The Change in Fibrinogen is Associated with Outcome in Patients with Acute Ischemic Stroke Treated with Endovascular Thrombectomy.

BACKGROUND: Fibrinogen has been identified as a modulator of the coagulation and inflammatory process. There is uncertainty about the relationship between the dynamic profile of fibrinogen levels and its impact on clinical outcomes in patients with acute ischemic stroke treated with endovascular thrombectomy. METHODS: We consecutively enrolled patients with acute ischemic stroke who underwent endovascular thrombectomy. Fibrinogen was measured on admission and during hospitalization. The change in fibrinogen (Δfibrinogen) was calculated as the highest follow-up fibrinogen minus admission fibrinogen, with a positive Δfibrinogen indicating an increase in fibrinogen level. Functional outcome was assessed by the modified Rankin Scale at 3 months. Poor outcome was defined as modified Rankin Scale > 2. RESULTS: A total of 346 patients were included (mean age 67.4 ± 13.6 years, 52.31% men). The median fibrinogen on admission was 2.77 g/L (interquartile range 2.30-3.39 g/L). The median Δfibrinogen was 1.38 g/L (interquartile range 0.27-2.79 g/L). Hyperfibrinogenemia (> 4.5 g/L) on admission was associated with an increased risk of poor outcome [odds ratio (OR) 5.93, 95% confidence interval (CI) 1.44-24.41, p = 0.014]. There was a possible U-shaped association of Δfibrinogen with outcomes, with an inflection point of - 0.43 g/L (p = 0.04). When Δfibrinogen was < - 0.43 g/L, a higher decrease in fibrinogen (lower Δfibrinogen value) was associated with a higher risk of poor outcome (OR 0.22, 95% CI 0.02-2.48, p = 0.219). When Δfibrinogen was > - 0.43 g/L, the risk of poor outcome increased with increasing fibrinogen (OR 1.27, 95% CI 1.04-1.54, p = 0.016). CONCLUSIONS: In patients with endovascular thrombectomy, hyperfibrinogenemia on admission was associated with poor functional outcomes at 3 months, whereas Δfibrinogen was associated with poor 3-month outcomes in a possible U-shaped manner.

Grazing alters the soil nematode communities in grasslands: A meta-analysis.

Grazing causes great disturbances in grassland ecosystems and may change the abundance, diversity, and ecological function of soil biota. Because of their important role in nutrient cycling and as good environmental indicators, nematodes are very representative soil organisms. However, the mechanisms by which grazing intensity, livestock type, duration, and environmental factors (e.g., climate and edaphic factors) affect soil nematodes remain poorly understood. In this study, we collected 1964 paired observations all over the world from 53 studies to clarify the grazing response patterns of soil nematodes and their potential mechanisms. Overall, grazing significantly decreased the abundance of bacterial-feeding (BF) nematodes (-16.54%) and omnivorous-predatory (OP) nematodes (-36.81%), and decreased nematode community diversity indices (Shannon-Weiner index: -4.33%, evenness index: -9.22%, species richness: -5.35%), but had no effect on ecological indices under a global regional scale. The response of soil nematodes to grazing varied by grazing intensity, animals, and duration. Heavy grazing decreased OP nematode abundance, but had no effect on the abundance of other trophic groups, or on diversity or ecological indices. Grazing by small animals had stronger effects than that by large animals and mixed-size animals on BF, fungal-feeding (FF), plant-feeding (PF) and OP nematodes, the Shannon-Wiener index, and the species richness index. The abundance of FF and OP nematodes influenced significantly under short-term grazing. The evenness index decreased significantly under long-term grazing (>10 years). Climate and edaphic factors impacted the effects of grazing on nematode abundance, diversity, and ecological indices. When resources (i.e., rain, heat, and soil nutrients) were abundant, the negative effects of grazing on nematodes were reduced; under sufficiently abundant resources, grazing even had positive effects on soil nematode communities. Thus, the influence of grazing on soil nematode communities is resource-dependent. Our study provides decision makers with grazing strategies based on the resource abundance. Resource-poor areas should have less grazing, while resource-rich areas should have more grazing to conserve soil biodiversity and maintain soil health.

Mariniradius sediminis sp. nov., a multi-xenobiotics degrading genes harbouring bacterium isolated from sediment of river.

A Gram-stain-negative, strictly aerobic, non-motile, catalase- and oxidase-positive, pink and rod-shaped strain, designated RY-2T, was isolated from sediment of Fuyang River located in Wuqiang County, Hengshui City, Hebei Province, PR China. The strain grew at 25-45 °C (optimum, 37 °C), pH 7.0-8.0 (optimum, pH 7.0) and in the presence of 0-1.5 % (w/v) NaCl (optimum, 1 %). From the phylogenetic analysis of the 16S rRNA gene sequence, strain RY-2T was affiliated to the genus Mariniradius, and had the highest 16S rRNA gene sequence similarity to Mariniradius saccharolyticus JCM 17389T (98.3 %) and the similarity values between strain RY-2T and other type strains was all below 89.3 %. The genome size of strain RY-2T was 4.75 Mb and the DNA G+C content was 46.6 %. Values of digital DNA-DNA hybridization and average nucleotide identity between strain RY-2T and the reference strain were 63.2 and 95.5 %, respectively. The major fatty acids (≥5.0 %) were iso-C15 : 0 (37.9 %), summed feature 9 (8.4 %, iso-C17 : 1 ω9c and/or C16 : 010-methyl), anteiso-C15 : 0 (8.2 %), iso-C17 : 0 3-OH (7.6 %) and summed feature 4 (5.2 %, iso-C17 : 1 I and/or anteiso-C17 : 1 B) and its sole menaquinone was MK-7. The polar lipids consisted of phosphatidylethanolamine, an unknown phosphoglycolipid, an unidentified phospholipid, two unidentified aminolipids, three unidentified glycolipids and nine unidentified lipids. Based on the results of biochemical, physiological, phylogenomic and chemotaxonomic analyses, strain RY-2T is considered to represent a novel species of the genus Mariniradius within the family Cyclobacteriaceae, for which the name Mariniradius sediminis sp. nov. is proposed. The type strain is RY-2T (=GDMCC 1.2781T=JCM 35631T).

