees [16], pear shoot canker [5], and dieback diseases involving 16 host genera [17] were reported in China. These references provided bounteous information for the study of Diaporthe in China.
During the investigation of fungal pathogens associated with Citrus sinensis in Zigui county, Diaporthe isolates were encountered based on morphology. Three Diaporthe-like isolates, YZU 181047, YZU 181403, and YZU 181223 were found pathogenic to fruit of C. sinenesis. The main objectives of this study were to identify them based on morphological observations and sequence analyses of multiple gene regions.
In 2018, diseased citrus fruit was collected from commercial orchards in Zigui county. Tissues from the margin of infected lesions were cut into segments, which contained both diseased and healthy parts. All segments were surface sterilized in 2% sodium hypochlorite for 2 min, followed by 75% ethanol for 30 s and rinsed in sterile distilled water for three times. All samples were dried with sterile filter paper and plated onto potato dextrose agar (PDA, Difco, USA). Plates were incubated at 25 °C in darkness until mycelia grow. Then, mycelia from colony margin were taken and transferred on fresh PDA plates. Pure cultures were stored in the Fungi Herbarium of Yangtze University (YZU) in Jingzhou, China.
Genomic DNA was extracted from mycelium developed on PDA medium according to Cenis [18]. The primers ITS4 and ITS5 [19] was used to amplify the ITS region of the nuclear ribosomal RNA operon, including the 3′ end of the 18S rRNA, the first internal transcribed spacer region, the 5.8S rRNA gene; the second internal transcribed spacer region and the 5′ end of the 28S rRNA gene. The primers EF1-728F and EF1-986R [20] were used to amplify part of the translation elongation factor 1-α (tef1) gene, and the primers Bt2a and Bt2b [21] were used to amplify the partial beta-tublin (tub2) gene. The PCRs were performed in a 25 μL reaction mixture consisted of 12.5 μL of 2 × Taq PCR StarMix (Genstar, Beijing, China), 2 μL genomic DNA, 1.25 μL of each primer, and 8 μL distilled water (ddH2O). The thermal cycling program was completed on a thermal cycler using the following conditions: initial denaturation at 94 °C for 2 min, followed by 35 cycles of denaturation at 94 °C for 60 s, annealing at (52 °C for ITS, and 56 °C for tef1, and 60 °C for tub2) for 30 s, extension at 72 °C for 60 s, with a final extension step at 72 °C for 5 min. Successful PCR amplification products were purified and sequenced at BGI (Beijing Genomics Institute).
All the obtained sequences were analyzed in the basic-local-alignment search tool (BLASTn) (http://blast.ncbi.nlm.nih.gov/) to retrieve the most similar taxa sequences. Relevant sequences were selected from the studies of Gomes et al. [3], Dissanayake et al. [16], Yang et al. [17], and Tibpromma et al. [22]. All sequences were aligned and combined in the MEGA 7.0 program [23]. Maximum Parsimony (MP) analysis was performed in PAUP version 4.0 b10 [24], generating a heuristic search option of 1000 random-addition replicates and a tree bisection-reconnection (TBR) as a branch-swapping algorithm. MaxTrees were set to 1000, branches of zero length collapsed, and all equally parsimonious trees were saved. Other scores in parsimony were calculated as tree length (TL), consistency index (CI), retention index (RI), and rescaled consistency (RC). The maximum likelihood (ML) [25] phylogeny of the combined dataset was constructed with 1000 bootstrap replicates using GTRGAMMAI model. Additionally, Bayesian (BI) analysis was conducted in MrBayes v. 3.2.6 with 1,000,000 Markov chain Monte Carlo (MCMC) generations and a sampling frequency of every 100th generations. The best-fit evolutionary model was determined via MrModelTest v. 2.3 [26]. At the end of the analysis, the first 25% of the samples were excluded as burn-in, and consensus trees were generated using the 50% majority-rule consensus tree criteria. The tree was viewed in Figtree v.1.3.1 [27]. Posterior probability (PP) values of BI analysis and bootstrap (BP) values of ML and MP analyses were shown at the nodes of branches. The out-group of the phylogeny was Diaporthella corylina CBS 121124.
