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Anthracnose, caused by Colletotrichum species, induces significant economic damages to crop plants annually, especially for Camellia oleifera . During infection, the counter-defence mechanisms of plant pathogens against ROS-mediated resistance, however, remain poorly understood. By employing Weighted Gene Co-expression Network Analysis (WGCNA), we identified ACTIVATOR PROTEIN-1 (AP-1), a bZIP transcription factor, as significant to infection. And deletion of CfAP1 inhibited aerial hyphae formation and growth under oxidative stress. Furthermore, RNA-seq analysis post H 2 O 2 treatment revealed 33 significantly down-regulated genes in the AP-1 deficient strain, including A12032, a dual specificity phosphatase (DSP) homologous to MSG5 from Saccharomyces cerevisiae . This Δ Cfmsg5 strain showed enhanced oxidative tolerance, reduced ROS scavenging, and negative regulation of the CWI MAPK cascade under oxygen stress, suggesting its involvement in oxidative signal transduction. Importantly, we provide evidence that CfMsg5 regulates growth, endoplasmic reticulum stress, and several unfolded protein response genes upregulated in Δ Cfmsg5 . Collectively, this study identified core components during C. fructicola infection and highlights a potential regulatory module involving CfAp1 and CfMsg5 in response to host ROS bursts. It provides new insights into fungal infection mechanisms and potential targets like CfAP1 and CfMSG5 for managing anthracnose diseases.

A novel actinobacterium strain, HUAS 14-6 T , was isolated from the healthy leaves of Camellia oleifera collected from Changde City, Hunan Province, China. Strain HUAS 14-6 T produced tight spiral spore chains consisting of oval or spherical spores with a smooth surface. 16S rRNA gene sequence analysis revealed that strain HUAS 14-6 T belonged to the genus Streptomyces and shared highest similarity to Streptomyces bungoensis DSM 41781 T (99.72%). Phylogenetic analysis based on 16S rRNA gene sequences indicated strain HUAS 14-6 T was in a clade with S. bungoensis DSM 41781 T . However, the ANIm and dDDH between strain HUAS 14-6 T and S. bungoensis DSM 41781 T were 88.16% and 31.2%, respectively, far less than the species-level thresholds. Phylogenetic trees based on the five housekeeping genes (atpD, gyrB, recA, rpoB and trpB) showed that strain HUAS 14-6 T formed a separate branch, indicating that this strain could belong to a potential new species. Pairwise MLSA distances between strain HUAS 14-6 T and all type strains exhibiting 16S rRNA gene sequence similarities of ≥98.7% to it were much higher than the maximum range of 0.014 recommended for delineating a new Streptomyces species. Based on polyphasic taxonomic study, HUAS 14-6 T represents a novel species within the genus Streptomyces for which the name Streptomyces albidocamelliae sp. nov. is proposed. The type strain is HUAS 14-6 T (=MCCC 1K08365 T =JCM 35920 T )., Competing Interests: Compliance with ethical standards. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to the Japan Antibiotics Research Association.)

Camellia oleifera is an important woody oil crop in China, and its seed oil has a high economic value. Anthracnose, one of the main diseases in C. oleifera , occurs in a wide range in the production areas, limiting the growth and development of plants and causing serious losses of oil production. With the rapid development of the C. oleifera industry in recent years, great progress has been achieved in the research on anthracnose in C. oleifera . This paper summarized the resistance mechanisms, the mining of resistance genes, and the evaluation of resistant germplasm resources, aiming to provide a theoretical basis for the prevention and control of anthracnose and the breeding of C. oleifera germplasm with resistance to anthracnose.

Bacillus tequilensis DZY 6715 was isolated from healthy leaves in Camellia oleifera, and the strain DZY 6715 significantly inhibited anthracnose disease resulting from Colletotrichum fructicola in C. oleifera, besides, its associated mechanism of disease resistance was explored. B. tequilensis DZY 6715 treatment controlled mycelial growth of C. fructicola in C. oleifera, and significantly decreased C. oleifera anthracnose incidence and disease index compared with the control group. B. tequilensis DZY 6715 has strong biofilm forming ability, and also secretes extracellular β-1, 3-glucanase and chitinase, which could cause cell membranes damage and increased cellular compound leakage. C.oleifera treated with DZY 6715 also effectively enhanced enzyme activities and stimulated the synthesis the substances related to phenylpropane metabolism and reactive oxygen metabolism. Moreover, transcript profiling analysis revealed more differentially expressed genes related to phenylpropanoid pathway metabolism and antioxidant system inducing by DZY 6715 compared with the control in C. oleifera. Thus, it can be concluded that B. tequilensis DZY 6715 is a suitable bio-control agent to control anthracnose disease in C. oleifera., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Anthracnose, caused by Colletotrichum spp., is a common disease of Camellia oleifera . In this study, a Bacillus amyloliquefaciens strain, GZY63, was isolated from fruit of the anthracnose-resistant cultivar of Ca. oleifera "Ganzhouyou7." Plate confrontation assays and field experiments demonstrated the strong inhibitory effect of GZY63 on anthracnose, and this strain exhibited broad-spectrum resistance to nine pathogenic Colletotrichum spp. This strain shows potential as a fungicide alternative, but genetic information on this strain is critical for its optimal use. Combining Illumina and Nanopore sequencing, we assembled a high-quality circular genome of GZY63 that contained no plasmids. The GZY63 complete genome was approximately 3.93 Mb and had an average guanine-cytosine content of 46.5%. The genome comprised 4,024 predicted coding sequences and 12 types of gene clusters involved in secondary metabolite production. This genome information provides insights into the mechanism underlying the antagonistic impact of the GZY63 strain on anthracnose and its symbiotic relationship with Ca. oleifera ., Competing Interests: The author(s) declare no conflict of interest.

