Desalination 282 (2011) 95103

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Desalination
journal homepage: www.elsevier.com/locate/desal

Urban wastewater treatment using vermi-bioltration system

Priyanka Tomar, Surindra Suthar
School of Environment & Natural Resources, Doon University, Dehradun-248001, India

a r t i c l e

i n f o

a b s t r a c t
This work illustrates the potential of a novel vermi-bioltration system in treatment of urban wastewater. A small-scale vermi-bioltration reactor was constructed using vertical subsurface-ow constructed wetlands (VSFCWs) aided with local earthworms Perionyx sansibaricus. The coco-grass: Cyprus rotundus (density 0.14 plants/in. 2) was used to construct VSFCW. Another reactor without earthworms acted as experimental control. The wastewater was treated through this system for a total of eight repetitive cycles and after each cycle the changes in pH, electrical conductivity (EC), total dissolved solids (TDS), and total suspended solids (TSS), 3 chemical oxygen demand (COD), NO3 and PO4 of water were measured. Vermi-bioltration caused signif 3 icant decrease in level of TSS (88.6%), TDS (99.8%), COD (90%), NO3 (92.7%) and PO4 (98.3%). There were 3 about 38.8, 20.8, 80.6, 50.8 and 144.6% more removal of TSS, TDS, NO3 , PO4 and COD, respectively in vermi-bioltration than control. Results thus suggested that vermin-bioltration system is more efcient than VSFCW in terms of contamination removal efcacy. However, this work provides a preliminary idea of using earthworms in wastewater treatment system and further detailed studies are required on some key issues (e.g., loading rate, ow alternation impacts and earthworm stocking density) of this system. 2011 Elsevier B.V. All rights reserved.

Article history: Received 3 June 2011 Received in revised form 8 September 2011 Accepted 9 September 2011 Available online 13 October 2011 Keywords: Wastewater treatment Perionyx sansibaricus COD TDS Vermi-bioltration Marshy plants

1. Introduction The urban runoff in general, carries organic load along with several hazardous chemicals which not only spoils the aesthetic sense of the river but at the same time also degrades the aquatic ecosystem. Due to high establishment and running cost of a sewage treatment plant (STP) the majority of urban centers in developing world dispose urban runoff and sewerage water directly into urban river without any treatments or with partial treatments. Several mechanical and chemical approaches are being applied widely for urban wastewater treatments systems in urban centers mainly by sewage treatment plants (STPs). Apart to construction costs the operation and maintenance problems in STPs has raised the question of sustainability [1]. Moreover, excess sewage sludge produced by STPs has been subjected to increasingly stringent limitations on discharge during the last few decades [2]. According to Sinha et al. [3] many developing countries cannot afford the construction of STP and therefore; there is growing concern over developing some ecologically safe and economically viable small-scale wastewater treatment technologies for onsite wastewater treatment. However, at this crucial juncture some ecologically engineered tools can solve issues related with safe and cost-effective wastewater treatments technologies. The majority of present wastewater treatment systems are a disposal-based liner system and they should be transformed into cyclical treatments [4]

Corresponding author. Tel.: + 91 135 2255103. E-mail address: suthariitd@gmail.com (S. Suthar). 0011-9164/$ see front matter 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.desal.2011.09.007

in order to conserve the water and nutrient resources. An economical and manageable wastewater treatment approach is often required and deserves to be explored [5]. Biological wastewater treatment process involves the potentials of some living organisms to remove contaminants and sludge from wastewater in order to make it suitable for surface irrigation and other industrial use. Biological wastewater treatment involves the transformation of dissolved and suspended organic contaminants to biomass and evolved gases: CO2, CH4, N2 and SO2 [6]. A variety of organism like aquatic plants, marshland plants, protozoa, nematodes, oligochaetes have been tested in both laboratory and eld conditions to develop a low-cost bioreactor for wastewater treatment and sludge reduction. The potential of oligochaetes for wastewater treatment and sludge has been explored widely in many parts of the world. In general, Oligochaetes can be divided into two distinct groups, rstly, microdrilli (aquatic and small sized worms) and, secondly, terrestrial oligochaetes (earthworms) [7]. The aquatic oligochaetes can be divided into two groups: (i) the large aquatic worms (Tubicidae, Lumbriculidae and the semi-aquatic or terrestrial Enchytraeidae) and, (ii) the small aquatic worms such as Naidids and Aeolosomatids [8]. In recent years, both aquatic and terrestrial oligochaetes have been tested by several authors under lab-based trials to remove water contaminants and excess quantity of sludge [3, 6, 8-13]. The major components and outcomes of previous experiments on vermi-bioltration are described in Table 1. The utilization of earthworms in wastewater or sludge treatment is called vermi-bioltration. It was rst advocated by the Prof. Jose Toha at the University of Chile in 1992 [22]. Vermi-bioltration is a