Three isolates were characterized for their colonial and conidial morphology. Agar plugs (6 mm diam.) of each isolate were taken from the edge of actively growing cultures and transferred onto the center of petri dishes (9 cm diam), containing potato dextrose agar (PDA) and oatmeal agar (OA) for cultural feature. Plates containing 2% water agar (WA) with autoclaved Citrus sinensis leave tissues were incubated at 25 °C under a 12-h near-ultraviolet light/12-h dark cycle to induce sporulation. The culture was checked periodically for the development of ascomata and conidiomata. Morphology was recorded including colony color, texture, microconidia, and sporocarps formation. Conidia were mounted in sterile water for microscopic observation using a light microscope (Nikon DS-Ri2, Tokyo, Japan), equipped with a Nikon DS-Ri2 digital camera. Conidia (n = 50) were measured for each species.
Pathogenicity tests were performed on fruit of Citrus sinensis. Mature and healthy fruit were surface-sterilized in 2% sodium hypochlorite for 2 min and washed three times with sterile distilled water. Fruit was wounded with a sterile scalpel around 6 mm × 6 mm in size. The mycelial plugs from 3 days old cultures grown on PDA were transferred onto the wounds. Controls were treated with sterile PDA. The inoculated fruits were maintained at 25 °C and 80 to 100% relative humidity (RH). The development of disease symptoms was checked daily for one week. The pathogen was re-isolated from the inoculated fruit and identified based on morphology to satisfy Koch's postulates. The pathogenicity tests were conducted with three replicates for each isolate and repeated three times.
A total of nine new sequences were generated and deposited in GenBank (Table 1). The combined multi-gene phylogeny (ITS, tef1, and tub2) contained 30 strains, of which 27 were obtained from NCBI (https://www.ncbi.nlm.nih.gov/) (Table 1, Figure 1). A total of 1276 characters (ITS 494, tef1 377, tub2 405) were included after alignment. Among them, 804 were constant, 190 were variable, and 282 were parsimony uninformative. The heuristic search generated 5 parsimonious trees (TL = 908, CI = 0.664, HI = 0.336, RI = 0.819, RC = 0.544). For the BI analysis, the HKY + I model was recommended by MrModeltest. The topology of ML phylogeny was identical to the results of BI and MP analyses, and it was used as a basal tree.
Table 1.
Isolates and GenBank accession numbers used in the phylogenetic analyses of Diaporthe.
| Species | Strain | Host/Locality | GenBank accession number |
||
|---|---|---|---|---|---|
| ITS | tef1 | tub2 | |||
| Diaporthe arecae | CBS 161.64 | Areca catechu/India | KC343032 | KC343758 | KC344000 |
| Diaporthe arengae | CBS 114979 | Arenga engleri/China, HongKong | KC343034 | KC343760 | KC344002 |
| Diaporthe batatas | CBS 122.21 | Ipomoea batatas/USA | KC343040 | KC343766 | KC344008 |
| Diaporthe citri | CFCC 53079 | Citrus sp./China | MK573940 | MK574615 | MK574635 |
| CFCC 53080 | Citrus sp./Brazil | MK573941 | MK574616 | MK574636 | |
| Diaporthe endophytica | CBS 133811 | Schinus terebinthifolius | KC343065 | KC343791 | KC344033 |
| Diaporthe eugeniae | CBS 444.82 | Eugenia aromatica/Indonesia | KC343098 | KC343824 | KC344066 |
| Diaporthe oxe | CBS 133186 | Maytenus ilicifolia/Brazil | KC343164 | KC343890 | KC344132 |
