Table Of ContentThe Japanese Society for Plant Systematics
ISSN 1346-7565 Acta Phytotax. Geobot. 65 (1): 37–42 (2014)
Short CommuniCation
Karyotype Analysis of Japanese Burmannia (Burmanniaceae)
mikio aoyama1,* and hiromi tSubota2
1Botanical Garden, Technical Center, Hiroshima University, 1-3-1, Kagamiyama, Higashi-Hiroshima 739-
8526, Japan. *[email protected] (author for correspondence); 2Miyajima Natural Botanical
Garden, Graduate School of Science, Hiroshima University, 1156-2, Miyajima, Hatsukaichi 739-0543, Japan.
Cytological studies on four myco-heterotrophic species of Burmannia (Burmanniaceae) in Japan were
carried out. The finding of a chromosome number of 2n = 176 = 88II (88 bivalents) in B. championii dif-
fered significantly from previous reports of 2n = 12, 64–72. Chromosome numbers of 2n = 120 for B.
cryptopetala, 2n = 60 for B. itoana and 2n = 24 for B. nepalensis are reported here for the first time. Wide
variation in chromosome number, chromosome length and karyotype at resting stage and metaphase was
observed among the four Japanese species of Burmannia.
Key words: Burmannia, Burmanniaceae, chromosome number, Dioscoreales, karyotype, myco-hetero-
troph
Approximately half of the members of the ge- shown in Table 1 and Fig. 1. For observations,
nus Burmannia are achlorophyllous and myco- young flower buds were collected and their ova-
heterotrophic plants. They exhibit evolutionary ries were cut into 1–2 mm thick sections. Pre-
pathways and/or speciation mechanisms that al- treatment, fixation, and chromosome staining fol-
low analysis of interesting biological processes. lowed Aoyama & Yokota (2012).
Furthermore, there is great cytological diversity Classification of the chromosome types at
in the genus, with chromosome numbers ranging resting and mitotic prophase stages follows Tana-
from n = 6, 2n = 12 to n = ca. 87–99, 2n = ca. 136 ka (1977). Terminology of chromosome mor-
(Ernst & Bernard 1912, Schoch 1920, Larsen phology at mitotic metaphase, based on the posi-
1963, Schinini 1975, Spitmann 1975, Rübsamen tion of the centromere, follows Levan et al.
1986). (1964).
Among the five species recorded in Japan, an
autotrophic, chlorophyllous species, Burmannia Results
coelestis, may now be extinct, although it is still
locally common in the rest of its range (Anony- Burmannia championii Thwaites, 2n = 176 =
mous, 2013). In this paper, detailed karyotype 88II, Table 2, Fig. 2
analyses of the remaining four species, B. cham- The chromosomes at resting stage appeared
pionii Thwaites, B. cryptopetala Makino, B. itoa- as numerous chromomeric granules and chroma-
na Makino and B. nepalensis (Miers) Hook.f. is tin blocks scattered throughout the nucleus.
reported and the cytological relationships of these Some chromatin blocks had a rough surface and
species discussed. stained lightly, while others were small, rounded
and stained darkly. The chromosomes at resting
Materials and Methods stage were intermediate between a simple chro-
mocenter type and a complex chromocenter type
Materials and their collecting localities are (Fig. 2A). The chromosomes at mitotic prophase
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Fig. 1. Flowers of myco-heterotrophic Burmannia in nature. A, B. championii. B, B. cryptopetala. C, B. itoana. D, B. nepalen-
sis. Bars = 10 mm.
table 1. Source of materials
Species Localities Altitude Voucher
B. championii Toyosaka, Higashi-hiroshima City, Hiroshima Pref. 400 m HIRO-MY 2522
B. cryptopetala Miyajima, Hatsukaichi City, Hiroshima Pref. 20 m HIRO-MY 6602
B. itoana Kunigami-son, Okinawa Is., Okinawa Pref. 450 m –
B. nepalensis Kirishima, Kirishima City, Kagoshima Pref. 590 m HIRO-MY 42530
Taruzakura, Tarumizu City, Kagoshima Pref. 640 m HIRO-MY 42531
formed early-condensed segments located in the Burmannia cryptopetala Makino, 2n = 120, Table
proximal and interstitial regions (Fig. 2B). A 2, Fig. 3
chromosome number of 2n = 176 was consistent- The chromosomes at resting stage and mitotic
ly counted at mitotic metaphase in four cells from prophase were similar in morphology to those of
three plants (Fig. 2C and D). Eighty-eight biva- Burmannia championii described above (Fig. 3A
lent chromosomes were counted in each of two and B). A chromosome number of 2n = 120 (the
embryo mother cells at meiotic metaphase I (Fig. first report for this species) was uniformly count-
2E). The chromosome number of 2n = 176 = 88II ed in three cells of two plants (Fig. 3C and D).
reported here differs from reports of n = 6 and 2n The chromosomes in the complement varied
= 12 (Ernst & Bernard 1912), and n = 32–36 and gradually in length from 2.2 to 0.7 μm (Fig. 3D).
