1. Introduction
Allelopathy describes the biological phenomenon where plants release biochemicals, known as allelochemicals, into their environment to influence the growth of neighboring flora. These compounds, which can originate from plants, microbes, viruses, and fungi, serve a dual role; they can either stimulate or suppress the development of agricultural systems. This influence excludes animal systems, and primarily pertains to plant growth and ecological balance [
1]. Sustainable agriculture aims to balance the need for food production with the preservation of environmental health. In this context, allelopathy offers a natural means of controlling weeds, pests, and diseases, reducing the reliance on chemical herbicides and pesticides. For example, the use of cover crops with strong allelopathic properties can suppress weed growth, thereby minimizing the need for chemical weed control [
2]. Weeds are a significant challenge in agriculture, particularly in organic farming systems, where they can drastically reduce crop yields [
3]. According to Lemerle et al. [
4], crop productivity is diminished by weeds by as much as 5% in the most developed nations and up to 25% in the least developed ones. The pervasive issue of weeds has increased reliance on pesticides, including herbicides, to safeguard crops. However, Fishel [
5] notes that the overuse of herbicides has given rise to weed resistance, undermining their effectiveness and necessitating the search for alternative weed management strategies. The exploration of allelopathy as a natural herbicide is well-founded, and the allelopathic potential of various crops and trees can be utilized to develop more sustainable weed control methods [
6,
7]. The issue of weeds extends beyond the loss of crop yields; as stated by Kubiak, Wolna-Maruwka et al. [
8], the rapid expansion of agriculture and the associated import and export of plants have inadvertently introduced invasive weeds. These weeds, now recognized as a significant threat to biodiversity, release allelochemicals that can be detrimental to crop and plantation growth. They can inhibit seed germination, disrupt the synthesis of essential proteins and carbohydrates, and impair metabolic pathways, resulting in diminished plant vigor and reduced agricultural output [
9,
10]. The allelopathic properties of certain weeds have been studied extensively. For instance,
Heliotropium species have been reported to exhibit inhibitory effects on the germination and growth of various seeds. Aqueous extract of
H. europaeum leaves impede the germination of
Cuscuta campestris (dodder) and
Raphanus raphanistrum subsp.
sativus (radish) seeds [
11], while ethanol extract of
H. indicum aerial parts could suppress the germination and growth of
Lactuca sativa (lettuce) and
Vigna mungo (black gram) [
12]. However, the specific growth-inhibitory substances within
H. indicum have not been isolated, highlighting an area ripe for further study. The concentration and efficacy of allelochemicals are influenced by many factors, including biotic and abiotic stressors, and they vary according to the plant’s age, cultivar, and specific organ. The action mechanisms of crops and allelochemicals are complex and not fully understood; some, such as sorgoleone, have been identified as photosynthesis inhibitors [
13]. Notably, contemporary crop cultivars tend to produce fewer allelochemicals than their wild relatives, a consequence of selective breeding for yield rather than defense [
6].
The cucurbitaceous vegetables are a significant group in the vegetable kingdom, adapting well to various environments, from arid to humid tropics [
14]. These plants are known for their richness in carotenoids, terpenoids, saponins, and phytochemicals. Cucurbit vegetables have been found to have positive effects on human health, with studies showing antioxidant, antidiabetic, anti-inflammatory, and purgative properties [
15]. Cucurbit cultivation is thriving in Bangladesh, with 2.57% of the land dedicated to it, yielding 3.73 million tons annually, helping address summer vegetable shortages [
14,
16].
However, the production of cucurbits faces challenges such as weed infestation, diseases, and insect pests. In some areas, farmers in Bangladesh spend about 25% of their cultivation costs solely on purchasing toxic pesticides [
17]. The repeated and prolonged use of these toxic insecticides has drawbacks, including high application costs, environmental pollution, and health hazards [
18]. Despite this, there is a lack of knowledge regarding weed allelopathy and its impact on cucurbit crops. It is crucial to fill this knowledge gap in order to better understand how different species of cucurbits respond to weeds. Considering the harmful effects of chemicals on non-target organisms and the environment, it is urgent to develop safer, cheaper, and eco-friendly weed management tools for cucurbit crops in Bangladesh. Hence, the present study aims to investigate the allelopathic effects of H. indicum on various cucurbit crops by focusing on the impact of leaf and root extracts on seed germination and subsequent seedling growth.
