**2.3 Symptomology**

HLB symptoms are more evident in cold weather conditions than in hot seasons [30]. It is difficult to specify the period between when the citrus tree is affected by HLB and the onset of disease symptoms. It will exhibit in different parts of the plants or only in infected sectors when it eventually manifests symptoms. It is, therefore, difficult to diagnose and control at the early stage of HLB disease [31]. The HLBinfected tree exhibits symptoms in various parts of the plant depending on the stage of infection. If infection occurs soon after propagation, the entire tree gets affected and turns yellow all over the canopy, which leads to a decline irrevocably. Both the symptoms and the causative organisms were restricted to the infected sector in the event of later infection [27]. Only the infected sector will exhibit symptoms in the case of citrus trees affected by HLB, while the remaining parts will show normal growth and good-quality fruits. The symptoms observed on the HLB-affected tree include a heavy drop in the leaf and out-of-season flushing and blooming. Chronically, HLB-affected trees displayed stunting growth, twig dieback, sparse yellow foliage, or severe fruit drop [24]. The initial stage of HLB is vein yellowing [32], and the secondary level includes (infected leaves) small, upright with various chlorotic patterns similar to that caused by nutrient deficiency, such as zinc, sulfur, iron, boron, manganese, and calcium [33, 34]. In severe cases, the leaves were utterly void of chlorophyll, except for rounded green spots located on the leaves at random places [24]. The most accurate diagnostic symptom for HLB is that the infected fruits are small, lopsided, and taste bitter and salty. HLB-affected trees with premature shedding of green fruit drops while remaining on the tree, in which fruits with yellow halo-like lesions were staying green on the shaded side, hence the name "greening" [7, 34]. Root systems are developed in severely infected trees that exhibit poorly formed roots with few fibrous roots due to undernourishment [24] and repression of new root growth and rootlets decay [10].

HLB disease is challenging to diagnose based on symptoms, particularly during the early stages of the disease. Numerous symptoms of HLB might occur, and citrus trees are often caused by other diseases or nutrient deficiencies that may lead to similar symptoms [11, 30, 35]. Symptoms could be aggravated by other pathogens being coinfected. Several reports from Asian countries postulated that HLB-affected citrus trees are commonly coinfected with the *Citrus tristeza virus* (CTV) [7]. Interestingly, some CTV isolates protect trees against HLB infection [36]. Blotchy mottle leaf is a principal diagnosis of HLB that could be misinterpreted with other diseases, such as stubborn citrus disease caused by *Spiroplasma citri*, a severe infection of CTV phytophthora root rot, zinc deficiency, and waterlogging. Furthermore, it can also be confused with symptoms of leaf-related stress and mineral deficiency [37]. Early stages of citrus blight are also associated with the symptoms of zinc deficiency [38]. For these confusing symptoms of the disease, an unequivocal diagnosis technique is needed for HLB disease.

#### **2.4 Method of HLB detection**

Early identification and isolation of *Canditatus* Liberibacter species-infected trees are effective management approaches used to limit the spread of HLB from invading HLB disease-free citrus orchards in local and international trade [39]. Visual examination is one of the most commonly employed approaches for detecting citrus HLB disease. Traditionally, early detection of HLB disease relied primarily on various symptoms in the field, such as blotchy mottle leaf, yellow shoot, aborted seed, and lopsided fruit with green color remaining at the stylar end [40]. Nevertheless, this approach is highly affected by subjective interpretation, diagnostic errors can be higher than 30%, and other biotic and abiotic stress-related problems may worsen diagnosis. HLB symptoms might be confused with diseases such as *Citrus Tristeza* 

*Devious Phloem Intruder* Candidatus *Liberibacter Species Causing Huanglongbing: History… DOI: http://dx.doi.org/10.5772/intechopen.105089*

*Closterovirus*, *Phytophthora* infection, citrus blight, and specific nutrient deficiencies [41]. Thus, the availability of advanced technologies that enable early and rapid detection of HLB pathogens is crucial [42]. Currently used methods for the diagnosis and confirmation of HLB disease include serology, enzymatic assay, enzyme-linked immunosorbent assays (ELISA), transmission electron microscopy, DNA probes, conventional polymerase chain reaction (PCR), quantitative PCR (qPCR), Fourier transform infrared spectroscopy (FTIR), and mid-infrared spectroscopy. Pereira et al. [43] developed a method for early diagnosis using X-ray fluorescence. The laserinduced breakdown spectroscopy (LIBS) combined with chemometric strategies is used to predict the condition of orchard plants infected with *Canditatus* Liberibacter species successfully. However, these methods did not provide early diagnosis except for the LIBS method. Recently, Tran et al. [44] reported a sensitive and selective label-free biosensor that combines the physical and chemical advantages of carbon nanomaterials such as single-walled carbon nanotubes (SWNTs) in a field-effect transistor (FET)/chemiresistor architecture with selective antibodies against Secdelivered effector 1 (SDE1), a secreted protein biomarker, for the detection of HLB. Detailed HLB detection techniques have recently been reviewed [42, 45].
