A NOTE ON THE HERB , Chelonopsis longipes

Kazuhiko Toyoda

What is C.longipes? Chelonopsis Miq. is one of the endemic genera to the eastern Asiatic floristic reagion which placed in the Lamiaceae (or Labiatae) and is perennial herbs and woody rhizomatous plants. The genus consists of 15 species distributed from eastern Tibet, continental China, Taiwan to Japan (Takhtajan 1986). The five species of subgenus Chelonopsis(Bostrychanthera deflexa,Chelonopsis chekiangensis, C. longipes, C. moschata, C. yagiharana) fall into two subclades. These five species belong to a Sino-Japanese distribution pattern(Wu & Wu 1996). All of the species possess an herbaceous habitat, rhizomes, pubescent leaves , 3/2-bilabiate calyx and leaves lacking capitates glandulaer trichomes (Xiang 2013). Three endemic species in the genus (C.longipes, C. moschata, C. yagiharana ) are found in Japan(Mutara and Yamazaki 1993), and grown in shaded moist rocky to moist forest floors places(Funamoto 2007). C.longipes longipes is distributed in Pacific side in central Honshu(western from Kanagawa Pref.), Shikoku and Kyushu Districts, and showed commonly the somatic chromosome number of 2n=32 (Fnamoto 2007). It is a rare species coming nearest to C.moschata Miq, but with leaves narrower, peduncles much longer and more slender, corolla smaller and intensely purplish. It is almost inodorous ever when we touch it(T.Makino 1899).

C.longopes in Mt. Nijo

I found the capsule of a herb in Mt.Nijo on winter in 2018. Then I didn't know what kind of the herb it was. I began to study this herb and continued to inquire it. The place where this herb grows is situated at Kamori in Mt. Nijo.When spring had come , I identified this herb a kind of Lamiaceae ,Chelonopsis longipes, for fresh leaf had grown up. Much of the forest in Mt. Nijo is a mixed forest of coniferous and deciduous trees such as cryptomeria , hinoki cypress and Japanese oaks. This herb grows in an evergreen coniferous forest as a summer green understory herb. I saw many stocks of the herb grew in shaded moist forest floor at spot A and B. There are 18 stocks of C.longipes at spot A (0.3/�u) and 4 stocks at spot B (0.05/�u).

April 2th 2018, fine day

I saw some seeds in its capsule that I never seen at Kamori during winter. Today I find the plant has fresh leaf. The plant may be Celonopsis longipes. It will not be identified exactly until I confirm to see the flower.

June 2th 2018, fine day

The rare herb, Chelonopsis longipes has many immature buds attached on the top of the needle-like stem , peduncle. Each bud is at the point where a leaf stalk joins the stem, axil.

June 16th 2018, fine day

I went and saw a herb, Chelonopsis longipes , which grew on the forest floor in Mt. Nijo to observe continuously. And I put the data-logger that had a function to get a forest temperature, intensity of sunlight and humidity at the forest floor near the herb. I will study a feature on the herb, Chelonopsis longipes. I picked up its peduncle is drown as follow.

June 17th 2018, fine day

I brought data-logger that I put it on a forest floor where a herb, Chelonopsis longipes, grows yesterday. The data analyzed is as same as the data recorded at the place where another kind of herb, Liparis nervosa, grows. Consequently the relative irradiation taken by that apparatus at where L.nervosa grows is as same as the place where C.longipes grows approximately. It is 0.02 comparing with sunny place. The place where the herb, Pyrola japonica, grows in a deciduous forest indicates the lesser value of relative irradiation than there. It is 0.013.

June 30th 2018, fine day but humid and hot.

I went to observe the herb, Chelonopsis longipes, which I would investigate continuously. I saw the tip of the peduncle growing larger than I had seen it. I picked up a peduncle to draw picture at my home. Around where they are growing I found some sprout of this herb. They might be derived from its own seeds dropped down on the forest floor last year.

July 21th 2018, fine day but humid and hot.

