scholarly journals Comparison of Molecular Procedures for Detection and Identification of Guignardia citricarpa and G. mangiferae

Plant Disease ◽  
2007 ◽  
Vol 91 (5) ◽  
pp. 525-531 ◽  
Author(s):  
N. A. Peres ◽  
R. Harakava ◽  
G. C. Carroll ◽  
J. E. Adaskaveg ◽  
L. W. Timmer
Keyword(s):  
Extraction Procedure ◽  
Citrus Fruit ◽  
Black Spot ◽  
Polymerase Chain Reaction Primer ◽  
Internal Transcribed Spacer Region ◽  
Guignardia Citricarpa ◽  
Detection And Identification ◽  
Specificity And Sensitivity ◽  
Fruit Spot ◽  
Primer Sets

Citrus black spot, caused by Guignardia citricarpa, is a serious fruit spot disease and is widely distributed in Asia, southern Africa, and South America, but does not occur in North America or the Mediterranean region. A nonpathogenic species, G. mangiferae, is cosmopolitan with a wide host range and can colonize citrus fruit and leaves saprophytically. Detection and identification of Guignardia spp. on citrus fruit is necessary for epidemiological, management, and regulatory purposes. In this study, we compared published and unpublished polymerase chain reaction primer sets for their specificity and sensitivity in the detection and differentiation of the two Guignardia spp. All primers evaluated successfully identified the two species using purified DNA from fungal cultures or mycelia as source materials. However, some primer sets were not highly effective in detecting G. citricarpa when DNA was extracted directly from single characteristic black spot lesions on fruit. Thus, new primer pairs for both species were designed from the internal transcribed spacer region that were highly sensitive and specific for detection of G. citricarpa using DNA recovered from single lesions on fruit by a rapid DNA extraction procedure.

Guignardia citricarpa. [Descriptions of Fungi and Bacteria].

1966 ◽  
Author(s):  
B. C. Sutton
Keyword(s):  
Geographical Distribution ◽  
Citrus Fruit ◽  
Black Spot ◽  
Minor Role ◽  
Latent Infections ◽  
Green Ring ◽  
Guignardia Citricarpa ◽  
Fiji Islands ◽  
Citrus Leaves ◽  
A Minor

Abstract A description is provided for Guignardia citricarpa. Information is included on the disease caused by the organism, its transmission, geographical distribution, and hosts. HOSTS: On Citrus spp. Also recorded in a non-pathogenic form on many other hosts in the following families: Anacardiaceae, Aquifoliaceae, Bignoniaceae, Burseraceae, Cunoniaceae, Dioscoreaceae, Gramineae, Lauraceae, Leguminosae, Liliaceae, Lythraceae, Magnoliaceae, Myrtaceae, Orchidaceae, Passifloraceae, Proteaceae, Rosaceae, Rutaceae, Solanaceae, Sterculiaceae, Theaceae (29: 208; 43: 1922a; Herb. IMI). DISEASE: Black spot of citrus. Kiely (1949) described 3 types of fruit lesion: Hard spot and shot-hole spot numerous, at first circular, brown with slight depressions, later more depressed in the centre which turns grey-white, margin black and surrounded by a ring of green rind tissue; Freckle spot develops after hard spot phase with abundant lesions, small, deep organge to brick red, finally brown, lacking a green ring, Virulent spot, irregular, confluent, rapidly spreading, black in the centre where pycnidia are produced, brown nearer the edge, finally brick red at the periphery forming the margin of the sunken lesion. McOnie found field temperatures affected symptom expression (44: 1556b). Also occurs on leaves, twigs and flowers of citrus and other hosts, often as latent infections. GEOGRAPHICAL DISTRIBUTION: Widely distributed on a large number of hosts. Countries where the fungus has been reported as a pathogen of citrus fruit in the field are designated below with an asterisk: Africa (? Egypt, Kenya, Mozambique, Nigeria, *Rhodesia, *South Africa, Uganda), Asia (Ceylon, *China, *Formosa (Taiwan), Hong Kong,? India, *Indonesia, Iran, Israel, *Japan, Korea,? Malaya, Okinawa,? Pakistan,? Philippines,? Singapore,? Thailand,? Vietnam); Australasia & Oceania, (*Australia,? Fiji Islands,? Hawaii, New Hebrides), Europe (Sicily, Spain, U.S.S.R. (Republic of Georgia)); North America (United States (Florida)); Central America and the Caribbean (Honduras, Jamaica, Trinidad); South America (*Argentine, *Brazil, *Peru, Venezuela). (CMI Map 53, ed. 3, 1961; 29: 208; 44, 701; Herb. IMI). TRANSMISSION: By air-borne ascospores from perithecia produced on cirrus leaf litter (43, 1922b). Conidia produced from pycnidia require water droplets for emergence and dispersal. They play a minor role by contributing to the infection of low hanging fruit. Mycelium in latent infections of citrus leaves remains viable up to 18 days and may be detected when the leaves are incubated at 30°C (Kiely, 1949). Nursery trees may also carry latent infection into other citrus growing areas (Kiely, 1949; Wager, 1953). Wounds on fruit made by insects may also provide infection courts (16: 247, 22: 19).

