Reviews in Agricultural Science
Online ISSN : 2187-090X
Intumescence: A Serious Physiological Disorder in Plants
Natassia Clara Sita, Iriawati, Yoshikazu Kiriiwa, Katsumi Suzuki
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Keywords: calcium, cell hypertrophy, cultivar, cuticle layer, high humidity, ultraviolet
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2024 Volume 12 Pages 182-212

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Abstract

Intumescence injury is a physiological disorder that occurs in various plants. Initially, this injury was observed in the field, and it has become a problem in greenhouse and plant factory cultivations in recent years. Many terms refer to this disorder, with intumescence and oedema as the main two that are still frequently used. Intumescence is characterized as hypertrophied and/or hyperplasia of epidermal cells, while oedema refers to a lesion caused by excess water accumulated in the plant tissues. Morphological characteristics of intumescence differ among plants. Environmental factors, such as light and humidity, mainly caused this problem, and recent reports associate intumescence with a calcium-related injury. In tomato plants, low ultraviolet combined with high humidity affected the cuticle layer, and the high humidity lowered the transpiration rate, decreasing the calcium uptake to the leaves. This combination leads to cell hypertrophy and intumescence injury. The severity of this disorder is different among cultivars within one species, and this phenomenon is highly genetically dependent. Some approaches are proposed to limit intumescence injury, which applies to the field or controlled environments. This review summarized the information regarding intumescence injury, as well as the future approaches regarding this study.

1. Introduction

Horticulture is a branch of agriculture that focuses on the cultivation of vegetables, fruits, and ornamental plants. Many problems arise with horticultural crop production in the field, in the greenhouse, or when using plant factories with artificial light (PFAL). Physiological disorder caused by abiotic stress is one of the significant problems in horticultural cultivation. In this review, we focused on one of the disorders called intumescence injury.

Intumescence injury is commonly found in horticultural crops. Some refer to this disorder as edema or oedema. It is characterized by hypertrophied and/or hyperplasia of epidermal cells. This phenomenon affects the growth and yield of plants, and it usually occurs in several parts of the plant, including leaves, stems, and flowers.

This physiological disorder occurs in the field and controlled environments, such as greenhouse and growth chambers. In the field and greenhouse, this problem arises under environmental changes, such as when humidity arises and low ultraviolet (UV) light during the rainy season. In Japan, intumescence injury increased when UV-cut film use increased and tomato cultivars were changed. UV-cut films are initially used in greenhouses for pest control [1, 2, 3]. In America, during the winter, low-light conditions outside cause the greenhouse to be supplied with high-pressure sodium (HPS) light, which lacks UV wavelength energy [4]. However, in these conditions, the UV irradiation is inhibited and leads to intumescence. In PFAL and closed nursery growing systems, recent technology used light-emitting diode (LED) light, which also lacks UV light as a substitute for fluorescent lights [5]. This situation also leads to an increase in the incidence of intumescence injury.

Many research studies have investigated intumescence injury for years; however, problems relating to this injury are continuing. Therefore, it is essential to understand the factors that cause this problem and how to countermeasure or prevent it. This review aims to provide comprehensive information about intumescence injury in plants, especially horticultural crops, including the factors, mechanisms, and countermeasures to minimize its occurrence in the field and controlled environments.

2. History and definition

2.1 Intumescence as one of the physiological disorders in crop

Studies about this physiological disorder have been performed since the late 1800s. This physiological disorder was first mentioned as intumescence by Sorauer in 1886. However, there are some other terms for this disorder: warts [6], oedema or edema [7], excrescences [8], neoplasms [9, 10], enations [11], galls, leaf lesions [12], and tumors [13, 14]. Among them, the most common terms are intumescence and oedema. We summarize the history and definition of this physiological disorder in Table 1.

The term intumescentia, a Latin word for intumescences, was first mentioned by Sorauer in 1886. It referred to tiny, wart-like eruptions on the leaves' surface, typically yellow and usually exhibiting atypical cell stretching [15, 16]. On the other hand, Atkinson [7] first mentioned the term oedema, who referred to this phenomenon as swelling in specific plant regions due to excessive water stretching the cell walls, making them very thin and the cells very large. The excess water may be so enormous that it causes the plant’s cell walls to disintegrate, negatively impacting surrounding areas. Oedema was also mentioned in Eisa and Dobrenz [17], and it was defined as irregular, protuberant growths resembling blisters on plants. Oedema was observed in Manihot spp. (Table 1) as elevations on the upper surface of the leaves that line up with depressions on the lower side and primarily develop in the spaces between the veins [17, 18].

Recent reports revealed that the term of intumescence was frequently used to indicate this phenomenon rather than oedema. Lang and Tibbitts proposed ‘intumescence’ as the suitable term to define lesions in tomatoes, as ‘oedema’ was defined as the watery swelling of plant organs or parts because of water accumulation in plant tissue [19]. Craver et al. [20] later supported this, and Williams et al. [21] stated that intumescence is a proper term to describe the lesions developed in tomato and ornamental sweet potatoes, and cuphea leaves, as intumescence is marked by the elongation of the epidermal cell as well as the stomatal guard cell. Williams et al. [21] propose that oedema is the appropriate term to describe the lesions in Pelargonium sp., where the elongation happens on the spongy parenchymal cells as a result of water congestion and causes the epidermal cells to stretch and tear. Other research has linked plant water relations to the occurrence of lesions in geranium. While the term ‘oedema’ is occasionally used in research articles, ‘intumescence’ has been the common term in several recent studies [3, 22, 23, 24, 25, 26, 27, 28].

Table 1: List of the plants in which intumescence injury (and/or all technical terms) occurre.

