Exploring Wild Plant Conservation from the Perspective of

Reproductive Biology: A Case Study of Rhododendron Excellens

Bo Cai

, ChangQiu Liu

2,*

and ShiXun Huang

School of Tourism and Landscape Architecture, Guilin University of Technology, 541006 Guilin, Guangxi, China

Guangxi Key Laboratory of Plant Conservation and Restoration Ecology in Karst Terrain, Guangxi Instituteof Botany,

Guangxi Zhuang Autonomous Region, Chinese Academy of Sciences, Guilin, China

1142210831@qq.com

hsx@gxib.cn

Keywords: Plant Conservation, Reproductive Biology, Rhododendron Excellens, Pollinators.

Abstract: Wild plant protection has always been one of the key issues at home and abroad, which is of great significance

to biodiversity protection and wild plant resource protection. The wild distribution of Rhododendron excellens

is of high ornamental value, but in recent years due to tourism development and man-made destruction, the

native plant population is shrinking, and the survival and reproduction of wild Rhododendron excellens is

threatened. From the perspective of reproductive biology, this paper discusses the endangered mechanism of

Rhododendron excellens, reveals the obstacles in the process of reproduction on the reproductive level, and

puts forward some strategies for the successful conservation and recovery of wild Rhododendron excellens.

1 INTRODUCTION

Wild plant resource is an important ecological

resource reserve and national property in our country,

and it is an indispensable and important resource in

the process of national development and

construction. The protection of wild plant resource is

not only related to the healthy development of human

beings, more impact on the sustainable development

of the global ecosystem. In recent years, with the

continuous development of China’s economic

construction, there have been numerous

environmental problems. Driven by economic

interests, unrestrained exploitation and destruction

have caused a large number of wild plant species to

be destroyed to varying degrees, even led to the risk

of extinction of many species, how to effectively

protect wild plant resources is a major problem to be

solved in the future.

Plant reproductive biology is the study of the laws

of plant reproductive behavior, which includes three

basic activities of plant growth, maintenance and

reproduction. The life cycle of wild angiosperms

mainly includes the stages of seed, seedling, adult

https://orcid.org/0000-0003-3014-1210

plant, and seed, and the whole process of flowering,

pollination, fertilization and seed-setting completes

the propagation of plants and the continuation of

species (Ren 2012). The natural pollination and seed

propagation of some wild plants are threatened by

human activities and other external factors, which

hinder the normal propagation of plants, the decline

of its own reproductive capacity is also a threat to its

survival.

Based on the theory of reproductive biology,

Field Investigation and experimental design, the

present situation of Rhododendron excellens

resources and pollination under natural conditions

were investigated, to examine whether pollination

failure is a potential risk factor for reproduction, to

study the reproductive potential and the endangered

mechanism of Rhododendron excellens under full

pollination, on the premise of deep understanding of

its biological characteristics and living environment,

the protection and utilization of Rhododendron

excellens are proposed.

160

Cai, B., Liu, C. and Huang, S.

Exploring Wild Plant Conservation from the Perspective of Reproductive Biology: A Case Study of Rhododendron Excellens.

DOI: 10.5220/0011192600003443

In Proceedings of the 4th International Conference on Biomedical Engineering and Bioinformatics (ICBEB 2022), pages 160-165

ISBN: 978-989-758-595-1

2 MATERIALS AND METHODS

2.1 Study Species and Sites

Rhododendron excellens (Rhododendron excellens

Hemsl. et Wils), evergreen shrub, 1.5-3 m high,

leaves evergreen, leathery, oblong-elliptic, corolla

broadly funnelform, flowers large, white aromatic.

Rhododendron excellens are found in mixed

evergreen and deciduous woodlands or thickets at

elevations of 1100-2400 m. The Rhododendron

excellens are mainly distributed in Yunnan Province,

and the distribution is small and concentrated, most

of which are distributed in Wenshan Zhuang and

Miao Autonomous Prefecture, southeast of Yunnan,

and also in Yuxi, Red River and Lincang (Tian 2011).

