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2022 ◽  
Author(s):  
Sagheer Atta

Citrus is a valuable crop in Pakistan because it is rich in vitamin C and antioxidants. Huanglongbing (HLB) has an influence on citrus production around the world caused by a bacterium “ Candidatus   liberibacter asiaticus ” (CLas), africanus and americanus. The structure and diversity of bacterial species in various ecosystems can be quickly examined using NGS. This approach is considerably quicker and more precise than outdated methods. Healthy or citrus greening infected leaf samples of Grapefruit,  Citrus aurantifolia , and  Citrus reticulata  Blanco was used for diversity analysis. In this study high throughput, NGS technique was used to access the population of both cultivable and non-cultivable bacterial endophytes from citrus leaves, by using PCR amplicons of 16S rDNA sequences (V5–V7 regions) with Illumina Hi seq. As a result, a total number of 68,722 sequences were produced from the test samples. According to the NGS-based diversity classification, the most common genera of exploited bacterial endophytes were Proteobacteria, Firmicutes, Bacteroides, Cyanobacteria, and Actinobacteria.  Citrus aurantifolia  and  Citrus paradisi  showed almost equal diversity, whereas  Citrus reticulata  Blanco had a higher proportion of Proteobacteria and Cyanobacteria in their leaves. To determine alpha diversity (AD), additional data was analyzed using statistical indices such as Shannon, Chao1, and Simpson. According to the inverse Simpson diversity index, the abundance of the microbial population in six different citrus samples was 0.48, 0.567, and 0.163, respectively. The metagenomics of microbiota in plant tissues was successfully recorded by NGS technology, which can help us learn more about the interactions between plants and microbes. This research is the first step toward a better understanding of 16SrRNA-based metagenomics from citrus in Pakistan using Illumina (Hi seq) Technology.


2022 ◽  
Vol 52 (3) ◽  
Author(s):  
Paulo Henrique Nascimento de Souza ◽  
Lilian Maria Arruda Bacchi ◽  
Walber Luiz Gavassoni ◽  
Wagner da Paz Andrade

ABSTRACT: In recent years, there have been reductions in the efficacy of the fungicidal control of Phakopsora pachyrhizi, thereby hindering the management of soybean rust and compromising crop yield. This study evaluated the effects of incorporating hydrogen peroxide (H2O2) in commercial fungicide formulations on the control of soybean rust. We conducted two experiments, one of which was performed in a greenhouse environment and the other under field conditions. In both environments, we examined the following four control programs using commercial fungicide formulations: (I) azoxystrobin + cyproconazole (quinone outside inhibitor [QoI] + demethylation inhibitor [DMI]); (II) picoxystrobin + cyproconazole (QoI + DMI); (III) pyraclostrobin + epoxiconazole + fluxapyroxad (QoI + DMI + succinate dehydrogenase inhibitor); and (IV) water (H2O) (program without fungicide application), combined with the incorporation of (i) H2O2; (ii) mancozeb (positive control I); (iii) chlorothalonil (positive control II); or (iv) water (H2O) alone. Analyses of infected leaf area and grain yield revealed that the addition of H2O2 to the formulations of DMI and QoI fungicides led to a reduction in disease severity of between 33% and 44% relative to the effects of these products used alone, as well as an increase in yield and SPAD values. The use of H2O2 and multi-site fungicides alone failed to provide effective control of soybean rust. In addition to enhancing the efficacy of disease control, the use of H2O2 associated with commercial fungicide mixtures was shown to be a potential tool for the management of fungicide resistance and reduction in losses from Asian soybean rust.


Agronomy ◽  
2021 ◽  
Vol 12 (1) ◽  
pp. 97
Author(s):  
Liang Gong ◽  
Chenrui Yu ◽  
Ke Lin ◽  
Chengliang Liu

