diversity of bacterial isolates during full scale rotary drum composting

Waste Management xxx (2013) xxx–xxx

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Waste Management

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Diversity of bacterial isolates during full scale rotary drum composting

Akansha Bhatia a,⇑, Sangeeta Madan b, Jitendra Sahoo c, Muntjeer Ali a, Ranjana Pathania c,Absar Ahmed Kazmi a

a Department of Civil Engineering, Indian Institute of Technology Roorkee (IITR), Roorkee 247 667, Indiab Department of Zoology and Environmental Sciences, G.K.V. Haridwar, Uttarakhand 249 404, Indiac Department of Biotechnology, Indian Institute of Technology Roorkee (IITR), Roorkee 247 667, India

a r t i c l e i n f o a b s t r a c t

Article history:Received 27 September 2012Accepted 29 March 2013Available online xxxx

Keywords:Rotary Drum16S rRNACulture- dependentMolecular analysisComposting

0956-053X/$ - see front matter � 2013 Elsevier Ltd. Ahttp://dx.doi.org/10.1016/j.wasman.2013.03.019

⇑ Corresponding author. Tel.: +91 9458108008.E-mail address: [email protected] (A. Bhati

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Bacterial diversity of full scale rotary drum composter from biodegradable organic waste samples wereanalyzed through two different approaches, i.e., Culture dependent and independent techniques. Cul-ture-dependent enumerations for indigenous population of bacterial isolates mainly total heterotrophicbacteria (Bacillus species, Pseudomonas species and Enterobacter species), Fecal Coliforms, Fecal Streptococci,Escherichia coli, Salmonella species and Shigella species showed reduction during the composting period.On the other hand, Culture-independent method using PCR amplification of specific 16S rRNA sequencesidentified the presence of Acinetobacter species, Actinobacteria species, Bacillus species, Clostridium species,Hydrogenophaga species, Butyrivibrio species, Pedobacter species, Empedobactor species and Flavobacteriumspecies by sequences clustering in the phylogenetic tree. Furthermore, correlating physico-chemical anal-ysis of samples with bacterial diversity revealed the bacterial communities have undergone changes, pos-sibly linked to the variations in temperature and availability of new metabolic substrates whiledecomposing organics at different stages of composting.

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1. Introduction particular, the appearance of certain microbes will reflect the qual-

Composting is an aerobic process, during which organic waste isbiologically degraded by micro-organisms to humus-like material.The end product should not contain pathogens or viable seeds andit should be stable and suitable for use as a soil amendment (Ep-stein, 1997). Many factors such as oxygen content, moisture, com-position of the feed, pH, and temperature, affect the compostingprocess and ultimately the end product. Furthermore, theseparameters are strongly connected to microbial communities,which are critical for the degradation of organic substrates suchas hemicellulose, cellulose, and lignin.

The initial phase of the composting process is characterized bythe activity and growth of mesophilic microbes, which in turnleads to a rapid increase in temperature. At the next stage, thermo-philic microbes become responsible for the degradation process.The final phase, which includes both cooling-down and maturingstages, is characterized by the development of a new mesophiliccommunity (Finstein and Morris, 1975; Ishii et al., 2000). There-fore, the optimization of compost quality is directly linked to thecomposition and succession of the microbial community in thecomposting process. Thus, the monitoring of a succession of micro-bial communities is important for effective management of thecomposting process, as microbes play key roles in this process; in

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a).

al. Diversity of bacterial isolat

ity of the maturing compost (Ryckeboer et al., 2003).Several protocols have been employed to date for the investiga-

tion of microbes in the composting process. Culture-dependent ap-proaches have been previously used to study microbes that areactive during the composting process (Ryckeboer et al., 2003;Strom, 1985a, 1985b; Beffa et al., 1996a, 1996b; Choi and Park,1998). However, only a small fraction of the microbes present inenvironment samples can be in fact cultured using the currenttechnology. As a result, important members of the compostingmicrobial community may have been missed. Furthermore, verylittle is known about microbial community structure at differentstages of the composting process. Therefore, culture-independentmethods have recently been employed to characterize the succes-sion of microbial communities during the composting process.

