Vol. 46, No. 2APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Aug. 1983, p. 406-4160099-2240/83/080406-11$02.00/0Copyright © 1983, American Society for Microbiology

Microbial Growth Associated with Granular Activated Carbonin a Pilot Water Treatment Facility

D. P. WILCOX,'* E. CHANG,2 K. L. DICKSON,1'2 AND K. R. JOHANSSON2

Institute ofApplied Sciences' and Biology Department,2 North Texas State University, Denton, Texas 76203

Received 21 December 1982/Accepted 6 June 1983

The microbial dynamics associated with granular activated carbon (GAC) in apilot water treatment plant were investigated over a period of 16 months.Microbial populations were monitored in the influent and effluent waters and onthe GAC particles by means of total plate counts and ATP assays. Microbialpopulations between the influent and effluent waters of the GAC columnsgenerally increased, indicating microbial growth. The dominant genera of micro-organisms isolated from interstitial waters and GAC particles were Achromobac-ter, Acinetobacter, Aeromonas, Alcaligenes, Bacillus, Chromobacterium, Cory-nebacterium, Micrococcus, Microcyclus, Paracoccus, and Pseudomonas.Coliform bacteria were found in small numbers in the effluents from some of theGAC columns in the later months of the study. Oxidation of influent waters withozone and maintenance of aerobic conditions on the GAC columns failed toappreciably enhance the microbial growth on GAC.

After the enactment of the Safe DrinkingWater Act in 1974, the U.S. EnvironmentalProtection Agency, acting under the authority ofthis legislation, promulgated the National Inter-im Primary Drinking Water Standards. Thesestandards set maximum contaminant levels forseveral inorganic elements, organic compounds,organic pesticides, radioactivity, and microbialconcentrations. These standards were amendedto include the establishment of maximum con-taminant levels of four halo-organic substancesknown as the trihalomethanes (THM) which areformed when humic compounds are chlorinated(2, 10, 23, 30, 36, 37, 43, 45). Since thesechlorinated compounds are carcinogenic or mu-tagenic (28, 39, 46), several methods for theirremoval from potable waters have been investi-gated (5, 11, 29, 38). One of the most promisingmethods consists of the removal ofTHM precur-sor compounds through their adsorption bygranular activated carbon (GAC).The idea of using GAC for the removal of

organics is not new. In Europe, GAC has beenused for years to remove organics associatedwith taste and odors, as well as for color remov-al (33, 42, 48; R. G. Rice and G. W. Miller,paper presented at the International Ozone Insti-tute Symposium on Advanced Ozone Technolo-gy, Toronto, Ontario, Canada, 1977). However,only recently has it been suggested that microor-ganisms, known to proliferate on GAC filters,may be important in the removal of organicsubstances from influent waters (3, 4, 15, 24, 25;D. van der Kooij, paper presented at the Confer-

ence on Oxidation Techniques in Drinking Wa-ter Treatment, Karlruhe, Germany, 1978).Many naturally occurring organic compounds

in water supplies, particularly humus, are rela-tively refractory to biodegradation (14, 21, 22,27). Various workers have suggested enhancingthe biodegradability of humic materials throughpreliminary ozonation (32; Rice and Miller, pa-per presented). This combined ozone-GACtreatment, used before chlorination, has beentermed the biological activated carbon (BAC)process. Proponents of this process contend thatit is capable of enhancing the adsorptive capaci-ty ofGAC (12, 32, 41) and extending the lifetimeof GAC columns almost indefinitely, providedmicrobiological colonization of GAC is main-tained (16, 50; R. C. Hoehn, R. C. Walter, A. J.Sullivan, and F. J. Neary, paper presented atthe 69th Annual Meeting of the American Insti-tute of Chemical Engineers, Cincinnati, Ohio,1971; H. Sontheimer, paper presented at theSeminar on Current Status of Wastewater Treat-ment and Disinfection with Ozone, Cleveland,Ohio, 1977). Treatment efficiencies during suchextended lifetimes ordinarily do not equal thoseof fresh carbon, but the proponents of the BACprocess contend that raising empty bed contacttimes allows the BAC process to achieve desir-able treatment efficiencies (16; Hoehn et al.,paper presented; Sontheimer, paper presented).European studies (19, 49; Sontheimer, paperpresented) have shown the operational life ofBAC filters to range from 1 to 3 years beforeregeneration is needed, whereas the time be-

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MICROBIAL BIOMASS ON GRANULAR ACTIVATED CARBON 407

tween regenerations for the regular GAC treat-ment process is much shorter (a few weeks toseveral months), although the frequency of re-generation depends on the quality of the influentwaters. The extended time between regenera-tions of the BAC system is attributed to microbi-al oxidation and utilization of absorbed com-pounds, resulting in continuous partialregeneration of the carbon surface and thus anextended service life.