2n = 64–72 (Schoch 1920). The chromosomes in Most chromosomes were median-centromeric or
the complement varied gradually in length from submedian-centromeric, although in a few of the
3.0 to 0.9 μm (Fig. 2D). Most chromosomes were smallest the position of the centromeres could not
median or submedian centromeric, although in a be determined with certainty. Burmannia cryp-
few of the smallest ones the position of the cen- topetala exhibited a similar karyotype to B.
tromeres could not be determined with certainty. championii.
Burmannia championii shows a homogeneous
karyotype with the chromosomes gradually vary- Burmannia itoana Makino, 2n = 60, Table 2, Fig.
ing in length and with symmetric centromeric 4
positions. The chromosomes at resting stage and mitotic
prophase were similar in morphology to those of
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Fig. 2. Chromosomes of Burmannia championii. A, resting stage. B, mitotic prophase. C, mitotic metaphase showing 2n = 176.
D, drawing of C. E, meiotic metaphase I showing 2n = 88II. Bar = 10 µm.
table 2. Comparison of chromosome characters in four species of Burmannia.
Chromosome length Karyotype
Species max.–min. average total at metaphase
at resting*
(µm) (µm) (µm) length, centromeric position
B. championii 3.0–0.9 1.6 288 intermediate gradual, symmetric
B. cryptopetala 2.2–0.7 1.1 132 intermediate gradual, symmetric
B. itoana 2.4–1.2 1.5 87 intermediate gradual, symmetric
B. nepalensis 5.9–2.7 4.0 96 complex gradual, symmetric and asymmetric
*complex; the complex chromocenter type, intermediate; intermediate type between simple chromocenter type and complex
chromocenter type (Tanaka 1977).
Burmannia championii described above (Fig. 4A sis at resting stage had a rough surface, stained
and B). A chromosome number of 2n = 60 (the lightly and were without the small round darkly
first report for this species) was uniformly count- stained chromatin blocks observed in the species
ed in two cells of two plants (Fig. 4C and D). The described above. The chromosomes at resting
chromosomes in the complement varied gradual- stage were of the complex chromocenter type
ly in length from 2.4 to 1.2 μm (Fig. 4D). Among (Fig. 5A). The chromosomes at mitotic prophase
the 60 chromosomes, 54 were median-centro- formed early-condensed segments that were lo-
meric and six were submedian-centromeric. Bur- cated in all regions of the prophase chromosomes
mannia itoana showed a similar karyotype to B. (Fig. 5B). A chromosome number of 2n = 24 (the
championii. first report for this species) was uniformly count-
ed in three cells of two plants from Kirishima and
Burmannia nepalensis (Miers) Hook.f., 2n = 24, two cells of one plant from Taruzakura (Fig. 5C
Table 2, Fig. 5 and D). The chromosomes in the complement
The chromatin blocks of Burmannia nepalen- varied gradually in length from 5.9 to 2.7 μm.
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Fig. 3. Chromosomes of Burmannia cryptopetala. A, resting stage. B, mitotic prophase. C, mitotic metaphase showing 2n =
120. D, drawing of C. Bar = 10 µm.
Fig. 4. Chromosomes of Burmannia itoana. A, resting stage. B, mitotic prophase. C, mitotic metaphase showing 2n = 60. D,
drawing of C. Bar = 10 µm.
Fig. 5. Chromosomes of Burmannia nepalensis. A, resting stage. B, mitotic prophase. C, mitotic metaphase showing 2n = 24.
D, drawing of C. E, chromosome alignment in order of decreasing length. Bars = 10 µm for A–D and 5 µm for E.
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Among the 24 chromosomes, four (Nos. 17, 19, tion in these three species.
20 and 22) were median-centromeric, four (Nos. The myco-heterotrophic B. nepalensis and B.
3, 4, 9 and 10) were subterminal-centromeric; the lutescens are placed together in a different clade
remaining 16 chromosomes were submedian- from the above three species. The chromosome
centromeric. The two largest chromosomes had number of 2n = 24 in B. nepalensis, and n = 6 and
secondary constrictions in the proximal region of 2n = 12 (Ernst & Bernard 1912) or n = 12 (Schoch
their short arms and formed large satellites 1.2 1920) in B. lutescens suggest a basic chromosome
μm in length (Fig. 5E). The karyotype of B. nep- number of x = 6 in this clade.
alensis was homogeneous with the chromosomes
varying gradually in length and was intermediate We sincerely thank Prof. R. D. Seppelt, of the Australian
Antarctic Division, and Mr. P. J. Dalton, University of
between symmetric and asymmetric centromere
Tasmania, for critically reading the manuscript. Dr. M.
positions.
Yokota, University of the Ryukyus, provided valuable
suggestions during the course of this study, and Mr. S.
Discussion Mukai, Mr. S. Nagatani, Ms. M. Kimura, Mr. T. Kama-
miya and Mr. A. Abe kindly helped in sampling the mate-
rials.
The four species of Burmannia in Japan differ
in cytological characteristics from each other
(Table 2). References
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Received March 28, 2013; accepted November 15, 2013
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