Cloning of the Cytochrome b Gene From the Tomato Powdery Mildew Fungus Leveillula taurica Reveals High Levels of Allelic Variation and Heteroplasmy for the G143A Mutation

Abstract

Leveillula taurica is a major pathogen of tomato and several other crops that can cause substantial yield losses in favorable conditions for the fungus. Quinone outside inhibitor fungicides (Q o Is) are routinely used for the control of the pathogen in tomato fields across California, but their recurrent use could lead to the emergence of resistance against these compounds. Here, we partially cloned the cytochrome b gene from L. taurica ( Lt cytb ) and searched within populations of the fungus collected from tomato fields across California for mutations that confer resistance to Q o Is. A total of 21 single nucleotide polymorphisms (SNPs) were identified within a 704 bp fragment of the Lt cytb gene analyzed, of which five were non-synonymous substitutions. Among the most frequent SNPs encountered within field populations of the pathogen was the G143A substitution that confers high levels of resistance against Q o Is in several fungi. The other four amino acid substitutions were novel mutations, whose effect on Q o I resistance is currently unknown. Sequencing of the Lt cytb gene from individual single-cell conidia of the fungus further revealed that most SNPs, including the one leading to the G143A substitution, were present in a heteroplasmic state, indicating the co-existence of multiple mitotypes in single cells. Analysis of the field samples showed that the G143A substitution is predominantly heteroplasmic also within field populations of L. taurica in California, suggesting that Q o I resistance in this fungus is likely to be quantitative rather than qualitative.

First characterization of the complete mitochondrial genome of fungal plant-pathogen Monilinia laxa which represents the mobile intron rich structure

“…The core gene content of the mitogenomes is largely conserved, but their relative genes order is highly variable between and within the major fungal phyla 13 – 15 . Furthermore, many mutations in mitogenome might be related to different traits such as virulence and drug resistance 16 – 19 . Thus, mitogenome information is important to find out many answers in view of the evolution and adaptation of plant pathogenic fungi.…”

Fungicide resistance characterised across seven modes of action inBotrytis cinereaisolated from Australian vineyards

“…In our study, the presence of this intron in an isolate (Bc-111) that also contains mutant (G143A) copies of CytB, could suggest the presence of a "fitness balance", whereby intron and G143A CytB copies are balanced to maintain the lowest possible fitness cost. Mitochondrial heteroplasmy with respect to the G143A mutation has been confirmed in numerous fungal plant pathogens; B. cinerea (Hashimoto et al 2015;Ishii et al 2009), Venturia inaequalis (Michalecka et al 2011;Villani and Cox 2014), Corynespora cassiicola (Ishii et al 2007), Mycovellosiella nattrassii (Ishii et al 2007), Colletotrichum gloeosporioides (Ishii et al 2007), and various powdery mildews (Fraaije et al 2002;Lesemann et al 2006;Mosquera et al 2019;Vielba-Fernández et al 2018). The widespread occurrence of mitochondrial heteroplasmy in fungal plant pathogens could be a mechanism by which some QoI resistant isolates overcome potential fitness costs associated with G143A.…”

Fungicide Resistance in Powdery Mildew Fungi

“…In addition, after 50 generations, two isolates still showed a reduced sensitivity to QoI fungicides [ 98 ]. Finally, regarding the tomato powdery mildew L. taurica , 48% of the isolates analysed in the US in 2015 were resistant to azoxystrobin [ 99 ] ( Table 2 ).…”

“…hordei [ 88 ], B. graminis f. sp. tritici [ 83 , 101 , 102 , 103 , 104 , 105 ], E. betae [ 96 , 97 ], E. necator [ 46 , 91 , 92 , 93 , 94 , 95 ], L. taurica [ 99 ], P. leucotricha [ 98 ] and P. xanthii [ 82 , 83 , 106 ]. This mutation does not seem to affect the activity of the enzyme, since resistant isolates can persist for several years in the absence of the application of QoI fungicides [ 83 , 87 , 94 , 98 , 104 ].…”

“…This mutation does not seem to affect the activity of the enzyme, since resistant isolates can persist for several years in the absence of the application of QoI fungicides [ 83 , 87 , 94 , 98 , 104 ]. A phenomenon called heteroplasmy, which is the coexistence of sensitive (G143) and resistant (A143) cytb alleles in the same fungal cell, appears to govern QoI resistance in some powdery mildew species, such as L. taurica [ 99 ], P. leucotricha [ 98 ] and P. xanthii [ 106 ]. However, other authors have detected the presence of an intron of 2.438 bp directly after codon 143 in P. leucotricha isolates collected from Austria, Belgium, France, Germany, Italy, the Netherlands, Spain and the UK [ 107 ].…”