The epiphytic and endophytic bacteria play an important role in the healthy growth of plants. Both plant species and growth environmental influence the bacterial population diversity, yet it is inconclusive whether it is the former or the latter that has a greater impact. To explore the communities of the epiphytic and endophytic microbes in Camellia oleifera, this study assessed three representative C. oleifera cultivars from three areas in Hunan, China by Illumina high-throughput sequencing. The results showed that the diversity and species richness of endophytic microbial community in leaves were significantly higher than those of microbial community in the epiphytic. The diversity and species richness of epiphytic and endophytic microbes are complex when the same cultivar was grown in different areas. The C. oleifera cultivars grown in Youxian had the highest diversity of epiphytic microbial community, but the lowest abundance, while the cultivars grown in Changsha had the highest diversity and species richness of endophytic microbes in leaves. It was concluded that the dominant phylum mainly included Proteobacteria, Actinobacteriota and Firmicutes through the analysis of the epiphytic and endophytic microbial communities of C. oleifera. The species and relative abundances of epiphytic and endophytic microbial community were extremely different at the genus level. The analysis of NMDS map and PERMANOVA shows that the species richness and diversity of microbial communities in epiphytes are greatly influenced by region. However, the community structure of endophytic microorganisms in leaves is influenced by region and cultivated varieties, but the influence of cultivars is more significant. Molecular ecological network analysis showed that the symbiotic interaction of epiphytic microbial community was more complex., (© 2024. The Author(s).)

A novel actinobacterium strain, designated HUAS 2-6  T , was isolated from the rhizosphere soil of Camellia oleifera Abel collected from Taoyuan County, Northwestern Hunan Province, South China. This strain was subjected to a polyphasic taxonomic study. Strain HUAS 2-6  T is characterized by morphology typical of members of the genus Streptomyces, with deep purplish vinaceous aerial mycelia and deep dull lavender substrate mycelia. Strain HUAS 2-6  T , based on the full-length 16S rRNA gene sequence analysis, exhibited the highest similarities to S. puniciscabiei S77 T (99.31%), S. filipinensis NBRC 12860  T (99.10%), S. yaanensis CGMCC 4.7035  T (99.09%), S. fodineus TW1S1 T (99.08%), S. broussonetiae CICC 24819  T (98.76%), S. achromogenes JCM 4121  T (98.69%), S. barringtoniae JA03 T (98.69%), and less than 98.70% with other validly species. In phylogenomic tree, strain HUAS 2-6  T was clustered together with S. broussonetiae CICC 24819  T , suggesting that they were closely related to each other. However, average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) between them were much less than the species cutoff values (ANI 96.7% and dDDH 70%). Moreover, in phenotypic and chemotaxonomic characteristics, strain HUAS 2-6  T is distinct from S. broussonetiae CICC 24819  T . On the basis of the polyphasic data, strain HUAS 2-6  T is proposed to represent a novel species, Streptomyces camelliae sp. nov. (= MCCC 1K04729 T  = JCM 35918  T )., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Camellia oleifera ( Ca. oleifera ) is a woody tree species cultivated for the production of edible oil from its seed. The growth and yield of tea-oil trees are severely affected by anthracnose (caused by Colletotrichum gloeosporioides ). In this study, the transcriptomic and metabolomic analyses were performed to detect the key transcripts and metabolites associated with differences in the susceptibility between anthracnose-resistant (ChangLin150) and susceptible (ChangLin102) varieties of Ca. oleifera . In total, 5001 differentially expressed genes (DEGs) were obtained, of which 479 DEGs were common between the susceptible and resistant varieties and further analyzed. KEGG enrichment analysis showed that these DEGs were significantly enriched in tyrosine metabolism, phenylpropanoid biosynthesis, flavonoid biosynthesis and isoquinoline alkaloid biosynthesis pathways. Furthermore, 68 differentially accumulated metabolites (DAMs) were detected, including flavonoids, such as epicatechin, phenethyl caffeate and procyanidin B2. Comparison of the DEGs and DAMs revealed that epicatechin, procyanidin B2 and arachidonic acid (peroxide free) are potentially important. The expression patterns of genes involved in flavonoid biosynthesis were confirmed by qRT-PCR. These results suggested that flavonoid biosynthesis might play an important role in the fight against anthracnose. This study provides valuable molecular information about the response of Ca. oleifera to Co. gloeosporioides infection and will aid the selection of resistant varieties using marker-assisted breeding.