Powdery mildew is a common crop disease and is one of the main diseases of cucumber in the middle and late stages of growth. Powdery mildew causes the plant leaves to lose their photosynthetic function and reduces crop yield. The segmentation of powdery mildew spot areas on plant leaves is the key to disease detection and severity evaluation. Considering the convenience for identification of powdery mildew in the field environment or for quantitative analysis in the lab, establishing a lightweight model for portable equipment is essential. In this study, the plant-leaf disease-area segmentation model was deliberately designed to make it meet the need for portability, such as deployment in a smartphone or a tablet with a constrained computational performance and memory size. First, we proposed a super-pixel clustering segmentation operation to preprocess the images to reduce the pixel-level computation. Second, in order to enhance the segmentation efficiency by leveraging the a priori knowledge, a Gaussian Mixture Model (GMM) was established to model different kinds of super-pixels in the images, namely the healthy leaf super pixel, the infected leaf super pixel, and the cluttered background. Subsequently, an Expectation–Maximization (EM) algorithm was adopted to optimize the computational efficiency. Third, in order to eliminate the effect of under-segmentation caused by the aforementioned clustering method, pixel-level expansion was used to describe and embody the nature of leaf mildew distribution and therefore improve the segmentation accuracy. Finally, a lightweight powdery-mildew-spot-area-segmentation software was integrated to realize a pixel-level segmentation of powdery mildew spot, and we developed a mobile powdery-mildew-spot-segmentation software that can run in Android devices, providing practitioners with a convenient way to analyze leaf diseases. Experiments show that the model proposed in this paper can easily run on mobile devices, as it occupies only 200 M memory when running. The model takes less than 3 s to run on a smartphone with a Cortex-A9 1.2G processor. Compared to the traditional applications, the proposed method achieves a trade-off among the powdery-mildew-area accuracy estimation, limited instrument resource occupation, and the computational latency, which meets the demand of portable automated phenotyping.


Plant Disease ◽  
2021 ◽  
Author(s):  
Wu Zhang ◽  
Xiu Li Song

The pygmy date palm (Phoenix roebelenii) is a popular ornamental plant widely cultivated in tropical regions as well as in China. In June 2018, a new leaf spot symptoms were observed on P. roebelenii in several different parks in Zhanjiang City of China. The early symptoms of infected leaves were presented with small, round, pale brown spots. As the size of these spots increased, they coalesced to form larger irregular necrotic lesions surrounded by dark brown edges, which eventually led to leaf wilted and defoliation. A filamentous fungus was consistently isolated from infected leaf samples. Colonies on PDA at 25°C (12 h light/dark) were initially white with abundant aerial mycelium, which turned fluffy and dark olivaceous after one-week culture. Pycnidial conidiomata were black and globose and formed on pine needles in water agar at 25°C (12 h light/dark) after 21 days. Conidiogenous cells were hyaline, cylindrical, holoblastic. The conidia was ovoid to ellipsoid, thick-walled, which was initially hyaline and aseptate, later turned into dark brown and 1-septate with a striate appearance to conidia, 11.6~25.0 μm×9.6~12.0 μm (av. 20.4 μm×10.1 μm). For molecular identification, the partial sequences of internal transcribed spacer (ITS) regions, translation elongation factor (EF-1α) and β-tubulin (TUB) genes of two representative isolates RYCK-1, RYCK-2 were amplified and sequenced using primer pairs ITS/ITS4 (White et al. 1990), EF-688F/EF-986R (Carbone and Kohn 1999), and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. The sequences of the above three loci of the two isolates (accession nos. ITS, OK329968 and OK329969; EF-1α, OK338067 and OK338068; TUB, OK338069 and OK338070) showed 98.4-100.0 % identity with the existing sequences of ex-type culture CBS 122528 of N. phoenicum. A multilocus phylogenetic analysis of the three loci concatenated sequences using the maximum likelihood method showed the isolates that belongs to N. phoenicum. Based on the morphological characteristics and molecular analysis of the isolates, the fungus was identified as N. phoenicum (Phillips et al. 2008). To confirm pathogenicity, five one-year-old potted plants were used for each isolate (RYCK-1 and RYCK-2) and the plants were inoculated by pricking the epidermis of the leaf with a needle. Five leaves of each plant were sprayed with 100 µl of a conidial suspension (1 × 106 conidia/ml) to the wounded surface for each plant. Sterilized distilled water was used as the control and the experiment was repeated. All the plants were incubated at 26 ± 2°C (12 h light/dark) and covered with plastic bags to maintain constant high humidity. After 14 days, all the inoculated leaves showed the same symptoms as those observed in the original diseased plants, but the control plants remained health. The reisolated fungus was identified as N. phoenicum by morphological and molecular characteristics. N. phoenicum is an important pathogen of Phoenix species plants worldwide, which have been reported to cause shoot blights and stalk rots on P. dactylifera and P. canariensis in Greece (Ligoxigakis et al. 2013) and root rot on P. dactylifera in Qatar (Nishad and Ahmed 2020). To our knowledge, this is first report N. phoenicum causing leaf spot on P. roebelenii in China.


