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Bea Parich oral history transcript
No full textA transcript of an oral history of Bea Parich on her experiences as a school counselor at Century High School
Volatile-mediated inhibitory activity of the biocontrol agent Lysobacter capsici AZ78 as a result of multiple factors interaction
No full textPlant beneficial rhizobacteria are able to inhibit the growth of soilborne phytopathogenic microorganisms through the release of a relevant number of volatile compounds. Based on this, we investigated the ability of the biocontrol agent Lysobacter capsici AZ78 (AZ78) to produce volatile organic compounds (VOCs) that may contribute to its efficacy in controlling soilborne phytopathogenic microorganisms. AZ78 significantly reduced the growth of Pythium ultimum, Rhizoctonia solani and Sclerotinia minor in split Petri dish assays. The GC-MS analysis revealed that AZ78 produce 22 VOCs and most of them were putatively identified as mono- and dialkylated methoxypyrazines. Exposure to 2,5-dimethylpyrazine, 2-ethyl-3-methoxypyrazine and 2-isopropyl-3-methoxypyrazine determined a drastic reduction of Pythium ultimum, Rhizoctonia solani and Sclerotinia minor mycelium growth in split Petri dish assays. However, the discrepancy of the toxicity between the quantity of pyrazines and the AZ78 lead us to further investigate the volatile-mediated inhibitory activity of the biocontrol bacterium. Further experiments revealed the ability of AZ78 cells to produce ammonia that caused the alkalinization of the physically separated culture medium in split Petri dishes assays. Results achieved in this work clearly demonstrated that VOCs, ammonia and the alkalinization of growth medium contribute to the overall inhibitory activity of AZ78 against soilborne phytopathogenic microorganisms
Volatile-mediated inhibitory activity of Rhizobacteria as a result of multiple factors interaction: the case of Lysobacter capsici AZ78
No full textPlant beneficial rhizobacteria may antagonize soilborne plant pathogens by producing a vast array of volatile organic compounds (VOCs). The production of these compounds depends on the medium composition used for bacterial cell growth. Accordingly, Lysobacter capsici AZ78 (AZ78) grown on a protein-rich medium was previously found to emit volatile pyrazines with toxic activity against soilborne phypathogenic fungi and oomycetes. However, the discrepancy between the quantity of pyrazines in the gaseous phase and the minimum quantity needed to achieve inhibition of plant pathogens observed, lead us to further investigate the volatile-mediated inhibitory activity of AZ78. Here, we show that, besides VOCs, AZ78 cells produce ammonia in increased amounts when a protein-rich medium is used for bacterial growth. The production of this volatile compound caused the alkalinization of the physically separated culture medium where Rhizoctonia solani was inoculated subsequently. Results achieved in this work clearly demonstrate that VOC, ammonia and the growth medium alkalinization contribute to the overall inhibitory activity of AZ78 against R. solani. Thus, our findings suggest that the volatile-mediated inhibitory activity of rhizobacteria in protein-rich substrates can be regarded as a result of multiple factors interaction, rather than exclusively VOCs productio
Characterization of the volatilome of Lysobacter capsici AZ78 and its bioactivity against soilborne plant pathogenic fungi and oomycetes
No full textAntibiotics, toxins and volatile organic compounds (VOCs), are some of the plethora of secondary metabolites produced by soil bacteria. These secondary metabolites have been shown to impact on microbial interactions in the soil. Here, we studied the activity and production of VOCs emitted by Lysobacter capsici AZ78, a soil bacterium, which produces non-volatile secondary metabolites toxic against plant pathogens [1]. Recently, it has been shown that VOCs produced by L. capsici DSM 19286 grown in a protein rich medium were highly active against Phytophthora infestans in vitro [2]. In contrast, the VOC-mediated inhibitory effect was attenuated when the strain was grown in a sugar rich medium. Based on these findings, we studied the effect of medium composition on the inhibition activity of L. capsici AZ78 against plant pathogens (e.g. Rhizoctonia solani). In parallel, GC-MS was combined with dynamic headspace (DHS) extraction and thermodesorption to investigate both type and relative amount of VOCs produced by the bacterium grown in media with crescent sugar (glucose) concentrations. Generally, VOC emission profiles exhibited mainly quantitative and not qualitative differences. The chemical group of pyrazines was the most abundant in the volatile profile of L. capsici AZ78 