九州大学総合研究博物館研究報告

Bull. Kyushu Univ. Museum
No. 5, 57-64, 2007

Dimorphism of Leaf Galls Induced by Pseudasphondylia neolitseae(Diptera: Cecidomyiidae) on Neolitsea sericea (Lauraceae) and Their
Distributional Patterns in Kyushu, Japan
Misako MISHIMA* and Junichi YUKAWA*†*The Kyushu University Museum, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan†Kyushu University, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan

Abstract Pseudasphondylia neolitseae (Diptera: Cecidomyiidae) induces two different types of gall (upperand lower types) on the leaves of Neolitsea sericea (Lauraceae). In the upper type, the adaxial side of the gallprojects more prominently than the abaxial side, and vice versa in the lower type. The lower type galls were foundmainly in northwestern and northern Kyushu, while the upper type were found mainly in southwestern and southern
Kyushu and their distributional patterns were parapatric with a little overlapping. The coexistence of both types on asingle host tree was relatively rare. Whenever they coexisted on a single tree, they always appeared together on thesame leaf, although the relative abundance of one type to another was biased toward one side. The ploidy level of hostplants was not related to the gall types, because almost all host trees examined were diploid. Therefore, the gall types
are not determined by the differentiation of ploidy level of host plant, and possibly determined by the gall midge. The
parapatric distribution pattern of the two gall types may suggest that P. neolitseae is now entering into the initial stageof speciation, if gall dimorphism leads to reproductive isolation.

Keywords: Cecidomyiidae, distribution pattern, gall dimorphism, Lauraceae,Neolitsea sericea, Pseudasphondylia neolitseae.

Introduction
Galls have been defined as all manifestations of growth,Hartigiola faggalli (Monzen) (Sato and Yukawa 2004),

whether positive or negative, and of abnormal differen-

and Pseudasphondylia elaeocarpini Tokuda and Yukawa

tiation induced on a plant by animal or plant parasites(Tokuda and Yukawa 2005), have been known to exhibit(Meyer, 1987). Cecidomyiidae (Diptera) includes thegall dimorphism (Yukawa and Masuda, 1996). Pseudas-largest number of gall-inducing species among arthropodphondylia neolitseae Yukawa (Diptera: Cecidomyiidae:taxa containing gall inducers, and many gall-inducing

Asphondyliini) (Yukawa, 1974) is also known to induce

cecidomyiids are monophagous or oligophagous within

two different types of gall (upper and lower types) on theone or a few plant genera (e.g. Yukawa and Rohfritsch,leaves of Neolitsea sericea (Bl.) Koidz. (Lauraceae) (Yu-2005). Cecidomyiid galls are diverse as a whole in shapekawa and Masuda, 1996). In the upper type, the adaxialand galling position, but the appearance and structure ofside of the gall projects more prominently than the abaxi-

each cecidomyiid gall is specific to a gall midge species

al side, and vice versa in the lower type (Fig. 1). Yukawa

in many instances (e.g. Yukawa and Rohfritsch, 2005).

and Masuda (1996) briefly mentioned that the upper andIn contrast, a few cecidomyiid species, such as Masa-

lower types could be seen, respectively, in southern andkimyia pustulae Yukawa and Sunose ( Sunose 1985) ,northern parts of Kyushu, Japan.
We have been considering that the gall dimorphism isCorrespondence : M. Mishima.
FAX : +81-92-642-4299
E-mail address: mishima@museum.kyushu-u.ac.jp

worth being paid attention as it may be a good model to2007 The Kyushu

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Materials and Methods

in the process of gall midge speciation. As the first step,
we tried to gather further information on distributional

Insect and plant materials

patterns of the two gall types induced by P. neolitseae inKyushu and to examine whether or not host plant traits,

The host plant, Neolitsea sericea, is an evergreen treesuch as the ploidy level, are related to the determinationof Lauraceae and widely distributed in Japan (Honshu,of gall types.

