Occurrence and diversity of microalgae in phytoplankton collected from freshwater community ponds of Hooghly District , West Bengal , India

A study on diversity, seasonal occurrence, distribution percentage of microalgal taxa and physicochemical parameters of five community ponds, located in Chinsurah town, Hooghly district of West Bengal, has been carried out. Correlation between occurrence of microalgal genera and some parameters of environment, physico-chemical nature of natural water bodies were explored by Canonical Correspondence Analysis (CCA). Diversity indices have been calculated using PAST software program. A total of 23 microalgal taxa belonging to four algal classes were recorded and the study indicated that the microalgal diversity vary with variation in physico-chemical parameters of water and light intensity. Out of these genera, occurrence of Chodatella sp. in late summer and Tetrallantos sp. and Synechocystis sp. in winter season were specific. In CCA, multiple variables (dissolved oxygen, water temperature, electrical conductivity, pH, light intensity and inorganic phosphorous) played a significant role in occurrence of microalgal taxa. Observation concluded that the Chlorophycean members were dominant throughout the study period and the Shannon–Wiener diversity index was highest for a site with large number of Chlorophycean member. This study will help in future to assess water quality.


Introduction
Unicellular and colonial microalgae form a very important component of freshwater phytoplankton.
These organisms have a critical role in primary production, nutrient cycling and food web in the aquatic ecosystem (1).Apart from that, microalgae in the aquatic ecosystems are indicators of changes that take place due to anthropogenic activities and environmental pollution.The diversity of these microalgae occurring as phytoplankton is influenced by various biotic and abiotic factors.Considering these aspects extensive studies have been carried out in India on the microalgae occurring as phytoplankton.These include seasonal distribution (2)(3)(4), diversity of fresh water algae (4,(5)(6)(7)(8)(9)(10)(11)(12)(13)(14) physico-chemical parameters of aquatic system (3,4,(15)(16)(17) and algal blooms (4,18).These works have been carried out with respect to both lotic and lentic ecosystems.
Studies concerning role of microalgae in water quality monitoring shows that changes in taxon diversity and composition indicate not only changes in physical and chemical variables of water, but also changes in biotic interactions (19).Ecological studies include seasonality of microalgal occurrence, natural variation of physico-chemical factors in water bodies and the relationship between occurrence and diversity of algal taxa and environmental factors (19,(20)(21)(22).To cope up with the changes, microalgal taxa have developed morphological and physiological adaptive strategies for surviving in different environments (19,23,24).Defined functional groups of microalgae with defined morphological and physiological traits were also studied in a given environment (19).It is also important to mention here that fresh water ecosystems are associated with human activity in the rural India.These are an important source of water for daily use and sustaining the livelihoods.The present study has been undertaken in some fresh water bodies, not studied earlier, in a town Chinsurah of district Hooghly of state of West Bengal.These fresh water bodies are located in the middle of the town and exposed to various pollutants.The prime objective of this study, on the occurrence and diversity of the microalgal component and their correlation with physico-chemical parameters, was to form a data bank of species diversity and growth condition for future prospect of using them in pollution monitoring.

(a) Study area:
The study area is located in Chinsurah town of Hooghly district, West Bengal, India.Five fresh water ponds were selected in this study area and their latitude and longitude were obtained from GPS data (Table 1).The five sites were marked as Site I, Site II, Site III, Site IV, Site V.The ponds were almost of the same size and located in area inhabited by people.They are used for household purposes and they received effluent through the drainage system (Photo plate 1).
(b) A random sampling method was used for the collection of samples from the selected sites over two different seasons-Late Summer and Winter.The samples were collected using a sampling bottle.
(c) After collection, three aliquots of sample each of 12 ml centrifuged and pellets were collected for species diversity study.Voucher sample was preserved in 4% formaldehyde for qualitative and quantitative analysis in laboratory.Microalgal species were identified on the basis of morphological features following published works (25)(26)(27)(28).10% glycerine was used as mounting material for microscopic study.

(d)
The organisms were photographed using Olympus CH20i.
(f) Temperature was noted by Eurolab ST9869B multi-thermometer, light intensity by LT Lutron LX-101 lux-meter, pH by Eutech instruments pH 700, for Electrical conductivity, Systronics conductivity meter 304 was used.
(g) For Inorganic phosphorous estimation, 50ml of water sample were taken and added 2 ml of ammonium molybdate followed by 5 drops of stannous chloride solution.After 5 min but before 12 min of the addition of the last reagent the absorbance was measured at 690 nm by UV-Visible spectrophotometer (UV-1800 Shimadzu).
(h) Dissolved oxygen was calculated by Winkler's Iodometric method.

