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Paavanen-Huhtala, Sari; Avikainen, Hanna; Yli-Mattila, Tapani

doi:10.1023/a:1008734519852

Cited by 34 publications

(8 citation statements)

References 19 publications

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“…into separate ITS groups, but no difference was observed among strains belonging to each group. Paavanen-Huhtala et al [13] also reported that the 28S, ITS1-5.8S-ITS2 and 18S regions amplified from 17 C. rosea strains, including J1446, were found to share identical sequences. Similarly, the partial D1/D2 and ITS1-5.8S-ITS2 regions amplified from C. rosea strain ACM941 and 88-710 in this work were 100% identical.…”

Section: Discussionmentioning

confidence: 99%

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A Universally Primed-Polymerase Chain Reaction (UP-PCR) Marker to Discriminate Clonostachys rosea ACM941 from Related Strains

Demissie

Brown²,

Loewen

2019

JoF

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Clonostachys rosea strain ACM941 is an effective biocontrol agent against several crop diseases including Fusarium head blight. In anticipation of its increased relevance going forward, the development of a reliable DNA-based molecular marker to track it is essential. Universally primed-PCR (UP-PCR) has been used successfully to differentiate other C. rosea strains. Herein, the development of a UP-PCR marker for ACM941 is described. A combination of two primers (AS15 and L45) produced a ~450 bp fragment that was unique to ACM941 compared to other commercial biocontrol agents. Primers subsequently designed based on the obtained fragment also produced a similarly unique band from ACM941 alone. BLAST analysis of the amplified sequence did not yield any homologous sequence in available online databases or within the closely related C. rosea IK726 and CBS125111 strains’ genomes. The specificity of this marker for ACM941 was validated against ten additional C. rosea strains isolated from Canada, with ACM941 producing the brightest band. Taken together, these results imply that the UP-PCR primers AS15 and L45 and the amplified fragment can be used to detect and monitor the ACM941 strain after its release into the environment.

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Section: Discussionmentioning

confidence: 99%

“…In contrast, Bulat et al [12] screened only seven UP-PCR primers and identified a strain-specific polymorphic DNA fragment in C. rosea strain GR5. Paavanen-Huhtala et al [13] also used UP-PCR to develop a strain-specific marker for isolate J1446.…”

Section: Introductionmentioning

confidence: 99%

A Universally Primed-Polymerase Chain Reaction (UP-PCR) Marker to Discriminate Clonostachys rosea ACM941 from Related Strains

Demissie

Brown²,

Loewen

2019

JoF

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“…In fact, TaqMan qPCR is also used for the detection and quantification of potential BCAs such as T. harzianum or Paecilomyces lilacinus , which helped to better understand their ability to colonize and suppress pathogens in the field (Atkins et al, 2005; López-Mondéjar et al, 2010). To our knowledge, only strain-specific markers are currently available for C. rosea , which were previously developed for the commercialized BCA C. rosea f. catenulata strain J1446 (Paavanen-Huhtala et al, 2000) and for C. rosea f. rosea strain GR5 (Bulat et al, 2000). The authors used universally primed-PCR (UP-PCR) and randomly amplified polymorphic DNA (RAPD) techniques to identify sequence-characterized amplified region (SCAR) markers.…”

Section: Discussionmentioning

confidence: 99%

TaqMan qPCR for Quantification of Clonostachys rosea Used as a Biological Control Agent Against Fusarium graminearum