growing in the various media. We additionally conducted experiments using a setup with Petri dishes having two compartments, where we measured the VOCs profile in the one compartment when the bacterium was growing on crescent sugar concentrations in the other. We confirmed the presence of the identified VOCs, thus giving an insight into which compounds could participate in L. capsici AZ78 bioactivity, exhibited during the pathogen inhibition assay. Currently, we are examining the inhibitory effects of the identified compounds against various plant pathogens in vitro, with the aim to understand the mechanisms of VOC-mediated microbe-microbe communications and to select bioactive VOCs for the further development of novel biopesticides
Identification and functional characterization of grapevine volatile organic compounds for the sustainable control of downy mildew
No full textIntroduction
Grapevine (Vitis vinifera) is one of the most widely cultivated fruit crops and is susceptible to various pathogens, such as Plasmopara viticola that causes downy mildew (Gessler et al., 2011). Wild grapevine species are resistant to P. viticola and breeding programs have introduced resistance traits to susceptible cultivars. Plant defence responses are based on different mechanisms and volatile organic compounds (VOCs) play a crucial role in the communication between plants and other organisms. Although the emission of VOCs upon P. viticola inoculation was shown in resistant grapevine genotypes (Algarra Alarcon et al., 2015), the molecular structure and functional role of these molecules in the grapevine defence was not yet investigated. The aim of this study was to identify and functionally characterize VOCs produced by resistant and susceptible grapevine genotypes in response to P. viticola in order to further develop innovative methods for the sustainable control of downy mildew.
Material and methods
The susceptible V. vinifera cultivar Pinot noir and four resistant genotypes (Kober 5BB, SO4, BC4 and Solaris) were grown for three months under greenhouse conditions. Plants were inoculated with a suspension of P. viticola sporangia as previously described (Perazzolli et al., 2012). Downy mildew severity was assessed at seven days after inoculation according to the OIV-452 descriptor and scores from 1 (the most susceptible) to 9 (the totally resistant) were assigned (Bellin et al., 2009). Leaf samples were collected before (T0) and six days (T1) after P. viticola inoculation and five replicates (plants) were analysed for each genotype at each time point. The complete experiment was carried out twice. Each sample was frozen in liquid nitrogen and ground to a fine powder. Leaf powder was weighed into 20 mL headspace vials and analysed by headspace solid phase microextraction-gas chromatography-mass spectrometry (HS-SPME/GC-MS; Weingart, et al. 2012). Eight of the identified VOCs were selected according to their emission profiles and pure compounds were tested against P. viticola by leaf disk assays. Downy mildew development was assessed on leaf disks at one, two and six days post inoculation (dpi) by aniline blue staining.
Results and discussion
VOC profiles measured by HS-SPME/GC-MS analysis were consistent between the two experiments. Terpenes, isoprenoids, aldehydes, alcohols, esters and heterocyclic compounds were found in both experiments in all five tested genotypes . In general, after P. viticola treatment (T1) the abundance of the detected VOCs was higher in resistant genotypes as compared with Pinot noir. Interestingly, the relative induction of VOC levels was found to be significantly higher in resistant genotypes compared to only small changes in Pinot noir. Compared to Pinot noir, treatment with P. viticola resulted in two sesquiterpenes being more abundant in all five resistant genotypes, while three other sesquiterpenes showed a higher abundance in three resistant genotypes (BC4, Kober 5BB and Solaris). At T1, Kober 5BB and Solaris showed also a higher abundance of one heterocyclic compound and one isoprenoid as compared with Pinot noir. Finally, the abundance of a C5 aldehyde was higher in Kober 5BB as compared with Pinot noir at T1. These eight pure VOCs were tested against P. viticola in liquid suspension and in air volume. The eight VOCs impaired the development of downy mildew symptoms at dosages that ranged from 0.1 to 10.0 g/L in liquid suspension. However, five of them also showed severe phytotoxic effects on leaf disks at the dosage of 10.0 g/L. Four pure VOCs (one isoprenoid, one alcohol, one C5 aldehyde and one heterocyclic compound) significantly reduced downy mildew symptoms at the dosage of 20.0 mg/L in air volume, when each VOC was applied to a filter paper disk and placed on the lid of the Petri dish.
