This study was designed to identify the pathogen causing soft rot of Pinellia ternata in Qianjiang of Hubei province and screen out the effective bactericides, so as to provide a theoretical basis for the control of soft rot of P. ternata. In this study, the pathogen was identified based on molecular biology and physiological biochemistry, followed by the detection of pathogenicity and pathogenicity spectrum via plant tissue inoculation in vitro and the indoor toxicity determination using the inhibition zone method to screen out bactericide with good antibacterial effects. The control effect of the bactericide against P. ternata soft rot was verified by the leave and tuber inoculation in vitro. The phylogenetic tree was constructed based on the 16 S rDNA, dnaX gene, and recA gene sequences, respectively, and the result showed that the pathogen belonged to the same branch as the type strain Dickeya fangzhongdai JS5. The physiological and biochemical tests showed that the pathogen was identical to D. fangzhongdai, which proved that the pathogen was D. fangzhongdai. The pathogenicity test indicated that the pathogen could obviously infect leaves at 24 h and tubers in 3 d. As revealed by the indoor toxicity test, 0.3% tetramycin, 5% allicin, and 80% ethylicin had good antibacterial activities, with EC_(50) values all less than 50 mg·L~(-1). Tests in tissues in vitro showed that 5% allicin exhibited the best control effect, followed by 0.3% tetramycin and 10% zhongshengmycin oligosaccharide, and their preventive effects were better than curative effects. Therefore, 5% allicin can be used as the preferred agent for the control of P. ternata soft rot, and 0.3% tetramycin and 10% zhongshengmycin oligosaccharide as the alternatives. This study has provided a certain theoretical basis for the control of P. ternata soft rot.
Phylogeny ; Pinellia/chemistry* ; Plant Leaves ; Plant Tubers

To further study the mechanism of sprout tumble caused by drought,drought stress was simulating with 30% PEG 6000,physiological,and then the morphological changes of Pinellia ternata cells at different treatment time were detected. The results indicated that,along with the period of drought stress continued,the contents of chlorophyll and water potential were decreased,relative electrical conductivity,contents of soluble sugar and MDA increased. Sprout tumble of P. ternata first occurred on the fourth day during drought stress,large scale of sprout tumble appeared on the eighth day with about 73% of tumble rate. The nuclei exposed to drought stress for 2 days were flattened,lobed,invalidated or irregular in shape and significant showed the apoptotic morphological characteristics. Adenylate transferase( ANT) gene expressions were inhibited by drought,with the rapid increase of Caspase-3 enzyme activity,the cell death rate increased. All this proves that the essence of sprout tumble caused by drought is programmed cell death,which may be a self dormancy protection mechanism of P. ternata against adverse environment.
Apoptosis ; Droughts ; Pinellia ; cytology ; Stress, Physiological

The wild Pinellia ternata resource was surveyed by field investigation, visiting surveys combine with literature study and changing factors of the wild P. ternata in the area of Huanghuai Plain were analyzed. The results indicated that Huanghuai Plain wild P. ternata resources were scarce and nearly extinct. The chemical farming methods in Huanghuai Plain have endangered the survival of the wild P. ternata.
China ; Conservation of Natural Resources ; Endangered Species ; Pinellia ; growth & development

The main factors which affected the isolation, purification and cultivation of Pinellia cordata protoplasts from leaves were studied. The results indicated that the optimum enzyme solution for P. cordata leaves was 13% CPW + 1.0% Cellulose +0.1% Pectolase, at pH 6.0, temperature (25-28 degrees C ) for 4 h. The sucrose density gradient centrifugation was adopted to purificate the protoplasts collected, when 25% sucrose was used as mediator, centrifugating at 500 rpm for 10 min. When the protoplasts were shallow liquid and liquid-solid double layer cultured on the medium of MS + 0.5 mg x L(-1) 6-BA + 0.25 mg x L(-1) NAA + 13% mannitol at the density of 2.5 x 104 protoplasts/mL, or fed and nursed cultured at the density of 100-500 protoplasts/mL, cell division could be observed for 3 days; granular calli appeared for 30 days. Calli was proliferated on the medium of MS + 0.5 mg x L(-1) 6-BA + 0.25 mg x L(-1) NAA solidified by 0.55% agar, and differentiated and regenerated after 5-6 months. Plant generation of P. cordata is successfully established.
Cell Separation ; methods ; Culture Media ; Pinellia ; physiology ; Protoplasts ; physiology ; Regeneration