Author(s):  
D. O. Etim ◽  
N. I. Okon

Momordica charantia Linn, commonly called bitter-melon or ampalaya, is a vigorous, tendril-bearing, frost tender, annual vine of the cucurbitacae family reported to play significant role in health and nutrition. Chlorotic spot symptoms were observed on this plant during a visit to some Gardens in Baccocco Cross River State, Nigeria in 2021. The aim of this study was to identify the virus infecting this plant and investigate the ability of Trichoderma viride to control the virus. Infected leaf samples of M. charantia were collected and maintained on young seedlings of cucumber through mechanical inoculation which was further used to test the ability of Trichoderma viride as biocontrol agent, the sample was further tested against RT-PCR. Result obtained from Gene sequence analysis revealed 87 % nucleotide sequence identity with Morroccan watermelon mosaic virus. This is the first report of MWMV infecting M. charantia in Nigeria. The result further showed that Trichoderma viride was very effective in the control of virus the pathogen.


Plant Methods ◽  
2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Leonardo Furci ◽  
David Pascual-Pardo ◽  
Jurriaan Ton

Abstract Background The bacterial leaf pathogen Pseudomonas syringae pv tomato (Pst) is the most popular model pathogen for plant pathology research. Previous methods to study the plant-Pst interactions rely on destructive quantification of Pst colonisation, which can be labour- and time-consuming and does not allow for spatial–temporal monitoring of the bacterial colonisation. Here, we describe a rapid and non-destructive method to quantify and visualise spatial–temporal colonisation by Pst in intact leaves of Arabidopsis and tomato. Results The method presented here uses a bioluminescent Pst DC3000 strain that constitutively expresses the luxCDABE operon from Photorhabdus luminescens (Pst::LUX) and requires a common gel documentation (Gel Doc) system with a sensitive CCD/CMOS camera and imaging software (Photoshop or Image J). By capturing bright field and bioluminescence images from Pst::LUX-infected leaves, we imaged the spatiotemporal dynamics of Pst infection. Analysis of bioluminescence from live Pst bacteria over a 5-day time course after spray inoculation of Arabidopsis revealed transition of the bacterial presence from the older leaves to the younger leaves and apical meristem. Colonisation by Pst:LUX bioluminescence was obtained from digital photos by calculating relative bioluminescence values, which is adjusted for bioluminescence intensity and normalised by leaf surface. This method detected statistically significant differences in Pst::LUX colonisation between Arabidopsis genotypes varying in basal resistance, as well as statistically significant reductions in Pst::LUX colonisation by resistance-inducing treatments in both Arabidopsis and tomato. Comparison of relative bioluminescence values to conventional colony counting on selective agar medium revealed a statistically significant correlation, which was reproducible between different Gel Doc systems. Conclusions We present a non-destructive method to quantify colonisation by bioluminescent Pst::LUX in plants. Using a common Gel Doc system and imaging software, our method requires less time and labour than conventional methods that are based on destructive sampling of infected leaf material. Furthermore, in contrast to conventional strategies, our method provides additional information about the spatial–temporal patterns of Pst colonisation.


Plant Disease ◽  
2021 ◽  
Author(s):  
Hafiz Muhammad Usman Aslam ◽  
Nasir Ahmad Khan ◽  
Syed Ismat Hussain ◽  
Yasir Ali ◽  
Muhammad Raheel ◽  
...  