Flavihumibacter fluminis sp. nov., a novel thermotolerant bacterium isolated from river silt.

A yellow, Gram-stain-positive, strictly aerobic, thermotolerant, non-motile and rod-shaped bacterial strain, designated RY-1T, was isolated from a silt sample of Fuyang River, Wuqiang County, Hengshui City, Hebei Province, PR China. Cells showed oxidase- and catalase-positive activities. Growth occurred at 20-45 °C (optimum, 37 °C) and pH 6.0-8.0 (optimum, pH 7.0), and in the presence of 0-1.5 % (w/v) NaCl (optimum, 0%). A phylogenetic tree based on 16S rRNA gene sequences revealed that strain RY-1T formed a phylogenetic lineage with Flavihumibacter members within the family Chitinophagaceae. A comparison of 16S rRNA gene sequences showed that strain RY-1T was most closely related to Flavihumibacter cheonanensis WS16T (98.6 %), Flavihumibacter sediminis CJ663T (97.7 %) and Flavihumibacter solisilvae 3-3T (97.6 %). The genome size of strain RY-1T was 4.71 Mb, and the DNA G+C content was 44.3  %. The average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values between strain RY-1T and reference strains were all lower than the threshold values for species delineation. Strain RY-1T contained menaquinone-7 and iso-C15 : 0, iso-C17 : 0 3-OH and iso-C15 : 1G as the sole respiratory isoprenoid quinone and major cellular fatty acids (≥5 %), respectively. The major polar lipids consisted of phosphatidylethanolamine, three unidentified aminolipids and four unidentified lipids. According to the results of phenotypic, phylogenetic and chemotaxonomic characteristics, strain RY-1T represents a novel species of the genus Flavihumibacter, for which the name Flavihumibacter fluminis sp. nov. is proposed. The type strain is RY-1T (=GDMCC 1.2775T=JCM 34870T).

Improved biosynthesis of heme in Bacillus subtilis through metabolic engineering assisted fed-batch fermentation.

BACKGROUND: Heme is an iron/porphyrin complex compound, widely used in the health care, food, and pharmaceutical industries. It is more advantageous and attractive to develop microbial cell factories to produce heme by fermentation, with lower production costs and environmentally more friendly procedures than those of the traditional extraction based on animal blood. In this study, Bacillus subtilis, a typical industrial model microorganism of food safety grade, was used for the first time as the host to synthesize heme. RESULTS: The heme biosynthetic pathway was engineered as four modules, the endogenous C5 pathway, the heterologous C4 pathway, the uroporphyrinogen (urogen) III synthesis pathway, and the downstream synthesis pathway. Knockout of hemX encoding the negative effector of the concentration of HemA, overexpression of hemA encoding glutamyl-tRNA reductase, and knockout of rocG encoding the major glutamate dehydrogenase in the C5 pathway, resulted in an increase of 427% in heme production. Introduction of the heterologous C4 pathway showed a negligible effect on heme biosynthesis. Overexpression of hemCDB, which encoded hydroxymethylbilane synthase, urogen III synthase, and porphobilinogen synthase participating in the urogen III synthesis pathway, increased heme production by 39%. Knockouts of uroporphyrinogen methyltransferase gene nasF and both heme monooxygenase genes hmoA and hmoB in the downstream synthesis pathway increased heme production by 52%. The engineered B. subtilis produced 248.26 ± 6.97 mg/L of total heme with 221.83 ± 4.71 mg/L of extracellular heme during the fed-batch fermentation in 10 L fermenter. CONCLUSIONS: Strengthening endogenous C5 pathway, urogen III synthesis pathway and downstream synthesis pathway promoted the biosynthesis of heme in B. subtilis. The engineered B. subtilis strain has great potential as a microbial cell factory for efficient industrial heme production.

Flavobacterium potami sp. nov., a multi-metal resistance genes harbouring bacterium isolated from shallow river silt.

Environmental pollution by heavy metals is becoming an increasing problem and has become a matter of great concern due to the adverse effects worldwide. In this study, we report a novel strain of multi-metal resistant bacteria. A Gram-stain-negative, strictly aerobic, non-motile, yellow, rod-shaped strain 17AT, was isolated from the shallow silt of Fuyang River located in Longdian town, Hengshui city, Hebei province, China. Strain 17AT grew at 20-35 °C (optimum, 30 °C), pH 5-10 (optimum, pH 7) and 0-2% (w/v) NaCl (optimum, 1%). Phylogenetic analyses of 16S rRNA gene sequences showed that strain 17AT was closely related to members of the genus Flavobacterium, and had the highest 16S rRNA gene sequence similarity with 'Flavobacterium panacis' DCY106T (97.5%), followed by Flavobacterium johnsoniae subsp. johnsoniae UW101T (97.3%), Flavobacterium cutihirudinis E89T (97.2%), Flavobacterium limi THG-AG6.4T (97.2%), Flavobacterium hibisci THG-HG1.4T (97.2%) and Flavobacterium johnsoniae subsp. aurantiacum DSM 6792T (97.1%). The genome size of strain 17AT was 5.4 Mb and the DNA G + C content was 34.0%. The average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values among strain 17AT and reference strains were in the ranges of 79.8-86.1%, 24.1-31.4% and 80.5-88.6%, respectively, lower than the threshold values for species delineation. Strain 17AT contained iso-C15:0 and C16:0 3-OH as the predominant fatty acids (≥ 10%). The main isoprenoid quinone of strain 17AT was identified as MK-6. The polar lipids consisted of phosphatidylethanolamine, three unidentified aminolipids, two unidentified aminophospholipids and six unidentified lipids. Comparative genomics analysis between strain 17AT and its reference type strains revealed that there are a number of metal-resistant genes in strain 17AT, which are located in 15 gene clusters responsible for the copper homeostasis, cobalt-zinc-cadmium resistance, copper resistance, and arsenic/antimony resistance, with the copper resistance protein NlpE being unique to 17AT. Combined data from phenotypic, phylogenetic and chemotaxonomic studies demonstrated that strain 17AT is a representative of a novel species within the genus Flavobacterium, for which the name Flavobacterium potami sp. nov. is proposed. The type strain is 17AT (= GDMCC 1.2723T = JCM 34833T).

