First Report of Postharvest Fruit Rot on Passion Fruit (Passiflora edulis) caused by Lasiodiplodia theobromae in the U.S.A

The commercial production of passion fruit is geographically limited (California, Florida, and Hawaii), but the development of cold-tolerant varieties could expand it beyond warm-climate states (Stafne et.al. 2023). With only few countries allowed to export fresh fruit to the U.S. (i.e., Australia, Bermuda, Chile, and New Zealand; USDA-APHIS 2022) and its high retail value ($3-5/fruit), domestic production of passion fruit has awakened the interest of producers. During 2019-2022, fruit samples showing rot symptoms were received at the Plant Diagnostic Clinic of the Tropical Research and Education Center (TREC) and diagnosed as affected by Lasiodiplodia theobromae based on symptomatology and on the morphology of the isolated fungus. In July 2023, similar symptoms were observed at TREC's experimental plot and at two local commercial farms (Homestead, Florida). Incidence in these locations was about 10-15% of the harvested fruit. Initial lesions were irregular in shape with a water-soaked greasy appearance. After 3 days of incubation, lesions enlarged into dark sunken blotches and were covered by globular pycnidia and greenish-gray hyphae. A cross section of the fruit revealed fungal colonization of the locular cavity and softening of the mesocarp. Infected tissue was plated (6 pieces/fruit, total of 6 fruits) and isolates which morphology matched that described for L. theobromae (Aleves et al. 2008) were recovered consistently. Conidia were initially hyaline, unicellular, apex rounded, thick-walled, and ellipsoid, becoming dark brown, bicellular with longitudinal striations at maturity, 25 ± 4 x 14 ± 2 µm (n=50). Two isolates (TREC-L-1 and DAK-L-2A) were selected for identification through the sequencing of the partial regions of the internal transcribed spacer (ITS), translational elongation factor 1α (TEF1α), B-tubulin (Bt), and RNA polymerase II second largest subunit (RPB2) genes (Glass and Donaldson 1995). The ITS (PP806573-PP806574), TEF1-α (PP812413- PP812414), Bt (PP812415- PP812416), and RPB2 (PQ807646- PQ807647) sequences, of both isolates, had the highest similarity with those generated from the type material of Lasiodiplodia theobromae (CBS 164.96) and the concatenated phylogenetic analysis confirmed the identification of the isolates. For the pathogenicity test, disinfested fruit that were either punctured with a sterile needle or non-wounded, were inoculated with an agar plug excised from the growing edge of a 5-day-old PDA (Potato Dextrose Agar) colony of the TREC-L-1 isolate or with 100 ul of a 10³ conidia suspension collected from pycnidia produced in 4-week-old cultures of the same isolate. Negative controls consisted of wounded and non-wounded fruit inoculated with sterile PDA plugs or sprayed with water. Each treatment consisted of ten fruits. The fruits were kept inside plastic boxes at 25°C, under darkness, and with a RH> 80%. After eight days, black sunken rot symptoms, pycnidia production, and external and internal mycelial growth were observed on the inoculated and wounded fruit while no symptoms were observed on the non-wounded plug-inoculated and on the mock inoculated fruit. Severity and fungal colonization were higher in wounded fruit. In the spore suspension inoculation experiment, all wounded fruit showed symptoms while in the non-wounded fruit, 8/10 showed symptoms. The fungi recovered from the fruit lesions were confirmed as L. theobromae by morphological characteristics. This disease has been reported in Taiwan and China (Zhang et al., 2020). To our knowledge this is the first formal report of this disease in the U.S.A. As the industry grows, management strategies to control this disease need to be explored.