Since knowledge about the BAC process hasbeen derived primarily from research efforts inEuropean waterworks, and because few of thesestudies reveal performance criteria which matchthose required by U.S. drinking water regula-tions, the U.S. Environmental Protection Agen-cy has sponsored several pilot-scale investiga-tions (including the one reported in this paper)concerned with the direct removal of THM orTHM precursors from drinking water throughthe BAC process (32). As an integral part of ourpilot study, which focused on the removal ofTHM precursors by GAC with and withoutpreozonation, microbiological studies were con-ducted to provide some insights into the follow-ing questions:

(i) What type of organisms are associated withthe BAC process?

(ii) Do microorganisms actually grow on GACsurfaces or are they simply filtered out, thusaccumulating to give the appearance of growth?

(iii) Do microorganisms of public health signif-icance grow in the BAC system?

MATERIALS AND METHODSPilot plant description and study site. The pilot

facility, housed in a 40-ft (ca. 1,219.20-cm) drop-frame

trailer, contained a treatment train that included alumflocculation, multimedia filtration, ozonation, andBAC adsorption processes. A schematic diagram ofthe pilot facility is shown in Fig. 1. Each GAC columncontained 37 kg of Filtrasorb-400 (Calgon Corp.) GAC.Based on the flow rates indicated in Fig. 1, the emptybed contact time for a pair of GAC columns wasapproximately 20 min.The pilot treatment facility was located at the Amiss

Water Treatment Facility, Shreveport, La., on theshore of Cross Lake. This study area was chosenbecause Cross Lake (Shreveport's main water supply)is known to have a high trihalomethane formationpotential resulting from runoff into the reservoir,which is high in naturally occurring organic com-pounds, such as humic and fulvic acids. These types ofsubstances, along with certain other dissolved organ-ics, are known precursors of THM (2, 10, 28, 30, 31).Sampling schedule. Once a month during the period

of January through November 1980, water and GACsamples were collected from the BAC pilot plant.Water samples were taken before and after eachexperimental carbon column, and GAC samples weretaken from sampling ports in the middle of eachcolumn (Fig. 2). These samples were immediatelytransported (on ice) to laboratories at North TexasState University in Denton, Tex., for analysis. Duringthe latter part of September and October 1980, addi-tional samples of GAC were taken to obtain moreinformation on the microorganisms associated withGAC. Sampling for microbial biomass present on GACparticles was begun in July 1980 and continuedthrough July 1981. Beginning in mid-November 1980,columns 1 and 2 were eliminated from the regularsampling scheme to allow their modification for use onother project objectives. All other columns continuedto be monitored, as before, through July 1981.ATP. The utility of the ATP assay as a measure of

viable microbial biomass in aqueous media has beenestablished by several investigators (7-9, 17, 18, 44).Mclllvaine buffer (ph 7.7) was used to extract ATP

FLOW -*Z~~~~

WATER WITH A3 MINUTE

OZONE CONTACT TIME

NON-OZONATEDWATE R

B FLOW RATE (GPM)GAC COLUMN

FIG. 1. Schematic flow diagram of the pilot water treatment facility, Shreveport, La.

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* Carbon Sampling PortO Other Sampling PortsA Water Sampling Site- Water Flow

Columns are245 cm in height,30.5 cm in diameter,and ore made of304 stainless steel

c0

-eU--

-0 ED "

u.x

_ I0

76-

FIG. 2. Flow diagram illustrating water and GACsampling points for a set ofGAC columns (identical forall column sets).

from both water and GAC samples. Extraction effi-ciency tests for ATP from GAC, similar to thoseperformed by Bullied (7) on sediments, indicated thatATP could be extracted from microorganisms associ-ated with GAC with better than 90% efficiency (Table1).

