Occurrence of Neopestalotiopsis clavispora Causing Leaf Blight on Acer oblongum in China.

Acer oblongum is native to Southwest China and is also distributed in Nepal and Northern India. It is an excellent garden ornamental tree species, suitable for solitary planting in courtyards and parks. From June to August 2022, severe leaf blight occurred on A. oblongum in Baihe Wetland Park (32°5'42" N, 112°28'13" E) in Nanyang City, China. The foliar disease rate reached 59% (n=100). Early signs were yellow spots on the leaves, mainly on the middle and edge parts. Then, the lesions gradually expanded, became amorphous, and turned yellowish brown, eventually led to necrosis on leaves and branches. Twenty diseased leaves were collected and the junction areas between infected and healthy tissues were cut into 5 x 5 mm2 pieces. The collected plant materials were sterilized in 75% ethanol and 1% NaClO for 30 s and 1 minute, respectively, followed by rinsing in sterile water, and placing on a potato dextrose agar (PDA) plate supplemented with 50 µg ml-1 streptomycin at 25 °C for 3 days. Colony edges were cut and transferred to new PDA plates for purification culture. A total of 18 purified fungal strains were obtained, which showed similar phenotypes in morphological characteristics. All colonies had spread radially with wavy surfaces, and dense cream to white aerial hyphae. After 14 days in culture, black fruiting bodies appeared. Conidia were fusiform to slightly clavate, with five cells and two or three setae, 4.2 to 7.9 µm × 17.5 to 25.4 µm in diameter (n = 100). The apical and basal cells and setae were colorless, three median cells were brown, and the middle cell was dark brown. Morphological characteristics of all 18 strains were consistent with the genus description of Neopestalotiopsis spp. (Maharachchikumbura et al. 2014). Further molecular identification showed that the ITS region sequences of all strains have extremely high homology with Neopestalotiopsis spp. The ß-tubulin gene (TUB), and the translation elongation factor 1-alpha gene (TEF1) were amplified for molecular identification (Shu et al. 2020). The sequences of three representative strains (FE-05, 09, 16) from different regions were deposited in GenBank with accession Nos. OQ867279, OQ867288, OQ867289 (ITS), OQ870207, OQ870208, OQ870209 (TUB), and OQ870204, OQ870205, OQ870206 (TEF1). BLASTn analysis of these sequences showed 99 to 100% identity to Neopestalotiopsis clavispora strains (OK655673, MZ648263 for ITS, ON000362, MZ286974 fr TUB, MH423941, MK512481 for TEF1). These morphological features and molecular identification indicated that the pathogen has the same characteristics as N. clavispora. Pathogenicity was tested on ten healthy 3-month-old seedlings using the three representative strains through in vivo experiments. For each strain, the conidial suspension (106 conidia ml-1) in absorbent cotton balls (50 µl of inoculum) were inoculated onto the healthy leaves of two seedlings, while a total of two other plants were served with sterile water as a blank control. The plants were potted in a climate incubator at 28°C and a relative humidity of approximately 90%. Symptoms consistent with natural lesions were observed on the inoculated leaves after 5 days while the control plants remained healthy. The strains of N. clavispora were reisolated from the symptomatic inoculated leaves, fulfilling Koch's postulates. N. clavispora is known to cause disease in a variety of plants in China, such as Dendrobium officinale (Cao et al., 2022), Fragaria ananassa (Shi et al., 2022), and Garcinia mangostana (Qiu et al., 2019). To the best of our knowledge, this is the first report of N. clavispora causing leaf blight on A. oblongum in China. The yellowing and falling off of leaves would seriously affects the garden landscape. It is necessary to further clarify the host range of the pathogen to select appropriate landscape matching plants in future planning.

First Report of Alternaria alternata Causing Leaf Spot on Catalpa bungei in China.