| Diaporthe perseae | CBS 151.73 | Persea americana/Netherlands | KC343173 | KC343899 | KC344141 |
| Diaporthe pseudomangiferae | CBS 101339 | Mangifera indica/Dominican Republic | KC343181 | KC343907 | KC344149 |
| Diaporthe pseudophoenicicola | CBS 462.69 | Phoenix dactylifera/Spain | KC343184 | KC343910 | KC344152 |
| Diaporthe siamensis | MFLUCC 10-0573a | Dasymaschalon sp./Thailand | JQ619879 | JX275393 | JX275429 |
| MFLUCC 10-0573 b | Dasymaschalon sp./Thailand | JQ619880 | JX275395 | JX275430 | |
| MFLUCC 10-0573c | Dasymaschalon sp./Thailand | JQ619881 | JX275396 | JX275431 | |
| MFLUCC 17-0591 | Pandanaceae/Thailand | MT908796 | MG646989 | MG646925 | |
| YZU 181403 | Citrus sinensis/China | MW160357 | MW160363 | MW160360 | |
| Diaporthe sojae | FAU 635 | Glycine max/USA | KJ590719 | KJ590762 | KJ610875 |
| Diaporthe taoicola | PSCG 292 | Pyrus pyrifolia/China | MK626871 | MK654800 | MK691232 |
| PSCG 386 | Pyrus pyrifolia/China | MK626868 | MK654797 | MK691222 | |
| PSCG 413 | Pyrus pyrifolia/China | MK626890 | MK654814 | MK691238 | |
| PSCG 485 | Pyrus pyrifolia/China | MK626869 | MK654812 | MK691238 | |
| MFLUCC 16-0117 | Prunus persica/China | KU557567 | KU557635 | KU557591 | |
| MFLUCC 16-0118 | Prunus persica/China | KU557568 | KU557636 | KU557592 | |
| MFLUCC 16-0119 | Prunus persica/China | KU557569 | KU557637 | KU557593 | |
| MFLUCC 16-0120 | Prunus persica/China | KU557570 | KU557638 | KU557594 | |
| YZU 181047 | Citrus sinensis/China | MW160355 | MW160361 | MW160358 | |
| YZU 181223 | Ctrus sinensis/China | MW160356 | MW160362 | MW160359 | |
| Diaporthe yunnanensis | LC 6168 | Coffea sp. | KX986796 | KX999188 | - |
| SAUCC 0254 | Unknown/China | MT376663 | MT376663 | MT376634 | |
| Diaporthella corylina | CBS 121124 | Corylus sp./China | KC343004 | KC343730 | KC343972 |
The present strains are shown in bold.
Figure 1.
Phylogram of Diaporthe strains based on combined gene sequences of ITS, tef1 and tub2. Values at the branch nodes indicated maximum parsimony bootstrap (MP BP ≥ 60%), Bayesian posterior (BI PP ≥ 0.6) and maximum likelihood bootstrap (ML BP ≥ 60%), respectively. The tree is rooted with Diaporthella corylina. Strains in the current study are in bold.
The phylogenetic tree showed that isolates YZU 181047 and YZU 181223 fell into a clade containing reference strains of Diaporthe taoicola supported with PP values of 0.93 and clustered together with D. taoicola PSCG 485 [5] with the BP or PP values of 94/1.0/95 (MP/BI/ML). The result indicated that both isolates were D. taoicola. However, the isolate YZU 181403 clustered with reference strains of D. siamensis with high BP or PP values of 87/1.0/95 (MP/BI/ML). The result showed that it was D. siamensis.
Morphological examination confirmed the phylogenetic results, the isolates YZU 181047 and YZU 181223 were identified as Diaporthe taoicola [5,16], and the isolate YZU 181403 was D. siamensis [22,28] based on the colony and conidia characteristics.
Diaporthe taoicola Dissanayake, X.H. Li & K.D. Hyde., Mycosphere 8: 543. (2017) (Figure 2)
Figure 2.
Diaporthe taoicola (YZU 181047). (a, b). Front and back view, respectively of colonies on PDA (a) and OA (b); (c). Pathogenicity test on Citrus sinensis fruit for 7 d; (d, e). Conidiomata; (f): Section view of conidiomata; (g). Conidiophores; (h): Alpha conidia. Scale bars: d, e, f = 100μm; g, h: 10 μm.
