scholarly journals Speciesspecific PCR primers for Guignardia citricarpa and Guignardia mangiferae

2006 ◽  
Vol 59 ◽  
pp. 141-145 ◽  
Author(s):  
K.R. Everett ◽  
J. Rees-George
Keyword(s):  
New Zealand ◽  
Venturia Inaequalis ◽  
Black Spot ◽  
Pathogenic Fungi ◽  
Pcr Primers ◽  
Colletotrichum Acutatum ◽  
Botryosphaeria Dothidea ◽  
Guignardia Citricarpa ◽  
Citrus Black Spot ◽  
Polymerase Chain

The plant pathogen Guignardia citricarpa causes citrus black spot and is not considered to be present in New Zealand Speciesspecific polymerase chain reaction (PCR) primers were designed to identify G citricarpa and G mangiferae a closely related saprotroph that is present in New Zealand These PCR primers were tested against a range of other saprotrophic and pathogenic fungi viz Botrytis cinerea Botryosphaeria dothidea B parva Cladosporium sp Colletotrichum acutatum C gloeosporioides Cryptosporiopsis sp Epicoccum sp Nigrospora sp Penicillium sp Pestalotia sp Phialophora sp Phlyctema sp Phoma sp Phomopsis sp Stemphylium sp and Venturia inaequalis The primers JRGGc were specific to G citricarpa and JRGGm to G mangiferae A 226 bp product was amplified from G mangiferae DNA using JRGGm primers and a 501 bp product was amplified from G citricarpa DNA using JRGGc primers These primers thus distinguished G citricarpa from G mangiferae and can be used to rapidly identify incursions by citrus black spot

scholarly journals High molecular diversity of the fungus Guignardia citricarpa and Guignardia mangiferae and new primers for the diagnosis of the citrus black spot

2009 ◽  
Vol 52 (5) ◽  
pp. 1063-1073 ◽  
Author(s):  
Danyelle Stringari ◽  
Chirlei Glienke ◽  
Daniel de Christo ◽  
Walter Maccheroni Jr. ◽  
João Lucio de Azevedo
Keyword(s):  
Rapd Markers ◽  
Geographical Origin ◽  
Black Spot ◽  
Polymerase Chain Reaction Analysis ◽  
Causal Agent ◽  
Specific Primers ◽  
Guignardia Citricarpa ◽  
Citrus Black Spot ◽  
Polymerase Chain ◽  
Sequence Characterized Amplified Regions

RAPD markers were used to investigate the distribution of genetic variability among a group of Guignardia citricarpa, G. mangiferae, and Phyllosticta spinarum isolates obtained from several hosts in Brazil, Argentina, Mexico, Costa Rica, Thailand, Japan, United States and South Africa. Pathogenic isolates G. citricarpa Kiely (anamorph form P. citricarpa McAlp Van Der Aa) are the etiological agent of the Citrus Black Spot (CBS), a disease that affects several citric plants and causes substantial injuries to the appearance of their fruits, thus preventing their export. Several previous studies have demonstrated the existence of an endophytic species with high morphological similarity to the causal agent of CBS that could remain latent in the same hosts. Consequently, the identification of the plants and fruits free from the causal agent of the disease is severely hampered. The RAPD analysis showed a clear discrimination among the pathogenic isolates of G. citricarpa and endophytic isolates (G. mangiferae and P. spinarum). In addition, a Principal Coordinate Analysis (PCO) based on a matrix of genetic similarity estimated by the RAPD markers showed four clusters, irrespective of their host or geographical origin. An Analysis of Molecular Variance (AMOVA) indicated that 62.8% of the genetic variation was found between the populations (G. citricarpa, G. mangiferae, P. spinarum and Phyllosticta sp.). Substantial variation was found in the populations (37.2%). Exclusive RAPD markers of isolates of G. citricarpa were cloned, sequenced and used to obtain SCARS (Sequence Characterized Amplified Regions), which allowed the development of new specific primers for the identification of G. citricarpa PCR (Polymerase Chain Reaction) analysis using a pair of primers specific to pathogenic isolates corroborating the groupings obtained by the RAPD markers, underscoring its efficiency in the identification of the causal agent of CBS.