First Report of Colletotrichum gloeosporioides Causing Anthracnose on Wisteria sinensis in China.

Wisteria (Wisteria sinensis) is a well-known ornamental plant for environmental protection in the garden, which also has a high value for medicinal use. In December 2021, leaf spots were observed on W. sinensis plants growing on the campus of Jiangxi Agricultural University in Nanchang, Jiangxi Province (28.45° N, 115.49° E), with a incidence rate of 40% plants were infested (n = 100 investigated plants). Initially leaf spots were small and pale brown (Approx. 2 mm in diameter), which gradually expanded into round or irregular dark brown spots as disease progressed, and lesions developed greyish-white necrotic tissues in the center at later stages, eventually causing the leaves to rot. To isolate the pathogen, tissues (5 × 5 mm) at the margin of lesions were cut from ten symptomatic leaves, surface disinfected with 75% ethanol for 30 s followed by 2% sodium chloride (NaClO) for 1 min, rinsed three times with sterile distilled water, and the dried tissues were cultured on potato dextrose agar (PDA) at 28 ± 1℃ in darkness for 3 days. After culture purification, five isolates were obtained and the representative single spore isolate (ZTTJ1) was used for subsequent identification tests. After 10 days of incubation on PDA medium, colonies had dense aerial mycelium with a gray center and dark gray-green mycelium outward, with orange-red conidial masses distributed in a ring on the surface. The underside of the colonies was light gray to dark gray. Conidia were cylindrical, with ends obtuse-rounded, 11.83 to 20.74 × 3.34 to 5.33 µm (av=16.11 µm × 4.26 µm, n = 50) in size. These morphological characteristics were consistent with Colletotrichum gloeosporioides (Shi et al, 2019). Six conserved regions of isolate (ZTTJ1), internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), calmodulin (CAL), ß-tublin (TUB), actin (ACT), and chitin synthase 1 (CHS1) gene regions were amplified using ITS1/ITS4 (Gardes et al, 1993), GDF/GDR (Templeton et al, 1992), CL1C/CL2C (Li et al, 2018), Bt2a/Bt2b (Prihastuti et al, 2009), ACT-512F/ACT-783R and CHS-79F/CHS-345R (Carbone et al, 1999) primers, respectively. Using BLAST, ITS, GAPDH, CAL, TUB, ACT, and CHS1 gene sequences (GenBank Accession No. OP703312, OP713773, OP713775, OP713776, OP713772, OP713774, respectively) were over 99% identical to C. gloeosporioides (GenBank Accession No. MK967281, MH594288, MT449307, MN624110, MN107239 and MN908602, respectively). A maximum likelihood (ML) phylogenetic analysis based on ITS-ACT-GAPDH-CHS1-CAL-TUB2 sequences using MEGA7.0, placed isolate (ZTTJ1) within C. gloeosporioides. To complete Koch's postulates, 10 µL spore suspension (1.0 × 106 conidia/mL) of ZTTJ1 (7-day-old culture on PDA medium) was dropped onto 10 leaves wounded with a sterilized needle and 10 non-wounded leaves, respectively. Ten wounded leaves were inoculated with sterile water as controls. All leaves were incubated at 28 ± 1 °C and 90 % relative humidity (12 h/12 h light/dark). After 7 days, all wounded leaves inoculated with C. gloeosporioides developed symptoms as previously observed, while the control and non-wounded leaves remained healthy. The fungus re-isolated from the inoculated leaves were identified as C. gloeosporioides by morphological and molecular identification; the pathogen causing disease in W. sinensis was determined to be C. gloeosporioides. To our knowledge, this is the first report of C. gloeosporioides causing anthracnose on W. sinensis in China. This work has identified the pathogenic species of the disease, which helps to take targeted measures to control its spread, providing a basis for the prevention and treatment of the disease.

First Report of Leaf Spot Disease on Chamaedorea elegans Caused by Fusarium oxysporum in China.

