Citrus Disease Research Articles

Cultural Management of Huanglongbing: Current Status and Ongoing Research.

Huanglongbing (HLB), formerly known as greening, is a bacterial disease restricted to some Asian and African regions until two decades ago. Nowadays, associated bacteria and their vectors have spread to almost all citrus-producing regions, and it is currently considered the most devastating citrus disease. HLB management can be approached in terms of prevention, limiting or avoiding pathogen and associated vectors to reach an area, or in terms of control, trying to reduce the impact of the disease by adopting different cultural strategies depending on infestation/infection levels. In both cases, control of psyllid populations is currently the best way to stop HLB spread. Best cultural actions (CHMAs, TPS system) to attain this goal and, thus, able to limit HLB spread, and ongoing research in this regard is summarized in this review.

Different Sweet Orange‒Rootstock Combinations Infected by Candidatus Liberibacter asiaticus under Greenhouse Conditions: Effects on the Roots

Grafting a scion onto a rootstock results in physical and physiological changes in plant growth and development, which can affect tree vigor, productivity, and tolerance to stress and disease. Huanglongbing (HLB) is one of the most destructive citrus diseases and has become endemic in Florida since its introduction in 2005. It is associated with the phloem-limited bacteria Candidatus Liberibacter asiaticus (CLas), which cause severe metabolic disruptions in affected plants. Although most scion cultivars are highly susceptible, some rootstock cultivars are tolerant and allow the grafted tree to cope better with the disease. The objectives of this study were to identify rootstock traits that can be used to assess cultivars under controlled greenhouse conditions in advance of longer-term field trials. We used 10 commercially important rootstocks with different genetic backgrounds and known field performance in graft combination with ‘Valencia’ sweet orange scion. Trees were graft-inoculated with CLas and compared against mock-inoculated trees. Tree health and CLas populations were assessed regularly, and root growth was monitored using a minirhizotron imaging system. Plants were excavated and destructively sampled 21 months after inoculation to assess biomass distributions and other CLas-induced effects. We found significant differences between healthy and infected trees for most variables measured, regardless of the rootstock. In contrast to leaf CLas titers, root titers were significantly influenced by the rootstock, and highest levels were measured for ‘Ridge’ sweet orange and sour orange. Root growth and root biomasses were reduced upon infection but differences among rootstocks did not always agree with reported field performances. Despite severe biomass reductions plants maintained their relative distribution of biomass among different components of the root system, and no dead roots were observed. Root respiration was reduced by CLas infection and was overall higher in tolerant cultivars suggesting its potential as a physiological marker. This study improves our knowledge about the strengths and weaknesses of assessing rootstock traits of grafted trees in a controlled greenhouse setting. Results from the study suggest that in addition to HLB tolerance, other rootstock traits will ultimately have major contributions to field survival and productivity of the grafted trees in an HLB endemic production environment.

Different Sweet Orange–Rootstock Combinations Infected by Candidatus Liberibacter asiaticus under Greenhouse Conditions: Effects on the Scion

The devastating citrus disease huanglongbing (HLB) associated with the phloem-limited bacteria Candidatus Liberibacter asiaticus (CLas) has caused a more than 70% reduction in citrus production since its discovery in Florida in 2005. Most citrus scion cultivars are sensitive to HLB, whereas some cultivars used as rootstocks are tolerant. Using such tolerant rootstocks can help trees to cope better with the disease’s impact. Evaluating rootstock effects on a grafted scion in the field takes many years, but shorter-term evaluation is imperative to aid in rootstock selection for an HLB-endemic production environment. In this study, we investigated grafted healthy and CLas-infected citrus trees under controlled greenhouse conditions. The objectives were to identify traits suitable for assessing grafted tree tolerance in advance of longer-term field studies and aiding in the selection of superior rootstock cultivars. We assessed 10 commercially important rootstocks grafted with ‘Valencia’ sweet orange scion and with known field performance. At 6, 9, 15, and 21 months after graft inoculation (mai), leaf CLas titers were determined and canopy health was evaluated. Plants were destructively sampled at 21 mai to assess plant biomasses and other physiological and horticultural variables. There was little influence of the rootstock cultivar on CLas titers. Surprisingly, few HLB foliar disease symptoms and no differences in soluble and nonsoluble carbohydrate concentrations were measured in infected compared with healthy plants, despite high CLas titers and significant reductions in plant biomasses. Most trees on rootstocks with trifoliate orange parentage were less damaged by HLB than other rootstocks, although results did not always agree with reported field performance. Among the different variables measured, leaf size appeared to be most predictive for grafted tree assessment of HLB sensitivity. The results of this study provide a better understanding of the strengths and weaknesses of assessing rootstock influence on grafted tree performance in a controlled greenhouse environment. Although such studies provide valuable information for cultivar tolerance to HLB, other rootstock traits will ultimately contribute to field survival and productivity in an HLB endemic production environment.