Catalpa bungei originates from China. It is fast-growing and possesses a vertically aligned trunk, rendering it a commendable construction material and a significant economic species. In July 2022, a serious leaf spot occurred in the LanLake farm (surveyed area of about 700 acres) in Nanyang (33°3'23" N, 112°28'50" E), Henan Province, China. The incidence rate of leaf disease reached 54% (n=100). The disease initially manifested as irregular round spots with a yellowish-brown hue, subsequently extending in all directions. Later, the lesion periphery exhibited a darkening effect, leading to yellowing. Twenty diseased leaves were randomly collected and cut into small pieces at the interfaces between infected and healthy tissues. The tissues were sterilized in a solution of 75% ethanol and 1% NaClO for 30 seconds and 1 minute, respectively. After rinsing in sterile water, the pieces were placed on potato dextrose agar (PDA) plates and incubated at 25°C for 5 days. A total of 29 purified fungal strains were acquired, exhibiting comparable phenotypes in terms of morphological characteristics. Three strains (QS1-1, QS1-2, and QS1-3) were isolated for subsequent investigations. The colony exhibited abundant aerial mycelium with shades ranging from dark green to grey-brown on the reverse side. To analyze the morphological characteristics of conidia, potato carrot agar (PCA) was used as the culture medium and incubated at 25°C with a 12-hour light/dark cycle. Conidia were obclavate or spheroidal, dark brown, with 3 to 5 transverse septa, and 1 to 4 longitudinal septa, measuring 12.4 to 36.7 × 4.4 to 9.0 µm (n=100), with conical beak lengths ranging from 0 to 4.3 µm. These morphological traits suggested that the pathogen shares similarities with the Alternaria species. The rDNA internal transcribed spacer (ITS), translation elongation factor 1-alpha gene (tef1), glyceraldehyde 3-phosphate dehydrogenase gene (gapdh), and RNA polymerase II second largest subunit (rpb2) were amplified for further molecular identification. The resultant sequences were submitted to GenBank with the following accession numbers: OR733559, OR742124, OR761873 (ITS), OR939796, OR939797, OR939798 (tef1), OR939801, OR939802, OR939803 (gapdh), and PP054846, PP054847, PP054848 (rpb2). A Phylogenetic tree was constructed of combined genes (ITS, tef1, gapdh, and rpb2) of sequences, alongside the sequences of the type strains by the neighbor-joining method. The three strains formed a clade with the strains CBS 121456 of Alternaria alternata in phylogenetic trees, being separated from other Alternaria spp. The morphological features and molecular analyses supported the strains as members of Alternaria alternata (Woudenberg et al. 2015). To validate pathogenicity, a conidial suspension (106 conidia ml-1) of all three strains was inoculated onto three healthy leaves of five seedlings, with 50 µl of inoculum absorbed with cotton balls. Another group of five plants received sterile water as a control. All plants were incubated in a climate chamber at 28°C and 90% relative humidity. Four days post-inoculation, lesions resembling natural phenomena were observed, whereas control plants showed no symptoms. Subsequent reisolation produced cultures that were morphologically and molecularly identical to the original strains, fulfilling Koch's postulates. Stem canker of C. bungei caused by Phytophthora nicotianae has been reported in China (Chang et al. 2022). This is the first report of A. alternata causing leaf spots on C. bungei in China. Further research is required on management options to control this disease and the host range still needs to be clarified for accurate disease management.

Inhibition of Verticillium Wilt in Cotton through the Application of Pseudomonas aeruginosa ZL6 Derived from Fermentation Residue of Kitchen Waste.

To isolate and analyze bacteria with Verticillium wilt-resistant properties from the fermentation residue of kitchen wastes, as well as explore their potential for new applications of the residue. A total of six bacterial strains exhibiting Verticillium wilt-resistant capabilities were isolated from the biogas residue of kitchen waste fermentation. Using a polyphasic approach, strain ZL6, which displayed the highest antagonistic activity against cotton Verticillium wilt, was identified as belonging to the Pseudomonas aeruginosa. Bioassay results demonstrated that this strain possessed robust antagonistic abilities, effectively inhibiting V. dahliae spore germination and mycelial growth. Furthermore, P. aeruginosa ZL6 exhibited high temperature resistance (42°C), nitrogen fixation, and phosphorus removal activities. Pot experiments revealed that P. aeruginosa ZL6 fermentation broth treatment achieved a 47.72% biological control effect compared to the control group. Through activity tracking and protein mass spectrometry identification, a neutral metalloproteinase (Nml) was hypothesized as the main virulence factor. The mutant strain ZL6ΔNml exhibited a significant reduction in its ability to inhibit cotton Verticillium wilt compared to the strain P. aeruginosa ZL6. While the inhibitory activities could be partially restored by a complementation of nml gene in the mutant strain ZL6CMΔNml. This research provides a theoretical foundation for the future development and application of biogas residue as biocontrol agents against Verticillium wilt and as biological preservatives for agricultural products. Additionally, this study presents a novel approach for mitigating the substantial amount of biogas residue generated from kitchen waste fermentation.

[Effects of Combined Stress of High Density Polyethylene Microplastics and Chlorimuron-ethyl on Soybean Growth and Rhizosphere Bacterial Community].

