Ralstonia solanacearum, a soil-borne pathogen with global distribution, infects over 310 plant species including economically important crops such as tomato, eggplant, and pepper, causing bacterial wilt and severe agricultural losses. Scarecrow-like protein 18 (SCL18), a member of the GRAS transcription factor family, primarily regulates plant development and signaling. This study explores the molecular mechanism by which NtSCL18 modulates bacterial wilt resistance in tobacco. CRISPR/Cas9-mediated knockout of NtSCL18 in tobacco cultivar K326 was conducted, with disease resistance evaluated through disease index analysis, lignin content determination, and ROS staining. Transcriptome profiling was performed 24 h post-inoculation with R. solanacearum. Results revealed that NtSCL18 knockout mutants significantly enhanced resistance to R. solanacearum56 and elevated lignin accumulation. Transcriptome analysis identified 833, 1756, and 1693 differentially expressed genes in the three mutants, respectively. The resistance mechanism involved affection of the Pattern-Triggered Immunity and MAPK signaling cascade, synergistic modulation of ethylene and calcium signaling pathways, and peroxidase-mediated lignin monomer synthesis to reinforce cell walls. In conclusion, NtSCL18 acts as a negative immune regulator; its knockout enhances resistance by coordinately affecting these pathways providing a novel target and theoretical foundation for breeding bacterial wilt-resistant tobacco. This demonstrates the potential of NtSCL18 knockout as a disease control strategy.

Meloidogyne incognita (M. incognita) is a highly destructive species of Meloidogyne spp., characterized by its ability to cause root-knot nematode (RKN) disease, which is difficult to control and severely inhibits plant growth. Temperature is one of the primary factors affecting M. incognita infection. However, the precise underlying mechanisms have not yet been clarified. The present study aims is to further explore the temperature-influenced resistance mechanisms to M. incognita. Antioxidant enzyme activities, osmotic regulation substance contents, tissue structure changes, and expression of the resistance gene (Rk) in the roots of two tobacco varieties were analyzed under three temperatures (15°C, 25°C, and 35°C) via artificial inoculation. A M. incognita-resistant variety (NC95) and a susceptible variety (CBH) was selected as experimental materials. The results showed that the activities of peroxidase (POD) and catalase (CAT), as well as the contents of soluble sugar, proline, and hydroxyproline-rich glycoprotein (HRGP), increased to varying degrees under M. incognita infection, while superoxide dismutase (SOD) activity decreased. Notably, the activities of POD and CAT, along with the contents of soluble sugar, proline, and HRGP, were all higher in NC95 than in CBH. Meanwhile, antioxidant enzyme activities and osmotic substance contents in both varieties varied most at 25°C and least at 35°C. No giant cells or oocysts were observed in the root tissues of NC95 at any temperature, whereas numerous giant cells and oocysts were present in CBH. The number of giant cells in CBH was highest at 25°C compared to 15°C and 35°C, and the degree of lignification in NC95 was also greater at 25°C. In addition, M. incognita infection induced the expression of Rk gene in NC95, with expression levels at 25°C and 15°C higher than at 35°C. The results indicated that SOD activity and osmotic regulatory substance contents decreased in the roots of the susceptible variety under M. incognita infection, accompanied by the appearance of numerous giant cells in the xylem, contributing to susceptibility. Conversely, the resistant tobacco variety exhibited stronger capabilities in reactive oxygen species (ROS) scavenging and osmotic regulation, no significant changes in root tissue structure, and upregulated expression of the Rk gene, all of which contributed to infection inhibition. Compared with the observations at 25°C, M. incognita infectivity on tobacco roots was effectively reduced by 35°C due to increased antioxidant enzyme activities, enhanced osmotic regulatory substance contents, and well-maintained root tissue structure. Additionally, Rk gene expression was not inactivated but only reduced at 35 °C, and it remained effective in inhibiting M. incognita infection.