In contrast to numerous studies that have analyzed the micro-biology of the composting processes, the microbiological charac-terization of finished compost is still in its infancy, and asystematic microbiological analysis of products from compostingfacilities is still lacking (Hassen et al., 2001; Tang et al., 2003).

The objective of this study was to detect, characterize and com-pare the most abundant bacteria in compost produced by full scalerotary drum composter and collect information about the stabilityand reproducibility of the bacterial community structure in com-posting materials. For the purpose, we carried out pour platingenumerations of bacterial populations based on the methods pre-viously published (Hassen et al., 2001). In parallel, we estimated

es during full scale rotary drum composting. Waste Management (2013),

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Table 1Waste composition and characteristics.

2 A. Bhatia et al. / Waste Management xxx (2013) xxx–xxx

the diversity of the community in each sample by molecular clon-ing approach.

Feedstock materials Weight (kg) Moisture content (%)

Mixed vegetable waste 70 85Cattle manure 15 70Saw dust 5 33Tree leaves 10 30Total weight of mixture 100 75

2. Materials and methods

2.1. Reactor configuration

In the continuous composting process study, a full-scale rotarydrum composter (demonstration scale) of 3.5 m3 capacity was in-stalled at Indian Institute of Roorkee, India (Fig. 1). The main unitof the composter, i.e. rotary drum of 3.7 m in length and 1.1 m indiameter, made up of a 4 mm thick metal sheet. The inner side ofthe drum was painted with anti-corrosive coating. The drum ismounted on four metal rollers attached to metal stand. A 7.5 kW mo-tor with gear reducer is used to turn the drum in clockwise directionat a speed of 2 rpm. In order to provide the appropriate mixing andagitation, 400 mm long angles (4 mm width � 150 mm height) werewelded longitudinally. These angles provided tumbling action andhelp to move the waste material along the drum. With regards tothe composting process, the main function of rotation is to exposethe material to air, add oxygen and release the heat and gaseousproducts of decomposition. Two main openings are provided at bothends for waste inlet and outlet zone. A 2.5 kW air blower fixed at theinlet end was used to suck the air from outlet end for aeration. It alsopromoted the escape of water vapors and foul gases generated dur-ing composting. Two ports are provided at the middle and outletzone of drum to drain possible excess water and to collect compostsamples. The shredded mixed organic waste is loaded into the drumby the means of plastic container on daily basis. To reach the stabil-ization phase, the retention time was kept as 8 days. Frequency ofone rotation per day was made to ensure the materials on the topportion moved to the central portion. Thereafter aerobic conditionswere maintained by switching on the air blower.

2.2. Collection of samples

During the study period, uncooked vegetable (Green leafy veg-etables, onion, Carrot, tomato, potato, etc.) waste was collectedfrom community vegetable market, shredded up to 1 cm size usingmetal shredder and mixed with the amendments (Cattle manure,tree leaves and Saw Dust), based on C: N ratios (26:1) (Table 1).Thereafter, homogenous mixture of ingredients was used as feed-stock (100 kg/d) for the composting process in full scale RotaryDrum composter continuously on the daily basis. About 500 g ofeach grab samples were collected at every four days interval,mostly at the mid span of three zones by compost sampler withoutdisturbing the adjacent materials. Triplicate samples were col-lected and stored at 4 �C for physico-chemical characterizationwhereas microbial analysis was preceded at the same day ofsampling.

Fig. 1. Temperature variations and microbial communities during rotary drumcomposting.