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Table 1 Earlier studies indicating potential of oligochaetes in sludge stabilization and wastewater treatment. Type of sludge/wastewater Worm species used Major observations Sludge reduction was 77% during the process TSS reduced up to 99% after treatment Worm effectively removes major pollutant from sludge There was drastic impact on sludge (46.4% reduction in rst stage), COD (more than 80% removal) and BOD (more than 81%) during the process 2040% sludge converted into worm biomass and nitrate as well as nitrite removed efciently References Hendrickx et al. [11] Elissen et al. [14] Wei et al. [15] Song et al. [16]

Aquatic worm ((Tubicidae, Lumbriculidae, Naidids and Aeolosomatids) Waste sludge produced in wastewater Lumbriculus variegatus treatment plant Waste sludge produced in wastewater Lumbriculus variegatus treatment plant Efuent of activated sludge process Lumbriculida hoffmeisteri Sludge from wastewater water Branchnria Sowerbyi, Limnodrilns

Sewage of domestic sludge

Lumbriculus variegatus

Buys et al. [17]

Earthworms Domestic wastewater (assessment of toxicity of ammonia on earthworm in vermi-bioltration system) Liquid waste products from dairy industry

Eisenia fetida

E. fetida

Domestic wastewater sludge Secondary liquid efuents from Gelatine Industry Raw sewage Treatment of sewerage and sludge Domestic wastewater treatment

E. fetida Lumbricus rubellus E. fetida, Perionyx excavatus, Eudrilus euginae E. fetida E. fetida

High salt concentration may cause damage to earthworms in vermitration units Removal of 5 day BOD by 98%, COD by 8090%, TDS by 9092% during the process Signicant reduction in pollutant during vermistabilization process Decrease in COD by 90% and BOD by 89%. Removal of COD by 8090% and BOD by 90% during vermi-bioltration Removal of COD by 8186% and BOD by 9098% during vermi-bioltration Removal of COD by 5566% and BOD by 4765% during process

Hughes et al. [10]

Sinha et al. [3]

Wang et al. [5] Ghatnekar et al. [18] Sinha et al. [19] Xing et al. [20] Xing et al. [21]

process that adapts traditional vermicomposting system into a passive wastewater treatment process by using potentials of epigeic earthworms. According to Komarowski [23] in vermi-bioltration system suspended solids are trapped on top of the vermilter and processed by the earthworms and fed to the soil microbes immobilized in the vermilter. The dissolved and suspended organic and inorganic solids are trapped by adsorption and stabilization through complex biodegradation processes that take place in the living soil inhabited by earthworm and the aerobic microbes. Intensication of soil processes and aeration by earthworms enable the soil stabilization and ltration system become effective and smaller in size [19]. In general, inoculated earthworms in vermibeds accumulate many organic pollutants from the surrounding soil environment, passive absorption through the body wall and also intestinal uptake during the passage of soil through the gut [24]. The efcacy of vermi-bioltration system is already described in literature (Table 1). Sinha and his group investigated the potential of vermi-bioltration system in treatment of dairy industry efuent [3]. They claimed that earthworms have been found to remove the 5 day BOD by over 98%, COD by 8090%, TDS by 9092% from any liquid wastes by the general mechanism of ingestion and biodegradation of organic wastes. According to a study conducted by Ghatnekar et al. [18] suggested that the vermi-bioltration system is efcient to remove COD and BOD load of wastewater generated from gelatin industry. They applied a three-tier biotechnology unit coupled with vermibioltration system to convert secondary liquid efuents from a gelatin manufacturing unit into bio-safe clean water. Results thus, suggested a signicant decrease in COD by 90% and BOD by 89%. Recently, Zhao et al. [2] studied the stabilization of domestic waste water sludge using earthworms and results have revealed that the presence of earthworms in the vermibeds to the signicant stabilization of the sludge. The volatile suspended solids (VSS) reduction in the vermibeds was in the ranges of 56.266.6% in different treatment units aided with earthworms. Similarly, Sinha et al. [19] developed a low-cost sustainable

technology over conventional systems to recycle the domestic wastewater with potential for decentralization facility for waste management. They claimed removal of 5 days' BOD (BOD5) by over 90%, COD by 8090%, total dissolved solids (TDS) by 9092%, and the total suspended solids (TSS) by 9095% from urban wastewater after the treatment with worms. The microbes play an important role in vermibioltration system and they also provide some extracellular enzymes to facilitate the earthworms for rapid degradation of organic substances in vermibeds [25]. Likewise, Zhao et al. [2] investigated the interactions between microorganism and earthworm in vermi-bioltration system. They demonstrated that earthworm biolm was dominated by the members of the phylum Proteobacteria and Pseudomonas sp. The majority of previous studies are available on either utilization of vermi-bioltration or only constructed wetland ltration system for removal of nutrients/pollutants from wastewaters, but no comprehensive report is available on utilizing potentials of both systems to develop an effective integrated system, comprising of earthworm and construction wetland system, for wastewater treatment. Although, Chiarawatchai [26] has conducted an interesting study on combining vertical sub-surface ow constructed wetlands (VSFCWs) with earthworm. The integration of these two ecological techniques (traditional wetlands system with vermi-bioltration mechanism) can be a cost effective and sustainable option for onsite wastewater treatment. The aim of this study was to assess the potential of an integrated vermi-bioltration system with VSFCWs constructed by using earthworm Perionyx sansibaricus and a wetland weed Cyprus rotundus (coco-grass or red nut sedge) under a small-scale laboratory experiment. C. rotundus is one of the most invasive weeds and have been reported from tropical and temperate regions of the world. It is a perennial plant and mainly occurs in gardens, agriculture plots, around stagnate water bodies etc. Few earlier studies have demonstrated the capabilities of C. rotundus in wastewater treatment and phytoremediation [2729].