Brown leaf spot of rice is one of the major seed-borne diseases and can diminish grain production up to 52% (Barnwal et al. 2013). In 2018, infected leaf samples showing the typical symptoms of brown spots were collected from the vicinity of the University of Agriculture, Faisalabad (31°26'10.3"N 73°03'35.1"E). The symptoms were brown-dark spots, with gray-light gray or brown centers surrounded by dark margins and with chlorotic halos and of oval or cylindrical shapes (5 to 9 mm in diameter). Disease incidence averaged 61% across the seven fields observed. Leaves were collected from the seven infected fields and symptomatic leaf tissues of 5 mm2 were excised from representative necrotic spots in each. These tissues were surface disinfected with 70% ethanol, rinsed with sterile distilled water (SDW), dried by blotting on paper, and placed on potato dextrose agar medium. For pathogen growth, the plates were placed at 25oC (±2oC) with a 12-hour photoperiod for 5 days. Five samples from each of the infected fields were taken for pathogen isolation and among them ten isolates were sub-cultured and purified by using the single spore method. The resulting fungal colonies were fluffy and ranged in color from grayish black/black to light brown. Fifteen conidia were measured that are olivaceous-brown to dark brown in color, elliptical to oblong with narrow (tapered) ends, with 3-10 septa and 35.6-65.4 µm in length x 13.1-25.7 µm in width. Conidiophores were yellowish-brown, geniculate, and solitary (Pratt 2003). For molecular studies, rDNA of the internal transcribed spacer (ITS) region, translation elongation factor (tef), RNA polymerase II second largest subunit (rpb2) and glyceraldehyde-3-phosphate dehydrogenase (gpd) gene were amplified by using the primers ITS1F/ITS4R (White et al. 1990), EF1-983F/EF1-2218R (Rehner and Buckley 2005), 5F2/7CR (O’Donnell et al. 2007), and GPD1/GPD2 (Berbee et al. 1999) respectively. The sequence of all the amplified gene regions of one SUL-1 isolate was deposited into GenBank with accession numbers MN314844 (ITS), MN326866 (tef), MN990457 (rpb2) and MN990456 (gpd). BLASTn queries of the obtained sequences (ITS, tef, rpb2 and gpd) showed 99-100% homology with the corresponding nucleotide sequences of B. sorokiniana (GenBank accession nos. GU480767, MF490855, LT715652 and MK558818 respectively). To fulfill the Koch’s postulates, twenty rice plants (cv. Basmati-385) were sprayed at 2 to 3 leaf stages by using the two representative isolates with a spore suspension of 105 spores/ml. SDW was sprayed on ten control plants. The plants were covered with polyethylene bags to keep the moisture contents and incubated at 25oC (±2oC) for 7 days. After a week, same symptoms as those described above were observed. In the repeated experiment, B. sorokiniana was re-isolated from the infected rice leaves and confirmed morphologically; fulfill the Koch’s postulates. With grave worry, the other species of the genus Bipolaris (B. oryzae, and B. victoriae) have also been found to the cause brown leaf spot of rice (Motlagh and Kaviani 2008). To our knowledge, this is the first report of Bipolaris sorokiniana causing brown leaf spot of rice in Pakistan. Because rice is highly consumable grain in Pakistan, so the rapid spread of this disease in the rice farming areas is of a serious concern.


Plant Disease ◽  
2021 ◽  
Author(s):  
Ross Joaquin Hatlen ◽  
Douglas S. Higgins ◽  
Julien Venne ◽  
J. Alejandro Rojas ◽  
Mary Hausbeck ◽  
...  

Halo blight of hop caused by Diaporthe humulicola has recently been reported in Michigan and Connecticut (Higgins et al. 2021, Allan-Perkins et al 2020). In August 2020 growers in Quebec, Canada reported necrotic foliar lesions and desiccation of the hop strobile (cone) on Chinook and Nugget cultivars. The foliar lesions were dry concentric circles with a chlorotic halo surrounding the lesions; no pycnidia were observed on leaves or cones. Up to 100% of the infected bract tissue was dry and easily shattered, the grower estimated that more than 90% of the plants in the hopyard exhibited symptoms. Twenty-six isolates were obtained from surface-sterilized leaf and cone tissue by plating the leading edge of lesions on potato dextrose agar. Fungal isolates were hyphal tipped and were incubated at 22°C with a 12 h photoperiod. After 21-days, all cultures were white to beige with pycnidia. DNA was extracted from cultures using the MagMAX Plant DNA Isolation Kit (Applied Biosystems, Foster City, CA). DNA amplification of a representative isolate (CD6C) was performed with primers ITS1/ITS4 (White et al. 1990) for the internal transcribed spacer (ITS), CYLH3F/H3-1b (Glass and Donaldson 1995) for histone 3 (HIS), and Ef1728f/EF1-986R (Carbone and Kohn 1999) for translation elongation factor 1-α (TEF). Amplification primers were used for bidirectional Sanger sequencing, reads were assembled using Geneious Prime (Biomatters, New Zealand), and identified using NCBI BLAST. BLAST results showed that the sequences for TEF, ITS, and HIS all had 100% pairwise identity to Diaporthe sp. 1-MI (MT909101, MT909099, MT909093, OK001342, MZ934713, OK001341). Futhermore, BLAST results showed that ITS and HIS have 100% pairwise identity D. humulicola (MN152929, MN180214). The TEF sequence also had 99.7% pairwise identity to D. humulicola (MN180209). Koch’s postulates were conducted by inoculating six 3-mo-old ‘Chinook’ plants with conidia harvested from 28-day-old cultures and spraying 50 ml of inoculum (6 x 105 conidia/ml) or water to each plant. Plants were then stored in a greenhouse at 100% relative humidity at 22°C with a 14-h photo period. Lesions appeared on the adaxial side of the leaf after 21 days. D. humulicola was re-isolated from all infected leaf tissue, but not from any water inoculated plants and identified by conidial morphology using descriptions from Higgins et al. (2021). So far, Diaporthe sp. 1-MI appears to be synonymous with Diaporthe humulicola, but currently two names are being utilized (i.e. Diaporthe leaf spot and halo blight). In Higgins et al., (2021) it was proposed that the name halo blight might be more appropriate because disease symptoms are not confined to the leaves and cause significant blighting of cones. Halo blight caused by D. humulicola appears widespread in Michigan and Canada and may become an issue in other eastern North American growing regions with humid conditions.