Shikoku, Kyushu, and the Ryukyus), southern Korea,
and Taiwan (Momiyama, 1989). In Kyushu, N. sericea isA

commonly seen everywhere except for higher mountains.
Pseudasphondylia neolitseae is fundamentally univoltine
and distributed in Honshu, Shikoku, Kyushu, and theRyukyus, Japan (Yukawa, 1974; Tokuda and Yukawa,2005). In spring, the females lay their eggs in the hostbuds one by one with the aciculate ovipositor. The first
instars hatch several days after oviposition. The firstBsigns of attack are small reddish spots on the upper surface of the fresh leaves. Later, apically rounded subconical swellings appear on the surface of the leaves accompanied by hemispherical swellings on the lower surface.
The galls mature in early summer and each gall contains
one midge larva (Yukawa, 1974; Yukawa and Akimoto,Fig. 1. Two gall types induced by Pseudasphondylia neolitseae.
A: Lower type. B: Upper type.

2006).

Table 1. Localities from which P. neolitseae galls were collected, gall type, the ploidy level of N. sericea, and the number of host plants surveyed.

Kurômaru

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Table 1. Continued

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Field survey and specimen acquisition

mens. Plant voucher specimens were collected from theWe searched for N. sericea trees that bore galled leaves

same plant individuals that bore the galls. All voucherat various localities in Kyushu in 2003, 2004, and 2005,specimens will be deposited to the Kyushu University

and collected fully developed galls from a total of 104

Museum.

localities during the season from October to April (Table

Geographical information and map description1). Each collection site is shown in Fig. 2 as a circle witha numeral that indicates the name of each locality in
Table 1. We distinguished the two gall types from eachAll localities were georeferenced by a handy GPS or map

other by their appearance. In some cases, galls exhibited

reading service provided by the Geographical Survey

an intermediate shape between the upper and lower type

Institute of Japan (http://watchizu.gsi.go.jp/). Geographi-galls. We treated such galls as the intermediate type. The

cal Information System (GIS) was applied for all map

galls collected were fixed in FAA (5% formaldehyde, 5%

description using ArcGIS (ESRI, Japan).

acetic acid, and 60% ethanol) solution as voucher speci-

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Flow cytometry

Results and Discussions

The ploidy level of individual host plants was determined

Distributional pattern of the two gall types

with a flow cytometer as follows. Each fresh ungalledleaf of N. sericea removed from the respective plants was

The lower type galls (Fig. 1-A) were found mainly in

wiped with a sheet of wet paper. Then, leaf tip area withnorthwestern and northern Kyushu, while the upper type

about 10mm×5mm was chopped up into small pieces

galls (Fig. 1-B) were found in southwestern and southernwith a razor blade in a 200μl solution A (Pertec) that wasKyushu (Fig. 3), as mentioned in Yukawa and Masuda

kept cool in a petri dish on ice. The solution, including(1996). In addition to the two gall types, we found the in-extracted intact nuclei, was filtered through 30μm nylon

termediate type of gall in three localities in northwesternmesh (Pertec) and then mixed with 1.5 ml of solution B

Kyushu, and some inverted type of galls (Fig. 4-B) on(Pertec) including DAPI. The mixture was analyzed witha few occasions. As a whole, our data indicated that thePA (Pertec) and the ploidy level was determined by read-two gall types were parapatrically distributed in Kyushu

ing the peak position.

with a little overlapping.
10°

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83 80
52
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Fig. 3. Distribution map of two gall types induced by Pseudasphondylia neolitseae on the leaves of Neolitsea sericea in Kyushu. Yellowcircle: upper type; Blue circle: lower type; White circle: intermediate
type. Dotted lines show the rough borders of two gall types.

Fig. 2. Localities from which material was collected. Each locality is
numbered for convenience of finding locations referred to in this paper(see also Table 1).