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C. rotundus was selected for vermi-bioltration system due its local and perennial availability, short length (up to 40 cm) and easy cultivation capabilities. 2. Materials and methods 2.1. Earthworm, plant collection and wastewater collection Individuals of earthworm Perionyx sansibaricus of different age group were collected from mud of a gray water drain in university campus, Dehradun, India. The stock of P. sansibaricus was cultured in laboratory using garden soil spiked with leaf litter and cow dung inappropriate ratios. Plant Cyprus rotundus used for construction of bioltration unit was originally obtained from moist soils around grey water drains in university campus. The identication was made using standard taxonomic key and conrmed by plant taxonomist in university. The urban wastewater was collected from a wastewater stream owing over nearby location of university campus. The wastewater was collected from main streamline of wastewater drain in largesize pre-cleaned circular plastic containers of 20 L capacity. Collected wastewater was brought immediately to laboratory and collected in large-size wastewater reservoir unit of the vermi-bioltration system. Wastewater was collected just before the starting of experimentation in order to avoid alternation in the wastewater characteristics mainly due to open storage of sample. Other accessories like aeration pump, ow control units, water pumps etc. were procured from a local sanitary engineering shop and scientic equipment supply rms. 2.2. Construction of vermi-bioltration and bioltration units The experimental vermi-bioltration /bioltration units were comprised of two reactors/batches: (i) long cylindrical unit: Reactor-I and (ii) rectangular unit: Reactor-II. Long unit was constructed with a traditional water ltration system using gravels and sand column at the base and a bioltration system at the top made of living individuals of wetland plant stand, i.e. Cyprus rotundus and earthworms in its root zone system. An aeration unit was also xed in the middle layer of the Reactor-I. A plastic circular cylinder of 80 L capacity was used to construct Reactor-I of vermi-bioltration/bioltration unit. Following materials/layers were used to ll (from bottom to top) the circular cylinder to construct the vermi-bioltration/bioltration unit: Layer I Large stones (1015 cm in diameter) up to 5-inch this layer creates a kind of air chamber system and for water storage in base of system. Layer II Thick layer of small stones and gravel (57 cm diameter) up to 2-inch acts as ltration unit and creates a kind of turbulence during water ow and provides space for aeration of wastewater. Aeration pipe (pierced 1 inch diameter and 15-inch length). Aeration pipe was covered with 1 inch layer of small pebbles. A ne plastic net was placed over the pebble layer aeration device was installed in order to remove BOD load of the wastewater. Layer III A thick layer of sawdust spread over the net (2-inch) saw dust acts as good absorbent for several kinds of inorganic pollutants of wastewater. Layer IV Dried leaves of Sal tree were placed over sawdust layer (2-inch) as natural adsorbent to remove nutrients from wastewater. It also acts as feed for microbial communities helping in wastewater mineralization. Fine net The ne plastic net (b0.5 mm pore-size) was placed over the leaf litter layer in order to check the entry of earthworm in deep layers of the vermi-bioltration system in order to avoid moving earthworm to deep bottom layers of the reactor.