One-liter water samples were collected before andafter each set of columns in bottles rinsed with waterfrom the appropriate sampling points. In the labora-tory (on site or at North Texas State University), thesesamples were subdivided into 250-ml portions andfiltered through 0.45-,um membrane filters. Each filterwas extracted for 3 to 5 min in boiling McIllvainebuffer and then frozen at -20°C.GAC was transferred from the middle sampling port

of each column (Fig. 2) with the assistance of a sterilespatula and low-pressure water from the column to a75-ml sterilized brown glass vial. The samples wereiced and transported to the laboratory, where three 2-ml (packed volume) portions of GAC were extractedfor 10 min in 10 ml of boiling Mclllvaine buffer on hotplates in 150-ml beakers covered with watch glasses.Each extract was decanted into a centrifuge tube andcentrifuged for 5 min at 3,000 rpm (Spinette centrifuge;International Equipment Co., Needham, Mass.) tosediment any carry-over particulates. The supernatantwas then diluted to a constant volume (10 ml) in a testtube and frozen at -200C.

All extracts were analyzed for ATP content by themethod of Chappelle and Picciolo (9) with an ATPphotometer (model 2000; JRB Co., La Jolla, Calif.).The data generated from these analyses are reportedas nanograms of ATP per gram (dry weight) of carbonor nanograms of ATP per liter of water.

Selective and total plate counts. Water samples fortotal plate counts were taken from three-way, sole-noid-operated sampling valves located before and aftereach carbon column. Interstitial water samples (e.g.,low-pressure water obtained during GAC removal)and GAC samples were removed in the same manneras ATP samples. All plate count samples were sealed

in 500-ml sterilized glass containers with screw-caplids, promptly iced, and transported to the NorthTexas State University laboratory, where they wereprocessed within 24 h.To maximize the recovery of microorganisms from

GAC surfaces, the samples were sonicated (Sonifercell disruptor, model W185D; Heat Systems-Ultrason-ics, Inc., Plainview, N.Y.) as recommended by McEl-haney et al. (25). Wet GAC (1 ml) was placed in asterile 10-ml graduated cylinder along with 9 ml ofsterile phosphate buffer (pH 7.0). This suspension wasthen transferred to a sterile 50-ml beaker for sonica-tion. The sonifer probe was disinfected with 95%isopropyl alcohol, and the carbon suspension was thensonicated in an ice bath for 1 min at 60 W. As soon aspossible (<1 h), the cold sonicate was analyzed micro-biologically.The basic procedure for total plate counts was

modified from that described in Standard Methods forthe Examination of Water and Wastewater (1). Basedon the recommendations of McElhaney (26), soil ex-tract agar was employed. Serial decimal dilutions ofthe sonicates or of the influent/effluent and interstitialwaters were prepared in test tubes (16 by 150 mm)containing 9 ml of sterile phosphate buffer. One millili-ter of each dilution was transferred to each of twosterile petri dishes (100 by 15 mm) and mixed thor-oughly with 15 ml of melted (45°C) soil extract agar.Other petri dishes were prepared with several selec-tive agar media: MacConkey agar (Difco Laboratories,Detroit, Mich.), Burk 1% glucose agar, Cooke rosebengal agar (Difco) with tetracycline (0.036 g/ml), andactinomycete isolation agar (Difco) with actidione(0.05 mg/ml). The latter medium was inoculated byusing the double-layer technique (1). Duplicate plateswere inoculated by spreading evenly 0.1 ml of undilut-ed or diluted sample over the agar surface with asterile, bent-glass rod (alcohol flamed). All plates wereinoculated, inverted, and promptly incubated aerobi-cally at 28°C for 7 days (14 days for actinomycetes).All colony counts were extrapolated from the mostcountable pair of plates and expressed as CFU permilliliter of water or per gram (dry weight) of carbon.

Identification of microbial isolates from GAC. Thesoil extract agar plates were examined under a stereo-scopic microscope (20x magnification) to determinethe dominant colonial types. Pure cultures of selecteddominant colonies were isolated from nutrient agar(Difco) streak plates and maintained by monthly trans-

TABLE 1. Calculation of percentage recovery ofATP from GAC with McIlvaine buffer

ATP (ng) in Extracted ATP (ng/g) ATP (ng)spikedrcGC GC

on GAC recovered fromextrct dry wt alone bacterial spikea(10 ml) (g)

84.5 1.% 76.8 7.786.3 2.05 75.5 10.886.7 2.10 77.9 8.888.3 1.89 74.7 8.6

a Mean (column 1 - column 3) = 8.97 ± 1.3. ATPcontent in the bacterial spike (1.0 ml) = 9.8 ng/liter.Therefore, percent recovery = 8.97/9.80 = 0.915 x100 = 91.5.

