Scientia Agricultura Sinica

Function of FCS-Like Zinc-Finger Protein OsFLZ18 in Regulating Rice Flowering Time

MA YaMei,ZHANG ShaoHong,ZHAO JunLiang,LIU Bin

Scientia Agricultura Sinica. 2022, 55(20): 3875-3884. doi:10.3864/j.issn.0578-1752.2022.20.001

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【Objective】Flowering time is an important agronomic trait which determines the yield and regional adaptability of rice, but the underlining molecular regulatory mechanism need further study. FCS-like Zinc finger proteins (FLZs) are a class of plant specific regulatory proteins which play essential roles in plant growth and stress response, but their functions in regulating flowering time have not been reported. This study aims to investigate the potential function of FLZ proteins in rice flowering time control. The finding will broaden our understanding on the molecular regulatory mechanism of rice flowering time.and provide new theoretical basis and gene resource for rice breeding. 【Method】Based on the target sequences published in RGAP database, OsFLZ18 overexpression vector and CRISPR-Cas9 vector were generated and introduced into Japonica variety Nipponbare by Agrobacterium tumefaciens-mediated genetic transformation assay. Homozygous CRISPR knockout mutants were screened by PCR and sequencing analyses. The quantitative real-time PCR (qRT-PCR) assay was used to examine the spatial-temporal expression and diurnal rhythmic expression of OsFLZ18, as well as the effects of OsFLZ18 on the transcription of several known flowering time-related genes. Yeast two-hybrid assay (Y2H) was used to test the interaction between OsFLZ18 and the flowering time-related regulatory proteins.【Result】OsFLZ18 was ubiquitously expressed in various rice tissues, with the highest expression level in 14 day-old seedling, followed by leaf sheaths and leaf blades at the tillering stage, and stem and young panicles at reproductive stages. The OsFLZ18-CRISPR vector was constructed and transformed into Nipponbare. Two independent homozygous OE lines (OE-2, OE-3) with higher OsFLZ18 expression level and two homozygous mutants (CRISPR-21, CRISPR-25) were selected for further study. Phenotypic observation showed that the OE lines flowered later than the wild-type plants under both natural long-day and short-day conditions in Guangzhou, while the CRISPR lines had no obvious differences in heading date when compared to the wild-type plants. The expression levels of Ehd1, Hd3a and RFT1 were significantly decreased in OE-2 plants compared with those in the wild-type plants under artificial short-day conditions, but no significant difference in the expression level of Hd1 was observed between them. The results of Y2H experiment showed that OsFLZ18 interacted with OsMADS51, a positive regulator of rice flowering time. Furthermore, OsFLZ18 exhibits a diurnal rhythmic expression profile, showing lower expression levels in the daytime and higher expression levels at night with a peak at midnight. 【Conclusion】Overexpression of OsFLZ18 delays rice flowering time.

Identification of the Root-Specific Soybean GmPR1-9 Promoter and Application in Phytophthora Root-Rot Resistance

YAN Qiang,XUE Dong,HU YaQun,ZHOU YanYan,WEI YaWen,YUAN XingXing,CHEN Xin

Scientia Agricultura Sinica. 2022, 55(20): 3885-3896. doi:10.3864/j.issn.0578-1752.2022.20.002

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【Objective】The objective of this study is to identify the root-specific promotors and the core regulatory sequence of soybean. Then evaluate the potential application of the synthetic promoter in Phytophthora root-rot resistance. 【Method】The genes which specifically expressed in roots with high expression levels were screened based on the transcriptome date of soybean root, stem and leaf tissues in the seedling stage. Based on the distribution of the cis elements, the promoter truncation approach was used to map the minimal promoter controlling root specific expression in soybean hairy roots. The obtained minimal promoter fragment was concatenated with the Phytophthora inducible promoter elements p4XD to construct the synthetic promoter. The synthetic promoter driven over-expression of Phytophthora resistance related gene GmNDR1 in soybean hairy roots, then the resistance level of transgenic tissue to Phytophthora and the expression profiles of GmNDR1 during the interaction had been analyzed. Furthermore, the transgenic Nicotiana benthamiana plants were generated to evaluate the resistance at plant level. 【Result】Though screening, six soybean PR1 homologues with significant root specific expression manner were identified, and GmPR1-9 had the highest promoter activity. Numbers of root specific expression related cis elements were identified in promoter sequence using the online tool PLACE. Truncation analysis of the promoter showed that serial 5’ end deletions L1, L2, L3, L4 and L5 had different GUS activities. The L5 (-166 to -1) fragment had 80% activity of the full-length promoter, and was able to drive GUS expression in roots of transgenic N. benthamiana. GUS enzyme activity was almost undetectable in three 3’ end deletions R1, R2 and R3, and the double terminal deletion mutant M1. When the fusion promoter p4XD-L5 driven GmNDR1 expression in soybean hairy roots, the resistance to P. sojae was significantly enhanced. The disease severity and lesion length were significantly reduced in the over-expression hairy roots when compared with control, and the relative biomass of Phytophthora decreased by 66.5% at 48 h post inoculation. GmNDR1 maintained high expression level in over-expression tissues, with 39.2 times of that in control tissues. The expressions were further up-regulated after inoculation, and reached the highest level at 36 h. In p4XD-L5::NDR1 transgenic N. benthamiana plants, the expression of GmNDR1 was significantly higher in roots than that in stems and leaves. Fifteen days after P. capsica inoculation, the plant height, root length and fresh weight of GmNDR1 over-expression plants were significantly higher, and meanwhile the leaf wilting rate and lesion length were significantly lower. 【Conclusion】This study obtained a soybean root specific promoter and identified the core regulation sequence. The strategy which driven the expression of GmNDR1 by the synthetic promoter p4XD-L5 combined the inducible and tissue-specific promoter core elements can significantly enhance the resistance of transgenic soybean hairy roots and Nicotiana benthamiana plants to Phytophthora pathogens.