Layer V Vermi-bioltration bed mainly constructed using thick bedding of soil mixed with small stones and pebbles along with complex root-zone system of surface plant Cyprus rotundus. The thickness of this layer was about 10 in. earthworm acts as biological agent to remove solid fractions of wastewater and mineralization of wastewater mainly driven by earthworm-microbe interactions in root-zone system. Layer VI Composed of surface vegetation stand of Cyprus. It was about 4 6 in. in length wetland plant provides air in root-zone system and removes nutrients from wastewater through general absorption, adsorption and translocation processes. Also provides shelters to benecial microbial communities responsible for N mineralization. The detail of vermi-bioltration/bioltration unit is given in Fig. 1. In the top layer of Reactor-I, i.e. Layer-V the fresh and viable specimens of Cyprus were planted in top soil layers. The roots of plant were planted deeply and surface layer was irrigated regularly (for one week) by tap water in order to x the planted Cyprus in top layer of vermireactor. The mean density of Cyprus in vermireactor-I was about19 plants/in.2. The open space between plant stand was lled with a thin layer of small stone to avoid direct hydraulic impact on the plant and earthworm. In this vermi-bioltration system efforts were made to create a kind of soil ecological system mainly comprised of thick soil layer spiked with complex rooting system of Cyprus rotundus. The pieces of stones and pebbles in this root-zone-ltration system create an appropriate space for air and inoculated earthworm in sub-soil system. Nonetheless, the root-zone system not only enhances the efciency of wastewater ltration but at the same time also provide shelters to bacterial communities (e.g., N-xers, ammonifying and denitrication bacteria) responsible for nutrient removal from wastewater. Reactor-II: Another unit of reactor, i.e. Reactor-II was introduced in order to enhance the removal efciency of the system. In Reactor-II the biological component of the ltration unit was of more importance therefore the majority of the reactor volume was lled with earthworm and plant root zone layers. A rectangular plastic container of size (23. 5-inch length 18-inch width 15-inch depth) was used to construct the second unit (Reactor-II) of vermin-ltration system. In Reactor-II there were two district layers: rstly, base layer (large pebbles; 1015 cm in diameter and height about 6-inches), and secondly, top layer (small pebbles; 57 cm in diameter mixed with ne sand and height up to 10-inches). The top layer acts as bedding substrate for earthworms in Reactor-II. A thin plastic net sheet was placed between the both layers to avoid movement of earthworms from top layers to base layer of the vermireactor. Fresh and viable specimens of plant: Cyprus rotundus were planted in top layer and thin patches of small stones were placed over the open spaces around Cyprus plant to avoid direct hydraulic impact of inow water in Reactor-II on plant stand and earthworms. The mean plant density in vermireactor was 0.14 plants/in. 2 (calculated using values of total surface area of Reactor-II and plant numbers in reactors). After plantation Cyprus stand was allowed to grow for one week and during this period adequate amount of tap water was supplied in vermireactor to facilitate the xing of roots of plant in top layers of Reactor-II. Initially, both reactors were run for twothree days using fresh tap water to wash and x the layers of vermibeds in proper functioning forms. After establishment of plant stands (after one week) the reactor was run for wastewater treatment experimentations. In both experimental vermi-bioltration systems, i.e. Reactor-I and Reactor-II individuals of earthworm P. sansibaricus were introduced over the top layer the reactors. Small passages were made in the surface layers of both reactors in order to facilitate worms to enter in the top soil layers of the vermireactor. The initial earthworm density in both vermi-bioltration systems was measured in the ranges of 22.024.5 g/L. The earthworms were allowed to settle in vermireactors for initial 23 days and thereafter, vermireactor was run for

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Water Sprinkling Device

Supply of wastewater from tank to reactor-I through pump Layer -VI

Layer - V Layer - IV Layer -III

Aeration pipe

Layer - II

Layer - I Aerator Wastewater Tank Outlet from Reactor-I Reactor-I

Layer -III

Layer -II

Layer -I Reactor -II Outlet (completion of cycle

Fig. 1. Vermi-bioltration system used for wastewater treatment.

experimentation. A reactor without earthworm (bioreactor) acted as experimental control for this study. 2.3. Observation and data collection The wastewater was used without any dilation for this experimentation. However, prior to putting wastewater in experimentation cycle a sample of wastewater (about 1 L) was separated from stock and analyzed for its physic-chemical characteristics (Table 2). As illustrated in Fig. 1, during experimentation cycle the stock wastewater was supplied in Reactor-I through a mechanical pump and a ow control device was also xed in main water-supply pipe. The wastewater was sprinkled over the surface of top layer of Reactor-I through a perforated plastic pipe and outlet of Reactor-I was closed to ll the reactor with wastewater. The wastewater was lled in reactor continuously up to the saturation level

of top layer, i.e. layer-V. The care was taken to avoid the overowing of water. The wastewater was retained for 1 h in Reactor -I and a continuous air was supplied during this period using an electronic aeration device. After that the outlet of Reactor-I was opened into Reactor-II and
Table 2 Characteristics of wastewater used for experimentations. Parameters pH EC (S/cm) TSS (mg/L) TDS (mg/L) NO3 (mg/L) 3 PO4 (mg/L) COD (mg/L) Range 7.61 0.10 922.0 5.29 216.67 7.64 56813.3 51.3 384.2 1.00 36.37 0.67 863.0 3.60