Order Family Species Common name Injury name First discovered (Year) References
Alismatales Araceae Philodendron pinnatifidum Comb-leaf philodendron Intumescence 1915 [14, 101]
Philodendron hastatum Schott Silver sword philodendron Intumescence 1957 [32, 102]
Asparagales Asparagaceae Yucca sp. Yucca Intumescence 1896 [36, 62]
Asterales Asteraceae Hieracium venosum Rattlesnake-weed Intumescence 1931 [37, 41]
Brassicales Brassicaceae Brassica oleracea var. botrytis Cauliflower Intumescence 1905 [16]
Brassica oleracea var. capitata Cabbage Intumescence 1918 [80]
Caryophyllales Caryophyllaceae Dianthus sp. Carnation Oedema 1892 [56, 82]
Dipsacales Adoxaceae Sambucus nigra Elder Intumescence 1915 [14, 101]
Ericales Balsaminaceae Impatiens capensis Nutt. Jewelweed Intumescence 1899 [16, 35]
Fabales Fabaceae Phaseolus vulgaris Common bean Intumescence 1896 [62]
Senna multiglandulosa (syn. Cassia tomentosa) Downy senna Intumescence 1899 [35, 41]
Cassia floribunda Golden Snowy Cassia Intumescence 1901 [36]
Vicia faba Broad bean Intumescence 1906 [61]
Pisum sativum Pea Intumescence, Tumor, Neoplastic growth 1906, 1969 [9, 10], [61]
Vigna unguiculata L. Walp. Cowpea Edema 2008 [55]
Gentianales Rubiaceae Mitchella repens Patridgeberry Intumescence 1931 [37, 41]
Geraniales Geraniaceae Pelargonium peltatum Ivy geranium Intumescence 1905 [36]
Pelargonium hortorum Ait. Garden geranium Oedema 1969 [63]
Pelargonium sp. Geranium Oedema 1976 [32, 64]
Pelargonium x ‘Caliente Coral’ Interspecific hybrid geranium Oedema 2014 [48]
Lamiales Acanthaceae Ruellia formosa Andr. Ruellias Intumescence 1905 [14, 41, 105]
Aphelandra porteana Morel Aphelandra Intumescence 1905 [14, 41, 105]
Verbenaceae Gmelina philippensis (syn. Gmelina hystrix Schult) Hedgehog bush Intumescence 1905 [36]
Malpighiales Euphorbiaceae Acalypha marginata Spreng. Copperleaf Intumescence 1905 [36]
Manihot spp. Cassava Oedema 1912 [17, 18]
Ricinus communis Castor oil bean Tumor 1917 [13, 14]
Salicaceae Populus tremula European aspen Intumescence 1903 [16, 69]
Populus tremuloides Quaking aspen Intumescence 1930 [40, 41]
Populus grandidentata Bigtooth aspen Intumescence 1930 [40, 41]
Populus spp. Poplar Intumescence 1933 [14, 41]
Malvales Malvaceae Hibiscus vitifolius L. Graped-leaved mallow Intumescence 1899 [57, 58, 59, 60]
Abelmoschus esculentus (syn. Hibiscus esculentus Linn.) Okra Intumescence 1905 [36]
Abelmoschus manihot (syn. Hibiscus manihot Linn.) Sunset hibiscus Intumescence 1905 [36]
Abelmoschus moschatus (syn. Hibiscus abelmoschus Linn.) Musk mallow Intumescence 1905 [36]
Pavonia arabica Hochst Arabian swamp mallow Intumescence 1905 [36]
Theobroma cacao Linn. Cacao tree Intumescence 1905 [36]
Gossypium thurberi (syn. Thurberia sp.) Dessert cotton Intumescence 1931 [37, 41]
Myrtales Lythraceae Cuphea spp. Cuphea Intumescence 1988 [68]
Cuphea llavea Bat-faced cuphea Intumescence 2014 [48]
Melastomataceae Kendrickia walkeri Hook   Intumescence 1905 [36]
Myrtaceae Eucalyptus coccifera L'Herit Tasmanian snow gum Intumescence 1886 [15] [37, 41]
Eucalyptus globulus Bluegum eucalyptus Intumescence 1899 [35]
Eucalyptus rostrata River red gum Intumescence 1899 [35]
Eucalyptus diversicolor F. Muell Karri Intumescence 1905 [36]
Eucalyptus melliodora A. Cunn Yellow box Intumescence 1905 [36]
Eucalyptus botryoides Sm Bangalay Intumescence 1905 [36]
Eucalyptus resinifera Sm Red mahogany Intumescence 1905 [36]
Eucalyptus saligna Sm Sydney blue gum Intumescence 1905 [36]
Eucalyptus cornuta Yate Intumescence 1931 [37, 41]
Eucalyptus spp. Eucalyptus Intumescence 1980 [30, 32]
Eucalyptus regnans F. Muell Mountain ash Intumescence 1992 [31, 32]
Eucalyptus nitens Shining gum Intumescence 2006 [32]
Piperales Piperaceae Piper ornatum N.E.Br. Celebes pepper Intumescence 1905 [36]
Peperomia sp. Radiator plants Edema 1969 [32, 87]
Rosales Moraceae Ficus elastica Roxb. Rubber fig Intumescence 1899 [16, 35]
Artocarpus incisa Breadfruit Intumescence 1915 [14, 101]
Rosaceae Rosa sp. Rose Intumescence 1896 [62]
Spiraea concinna Intumescence 1915 [14, 101]
Malus sylvestris Crab apple Intumescence 1926 [38]
Malus domestica cv. 'White Transparent' (syn. Pyrus malus) Transparent Apple Intumescence 1926 [38]
Sapindales Sapindaceae Aesculus hippocastanum European horse-chestnut Intumescence 1915 [14, 101]
Saxifragales Grossulariaceae Ribes aureum Golden Currant Intumescence 1896 [62]
Solanales Convolvulaceae Stictocardia laxiflora var. woodii (syn. Ipomoea woodii) Intumescence 1901 [60]
Ipomoea batatas Sweet potato (ornamental, edible) Intumescence 1904 [20], [79]
Ipomoea aquatica Forsk Water spinach Intumescence 2019 [72]
Solanaceae Solanum tuberosum Potato Warts, Intumescence, Lesion, Enation 1878, 1907, 1971, 1974 [6], [11], [12], [56]
Solanum lycopersicum (syn. Lycopersicon esculentum) Tomato Oedema, Enation, Tumor, Neoplasm, Intumescence 1893, 1967, 1977, 1983, 2014 [7], [19], [32, 43], [32, 44], [42], [48]
Solanum aculeatissimum Dutch eggplant Intumescence 1905 [36]
Solanum phyracanthum Porcupine tomato Intumescence 1905 [36]
Solanum chilense (syn. Lycopersicon chilense) Wild tomato Tumor 1967 [42]
Solanum melongena L. Eggplant Oedema 1971 [17]
Solanum habrochaites (syn. Lycopersicon hirsutum) Hairy wild tomato Neoplasm, Tumor 1977, 1988 [32, 44], [47]
Capsicum annuum Pepper Edema 2008 [55]
Vitales Vitaceae Vitis rotundifolia Michx Muscadine Intumescence 1879 [36, 104]
Parthenocissus quinquefolia Virginia creeper Intumescence 1879 [36, 104]
Vitis vinifera Common grape vine Intumescence 1899 [35, 36]
Ampelopsis glandulosa var. heterophylla Porcelain berry Intumescence 1905 [36]
Parthenocissus tricuspidata Boston Ivy Intumescence 1905 [36]
Pinales Araucariaceae Araucaria bidwillii Bunya pine Excrescences 1920 [8]
Pinaceae Pinus ponderosa Ponderosa pine Excrescences 1915 [8]
Pinus coulteri Coulter pine Excrescences 1920 [8]
Pinus rigida Pitch pine Excrescences 1920 [8]
Pinus resinosa Red pine Excrescences 1920 [8]
Pinus banksiana Jack pine Excrescences 1920 [8]
Pinus virginiana Virginia pine Excrescences 1920 [8]
Pinus sylvestris Scotch pine Excrescences 1920 [8]
Pinus caribaea Caribbean pine Excrescences 1920 [8]
Pinus strobus Eastern white pine Excrescences 1920 [8]
Pinus monticola Western white pine Excrescences 1920 [8]
Pinus excelsa Himalayan pine Excrescences 1920 [8]
Picea canadensis White spruce Excrescences 1920 [8]
Picea rubens Red spruce Excrescences 1920 [8]
Picea mariana Black spruce Excrescences 1920 [8]
Picea pungens Blue spruce Excrescences 1920 [8]
Abies balsamea Balsam fir Excrescences 1920 [8]
Tsuga canadensis Eastern hemlock Excrescences 1920 [8]
Larix laricina Tamarack Excrescences 1920 [8]
Taxaceae Taxus cuspidata Japanese yew Excrescences 1920 [8]
Taxus brevifolia Pacific yew Excrescences 1920 [8]

2.2 History of intumescence discovery

From the 19th century to the early 20th century, it was reported that this physiological disorder occurred in various plants. In addition, the relationship between high humidity, temperature, and amount of light was studied. This phenomenon was first reported by Masters in 1878 in Solanum tuberosum as warts that occurred in a high-humidity environment [6]. During the late 1800s to early 1900s, this disorder was also reported in about 80 species (Table 1).

In the mid-20th century, artificial weather machines that were used for growing plants revealed the relationship between various environmental factors and light quality to intumescence occurrence. Morphological studies and the effect of culture solutions were also conducted to clarify the developmental process in more detail. The terms oedema and intumescence were still used interchangeably, with some of the main objects of the research being from the Solanaceae family, Pelargonium sp., and Cuphea sp. (Table 1).

In the 21st century, with the development of LEDs, a detailed relationship with light quality has been clarified. Furthermore, comprehensive gene expression and genetic analysis using recombinant inbred line (RIL) have been conducted, and the developmental mechanisms and relationships among related genes have been clarified. In addition, the relationship between the cuticle layer and intumescence development and the effect of calcium (Ca) on intumescence has also been clarified, as well as other physiological studies associated with this phenomenon.

Then, we will explain the plant species reported, morphological features, environmental factors, mechanisms of intumescence development, genetic factors, and countermeasures against intumescence injury.

2.3 Occurrence of this physiological disorder in plants

In this subchapter, we would like to summarize the plant families and species in which intumescences (and all technical terms that refer to this physiological disorder), such as oedema, have been reported. They are found in approximately 30 families, both naturally and artificially induced (Table 1).

Intumescences are mostly discovered in Gymnosperms, especially in the family of Pinaceae. Pinaceae is a family of monoecious evergreen (some deciduous) trees with smooth or scaly bark and ectomycorrhizal roots [29]. Hypertrophied lenticels on the main tap roots and lateral roots characterize intumescences in Pinaceae. Intumescence injury in this family is known as excrescences, and the main factor that affects this phenomenon is high humidity conditions. Excrescences vary in size and shape, from 0.5 mm to 5–8 mm in diameter, from circular to wart-like patches [8]. This disorder was primarily observed in Pinus genera, while also observed in some spruce (Picea), fir (Abies), hemlock (Tsuga), and tamarack (Larix laricina) (Table 1).