Arbuscular mycorrhizal (AM) fungi play an important role in the acquisition of phosphorus (P) by plants. The external hyphae of AM fungi function as an extension of plant roots and may downregulate related functions in the roots. It is not clear whether the ability of AM fungi to mineralize organic P affects root phosphatase activities. A pot experiment was conducted to investigate the effect of Funneliformis mosseae on soil organic P mineralization under phytate application and to explore root phosphatase activities, P uptake, and growth in Camellia oleifera Abel. The plants and their growth substrates were harvested 4 and 8 months after planting. The results showed that organic P application had no effect on the total dry mass of nonmycorrhizal plants, but differences in dry mass under P application were observed in mycorrhizal plants in both harvests. Inoculation with F .  mosseae increased soil acid phosphatase, phytase, and alkaline phosphatase activities and reduced the soil organic P content. Mycorrhizal plants had higher root activity, shoot and root P contents and root acid phosphatase and phytase activities than nonmycorrhizal plants irrespective of organic P application. In conclusion, AM fungi enhanced the mineralization of soil organic P and positively affect root phosphatase activities.

Food contamination caused by microorganisms has become a threat to consumers' health. Exploring antagonistic endophytes from plants of food raw-material and applying bioactive metabolites to inhibit the contamination has been an alternative and safer solution. In this study, we isolated and screened potential antagonistic endophytes from fresh Camellia assamica leaves, which were widely used in tea beverage production. We focused on a strain that showed visible inhibitory activity to Gram-positive bacteria, Gram-negative bacteria, and fungi. It was identified as a member of Bacillus velezensis and named FZ06. The results of genome analysis showed the strain FZ06 had 167 single-copy specific genes, much higher than those of most related strains. Also, 11 potential gene clusters of antimicrobial metabolites were found. Three groups of lipopeptides (surfactin, iturin, and fengycin) were identified by UPLC-MS/MS in purified antimicrobial methanol fraction of strain FZ06. The results of minimum inhibitory concentration (MIC) test proved the lipopeptide extract showed significant inhibitory effect on food spoilage bacteria (MIC 512-2048 μg/mL) and toxigenic fungi (MIC 128-256 μg/mL). In conclusion, this study suggests that the endophytic B. velezensis FZ06 and its lipopeptide extract hold great potential applications in the inhibition of food spoilage bacteria and toxic fungi in food industry., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Agricultural production is dependent on inputs of nitrogen (N) whose cycle relies on soil and crop microbiomes. Crop diversification has increased productivity; however, its impact on the expression of microbial genes involved in N-cycling pathways remains unknown. Here, we assessed N-cycling gene expression patterns in the root and rhizosphere microbiomes of five oilseed crops as influenced by three 2-year crop rotations. The first phase consisted of fallow, lentil or wheat, and the second phase consisted of one of five oilseed crops. Expression of bacterial amoA, nirK and nirS genes showed that the microbiome of Ethiopian mustard had the lowest and that of camelina the highest potential for N loss. A preceding rotation phase of lentil significantly increased the expression of nifH gene by 23% compared with wheat and improved nxrA gene expression by 51% with chemical fallow in the following oilseed crops respectively. Lentil substantially increased biological N 2 fixation and reduced denitrification in the following oilseed crops. Our results also revealed that most N-cycling gene transcripts are more abundant in the microbiomes associated with roots than with the rhizosphere. The outcome of our investigation brings a new level of understanding on how crop diversification and rotation sequences are related to N-cycling in annual cropping systems., (© 2020 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The family Sclerotiniaceae includes important phytopathogens, such as Botrytis cinerea and Sclerotinia sclerotiorum , that activate plant immune responses to facilitate infection propagation. The mechanisms of plant resistance to these necrotrophic pathogens are still poorly understood. To discover mechanisms of resistance, we used the Ciborinia camelliae (Sclerotiniaceae)- Camellia spp. pathosystem. This fungus induces rapid infection of the blooms of susceptible cultivar Nicky Crisp ( Camellia japonica × Camellia pitardii var. pitardii ), while Camellia lutchuensis is highly resistant. Genome-wide analysis of gene expression in resistant plants revealed fast modulation of host transcriptional activity 6 h after ascospore inoculation. Ascospores induced the same defense pathways in the susceptible Camellia cultivar but much delayed and coinciding with disease development. We next tested the hypothesis that differences in defense timing influences disease outcome. We induced early defense in the susceptible cultivar using methyl jasmonate and this strongly reduced disease development. Conversely, delaying the response in the resistant species, by infecting it with actively growing fungal mycelium, increased susceptibility. The same plant defense pathways, therefore, contribute to both resistance and susceptibility, suggesting that defense timing is a critical factor in plant health, and resistance against necrotrophic pathogens may occur during the initial biotrophy-like stages.

Endophytic bacteria, which are common in plant tissues, may help to control plant pathogens and enhance plant growth. Camellia oleifera, an oil-producing plant, is widely grown in warm, subtropical, hilly regions in China. However, C. oleifera is strongly negatively affected by C. oleifera anthracnose, which is caused by Colletetrichum fructicola. To find a suitable biocontrol agent for C. oleifera anthracnose, 41 endophytes were isolated from the stems, leaves, and roots of C. oleifera. Bacterial cultures were identified based on analyses of 16S rDNA sequences; most strains belonged to the genus Bacillus. The antagonistic effects of these strains on C. fructicola were tested in vitro. In total, 16 strains inhibited C. fructicola growth, with B. subtilis strain 1-L-29 being the most efficient. Strain 1-L-29 demonstrated antagonistic activity against C. siamense, C. asianum, Fusarium proliferatum, Agaricodochium camellia, and Pseudomonas syringae. In addition, this strain produced indole acetic acid, solubilized phosphate, grew on N-free media, and produced siderophores. To facilitate further microecological studies of this strain, a rifampicin-resistant, green fluorescent protein (GFP)-labeled strain, 1-L-29gfpr, was created using protoplast transformation. This plasmid had good segregational stability. Strain 1-L-29gfpr was re-introduced into C. oleifera and successfully colonized root, stem, and leaf tissues. This strain remained at a stable concentration in the root more than 20 d after inoculation. Fluorescence microscopic analysis showed that strain 1-L-29gfpr thoroughly colonized the root surfaces of C. fructicola as well as the root vascular tissues of Arabidopsis thaliana., Competing Interests: The authors have declared that no competing interests exist.

