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Pitaya proteomics LC-MS/MS

ABSTRACT: The pathogenic fungus Neoscytalidium dimidiatum (Nd) is the causal agent of pitaya canker and causes significant yield losses. The mechanism by which Nd invades pitaya stems remains largely unknown. Here, quantitative proteomic analysis was employed to investigate pitaya immune responses against Nd infection. A total of 2766 proteins including 244 differentially expressed proteins (DEPs) were identified during the infection cycle. Bioinformatics analysis indicated that the DEPs were associated with photosynthesis, phytohormone activity, reactive oxygen species (ROS) homeostasis and pathogenic defense responses. This study demonstrates that comparative proteomics is an effective strategy for providing new perspective on biotic stress from fungal pathogens in pitaya.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Stenocereus Stellatus

TISSUE(S): Plant Cell, Leaf

DISEASE(S): Fungal Infectious Disease

SUBMITTER: qiang Fu

LAB HEAD: Li Mei Pan

PROVIDER: PXD007983 | Pride | 2022-12-05

REPOSITORIES: Pride

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Project description:Pathogenic biofilms have been associated with persistent infections due to high resistance to antimicrobial agents while commensal biofilms often fortify host immune system. Hence, controlling biofilm formation of both pathogenic bacteria and commensal bacteria is important in bacteria-related diseases. We investigated the effect of plant flavonoids on biofilm formation of both enterohemorrhagic Escherichia coli O157:H7 and three commensal E. coli K-12 strains. Phloretin abundant in apples markedly reduced E. coli O157:H7 biofilm formation without affecting the growth of planktonic cells while phloretin did not harm commensal E. coli K-12 biofilms. Also, phloretin reduced E. coli O157:H7 attachment to human colon epithelial cells. Global transcriptome analyses revealed that phloretin repressed toxin genes (hlyE and stx2), autoinducer-2 importer genes (lsrACDBF), a curli gene (csgA), and a dozens of prophage genes in E. coli O157:H7 cells. Electron microscopy confirmed that phroretin reduced the curli production in E. coli O157:H7. In addition, phloretin suppressed TNF-Î±-induced inflammatory response in vitro using human colonic epithelial cells. Moreover, in the trinitrobenzene sulfonic acid (TNBS)-induced rat colitis model, phloretin significantly ameliorated colon inflammation and body weight loss. Taken together, our results suggest that phloretin may act as an inhibitor of E. coli O157:H7 biofilm formation as well as anti-inflammatory agent on inflammatory bowel diseases while leaving beneficial commensal E. coli biofilm intact. For the microarray experiments, E. coli O157:H7 EDL933 was inoculated in 25 ml of LB in 250 ml flasks with overnight cultures that were diluted at 1:100. Cells were shaken at 100 rpm and 37Â°C for 7 hrs. Cells were immediately chilled with dry ice and 95% ethanol (to prevent RNA degradation) for 30 sec before centrifugation in 50 ml centrifuge tubes at 13,000 g for 2 min; cell pellets were frozen immediately with dry ice and stored -80Â°C. RNA was isolated using Qiagen RNeasy mini Kit (Valencia, CA, USA). To eliminate DNA contamination, Qiagen RNase-free DNase I was used to digest DNA. RNA quality was assessed by Agilent 2100 bioanalyser using the RNA 6000 Nano Chip (Agilent Technologies, Amstelveen, The Netherlands), and quantity was determined by ND-1000 Spectrophotometer (NanoDrop Technologies, Inc., DE, USA).

Project description:Honey has been widely used against bacterial infection for centuries. Previous studies suggested that honeys in high concentrations inhibited bacterial growth due to the presence of anti-microbial compounds, such as methylglyoxal, hydrogen peroxide, and peptides. In this study, we found that three honeys (acacia, clover, and polyfloral) in a low concentration as below as 0.5% (v/v) significantly suppress virulence and biofilm formation in enterohemorrhagic E. coli O157:H7 affecting the growth of planktonic cells while these honeys do not harm commensal E. coli K-12 biofilm formation. Transcriptome analyses show that honeys (0.5%) markedly repress quorum sensing genes (e.g., AI-2 import and indole biosynthesis), virulence genes (e.g., LEE genes), and curli genes (csgBAC). We found that glucose and fructose in honeys are key compounds to reduce the biofilm formation of E. coli O157:H7 via suppressing curli production, but not that of E. coli K-12. Additionally, we observed the temperature-dependent response of honeys and glucose on commensal E. coli K-12 biofilm formation; honey and glucose increase E. coli K-12 biofilm formation at 37Â°C, while they decrease E. coli K-12 biofilm formation at 26Â°C. These results suggest that honey can be a practical tool for reducing virulence and colonization of the pathogenic E. coli O157:H7, while honeys do not harm commensal E. coli community in the human. For the microarray experiments, E. coli O157:H7 EDL933 was inoculated in 250 ml of LB in 1000 ml flasks with overnight cultures that were diluted at 1:100. Cells were shaken with 10g of glass wool at 100 rpm and 37Â°C for 7 hrs. Cells were immediately chilled with dry ice and 95% ethanol (to prevent RNA degradation) for 30 sec before centrifugation in 50 ml centrifuge tubes at 13,000 g for 2 min; cell pellets were frozen immediately with dry ice and stored -80Â°C. RNA was isolated using Qiagen RNeasy mini Kit (Valencia, CA, USA). To eliminate DNA contamination, Qiagen RNase-free DNase I was used to digest DNA. RNA quality was assessed by Agilent 2100 bioanalyser using the RNA 6000 Nano Chip (Agilent Technologies, Amstelveen, The Netherlands), and quantity was determined by ND-1000 Spectrophotometer (NanoDrop Technologies, Inc., DE, USA).

			Action	DRS
	pitaya.fasta	Fasta
	pitaya_1.msf	Msf
	t1_1.raw	Raw
	t1_10.raw	Raw
	t1_2.raw	Raw

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Pitaya proteomics LC-MS/MS

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