Leaf senescence is a vital aspect of plant physiology and stress responses and is induced by endogenous factors and environmental cues. The plant-specific NAC (NAM, ATAF1/2, CUC2) transcription factor family influences growth, development, and stress responses in Arabidopsis (Arabidopsis thaliana) and other species. However, the roles of NACs in tobacco (Nicotiana tabacum) leaf senescence are still unclear. Here, we report that NtNAC56 regulates leaf senescence in tobacco. Transgenic plants overexpressing NtNAC56 (NtNAC56-OE) showed induction of senescence-related genes and exhibited early senescence and lower chlorophyll content compared to wild-type (WT) plants and the Ntnac56-19 mutant. In addition, root development and seed germination were inhibited in the NtNAC56-OE lines. Transmission electron microscopy observations accompanied by physiological and biochemical assays revealed that NtNAC56 overexpression triggers chloroplast degradation and reactive oxygen species accumulation in tobacco leaves. Transcriptome analysis demonstrated that NtNAC56 activates leaf senescence-related genes and jasmonic acid (JA) biosynthesis pathway genes. In addition, the JA content of NtNAC56-OE plants was higher than in WT plants, and JA treatment induced NtNAC56 expression. We performed DNA affinity purification sequencing to identify direct targets of NtNAC56, among which we focused on LIPOXYGENASE 5 (NtLOX5), a key gene in JA biosynthesis. A dual-luciferase reporter assay and a yeast one-hybrid assay confirmed that NtNAC56 directly binds to the TTTCTT motif in the NtLOX5 promoter. Our results reveal a mechanism whereby NtNAC56 regulates JA-induced leaf senescence in tobacco and provide a strategy for genetically manipulating leaf senescence and plant growth.

Aspartic proteases (APs) constitute a large family in plants and are widely involved in diverse biological processes, like chloroplast metabolism, biotic and abiotic stress responses, and reproductive development. In this study, we focused on overall analysis of the APs genes in tobacco. Our analysis included the phylogeny and cis-elements in the cell wall-associated promoters of these genes. To characterize the expression patterns of APs genes in stem vascular development. The tissue expression analysis showed that NtAED3-like was preferentially expressed in the differentiating xylem and phloem cells of the vascular system. Based on histochemical staining analysis showed that the NtAED3-like gene was specifically expressed in stem vascular tissue, root vascular tissue, and petiole vascular tissue. The TdT-mediated dUTP nick-end labeling (TUNEL) assay illustrated a delayed progression of programmed cell death (PCD) within the xylem of the ko-ntaed3a-like mutant, relative to the wild type. The mutant ko-ntaed3a-like exhibited a phenotype of thinning stem circumference and changed in xylem structure and lignin content. In addition, the two-dimension heteronuclear single quantum coherent nuclear magnetic resonance (2D-HSQC) analysis of three milled wood lignins (MWLs) showed that the content of β-O-4 connection in ko-ntaed3a-like decreased slightly compared with wild type. In conclusion, this study provides our understanding of the regulation of vascular tissue development by the NtAED3-like gene in tobacco and provides a better basis for determining the molecular mechanism of the aspartic protease in secondary cell wall (SCW) development.

综述了烤烟碳氮代谢途径和关键酶的研究进展,研究了移栽期、光温、施肥、外源小分子有机物对碳氮代谢及烟叶品质的影响,分析了碳氮代谢与烟草抗逆性的关系,并展望了碳氮代谢的重点研究方向.阐明了碳氮代谢协调平衡分子机理机制、筛选碳氮代谢关键基因以及应用碳氮代谢及抗逆性的关系,以期推动碳高效、氮高效的高光合效能、高品质的烤烟新品种选育,达到降本增效,实现烤烟生产可持续发展.

比较不同抗性烟草品种在赤星病胁迫下的光合生理响应差异可揭示赤星病和光合作用之间的关系,有助于进一步明确烟草赤星病的抗性机理。本实验采用盆栽方法,分析赤星病胁迫对2个烟草抗、感病品种JYH和CBH的光合作用、叶绿素荧光特性、光合色素、光合碳同化关键酶以及叶黄素循环的影响。结果显示,在5 d时,赤星病胁迫对JYH的一系列光合生理活动均未产生不利影响。赤星病胁迫明显抑制9d时的JYH和5～9 d时的CBH的叶绿素荧光参数、光合色素含量、光合碳同化关键酶活性及叶黄素循环功能,最终表现为光合作用下降。JYH的光合作用降低是由气孔限制因素引起,而非气孔限制因素是CBH的光合作用被抑制的主要原因。与CBH相比, JYH具有较高的PSII最大光化学效率（Fv/Fm）、PSII实际光化学效率（ΦPSII）、光化学淬灭系数（qp）、非光化学淬灭系数（NPQ）和叶绿素a、叶绿素a+b、类胡萝卜素含量以及核酮糖-1,5-二磷酸羧化/加氧酶（Rubisco）、景天庚酮糖-1,7-二磷酸酯酶（SBPase）、果糖-1,6-二磷酸酯酶（FBPase）活性,且其花药黄质（A）、玉米黄质（Z）、叶黄素循环库（A+V+Z）含量与叶黄素循环脱环氧化状态（A+Z）/（A+V+Z）也显著大于CBH。研究结果表明,赤星病胁迫对抗病品种光合生理活动的危害程度小于感病品种,有利于保持较高的光合作用强度,从而具有较强的抗病性。