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ow of outlet was controlled using a mechanical ow control device. The outlet water from Reactor-I was sprinkled over the surface of Reactor-II using a perforated pipe system. In second cycle of treatment (i.e., treatment in Reactor-II) the water retention mechanism and time framework was same as used in Reactor-I. After that the outlet of Reactor-II was opened to release the water from second reactor, i.e. Reactor-II. The complete passing of water from both reactors was counted as one treatment cycle and water after each cycle was putted back into new cycle. The wastewater was repeatedly passed through both units of vermiltration system for complete 8 cycles. An interval (stabilization period) of 24 h was kept between two subsequent treatment cycles in order to stabilize the microbial environment and earthworm population in sub-surface of the vermireactor after each cycle. A sample of wastewater was collected in pre-cleaned and sterilized polythene bottle of 1 L capacity from outlet of Reactor-II after each treatment cycle and stored at 4 C for further investigations on changes in physico-chemical characteristics of wastewater during each cycle. 2.4. Chemical analysis The chemical characteristics of wastewater samples collected after each treatment recycle were analyzed for different physic-chemical parameters by following methods as described by APHA-AWWA-WPFC [30]. pH was measured using digital pH meter (Metrohm, Swiss-made). Conductivity was measured using digital conductivity meter (Remi, India). Total dissolved solids (TDS) and total suspended solids (TSS) was measured was measured ltration and gravimetric and oven drying methods. Chemical oxygen demand (COD) was measured using potassium dichromate oxidation method. Nitrate, sulphate and phosphate contents in water were analysed spectrophometrically by following methods as described by APHA-AWWA-WPFC [30]. 2.5. Statistical analysis A paired sample t-test between control (without earthworm) and experimental (with earthworm) vermi-bioltration unit was performed for each chemical parameter to analyze the differences. One-way analysis of variance (ANOVA) was also preformed to measure the difference among different cycles for each physic-chemical parameter of wastewater. SPSS statistical package (Window Version 13.0) was used for data analysis. All statements reported in this study are at the p b 0.05 levels. 3. Results and discussion The quality of wastewater in terms of phyico-chemical characteristics is described in Table 2. The collected sample of urban wastewater showed relatively high values of some key pollution indicating 2 parameters of water: TDS (50813 mg/L), NO3 (384.2 mg/L), PO4 2 (36.37 mg/L), SO4 (293.3 mg/L) and COD (863.3 mg/L). The wastewater after vermi-bioltration process showed a drastic change in its major physico-chemical parameters, after each treatment cycle. Although, there was signicant reduction in key pollutants of urban water in both bioltration (without earthworm) and vermi-bioltration (with earthworm), but difference was more prominent in water from vermi-bioltration unit than initial levels. The changes in all reactors could be attributed to the development of biological communities within reactors [26]. 3.1. pH The change in pH during different treatment cycle is illustrated in Fig. 2. In bioltration system (control) a trend of slight increment in pH was observed till last observation. On the other hand, in vermibioltration system pH decreased sharply up to 3 rd cycle of treatment thereafter, a trend of gradual increment was observed up to

8.4 8.2 8 7.8 7.6 7.4 7.2 7 6.8 0 1 2 3

pH

Experiment Control

Treatment cycle
Fig. 2. Changes in pH during different cycles of treatment in control and experimental reactor.

7th cycle followed by pH stabilization state during last treatment cycle. pH of water mainly depends upon a variety of chemical factors, e.g., dissolved gases, organic acids, humic fractions and inorganic salts. The decomposition of organic fractions of wastewater, mainly by microbes in water, produces some acidic species of mineralized or ganic materials (CO2, ammonia, NO3 and organic acids) which plays an important role in shifting of pH scale of treated water. Probably the reduction in the level of ammonia, NO3 during bioltration and vermi-bioltration treatment caused sight changes in pH. There was statistically signicant difference between bioltration and vermibioltration process for pH level (t-test: p b 0.05) of efuent from reactors. The pH value of efuent obtained at the end of treatment process was 7.81 in bioltration (about 9.1% more than initial) and 8.15 in vermi-bioltration (about 13. 8% more than initial) reactor. The different between control and experimental reactor for pH could be related to earthworm mediated rapid mineralization of organic fractions of wastewater. Also few earlier researchers have reported increase in pH after vermi-bioltration processes [19, 21]. 3.2. Electrical conductivity (EC) Electrical conductivity (EC) of wastewater showed signicant changes after treatment through ltration system in both bioltration and vermibioltration processes. The conductivity of treated water was: 1230.0 S/cm in bioltration and 984.7 S/cm in vermi-bioltration system. The difference between control and experimental reactor was statistically signicant (t-test: p =0.002). The changes in EC during different cycles are described in Fig. 3. In bioltration system EC showed a linear trend of increase till last observation while in vermi-bioltration reactor EC showed different patterns of uctuation during experimental processes. In vermi-bioltration system EC value of efuent showed increasing pattern up to 5th cycle of treatment thereafter, it reduced sharply till last observation. The increasing EC could be attributed to mineralization of organic waste fractions of wastewater through microbial and
1400 1300 1200

EC

Experiment Control

S/cm

1100 1000 900 800 700 600 0 1 2 3 4 5 6 7 8

Treatment cycle
Fig. 3. Changes in EC during different cycles of treatment in control and experimental reactor.

