Adnan Uzunovic

Appropriateness of Probit-9 in the Development of Quarantine Treatments for Timber and Timber Commodities

ABSTRACT Following the increasing international phasing out of methyl bromide for quarantine purposes, the development of alternative treatments for timber pests becomes imperative. The international accreditation of new quarantine treatments requires verification standards that give confidence in the effectiveness of a treatment. Probit-9 mortality is a standard for treatment effectiveness that has its origin in fruit fly research, and has been adopted by the United States Department of Agriculture for fruit flies and several other pests. Following this, the probit-9 standard has been adopted as a benchmark for many quarantine treatments worldwide. This article discusses aspects of the application of this concept for a range of timber pests. Problematic issues include the often small pest populations available for testing, the limits of modeling pest responses to a treatment in the absence of sufficient numbers for treatment verification, the species diversity of pests and host materials and the physical and chemical conditions of host material or treatment conditions. Where treatment verification by killing large numbers of individuals is impossible, data collected from small populations or under specific conditions must be interpreted with caution. We discuss possible alternative approaches to probit-9 as a treatment efficacy standard.

Molecular Detection of 10 of the Most Unwanted Alien Forest Pathogens in Canada Using Real-Time PCR

Invasive alien tree pathogens can cause significant economic losses as well as large-scale damage to natural ecosystems. Early detection to prevent their establishment and spread is an important approach used by several national plant protection organizations (NPPOs). Molecular detection tools targeting 10 of the most unwanted alien forest pathogens in Canada were developed as part of the TAIGA project (http://taigaforesthealth.com/). Forest pathogens were selected following an independent prioritization. Specific TaqMan real-time PCR detection assays were designed to function under homogeneous conditions so that they may be used in 96- or 384-well plate format arrays for high-throughput testing of large numbers of samples against multiple targets. Assays were validated for 1) specificity, 2) sensitivity, 3) precision, and 4) robustness on environmental samples. All assays were highly specific when evaluated against a panel of pure cultures of target and phylogenetically closely-related species. Sensitivity, evaluated by assessing the limit of detection (with a threshold of 95% of positive samples), was found to be between one and ten target gene region copies. Precision or repeatability of each assay revealed a mean coefficient of variation of 3.4%. All assays successfully allowed detection of target pathogen on positive environmental samples, without any non-specific amplification. These molecular detection tools will allow for rapid and reliable detection of 10 of the most unwanted alien forest pathogens in Canada.

Control of Black Stain Fungi with Biocides in Semitransparent Wood Coatings

One of the frequent signs of early failure in semitransparent wood coatings is colonization by “black stain” fungi, such as Aureobasidium pullulans. A laboratory method evaluating the ability of va...

Ultrasonic phytosanitation of pinewood nematode infected wood

Pinewood nematode (Bursaphelenchus xylophilus) mortality was investigated after ultrasonic treatment at 20 and 40 kHz frequency. Experiments were conducted with infected small wood specimens that were ultrasonically treated for 1, 3, 5 and 7 hours and two variable temperature conditions, namely, a gradually increasing from ambient to a maximum of 70oC and a decreasing from ambient to a minimum of 5oC. The results revealed that the ultrasonic treatment itself had no significant effect on the nematode mortality at the 5oC level, while at the 70oC level, considerable nematode mortality was observed in short time periods and at 7 hours of sonic exposure it reached 100%. Therefore, certain combinations of timing and frequency of ultrasonic waves and produced heat can be effective in killing pinewood nematodes thus resulting in phytosanitized wood.

The Effect of Fungal Preinfection on the Efficacy of Mold and Sapstain Control Products

ABSTRACT Efficient use of sapstain control products requires information on the effectiveness of the actives and formulations against different types of fungal challenges. The present work examines...

Genome-Enhanced Detection and Identification (GEDI) of plant pathogens

Plant diseases caused by fungi and Oomycetes represent worldwide threats to crops and forest ecosystems. Effective prevention and appropriate management of emerging diseases rely on rapid detection and identification of the causal pathogens. The increase in genomic resources makes it possible to generate novel genome-enhanced DNA detection assays that can exploit whole genomes to discover candidate genes for pathogen detection. A pipeline was developed to identify genome regions that discriminate taxa or groups of taxa and can be converted into PCR assays. The modular pipeline is comprised of four components: (1) selection and genome sequencing of phylogenetically related taxa, (2) identification of clusters of orthologous genes, (3) elimination of false positives by filtering, and (4) assay design. This pipeline was applied to some of the most important plant pathogens across three broad taxonomic groups: Phytophthoras (Stramenopiles, Oomycota), Dothideomycetes (Fungi, Ascomycota) and Pucciniales (Fungi, Basidiomycota). Comparison of 73 fungal and Oomycete genomes led the discovery of 5,939 gene clusters that were unique to the targeted taxa and an additional 535 that were common at higher taxonomic levels. Approximately 28% of the 299 tested were converted into qPCR assays that met our set of specificity criteria. This work demonstrates that a genome-wide approach can efficiently identify multiple taxon-specific genome regions that can be converted into highly specific PCR assays. The possibility to easily obtain multiple alternative regions to design highly specific qPCR assays should be of great help in tackling challenging cases for which higher taxon-resolution is needed.

Biosurveillance of forest insects: part I—integration and application of genomic tools to the surveillance of non-native forest insects

Invasive species pose significant threats to forest ecosystems. Early intervention strategies are the most cost-effective means to control biological invasions, but are reliant on robust biosurveillance. State-of-the-art genomic approaches can provide an unprecedented opportunity to access detailed information on the invasion process and adaptive potential of invasive insects that pose an immediate threat to forests environments. Genomics can improve diagnostics of the invader and identify its route of invasion by determining the source population(s), assess its probability of establishment and patterns of spread, as well as provide evidence of adaptation. Applied biosurveillance efforts by plant health regulatory agencies will benefit substantially from the detailed insights that genomic data bring to our understanding of biological invasions.

Rapid Screening of Sulfuryl Fluoride as a Potential Phytosanitary Treatment for a Broad Selection of Fungi Relevant to Forestry

Abstract Increasing restrictions on the use of methyl bromide have created an urgent need to find suitable replacements that are effective in eliminating exotic pest establishments via trade in woo...