With the vigorous development of agriculture in China， plastic mulch film and pesticides are widely used in agricultural production. However， the accumulation of microplastics （formed by the degradation of plastic mulch film） and pesticides in soil has also caused many environmental problems. At present， the environmental biological effects of microplastics or pesticides have been reported， but there are few studies on the combined effects on crop growth and the rhizosphere soil bacterial community. Therefore， in this study， the high density polyethylene microplastics （HDPE， 500 mesh） were designed to be co-treated with sulfonylurea herbicide chlorimuron-ethyl to study their effects on soybean growth. In addition， the effects of the combined stress of HDPE and chlorimuron-ethyl on soybean rhizosphere soil bacterial community diversity， structure composition， microbial community network， and soil function were investigated using high-throughput sequencing technology， interaction network， and PICRUSt2 function analysis to clarify the combined toxicity of HDPE and chlorimuron-ethyl to soybean. The results showed that the half-life of chlorimuron-ethyl in soil was prolonged by the 1% HDPE treatment （from 11.5 d to 14.3 d）， and the combined stress of HDPE and chlorimuron-ethyl had more obvious inhibition effects on soybean growth than that of the single pollutant or control. The HiSeq 2 500 sequencing showed that the rhizosphere bacterial community of soybean was composed of 20 phyla and 312 genera under combined stress， the number of phyla and genera was significantly less than that of the control and single pollutant treatment， and the relative abundances of bacteria with potential biological control and plant growth-promoting characteristics （such as Nocardioides and Sphingomonas） were reduced. Alpha diversity analysis showed that the combined stress significantly reduced the richness and diversity of the soybean rhizosphere bacterial community， and Beta diversity analysis showed that the combined stress significantly changed the structure of the bacterial community. The dominant flora of the rhizosphere bacterial community were regulated， and the abundances of secondary functional layers such as amino acid metabolism， energy metabolism， and lipid metabolism were reduced under combined stress by the analysis of LEfSe and PICRUSt2. It was inferred from the network analysis that the combined stress of HDPE and chlorimuron-ethyl reduced the total number of connections and network density of soil bacteria， simplified the network structure， and changed the important flora species to maintain the stability of the network. The results above indicated that the combined stress of HDPE and chlorimuron-ethyl significantly affected the growth of soybean and changed the rhizosphere bacterial community structure， soil function， and network structure. Compared with that of the single pollutant treatment， the potential risk of combined stress was greater. The results of this study can provide guidance for evaluating the ecological risks of polyethylene microplastics and chlorimuron-ethyl and for the remediation of contaminated soil.

Three new Dioszegia species (Bulleribasidiaceae, Tremellales) discovered in the phylloplane in China.

The genus Dioszegia is comprised of anamorphic basidiomycetous yeasts and is classified in the family Bulleribasidiaceae of the order Tremellales. Currently, 24 species have been described and accepted as members of the genus, although its diversity and global distribution have not been thoroughly investigated. In this study, yeasts were isolated from plant leaves collected in the Guizhou and Henan Provinces of China and identified through a combination of morphological and molecular methods. Phylogenetic analyses of the combined ITS and LSU sequences coupled with morphological studies revealed three novel species, D.guizhouensissp. nov., D.foliicolasp. nov., and D.aurantiasp. nov., proposed here. Additionally, our phylogenetic analyses suggest that the recently discovered species D.terrae is a synonym of D.maotaiensis. This study presents detailed descriptions and illustrations of three new Dioszegia species and highlights distinctions between them and their close relatives. The findings of this study contribute to our knowledge of the biodiversity of Dioszegia, offering a foundation for future research.

5-HT1A receptors within the intermediate lateral septum modulate stress vulnerability in male mice.

Chronic stress is a major risk factor for psychiatric disorders. However, certain individuals may be at higher risk due to greater stress susceptibility. Elucidating the neurobiology of stress resilience and susceptibility may facilitate the development of novel strategies to prevent and treat stress-related disorders such as depression. Mounting evidence suggests that the serotonin (5-HT) system is a major regulator of stress sensitivity. In this study, we assessed the functions of 5-HT1A and 5-HT2A receptors within the lateral septum (LS) in regulating stress vulnerability. Among a group of male mice exposed to chronic social defeat stress (CSDS), 47.2% were classified as stress-susceptible, and these mice employed more passive coping strategies during the defeat and exhibited more severe anxiety- and depression-like behaviors during the following behavioral tests. These stress-susceptible mice also exhibited elevated neuronal activity in the LS as evidenced by greater c-Fos expression, greater activity of 5-HT neurons in both the dorsal and median raphe nucleus, and downregulated expression of the 5-HT1A receptor in the intermediate LS (LSi). Finally, we found the stress-induced social withdrawal symptoms could be rapidly relieved by LSi administration of 8-OH-DPAT, a 5-HT1A receptor agonist. These results indicate that 5-HT1A receptors within the LSi play an important role in stress vulnerability in mice. Therefore, modulation of stress vulnerable via 5-HT1A receptor activation in the LSi is a potential strategy to treat stress-related psychiatric disorders.

