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Ultrastructure of Hirschmanniella diversa early-stage infection in browning rhizomes of Indian lotus

Shigeru Uematsu
Shigeru Uematsu
, 
Tetsuo Yabu
Tetsuo Yabu
, 
Mitsuyoshi Yao
Mitsuyoshi Yao
, 
Takayuki Kurihara
Takayuki Kurihara
 und 
Hironori Koga
Hironori Koga
   | 06. Juli 2020
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Online veröffentlicht: 06. Juli 2020
Seitenbereich: 1 - 9
Eingereicht: 10. Juni 2019
DOI: https://doi.org/10.21307/jofnem-2020-055
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© 2020 Shigeru Uematsu et al., published by Sciendo.
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Figure 1:

Inoculation of H. diversa Sher, 1968 into young lotus rhizome tissues. (A) Apices of excised young lotus rhizome tissue approximately 1 cm in length in the center of petri dishes containing 4.0 ml of 0.4% water agar. (B) Approximately 100 to 300 nematodes were added by pipette to each dish. (C) Petri dishes containing the young lotus rhizome, water agar, and nematodes were frozen in liquid nitrogen at 4.5, 24, 48, and 72 hr after inoculation. (D) Apices of young lotus rhizome tissues (box) were observed under a stereomicroscope after thawing and removal of water agar around the nematodes.
Inoculation of H. diversa Sher, 1968 into young lotus rhizome tissues. (A) Apices of excised young lotus rhizome tissue approximately 1 cm in length in the center of petri dishes containing 4.0 ml of 0.4% water agar. (B) Approximately 100 to 300 nematodes were added by pipette to each dish. (C) Petri dishes containing the young lotus rhizome, water agar, and nematodes were frozen in liquid nitrogen at 4.5, 24, 48, and 72 hr after inoculation. (D) Apices of young lotus rhizome tissues (box) were observed under a stereomicroscope after thawing and removal of water agar around the nematodes.

Figure 2:

Stereoscope micrograph of H. diversa Sher, 1968 invading the apex of a young lotus rhizome 4.5 hr after inoculation. Note that many nematodes remained at the apex of the rhizome even after the water agar was removed. Ne, nematode.
Stereoscope micrograph of H. diversa Sher, 1968 invading the apex of a young lotus rhizome 4.5 hr after inoculation. Note that many nematodes remained at the apex of the rhizome even after the water agar was removed. Ne, nematode.

Figure 3:

Light micrographs showing cross sections of the apices of young rhizomes inoculated with H. diversa Sher, 1968 at 24 hr (A, B) and 72 hr (C, D) after inoculation. (A) Young rhizome apex invaded by nematodes. Note that nematodes (Ne) invaded the epidermis (Ep) and the cavity (Ca), and several layers of cortical cells disappeared. (B) Cortical cell invaded by a nematode. Note that the host cell wall collapsed in the cell invaded by the nematode. (C) Young rhizome invaded by nematodes. Note that invading nematodes stopped penetrating in a vertical direction relative to the surface at about 1 mm depth from the surface. (D) Magnification of Fig. 3C, box. Host tissues surrounding the invading nematodes disappeared and cavities were observed.
Light micrographs showing cross sections of the apices of young rhizomes inoculated with H. diversa Sher, 1968 at 24 hr (A, B) and 72 hr (C, D) after inoculation. (A) Young rhizome apex invaded by nematodes. Note that nematodes (Ne) invaded the epidermis (Ep) and the cavity (Ca), and several layers of cortical cells disappeared. (B) Cortical cell invaded by a nematode. Note that the host cell wall collapsed in the cell invaded by the nematode. (C) Young rhizome invaded by nematodes. Note that invading nematodes stopped penetrating in a vertical direction relative to the surface at about 1 mm depth from the surface. (D) Magnification of Fig. 3C, box. Host tissues surrounding the invading nematodes disappeared and cavities were observed.

Figure 4:

SEM secondary electron images of the surfaces of young lotus rhizome apices invaded by H. diversa Sher, 1968 at 4.5 hr (A-C) and 24 hr (D-F) after inoculation. (A) Nematodes (Ne) invading the epidermis (Ep) of the apex. (B) Magnified micrograph of the white box in Fig. 4A. The forepart of the nematode burrowed under the peeled part of the epidermis of the apex. (C) Magnified micrograph of the black box in Fig. 4A. Clusters of nematodes penetrated the epidermis of the apex through narrow indentations. (D) Clusters of nematodes on the tissues where the epidermis was peeled off accompanied by larger indentations than those seen at 4.5 hr after inoculation. (E) Magnified micrograph of the white box in Fig. 4D. A few clusters of nematodes have invaded the tissues from the peeled part of the epidermis. Note that host tissues around invading nematodes disappeared into indentations. (F) Magnified micrograph of the box in Fig. 4E. Only the posterior parts of most nematodes were observed, suggesting that they had penetrated the tissues deeply.
SEM secondary electron images of the surfaces of young lotus rhizome apices invaded by H. diversa Sher, 1968 at 4.5 hr (A-C) and 24 hr (D-F) after inoculation. (A) Nematodes (Ne) invading the epidermis (Ep) of the apex. (B) Magnified micrograph of the white box in Fig. 4A. The forepart of the nematode burrowed under the peeled part of the epidermis of the apex. (C) Magnified micrograph of the black box in Fig. 4A. Clusters of nematodes penetrated the epidermis of the apex through narrow indentations. (D) Clusters of nematodes on the tissues where the epidermis was peeled off accompanied by larger indentations than those seen at 4.5 hr after inoculation. (E) Magnified micrograph of the white box in Fig. 4D. A few clusters of nematodes have invaded the tissues from the peeled part of the epidermis. Note that host tissues around invading nematodes disappeared into indentations. (F) Magnified micrograph of the box in Fig. 4E. Only the posterior parts of most nematodes were observed, suggesting that they had penetrated the tissues deeply.