Chamaedorea elegans, native to Mexico and Guatemala, is a commonly planted indoor and small-scale garden ornamental due to its stately appearance, tolerance of low light levels, and its ability to improve air quality (El-Khateeb et al. 2010). In December 2021, an unknow leaf-spot disease was observed on C. elegans in Ganzhou City of Jiangxi Province, China (25.83 °N, 114.93 °E). The symptoms were small brown spots on the leaves, gradually expanded into irregular dark brown spots with necrotic tissue forming in the center of the lesions (Figure 2 A-1 and A-2). To isolate the pathogen, the diseased leaves were surface sterilized in 75% ethanol for 30 s. Small pieces of tissue (5 × 5 mm) were taken from the margin between diseased and healthy tissue, disinfected 1% NaClO for 45 s, washed three times in sterile water, and then placed on PDA at 25 ± 1°C for 5 days. Later, five isolates were purified from single spores and each of the five isolates has the same properties as described below. The isolates had abundant pale purple flocculent hyphae with purple pigmentation (Figure 2 C-1 and C-2). Macroconidia were falciform, straight or slightly curved, 1-2 septate, 11.75 to 22.99 × 3.06 to 4.44 µm (µ=16.08 µm × 3.37 µm, n=50) (Figure 2 D-1). Microconidia were oval or elliptical, a septate, 4.03 to 9.19 × 1.92 to 3.73 µm (µ=5.88 µm × 2.66 µm, n=50) (Figure 2 D-2). Chlamydospores formed singly or in pairs, and were terminal or intercalary in hyphae (Figure 2 D-3). Based on morphological characteristics, the fungus was preliminarily identified as a Fusarium sp. (Leslie et al. 2006). To confirm the identification, primers ITS1/ITS4 (White et al. 1990), RPB2-5f2/RPB2-7cr (O'Donnell et al. 2010; Liu et al. 1999) and TEF 1-αF/TEF 1-αR (O'Donnell et al. 2000) were used to amplify and sequence apportion of the ITS, RPB2 and TEF (Table 1). The sequences (Genebank accession number: OM780148, OM782679, OM782680) shared 100% idnetity with Fusarium oxysporum (Genebank accession number: MH866024.1, MH484930.1, MH485021.1). The maximum likelihood (ML) phylogenetic analysis of the concantenated ITS, RPB2 and TEF sequences was performed in MEGA7.0. (Sudhir et al. 2016), assigning the isoaltes to the F. oxysporum species complex (Figure 1). To confirm the pathogenicity, nine pots of healthy 3-year-old C. elegans plants were inoculated in the greenhouse (12 h light/12 h dark cycle, RH 90 %, three for wounded inoculation, three for nonwounded inoculation and three for control). Fifty disinfected leaves were wounded with sterile needles and fifty remained unwounded. The wounded (Figure 2 B-1 and B-2) and unwounded leaves were inoculated with a 10 µL spore suspension (1.0 × 106 conidia/ml) which was taken from each of the five isolates cultured for 7 days. Fifty leaves were mock-inoculated with sterile water (Figure 2 B-3 and B-4). After incubation for 7 days, the wounded leaves inoculated with the spore suspension had similar symptoms to the original diseased leaves, while the unwounded leaves and the control leaves did not develop symptoms. The experiment was repeated three times and the pathogens was reisolated from wound-inoculated leaves with the same morphological characteristics to the original pathogens, and identified as F. oxysporum by morphological and molecular analysis, completing Koch's postulates. F. oxysporum, a pathogen with a broad spectrum of hosts, ranks 5th among the top 10 fungal plant pathogens (Amjad et al. 2018.) and has been reported to Carpinus betulus, Citrullus lanatus, Pinus pinea (Mao et al. 2021; Muhammad et al. 2021; Monther et al. 2021). To our knowledge, this is the first report of leaf spot disease on C. elegans caused by F. oxysporum in China. C. elegans is an important ornamental plant in China with high economic value, so the disease has the potential to be a threat to its cultivation industry.

Lysobacter selenitireducens sp. nov., isolated from river sediment.

A Gram-stain-negative, yellow-pigmented, motile, flagellated and rod-shaped bacterium, designated as 13AT, was isolated from a river sediment sample of Fuyang River in Hengshui City, Hebei Province, PR China. Strain 13AT grew at 10-37 °C (optimum, 30 °C), at pH 5.0-11.0 (optimum, pH 7.0) and at 0-7 % (w/v) NaCl concentration (optimum, 0 %). Phylogenetic analysis based on the 16S rRNA gene sequence showed that strain 13AT belongs to the genus Lysobacter, and was most closely related to Lysobacter spongiicola DSM 21749T (97.8 %), Lysobacter concretionis DSM 16239T (97.5 %), Lysobacter daejeonensis GIM 1.690T (97.3 %) and Lysobacter arseniciresistens CGMCC 1.10752T (96.9 %). Meanwhile, the type species Lysobacter enzymogenes ATCC 29487T was selected as a reference strain (95.2 %). The genomic size of strain 13AT was 3.0 Mb and the DNA G+C content was 69.0 %. The average nucleotide identity values between strain 13AT and each of the reference type strains L. spongiicola DSM 21749T, L. concretionis DSM 16239T, L. daejeonensis GIM 1.690T, L. arseniciresistens CGMCC 1.10752T and L. enzymogenes ATCC 29487T were 75.9, 76.1, 77.7, 78.0 and 73.2 %, respectively. The digital DNA-DNA hybridization values between strain 13AT and each of the reference type strains were 21.7, 22.2, 21.9, 22.7 and 23.2 %, respectively. The average amino acid identity values between strain 13AT and each of the reference type strains were 72.5, 72.9, 72.3, 75.0 and 69.2 %, respectively. The major fatty acids were iso-C15 : 0, iso-C16 : 0 and summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 0 10-methyl). The sole respiratory quinone was identified as ubiquinone-8. The polar lipid profile contained phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, an unidentified aminolipid, an unidentified lipid, four unidentified phospholipids and two unidentified glycolipids. Based on the phenotypic, physiological, phylogenetic and chemotaxonomic data, strain 13AT represents a novel species of the genus Lysobacter, for which the name Lysobacter selenitireducens sp. nov. is proposed. The type strain is 13AT (=JCM 34786T=GDMCC 1.2722T).

Flavobacterium selenitireducens sp. nov., isolated from rhizosphere soil of ancient mulberry.