In Angiosperms, the physiological disorders are mostly reported in Myrtacae family, especially Eucalyptus [30, 31, 32, 33]. Eucalyptus is a genus that is mostly native to Australia. They have simple, petiolate, and alternately arranged leaves, with entire leaf blades along the margins, acuminate at the apex, acute to attenuate at the base, and glabrous and smooth on both surfaces [34]. It has a slightly striated cuticle on the leaf epidermal surfaces. The disorders found on leaves of Eucalyptus is referred to as intumescence (Table 1) [15, 30, 31, 32, 35, 36, 37]. Some other publications on intumescence in woody plants were reported by Wallace, who studied intumescence on apple and transparent apple twigs [38, 39]; by La Rue, who found intumescence on Populus sp. in 1930 [14, 40, 41], on Hieracium venosum, Gossypium thurberi (Thurberia sp.), and Mitchella repens in 1931 (Table 1) [37, 41].

In crops, it has been observed that Solanaceous crops are most affected by intumescences, followed by Malvaceae and Fabaceae (Table 1). There are three significant genera of Solanaceae: Solanum, Capsicum, and Nicotiana. Intumescence is mainly observed in the genus Solanum, including tomatoes and wild tomatoes [3, 19, 23, 24, 25, 26, 27, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52], potatoes [11, 12, 22, 53, 54], and eggplants [17]. Intumescence has also been reported in pepper [55]. The symptoms mainly appear on the abaxial surfaces of leaves. However, in some species like Solanum melongena L. [17] and Lycopersicon esculentum (Solanum lycopersicum) [19, 46], it also appears on the adaxial surface of the leaves (Table 2). In some species, such as hairy wild tomatoes [19, 46] and potatoes [56], both sides of the leaves are affected by this physiological disorder. Intumescences were also observed in the petioles of eggplant and tomato leaves and the stems of some tomatoes and potatoes (Table 2). In Capsicum annuum, oedema is observed in the leaf (adaxial surface), flower sepals, and shoot apex (Table 2) [55].

Among the Malvaceae species, investigation on intumescence was mainly focused on Hibiscus vitifolius L. (Table 1). The symptom appeared on both surfaces of the leaves, flower sepals, fruit, petioles, and stem (Table 2) [36, 57, 58, 59, 60]. Intumescence in Fabaceae species is commonly found on their pods, such as in Pisum sativum [10] and Vicia faba [61]; meanwhile, in Phaseolus vulgaris, it was found in the stem [62]. In Cassia floribunda, intumescence was found on the adaxial surface of the leaves (Table 2) [36]. In ornamental plants, researchers focused on investigating oedema in Pelargonium sp. [48, 63, 64, 65, 66, 67] and intumescence in Cuphea sp. [48, 68].

The location of the injury varies depending on plant species (Table 2). For example, excrescence is an injury observed in the roots of Araucariaceae, Pinaceae, and Taxaceae. On the other hand, most of the intumescence found in horticultural crops, such as Solanaceae, was found on their leaves. The differences in the position of this injury might be related to the plant’s characteristics. Conifers, like pines, spruces, and yews, have needle-like leaves with thick cuticle layers. Therefore, it is possible that intumescence does not occur on their leaves but occurs on their roots. Solanaceae leaves also have a cuticle layer; however, some species are sensitive to intumescence injury. There might be a possibility that there are differences in cuticle layer thickness among species and families.

Some studies investigated the differences in intumescence injury sensitivities among several species and/or cultivars [17, 23, 27, 68]. Under the same environmental conditions, there are differences in susceptibility to intumescence injury, which suggests that more complex factors cause this phenomenon. Suppose various cultivars within the same species or various species within the same genus could result in different susceptibility. In that case, it is also possible that the sensitivity of intumescence injury is different between families.

3. Characteristics and morphological features

3.1 Characteristics of intumescence injury

As described in Chapter 1, Williams et al. [21] proposed the term ‘oedema’ to describe this phenomenon of leaf cell hypertrophy, which did not involve epidermal cell enlargement in Pelargonium sp. On the other hand, intumescence starts with one to a few epidermal cells undergoing hypertrophy. Consequently, the affected area expanded as lesions developed, as shown in tomato, ornamental sweet potato, and cuphea disorders. Therefore, in this chapter, we will introduce various types of intumescence in some plants.

Intumescence is characterized as a hypertrophied outgrowth of epidermal cells beginning at a stoma [60]. In contrast to oedema, which is characterized by the stretching of numerous epidermal cells over what appears to be a pocket of water or solutes to form single, blister-like oedema, intumescences include the swelling of multiple individual cells in a specific location.

Küster described two phenomena among diseases with abnormal cell growth: ‘hyperplasia’ and ‘hypertrophy’ [69]. ‘Hyperplasia’ is characterized by cell division, while ‘hypertrophy’ refers to the enlargement of the existing cells with little or no cell division [36]. Intumescence in leaves varied from exclusively hypertrophied (such as in potatoes), to hypertrophy followed by hyperplasia, and the incidence on stems was mainly classified into the latter groups [36].

Intumescences observed in cauliflower [16] and young potato plants [56] usually appeared on the adaxial and abaxial surfaces of the leaves (Table 2). Under direct observation, initially, it came out around the area of the main veins before spreading all over the surfaces. It appeared as small greenish-yellow dots, smooth and glossy, then developing in size, turning into yellow-whiteish, roughened, before becoming dry and wilted in two days [56].

One of the distinctive characteristics of intumescence was discovered in Eucalyptus species. Pinkard et al. [32] experimented with inducing intumescence in Eucalyptus globulus and Eucalyptus nitens leaves under high-humidity environments. The findings indicated that the phenomenon was non-pathological and was known as environmentally induced lenticel-like structures (ERLS). In the early stage of the development, this injury is shown as hyperplasia of spongy parenchyma cells in E. globulus, followed by hypertrophied epidermal and parenchymal cells, and by the later stage, the epidermal cells erupted, and the affected area of palisade parenchymal cells became necrotic. Eucalyptus nitens, on the other side, has distinctive features where palisade parenchymal cells are developed both in the adaxial and abaxial sides of the leaves. When intumescence develops, it develops alongside the second upper epidermis and lower epidermis. The cell division of parenchyma cells and hypertrophied cells resembled E. globulus. However, the upper epidermal cells in this species remained intact [32].

It has been reported that physiological disorders such as leaf hypertrophy or epidermal cell elongation and subsequent browning occur in various plants. In this review, we would like to explain the morphological features of intumescence in vegetables such as tomatoes.

3.2 Morphological features

As mentioned before, the first characteristic of intumescence is hypertrophy of the leaf epidermal cells. Intumescence occurs on both the adaxial and abaxial sides, but for example, in tomato leaves, the abaxial side is more sensitive than the adaxial side. Even in the same genus of Solanum, it has been reported that Lycopersicon hirsutum (Solanum habrochaites) has a gall-like external morphology, while L. esculentum (Solanum lycopersicum) has a callus-like external morphology [46].

Suzuki et al. [3] investigated the stages in which intumescence is most likely to occur, and it was found that it was less likely to occur in fully expanded mature leaves and young, immature leaves in the early stages of development, while it was more sensitive in expanding leaves. In experiments using leaf disks, fully expanded leaves were used to induce intumescence in several species, including tomatoes, and earlier or later stages of development exhibited little or no injury [70].

We observed intumescence development using a scanning electron microscope (SEM) and light microscope. By SEM, intumescence is confirmed by the hypertrophy of epidermal cells, which had previously covered the surface in a flat, mosaic pattern. It bulged in a semicircular shape (Fig. 1E). The leaf blade, vein, and guard cells are also enlarged in the leaves. Afterward, as the symptoms progress, the enlargement becomes more extensive in a circular shape. The cells inside also enlarge, resulting in a mass of cells protruding from the leaf (Fig. 1E). The enlarged area is not green but translucent white visually (Fig. 1A). Epidermal cell enlargement is observed even in cross-section using a light microscope. In addition, it can be observed that the spongy tissue and the spongy tissue inside it are also enlarged (Fig. 1G, J). Inside the enlarged cells, cytoplasm was observed only around the cell wall, and the interior was enlarged and occupied by large vacuoles. The blistering area in plants such as tomatoes comprises enlarged epidermal and internal parenchyma cells.

