Deciphering Molecular Determinants Underlying Penicillium digitatum’s Response to Biological and Chemical Antifungal Agents by Tandem Mass Tag (TMT)-Based High-Resolution LC-MS/MS

Penicillium digitatum is a widespread pathogen responsible for the postharvest decay of citrus, one of the most economically important crops worldwide. Currently, chemical fungicides are still the main strategy to control the green mould disease caused by the fungus. However, the increasing selection and proliferation of fungicide-resistant strains require more efforts to explore new alternatives acting via new or unexplored mechanisms for postharvest disease management. To date, several non-chemical compounds have been investigated for the control of fungal pathogens. In this scenario, understanding the molecular determinants underlying P. digitatum’s response to biological and chemical antifungals may help in the development of safer and more effective non-chemical control methods. In this work, a proteomic approach based on isobaric labelling and a nanoLC tandem mass spectrometry approach was used to investigate molecular changes associated with P. digitatum’s response to treatments with α-sarcin and beetin 27 (BE27), two proteins endowed with antifungal activity. The outcomes of treatments with these biological agents were then compared with those triggered by the commonly used chemical fungicide thiabendazole (TBZ). Our results showed that differentially expressed proteins mainly include cell wall-degrading enzymes, proteins involved in stress response, antioxidant and detoxification mechanisms and metabolic processes such as thiamine biosynthesis. Interestingly, specific modulations in response to protein toxins treatments were observed for a subset of proteins. Deciphering the inhibitory mechanisms of biofungicides and chemical compounds, together with understanding their effects on the fungal physiology, will provide a new direction for improving the efficacy of novel antifungal formulations and developing new control strategies.

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Intracellular and Extracellular Markers of Lethality in Osteogenesis Imperfecta: A Quantitative Proteomic Approach

International Journal of Molecular Sciences ◽

10.3390/ijms22010429 ◽

2021 ◽

Vol 22 (1) ◽

pp. 429

Author(s):

Luca Bini ◽

Domitille Schvartz ◽

Chiara Carnemolla ◽

Roberta Besio ◽

Nadia Garibaldi ◽

...

Keyword(s):

Osteogenesis Imperfecta ◽

Type I Collagen ◽

Type I ◽

Tandem Mass ◽

Type Ii ◽

Type Iii ◽

Bioinformatic Tools ◽

Proteomic Approach ◽

Fibrillin 1 ◽

Heritable Disorder

Osteogenesis imperfecta (OI) is a heritable disorder that mainly affects the skeleton. The inheritance is mostly autosomal dominant and associated to mutations in one of the two genes, COL1A1 and COL1A2, encoding for the type I collagen α chains. According to more than 1500 described mutation sites and to outcome spanning from very mild cases to perinatal-lethality, OI is characterized by a wide genotype/phenotype heterogeneity. In order to identify common affected molecular-pathways and disease biomarkers in OI probands with different mutations and lethal or surviving phenotypes, primary fibroblasts from dominant OI patients, carrying COL1A1 or COL1A2 defects, were investigated by applying a Tandem Mass Tag labeling-Liquid Chromatography-Tandem Mass Spectrometry (TMT LC-MS/MS) proteomics approach and bioinformatic tools for comparative protein-abundance profiling. While no difference in α1 or α2 abundance was detected among lethal (type II) and not-lethal (type III) OI patients, 17 proteins, with key effects on matrix structure and organization, cell signaling, and cell and tissue development and differentiation, were significantly different between type II and type III OI patients. Among them, some non–collagenous extracellular matrix (ECM) proteins (e.g., decorin and fibrillin-1) and proteins modulating cytoskeleton (e.g., nestin and palladin) directly correlate to the severity of the disease. Their defective presence may define proband-failure in balancing aberrances related to mutant collagen.

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The Tripod for Bacterial Natural Product Discovery: Genome Mining, Silent Pathway Induction, and Mass Spectrometry-Based Molecular Networking

mSystems ◽

10.1128/msystems.00160-17 ◽

2018 ◽

Vol 3 (2) ◽

Cited By ~ 14

Author(s):

Daniela B. B. Trivella ◽

Rafael de Felicio

Keyword(s):

Mass Spectrometry ◽

Natural Products ◽

Natural Product ◽

Biosynthetic Pathway ◽

Genome Mining ◽

Chemical Compounds ◽

Gene Clusters ◽

Tandem Mass ◽

Molecular Networking ◽

Natural Product Discovery

ABSTRACT Natural products are the richest source of chemical compounds for drug discovery. Particularly, bacterial secondary metabolites are in the spotlight due to advances in genome sequencing and mining, as well as for the potential of biosynthetic pathway manipulation to awake silent (cryptic) gene clusters under laboratory cultivation. Further progress in compound detection, such as the development of the tandem mass spectrometry (MS/MS) molecular networking approach, has contributed to the discovery of novel bacterial natural products. The latter can be applied directly to bacterial crude extracts for identifying and dereplicating known compounds, therefore assisting the prioritization of extracts containing novel natural products, for example. In our opinion, these three approaches—genome mining, silent pathway induction, and MS-based molecular networking—compose the tripod for modern bacterial natural product discovery and will be discussed in this perspective.

