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Contamination control and removal are very important technical aspects of microbiological research. Bacterial contamination is very common in fungal cultures. Currently, the commonly used approach for inhibiting bacteria is antibiotic treatment; however, there are drawbacks to using antibiotics, including incomplete removal, limited antibacterial spectra, tendency toward recontamination, effects to fungal strains, and potential risks to the environment. Therefore, in the present work, we developed a new method for bacterial removal from fungi cultured on solid medium, the Cabin-Sequestering (CS) method, based on the different culture characteristics between fungi and bacteria. First, 3–5 mm round or square holes (the "cabin") are excavated on a solid medium plate. The fungal strain containing possible bacterial contamination is inoculated into the cabin. The cabin is then covered with a sterilized coverslip, followed by incubation at the appropriate temperature. After 7–10 days of culturing, fungal hyphae grow out along the edge of the coverslip; however, the contaminating bacteria cannot pass through the space formed between the medium and the coverslip and, thus, remain in the cabin. The newly grown fungal hyphae around the coverslip are re-inoculated into fresh culture plates, where they form bacteria-free fungal colonies. The CS method is easy handling, with a short experimental cycle and rare recontamination. When necessary, it can also be used in combination with antibiotics in bacterial removal operations. [ABSTRACT FROM AUTHOR]

Bacillus strains are broadly studied for their beneficial role in plant growth and biological control of plant disease and pest; however, little is known about their underlying mechanisms. In this study, we assessed the controlling and defence-related mechanisms of three Bacillus strains including rice seed-associated strain B. subtilis A15, rhizobacterial strains B. amyloliquefaciens D29 and B. methylotrophicus H8, all of which are against bacterial leaf blight (BLB) caused by Xanthomonas oryzae pv. oryzae. Results indicated that all three strains showed strong biofilm formation ability. The culture filtrates of each strain significantly suppressed the growth and biofilm formation of X. oryzae, while changes in bacterial cell morphology such as cell swell and severe cell wall alterations were observed through the transmission electron microscopy images. PCR analysis revealed that all three strains harbour the antimicrobial-associated genes that are responsible for biosynthesis of bacillomycin, fengycin, iturin and surfactin. Subsequent real-time qPCR analysis revealed the upregulated expression of fenD and srfAA genes in D29 and H8, and fenD and ituC genes in A15 during their in vitro interaction with X. oryzae. It suggests that the antibacterial mechanisms of the three strains may be at least partially associated with their ability to secrete corresponding lipopeptides. Interestingly, the applications of the three strains in greenhouse conditions were found to be effective in controlling the BLB disease, which was achieved through the activation of inducing systemic resistance resulted from the enhanced activities of defence-related enzymes. This is the first report of demonstration of the mode of antibacterial effect of Bacillus strains against X. oryzae. Overall, data from the current study provide valuable information for biological control of BLB disease in rice. [ABSTRACT FROM AUTHOR]

This present study deals with synthesis, characterization and antibacterial activity of cross-linked chitosan-glutaraldehyde. Results from this study indicated that cross-linked chitosan-glutaraldehyde markedly inhibited the growth of antibiotic-resistant Burkholderia cepacia complex regardless of bacterial species and incubation time while bacterial growth was unaffected by solid chitosan. Furthermore, high temperature treated cross-linked chitosan-glutaraldehyde showed strong antibacterial activity against the selected strain 0901 although the inhibitory effects varied with different temperatures. In addition, physical-chemical and structural characterization revealed that the cross-linking of chitosan with glutaraldehyde resulted in a rougher surface morphology, a characteristic Fourier transform infrared (FTIR) band at 1559 cm-1, a specific X-ray diffraction peak centered at 2? = 15°, a lower contents of carbon, hydrogen and nitrogen, and a higher stability of glucose units compared to chitosan based on scanning electron microscopic observation, FTIR spectra, X-ray diffraction pattern, as well as elemental and thermo gravimetric analysis. Overall, this study indicated that cross-linked chitosan-glutaraldehyde is promising to be developed as a new antibacterial drug. [ABSTRACT FROM AUTHOR]