It has not been rain for several days and very hot,38�� approximately. I went to the place where I continued to observe the herb, Chelonopsis longipes. I saw a purple flower that was on the leaf. I took it a picture, but I missed the flower which might drop down on forest floor. Then I found a sepal(calyx) on which had been the flower I had missed.

August 5th 2018, fine day and extremely hot.

The temperature by the road indicates 30�� at 8:00 AM. I went to see the herb, C.longipes. Grasses that disturbs me to reach my destination have cut off. It becomes easier to pass than it was. The herbs had only one flower and almost are young buds. After I went back my home, I observed a pollen and a style of the flower, C. longipes. Pollen has cleavage, may be three cleaveges reason why some closely related one have three. Style is 15.1mm long and anther 2.5mm. This flower has four stamens and one pistil. One of these herbs has several twigs of which leaf arrangement are opposite phyllotaxis. Individual has eight twigs and it has 11.8 receptacles as mean. I saw only one flower of them. Number of receptacle per herb is shown as follows:

Number of each twig has

Summary
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Individual	A:12,8,14,14,12,12,12,10	receptacle respectively	(sum:94,mean:11.8)
Individual	B:8,6,10,8,8,8	receptacle respectively	(sum:48,mean:8.0)
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Why does only one flower open among many buds? The ratio is 1/94=0.01. I have ever seen only one flower on the same herb on 21th in July.

August 18th 2018,fine day,8:00-9:30,25��

I felt a refreshing air in the morning. A thermometer along the road that I run by bike indicated 22��,Although It was very hot and humid yesterday. I arrived at the place where the herb, C. longipes, grows. I didn't find a flower at all. Some sepals(calyx) that grew up were seen.�@It imples that flowers might drop down. Many of buds ripen better than they were on 5th in August. There are three ripe sepals (calyces) on the top of a receptacle that I have never seen. I want to know when the flowers open and how long they continue to open.

August 25th 2018 at Kamori, 30centigrade, fine day ,7:30-8:30

The Thermometer set up near the road 165 indicated 30 centigrade when I went to , 31 when I went back. It was too hot and humid. The herb, Chelonopsis longipes, has changed little than it was in last week. I also found a bud and young one (seedling) that were around the adult one. I took a stack off which had the bud to observe the growth of the bud at my home. �@The young herbs are implied that the seeds might germinate at high ratio. Then why does this herb become a endangered plant? Does it live in the strict environment, the suitable temperature, humidity and sun light? I put a tip of the stem in water to water-cultivate and record a moving image with a time-lapse camera. But the result disappointed me very much. This water-potted herb, C. longipes withered and died in a day. From this consequence I think that it might be very sensitive to environmental change. It may be one of the factors that C. longipes may be the endangered plant.

September 15th 2018 at Kamori, room temperature 23-25 centigrade, cloudy

Eleven flowers of Chelonopsis longipes bloom among three herbs, of which are mature. Five flowers were measured which have picked and taken at random. The mean of length reveals 37.7�}2.1mm long for corolla ,30.6�}1.5mm,for style, 29.3�}1.8mm For long stamen,25.6�}1.7mm for short stamen respectively. Pollens are able to adhere on stigma, for the anther of the long stamen is near the stigma. I saw many pollens with a microscope , which have been on the surface of style. This could indicate an inbreeding. Besides, I think that anthers possess many pollens in the bud, despite it is less than 20mm longitudinally. This implies a protandry.As Makino Tomitaro describes in his treatise that it is almost inodorous even when we touch it. I didn't feel a oder from the herb, even from its leaf , stem and flower. The anther is mature for I see many pollens on it. I don't know whether stigma is mature or not. A nectar is sufficiently secreted which I feel sweat when I lap it.

