Youbo Su

Mould contamination during cigar tobacco fermentation can cause spoilage and aflatoxin accumulation. This study isolated culturable fungi associated with mouldy cigar tobacco and evaluated Bacillus amyloliquefaciens JDF-1 as a candidate biocontrol strain under controlled fermentation conditions. A total of 215 fungal isolates were obtained using six culture media and identified by colony morphology and ITS sequencing. Antagonistic bacteria isolated from healthy tobacco leaves were screened by dual-culture assays and identified using physiological-biochemical characteristics and 16 S rRNA gene sequencing. Candidate compounds tentatively assigned from the cell-free JDF-1 fermentation supernatant were evaluated in a comparative antifungal screening assay against Aspergillus flavus. Four treatments—sterile-water control (CK), JDF-1 inoculation (TA), A. flavus inoculation (TH), and co-inoculation with JDF-1 and A. flavus (THA)—were compared after 35 days of simulated fermentation. (A) flavus was the predominant culturable fungal species, accounting for 48.84% (105/215) of all isolates. JDF-1 produced the largest inhibition zone (9.5 mm) among the screened bacterial isolates and was preliminarily identified as (B) amyloliquefaciens. Among the tested candidate compounds, cyclo(Leu-Pro) showed the lowest EC₅₀ against A. flavus (101.66 ± 6.55 µg/mL), but its contribution to the activity of the native supernatant was not established. The TH treatment showed pronounced mould development and the highest concentrations of aflatoxins B1, B2, G1, and G2. Co-inoculation with JDF-1 substantially reduced visible mould development and aflatoxin accumulation relative to TH, whereas TA showed a mould incidence of 0.71% and no detectable aflatoxins. JDF-1-containing treatments were also associated with changes in major tobacco chemical constituents and metabolite profiles. Untargeted metabolomics and KEGG enrichment identified treatment-associated LC–MS features and pathway annotations but did not establish pathway regulation, metabolic flux, or causal mechanisms. These findings support JDF-1 as a candidate biocontrol strain for reducing A. flavus contamination and aflatoxin risk during cigar tobacco fermentation under controlled conditions. Further targeted metabolite validation, fungal-biomass quantification, and pilot-scale evaluation are required.

Arbuscular Mycorrhizal Fungi (AMF) are key species in plant-microbe interactions, and this study is the first to discover their dynamic survival in the fermentation system of cigar tobacco. To explore the functional significance of AMF in cigar tobacco fermentation, this study focused on the Yunxue variety of cigar tobacco. We combined multi-time point sampling over a 35-day fermentation process and used Internal Transcribed Spacer (ITS) gene high-throughput sequencing to analyze the AMF community structure. Diversity indices, species correlation networks, and Mantel tests were employed to explore the relationship between AMF and chemical components. The study revealed a significant dynamic succession within the arbuscular mycorrhizal fungi (AMF) community throughout the fermentation process, identifying 22 species (comprising 524 operational taxonomic units [OTUs]), with Paraglomus being the predominant species. Core functional flora included OTU217 and OTU88, whose abundance variations aligned with the generation of volatile flavor compounds. AMF diversity peaked during the mid-fermentation stage and exhibited a negative correlation with total nitrogen (TN), total sulfur (TS), and reducing sugars (RS), indicating that sugar and nitrogen metabolism were driving factors in the reorganization of the AMF community. Notably, Glomus-group-B-Glomus-lamellosu-VTX00193 demonstrated a marked increase in abundance towards the end of fermentation, suggesting its crucial role in the degradation of complex organic compounds. Analysis specific to different tobacco varieties revealed a significant increase in the number of OTUs unique to Yunxue 6, with fluctuations in total acidity (TA) content significantly associated with changes in AMF abundance. The findings highlight the regulatory role of AMF in modulating the chemical composition of tobacco leaves through carbon and nitrogen metabolism, with Paraglomus and Glomusidentified as core functional flora. These results offer a foundational framework for targeted manipulation of AMF communities and the design of innovative fermentation processes.