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Peach Brown Rot: Still in Search of an Ideal Management Option

Agriculture ◽

10.3390/agriculture8080125 ◽

2018 ◽

Vol 8 (8) ◽

pp. 125 ◽

Cited By ~ 13

Author(s):

Vitus Ikechukwu Obi ◽

Juan José Barriuso ◽

Yolanda Gogorcena

Keyword(s):

Fungal Pathogens ◽

Crop Production ◽

Control Strategies ◽

Brown Rot ◽

Management Option ◽

Management Options ◽

Pests And Diseases ◽

Resistant Varieties ◽

Rot Disease ◽

Post Harvest Loss

The peach is one of the most important global tree crops within the economically important Rosaceae family. The crop is threatened by numerous pests and diseases, especially fungal pathogens, in the field, in transit, and in the store. More than 50% of the global post-harvest loss has been ascribed to brown rot disease, especially in peach late-ripening varieties. In recent years, the disease has been so manifest in the orchards that some stone fruits were abandoned before harvest. In Spain, particularly, the disease has been associated with well over 60% of fruit loss after harvest. The most common management options available for the control of this disease involve agronomical, chemical, biological, and physical approaches. However, the effects of biochemical fungicides (biological and conventional fungicides), on the environment, human health, and strain fungicide resistance, tend to revise these control strategies. This review aims to comprehensively compile the information currently available on the species of the fungus Monilinia, which causes brown rot in peach, and the available options to control the disease. The breeding for brown rot-resistant varieties remains an ideal management option for brown rot disease control, considering the uniqueness of its sustainability in the chain of crop production.

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STRUCTURE AND FUNCTION OF THE FRUIT MICROBIOME IN HEALTHY AND DISEASED KIWIFRUIT

The Pakistan Journal of Agricultural Sciences ◽

10.21162/pakjas/19.8820 ◽

2019 ◽

Vol 56 (03) ◽

pp. 577-585

Author(s):

Wenneng Wu

Keyword(s):

Microbial Diversity ◽

Relative Abundance ◽

Foodborne Pathogens ◽

Fungal Pathogens ◽

Sugar Metabolism ◽

Amino Sugar ◽

Disease Diagnosis ◽

Postharvest Disease ◽

Pathogen Infection ◽

Microbiome Composition

The fruit surface is an infection court where foodborne pathogens compete with indigenous microbiota for microsites to invade the fruits for nutrients acquisition. However, our current understanding of the structure and functions of fruit microbiome visa-vis postharvest pathogen infection is still nascent. Here, we sequenced the metagenomic DNA to understand the structural and functional attributes of healthy and diseased kiwifruit microbiome. The healthy fruits exhibited higher microbial diversity and distinct microbiome composition compared with diseased fruits. The microbiome of diseased fruit was dominated by fungal pathogens Neofusicoccum parvum and Diplodiaseriata, while the microbiome of healthy fruits were enriched by bacteria from Methylobacteriaceae, Sphingomonadaceae, Nocardioidaceae and fungi in Pleosporaceae. Importantly, the healthy fruit microbiome had a higher relative abundance of genes related to ABC transporter, two-component system, bacterial chemotaxis, bacterial secretion system, but had a lower relative abundance of genes associated with polycyclic aromatic hydrocarbon degradation, amino sugar and nucleotide sugar metabolism, glycine, serine and threonine metabolism compared with diseased fruits. Our results indicate that pathogen infection disrupts the fruit microbiome. The changes in microbiome composition and functions could also increase the possibility of secondary pathogen infection as the reduced microbial diversity may demonstrate less resistance to pathogens infection. Therefore, monitoring the microbiome dynamics and their functions using metagenomic approaches could be useful to build a predictive understanding of accurate postharvest disease diagnosis and management in the future

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Identification of the Fungal Pathogens of Postharvest Disease on Peach Fruits and the Control Mechanisms of Bacillus subtilis JK-14

