A quantitative PCR based method using propidium monoazide for specific and sensitive detection of Pectobacterium carotovorum ssp. carotovorum in kimchi cabbage (Brassica rapa L. subsp. pekinensis)

Jeong, Sang‐Ho; Lee, Jae Yong; Yoon, So‐Ra; Moon, Eun Woo; Ha, Ji‐Hyoung

doi:10.1016/j.lwt.2019.108327

Cited by 8 publications

(5 citation statements)

References 32 publications

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“…qPCR has been widely used as a quantitative test for pathogenic bacteria, including Pectobacterium odoriferum [35], Pectobacterium parmentieri [8], Pectobacterium carotovorum ssp. Carotovorum [36], and Pbr [37]. However, the presence of residual DNA from deceased pathogens can potentially restrict the accurate detection of pathogen numbers via qPCR [37][38][39].…”

Section: Discussionmentioning

confidence: 99%

“…Similarly, once the surviving Pbr pathogen remains on the surface of potato tubers, it can also induce soft rot symptoms in newly sown potatoes. Although the traditional plate isolation method can isolate and identify pathogenic bacteria from seeds, it is unable to meet the current requirements for detecting and providing early warnings of bacterial soft rot disease, which is primarily due to interference caused by non-pathogenic bacteria and the time and costs involved in identifying pathogenic bacteria using this method [36,43]. Whether the constructed PMA-qPCR detection system can accurately and effectively detect live disease-causing bacteria under practical application scenarios is a criterion reflecting the necessity of system construction.…”

Section: Discussionmentioning

confidence: 99%

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Rapid Detection and Quantification of Viable Cells of Pectobacterium brasiliense Using Propidium Monoazide Combined with Real-Time PCR

Li,

Chen,

Yang

et al. 2023

Microorganisms

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Pectobacterium brasiliense (Pbr) has caused significant economic losses in major vegetable production areas in Northern China by causing bacterial soft rot in cash crops such as potatoes and cucumbers. This study aimed to establish a PMA-qPCR detection method for Pbr by screening specific and sensitive primers based on the glu gene and the conserved region of the 23S rRNA gene. Based on the optimized PMA pretreatment conditions, a standard curve was designed and constructed for PMA-qPCR detection (y = −3.391x + 36.28; R2 = 0.99). The amplification efficiency reached 97%, and the lowest detection limit of viable cells was approximately 2 × 102 CFU·mL−1. The feasibility of the PMA-qPCR method was confirmed through a manually simulated viable/dead cell assay under various concentrations. The analysis of potato tubers and cucumber seeds revealed that nine naturally collected seed samples contained a range from 102 to 104 CFU·g−1 viable Pbr bacteria. Furthermore, the system effectively identified changes in the number of pathogenic bacteria in cucumber and potato leaves affected by soft rot throughout the disease period. Overall, the detection and prevention of bacterial soft rot caused by Pbr is crucial.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Rapid Detection and Quantification of Viable Cells of Pectobacterium brasiliense Using Propidium Monoazide Combined with Real-Time PCR

Li,

Chen,

Yang

et al. 2023

Microorganisms

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“…The Petri dishes were incubated at 28°C for 20 h, and the PCC colonies appearing on the plates were counted using the standard plate count method and expressed as CFU/g. Additionally, we quantified the viable PCC cells in cabbage samples by quantitative reverse transcriptase PCR assays combined with an intercalating propidium monoazide dye [ 23 ]. PCC bacterial DNA was not detected in some cabbage samples, or PCC bacterial DNA was detected in no cabbage samples (data not shown).…”

Section: Methodsmentioning

confidence: 99%

Volatile Metabolic Markers for MonitoringPectobacterium carotovorumsubsp.carotovorumUsing Headspace Solid-Phase Microextraction Coupled with Gas Chromatography-Mass Spectrometry

