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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV230756	1/2	Xylella fastidiosa	9a5c	(d)(U)C Filtration	Mendes JS	2016	56%
Study summary Full title Determination of Extracellular Proteins from . All authors Mendes JS, Santiago AS, Toledo MA, Horta MA, de Souza AA, Tasic L, de Souza AP Journal Front Microbiol Abstract The phytopathogen causes economic losses in important agricultural crops. Xylem vessel occlusion ca (show more...)The phytopathogen causes economic losses in important agricultural crops. Xylem vessel occlusion caused by biofilm formation is the major mechanism underlying the pathogenicity of distinct strains of . Here, we provide a detailed characterization of the extracellular proteins of . Based on the results, we performed a comparison with a strain J1a12, which cannot induce citrus variegated chlorosis symptoms when inoculated into citrus plants. We then extend this approach to analyze the extracellular proteins of in media supplemented with calcium. We verified increases in extracellular proteins concomitant with the days of growth and, consequently, biofilm development (3-30 days). Outer membrane vesicles carrying toxins were identified beginning at 10 days of growth in the 9a5c strain. In addition, a decrease in extracellular proteins in media supplemented with calcium was observed in both strains. Using mass spectrometry, 71 different proteins were identified during 30 days of biofilm development, including proteases, quorum-sensing proteins, biofilm formation proteins, hypothetical proteins, phage-related proteins, chaperones, toxins, antitoxins, and extracellular vesicle membrane components. (hide) EV-METRIC 56% (90th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles outer membrane vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (Differential) (ultra)centrifugation Filtration Protein markers EV: XF2491 non-EV: not specified Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Xylella fastidiosa Sample Type Cell culture supernatant EV-producing cells 9a5c EV-harvesting Medium Serum free medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 180 Pelleting: rotor type L8-80 M Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 40 Wash: time (min) 120 Wash: speed (g) 100000 Filtration steps 0.2 or 0.22 Î¼m Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins XF2491 Not detected EV-associated proteins not specified Detected contaminants not specified Not detected contaminants not specified Proteomics database Unicamp Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Transmission-EM Image type Wide-field

	EV230756	2/2	Xylella fastidiosa	J1a12	(d)(U)C Filtration	Mendes JS	2016	44%
Study summary Full title Determination of Extracellular Proteins from . All authors Mendes JS, Santiago AS, Toledo MA, Horta MA, de Souza AA, Tasic L, de Souza AP Journal Front Microbiol Abstract The phytopathogen causes economic losses in important agricultural crops. Xylem vessel occlusion ca (show more...)The phytopathogen causes economic losses in important agricultural crops. Xylem vessel occlusion caused by biofilm formation is the major mechanism underlying the pathogenicity of distinct strains of . Here, we provide a detailed characterization of the extracellular proteins of . Based on the results, we performed a comparison with a strain J1a12, which cannot induce citrus variegated chlorosis symptoms when inoculated into citrus plants. We then extend this approach to analyze the extracellular proteins of in media supplemented with calcium. We verified increases in extracellular proteins concomitant with the days of growth and, consequently, biofilm development (3-30 days). Outer membrane vesicles carrying toxins were identified beginning at 10 days of growth in the 9a5c strain. In addition, a decrease in extracellular proteins in media supplemented with calcium was observed in both strains. Using mass spectrometry, 71 different proteins were identified during 30 days of biofilm development, including proteases, quorum-sensing proteins, biofilm formation proteins, hypothetical proteins, phage-related proteins, chaperones, toxins, antitoxins, and extracellular vesicle membrane components. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles outer membrane vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (Differential) (ultra)centrifugation Filtration Protein markers EV: not specified non-EV: not specified Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Xylella fastidiosa Sample Type Cell culture supernatant EV-producing cells J1a12 EV-harvesting Medium Serum free medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 180 Pelleting: rotor type L8-80 M Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 40 Wash: time (min) 120 Wash: speed (g) 100000 Filtration steps 0.2 or 0.22 Î¼m Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins not specified Not detected EV-associated proteins XF2491 Detected contaminants not specified Not detected contaminants not specified Proteomics database Unicamp Characterization: Lipid analysis No Characterization: Particle analysis None

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EV-TRACK ID	EV230756
species	Xylella fastidiosa
sample type	Cell culture
cell type	9a5c	J1a12
condition	Control condition	Control condition
separation protocol	dUC/ Filtration	dUC/ Filtration
Exp. nr.	1	2
EV-METRIC %	56	44