Microscope observations with aniline blue staining revealed marked morphological changes in VOC-treated leaf disks after P. viticola inoculation. The number of pathogen structures was reduced in leaf disks treated with one isoprenoid, one alcohol and one heterocyclic compound as compared to control disks at one, two and six dpi. Moreover, no P. viticola structures were visible on leaf disks treated with the C5 aldehyde. This aldehyde and one isoprenoid were also able to reduce the diameter of P. viticola sporangia.
In conclusion, downy mildew increased the production of VOCs (terpenes, isoprenoid, alcohols, aldehydes and heterocyclic compounds) in resistant but not in the susceptible genotype and these molecules are associated to the activation of grapevine defence mechanisms. Moreover, VOCs of resistant genotypes have the porential to contribute to grapevine resistance and significantly reduced downy mildew symptoms on susceptible leaf disks, indicating that they can be further developed as sustainable control molecules
Volatile organic compounds from Lysobacter capsici AZ78 as potential candidates for biological control of soilborne plant pathogens
No full textThe genus Lysobacter includes several bacterial species which show potential for being used in biological control of plant diseases. It was shown recently that several Lysobacter type strains produce volatile organic compounds (VOCs) which controlled the growth of Phytophthora infestans in vitro when the bacteria were grown on a protein rich medium. In the present study, Lysobacter capsici AZ78 (AZ78) has been tested for its potential to produce VOCs that may contribute to the bioactivity against soilborne plant pathogens. To this end, split Petri dish assays of bacterial cultures have been combined with GC-MS measurements with the aim to reveal the identity of the VOCs which inhibit the growth of Pythium ultimum Rhizoctonia solani, and Sclerotinia minor. While AZ78 completely suppressed the growth of P. ultimum and S. minor, the growth of R. solani was still reduced significantly. The GC-MS analysis revealed 22 VOCs to be produced by AZ78, the majority of which were (putatively) identified as mono- and dialkylated methoxypyrazines. Based on additional cultivation and GC-MS experiments, 2,5-dimethylpyrazine, 2-ethyl-3-methoxypyrazine and 2-isopropyl-3-methoxypyrazine were selected as presumable bioactive compounds. Further bioassays employing indirect exposure to standard solutions (1-10 mg per Petri dish) of the synthetic compounds via the gas phase, revealed that each of these pyrazines was able to suppress the growth of the pathogens under investigation. The results of this study highlight the possible future implementation of pyrazine derivatives in the control of soilborne plant diseases and further support the biocontrol potential of L. capsici AZ78
Identification of volatile organic compounds in different grapevine genotypes after inoculation with Plasmopara viticola
No full textThe grapevine Vitis vinifera cv Pinot noir is susceptible to several pathogens including Plasmopara viticola that is the causal agent of downy mildew [1]. Hybrids of V. berlandieri and V. riparia (SO4 and Kober 5BB) and hybrids of Muscadinia rotundifolia and V. vinifera (BC4) and others such as Solaris are resistant or tolerant to downy mildew. It has been demonstrated recently [2] that resistant in vitro hybrids SO4 and Kober 5BB emit volatile organic compounds (VOCs) in response to P. viticola infection. In particular, the most interesting class of VOCs constitutes terpenoids (mono- and sesquiterpenes) emitted by the resistant cultivars, whereas for Pinot noir no terpenes have been detected under the tested conditions.
In the present study we have used gas chromatography coupled with mass spectrometry (GC-MS) to study in more detail the chemical identity of the compounds produced by selected plants of the five genotypes Pinot noir, Kober 5BB, SO4, BC4 and Solaris. All the genotypes were cultured in the greenhouse and leaves were harvested immediately (0 dpi) and six (6 dpi) days after the inoculation with P. viticola. All samples were immediately frozen and homogenized under cooled conditions. VOCs were extracted by using solid phase microextraction (SPME) and analyzed by GC-MS. Mass spectral deconvolution and annotation / identification of volatile compounds was based on comparison of mass spectra and retention indices with reference values and performed by Metabolite Detector software [3].
Preliminary results showed increased levels compared to day zero of sesquiterpenes in resistant cultivars six days after inoculation, demonstrating that terpenes could play an important role in plant resistance against downy mildew in resistant genotypes.