(i)
Canonical Correspondence Analysis: Canonical Correspondence Analysis (CANOCO version 4.5; terBraak, 1986) was used to understand the correlation between physicochemical parameters of fresh water and fresh water microalgal component in late summer and winter seasons.Biplot scores of the environmental parameters were noted.CCA ordination was tested for significance with Monte Carlo test (500 runs) using CANOCO software.
(j) Venn diagrams were generated by using the tool http://www.interactivenn.netto represent the unique and common microalgal taxa in different sites in late summer and winter seasons.

Results and Discussion
The microalgal components of phytoplankton community, collected from five ponds in the study area, were diverse.A total number of 23 taxa, identified up to genus, were recorded.These microalgae belong to 4 different classes, namely Cyanophyceae, Chlorophyceae, Bacillariophyceae  1).A similar result was obtained earlier in another study (29).Relative abundance of microalgal taxa was high in winter season in four sites except site IV.Diversity of taxa has been found to be much more in winter as compared to late summer except Site IV (Fig. 2).Physico-chemical parameters were recorded both in winter and late summer.Table 3 shows value of physico-chemical parameters in all the sites and in both the seasons.There is significant variation with respect to light intensity and water temperature in these two seasons at all the sites.Though other parameters show very little variations, inorganic phosphorous has higher concentration (1.15mg/l and 0.85mg/l) in site III and site V, electrical conductivity shows high value (0.8ms/cm) in site II except in few cases.In late summer, value of light intensity and water temperature are higher than winter season.pH was more alkaline in winter season than late summer and winter except Site II.
Euglenophycean genera were present in Site III, Site IV and Site V in late summer and winter.In Site III both Euglenophycean genera, Euglena sp., Phacus sp., were present in winter season and also noticed that inorganic phosphorous concentration was high.Same result found in CCA analysis also.Euglenophycean genera plays a role as biological indicator of organic pollution (31).In Site I Synechocystis sp. was present in winter season and pH (8) was found to be high when compared to other sites.In Site II Tetrallantos sp. was present in winter season and the physico-chemical parameter, electrical conductivity value (0.8) was found to be high than other sites.Pediastrum sp. was found in high pH (7.5-8) sites I, sites III, site IV sites V and the same result was observed in CCA analysis.Chlorococcum sp., was present in winter season except in Site IV.Tetraedron sp., was present in both the seasons in Site IV.
Canonical correspondence analysis showed correlation between species (here species represents taxa) occurrence and environmental parameters both at late summer and winter.In the graph (Fig. 3 and 4) 'Δ' represents occurrence of species and ' ' represents environmental → variables.The result from the ordination diagram indicates that the relative weight, as denoted by extra fit values, for the environmental variables indicates a narrow range of variation (0.3359 -0.5126) with a p-value of 0.1160 in late summer sampling (Fig. 3) and (0.1612-0.3467) with a pvalue of 0.0580 in winter season, (Fig. 4) respectively.Therefore, result for winter season is statistically more reliable.The eigenvalues for corresponding axes indicate that the first axis is contributing the most in explaining the species environment relationship (0.602) in case of late summer and (0.396) in winter.The second axis is contributing more than remaining two axes in both seasons.In late summer, Axis 1 has the strongest positive correlation with light intensity and weakest positive correlation with water temperature.Axis 2 has the strongest positive correlation with dissolved oxygen and weakest positive correlation with light intensity.Axis 1 shows negative correlation with water pH.Axis 2 shows negative correlation with water   temperature, water pH and electrical conductivity, whereas, in winter Axis 1 has the strongest positive correlation with electrical conductivity.Axis 2 has the strongest positive correlation with inorganic phosphorous and weakest positive correlation with light intensity.Axis 1 shows negative correlation with light intensity, water temperature, water pH, dissolved oxygen, inorganic phosphorous.Axis 2 shows negative correlation with water temperature, water pH, dissolved oxygen.All the four eigenvalues reported above are canonical and correspond to axes that are constrained by the environmental variables in both seasons.In late summer (Fig. 3), Cosmarium sp. and Actinochloris sp.showed affinity with dissolved oxygen, inorganic phosphate and light intensity in axis 1. Schroederia sp., Scenedesmus sp., Merismopedia sp., Crucigenia sp., Phacus sp., Chodatella sp., Ankistrodesmus sp. are positive correlated with dissolved oxygen.In axis 2 the species have strong negative correlation with electrical conductivity.Chlorococcum sp., Tetraedron sp., Oocystis sp., Pinnularia sp., Navicula sp. are aline with electrical conductivity.
Following common microalgal taxa were observed in different sites (Fig. 5): one in Site II and Site IV (Chlorella sp.), three in Site III, IV and V (Scenedesmus sp., Schroederia sp., Merismopedia sp.).Another three found in Site III and IV (Tetrastrum sp., Closterium sp., Euglena sp.), two in Site I and IV (Navicula sp., Pinnularia sp.), one in Site I, IV and V (Pediastrum sp.) and another one in Site II, IV and V (Cosmarium sp.).
Following common microalgal taxa were recorded in different sites (Fig. 6): one in the following sites representing with different taxa: Site II, III and IV (Chlorella sp.), Site III and V (Euglena sp.), Site III and IV (Phacus sp.), Site I, III and IV (Navicula sp.), Site I, IV and V (Scenedesmus sp.), Site II, IV and V (Closterium sp.), Site I, II and V (Chlorococcum sp.), Site I, III and V (Crucigenia sp.), Site I and III (Pinnularia sp.), SITE I -V (Cosmarium sp.).Two common taxa were noted in Site I, III, IV and V (Pediastrum sp.Merismopedia sp.), Site IV and V (Selenastrum sp.,Tetraedron sp.), Site I, II, IV and V (Schroederia sp., Actinochloris sp.).
Moreover, these ponds are the sink of all domestic pollution because of through open drainage system from which brings the domestic waste of nearby settlements enters to the specific site.Shannon's diversity values depicted minor fluctuations with respective sites because there was no remarkable difference in the number of taxa composing dominant species at different sites.It has been observed (30,31) that incase of polluted areas, there was a decrease in biomass of the microalgae in phytoplankton associated with increase in species diversity.Diversity index is a good index for assessing and ranking the water quality.Analysis of diversity of microalgal flora of the lake leads to conclusion that the mean values did not differ from each other remarkably at the stations in different seasons studied, which can be directly related to the eutrophication process of aquatic system (32).Wetland ecosystems are the primary shelter for aquatic biodiversity including aquatic flora, fauna and other microorganisms.Phytoplankton is one of the important components of aquatic ecosystems (4).Fresh water ponds play an important role in the social ecology of the region in which they are located.Phytoplankton ecology plays an important role for indicating the eutrophication (33) as well as water quality and further way forward towards ecosystem management.
Major outcome: (i) The ponds located very near to the human residential complexes show a rich microalgal flora dominated by Chlorophycean members belonging to the order Chlorococcales and desmids (Desmidales).(ii) The microalgal genera are more abundant in winter season than in late summer.(iii) Two genera have been found to be season specific Synechocystis sp., Tetrallantos sp.