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MICROBIAL BIOMASS ON GRANULAR ACTIVATED CARBON 409

fer in the same medium. All isolates were character-ized according to the basic determinative schemedisplayed in Fig. 3. Identification to the generic levelof the isolates was facilitated by several guides (6, 20,34, 35, 40). However, Bergey's Manual ofDetermina-tive Bacteriology (6) was the final authority.

Coilform tests. Parallel total and fecal coliformcounts in interstitial and influent/effluent water sam-ples were obtained by the standard membrane filtermethods (1, 47).

RESULTSThe data in Table 2 reveal the numbers and

types of microorganisms on GAC and in theinterstitial waters of the GAC columns. Thedominant isolates from soil extract agar wereidentified as Pseudomonas spp. and Bacillusspp. Nine other genera, all gram negative, wereregularly encountered. CFU in interstitial waterranged from 1.8 x 104 to 1.8 to 107/ml. Thenumbers of CFU per gram of GAC ranged from3 x 5s to 6 x 1. No effect of ozonation on thenumbers of microorganisms in the interstitialwaters and on GAC was apparent.

Plating of GAC sonicates on several selectivemedia (Table 3) revealed the most abundantmicroorganisms to be gram-negative bacteriawhich generally grow well on MacConkey agar.The latter counts ranged between 100 and15,000/g (dry weight) of GAC. Actinomyceteswere present in numbers from 200 to 4,400CFU/g (dry weight). Low numbers offungi wereencountered on Cooke rose bengal agar. Free-living, nitrogen-fixing bacteria appeared occa-sionally on Burk medium. Ozonation appeared

to have little qualitative effect on the GACmicroflora.

Microbial biomass and numbers were moni-tored in the influent waters to and effluentwaters from the three pairs of carbon columns.One set of columns received influent waterswhich had been ozonated (2 to 3 mg/liter) with acontact time of 3 min. Another set was exposedto ozone (2 to 3 mg/liter) for 40 min. A controlset of columns received nonozonated influentwater. Between February 1980 and July 1981,monthly samples of influent and effluent watersfrom the three sets of columns were collectedand assayed for ATP and microbial CFU (Fig.4). In general, ATP levels and the number ofbacteria were lower in the ozonated than innonozonated influents. With few exceptions, thenumber of CFU and levels of ATP were higherin the effluents, from all sets of columns, than inthe influents, indicating proliferation of microor-ganisms in the GAC columns. Generally, theeffluents from the ozonated columns did notcontain any higher concentrations of bacteriathan the effluents from columns receiving nono-zonated water, suggesting that microbiologicalactivity is not greatly enhanced by ozonation.

Levels of ATP in the GAC particles in thethree sets of carbon columns ranged from 5 to490 ng/g (dry weight) of carbon (Fig. 5). Ozona-tion did not consistently affect ATP levels of theGAC biomass. Likewise, the numbers of CFUpresent on GAC in the ozonated and the nono-zonated carbon columns did not consistentlydiffer (Fig. 6). Figure 6 also reveals greater GAC

Dominant Colony

Isolate(Nutrient Agar)

Pure Culture

Gram Stain

Gross PhysicalMorphology

I

G(-)

I. Growth on MacConkey Agar2. Oxidose Test3. OXI/FERM Tube (Roche)4. Pigment Formation5. Odor

G(+

CoccusII. Catolose Test Rod2. Carbohydrate Reaction I. Spore Sta in3. Coagulase Test 2. Catalose4. Pigment Formation

I.

Additional Tests: Motililty Test* Hemolysis aH2S Production* Optochin Susceptibility* Bile* Esculin* Salt Tolerance (6.5% NaCI)* Bacitracin Susceptibility

FIG. 3. Flow chart used to help identify the bacteria isolated in this study. G(+), Gram positive; G(-), gramnegative.