[1] C. Gessler, I. Pertot, M. Perazzolli. (2011). Plasmopara viticola: a review of knowledge on downy mildew of grapevine and effective disease management. Phytopathologia Mediterranea, 50, p. 3-44
[2] A. Algarra Alarcon, V. Lazazzara, L. Cappellin, P. L. Bianchedi, R. Schuhmacher, G. Wohlfahrt, I. Pertot, F. Biasioli and M. Perazzolli. (2015). Emission of volatile sesquiterpenes and monoterpenes in grapevine genotypes following Plasmopara viticola inoculation in vitro. Journal of Mass Spectrometry, 50, p. 1013–1022
[3] K. Hiller, J. Hangebrauk, C. Jäger, J. Spura, K. Schreiber, D. Schomburg. (2009) MetaboliteDetector: comprehensive analysis tool for targeted and nontargeted GC/MS based metabolome analysis. Analytical Chemistry, 81(9), p. 3429-343
Downy mildew symptoms on grapevines can be reduced by volatile organic compounds of resistant genotypes
No full textVolatile organic compounds (VOCs) play a crucial role in the communication of plants with other
organisms and are possible mediators of plant defence against phytopathogens. Although the role
of non-volatile secondary metabolites has been largely characterised in resistant genotypes, the
contribution of VOCs to grapevine defence mechanisms against downy mildew (caused by Plasmopara
viticola) has not yet been investigated. In this study, more than 50 VOCs from grapevine leaves
were annotated/identifed by headspace-solid-phase microextraction gas chromatography-mass
spectrometry analysis. Following P. viticola inoculation, the abundance of most of these VOCs was
higher in resistant (BC4, Kober 5BB, SO4 and Solaris) than in susceptible (Pinot noir) genotypes.
The post-inoculation mechanism included the accumulation of 2-ethylfuran, 2-phenylethanol,
β-caryophyllene, β-cyclocitral, β-selinene and trans-2-pentenal, which all demonstrated inhibitory
activities against downy mildew infections in water suspensions. Moreover, the development of
downy mildew symptoms was reduced on leaf disks of susceptible grapevines exposed to air treated
with 2-ethylfuran, 2-phenylethanol, β-cyclocitral or trans-2-pentenal, indicating the efcacy of these
VOCs against P. viticola in receiver plant tissues. Our data suggest that VOCs contribute to the defence
mechanisms of resistant grapevines and that they may inhibit the development of downy mildew
symptoms on both emitting and receiving tissues
Identification of volatile organic compounds emitted by different grapevine genotypes in response to downy mildew infection
No full textVitis vinifera is susceptible to several pathogens including Plasmopara viticola, the causal agent of downy mildew. American grapevine species are resistant or tolerant to P. viticola and breeding programs have introduced resistance traits to susceptible cultivars. Although grapevine resistance to P. viticola has been widely characterized in resistant genotypes, the possible contribute of volatile organic compounds (VOCs) was not yet investigated. The aim of this work was the characterization of VOCs produced by resistant and susceptible genotypes in response to P. viticola inoculation, in order to identify VOCs associated to grapevine resistance. The susceptible V. vinifera cultivar Pinot noir, and the resistant genotypes Kober 5BB, SO4, BC4 and Solaris were grown under greenhouse conditions and they were subsequently inoculated with P. viticola. Leaves were harvested immediately before (0 dpi) and six days (6 dpi) after inoculation, and the lower disease severity in resistant genotypes as compared with Pinot noir was confirmed. A solid-phase microextraction-gas chromatography-mass spectrometry approach (SPME/GC-MS) was used to analyze VOCs emitted by the five genotypes studied. GC-MS chromatograms showed specific VOC emission profiles of the four resistant genotypes as compared with Pinot noir at 6 dpi. VOCs specifically emitted by resistant genotypes were then selected, and pure compounds were tested against P. viticola by leaf discs assays. Particularly, three sesquiterpenes, two aldehydes and one heterocyclic compound significantly reduced downy mildew severity on Pinot noir, demonstrating that VOCs could play an important role in the resistance against downy mildew by direct toxicity against P. viticola
Going Beyond Counting First Authors in Author Co-citation Analysis
No full textThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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