Toxins ◽

10.3390/toxins11060322 ◽

2019 ◽

Vol 11 (6) ◽

pp. 322 ◽

Cited By ~ 11

Author(s):

Shuwu Zhang ◽

Qi Zheng ◽

Bingliang Xu ◽

Jia Liu

Keyword(s):

Biological Control ◽

Bacillus Subtilis ◽

Fungal Pathogens ◽

Biological Control Agent ◽

Storage Period ◽

Control Agent ◽

Antagonistic Activity ◽

Postharvest Disease ◽

Economic Losses ◽

Control Mechanisms

Postharvest fungal disease is one of the significant factors that limits the storage period and marketing life of peaches, and even result in serious economic losses worldwide. Biological control using microbial antagonists has been explored as an alternative approach for the management of postharvest disease of fruits. However, there is little information available regarding to the identification the fungal pathogen species that cause the postharvest peach diseases and the potential and mechanisms of using the Bacillus subtilis JK-14 to control postharvest peach diseases. In the present study, a total of six fungal isolates were isolated from peach fruits, and the isolates of Alternaria tenuis and Botrytis cinerea exhibited the highest pathogenicity and virulence on the host of mature peaches. In the culture plates, the strain of B. subtilis JK-14 showed the significant antagonistic activity against the growth of A. tenuis and B. cinerea with the inhibitory rates of 81.32% and 83.45% at 5 days after incubation, respectively. Peach fruits treated with different formulations of B. subtilis JK-14 significantly reduced the mean disease incidences and lesion diameters of A. tenuis and B. cinerea. The greatest mean percent reduction of the disease incidences (81.99% and 71.34%) and lesion diameters (82.80% and 73.57%) of A. tenuis and B. cinerea were obtained at the concentration of 1 × 107 CFU mL−1 (colony forming unit, CFU). Treatment with the strain of B. subtilis JK-14 effectively enhanced the activity of the antioxidant enzymes-superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) in A. tenuis and B. cinerea inoculated peach fruits. As such, the average activities of SOD, POD and CAT were increased by 36.56%, 17.63% and 20.35%, respectively, compared to the sterile water treatment. Our results indicate that the isolates of A. tenuis and B. cinerea are the main pathogens that cause the postharvest peach diseases, and the strain of B. subtilis JK-14 can be considered as an environmentally-safe biological control agent for the management of postharvest fruits diseases. We propose the possible mechanisms of the strain of B. subtilis JK-14 in controlling of postharvest peach diseases.

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Application of Plant Extracts to Control Postharvest Gray Mold and Susceptibility of Apple Fruits to B. cinerea from Different Plant Hosts

Foods ◽

10.3390/foods9101430 ◽

2020 ◽

Vol 9 (10) ◽

pp. 1430 ◽

Cited By ~ 1

Author(s):

Lina Šernaitė ◽

Neringa Rasiukevičiūtė ◽

Alma Valiuškaitė

Keyword(s):

Antifungal Activity ◽

Fungal Pathogens ◽

Plant Protection ◽

Gray Mold ◽

Postharvest Disease ◽

Cinnamon Extract ◽

Wide Range ◽

Plant Hosts ◽

Apple Fruits

Sustainable plant protection can be applied on apples against fungal pathogens such as Botrytis cinerea (which is responsible for gray mold)—a significant global postharvest disease. This pathogen can affect a wide range of hosts; and fruits may have variable susceptibilities to B. cinerea from different plant hosts. New possibilities to control gray mold in food production are under demand due to the emergence of resistance against antifungal agents in fungal pathogens. Cinnamon, pimento, and laurel extracts were previously assessed for antifungal activities under in vitro conditions and were found to have the potential to be effective against postharvest gray mold. Therefore, this study aimed to investigate the antifungal activity of cinnamon, pimento, and laurel extracts in vitro and against postharvest gray mold on apples to determine the susceptibility of apple fruits to B. cinerea from different plant hosts, and to analyze the chemical composition of the extracts. Apples (cv. “Connell Red”) were treated with different concentrations of extracts and inoculated with B. cinerea isolates from apple and strawberry followed by evaluation of in vitro antifungal activity. The results reveal that most of the concentrations of the extracts that were investigated were not efficient enough when assessed in the postharvest assay, despite having demonstrated a high in vitro antifungal effect. Apples were less susceptible to B. cinerea isolated from strawberry. To conclude, cinnamon extract was found to be the most effective against apple gray mold; however, higher concentrations of the extracts are required for the efficient inhibition of B. cinerea in fruits during storage.