Background: Tea-oil tree (Camellia oleifera) is a unique edible-oil tree in China, and anthracnose occurs in wherever it is cultivated, causing great economic losses each year. We have previously identified the Ascomycete fungus Colletotrichum fructicola as the major pathogen of anthracnose in Ca. oleifera. The purpose of this study was to characterize the biological function of Snf1 protein, a key component of the AMPK (AMP-activated protein kinase) pathway, for the molecular pathogenic-mechanisms of C. fructicola., Results: We characterized CfSnf1 as the homolog of Saccharomyces cerevisiae Snf1. Targeted CfSNF1 gene deletion revealed that CfSnf1 is involved in the utilization of specific carbon sources, conidiation, and stress responses. We further found that the ΔCfSnf1 mutant was not pathogenic to Ca. oleifera, resulting from its defect in appressorium formation. In addition, we provided evidence showing crosstalk between the AMPK and the cAMP/PKA pathways for the first time in filamentous fungi., Conclusion: This study indicate that CfSnf1 is a critical factor in the development and pathogenicity of C. fructicola and, therefore, a potential fungicide target for anthracnose control.

Camellia taliensis (W. W. Smith) Melchior is a wild tea plant endemic from the west and southwest of Yunnan province of China to the north of Myanmar and is used commonly to produce tea by the local people of its growing areas. Its chemical constituents are closely related to those of C. sinensis var. assamica, a widely cultivated tea plant. In this study, the α diversity and phylogeny of endophytic fungi in the branches of C. taliensis were explored for the first time. A total of 160 fungal strains were obtained and grouped into 42 species from 29 genera, which were identified based on rDNA internal transcribed spacer sequence analysis. Diversity analysis showed that the endophytic fungal community of the branches of C. taliensis had high species richness S (42), Margalef index D' (8.0785), Shannon-Wiener index H' (2.8494), Simpson diversity index D S (0.8891), PIE index (0.8947) and evenness Pielou index J (0.7623) but a low dominant index λ (0.1109). By contrast, that in the branches of C. taliensis had abundant species and high species evenness. Diaporthe tectonigena, Acrocalymma sp. and Colletotrichum magnisporum were the dominant endophytic fungi. The phylogenetic tree was established by maximum parsimony analysis, and the 11 orders observed for endophytic fungi belonging to Ascomycota and Basidiomycota were grouped into 4 classes.

Camellia oleifera is an important Chinese commercial crop. Camellia oleifera can display abnormal leaves due to infection by the parasitic fungus Exobasidium gracile . Exobasidium gracile was isolated from infected leaves and used in fermentation, and exopolysaccharides EP0-1 and EP0.5-1 were purified from the fermentation broth. EP0-1 was an alkaline polysaccharide consisting mainly of the linkages α-d-Manp(1→, →2)-α-d-Manp(1→ and →6)-α-d-Manp(1→, →3)-α-d-Glcp(1→ and→4)-α-d-Glcp(1→, terminal β-d-Galf, (1→5)-β-d-Galf, and terminal β-D-GlcN(1→. EP0.5-1 was an acidic galactofuranose-containing polysaccharide. It contained the linkages of α-d-Manp(1→, →2)-α-d-Manp(1→, →6)-α-d-Manp(1→,→2, 6)-α-d-Manp(1→, →4)-α-d-Glcp(1→, and →4)-α-d-GlcUA(1→. Galactofuranose linkages were composed of terminal β-d-Galf, (1→6)-β-d-Galf and (1→2)-β-d-Galf. Exobasidium gracile exopolysaccharides displayed significant immunoregulatory activity by activating macrophages. This research indicates that infected leaves from Camellia oleifera including the exopolysaccharides produced by the parasitic fungus Exobasidium gracile by are worth further investigation as a functional product.