Abstract: Inoculation methods for rice blast fungus Magnaporthe oryzae to Brachypodium distachyon were developed to investigate the infection process and symptom development in comparison with those on rice (Oryza sativa) and barley (Hordeum vulgare). M. oryzae could infect leaves, sheathes, stems and panicles of B. distachyon and cause blast disease. Spraying conidial suspension on either intact seedlings or leaf segments induced typical symptoms on B. distachyon. During the intact seedling inoculation, the symptom developed on B. distachyon leaves closely resembled that on rice; but the lesions on B. distachyon had better uniformity in shapes and sizes than those on rice or barley. In the leaf segments inoculation, only initial and low-developed lesions could be found on rice, while normal symptoms on B. distachyon and barley. Inoculated with low-virulent mutants of M. oryzae, B. distachyon produced low-level symptoms. The symptom level of each mutant on B. distachyon corresponded well to that on rice. In addition, typical infection processes presented on B. distachyon leaves: forming melanized appressoria, penetrating into host epidermis and then forming hyphae in epidermal cells. According to these results, B. distachyon can be used as a candidate for studying fungus-plant interactions and as a probable source of disease resistance. [Copyright &y& Elsevier]

Abstract: To find new genes involved in fungal pathogenicity, a mutant (B11) exhibiting enhanced pathogenicity was isolated from an Agrobacterium-mediated transformed Magnaporthe oryzae mutant library. Southern blotting analysis showed that T-DNA insertion in the B11 genome was a single copy. TAIL-PCR and sequence alignment analyses revealed that a putative gene locus MG01679 was interrupted by the T-DNA fragment. By using the PCR-based method, the DNA and cDNA of the mutant gene MG01679 was cloned and sequenced. The open reading frame of MG01679 includes one intron and two exons, and the coding sequence is 696 bp in length and encodes a 231 amino acid peptide. Protein similarity analysis indicated that the gene belongs to the ThiJ/Pfp I protein family, and the gene was thus designated MgThiJ1. MgThiJ1 showed 57% similarity to FOXG_09029 from Fusarium oxysporum and 54% similarity to FGSG_08979 from F. graminearum in protein sequence. MgThiJ1 gene might act as a negative regulator in vegetative growth and pathogenesis in filamentous fungi, and its specific mechanism needs to be studied further. [Copyright &y& Elsevier]

Abstract: Global warming, which is caused by greenhouse gas emissions, makes food crops more vulnerable to heat stress. Understanding the heat stress-related mechanisms in crops and classifying heat stress-related genes can increase our knowledge in heat-resistant molecular biology and propel developments in molecular design breeding, which can help rice cope with unfavorable temperatures. In this study, we carried out a physiological analysis of rice plants after heat stress. The results show a dramatic increase in malondialdehyde contents and SOD activities. We successfully isolated 11 heat-related rice genes with known function annotation through DNSH, which is an improved SSH method for screening long cDNA fragments. The reanalysis of microarray data from public database revealed that all these genes displayed various expression patterns after heat stress, drought, cold and salt. Quantitative real-time reverse transcription PCR was also performed to validate the expression of these genes after heat stress. The expressions in 10 genes were all significantly changed except for contig 77, which is a CBL-interacting protein kinase. Several reports have been published about the members of the same gene family. [Copyright &y& Elsevier]

Peroxisomes have been proved playing roles in infection of several plant pathogens. Although the contribution of a portion of peroxins in pathogenicity was demonstrated, most of them are undocumented in fungi, especially, Botrytis cinerea . The homologs of Pex8, Pex10, and Pex12 in B. cinerea were functionally characterized in this work using gene disruption strategies. Compared with the wild-type strain (WT), the Δ bcpex8 , Δ bcpex10 , and Δ bcpex12 mutants exhibited significant reduction in melanin production, fatty acid utilization, and decreased tolerance to high osmotic pressure and reactive oxygen species (ROS). The mycelial growth and conidiation of were significantly inhibited in Δ bcpex8 , Δ bcpex10 , and Δ bcpex12 strains. The mycelial growth rates of Δ bcpex8 , Δ bcpex10 , and Δ bcpex12 were reduced by 32, 35, and 34%, respectively, compared with WT and ectopic transformant (ET), and the conidiation was reduced by approximately 89, 27, and 88%, respectively. The conidial germination, germ tube elongation, and the formation of initiate infection structures (IFSs) were also reduced by the deletion of the genes. The pathogenicity was tested on the leaves of tobacco and strawberry, and fruits of tomato. On the leaves of tobacco and strawberry, the Δ bcpex8 , Δ bcpex10 , and Δ bcpex12 mutants could not induce necrotic lesions, and the lesions on tomato fruits infected with the mutants were significantly reduced than those of the wide type. The results indicated that BcPEX8 , BcPEX10 , and BcPEX12 are indispensable for the development and pathogenicity of B . cinerea ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Li, Yu, Guo, Hao, Zhang, Lu, Wang, Zhu, Wang, Chen, Sun and Lin.)