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Protein profiling of sickle cell versus control RBC core membrane skeletons by ICAT technology and tandem mass spectrometry

Cellular & Molecular Biology Letters ◽

10.2478/s11658-006-0026-2 ◽

2006 ◽

Vol 11 (3) ◽

Cited By ~ 18

Author(s):

Jose Chou ◽

Pankaj Choudhary ◽

Steven Goodman

Keyword(s):

Mass Spectrometry ◽

Tandem Mass Spectrometry ◽

Sickle Cell ◽

Membrane Skeleton ◽

Protein Profiling ◽

Tandem Mass ◽

Proteomic Approach ◽

Significant Difference ◽

The Mean ◽

Cell Versus

AbstractA proteomic approach using a cleavable ICAT reagent and nano-LC ESI tandem mass spectrometry was used to perform protein profiling of core RBC membrane skeleton proteins between sickle cell patients (SS) and controls (AA), and determine the efficacy of this technology. The data was validated through Peptide/Protein Prophet and protein ratios were calculated through ASAPratio. Through an ANOVA test, it was determined that there is no significant difference in the mean ratios from control populations (AA1/AA2) and sickle cell versus control populations (AA/SS). The mean ratios were not significantly different from 1.0 in either comparison for the core skeleton proteins (α spectrin, β spectrin, band 4.1 and actin). On the natural-log scale, the variation (standard deviation) of the method was determined to be 14.1% and the variation contributed by the samples was 13.8% which together give a total variation of 19.7% in the ratios.

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A Tomato Fruit Rot Caused by Trichothecium roseum in Brazil

Plant Disease ◽

10.1094/pdis-06-11-0464 ◽

2011 ◽

Vol 95 (10) ◽

pp. 1318-1318 ◽

Cited By ~ 6

Author(s):

C. A. Inácio ◽

R. C. Pereira-Carvalho ◽

F. G. A. Morgado ◽

M. E. N. Fonseca ◽

L. S. Boiteux

Keyword(s):

Fungal Pathogens ◽

Tomato Fruit ◽

Its Region ◽

The United States ◽

Fruit Rot ◽

Control Treatment ◽

Postharvest Disease ◽

Potato Dextrose Agar Medium ◽

Trichothecium Roseum ◽

Greenhouse Tomatoes

Fruit rots caused by distinct fungal pathogens are commonly observed on tomatoes (Solanum lycopersicum L.) throughout all major production areas in Brazil. Samples of fruits displaying white mycelial growth associated with a profuse salmon-color sporulation were collected in greenhouse-grown tomatoes in Brasília-DF in February 2011. The isolated fungus displayed pink-to-white colonies containing several conidiophores with conidia. Mycelia displayed hyaline hyphae as much as 4 μm in diameter; conidiophores were simple or branched, 112 to 300 (360) μm long, and 2 to 4 μm wide. Conidia were produced in basipetal chains (frequently clustered), were ellipsoidal to pyriform with oblique and prominent truncate basal scars, two-celled, hyaline, and (14-) 16 to 26 (-28) × (6-) 7 to 10 (-12) μm. These characteristics allocated the specimen to Trichothecium roseum (Pers.). Koch's postulates were fulfilled for one fungal isolate by either spraying 10 intact fruits or by placing a drop of a spore suspension (adjusted to 105 conidia/ml) into three to five wounds created on 10 mature fruits of each of two tomato cultivars (Santa Clara and Dominador) by puncturing each fruit with a sterile needle. Five fruits of each cultivar were treated with sterile water as the mock-inoculated control treatment. Identical symptoms to those of the original fruit were observed only in the T. roseum-inoculated samples 5 to 7 days after using both inoculation procedures. Total DNA was extracted from a pure colony of the fungus growing on potato dextrose agar medium and used as template in PCR assays with the internal transcribed spacer (ITS)-4 (5′-TCCTCCGCTTATTGATATGC-3′) and ITS-5 (5′-GGAAGTAAAAGTCGTAACAAGG-3′) primer pair (2). A single amplicon of approximately 630 bp was observed and directly sequenced. Sequence analysis of the Brazilian isolate (GenBank No. JN081877) indicated identity levels of 99% with T. roseum isolates reported on Leucadendron xanthoconus in South Africa (GenBank No. EU552162) and isolates from strawberry fruits in South Korea (GenBank No. HM355750). However, phylogenetic analysis was unable to discriminate isolates of T. roseum from Passalora (GenBank No. EF432764) and Fusarium (GenBank No. GU183369) isolates, confirming the low genetic variability of the ITS region in Hypocreales (3). T. roseum has been reported to be infecting greenhouse tomatoes in the United States (4) and causing postharvest disease of tomatoes in Argentina (1). To our knowledge, this is the first report of T. roseum infecting greenhouse tomatoes in Brazil. References: (1) G. Dal Bello. Australas. Plant Dis. Notes 3:103, 2008. (2) N. L. Glass and G. C. Donaldson. Appl. Environ. Microbiol. 61:1323, 1995. (3) L. Lombard et al. Stud. Mycol. 66:31, 2010. (4) A. W. Welch, Jr. et al. Plant Dis. Rep. 59:255, 1975.

