Sharma, Ashwani Kumar

Candidatus Liberibacter asiaticus (CLas) is a host-dependent obligate bacteria that causes citrus greening disease in citrus plants. The prolonged association of CLas with the host led to the loss of essential metabolic genes, including transaldolase in the pentose phosphate pathway, hindering the development of axenic culture in the host-free environment. In this study, in silico functional annotation of pyrophosphate-fructose 6-phosphate 1-phosphotransferase (PFP) protein was performed by utilizing sequence analysis and molecular modeling, revealing its potential role in bypassing the loss of transaldolase. The findings suggest how PFP and fructose bisphosphate aldolase (FBPA) can reconstruct the disrupted central carbon metabolism and help CLas in ATP production. The molecular modeling of PFP showed binding affinity towards fructose 6-phosphate and sedoheptulose 7-phosphate, showing its dual function. The sequence-based analysis and metabolic modeling study also highlight the presence of other alternate enzymes or transport proteins in CLas, elucidating the broader metabolic resilience. This is the first report to provide the structural insights of PFP in compensating for the lost metabolic function and survival strategies adopted by CLas despite the restrained metabolic capability.

Citrus, mainly mandarin (Citrus reticulata Blanco), is an economically important fruit crop in Bhutan. Despite having favorable agroclimatic conditions for citrus cultivation, the early decline of fruit-bearing orchards coupled with low crop productivity is a major concern among citrus growers. During a recent survey, an association of ‘Candidatus Liberibacter asiaticus’ (citrus greening) and citrus tristeza virus (CTV), either singly or as mixed infections in declined citrus trees, was recorded in all four major citrus-growing districts (Tsirang, Dagana, Zhemgang, and Sarpang). Using PCR-based diagnosis, a higher incidence of citrus greening (27.45%) and tristeza (70.58%) was observed in symptomatic field samples. Detection and characterization of ‘Ca. L. asiaticus’ was performed based on the 16S ribosomal DNA, prophage gene, 50S ribosomal rplA-rplJ gene, and tandem repeats of the CLIBASIA_01645 locus. Similarly, the coat protein, p23, and p18 genes were used as genetic markers for the detection and characterization of Bhutanese CTV. The ‘Ca. L. asiaticus’ isolates from Bhutan segregated into classes II and III based on the CLIBASIA_01645 locus, analogous to Indian isolates from the northeast region and Term-A based on the CLIBASIA_05610 locus. CTV isolates of Bhutan were observed as closely related to the VT strain, which is considered to be the most devastating. To the best of our knowledge, this is the first study on molecular characterization of ‘Ca. L. asiaticus’ and CTV isolates and their association with citrus decline in Bhutan.