When intumescence development in tomatoes progressed (Fig. 1B-D), the enlarged area turned yellowish-brown, and the epidermal cells on the abaxial side ruptured (Fig. 1B, D). When intumescence is severe, it also affects the adaxial side (Fig. 1C), where the leaf surface’s color changes. Using SEM, we can observe the surface of the abaxial side, and it was shown that some cells are ruptured (Fig. 1F). Under the light microscope, it was clear that epidermal cells on the abaxial side rupture, cells in spongy tissue do not maintain their original shape (Fig. 1G, J). More than half of the cells around palisade tissue, which were still unaffected in Fig. 1G, became elongated, and the cell structure changed, resulting in the change of cell shape (Fig. 1J). It is thought to be mechanical damage in intumescence parts, which causes the leaf tissue to break down and become necrotic (Fig. 1B-D).

Our observation showed similar patterns in jute leaves (Corchorus olitorius) seedlings (Fig. 2D, E, I, J, K, M, O). In contrast to tomato, intumescence injury in jute leaves developed more on the adaxial surface (Fig. 2D) than on the abaxial surface (Fig. 2E). Cell enlargements develop from the leaf blade around the midrib to the veins. In the early stage of development, the enlarged cells were greenish-white. Using SEM, the morphological characteristic of intumescences on jute leaves was similar to that in tomato leaves (Fig. 2J, K), where semicircular shapes developed on the leaves. In the later stage, the enlarged cell ruptured. Under light microscope observation, initial cell hypertrophy was observed in the adaxial epidermal cells (Fig. 2P) compared to the normal cells (Fig. 2M). In addition, the later development of elongated cells was observed in the abaxial epidermal cells (Fig. 2P).

On the other hand, in sweet potatoes, cell division was observed in intumescence parts [71], and our observations also showed an increase in cell number in water spinach (Fig. 2O). Unlike tomatoes and jute leaves, intumescence in water spinach developed differently. Under direct observation, it started as white-translucent protrusions on the abaxial and adaxial surfaces of leaves, petioles, and stems (Fig. 2A, B, C). It later turns yellowish and brown before it breaks and becomes necrotic. Under SEM observation, cell elongation and division were found on the leaf blade’s surface. Intumescence in water spinach is single-like (Fig. 2G, H), unlike in jute leaves, whose intumescences are clustered together (Fig. 2J, K). Cell hypertrophy was observed using a light microscope on the epidermal adaxial and abaxial cells (Fig. 2N). When intumescence development progressed, it was speculated that cell division might occur (Fig. 2O). Cell divisions were also observed on water spinach stems [72].

In addition, in our previous paper, when the cross sections of resin-embedded samples of tomato leaves that had developed intumescence were observed, it was found that in normal leaves, resin detached from the cuticle layer of the leaf surface [3]. However, it was noticed that no such detachment of the resin and epidermis occurred in intumescence parts and that it was distinguishable from the normal parts (Fig. 1G). When a transmission electron microscope was used to observe the normal site and where the intumescence occurred, a three-layered cuticle layer in the normal epidermal cells was observed (Fig. 3A, B). However, an abnormality occurred in the cuticle layer in the intumescence area (Fig. 3C, D, E, F, G).

It was demonstrated above that intumescence is accompanied by an anomaly in the cuticle layer. The percentage of intumescence areas can be determined by using a computer scanner to scan the leaves and measuring the area of the toluidine blue-stained areas. Until now, research about intumescence has often been determined visually to determine the score of the intumescence, but this staining method can more accurately indicate the degree of intumescence [3]. When leaves with intumescence were stained with a toluidine blue aqueous solution, it did not stain the cuticle layer but stained the cell walls. Thus, it was found that only the intumescence areas were stained. (Fig. 1H, I, K).

Leaves with advanced intumescence symptoms lose moisture and wilt, eventually turning brown and dying [3]. Morphological studies suggest that cell hypertrophy associated with intumescence compresses vascular bundles and inhibits ductal water transport [46]. The vascular tissue of tomatoes exhibited tyloses, which occluded xylem vessels, apparently blocking water transport and causing wilting and abscission of leaves. In addition, increased water damage from the leaf surface due to loss of the cuticle layer is also thought to be a factor in wilting.

Figure 1: (A) Intumescence injury area (white arrow) on the abaxial side of tomato leaf; (B, C, D) Advanced area where the intumescence injury became severe, and the surface color turned brown (D: magnified); (E) SEM of epidermal cells that occurred intumescence injury (red arrows) on tomato abaxial leaf. Bar = 200 μm; (F) SEM of the advanced area (red arrows) where the intumescence injury became severe on the tomato abaxial leaf surface. Bar = 20 μm; (G, J) Light micrograph of tomato leaf with intumescence injury. The arrow indicates the position of the boundary between the normal and abnormal areas, ab-e: abaxial epidermis of the leaf, ad-e: adaxial epidermis of the leaf, i-ab-e: abaxial epidermal cells apparent during intumescence with elongation, i-pt: palisade tissue apparent during intumescence with elongation, i-st: spongy tissue apparent during intumescence with elongation, pt: palisade tissue, st: spongy tissue, vb: vascular bundle. Bar = 100 μm; (H, I, K) Leaves with intumescence injury stained with toluidine blue O aqueous solution; (H) Abaxial side of a compound leaflet with intumescence before staining, black arrows: intumescence areas; (I) Abaxial side of the compound leaflet with intumescence after staining, black arrows: intumescence areas stained purple; (K) Abaxial side of all leaves with intumescence after staining, black arrows: intumescence areas stained purple. I and J are shown at the same magnification, Bar = 1 cm. K, Bar = 1 cm; B, D, E, G, I, J, K were adopted from Suzuki et al. [3].
Figure 2: Intumescence injury area (white arrow) on the adaxial (A) and abaxial (B) side of the water spinach leaf. Bar = 1 cm; (C) Intumescence injury (white arrows) on the petioles and stem of water spinach leaf. Bar = 2 cm; Intumescence injury area (white arrows) on the adaxial (D) and abaxial (E) side of the jute leaf. Bar = 1 cm; SEM of epidermal cells of normal abaxial surface of water spinach (F) and jute leaves (I). Bar = 100 μm; SEM of epidermal cells that occurred intumescence injury (red arrows) on adaxial (G) and abaxial (H) surfaces of water spinach leaves. Bar = 200 μm; Epidermal cells that occurred intumescence injury (red arrows) on adaxial (J) and abaxial (K) surfaces of jute leaves. Bar = 200 μm; Light micrograph of normal cells of water spinach (L) and jute (M) leaves. Bar = 100 μm; Light micrograph of water spinach (N, O) and jute (P) leaf with intumescence injury. Bar = 100 μm; The arrow indicates the position of the boundary between the normal and abnormal areas; ab-e: abaxial epidermis of the leaf; ad-e: adaxial epidermis of the leaf; i-ab-e: abaxial epidermal cells apparent during intumescence with elongation; i-pt: palisade tissue apparent during intumescence with elongation; i-st: spongy tissue apparent during intumescence with elongation; pt: palisade tissue; st: spongy tissue; vb: vascular bundle; asterisk: indicates the initial cell hyperplasia.
Figure 3: TEM micrographs showing the epidermal cell surface on the abaxial side of the tomato leaf that is unaffected (A, B) and injured by intumescence (C-G). ab-e: epidermal cell on the leaf's abaxial side; i-ab-e: epidermal cell on a leaf's abaxial side affected by intumescence; cu1: the cuticle layer composed of surface wax layer; cu2: an intermediate layer with substantial cutin below it; cu3: a layer that consists of a combination of cutin and cell wall polysaccharides; cw: cell wall of the epidermis; (C) arrows: indicate the region where the outermost surface wax layer disappeared; (E) asterisk: indicates the lower electron density region of the intermediate layer. (A) Bar = 5 μm, (B, F, G) Bar = 1 μm, (C, E) Bar = 2 μm. Adopted from Suzuki et al. [3].

4. Factors affecting intumescence injury

Intumescence injuries are influenced by many factors, one of which is environmental factors, including light, humidity, and temperature. In early reports, it was mentioned that this problem commonly occurred in plants cultivated in a greenhouse. Other factors such as nutrient status, chemical injury, excess water, and air quality also affect the incidence of intumescence injury. Most of the plants are affected with intumescence by the combination of several factors. The severity of this disorder increases with multiple factors affecting each other, and when one of the factors is controlled or in normal condition, the severity can be decreased. Other factors individually contribute to the appearance of intumescence. In this chapter, the individual factors will be explained, and we summarize the factors that induce intumescences in Table 2.