Please cite this article in press as: Bhatia, A., et al. Diversity of bacterial isolathttp://dx.doi.org/10.1016/j.wasman.2013.03.019

2.3. Physico-chemical characteristics

Following physico-chemical parameters were analyzed, i.e.,Temperature, moisture content, pH, total carbon and total nitro-gen. Temperature was monitored on the basis of 24 h time intervalusing a digital thermometer throughout the composting period byinserting the thermometer into the composting mass in three dif-ferent locations. Moisture content was determined by weight lossof compost sample (105 �C for 24 h) using the gravimetric method(APHA, 2005). pH was measured using a pH meter with a glasselectrode (BIS, 1982). Total Carbon (%) and total nitrogen (%) wasanalyzed through Vario ELIII CHNSO Elemental analyzer on drymatter basis. The total organic matter (expressed in%) was calcu-lated according to the equation % carbon � 1.723.

2.4. Culture-dependent bacteriological enumeration

10 g of fresh waste or compost was dispersed into 90 mL of ster-ile distilled water and shaked mechanically for 2 h at 100 rpm inorder to remove the maximum of microorganisms from their org-ano-mineral substrates. Finally, the waste suspensions preparedwas used for bacterial counts. Pour Plate technique preparing serialdilutions of samples was performed to enumerate the bacterialcommunities during the composting period. Total heterotrophicbacterial count was determined using Nutrient Agar Medium(NAM). Plates were incubated in an inverted position for 24–48 hat 37 �C (Hassen et al., 2001). Indicator Organisms (Fecal Thermo-tolerant Coliforms and Fecal Streptococci) were monitored usingthe Most Probable Number (MPN) method (APHA, 2005) Esche-richia coli number was counted for serial dilutions of sample thatwere plated on Mac Conkey Agar medium and incubated in an in-verted position for 24–48 h at 37 �C. For Salmonella pathogenic spe-cies, samples were cultured on the plates of Modified SemisolidRappaport–Vassiliadis (MSRV) Medium and incubated for 17 h at42 �C. Suspected colonies were sub-cultured for confirmation onXylose Lysine Deoxycholate (XLD) agar with 21 h incubation at35 �C. Finally, Triple Sugar Iron agar (TSI) and Xylose Lysine Decar-boxylase biochemical tests were performed (USEPA, 2005a).Formorphological analysis Hi-media biochemical bacterial kit wasused for performing IMVic test, Urease test, Catalase test, fermen-tation of carbohydrates and gram staining was performed withhimedia-gram stains (Aneja, 1999).

2.5. Culture-independent analysis (molecular method)

2.5.1. Total genomic DNA extraction from samplesGenomic DNA was extracted from samples with beat-beating

method, using a DNA extraction Kit (Hi PurA™ Soil DNA kit) asper manufacturer’s instruction. Extracted yield DNA was measuredthrough spectrophotometer. The kit was designed in such a waythat apart from soil DNA, it was efficient to isolate DNA from othersources like compost, sludge, etc. In addition, lysis time period waselongated, as a pretreatment for DNA extraction process. The integ-rity and purity of the DNA was checked by horizontal gel electro-phoresis in 0.7% agarose gel.



A. Bhatia et al. / Waste Management xxx (2013) xxx–xxx 3

2.5.2. PCR amplification and 16S rRNA clone library constructionA stretch of 1465 bp was amplified from the total genomic DNA

isolated from compost sample using eubacterial specific 16 S rRNAuniversal primers 1492r (50-TAC CTT GTT ACG ACT T-30) and 27f

Fig. 2. Bacterial isolates analyzed thro

Table 2Physico-chemical characteristics of Rotary drum compost.

CompostingTime(Days)

Temperature (�C) Moisture content (%) pH

Inletzone

Middlezone

Outletzone

Inletzone

Middlezone

Outletzone

Inletzone

0 35 75 7.614 55 47 72.3 69 8.018 48 45 72 70 7.44

12 64 48 71 70 8.1116 62 48 34 79 77.5 76 7.7620 58 47 33 79.5 75.5 74.4 7.9124 60 45 36 75.5 75 71.5 8.3628 61 44 34 77.5 76.1 74.9 7.9532 63 45 35 74.5 74.5 71.5 8.3436 61 43 34 75.5 73.5 73 7.9540 62 42 34 74 70 64 7.8644 53 33 35 70 68 60 7.5848 59 42 34 65 62 56 8.352 61 41 34 72 70 64 7.7256 54 40 34 71.5 71 66 7.2760 48 34 33 69.5 70 65 7.7864 54 41 32 72.5 70.5 62 6.6568 62 39 33 71.5 71 66 8.3372 57 39 33 66 65 61 6.7576 59 42 34 69 65.5 63 6.7880 60 42 34 71.5 71 66 6.7584 57 42 34 72.5 70.5 62.5 7.2788 53 42 34 71.5 71 66 6.6592 45 31 26 72 70 58 8.7196 33 26 21 73 64 57 7.09