Table 1
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Sample	Calyx	Corolla	style	stamen1	stamen2
S1	13.0	40.0	31.0	28.5	24.5
S2	12.0	38.5	33.0	32.5	28.5
S3	15.0	38.5	30.0	28.5	25.5
S4	10.5	34.5	29.0	28.5	25.0
S5	8.0	37.0	30.0	28.5	24.5
Mean	11.7�}2.6	37.7�}2.1	30.6�}1.5	29.3�}1.8	25.6�}1.7
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September 17th 2018 at Kamori, room temperature 24-25 centigrade, cloudy sometimes sunny

I went to Kamori again to collect the time-lapse camera which has worked for two days from 15th in the morning to 17th in the morning. I saw 26 flowers, Chelonopsis longipes, open. 15 flowers have opened since 15th Sept. for I found 11 flowers on 15th. The record gotten from the time-lapse camera shows that mature flower dropped on a forest floor during a night and a bud grows to change the length from 19.7mm to 24.6mm,24.6mm to 29.5mm next day respectively. The bud might be assumed to grow 4.9mm/day(0.2mm/hour). Let us presume the juvenile bud may be 10mm long in the beginning, the formula of the bud growth could be linear as follow: y=4.9x + 10 y is a longitudinal length of bud and x is a day. Or y=0.2x+10 y is a longitudinal length of bud and x is a hour. As length of mature flower, which sufficiently has grown up, is about 40mm long, it takes 6days for the bud to flower grown up.

October 1th 2018 at Kamori in Mt.�@Nijo,�@room temperature:23 centigrade ,fine day

I went to see a herb,�@Chelonopsis longipes. It was storm of Taifu24 last night. The strong wind and intense rain may make the flowers fall down. On the way to the spot where the flowers grow. There were many purple flowers which had been buds last week. These are Isodon longitubus . I think dozen of individuals may be there. I collected them and another plant, Tubocapsicum anomalum. I saw many flowers of C. longipes, which could count dozens of flowers. I picked up five flowers to investigate them. After a dissection, I found many fallen flowers that might be sufficiently mature and the length of the long stamen grew over the style. In addition to the fact, the style was caught between long stamens. This suggests that the herb might have a possibility of self-pollination. Moreover the anther has dense hairs to may stick many pollens. As the stamen got long after style had gotten long, the style could receive pollens from the hairs. This herb could increase by means of self-fertilization.

I will experiment using a net to cover those flowers, which disturbs a invitation of pollinators. If the flowers in the net have no seeds, it needs pollination, after all, it doesn't increase by means of self-fertilization. I also studied the process that the flowers were growing from immatured corolla to matured one by means of dissecting them. The result is showed next table. The style might have grown up before stamen did could employ a protogyny. The anther of juvenile stamen already possesses many pollens, even though the length of corolla is 11.5mm at least.

Table 1
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Sample	Calyx	Corolla	style	stamen1	stamen2
S6		43.0	33.0	35.0	30.0
S7	12.0	41.0	34.0	34.0	30.0
S8	11.5	39.0	33.0	32.0	29.0
S9		40.0	34.0	35.0	30.0
S10		40.0	34.5	35.0	30.0
Mean		40.6�}1.5	33.7�}0.7	34.2�}1.3	29.8�}0.4
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Table 1
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Sample	Calyx	Corolla	style	stamen1	stamen2
S11		22.0	20.0	15.0	9.5
S12		14.0	13.0	12.0	8.5
S13		26.0	26.5	18.0	10.0
S14		23.5	22.5	15.5	10.0
S15		18.5	17.5	14.0	10.0
S16		16.5	15.5	12.5	9.5
S17		34.0	32.0	30.5	12.0
S18		11.5	9.5	9.0	8.5
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Oct.6th 2018 at Kamori in Mt. Nijo. It clouds sometimes rains.

