**3.4 Effect on mitotic phase frequency**

A study of the mitotic phase frequency revealed that the total number of cells in wastewater of different locations was less than the cells present in distilled water treatment. It exhibited that dividing cells present in distilled water was higher than the remaining three sources of wastewater. In the anaphase stages, it was noticed in the sample of Mathpara to have a higher number of metaphase stages as compared to Preonagar, Harishnagar and distilled water samples (**Figure 4**).

#### **Figure 1.**

*Sprouting shoots along with floral parts of the A. cepa* L. *(a) Sprouting shoots of the onion (15 days old) (b) Cylindrical fleshy leaf with Peduncle (45 days old ) (c) Very good No. of Peduncle with Inflorescence (70 days Old) (d) Immature florets (e) Pedicel of a about equal length all arising from the apex of the Peduncle with single open floret (f) Onion inflorescence at mature bud and bloom stage 98% (100 days old) (g) Two florets are attach with single pedicel (h) Distinctly showing the two parts attached at lower part of pedicel (i) Pistil of the floret (j) Petals, Pistil and Stamens of the floret (k, l, m, n) Vector/pollinator (arrow) observed on the individual flower (o, p) Floral nectar (arrows).*

*Pollination Biology and Environmental Water Pollution Indicator of Onion (*Allium cepa *L.) DOI: http://dx.doi.org/10.5772/intechopen.111475*

#### **Figure 2.**

*Normal division and Chromosomal abnormalities observed in A. cepa L. (2n=16) meristematic cells exposed to three different polluted waters with control (distilled water) (a) Normal Late Prophase (b) Normal Metaphase (c) Normal Early Anaphase (d) Normal Telophase respectively treated with distilled water (e) Telophase with chromosome break (f) Micronucleus of large size in Interphase (g) Multipolar telophase with chromosome bridge (h) Tripolar anaphase respectively treated with Preonagar sample (i) Anaphase bridge with laggard chromosome (j) Fragmented chromosome (k) Anaphase with multi chromosome bridges (l) Bi-nucleus cell, respectively treated with Mathpara sample and (m) Multi nucleus cell (n) Fragmented chromosome (o) Metaphase with chromosome adherence and chromosome break (p) C-metaphase chromosome respectively treated with Harishnagar sample.*

#### **3.5 Chromosomal abnormalities (CA)**

Meristematic cells present in root tips of *A. cepa* L. were exposed to 72 hours of the three different locations water sample and exhibited various chromosomal aberrations induced by chemical agents in the form of changes either in chromosomal structure or in the total numbers of chromosomes compared with distilled water as control. The different types of observed abnormalities included telophase with chromosome break, micronucleus, multipolar telophase with chromosome bridge, tripolar anaphase, anaphase bridge with laggard chromosome, fragmented chromosome, anaphase with multi chromosome bridges, bi-nucleus cell, multi nucleus cell, metaphase with chromosome adherence and chromosome break and c-metaphase chromosome (**Figure 2**). Studies indicated that all the toxic elements present in the different locations sample water caused a significant increase in the total aberrant cells except distilled water (**Figure 3**). The toxic elements in wastewater samples induced the highest aberrant cells frequency of about 18% which was observed to be in Mathpara, whereas medium in Preonagar (11%) and the lowest frequency in Harishnagar (8%) water samples (**Figure 3**). The analysis of chromosomal abnormalities was made mainly on the anaphase and telophase stages of the cell cycle division.

#### **Figure 3.**

*Effect of Distilled water, Preonagar, Mathpara, and Harishnagar water samples on Germination %, Disinhibition Root length %, Chromosomal Aberration (CA) frequency % and Mitotic Index (MI) % in A. cepa* L.

#### **Figure 4.**

*Frequencies of different types of cells after treatment with different water samples (Distilled water, Preonagar, Mathpara and Harishnagar water samples).*

The upward direction of Chromosomal aberrations (CA) of the *A. cepa* root tips meristematic cells reflected a significant decrease in the Mitotic Index (MI) due to taxological elements existing in the wastewater samples (**Figure 3**).

*Pollination Biology and Environmental Water Pollution Indicator of Onion (*Allium cepa *L.) DOI: http://dx.doi.org/10.5772/intechopen.111475*

### **4. Discussion**

Environment and its interaction with the genome help in the expression of physio-morphological characters of all organisms. The inflorescences *A. cepa* L. is of dull white colour whereas *Allium aflatunense* bears violet colour then slowly turned into purple colour [15]. *Allium giganteum* inflorescences produce red-coloured florets from the purple colour [16]. Nevertheless, the onion inflorescence displays a very strong flavour and odour due to the presence of chemical alteration of a volatile secondary metabolite like S-methyl cysteine sulphoxide compound [17]. Temperature strongly influences the reproductive structure of the onion [18]. Interspecies relation may be a cause of the very good method for cross-pollination in the different types of organisms [19]. It is no wonder that nectar offers as a food reward to the insect vectors pollinating the plant to their pollen vectors [20]. In the present investigation, different species of the Genus *Apis* viz., Apis dorsata*, A. cerana,* Apis mellifera and *A. florea* visit the onion inflorescence florets as a gatherer of pollens and nectar (**Figure 4**) [21].

The present study establishes a steady relationship between the wastewater heavy metal toxicity and abnormal cellular behaviour of onions having been indicated by frequencies of the different mitotic phases, MI and the types of structural chromosomal abnormalities. The cytological studies of the meristematic cells of the *A. cepa* L. roots assay provide one of the most reliable and useful protocols for the investigation of environmental pollution, biological monitoring and determination of the toxicity of the different elements present in the different sources.

The lower level of the MI in the *A. cepa* L. meristematic cells can be indicated as a reliable process to determine the presence of cytotoxic agents such as heavy metals in the environment and considered as a real test to evaluate the pollution level in the natural water bodies [22]. Our observation corroborates earlier investigations using the Mitotic Index evaluation as a tool for the detection of cytotoxicity mediators and pollution agents present in the environment [10, 23, 24].

Fiskesjo [25] has established drinking water contamination by copper using the *A. cepa* L. test. Subsequently, the author has successfully extended the same test for others to detect different toxic metals such as Hg, Ni, Cu, Cd, Ne, Al, Mn and Li, establishing chromosomal abnormalities in the form of C-metaphase to be associated with heavy metal Ni [26].

*A. cepa* L. meristematic root cells assay efficiently evaluates different aqueous concentrations of copper mine waste causing cytogenetic effects such as chromosomal abnormalities. The 100 percent concentration of copper mine waste i.e., raw sample presented the highest toxicity and exhibit a relationship with the inhibition of MI along with chromosome breaks, delay, bridges and adherence were the most frequent Chromosomal aberration (CA) observed [27].

Borboa and Torre [28] have assessed heavy metals Zinc and Cadmium genotoxicity linked with chromosomal abnormalities in the *A. cepa* L. test system with cadmium inflicting a greater genotoxic effect. Seth et al. [29], in evaluation studies of the genotoxicity effects of cadmium by the *A. cepa* L test, have revealed inhibition of CA with MN induction.

Contamination by tannery effluents found in river water causes significant frequencies of chromosomal abnormalities and micronucleus in *A. cepa* L. meristematic cells noticed [30].