Analysis of Cross Compatibility Variation Among Diverse Sesamum Species and Biological Characteristics of the Interspecific Hybrid Progenies

JU Ming, MIAO HongMei, HUANG YingYing, MA Qin, WANG HuiLi, WANG CuiYing, DUAN YingHui, HAN XiuHua, ZHANG HaiYang

Scientia Agricultura Sinica. 2022, 55(20): 3897-3909. doi:10.3864/j.issn.0578-1752.2022.20.003

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【Objective】 The research aims to explore the cross compatibility between different Sesamum species and analyze the biological characteristics of interspecies hybrid progeny so as to supply the foundation for efficient application of wild sesame species. 【Method】 A sesame cultivar Yuzhi 11 (S. indicum, 2n=26) and 4 wild Sesamum species including S. latifolium (2n=32), S. calycinum (2n=32), S. angustifolium (2n=32), and S. radiatum (2n=64) were applied to construct interspecies cross combinations using diallel hybridization method by artificial pollination in the field. Embryo rescue method was also used to obtain interspecific hybrid F₁. Interspecific hybrid compatibility was compared based on hybrid capsule formation rate. Botanical characters of hybrids were observed during flowering and mature stages. Pollen fertility was assessed using Alexander staining method. Chromosome number and karyotype characteristics of root somatic cells of hybrids were observed using smear chromosome preparation technique. Specific and polymorphic SSR primers in Sesamum were used to analyze the molecular difference in interspecific hybrids.【Result】 Twenty positive and reciprocal cross combinations were constructed for the 5 Sesamum species. A total of 2091 flowers were pollinated and 370 hybrid capsules were harvested. As to the female parents with more chromosomes, hybrid capsules were more easily obtained. The cross compatibility among the 5 Sesamum species significantly varied from 1.18% (S. radiatum×S. calycinum) to 63.33% (S. calycinum×S. angustifolium). F₁ plants of 9 combinations produced hybrid seeds, while the ratio of pollen sterility of F₁ progeny ranged from 35.21%-100.00%. The cross S. calycinum×S. angustifolium presented the highest sterility ratio to 87.68%. Hybrid progeny exhibited the obvious heterosis over parents in plant height, plant type, and some key agronomic traits. As to the positive and reciprocal hybrid F₁ derived from sesame cultivar and the wild species, leaf shape, flower shape, and flower color showed partial characters of both parents. The cross compatibility between sesame cultivar (n=13) and the 3 Sesamum species with chromosome group n=16 ranked as S. angustifolium>S. calycinum>S. latifolium. The cross compatibility between wild species S. radiatum (n=32) and the 3 species with n=16 ranked as S. calycinum>S. angustifolium>S. latifolium. Among the 5 Sesamum species, the genetic relationship between S. calycinum and S. angustifolium is relatively closest. The chromosome number of root tip cells of some hybrid plants is consistent with the theoretical value calculated from the parents. Screening results of the 3 pairs of polymorphic SSR primers indicated that 99.66% of obtained F₁ plants are true hybrid. Chromosome karyotype and SSR marker screening results reflected the genetic difference and characters of Sesamum species. 【Conclusion】Among the 5 Sesamum species, the cross compatibility varies significantly and the heterosis of interspecific hybrid is obvious. Of which only S. calycinum and S. angustifolium have the relatively closest genetic relationship and could be directly applied for elite germplasm creation and interspecific hybrid breeding in Sesamum. Reproductive isolation barriers exist in other cross combinations. Some techniques including embryo rescue and molecular marker application should be used to achieve the utilization of wild Sesamum species for sesame breeding.

Effects of Two Mechanical Planting Methods on the Yield and Quality of High-Quality Late Indica Rice

GUO BaoWei,TANG Chuang,WANG Yan,CAI JiaXin,TANG Jian,ZHOU Miao,JING Xiu,ZHANG HongCheng,XU Ke,HU YaJie,XING ZhiPeng,LI GuoHui,CHEN Heng

Scientia Agricultura Sinica. 2022, 55(20): 3910-3925. doi:10.3864/j.issn.0578-1752.2022.20.004

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【Objective】The experiment was conducted to clarify the differences in the yield and quality of double cropping high-quality late indica rice and the selection criteria for variety tapes under mechanical transplanting (MT) and mechanical direct seeding (MS).【Method】Early-maturing late indica rice, medium-maturing late indica rice, moderately late-maturing late indica rice, late-maturing late indica rice and extremely late-maturing late indica rice were used as materials, and two planting methods including MT and MS were set to study the yield and quality characteristics of high-quality late indica rice.【Result】(1) The seed setting rate and the number of spikelets in the population of the high-quality late indica rice under MT were significantly higher than that under MS, and the 1000-grain weight under MT also increased, finally the actual yield was significantly increased. For MT, the medium-maturing late indica rice had the highest yield, followed by moderately late-maturing late indica rice; while for MS, the early-maturing late indica rice had the highest yield, followed by medium-maturing late indica rice. (2) Compared with MS, the processing quality of high-quality late indica rice varieties under MT was better for the significantly higher brown rice rate, milled rice rate and head rice rate, but the appearance quality deteriorated because of the increased chalky rice rate and chalkiness degree. The cooking quality improved for higher gel consistency, breakdown, taste value and lower amylose content. The highest brown rice rate, milled rice rate, and head rice rate were all found in medium-maturing late indica rice under two planting methods, chalky rice rate and chalkiness degree among different varieties showed the trend of extremely late-maturing late indica rice

Improvement of Row Detection Method Before Wheat Canopy Closure Using Multispectral Images of UAV Image

MA Xiao,CHEN PengFei

Scientia Agricultura Sinica. 2022, 55(20): 3926-3938. doi:10.3864/j.issn.0578-1752.2022.20.005

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【Objective】In order to make precise management of wheat, it is of great significance to accurately identify the location of its rows. In this study, the aim was to improve the traditional Hough transform-based methods and the green pixel accumulation- based methods respectively, and then to analyze the different methods before and after the improvement for wheat row detection, so as to provide the technical support for accurate detection of wheat rows.【Method】A wheat water-nitrogen coupling experiment was established. During the early jointing stage, the multispectral images of wheat under different growth conditions was obtained by the RedEdge M sensors mounted on a four rotor unmanned aerial vehicle (UAV). First, based on these data, these images of the excess green minus excess red (ExGR) index were calculated, and then the Otsu method was performed on it to classify image pixels into vegetation and soil. Second, 3×1 line template was used for morphological opening operation to reduce boundary irregularity and remove noise. Third, according to the characteristics of the wheat rows in the image, the modified wheat row extraction methods were proposed based on the optimization of the peak detection process of the traditional Hough transform-based method and the angel detection process of the traditional green pixel accumulation-based method, respectively. Finally, the detection results before and after improving the two methods were compared with the result of visually interpreted wheat rows, and their performances were evaluated by using the detection rate and crop row detection accuracy (CRDA). 【Result】The combination of the ExGR index and OTSU method could accurately identify the vegetation pixels and soil pixels automatically, and the overall accuracy was 93.75% and the Kappa was 0.87. The morphological opening operation could remove the pattern noise and reduce the error of later crop row detection. The modified Hough transform-based method effectively improved the peak detection accuracy by restricting the peak detection range with prior knowledge; Compare with traditional Hough transform-based methods, the averaged detection rate has increased from 30% to 67%, and the averaged CRDA has increased from 0.22 to 0.44. The modified green pixel accumulation-based method investigated the accumulation characteristics of the green pixels of the entire image, which effectively improved the angel detection accuracy; Compare with traditional green pixel accumulation-based method, the averaged detection rate has increased from 14% to 93%, and the average CRDA has increased from 0.12 to 0.69. The identification accuracy of the four methods from high to low were the modified green pixel accumulation-based method, the modified Hough transform-based method, the traditional Hough transform-based method, and the traditional green pixel accumulation-based method. 【Conclusion】This study improved the traditional crop row detection methods, which provided the technical support for the identification of wheat rows with high planting density and small row spacing.