4.1 Light

During the early eras of intumescence study, experiments were conducted in the field, in a glasshouse, and under colorful glasses to investigate whether light affects the occurrence of intumescence. Reports showed that some plants require light to induce intumescence injury.

Dale mentioned that intumescences were not formed in H. vitifolius Linn. when the light is absent [60], as well as in P. sativum [61] and potato plants [56] (Table 2). In contrast, other reports found that lower light intensity increased the incidence of this disorder, such as on tomato leaves grown in glasshouses [7] and grape leaves [56, 71]. However, an investigation on grape leaves found that intumescence formed due to excess light in high humidity [16, 74]. Oedema in geranium is also produced under high light intensity [66].

In this way, early reports showed that intumescence occurred in various light conditions depending on the crop. However, the relationship between light wavelength and intumescence was gradually clarified with the development of artificial light, such as LEDs and photoselective films, in plant factories and greenhouses. Further information regarding these conditions will be explained below.

4.1.1 Red, far-red and yellow light

Tracing back from the early investigation of intumescence injury in plants, red and yellow light were produced by passing the light through red and yellow glass, and plants were conditioned under the glass. Dale found that under red and yellow light, intumescence developed in H. vitifolius Linn., thus speculated this incidence with assimilation, where it was found that starch is accumulated in plants treated with yellow light, followed by red light [60].

Intumescence in wild tomatoes was produced on leaf discs by Morrow and Tibbitts [47] and was conditioned under red, blue, and green lights. This study proposed that two separate irradiance responses were involved in the intumescence development in wild tomatoes: the prolonged red response and the reversible red/far-red response. The prolonged red irradiance was responsible for inducing intumescence or neoplasm development, while the latter was responsible for the regulation of the expression of intumescence development. These phenomena suggested the involvement of phytochrome [47], a red/far-red photoreceptor that plays a role in plant growth and development [75].

High red and blue light ratios in a controlled environment also increased intumescence injury incidence. Intumescence was found in cowpea leaves under 10–15% blue light LED and developed in ‘Triton’ pepper under 15% blue LED light [55]. Wollaeger and Runkle also found that tomatoes grown under 100% red LED showed intumescence on most leaves [49].

4.1.2 Blue and green light

Experiments using H. vitifolius Linn, showed that intumescences did not form under blue and green light, although being conditioned in high relative humidity (RH) [60]. Morrow and Tibbitts found that the intumescence of tomato produced on leaf discs was only 3% under green light and none under blue light [47]. Intumescences were also repressed when grown under blue and green light treatments, and a 1:1 green: red light ratio could suppress intumescence until 40% than 100% red light [49]. However, some reports showed opposition to this theory, such as Seabrook and Douglas, who mentioned the involvement of blue-green light (380–525 nm) in the occurrence of intumescence in potato cultivar ‘AC Brador’ and ‘Shepody’ [54].

Blue LED irradiance mitigated the intumescence injury on cowpea leaves while not significantly suppressing this injury on ‘Triton’ pepper [55]. Oedema on basil was also decreased when treated under high levels of blue light [5, 76]. Growing tomatoes under a high blue-to-red light ratio will decrease the occurrence of this injury. A possible mechanism of blue light irradiance is cell enlargement and cell division inhibition [51, 77]. Applying a 75% blue photon flux (PF) ratio during the photoperiod and EOD-FR light could suppress the intumescence injury in tomato leaves until 5% [51].

4.1.3 Ultraviolet (UV)

Ultraviolet light ranges from UV-A (315–400 nm), UV-B (280–315 nm), and UV-C (100–280 nm) [5]. Lack of UV is known to induce intumescence injury. This phenomenon was observed in pea pods, which showed less intumescence incidence when exposed to near-UV wavelengths of light and occurred when it was absent [9, 10]. This phenomenon was also observed in the leaves of diploid potatoes [12]. Intumescences were prevented by UV-B light irradiating in tomatoes [3, 19, 44, 52, 78] and sweet potato leaves [48].

4.2 Humidity

Plants in high-humidity environments are likelier to develop intumescence injury than those in lower-humidity conditions. This was confirmed by old reports that mentioned this phenomenon in the leaves of potatoes [6], common beans [62], sweet potatoes [79], cabbage [80], tomatoes [42], geraniums [63], eggplant [17], and peas [10] (Table 2). In coniferous plants, high humidity is one of the primary triggers of hypertrophied lenticels on Pinus ponderosa stems [8]. In tomato leaves, intumescences were more severe under 90% RH than 70% RH in a controlled environment [3].

Under high RH conditions, plants require more time transpiring. When the transpiration rate is reduced, it leads to excessive turgor pressure within the leaf cells, which can cause intumescence on the leaf [25, 45, 81]. In some cases, light combined with humidity contributes to the appearance of oedema, for example, in Pelargonium hortorum Ait. Transpiration rates are reduced when plants are grown in lower light intensity and high RH, which increases the occurrence of oedema [66]. However, some publications contradict this finding, such as Lang and Tibbitts, which showed that intumescence injury in L. esculentum (S. lycopersicum) and L. hirsutum (S. habrochaites) also developed under 30% RH [19]. This suggested that some plants and cultivars are prone to intumescence, regardless of humidity.

4.3 Temperature

Early publications stated that high air temperature contributes to intumescence injury, such as in H. vitifolius [57], Ficus elatica, and Impatiens capensis Nutt. (Impatiens fulva) [16, 35]. Intumescence in pea formed when the temperature was around 25–30 °C [61]. It was suggested that high temperatures combined with high humidity and excess water could induce intumescence injury [7, 16, 35, 73, 79, 82]. However, later experiments contradicted the previous ones, such as the one on cabbage [66, 83]. Lang and Tibbitts mentioned that intumescence injury could also be induced under 20 and 25 °C in tomatoes [19], which was not aligned with the previous report that oedema in eggplants was induced at high temperatures [17]. In conclusion, intumescence injury incidents usually occur when the temperature is around 25–30 °C, although low to high temperatures affect differently depending on the plant species and families.

4.4 Nutrient (calcium) supply

Oedema in geranium is affected by low nutrition in the soil [66]. Lack of nutrient supply also increases intumescence in potato leaves [53, 56] (Table 2). Schabow and Palta found that intumescence on the leaves of ‘Russet Burbank,’ a susceptible potato cultivar, can be mitigated by an adequate supply of calcium (Ca) [22]. Intumescence in several tomato cultivars was also found to be a calcium-related physiological disorder, as it occurred when the calcium was insufficient in nutrient solutions [27].

Ca has a function to stabilize cell walls [84]; thus, cell wall structures are prone to loosening when Ca in growing plant tissues decreases. Loosening of the cell walls may be associated with cell hypertrophy in intumescence. Moreover, plant transpiration rate impacts Ca transfer to the leaves. Low transpiration causes less Ca to be transported to the leaves, which lowers the Ca content of the leaves. Sita et al. [27] speculated that under high humidity conditions, transpiration rate and calcium uptake decreased, leading to insufficient calcium content in the leaves, which caused the intumescence to develop.

4.5 Chemical stimulation

Early investigation revealed that intumescence in cauliflower was induced when sprayed using copper ammonium carbonate solution [16]. Other reports also found that intumescence in potato leaves could be induced by using copper nitrate spray [56]. The recent investigation also observed intumescence injury in tomato leaves because of topical treatments of Ca-chelator, EGTA and Ca-inhibitor, LaCl3, and Nifedipine [85].

4.6 Other factors

Excess water is known to be associated with intumescence injury in H. vitifolius [57], potato [56], tomato [7], rose [62], and geranium [63]. Air contaminant is also suggested as the factor causing intumescence in potatoes [53] and tomatoes [19, 32, 43]. Intumescence in cabbage can be induced by wind-blown sand, a mechanical injury [80]. Recent reports also found that intumescence was severe in plants with a high shoot/root (S/R) ratio [23].

Intumescence and/or oedema were not clearly defined as a physiological disorder during the early investigation. This was reported by several reports who refer to them as damage caused by insect or pathological injury (Table 2). For example, oedema in the carnation is related to insect injury (aphid) [86]. Others also reported that intumescence in Peperomia sp. was caused by fungal infection [32, 87]. However, recent reports referred to this phenomenon as a physiological disorder.

