Single berry sampling reveals novel insights into transcriptomic changes induced by leafroll virus infections in grapevine (V. vinifera)

2020 ◽  
Author(s):  
Sana Ghaffari ◽  
Jean Sébastien Reynard ◽  
Markus Rienth

Abstract Background Grapevine leafroll associated viruses (GLRaV), the causing agents of grapevine leafroll disease (GLD) are amongst the most devastating graft and vector transmitted pathogens in viticulture and responsible for important economic losses in the wine industry. Reported fruit alterations caused by GLD consist in a delay of ripening, a reduction in anthocyanins, aroma compounds and sugar concentration. The molecular interactions underlying the quality deteriorating effects are not well understood so far. The few conducted molecular studies on infected berries did associate the lack of anthocyanin and sugar content with a repression of key genes of the respective pathways. However, sampling protocols in such studies never accounted for berry heterogeneity and potential virus induced phenological shifts. Thus, such commonly used sampling strategies could have introduced unquantifiable biases in gene expression studies which would impede the discovery of novel molecular information. The aim of the present study was to investigate the effects of GLRaV-1 and GLRaV-1&3 infections on berry physiology of Pinot Noir vines. Berries of different treatments were individually sampled and analyzed for sugar and organic acids. According to this biochemical analysis 2 homogenous stages were reconstituted to circumvent berry heterogeneity and compensate for virus induced phenological shifts. RNA of 18 reconstituted samples (2 stages and 3 treatments) was extracted, sequenced and analyzed for differentially expressed genes (DEGs). Results A total of 2136 transcripts were modulated by GLRaV-1 and GLRaV-1&3 infections. Interestingly the transcriptome of the early ripening berry was much more affected as the later ripening stage. Several pathways related to abiotic and biotic stress, defense mechanism as well as plant immunity showed a virus-induced upregulation in dependence to ripening stage and infection severity. Surprisingly, the previously reported repression of anthocyanin biosynthesis and sugar metabolism could not be confirmed by gene expression. This illustrates that the main damaging effect on GLRaV infection is rather related to a phenological shift than to a direct impact on metabolism. Conclusions The here reported results give new insight in the mechanism of leafroll infection and emphasize the importance of the sampling protocol of molecular studies investigating berry metabolism.

2021 ◽  
pp. 1-17
Author(s):  
Chau Thi Thu Nguyen ◽  
Ji Hyun Lee ◽  
Phu Tri Tran

BACKGROUND: The sweetness of highbush blueberries is a sensory quality standard for consumers. Changes in metabolites and expression of associated genes induce variation in the sensory quality of blueberries. OBJECTIVE: This work investigated the ripening mechanism of blueberries by measuring changes in organic acids and sugar content as well as the gene expression levels associated with sugar accumulation, such as β amylase, invertase, sucrose phosphate synthase, and tonoplast monosaccharide transporter genes, before and after harvesting. METHODS: We used three different blueberry cultivars (Duke, Sierra, and Elizabeth) that we harvested at two ripening stages in a randomized complete block design and then kept at room temperature or 4°C for 7 days. The organic acids and sugar content were measured via HPLC, and gene expression levels were analyzed by qPCR. RESULTS: Cultivars, ripening stage, and storage conditions affected the synthesis of sugars and degradation of organic acids to determine the sweetness in blueberries and the expression of tagged genes and analyses of compounds involved in the metabolic mechanism. CONCLUSIONS: The results provide insights into the mechanism underlying the ripening and the postharvest quality. This study may support the selection of suitable blueberry cultivars that meet customer demand.


2021 ◽  
Vol 12 ◽  
Author(s):  
Demetrio Marcianò ◽  
Valentina Ricciardi ◽  
Elena Marone Fassolo ◽  
Alessandro Passera ◽  
Piero Attilio Bianco ◽  
...  