100 50 34 26 70.5 60 58 6.5


(50-AGA GTT TGA TCC TGG CTC AG-30) [14] .The PCR reaction wasdone using 2 lM of both primers, 50 ng of metagenomic DNA,200 lM of each dNTPs, 1� PCR buffer, 2 mM MgCl2, 2.5 units ofTaq DNA polymerase (Biochem Biotech products). The PCR reaction

ugh culture-dependent approach.

Total carbon (%) Total nitrogen (%)

Middlezone

Outletzone

Inletzone

Middlezone

Outletzone

Inletzone

Middlezone

Outletzone

30 1.437.54 24 25 1.51 1.547.35 7.8 29 26 1.27 1.467.48 8.8 29 35 31 1.48 1.48 1.547.84 8.46 32 32 30 1.36 1.56 1.518.27 8.37 32 30 28 1.2 1.6 1.408.41 8.65 30 29 27 1.3 1.43 1.568.6 8.76 29 30 26 1.48 1.47 1.58.66 8.12 33 26 29 1.39 1.29 1.218.57 8.65 34 26 31 1.25 1.3 1.278.23 8.5 35 28 26 1.30 1.24 1.368.12 8.34 32 29 26 1.46 1.54 1.678.32 7.41 34 26 33 1.36 1.42 1.508.38 8.98 32 26 24 1.09 1.91 1.318.68 8.86 33 29 29 1.23 1.35 1.528.43 8.93 27 29 25 1.2 1.26 1.398.56 7.81 24 29 26 1.436 1.31 1.078.6 7.89 25 24 14 0.942 1.44 1.287.48 7.52 29 19 20 1.256 1.41 1.057.5 7.56 27 16 1 1.2 1.26 1.397.48 7.52 28 17 15 1.436 1.31 1.078.68 8.86 27 18 25 0.942 1.44 1.288.56 7.81 27 19 25 1.09 0.91 1.318.51 7.53 30 20 18 1.361 1.13 1.967.5 7.59 26 20 16 1.34 1.42 2.017.5 7.57 34 28 18 1.407 1.46 2.14




was carried out at 94 �C for 2 min, followed by 35 cycles at 94 �Cfor 1 min, then at 48 �C for 1 min and 72 �C for 1.5 min Final elon-gation step was carried at 72 �C for 10 min (Acinas et al., 2004). Theamplified products corresponding to 1465 bp were electrophoreti-cally separated and gel purified using Gel extraction kit (Qiagencommercial kit). The purified PCR products were used for ligationusing vector pTZ57R-T (Fermentas) as per manufacturer instruc-tions. The ligated product was transformed into E. coli DH5 alphachemical competent cells. The positive white clones were selectedon LB, ampicillin and X gal, IPTG plates (Sambrook et al., 1989).

2.5.3. DNA sequencing and Insilco analysisClones (10 from inlet zone, 6 from out let, and one from middle

zone) were sent for partial sequencing of 16S rRNA gene using 27fforward primer to SciGenom Labs Private Ltd. (Kochi, India).DNAsequencing reactions were carried out using big dye terminationcycle sequencing kit version. Partial 16S rRNA gene sequenceswere initially analyzed for chimera, using pintail version 1.1. Iden-tification of clones are performed using BLASTn search facility.

2.5.4. Phylogenetic tree constructionPhylogenetic tree for the representative sequences were con-

structed using evolutionary distances and Neighbor-Joining meth-od implemented through NEIGHBOR (DNADIST) from the PHYLIPversion 3.61 packages. The tree was constructed with bootstrapanalysis with 1000 iterations.

