Molecular and biological properties of phytoplasmas

Abstract

Phytoplasmas, a large group of plant-pathogenic, phloem-inhabiting bacteria were discovered by Japanese scientists in 1967. They are transmitted from plant to plant by phloem-feeding insect hosts and cause a variety of symptoms and considerable damage in more than 1,000 plant species. In the first quarter century following the discovery of phytoplasmas, their tiny cell size and the difficulty in culturing them hampered their biological classification and restricted research to ecological studies such as detection by electron microscopy and identification of insect vectors. In the 1990s, however, tremendous advances in molecular biology and related technologies encouraged investigation of phytoplasmas at the molecular level. In the last quarter century, molecular biology has revealed important properties of phytoplasmas. This review summarizes the history and current status of phytoplasma research, focusing on their discovery, molecular classification, diagnosis of phytoplasma diseases, reductive evolution of their genomes, characteristic features of their plasmids, molecular mechanisms of insect transmission, virulence factors, and chemotherapy.

Keywords: genome; host specificity; mycoplasma-like organism; pathogenicity; phytoplasma; plant pathology.

Figure 1.

The earliest record of a phytoplasma disease is evident in peonies, attributed to Zhao Chang, a court painter of the Song Dynasty of China. The two flowers on the middle-right exhibit floral virescence symptoms. Printed with permission of the Museum of the Imperial Collections, Sannomaru-Shozokan.

Figure 2.

Various symptoms caused by yellows diseases. (A) Mulberry dwarf. (B) Paulownia witches’ broom. (C) Rice yellow dwarf. (D) Aster yellows. (E) Coconut lethal yellowing. (F) Poinsettia witches’ broom. (G) Hydrangea phyllody. (B, C, F, G) right side: infected plants; left side: healthy plants. The photographs were kindly provided by Drs. Akira Shirata (A), Norio Nishimura (B), and Akira Shinkai (C, D).

Figure 3.

The discovery of mycoplasma-like organisms (MLOs). (A) MLO cells observed via electron microscopy in an ultrathin section of a plant phloem cell. (B) Tetracycline-mediated recovery of mulberry from dwarf symptoms. Symptomatic mulberries infected with mulberry dwarf MLO (left panel) were treated with tetracycline (100 ppm) via foliage spraying (20 mL/pot) and soil drenching (200 mL/pot) at intervals of 2 or 3 days (12 applications). One month later, the plants had recovered (right panels). The photographs in (B) were originally published in ref. .

Figure 4.

Phylogenetic tree of life. MLOs are included in class Mollicutes (together with mycoplasmas), but they form a distinct group designated as the new genus ‘Ca. Phytoplasma’. More than 40 species have been described to date. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 5.

Establishment of the first phytoplasma mutant lines. (A) The mildly pathogenic mutant OY-M was isolated after 20 years of maintenance of the wild-type OY-W line in plant and insect hosts. (B) The non-insect-transmissible mutant line OY-NIM was isolated via 2-year grafting maintenance of OY-M (in the absence of the insect host). (C) Healthy plant and plants infected with OY-W and the mutant lines OY-M and OY-NIM. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 6.

Reductive evolution of phytoplasma metabolic pathways. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 7.

The chimeric “Rep” proteins of phytoplasma plasmids may be the missing link between bacterial plasmids and DNA viruses. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 8.

The life cycle of phytoplasmas; these are acquired by insects from plant phloem (via the stylets) and then enter the insect gut (acquisition feeding). Phytoplasmas must overcome three insect barriers to phytoplasma transmission (gut, hemocoel, and salivary gland barriers) if they are to be transmitted to plants. Phytoplasmas are transmitted from the insect to the phloem (of another plant) via inoculation feeding, and then multiply and establish a systemic infection, causing many unique symptoms. PP: phloem parenchymal cell, CC: companion cell, SE: sieve element. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 9.

The immunodominant membrane proteins of phytoplasmas and their localization. Most of the phytoplasma cell surface is covered with three membrane proteins termed immunodominant membrane proteins (IMP, AMP, and IDPA (immunodominant membrane protein A)), which are thought to be involved in interactions with both the insect and plant hosts. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 10.

The insect host ranges of OY phytoplasmas are determined by interactions between AMP and insect microfilaments. (A) Phytoplasma AMP co-localizes with microfilaments in the guts of insect vectors. Green: AMP detected with the aid of Alexa 488-conjugated IgG; Red: microfilaments stained with Alexa 546-coupled phalloidin. (B) Insect vector-specific formation of an AMP-microfilament complex. The photographs were kindly provided by Drs. Toshiki Shiomi (Ophiola flavopicta), Norio Nishimura (Hishimonoides sellatiformis), and Akira Shinkai (Hishimonus sellatus). This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 11.

Virus-vector-mediated expression of TENGU-peptide-induced “tengu-su” (witches’ broom) symptoms. Originally published in ref. .

Figure 12.

Phyllody symptom induction. A phyllogen protein expressed by phytoplasmas binds to and induces degradation of A- and E-class, MADS-domain-containing transcription factors (MTFs) via the 26S proteasome pathway, downregulating B-class MTF expression. Thus, phyllogens disturb the floral quartet model in an organ-specific manner. This figure was reproduced with modifications based on the original literature.¹²⁴⁾

Figure 13.

Phyllogen converts the flowers of all plants to leaves. (A) Virus-vector-mediated expression of a phyllogen peptide induced phyllody symptoms in eudicot plants. (B) Phyllogen induced degradation of YFP-fused MTFs not only of eudicots but also of monocots, gymnosperms, and ferns. This figure was reproduced with modifications based on the original literature.¹²⁴⁾ The sunflower photographs were originally published in ref. .

Figure 14.

The universal LAMP kit for phytoplasma detection. (A) The LAMP kit is 1000-fold more sensitive than conventional PCR. (B) The reagents are dry; the kit can thus be transported and stored at room temperature. (C) Phytoplasmas detected in the field from sawdust of coconut trees within 1 h without the need for electrical equipment. This figure was reproduced with modifications based on the original literature.¹²⁴⁾ The photographs in (C) were kindly provided by Dr. Toshiro Shigaki.

Figure 15.

Identification of antimicrobials that control phytoplasma diseases. (A) The screening protocol. Phytoplasma-infected micropropagated shoots are cultured on Murashige and Skoog medium containing antimicrobials; anti-phytoplasma compounds are thus selected. (B) The targets of the antimicrobials. Phytoplasmas exhibit few such targets because they have lost most metabolic pathways, including those for cell wall biosynthesis, during reductive evolution. (C) Antimicrobials effective against phytoplasmas. The photographs were taken 4 weeks after beginning antimicrobial treatment. Phytoplasmas were not detected from host plants after tetracycline or rifampicin treatment for 4 months. This figure was reproduced with modifications based on the original literature.¹²⁴⁾