Table 2: Agents/factors, location in plants, and features of intumescence injury in plants

Intumescence Agent/Factor Species Location of Intumescence Features References
Air quality and airborne factor(s) Eucalyptus spp. Leaf [30, 32]
Populus spp. [14, 41]
Solanum lycopersicum Leaf (adaxial & abaxial), Stem*, Petioles* callus*, cell elongation [7], [19]*, [32, 43], [42], [45]*, [46]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [53], [56]
Chemical injury Brassica oleracea var. botrytis Leaf (adaxial & abaxial) cell elongation [16], [13, 14]
Brassica oleracea var. capitata Leaf (adaxial & abaxial) cell division, cell elongation [41, 83], [80]
Ricinus communis Stem [13, 14]
Solanum lycopersicum Leaf (abaxial) cell division*, cell elongation* [48]*, [85]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [56]
Excess water Abies balsamea [8]
Aphelandra porteana Morel [41, 103]
Eucalyptus spp. Leaf [30, 32]
Ficus elastica Roxb. [16, 35]
Hibiscus vitifolius L. Flower sepals, Fruit, Leaf (adaxial & abaxial), Petioles, Stem cell division, cell elongation [57, 58, 59, 60]
Impatiens capensis Nutt. [16, 35]
Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63]
Pelargonium sp. [32, 64]
Picea mariana Root cell elongation [8]
Picea rubens Root cell elongation [8]
Pinus ponderosa Root, Stem cell elongation [8]
Pinus rigida Root cell elongation [8]
Pinus sylvestris Root cell elongation [8]
Pinus virginiana Root cell elongation [8]
Pisum sativum Pods cell division [10, 61]
Rosa sp. Bark, Leaf cell elongation [62]
Ruellia formosa Andr. [41, 103]
Solanum lycopersicum Leaf (abaxial), Stem* cell elongation [7], [32, 43], [45]*
Solanum melongena L. Leaf (adaxial), Petioles cell elongation [17]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [56]
Tsuga canadensis Root cell elongation [8]
Fungal infection Eucalyptus regnans F. Muell Leaf [31, 32]
Peperomia sp. [32, 87]
Genetics Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63], [65, 66]
Solanum chilense Leaf (abaxial) cell elongation [42]
Solanum habrochaites Leaf (adaxial & abaxial) galls, cell elongation* [19], [46]*, [47]
Solanum lycopersicum Leaf (adaxial & abaxial), Stem*, Petioles* callus*, cell division**, cell elongation** [19]*, [24], [42], [46], [48]**, [52]
Solanum melongena L. Leaf (adaxial), Petioles cell elongation [17]
Hormones (including ethylene) Malus domestica cv. 'White Transparent' Buds, Internodes, Proximal or distal ends cell division, cell elongation [38]
Malus sylvestris Buds, Internodes, Twigs cell division, cell elongation [38]
Populus grandidentata Leaf (adaxial & abaxial) cell elongation [40, 41], [105]
Populus tremuloides Leaf (adaxial & abaxial) cell elongation [40, 41], [105]
Solanum habrochaites Leaf (adaxial & abaxial) galls, cell elongation* [19], [46]*, [47]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [11], [53], [56]
Humidity (air and soil) Aesculus hippocastanum Stem [14, 101]
Aphelandra porteana Morel [14, 103]
Artocarpus incisa Stem [14, 101]
Brassica oleracea var. capitata Leaf (adaxial & abaxial) cell division, cell elongation [41, 83], [80]
Eucalyptus globulus Leaf (abaxial) [32], [35]
Eucalyptus nitens Leaf [32]
Hibiscus vitifolius L. Flower sepals, Fruit, Leaf (adaxial & abaxial), Petioles, Stem cell division, cell elongation [57, 58, 59, 60]
Ipomoea batatas Leaf (adaxial & abaxial) cell division, cell elongation [71], [79]
Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63], [66]
Phaseolus vulgaris Stem cell division, cell elongation [62]
Philodendron pinnatifidum Stem [14, 101]
Pinus ponderosa Root cell elongation [8]
Pisum sativum Pods cell division [10]
Populus grandidentata Leaf (adaxial & abaxial) cell elongation [14], [40, 41]
Populus tremuloides Leaf (adaxial & abaxial) cell elongation [14], [40, 41]
Ruellia formosa Andr. [14, 103]
Sambucus nigra Stem [14, 101]
Solanum chilense Leaf (abaxial) cell elongation [42]
Solanum lycopersicum Leaf (adaxial & abaxial), Stem*, Petioles* callus*, cell division**, cell elongation** [3], [19]*, [24], [42], [46], [48]**, [52]
Solanum melongena L. Leaf (adaxial), Petioles cell elongation [17]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [6], [56]
Spiraea concinna Stem [14, 101]
Vitis vinifera Leaf (adaxial & abaxial) cell elongation [16, 74], [35, 36], [56, 73]
Yucca sp. Leaf [36, 62]
Insect injury Dianthus sp Leaf cell elongation [86]
Picea canadensis Root cell elongation [8]
Picea pungens Root cell elongation [8]
Populus grandidentata Leaf (adaxial & abaxial) cell elongation [40, 41]
Populus tremula Leaf cell elongation [16, 69]
Populus tremuloides Leaf (adaxial & abaxial) cell elongation [40, 41]
Light availability and intensity Dianthus sp Leaf cell elongation [56, 82], [86]
Hibiscus vitifolius L. Flower sepals, Fruit, Leaf (adaxial & abaxial), Petioles, Stem cell division, cell elongation [57, 58, 59, 60]
Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63], [66]
Pisum sativum Pods cell division [10], [61]
Populus tremula Leaf cell elongation [16, 69]
Solanum lycopersicum Leaf (abaxial), Stem* cell elongation [7], [32, 43], [45]*
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [56]
Vitis vinifera Leaf (adaxial & abaxial) cell elongation [16, 74], [35, 36], [56, 73]
Light quality (wavelength, UV) Capsicum annuum Flower sepals, Leaf, Shoot apex   [55]
Hibiscus vitifolius L. Flower sepals, Fruit, Leaf (adaxial & abaxial), Petioles, Stem cell division, cell elongation [57, 58, 59, 60]
Ipomoea aquatica Forsk Stem cell division [72]
Pisum sativum Pods cell division [9, 10], [10]
Solanum habrochaites Leaf (adaxial & abaxial) galls, cell elongation* [19], [32, 44], [46]*, [47]
Solanum lycopersicum Leaf (adaxial & abaxial), Stem*, Petioles* callus*, cell division**, cell elongation** [3], [19]*, [32, 44], [42], [46], [48]**, [49]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [12], [56]
Vigna unguiculata L. Walp. Leaf [55]
Mechanical injury Brassica oleracea var. capitata Leaf (adaxial & abaxial) cell division, cell elongation [41, 83], [80]
Hieracium venosum Leaf [37, 41], [32, 106]
Nutrient status Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63], [66]
Populus spp. [14, 41]
Solanum lycopersicum Leaf (abaxial) cell division*, cell elongation* [27], [48]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [22], [53], [56]
Temperature (air and soil) Brassica oleracea var. capitata Leaf (adaxial & abaxial) cell division, cell elongation [41, 66, 83], [80]
Ficus elastica Roxb. [16, 35]
Hibiscus vitifolius L. Flower sepals, Fruit, Leaf (adaxial & abaxial), Petioles, Stem cell division, cell elongation [57, 58, 59, 60]
Impatiens capensis Nutt. [16, 35]
Pelargonium hortorum Ait. Leaf (abaxial), Petioles cell elongation [63], [66]
Philodendron hastatum Schott Leaf [32, 102]
Pisum sativum Pods cell division [10], [61]
Solanum aculeatissimum Leaf (abaxial) [36]
Solanum lycopersicum Leaf (abaxial), Stem* cell elongation [7], [32, 43], [45]*
Solanum melongena L. Leaf (adaxial), Petioles cell elongation [17]
Solanum phyracanthum Leaf (abaxial) [36]
Solanum tuberosum Leaf (adaxial & abaxial), Stem cell division, cell elongation [56]
Unknown Abelmoschus esculentus Leaf (abaxial) cell division, cell elongation [36]
Abelmoschus manihot Leaf (abaxial) cell division, cell elongation [36]
Abelmoschus moschatus Leaf (abaxial) cell division, cell elongation [36]
Abies balsamea Root cell elongation [8]
Acalypha marginata Spreng. Leaf (abaxial) cell division, cell elongation [36]
Ampelopsis glandulosa var. heterophylla Young and/or developing leaves [36]
Cassia floribunda Leaf (abaxial) [36]
Cuphea llavea Leaf (abaxial) cell division, cell elongation [48]
Cuphea spp. Leaf (abaxial) [68]
Eucalyptus botryoides Sm Leaf cell division, cell elongation [36]
Eucalyptus coccifera L'Herit Leaf [15], [37, 41]
Eucalyptus cornuta Leaf [37, 41]
Eucalyptus diversicolor F. Muell Leaf cell elongation [36]
Eucalyptus melliodora A. Cunn Leaf [36]
Eucalyptus resinifera Sm Leaf [36]
Eucalyptus rostrata Leaf (abaxial) [35]
Eucalyptus saligna Sm Leaf [36]
Gmelina philippensis Leaf [36]
Kendrickia walkeri Hook Leaf [36]
Larix laricina Root cell elongation [8]
Manihot spp. Leaf (adaxial) [17, 18]
Mitchella repens Leaf [37, 41]
Parthenocissus quinquefolia Petioles, Young and/or developing leaves cell division, cell elongation [36, 104]
Parthenocissus tricuspidata Young and/or developing leaves cell division, cell elongation [36]
Pavonia arabica Hochst Leaf (abaxial), Stem cell elongation [36]
Pelargonium peltatum Leaf (abaxial) cell elongation [36]. [67]
Pelargonium x ‘Caliente Coral’ Leaf (adaxial & abaxial) cell division, cell elongation [48]
Pinus banksiana Root cell elongation [8]
Pinus caribaea Root cell elongation [8]
Pinus coulteri Root cell elongation [8]
Pinus excelsa Root cell elongation [8]
Pinus monticola Root cell elongation [8]
Pinus resinosa Root cell elongation [8]
Pinus strobus Root cell elongation [8]
Piper ornatum N.E. Br. Leaf (abaxial), Young and/or developing leaves [36]
Ribes aureum Bark, Shoot cell division, cell elongation [62], [107, 108]
Stictocardia laxiflora var. woodii Fruit, Leaf (abaxial), Petioles cell division, cell elongation [60]
Vicia faba Pods [61]
Vitis rotundifolia Michx Leaf (adaxial & abaxial)   [36, 104]