et al. 2019

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Clonostachys rosea is a biological control agent against Fusarium graminearum in small grain cereals and maize. Infections with F. graminearum do not only reduce the yield but, due to the production of mycotoxins, also affect the entire value chain of food and feed. In addition, production of other secondary metabolites such as hydrophobins, also known as gushing inducers, may cause quality challenges for the malting and brewing industry. Sustainable disease control strategies using C. rosea are treatment of infected residues of the previous crop, direct treatment of the actual cereal crop or post-harvest treatment during malting processes. Follow-up of growth and survival of biocontrol organisms during these different stages is of crucial importance. In the current study, we developed a quantitative real-time PCR detection method that amends the currently available culture-dependent techniques by using TaqMan chemistry with a highly specific primer and probe set, targeting the actin gene. We established a sensitive assay that detects the biological control agent down to 100 genome copies per reaction, with PCR efficiencies between 90 and 100%. The specificity of the assay was confirmed against a panel of 30 fungal and 3 bacterial species including 12 members of the Fusarium head blight complex and DNA of barley, maize and wheat. The DNA of C. rosea was detected in Fusarium -infected maize crop residues that were either treated in the laboratory or in the field with C. rosea and followed its DNA throughout the barley malting process to estimate its growth during grain germination. We used a standardized DNA extraction protocol and showed that C. rosea can be quantified in different sample matrices. This method will enable the monitoring of C. rosea during experiments studying the biological control of F. graminearum on cereal crop residues and on cereal grains and will thus contribute to the development of a new disease control strategy.

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“…could lead to underestimating the actual population of the fungus (Black and Foarde, 2007;Martin-Laurent et al, 2001). SCAR markers derived from an RAPD assay have been used to detect different organisms at isolate or strain levels, such as Gliocladium catenulatum (Paavanen-Huhtala et al, 2000) and Fusarium oxysporum (del Mar Jiménez-Gascó and Jiménez-Díaz, 2003;Pasquali et al, 2006;Lievens et al, 2008). In addition, SCAR markers derived from an RAPD assay have been also previously reported and used to detect different Trichoderma species or strains in different samples with varying detection limits.…”

Section: Discussionmentioning

confidence: 99%

Designing a SCAR molecular marker for monitoring Trichoderma cf. harzianum in experimental communities

Perez

Verdejo

Gondim-Porto

et al. 2014

J. Zhejiang Univ. Sci. B

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Several species of the fungal genus Trichoderma establish biological interactions with various micro-and macro-organisms. Some of these interactions are relevant in ecological terms and in biotechnological applications, such as biocontrol, where Trichoderma could be considered as an invasive species that colonizes a recipient community. The success of this invasion depends on multiple factors, which can be assayed using experimental communities as study models. Therefore, the aim of this work is to develop a species-specific sequence-characterized amplified region (SCAR) marker to monitor the colonization and growth of T. cf. harzianum when it invades experimental communities. For this study, 16 randomly amplified polymorphic DNA (RAPD) primers of 10-mer were used to generate polymorphic patterns, one of which generated a band present only in strains of T. cf. harzianum. This band was cloned, sequenced, and five primers of 20-23 mer were designed. Primer pairs 2F2/2R2 and 2F2/2R3 successfully and specifically amplified fragments of 278 and 448 bp from the T. cf. harzianum BpT10a strain DNA, respectively. Both primer pairs were also tested against the DNA from 14 strains of T. cf. harzianum and several strains of different fungal genera as specificity controls. Only the DNA from the strains of T. cf. harzianum was successfully amplified. Moreover, primer pair 2F2/2R2 was assessed by quantitative real-time polymerase chain reaction (PCR) using fungal DNA mixtures and DNA extracted from fungal experimental communities as templates. T. cf. harzianum was detectable even when as few as 100 copies of the SCAR marker were available or even when its population represented only 0.1% of the whole community.

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Cited by 34 publications

References 19 publications

A Universally Primed-Polymerase Chain Reaction (UP-PCR) Marker to Discriminate Clonostachys rosea ACM941 from Related Strains

A Universally Primed-Polymerase Chain Reaction (UP-PCR) Marker to Discriminate Clonostachys rosea ACM941 from Related Strains

TaqMan qPCR for Quantification of Clonostachys rosea Used as a Biological Control Agent Against Fusarium graminearum

Designing a SCAR molecular marker for monitoring Trichoderma cf. harzianum in experimental communities

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