Forests ◽  
2021 ◽  
Vol 12 (11) ◽  
pp. 1574
Author(s):  
Thomas R. Gaskin ◽  
Max Tischendorf ◽  
Ines Günther ◽  
Marius Rehanek ◽  
Carmen Büttner ◽  
...  

We identified a novel virus in diseased European ash (Fraxinus excelsior) and manna ash (F. ornus) trees exhibiting chlorotic ringspots, mottle and leaf deformation such as curling and shoestring symptoms. High-throughput sequencing (HTS, Illumina RNASeq) of total RNA isolated from diseased leaf material in combination with RT-PCR-based amplification techniques and Sanger sequencing determined five complete genome segments, each encoding a single open reading frame. Sequence analyses of RNA1–RNA5 revealed a genome organization typical for emaraviruses, i.e., (i) conserved and complementary terminal 5′and 3′termini of each genome segment (ii) proteins showing significant homologies to the RNA-dependent RNA polymerase (RdRP) encoded by RNA1, the glycoprotein precursor (GPP) encoded by RNA2, the viral nucleocapsid protein (N, RNA3), the movement protein (MP, RNA4), and a protein of 26 kDA (P26, RNA5) highly similar to proteins of unknown function encoded by other emaraviruses. Furthermore, we identified spherical particles (double-membrane bodies, DMB) of different sizes (70–80 nm in diameter) which are typical for emaraviruses exclusively in virus-infected leaf tissue exhibiting mottle and leaf deformation. Sequence comparison and phylogenetic analyses confirmed the identified novel virus as a new member of the genus Emaravirus. We established a species-specific RT-PCR detection protocol and could associate the observed disease symptoms with the infection of the novel emaravirus in F. excelsior and F. ornus. Therefore, we propose the name ash shoestring-associated emaravirus (ASaV). Investigation of ASaV-infected sample trees originating from different locations in Switzerland, Germany, Italy and Sweden provided a wide geographical distribution of the virus in affected ash species. To our knowledge, this is the first confirmation of an emaravirus affecting ash tree species with shoestring symptoms of leaves in Europe.


Author(s):  
Bharathi C

Abstract: The main walk of life of our Country is Agriculture. More than 70 % of the population’s lives depend upon agriculture. It is also a great source of country’s economy. In order to make this filed more profitable for farmers proper crops have to be grown in their fields. The prevalent problem among the farmers is Crop choice depending upon the soil in their farmlands. Another challenge faced by farmers is choosing the right fertilisers for their crops, which plays a very important role in getting a good and profitable yield. There is another major problem which they have to give more attention is the pest control or the diseases to which the plants may limit their growth. The above listed problems may solved using the advanced techniques of Precision Agriculture and data mining. Precision Agriculture is modern technique which can be used for farming. The main objective of is to solve above problems using data mining techniques and build a decision system which would help farmers to choose right crops for their farm , fertiliser recommendation for the crops grown and also to help the farmers in detecting the diseases by using the infected leaf images.. Keywords: Precision Agriculture, Data Mining, Crop, fertilizer recommendation system, ML Algorithms


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