Sexual morph: Not observed. Asexual morph: Conidiomata 160–230 μm in size, pycnidial, subcuticular, scattered to confluent, dark brown to black, uniloculate, broadly spherical to flattened, cream conidial droplets exuding from central ostioles (Figure 2(d–f)). Conidiophores 16–28 × 2–3 μm, hyaline, smooth, densely aggregated, cylindrical, straight, or slightly curved, tapering toward the apex (Figure 2(g)). Alpha conidia 6–9 × 2–3 μm (av. 8 × 2.7 μm) hyaline, smooth, fusiform to ellipsoid, tapering toward both ends, straight (Figure 2(h)).
Colony morphology: Colonies on PDA covering the entire Petri dishes after 7 days, ropey with abundant tufted white aerial mycelium, reverse buff with zonate and irregular lines (Figure 2(a)), 79–81 mm in diam., with aerial mycelium dense in the center and sparse at the marginal area. Colonies on OA flat with white felty aerial mycelium, turning white to dark brown aerial mycelium, conidiomata irregularly distributed on the medium surface after 15-day incubation (Figure 2(b)).
Materials examined. China, Hubei province, Zigui county, on fruit of Citrus sinensis, August, 2018, M. J. Cui (cultures YZU 181047 and YZU 181223).
Notes: The size and shape of Alpha conidia from the present isolates were identical to that firstly reported by Dissanayake et al. (7–9 × 2–3 μm) [16]. However, Beta conidia were not observed during the present study, which was same as the strains found on pear shoot canker by Guo et al. [5].
Diaporthe siamensis Udayanga, X.Z. Liu & K.D. Hyde., Cryptogamie Mycologie, 33(3): 295-309 (2012) (Figure 3)
Figure 3.
Diaporthe siamensis (YZU 181403). (a, b). Front and back view, respectively of colonies on PDA (a) and OA (b); (c). Pathogenicity test on Citrus sinensis fruit for 7 d; (d, e). Conidiomata; (f): Section view of conidiomata; (g). Conidiophores; (h): Alpha conidia. Scale bars: d, e, f = 100μm; g, h: 10 μm.
Sexual morph: Not observed. Asexual morph: Conidiomata 130–240 μm wide, 94–200 μm high, solitary, single conical neck erumpent through leave tissues, 80–160 × 54–85 μm in size (Figure 3(d–f)). Conidiophores 11–23 × 1–2.5 μm, cylindrical, hyaline, straight, or curved, tapering toward the apex (Figure 3(g)). Alpha conidia 6–8 × 3–3.5 μm (av. 7 × 3.2 μm) hyaline, aseptate, ellipsoidal to oval, biguttulate, rounded at both ends (Figure 3(h)).
Colony morphology: Colonies on PDA mycelia growing full of Petri-dishes after 7 d with zones of the dirty white and umber, reverse umber patches (Figure 3(a)). Colonies on OA flat with white felty aerial mycelium, turning white to reddish-brown, with irregular black zones (Figure 3(b)).
Materials examined. China, Hubei province, Zigui county, on fruit of Citrus sinensis, August, 2018, M. J. Cui (culture YZU 181403).
Note: Alpha conidia of the present isolate were identical to that firstly reported by Udayanga et al. with (3.5–)4–5(–6) × (2–)2.5(–3) μm in size, collected from diseased leaves of Dasymaschalon sp. (Annonaceae) [28]. Besides, its cultural characteristics on PDA were identical to D. siamensis reported by Tibpromma et al., as an endophytic fungus from a Pandanaceae host (Pandanus sp.) [22]. Unfortunately, Beta conidia and Gamma conidia were not observed in the present study.
In the pathogenicity tests, all isolates caused brown fruit rot (Figures 2(c), 3(c)) on Citrus sinensis, exposing mycelia on surface and severe rotting inside. The initial symptoms appeared as tiny, watery lesions, which gradually expanded eventually led to fruit rot on 7th day. However, the diameters of the lesions varied among different species; D. siamensis caused larger lesions (33–37 mm, av. 34 mm) than D. taoicola (26–30 mm, av. 28 mm) during the tests. In parallel, no lesions developed on the fruit that were inoculated with PDA disks as control. These results showed that all the present isolates were responsible agents for fruit rot of Citrus sinensis.