The experiment was conducted in the wild population

of Ganbazi, Xiachang Township, Malipo County,

Yunnan Province. Population coordinates of Ganbazi

(N: 23 ° 07’27.47”, E: 104 ° 49’21.63”), altitude

1850-1890m.

Figure 1: A photographs of Rhododendron excellens.

2.2 Research Methodology

2.2.1 Observation Record of Pollination

Media and Determination of Visiting

Frequency

Observation records of pollinator species and flower

visiting frequency were conducted in the wild

population of Ganbazi, Xiakinchang Township,

Malipo County, Wenshan Zhuang and Miao

Autonomous Prefecture, in May 2019, May 2020 and

2021 May 2020. The experiment was conducted in

the blooming period of Rhododendron excellens for

three years. During the field observation in the wild

population of Ganbazi, a cluster of healthy growing

plants was randomly selected, and 20 flowers were

selected to be in full bloom, each cluster was

observed for one hour, from 9:00 to 12:00 in the

morning, from 2:00 to 6:00 in the afternoon, and from

7:30 to 10:00 in the evening. Each cluster was

observed for about 10 hours a day and recorded at

half an hour, change of observation site one hour after

completion. In each time period, the number and

species of successive visiting flowers were recorded

in detail. Squatting on the ground at night to view the

sky as a background from the bottom up, with a red

flashlight to observe the weak red light.

Figure 2: Pollinators of Rhododendron excellens.

2.2.2 Self-and Cross-pollination

Experiments and Control Experiments

In May 2020, experiments were conducted in the wild

population of Ganbazi in Xiakinchang Township to

study the affinity, outcrossing potential and natural

pollination of Rhododendron excellens, there were

artificial self-pollination experiment, artificial cross-

pollination experiment and control experiment. More

than ten healthy plants were selected for each

experiment, and a cluster of healthy inflorescences

was selected for each treatment. Artificial self-

pollination experiment and artificial cross-pollination

experiment were carried out in the bud stage before

the flower opened, and then the bag was taken off

when the flower opened for self-pollination and

cross-pollination, the flowers were then bagged until

they wilted; the control experiment was listed at the

bud stage, and the rest were left untreated.

In early November 2020, the three treatments

were bagged at the experimental site, as some of the

fruits were not yet ripe and early bagging prevented

seed loss when the fruits cracked. At the beginning of

December 2020, the fruits were basically ripe. The

fruits were collected from three experimental

treatments at the experimental site, and then brought

back to the room for drying and waiting for the fruits

to open naturally, fruit setting rate and seed number

per fruit were counted in laboratory, in which fruit

setting rate = fruit number/flower number × 100%.

Seed number per fruit was weighed by 100-grain

weight (Yang2020).

Exploring Wild Plant Conservation from the Perspective of Reproductive Biology: A Case Study of Rhododendron Excellens

161

2.3 Statistical Analysis

In order to compare the frequency of flower-visiting,

fruit-setting and seed-setting of different pollination

treatments, Microsoft Excel 2020 was used to

arrange, statistic and plot the data, and SPSS23.0

software was used to analyze the data, the difference

of each group was compared by independent sample

t test, the significance level was P < 0.05.

3 RESULTS AND ANALYSIS

3.1 Species of Pollinators

Over the course of three years, from 2020 to 2021, we

counted about 90 hours of observations, observed

different pollinators during the day and night, and

recorded the frequency of flower visits in detail. The

major flower-visiting insects in Rhododendron

excellens are Hymenoptera, among which bumblebee

is the main flower-visiting insect, bee is the

secondary flower-visiting insect, butterfly and

Drosophila are negligible due to the low frequency of

flower-visiting. Four species of long-billed hawk

moth, Cechetra Scotti, Notonagemia analis,

Acosmeryx Naga and Daphnis Thous hypocrameri,

have been observed visiting flowers at night in

Rhododendron excellens. Flower visiting insects

from the Rhododendron excellens visit more

frequently when the weather is fine, all insects stop

their activity when it is rainy, and all observations are

counted on non rainy days.