The data-loggers for solar radiation set on at 8:30 Am in the forest where C. longipes are seen. And other data-loggers set on at 9:30 at open place where doesn't disturb solar radiation. Another equipment to take a picture every 5 minutes automatically set on there. I measured the length of each part of flower which has fallen down on the forest floor. 19 of flowers that I picked up from the floor are situated at A spot (Table1),6flowers are B spot(Table2). I confirmed how many flowers there are at spot A today. I took a picture of every twig which has flowers. Looking at these pictures, I counted flowers, buds, calyx, which remained on the receptacle after the corolla had fallen down and would make seed in it. The results are shown as follow:

Table 1
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Sample	Calyx	Corolla	style	stamen1	stamen2
S19		43.0	34.5	35.5	31.0
S20		42.5	34.5	35.2	30.5
S21		39.0	31.0	32.5	27.5
S22		38.5	32.5	32.0	27.0
S23		40.0	32.0	31.0	27.5
S24		39.5	31.5	31.5	26.5
S25		39.0	32.0	32.5	28.5
S26		37.5	29.0	30.5	27.0
S27		40.0	34.0	32.0	27.5
S28		35.0	30.5	30.0	25.0
S29		40.0	32.0	32.0	29.0
S30		37.0	29.0	29.0	24.0
S31		40.0	32.5	32.0	27.0
S32		40.5	32.5	33.0	28.0
S33		38.0	31.5	31.0	26.5
S34		39.5	32.5	31.0	27.0
S35		40.0	32.5	31.5	27.5
S36		39.5	32.0	32.0	28.0
S37		38.5	30.0	30.5	26.0
Mean		39.5�}1.8	31.9�}1.6	31.8�}1.6	27.4�}1.6
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Table 1
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Sample	Calyx	Corolla	style	stamen1	stamen2
S38		38.0	30.5	31.0	27.0
S39		35.5	27.0	28.0	24.5
S40		35.0	27.0	27.0	24.5
S41		37.5	30.0	30.0	26.5
S42		32.5	24.5	24.5	21.0
S43		35.0	28.5	28.5	25.5
Mean		35.6�}2.0	27.9�}2.2	28.2�}2.3	24.8�}2.1
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The herb at spot A grow earlier than the one at spot B, because a ratio of calyx at spot A is greater and buds are lesser than spot B. Another herb, Isodon longitubus, was encountered at Spot B which might be two individuals. The location is N34 31 53, E135 41 4. The structure of this flower resembles C.longipes , two pair of stamens and one style. But the anthers are different from the one of C.longipes.

Table 1
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Location	Flower	Bud	Calyx
Spot A	72(41%)	43(24%)	63(35%)
Spot B	25(45%)	19(34%)	12(21%)
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October 8th 2018 at Kamori in Mt. Nijo, room temperature 20-22 centigrade, fine day

I collected the equipments that I have set them on spot A at Kamori three days ago. Surprisingly, two of six equipments have been broken by a animal, which might be a wild boar. Fortunately the data recorded by them are approximately safe to use. From the data, consequently a relative light intensity is 1.4 �} 0.4(%),n=3. The second purpose to come there is the work that I put a net on a bud to defend its pollination. I covered 8 twigs with net, which put out some buds. A moth, which may be the hawkmoth, Macroglossum pyrrhosticta, visited the flower on one occasion at about 9:00 Am. I confirmed the growth of two flowers from the time-lapse camera. One of the flower length increased from 20mm to 37mm during 7 hours, Oct. 6th 22h-Oct.7th 6h,another from 23mm to 40mm.

October 21th 2018 at Kamori in Mt. Nijo, temperature 13��- ,fine day

I found no flowers at point B , N34.5308,E135.6842,at all. I took pictures of each twig which possesses some calyx. Thirteen flowers were found at point A, N34.5311, E135.6831, but almost twigs have only calyx, not flowers. The twigs covered with a bag made by net were safe. I could see some flowers fallen in the bottom of bag. I was able to see the flowers in the sunny point there at 10:00 AM. I have never seen a pollinator at all since then. Whether calyces might have seeds or ovary or not, I counted to confirm the existence of calyx there. I searched 50 calyxes and I could see 50 ovaries on the receptacle. Its ratio is 100%. I didn't see any pollinator during two hours.

