Fattah-Hosseini, Somayeh

ABSTRACT “ Candidatus Liberibacter asiaticus” is the causative bacterium associated with citrus greening disease. “ Ca . Liberibacter asiaticus” is transmitted by Diaphorina citri more efficiently when it is acquired by nymphs rather than adults. Why this occurs is not known. We compared midguts of D. citri insects reared on healthy or “ Ca . Liberibacter asiaticus”-infected citrus trees using quantitative PCR, confocal microscopy, and mitochondrial superoxide staining for evidence of oxidative stress. Consistent with its classification as propagative, “ Ca . Liberibacter asiaticus” titers were higher in adults than in nymphs. Our previous work showed that adult D. citri insects have basal levels of karyorrhexis (fragmentation of the nucleus) in midgut epithelial cells, which is increased in severity and frequency in response to “ Ca . Liberibacter asiaticus.” Here, we show that nymphs exhibit lower levels of early-stage karyorrhexis than adults and are refractory to the induction of advanced karyorrhexis by “ Ca . Liberibacter asiaticus” in the midgut epithelium. MitoSox Red staining showed that guts of infected adults, particularly males, experienced oxidative stress in response to “ Ca . Liberibacter asiaticus.” A positive correlation between the titers of “ Ca . Liberibacter asiaticus” and the Wolbachia endosymbiont was observed in adult and nymph midguts, suggesting an interplay between these bacteria during development. We hypothesize that the resistance of the nymph midgut to late-stage karyorrhexis through as yet unknown molecular mechanisms benefits “ Ca . Liberibacter asiaticus” for efficient invasion of midgut epithelial cells, which may be a factor explaining the developmental dependency of “ Ca . Liberibacter asiaticus” acquisition by the vector.

AbstractCandidatus Liberibacter asiaticus (CLas) is a phloem-limited, gram-negative, fastidious bacterium that is associated with the development of citrus greening disease, also known as Huanglongbing (HLB). CLas is transmitted by the Asian citrus psyllid (ACP) Diaphorina citri, in a circulative manner. Two major barriers to transmission within the insect are the midgut and the salivary glands. We performed a thorough microscopic analysis within the insect midgut following exposure to CLas-infected citrus trees. We observed changes in nuclear architecture, including pyknosis and karyorrhexis as well as changes to the actin cytoskeleton in CLas-exposed midgut cells. Further analyses showed that the changes are likely due to the activation of programmed cell death as assessed by Annexin V staining and DNA fragmentation assays. These results suggest that exposure to CLas-infected trees induces apoptotic responses in the psyllid midgut that should be further investigated. Understanding the adaptive significance of the apoptotic response has the potential to create new approaches for controlling HLB.