Grading Criterion of Maize Root Rot Based on Aboveground Symptoms and the Relationship Between Severity and Agronomic Traits

LIU ShuSen,SUN Hua,SHI Jie,GUO Ning,MA HongXia,ZHANG HaiJian

Scientia Agricultura Sinica. 2022, 55(20): 3939-3947. doi:10.3864/j.issn.0578-1752.2022.20.006

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【Objective】This study aimed to establish a standardized and easy-to-operate grading criterion for maize root rot severity based on the aboveground symptoms at the seedling stage, and to clarify the effects of different disease severities on maize growth and yield.【Method】The maize was planted with hole-sowing, and the inoculum of Fusarium graminearum was placed between the seeds of two holes for root rot inoculation. After emergence, the symptoms of root rot were observed continuously, and the grading criterion of disease severity was established based on the symptoms of the aboveground parts at the V5 stage. The agronomic traits of plant height, stem base diameter, stem diameter, and ear height of plants with different grades were measured at the V11 stage and dough stage, respectively. At the same time, the incidence of stalk rot, which developed from root rot, was investigated at the dough stage. After harvesting, yield-related traits, including ear diameter, ear length, bare top length, row number per ear, kernel number per row, 100-kernel weight, and kernel weight per ear, were measured. Then, the relationship between root rot severity and yield-related traits was analyzed.【Result】At the V1 stage, the symptoms of yellowing leaf and wilting tip, seminal root and mesocotyl with yellowish-brown lesions, and some mesocotyl with browned vascular tissue were observed. At the V3 stage, the mesocotyl and root system of the diseased plants were necrotic to varying degrees, and the aboveground part of some plants was dead due to the necrotic mesocotyl. At the V5 stage, it can be classified into the following three types, i.e., leaves showed yellowing to varying degrees, or even gradually withered; the plant appearance was a dwarf and developed slowly and stagnantly; the plant died. Furthermore, the severity of root rot was divided into grades 1, 3, 5, 7 and 9 based on the aboveground symptoms at the V5 stage. Investigation of agronomic traits showed that plant height, stem base diameter, stem diameter and ear height decreased gradually with the increase of disease grade of root rot at the V11 stage and dough stage. At the same time, the development of most 9-grade plants was inhibited or stagnated, and showed tillering or deformity. The investigation of stalk rot indicated that some of the root rot plants developed into stalk rot in the late growth stage. The incidence of stalk rot showed a decreasing trend with the increase of root rot grade. The analysis of yield traits showed that the grade of root rot disease was negatively correlated with seven yield-related traits. It could cause more than 50% of the yield loss at the 5-grade, and even no grain at the 9-grade.【Conclusion】Based on the observation and description of the symptoms of maize root rot, an efficient and easy-to-operate grading criterion reflecting disease severity was formulated according to the aboveground symptoms at the V5 stage. Root rot disease can reduce the plant height, stem diameter and ear height, and easily lead to cause stalk rot. The severity of root rot is negatively correlated with yield-related traits. The results of this study are expected to guide resistance evaluation of varieties against root rot, disease control and yield loss assessment.

Mutation Analysis of Insecticide Target Genes in Populations of Spodoptera frugiperda in China

WANG ShuaiYu,ZHANG ZiTeng,XIE AiTing,DONG Jie,YANG JianGuo,ZHANG AiHuan

Scientia Agricultura Sinica. 2022, 55(20): 3948-3959. doi:10.3864/j.issn.0578-1752.2022.20.007

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【Background】The fall armyworm (Spodoptera frugiperda) is a major migratory agricultural pest in the tropical and subtropical areas of America, it has rapidly expanded to more than 100 countries around the world since 2016, and has now invaded 27 provinces (Autonomous Region, Municipality) in China, posing a great threat to food security. Resistance may become an increasingly serious problem in many regions with the heavy reliance on insecticides for several years and lead to low control efficiency. S. frugiperda had spread to the north of the Great Wall rapidly. However, there are few studies on insecticide resistance and annual variation in China, especially the Great Wall protection zone in 2020 and 2021.【Objective】The objective of this study is to clarify the susceptibility and differences of S. frugiperda populations to insecticides in the Great Wall protection zone and other areas of China, and to provide guidance for scientific insecticide use for the prevention and control of S. frugiperda.【Method】Samples of S. frugiperda were collected from maize fields in 13 provinces (Autonomous Region, Municipality) in China, and the genomic DNA was extracted. The mutations of target genes of carbamate, pyrethroid and diamide insecticides in 362 individuals collected from 47 cities (districts) of 13 provinces (Autonomous Region, Municipality) in 2020 and 2021 were analyzed through direct sequencing of the PCR products individually.【Result】There were 6 genotypes of mutations in ace-1 to carbamate insecticides in all the samples. A201S and F290V mutations were detected in ace-1. The frequency of A201S heterozygous mutation was 8.4%, and the frequency of F290V homozygous and heterozygous mutations was 14.9% and 25.7%, respectively. No G227A mutation was detected. The mutation frequency at F290V in the key prevention region was higher than that in the migratory transition region and annual breeding region, while the mutation frequency at A201S was lower than that in the above two regions. The average frequency of ace-1 mutation was high in the Great Wall protection zone, Huang-Huai-Hai interception zone, Yangtze River Basin and Southwest and South China protection zone. There was no significant difference in the frequency of ace-1 mutation between the populations in 2021 and 2020. No mutations were detected in pyrethroid vgsc and diamide ryr.【Conclusion】A201S and F290V mutations with high frequency in ace-1 is one of the mechanisms conferring carbamate resistance in many populations in China. The use of carbamates should be restricted in the control of S. frugiperda. No mutations had been detected in the vgsc and ryr, suggesting susceptibility to pyrethroids and diamides for rational rotation, however, the low frequency of mutation in S. frugiperda and common mutation in Spodoptera exigua (relatives of S. frugiperda) to pyrethroids and diamides have been detected. In the future, the effect of high insecticide selection stress in the lab and field populations fed by different hosts on the target gene mutation should be evaluated, and the research of rapid diagnostics methods and products of insecticide resistance should be accelerated.

