- HUANGLONGBING: DEVASTATING DISEASE OF CITRUS 329
A. Anatomical
Early light microscope studies of citrus leaves with HLB symptoms
revealed localized pockets of necrotic phloem, massive accumulation
of starch in plastids, aberrations in cambial activity and subsequent dif-
ferentiation of cambial derivatives, thickening of cytoplasm, increased
staining of cell walls, and excessive phloem formation in aerial plant
parts (Schneider 1968). Although Schneider’s work was conducted with
the African form of HLB (associated with CLaf), similar results have
been reported repeatedly for trees infected with CLas (Etxeberria et al.
2009; Kim et al. 2009; Achor et al. 2010; Folimonova and Achor 2010;
Shokrollah et al. 2010; Fan et al. 2012; Koh et al. 2012; Aritua et al.
2013; Fan et al. 2013; Brodersen et al. 2014; Fu et al. 2015).
Kim et al. (2009) and Achor et al. (2010) demonstrated that callose
accumulates around sieve pores in the midvein tissues of CLas-infected
leaves. Koh et al. (2012) found that plasmodesmata pore units (PPUs)
connecting companion cells and sieve elements were stained with a
callose-specific dye in the CLas-infected leaf phloem cells. When exam-
ined by transmission electron microscopy, PPUs with abnormally large
callose deposits were more abundant in the Liberibacter-infected sam-
ples than in the uninfected samples. The PPUs in the CLas-infected, but
not HLB symptomatic, leaves were stained at levels similar to the PPUs
in the HLB-symptomatic leaves. Sieve plates in CLas-negative samples
were stained with aniline blue, but callose staining in PPUs was rarely
detected. Surprisingly, among the numerous published transcriptomic
studies regarding the effects of CLas, only one (Fan et al. 2012) reported
that callose synthase gene transcripts were more abundant in CLas-
infected than in non-infected plants. Enrique et al. (2011) demonstrated
that RNAi knockout of callose synthase in lemon resulted in plants more
susceptible to Asiatic citrus canker (causal agentXanthomonas citripv.
citri) than were wild type plants. It would be interesting to know how
knocking out callose synthase affects susceptibility to HLB.
Anatomical changes in response to CLas infection are not as pro-
nounced in roots as in aerial plant parts. At the light microscope level,
phloem collapse and plugging are not apparent in roots (Aritua et al.
2013; Johnson et al. 2014). Examination of root sections with transmis-
sion electron microscopy did reveal thickening of the middle lamella
along with thickening and collapse of sieve elements, however, the
response was not as pronounced as in aerial plant parts (Aritua et al.
2013).
According to Folimonova and Achor (2010), ultrastructural changes
were often observed in samples from recent growth flushes in
CLas-inoculated sweet orange and grapefruit leaves prior to the