DETERMINATION OF ANTIBACTERIAL ACTIVITY OF PHYTOCHEMICALS TOWARDS XANTHOMONAS CITRI PV. CITRI CAUSING CITRUS CANKER

Citrus canker, caused by Xanthomonas citri pv. citri, is the most destructive disease of citrus throughout the world. Synthetic chemicals have so far proved ineffective and harmful to the environment. That is why in current study, phyto-extracts were examined due to their less hazardous and eco-friendly nature. For the management of citrus canker, nine medicinal phytoextract (C. colocynthis, N. sativa, C. tamala, Z. officinale, P. nigrum, C. verum, S. aromaticum, E. globu, T. Graecum) were evaluated under lab condition. Maximum inhibition zone was produced by C. colocynthis (25.71mm) followed by N. sativa (21.52mm), C. tamala (18.63mm), Z. officinale (20.42mm), P. nigrum (18.99mm). C. verum (15.73mm), S. aromaticum (18.75mm), E. globu (15.37mm), T. Graecum (14.57mm) as compared to control under lab conditions. The most effective phytoextract (C. colocynthis, N. sativa) were evaluated under greenhouse and field condition. Minimum disease severity (12.62%) was observed when of (C. colocynthis + N. sativa) were applied in combination followed by C. colocynthis (23.23%) and N. sativa (28.31%) as compared to the control under greenhouse condition. Maximum disease incidence was noticed by N. sativa (42.18%), C. colocynthis (36.79%) and minimum disease incidence (27.40%) was observed when (C. colocynthis+ N. sativa) were applied in combinations under field condition as compared to the control and reduce the disease severity up to 50%. The results showed that C. colocynthis+ N. sativa can be implicated for the management of citrus canker.

DETERMINATION OF ANTIBACTERIAL ACTIVITY OF PHYTOCHEMICALS TOWARDS XANTHOMONAS CITRI PV. CITRI CAUSING CITRUS CANKER

Citrus canker, caused by Xanthomonas citri pv. citri, is the most destructive disease of citrus throughout the world. Synthetic chemicals have so far proved ineffective and harmful to the environment. That is why in current study, phyto-extracts were examined due to their less hazardous and eco-friendly nature. For the management of citrus canker, nine medicinal phytoextract (C. colocynthis, N. sativa, C. tamala, Z. officinale, P. nigrum, C. verum, S. aromaticum, E. globu, T. Graecum) were evaluated under lab condition. Maximum inhibition zone was produced by C. colocynthis (25.71mm) followed by N. sativa (21.52mm), C. tamala (18.63mm), Z. officinale (20.42mm), P. nigrum (18.99mm). C. verum (15.73mm), S. aromaticum (18.75mm), E. globu (15.37mm), T. Graecum (14.57mm) as compared to control under lab conditions. The most effective phytoextract (C. colocynthis, N. sativa) were evaluated under greenhouse and field condition. Minimum disease severity (12.62%) was observed when of (C. colocynthis + N. sativa) were applied in combination followed by C. colocynthis (23.23%) and N. sativa (28.31%) as compared to the control under greenhouse condition. Maximum disease incidence was noticed by N. sativa (42.18%), C. colocynthis (36.79%) and minimum disease incidence (27.40%) was observed when (C. colocynthis+ N. sativa) were applied in combinations under field condition as compared to the control and reduce the disease severity up to 50%. The results showed that C. colocynthis+ N. sativa can be implicated for the management of citrus canker.

A New Perspective on Detection of Green Citrus Fruit in the Grove Using Deep Learning Neural Networks

The entire citrus industry is under attack by terminal citrus diseases such as Huanglongbing (HLB) or citrus greening (Candidatus Liberibacter asiaticus), and citrus canker (Xanthomonas axonopodis pv. citri). There is a need to develop fruit quality assessment techniques that can assess the quality of green citrus fruits during the growing season when green fruits are still on the trees. Learning technique is one of the fundamental techniques for fruit quality evaluation.