Spatial-Temporal Variation of Cultivated Land Soil Basic Productivity for Main Food Crops in China

LI YuHao,WANG HongYe,CUI ZhenLing,YING Hao,QU XiaoLin,ZHANG JunDa,WANG XinYu

Scientia Agricultura Sinica. 2022, 55(20): 3960-3969. doi:10.3864/j.issn.0578-1752.2022.20.008

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【Objective】 Soil basic productivity is the cornerstone of realizing high and stable yield for food crops. The temporal change trends and spatial variation characteristics of cultivated land productivity for main food crops were defined, so as to provide the important theoretical support for ensuring food security and improve cultivated land quality in China. 【Method】 In this study, based on the national long-term positioning monitoring network of cultivated land quality from 1988 to 2019, the long-term monitoring data of the check area were selected with non-fertilization treatment and the conventional area with farmers' fertilization treatment in the first 3-5 years since the establishment of each monitoring point. The temporal and spatial changes in yield of maize, rice and wheat and soil productivity contribution rates were analyzed in China. 【Result】 In the past 30 years, the grain crops’ yield and soil productivity contribution rates showed an overall increasing trend with time, and the annual growth rate of crop yield showed the change law of non-fertilizer area < conventional area, rice < wheat < maize. The yield of maize, wheat and rice in the non-fertilizer area increased from 2 370, 1 712 and 3 111 kg·hm^-2 in 1988 to 4 852, 3 258 and 4 167 kg·hm^-2 in 2019, respectively, and increased by 104.7%, 90.2% and 34.0%, respectively. The yield of maize, wheat and rice in the conventional area increased from 5 356, 3 296 and 5 970 kg·hm^-2 in 1988 to 8 859, 6 515 and 7 825 kg·hm^-2 in 2019, respectively, with the increment of 65.4%, 97.6% and 31.0%, respectively. The contribution rate of soil productivity for the three major food crops in China from 2015 to 2019 was 52.7%, which was significantly increased by 7.3% compared with 45.4% in 1988-1994. Among them, the contribution rate from maize was 54.3%, which was 12.2% higher than that of 42.1% in 1988-1994. The contribution rate from rice was 53.3%, which was 6.7% higher than that of 46.6% in 1988-1994. The soil productivity contribution rate from wheat increased with the year as a whole, and was lower than that in maize and rice as a whole. The spatial distribution of soil productivity contribution rate for the three major grain crops was quite different. The Northeast region and Yellow River and Huaihai region were higher, which were 56.5% and 54.1%, respectively, followed by the Southwest region and South region, which were 53.7% and 52.9%, respectively. Gan Xin region and Qinghai-Tibet region were the lowest, only 38.7% and 40.4%, respectively. The random forest model was used to rank the soil factors affecting the basic soil productivity contribution rate in the three major grain crop systems. Among them, soil available potassium, organic matter content and soil bulk density were the key factors affecting the spatial distribution of maize basic soil fertility contribution rate; soil available phosphorus, available potassium and organic matter content were the key factors affecting the spatial distribution of wheat basic soil fertility contribution rate; soil pH, soil available phosphorus and organic matter content were the key factors affecting the spatial distribution of rice basic soil fertility contribution rate.【Conclusion】 Over the past 30 years, the soil basic productivity for three major grain crops in China has been continuously improved, but there were great differences among regions and the overall level was still low, which was far lower than that of developed countries in Europe and United States. Soil available potassium content, soil available phosphorus content and soil pH are the most key factors affecting the spatial distribution of basic soil fertility contribution rate of maize, wheat and rice, respectively.

Preparation and Properties of Bionic Modified Water-Based Polymer Coated Urea

CHEN ChunYu,CHEN SongLing,HAN YanYu,REN LiJun,ZOU HongTao,ZHANG YunLong

Scientia Agricultura Sinica. 2022, 55(20): 3970-3982. doi:10.3864/j.issn.0578-1752.2022.20.009

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【Objective】The water-based polymer material is widely used in slow-release fertilizers due to its degradability, non-toxicity and good film-forming properties, but due to its poor water resistance, resulting in poor slow-release effect of the prepared coated fertilizer. In order to improve its water resistance ability, according to the principle of bionics, the nano silica and 1H,1H,2H,2H-perfluorodecyltrimethoxysilane were used for hydrophobic modification, the optimal modification ratio and mechanism were determined, and then the environmentally friendly coated urea with better slow-release effect was prepared. 【Method】 This experiment used a three-factor three-level L₉ (3³) orthogonal design to explore the content of chitosan (0.5%, 1.0%, and 1.5%), starch content (0.5%, 1.0%, and 1.5%) and polyvinyl alcohol (PVA) content ( 2%, 3%, and 4%) on the performance of water-based polymer films, the optimal ratio with better hydrophobic effect was screened through water absorption and range analysis. Furthermore, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, and 3.0% of nano silica was add to the preferred coating materials for hydrophobic modification, respectively. The optimal amount of nano silica was confirmed by measuring the water absorption and permeability of the nano-modified water-based polymer films. Subsequently, the preferred nano-modified water-based polymer film was placed in the nhexane solution containing 0.5%, 1.0%, 1.5%, and 2.0% 1H,1H,2H,2H-perfluorodecyltrimethoxysilane for self-assembly modification, and then the effect of assembly concentration on the properties of films was explored. The modification effect was clarified by the changes of the film's hydrophobicity and permeability, and the hydrophobic modification mechanism was explored through the infrared spectrum characteristics and surface microstructure changes of the films before and after modification. The nutrient release experiments in soil were conducted to explore the nutrient release characters of the biomimetic modified water-based copolymer coated urea. The results of the nutrient release experiments in soil were used to investigate the sustained nutrient release period of nano-modified and biomimetic modified water-based copolymer-coated urea.【Result】 When the content of chitosan was 0.5%, the content of starch was 1.5%, and the content of PVA was 4.0%, the water absorption of the prepared water-based polymer film was the lowest, which was 42.50%. Compared with water-based polymers, the water absorption rates of nano-silica modified water-based copolymer, nano-silica and FAS dual-modified water-based copolymer membrane materials were reduced by 38.54% and 55.98%, respectively, the water permeability were reduced by 36.14% and 60.98%, respectively, and the NH₄⁺ permeability were reduced by 24.14% and 44.58%, respectively. Infrared spectroscopy results showed that the amount of -OH in the nano-silica modified water-based copolymer membrane material was reduced, and Si-O-Si swing vibration and anti-symmetric stretching vibration were observed. The water after the double modification of nano-silica and FAS C-F bonds were observed on the surface of the base copolymer material. The scanning electron microscopy and energy spectrum analysis results showed that Si element appeared on the surface of nano-silica modified water-based copolymer membrane material, and F element appeared on the surface of nano-silica and FAS dual-modified water-based copolymer membrane material and observed a rough surface structure, the water contact angles were increased from 62.5º to 118.6º, and then the coating material could slow down the release effect. In addition, the results of soil culture experiments showed that the control release period of nutrients for nano-silica modified water-based copolymer-coated urea (NWCU), nano-silica and FAS dual-modified water-based copolymer-coated urea (SNWCU) was significantly prolonged. The control release period of nano-silica and FAS dual-modified water-based copolymer-coated urea was increased to about 28 days compared with those of WCU about 10 days. 【Conclusion】 According to the principle of biomimetics, the combined modification with nano-silica and FAS could significantly improve the water resistance and permeability of water-based polymer films, which the prepared biomimetic modified water-based copolymer coated nitrogen fertilizer had a good slow-release effect.