5. Mechanism of intumescence development

In Chapter 2, morphological observation reveals that intumescence is characterized by cell hypertrophy. Why does cell hypertrophy occur? Since intumescence occurs even when fully expanded leaf discs are incubated under UV cut light [53], these changes are considered to occur at the site of each cell. Various physiological studies have been conducted regarding the mechanism of occurrence of intumescence. Photosynthesis was shown to be reduced in intumescence [14, 17, 46]. This is also supported by a comprehensive gene analysis showing that photosynthesis-related genes are affected [52].

A study that uses tomato plants to limit UV radiation exposure to examine the transcriptome analysis of intumescence has recently been published [52]. Extensive intumescences were induced in leaves grown under the blocked UV condition. A comparison of the gene expression profiles of leaf tissues with and without intumescences indicated that different expression patterns for 1604 genes were found, including cell wall biosynthesis, metabolic pathways, hormonal response, and DNA synthesis and repair. Furthermore, most photosynthesis-associated genes were uniformly repressed in the leaves with intumescences. Therefore, it is probable that the hypertrophied cells in intumescent leaves are genetically programmed to express fewer genes involved in photosynthesis and to produce fewer chloroplasts [52]. It was also discovered that genes involved in signaling, ethylene generation, and biotic and abiotic stress response genes were upregulated. Wu et al. [52] indicated that all the signaling and ethylene synthesis-related genes examined were expressed at higher levels in the intumescent leaves. Thus, their findings suggest that ethylene signaling may also play a role in intumescence (Fig. 4). An important gene, 3-beta hydroxysteroid dehydrogenase (3β – HSD), might play a key role in UV inhibition of intumescence development [52].

A water supply is also necessary for intumescence incidence, as intumescences were generally more significant on the disk surface in contact with water [70]. It was thought to be caused by excessive water flowing into the cells at the site of intumescence. In their investigation of the xylem pressure potential, Miyama and Yasui discovered that cells are more prone to rupture when the water potential rises following a change from dry to wet conditions [23]. They suggested that an environment with (combined) elevated temperature and humidity could cause a higher water absorption by plant tissues compared with the water loss by transpiration, which could induce intumescence. The xylem pressure potential decreased under dry conditions to the S/R ratio and intumescence incidence but increased rapidly after exposure to wet conditions. Tomato varieties with large S/R ratios showed significant changes in their water potential in response to changes in the surrounding water environment, and it is thought that cells are more likely to rupture when water potential increases after a transition from dry to wet conditions (Fig. 4).

Recently, there have been reports about the relationship between the Ca content in leaves and the occurrence of intumescence in potatoes and tomatoes [22, 27]; Sita et al. [27] reported that when the Ca content in tomato leaves decreased, the incidence of intumescence increased. Ca is a crucial macronutrient that acts as an intracellular messenger in the cytosol and gives cell walls and membranes their structural function [88, 89]. Ca is taken from the soil and moved by the xylem from roots to shoots in the cation form (Ca2+). The rate of transpiration in plants affects the apoplastic movement of Ca in plants [90, 91]. As cells grow, pectin methylesterase gradually esterifies cell wall pectin, which is then cross-linked by calcium and branches. Temporary insufficiency of calcium in developing plant tissues leads to increased membrane leakage because calcium stabilizes cell walls, which makes them more prone to loosening [84]. The loosening of cell walls may be associated with hypertrophy during intumescence. Under transmission electron microscopy and light microscope examination, expanded cell walls were wavy, indicating a loss of cell integrity (Fig. 3) [3]. In potato plants, intumescence injury has also been linked to cell wall rupture [22].

Ca transport to the leaves is influenced by plant transpiration rate. Since Ca is an immobile macronutrient, it must move from roots to leaves faster at transpiration [92, 93]. Low transpiration causes less Ca to be transported to the leaves, which lowers the Ca content of the leaves. High humidity is a factor that induces intumescence injury in plants [7, 19, 22, 27, 60]. Transpiration rate and Ca uptake decreased under these conditions [94]. Consequently, it is speculated that a Ca deficiency in tomato leaves may be associated with increased intumescence injury under high RH (Fig. 4).

The cuticle layer is important to protect plants from UV and drought stress, and it develops well under high UV and low humidity environments [3, 95, 96]. When UV lacks, and humidity is high, the formation of the cuticle layer also decreases, and the cuticle layer becomes abnormal due to cell enlargement. The epidermal and internal parenchymal cells enlarge and lose their photosynthetic function. It is thought that the loss of water from the epidermis increases, which has caused the loss of function of the cuticle layer, causing necrosis of both the leaf epidermal cells and the internal cells. Eventually, the entire leaf wilt, leading to death [3].

Figure 4: Mechanism of intumescence injury development. Green boxes refer to environmental factors inducing intumescence injury. Blue box: intumescence injury. Arrows show the flow of the mechanism of intumescence injury.

In summary, there are three main factors in which their mechanism of intumescence occurrence can be explained. In UV-blocked leaves, cell hypertrophy is induced, leading to intumescence and reduced photosynthetic rate (Fig. 4). High humidity also contributed to the development of intumescence, as the pressure potential of the leaf increases under high humidity. When humidity increases, the amount of transpiration decreases, resulting in a decrease in Ca influx into the leaves and a decrease in Ca concentration. As a result, the Ca in the cell walls decreases, making the cell walls more likely to loosen, enlarge, and easily rupture, resulting in intumescence (Fig. 4). Under insufficient UV combined with high humidity conditions, the formation of the cuticle layer also decreases, and the cuticle layer becomes abnormal due to cell enlargement. The epidermal and internal parenchymal cells enlarge and lose their photosynthetic function. It is thought that the loss of water from the epidermis increases, which has lost the function of the cuticle layer, causing necrosis of both the leaf epidermal cells and the internal cells. Eventually, intumescence occurred, and the entire leaf wilt, leading to death.