Species of Botryosphaeriaceae associated with citrus branch diseases in China.

Citrus is an important and widely cultivated fruit crop in South China. Although the species of fungal diseases of leaves and fruits have been extensively studied, the causal organisms of branch diseases remain poorly known in China. Species of Botryosphaeriaceae are known as important fungal pathogens causing branch diseases on citrus in the USA and Europe. To determine the diversity of Botryosphaeriaceae species associated with citrus branch diseases in China, surveys were conducted in the major citrus-producing areas from 2017 to 2020. Diseased tissues were collected from twigs, branches and trunks with a range of symptoms including cankers, cracking, dieback and gummosis. Based on morphological characteristics and phylogenetic comparison of the DNA sequences of the internal transcribed spacer region (ITS), the translation elongation factor 1-alpha gene (tef1), the β-tubulin gene (tub2) and the DNA-directed RNA polymerase II second largest subunit (rpb2), 111 isolates from nine provinces were identified as 18 species of Botryosphaeriaceae, including Botryosphaeria dothidea, B. fabicerciana, Diplodia seriata, Dothiorella alpina, Do. plurivora, Lasiodiplodia citricola, L. iraniensis, L. microconidia, L. pseudotheobromae, L. theobromae, Neodeightonia subglobosa, Neofusicoccum parvum, and six previously undescribed species, namely Do. citrimurcotticola, L. guilinensis, L. huangyanensis, L. linhaiensis, L. ponkanicola and Sphaeropsis linhaiensis spp. nov. Botryosphaeria dothidea (28.8 %) was the most abundant species, followed by L. pseudotheobromae (23.4 %), which was the most widely distributed species on citrus, occurring in six of the nine provinces sampled. Pathogenicity tests indicated that all 18 species of Botryosphaeriaceae obtained from diseased citrus tissues in this study were pathogenic to the tested Citrus reticulata shoots in vitro, while not all species are pathogenic to the tested Cocktail grapefruit (C. paradisi × C. reticulata) shoots in vivo. In addition, Lasiodiplodia was the most aggressive genus both in vitro and in vivo. This is the first study to identify Botryosphaeriaceae species related to citrus branch diseases in China and the results provide a theoretical basis for the implementation of prevention and control measures. Citation: Xiao XE, Wang W, Crous PW, et al. 2021. Species of Botryosphaeriaceae associated with citrus branch diseases in China. Persoonia 47: 106-135. https://doi.org/10.3767/persoonia.2021.47.03.

Understanding Citrus Greening Disease and Its Possible Management Strategies in Nepal

Huanglongbing (HLB), also known as citrus greening, is a devastating disease of citrus that has decimated several citrus orchards throughout the world. The disease is associated with three species of unculturable and phloem-limited bacteriae, Candidatus Liberibacter asiaticus, Candidatus Liberibacter africanus and Candidatus Liberibacter americanus. The most common species of bacteria found in Nepal is Candidatus Liberibacter asiaticus which is transmitted by an insect vector, Asian citrus psyllid (Diaphorina citri). This disease has been detected in several economically important citrus production areas of Nepal, which resulted in heavy yield loss. No cure for the disease has been discovered yet and it is essential to practice proper management strategies to maintain citrus health and sustain citrus production under HLB pressure. Several disease management approaches such as pathogen-free nursery establishment, use of disease tolerant rootstock cultivars, proper irrigation and nutrient supply, removal of HLB affected trees, and control of psyllid with frequent insecticide application are widely practiced throughout the world. This review article highlights the characteristics of the citrus greening disease and its insect vector and gives insights into their management techniques. Several technologically advanced options available to minimize the HLB infection might not be feasible currently in Nepal due to economic and topographic constraints. This article also aims to bring into focus the cost-effective methods that growers in Nepal can practice to mitigate the impact of HLB disease in their citrus orchards. Int. J. Appl. Sci. Biotechnol. Vol 9(4): 227-238.

