Assessing the Functionality of RNA Interference (RNAi) in the Phloem-feeding Maize pest Dalbulus maidis

Dalbulus maidis [(DeLong & Wolcott), corn leafhopper], a phloem-feeding insect, is the most efficient vector of maize stunting pathogens (Spiroplasma kunkelii, Maize bushy stunt phytoplasma, and Maize rayado fino virus) in the Americas. Studies involving gene editing in insects are rapidly providing information that can potentially be used for insect vector and plant disease control. RNA interference (RNAi), a sequence-specific gene silencing method, is one of the most widely used molecular tools in functional genomics studies. RNAi uses exogenous double-stranded RNA (dsRNA) or small interfering RNA (siRNA) to prevent the production of proteins by inhibiting the expression of their corresponding messenger RNA (mRNA). In this study, we measured the efficacy of gene silencing, and its effects on D. maidis mortality as proof of concept that RNAi is a viable tool for use in genetic pest control of phloem-feeding insects. Oral delivery of dsRNA using an artificial diet was used to silence two key insect genes, vacuolar ATP synthase subunit B, and subunit D (V-ATPase B and V-ATPase D). Our results showed reduced gene expression of V-ATPase B and V-ATPase D after ingestion of dsRNA, and significantly higher mortality, and wing deformation, associated with reduced gene expression, compared to control insects that were not orally fed dsRNA. These results reveal RNAi as a viable tool for use in genetic pest control of phloem-feeding insects, and a way for further functional genomic studies, such as identification of potential target genes for either population suppression or population replacement of this vector of maize diseases.

Corn Stunt Disease: An Ideal Insect–Microbial–Plant Pathosystem for Comprehensive Studies of Vector-Borne Plant Diseases of Corn

Over 700 plant diseases identified as vector-borne negatively impact plant health and food security globally. The pest control of vector-borne diseases in agricultural settings is in urgent need of more effective tools. Ongoing research in genetics, molecular biology, physiology, and vector behavior has begun to unravel new insights into the transmission of phytopathogens by their insect vectors. However, the intricate mechanisms involved in phytopathogen transmission for certain pathosystems warrant further investigation. In this review, we propose the corn stunt pathosystem (Zea mays–Spiroplasma kunkelii–Dalbulus maidis) as an ideal model for dissecting the molecular determinants and mechanisms underpinning the persistent transmission of a mollicute by its specialist insect vector to an economically important monocotyledonous crop. Corn stunt is the most important disease of corn in the Americas and the Caribbean, where it causes the severe stunting of corn plants and can result in up to 100% yield loss. A comprehensive study of the corn stunt disease system will pave the way for the discovery of novel molecular targets for genetic pest control targeting either the insect vector or the phytopathogen.

Characterization of Components of Resistance to Corn Stunt Disease

Considering the occasional but increasing presence of corn stunt disease in the subtropical region of Argentina, the objective of this research was to devise an effective strategy to screen disease-resistant genotypes in the absence of high and constant natural pressures. To do so, the presence of antixenosis and antibiosis as components of resistance to vector Dalbulus maidis (DeLong 1923) as well as resistance to the pathogen Spiroplasma kunkelii (Whitcomb et al. 1986) under artificial inoculation conditions were investigated in four widely-distributed maize hybrids in Argentina. The hybrids shown differences in the levels of resistance and target organisms (either the insect vector or the pathogen). Antixenosis and antibiosis to D. maidis were observed in DK72-10. Resistance of DK79-10 to S. kunkelii was evidenced by a delayed onset of symptoms, and DKB390 was antixenotic to D. maidis and highly resistant to S. kunkelii. A good association was found between symptom severity and yield, but not between symptom severity and accumulation of pathogen S. kunkelii. In conclusion, the proposed methodology was efficacious and can aid the screening of resistant genotypes in breeding programs to reduce the impact of corn stunt disease, ensuring that hybrids with good resistance level will be planted by farmers whenever disease occurs.

