Materials and Methods
This study was conducted from April to July 2019 under greenhouse
conditions (had a temperature of 25±5oC with 60%
relative humidity) to simulate the natural condition. The greenhouse is
located at Jiangsu University, Zhenjiang, China (32.20°N, 119.45°E).Wedelia trilobata and Wedelia chinensis were chosen in
this study. Wedelia trilobata (WT) is an annual
invasive plant species in China, while Wedelia chinensis(WC) is its congener native species, belonging to theAsteraceae family. WT is a clonal evergreen
creeping herb that was also found in the southern region of China in the
1970s on a large scale (Qi et al., 2014).
In China, initially, it was introduced as a groundcover species but
later it spread rapidly from gardens to roadsides, and then to the
agricultural fields, and nowadays, it is found near riverside as well
(Song et al., 2010,
Talukdar and Talukdar, 2013).
WC is the native congener of WT, and is
mostly used as a medicinal plant. The growth rate of WCis very slow as compared to WT(Dai et al., 2016a,
Talukdar and Mukherjee, 2008).
WT in China was mostly found in the arid and semi-arid
regions. Some of the population of WT was also found
near the Yangzi River in Jiangsu province of China, which probably
indicates that water fluctuation and nutrient enrichment made
WT successful in these environments. It prefers growing
in nutrient and moisture-rich soils (Dai et
al., 2016a). Ramets of WT and WC were
collected from the same study site of Jiangsu University for the
experiment. Ramets of WT and WC were
growing in the seedling tray with sand as the growth medium. These trays
were placed in a greenhouse. The ramets were irrigated with distilled
water every day, while nutrition was provided weekly through Hoagland
solution. When ramets had two fully expanded leaves, these were
transferred to plastic pots (height = 10 cm, diameter = 13 cm)
containing sand as a growing-medium. The ramets in the pots were placed
in the greenhouse for one week to let them adapt to the greenhouse
condition. The two plant species were left growing under two cultures,
i.e. mono and mixed cultures. In monoculture, one plant of each species
grew; in mixed culture one plant of WT and
WC grew together. Subsequently, the treatments were
implemented as three levels of nitrogen (control = 0.043 g, denoted as
CK; additional = 0.130 g, denoted as N and double additional = 0.261 g,
denoted as 2N) and two levels of water (normal water = 0.450 L/week and
flooding = 0.9 L/week) (Fig. 1). Nitrogen treatments prepared according
to (Wan et al., 2018), comprised of equal
proportions of KNO3 and NH4Cl, and water
treatment was made according to (Rahlao et
al., 2010). Nitrogen treatments were renewed once a week. The required
water amount for water treatments were given three times a week. Two
environmental factors were subjected to all pots after transplanting
based on factorial design: water (normal or flooding), nitrogen
(control, additional, or double additional), and three cultures (two
monocultures and one mixed culture), with five replicates. According to
this experimental design, there
were 90 pots in total (3 nitrogen levels x 2 water levels x 3 cultures x
5 replicates).
Growth and physiological
traits measurement
Two months after the treatment, i.e. in the month of July, leaf
chlorophyll content (CHI) and leaf nitrogen of both species were
measured with portable chlorophyll meter, SPAD; Oakoch OK-Y104, China.
Leaf area was measured with ImageJ software every plant with five
replicates. Plant height of every plant with five replicates was
measured with a measuring scale. The dry weight of above ground (leaf
and stem) and below ground (root) biomass of each individual was
measured separately, after oven drying to constant weight at 72oC for 48 hours (Parepa
et al., 2019).