The team of the Science Center for the Development of the (CCD-CROP-IAC) project has reaped the first results of the project “Biotechnological and genomic strategies for quality, productivity and sustainable management of citrus, coffee and sugarcane in the State of São Paulo” as to the tolerance of the citrandarins rootstocks (hybrids of Citrus sunki x Poncirus trifoliata) to the huanglongbing (HLB) to pest or citrus greening. This study paves the way for new research works aimed at managing the HLB utilizing combinations of canopy/rootstocks more resilient to the harmful effects of the disease to the root system of the plants.

The tolerance of the citrandarins genotypes to the bacterium Candidatus Liberibacter asiaticus (CLas), causal agent of the HLB in citrus, was studied by master’s degree holder Thaís Magni Cavichioli supported by Fapesp (process 2019/06412-5), under the supervision of the Agronomic Institute (IAC) researcher Helvécio D. Coletta-Filho. The research was conducted during the graduation course on Plant Genetics and Enhancement at the Veterinary and Agrarian Science Course at the São Paulo State University “Júlio de Mesquita Filho (UNESP–Jaboticabal). The dissertation was submitted and defended in December 2021.

“In the study, 14 citrandarins genotypes were challenged under continued exposure to the HLB bacterium, grafting them with the canopy of the Citrus sinensis plant (Valência orange) heavily infected with HLB bacteria (Figure 1). After 360 days of infection, in four genotypes of these hybrids, there was a reduction from 50% to 83% in the number of plants infected by the HLB bacterium, ending up with the absence of any positive diagnosis in all the analyzed repetitions for the citrandarin H106, as determined by quantitative PCR in real time (qPCR)”, Dr. Coletta-Filho explains. Furthermore, in that hybrid low concentrations of starch (17 mg/g tissue) were observed and the formation of callose (8 fluorescence units/mm²), which indicate the severity of the disease.

Figure 1. Illustration of the citrandarins genotype graftings challenging Candidatus Liberibacter asiaticus – CLas infections. A. Sweet orange plants which received citrandarins graftings. B. Moist chamber utilized to stimulate the graft to start growing in the form of the fork method. C. General view of the experiment.

In another experiment the citrandarin genotype (H222), previously selected as tolerant to HLB, was used as rootstock and interspecies grafting for sweet orange trees infected with the HLB bacterium. According to the researcher, the concentration parameters of the bacterium in the canopy and in the root were also defined, both of them determined by qPCR, volume, dry and wet weight of the infected root system of the plants and not infected with CLas.

“The results attested that the interspecies grafts with the genotype H222 between the sweet orange canopy and the citrumelo Swingle as rootstock did not result in any gain for the root system of the plants. However, the citrandarin H222 as rootstock showed absence of HLB bacterium infection in its root system in three out of four repetitions”, he comments.

As a result, the roots of the citrandarin H222 rootstock, though having the Valência as infected canopy by the HLB bacterium, presented, on average, 25-percent gain in development rates compared with the combination Valência orange/citrumelo Swingle, also infected with HLB (Figure 2A).

The research reveals that this minor HLB impact on the citrandarin H222 root system, at an analysis that involves all the parameters, made it comparable to the results of this hybrid with the ones achieved by the noninoculated plants (Figure 2B). “However, it should be stressed that the HLB disease affected the development of the root system of the rootstocks, regardless of the variety, reaching rates of 70%, like what happened to the citrumelo Swingle”, the Dr. Coletta-Filho highlights.

Figure 2. Response of the hybrid H222 as interspecies graft and rootstock to the infection by Candidatus Liberibacter asiaticus – CLas. A. Aspect of the volume of the root system of the treatments where the Citrus sinensis canopy presented a high concentration of CLas. B. Analysis of the clusterization process of the analyzed variables (diameter of the ground portion of the main stem, volume and dry weight of the root system and C_T values (cycle threshold) as a response to treatments T1, T2, T3 and T4 to infection by CLas if compared with non-infected treatments. The variables were jointly analyzed using the multivariate technique.