A Gram-stain-negative, non-motile, aerobic, yellow, convex, rod-shaped mesophilic bacterial strain, designated strain D33T, was isolated from rhizosphere soil of ancient mulberry in Dezhou city, Shandong province, PR China. The strain grew at 8-37 °C (optimum, 30 °C), pH 4-9 (optimum, pH 7) and growth occurred at 0.5-5.5 % (w/v) NaCl (optimally at 1 %). The results of the phylogenetic analyses of 16S rRNA gene and whole genome sequences indicated that D33T was closely related to members of the genus Flavobacterium and had the highest 16S rRNA gene sequence similarity with 'Flavobacterium agri' KACC 19300 (95.4 %), Flavobacterium ichthyis NST-5T (94.6 %), Flavobacterium ahnfeltiae KCTC 32467T (93.6 %) and Flavobacterium longum JCM 19141T (93.6 %). The genome size of D33T was 3.8 Mb and the DNA G+C content was 48.0 mol%. The average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH) and average amino acid identity (AAI) values among D33T and reference strains were lower than the threshold values for species delineation. The only respiratory quinone of D33T was menaquinone 6 (MK-6). The predominant fatty acids (>5 %) were C15:0, C16 : 0, C18 : 0, iso-C15:0, iso-C17 : 0 3-OH, anteiso-C15 : 0 and summed feature 9 . The polar lipid profile contained phosphatidylethanolamine, two unidentified aminophospholipids, three unidentified aminolipids and two unidentified lipids. Combined data from phenotypic, phylogenetic and chemotaxonomic studies indicated that D33T is a representative of a novel species of the genus Flavobacterium, for which the name Flavobacterium selenitireducens sp. nov. is proposed. The type strain is D33T (=GDMCC 1.1946T=KACC 22131T).

First Report of Postharvest Fruit Rot in Solanum muricatum Aiton Caused by Alternaria alternata in southwest China.

Solanum muricatum is native to South America and well known for its sweet, attractive, nutritious fruits. S. muricatum has been cultivated in China since the 1980s and increasingly popular (Li et al. 2015). In November 2021, an unknown fruit rot was observed in Shilin County of Yunnan Province (24.77 °N, 103.28 °E). The incidence of this disease was about 16% of 500 postharvest S. muricatum fruits after 7 d in storage room (25°C, 90% relative humidity). The initial symptoms were small brown spots on the fruit surface, which gradually expanded into irregular brown or black lesions, and gray-white mold developed in the center of the lesions, eventually the fruit turned rot. To isolate the pathogen, ten fruits with typical symptoms were collected and surface-sterilized with 75% ethanol for 45 s. Small fragments (5 × 5 mm) from the margin of lesions on fruit were disinfected with 1% sodium hypochlorite for 60 s, washed three times with sterile water then transferred to potato dextrose agar (PDA), and incubated at 28 ± 1℃ for 3 days (Li et al. 2022). Two fungal isolates with the same morphology were obtained and purified by single-spore isolation method. The colony was covered with thick fluffy aerial mycelia and the center was dark brown or black with white margins. Conidia were brown, pyriform or ellipsoid, with 1 to 3 longitudinal and 2 to 6 transverse septa, 15.12 to 34.01 × 6.90 to 12.73 µm (21.22 × 9.69 µm on average, n=50) in size. These morphological characteristics were consistent with Alternaria alternata (Li et al. 2015; Xiang et al. 2021; Alberto. 1992). For molecular identification, genomic DNA was extracted from a representative isolate, and primers ITS1/ITS4 (Gardes et al. 1993), TEF-F/TEF-R (Lawrence et al. 2013), Alt-F/Alt-R (Hong et al. 2005), GPD-F/GPD-R (Berbee et al. 1999) and EPG-F/EPG-R (Peever et al. 2004) were used to amplify the internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF), Alternaria major allergen (Alt a1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and endo-polygalacturonase (endoPG), respectively. The obtained DNA sequences (ITS: OM049821; TEF: OM069656; Alt a1: OM069655; GAPDH: OM069654 and endoPG: OM069653) showed over 99% homology with that of A. alternata (GenBank Accession No. MN856355.1 (565/573 bp); MN258023.1 (267/267 bp); KY923227.1 (491/501 bp); LC131645.1 (608/609 bp) and MN698284.1 (452/454 bp)). A phylogenetic tree based on the combined ITS, TEF, Alt a1, GAPDH, and endoPG sequences using the maximum likelihood methods with Kimura 2-parameter model, bootstrap nodal support for 1000 replicates in MEGA7.0 (Li et al. 2019) revealed that the isolate was assigned to A. alternata. To confirm pathogenicity, 10 µL spore suspension (1.0 × 106 conidia/ml) obtained from 7-day-old PDA cultures of each isolate were inoculated on 15 needle-wounded and 15 non-wounded surface-disinfected fruits, respectively. Healthy fruits were inoculated with sterile water as controls and the experiment was repeated 3 times. All fruit were incubated at 25 ± 1℃, 90% relative humidity. After 7 days, all the wounded and non-wounded fruit inoculated with A. alternata showed similar symptoms to those observed on the previously fruits, while the control fruits remained healthy. The same pathogen was again isolated from the inoculated fruits, thus Koch's postulates were fulfilled. A. alternata causing fruit rot of Prunus avium and Mangifera indica in China were reported in previous studies (Ahmad et al. 2020; Liu et al. 2019). As far as we know, this is the first report of postharvest fruit rot on S. muricatum caused by A. alternata in southwest China. This work provides a basis for the development of control strategies of the disease in the future.

First report of Fusarium tricinctum causing fruit rot on navel orange (Citrus sinensis (L.) Osbeck) in China.