Bacillus megaterium strain JX285, isolated from rhizosphere red soil sample, can solubilize inorganic phosphorus, which increases the amount of available phosphorus and promotes plant growth. To investigate the mechanisms underlying phosphate solubilization, we sequenced the entire genome of B. megaterium strain JX285 (CGMCC 1.1621), which comprises a circular chromosome of 5,066,463 bp and seven plasmids of 167,030, 128,297, 60,905, 134,795, 9,598, 37,455, and 6332 bp, respectively. The whole genome sequence includes 5948 protein-coding genes, 124 tRNAs, and 29 rRNAs, and has been deposited at Genbank/EMBL/DDBJ with accession numbers CP018874-CP018881. We detected genes associated with organic acid production, which may be vital for phosphate conversion. Furthermore, phosphatase-encoding genes were also detected. The information embedded in the genome will assist in studying the mechanisms of phosphate solubilization. In conclusion, analysis of the JX285 genome will further our knowledge regarding this strain and may contribute to its biotechnological application., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The saponins, as components of tea seed meal, are undesirable hemolytic components and should be degraded for reducing their hemolytic activity in order to be used in animal feed. In this study, β-glucuronidase was verified to be a potent hydrolase of tea seed saponins to reduce their hemolytic activity and a β-glucuronidase-producing Lactobacillus crustorum strain was screened from raw bovine milk. Next, solid-state fermentation with the isolated L. crustorum and a Bacillus subtilis natto strain, which can produce cellulase and hence improve the fermentation performance of tea seed meal, was carried out for detoxification of tea seed meal. The 50% hemolytic dosage (HD50) value of tea seed saponins was increased from 6.69 to 27.43 μg mL-1. The results of LC-MS analysis showed that the percentage of saponin aglycones increased from 30.95 to 84.25% after the fermentation. According to the roles of sugar moieties in hemolytic activity, and the enzymatic hydrolysis characteristics of β-glucuronidase, the degradation of tea seed saponins from glucosides to aglycones may contribute to the reduction of hemolytic activity. Therefore, tea seed meal may be used as animal feed after fermentation with the tested saponin-degrading microbial strains.

A total of 124 Pestalotiopsis-like isolates associated with symptomatic and asymptomatic tissues of Camellia sinensis and other Camellia spp. from eight provinces in China were investigated. Based on single- and multi-locus (ITS, TEF, TUB2) phylogenies, as well as morphological characters, host associations and geographical distributions, they were classified into at least 19 species in three genera, i.e. Neopestalotiopsis, Pestalotiopsis and Pseudopestalotiopsis. Eight novel species in Pestalotiopsis and three novel species in Pseudopestalotiopsis were described. Our data suggested that the currently widely used loci in Pestalotiopsis-like genera do not consistently provide stable and sufficient resolution tree topologies, especially for Neopestalotiopsis. Moreover, the number, branch pattern and length of the conidial basal appendages were revealed to be phylogenetically informative characters in Pestalotiopsis.

The use of oligonucleotide-coupled fluorescent microspheres is a rapid, sequencing-independent, and reliable way to diagnose bacterial diseases. Previously described applications of oligonucleotide-coupled fluorescent microspheres for the detection and identification of bacteria in human clinical samples have been successfully adapted to detect and differentiate "Ca. Phytoplasma" species using as a target the chaperonin 60-encoding gene. In this chapter, we describe in detail the design and validation of oligonucleotide capture probes, and their application in the assay aiming to differentiate phytoplasma strains infecting Brassica napus and Camelina sativa plants grown in the same geographic location at the same time.

A novel bacterium, designated strain CF1T, was isolated from a soil sample of a tea plantation and its taxonomic position was determined using a polyphasic approach. Strain CF1T was a Gram-stain-negative, facultatively anaerobic, non-sporulating, non-motile and rod-shaped bacterium. Optimum growth occurred at 25 °C and pH 6.0. Comparative analysis of the 16S rRNA gene sequence showed that the isolate belongs to the genus Paraburkholderia, showing highest levels of similarity with respect to Paraburkholderia sediminicola LMG 24238T (98.44 %). Additionally, strain CF1T, P. sediminicola LMG 24238T and Paraburkholderia aspalathi LMG 27731 formed a distinct group in the phylogenetic tree based on 16S rRNA gene sequences. The predominant ubiquinone was Q-8, and the polar lipid profile consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one unidentified aminophospholipid, two unidentified aminolipids and two unidentified polar lipids. The DNA G+C content was 60.2 mol%, and the major fatty acids were C16 : 0, summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The DNA-DNA relatedness values between strain CF1T and its close relatives including P. sediminicola LMG 24238T and P. aspalathi LMG 27731 49.3±0.4 % and 38.3±0.5 %, respectively. On the basis of phylogenetic analysis, phenotypic and genotypic data, it is concluded that the isolate represents a novel species of the genus Paraburkholderia, for which the name Paraburkholderia caffeinilytica sp. nov. is proposed. The type strain is CF1T (=LMG 28690T=CGMCC 1.15103T).

The filamentous fungus Colletotrichum fructicola is found in all five continents and is capable of causing severe diseases in a number of economically important plants such as avocado, fig, cocoa, pear, and tea-oil trees. However, almost nothing is known about its patterns of genetic variation and epidemiology on any of its host plant species. Here we analyzed 167 isolates of C. fructicola obtained from the leaves of tea-oil tree Camellia oleifera at 15 plantations in seven Chinese provinces. Multilocus sequence typing was conducted for all isolates based on DNA sequences at fragments of four genes: the internal transcribed spacers of the nuclear ribosomal RNA gene cluster (539 bp), calmodulin (633 bp), glutamine synthetase (711 bp), and glyceraldehyde-3-phosphate dehydrogenase (190 bp), yielding 3.52%, 0.63%, 8.44%, and 7.89% of single nucleotide polymorphic sites and resulting in 15, 5, 12 and 11 alleles respectively at the four gene fragments in the total sample. The combined allelic information from all four loci identified 53 multilocus genotypes with the most frequent represented by 21 isolates distributed in eight tea-oil plantations in three provinces, consistent with long-distance clonal dispersal. However, despite evidence for clonal dispersal, statistically significant genetic differentiation among geographic populations was detected. In addition, while no evidence of recombination was found within any of the four gene fragments, signatures of recombination were found among the four gene fragments in most geographic populations, consistent with sexual mating of this species in nature. Our study provides the first insights into the population genetics and epidemiology of the important plant fungal pathogen C. fructicola.