Downy mildew, caused by the oomycete Plasmopara viticola, is one of the diseases causing the most severe economic losses to grapevine (Vitis vinifera) production. To date, the application of fungicides is the most efficient method to control the pathogen and the implementation of novel and sustainable disease control methods is a major challenge. RNA interference (RNAi) represents a novel biotechnological tool with a great potential for controlling fungal pathogens. Recently, a candidate susceptibility gene (VviLBDIf7) to downy mildew has been identified in V. vinifera. In this work, the efficacy of RNAi triggered by exogenous double-stranded RNA (dsRNA) in controlling P. viticola infections has been assessed in a highly susceptible grapevine cultivar (Pinot noir) by knocking down VviLBDIf7 gene. The effects of dsRNA treatment on this target gene were assessed by evaluating gene expression, disease severity, and development of vegetative and reproductive structures of P. viticola in the leaf tissues. Furthermore, the effects of dsRNA treatment on off-target (EF1α, GAPDH, PEPC, and PEPCK) and jasmonic acid metabolism (COI1) genes have been evaluated. Exogenous application of dsRNA led to significant reductions both in VviLBDIf7 gene expression, 5 days after the treatment, and in the disease severity when artificial inoculation was carried out 7 days after dsRNA treatments. The pathogen showed clear alterations to both vegetative (hyphae and haustoria) and reproductive structures (sporangiophores) that resulted in stunted growth and reduced sporulation. Treatment with dsRNA showed signatures of systemic activity and no deleterious off-target effects. These results demonstrated the potential of RNAi for silencing susceptibility factors in grapevine as a sustainable strategy for pathogen control, underlying the possibility to adopt this promising biotechnological tool in disease management strategies.


2020 ◽  
Vol 139 ◽  
pp. 153-160
Author(s):  
S Peeralil ◽  
TC Joseph ◽  
V Murugadas ◽  
PG Akhilnath ◽  
VN Sreejith ◽  
...  

Luminescent Vibrio harveyi is common in sea and estuarine waters. It produces several virulence factors and negatively affects larval penaeid shrimp in hatcheries, resulting in severe economic losses to shrimp aquaculture. Although V. harveyi is an important pathogen of shrimp, its pathogenicity mechanisms have yet to be completely elucidated. In the present study, isolates of V. harveyi were isolated and characterized from diseased Penaeus monodon postlarvae from hatcheries in Kerala, India, from September to December 2016. All 23 tested isolates were positive for lipase, phospholipase, caseinase, gelatinase and chitinase activity, and 3 of the isolates (MFB32, MFB71 and MFB68) showed potential for significant biofilm formation. Based on the presence of virulence genes, the isolates of V. harveyi were grouped into 6 genotypes, predominated by vhpA+ flaB+ ser+ vhh1- luxR+ vopD- vcrD+ vscN-. One isolate from each genotype was randomly selected for in vivo virulence experiments, and the LD50 ranged from 1.7 ± 0.5 × 103 to 4.1 ± 0.1 × 105 CFU ml-1. The expression of genes during the infection in postlarvae was high in 2 of the isolates (MFB12 and MFB32), consistent with the result of the challenge test. However, in MFB19, even though all genes tested were present, their expression level was very low and likely contributed to its lack of virulence. Because of the significant variation in gene expression, the presence of virulence genes alone cannot be used as a marker for pathogenicity of V. harveyi.


2021 ◽  
Vol 22 (2) ◽  
pp. 522
Author(s):  
Noreen Falak ◽  
Qari Muhammad Imran ◽  
Adil Hussain ◽  
Byung-Wook Yun

Plants are in continuous conflict with the environmental constraints and their sessile nature demands a fine-tuned, well-designed defense mechanism that can cope with a multitude of biotic and abiotic assaults. Therefore, plants have developed innate immunity, R-gene-mediated resistance, and systemic acquired resistance to ensure their survival. Transcription factors (TFs) are among the most important genetic components for the regulation of gene expression and several other biological processes. They bind to specific sequences in the DNA called transcription factor binding sites (TFBSs) that are present in the regulatory regions of genes. Depending on the environmental conditions, TFs can either enhance or suppress transcriptional processes. In the last couple of decades, nitric oxide (NO) emerged as a crucial molecule for signaling and regulating biological processes. Here, we have overviewed the plant defense system, the role of TFs in mediating the defense response, and that how NO can manipulate transcriptional changes including direct post-translational modifications of TFs. We also propose that NO might regulate gene expression by regulating the recruitment of RNA polymerase during transcription.