Influence of Plastic Film on Agricultural Production and Its Pollution Control

ZHANG JinRui,REN SiYang,DAI JiZhao,DING Fan,XIAO MouLiang,LIU XueJun,YAN ChangRong,GE TiDa,WANG JingKuan,LIU Qin,WANG Kai,ZHANG FuSuo

Scientia Agricultura Sinica. 2022, 55(20): 3983-3996. doi:10.3864/j.issn.0578-1752.2022.20.010

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Plastic film has been widely used in the farmland all over the world especially in arid and semi-arid areas because of its remarkable agricultural benefits, such as increasing soil temperature and moisture, reducing weed and pest damage, extending crop-growing areas, and consequently improving crop yield and quality. However, the degradation rate of plastic film is extremely slow, and the recovery of plastic film is also relatively backward in China, which lead to a large number of plastic debris in the farmland, causing plastic residues and microplastics pollution in the soil environment. Based on literature, investigation and statistical data, this research reviewed and prospects the impact of plastic film on agricultural production and pollution control in China. Plastic residues and microplastics have been reported to change the physical and chemical properties of soil, restrict soil water and nutrient transport, do harm to the growth, development and reproduction of soil animals and plants, change the abundance and community structure of soil microorganisms, and damage the soil health. In the long-term, plastic residues and microplastics pollution will cause a decline in crop yield and quality. Microplastics had the potential to be absorbed by plants, enter the human body through the food chain and pose a threat to human health. In addition, the large specific surface area of microplastics enabled them to become carriers of other pollutants (e.g. heavy metals, pesticides and antibiotics), causing combined pollution to the soil ecological environment. The standard of plastic film production and use in China was gradually being improved, however, there was still a certain gap compared with developed countries and regions. In addition, a sustainable recycling system of plastic film and the policy of preventing plastic residue and microplastics pollution have not been well formed in China, and the study of microplastics pollution in Chinese farmland soil was still very limited. Therefore, it is critical to solve the problem of plastic residues and microplastics pollution in the soil by evaluating the present situation of plastic residues and microplastics pollution in soils, quantifying the effects of microplastics on the soil environment, and evaluating risks of microplastics to the soil ecosystems, as well as exploring the measures of controlling soil plastic residues and microplastics pollution, and formulating relevant policies and regulations of preventing these pollution.

Transcriptome and Metabolome Integrated Analysis of Epistatic Genetics Effects on Eggplant Peel Color

SUN BaoJuan,WANG Rui,SUN GuangWen,WANG YiKui,LI Tao,GONG Chao,HENG Zhou,YOU Qian,LI ZhiLiang

Scientia Agricultura Sinica. 2022, 55(20): 3997-4010. doi:10.3864/j.issn.0578-1752.2022.20.011

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【Objective】Peel color is closely related to the appearance and value of eggplant. Anthocyanin is one of the natural pigments that determines an eggplant’s peel color. Through the comparison of gene expression and metabolites as well as the analysis on the mechanism of epistatic gene interaction and regulation of anthocyanin synthesis in eggplant peel, this paper provided a theoretical basis for eggplant breeding with different peel colors.【Method】The white-peel female parent 19141 with mutation at the structural gene ANS of anthocyanin biosynthesis pathway, white-peel male parent 19147 with mutation at regulatory gene MYB1 of anthocyanin biosynthesis pathway, and their F₁hybrid E3316 with reddish-purple-peel were used as test materials, while transcriptome sequencing and wide-targeted metabolome analysis were performed on peels of commercial eggplant.【Result】The transcriptome sequencing analysis showed that: 19141_vs_19147 had the most differentially expressed genes (DEGs), followed by E3316_vs_19141. The two comparison groups both had the significant enrichment of DEGs in the flavonoid pathway. The DEGs of E3316_vs_19147 were the least and were not enriched in the flavonoid pathway. A total of 218 metabolites were detected by wide-targeted metabolome analysis. A total of 113 differential accumulated metabolites (DAMs) were detected in E3316_vs_19141, and total 98 DAMs were detected in E3316_vs_19147. The combined analysis of transcriptome and metabolome found that the relative expression levels of key structural genes CHS, CHI, F3H, DFR and ANS, key regulatory genes MYB1, AN1 and AN11, modifier genes 3GT, 5GT, AT and OMT, and transporter gene AN9 (GST) involving anthocyanin biosynthesis pathway had the relation of 19141>E3316>19147. The contents of anthocyanin metabolite cyanidin (CAS: 528-58-5) and cyanin (CAS: 20905-74-2) related to peel coloration had the relation of E3316>19147>19141. The combined analysis of transcriptome and metabolome both showed that the difference of E3316_vs_19141 was bigger than E3316_vs_19147. Under the regulation of epistatic genes, F3'H and F3'5'H genes were expressed simultaneously in eggplant peel, but the expression level of F3'H was much higher than that of F3'5'H. The anthocyanin content of E3316 was higher than that of its parents, while the chlorogenic acid content was lower than that of its parents. 【Conclusion】Under the interaction of epistatic genes with controlling the peel color, the locus (type) and mutation mode of mutant gene in parent determined the trend of gene expression in the whole anthocyanin metabolism pathway and the inhibition mode of peel pigmentation. The reddish-purple-peel F₁ hybrid of two white-peel parents was the result of the function complementation of two mutated gene involved in anthocyanin biosynthesis pathway. The highly expressed F3'H was the main cause for synthesis of cyanidin, and the relationship between anthocyanin and chlorogenic acid biosynthesis was competitive in peels.