Citrus sinensis (L.) Osbeck is popular with consumers for its delicious taste. In December 2020, a rot symptom causing about 15% losses of a total of 450 fruits was observed on 'Newhall' navel oranges after 70 d storage (20℃, 85%-90% RH) at Jiangxi Key Laboratory for Postharvest Technology and Nondestructive Testing of Fruits & Vegetables (28.68° N, 115.85° E). The fruits were harvested from an orchard in Ganzhou City, Jiangxi Province, China (25.53° N, 114.79° E). Incipiently, the pedicles of infected fruits were brown, the peels became softened and showed yellowish-brown lesions which, gradually expanded and had white hyphae (Fig. S1A). To isolate the pathogen, the surface of diseased fruits was disinfected with NaClO (2%) for 2 min and ethanol (75%) for 0.5 min, then washed with sterile water three times. Tissues (5 × 5 mm) around the lesion were incubated on potato dextrose agar (PDA) at 28 ± 1℃ (L: D=12: 12) for 5 days. Five cultures with similar morphology were obtained and colonies initially produced white aerial hyphae and became khaki then turned pink on PDA (Fig. S1F, G, H). Abundant microconidia, macroconidia and rare chlamydospores were observed after 10 days on PDA and no glucose PDA media (Zhang et al. 2020). Macroconidia were falciform and curved to lunate, 2-4 septa, 29.38 × 3.75 µm in size (n=50) (Fig. S1K, Fig. S3). Microconidia were oval, napiform or pyriform, 0-1 septa, 12.00 × 3.43 µm in size (n=50) (Fig. S1L1 to L4, Fig. S3). Chlamydospores were found in hyphae, ellipsoidal or orbicular (Fig. S1I-1 to I-2, J-1 to J-2). The morphological features of five isolates were similar to Fusarium (Leslie and Summerell 2006). Genomic DNA of five isolates was extracted with DNA Extraction Kit (Yeasen, Shanghai, China), ITS1/ITS4, EF1Ha/EF2Tb and fRPB2-5F/fRPB2-7cR primers were used to amplify the internal transcribed spacer region (ITS), and the transcriptional elongation factor-1 alpha (TEF-1α), and RNA polymerase II (RPB2) gene sequences (White et al. 1990; Carbone and Kohn 1999; Liu et al. 1999). The ITS, TEF-1α and RPB2 sequences of five isolates were deposited in GenBank and showed 99-100% identity with corresponding sequences from F. tricinctum (Table S1). A phylogenetic tree was constructed with ITS-TEF-1α-RPB2 concatenated sequences in MEGA7.0 (Li et al. 2021) and all five isolates were placed in F. tricinctum clade with 100% bootstrap support (Fig. S2). To confirm pathogenicity, ten healthy C. sinensis fruits were surface-sterilized with 75% ethanol and inoculated with 10 µL spore suspension (1.0 × 106 spore/mL) including five wounded (with sterilized needle) and five unwounded (Fig. S1B to E). Control fruits were inoculated with 10 µL sterile water. All fruits were incubated at 28 ± 1℃, 90% RH for 7 days. The experiment was conducted three times. The lesion diameter of inoculated wounded fruits was 21.01 ± 2.52 mm and showed similar symptoms to original rotten fruits. However, the control and unwounded fruits remained healthy. To fulfill Koch's postulates, F. tricinctum was re-isolated from the inoculated fruits and deposited in Collaborative Innovation Center of Postharvest Key Technology and Quality Safety of Fruits and Vegetables in Jiangxi Province. To our knowledge, F. tricinctum has been reported on apple tree and kiwi plant in China (Zhang et al. 2021; Ma et al. 2022), but this is the first report of F. tricinctum causing fruit rot on navel orange in China. This finding provides important information for preventing postharvest disease of citrus.

First Report of Colletotrichum karstii Causing Anthracnose of Begonia lanternaria Irmsch. in China.

Begonia lanternaria Irmsch., an ornamental plant endemic in China, which is commonly used in landscape and interior decoration. In March 2021, an estimated 30% B. lanternaria plants were observed with anthracnose-like symptoms at a botanical garden conservation greenhouse in Mengla County of Yunnan Province (21.91° N, 101.21°E). Initially, small black spots developed on the disease leaves, which gradually expanded into irregular necrotic lesions surrounded by a yellowish halo, eventually turned wilting and defoliating. Twenty diseased leaves were collected and surface-disinfested with 75% ethanol for 30 s. Small fragments (5 × 5 mm) from the margin of lesions were disinfected with 1% NaClO for 120 s, washed with sterile water three times, and cultured on potato dextrose agar (PDA) at 28 ± 1℃. After 3 days single spores from four fungal colonies with identical morphology were isolated. Colonies on PDA were 70-75 mm diam in 7 d (7.5-10.6 mm/d), with dense white to gray-white mycelia attached with brown to black-brown acervulus. The underside of the culture was yellow to yellowish-brown concentric circle. Conidia were single-celled, hyaline, straight to slightly curved, cylindrical, 12.88 to 16.66 × 6.25 to 7.97 µm (av=14.65 µm × 7.22 µm, n=50) in size. For molecular identification, genomic DNA was extracted from a representative isolate, and the internal transcribed spacer, glyceraldehyde-3-phosphate dehydrogenase, calmodulin gene, ß-tublin, actin, and chitin synthase 1 genes were amplified with ITS1/ITS4 (Gardes et al, 1993), GDF/GDR (Templeton et al, 1992), CL1C/CL2C (Li et al, 2018), Bt2a/Bt2b (Prihastuti et al, 2009), ACT-512F/ACT-783R and CHS-79F/CHS-345R (Carbone et al, 1999) primers, respectively. The obtained DNA sequences showed over 99% homology with Colletotrichum karsti (GenBank Accession No. ITS: NR144790; GAPDH: KX578772; CAL: KY039988; TUB2: KX578804; ACT: LC412408; CHS1: KU251855), and the results of sequences were deposited into GenBank with accession No. MZ496954 (522/522 bp), MZ504978 (238/238 bp), MZ504979 (737/737 bp), MZ504982 (472/472 bp), MZ504981 (273/273 bp), MZ504980 (282/284 bp). The phylogenetic tree combined with ITS-ACT-GAPDH-CHS 1-CAL-TUB2 concatenated sequences using the maximum likelihood methods showed that the isolate was C. karsti. To confirm pathogenicity, Koch's postulates were conducted on intact plants, 10 µl spore suspension (1.0 × 106 conidia/ml) of each of four isolates (7-day-old culture on PDA) was inoculated on 15 wounded with a sterilized needle or non-wounded healthy living leaves, and 15 wounded leaves were inoculated with sterile water as controls. All leaves were incubated at 28 ± 1°C and 90% relative humidity (12 h/12 h light/dark). After 5 days, all wounded leaves inoculated with C. karsti showed symptoms similar to those previously observed, while the control and non-wounded leaves remained healthy. Colletotrichum karsti was re-isolated from inoculated leaves. C. karsti was previously reported to cause disease on Nicotiana tabacum L. (Zhao et al, 2020), Stylosanthes guianensis (Jia et al, 2017) and Fatsia japonica (Xu et al, 2020) in China. To our knowledge, this is the first report of C. karsti causing anthracnose of B. lanternaria Irmsch. in China. This disease reduces the ornamental and economic value of B. lanternaria Irmsch., and this work will provide a basis for the prevention and treatment of the disease in the future.