Author(s):  
Shiyi Tian ◽  
Jue Wang ◽  
Jing Wang ◽  
Weiyun Zhu

Recently, we have proved that the early-life galactooligosaccharides (GOS) intervention could improve the colonic function by altering the bacterial composition in the suckling piglets. However, whether the early-life GOS (ELG) intervention could have a long influence of the colonic microbiota, and the ELG and post-weaning GOS (PWG) combined intervention would have an interaction effect on maintaining colonic health in weaning piglets remain to be explored. Thus in this study, we illustrated the differential effect of ELG and PWG intervention on colonic microbiota and colonic function of weaning piglets. Our results showed that both the ELG and PWG intervention decreased the diarrhea frequency of weaning piglets, while the PWG intervention increased colonic indexes. After 16S rRNA MiSeq sequencing of gut bacteria belonged to colonic niches (mucosa and digesta), the PWG increased the α-diversity of colonic mucosal bacteria was revealed. In addition, we found both the ELG and PWG intervention enriched the abundance of short chain fatty acids (SCFAs) producer in different colonic niches and increased total SCFAs concentrations in colonic digesta. These changes selectively modulated the mRNA expression of pattern recognition receptors and barrier proteins in the colonic mucosa. Of note, the combined effect of ELG and PWG effectively enhanced colonic SCFAs producer enrichment and up-regulated the butyrate concentration. Meanwhile, the gene expression of MyD88-NFκB signaling and the pro-inflammatory cytokines contents were markedly reduced under the combined effect of ELG and PWG. Importance Reducing the disorders of gut ecosystem is an effective way to relieve weaning stresses of piglets and save economic losses in the modern swine industry. To this end, prebiotics were often added in diet during the weaning transition. In present study, we demonstrated that the ELG and PWG intervention had shown different effects on the bacterial composition of different colonic niches and colonic function in the weaning piglets. Especially under the combined effect of ELG and PWG intervention, the gene expression of MyD88-NFκB signaling and the contents of pro-inflammation cytokines decreased with the increasing concentration of butyrate, which is one of the important microbial metabolites in the colon of weaning piglets. These findings further provided new insights into nutritional interventions to alleviate intestinal ecosystem dysbiosis and gut dysfunction in the piglets during the weaning transition.


Genes ◽  
2018 ◽  
Vol 9 (12) ◽  
pp. 591
Author(s):  
Tsung-Lin Tsai ◽  
Chen-Chang Su ◽  
Ching-Chi Hsieh ◽  
Chao-Nan Lin ◽  
Hui-Wen Chang ◽  
...  

In 2013, the outbreak of porcine epidemic diarrhea (PED) in Taiwan caused serious economic losses. In this study, we examined whether the variations of the cis-acting elements between the porcine epidemic diarrhea virus (PEDV) Taiwan (TW) strain and the prototype strain CV777 alter gene expression. For this aim, we analyzed the variations of the cis-acting elements in the 5’ and 3’ untranslated regions (UTRs) between the PEDV TW, CV777, and other reference strains. We also determined the previously unidentified transcription regulatory sequence (TRS), a sequence motif required for coronavirus transcription, and found that a nucleotide deletion in the TW strain, in comparison with CV777 strain, immediately downstream of the leader core sequence alters the identity between the leader TRS and the body TRS. Functional analyses using coronavirus defective interfering (DI) RNA revealed that such variations in cis-acting elements for the TW strain compared with the CV777 strain have an influence on the efficiency of gene expression. The current data show for the first time the evolution of PEDV in terms of cis-acting elements and their effects on gene expression, and thus may contribute to our understanding of recent PED outbreaks worldwide.


2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
ROSSANA FRANZIN ◽  
Fabio Sallustio ◽  
Claudia Curci ◽  
Simona Simone ◽  
Angela Picerno ◽  
...  