0.1

Acinetobacter rad

Acinetobacter bau68

Moraxellaceae b23

Acinetobacter sp.

47

Acinetobacter sp.

39

Uncultured bacte

CloneACD1285

Acinetobacter sp. 82

35

Acinetobacter sp

Arthrobacter sp95

CloneACD11 79

100

64

100

CloneACD8

100

Lampropedia

Clone100

96

Uncultured

Uncultured C83

Clone ACD156

Uncultured rume

Uncultured rum90

100

Uncultured rum

Clone ACD16 68

Rumen bacterium [H

Lachnospiraceae b74

98

Clone ACD13

100

66

CloneACD1

Fig. 3. Rooted phylogenetic tree constructed from partial 16S rRNA gene derived from Inlare given at the branch point.


3. Results

3.1. Physico-chemical characteristics

Temperature ranged 64–32 �C in inlet zone, 72–31 �C in middlezone and 34–21 �C in outlet zone of rotary drum (Table 2). Thesevariations could be attributed to heat produced as a result ofmicrobial activities. Moisture content of inlet zone (75%) was re-duced to 60% in the middle zone and 58% in outlet zone of rotarydrum composter. The reduction of moisture content might beattributed to higher degree of temperature and evaporation lossto the surrounding air. Moisture loss during the active compostingphase can be viewed as an index of decomposition rate, since theheat generation which accompanies decomposition drives thevaporization or moisture loss is there (Liao et al., 1996). pH valuesof three zones of rotary drum ranged from neutral pH to slightlybasic nature. Initial amount of total carbon, 30% reduced to 28%and 18% in middle zone and outlet zone of the rotary drum, respec-tively. Total nitrogen values increased from initial values 1.43% to1.46% middle zone and 2.14% in outlet zone composts (Table 2).

3.2. Enumeration analysis

Total heterotrophic bacterial count in the feedstock was2.3 � 104 CFU/g. Finally, number reduced to 2.5 � 102 CFU/g and2.3 � 102 CFU/g in middle zone and outlet zone, respectively(Fig. 2). Presence of coliforms is often used as an indicator of

ioresistens [HQ908727]

mannii [GU319977]

acterium [EU169142]

[HQ407290

[GU566361]

rium [AM500790]

[EU073077 ]

. [AY673994]

.[ EF412972]

Clone ACD14

CloneACD9

hyaline [AY291121]

ACD17

Clostridiales bacterium [AB198575]

lostridiaceae bacterium [AB089035]

0

n bacterium clone [DQ673514]

en bacterium clone [EU259469]

en bacterium [AB034116]

M597706]

acterium [EU728736]

5

et zone of clone library. Bootstrap confidence values obtained with 1000 resamplings




overall sanitary quality of the compost. For compost hygenisation,the recommended Fecal Coliforms (FC) and Fecal Streptococci (FS)densities are 5 � 102 bacteria/g and 5 � 103 bacteria/g, respec-tively (Vuorinen and Saharinen, 1997). The average number of Fe-cal Coliforms showed a decrement with time from 9.3 � 105 MPN/g to 1.5 � 103 MPN/g and 1.5 � 102 MPN/g, in the middle zone andoutlet zone composts. Population of Fecal Streptococci decreasedfrom 9.3 � 104 MPN/g in inlet zone to 2.3 � 103 MPN/g in middleand 2.3 � 102 MPN/g in outlet zone The decrement in E. coli num-ber was observed from 2.3 � 104 CFU/g to 1.5 � 103 CFU/g and1.5 � 102 MPN, in the middle zone and outlet zone composts. Sal-monella isolates varied from 18 MPN/4 g in inlet zone to nil in mid-dle and outlet zone respectively. Similarly, Shigella species reducedto nil in the outlet zone compost (Fig. 2).