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the two gall types on a single leaf would be derived fromThe two dividing lines could be drawn on the map tooviposition by a single female, as has been observed byshow the parapatric distribution pattern of the two gall

Yukawa et al. (1976) and Yukawa and Akimoto (2005).

types (Fig. 3). One line runs in northwestern Kyushu

The minor gall type might be caused by some kinds offrom the vicinity of Tanomoto (81, 82), Nagasaki Pref.,behavioral mistake by the gall midge or by generic varia-

through Ureshino (18), to Kashima (11), Saga Pref. An-tion during the process of gall induction.

other line runs in central Kyushu from the Usuki (22),

Ploidy of host plants

Oita Pref., through the southern foot of Mt. Aso (39-42,47), to Toyono (52-54), Kumamoto Pref. At the westernend of the latter line, the upper type galls replaced theAll host plants examined were diploid, except one trip-

lower type galls between Ue Island (65) and Shimo Is-

loid individual. This result indicates that the ploidy level

land (71) of the Amakusa Islands.

of host plants was not related to the types of gall. The
triploid individual (Fig. 5) was found in Takamori (42),Coexistence of the two gall types on a single tree

AB

Kumamoto Pref. and seems to be a natural polyploid. It
bore lower type galls, but we cannot conclude that trip-Fig. 4. Coexistence of two different gall types induced by
Pseudasphondylia neolitseae
on a leaf of Neolitsea sericea.
A : Upper surface of the hostleaf. White arrow points toward
the upper part of a lower typegall that coexists with uppertype galls. B : Under surfaceof the host leaf. White arrow
points toward the lower part ofa lower type gall that coexists
with upper type galls. Blackarrow points toward inverted
type galls. The upper part of theinverted galls are projected tothe under side of the leaf.

loid individuals always have lower type galls, because
triploid individuals are very rare among wild N. sericea
trees. We will report elsewhere more about the cecidomyiid galls induced on the triploid individual.

Usually the host trees bearing different gall types wereFig. 5. Histograms showing DNA contents of diploid and triploid individuals of Neolitsea sericea that were analyzed with a flow cytometer.
The numeral 1 indicates a peak of diploid and 2 indicates that of triploid.

growing several km apart from each other, but in a few
cases they stood near by in Tanomoto (81 and 82) orwithin a distance of 20 m in Takamori (41 and 42). It
might be possible for the females to fly for at least 20 m,
but every host tree had either upper or lower type galls.
The coexistence of both types on a single host tree wasrelatively rare. Whenever they coexisted on a single tree,

Because the shape of galls is often specific to a gall-

the two types of gall always appeared together on theinducing species, the galls have been regarded as an ex-same leaf, although the relative abundance of one type

tended phenotype of the gall inducers (e.g. Fukatsu et al.,to another was extremely biased toward one side (Fig.

1994; Stone & Schönrogge, 2003; Yukawa & Rohfritsch,4). Therefore, we have never seen any segregate utiliza-2005). Therefore, the parapatric distribution pattern oftion of leaves or shoots by the two different gall types.

the two gall types may suggest that P. neolitseae is now

These observations may suggest that the coexistence ofentering into the initial stage of speciation, if gall dimor-

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phism leads to reproductive isolation. In order to confirm

giola faggalli (Monzen) comb. n. (Diptera: Ceci-

such a hypothesis, we need to clarify the mechanism ofdomyiidae) inducing leaf galls on Fagus crenata

gall shape differentiation. The mechanism is possibly(Fagaceae) in Japan. Esakia 44: 13-26.

related to behavioral traits of the gall midge, such as theStone, G.N. and Schönrogge, K. 2003. The adaptive sig-position of egg-laying (upper or under surface of leaves

nificance of insect gall morphology. Trends inthat are folded in a host bud), the direction of the larval

Ecology and Evolution, 18: 512-522.

head to feed on upper or under surface of the leaves, andSunose, T. 1985. Geographical distribution of two gallso on. In addition, intraspecific mating tests and com-types of Masakimyia pustulae Yukawa & Su-

parison of DNA sequencing data are required between

nose (Diptera, Cecidomyiidae) and reproductive

the two different populations. Then, we have to gather

isolation between them by a parasitoid. Kontyû

phenological data of gall midge emergence from differ-53: 677-689.

ent gall types to examine whether or not a chronological

Tokuda, M. and Yukawa, J. 2005. Two new and threeisolation mechanism is operating on the different popula-known Japanese species of genus Pseudasphon-tions.

dylia Monzen (Diptera: Cecidomyiidae: Asphondyliini) and their life history strategies. Annals

Acknowledgementsof the Entomological Society America, 98: 259272.