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.

scholarly journals Validation of Quantitative and Digital Polymerase Chain Reaction Assays Targeting the Mating Types of Phyllosticta citricarpa, the Causal Agent of Citrus Black Spot

2021 ◽  
Author(s):  
Hector Urbina ◽  
Taylor Smith ◽  
Callie Jones ◽  
Xiaoan Sun ◽  
John McVay ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Mating Type ◽  
Citrus Fruit ◽  
Black Spot ◽  
Dispersal Ability ◽  
Mating Types ◽  
Chain Reaction ◽  
Citrus Black Spot ◽  
Limited Dispersal ◽  
Polymerase Chain

Citrus black spot (CBS) is a disease caused by the ascomycetous fungus Phyllosticta (formerly Guignardia) citricarpa (Botryosphaeriales, Pezizomycotina) currently present in citrus groves in five counties in southwest Florida. Within Florida, P. citricarpa shows limited reproduction via asexual sporulation due to the presence of only one (MAT1-2-1) of the two required mating types for sexual reproduction. Here we present two novel polymerase chain reaction (PCR) assays standardized in quantitative (qPCR) and digital (dPCR) platforms to distinguish both mating types (MAT1-1-1 and MAT1-2-1) of P. citricarpa, to monitor for the potential introduction of the MAT1-1-1 mating type into Florida and a novel protocol for DNA extraction from asymptomatic leaves. During citrus harvesting season 2018−2019, fruit lesions, as well as asymptomatic leaves adjacent to symptomatic fruit and asymptomatic trees in CBS-infected groves were surveyed for P. citricarpa presence and mating types. Results support the presence of only MAT1‑2-1 mating type in Florida, after surveying more than 1,145 citrus fruit lesions. We also confirmed the limited dispersal ability of the asexual state of P. citricarpa in Florida in ten groves using the enhanced capabilities of the dPCR platform in the detection of P. citricarpa directly from asymptomatic leaves with low pathogen inoculum.

scholarly journals Pathogenic and Nonpathogenic Lifestyles in Colletotrichum acutatum from Strawberry and Other Plants

2001 ◽  
Vol 91 (10) ◽  
pp. 986-992 ◽  
Author(s):  
Stanley Freeman ◽  
Sigal Horowitz ◽  
Amir Sharon
Keyword(s):  
Plant Species ◽  
Fungal Growth ◽  
Black Spot ◽  
Crown Rot ◽  
Colletotrichum Acutatum ◽  
Disease Symptoms ◽  
Root Necrosis ◽  
Leaf Tissues ◽  
Polymerase Chain ◽  
Number Of Plant Species

Anthracnose is one of the major fungal diseases of strawberry occurring worldwide. In Israel, the disease is caused primarily by the species Colletotrichum acutatum. The pathogen causes black spot on fruit, root necrosis, and crown rot resulting in mortality of transplants in the field. The host range and specificity of C. acutatum from strawberry was examined on pepper, eggplant, tomato, bean, and strawberry under greenhouse conditions. The fungus was recovered from all plant species over a 3-month period but caused disease symptoms only on strawberry. Epiphytic and endophytic (colonization) fungal growth in the different plant species was confirmed by reisolation from leaf tissues and by polymerase chain reaction (PCR)-specific primer amplification. C. acutatum was also isolated from healthy looking, asymptomatic plants of the weed genera Vicia and Conyza. Isolates that were recovered from the weeds caused disease symptoms on strawberry and were positively identified as C. acutatum by PCR. The habitation of a large number of plant species, including weeds, by C. acutatum suggests that, although it causes disease only on strawberry and anemone in Israel, this fungus can persist on many other plant species. Therefore, plants that are not considered hosts of C. acutatum may serve as a potential inoculum source for strawberry infection and permit survival of the pathogen between seasons.

scholarly journals Application of Azoxystrobin for Control of Benomyl-Resistant Guignardia citricarpa on ‘Valencia’ Oranges in South Africa

Plant Disease ◽  
2003 ◽  
Vol 87 (7) ◽  
pp. 784-788 ◽  
Author(s):  
G. C. Schutte ◽  
R. I. Mansfield ◽  
H. Smith ◽  
K. V. Beeton
Keyword(s):  
South Africa ◽  
Disease Control ◽  
Untreated Control ◽  
Black Spot ◽  
Mineral Oil ◽  
Guignardia Citricarpa ◽  
Citrus Black Spot ◽  
Oil In Water ◽  
Small Plot