Nutritional correlations between plants and pathogens can crucially affect disease severity. As an essential macronutrient, the availability of nitrogen (N) and the types of N content play a fundamental part not only in energy metabolism and protein synthesis but also in pathogenesis. However, a direct connection has not yet been established between differences in the level of resistance and N metabolism. Pertinently, former studies hold ammonia (NH3) accountable for the development of diseases in tobacco (Nicotiana tabacum L.) and in some post-harvest fruits. With a purpose of pinpointing the function of NH3 volatilization on Alternaria alternata (Fries) Keissl pathogenesis and its correlation with both N metabolism and resistance differences to Alternaria alternata infection in tobacco, leaf tissue of two tobacco cultivars with susceptibility (Changbohuang; CBH), or resistance (Jingyehuang; JYH) were analyzed apropos of ammonia compensation point, apoplastic NH4+ concentration, pH value as well as activities of key enzymes and N status. At the leaf age of 40 to 60 d, the susceptible cultivar had a significantly higher foliar apoplastic ammonium (NH4+) concentration, pH value and NH3 volatilization potential compared to the resistant one accompanied by a significant reduction in glutamine synthetase (GS), which in particular was a primary factor causing the NH3 volatilization. The NH4+ concentration in CBH was 1.44 times higher than that in JYH, and CBH had NH3 compensation points that were 7.09, 6.15 and 4.35-fold higher than those of JYH at 40, 50 and 60 d, respectively. Moreover, the glutamate dehydrogenase (GDH) activity had an upward tendency related to an increased NH4+ accumulation in both leaf tissues and apoplast but not with the NH3 compensation point. Collectively, our results strongly suggest that the accumulation of NH3 volatilization, rather than NH4+ and total N, was the primary factor inducing the Alternaria alternata infection in tobacco. Meanwhile, the susceptible cultivar was characterized by a higher N re-transfer ability of NH3 volatilization, in contrast to the disease–resistant cultivar, and had a stronger capability of N assimilation and reutilization. This study provides a deeper understanding of the pathogenicity mechanism induced by Alternaria alternata, which is useful for breeding Alternaria alternata-resistant varieties of tobacco, at the same time, our research is also conducive to control tobacco brown spot caused by Alternaria alternata in the field.

为进一步明确烟株内源激素与赤星病抗性的关系,以抗赤星病烟草品种净叶黄和感病品种长脖黄为研究对象,通过盆栽种植并接种赤星病菌,比较不同抗性品种内源激素含量对赤星病胁迫的响应差异.研究结果显示,在赤星病胁迫下,净叶黄、长脖黄的脱落酸(ABA)含量和乙烯(ET)释放量均呈上升趋势,而生长素(IAA)以及细胞分裂素(CTK)含量表现为降低.同时,赤星病胁迫导致净叶黄的赤霉素(GA)、茉莉酸(JA)含量增加,引起长脖黄的JA含量降低,GA含量则先增加后降低.与净叶黄相比,长脖黄的ABA含量增幅较小,而GA、IAA、CTK与JA含量降幅较大,并且ET释放量较高.综上,在赤星病胁迫下,内源激素的含量差异以及变化幅度可以反映不同品种的抗病能力.

为烤烟新品系在黄平及类似烟区生产上的推广应用提供科学依据,采用田间试验方法,以当地主栽品种云烟116为对照,分析比较烤烟新品系6036和6028在黄平烟区的生育期、农艺性状、田间抗病性、经济性状、烤后烟叶外观质量和化学成分含量的差异.结果 表明:田间抗病性6028最好,6036其次,云烟116最差;6036和6028的生育期与云烟116相当,农艺性状表现较云烟116略差,但田间长势强、整齐度好,易烘烤;烟叶化学成分的协调性优于云烟116,经济性状表现6036最好,云烟116其次,6028略差.6036可在黄平烟区推广种植;6028田间抗病性强,可在黄平烟区易发生根茎性病害区域作为优质烤烟补充品系推广种植.

