Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/4/2007
Publication Date: 3/1/2008
Citation: Keith, R.C., K.A. Nishijima, L.M. Keith, M.M. Fitch, W.T. Nishijima, M.M. Wall. 2008. Atypical internal yellowing of papaya fruit in Hawaii caused by Enterobacter sakazakii. Plant Disease. 92:487. Interpretive Summary: This is the first report of a new bacterial disease of papaya fruit, associated with atypical internal yellowing symptoms. The bacterium, Enterobacter sakazakii was isolated and identified as the causal agent of the symptoms (a dull greenish-yellow discoloration of the flesh near the seed cavity). E. sakazakii can be considered a cross-domain pathogen that causes plant disease and also is implicated in human illness. This bacterium has the potential to affect the quality and safety of fresh and processed papaya products.
Technical Abstract: Internal yellowing (IY), characterized by yellow discolored tissue around the papaya (Carica papaya) seed cavity, diffuse margins and the presence of a distinctly rotten odor, was first reported in 1987. These symptoms were associated with the causal agent Enterobacter cloacae. Here we report the formation of atypical internal yellowing (AIY) in fully ripened papaya caused by the bacterium Enterobacter sakazakii. In surveys conducted in 2006 to 2007, 'Kapoho' papayas were obtained from various packinghouses on the island of Hawaii. Among papayas examined that were asymptomatic for IY, a dull greenish-yellow discoloration of the flesh with a distinct margin extending from the seed cavity into the pericarp was noted, along with a pungent odor. These symptoms occurred in 5 of 500 fruit surveyed and bacterial populations were at 100 to 1000 cfu/g. Areas exhibiting discoloration were aseptically excised, macerated, serially diluted and plated onto PT-M4 media. Colonies on PT-M4 were orange-red in color, slightly convex, and circular with a somewhat opaque 1 mm margin surrounding the entire colony after 48 h. Five purified strains, inoculated into oxidation/fermentation-glucose (OF) tubes and API 20E strips incubated at 30C, were shown to be facultative anaerobes and identified as E. sakazakii with a 98.4% certainty. Colonies plated onto tryptic soy agar (TSA) and incubated in light for 72 h at 25C produced bright yellow pigmentation, indicative of E. sakazakii. Amplification using the polymerase chain reaction (pcr) with E. sakazakii specific primers yielded a 929 bp fragment which was absent from E. cloacae and Pseudomonas aeruginosa samples. To confirm pathogenicity, 'Kapoho' papayas were inoculated with cell suspensions of putative E. sakazakii strains, as well as E. cloacae strain YPV-5B. Koch’s postulates were fulfilled by re-isolation from the tissue exhibiting symptoms. E. cloacae inoculated sites produced typical IY symptoms, while the sites inoculated with the putative E. sakazakii strains produced greenish-yellow discolored tissue (AIY). The identification of the re-isolated bacterial strains was confirmed with API 20E testing, PCR amplification and pigment production on TSA. Although less prevalent (1% incidence) than the typical IY produced by E. cloacae, this bacterium has the potential to affect the quality and safety of fresh and processed papaya products.