qPCR Assays for the Detection of <i>Cylindrocladium buxicola</i> in Plant, Water, and Air Samples
Bibliographic record
Abstract
Cylindrocladium buxicola (syn. C. pseudonaviculatum; teleomorph Calonectria pseudonaviculata) is an important fungal pathogen of Buxus spp. Although widespread in Western Europe, this pathogen has only recently been introduced into North America, where it represents a significant threat to the U.S. and Canadian boxwood industries. Trade of latently infected nursery stock is an important mode of long-distance dissemination and introduction of this pathogen but no methods for detection of latently infected material are available. Also, the pathways for short-distance dispersal of C. buxicola have not been adequately studied. Improved detection methods of this pathogen in air and water samples would benefit future research in this area. We have developed real-time polymerase chain reaction assays for the detection of C. buxicola based on the ribosomal DNA internal transcribed spacer 1 (ITS) and the β-tubulin 2 gene (TUB). Using a TaqMan probe conjugated with a 3' minor groove binding group (TaqMan MGB probe), the ITS-based assay could reliably detect as little as 10 fg of genomic DNA or 20 copies of cloned target DNA and was approximately 70 times more sensitive than the SYBR Green TUB-based assay. The ITS-based assay provided good but not complete specificity, and is well suited for epidemiological studies. The TUB-based assay, however, proved to be fully specific and can be used for diagnostics. We developed and optimized sample processing and DNA extraction methods for detection of latently present C. buxicola in boxwood plants and quantification of conidia in water and air samples. C. buxicola could be detected in 20 g of plant material, of which only 1 ppm of the tissue was infected, in 10-ml water samples containing as low as 1 conidium/ml, and on Melinex tape pieces representing 12 h of air sampling containing 10 or more conidia. The applicability of the techniques to plant, water, and air samples of practical size was demonstrated.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".