Yang¹,

Lee²,

Song³

et al. 2021

J. Microbiol. Biotechnol.

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Identifying the extracellular metabolites of microorganisms in fresh vegetables is industrially useful for assessing the quality of processed foods. Pectobacterium carotovorum subsp. carotovorum (PCC) is a plant pathogenic bacterium that causes soft rot disease in cabbages. This microbial species in plant tissues can emit specific volatile molecules with odors that are characteristic of the host cell tissues and PCC species. In this study, we used headspace solid-phase microextraction followed by gas chromatography coupled with mass spectrometry (HS-SPME-GC-MS) to identify volatile compounds (VCs) in PCC-inoculated cabbage at different storage temperatures. HS-SPME-GC-MS allowed for recognition of extracellular metabolites in PCC-infected cabbages by identifying specific volatile metabolic markers. 4-Ethyl-5-methylthiazole and 3-butenyl isothiocyanate were identified as markers of fresh cabbages, whereas 2,3-butanediol and ethyl acetate were identified as markers of soft rot in PCC-infected cabbages. These analytical results demonstrate a suitable approach for establishing non-destructive plant pathogen-diagnosis techniques as alternatives to standard methods, within the framework of the development of rapid and efficient analytical techniques for monitoring plant-borne bacterial pathogens. These techniques also demonstrate promising applications for the freshness management and quality control of cabbages.

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“…In each treatment group (F1, F2, P1, and P2), 45 pieces were used for calibration groups (total of 180 samples; F1: 45 samples, F2: 45 samples, P1: 45 samples, and P2: 45 samples) to develop the classi cation models and 15 pieces for the validation group (total of 60 samples; F1: 15 samples, F2: 15 samples, P1: 15 samples, and P2: 15 samples) to verify the tness of the calibration models. For diagnostic analyses of the soft rot disease in cabbage, viable PCC cells were detected using PMA/RT-qPCR assay 25 , water content was determined using the gravimetric method 26 , and extracellular metabolites of the soft rot microorganisms were identi ed using headspace solid-phase microextraction followed by gas chromatography coupled with mass spectrometry (HS-SPME-GC-MS) 4 .…”

Section: Napa Cabbage Preparationmentioning

confidence: 99%

Nondestructive classification of soft rot disease in napa cabbage using hyperspectral imaging analysis

Song

Yoon

Dang

et al. 2022

Preprint

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Identification of soft rot disease in napa cabbage, an essential ingredient of kimchi, is challenging at the industrial scale. Therefore, nondestructive imaging techniques are necessary. Here, we investigated the potential of hyperspectral imaging (HSI) processing in the near-infrared region (900–1,700 nm) for classifying napa cabbage quality using nondestructive measurements. We determined the microbiological and physicochemical qualitative properties of napa cabbage for intercomparison of HSI information, extracted HSI characteristics from hyperspectral images to predict and classify freshness, and established a novel approach for classifying healthy and rotten napa cabbage. Golay’s derivative method for data preprocessing was implemented, followed by wavelength selection using variable importance in projection scores. For multivariate data of the classification models, partial least square discriminant analysis, support vector machine (SVM), and random forests were used for predicting cabbage conditions. The SVM model accurately distinguished the cabbage exhibiting soft rot disease symptoms from the healthy cabbage. This study presents the potential of HSI systems for separating soft rot disease-infected napa cabbages from healthy napa cabbages using the SVM model, especially under the most effective wavelengths (970, 980, 1,180, 1,070, 1,120, and 978 nm), prior to processing. These results are applicable to industrial multispectral images.

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A quantitative PCR based method using propidium monoazide for specific and sensitive detection of Pectobacterium carotovorum ssp. carotovorum in kimchi cabbage (Brassica rapa L. subsp. pekinensis)

Cited by 8 publications

References 32 publications

Rapid Detection and Quantification of Viable Cells of Pectobacterium brasiliense Using Propidium Monoazide Combined with Real-Time PCR

Rapid Detection and Quantification of Viable Cells of Pectobacterium brasiliense Using Propidium Monoazide Combined with Real-Time PCR

Volatile Metabolic Markers for MonitoringPectobacterium carotovorumsubsp.carotovorumUsing Headspace Solid-Phase Microextraction Coupled with Gas Chromatography-Mass Spectrometry

Nondestructive classification of soft rot disease in napa cabbage using hyperspectral imaging analysis

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