Presently, the identification of Diaporthe is mainly based on morphological characters and phylogenetic analysis [3,28]. In recent reported studies, nearly 65 Diaporthe species were associated with Chinese hosts, from which 15 were founded on Citrus spp. (Table. 2). According to Huang et al. [15] and Li et al. [36], phylogeny inferred from combined gene loci of ITS, tef1, and tub2 could be used for further identification of Diaporthe species.
Table 2.
Diaporthe species isolated from various hosts in China.
| Species | Authority | Host | Locality (Province) | Reference |
|---|---|---|---|---|
| Diaporthe acerigena | C.M. Tian & Q. Yang, | Acer tataricum | Shaanxi | Yang et al. [17] |
| D. acuta | Y.S. Guo & G.P. Wang | Pyrus pyrifolia | Hubei | Guo et al. [5] |
| D. acutispora | Y.H. Gao & L. Cai | Coffea sp. | Yunnan | Gao et al. [4] |
| D. alangii | C.M. Tian & Q. Yang | Alangium kurzii | Zhejiang | Yang et al. [17] |
| D. amygdali | Udayanga, Crous & K.D. Hyde | Pyrus pyrifolia | Jiangxi, Yunnan | Bai et al. [30] |
| Camellia sp. | Sichuan | Gao et al. [4] | ||
| D. apiculata | Y.H. Gao & L. Cai | Camellia sp. | Jiangxi, Guangxi | Gao et al. [4] |
| D. aquatica | D.M. Hu, L. Cai & K.D. Hyde | aquatic habitats | Guizhou | Hu et al. [31] |
| D. betulae | C.M. Tian & X.L. Fan, | Betula platyphylla | Sichuan | Du et al. [8] |
| D. betulicola | C.M. Tian & Z. Du, | Betula albo-sinensis | Shaanxi | Du et al. [8] |
| D. betulina | C.M. Tian & Q. Yang | Betula sp. | Heilongjiang | Yang et al. [17] |
| D. biconispora | F. Huang, K.D. Hyde & H.Y. Li | Citrus sinensis | Jiangxi, Guangxi, Fujian | Huang et al. [12] |
| D. biguttulata | F. Huang, K.D. Hyde & H.Y. Li | Citrus limon | Yunnan | Huang et al. [12] |
| Juglans regia | Zhejiang | Yang et al. [17] | ||
| D. biguttusis | Y.H. Gao & L. Cai | Lithocarpus glabra | Zhejiang | Gao et al. [4] |
| D. camptothecicola | C.M. Tian & Qin Yang | Camptotheca acuminata | Jiangsu | Yang et al. [32] |
| D. caryae | C.M. Tian & Q. Yang | Carya illinoensis | Jiangsu | Yang et al. [32] |
| Pyrus pyrifolia | Jiangsu | Guo et al. [5] | ||
| D. cercidis | C.M. Tian & Q. Yang | Carya illinoensis | Jiangsu | Yang et al. [17] |
| Pyrus pyrifolia | Shandong, Zhejiang, et al. | Guo et al. [5] | ||
| D. chensiensis | C.M. Tian & Q. Yang | Abies chensiensis | Shaanxi | Yang et al. [17] |
| D. chongqingensis | Y.S. Guo & G.P. Wang | Pyrus pyrifolia | Chongqing | Guo et al. [5] |
| D. cinnamomi | C.M. Tian & Q. Yang | Cinnamomum sp. | Zhejiang | Yang et al. [17] |
| D. citri | F.A. Wolf | Citrus sp. | Zhejiang, Huangyan, Jiangxi | Huang et al. [15] |
| D. citriasiana | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Shaanxi, Jiangxi, Zhejiang | Huang et al. [15] |
| D. citrichinensis | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Shaanxi, Guangxi, Fujian | Huang et al. et al. [15] |
| D. compacta | Y.H. Gao & L. Cai | Camellia sp. | Jiangxi | Gao et al. [4] |
| D. conica | C.M. Tian & Q. Yang | Alangium chinense | Zhejiang | Yang et al. [17] |
| D. discoidispora | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Jiangxi | Huang et al. [12] |