3. Results
Table 1 shows that most cucurbit crops had high germination percentages for the control groups (T0), with the exception of
L. siceraria (70.00 ± 0.81) and
B. hispida (60.00 ± 0.71). Other cucurbit crops that showed high percentages were
L. acutangula (80.00 ± 0.76),
L. aegyptiaca (90.00 ± 0.80),
M. charantia (90.00 ± 0.85),
T. cucumerina (100.00 ± 0.00),
C. melo (100.00 ± 0.00), and
C. moschata (60.00 ± 0.78). Treatment groups T1 to T4, which were treated with aqueous extract (AQE), generally showed a reduction in germination percentage across all crops, compared to the control. For example,
L. siceraria dropped to 10.66 ± 0.46% in T1 and remained around this value through T4, indicating a potential inhibitory effect of the aqueous extract on germination (
Figure 3).
The methanol extract (MTE) treatment groups T5 to T9 showed similar variable percentages of germination (
Figure 3). In T5,
T. cucumerina exhibited 50.00 ± 0.81% of germination, but in T4 it increased slightly to 50.66 ± 0.71%, indicating some variation in the response to the methanol extract that is statistically significant (
p < 0.001), compared to the control group. The most drastic reductions in germination were observed in
C. moschata, with T8 showing a germination percentage of 10.60 ± 0.47, which is notably lower than the control (
Table 1 and
Figure 3).
Table 2 showcases the shoot elongation of eight different receptor cucurbit crops when exposed to the control (T0) and varying concentrations of aqueous and methanol leaf extracts of
H. indicum at 9 days (
Figure 4). For the control treatment (T0), which is the baseline with just distilled water, the shoot lengths were as follows:
L. siceraria (10.2 ± 0.16 cm),
B. hispida (13.9 ± 0.08 cm),
L. acutangula (13.1 ± 0.08 cm),
L. aegyptiac (11.4 ± 0.08 cm),
M. charantia (22.4 ± 0.08 cm),
T. cucumerina (16.9 ± 0.08 cm),
C. melo (12.4 ± 0.08 cm), and
C. moschata (11.03 ± 0.12 cm).
Upon treatment with aqueous extract (T1–T4), there was a general decline in shoot elongation across all crops, compared to the control. For instance, L. siceraria’s shoot length reduced from 10.2 cm in the control to 8.53 cm (p < 0.05) in T1, and had further decreased to 4.53 cm (p < 0.01) by T4.
Methanol extract treatments (T5–T9) also affected shoot elongation variably. For example,
T. cucumerina had a shoot length of 11.1 cm in T5, which was lower than the control, and then further decreased to 6.1 cm in T6. Notably,
L. acutangula showed an increase in shoot length in treatment T1 to 19.0 ± 0.16 cm, which is higher than the control, but this effect was not maintained in the subsequent treatments. The greatest reduction in shoot length was seen in
B. hispida in treatment T6, with a length of 2.1 ± 0.14 cm (
p < 0.001), significantly lower than the control length (
Table 2 and
Figure 4).
Table 3 outlines the effects of various treatments with aqueous and methanol
H. indicum leaf extracts on the root length of eight cucurbit crops, at 9 days (
Figure 4). On the other hand, the control treatment (T0), untreated shows the initial root lengths:
L. siceraria (8.1 ± 0.21 cm),
B. hispida (9.13 ± 0.12 cm),
L. acutangula (7.06 ± 0.04 cm),
L. aegyptiaca (8.1 ± 0.08 cm),
M. charantia (6.06 ± 0.04 cm),
T. cucumerina (5.43 ± 0.09 cm),
C. melo (6 ± 0.08 cm), and
C. moschata (10.23 ± 0.20 cm). For the aqueous-extract treatments (T1–T4), the root lengths varied, with some reductions and some increases, compared to the control. In the case of
L. siceraria, the root length decreased in T1 to 6.46 ± 0.12 cm (
p < 0.001) but had increased to 8.36 ± 0.12 cm by T4, which is close to the control length.