scholarly journals Chitosan reduces infection by Guignardia citricarpa in postharvest 'Valencia' oranges

2009 ◽  
Vol 52 (3) ◽  
pp. 513-521 ◽  
Author(s):  
Maria Cristina Canale Rappussi ◽  
Sérgio Florentino Pascholati ◽  
Eliane Aparecida Benato ◽  
Patrícia Cia
Keyword(s):  
Citric Acid ◽  
Biochemical Analysis ◽  
Control Measure ◽  
Citrus Fruit ◽  
Black Spot ◽  
Resistance Induction ◽  
Guignardia Citricarpa ◽  
Black Spots ◽  
Room Condition

Citrus fruits are affected by the black spot disease caused by Guignardia citricarpa. Resistance induction is an alternative control measure and chitosan exhibits potential as resistance inducer. The effect of chitosan on G. citricarpa was evaluated in vitro and in 'Valencia' oranges. Citrus fruit were immersed into different chitosan concentrations. Chitosan (2%), combined with or without thiabendazole and the citric acid was also investigated. All the chitosan concentrations inhibited G. citricarpa mycelial growth and affected morphologically the conidial germination and appressorium formation. Chitosan inhibited the development of new lesions in oranges at room condition or under refrigeration. Thiabendazole and citric acid did not reduce the formation of lesions. Biochemical analysis revealed that chitinase, β-1,3-glucanase, peroxidase and polyphenoloxidase activities were increased in chitosan-treated fruits. Thus, the effect of chitosan on the reduction of black spots in 'Valencia' oranges could be due to the germicidal effect on the pathogen and/or resistance induction in the fruit.

scholarly journals Multiplex End-Point PCR for the Detection of Three Species of Ophiosphaerella Causing Spring Dead Spot of Bermudagrass

Plant Disease ◽  
2019 ◽  
Vol 103 (8) ◽  
pp. 2010-2014 ◽  
Author(s):  
J. Francisco Iturralde Martinez ◽  
Francisco J. Flores ◽  
Alma R. Koch ◽  
Carla D. Garzón ◽  
Nathan R. Walker
Keyword(s):  
Rna Polymerase Ii ◽  
Elongation Factor ◽  
Temperature Cycling ◽  
Molecular Technique ◽  
Correct Identification ◽  
Internal Transcribed Spacer Region ◽  
Spring Dead Spot ◽  
End Point ◽  
Specificity And Sensitivity ◽  
Species Specific

A multiplex end-point polymerase chain reaction (PCR) assay was developed for identifying the three-fungal species in the genus Ophiosphaerella that cause spring dead spot (SDS), a devastating disease of bermudagrass. These fungi are difficult to identify by morphology because they seldom produce pseudothecia. To achieve species-specific diagnosis, three pairs of primers were designed to identify fungal isolates and detect the pathogen in infected roots. The internal transcribed spacer region, the translation elongation factor 1-α, and the RNA polymerase II second-largest subunit were selected as targets and served as templates for the design of each primer pair. To achieve uniform melting temperatures, three to five random nucleotide extensions (flaps) were added to the 5′ terminus of some of the designed specific primers. Temperature cycling conditions and PCR components were standardized to optimize specificity and sensitivity of the multiplex reaction. Primers were tested in multiplex on DNA extracted from axenic fungal cultures and from field-collected infected and uninfected roots. A distinct amplicon was produced for each Ophiosphaerella sp. tested. The DNA from Ophiosphaerella close relatives and other common bermudagrass pathogens did not amplify during the multiplex assay. Metagenomic DNA from infected bermudagrass produced species-specific amplicons while DNA extracted from noninfected roots did not. This multiplex end-point PCR approach is a sensitive and specific molecular technique that allows for correct identification of SDS-associated Ophiosphaerella spp. from field-collected roots.