Azoxystrobin was evaluated in replicated small-plot trials from 1995 to 1999 for control of citrus black spot (CBS) on ‘Valencia’ oranges caused by Guignardia citricarpa. Applications of different rates of tank mixes of azoxystrobin and mancozeb during the susceptible period from October to January were compared with an untreated control as well as the standard four applications of mancozeb with or without mineral oil (1.20 g a.i./liter + 0.5% [vol/vol]/liter and 1.60 g a.i./liter of water, respectively). Two applications of azoxystrobin in tank mixtures with mancozeb and mineral oil (0.5% [vol/vol]/liter) in mid-November and mid-January at rates of 0.10, 0.15, and 0.20 g a.i./liter controlled CBS by more than 98 to 99%, 99 to 100% and 95 to 98%, respectively. Concomitantly, where mineral oil was not added to the fungicide mixture, azoxystrobin and mancozeb resulted only in 73 to 95%, 74 to 93% and 92.2 to 92.3% CBS control, respectively. Tank mixtures of benomyl, mancozeb, and mineral oil reduced CBS by only 29%, which could be attributed to the presence of benomyl-resistant pathogen isolates in the experimental orchard. Azoxystrobin applied at rates of 0.05, 0.075, and 0.10 g a.i./liter in tank mixtures with mancozeb (1.2 g a.i./liter) and mineral oil (0.5% [vol/vol]/liter of water) or Agral 90 (0.5% [vol/vol]/liter of water) were equally effective, reducing CBS by more than 99%. When mineral oil was compared to different adjuvants in tank mixtures with azoxystrobin and mancozeb, only mineral oil resulted in 100% clean exportable fruit. There was no difference between Sunspray 6E and Bac oil when mixed with azoxystrobin and mancozeb on the degree of disease control. Furthermore, the concentration of mineral oil in water can be lowered from 0.5% (vol/vol)/liter of water to 0.3% (vol/vol)/liter of water without a loss in efficacy against CBS. It is therefore, recommended that azoxystrobin (0.075 g a.i./liter) must be applied in tank mixtures with mancozeb (1.2 g a.i./liter) and mineral oil, which can be applied at either 0.5% (vol/vol)/liter of water or 0.3% (vol/vol)/liter of water.

scholarly journals Growth supression of plant pathogenic fungi using bryophite extracts

2019 ◽  
Vol 35 (4) ◽  
Author(s):  
Nedeljko Latinovic ◽  
Marko S. Sabovljevic ◽  
Milorad Vujicic ◽  
Jelena Latinovic ◽  
Aneta D. Sabovljevic
Keyword(s):  
Adverse Effects ◽  
Food Production ◽  
Environmentally Friendly ◽  
Pathogenic Fungi ◽  
Colletotrichum Acutatum ◽  
Plant Pathogenic Fungi ◽  
Monilinia Laxa ◽  
Botryosphaeria Dothidea ◽  
Phomopsis Viticola ◽  
Long Time

Chemicals are often used in attempts to control diseases caused by plant pathogenic fungi during food production. However, chemicals can have adverse effects not just on food, but they also remain active for a long time within ecosystems, and thus are not environmentally friendly. Therefore, development of bio-treatment and avoiding use of chemicals are urgently needed. With the aim of studying and developing new environmentally-friendly treatments, we tested extracts from selected bryophyte species (Porella platyphylla, Cinclidotus fontinaloides and Anomodon viticulosus) on five plant pathogenic fungi under controlled conditions. The fungi (Botryosphaeria dothidea, Phomopsis viticola, Calosphaeria sp., Colletotrichum acutatum and Monilinia laxa) were selected based on common diseases they cause on fruits and grapevine. They were isolated in cultures and treated with bryophyte extracts. Bryophyte extracts were shown to be effective in suppression of certain plant pathogenic fungi growth and to have a huge potential in development of novel biotechnological treatments and biofungicides. The best results were achieved in inhibition of B. dothidea, P. viticola and Calosphaeria sp.

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