Methanol extract treatments (T5–T9) also varied in their impact.
M. charantia’s root length decreased to 3.3 ± 0.21 cm (
p < 0.001) in T5, which is significantly less than the control, but by T8 the root length had increased to 9.03 ± 0.12 cm, which is comparable to the control. Notably, some treatments, like T3 for
L. acutangula and T8 for
B. hispida, showed an increase in root length compared to the control, with 8.0 ± 0.08 cm and 9.43 ± 0.16 cm, respectively. However, in some cases, such as
B. hispida in T6, a significant reduction in root length to 4.01 ± 0.08 cm (
p < 0.001) was observed, indicating a strong effect of the methanol extract at this concentration (
Table 3 and
Figure 4).
Table 4 reports the germination percentage of eight cucurbit crops treated with distilled water (T0) and different concentrations of aqueous and methanolic root extracts of
H. indicum at 9 days (
Figure 3). The control group (T0) had the following germination percentages:
L. siceraria (70.00 ± 0.81),
B. hispida (60.66 ± 1.69),
L. acutangula (80.00 ± 0.81),
L. aegyptiaca (90.00 ± 0.81),
M. charantia (90.00± 0.85),
T. cucumerina (90.66 ± 0.47),
C. melo (90.033 ± 0.47), and
C. moschata (60.00 ± 0.81). Treatments with aqueous extract (T1–T4) displayed a general decline in germination percentages for all species, compared to the control. For example,
L. siceraria showed a decrease to 40.00 ± 0.77% (
p < 0.05) in T1 and further, to 20.00 ± 0.78% (
p < 0.05), by T4.
Methanol extract treatments (T5–T8) continued this trend of reduced germination percentages.
T. cucumerina dropped from a control value of 90.66 ± 0.85 to 10.66 ± 0.47% (
p < 0.001) in T7. Certain treatments, like T8 for
C. melo and
L. acutangula, resulted in a substantial germination percentage decrease, compared to the control (20.33 ± 0.47% and 20.00 ± 0.81%, (
p < 0.05), respectively), suggesting some specific concentrations of the extracts might not significantly affect germination (
Table 4 and
Figure 3).
Table 5 presents the shoot length of eight cucurbit crops when treated with distilled water (T0) and various concentrations of aqueous and methanolic root extracts of
H. indicum, at 9 days (
Figure 5). The control group (T0) shows standard shoot lengths:
L. siceraria (10.2 ± 0.16 cm),
B. hispida (14.2 ± 0.16 cm),
L. acutangula (13 ± 0.08 cm),
L. aegyptiaca (11.4 ± 0.08 cm),
M. charantia (22.4 ± 0.08 cm),
T. cucumerina (17.1 ± 0.08 cm),
C. melo (12.3 ± 0.16 cm), and
C. moschata (11.1 ± 0.08 cm). Treatment T1, which involved aqueous extract, increased the shoot length in
L. siceraria to 14.66 ± 0.12 cm, but decreased
B. hispida to 10.33 ± 0.04 cm (
p < 0.01). Subsequent treatments (T2–T8) showed fluctuations in shoot lengths across the species. For instance,
L. acutangula experienced a notable increase to 15.30 ± 0.08 cm at T4.
Methanol extract treatments (T5–T9) also showed variability in their effects, with some treatments resulting in shoot lengths that were shorter than the control. For example, C. moschata decreased to 3.46 ± 0.12 cm (p < 0.001) at T7.
Many treatments resulted in significant changes in shoot length compared to the control, such as the decrease in
M. charantia to 4.06 ± 0.04 cm (
p < 0.001) at T4. Some treatments, such as T8, had less of an impact, as seen in the shoot length of
L. siceraria at 3.6 ± 0.08 cm (
p < 0.001), which was closer to the control measurement (
Table 5 and
Figure 5).