Magnaporthe oryzae , a fungal pathogen that causes rice blast, which is the most destructive disease of rice worldwide, has the potential to perform both asexual and sexual reproduction. MAT loci, consisting of MAT genes, were deemed to determine the mating types of M. oryzae strains. However, investigation was rarely performed on the development and molecular mechanisms of the sexual reproduction of the fungus. In the present work, we analyzed the roles of two MAT loci and five individual MAT genes in the sex determination, sexual development and pathogenicity of M. oryzae . Both of the MAT1-1 and MAT1-2 loci are required for sex determination and the development of sexual structures. MAT1-1-1 , MAT1-1-3 and MAT1-2-1 genes are crucial for the formation of perithecium. MAT1-1-2 impacts the generation of asci and ascospores, while MAT1-2-2 is dispensable for sexual development. A GFP fusion experiment indicated that the protein of MAT1-1-3 is distributed in the nucleus. However, all of the MAT loci or MAT genes are dispensable for vegetative growth, asexual reproduction, pathogenicity and pathogenicity-related developments of the fungus, suggesting that sexual reproduction is regulated relatively independently in the development of the fungus. The data and methods of this work may be helpful to further understand the life cycle and the variation of the fungus.

Peroxisomes are cellular organelles present ubiquitously in eukaryotic cells and are involved in β-oxidation, glyoxylate cycle and a variety of biochemical metabolisms. Recently peroxisomes have been demonstrated to play vital roles in the host infection processes by plant fungal pathogens. The biogenesis of peroxisomes requires a category of proteins named peroxins, which are encoded by the PEX genes. So far, more than 10 PEX genes were isolated in phytopathogenic fungi, and significant research efforts are focused on the mechanism of peroxisome formation and the roles of peroxisome in the development and pathogenicity of fungal pathogens. In this review, we summarize the latest advances in peroxisome biogenesis and functions in pathogenic fungi, including the roles of PEXs in life cycle of peroxisome, peroxisome related metabolisms, and fungal development, infection and pathogenicity, in order to provide references for future studies in plant pathogenic fungi and the control of disease.

This present study deals with synthesis, characterization and antibacterial activity of cross-linked chitosan-glutaraldehyde. Results from this study indicated that cross-linked chitosan-glutaraldehyde markedly inhibited the growth of antibiotic-resistant Burkholderia cepacia complex regardless of bacterial species and incubation time while bacterial growth was unaffected by solid chitosan. Furthermore, high temperature treated cross-linked chitosan-glutaraldehyde showed strong antibacterial activity against the selected strain 0901 although the inhibitory effects varied with different temperatures. In addition, physical-chemical and structural characterization revealed that the cross-linking of chitosan with glutaraldehyde resulted in a rougher surface morphology, a characteristic Fourier transform infrared (FTIR) band at 1559 cm⁻¹, a specific X-ray diffraction peak centered at 2θ = 15°, a lower contents of carbon, hydrogen and nitrogen, and a higher stability of glucose units compared to chitosan based on scanning electron microscopic observation, FTIR spectra, X-ray diffraction pattern, as well as elemental and thermo gravimetric analysis. Overall, this study indicated that cross-linked chitosan-glutaraldehyde is promising to be developed as a new antibacterial drug.

Aim: To clarify whether -238G/A polymorphism of tumor necrosis factor-alpha (TNF-alpha) gene promoter region was associated with outcomes of hepatitis B virus (HBV) infection in Han population of northern China, and to analyze the gene-environment interaction between -238G/A polymorphism and cigarette smoking or alcohol consumption., Methods: A case-control study was conducted to analyze the association of TNF-alpha gene promoter polymorphism with HBV infection outcomes. A total of 207 patients with chronic hepatitis B (HB) and 148 cases of self-limited HBV infection from Ditan Hospital and Shunyi District Hospital in Beijing, respectively were recruited. History of smoking and alcohol drinking was inquired by a questionnaire. The -238G/A polymorphism of TNF-alpha gene promoter was genotyped by polymerase chain reaction-restricted fragment length polymorphism (PCR-RFLP)., Results: The frequencies of GG and GA genotypes were 98.07% and 1.93% in chronic HB patients and 93.24% and 6.76% in self-limited HBV infection individuals, respectively (chi(2)=5.30, P=0.02). The frequency of G allele was significantly higher in patients with chronic HB that in individuals with self-limited HBV infection (99.03% vs 96.62%, chi(2)=5.20, P=0.02). Only modestly increased risk of onset of chronic HB was found in smokers (OR=1.40, 95% CI: 0.87-2.28, P=0.14) and drinkers (OR=1.26, 95%CI: 0.78-2.05, P=0.32). There was a positive interaction between genotype GG and cigarette smoking with an interaction index (II) of 2.95, or alcohol consumption with an II of 1.64., Conclusion: The -238G/A polymorphism of TNF-alpha gene promoter region is independently associated with different outcomes of HBV infection.