Diversity of Wickerhamomyces (Wickerhamomycetaceae, Saccharomycetales) in China with the description of four new species.

Wickerhamomyces is a well-known genus of the family Wickerhamomycetaceae in the class Ascomycetes. These fungi can survive in a variety of substrates and environments and perform many valuable roles in both industrial processes and the natural ecosystems. During our investigation of yeast diversity associated with plant materials, 53 Wickerhamomyces isolates were obtained from rotting wood and plant leaves collected in Fujian, Guizhou, Henan, and Yunnan Provinces of China. Isolates were identified as 14 Wickerhamomyces species, including 1 species known previously to occur in China (W. anomalus), 9 new record species in China (W. arborarius, W. ciferrii, W. edaphicus, W. lynferdii, W. pijperi, W. subpelliculosa, W. xylosica, W. strasburgensis, and W. sydowiorum), and 4 novel species (W. guiyangensis sp. nov., W. paramyanmarensis sp. nov., W. quanzhouensis sp. nov., and W. phyllophilus sp. nov.). This study presents a detailed account of these new species, illustrating their morphology and analyzing their phylogenetic relationships with other Wickerhamomyces species. Our study is the first comprehensive study on Wickerhamomyces species associated with plant materials from tropical and subtropical China. The results of this study update our understanding of the phylogenetic relationships, systematics, and ecology of Wickerhamomyces.

Ultrasonic-Assisted Extraction of Antioxidants from Perilla frutescens Leaves Based on Tailor-Made Deep Eutectic Solvents: Optimization and Antioxidant Activity.

The development of natural antioxidants to replace synthetic compounds is attractive. Perilla frutescens leaves were proven to be rich in antioxidants. The extraction of antioxidants from Perilla leaves via ultrasonic-assisted extraction (UAE) based on choline chloride-based deep eutectic solvents (DESs) was studied. Firstly, several DESs were prepared, and their extraction effects were compared. Secondly, the extraction process was optimized by single-factor experiments and response surface methodology (RSM). Finally, the optimization results were verified and compared with the results of traditional solvent-based UAE. The effects of solvents on the surface cell morphology of Perilla frutescens leaves were characterized by scanning electron microscopy (SEM). Choline chloride-acetic acid-based DES (ChCl-AcA) extract showed a relatively high ferric-reducing antioxidant activity (FRAP) and 2,2-diphenyl-1-picrylhyldrazyl radical scavenging rate (DPPH). Under the optimal operating conditions (temperature 41 °C, liquid-solid ratio 33:1, ultrasonic time 30 min, water content 25%, ultrasonic power 219 W), the experimental results are as follows: DPPH64.40% and FRAP0.40 mM Fe(II)SE/g DW. The experimental and predicted results were highly consistent with a low error (<3.38%). The values of the DPPH and FRAP were significantly higher than that for the water, ethanol, and butanol-based UAE. SEM analysis confirmed that ChCl-AcA enhanced the destruction of the cell wall, so that more antioxidants were released. This study provides an eco-friendly technology for the efficient extraction of antioxidants from Perilla frutescens leaves. The cytotoxicity and biodegradability of the extract will be further verified in a future work.

Genomic Sequence Resource of Talaromyces albobiverticillius, the Causative Pathogen of Pomegranate Pulp Rot Disease.

Talaromyces albobiverticillius, a prominent pathogen responsible for pomegranate pulp rot disease, inflicts significant damage on Punica granatum L. Besides its pathogenicity, this fungus possesses the potential to produce substantial amounts of red pigments, making it promising for industrial applications. This study presents the genome annotation of T. albobiverticillius field strain Tp-2, isolated from pomegranates. The genome assembly, generated through a combination of Oxford Nanopore and Illumina sequencing reads, yielded a high-quality assembly with 14 contigs, featuring an N50 length of 4,594,200 bp. The complete genome of strain Tp-2 spans 38,354,882 bp, with a GC content of 45.78%. Importantly, the assembly exhibits remarkable integrity, with 98.3% of complete Benchmarking Universal Single-Copy Orthologs validating genome completeness. Genome prediction analysis reveals the presence of 10,380 protein-coding genes. To our knowledge, this study is the first report on the genome sequence of T. albobiverticillius, offering valuable insights into its genetic variation and molecular mechanisms of pigment production.

Bacillus sp. YC7 from intestines of Lasioderma serricorne degrades nicotine due to nicotine dehydrogenase.