Paraflavitalea devenefica sp. nov., isolated from urban soil.

A Gram-stain-negative, rod-shaped, mesophilic, milky white-pigmented, aerobic, non-spore-forming and non-flagellated bacterium, designated strain X16T, was isolated from urban soil of Zibo, Shandong, China. According to 16S rRNA gene sequence analysis, the isolate showed highest similarities with Paraflavitalea soli 5GH32-13T (97.6 %), Pseudoflavitalea soli KIS20-3T (96.2 %), Pseudobacter ginsenosidimutans Gsoil 221T (96.0 %) and Pseudoflavitalea rhizosphaerae T16R-265T (95.8 %). The neighbour-joining tree based on 16S rRNA gene sequences showed that strain X16T formed a subcluster with Paraflavitalea soli 5GH32-13T, and the subcluster was closely related to Pseudoflavitalea soli KIS20-3T, Pseudobacter ginsenosidimutans Gsoil 221T and Pseudoflavitalea rhizosphaerae T16R-265T. Strain X16T also formed a subcluster with Paraflavitalea soli 5GH32-13T in phylogenetic tree based on genomic sequences. The polar lipids are phosphatidylethanolamine, two unknown aminolipids, two unknown aminophospholipids, two unknown lipids and two unknown phospholipids. The major quinone of strain X16T is menaquinone-7 and the main fatty acids (>10 % of total fatty acids) of strain X16T are iso-C15 : 0, iso-C17 : 0 3-OH and iso-C15 : 1 G. The genome length of strain X16T is 8.7 Mb with a DNA G+C content of 47.4 %. ANI values among strain X16T and strain Paraflavitalea soli 5GH32-13T, Pseudobacter ginsenosidimutans Gsoil 221T, and Pseudoflavitalea rhizosphaerae T16R-265T are 78.1, 70.7, 70.6 %, respectively. On the basis of the results of the polyphasic characterization presented in this study, it is concluded that strain X16T represents a novel species. Besides, strain X16T can detoxify high toxicity selenite [Se(IV)] to low toxicity elemental selenium [Se(0)], for which the name Paraflavitale devenefica sp. nov. is proposed. The type strain is X16T (=KACC 21698T=GDMCC1.1757T).

Association between low-density cholesterol change and outcomes in acute ischemic stroke patients who underwent reperfusion therapy.

BACKGROUND: Low-density lipoprotein cholesterol (LDL-C) can increase cardiovascular risk. However, the association between LDL-C change and functional outcomes in acute ischemic stroke (AIS) patients who underwent reperfusion therapy remains unclear. METHODS: Patients who received reperfusion therapy were consecutively enrolled. LDL-C measurement was conducted at the emergency department immediately after admission and during hospitalization. The change of LDL-C level (ΔLDL-C) was calculated by subtracting the lowest LDL-C among all measurements during hospitalization from the admission LDL-C. Poor functional outcome was defined as modified Rankin Scale (mRS) > 2 at 90 days. RESULTS: A total of 432 patients were enrolled (mean age 69.2 ± 13.5 years, 54.6 % males). The mean LDL-C level at admission was 2.55 ± 0.93 mmol/L. The median ΔLDL-C level was 0.43 mmol/L (IQR 0.08-0.94 mmol/L). A total of 263 (60.9 %) patients had poor functional outcomes at 90 days. There was no significant association between admission LDL-C level and functional outcome (OR 0.99, 95 % CI 0.77-1.27, p = 0.904). ΔLDL-C level was positively associated with poor functional outcome (OR 1.80, 95 % CI 1,12-2.91, p = 0.016). When patients were divided into tertiles according to ΔLDL-C, those in the upper tertile (T3, 0.80-3.98 mmol/L) were positively associated with poor functional outcomes compared to patients in the lower tertile (T1, -0.91-0.13 mmol/L) (OR 2.56, 95 % CI 1.22-5.36, p = 0.013). The risk of poor functional outcome increased significantly with ΔLDL-C tertile (P-trend = 0.010). CONCLUSIONS: In AIS patients who underwent reperfusion therapy, the decrease in LDL-C level during hospitalization was significantly associated with poor functional outcomes at 90 days.

First Report of Alternaria alternata causing fruit Rot on Tetradium ruticarpum in China.

