A Comparison of Pathogen Isolation in Culture and Injection–infiltration Bioassay of Citrus Leaves for DetectingXanthomonas citrisubsp.citri

Abstract

Citrus canker [caused by Xanthomonas citri subsp. citri (Xcc)] can cause yield loss of susceptible citrus and result in trade restrictions of fresh fruit. For both regulatory purposes and epidemiological studies, accurate detection and quantification of viable inoculum are critical. Two accepted methods used to detect and quantify Xcc are injection–infiltration bioassay and culture, but these two methods have not been directly compared using field‐obtained samples. The two methods were compared using washates of lesions taken from fruit, leaves and shoots in a commercial orchard in Florida in 2009–2010 and 2010–2011, with bioassay being the assumed standard. Despite some misclassifications, true positives (sensitivity) and true negatives (specificity) were the dominant classes using culture. False positives for lesions from shoots ranged from 13.1 to 21.4% in 2009–2010 and 2010–2011, respectively, and false positives for lesions from fruit and leaves ranged from 4.3 to 15.7%, in the two seasons, respectively. The false positive rate for culture compared with injection–infiltration bioassay was highest (0.16–0.55), due to more frequent recovery of Xcc by culture at ≤103 colony‐forming units (CFU) Xcc per ml. The false negative rate was consistently lower (0.02–0.21), confirming that in only a few cases did culture fail to detect Xcc when it was present. The area under the curve for receiver operator characteristic analysis ranged from 0.80 to 0.97, confirming that culture provided an accurate diagnosis in most cases. There was a higher frequency of lesions from shoots with a CFU ≤103 Xcc compared with lesions from fruit or leaves, making culture more effective at detecting these. The data demonstrate that culture is a reliable way to detect and quantify Xcc compared with injection–infiltration bioassay, particularly when the CFU is ≤103 Xcc per ml.

Canine Olfactory Detection of a Non-Systemic Phytobacterial Citrus Pathogen of International Quarantine Significance

“…Ruby Red Grapefruit seedlings were inoculated using a needless syringe tightly appressed to the abaxial laminar surface of 1/2 to 2/3 expanded leaves and forcing inoculum (≈10 5 cfu/mL) into the lamina via pressure injection infiltration; then, they were allowed to develop citrus canker symptoms for 4 weeks prior to canine interrogation [ 35 ]. Non-inoculated trees were arrayed in a 100 tree, 10 × 10 grid, with ≈3 m between trees within row and between rows, and with 1 to 10 Xcc-infected trees randomly placed within the grid.…”