| D. elaeagni-glabrae | Y.H. Gao & L. Cai | Elaeagnus glabra | Jiangxi | Gao et al. [29] |
| D. ellipicola | Y. H. Gao & L. Cai | Lithocarpus glabra | Zhejiang | Gao et al. [4] |
| D. endophytica | R.R. Gomes, C. Glienke & Crous | Citrus sp. | Fujian | Huang et al. [12] |
| D. eres | Nitschke | Aralia elata | northeastern China | Bai et al. [30] |
| Citrus sp. | Guangxi, Jiangxi, Zhejiang | Huang et al. [12] | ||
| Vitis vinifera | Beijing, Zhejiang | Dissanayake et al. [16] | ||
| Juglans regia | Zhejiang | Yang et al. [17] | ||
| Camellia sp. | Sichuan | Gao et al. [29] | ||
| D. fraxinicola | C.M. Tian & Q. Yang, | Fraxinus chinensis | Shaanxi | Yang et al. [17] |
| D. fulvicolor | Y.S. Guo & G.P. Wang | Pyrus pyrifolia | Hubei | Guo et al. [5] |
| D. fusicola | Y.H. Gao & L. Cai | Pyrus pyrifolia | Jiangxi, Fujian, Zhejiang | Guo et al. [5] |
| D. ganjae | R.R. Gomes | Pyrus pyrifolia | Guizhou | Guo et al. [5] |
| D. hongkongensis | R.R. Gomes, C. Glienke & Crous | Citrus sp. | Zhejiang, Guangxi | Huang et al. [12] |
| Vitis vinifera | Beijing | Dissanayake et al. [16] | ||
| Camellia sp. | Guangxi | Gao et al. [4] | ||
| D. incompleta | Y.H. Gao & L. Cai | Camellia sinensis | Yunnan | Gao et al. [4] |
| D. juglandicola | C.M. Tian & Q. Yang | Juglans mandshurica | Beijing | Yang et al. [32] |
| D. kadsurae | C.M. Tian & Q. Yang | Kadsura longipedunculata | Jiangxi | Yang et al. [17] |
| D. lithocarpus | Y.H. Gao, W. Sun & L. Cai | Lithocarpus sp. | Zhejiang | Gao et al. [4] |
| D. longicolla | (Hobbs) J.M. Santos, Vrandečić & A.J.L. Phillips | Pyrus pyrifolia | Jiangxi, Fujian, Hubei | Bai et al. [30] |
| D. mahothocarpus | Y.H. Gao, W. Sun & L. Cai | Lithocarpus sp. | Zhejiang | Gao et al. [4] |
| D. multigutullata | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Fujian | Huang et al. [12] |
| D. neotheicola | A.J.L. Phillips & J.M. Santos | Pyrus bretschneideri | Yunnan, Jiangxi, Fujian | Bai et al. [30] |
| D. oraccinii | Y.H. Gao & L. Cai | Camellia sp. | Jiangxi | Gao et al. [29] |
| D. ovalispora | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Yunnan | Huang et al. [12] |
| D. ovoicicola | Y. H. Gao & L. Cai | Citrus sp. | Zhejiang | Gao et al. [4] |
| D. padina | C.M. Tian & Q. Yang | Padus racemosa | Jiangxi | Yang et al. [17] |
| D. parvae | Y.S. Guo & G.P. Wang | Pyrus pyrifolia | Yunnan | Guo et al. [5] |
| D. pentriteum | Y.H. Gao & L. Cai | Camellia sp. | Jiangxi | Gao et al. [29] |
| D. pescicola | Dissanayake et al. | Pyrus bretschneideri | Yunnan | Guo et al. [5] |
| D. phaseolorum | (Cooke & Ellis) Sacc. | Vitis vinifera | Beijing | Huang et al. [12] |
| D. phragmitis | Crous | Phragmitis australis | Beijing | Crous et al. [33] |
| D. podocarpi-macrophylli | Y.H. Gao & L. Cai | Podocarpus macrophyllus | Zhejiang | Gao et al. [29] |
| D. rostrata | C.M. Tian, X.L. Fan & K.D. Hyde | Juglans mandshurica | Gansu | Fan et al. [34] |
| D. sambucusii | C.M. Tian & Q. Yang | Sambucus williamsii | Heilongjiang | Yang et al. [35] |
| D. schisandrae | C.M. Tian & Q. Yang | Schisandra chinensis | Heilongjiang | Yang et al. [35] |
