ABSTRACT
Tobacco (Nicotiana tabacum L.) is one of the most widely cultivated industrial crops worldwide. From April to July 2023, about 40% of tobacco seedlings in the greenhouse exhibited irregular taupe lesions in Zhengzhou, Henan Province, China. At an early stage of the lesion development, light grey spots with the diameter of 1-2 mm were observed, these spots gradually expanded and connected into large irregular lesions causing leaf wrinkling or withered. A total of 12 infected leaf tissues were sterilized with 75% ethanol for 45 s, rinsed three times in sterilized water and then plated on potato dextrose agar (PDA) medium for 10 days at 28°C in darkness. Seven fungal colonies that show the similar appearance were isolated and three of them (MB-1, MB-2 and MB-3) were used for subsequent identification. Colonies of these strains on PDA with loose mycelium and orange-red pigment on the underside, white aerial in the center and light yellow hyphae near the periphery, formed in the shape of a concentric ring pattern. Ascomata appeared from the 14th day, were black, spherical or ellipsoid with walls of textura angularis, and size was 53.8-101.1 µm × 50.3-104.3 µm (n=30). Terminal hairs were brown and straight, gradually tapering toward the tips. Asci clavate or fusiform, spore bearing part 16.2-29.2 × 7.3-11.4 µm (n=21), with 8 irregularly arranged ascospores, evanescent. Ascospores are brown at maturity, biapiculate, navicular or fusiform shapes with size of 8.7-12.8 µm × 4.8-6.9 µm (n=100), and more or less inaequilateral. Single spore strains derived from these strains exhibited the morphological features consistent with the original strains. The morphological characteristics of the fungus were consistent with the description of Arcopilus aureus (Chivers) X.W. Wang & Samson (= Chaetomium aureum Chivers) (Lee et al. 2019). Furthermore, the sequences of RPB2 region were amplified from these strains and the result sequences (GenBank accession no. OR513105-OR513108) all showed a 100.00% identity with A. aureus strain CBS 538.73 (GenBank accession no. KX976807.1). It was reported that the RPB2 gene was efficient in discriminating Arcopilus species (Tavares et al. 2022), thus a maximum likelihood (ML) phylogenetic tree based on the RPB2 gene sequences were constructed using MEGA 7.0 with 1000 replications of bootstrapping (Kumar et al. 2016), which revealed that these strains formed a well-supported clade with A. aureus strains of (CBS 153.52 and CBS538.73) (Wang et al. 2022). Pathogenicity analysis were performed on healthy flue-cured tobacco seedlings leaves (cv Y85) by using mycelial agar plugs (5 mm in diameter) and spore suspension (1×106 spores/mL), and the PDA plugs and sterile water were used for control group, respectively. Tobacco seedlings were incubated in a 25°C and 70% RH growth chamber. After seven days, the leaves showed obvious symptoms, with taupe lesions and yellow halos on the periphery, whereas no symptoms were found on the control leaves. The A. aureu was then reisolated from inoculated diseased leaves. Previously, A. aureus has been only reported to cause leaf black disease on Pseudostellaria heterophylla in China (Yuan et al. 2021). To our knowledge, this is the first reported of A. aureus causing tobacco leaf grey spot worldwide. Arcopilus aureus has been reported as a plant biocontrol fungus (Wang et al. 2013). However, due to the potential serious damage in tobacco seedlings caused by this fungus, the use of A. aureus as a plant biocontrol agent needs to be given more attention, and disease control measures of this pathogen should be developed.
ABSTRACT
A novel strain Serratia marcescens HB-4 with high Cadmium adsorption capacity was isolated from heavy metal contaminated soil in Hunan province, China. S. marcescens HB-4 reduced the concentration of Cd present in wastewater to less than 0.1â¯mg/L when the inlet stream contained no higher than 5.0â¯mg/L Cd. After treatment, wastewater meets Integrated Wastewater Discharge Standard of China (GB8978-1996). The naturally dead S. marcescens HB-4 still maintained over 80% of its Cd adsorption capacity. Scanning electron microscope (SEM), transmission electron microscope (TEM) and energy dispersive spectroscopy (EDS) results suggested that the mechanism of Cd adsorption can be explained as the synergy of extracellular adsorption, periplasm accumulation and intracellular absorption. The size of the accumulated Cd particular is at the nanometer scale, which can be washed out by EDTA without damaging cell integrity. SDS-polyacrylamide gel electrophoresis experiment showed that the heavy metal binding protein (especially Fe binding protein), transporter, amino acid and histidine periplasmic binding proteins and oxidoreductases were responsible for Cd removal. The pot experiment of S. marcescens HB-4 combined with Houttuynia cordata to detoxify Cd contaminated soil showed that the cadmium content in the aboveground and underground parts of Houttuynia cordata increased by 34.48% and 59.13% (w/w), respectively. The cadmium accumulation in Houttuynia cordata increased by 44.27% compared with the blank group which was not combined with S. marcescens HB-4. This work demonstrates that microbial synergistic phytoremediation has a significant potential to treat heavy metal contaminated soil.