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earthworm activities in reactors. The higher EC of efuent from verminbioltration than experimental control was possibly due to high mineralization processes driven by inoculated worm community in reactor [31]. In general, when organic waste transits through earthworm gut some fraction of it is then converted into plant available forms [25]. Probably the release of different mineral ions, such as phosphate, ammonium and potassium results in increased EC of substrate. The EC reects the salinity of any material and it is a good indicator of the mineralize fraction of wastewater. However, after 56 cycles of treatment EC of efuent water from vermi-bioltration reactor showed sharp decrement till last observation. This could be due to adsorption and/or absorption of inorganic constituents of water by different biological or non-biological components [32] of vermi-bioltration system. Moreover, accumulation of salts by inoculated worms, during later ltration in vermi-bioltration process, might be responsible for reduction in EC. 3.3. COD COD is an important indicator of organic load of urban wastewater. The COD load in efuents from bioltration and vermi-bioltration system was signicantly low than initial levels, but vermi-bioltration showed more removal efciency than bioltration reactor (t-test: p b 0.001). The removal rate was 90% (as compared to initial level) in vermi-bioltration system and 36.8% in bioltartion system. This could be because earthworms and aerobic microbes act symbiotically to accelerate and enhance the decomposition of organic matter [33]. Results clearly indicated the potential of worms in removal of organic load from wastewater through direct feeding of solid fractions of water or by promoting microbial-mediated organic decomposition process. The COD removal rate was 84% in vermi-bioltration system after completion of 8 cycles of treatment. The changed in COD load of wastewater during different treatment cycles is illustrated in Fig. 4. In control bioltration system COD reduced gradually during treatment cycles while in vermi-bioltration system COD level of efuent water reduced rapidly after 1st treatment cycle (Fig. 4). Earlier worker have also reported signicant reduction in the COD load during bioltration and vermibioltration processes [3, 21]. Sinha et al. [3] studied the vermiltration of wastewater originated from dairy industry under a pilot-scale project. They claimed the average COD reduction in the ranges of 8090% at the end. Under another laboratory trial of urban wastewater treatment through vermiltration, Sinha and his associates reported about 45% reduction in COD load after treatment [19] and removal rate was signicantly high in experimental reactor than control one (without worms). In general, the geological and microbial system in control bioltration unit is responsible for COD reduction while in vermi-bioltration system enzymes, secreted by earthworm and gut-associated microora, reduce the those chemicals which otherwise cannot be decomposed by microbes [19, 25]. Recently Xing et al. [21] have reported signicant COD

reduction (47 58% than initial) during vermi-bioltration of domestic wastewater. Also Ghatnekar et al. [18] have investigated the impact of vermi-bioltration system on chemical characteristics of wastewater generated from gelatin industry. They claimed about 90% reduction in level of COD at the end of process. Wang et al. [5] also reported 90.2% average removal efciencies of vermi-bioltration system for COD of a domestic wastewater. The microbial association with worms in vermiltration system could be important for removal of organic load form wastewater. The presence of earthworm also promotes the microbial colonization in vermibeds and evidences from recent investigation supports this hypothesis [2, 34]. Zhao et al. [2] investigated the earthworm-microorganism interaction during wastewater sludge treatments and results suggested about 46% reduction in the contents of volatile suspended solids due to earthworm-microbial action after treatment process. The easy assimilable source of carbon and other available nutrients from earthworm products, i.e. casts and mucus accelerates the microbial colonization in earthworm-containing vermibeds. According to Singleton et al. [35] earthworm hosts millions of decomposer microbes in their gut and excreate them in soil along with nutrients in worm casts. Such nutrients further enhance the microbial quality and quality of the vermibeds. Moreover the formation of biolms of decomposer microbes in the geological system of the vermireactor also promotes COD reduction during vermiltration process [3]. Chiarawatchai and Nuengjamnong [36] and Chiarawatchai [26] suggested that earthworms contributed to the wastewater remediation during the treatment process within the VSFCWs. Results thus, clearly indicate that vermi-bioltration may be an efcient treatment tool for designing of a low-cost domestic wastewater treatment facility.
3.4. NO3 N

Nitrate is an important indicator of water pollution and its high concentration in freshwater bodies leads to eutrophication problem. In this study there was signicant impact on nitrate concentration in efuents after treatment in both experimental reactors. But NO3 reduction arte was prominent in vermi-bioltration unit than biol tration system (Table 3). The level of NO3 in efuent after nal treatment cycle was 27.9 mg/L for vermi-bioltration unit and 186.9 mg/L for bioltration unit. There was about 92.7% total removal of NO3 N in vermi-bioltration unit that was signicantly higher than total removal in bioltration unit (51.3% removal) (t-test: p b 0.001). In traditional wetland bioltration system the nutrients and metals may be removed from the polluted water and retained in the sediment and taken up by the plants and by microorganisms associated on the surface of the roots and sediments, by immobilization in sediments via., mechanism such as adsorption on ion exchange sites binding to organic matter, incorporation into lattice structure and precipitates into insoluble compounds [37]. The results of removal efcacy of bioltration system were similar to those observed in constructed wetlands by other researchers [28, 32] and were moderately higher than those detected in horizontal ow constructed wetlands planted
Table 3 Chemical characteristics of outlet from bioltration (control reactor) and vermi-bioltration (experimental) at the end of process. Parameters pH EC (S/cm) TSS (mg/l) TDS (mg/l) NO3 (mg/l) 3 PO4 (mg/l) COD (mg/l)
a

Control reactor a 7.81 0.01 1230.0 8.0 78.40 1.00 9875.0 15.0 186.90 1.02 12.67 0.03 545.6 5.0

Experimental reactor b 8.15 0.01 984.67 11.7 24.8 2.82 91.3 5.13 27.92 2.93 0.62 0.04 86.67 2.03

t-test c (t-coefcient value) 51.50 22.86 1179.71 26.113 69.55 374.86 160.93

Signicance level P b 0.001 P = 0.002 P b 0.001 P b 0.001 P b 0.001 P b 0.001 P b 0.001

Fig. 4. Changes in COD during different cycles of treatment in control and experimental reactor.

b c

Reactor without earthworms (vermi-bioltration). Reactor with earthworms (bioltration). Paired sample t-test between control and experiment.