Camellia japonica L. is an acidophilic ornamental shrub of high economic value that has its center of origin in Japan and has been introduced in several European environmental niches. This exotic species forms arbuscular mycorrhizas (AM), known for their ability to positively affect plant growth. However, AM fungal communities associated to C. japonica in the field have never been characterized. For the first time, the AM fungal community naturally selected by C. japonica was screened in three sites located on the shores of Lake Maggiore (Italy), where specimens of this plant were introduced in the nineteenth century. Mycorrhizal levels were assessed, and the AM fungal communities associated to roots and soil were molecularly characterized based on the small subunit (SSU) rDNA region. The frequency of mycorrhizal roots was high in all sampled root systems (>90 %). Overall, 39 Operational Taxonomic Units (OTUs; 22 Glomerales, 9 Paraglomerales, 7 Archaeosporales, and 1 Diversisporales) were found in the root and soil samples. OTU richness did not significantly differ between the root and the soil niche (5.7 ± 0.6 and 8.0 ± 1.1 average OTUs per sample, respectively) and the three sites analyzed (7.5 ± 0.7, 5.2 ± 1.0, and 7.8 ± 1.5 average OTUs per sample in the three sites, respectively). The AM fungal community composition significantly differed between root-colonizing and soil-dwelling communities and among the three sites under study. Data show a major involvement of edaphic factors, such as available N sources, P, Mg, and K content in soil and soil compaction, in the structuring of the AM fungal communities.

Phytoplasmas ('Candidatus Phytoplasma' spp.) are insect-vectored bacteria that infect a wide variety of plants, including many agriculturally important species. The infections can cause devastating yield losses by inducing morphological changes that dramatically alter inflorescence development. Detection of phytoplasma infection typically utilizes sequences located within the 16S-23S rRNA-encoding locus, and these sequences are necessary for strain identification by currently accepted standards for phytoplasma classification. However, these methods can generate PCR products >1400 bp that are less divergent in sequence than protein-encoding genes, limiting strain resolution in certain cases. We describe a method for accessing the chaperonin-60 (cpn60) gene sequence from a diverse array of 'Ca.Phytoplasma' spp. Two degenerate primer sets were designed based on the known sequence diversity of cpn60 from 'Ca.Phytoplasma' spp. and used to amplify cpn60 gene fragments from various reference samples and infected plant tissues. Forty three cpn60 sequences were thereby determined. The cpn60 PCR-gel electrophoresis method was highly sensitive compared to 16S-23S-targeted PCR-gel electrophoresis. The topology of a phylogenetic tree generated using cpn60 sequences was congruent with that reported for 16S rRNA-encoding genes. The cpn60 sequences were used to design a hybridization array using oligonucleotide-coupled fluorescent microspheres, providing rapid diagnosis and typing of phytoplasma infections. The oligonucleotide-coupled fluorescent microsphere assay revealed samples that were infected simultaneously with two subtypes of phytoplasma. These tools were applied to show that two host plants, Brassica napus and Camelina sativa, displayed different phytoplasma infection patterns.

Endophytic fungi play an important role in terrestrial ecosystem, while little is known about those in hemi-parasitic plants, a group of special plants which absorb nutrients from its hosts by haustoria. The relationship of the endophytes in the two parts of the bipartite systems (hemiparasites together with their hosts) is also poorly understood. Endophytic fungi of a hemi-parasitic plant Macrosolen tricolor, and its host plant Camellia oleifera were investigated and compared in this study. M. tricolor contained rich and diversified endophytic fungi (H' = 2.829), which consisted mainly of ascomycetes, distributed in more than ten orders of four classes (Sordariomycetes, Dothideomycetes, Leotiomycetes and Eurotiomycetes) besides Incertae sedis strains (23.2 % of total). In addition, 2.2 % of isolates were identified to be Basidiomycota, all of which belonged to Agaricomycetes. Obvious differences were observed between the endophytic fungal assembles in the leaves and those in the branches of M. tricolor. The endophytic fungi isolated from C. oleifera distributed in nearly same orders of the four classes of Ascomycota and one class (Agaricomycetes) of Basidiomycota as those from M. tricolor with similar proportion. For both M. tricolor and C. oleifera, Valsa sp. was the dominant endophyte species in the leaves, Torula sp. 1 and Fusarium sp. 1 were the dominant endophytic fungi in the branches. The similarity coefficient of the endophyte assembles in the two host was 64.4 %. Canonical correspondence analysis showed that the endophyte assembles of M. tricolor and C. oleifera were significantly different (p < 0.01).

This paper concentrates on the separation of three conjugated linoleic acid (CLA) isomers (trans-9,trans-11 CLA, trans-10,cis-12 CLA and cis-9,trans-11 CLA) by β-cyclodextrin (β-CD) encapsulation using countercurrent chromatography from Camellia oleifera Abel cake fermented by lactic acid bacteria Lactobacillus sp. LL-ZSDS001. The elution sequence of the CLA isomers, the mixing zones and mechanism of separation are discussed. The separation of 305.9mg of the crude sample yielded three isomeric compounds: 91.3mg of trans-9,trans-11 CLA, 84.1mg of trans-10,cis-12 CLA and 79.7mg of cis-9,trans-11 CLA at high purities of 98%, 94% and 96%, respectively., (Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.)