Discrete bacterial colonies cultured from all the samples werecharacterized morphologically by using phase contrast microscopeafter staining and identified the bacterial colonies have differentforms: rod, circular and irregular shape (Holt et al., 1994),while

PHYLIP_1 67

Flavoba

Uncultu100

58

uncultu

Chryseo100

100

FlavobaClone A

Fig. 4. Rooted phylogenetic tree constructed from partial 16S rRNA gene derived fromresamplings are given at the branch point.

0.1

Actinobacte

Rhodoluna64

Candidatus64

Uncultured

52

Candida

100

Cellu

100

Clone ACD6

99

100

83

U

75

U

100

CloneAC97

Clone

99

Fig. 5. Rooted phylogenetic tree constructed from partial 16S rRNA gene derived froresamplings are given at the branch point.


the color of the isolated colonies ranged from white, off white, yel-low, orange, or pink. For gram staining reactions, the majority ofthe bacterial isolates (78 out of 104) were gram positive rods; theyoccured singly or in chains, while six bacterial isolates were grampositive coccobacilli and rest were gram negative. A total 104 bac-terial species were isolated from all three zones samples which in-cludes Bacillus species, Micrococcus species, Pseudomonas species,Clostridium species. A large majority (33%) of total number of iso-lates were members of genus Bacillus.

3.3. Molecular analysis

All the non chimaric and non redundant 16S rRNA gene se-quences were submitted in NCBI for obtaining accession numbersJN872746–JN872756. The 16S rRNA analysis of the compostsamples gathered clones sequences that were clustered into eightgroups using phylogenetic analysis. The groups Actinobacteria,Bacillus, Clostridium, Acidovorax, Butyrivibrio, Pedobacter, Empedobactor

cterium sp. NL124 [AB636296]

red bacterium clone Q7375-HYSA [JN391863] red bacterium clone nbt96c04 [EU538971]

bacterium sp. UOF CM895 [AY468453]

cteriaceae bacterium KHS1 [AB636296] CD1

Middle zone of clone library. Bootstrap confidence values obtained with 1000

rium [AB426566]

sp. [AB607304]

Rhodoluna [AJ565417]

actinobacterium [JN656853]

tus Aquiluna [AM999985]

lomonas sp.

Uncultured Fusobacterium sp.[ AM159386]

Uncultured Fusobacterium sp.[ AM159386]100

100

Hydrogenophaga sp.

CloneACD255

Uncultured beta proteobacterium clone[DQ230946]

97

ncultured Hydrogenophaga sp[HQ183876]

ncultured Burkholderiales bacterium clone [EF648078]

D7

CloneACD3

ACD4

CloneACD5

m outlet zone of clone library. Bootstrap confidence values obtained with 1000



Table 3Similarity between bacterial isolates with MTCC bank gene and accession numbers.

Compostsamples

16S rDNA basedidentification

Closest identitybased on partialsequencehomology (%)

Gen bankaccessionno.

ACD1 Chryseobacterium sp. 89 JN872746ACD2 Hydrogenophaga sp. 99 JN872747ACD3 Flavobacterium sp. 97 JN872748ACD4 Uncultured bacterium clone 97 JN872749ACD5 Uncultured bacterium clone 94 JN872750ACD6 Actinobacterium bacterium clone 96 JN872751ACD7 Empedobactor sp. 88 JN872752ACD8 Acinetobactor sp. 89 JN872753ACD9 Uncultured bacterium clone 88 JN872754ACD10 Acinetobactor sp. 98 JN872755ACD12 Uncultured rumen bacterium 87 JN872756


