Primavera otoño 2020 (Año LXIII Núms. 122-123)

horizontes@pucpr.edu Año LXIV Núm. 124-125 horizontes PRIMAVERA / OTOÑO 2021 PUCPR 62 eradicate CLas. Eradication is critical to the citrus nursery industry. The utilization of clean nursery stock is part of the standard procedure for HLB management. Because CLas cannot be cultured, Bao et al. (2020) used information from whole-genome sequence strains to improve the detection and the sensitivity of polymerase chain reaction to detect HLB. They developed a new polymerase chain reaction based on a four-copy locus. The oligonucleotide 1 had a missing G base (as in A, G, C, T/U bases in DNA and RNA) that they replaced within primers to identify CLas. Studies showed that the whole genome sequencing information improved CLas detection accuracy and sensitivity. Researchers had three objectives: 1) to re-evaluate the sequence integrity of OI1/HLB based on the whole genome sequencing the data and to evaluate the impact of the missing G for CLas detection; 2) to evaluate the use of RNR-PCR to assist differentiation between a low CLas titer and no CLas in the samples with high Ct values to form the HLBas-PCR; 3) to develop a new PCR system based on a four-copy genomic locus and the evaluation of the potential to detect CLas. Other researchers investigated the transcriptome (the range of mRNA) of the sweet orange seedling trees infected by CLas. When Clas was inoculated with Citrus tristeza virus strains, both mild and severe Citrus tristeza virus strains became established in the trees. Selvaraj et al. (2018) demonstrated pathogen quantification using multiple copies in a duplex droplet digital polymerase chain reaction assay. The duplex assay used specific primers for the rRNA 16s gene and the RNR gene, in which quantitative polymerase chain reaction and duplex droplet digital polymerase chain reaction improved detection precision. The CLas duplex droplet digital polymerase chain reaction technology provided a robust method for quantitative detection to obtain more accurate results than the quantitative polymerase chain reaction assay without using standard curves. Shi-min et al. (2019) applied extensions of soft citrus tristeza virus strains to protect citrus fruits against citrus tristeza virus strains and trees infected with both citrus tristeza virus and CLas. They studied each infection model, in addition to coinfection models. Due to the different strains of citrus tristeza virus isolated from CLas and a variety of citrus fruits, the presence and duration of citrus tristeza virus establishment were significant before being inoculated with CLas. The metagenomic studies of HLB, which typically occurred in citric tissue or the Asian citrus psyllids models, were suspected of being infected with CLas. The detection of CLas was hampered by the limited information available. The lack of G in HLBas-PCR for the detection of CLas was insignificant in samples with a high titer of CLas; however, the low titer of no CLas could not be distinguished from CLas or the Asian citrus psyllid samples. Researchers carried out high-throughput screening to identify compounds inhibiting the activity of the transcriptional activator CLas without substantially inhibiting bacterial growth because CLas cannot be cultured. Merfa and de La Fuente (2020) discussed the cultivation of bacteria that were not ordinarily cultivable from different environmental sources. For instance, axenically (single species) cultured bacteria is a foundation of bacteriology. They discussed the cultivation of pure cultures marine bacteria through encapsulation. The l-tip methodology allowed cell and chemical compounds to diffuse into the I-tip to promote bacterial growth. Overall, research indicates that it is impossible to identify CLas lead compounds using an in vivo fluorescence baseline screen in the host. The goal is to use molecular techniques to develop CLas-resistant varieties. The