A large number of nicotine-containing wastes produced during the tobacco manufacturing process are seriously harmful to the environment and human health. The degradation and transformation of nicotine-containing environmental contaminants to harmless substances has become an urgent requirement. Lasioderma serricorne can grow and reproduce in nicotine-rich sources, and their intestinal microbiota show promising potential to degrade and utilize nicotine. The purpose of this study is to screen and identify nicotine-degrading bacteria from the intestines of L. serricorne and explore their degradation characteristics. A dominant strain, YC7, with significant nicotine degradation capabilities was isolated from the intestines of L. serricorne. The strain was identified as Bacillus using a polyphasic approach. The test results showed it can produce multiple enzymes that include ß-glucosidase, cellulase, proteases, and amylases. The nicotine-degrading bacteria were functionally annotated using databases. Nicotine dehydrogenase (NDH) was found by combining an activity tracking test and protein mass spectrometry analysis. The YC-7 NDH in the pathway was molecularly docked and functionally verified via the gene knockdown method. The binding ability of nicotine to nicotine-degrading enzymes was investigated using molecular docking. A high-efficiency nicotine-degrading bacteria, YC-7, was isolated and screened from tobacco, and the gene functions related to degradation were verified. This investigation provides a new hypothesis for screening nicotine-degrading bacteria and increases our knowledge of potential nicotine-degrading microbial sources.

Effect of volatile compounds produced by the cotton endophytic bacterial strain Bacillus sp. T6 against Verticillium wilt.

BACKGROUND: Verticillium wilt, caused by the fungus Verticillium dahliae, leads to significant losses in cotton yield worldwide. Biocontrol management is a promising means of suppressing verticillium wilt. The purpose of the study was to obtain and analyze endophytic bacteria with Verticillium wilt-resistant activities from the roots of Gossypium barbadense 'Xinhai15' and to explore the interactions between the soil and plants. RESULTS: An endophytic bacterium Bacillus sp. T6 was obtained from the Verticillium wilt-resistant cotton G. barbadense 'Xinhai15', which showed significant antagonistic abilities against cotton Verticillium wilt. The bioassay results indicated that the strain possessed strong antagonistic abilities that inhibited V. dahliae spore germination and mycelial growth without contact, and thus it was speculated that the active factor of the bacteria might be volatile compounds. A total of 46 volatile substances were detected via headspace solid-phase microextraction and gas chromatography-mass spectrometry analysis. The pure product verification experiment confirmed that the styrene produced by the T6 strain was the main virulence factor. Transcriptome analysis showed that following styrene induction, 247 genes in V. dahliae, including four hydrolase genes, eight dehydrogenase genes, 11 reductase genes, 17 genes related to transport and transfer were upregulated. Additionally, 72 genes, including two chitinase genes, two protease genes, five transport-related genes, and 33 hypothetical protein genes, were downregulated. The quantitative real-time PCR results confirmed that the expression of the four genes VDAG_02838, VDAG_09554, VDAG_045572, and VDAG_08251 was increased by 3.18, 78.83, 2.71, and 2.92 times, respectively, compared with the uninduced control group. CONCLUSIONS: The research provides a new reference for the development and application of the volatile compounds of endophytic bacteria as new biocontrol agents for the control of Verticillium wilt and as biological preservatives for agricultural products.

Increased Lactobacillus Abundance Contributes to Stress Resilience in Mice Exposed to Chronic Social Defeat Stress.

INTRODUCTION: Accumulating evidence indicates that abnormalities in the composition of gastrointestinal (GI) microbiota play a vital role in stress-related disorders. Both human beings and animals perceive stressful events differently, i.e., resilience or susceptibility. However, the role of GI microbiota in stress resilience/susceptibility and the underlying mechanisms remain largely unknown. METHODS AND RESULTS: Sixty male C57BL/6J mice were exposed to 10-day chronic social defeat stress (CSDS), and 28 were found to be resilient to CSDS. We next analyzed microbiota compositions in the cecum using 16S rDNA gene sequencing, which revealed a significant increase in the relative abundance of Lactobacillus at the genus level in the resilient mice. In subsequent experiments, we found that oral administration of a strain of Lactobacillus (Lactobacillus murinus) for 2 weeks attenuated the increased levels of stress-induced corticosterone and anxiety-like behavior in stress-susceptible mice. The mRNA expression of tryptophan hydroxylase 2 (a rate-limiting enzyme in serotonin [5-HT] synthesis) was also significantly increased in the dorsal raphe nucleus (DR) of stress-susceptible mice. CONCLUSIONS: Lactobacillus contributes to stress resilience, and the DR 5-HT system may play an important role during this process. The above results suggest that certain organisms in the GI tract may play an essential role in stress response and be useful in the prevention and treatment of some stress-related psychiatric disorders, such as depression.