Respuesta agronómica del cultivo de maíz (Zea mays): aplicación de insecticidas para el control del vector de la cinta roja (Spiroplasma kunkellii)

El cultivo del maíz (Zea mays), es uno de los más investigados en el mundo y cada año el potencial productivo de los cultivares ha aumentado. Sin embargo, en varios países el rendimiento medio está todavía muy por debajo de lo que se puede producir. La cinta roja es una enfermedad causada por patógenos específicamente por el micoplasma, espiroplasma y virus los mismos que los transmite un insecto vector conocido como la cigarrita (Dalbulus maidis) del orden homóptero, los agricultores han venido realizando sus controles por años con insecticidas convencionales, tales como cipermetrina, clorpirifos, malathion. No existe ningún tipo de control a la hora de aplicar en los cultivos de maíz. El objetivo ha sido evaluar la aplicación de insecticidas químicos en el control del vector de cinta roja (Spiroplasma kunkelii), valorando las características agronómicas del cultivo de maíz, en base a los tratamientos de estudio y determinando el insecticida más eficiente en el control del vector de la enfermedad. La investigación fue de tipo experimental con enfoque investigativo de campo, con diseño experimental, con el propósito de validar el mejor tratamiento para el control del vector de la cinta roja. Los resultados obtenidos demuestran que el tratamiento aplicado, con tres frecuencias de aplicación reduce el ataque del insecto trasmisor del virus causante de la cinta roja, obteniendo el mayor rendimiento por hectáreas de 7834kg/ha. Se concluyó que en la altura de planta, inserción de mazorca, longitud de mazorca no se encontró diferencia significativa entre los tratamientos.

Complete Genome Sequence of Spiroplasma kunkelii Strain CR2-3x, Causal Agent of Corn Stunt Disease in Zea mays L.

Spiroplasma kunkelii causes corn stunt disease of Zea mays L. in the Americas. Here, we report the nucleotide sequence of the 1,463,926-bp circular chromosome and four plasmids of strain CR2-3x. This information will facilitate studies of Spiroplasma pathogenicity and evolutionary adaptations to transkingdom parasitism in plants and insect vectors.

An Analysis of the Genomic Variability of the Phytopathogenic Mollicute Spiroplasma kunkelii

Corn stunt disease has become a factor limiting maize production in some areas of the Americas in recent years. Although resistant maize genotypes have been developed in the past, this resistance has been unstable over time or in some geographical locations. To better understand disease components that could affect the stability of host resistance, we assessed the genome variability of the etiologic agent, Spiroplasma kunkelii. Isolates were obtained from a number of areas, and characterized molecularly by amplification of several regions of the spiroplasma chromosome and sequencing of specific gene fragments. The degree of polymorphism between isolates of different geographic origins was low, and the level of genomic variability was similar within isolates of different countries. Polymorphism among isolates was found in viral insertions and in the sequence of Skarp, a gene that encodes a membrane protein implicated in attachment to insect cells. The results suggest that the genome composition of this species is highly conserved among isolates. Hence, it is unlikely that the instability of maize resistance is due to generation of new pathotypes of S. kunkelii. Instead, other components of this complex pathosystem could account for the breakdown of resistance.

Spiroplasma kunkelii. [Distribution map].

A new distribution map is provided for Spiroplasma kunkelii Whitcomb, Chen et al. Bacteria. Hosts: maize (Zea mays), sweetcorn (Zea mays subsp. mays), teosinte (Zea mexicana) and perennial teosinte (Zea perennis). Information is given on the geographical distribution in North America (Mexico, USA, California, Florida, Louisiana, Michigan, Mississippi, Ohio, Texas), Central America and Caribbean (Belize, El Salvador, Guatemala, Honduras, Jamaica, Nicaragua, Panama), South America (Argentina, Bolivia, Brazil, Mato Grosso do Sul, Minas Gerais, Colombia, Paraguay, Peru, Venezuela).