The serial and generation intervals from SARS-CoV-2 transmission dynamics and their potential application in the epidemiology of two citrus diseases

<p>Since the start of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing the coronavirus disease 2019 (COVID-19) pandemic, the concepts of serial and generation intervals have been used as key epidemiological measures to understand the transmission dynamics of the disease. We carefully examined and repurposed these concepts to the understanding of the transmission chain and dynamics of two major citrus diseases: tristeza virus (caused by Citrus tristeza virus, CTV) and Huanglongbing (caused by <em>Candidatus</em> Liberibacter asiaticus). Following the fundamental definition of the concepts, the review delineates the transmission chain in the SARS-CoV-2 and that of CTV and CLas, pointing out their major similarities and differences. Then, it discusses estimation of the serial and generation intervals and their distributions for both plant diseases. Identification of infector-infectee tree pairs in a transmission chain within orchards is proposed through use of disease incidence data from intensive mapping, spatial pattern analysis, conditional probability, and simulation approaches. Like in SARS-CoV-2 dynamics, pre-symptomatic transmission in these two plant pathosystems is of epidemiological significance. Hence, estimation of the serial and generation interval can lay the foundations to understanding of early disease transmission dynamics, thus the implementation of vector control measures or eradication of infected trees. We hope this review motivates discussions on estimation and usage of these concepts to enhance understanding of the epidemiology of both of the herein examined citrus diseases.</p>

Wind Speed and Direction Drive Assisted Dispersal of Asian Citrus Psyllid.

Wind directly influences the spread of vector-borne plant pathogens by driving the passive dispersal of vectors to potentially new areas. Here, we evaluated the effect of wind speed and direction on the dispersal of the Asian citrus psyllid (ACP), Diaphorina citri (Kuwayama) (Hemiptera: Psyllidae), the vector of the bacteria causing huanglongbing (HLB), a lethal disease of citrus. The effect of different wind speeds on short or long-distance dispersal of ACP was investigated using a high-speed wind tunnel under laboratory conditions. The effect of wind direction on ACP dispersal under field conditions was evaluated using custom-made wind vane-style traps. In wind tunnel assays, ACP remained on plants until wind treatments reached ≥48 km/h when psyllids were mostly dislodged from plants and moved by the wind. For a short-distance, wind-driven movement (movement by the wind from one plant to another), the effect of wind speed was not significant at any of the wind speed treatments tested. Wind vane traps placed in a Florida citrus grove captured significantly more ACP on the windward side, suggesting that ACP were moved with the wind. The number of ACP found on the windward side of traps was significantly higher from May to August. These results indicate that ACP is likely to disperse with prevailing wind direction and that settled ACP may become dislodged and moved at random by high wind speeds occurring in areas of significant citrus production (southern California, Florida, or Texas).

Molecular signatures between citrus and Candidatus Liberibacter asiaticus.

Citrus Huanglongbing (HLB), also known as citrus greening, is one of the most devastating citrus diseases worldwide. Candidatus Liberibacter asiaticus (CLas) is the most prevalent strain associated with HLB, which is yet to be cultured in vitro. None of the commercial citrus cultivars are resistant to HLB. The pathosystem of Ca. Liberibacter is complex and remains a mystery. In this review, we focus on the recent progress in genomic research on the pathogen, the interaction of host and CLas, and the influence of CLas infection on the transcripts, proteins, and metabolism of the host. We have also focused on the identification of candidate genes for CLas pathogenicity or the improvements of HLB tolerance in citrus. In the end, we propose potentially promising areas for mechanistic studies of CLas pathogenicity, defense regulators, and genetic improvement for HLB tolerance/resistance in the future.

Detection of citrus diseases using a fuzzy neural network

The objective is to use AI techniques to build a citrus image recognition system and to produce an integrated program that will assist plant protection professionals in determining whether the disease is infected and early detection for the purpose of taking the necessary preventive measures and reducing its spread to other plants. In this research, the RBF and FRBF networks were used and applied to 830 images, to detect whether citrus fruits were healthy or ill. At first, the preprocessing of these images was done, and they were reduced to 250 x 250 pixels, and the features were extracted from them using the co-occurrence matrix method (GLCM) after setting the gray level at 8 gradients and 1 pixel distance, 21 statistical features were derived, and then these features were introduced to RBF after determine the number of input layer nodes by 21 , 20 for the hidden layer and 1 node for output layer, the centers were randomly selected from the training data and the weights were also randomly selected and trained using the Pseudo Inverse method. The RBF network was hybridized with the fuzzy logic using the FCM method, the fuzziness parameter = 2.3 was selected, and a new network called FRBF was acquired. These networks were trained and tested in training data (660 images) and testing (170 images) for citrus fruits. The detection rate was then calculated, and the results showed that the (FRBF) had a higher accuracy of 98.24% compared to RBF of 94.71%.