11 Photo plate 1 .Fig. 1 .
Photo plate 1. Location of study area and sampling sites

Fig. 2 .
Fig. 2. Occurrence of microalgal genera in late summer (LS) and winter (W) in five sites.

Fig. 3 .
Fig. 3. CCA ordination biplot based on PCA (vide table 4) showing the relative position of species-environment correlation between species occurrence and physico-chemical parameters in late summer season.

Fig. 4 .
Fig. 4. CCA ordination biplot based on PCA (vide table 5) showing the relative position of species-environment correlation between species occurrence and physico-chemical parameters in winter season.

14 Fig. 5 .
Fig. 5. Graphical presentation of all the microalgal taxa from the five study sites (Site I, II, III, IV & V) in late summer based on commonality of occurrence.

Fig. 6 .
Fig. 6.Graphical presentation of all the microalgal taxa from the five study sites (Site I, II, III, IV & V) in winter based on commonality of occurrence.

Table 1 .
List of sampling sites in Chinsurah, Hooghly the state of West Bengal India

Table 2 )
. Microphotographs of some of the organisms were given in Photo plate 2. Out of these genera occurrence of two genera belonging to Chlorophyceae is season specific -while Chodatella sp. was recorded in late summer, Tetrallantos sp.
Two Cyanophycean genera have been recorded namely Merismopedia sp. and Synechocystis sp.While Merismopedia sp. was recorded in both the seasons, Synechocystis sp. was recorded only in winter season.Two genera under Bacillariophyceae recorded were Navicula sp. and Pinnularia sp.Two genera of Euglenophyceae recorded were Euglena sp. and Phacus sp.

Table 4 .
CCA ordination score plot for late summer season based on first two principal components

Table 5 .
CCA ordination score plot for winter season based on first two principal components

Table 6 .
Diversity indices of microalgal class component in five sites under that investigation (L.S: Late summer, W: Winter)