Diaporthe eres is considered one of the most important causal agents of many plant diseases, with a broad host range worldwide. In this study, multiple sequences of ribosomal internal transcribed spacer region (ITS), translation elongation factor 1-α gene (EF1-α), beta-tubulin gene (TUB2), calmodulin gene (CAL), and histone-3 gene (HIS) were used for multi-locus phylogenetic analysis. For phylogenetic analysis, maximum likelihood (ML), maximum parsimony (MP), and Bayesian inferred (BI) approaches were performed to investigate relationships of D. eres with closely related species. The results strongly support that the D. eres species falls into a monophyletic lineage, with the characteristics of a species complex. Phylogenetic informativeness (PI) analysis showed that clear boundaries could be proposed by using EF1-α, whereas ITS showed an ineffective reconstruction and, thus, was unsuitable for speciating boundaries for Diaporthe species. A combined dataset of EF1-α, CAL, TUB2, and HIS showed strong resolution for Diaporthe species, providing insights for the D. eres complex. Accordingly, besides D. biguttusis, D. camptothecicola, D. castaneae-mollissimae, D. cotoneastri, D. ellipicola, D. longicicola, D. mahothocarpus, D. momicola, D. nobilis, and Phomopsis fukushii, which have already been previously considered the synonymous species of D. eres, another three species, D. henanensis, D. lonicerae and D. rosicola, were further revealed to be synonyms of D. eres in this study. In order to demonstrate the genetic diversity of D. eres species in China, 138 D. eres isolates were randomly selected from previous studies in 16 provinces. These isolates were obtained from different major plant species from 2006 to 2020. The genetic distance was estimated with phylogenetic analysis and haplotype networks, and it was revealed that two major haplotypes existed in the Chinese populations of D. eres. The haplotype networks were widely dispersed and not uniquely correlated to specific populations. Overall, our analyses evaluated the phylogenetic identification for D. eres species and demonstrated the population diversity of D. eres in China.

Melanose disease caused by Diaporthe citri is considered as one of the most important and destructive diseases of citrus worldwide. In this study, isolates from melanose samples were obtained and analyzed. Firstly, the internal transcribed spacer (ITS) sequences were used to measure Diaporthe-like boundary species. Then, a subset of thirty-eight representatives were selected to perform the phylogenetic analysis with combined sequences of ITS, beta-tubulin gene (TUB), translation elongation factor 1-α gene (TEF), calmodulin gene (CAL), and histone-3 gene (HIS). As a result, these representative isolates were identified belonging to D. citri, D. citriasiana, D. discoidispora, D. eres, D. sojae, and D. unshiuensis. Among these species, the D. citri was the predominant species that could be isolated at highest rate from different melanose diseased tissues. The morphological characteristics of representative isolates of D. citri were investigated on different media. Finally, a molecular tool based on the novel species-specific primer pair TUBDcitri-F1/TUBD-R1, which was designed from TUB gene, was developed to detect D. citri efficiently. A polymerase chain reaction (PCR) amplicon of 217 bp could be specifically amplified with the developed molecular tool. The sensitivity of the novel species-specific detection was upon to 10 pg of D. citri genomic DNA in a reaction. Therefore, the D. citri could be unequivocally identified from closely related Diaporthe species by using this simple PCR approach.

Peach scab is a fungal disease caused by Venturia carpophila, which can significantly reduce peach yield and quality. Fungicide application is the main control measure for peach scab worldwide. For better understanding of the fungicide resistance status and making suitable management strategies, the sensitivity of 135 single-spore V. carpophila isolates to commonly used fungicides carbendazim, iprodione, propiconazole, azoxystrobin and boscalid were determined using microtiter plate test method. Results showed that the mean EC50 values of tested isolates to iprodione, propiconazole, azoxystrobin and boscalid were 16.287 µg/ml, 0.165 µg/ml, 0.570 µg/ml and 0.136 µg/ml, respectively. The EC50 values of V. carpophila isolates to four fungicides displayed unimodal frequency distributions, indicating no resistance occurred to these fungicides. On the other hand, bimodal frequency distribution was observed for carbendazim, indicating that the V. carpophila developed resistance to carbendazim. Actually, the resistance was widely detected from all of the 14 provinces. Molecular analysis showed that the point mutation E198K of the TUB2 gene determined the high resistance, while the E198G conferred the moderate resistance. Both moderate and high resistances were stable, and the resistant isolates did not show significant fitness penalties. On the contrary, some resistant isolates showed better competitiveness under certain stresses. This is the first report to detect the sensitivity of V. carpophila to fungicides which enables future monitoring of fungicide resistance and provides the basic information for making suitable peach scab management strategies.

Colletotrichum is regarded as one of the 10 most important genera of plant pathogens in the world. It causes diseases in a wide range of economically important plants, including peaches. China is the largest producer of peaches in the world but little is known about the Colletotrichum spp. affecting the crop. In 2017 and 2018, a total of 286 Colletotrichum isolates were isolated from symptomatic fruit and leaves in 11 peach production provinces of China. Based on multilocus phylogenetic analyses (ITS, ACT, CAL, CHS-1, GAPDH, TUB2, and HIS3) and morphological characterization, the isolates were identified to be C. nymphaeae, C. fioriniae, and C. godetiae of the C. acutatum species complex, C. fructicola and C. siamense of the C. gloeosporioides species complex, C. karsti of the C. boninense species complex, and one newly identified species, C. folicola sp. nov. This study is the first report of C. karsti and C. godetiae in peaches, and the first report of C. nymphaeae, C. fioriniae, C. fructicola, and C. siamense in peaches in China. C. nymphaeae is the most prevalent species of Colletotrichum in peaches in China, which may be the result of fungicide selection. Pathogenicity tests revealed that all species found in this study were pathogenic on both the leaves and fruit of peaches, except for C. folicola, which only infected the leaves. The present study substantially improves our understanding of the causal agents of anthracnose on peaches in China.

BACKGROUND Peach scab, caused by Venturia carpophila, can significantly reduce both the yield and quality of peach fruit. It is difficult to distinguish peach scab from other peach diseases such as black spot and bacterial spot. An efficient assay is needed to identify V. carpophila in order to develop scientific management strategies. RESULTS A set of loop‐mediated isothermal amplification (LAMP) primers was designed based on the internal transcribed spacer (rDNA‐ITS) sequence to detect V. carpophila. Compared with the conventional polymerase chain reaction (PCR) method, the LAMP method not only exhibited higher sensitivity and specificity in the detection of V. carpophila, but also required simpler equipment and less operational time. The minimum detectable concentration of V. carpophila genomic DNA with the LAMP method was 56.6 fg μl–1, which was 100 times lower than with the conventional PCR method. When eight fungal species including V. carpophila (23 isolates from 14 provinces) and one bacterial species were used with LAMP detection, only V. carpophila showed a color change, from brown to yellowish green, and ladder‐like bands in electrophoresis, indicating successful amplification. Moreover, when crude DNAs of peach fruit samples were used in LAMP detection, amplification was observed only from diseased fruits, and not from healthy fruits and the negative control. CONCLUSION The LAMP assay shows simplicity, rapidity, high sensitivity and specificity, and will be useful in distinguishing scab caused by Venturia carpophila from other diseases with similar symptoms. © 2020 Society of Chemical Industry