3.2 Frequency of Flower Visits by

Pollinators

Flower visiting frequency refers to the number of

flower visiting insects in a unit time. When observing

in the field, a certain number of flowers were selected

and the number and species of flower visiting insects

were recorded, thus the number of insects that visit a

single flower per unit of time (Li 2014) is obtained.

The number of insects and the frequency of visiting

flowers varied greatly in different observation years,

but there was no significant difference between 2019

and 2020, and the number of insects visiting flowers

by day and night increased significantly in 2021

compared with the previous two years, the number

and frequency of bumblebees were significantly

higher in the daytime than in the previous two years.

Since the 2019 observations are scattered, this is

a detailed analysis of the 2020 and 2021 data. In

2020, the frequency of flower visiting of bumblebee

and honeybee is relatively low, the frequency of

flower visiting of Bumblebee is 0.144

times/flower/hour, the frequency of flower visiting of

honeybee is 0.128 times/flower/hour, the frequency

of flower visiting of bumblebee is slightly higher than

that of Honeybee, the frequency of butterflies and day

moths is too low to be considered. During the

observation period, the number and species of flower-

visiting insects are relatively low and there is no

obvious peak activity. In 2020,0.079 flower visits per

hour were observed at night, and there was a more

active peak from 7:30 to 8:30 at night. The frequency

of flower visits at night was not high.

Bumblebees and honeybees were still the main

flower-visiting insects in 2021. The frequency of

flower-visiting of bumblebees was 1.167 and 0.198

respectively. The frequency of flower-visiting of

bumblebees was significantly higher than that of

honeybees, the frequency of flower-visiting of other

insects is so low that it can be neglected. Bumblebees

have a peak of flower-visiting at 5:00 pm, while bees

have a peak of activity at 3:00 pm, as at 4:00 pm, the

bloom-visiting activities of bumblebees and

honeybees decreased dramatically at 12:00 p.m. --

2:00 p.m., possibly due to higher temperatures and

less insect activity. The frequency of flower visits

was 0.3692021 at night, and there was an active peak

between 7:30 and 8:30 at night.

3.2.1 Comparison of Flower Visiting

Frequency of Pollination Media in

Different Years

Data from 2020 and the 2021 show that the frequency

of flower visits by pollinators varies significantly

from year to year. In 2020, the flower visiting

frequency of bumblebees was 0.144 times per flower

per hour, which was significantly lower than that of

2021 bumblebees (1.167 times per flower per hour)

(p < 0.05). There was no significant difference

between bumblebees and 2021 bumblebees in 2020,

0.128 times per half hour and 0.198 times per hour,

respectively. Compared with 2020, the flower

visiting frequency of 2021 Bombus showed an

explosive increase, while the flower visiting

frequency of honeybee increased slightly. In 2020,

the frequency of flower visiting was 0.079 times per

flower hour at night, which was significantly lower

than that of 0.369 times per flower hour at night (p <

0.05), and the frequency of flower visiting was also

significantly increased for the 2021.

ICBEB 2022 - The International Conference on Biomedical Engineering and Bioinformatics

162

Figure 3: Frequency of insects visiting flowers in different

years.

3.3 Results of Pollination Experiment

and Control Experiment

The percentage of fruit set and the average number of

seeds per fruit were obtained by treating the fruits and

seeds of artificial self-pollination experiment, cross-

pollination experiment and control experiment in

laboratory, the fruit setting rate of the artificial cross-

pollination experiment was 86%, and that of the

control experiment was 82%. There was no

significant difference among the three treatments (p

< 0.05). The results showed that Rhododendron

excellens had a certain degree of self-compatibility,

and self-pollination also produced higher fruit-setting

rate, the fruit-setting rate of artificial cross-

pollination was slightly higher than that of the

control.