Abstract Background and Aims Cisplatin, is a nonspecific cytotoxic agent that primarily interferes with cellular DNA replication and the cell cycle, nevertheless it lacks tumor selectivity and acts also in normal cells. The most serious adverse reaction of cisplatin is Acute Kidney Injury (AKI), limiting its use and efficacy in chemotherapy. Cisplatin nephrotoxicity is observed in more than 30% of older patients, however the mechanism of nephrotoxicity remains unclear and specific preventive measures are not available. Today, there is an urgent need for specific nephroprotective strategies to be used during cisplatin chemotherapy. Recently, we found that tubular stem/progenitor cells (tARPC) are able to protect the tubular epithelial (RPTEC) from cisplatin induced injury, preserving their proliferation and inhibiting apoptosis. The aim of this study was to identify the molecular mechanisms involved in tARPC-mediated resistance to cisplatin. Method Co-cultures of RPTEC cells and tARPCs were exposed to cisplatin (2.5 µM) for 6 h and then kept in culture for 96 h. Gene expression profile was obtained from tARPCs and RPTECs by Agilent SurePrint G3 Human Gene Expression Microarrays. Genespring and R software were used for the analysis. Gene expression data were validated by Real-time PCR. Extracellular vesicles were isolated from cell culture supernatant by miRCURY Exosome Cell/Urine/CSF Kit (Qiagen) and RNA contained in extracellular vesicles was purified, analyzed in quality by Bioanalyzer (RNA nano) and evaluated by qPCR. The BrdU assay and caspase 3 were used to measure proliferation and apoptosis levels. Immunohistochemical expression of activated caspase-3 was used as a marker of apoptosis in RPTECs. Results By a whole-genome gene expression analysis, we found 107 genes specifically modulated by RPTECs in response to cisplatin and, among these, 30 genes induced by ARPCs following the cisplatin damage. In particular, we found a strong upregulation of the CYP1B1 gene (false discovery rate corrected p value <0.05; fold change=4,1). The qPCR confirmed the increase in CYP1B1 levels in the co-cultures with respect to the respective basal conditions (p <0.05). Interestingly, the CYP1B1 mRNA was also enveloped in Extracellular Vesicles released in the cell co-culture media by tARPC and RPTEC after cisplatin exposition. The CYP1B1 gene encodes a member of the cytochrome P450 superfamily of enzymes and the produced enzyme metabolizes procarcinogens, such as polycyclic aromatic hydrocarbons. CYP1B1 has been shown to be active within tumors and is also capable of metabolizing a structurally diverse range of anticancer drugs. It is responsible for the resistance to docetaxel, cisplatin, tamoxifen and nucleoside analogues. CYP1B1 is involved in the detoxification of the body by various exogenous toxic agents, including cisplatin. We found that CYP1B1 gene was expressed at low levels in RPTECs and in cisplatin-damaged RPTECs. Moreover, 96 h days after 2.5 μM exposure to cisplatin, RPTECs reduced the proliferation and underwent in apoptosis, as showed by caspase 3. However, in co-culture with ARPCs, ARPC cellular and extracellular vesicles CYP1B1 gene expression significantly increased, the apoptotic process was stopped and RPTECs increased their proliferation rate. These data support the hypothesis that ARPCs are sensor of cisplatin damaged-RPTEC and confers cisplatin resistance by transferring CYP1B1 gene in extracellular vesicles. Conclusion This is the first evidence of a cisplatin-induced overexpression of CYP1b1 in renal epithelial cells as a defense mechanism against cisplatin toxicity. This is consistent with our previous data showing that renal progenitors are resistant to cisplatin. The findings may have biological and clinical significance in terms of their implications in cellular communications and potential use of CYP1B1 as biomarkers for AKI induced by cisplatin or as protective agent.


2021 ◽  
Author(s):  
Taylor Reiter ◽  
Rachel Montpetit ◽  
Ron Runnebaum ◽  
C. Titus Brown ◽  
Ben Montpetit

AbstractGrapes grown in a particular geographic region often produce wines with consistent characteristics, suggesting there are site-specific factors driving recurrent fermentation outcomes. However, our understanding of the relationship between site-specific factors, microbial metabolism, and wine fermentation outcomes are not well understood. Here, we used differences in Saccharomyces cerevisiae gene expression as a biosensor for differences among Pinot noir fermentations from 15 vineyard sites. We profiled time series gene expression patterns of primary fermentations, but fermentations proceeded at different rates, making analyzes of these data with conventional differential expression tools difficult. This led us to develop a novel approach that combines diffusion mapping with continuous differential expression analysis. Using this method, we identified vineyard specific deviations in gene expression, including changes in gene expression correlated with the activity of the non-Saccharomyces yeast Hanseniaspora uvarum, as well as with initial nitrogen concentrations in grape musts. These results highlight novel relationships between site-specific variables and Saccharomyces cerevisiae gene expression that are linked to repeated wine fermentation outcomes. In addition, we demonstrate that our analysis approach can extract biologically relevant gene expression patterns in other contexts (e.g., hypoxic response of Saccharomyces cerevisiae), indicating that this approach offers a general method for investigating asynchronous time series gene expression data.ImportanceWhile it is generally accepted that foods, in particular wine, possess sensory characteristics associated with or derived from their place of origin, we lack knowledge of the biotic and abiotic factors central to this phenomenon. We have used Saccharomyces cerevisiae gene expression as a biosensor to capture differences in fermentations of Pinot noir grapes from 15 vineyards across two vintages. We find that gene expression by non-Saccharomyces yeasts and initial nitrogen content in the grape must correlates with differences in gene expression among fermentations from these vintages. These findings highlight important relationships between site-specific variables and gene expression that can be used to understand, or possibly modify, wine fermentation outcomes. Our work also provides a novel analysis method for investigating asynchronous gene expression data sets that is able to reveal both global shifts and subtle differences in gene expression due to varied cell – environment interactions.