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with Phragmites by Vymazal [38] and with Canna and Heliconia by Konnerup et al. [39]. The high NO3 N removal efciency of current bioltration system than previous reports could be explained in terms of oxygen supply in rhizosphere of bioltration system. In general, oxygen is released by roots of plants in constructed wetlands and it should be sufcient to meet the demand for the aerobic bacterial communities which are mainly responsible for NO3 N removal from wastewater. Therefore, addition of aeration device in current bioltration system was an advantage over traditional bioltration or wetland ltration systems. Vymazal [38] also pointed out that in most system designed for the treatment of domestic or municipal sewage the supply of dissolved organic matter is sufcient and aerobic degradation is limited by oxygen availability. The supply of oxygen also promotes the activities of heterotrophic and ammonifying bacteria which are mainly responsible for nitrate removal from wastewater. In general, denitrication processes involved the initial NO3N reduction to NO2N, followed by further reduction to nitric oxide (NO), nitrous oxide (N2O) and nally to molecular nitrogen (N2) [40]. However, NO3 N reduction rate was relatively high in this study than previous reports and probably that attributed to oxygen supply in the system. Wang et al. [5] reported efcient removal of + NH4 N (with 85.797.1% of removal rate) while studying wastewater treatment using an earthworm-based ecological lter integrated constructed rapid inltration (Eco-CRI) system. Xing et al. [21] reported about 7.63 to 14.9% total N and 21.0 to 62.3% NH4N removal in wastewater after treating through vermi-bioltration system. The different between bioltration system and vermiltration system for removal rate should be explained in terms of the population and activities of nitrogen metabolizing bacteria. Chiarawatchai [26] reported signicant reduction in level of nitrate in efuents obtained from a labscale microcosm wastewater treatment unit than efuent from reactor without worms. The presence of earthworm in rhizosphere sub-system has some advantages over traditional bioltration system because of the direct impact of earthworms on aerobic heterotrophic bacterial communities which are mainly responsible for N-mineralization in wastewater bioltration systems. The trend of changing NO3 -N level during the treatment cycles is described in Fig. 5. There was rapid removal in vermi-bioltration unit than the bioltration unit in the rst and second cycle of the treatments and that could be due to ltration of suspended substances during rst cycle of treatment which are considered to be feed materials for earthworms in vermi-bioltration system. Moreover, earthworm-mediated rapid nitrogen transformation leads to rapid NO3 -N loss from wastewater.
3 3.5. Phosphate (PO4 )

in wastewater is household drains and urban runoff water containing excreta and other organic substances [21]. The nal efuent from vermi3 bioltration system showed low concentration of PO4 (0.62 mg/L) than sample collected from nal stage of bioltration system (12.67 mg/L). 3 The PO4 removal efciency of vermi-bioltration was recorded 98.3% and that was signicantly higher than removal efciency of bioltration 3 system (65.2%) (t-test: pb 0.001). The patterns of PO4 removal during different treatment cycles is described in Fig. 6. It is clear that in bioltra3 tion system the removal trend for PO4 is slow and linear but in vermi3 bioltration reactor there was a trend of sharp PO4 removal up to 56 cycles of treatments thereafter; removal rates declined sharply. The ligand exchange reactions and physical adsorption or sorption sites rapidly removes phosphorous from the soil solution. In soil column the hydroxides and oxides of Al and Fe, calcium carbonate and layer silicate minerals are important sites for sorption of phosphate anions [41, 42]. 3 The PO4 removal efciency of current bioltration system with Cyprus stand was relatively higher than those detected in horizontal ow constructed wetlands planted with Phragmites by Vymazal [38]. In current vermitration system the top layer composed of sandy soils along with mixtures of large stones and pebbles. Probably the sand mixed column of current bioltration reactor was advantage over the previous bioltration systems. Preetha and Kumar [43] demonstrated more 3 than 99% removal of PO4 from wastewater using sand-column treatments device. According to Bostrom et al. [44] aerobic conditions are more favourable for P sorption and co-precipitation therefore; it is sug3 gested that high PO4 removal could be due to addition of aeration device in our vermi-bioltration system. However, results of phosphorus removal contrasts with nding of Chiarawatchai [26] who reported least impact of earthworm inoculation on phosphorous removal from wastewater during vermi-bioltration process. He suggested some technical improvements like replacement of substrates from gravel or sand to ones with high phosphorus adsorption capacities to enhance phosphorous removal capability of vermi-bioltration unit. However, better results of this study than previous report could be attributed to substrate quality, design and biological components (earthworm species, plant type etc.). Moreover, activities of earthworm and associated microora in vermibeds also promote rapid P-mineralization in the sys3 tem. The level of PO4 in treated efuent from bioltration and vermibioltration is of prime concern because high concentration of such substance is responsible for eutrophication in surface freshwater re3 sources. Although, the level of PO4 in nal efuent from bioltration system was comparatively high than prescribed limit, i.e. 5.0 mg/L as decided by national pollution monitoring agency, i.e. Central Pollution Control Board (CPCB) for surface discharges of treated water. 3.6. TSS and TDS