Figure 5:

SEM secondary electron images of cross sections of the site shown in Fig. 4E, where a cluster of nematodes invaded the inner apex of a young rhizome. (A) Cross section of the apex of a young rhizome invaded by nematodes (Ne). Note that the nematodes invaded the tips of the tissue to depths of up to about 1 mm. (B) Magnified micrograph of the upper region of the young rhizome shown in Fig. 4A. A cluster of nematodes invaded the apex tissue of a young rhizome. Note that a cavity (Ca) formed around the invading nematodes. (C) Magnified micrograph of the lower region of the young rhizome shown in Fig. 4A. Host tissues at the sides of the cavity around the invading nematodes disappeared as if they had disintegrated.
SEM secondary electron images of cross sections of the site shown in Fig. 4E, where a cluster of nematodes invaded the inner apex of a young rhizome. (A) Cross section of the apex of a young rhizome invaded by nematodes (Ne). Note that the nematodes invaded the tips of the tissue to depths of up to about 1 mm. (B) Magnified micrograph of the upper region of the young rhizome shown in Fig. 4A. A cluster of nematodes invaded the apex tissue of a young rhizome. Note that a cavity (Ca) formed around the invading nematodes. (C) Magnified micrograph of the lower region of the young rhizome shown in Fig. 4A. Host tissues at the sides of the cavity around the invading nematodes disappeared as if they had disintegrated.

Figure 6:

Cross section through the apex of a young rhizome inoculated with H. diversa Sher, 1968 at 24 hr after inoculation. (A) uninoculated cortex (control). Note that artifacts generated by freezing specimens in liquid nitrogen were observed as fragmentation of plasma membranes and collapse of cytoplasm, while cell walls (CW) and starch grains (Sg) of cortical cells (CC) show minimal damage. (B) Cross section through the epidermis (Ep) of an apex and invading nematodes (Ne). Epidermal cells neighboring the invading nematodes degraded and collapsed, and the area surrounding the nematodes formed a cavity (Ca). (C) Cross section of a cortex invaded by a nematode. The cortical cells close to the nematode disappeared, and the surrounding host cells degraded.
Cross section through the apex of a young rhizome inoculated with H. diversa Sher, 1968 at 24 hr after inoculation. (A) uninoculated cortex (control). Note that artifacts generated by freezing specimens in liquid nitrogen were observed as fragmentation of plasma membranes and collapse of cytoplasm, while cell walls (CW) and starch grains (Sg) of cortical cells (CC) show minimal damage. (B) Cross section through the epidermis (Ep) of an apex and invading nematodes (Ne). Epidermal cells neighboring the invading nematodes degraded and collapsed, and the area surrounding the nematodes formed a cavity (Ca). (C) Cross section of a cortex invaded by a nematode. The cortical cells close to the nematode disappeared, and the surrounding host cells degraded.

Figure 7:

Cross section through the cortex of a young rhizome apex invaded by nematodes. (A) Cortical cells close to invading nematodes (Ne). Note that the upper portions of cortical cells disappeared, and the cell wall (CW) and cytoplasm of the remaining cell degraded and became electron-dense. (B) Magnified micrograph of the box in Fig. 7A. Weakly electron-dense materials (thick arrowheads) are observed close to the nematode. Note that the host cells close to the nematode disappeared linearly (thin arrows). (C) Magnified micrograph of the box in Fig. 7B. The weakly electron-dense materials are composed of fine granular materials (FGM), which are denser closer to the nematode. Note that outlines of the cell wall (CW) and the cellular content (CC) are indistinct.
Cross section through the cortex of a young rhizome apex invaded by nematodes. (A) Cortical cells close to invading nematodes (Ne). Note that the upper portions of cortical cells disappeared, and the cell wall (CW) and cytoplasm of the remaining cell degraded and became electron-dense. (B) Magnified micrograph of the box in Fig. 7A. Weakly electron-dense materials (thick arrowheads) are observed close to the nematode. Note that the host cells close to the nematode disappeared linearly (thin arrows). (C) Magnified micrograph of the box in Fig. 7B. The weakly electron-dense materials are composed of fine granular materials (FGM), which are denser closer to the nematode. Note that outlines of the cell wall (CW) and the cellular content (CC) are indistinct.
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