Compatibility of a Beta-cyfluthrin-Based 'Attract-and-Kill' Device with Tamarixia radiata (Hymenoptera: Eulophidae) for Suppression of Diaphorina citri (Hemiptera: Liviidae) on Residential Citrus.

The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), is a serious threat to the U.S. citrus industry because it spreads huanglongbing (HLB), a bacterial and incurable citrus disease. The nymphal parasitoid, Tamarixia radiata Waterston (Hymenoptera: Eulophidae), has been extensively released for biological control of D. citri in South Texas since 2010 but provides insufficient control. An 'attract-and-kill' (AK) device was evaluated for its compatibility with T. radiata for suppression of D. citri on dooryard citrus. The AK device is visually attractive to D. citri adults because it is the same color as young citrus flush and kills individuals on contact with the toxicant beta-cyfluthrin. This study evaluated 1) lethality of AK devices to T. radiata adults under lab conditions; 2) efficacy of AK devices for year-round psyllid suppression on individual dooryard lemon trees; 3) discovery and parasitism of D. citri colonies by T. radiata on lemon trees with or without AK devices. Contact with AK devices for 5 s or more was lethal to adult parasitoids. Deployment of 20 AK devices per tree provided significant year-round suppression of D. citri on infested lemon trees and reduced mean attack intensity (cumulative psyllid-days) of adults by 66% and nymphs by 82%. Discovery and parasitism rates of D. citri colonies by T. radiata were similar on control trees and trees protected by AK devices. An AK device that targets adult psyllids could be used to effectively complement biological control of D. citri by T. radiata in residential landscapes.

Parasitoid vectors a plant pathogen, potentially diminishing the benefits it confers as a biological control agent

Huanglongbing (HLB) is a destructive disease of citrus primarily transmitted by the Asian citrus psyllid (ACP). Biocontrol of ACP is an environmentally sustainable alternative to chemicals. However, the risk of parasitoid rational application in ACP biocontrol has never been evaluated. Here we show, the dominant parasitoid of ACP, Tamarixia radiata, can acquire the HLB pathogen Candidatus Liberibacter asiaticus (CLas) and transmit it horizontally when probing ACP nymphs. If these ACP nymphs survive the probing, develop to adults and move to healthy plants, CLas can be transmitted to citrus leaves during feeding. We illustrate the formerly unrecognized risk that a parasitoid can potentially serve as a phoretic vector of the pathogen transmitted by its host, thus potentially diminishing some of the benefits it confers via biocontrol. Our findings present a significant caution to the strategy of using parasitoids in orchards with different infection status of insect-vectored pathogens.

First Report of Xanthomonas citri subsp. citri causing Citrus Canker on lime in Rio Grande do Norte, Brazil.