scholarly journals First Report of Black Spots on Avocado Fruit Caused by Neofusicoccum parvum in Mexico

Plant Disease ◽  
2012 ◽  
Vol 96 (2) ◽  
pp. 287-287 ◽  
Author(s):  
E. Molina-Gayosso ◽  
H. V. Silva-Rojas ◽  
S. García-Morales ◽  
G. Avila-Quezada
Keyword(s):  
Average Length ◽  
Black Spot ◽  
Persea Americana ◽  
Universal Primers ◽  
Internal Transcribed Spacer Region ◽  
Pathogenic Potential ◽  
Neofusicoccum Parvum ◽  
First Report ◽  
Black Spots ◽  
Avocado Fruit

Avocado (Persea americana L.) production for export markets has increased in Mexico during the past 10 years. The production system, however, is affected by several sanitation factors, including diseases. During the spring of 2009, smooth, black, circular spots were noted on the surface of avocado fruit. A study was conducted during the winter of 2010 to ascertain the etiology and identify the fungus associated with black spot symptoms on avocado fruit in orchards of Nuevo Parangaricutiro County (19°25′00″ and 102°07′43″) in Michoacan, Mexico. Several fungal isolates were obtained on potato dextrose agar (PDA) from the margin of lesions on immature fruit. The internal transcribed spacer region (ITS) of the rDNA from representative isolates was sequenced with universal primers ITS5 and ITS4 (2). BLAST searches in GenBank showed 100% similarity of the nucleotide sequences with Neofusicoccum parvum (Pennycook & Samuels) Crous, Slippers & A.J.L. Phillips, GenBank Accession Nos. GU188001 to GU188007 and GU187985 to GU187987. A representative nucleotide sequence of this region was deposited in GenBank under the Accession No. JN203129. Strains of N. parvum produced aerial and compact mycelium on acidified PDA, the anamorph state of Botryosphaeria parva. Mycelium was initially white, turning gradually gray to black. Conidia were one celled, hyaline, ellipsoidal to fusiform, externally smooth, thin walled, nonseptate, with one or two septa with age, and an average length and width of 14.5 (9.5 to 19) × 5.8 (4.0 to 7.2) μm (n = 100). Pathogenicity tests were conducted with six avocado fruit cv. Hass. Fruit were inoculated at three evenly spaced locations on the fruit surface, either by wounding the tissue with a needle that had been dipped in a conidial mass from an 8-day-old monoconidial culture of N. parvum strain CIAD-021-11 or by placing 5 μl of 1 × 106 conidia ml–1 suspension on each inoculation site. Inoculated fruit were maintained in a moist chamber at 25°C for 2 weeks. Black lesions appeared on all wounded sites 2 days postinoculation (dpi) and on unwounded sites 4 dpi. The delay of symptom development was likely due to penetration through the lenticels, which took longer to initiate infection. No symptoms were observed in the control fruit. The pathogen was reisolated from the lesions of all inoculated fruit, thus fulfilling Koch's postulates. The results confirmed the pathogenic potential of this fungus and indicated its possible involvement in the etiology of black spot on avocado fruit. N. parvum is a cosmopolitan, plurivorous pathogen causing disease in several hosts of economic importance, such as grapes and kiwi, as well as causing stem-end rot of avocado fruit in New Zealand (1) and avocado twigs in Spain (3). To our knowledge, this is the first report of N. parvum causing black spots on avocado fruit in Mexico. References: (1) W. F. T. Hartill et al. N.Z.J. Crop Hortic. Sci. 30:249. 2002. (2) T. J. White et al. Page: 315 in: PCR Protocols: A Guide to Methods and Application. M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990. (3) T. Zea-Bonilla et al. Plant Dis. 91:1052, 2007.

scholarly journals Molecular beacons allow specific RT-LAMP detection of B.1.1.7 variant SARS-CoV-2

2021 ◽  
Author(s):  
Scott Sherrill-Mix ◽  
Gregory D. Van Duyne ◽  
Frederic D. Bushman
Keyword(s):  
Genetic Variants ◽  
Molecular Beacons ◽  
Spike Protein ◽  
Detection Methods ◽  
Gene Encoding ◽  
Detection And Identification ◽  
Rapid Spread ◽  
The World ◽  
Primer Sets ◽  
Current Detection