Tipburn Injury and Nutritional Quality of Lettuce Plants as Affected by Humidity Control During the Light Period in A Plant Factory

LI YangMei,LIU Xin,JIA MengHan,TONG YuXin

Scientia Agricultura Sinica. 2022, 55(20): 4011-4019. doi:10.3864/j.issn.0578-1752.2022.20.012

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【Background】In the recent years, the plant factories have been developing very fast in the world due to the incomparable advantages, such as labor-saving, pesticide-free, clean, safe and annual planned leafy vegetables production with vertical multi-layer cultivation systems. However, because of the faster growth rate and the weaker transpiration rate of the newly developed leaves, the vegetables in plant factories are very susceptible to tipburn caused by calcium deficiency, and therefore the appearance and nutritional qualities of leafy vegetable are significantly reduced. 【Objective】In this study, the effects of different relative humidity levels on the tipburn injury, growth and nutritional qualities of hydroponically grown lettuce were investigated. The objective of this study was to find out the optimum relative humidity levels for preventing or alleviating tipburn occurrence of vegetable in plant factories. Furthermore, this study also could provide the technical and theoretical support to prevent the tipburn occurrence, and improve vegetable quality and economic benefits of plant factories. 【Method】‘Tiberius’ lettuce, which was susceptible to calcium deficiency, was used in this experiment. Three different air relative humidity levels of 50% (RH50), 70% (RH70) and 90% (RH90) were set up during the light period (16 h), while the relative humidity was maintained at 70% during the dark period (8 h). The other environmental factors, such as light environment, air temperature and carbon dioxide concentration, were kept constant during this study. The tipburn occurrence, yield, photosynthetic parameters and nutritional quality of the lettuce plants were investigated. 【Result】Compared with RH70 and RH90, the calcium content in the newly developed lettuce leaves plants under RH50 was increased significantly, the percentage of lettuce plants with tipburn injury was decreased while harvesting, and the initial tipburn occurrence time was postponed significantly. The stomatal conductance and transpiration rate of lettuce plants increased significantly, which in turn enhanced transpiration and accelerated the absorption and transportation of Ca²⁺, under the lower relative humidity. The photosynthesis rate and fresh weight of lettuce plants were not significantly affected by different relative humidity levels. Moreover, the lower relative humidity also improved the nutritional quality of lettuce plants, including improving the contents of starch, ascorbic acid, and soluble protein. 【Conclusion】The high humidity environment in a plant factory was also one of the reasons for the frequent tipburn occurrence of lettuce plants. Based on the above analysis, it could be concluded that the tipburn occurrence of lettuce plants could be alleviated, the nutritional and commercial qualities of lettuce plants could be improved without affecting its yield and photosynthetic capacity by appropriately controlling the air relative humidity in a plant factory.

Transcriptome Analysis During Flower Bud Differentiation of Red Globe Grape

LIU Xin,ZHANG YaHong,YUAN Miao,DANG ShiZhuo,ZHOU Juan

Scientia Agricultura Sinica. 2022, 55(20): 4020-4035. doi:10.3864/j.issn.0578-1752.2022.20.013

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【Objective】 Grape is an important fruit tree species of China, and the flower bud differentiation directly affects the quality and quantity of this grape. In this study, Red Globe grape developmental stages during flower bud differentiation under natural growth conditions were compared and analyzed, the mechanism of flower bud differentiation of this grape variant was evaluated, and the essential genes were mined, so as to provide a theoretical basis for understanding the flower bud differentiation of this grape species. 【Method】During flower bud differentiation of the Red Globe grape, the morphological observation, phytohormone determination, and transcriptome sequencing analysis were performed on the buds of four developmental stages, including S1 (undifferentiated), S2 (flower anlagen development), S3 (formation of the main cob of inflorescence), and S4 (formation of the second cob of inflorescence). 【Result】A total of 13 729 differentially expressed genes were determined during the flower bud differentiation process of the ‘Red Globe’ grape, which included 4 158, 2 050, 3 425, and 7 652 genes in S1-S2, S2-S3, S3-S4, and S1-S4, respectively. In the enrichment-regulation network of the S1-S4 differential genes, those differential genes were found to be enriched in the hormone-mediated signaling pathways, abscisic acid metabolism, acid chemical reactions, plant cell wall tissues, or biogenesis. Several genes related to auxin, gibberellin, and abscisic acid were detected in the hormone-mediated signaling pathway. The results revealed that the content of auxin was the highest in S2 and the lowest in S3 and S4. The gibberellin content decreased continuously during flower bud differentiation, 80% of that at S4 in S1; the abscisic acid content was higher in S1 and S4 and the lowest in S2. In addition, the S1-S4 differential genes belonged to the transcription factor families (MYB, ERF, bHLH, and MADS-box), indicating that these family genes were involved in the flower bud differentiation of the Red Globe grape. The further analysis of the 13 differentially expressed MADS-box genes revealed upregulated expressions of MADS8, AGL65, AGL15, AGL12, and MADS2 during flower bud differentiation. In contrast, the expressions of AGL30, LeMADS, FBP24, AGL14, and MADS3 were downregulated. These MADS-box genes were verified via quantitative reverse transcription polymerase chain reaction, and the expression trend was found to be consistent with the corresponding transcriptome. 【Conclusion】The flower bud differentiation of the Red Globe grape was a complex biological process. The plant hormone-mediated signaling pathways and the MADS-box family genes played essential roles in flower bud differentiation. These results provided information about transcription factors, genes, and hormones to help understand this complex developmental process and provided a theoretical basis for establishing a comprehensive model for flower bud differentiation in the Red Globe grape.

Comprehensive Evaluation and Selection of Hybrid Offsprings of Large-Flowered Tea Chrysanthemum

HU Xin, ZHANG ZhiLiang, ZHANG Fei, DENG Bo, FANG WeiMin

Scientia Agricultura Sinica. 2022, 55(20): 4036-4051. doi:10.3864/j.issn.0578-1752.2022.20.014

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【Objective】The purpose was to establish an evaluation system to screen out elite hybrid lines of large-flowered tea chrysanthemum, so as to provide a theory reference and material base for the breeding of new varieties. 【Method】A total of 20 factors representative of agronomic traits, sensory quality, and active ingredient were investigated for 46 F₁ offsprings derived from maternal large-flowered tea chrysanthemum Jinsi Huangju and for five parental cultivars, including dry flower weight, ray floret number, florescence, plant height, crown diameter, number of flowers per plant, yield of dry flower, drought resistance, disease resistance, dry flower diameter, dry flower type, dry flower color, aroma, liquor flower type, liquor flower color, taste, total flavones, chlorogenic acid, luteolin glycosides, and iso-chlorogenic acid A. The analytic hierarchy process and K-Means cluster analysis were used to establish a comprehensive evaluation system for assessing the hybrid offsprings. 【Result】Through the observation and analysis of the characters of F₁ offsprings of large-flowered tea chrysanthemum, the results showed that the flowering period of most F₁ offsprings was relatively late. Compared with ‘Jinsi Huangju’, the drought resistance of F₁offsprings was improved to a certain extent, but the disease resistance was average. The inflorescence-related ornamental traits of the most F₁ offspring were generally improved, but the flower diameter was not significantly increased, and the performances of growth, yield and taste traits were not improved as expected. The weight of the 20 factors was determined by analytic hierarchy process, and the result showed that dry flower diameter shared the largest weight value of 0.148, followed by liquor flower type, liquor flower color, and taste with a weight value of 0.084, florescence and yield of dry flower with a weight value of 0.080. The K-Means cluster analysis divided the investigated materials into 3 grades, i.e., 6 superior, 29 good, and 16 poor accessions, accounted for 11.76%, 56.86%, and 31.37%, respectively. Among them, compared with Jinsi Huangju, the superior F₁ offsprings showed heavier fresh flower weight, wider and thicker petals, and more petals, more pure and bright color, higher active ingredients, which could be further conducted regional test. 【Conclusion】The evaluation system of large-flowered tea chrysanthemum was established by the analytic hierarchy process and K-Means cluster analysis, and 5 elite hybrid lines with good agronomic traits, strong resistance, high quality were screened out for future breeding use.