Citrus canker caused by Xanthomonas citri subsp. citri is one of the most important citrus diseases in the world (Gottwald et al. 2002), mainly for citrus-producing countries with humid sub-tropical regions such as United States, Argentina, and Brazil, where losses may be significant (Behlau et al. 2020). In the state of Rio Grande do Norte (RN), Brazil, citrus production is expanding and shows social and economic importance for small farmers, which produced approximately 297 tons of lime in this state in 2019 (IBGE 2021). In December 2019, we observed symptoms of erumpent lesions with margins surrounded by yellow haloes on leaves and fruit of the lime (Citrus aurantifolia cv. 'Galego') (about 5% incidence) in a plantation located in the municipality of Mossoró, RN (05°12'21.1"S, 37°19'16"W). Samples were collected from the lime orchard, and five bacterial strains (CCRMXC01 to CCRMXC05) showing yellow, convex, mucoid colonies were isolated in a nutrient-yeast-dextrose-agar medium (NYDA). Pathogenicity tests were performed on sweet orange (C. sinensis cv. 'Pêra') and lime (C. latifolia cv. 'Tahiti') seedlings. Four wounds per leaf (upper side) were carried out with an entomological pin and 10 µl of a bacterial suspension (108 CFU mL-1) were deposited on each wound. The negative control consisted of leaves treated with sterile distilled water (SDW). For each citrus species, we used four replicates per strain and one leaf with four wounds per replicate. Inoculated leaves developed erumpent lesions with margins surrounded by yellow haloes six days after inoculation (DAI) in both citrus species, while leaves treated with SDW remained symptomless. Nine DAI, we reisolated the pathogen and performed rep-PCR (REP, ERIC, and BOX-PCR) analyses (Gama et al. 2018) with the strains inoculated and reisolated to confirm the identity of the strains and to fulfill Koch's postulates. The strains were stored at the Culture Collection Rosa Mariano (CCRM) of the Phytobacteriology Laboratory at the Universidade Federal Rural de Pernambuco. The five strains reisolated showed the same REP, ERIC, and BOX-PCR profiles as the strains used for inoculations. The molecular identification was performed sequencing the dnaK, fyuA, gyrB, and rpoD genes (Young et al. 2008). Each fragment was sequenced in both the forward and reverse directions. Using the BLASTn tool, we observed that sequences of the dnaK (GenBank MW218913 to MW218917), fyuA (GenBank MW218918 to MW218922), and rpoD (GenBank MW218928 to MW218932) genes of the strains CCRMXC01 to CCRMXC05 showed 100% of identity with the sequences of these genes from the type strain (ICMP 24T) and of other strains of X. citri subsp. citri (ICMP 21 and ICMP 7493), while sequences of gryB (GenBank MW218923 to MW218927) of the former strains showed 100% identity with the gyrB sequence of the strains ICMP 24T and ICMP 7493 and 99,85% identity with strain ICMP 21. This short variation in the sequence of the gyrB gene also may be observed among strains of X. citri subsp. citri available in NCBI database (https://www.ncbi.nlm.nih.gov/). The phylogenetic analysis performed using Bayesian inference and the concatenated sequence of all the type or representative strains of species and pathovars of Xanthomonas available in GenBank showed that the strains CCRMXC01 to CCRMXC05 clustered together with strain ICMP 24T with 1.0 posterior probability. To our information, this is the first report of X. citri subsp. citri causing citrus canker on lime in RN state, Brazil.

Parasitism of Diaphorina citri (Hemiptera: Liviidae) by Tamarixia radiata (Hymenoptera: Eulophidae) on residential citrus in Texas: Importance of colony size and instar composition

Diaphorina citri (Hemiptera: Liviidae) is an invasive citrus pest that colonizes young flush shoots and spreads huanglongbing, a lethal citrus disease. Effects of D. citri colony size (nymphs per shoot) and instar composition on parasitism rates by the nymphal ectoparasitoid, Tamarixia radiata (Hymenoptera: Eulophidae), were evaluated on residential citrus in South Texas. From 2016 to 17, D. citri colony sizes and apparent parasitism (T. radiata adult emergence) were monitored monthly on grapefruit, lemon, lime, and orange trees at 12 residential sites. In 2019, size and instar composition of colonies and actual parasitism (mummies and nymphs with parasitoid larvae or eggs) were monitored biweekly on citrus trees at a residential park. Psyllid nymphs were found year-round at these sites with highest densities from late summer to fall and lowest densities in winter. Mean apparent parasitism was independent of citrus species and ranged from 23% in April 2016 to 4% in February 2017. Mean actual parasitism ranged from 1.5% in August to 33.4% in November 2019. Because actual parasitism of 3rd or younger instars was very rare, a ‘host patch’ was defined as the cluster(s) of 4th to 5th instars within a colony infesting a shoot. Based on apparent or actual parasitism, host density was positively related to discovery of D. citri colonies but inversely related to percent parasitism by T. radiata. The inability of T. radiata to achieve high rates of field parasitism under a wide range of host densities could explain, in part, its low impact against D. citri on residential citrus.

Metabolomic analysis elucidates how shade conditions ameliorate the deleterious effects of greening (Huanglongbing) disease in citrus.