To screen new tobacco material to meet the demand of cigarette industry,the optimum planting density and fertiliz-ing amount of new tobacco material Y28,a hybrid of sun-cured and flue-cured tobacco,was studied in the present study. The results showed that,for agronomic traits of Y28 seedlings,the best density is 1300 plants combined with 5 kg nitrogen per 666.7 m2. When 1400 plants were planted and 5 kg nitrogen were applied per 666.7 m2,the appearance characteristic and chemical composition of flue-cured tobacco leaves were better than other treatments with high sugar and potassium content, low-nicotine content,appropriate values for reducing sugar to total sugar ratio,nitrogen/nicotine and potassium/chloride. For flue-cured tobacco leaves sensory quality,two optimum cultivation combinations are 1300 plants with 3 kg nitrogen and 1400 plants with 4 kg nitrogen per 666.7 m2,respectively. Moreover,the heterosis variety of Y28 are with unique characteristics of better smoking aroma quality,enough smoking aroma,less offensive odor, sweet and pure taste, which can meet the increasing demand of cigarette industry for new tobacco raw material.

In order to increase baking quality and usability of upper tobacco leaves, the effects of different temperature contrast between dry-bulb and wet-bulb in yellowing stage on chemical composition, aroma components and economic characters of upper leaves under one-time harvest were studied, and the method of combining field test with laboratory test was adopted.Results: When the dry-bulb temperature was 38℃ in yellowing stage, compared with other treatments, the temperature contrast of 1.5℃ between dry-bulb and wet-bulb was most suitable, it could enhance the coordination of chemical components and aroma substances contents in upper leaves, improve the appearance quality sensory evaluation quality, and the economic characters also increased, which was helpful to improve the yield and quality of upper tobacco leaves.

The contamination constants of soil,irrigation,rainfall and specific base manure to heavy metal content in flue-cured tobacco were analyzed by using the artificial simulation experiment method and mathematical statistics,and then the heavy metal source and content in flue-cured tobacco were analyzed and predicted by using contamination constants as basic parameters to provide the scientific basis for rational regulation of heavy metal content in flue-cured tobacco.Results:There are obvious differences in Cd,Hg,Pb and Cr contamination constant among four factors.The contribution ratio of heavy metal content in flue-cured tobacco is rainfall ＞ irrigation ＞ soil ＞ specific base manure under the same heavy metal concentration.The heavy metals in flue-cured tobacco are from soil and specific base manure in Guizhou tobacco-growing area mainly.66.82％ Cd,63.98％ As,92.90％ Pb and 96.91％ Cr of flue-cured tobacco are from soil and 33.01％ Cd,35.76 ％ As,7.05 ％ Pb and 3.06 ％ Cr of flue-cured tobacco are from specific base manure.99.64％ Hg of flue-cured tobacco is from specific base manure.The mean deviation ratio of predicted value of different elements is about 30 ％ by the established prediction model of heavy metal content in flue-cured tobacco and the prediction accuracy rate of the prediction model reaches above 60％,which indicates the prediction model can be basically used in rough estimation of heavy metal content in flue-cured tobacco.

To clarify the influences of cement production on heavy metal contents in tobacco leaves and surrounding environment,two cement plants located in Huanghuai Plain and Southwest mountainous area were studied. The variations of deposition fluxes of heavy metals in the surrounding atmosphere and the heavy metal contents in surrounding farmland soil and tobacco leaves along with the distance from the cement plants were analyzed. The results showed that the effect of cement production on heavy metal contents in surrounding farmland soil was relatively light. The deposition fluxes of heavy metals in the atmosphere raised at 1.5-3.5 km and 1.0 km away from the cement plants in Huanghuai Plain and Southwest mountainous area,respectively. The maximum/minimum ratios of Pb,Cd,As,Cr and Ni deposition fluxes in the atmosphere were in the range of 1.7-2.8. Comparing with the upper tobacco leaves grown in the upwind direction,the highest contents of Pb,Cd, As,Cr and Ni in upper tobacco leaves in the downwind direction of the cement plant in Huanghuai Plain were 2.26,3.09,2.44,3.72 and 1.78 times,respectively; and the peaks of heavy metal contents in upper tobacco leaves were at 1.0-4.0 km downwind of the cement plant. However,in places surrounding the cement plant in Southwest area,the maximum contents of Pb,Cd,As,Cr and Ni were 1.34,1.19,2.32,3.41 and 4.66 times of the minimum in upper leaves,respectively; and the peaks were at 1.0-1.5 km away from the cement plant. In conclusion,tobacco is a sensitive indicator of atmospheric environment,and the influences of cement plant on plain at monsoon region are larger than those on mountainous area.