| D. sojae | Lehman | Vitis vinifera | Beijing | Huang et al. [12] |
| Citrus sp. | Shaanxi | Huang et al. [12] | ||
| Pyrus pyrifolia | Guizhou, Jiangsu | Guo et al. [5] | ||
| D. spinosa | Y.S. Guo & G.P. Wang | Pyrus pyrifolia | Zhejiang, Jiangsu, Guizhou | Guo et al. [5] |
| D. subclavata | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Fujian, Guangdong | Huang et al. [12] |
| D. taoicola | Dissanayake, X.H. Li & K. D | Prunus persica | Hubei | Dissanayake et al. [16] |
| Pyrus pyrifolia | Zhejiang, Jiangsu, Guizhou | Guo et al. [5] | ||
| D. ternstroemia | Y.H. Gao, W. Sun & L. Cai | Ternstroemia sp. | Zhejiang | Gao et al. [4] |
| D. unshiuensis | F. Huang, K.D. Hyde & H.Y. Li | Citrus sp. | Guangxi | Huang et al. [12] |
| Carya illinoensis | Jiangsu | Yang et al. [17] | ||
| D. velutina | Y.H. Gao & L. Cai | Pyrus pyrifolia | Fujian, Guizhou | Guo et al. [5] |
| D. xishuangbanica | Y.H. Gao & L. Cai | Camellia sinensis | Yunnan | Gao et al. [29] |
| D. yunnanensis | Y.H. Gao & L. Cai | Coffea sp. | Yunnan | Gao et al. [29] |
| D. zaobaisu | Y.S. Guo & G.P. Wang | Pyrus bretschneideri | Yunnan | Guo et al. [5] |
Diaporthe taoicola was firstly isolated from diseased shoots of Prunus persica in Hubei province, China, 2017, proved being able to cause necrotic lesions on detached peach shoots [16]. From then on, it had been only reported on Pyrus pyrifolia causing shoot canker symptoms in China with high phylogenetic diversity [5]. Guo et al. [5] inoculated D. taoicola from pear shoots on wounded twigs of different fruit crops to evaluate its host range, which could induce symptoms on citrus, apple, peach, and kiwifruit. It is worth noting that D. taoicola might pose threats to fruit trees in China. The present study firstly confirmed that it also pathogenic to Citrus sinensis fruit.
Diaporthe siamensis had been reported on diseased leaves of Dasymaschalon sp. in the family of Annonaceae in Thailand [28], but without pathogenicity test on the host plant. Then, it was also found as an endophytic fungus from Pandanus sp. (Pandanaceae) in Thailand [22], also probably as endophyte on Garcinia parvifolia from Malaysia [28]. Regretfully, the pathogenicity evaluation of the species remained a lack in previous studies. Except a detailed description of D. siamensis given in this study, the pathogenicity tests revealed that it could induce fruit rot on Citrus sinensis, stronger than D. taoicola. To the best of our knowledge, this is the first report of Diaporthe taoicola and D. siamensis from Citrus sinensis in China, which could induce fruit rot on the host.
Funding Statement
The research was supported by Guangxi Key Laboratory of Biology for Crop Disease and Insect Pests [2019-KF-01].
Compliance with ethical standards
The research does not contain any studies with Human Participants and/or Animals, and the authors declare that they have no conflict of interest.
Disclosure statement
No potential conflict of interest was reported by the author(s).
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