Table 6 details the root length of eight cucurbit crops subjected to distilled water (control) and various concentrations of aqueous and methanolic root extracts of
H. indicum, at 9 days. The control (T0) readings show the baseline root lengths for the crops:
L. siceraria (8.13 ± 0.12 cm),
B. hispida (9.1 ± 0.08 cm),
L. acutangula (7.1 ± 0.08 cm),
L. aegyptiaca (8.1 ± 0.08 cm),
M. charantia (6.1 ± 0.08 cm),
T. cucumerina (5.36 ± 0.09 cm),
C. melo (6.13 ± 0.12 cm), and
C. moschata (10.4 ± 0.08 cm). Aqueous extract treatments (T1–T4) influenced the root length in varying ways. For instance,
B. hispida’s root length increased from 9.1 cm in the control to 9.16 ± 0.12 cm in T7, which was non-significant and almost the same result. In
L. acutangula, the root length decreased to 4.4 ± 0.08 cm (
p < 0.001) in T2, but then increased to 10.2 ± 0.08 cm in T4.
Methanol extract treatments (T5–T9) also showed a differential impact on root length. M. charantia’s root length decreased to 3.1 ± 0.08 cm (p < 0.001) in T7, but was higher in T4, at 10.13 ± 0.12 cm.
Notable significant changes in root length include the increase in
L. siceraria to 9.13 ± 0.12 cm in T6 and the decrease in
C. melo to 1.6 ± 0.08 cm (
p < 0.001) in T7. A treatment like T5 did not significantly affect the root length for certain species, such as
C. moschata, suggesting that the effect of the extract may vary, depending on the concentration and the specific crop (
Table 6 and
Figure 5).
3.1. Effect of Leaf Extract of H. indicum on Chlorophyll Content of Tested Crops
The results from
Figure 6 reveal the differential impact of aqueous (T1–T4) and methanol (T5-T8) extracts of
H. indicum leaves on the chlorophyll content in eight plant species. For
L. siceraria (A), the treatments showed variability in the chlorophyll content, with several treatments significantly differing from the control (T0). Treatment T3 resulted in the highest chlorophyll a content, with
p < 0.001, indicating a highly significant difference. Chlorophyll b also showed substantial variations, especially in T3 and T6. In the case of
B. hispida (B), treatments T1, T2, T3 and T7 notably increased chlorophyll a content significantly, with significance (
p < 0.001). Chlorophyll b levels followed a similar trend in the aqueous extract, but in methanolic extracts highly significant differences were found. Again, for
L. acutangula (C), no significant differences (ns) were observed for any treatments in chlorophyll a and b content, suggesting that this species might be less responsive to the extracts.
L. aegyptiaca (D) displayed a complex response pattern. While T5 had a highly significant effect on chlorophyll a content (
p < 0.0001), T6 presented a very significant effect (
p < 0.001) on chlorophyll b content. Other treatments varied, with T8 showing a significant impact (
p < 0.05) on chlorophyll b.
Furthermore, in M. charantia (E), the majority of treatments did not significantly alter chlorophyll a levels, whereas chlorophyll b levels were significantly increased by treatment T5 (b, p < 0.001). Other treatments were not significant. For T. cucumerina (F), treatments T2, T3, T5, T6, and T7 resulted in a substantial increase in chlorophyll a content (a, p < 0.0001), with T7 also showing a very significant increase in chlorophyll b content (b, p < 0.001). Treatment T8 significantly affected chlorophyll a content (c, p < 0.05). In the case of C. melo (G) exhibited significant increases in chlorophyll a content with treatments T2, T3, T5, and T6 (a, p < 0.0001), while chlorophyll b content did not significantly change with any treatment. Finally, C. moschata (H) showed a very significant increase in chlorophyll a content with treatments T3 and T7 (b, p < 0.001), and a significant increase with T5 and T6 (c, p < 0.05). Chlorophyll b content was significantly increased by treatment T7 (c, p < 0.05), with other treatments not showing significant changes.
