Xanthomonas arboricola pv. pruni (Xap) causes bacterial spot, a major worldwide disease of Prunus species. Very few chemical management options are available for this disease and frequent applications of oxytetracycline (OTC) in the United States peach orchards have raised concerns about resistance development. During 2017–2020, 430 Xap strains were collected from ten peach orchards in South Carolina. Seven OTC-resistant (OTC^R) Xap strains were found in 2017 and 2020 from four orchards about 20–270 km apart. Interestingly, the seven strains were also resistant to streptomycin (STR). Six strains grew on media amended with ≤100 μg/mL OTC, while one strain, R1, grew on ≤250 μg/mL OTC. Genome sequence analysis of four representative OTC^R strains revealed a 14–20 kb plasmid carrying tetC, tetR, and strAB in each strain. These three genes were transferable to Xanthomonas perforans via conjugation, and they were PCR confirmed in all seven OTC^R Xap strains. When tetC and tetR were cloned and expressed together in a sensitive strain, the transconjugants showed resistance to ≤100 μg/mL OTC. When tetC was cloned and expressed alone in a sensitive strain, the transconjugants showed resistance to ≤250 μg/mL OTC. TetC and tetR expression was inducible by OTC in all six wild-type strains resistant to ≤100 μg/mL OTC. However, in the R1 strain resistant to ≤250 μg/mL OTC, tetR was not expressed, possibly due to the presence of Tn3 in the tetR gene, and in this case tetC was constitutively expressed. These data suggest that tetC confers OTC resistance in Xap strains, and tetR regulates the level of OTC resistance conferred by tetC. To our knowledge, this is the first report of OTC resistance in plant pathogenic xanthomonads.

In several host–pathogen systems, the detection of an individual quantitative trait locus (QTL) differed depending on the specific pathogen isolate or phenotypic assay that was used. Moreover, the contribution of a QTL toward yield is an important factor for selecting specific QTL candidates for resistance breeding. The first experiment in this study compared the QTL identified previously in a ‘Conrad’ × ‘Sloan’ F_4:6 soybean [Glycine max (L.) Merr.] population challenged with Phytophthora sojae Kaufmann and Gerdemann isolate 1.S.1.1 to the QTL identified when this same population was challenged with isolates PT2004C2.S1 and OH25 using a tray test assay. In the second experiment, QTL were mapped in the same population with P. sojae isolate 1.S.1.1 using a different phenotypic assay, the layer test. In the third experiment, the impact of these QTL on soybean yield was evaluated using 20 selected recombinant inbred lines (RILs) grown in a field infested with P. sojae. Resistance QTL with smaller effects, especially those from the susceptible parent, were not consistently detected with the three isolates or the two phenotypic assays. Composite interval mapping identified a major Conrad QTL on chromosome 18 and one on 19 that were detected with all three isolates and both phenotypic assays. Recombinant inbred lines with resistance alleles at these QTL had significantly higher yields (∼1000 kg ha⁻¹ more) than RILs with susceptible alleles. These results indicate the important role these QTL play in conferring partial resistance to P. sojae.

Xanthomonas fragariae causes angular leaf spot of strawberry, an important disease in strawberry growing regions worldwide. To better understand how X. fragariae multiplies and moves in strawberry plants, a green fluorescent protein (GFP)-labeled strain was constructed and used to monitor the pathogen’s presence in leaf, petiole, and crown tissue with fluorescence microscopy following natural and wound inoculation in three strawberry cultivars. Taqman PCR was used to quantify bacterial densities in these same tissues regardless of the presence of GFP signal. Results showed X. fragariae colonized leaf mesophyll, the top 1 cm portion of the petiole adjacent to the leaf blade, and was occasionally found colonizing xylem vessels down to the middle of the petioles. The colonization of vascular bundles and the limited systemic movement that was observed appeared to be a passive process, of which the frequency increased with wounding and direct infiltration of bacteria into leaf veins. X. fragariae was able to directly enter petioles and colonize the space under the epidermis. Systemic movement of the bacteria into crown and other uninoculated tissues was not detected visually by GFP. However, X. fragariae was occasionally detected in these tissues by qPCR, but at quantities very near the qPCR detection limit. Petiole tissue harboring bacteria introduced either by direct entry through natural openings or wounds, or by systemic movement from infected foliar tissue, likely serves as a main source of initial inoculum in field plantings.

Cotton leafroll dwarf virus (CLRDV, family: Luteoviridae, genus: Polerovirus) was recently reported in cotton (Gossypium hirsutum L.) plants affected by cotton leafroll dwarf disease (CLRDD) in the...

Xanthomonas fragariae causes angular leaf spot in strawberry. The pathogen’s association with its host tissue is thought to be a condition for its survival. Consequently, transmission of the pathogen to field production sites occurs almost exclusively through the movement of contaminated planting stock. The aim of this study was to develop a propidium monoazide (PMA)-quantitative PCR (qPCR) protocol for specific detection of viable X. fragariae cells. The qPCR procedure was developed for two different primer pairs: one producing a long amplicon (863 bp) and the other a short amplicon (61 bp). Both pairs were tested on mixtures of viable and heat-killed bacteria cells, bacteria-spiked strawberry petiole samples, and petioles collected from symptomatic, inoculated plants. The results showed that long-amplicon PMA-qPCR enabled specific and sensitive detection of X. fragariae with a detection limit of 10³ CFU/ml, and it significantly improved PMA efficiency in differentiating viable from dead bacterial cells relative to short-amplicon PMA-qPCR. Based on the delta threshold cycle (C_t) values (i.e., the difference in C_t values between PMA-treated and nontreated samples), the long-amplicon PMA-qPCR was able to suppress the detection of dead X. fragariae cells 1.9- to 3.1-fold across all petiole samples tested. The quantification results from PMA-qPCR for mixtures of viable and dead cells were highly correlated with the predicted bacterial concentrations in a linear relationship (R² = 0.981). This assay can be useful for identifying inoculum sources in the strawberry production cycle, which may lead to improved disease management strategies.

A quantitative PCR (qPCR)-assay was developed to detect airborne inoculum of Nothopassalora personata, causal agent of late leaf spot (LLS) on peanut, collected with a modified impaction spore trap. The qPCR assay was able to consistently detect as few as 10 spores with purified DNA and 25 spores based on crude DNA extraction from rods. In 2019, two spore traps were placed in two peanut fields with a history of LLS. Sampling units were replaced every 2 to 4 days and tested with the developed qPCR assay, while plots were monitored for symptom development. The system detected inoculum 35 to 56 days before visual symptoms developed in the field, with detection related to environmental parameters affecting pathogen life-cycle and disease development. This study develops the framework of the qPCR spore trap system and represents the initial steps towards validation of the performance of the system for use as a decision support tool to complement integrated management of LLS.