Under tropical and subtropical environments, citrus leaves are exposed to excess sunlight, inducing photoinhibition. Huanglongbing (HLB, citrus greening), a devastating phloem-limited disease putatively caused by Candidatus Liberibacter asiaticus, exacerbates this challenge with additional photosynthetic loss and excessive starch accumulation. A combined metabolomics and physiological approach was used to elucidate whether shade alleviates the deleterious effects of HLB in field-grown citrus trees, and to understand the underlying metabolic mechanisms related to shade-induced morpho-physiological changes in citrus. Using metabolite profiling and multinomial logistic regression, we identified pivotal metabolites altered in response to shade. A core metabolic network associated with shade conditions was identified through pathway enrichment analysis and metabolite mapping. We measured physio-biochemical responses and growth and yield characteristics. With these, the relationships between metabolic network and the variables measured above were investigated. We found that moderate-shade alleviates sink limitation by preventing excessive starch accumulation and increasing foliar sucrose levels. Increased growth and fruit yield in shaded compared with non-shaded trees were associated with increased photosystem II efficiency and leaf carbon fixation pathway metabolites. Our study also shows that, in HLB-affected trees under shade, the signaling of plant hormones (auxins and cytokinins) and nitrogen supply were downregulated with reducing new shoot production likely due to diminished needs of cell damage repair and tissue regeneration under shade. Overall, our findings provide the first glimpse of the complex dynamics between cellular metabolites and leaf physiological functions in citrus HLB pathosystem under shade, and reveal the mechanistic basis of how shade ameliorates HLB disease.

Smartphone assist deep neural network to detect the citrus diseases in Agri-informatics

The citrus family provides healthy fruits to humans. The quality and quantity of the citrus fruits depend on the quality of the citrus leaves. Due to the diseases of citrus leaves, the quality and productivity of citrus fruit are degraded. This paper provides a computer automation system to detect the diseases of the citrus leaves using machine learning and deep learning techniques. In this paper, images of citrus leaves are captured using an Android Smartphone in natural environmental light. Three different classes of citrus leaves are collected using the Smartphone and destructive method. These are citrus healthy, citrus greening, and citrus CTV (Citrus Tristeza virus). Citrus images are processed for image resizing, image noise removal, image enhancement, and feature extraction. Features of the images are calculated using Gray Level Co-occurrence Matrix in the color and gray domain of the images. Finally, the detection and classification are done using K- Nearest Neighbor (KNN) and Deep Neural Network (DNN) classifier. The accuracy of the KNN classifier is 89.9% for the K value 3 whereas the accuracy of DNN is 99.89% with an error of 0.0219. The proposed system helps the farmer to detect diseases at an early stage.

Photodynamic control of citrus crop diseases.

Citrus are economically important fruit crops to which infectious diseases like citrus canker caused by Xanthomonas citri subs. citri, citrus variegated chlorosis caused by Xylella fastidiosa, "huanglongbing" associated with the presence of Candidatus liberibacter species, anthracnose caused by Colletotrichum gloeosporioides and citrus black spot caused by Phyllosticta citricarpa, impose significant losses. Control measures involve chemical treatment of orchards but often, eradication of infected plants is unavoidable. To circumvent the environmental impacts of pesticides and the socio-economic impacts of eradication, innovative antimicrobial approaches like photodynamic inactivation are being tested. There is evidence of the susceptibility of Xanthomonas citri subs. citri and C. gloeosporioides to photodynamic damage. However, the realistic assessment of perspectives for widespread application of photodynamic inactivation in the control of citrus diseases, necessarily implies that other microorganisms are also considered. This review intends to provide a critical summary of the current state of research on photodynamic inactivation of citrus pathogens and to identify some of the current limitations to the widespread use of photodynamic treatments in citrus crops.

Sensitive and Rapid Detection of Citrus Scab Using an RPA-CRISPR/Cas12a System Combined with a Lateral Flow Assay.

Citrus is the most extensively produced fruit tree crop in the world and is grown in over 130 countries. Fungal diseases in citrus can cause significant losses in yield and quality. An accurate diagnosis is critical for determining the best management practices and preventing future losses. In this study, a Recombinase polymerase amplification (RPA)-clustered regularly interspaced short palindromic repeats (CRISPR)/associated (Cas) system was established with the integration of a lateral flow assay (LFA) readout system for diagnosis of citrus scab. This detection can be completed within 1 h, is highly sensitive and prevents cross-reactions with other common fungal citrus diseases. Furthermore, the detection system is compatible with crude DNA extracted from infected plant tissue. This RPA-CRISPR/Cas12a-LFA system provides a sensitive, rapid, and cost-effective method with promising and significant practical value for point-of-care diagnosis of citrus scab. To our knowledge, this is the first report to establish an RPA- and CRISPR-based method with LFA for fungal diseases in plants.