As described in Table 3, there was signicant different between 3 inlet and outlet water for PO4 concentration in both treatment reactors, i.e. bioltration and vermi-bioltration. The source of phosphate

Total suspended solids (TSS) and total dissolved solids (TDS) showed drastic reduction during bioltration and vermi-bioltration

Fig. 5. Changes in NO3 during different cycles of treatment in control and experimental reactor.

3 Fig. 6. Changes in PO4 during different cycles of treatment in control and experimental reactor.

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Fig. 7. Changes in TSS during different cycles of treatment in control and experimental reactor.

Fig. 9. Changes in different parameters of wastewater during different cycles of treatment in control and experimental reactor.

process (Fig. 7 and 8, respectively). The total reduction in TDS content was about 99.8% in vermi-bioltration unit and that was signicantly higher than total removal in bioltration system, i.e. 82.6% (t-test: p b 0.001). Results thus clearly suggested the capability of earthworms to remove solid fractions of wastewater during vermi-bioltration processes. Similarly TSS also reduced signicantly in wastewater obtained from both experimental reactors (bioltration and vermibioltration) at the end of process. The removal rate was high in vermi-bioltration unit (88.6% than initial level) than bioltration system (63.8%) (t-test: p b 0.001) (Table 3). The removal pattern of TDS in both ltration systems is described in Fig. 7. The control (bioltration) system showed a gradual removal of TDS during different cycles of treatments process while in vermi-bioltartion system TDS removed sharply during initial 34 cycles thereafter; the removal process was more or less steady till last observation. The difference between both systems could be due to difference in biological components and working capabilities of both reactors. According to Cooper et al. [45] and Vymazal et al. [46] the suspended solids that are not removed in pre-treatment system are effectively removed by ltration and settlement processes. Mustafa et al. [47] reported the potential of integrated constructed wetland system with Typha latifolia, Carex riparia, Glyceria maxima, Philarius arundiraecae and Juncus effuses in wastewater treatments. They have reported signicant removal of TSS, i.e. 93.7% after treatment process. According to a study conducted by Prabu and Udayasoorian [28] Phragmitis australis, Cyperus pangorei and Typha latifolia planted bioltration system removed about 77, 72 and 67%, respectively TSS from wastewater after treatments. The efcacy of vermi-bioltration system in TDS and TSS removal is also reported by earlier authors. Sinha et al. [3] reported total removal of TSS and TDS in the ranges of 9092% and 9095%, respectively. They have attributed the TSS removal to continuous consumption by earthworms. Xing et al. [21] demonstrated the results of a small-scale vermiltration unit for

domestic wastewater treatment. According to this study earthworm presence in treatment system caused about 57 to 79% reduction in total content of suspended solids in wastewater. The results of present study corroborates with the ndings of other scientists who claimed importance of earthworm in vermiltration system. However, more detailed is needed to establish the relationship between removal of solids and earthworm working mechanism in vermi-bioltration system. 4. Conclusions This work provides an opportunity to explore the efciency of a vermi-bioltration system (mainly constructed by using a wetland weed Cyprus rotundus and live biomass of a local earthworm P. sansibaricus) in treatment of urban wastewater. Earlier scientic approaches were based upon the use of either plant or earthworm in bioltration unit design. But in this study the integration of these two components (traditional constructed wetlands system and earthworms has been applied to design a cost effective and sustainable option for onsite wastewater treatment. Results clearly suggested that integrated vermi-bioltration reactor was more efcient than traditional bioltration system in terms of removal of key chemical pollutant from wastewater (Fig. 9). Although, results clearly indicates the efcacy of vermibioltration system in wastewater treatment but further detailed studies are still required to answer few key issues of this system, e.g. hydrolic load, retention time impact, microbial ecology in vermibeds, earthworm-microbial interaction etc. Acknowledgement We would like to thank four anonymous reviewers for critical comments and fruitful suggestions on earlier version of the manuscript. The kind cooperation of laboratory staffs (Mr. Ganesh Bahuguna, Namita Tiwari, Kamal and Digpal Negi) during experimentation is also acknowledged here. References
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Fig. 8. Changes in TDS during different cycles of treatment in control and experimental reactor.

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