In summer in 2020, Pinellia ternata in many planting areas in Hubei suffered from serious southern blight, as manifested by the yellowing and wilted leaves and rotten tubers. This study aims to identify the pathogen, clarify the biological characteristics of the pathogen, and screen fungicides. To be specific, the pathogen was isolated, purified, and identified, and the pathogenicity was detected according to the Koch's postulates. Moreover, the biological characteristics of the pathogen were analyzed. Furthermore, PDA plates and seedlings were used to determine the most effective fungicides. The results showed that the mycelia of the pathogen were white and villous with silk luster, which produced a large number of white to black brown sclerotia. The pathogen was identified as Athelia rolfsii by morphological observation and molecular identification based on LSU and TEF gene sequences. The optimum growth conditions for A. rolfsii were 30 ℃ and pH 5-8, and the optimum conditions for the germination of sclerotia were 25 ℃ and pH 7-9. Bacillus subtilis, difenoconazole, and flusilazole were identified as effective fungicides with PDA, and their half maximal effective concentration(EC_(50)) was all less than 5 mg·L~(-1). The effective fungicides screened with the seedlings were hymexazol and difenoconazole. Based on the screening experiments, difenoconazole can be used as the main agent for the prevention and treatment of southern blight.
Pinellia/genetics* ; Fungicides, Industrial/pharmacology* ; Seedlings ; Bacillus subtilis ; Mycelium

The study aims at investigating the grade standard and the quality characteristic of Pinelliae Rhizoma for commodity specification, which provides the reference for its grade standard formulation. 42 Pinelliae Rhizoma simples were collected from 5 medicinal materials markets and 2 producing areas. Based on the previous herbalogical study and market investigation, we combined with the data analysis to select the grading indicators using SPSS software for descriptive statistical analysis, analysis of variance, K-cluster analysis and correlation analysis. According to the actual production condition, we developed the grading standards of Pinelliae Rhizoma. Moreover, we compared the internal indicators(water, total ash, leachate and guanosine) of Pinelliae Rhizoma at various grade levels, and analyzed the correlation between appearance traits and internal indicators. The herbalogical study and market research found that the Pinelliae Rhizoma was better in large, solid and white. The results from descriptive and variance analysis showed that the appearance of Pinelliae Rhizoma was significantly different in weight per grain and grain number of 500 g. Referring to the 2015 Chinese Pharmacopoeia and the production practice, we use the length, weight per grain and grain number of 500 g as the classification index of Pinelliae Rhizoma. The results from correlation analysis showed that there was no significant correlation between the appearance of Pinelliae Rhizoma and the intrinsic quality index. In addition, we found there was no significant difference in the content of the intrinsic index except for the total ash and the extract. The current study established the classification index of the product specification and grade standard of Pinelliae Rhizoma with length, weight per grain and grain number of 500 g as the index, which can provide the basis for the classification of the product specification and grade of Pinelliae Rhizoma market.
Drugs, Chinese Herbal ; standards ; Pinellia ; chemistry ; Rhizome ; chemistry

The evaluation of germplasm resources is the prerequisite for the development, utilization, and conservation of Chinese medicinal resources. The selection of excellent germplasm is the key to the breeding and orderly production of Pinellia ternata. In this study, 21 germplasm materials of P. ternata from major production areas in China were collected and analyzed for population diversity after phenotypic preliminary screening. The results have revealed that the P. ternata population has abundant phenotypic variation, and the phenotypic changes could be divided into five phenotypes in terms of organ trait variation. Further analysis of variation in 20 quantitative traits of the population revealed that the coefficient of variation for adenosine content(339.05%) was the largest, while the coefficient of variation for the underground plant height(16.35%) was the smallest. Correlation analysis showed that there was a strong correlation among various traits, with 52 pairs of traits showing highly significant correlation(P<0.01) and 19 pairs of traits showing a significant correlation(P<0.05). The 21 germplasms in the test could be classified into three major clusters by cluster analysis, with Cluster Ⅱ having the highest number and content of nucleosides, making it suitable for the selection and breeding of P. ternata varieties with high content of nucleosides. The yield in Cluster Ⅲ was higher than that in other groups, making it suitable for the selection and breeding of P. ternata varieties with a high yield. All trait indicators could be simplified into five principal component factors through principal component analysis, and the cumulative contribution rate was up to 86.04%. Further, comprehensive analysis using membership function and stepwise regression analysis identified nine traits, such as plant height, main leaf length, and underground plant height as characteristic indicators for the comprehensive evaluation of germplasm resources of P. ternata. BX007, BX008, and BX005 were identified as germplasms with both high yield and high uridine content, with BX007 having the highest uridine content of 479.51 μg·g~(-1). It belonged to the germplasm of P. ternata with double bulbils and could be cultivated as a potential good variety. Based on the phenotypic classification of P. ternata, systematic resource evaluation was carried out in this study, which could lay a foundation for the excavation of genetic resources and the breeding of new varieties of P. ternata.
Plants, Medicinal ; Pinellia/genetics* ; Plant Breeding ; Phenotype ; Uridine