We thank Dr. K. M. Harris (former Director of IIE, UK)

Yukawa, J. 1974. Descriptions of new Japanese gallfor his critical reading of an early draft. Dr. T. Fujita, Dr.

midges (Diptera, Cecidomyiidae, Asphondyliidi)

M. Nakazawa, Mr. S. Tagane, Mr. H. Tohyama, Mr. S.

causing leaf galls on Lauraceae. Kontyû 42: 293-

Watanabe, and Ms. Y. Watanabe helped us in collecting

304.

materials, to whom we are indebted. This research was

Yukawa, J. and Akimoto, K. 2006. Influence of synchro-partially supported by a Grant-in-Aid for Young Scien-nization between adult emergence and host planttists (B) from the Japan Ministry of Education, Science,

phenology on the population density of Pseudas-Sports, and Culture to MM (No. 15770059).

phondylia neolitseae (Diptera: Cecidomyiidae)
inducing leaf galls on Neolitseae sericea (Lauraceae). Population Ecology 48: 13-21.

References

Yukawa, J. and Masuda, H. 1996. Insect and mite galls
Fukatsu, T., Aoki, S., Kurosu, U. and Ishikawa, H. 1994.

of Japan in colors. Zenkoku Nôson Kyôiku

Phylogeny of Cerataphidini aphids revealed theirKyôkai, Tokyo.

symbiotic microorganisms and basic structure

Yukawa, J. and Rohfritsch, O. 2005. Biology and ecolo-of their galls: implications for host-symbiont co-gy of gall-inducing Cecidomyiidae (Diptera). In:

evolution and evolution of sterile soldier castes.

Biology, ecology, and evolution of gall-inducing

Zoological Science, 11: 613-623.

arthropods. Eds. A. Raman, C.W. Schaefer, andMeyer, J. 1987. Plant galls and gall inducers. Gebruder

T.M. Withers. Science publishers, Inc. Enfield(NH), USA. pp. 273-304.

Borntraeger, Berlin, Stuttgart.
Momiyama, Y. 1989. Lauraceae. In: Satake et al. (eds)

Yukawa, J., Takahashi, K. and Ohsaki. N. 1976. Popula-Wild flowers of Japan, Woodyplants I. Heibon-tion behaviour of the neolitsea leaf gall midge,sha, Tokyo, pp. 113-125.

Pseudasphondylia neolitseae Yukawa (Diptera,Cecidomyiidae). Kontyû, 44: 358–365.

Sato, S and Yukawa, J. 2004. Redescription of Harti-

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和文要旨
シロダモタマバエ（双翅目：タマバエ科）
によってシロダモ（クスノキ科）の葉に形成されるゴール２型と九州におけるそれらの地理的分布様式
三島美佐子*・湯川淳一*†*九州大学総合研究博物館
九州大学†（〒812-8581福岡市東区箱崎6-10-1）タマバエ科のシロダモタマバエは、クスノキ科の常緑樹であるシロダモの葉にゴールを形成する。ゴールには2型があり、一方は葉の表面側がより突出しており（表型）、他方は葉の裏面側がより肥大している（裏型）。我々は、九州におけるそれぞれのゴール型の詳しい分布パターンを調べた。それぞれのゴール型はやや重なる側所的な分布域を持ち、九州北西部と中央部に２つの分布境界を持っていた。通常、1個体の寄主植物上には、どちらか一方のゴール型しか出現しないが、2型が共存している場合もあった。その場合は、同一葉上に一方の型が多数存在するなかに、他方の型がわずかにみられるというものであった。同じ寄主植物上で、異なるシュートや葉を、どちらか一方の型のゴールが占めている状態で共存しているようなケースは、まったくなかった。調査したシロダモはほとんど2倍体で、1個体のみ3倍体であった。同じ倍数レベルのシロダモに、異なるゴール型が出現していることから、植物の倍数性は、これらのゴール型の決定には関与していないと結論づけられた。今回明らかになった
ゴール2型の側所的な分布様式は、タマバエの行動学的、生態学的、遺伝学的な違いが、2型の発現に何らかの形で関与した結果であると推察される。このことは、シロダモタマバエが種分化のきわめて初期の段階に移行しているという可能性を示唆しているかも知れない。

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