Tetradium ruticarpum, previously and commonly known as Evodia rutaecarpa, is a tree that produces a fruit which is one of the most important traditional Chinese medicine herbs in China (Zhao et al. 2015). In July 2019, an investigation of diseases of T. ruticarpum was conducted in the farmland of Ruichang County (29.68° N, 115.65° E), Jiujiang City, China. An unknown fruit rot disease was observed and the incidence rate was estimated to be 60% to 70% within a 5,000 m2 area. The early symptoms appeared as small circular to irregular dark brown or black spots on the fruit, which gradually coalesced to a light brown-to-black discoloration and caused fruit rot. To identify the causal agent of the disease, 10 diseased fruits were collected and surface disinfected with 2% sodium hypochlorite for 2 min, 70% ethanol for 30 s, rinsed in sterile water and dried on filter paper. Tissues from non-symptomatic tissue as well as from the margin between healthy and affected edge were incubated on potato dextrose agar (PDA) at 25±1°C (12 h light/dark) with 90% relative humidity for 5 days. The colonies were brown to black with abundant whitish margins. Conidiophores were brown and measured 20.40 - 43.10×1.30 - 4.20 µm (25.47 × 2.35 µm on average, n=50). Conidia produced in single or branched chains, were obclavate or ovoid, approximately 9.90 - 32.80×6.50 - 14.50 µm (28.75×12.57 µm on average, n=50) with 2 to 5 transverse septa and 0 to 3 longitudinal septa. The colonies were consistent with Alternaria alternata (Simmons 2007). For molecular identification, the f partial internal transcribed spacer (ITS) regions, Glyceraldehyde-3-phosphate dehydrogenase (gapdh) genes, translation elongation factor 1-alpha (TEF) and Alternaria major allergen (Alt a1) gene of the isolate were amplified using primers ITS1/ITS4 (White et al. 1990), GDF/GDR (Templeton et al. 1992), EF1-728F/EF1-986R (Carbone and Kohn 1999) and Alt-for/Alt-rev (Hong et al. 2005). Sequence data showed 100% homology to A. alternata (GenBank accessions No.MN625176.1 (570/570 bp), MK683866.1 (618/618 bp), MK637432.1 (281/281 bp), KT315515.1 (488/488 bp)), respectively and the sequence data were deposited into GenBank with accession numbers MN897753 (ITS), MT041998 (gapdh), MT041999 (TEF), and MT042000 (Alt a1). Based on both morphological and molecular characteristics, the pathogen was identified as A. alternata. To confirm pathogenicity, 10 µl of a spore suspension (1.0 × 106 conidia/ml) obtained from 5-day-old PDA cultures of the strain were inoculated on 20 wounded (using sterile needle) and 20 nonwounded healthy T. ruticarpum fruits previously disinfected in 75% ethanol. Control fruits including 20 wounded fruits and 20 nonwounded fruits were inoculated with sterilized water. All fruits were incubated at 25±1°C (12 h light/dark) with 90% relative humidity. Four days later, all the wounded and non-wounded fruits showed the initial symptoms of black rot which was similar to that observed in the field, while the wounded and nonwounded fruits treated with sterile water remained healthy. The same pathogen was again isolated from the inoculated fruits. The pathogenicity experiment was repeated three times with the same results. As far as we know, this is the first report of A. alternata causing fruits rot on T. ruticarpum in China, and the identification of the pathogen will provide useful information for developing effective control strategies.

Soil nitrogen availability drives the response of soil microbial biomass to warming.

Although soil microbial biomass responses to experimental warming have been extensively studied, the mechanisms through which elevated temperatures influence soil microbial biomass remain unclear. In this study, we performed a global meta-analysis to quantify the global pattern of soil microbial biomass in response to warming. Our findings suggest that global warming effect is not apparent when all the data are pooled together, while warming does increase microbial biomass under specific conditions (Δ°C ≥ 2 °C). This constructive influence is particularly accentuated under certain circumstances, including high precipitation levels (>800 mm), short treatment durations (<1 year), and within agricultural ecosystems. More importantly, our findings suggest that the impact of global warming on soil microbial biomass is largely mediated by changes in soil nitrogen availability. These findings underscore the pivotal role of nitrogen availability in modulating the response of soil microbial biomass to warming, while also emphasizing the intricate influence between multiple factors such as temperature, duration, and precipitation in shaping the patterns of warming effects.

Fabrication of a Three-Dimensional Microfluidic System from Poly(methyl methacrylate) (PMMA) Using an Intermiscibility Vacuum Bonding Technique.

In this study, the fabrication of microfluidic chips through the bonding of poly (methyl methacrylate) (PMMA) boards featuring designed patterns to create a three-dimensional sandwich structure with embedded microchannels was explored. A key focus was optimization of the interface quality of bonded PMMA pairs by adjusting the solvent, such as such as acetone, alcohol, and their mixture. Annealing was conducted below 50 °C to leverage the advantages of low-temperature bonding. Because of the differences in the chemical reactivity of PMMA toward acetone, alcohol, and their combinations, the resulting defect densities at the bonding interfaces differed significantly under low-temperature annealing conditions. To achieve the optimal sealing integrity, bonding pressures of 30 N, 40 N, and 50 N were evaluated. The interface was analyzed through microstructural examination via optical microscopy and stress measurements were determined using digital photoelasticity, while the bonding strength was assessed through tensile testing.

Soil microbial community variation among different land use types in the agro-pastoral ecotone of northern China is likely to be caused by anthropogenic activities.

There are various types of land use in the agricultural and pastoral areas of northern China, including natural grassland and artificial grassland, scrub land, forest land and farmland, may change the soil microbial community However, the soil microbial communities in these different land use types remain poorly understood. In this study, we compared soil microbial communities in these five land use types within the agro-pastoral ecotone of northern China. Our results showed that land use has had a considerable impact on soil bacterial and fungal community structures. Bacterial diversity was highest in shrubland and lowest in natural grassland; fungal diversity was highest in woodland. Microbial network structural complexity also differed significantly among land use types. The lower complexity of artificial grassland and farmland may be a result of the high intensity of anthropogenic activities in these two land-use types, while the higher structural complexity of the shrubland and woodland networks characterised by low-intensity management may be a result of low anthropogenic disturbance. Correlation analysis of soil properties (e.g., soil physicochemical properties, soil nutrients, and microbiomass carbon and nitrogen levels) and soil microbial communities demonstrated that although microbial taxa were correlated to some extent with soil environmental factors, these factors did not sufficiently explain the microbial community differences among land use types. Understanding variability among soil microbial communities within agro-pastoral areas of northern China is critical for determining the most effective land management strategies and conserving microbial diversity at the regional level.