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Comparative Analysis of Herbaceous and Woody Cell Wall Digestibility by Pathogenic Fungi

Molecules ◽

10.3390/molecules26237220 ◽

2021 ◽

Vol 26 (23) ◽

pp. 7220

Author(s):

Yanhua Dou ◽

Yan Yang ◽

Nitesh Kumar Mund ◽

Yanping Wei ◽

Yisong Liu ◽

...

Keyword(s):

Cell Wall ◽

Plant Cell ◽

Fungal Pathogens ◽

Plant Cell Wall ◽

Apple Tree ◽

Plant Biomass ◽

Genomic Analysis ◽

Pathogenic Fungi ◽

Cell Wall Degrading Enzymes ◽

Enzyme Activity Assay

Fungal pathogens have evolved combinations of plant cell-wall-degrading enzymes (PCWDEs) to deconstruct host plant cell walls (PCWs). An understanding of this process is hoped to create a basis for improving plant biomass conversion efficiency into sustainable biofuels and bioproducts. Here, an approach integrating enzyme activity assay, biomass pretreatment, field emission scanning electron microscopy (FESEM), and genomic analysis of PCWDEs were applied to examine digestibility or degradability of selected woody and herbaceous biomass by pathogenic fungi. Preferred hydrolysis of apple tree branch, rapeseed straw, or wheat straw were observed by the apple-tree-specific pathogen Valsa mali, the rapeseed pathogen Sclerotinia sclerotiorum, and the wheat pathogen Rhizoctonia cerealis, respectively. Delignification by peracetic acid (PAA) pretreatment increased PCW digestibility, and the increase was generally more profound with non-host than host PCW substrates. Hemicellulase pretreatment slightly reduced or had no effect on hemicellulose content in the PCW substrates tested; however, the pretreatment significantly changed hydrolytic preferences of the selected pathogens, indicating a role of hemicellulose branching in PCW digestibility. Cellulose organization appears to also impact digestibility of host PCWs, as reflected by differences in cellulose microfibril organization in woody and herbaceous PCWs and variation in cellulose-binding domain organization in cellulases of pathogenic fungi, which is known to influence enzyme access to cellulose. Taken together, this study highlighted the importance of chemical structure of both hemicelluloses and cellulose in host PCW digestibility by fungal pathogens.

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Study on the Infection Mechanism of Penicillium Digitatum on Postharvest Citrus (Citrus Reticulata Blanco) Based on Transcriptomics

Microorganisms ◽

10.3390/microorganisms7120672 ◽

2019 ◽

Vol 7 (12) ◽

pp. 672 ◽

Cited By ~ 4

Author(s):

Qiya Yang ◽

Xin Qian ◽

Solairaj Dhanasekaran ◽

Nana Adwoa Serwah Boateng ◽

Xueli Yan ◽

...

Keyword(s):

Iron Transport ◽

Bioinformatics Analysis ◽

Control Strategies ◽

Penicillium Digitatum ◽

Citrus Reticulata ◽

Postharvest Diseases ◽

Cell Wall Degrading Enzymes ◽

Infection Mechanism ◽

Citrus Reticulata Blanco ◽

Pathogenic Factors

Penicillium digitatum is one of the most important pathogens known widely to cause postharvest losses of citrus. It is significant to explore its infection mechanism to improve the control technology of postharvest diseases of citrus. This research aimed to study the changes in gene expression of P. digitatum at its early stages of citrus infection by transcriptomics sequencing and bioinformatics analysis in order to explore the molecular mechanism of its infection. The results showed that genes associated with pathogenic factors, such as cell wall degrading enzymes, ethylene, organic acids, and effectors, were significantly up-regulated. Concurrently, genes related to anti-oxidation and iron transport were equally up-regulated at varying degrees. From this study, we demonstrated a simple blueprint for the infection mechanism of P. digitatum in Citrus reticulata Blanco, which provided a new direction for subsequent pathological research and paves the way for developing new control strategies.

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