6. Genetic factors

In some plants, such as geraniums, tomatoes, and eggplants, intumescence was reported differently among several cultivars within one species when treated under the same environmental conditions. Many researchers, especially recent studies, are researching why this phenomenon happened. In this chapter, we will explain the genetic factors involved in the incidence of intumescence injury.

6.1 Differences among species and/or cultivars

As described in Chapter 1, we understood that intumescence appeared in some species within the same genus or family. In fields or production areas, it was found that intumescences sometimes appeared when farmers changed the production crops into different cultivars, while intumescence did not appear when growing other cultivars. One question arises from this phenomenon: Does intumescence injury differ within cultivars?

The differences in intumescence incidence among species and/or cultivars have been an object of study since the 1960s. Metwally et al. [66] conducted an experiment using two geranium cultivars to determine the environmental and cultural practice factors affecting the incidence of oedema. Results showed that the ‘Dark Red Irene’ cultivar was more susceptible than the other, ‘Princess Irene,’ though both were conditioned under the same treatment. The ‘Dark Red Irene’ transpiration rate was higher than the other, resulting in excess water in the leaf cells and leading to oedema formation [66]. The differences between these two cultivars were speculated to be genetically involved, as shown in their cell structures, stomatal behavior, ability to increase their cell water content and resist turgor pressure, and cell wall compositions [65, 66]. The cultivar with fewer xylem components in the stems and petioles, more compact tissues, thicker cell walls, and smaller cells [65] may result in less water translocation to the leaves. It also had larger substomatal chambers [65], larger apertures, and reduced stomatal diffusional resistance [97]. As a result, there was less oedema susceptibility due to increased transpiration. The differences in intumescence severity were also observed in several Cuphea species [68].

A similar trend was observed in vegetables. For example, in eggplant, intumescences were observed in two out of six cultivars investigated by Eisa and Dobrenz [17]. Intumescences in eggplant resembled those found in tomatoes and potatoes, although they were morphologically smaller and more uniform. The differences between susceptible and strong cultivars are speculated to be genetically related [17]. Similar phenomena were also observed in potatoes, where the ‘Russet Burbank’ cultivar is more susceptible than the ‘Atlantic’ cultivar. These differences could be related to ‘Atlantic’s ability to absorb and utilize calcium, which was more efficient than ‘Russet Burbank’s [22].

Miyama and Yasui compared several cultivars and found that intumescence severity is related to the S/R ratio [23]. Cultivars with high S/R ratios were prone to sudden changes in water conditions, in which cells become easily ruptured and cause intumescence. Our study was also conducted using twelve cultivars subjected to different calcium nutrient conditions [27]. It was discovered that some cultivars showed intumescence incidence under insufficient calcium, such as ‘Reika’ and ‘CF Rinka 409’, while others remained healthy.

6.2 Genetic analysis

In order to identify genetic loci associated with intumescence and determine whether maternal effects via inheritance of plastids play a role as well, Prinzenberg et al. [24] grew tomato plants of four different RIL populations under HPS and red/blue LED supplemental lighting in a greenhouse. They also measured the severity of intumescence on 4-week-old plants. It was concluded that genetics, including the maternal background, determined intumescence strongly. A large quantitative trait locus (QTL) was found at the exact location on chromosome 01 in two of the three RIL populations, designating this locus as a breeding target for tomato plants with reduced intumescence susceptibility. Across the three replicates in time, the main QTLs are repeatable. Furthermore, they are visible in both light settings, demonstrating the QTLs’ repeatability. The observed QTL and the maternal impact did not interact, which may facilitate intumescence-resistant breeding. It is possible to select an ideal maternal genotype separately and cross it with a genotype with an allele conferring reduced susceptibility to intumescence. A resistant locus on chromosome 01 and independent selection of maternal background could significantly reduce the incidence and symptoms of intumescence. At this location, extensin-like proteins may be potential genes underlying the intumescence QTL [24].

7. Method to suppress or control the intumescence injury

Chapter 3 explains what factors can induce intumescence injury, both naturally in the field, in a greenhouse, and in a plant factory with a controlled environment. In this chapter, we tried to find some approaches to control or suppress the occurrence of intumescence injury in plants. Based on the explanation of the environmental factors that cause intumescence, the mechanism of intumescence occurrence, and the cultivar differences, the following measures are useful for preventing intumescence. In general, some approaches included environmental conditionings (light, humidity), methods related to the supply of nutrients, and selection of cultivars.

7.1 Supply sufficient UV rays and light conditioning

The UV-transparent film is effective for Solanaceae cultivation, where intumescence can easily occur in the greenhouse. In controlled environments such as greenhouses and plant factories, use light that includes UV. In Japan, a commercial product can be used to prevent intumescence development. It is a UV-B light that has effectively controlled powdery mildew disease in cucumbers and tomatoes [98]. Therefore, it might be effective for controlling intumescence as well. Studies showed that tomato rootstock seedlings were protected against intumescence injury by daily exposure to UV-B at 6.7 mmol m–2 d–1 (2.7 kJ m−2 d−1) [78].

Besides UV light, applying blue light and end-of-day far-red light (EOD-FR) also inhibits intumescence development. Eguchi et al. [50, 51] found that tomato plants cultivated under red and blue LEDs lacking UV-B radiation experienced less intumescence damage after receiving EOD-FR light treatment. The mitigation response was saturated at EOD-FR doses as low as 1.1 mmol m–2 d–1, which was acquired by 5.3 mmol m–2 s–1 FR irradiance for 3.3 min. The intumescence injury can be effectively mitigated by combining intense blue photon flux (50% of PPF or greater) during the photoperiod with a tiny dosage (1 mmol m–2 d–1) of far-red lighting after each photoperiod [50, 51, 78].

7.2 Avoid high humidity

High humidity induces intumescence, so to countermeasure this problem, plants should be kept at low humidity and not left in high-humidity environments for longer than two days to prevent intumescence when cultivated under artificial settings. High humidity is related to the decrease in transpiration rate, which also affects the translocation. Therefore, low humidity and a high transpiration rate are necessary to reduce intumescence [3].

7.3 Provide sufficient calcium

Exogenous Ca2+ administration may mitigate intumescence harm in ‘Russet Burbank’ potato [22]. Controlling humidity, providing extra UV, and adding extra Ca2+ to avoid intumescence development in plants cultivated in greenhouses were also suggested. Additionally, intumescence damage could be effectively reduced by raising the Ca content in the shoots. Another efficient way to raise the Ca concentration in shoots is to grow them in a nutrient solution with a high Ca concentration [27]. However, caution is necessary because increased Ca concentration may prevent potassium (K) absorption, leading to K insufficiency [99, 100]. To prevent intumescence harm, Ca solution can be sprayed often onto growing leaves by foliar spraying [27].

7.4 Use tolerant cultivars

On the production scale, using cultivars sensitive to intumescence injury could affect the yield and outcome when this injury happens. Therefore, using cultivars resistant to such injury is also significant since the prevalence of intumescence injury varies throughout cultivars [22, 23, 27].

The combination of methods explained above is important to control or prevent intumescence injury. The most important is that if all methods are used together, intumescence might not appear. Some countermeasures can be applied generally, while others need to be applied case by case. Applying the right method would efficiently reduce this problem from a small scale to a production area. The expense of investing in preventative measures may be worthwhile if the crop is suffering from severe leaf senescence and low yields. In some cases, the occurrence of this disorder may not be sufficient to justify the cost of fully renovating a greenhouse.

8. Conclusion and future prospect

The characteristics, environmental conditions, mechanisms, and genetic backgrounds of intumescence injury have been cleared up above. Some effective countermeasures, especially Ca supply, have been proposed that can be applied in controlled conditions and open fields. However, the problems below remain.

  • 1) There are many studies about intumescence injury in some plant species, like Solanaceae, but many species have little information on whether intumescence occurs or not.

    2) The cell elongation mechanism during intumescence injury development is still unknown. Plant cell structure is determined by the intracellular cytoskeleton and the direction of cell wall microfibrils, but the relationship to intumescence is unclear. It is necessary to clarify the relationship between some plant hormones and Ca in the cell walls with cell structure changing mechanism during intumescence occurrence.

    3) Intumescence severity differed in some cultivars, but there is still little research about the genes related to this tolerant trait, expression, and physiological mechanisms.

Studying intumescence injuries to resolve unknown problems leads to strengthening the epidermal tissue of plants. When an epidermal cell is strong, it may protect the plants from abiotic and biotic stress. It is thought to have a significant impact on agriculture.

References
 
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