The present study investigated the effect of immersion in the excipient lime water on the toxic component lectin protein and explained the scientific connotation of lime water detoxication during the processing of Pinelliae Rhizoma Praeparatum. Western blot was used to investigate the effects of immersion in lime water with different pH(pH 10, 11, and 12.4), saturated sodium hydroxide, and sodium bicarbonate solution on the content of lectin protein. The protein compositions of the supernatant and the precipitate after immersing lectin protein in lime water of different pH were determined by the SDS-PAGE method combined with the silver staining technique. The MALDI-TOF-MS/MS technique was used to detect the molecular weight distribution of peptide fragments in the supernatant and precipitate after immersing lectin protein in lime water of different pH, and circular dichroism spectroscopy was used to detect the ratio changes in the secondary structure of lectin protein during the immersion. The results showed that immersion in lime water at pH>12 and saturated sodium hydroxide solution could significantly reduce the content of lectin protein, while immersion in lime water at pH<12 and sodium bicarbonate solution had no significant effect on lectin protein content. The corresponding lectin protein bands and molecular ion peaks were not detected at the 12 kDa position in the supernatant and precipitate after immersing the lectin protein in lime water at pH>12, which was attributed to the fact that lime water immersion at pH>12 could significantly change the ratio of the secondary structure of lectin protein, resulting in irreversible denaturation, while lime water immersion at pH<12 did not change the ratio of the secondary structure of lectin protein. Therefore, pH>12 was the key condition for the detoxication of lime water during the processing of Pinelliae Rhizoma Praeparatum. Lime water immersion at pH>12 could cause irreversible denaturation of lectin protein, resulting in a significant decrease in the inflammatory toxicity of Pinelliae Rhizoma Praeparatum, which played a key role in detoxification.
Lectins ; Pinellia ; Sodium Bicarbonate ; Sodium Hydroxide ; Tandem Mass Spectrometry ; Water

OBJECTIVETo assess population genetic diversity of Pinellia ternata with different phenotype and from different habitat by sequence-related amplified polymorphism (SRAP) technique.

METHODFourteen appropriate primer pairs were selected out from a total of 80 pairs for SRAP PCR amplification. A Jaccard's genetic similarity matrix and a dendrogram were established using SPSS 16.0 software.

RESULTThe 14 primer pairs could amplify 171 bands, and the percentage of polymorphic bands was 15.8%. Based on the dendrogram, P. ternata in the same habitat clustered in a clade.

CONCLUSIONThese results suggested that SRAP markers could be used as an effective molecular technique for the diversity study of P. ternata and the habitat was more important than the phenotype in identification of P. ternata germplasm resource.

DNA Fingerprinting ; Ecosystem ; Genetic Variation ; Geography ; Phylogeny ; Pinellia ; classification ; genetics ; Polymerase Chain Reaction

OBJECTIVEIn order to clarify the genetic background of Pinellia ternata germplasm resources in China, the chromosomal constitution and cytogeographical distribution of P. ternata were investigated in 27 different populations among 16 provinces and regions in China systematically.

METHODCytological and cytogeographical methods were used in the study.

RESULTP. ternata in China is a polyploid complex, which contains septuploid (2n = 7x = 91) , octoploid (2n = 8x = 104) , nonuploid (2n = 9x = 117) and decaploid (2n = 10x = 130). Meanwhile the aneuploid series (2n = 92, 103, 105, 115) of a minority of P. ternata were also found.

CONCLUSIONThe genetic differentiation and the phenomenon of ploidy miscellany commonly exist in the species of P. ternata in China, both for natural populations and cultivated populations. Toxicity and chemical components of different ploidy P. ternata should be clarified before the superior multiploid is selected for normalized plantation of the plant.

Aneuploidy ; China ; Chromosomes, Plant ; genetics ; Ecosystem ; Genetic Variation ; Pinellia ; genetics ; Plants, Medicinal ; genetics ; Polyploidy