and Flavobacterium were named by sequences clustering in the phy-logenetic tree (Figs. 3–5). Based on the species concept of microbialclassification about 40% of the sequences in the cloned library wereidentified as uncultured type and most of them are not classified.Microbial activity raised the temperature and changes in commu-nity structure of bacterial isolates were observed. .At the feeding (in-let) zone of the drum, dominant bacterial groups found weresignificantly related to Acinetobactor species, Bacillus species andClostridium species (Fig. 3), Bacteriodetes species, Chryseobacteriumspecies, Bacillus species were found mostly in the middle zone(Fig. 4) while Actinobacterium species, Hydrogenophaga species andFlavobacterium species were observed to be abundant in outlet zonecompost samples (Fig. 5). Dominance of these species might be dueto their living habits under meso- and thermophilic temperatures(Sneath, 1986). Although composting is supposed to be an aerobicprocess, members of Bacillus and Thermoactinomyces genera are fac-ultative anaerobic and members of Clostridium and Desulfomalacumgenera are strictly anaerobic (Sneath, 1986; Jones and Collins, 1986).Table 3 signifies the similarity between bacterial isolates with MTCCbank gene and accession numbers provided.

4. Discussion

Temperature is one of the key indicators of composting anddetermines the rate at which many of the biological processestakes place and plays a selective role on evolution and successionon the microbiological communities (Hassen et al., 2001). Temper-ature and microbial activity of Rotary Drum was observed to be inthe following sequence: inlet < middle > outlet zone. This could beattributed to the rapid intense microbial activity leading to fastercomposting, shorter residence times and partial pasteurisation ofthe compost causing death of many pathogens. As a result of theseactivities and subsequent composting, the numbers of indicatororganisms and pathogenic species in final rotary drum compostswere extremely low. While degrading organic compounds, mi-crobes convert 60–70% carbon to CO2 and utilize remaining 30–40% into their body as cellular components (Kalamdhad and Kazmi,2009). Organic matter mineralized after composting, mostly due tothe degradation of easily degradable compounds such as proteins,cellulose and hemi-cellulose are utilized by microorganisms as Cand N sources (Barington et al., 2002). Increase in value of totalnitrogen could be due to the net loss of dry mass in terms of carbondioxide and water loss by evaporation caused by heat evolved dur-ing oxidization of organic matter (Huang et al., 2004). The quicklychanging physicochemical conditions in composting processes arelikely to select for a succession of different microbial communitiesand it could be expected that temperature and the available sub-strates, are the main factors (Paul and Clark, 1996).


From the culture-dependant method, it was found the total bac-terial count decreased during the composting process. Interest-ingly, there was a gradual decrease in the load of pathogens,such as Salmonella and Shigella species during the process of com-posting. The decrease was presumably the result of the high tem-perature and unfavorable conditions established during thethermophilic phase (Hassen et al., 2001). From molecular analysis,it could be reported that number of pathogenic species decreasedduring the end of composting as compared to beneficial bacterialspecies. At the outlet zone, bacterial sp. such as Flavobacteriumsp. was found responsible for denitrification (Trios et al., 2010).Actinobacteria found in the drum are filamentous in nature andresponsible for degradation of cellulose, hemicelluloses and lignin(Perez et al., 2002). The middle zone of the rotary drum whosetemperature ranged from 26–48 �C creates a favorable conditionfor the growth of actinomycetes and other heat tolerating bacteria.High temperature intern accelerates breakdown of protein, fat andcomplex polymers. Rumen bacterium found in the solid matter issolely responsible for production of Volatile Fatty Acids (VFA)(Vuorinen and Saharinen, 1997) during the process of composting.Hydrogenophaga species are also involved in carbonaceous matterbreakdown and formation of volatile acids.

5. Conclusions

Aforementioned results concluded growth of significant micro-bial communities ensuing the higher rate of organics degradation.Bacterial diversity revealed in the composting process undergonethe changes, possibly linked to the variations in temperature andavailability of new metabolic substrates while decomposing organ-ics at different stages of composting. In addition, both Culture-dependent and independent approach gave an idea about the ma-jor bacterial populations responsible for the high rate rotary drumcomposting process. Hence, Rotary drum is proved to be a viable,efficient and appropriate decentralized composting technology inwhich most of the organic waste combination was primarily stabi-lized successfully within 7 days period in drum composting.

Acknowledgements

The authors would like to express thanks to Department of Sci-ence and Technology (DST) India for their financial assistance andDr.Ankur Rajpal for technical support.

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diversity of bacterial isolates during full scale rotary drum composting

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