2020 ◽  
Vol 7 (16) ◽  
pp. 193-208
Author(s):  
Catarina de Bortoli Munhae ◽  
Vanderlei Geraldo Martins ◽  
Cintia Martins ◽  
Odair Correa Bueno

Eurhizococcus brasiliensis Wille, 1922 (Hemiptera: Margarodidae), also known as ground pearl, is a root cochineal found in viticultures in several Brazilian states. It causes economic losses and is difficult to control. Given the difficulty in morphologically identifying the group, the current study aims to trace the ground pearl molecular profile by assuming that there is more than one species of this cochineal in the country. Ground pearl samples were collected in Rio Grande do Sul (Flores da Cunha and Pinto Bandeira Counties), Santa Catarina (Videira and Pinheiro Preto Counties), São Paulo (São Roque, São Miguel Arcanjo, Louveira and Indaiatuba Counties) and Pernambuco States (Petrolina County). Subsequently, the COI, ITS and 28S genes as well as the 18 RAPD primers were analyzed. The COI and ITS gene primers were not successfully amplified. The 28S gene analysis formed two clades and the RAPD analysis formed three groups. The genetic and geographic distances among the herein analyzed samples were positive. Results allow inferring the existence of at least two ground pearl groups in the analyzed areas. Two hypotheses are presented in order to explain the cochineal groups' separation: (i) the founder effect action and (ii) the regional species independence. However, it is necessary to conduct complementary molecular studies using other genes as well as studies on the group taxonomy and basic biology to explain the ground pearl phylogeny.


2020 ◽  
Author(s):  
Lida Hashemi ◽  
Ahmad Reza Golparvar ◽  
Mehdi Nasr Esfahani ◽  
Maryam Golabadi

AbstractPhytophthora melonis is the causal agent of damping-off or crown rot, one of the most destructive cucumber diseases that causes severe economic losses in Iran and some other parts of the world. Despite intense research efforts made in the past years, no permanent cure currently exists for this disease. With the aim to understand the molecular mechanisms of defense against P. melonis, root collars and leaves of four cucumber genotypes consisting of resistant Ramezz; moderately resistant Baby and very susceptible Mini 6-23 and Extrem, were monitored for quantitative gene expression analysis of five antifungal and/or anti-oomycete genes (CsWRKY20, CsLecRK6.1, PR3, PR1-1a and LOX1) at three points after inoculation with P. melonis. The gene expression analysis indicated that P. melonis strongly enhanced the expression of these genes after inoculation in both leaves and root collars. Further, not only the transcript levels of these genes were significantly higher in the resistant and moderately resistance genotypes, but also the time point of the highest relative expression ratio for the five genes was different in the four cucumber genotypes. CsWRKY20 and PR3 showed the maximum expression in Ramezz at 48 hours post inoculation (hpi) while CsLecRK6.1, and LOX1 showed the highest expression at 72 hpi. In addition, PR1-1a showed the maximum expression in the Baby at 72 hpi. Root collars responded faster than leaves and some responses were more strongly up-regulated in root collars than in leaves. The genes found to be involved in disease resistance in two different organs of cucumber after pathogen infection. The results suggest that increased expression of these genes led to activation of defense pathways and could be responsible for a reduced P. melonis colonization capacity in Ramezz and Baby. Overall, this work represents a valuable resource for future functional genomics studies to unravel the molecular mechanisms of C. sativus- P. melonis interaction.


Sign in / Sign up

Export Citation Format

Share Document