a-contrast-three-inoculation-techniques-used-to-determine-race

A Contrast of Three Inoculation Techniques used to Determine the Race of Unknown <em>Fusarium oxysporum</em> f.sp. <em>niveum</em> Isolates

Fusarium wilt of watermelon (Citrullus lanatus), caused by Fusarium oxysporum f. sp. niveum (Fon), has reemerged as a major production constraint in the ...

University of Florida

The genus  includes pathogenic and nonpathogenic species. We report the 5.57-Mb genome sequences of two  strains, G18-1365 and G18-1376, isolated from symptomatic plantain plants in Haiti. These strains are genetically closely related (average nucleotide identity [ANI] > 99%) to the previously described type strain of , DSM 15968.

Draft Genome Sequences of Plant-Pathogenic Klebsiella variicola Strains Isolated from Plantain in Haiti.

Here, we announce the draft genome sequences of three  isolates that were used to inoculate slash pines () at the U.S. Forest Service Resistance Screening Center in Asheville, North Carolina. The genomes of these isolates were similar to other publicly available genomes, with average nucleotide identity values of >0.98.

Draft Genome Sequences of Three Fusarium circinatum Isolates Used To Inoculate a Pedigreed Population of  Seedlings.

Before 1991,  was the causal agent of bacterial spot of tomato in Florida but was quickly replaced by . The  population has changed in genotype and phenotype despite lack of a clear selection pressure. To determine the current  population in Florida, we collected 585  strains from 70 tomato fields, representing 22 farms across eight counties, in the Florida tomato production region. Strains were isolated from 23 cultivars across eight seed producers and were associated with eight transplant facilities during the fall 2017 season. Our collection was phenotypically and genotypically characterized. Only  was identified, and all strains except one (99.8%) were tolerant to copper sulfate and 25% of strains were resistant to streptomycin sulfate. Most of the strains (99.3%) that were resistant to streptomycin sulfate were sequence type 1. The  population consisted of tomato races 3 (8%) and 4 (92%) and all three previously reported sequence types, ranging from 22 to 46% frequency. Approximately half of all strains, none of which were sequence type 2, produced bacteriocins against . Effector profiles were highly variable among strains, which could impact the strains' host range. The effector , which was previously thought to be conserved in  tomato pathogens, was absent in 19 strains. Nonmetric multidimensional scaling and network analyses show how strains and strain traits were associated with production system variables, including anonymized farms and transplant facilities. These analyses show that the composition of the Florida  population is diverse and complex.

Assessing Changes and Associations in the  Population Across Florida Commercial Tomato Fields Via a Statewide Survey.

Fusarium wilt of watermelon, caused by  f. sp.  (FON), is pathogenic only to watermelon and has become one of the main limiting factors in watermelon production internationally. Detection methods for this pathogen are limited, with few published molecular assays available to differentiate FON from other formae speciales of . FON has four known races that vary in virulence but are difficult and costly to differentiate using traditional inoculation methods and only race 2 can be differentiated molecularly. In this study, genomic and chromosomal comparisons facilitated the development of a conventional polymerase chain reaction (PCR) assay that could differentiate race 3 from races 1 and 2, and by using two other published PCR markers in unison with the new marker, the three races could be differentiated. The new PCR marker, FNR3-F/FNR3-R, amplified a 511 bp region on the "pathogenicity chromosome" of the FON genome that is absent in race 3. FNR3-F/FNR3-R detected genomic DNA down to 2.0 pg/µL. This marker, along with two previously published FON markers, was successfully applied to test over 160 pathogenic FON isolates from Florida, Georgia, and South Carolina. Together, these three FON primer sets worked well for differentiating races 1, 2, and 3 of FON. For each marker, a greater proportion (60 to 90%) of molecular results agreed with the traditional bioassay method of race differentiation compared to those that did not. The new PCR marker should be useful to differentiate FON races and improve Fusarium wilt research.

Marker Development for Differentiation of  f. sp.  Race 3 from Races 1 and 2.

Fusarium wilt of watermelon (Citrullus lanatus) caused by Fusarium oxysporum f. sp. niveum (Fon), has become an increasing concern of farmers in the southeastern USA, especially in Florida. Management of this disease, most often through the use of resistant cultivars and crop rotation, requires an accurate understanding of an area's pathogen population structure and phenotypic characteristics. This study improved the understanding of the state's pathogen population by completing multilocus sequence analysis (MLSA) of two housekeeping genes (BT and TEF) and two loci (ITS and IGS), aggressiveness and race-determining bioassays on 72 isolates collected between 2011 and 2015 from major watermelon production areas in North, Central, and South Florida. Multilocus sequence analysis (MLSA) failed to group race 3 isolates into a single large clade; moreover, clade membership was not apparently correlated with aggressiveness (which varied both within and between clades), and only slightly with sampling location. The failure of multilocus sequence analysis using four highly conserved housekeeping genes and loci to clearly group and delineate known Fon races provides justification for future whole genome sequencing efforts whose more robust genomic comparisons will provide higher resolution of intra-species genetic distinctions. Consequently, these results suggest that identification of Fon isolates by race determination alone may fail to detect economically important phenotypic characteristics such as aggressiveness leading to inaccurate risk assessment.

Phylogenetic and phenotypic characterization of Fusarium oxysporum f. sp. niveum isolates from Florida-grown watermelon.

Watermelon is an important commercial crop in the Southeastern United States and around the world. However, production is significantly limited by biotic factors including fusarium wilt caused by the hemibiotrophic fungus  forma specialis  (Fon). Unfortunately, this disease has increased significantly in its presence over the last several decades as races have emerged which can overcome the available commercial resistance. Management strategies include rotation, improved crop resistance, and chemical control, but early and accurate diagnostics are required for appropriate management. Accurate diagnostics require molecular and genomic strategies due to the near identical genomic sequences of the various races. Bioassays exist for evaluating both the pathogenicity and virulence of an isolate but are limited by the time and resources required. Molecular strategies are still imperfect but greatly reduce the time to complete the diagnosis. This article presents the current state of the research surrounding races, both how races have been detected and diagnosed in the past and future prospects for improving the system of differentiation. Additionally, the available Fon genomes were analyzed using a strategy previously described in separate formae speciales avirulence gene association studies in  races.

Article	Year
Draft Genome Sequences of Plant-Pathogenic Klebsiella variicola Strains Isolated from Plantain in Haiti. Microbiology resource announcements\| PubMed ID: 32675179	2020
Draft Genome Sequences of Three Fusarium circinatum Isolates Used To Inoculate a Pedigreed Population of Seedlings. Microbiology resource announcements\| PubMed ID: 32703836	2020
Assessing Changes and Associations in the Population Across Florida Commercial Tomato Fields Via a Statewide Survey. Phytopathology\| PubMed ID: 33048630	2021
Marker Development for Differentiation of f. sp. Race 3 from Races 1 and 2. International journal of molecular sciences\| PubMed ID: 33467563	2021
Phylogenetic and phenotypic characterization of Fusarium oxysporum f. sp. niveum isolates from Florida-grown watermelon. PloS one\| PubMed ID: 33764995	2021
f. sp. Molecular Diagnostics Past, Present and Future. International journal of molecular sciences\| PubMed ID: 34575897	2021

James C. Fulton

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