Ciborinia camelliae is the causal agent of Camellia flower blight. This fungal pathogen is a significant pest of the Camellia floriculture industry because it specifically infects the floral tissue of ornamental camellia cultivars leading to the rapid development of necrotic lesions and blight. This study aims to characterize natural resistance to Ciborinia camelliae within a selection of Camellia spp. Based on macroscopic lesion development, Camellia 'Nicky Crisp' and Camellia lutchuensis were chosen as compatible and incompatible hosts, respectively. Microscopic analyses of the incompatible Camellia lutchuensis-Ciborinia camelliae interaction revealed several hallmarks of induced plant resistance, including papillae formation, H2O2 accumulation, and localized cell death. The compatible Camellia Nicky Crisp-Ciborinia camelliae interaction failed to trigger a similar resistance response. Ciborinia camelliae growth in compatible tissue demonstrated a switch from biotrophy to necrotrophy, evident from the simultaneous development of secondary hyphae and necrotic lesions. Extension of resistance analyses to 39 additional Camellia spp. identified variable levels of resistance within the Camellia genus. The evidence presented supports a resistance breeding strategy for controlling Ciborinia camelliae on ornamental Camellia hybrids.

Fungal assemblages in live, newly shed and partly decomposed leaves of Camellia japonica were investigated with a clone library analysis to assess the fungal diversity and succession in a subtropical forest in southern Japan. Partly decomposed leaves were divided into bleached and adjacent nonbleached portions to estimate the fungi functionally associated with lignin decomposition in the bleached portions, with an emphasis on Coccomyces sinensis (Rhytismataceae, Ascomycota). From 144 cloned 28S ribosomal DNA (rDNA) sequences, 48 operational taxonomic units (OTUs) were defined based on a sequence similarity threshold of 98%. Forty-one (85%) of the 48 OTUs belonged to the Ascomycota and seven OTUs (15%) to the Basidiomycota. Twenty-six OTUs (54%) were detected only once (singletons). The number of OTUs and the diversity indices of the fungal assemblages in the different leaves were in this order: live leaves > newly shed leaves > bleached portions > nonbleached portions of partly decomposed leaves. The fungal assemblages were similar in newly shed leaves and the bleached portions of partly decomposed leaves. Ligninolytic fungi of the genera Coccomyces, Lophodermium and Xylaria were frequently detected in the bleached portions. OTU3, identified as Coccomyces sinensis, was detected in live and newly shed leaves and the bleached portions of partly decomposed leaves, suggesting that this fungus latently infects live leaves, persists after leaf fall and takes part in lignin decomposition.

Two new oxysporone derivatives, pestalrone A (1) and pestalrone B (2), along with two known structurally related compounds 3, 4, were from the fermentation broth of the endophytic plant fungus Pestalotiopsis karstenii isolated from stems of Camellia sasanqua. Their structures and relative configurations were elucidated by extensive spectroscopic analysis and comparison of chemical shifts with related known compounds. Compound 2 exhibited significant activities agains HeLa, HepG2 and U-251 with IC₅₀ values of 12.6, 31.7 and 5.4 µg/mL, respectively.

The prediction model of chlorophyll content of leaves in canopies of oil camelliae under disease was explored and built by analyzing the Vis/NIR spectroscopy characteristics of oil camelliae canopies after being injected with anthracnose. Through field survey of disease index (DI), chlorophyll content and spectral data of leaves in canopies surviving different severity of disease were acquired. The first order differential of spectral data combined with moving average filter was pretreated. The prediction model of BP neural network of chlorophyll content was built by extracting sensitive wave band from spectral resample data. The results showed that with the disease being aggravated, reflection peaks and valleys of spectra of oil camelliae canopies in visible-light region vanished gradually, steep red edges from red light to near infrared leveled little by little, and reflectivity of healthy oil camelliae was far larger than that of ill ones. The sensitive wave band of absorption and reflection of chlorophyll lay in the region of 84-512, 533-565, 586-606 and 672-724 nm. The correlation coefficient r and RMSE between predictive values calculated from BP neural network using sensitive wave band as input variables and observed values was 0.9921 and 0.0458 respectively. It was therefore feasible to utilize Vis/NIR spectroscopy technology to forecast the chlorophyll content of oil camelliae after being infected with anthracnose.

The aim of this study was to evaluate tea polyphenol and purine alkaloid contents of pu-erh tea (Camellia assamica) in a fermentation solid system with Aspergillus niger and Aspergillus fumigatu. In addition, the objective was to find the major intermediate product during fermentation by HPLC-MS(n) analysis. The results showed the change of catechin, ester-catechins and gallic acid by quantitative analysis. In the early stages, the contents of ester-catechins were lightly increased. Then, ester-catechins were gradually degraded to produce catechins and gallic acid. Furthermore, a major metabolic intermediate compound of catechins was observed and elucidated by HPLC-DAD-MS(n) analysis. This study provided a reliable dynamic data description and metabolic pathway of tea polyphenols for postfermented pu-erh tea.