Effects of the Number of Subclinical Mastitis and Somatic Cell Score in Milk of Parity 1 on Somatic Cell Score of Holstein Cows for Parity 2

XIA YuXin,LIANG Yan,WANG HaiYang,GUO MengLing,ZHOU Bu,DAI Xu,YANG ZhangPing,MAO YongJiang

Scientia Agricultura Sinica. 2022, 55(20): 4052-4064. doi:10.3864/j.issn.0578-1752.2022.20.015

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【Objective】There was correlation between physiological status of different parities of cows. The objective of this research was to explore the effect of the number of subclinical mastitis (SCM) and somatic cell score (SCS) in milk of parity 1 on SCS of Holstein cows for parity 2. 【Method】This study collected 162 509 DHI records of Holstein cows with parity 1 and 2 from 2015 to 2020 in 13 farms in Jiangsu Province. Excel 2019 was used to pre-process and filter DHI records firstly. The mixed model of SAS (Ver 9.4) was used to explore the effects of farm size, sampling year, calving season, lactation month, the number of SCM on parity 1 and each average lactation stage of SCS on parity 1 on SCS of parity 2. At the same, the correlation coefficients between the number of SCM and the SCS for each lactation stage of parity 1 with SCS in each lactation month for parity 2 of Holstein cows were calculated.【Result】The farm size, sampling year, calving season, lactation month, the number of SCM on parity 1 and average SCS on parity 1 in different periods had extremely significant effects on the SCS of parity 2 (P<0.01). Among them, SCS with farm size over 5 000 was lower than other farm size (P<0.05). SCS in 2020 was higher than other sampling year (P<0.05). SCS of cows calving in summer and spring was higher than other calving season (P<0.05), while SCS of cows calving in winter was lower than other calving season (P<0.05). SCS in the 9th and 10th lactation month was higher than other lactation months (P<0.05), while SCS in the second lactation month was lower than other lactation months (P<0.05). The SCS in each lactation month of parity 2 showed downward firstly and then upward trend for the cows with the number of SCM less than 2 for parity 1. The SCS for parity 2 in each lactation month fluctuated greatly for the cow with the number of SCM with 3 or more than in parity 1. When the average lactation of SCS, early lactation of SCS, mid lactation of SCS, and late lactation of SCS for the cows in parity 1 were 0, 1, 2, and 3, respectively, the SCS for each lactation month in parity 2 showed downward firstly and then upward trend. When the average lactation of SCS for the cow in parity 1 was more than 3, the SCS in parity 2 in each lactation fluctuated greatly. Overall, the number of SCM and the SCS for each lactation stage of cows in parity 1 increased, and the SCS in each lactation month for parity 2 gradually also increased. There was an extremely significant positive correlation between the number of SCM on parity 1 and SCS on parity 2 in each lactation month (P<0.01). The correlation coefficient of average SCS in the lactation for parity 1 with the SCS in parity 2 lactation month was the maximum (0.238). The correlation coefficient of average SCS in the early lactation for parity 1 and the SCS in parity 2 in each lactation month was the minimum (0.104). Among them, the correlation coefficient of the number of SCM for parity 1 with the SCS for parity 2 in the 3rd lactation month was the maximum. The correlation coefficient of the number of SCM for parity 1 with the SCS for parity 2 in the 10th lactation month was the minimum. The correlation coefficient of average SCS in the lactation for parity 1 with the SCS for parity 2 in the 5th lactation month was the maximum. The correlation coefficient of average SCS in the lactation for parity 1 with the SCS for parity 2 in the 5th lactation month was the minimum. The correlation coefficient of early lactation of SCS for parity 1 with the SCS for parity 2 in the 5th lactation month was the maximum, however, the SCS for parity 2 in the 1st and 10th lactation month was the minimum. The correlation coefficient of mid-lactation of SCS for parity 1 with the SCS for parity 2 in the 5th lactation month was the maximum, while the SCS for parity 2 in the 1st lactation month was the minimum. The correlation coefficient of late lactation of SCS for parity 1 with the SCS for parity 2 in the 5th lactation month was the maximum, while the SCS for parity 2 in the 1st lactation month was the minimum. 【Conclusion】The number of SCM and each average lactation stage of SCS for parity 1 had extremely significant effects on SCS in lactation for the cows of parity 2. There was an extremely significant positive correlation between the numbers of SCM for parity 1 with the SCS for parity 2 in each lactation month. The results provided references for improving quality of raw milk of Holstein cows in parity 2 in the future.

Effects of Amino Acid By-Products on Fermentation Quality and Digestibility of White Sorghum Silage

NUERHATI·Silafuer ,WUSIMAN·Yimiti

Scientia Agricultura Sinica. 2022, 55(20): 4065-4074. doi:10.3864/j.issn.0578-1752.2022.20.016