The average number of seeds per fruit was 734 ±

50 in the artificial self-pollination experiment, among

which the number of seeds of some fruits was 0, the

minimum number of seeds was 120, and the

maximum number of seeds was 4436 The average

number of seeds per fruit was 5472 ± 452 in artificial

cross-pollination experiment. There was only one

fruit with 0 seeds, the next was 256 seeds, and the

most was 15103 seeds In the control experiment, the

average number of seeds per fruit was 3510 ± 312,

part of the seeds was 0, the minimum number of seeds

was 158, and the maximum number of seeds was

10378. There was significant difference (p < 0.05) in

the number of seeds of each fruit in the three groups,

and there was no significant synchronism between the

number of seeds of each fruit, there was a great

significance between the most and least number of

seeds, some seeds did not develop, and the number of

seeds produced was 0.

Figure 4. Average number of seeds produced per fruit under

different treatments.

3.4 Analysis of Pollination Agents and

Flower Visiting Frequency

Based on the results of field experiments, we can

confirm that there are three theoretical effective

pollinators of Rhododendron excellens, mainly

bumblebees and honeybees during the day and Hawk

Moths at night. Due to the integrated flower

characteristics of the Rhododendron excellens,

further quantitative tests of pollination effectiveness

are needed to measure the contribution of different

flower visitors to Rhododendron excellens fitness

(Huang 2007). According to the frequency of flower

visits by pollinators in different years, the frequency

of insect visits by day and night in 2021 was

significantly higher than that in 2020, especially the

frequency of flower visits by day pollinators

increased significantly, the frequency of flower

visiting in night pollination medium also increased

significantly. The increase in the frequency of visits

to pollinators has ensured that Rhododendron

excellens pollination is assured and has reduced

pollination failures due to pollinator restrictions.

3.5 Analysis of Experimental Results

There was no significant difference in fruit setting

rate between artificial self-pollination, cross-

pollination and control, and there was partial self-

compatibility in Rhododendron excellens, although

there was no significant difference in the percentage

Exploring Wild Plant Conservation from the Perspective of Reproductive Biology: A Case Study of Rhododendron Excellens

163

of fruit set, there was a significant difference in the

average number of seeds produced by different types

of fruit, the number of seeds produced by artificial

self-flowering experiment was significantly less than

that by cross-pollination experiment and control

experiment, which indicated that the propagation of

Rhododendron excellens in natural state needed

pollination medium to be carried out, although a low

frequency of flower visits by pollinators was

observed in 2020, it may be that the influence of

human factors on pollinators is less than the threshold

of the minimal pollinators that maintain plant sexual

reproduction, therefore, in the control experiment, the

fruit setting rate and seed number of plants showe d

no obvious limitation of pollination, and the cross-

pollination experiment showed the most fruit setting

rate and seed number, it also shows the reproductive

potential of the Rhododendron excellens in the

presence of sufficient pollinators, and the results of

the present study suggest that there is no pollinator

restriction in the Rhododendron excellens, however,

the impact of human factors on the ecological

environment of the wild population of the megaphone

is still a major problem that needs to be solved

urgently, and efforts should be made to regularize the

construction of the MALIPO national nature reserve

and limit all activities that cause human interference

and destruction, maintaining the original habitat

conditions of wild animals and plants is the

responsibility and goal that must be fulfilled to

protect the animal and plant resources in the future.