Gray Blight Disease on Euonymus japonicas Caused by Pestalotiopsis disseminata in China.

Euonymus japonicas is widely planted as an important landscape species throughout China. In June 2021, a serious gray blight disease was detected on E. japonicas in Henan Province (32°30'58" N, 112°19'44" E), causing severe defoliation of infected trees with a foliar disease incidence of 52 to 70% (n = 100). Gray spots initially appeared on leaves, gradually expanded into irregular white blotches with dark brown borders, eventually leading to wilting and death of the leaves. The junctions between the lesion and healthy tissue of infected leaves were cut into 3 × 3-mm pieces, surface sterilized with 1% NaClO solution for 1 min, rinsed in sterile water, and placed on PDA plates with 50 µg/ml of streptomycin. Three isolates (HY94, HY95, and HY98) were selected for subsequent experiments. The colonies reached 80-85 mm diam after 7 days at 25°C, with undulated margins, white to pale in color, with moderate aerial mycelium on the surface. Conidiomata were globose, solitary, and dark black. Conidia were ellipsoid, straight to slightly curved, 4-septate, 19 to 26.4 × 5 to 7.5 µm (n=100). The apical cell was cylindrical and hyaline, with 2 to 3 tubular apical appendages, unbranched, filiform, 2.5 to 3.5 µm in length. The basal appendage was single, unbranched, centric, 1.5 to 3 µm long. The characteristics were close to those of Pestalotiopsis spp. (Maharachchikumbura et al. 2013). The genomic DNA was extracted, and the rDNA internal transcribed spacer (ITS), the ß-tubulin gene (TUB), and the translation elongation factor 1-alpha gene (TEF1) were amplified by primers ITS1/ITS4, Bt2a/Bt2b, and EF1-728F/EF1-986R, respectively (Carbone and Kohn, 1999). Sequences were submitted to GenBank with accession numbers OL840327-OL840329(ITS), OL961454-OL961456(TUB), and OL961448-OL961450 (TEF1). BLASTn analyses of ITS, TUB, and TEF1 sequences exhibited 99.46, 99.05, and 96.53% similarity to the sequences of Pestalotiopsis disseminata strain MEAN1166 (ITS, 548/551 bp; MT374688) (Silva et al. 2020), PSH2000I-066 (TUB, 418/422 bp; DQ333575), and TAP29O082 (TEF1, 250/259 bp; AB453850), respectively in GenBank. The three isolates formed a clade with the type strains, MEAN 1166 and MAFF238347 of P. disseminata in phylogenetic trees, being clearly seperated from other Pestalotiopsis spp. Based on morphological and molecular evidence, the pathogen was identified as P. disseminata (Maharachchikumbura et al. 2011). To fulfill Koch's postulates, pathogenicity was tested with three isolates. Ten healthy leaves of 5-year-old intact plants were used per isolate and inoculated with mycelial plugs on both nonwounded and wounded leaves. Control leaves were inoculated with agar plugs. The inoculated plants were placed at 28°C in a greenhouse (90% relative humidity). Distinct lesions were observed after 10 days. The pathogen reisolated was identical to that of the original cultures according to phenotype and ITS sequences. The control leaves showed no obvious symptoms. P. disseminata is known to cause disease on several important plants in China, such as Camellia japonica (Zhang et al. 2012), Pinus armandii (Hu et al. 2007), and Tripterygium wilfordii (Kumar et al. 2004). This is the first report of gray blight disease caused by P. disseminata on E. japonicas in China and worldwide. The fungal pathogen identification will provide valuable information for prevention and management of gray blight disease associated with E. japonicas.

Yeast Diversity in the Qaidam Basin Desert in China with the Description of Five New Yeast Species.

The Qaidam Basin is the highest and one of the largest and driest deserts on Earth. It is considered a mars analog area in China. In contrast to numerous studies concerning its geology, geophysical, and chemistry, relatively few studies have reported microbial diversity and distribution in this area. Here, we investigated culturable yeast diversity in the northeast Qaidam Basin. A total of 194 yeast strains were isolated, and 12 genera and 21 species were identified, among which 19 were basidiomycetous yeasts. Naganishia albida, N. adeliensis, and Filobasidium magnum were the three most dominant species and were distributed in thirteen samples from eight locations. Five new species (Filobasidium chaidanensis, Kondoa globosum, Symmetrospora salmoneus, Teunia nitrariae, and Vishniacozyma pseudodimennae) were found and described based on ITS and D1D2 gene loci together with phenotypic characteristics and physiochemical analysis. Representative strains from each species were chosen for the salt-tolerant test, in which species showed different responses to different levels of NaCl concentrations. Further, the strain from soil can adapt well to the higher salt stress compared to those from plants or lichens. Our study represents the first report of the yeast diversity in the Qaidam Basin, including five new species, and also provides further information on the halotolerance of yeasts from the saline environment in mars analog.