Nobember 23th 2018 at Kamori in Mt. Nijo, temperature 10��- ,fine day

It was cold today morning. The thermometer by the road route 165 indicated 10��. I went to a spot in Kamori to observe a herb, Chelonopsis longipes. Many of their capsules were swollen like a rugby ball of which length was measured to 14.4 �} 0.67(n=6;see table1) . There are 4 ovules grown enough in that capsule. The sizes of them are shown next. (table2) The result are 6.9�} 0.35 for length, 5.3�} 0.49 for width, 3.6�} 0.39 for thickness, 69�} 15 for volume(?). The volume is calculated by means of next formula ; Volume=4��(length/2)(width/2)(thickness/2)/3 If its density is as much as water, its weight may be calculated to be 0.069g, because weight of water is 1g par 1000 ?.

Table 1 Size of fruit(mm)
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Sample	Length	Width	Thickness	Volume
S1	7.2	5.8	3.6	78.7
S2	6.8	5.1	3.6	65.3
S3	6.7	5.0	3.4	59.6
S4	6.2	4.5	3.3	48.2
S5	7.2	5.5	3.2	66.3
S6	7.1	5.9	4.4	96.5
Mean	6.9�}0.35	5.3�}0.49	3.6�}0.39	69�}15
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Table 2 Length of Calyx (mm)
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Sample	Length
S1	15.5
S2	15.0
S3	14.5
S4	14.0
S5	14.0
S6	13.5
Mean	14.4�}0.67
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Dec.8th 2018 at Kamori,5��~10��,8:30~11:00 AM, fine

I cut and picked up two twigs of C.longipes at spot A and one twig at spot B. I found new stock at sopt B where was approximately 10m away from the old one. Each twig had many calices. A twig had 5 calices and another did 9 calices at spot A. I saw that they had no seed at all. To study morphological feature of the seed , I picked up many calices which possessed the seeds. A calyx of C.longipes has 4 seeds. I didn't find all of them were matured perfectly. One of them possessed two matured seeds and two immatured one. Another did one matured and three immature, and so on. I measured each length, width and thickness of the seed in the capsule A-H. I calculated the volume from them. The result is shown next;

Table 2 (mature fruit , immature fruit) per Calyx
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Sample	(mature fruit , immature fruit)	number of calyces
S1	(1,3)	3
S2	(2,2)	2
S3	(3,1)	2
S4	(4,0)	0
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Table 3 mature fruit and immature fruit per Calyx
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Sample	mature fruit	immature fruit
A	1	3
B	3	1
C	2	2
D	3	1
E	1	2
F	1	3
G	2	2
H	1	3
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Table 4 Size of fruit A-H
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Sample	Length	Width	Thickness	Volume
A1	5.70	4.90	3.10	45
A2	5.30	3.30	0.70	6
A3	4.90	2.80	0.60	4
A4	4.80	2.95	0.65	5
B1	6.55	4.15	2.10	30
B2	6.35	3.85	2..05	26
B3	6.35	3.60	2.20	26
B4	6.10	3.30	1.30	14
C1	6.35	4.35	3.10	45
C2	6.10	3.60	2.65	30
C3	5.20	2.75	1.00	7
C4	5.30	2.65	0.85	6
D1	7.05	6.30	3.10	72
D2	6.25	5.30	4.60	80
D3	7.55	4.80	2.85	54
D4	6.30	3.45	1.80	20
E1	7.35	4.95	3.10	59
E2	5.40	3.25	0.75	7
E3	4.20	2.80	0.35	2
F1	6.50	5.05	3.20	55
F2	5.70	2.70	0.95	8
F3	5.40	2.40	1.10	7
F4	5.50	2.20	0.85	5
G1	6.70	5.50	3.30	64
G2	6.80	5.40	3.50	67
G3	6.30	3.60	0.95	11
G4	6.05	3.50	0.75	8
H1	6.95	6.30	3.50	80
H2	6.35	3.50	1.30	15
H3	6.40	2.85	1.05	10
H4	5.40	2.40	0.90	6
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Dec.15th 2018 at Kamori,5��~7��,8:30~12:00 AM, fine and sometimes cloudy day

I went to the spot A and B in Kamori to ascertain whether capsule of C.longipes , which has been bagged before ,possesses a seed or not and a ratio of fruition on all capsule. There are 18 stocks of the herb at spot A. I searched 6 stocks of them , which have been bagged in September .The stock A ,of which twigs aren't bagged, possesse 13 capsule with seeds and 102 capsule without seeds and other twigs bagged possess 1 capsule with seeds ,9 capsule without seeds respectively.