4 CONCLUSIONS

Our experiments show that Rhododendron excellens

can not self pollinate and therefore must rely on

pollinators for reproduction. Moreover, this species

has a low degree of self-compatibility, similar to

many other rhododendron plants (Weber 2004), so it

must visit the flowers of animals for the alienation of

pollination for full reproduction. Although sexual

reproduction difficulties are common in some other

endangered plants (Johnson 2004; Walsh 2019), we

found that the natural fruit setting and seed setting

rates of Rhododendron excellens are not low,

indicating that it attracts pollinators and achieves a

considerable degree of reproductive success, so we

can rule out reproductive difficulties as a possible risk

factor for the species. The results showed that

environmental factors had no significant effect on the

reproductive success of Rhododendron excellens, but

the negative effects of limiting human factors did

increase the frequency of bumblebees’ visits to

flowers during the day, Rhododendron excellens

increases the chances of successful reproduction, so

the importance of in situ conservation of wild plants

needs to be addressed.

The Rhododendron excellens has a remarkable

combination of pollinating characteristics, such as

white corolla, aromatic smell and thin and rich nectar

(Martins 2013). Its large funnel-shaped corolla is

characteristic of a subtype of Hyphantria, which has

been reported in the genera Lilium and Cactaceae

(Liu 2019). It was found that the moth was indeed

able to be visited, and the flower visiting behavior

indicated that the moth was probably an effective

pollinator (fig. 2). On the other hand, Rhododendron

excellens flowers last for several days and are visited

by a large number of bees during the day, so their

pollination patterns may not be specialized. We don’t

know which of the daytime and nighttime visitors is

effective or whether both are effective, but further

experiments are needed to confirm this.

5 DISCUSSION

Experimental Underground Jinchangxiang Ganbazi

wild population belongs to the MALIPO National

Nature Reserve, which has extremely rich animal and

plant resources and excellent ecological environment,

and provides important conditions for the

conservation and reproduction of species, however,

due to the recent establishment of the reserve, various

rules and regulations, regulations on management

and protection are still in the stage of perfection. In

previous years, human activities in the reserve were

more frequent, there are many recreational facilities

in the reserve, such as agricultural land and farm

recreation facilities. Local residents have a serious

grazing phenomenon. Tourists also barbecue and

entertain in the reserve. random picking of plants and

trampling of plants cause serious pollution, the

structure and stability of animal and plant habitat

were destroyed to a certain extent, which affected the

abundance of pollinators and destroyed the stable

relationship of mutualism between plants and

pollinators. The paucity of pollination media may

lead to difficulties in sexual reproduction in plants

after 2020 due to the impact of a new global

coronavirus, measures such as strict access control

and registration of commuters have greatly reduced

the number of tourists and idlers in Malipo’s national

nature reserves, and places of entertainment such as

farmhouses are banned, the original habitat in the

reserve has been restored and protected to the greatest

extent under the combined effect of various factors,

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and the original damaged plant and animal

communities have also been restored to the greatest

extent under the condition of sharp reduction of

human disturbance, this may be an important reason

for the large increase in the number and frequency of

2021 pollinators, further explaining the importance of

protecting the habitat of wild plants, as well as the

negative effects of man-made factors on wild animals

and plants.

From the perspective of reproductive biology, this

paper analyzed the visiting frequency of pollination

media in different years and the seed setting rate and

seed setting quantity under different experiments in

the wild Rhododendron excellens, the natural

reproductive potential and pollination of the wild

horn were fully discussed, and the negative effects of

human activities on pollination media were revealed

through the experimental results, and some

suggestions for solving the related problems were put

forward, the purpose of this study is to provide a

scientific basis for the propagation and growth of

wild Rhododendron excellens. The deficiency of this

study is that there is no quantitative test for the

pollination effectiveness of different flower visitors

and no experiment for seed germination, etc., follow-

up related experiments have yet to be conducted.

Pollination is one of the most important processes in

the conservation of wild plants, and is an essential

step in plant reproduction and species continuation.

However, most of the research on wild plants focuses

on the investigation and analysis of the original

habitats of plants, in the wild plant pollination

research is relatively few, this paper expects to wild

plant protection in the pollination of specific research

to make reference value.

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