Quinoa (Chenopodium quinoa) is an Andean allotetraploid pseudocereal crop with higher protein content and balanced amino acid composition in the seeds. Ammonium (NH₄⁺), a direct source of organic nitrogen assimilation, mainly transported by specific transmembrane ammonium transporters (AMTs), plays important roles in the development, yield, and quality of crops. Many AMTs and their functions have been identified in major crops; however, no systematic analyses of AMTs and their regulatory networks, which is important to increase the yield and protein accumulation in the seeds of quinoa, have been performed to date. In this study, the CqAMTs were identified, followed by the quantification of the gene expression, while the regulatory networks were predicted based on weighted gene co-expression network analysis (WGCNA), with the putative transcriptional factors (TFs) having binding sites on the promoters of CqAMTs, nitrate transporters (CqNRTs), and glutamine-synthases (CqGSs), as well as the putative TF expression being correlated with the phenotypes and activities of GSs, glutamate synthase (GOGAT), nitrite reductase (NiR), and nitrate reductase (NR) of quinoa roots. The results showed a total of 12 members of the CqAMT family with varying expressions in different organs and in the same organs at different developmental stages. Complementation expression analyses in the triple mep1/2/3 mutant of yeast showed that except for CqAMT2.2b, 11/12 CqAMTs restored the uptake of NH₄⁺ in the host yeast. CqAMT1.2a was found to mainly locate on the cell membrane, while TFs (e.g., CqNLPs, CqG2Ls, B3 TFs, CqbHLHs, CqZFs, CqMYBs, CqNF-YA/YB/YC, CqNACs, and CqWRKY) were predicted to be predominantly involved in the regulation, transportation, and assimilation of nitrogen. These results provide the functions of CqAMTs and their possible regulatory networks, which will lead to improved nitrogen use efficiency (NUE) in quinoa as well as other major crops.

Introduction: The air-curing process of cigar tobacco, as a key step in enhancing the quality of cigars, is often susceptible to contamination by mold spores, which severely constrains the quality of cigar tobacco.Methods: This study employed high-throughput Illumina sequencing technology and a continuous flow analysis system to analyze the differences between the microbial communities and physicochemical components of moldy and healthy cigar tobacco leaves. Furthermore, correlation analysis was performed to reveal the impact of mold on the quality of cigar tobacco.Results: The differences between the microbial flora and physicochemical compositions of moldy (MC) and healthy (HC) tobacco leaves were analyzed, revealing significant disparities between the two groups. Aspergillus spp. represented the dominant mold in MC, with nine out of twelve isolated molds showing higher quantities on MC than on HC. Mold contamination notably decreased the total nitrogen (TN), total phosphorus (TP), total alkaloids (TA), starch, protein, and flavor constituents while increasing the total fatty acid esters (TFAA), which was accompanied by a shift towards weakly acidic pH in the leaves. Fungal community analysis indicated a significant reduction in the fungal operational taxonomic unit (OUT) numbers and diversity indices in MC, contrasting with the bacterial trends. Aspergillus exhibited significantly higher relative abundance in MC, with LEfSe analysis pinpointing it as the primary driver of differentiation. Furthermore, significant negative correlations were observed between Aspergillus and TP, starch, TA, and protein, while a significant positive association was evident with TFAA. Network analysis underscored the pivotal role of Aspergillus as the species influencing disparities between HC and MC, with its abundance serving as a critical determinant during the air-curing process.Discussion: This study elucidated substantial quality distinctions between MC and HC during air-curing, with Aspergillus emerging as the key species contributing to leaf mold.

Wood vinegar, a by-product of charcoal biomass pyrolysis, has been used as a biofungicide in plant disease management because of its antimicrobial properties. However, the physiological and biochemical mechanisms through which wood vinegar alleviates biotic stress are poorly understood. In this study, pot experiments were conducted to investigate the resistance and regulation mechanism of wood vinegar prepared from different raw materials (ZM) and from a single raw material (SM) in controlling tomato (Solanum lycopersicum “Bonny Best”) Fusarium wilt at different concentrations (0.3%, 0.6%, 0.9%, 1.2%, and 1.5%). The results showed that ZM and SM had significant control effects on tomato fusarium wilt under different concentrations in the same growth cycle. Under biotic stress, the two kinds of wood vinegar significantly increased the plant height, stem diameter, leaf area and yield of tomato under the concentration of 0.3%, 0.6%, 0.9% and 1.2%, and significantly reduced the content of malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) in tomato leaves. The effect of 0.9% treatment was the most significant, ZM and SM significantly increased tomato yield by 122% and 74%, respectively, compared with CK under 0.9% treatment. However, the plant height, stem diameter and leaf area of tomato were significantly reduced under 1.5% treatment, but the content of soluble sugar, soluble protein and vitamin C in tomato fruit was the best. Compared with CK, ZM significantly increased by 14%, 193% and 67%, respectively, and SM significantly increased by 28%, 300% and 159%, respectively. Except for 0.3% treatment, both significantly increased the activities of catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) in tomato leaves. The response intensity of two kinds of wood vinegar—physiological and biochemical—to tomato disease resistance, growth and development, showed ZM > SM. The disease index of tomato showed highly significant negative correlation with plant height, stem thickness, leaf area and antioxidant physiology CAT, and highly significant positive correlation with MDA and H₂O₂ content. In conclusion, ZM was more effective than SM in enhancing tomato disease resistance by promoting tomato growth and development, decreasing leaf MDA and H₂O₂ content, and inducing antioxidant enzyme activity in leaves at moderate concentrations.