Genetic Characterization ofDidymella bryoniaeIsolates Infecting Watermelon and Other Cucurbits in Florida and Georgia

Abstract

Gummy stem blight caused by Didymella bryoniae (anamorph Phoma cucurbitacearum) is a major fungal disease of watermelon (Citrullus lanatus) and other cucurbits. Thirty-five isolates of Didymella and Phoma spp. associated with symptoms of gummy stem blight on watermelon, Canary melon (Cucumis melo), muskmelon (C. melo), and winter squash (Cucurbita maxima) from Florida and Georgia were characterized based on morphology on agar media, pathogenicity on ‘Melody’ watermelon, the internal transcribed spacer (ITS) sequence of ribosomal DNA (rDNA), random amplified polymorphic DNA (RAPD) analysis, and polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) analysis. All of the isolates were pathogenic on watermelon but differed in virulence. RAPD and ITS sequence analysis indicated genetic variability among the isolates but PCR-RFLP analysis did not show any variability. ITS sequence phylogenetic analysis identified two isolates, DB-05 and DB-33, which had a greater identity to that of D. bryoniae isolates from China (98 to 100% sequence homology) than other isolates from Florida and Georgia (95 to 98%). These two isolates possessed a single nucleotide substitution of A to G at position 131 of the ITS1 region. The study characterized the genetic profile of a collection of D. bryoniae isolates from Florida and Georgia in relation to isolates from other U.S. states and countries.

Stagonosporopsis cucurbitacearum the causal agent of gummy stem blight of watermelon in India

“…Results confirmed that the fungal isolate was sharing 99% homology with the S. cucurbitacearum isolate (NCBI accession no.KC460840.1) available in the database submitted by Liu et al (2013). Isolated DNA was also used to amplify with SCAR primers (RG-I-F 5'-TGTCGTTGACATCATTCCAGC-3′ and RG-I-R 5'-ACCACTCTGCTTAGTATCTGC-3′) which are specific to S. cucurbitacearum (Babu et al 2015). The amplicon obtained with this primer was of 735 bp size.…”

Rapid and Sensitive Detection of Didymella bryoniae by Visual Loop-Mediated Isothermal Amplification Assay

“…Assignment to the RGI and RGII genotypes of D. bryoniae strains in this study was performed by an RG-specific PCR according to previously published protocols ( Keinath et al, 2001 ; Somai et al, 2002 ; Babu et al, 2015 ). In brief, PCRs were performed in a 25 μL reaction mixture containing 40 ng DNA, 2.0 μL 10x reaction buffer [50 mM Tris/HCl (pH 8.8), 1.5 mM MgCl 2 , 15 mM (NH 4 ) 2 SO 4 and 0.1% Triton X-100], 200 μM dNTPs, 2.5 U Taq DNA polymerase (5 U μL -1 ), and 0.25 μM each primer.…”

“…In brief, PCRs were performed in a 25 μL reaction mixture containing 40 ng DNA, 2.0 μL 10x reaction buffer [50 mM Tris/HCl (pH 8.8), 1.5 mM MgCl 2 , 15 mM (NH 4 ) 2 SO 4 and 0.1% Triton X-100], 200 μM dNTPs, 2.5 U Taq DNA polymerase (5 U μL -1 ), and 0.25 μM each primer. The thermal cycling program was: 94°C for 2 min, and 35 cycles of 94°C for 1 min, 62°C for 1 min, and 72°C for 2 min ( Babu et al, 2015 ). The DB17 primer set was used to amplify the conserved SCAR common to both genotypes of D. bryoniae ( Table 2 ) ( Ling et al, 2010 ).…”

Simultaneous determination of difenoconazole, trifloxystrobin and its metabolite trifloxystrobin acid residues in watermelon under field conditions by GC–MS/MS

“…Asia occupied more than 80% watermelon production all over the world. However, while anthracnose (Zhou & Everts, 2012), gummy stem blight (Babu et al, 2015) and powdery mildew (Keinath & DuBose, 2004) impacted on watermelon production for a long time, some pesticides should be applied to control the fungal threat. As two kinds of broad-spectrum fungicides, strobilurin fungicides and demethylation inhibiting fungicides were effective to above fungal threats (Gopinath, Radhakrishnan, & Jayaraj, 2006;Reuveni, 2000;Thomas, Langston, Sanders, & Stevensosn, 2012) on many fruits and vegetables, e.g.…”