The C(4) plants, whose first product of photosynthetic CO(2) fixation is a four-carbon acid, have evolved independently many times. Crassulacean acid metabolism (CAM) is a biological mechanism known to exhibit some C(4) characteristics such as the C(3) cycle during daylight and demonstrates the C(4) cycle at night. There are also various C(3)-CAM intermediates, whose CAM pathway can be induced by environmental changes. However, neither fungus-induced CAM nor Theaceae CAM have been reported previously. Here, we show that CAM could be generated by fungus infection in Camellia oleifera Abel. young leaves, even at a location of a single leaf where the upper part had been transformed into a succulent one, while the lower part remained unchanged. The early photosynthetic products of dark-grown C. oleifera succulent leaves were malate, whereas C. oleifera normal leaves and light-grown succulent leaves incorporated most of (14)C into the primary photosynthetic product 3-phosphoglycerate. Camellia oleifera succulent leaves have a lower absolute δ(13)C value, much lower photorespiration rates and lower transpiration rates during the day than those of C. oleifera normal leaves. Like a typical CAM plant, stomata of C. oleifera succulent leaves closed during the daylight, but opened at night, and therefore had a detectable CO(2) compensation point in darkness. Net photosynthetic rates (P(n)) fluctuated diurnally and similarly with stomatal aperture. No light intensity saturation could be defined for C. oleifera succulent leaves. C(4) key enzymes in C. oleifera succulent leaves were increased at both the transcriptional/translational levels as well as at the enzyme activity level.

To improve control of camellia twig blight (CTB) using sanitation methods, a more complete epidemiologic understanding of this disease is necessary. Three CTB disease stages were modeled using recurrent event analysis. Wound inoculated stems were observed at regular intervals for appearance of disease symptoms. Survival times (time from inoculation until symptom appearance) for the three disease stages (mild, moderate, and severe) were regressed against stem diameter, monthly mean hours/day within a specified temperature range (15 to 30 degrees C), and season (spring, summer, fall, and winter). For all three CTB disease stages, stem diameter had a protective effect on survival times, while monthly mean hours/day in the specified temperature range and warmer seasons were risk factors. Based upon median ratios, the mild disease stage developed 2 to 3 times faster in spring, summer, and fall than in winter. Similarly, moderate and severe disease stages developed 2 to 2.5 times faster. For all three disease stages, seasonal differences in stage development were smaller among fall, spring, and summer, varying from 1 to 1.6 times faster. Recurrent event modeling of CTB progression provides knowledge concerning developmental expression of this disease, information necessary for creating a comprehensive, integrated disease management program.

Seasonal and leaf age-dependent variations in the endophytic and epiphytic phyllosphere fungal assemblages of Camellia japonica were examined and compared. Live leaves of C. japonica were collected in four seasons (May, Aug, Nov, Feb), and fungi were isolated from healthy-looking leaves of 0, 1, 2 and 3 y old. The infection rate and total number of endophytic fungi increased May-Feb, and species richness of endophytes increased as leaves aged. In contrast the infection rate of epiphytic fungi was 100% for all leaf ages at every sampling date. The total number of epiphytic fungi isolated was greatest in May and lowest in Aug. The species richness of epiphytes did not differ significantly by season or leaf age. Eight fungal species were recorded as major phyllosphere fungi of C. japonica. Seasonal variations were detected for the frequencies of Colletotrichum gloeosporioides, C. acutatum, and epiphytes Pestalotiopsis sp.1, Aureobasidium pullulans, Phoma sp.1 and Ramichloridium sp., whereas the frequency of the endophyte Geniculosporium sp.1 varied with leaf age. The frequency of the epiphyte Cladosporium cladosporioides varied with both season and leaf age.

The microbial transformation of five flavans (1-5) by endophytic fungi isolated from the tea plant Camellia sinensis was investigated. It was found that the endophytic filamentous fungus Diaporthe sp. oxidized stereoselectively at C-4 position of (+)-catechin (1) and (-)-epicatechin (2) to give the correspondent 3,4-cis-dihydroxyflavan derivatives (6, 10), respectively. (-)-Epicatechin 3-O-gallate (3) and (-)-epigallocatechin 3-O-gallate (4) were also oxidized by the fungus into 3,4-dihydroxyflavan derivatives (10, 12) via (-)-epicatechin (2) and (-)-epigallocatechin (11), respectively. Meanwhile, (-)-gallocatechin 3-O-gallate (5), (-)-catechin (ent-1) and (+)-epicatechin (ent-2), which possess a 2S-phenyl substitution, resisted the biotransformation.

The taxonomic position of a rhizosphere isolate, strain 52108a(T), was determined using a polyphasic approach. The strain was found to have chemical and morphological properties consistent with its assignment to the genus Kitasatospora. An almost complete 16S rRNA gene sequence determined for the strain was aligned with corresponding sequences of representatives of the genus Kitasatospora and related taxa using three tree-making algorithms. The organism formed a distinct phyletic line within the Kitasatospora clade and was most closely related to Kitasatospora arboriphila (98.9 %), Kitasatospora kifunensis (99.0 %), Kitasatospora paracochleata (98.4 %) and Kitasatospora terrestris (98.2 %), but was readily distinguished from representatives of these species using a combination of phenotypic properties. The combined genotypic and phenotypic data show that the strain should be classified in the genus Kitasatospora as a novel species. The name proposed is Kitasatospora viridis sp. nov., with the type strain 52108a(T) (=AS 4.1878(T)=DSM 44826(T)).