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【Objective】 The aim of this study was to investigate the effects of appropriate amino acid by-products (ABP) on fermentation quality and digestibility of white sorghum, so as to provide ideas for reducing environmental pollution as well as developing and utilizing new feed additives.【Method】In the study, the control group without any additives and the two experimental groups with ABP and ABP+ forage bacteria were used to carry out the experiment of white sorghum silage fermentation. The effects of ABP on the fermentation quality and digestibility of silage were obtained by measuring the feed composition and in vitro digestibility, and scanning electron microscopy (SEM) was used to observe the mechanism of ABP on improving feed fermentation quality and digestibility.【Result】The study has shown that the addition of 2.0% ABP to white sorghum straw could reduce the pH of the feed to 3.65, which was significantly different from the control group (5.13) (P<0.05). The sensory score belonged to the quality silage interval. The lactic acid content of each experimental group (ABP:11.95 g·kg^-1; MIX:15.14 g·kg^-1) was significantly higher than that of the control group (3.54 g·kg^-1) (P<0.01), the content of acetic acid and butyric acid (AA: ABP:2.87 g·kg^-1, MIX:2.75 g·kg^-1; BA: ABP:0.72 g·kg^-1, MIX:0.78 g·kg^-1) was significantly lower than that of the control group (acetic acid:3.85 g·kg^-1; butyric acid: 1.39 g·kg^-1) (P<0.05), and the lactic acid content of the experimental group of ABP+ forage bacteria was 327.85% higher than that of the control group; the content of dry matter (DM) in each group did not change significantly (P>0.05). Neutral detergent fiber (NDF) (ABP:58.67%; MIX:57.67%), acid detergent lignin (ADL)(ABP:4.77%; MIX:4.27%) and ash (Ash) (ABP : 1.56%; mixed: 2.04%) lower than the control group (NDF:63.66%; ADL:5.15%; Ash:2.76%), but the difference was not significant (P>0.05), the acid detergent fiber (ADF) (ABP:35.77%; MIX:28.63%) was significantly lower than that of the control group (40.58%) (P<0.01), and the crude protein content (ABP: 9.65%, MIX:9.67%) was significantly higher than the control group (6.88%) (P<0.01); the in vitro digestibility of each experimental group was DM (ABP: 74.66%; MIX: 80.03%), NDF (ABP: 72.74%; MIX: 83.08%) and ADF (ABP: 68.29%; MIX: 79.56%), which were significantly higher than the control group (DM: 60.67%, NDF: 48.06%; ADF: 44.81%) (P<0.05); the results showed that ABP significantly improved and increased the fermentation quality and digestibility of silage. From the SEM results, it was found that the cross-section and surface structure of the control group were small, and the number of adhering microorganisms was small too, while the wax layer of the surface structure of the treatment group was destroyed and adhered to a large number of forage bacteria, cross-section cells or a large amount of forage bacteria adhered inside the tissue. Therefore, it was preliminarily informed that ABP improved and increased the fermentation quality and digestibility of silage. In addition, the carbon and nitrogen sources provided by the feed bacteria, the wax layer on the surface of the feed was destroyed to promote the adhesion of the feed bacteria and degrade the cell wall cellulose.【Conclusion】2.0% ABP added to white sorghum silage could significantly improve the fermentation quality and digestibility, and had great economic and social significance for the reuse of ABP, the reduction of environmental pollution, and the development and utilization of new feed additives.

Evolution of Human H9N2 Avian Influenza Virus in China from 1998 to 2021

WANG YanWen,WANG MengJing,ZHANG Hong,GAO XinXin,GUO Jing,LI XuYong

Scientia Agricultura Sinica. 2022, 55(20): 4075-4090. doi:10.3864/j.issn.0578-1752.2022.20.017

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【Objective】 The epidemiological characteristics of H9N2 avian influenza virus were clarified by analyzing the onset time, province, age, gender and other information of human cases infected with H9N2 avian influenza from 1998 to 2021 in China. By analyzing the genetic characteristics of human H9N2 subtype avian influenza virus, the genetic evolution law of human H9N2 subtype avian influenza virus was elucidated. This study could provide data support for the early warning and control of H9N2 subtype avian influenza virus cross-species transmission. 【Method】 According to the case report, literature data and gene bank, the case information and strain sequence data of human infection with H9N2 subtype avian influenza virus in China from 1998 to 2021 were obtained. The infection cases were analyzed from the time distribution, space distribution, gender and age distribution, and the epidemiological characteristics of human-derived H9N2 subtype avian influenza virus infection were found. The MegAlign software in DNASTAR were used to analyze Nucleotide sequence of the homology of various gene fragments of human H9N2 isolates, a phylogenetic tree was constructed by MEGA7.0 software, and the key sites of viral proteins were analyzed. The genetic evolution trend of viral proteins and the variation of key amino acid sites were clarified. The 2019-2021 H9N2 subtype avian influenza virus nucleotide sequence in China was downloaded from the GISAID website. After mafft alignment, the key amino acid site mutation differences between human and avian H9N2 viruses were examined in MEGA7.0, the potential risks brought by current human and avian H9N2 viruses were revealed. 【Result】 From1998 to 2021, a total of 71 cases of human infection with the H9N2 subtype avian influenza virus occurred in China. From the spatial distribution analysis, the cases were distributed in 16 provinces and cities, of which 91.55% of the cases were from 12 provinces and cities in the south. In terms of time, the number of reported infections has been on the rise after 2013, and the cumulative number of infections from 2013 to 2021 accounted for 61.97% of the total number of infections. From the analysis of gender and age distribution, the ratio of male to female was 1:1.68. Human infection with H9N2 virus was mainly seen in infants and young children, accounting for 74.14% of the total number of cases. Genome comparison analysis of human H9N2 viruses found that these viruses belonged to the Eurasian branch, but the nucleotide sequence of homology of these viruses gene fragments was quite different. The nucleotide homology of HA, NA, PB2, PB1, PA, NP, M, and NS were 75.3%-100%, 80.1%-100%, 78.7%-100%, 82.5%-100%, 72.6%-100%, 74.1 -100%, 65.5%-100%, and 82.0%-100%, respectively. According to the genealogy of human isolates, 22 human isolates with complete gene fragments could be divided into 8 genotypes, genotypes in 2003, 2008, and 2013 were significantly different from genotypes in 1999. A total of 42 human H9N2 virus strains uploaded HA sequences from 1998 to 2021, and 38 of them had the mutation of Q226L in the HA protein; a total of 30 human virus strains uploaded PB2 sequence from 1998 to 2021, and 9 strains of which had the mutation of E627V in the PB2 protein, 1 of which had the mutation of E627K in the PB2 protein; the 701 site of the PB2 protein of one virus had the mutation of D701N, a total of 31 virus strains uploaded NS and M sequence from 1998 to 2021, the 42nd positions of the NS1 protein were all S, and the amino acids at positions 30th and 215th of the M1 protein were D and A, respectively. 【Conclusion】 Since 2013, the number of human infections with H9N2 subtype avian influenza reported in China has been on the rise, with significant differences in geographic, age and gender distribution. Since 1998, the genetic homology between human H9N2 isolates was quite different, and the viral gene rearrangement between different branches was frequent, forming a complex genotype. It was suggested that the H9N2 subtype avian influenza virus was constantly evolving. Mutations in key amino acid sites of human H9N2 virus and mutation rate of human viruses was higher than avian H9N2 viruses from 2019 to 2021, suggesting that the potential of H9N2 subtype avian influenza virus to infect humans was gradually increasing. This result enriched the knowledge of human H9N2 virus and provided an important reference for the prevention and control of H9N2 subtype avian influenza virus.

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