Characterization of a novel chitinolytic Serratia marcescens strain TC-1 with broad insecticidal spectrum.

The Gram-negative rod-shaped bacterium Serratia marcescens is an opportunistic pathogen of many organisms, including insects. We report the identification and optimal in vitro chitinase production conditions of a novel chitinolytic S. marcescens strain TC-1 isolated from a naturally infected white grub (Anomala corpulenta) collected from a peanut field at Nanyang city, Henan province, China. Strain identification was conducted by morphological, physiological, biochemical and molecular analyses. The amplified 16S rRNA gene of TC-1 showed a similarity greater than 99% with multiple strains of S. marcescens. Based on Neighbor-joining phylogenetic tree analysis of bacterial 16S rRNA gene sequences, TC-1 formed a clade with S. marcescens, clearly separated from other Serratia spp. The strain TC-1 showed larvicidal activities against five insect species (A. corpulenta, Plutella xylostella, Spodoptera exigua, Helicoverpa armigera, Bombyx mori) and the nematode Caenorhabditis elegans, but not against S. litura. The operating parameters of chitinase production by TC-1 were optimized by response surface methodology using a three-factor, three-level Box-Behnken experimental design. The effects of three independent variables i.e. colloidal chitin concentration (7-13 g l-1), incubation time (24-72 h) and incubation temperature (24-32 °C) on chitinase production by TC-1 were investigated. A regression model was proposed to correlate the independent variables for an optimal chitinase activity predicted as 20.946 U ml-1, using a combination of colloidal chitin concentration, incubation time and incubation temperature of 9.06 g l-1, 63.83 h and 28.12 °C, respectively. The latter agreed well with a mean chitinase activity of 20.761 ± 0.102 U ml-1 measured in the culture supernatants of TC-1 grown under similar conditions with a colloidal chitin concentration, incubation time and incubation temperature of 9 g l-1, 64 h and 28 °C, respectively. Our study revealed the S. marcescens strain TC-1 with potential as a biocontrol agent of insect pests and nematodes and demonstrated the proposed regression model's potential to guide chitinase production by this strain.

Phytobacter diazotrophicus from Intestine of Caenorhabditis elegans Confers Colonization-Resistance against Bacillus nematocida Using Flagellin (FliC) as an Inhibition Factor.

Symbiotic microorganisms in the intestinal tract can influence the general fitness of their hosts and contribute to protecting them against invading pathogens. In this study, we obtained isolate Phytobacter diazotrophicus SCO41 from the gut of free-living nematode Caenorhabditis elegans that displayed strong colonization-resistance against invading biocontrol bacterium Bacillus nematocida B16. The colonization-resistance phenotype was found to be mediated by a 37-kDa extracellular protein that was identified as flagellin (FliC). With the help of genome information, the fliC gene was cloned and heterologously expressed in E. coli. It could be shown that the B. nematocida B16 grows in chains rather than in planktonic form in the presence of FliC. Scanning Electronic Microscopy results showed that protein FliC-treated B16 bacterial cells are thinner and longer than normal cells. Localization experiments confirmed that the protein FliC is localized in both the cytoplasm and the cell membrane of B16 strain, in the latter especially at the position of cell division. ZDOCK analysis showed that FliC could bind with serine/threonine protein kinase, membrane protein insertase YidC and redox membrane protein CydB. It was inferred that FliC interferes with cell division of B. nematocidal B16, therefore inhibiting its colonization of C. elegans intestines in vivo. The isolation of P. diazotrophicus as part of the gut microbiome of C. elegans not only provides interesting insights about the lifestyle of this nitrogen-fixing bacterium, but also reveals how the composition of the natural gut microbiota of nematodes can affect biological control efforts by protecting the host from its natural enemies.

Stenotrophomonas nematodicola sp. nov., isolated from Caenorhabditis elegans.