3.2. Effect of Root Extract of H. indicum on Chlorophyll Content of Tested Crops
The impact of aqueous and methanol extracts of
H. indicum root on the chlorophyll content of eight cucurbit plants was investigated, with the results shown in
Figure 7. For
L. siceraria (A), the control (T0) exhibited the highest chlorophyll a and b content. Upon treatment, a marked decline in both chlorophyll a and b was observed, with the aqueous extracts (T1–T4) showing a highly significant reduction (
p < 0.0001); in the case of
B. hispida (B), the chlorophyll content in the control group remained high. Interestingly, the treatments did not significantly affect the chlorophyll a and b levels at T1 and T2. Still, for chlorophyll a and b, a significant decrease was observed in the T5–T8 methanol extract treatments (
p < 0.0001), suggesting a differential sensitivity between the chlorophyll types to the methanol extracts. In
L. acutangular (C), the control group showed high levels of both chlorophylls. Following treatment, the chlorophyll a level showed a significant decline at T3 and T4 (
p < 0.05), and a more pronounced effect at T5–T8 (
p < 0.0001). Chlorophyll b levels followed a similar pattern, but with less variance among treatments. For
L. aegyptiaca (D), chlorophyll levels in the control group were relatively higher, compared to treated groups. The treatments caused a significant reduction in chlorophyll content, particularly with the methanol extracts (T5–T8), where the decrease in chlorophyll a and b was highly significant (
p < 0.0001).
Further, in M. charantia (E), the control exhibited the highest chlorophyll a and b content. A dramatic reduction in chlorophyll content was observed in response to the treatments, with the methanol extracts (T5–T8) showing a highly significant reduction (p < 0.0001) for both chlorophyll types. Again, for T. cucumerina (F), chlorophyll a and b levels in the control remained high, similar to M. charantia. Post treatment, there was a pronounced decrease in chlorophyll content, with methanol extracts (T5–T8) exhibiting a highly significant reduction (p < 0.0001) in both chlorophyll a and b. For C. melo (G), the control group showed the highest chlorophyll a and b levels. Upon treatment, chlorophyll levels did not differ significantly (ns), but chlorophyll b levels were affected considerably by methanol extracts (T5–T8), with a notable reduction (p < 0.0001). In the case of C. moschat (H), the control group showed the highest chlorophyll a and b content. Treatments led to a significant decline in chlorophyll a and b, particularly with the methanol extract treatments (T5–T8), where the decrease was highly significant (p < 0.0001).
3.3. MDA Levels
Figure 8 illustrates the malondialdehyde (MDA) levels observed in this study for different plants and treatments. For
L. siceraria (A), both leaf and root extracts showed a significant increase in MDA content across all treatments, compared to the control, with the highest significance observed in T3, T5, and T7 for both, and in T2 and T7 for root extracts (
p < 0.0001). In
B. hispida (B), the MDA levels also increased significantly in response to the treatments, with T1, T7, T4 and T8 (leaf) and T1, T2, T3 and T5 (root) showing the most significant effects (
p < 0.0001). For
L. acutangula (C), the results were more varied; some treatments did not significantly affect T2 (leaf) MDA levels (ns), while others, like T5, and T6 (root) showed high significance (
p < 0.001). Interestingly, T1 demonstrated a significant decrease in MDA content (
p < 0.05).
L. aegyptiaca (D) displayed a pattern where the leaf extract treatments (T1–T4) generally resulted in higher MDA levels (
p < 0.05 to
p < 0.001), whereas T3 and T4 of root extracts showed extremely significant increases (
p < 0.0001) of MDA levels, compared to control.
In M. charantia (E), significant increases in MDA content were observed in treatments T2, T4, T5, and T7 (p < 0.05 to p < 0.001), with the highest significance in T4 (p < 0.001), for both extracts. Conversely, several treatments did not significantly affect MDA levels (ns). For T. cucumerina (F), the MDA response was profound, particularly in treatments T4, T6, T7, and T8, with all showing extremely significant increases (p < 0.0001). However, some treatments did not significantly alter MDA content (ns). C. melo (G) showed significant variation in MDA levels, with the most pronounced increases in T5 and T6 (p < 0.0001) in both cases. Other treatments also significantly elevated MDA content (p < 0.01), while T1, T2 (leaf), and T4 (both) were less impactful, yet still significant (p < 0.05). Lastly, C. moschata (H) exhibited significant increases in MDA content with treatments T5 to T8 (p < 0.05 to p < 0.0001), indicating a strong response to both aqueous and methanol leaf extracts. Notably, T1 and T4 did not significantly affect the MDA levels (ns).