AbstractOver the course of the COVID-19 pandemic, several SARS-CoV-2 genetic variants of concern have appeared and spread throughout the world. Detection and identification of these variants is important to understanding and controlling their rapid spread. Current detection methods for a particularly concerning variant, B.1.1.7, require expensive qPCR machines and depend on the absence of a signal rather than a positive indicator of variant presence. Here we report an assay using a pair of molecular beacons paired with reverse transcription loop mediated amplification to allow isothermal amplification from saliva to specifically detect B.1.1.7 and other variants which contain a characteristic deletion in the gene encoding the viral spike protein. This assay is specific, affordable and allows multiplexing with other SARS-CoV-2 LAMP primer sets.

scholarly journals A One-Day Sensitive Method to Detect and Distinguish Between the Citrus Black Spot Pathogen Guignardia citricarpa and the Endophyte Guignardia mangiferae

Plant Disease ◽  
2006 ◽  
Vol 90 (1) ◽  
pp. 97-101 ◽  
Author(s):  
L. Meyer ◽  
G. M. Sanders ◽  
R. Jacobs ◽  
L. Korsten
Keyword(s):  
South African ◽  
Sequence Data ◽  
Black Spot ◽  
The United States ◽  
Guignardia Citricarpa ◽  
Competitive Edge ◽  
Citrus Black Spot ◽  
Species Specific ◽  
Time Required ◽  
Slow Growing

If South African citrus exporters wish to retain their competitive edge in the European market and access new markets such as the United States of America, it is of quarantine importance to distinguish between the citrus black spot pathogen, Guignardia citricarpa, and the harmless endophyte, G. mangiferae. The endophyte is not a sanitary or phytosanitary concern. This paper describes the design of species-specific primers that are able to detect and distinguish between these two Guignardia species. Application of the primer set CITRIC1 and CAMEL2 in conjunction with the ITS4 primer yielded polymerase chain reaction (PCR) amplicons of approximately 580 bp and 430 bp for G. citricarpa and G. mangiferae, respectively. Results obtained with these primers are in accordance with sequence data, and repeated tests verified accuracy and sensitivity. A BLAST search revealed no matches other than G. citricarpa and G. mangiferae, and no positive PCR results were obtained with Colletotrichum gloeosporioides, which is the most common contaminant in black spot lesions. We are, therefore, able to distinguish G. citricarpa and G. mangiferae unequivocally using a PCR-based method. This method was further improved to directly isolate DNA from fruit lesions by means of the DNeasy Plant Mini Kit (Qiagen). This eliminates the prior need for culturing the slow-growing organism, thereby shortening the time required to one day to test for and verify the presence or absence of the pathogenic G. citricarpa in export consignments.

A novel molecular diagnostic method for the gut content analysis of Philaenus DNA

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Isabel Rodrigues ◽  
Vítor Ramos ◽  
Jacinto Benhadi-Marín ◽  
Aránzazu Moreno ◽  
Alberto Fereres ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Control Strategies ◽  
Control Measure ◽  
Coi Gene ◽  
Molecular Diagnostic ◽  
Generalist Predators ◽  
Specificity And Sensitivity ◽  
Vector Borne ◽  
Polymerase Chain ◽  
Primer Sets

AbstractPhilaenus spumarius is a vector of Xylella fastidiosa, one of the most dangerous plants pathogenic bacteria worldwide. There is currently no control measure against this pathogen. Thus, the development of vector control strategies, like generalist predators, such as spiders, could be essential to limit the spread of this vector-borne pathogen. In this study, a polymerase chain reaction (PCR)-based approach was developed to principally detect DNA of P. spumarius in the spider’s gut. Accordingly, 20 primer pairs, targeting the mitochondrial cytochrome oxidase I (COI) and cytochrome b (cytB) genes, were tested for specificity, sensitivity, and efficiency in detecting P. spumarius DNA. Overall, two primer sets, targeting COI gene (COI_Ph71F/COI_Ph941R) and the cytB gene (cytB_Ph85F/cytB_Ph635R), showed the highest specificity and sensitivity, being able to amplify 870 pb and 550 bp fragments, respectively, with P. spumarius DNA concentrations 100-fold lower than that of the DNA of non-target species. Among these two primer sets, the cytB_Ph85F/cytB_Ph635R was able to detect P. spumarius in the spider Xysticus acerbus, reaching 50% detection success 82 h after feeding. The feasibility of this primer set to detect predation of P. spumarius by spiders was confirmed in the field, where 20% of the collected spiders presented positive amplifications.