A Gram stain-negative, coccoid rod-shaped, motile by gliding, facultatively aerobic bacterium, designated W5, was isolated from Caenorhabditis elegans samples in Baotianman Natural Reserve (33° 27' 47'' N; 111° 48' 32'' E), Nanyang, China. The isolate was characterized taxonomically using a polyphasic approach. The 16S rRNA gene of strain W5 exhibited 98.1-99.7% similarity to the 16S rRNA genes of members of the genus Stenotrophomonas, and < 98.0% similarities to those of other bacterial species in the family Lysobacteraceae. The most closely related strains were Stenotrophomonas rhizophila JCM 13333T (99.7%) and Stenotrophomonas bentonitica DSM 103927T (99.2%). The predominant respiratory quinone of the isolate is Q-8. The major fatty acids are iso-C15:0 (38.2%) and antesio-C15:0 (16.6%). The draft genome of strain W5 had a length of 4,402,751 bp and a DNA G + C content of 67.3 mol%. The ANI values between the draft genomes of strain W5 and its closest phylogenetic neighbors S. rhizophila JCM 1333T and S. bentonitica DSM 103927T were 84.7% and 85.0%, respectively. The DDH value between W5 and S. rhizophila JCM 13333T was 30.8%, which was the highest DDH level. We propose that strain W5 represents a novel bacterial species with the name Stenotrophomonas nematodicola sp. nov. and W5 as the type strain. The type strain is W5 (= CPCC 101271T = CGMCC 19401T = KCTC XXXT).

The enhanced mechanisms of Hansschlegelia zhihuaiae S113 degrading bensulfuron-methyl in maize rhizosphere by three organic acids in root exudates.

The residues of bensulfuron-methyl (BSM), a sulfonylurea herbicide, in soil have caused serious damage to the rotation of susceptible crops. Many studies have reported that the removal of BSM in soil was achieved by adding degrading bacteria. However, the mechanisms used by bacteria to degrade BSM in the crop rhizosphere remain unclear. In this study, a BSM-degrading bacterium, Hansschlegelia zhihuaiae S113, was applied to investigate the enhancement of effects mediated by organic acids during the bioremediation of BSM-contaminated maize rhizosphere soil. Organic acids, such as L-malic acid, tartaric acid, and fumaric acid, identified in maize root exudates, significantly stimulated the expression of cheA, which encoded the histidine kinase in strain S113 and contributed to the chemotactic response. This process accelerated the accumulation of strain S113 around the maize roots and promoted the colonization process on maize roots. The growth of strain S113 was significantly increased by L-malic acid but not tartaric acid or fumaric acid. After the S113 suspension was root-irrigated to BSM-contaminated soil, the density of strain S113 colonizing root surfaces and in rhizosphere soil reached 1.1 × 104 cells/g for roots and 4.9 × 104 cells/g in dry soil at 15 d, leading to 80.9% BSM degradation efficiency. The treatment with the addition of a mixture of S113 and L-malic acid completely degraded BSM in rhizosphere soil due to the strong attraction and growth promotion of strain S113 by L-malic acid, with a higher efficiency than that with the extra addition of fumaric acid (89.7%) or tartaric acid (87.0%). This paper revealed the enhancement effects of organic acids identified in root exudates for the in situ bioremediation of BSM-contaminated rhizosphere soil.

Stenotrophomonas strain CPCC 101271, an intestinal lifespan-prolonging bacterium for Caenorhabditis elegans that assists in host resistance to "Bacillus nematocida" colonization.

The soil-dwelling, opportunistic pathogenic bacterium "Bacillus nematocida" B16 exhibits strong killing activities against a variety of pathogenic nematodes via a "Trojan horse" mechanism that can kill worm species like Caenorhabditis elegans. The bacterial strain CPCC 101271 was previously isolated from the intestines of C. elegans that were recovered from natural habitats and can serve as a probiotic for C. elegans, while also assisting in resistance to infection by the pathogenic strain B16. In this study, the lifespan of C. elegans fed with strain CPCC 101271 cells was extended by approximately 40% compared with that of worms fed with Escherichia coli OP50 cells. In addition, the colonization of C. elegans by the pathogenic bacterium "B. nematocida" B16 was inhibited when pre-fed with strain CPCC 101271. Metagenomic sequence analysis of intestinal microbiota of C. elegans fed with strain CPCC 101271 and infected with B16 revealed that pre-feeding worms with CPCC 101271 improved the diversity of the intestinal bacteria. Moreover, community structure significantly varied in coordination with Stenotrophomonas spp. and Bacillus spp. abundances when competition between strains CPCC 101271 and B16 was evaluated. In conclusion, the nematode microbiota strain CPCC 101271 assisted in its host resistance to colonization by the pathogen "Bacillus nematocida" and can also promote life span-prolongation in C. elegans. These results underscore that understanding the interactions between C. elegans microbiota and pathogens can provide new insights into achieving effective biological control of agricultural pests.