The result of fruition at sopt A

Table 1 Result of fruit set
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Bagged twig of	With fruit	Widthout fruit	percentage
a	1	9	10
b	2	6	25
c	0	4	0
d	0	3	0
e	0	4	0
f	0	5	0
Total	3	31	5.8�}9.3
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Table 2 Result of fruit set
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Control twig of	With fruit	Widthout fruit	percentage
a	13	102	13
b	10	14	42
c	1	18	5
d	2	17	11
e	12	83	13
f	19	75	20
Total	57	309	1 6.9�}11.9
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Table 2 The data for GMLL
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ID	TR	FL	FR
a	T1	115	13
a	T2	10	1
b	T1	24	10
b	T2	8	2
c	T1	19	1
c	T2	4	0
d	T1	19	2
d	T2	3	0
e	T1	95	12
e	T2	4	0
f	T1	94	19
f	T2	5	0
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glmmML (formula = cbind (FR, FL - FR) ~ TR, family = binomial, data = ex1, cluster = ID)

Table 1 The result analysed by the way of GMLL
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	coef	se(coef)	z	Pr(>\|z\|)
intercept	-1.7020	0.3007	-5.659	1.52e-08
TRT2	-0.8713	0.6513	-1.338	1.81e-01
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Scale parameter in mixing distribution: 0.5849 gaussian
Std. Error: 0.2816
LR p-value for H_0: sigma = 0: 0.01808
Residual deviance: 15.95 on 9 degrees of freedom AIC: 21.95

While the ratio of fruition taken from the twig a to f , which has not been bagged, is 16.9�}11.9%(n=366), on the case of bagged twig shows 5.8�}9.3%(n=34). The result with an analysis GLMM reveals -0.8731 for TRT2coef and 0.181 for Pr. This implies that the bagging is effective to disturb the fruition with 82% certainty.

The result of fruition at sopt B (unbagged)

Table 1 The result data at spot B
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Twig of	With fruit	Widthout fruit	percentage
1	1	28	3.4
2	1	9	10
3	1	4	20
4	1	2	33
Total	4	43	16.6�}11.2
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Dec.24th 2018 at Kamori,7��~9��,9:00~10:30 AM, fine day

The green leaves of C.longipes have turned yellow and many of them have fallen down on the forest floor at spot A. There I put 5 photo-data-logger made by myself and another logger RS-13L on the floor. On the other hand I put same loggers on the sunny place of my house to compare with the one at spot A as the control.

Dec.28th 2018 at Kamori,4��~4��,9:00~10:30 AM, fine day

I collected the data-logger that I had put them on 24th Dec. from spot A in Mt. Nijo. The result of the relative light intensity recorded is 0.023 �} 0.004(n=5) of which data is recorded for 500 minutes from Dec.24th 9:30 to 24th 17:00 (Table 1). While the result recorded on 6th in Octber was 0.014 �} 0.004 (n=3) and on 17th in Jun was 0.02 (n=1).

Table 1 Relative light intensity at Spot A
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Set point(Deta-logger)	light intensity(Lux)	relative light intensity
C(B1)	16710	0.017
11(B2)	22040	0.022
6(B4)	20926	0.021
A(B5)	25118	0.025
D(RS-13L)	26618	0.027
Control(RS-13L)	985652	1
Mean and SD	n=5	0.023�}0.004
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The result on mean temperature obtained from 24th 9:30 to 28th 9:19 is 4.6 �� and mean humidity is 80.4% in spot A. On the other hand the result on mean temperature and humidity obtained by another data logger ,which put on a sunny place as the control, shows 8.1 �� and 60.7% respectively.
* The batteries for the data loggers have exhausted completely for 4 days.
The leaves of C. longipes have been completely yellow. Almost of them have fallen but some remain on the twig.