scholarly journals Polymerase chain reaction primer sets for the detection of genetically diverse human sapoviruses

2020 ◽  
Vol 165 (10) ◽  
pp. 2335-2340
Author(s):  
Tomoichiro Oka ◽  
Seiji P. Yamamoto ◽  
Nobuhiro Iritani ◽  
Shigenori Sato ◽  
Chika Tatsumi ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Primer ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Primer Sets

scholarly journals Effect of temperature, leaf wetness, and rainfall on the production of Guignardia citricarpa ascospores and on back spot severity on sweet orange

2006 ◽  
Vol 31 (1) ◽  
pp. 29-34 ◽  
Author(s):  
Renato F. Reis ◽  
Lavern W. Timmer ◽  
Antonio de Goes
Keyword(s):  
Sweet Orange ◽  
Climatic Factors ◽  
Black Spot ◽  
Strong Relationship ◽  
Effect Of Temperature ◽  
Leaf Wetness ◽  
Total Rainfall ◽  
Spore Trap ◽  
Guignardia Citricarpa ◽  
Ascospore Production

The black spot of citrus (Citrus sp.) is caused by Guignardia citricarpa with ascospore production depending on temperature, leaf wetness, and rainfall. The number of ascospores produced was monitored using a spore trap and climatic factors were recorded using an automated meteorological station of 'Natal' and 'Valencia' sweet orange (Citrus sinensis) orchards in Mogi Guaçu in the state of São Paulo, Brazil, from November 2000 to March 2001. The fruits were bagged to prevent infection and the bags removed from different sets of fruit for one week during each of the 18 weeks of the season in both orchards. Ascospores were produced during the entire experimental period, from spring through summer, primarily after rain events. In both orchards, ascospore production reached a peak in January and February. Ascospore production was related to leaf wetness only in the Natal orange orchard but was not related to total rainfall or temperature in either orchard. Disease was most severe on fruit exposed the 7th, 8th, and 13th weeks after beginning the experiment in both cultivars as well as after the 16th week for 'Natal'. There was a strong relationship between disease severity and total rainfall for both orchards and a weak correlation between temperature and severity in the 'Natal' block only. There was no relationship between severity and leaf wetness or ascospore numbers.

scholarly journals Applications of Nanotechnology in Sensor-Based Detection of Foodborne Pathogens

Sensors ◽  
2020 ◽  
Vol 20 (7) ◽  
pp. 1966 ◽  
Author(s):  
Harsh Kumar ◽  
Kamil Kuča ◽  
Shashi Kant Bhatia ◽  
Kritika Saini ◽  
Ankur Kaushal ◽  
...  
Keyword(s):  
Foodborne Pathogens ◽  
Pathogen Detection ◽  
Foodborne Pathogen ◽  
Health Issues ◽  
Detection And Identification ◽  
Specificity And Sensitivity ◽  
Food Borne ◽  
Contaminated Food ◽  
Working Principle ◽  
Effective Use

The intake of microbial-contaminated food poses severe health issues due to the outbreaks of stern food-borne diseases. Therefore, there is a need for precise detection and identification of pathogenic microbes and toxins in food to prevent these concerns. Thus, understanding the concept of biosensing has enabled researchers to develop nanobiosensors with different nanomaterials and composites to improve the sensitivity as well as the specificity of pathogen detection. The application of nanomaterials has enabled researchers to use advanced technologies in biosensors for the transfer of signals to enhance their efficiency and sensitivity. Nanomaterials like carbon nanotubes, magnetic and gold, dendrimers, graphene nanomaterials and quantum dots are predominantly used for developing biosensors with improved specificity and sensitivity of detection due to their exclusive chemical, magnetic, mechanical, optical and physical properties. All nanoparticles and new composites used in biosensors need to be classified and categorized for their enhanced performance, quick detection, and unobtrusive and effective use in foodborne analysis. Hence, this review intends to summarize the different sensing methods used in foodborne pathogen detection, their design, working principle and advances in sensing systems.

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