Topography influences the density of soil sampling points for precise fertilization. In the case of high phosphorus and potassium fertilization for flue-cured tobacco, the soils in areas of plain, broad valley, narrow valley and gentle slope were sampled by 50 m×50 m grids and the available nutrients in soil samples were tested. The results showed that by Cochran formula, individual sample for plain, valley and gentle slope represented an area of 1.92, 1.01-1.56 and 0.73 ha, respectively at a confidence level of 90% and a relative error of 15%; by geostatistical method, the maximum sampling space for precise nutrient management were 234.0, 134.4-177.5 and 58.2 m for plain, valley and gentle slope, respectively. It indicated that sampling density should be increased gradually from flat to slope.

Tobacco brown spot caused by Alternaria alternata is a devastating disease of tobacco worldwide. Phenotypic characterization of the pathogen was investigated to provide some basic information for biology and pathology by using BIOLOG Phenotype Microarray (PM). Using PM plates 1-10, 950 different growth conditions were tested. Results exhibited that the pathogen was able to metabolize 24.74% of tested carbon sources, 85.26% of nitrogen sources, 97.14% of sulfur sources and 89.83% of phosphorus sources. Most informative utilization patterns for carbon sources of A. alternata were carbohydrates and for nitrogen were various amino acids. The pathogen presented 274 different nitrogen pathways. It had wide range adaptabilities in osmolytes with up to 10% sodium chloride, up to 6% potassium chloride, up to 5% sodium sulfate, up to 20% ethylene glycol, up to 6% sodium formate, up to 6% urea, up to 12% sodium lactate, up to 200 mM sodium phosphate, up to 100 mM ammonium sulfate, up to 100 mM sodium nitrate and up to 20 mM sodium nitrite. It also exhibited active metabolism in the range of pH values between 3.5 and 10, with optimal pH of around 6.0. The pathogen showed active decarboxylase activity, whereas no deaminase activity in the presence of various amino acids.

To explore the validity changes of heavy metal cadmium in the planting tobacco's soil and the accumulation and distribution characteristics of Cd in plant organs,a pot experiment was conducted with lime soil treated with different concentrations of cadmium.The results showed that the performance of Cd content in tobacco organs were lower leaf> middle leaf> upper leaf> stem,root;With the growth and promotion,cadmium content of lower leaves increased first,then decreased,but a decline of 50%~80%Cd in other organs reduce in different level.With the increasing of cadmium concentration in soil,the cadmium content increased significantly in different organs of tobacco,and cadmium accumulation also increased(p<0.05).More than 90% of the Cd absorbed by tobacco was saved in the leaves.Accumulation of cadmium accounted for about 95% of the total during reproductive stage.When exogenous cadmium in soil,its effectiveness decreased by about 30%,but still a lot of absorption of tobacco plants.Therefore, exogenous heavy metal was strictly forbidden getting into soil-tobacco system in the process of tobacco production.

Stem rot in flue-cured tobacco (Nicotiana tabacum L.) was observed in tobacco seedlings in 2012. Its pathogen was identified as Fusarium kyushuense according to morphology, pathogenicity and rDNA-ITS and EF-1αgene analysis. The temperature for the pathogen to grow on LBA ranged from 5℃to 42℃with an optimum temperature of 30℃, ranged from 10℃to 35℃with optimum temperature of 35℃;and ranged from 5℃to 42℃with optimum temperature of 30℃to 35℃for conidial germination. The suitable relative humidity for conidial germination of pathogen ranged from 70%to 100%. The pathogen could grow under pH 2-11 with optimum pH 5-7. Light had no effect on mycelial growth and conidial germination. Light and dark alternation reduced mycelial growth, but enhanced sporulation. Among the 95 carbon resources in Biolog FF Microplate, 89 ones could be utilized by F. kyushuense. Carbon resources which could not be used by pathogen include N-acetyl-D-galactosamine, N-acetyl-?-D-glucosamine, glucuronamide, sebacic acid, L-serine and adenosine-5’-monophosphate.