Additional Tests:* Motility* Antibiotic Sensitivitye Gelotinasee Lysine Decarboxylasee ONPG* Anaerobic Growth* Salt Tolerance (6.5% NoCI)e Growth at 42°C

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TABLE 2. Bacterial colony counts and dominant genera in GAC columns (June to October 1980)CFU x 106 Dominant generac

(1980) Column no.' per ml of perg (dry wt) Interstitialinterstitial water of carbonb water Carbon

6/13 1 0.11 2.0 12 1, 2, 4, 13

7/10

8/1

8/15

8/29

9/12

10/1

23

135

I23456

123456

123456

1234S6

123456

10/27 123456

1.800.04

0.504.31.80

4.00.656.00.60.760.3

6.21.6

10.00.51.40.77

18.03.78.01.80.32.0

1.20.820.40.40.180.1

1.30.90.350.60.70.25

1.00.310.650.070.050.018

6.62.0

12.014.036.0

20.04.38.02.01.62.0

40.018.020.011.015.033.0

60.010.023.08.02.02.6

23.015.033.04.72.02.6

4.56.65.50.50.92.7

2.70.71.40.50.30.3

2, 9, 1312

11, 1212, 1313

6, 11, 126, 126, 126. 123, 6, 7, 9, 129, 12

11, 12125, 12127, 9, 127, 12

6, 8, 11, 124, 6, 8, 128, 11, 126, 138, 126, 8

12, 1312, 134, 6, 1212, 1310, 12, 1312, 13

6, 134, 6, 124, 6, 122, 6, 136, 122, 6

6, 134, 6, 124, 6, 122, 6, 136, 122, 6

122, 6, 9

2, 9, 121212

11, 12, 132, 6, 12, 132, 6, 125, 12, 135, 12, 1312

9, 129, 1212127, 127, 12

6, 8, 11, 126, 8, 126, 8, 126, 86, 88, 12

1, 12, 136, 12, 136, 6, 1212, 139, 1210, 12

6, 12, 136, 124, 6, 126, 126, 12, 136, 13

6, 12, 136, 124, 6, 126, 126, 12, 136, 13

a Column no. 1 received water with an ozone contact time of 3 min; column no. 3 received water with an ozonecontact time of 40 min; column no. 5 received nonozonated water; columns no. 2 and 4 received water fromcolumns 1 and 3, respectively; column no. 6 received water from column 5.

b Deduced from a standard plot relating packed volume of carbon to dry weight.c Key: 1, Achromobacter; 2, Acinetobacter; 3, Aeromonas; 4, Alcaligenes; 5, Alcaligenes-like; 6, Bacillus; 7,

Chromobacterium; 8, Corynebacterium; 9, Micrococcus; 10, Microcyclus; 11, Paracoccus; 12, Pseudomonas;13, Pseudomonas-like.

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MICROBIAL BIOMASS ON GRANULAR ACTIVATED CARBON 411

TABLE 3. Colony counts obtained from carbon particles by using type-specific media and sonificationmethodologiesa

CFU x 102SampledateCFx10

(1980) Column no. Actinomycete Cooke rose(1980) ~~~~~~MacConkey aigail isolation agar" Burk agarb' Cookel rose

6/13 1 9.0 5.0 2.5 1.02 1,200.0 2.0 0 1.03 1,500.0 1.2 0.3 0.1

7/10 1 90.0 40.0 47.0 2.03 300.0 10.0 1.3 0.25 400.0 2.0 4.0 0.1

8/1 1 40.0 4.0 1.0 02 0.1 4.0 1.0 7.03 1.0 2.0 0 04 18.0 4.0 0.2 05 35.0 0.6 0 06 19.0 4.0 0 0.5

8/15 1 3.0 6.0 10.0 1.02 4.0 7.6 3.0 2.03 13.0 4.0 0 0.54 12.0 26.0 0 0.85 35.0 14.0 0 0.26 60.0 40.0 1.0 3.0

8/29 1 9.0 12.0 0 0.22 3.5 44.0 0 1.03 4.0 10.0 0 2.04 11.0 1.2 0 05 4.0 10.0 0 0.26 2.0 6.0 0 0.5

9/12 1 5.0 10.0 0 4.02 10.0 7.5 0 03 1.2 0.5 0 04 13.0 0.2 0 2.05 3.0 2.0 0 0.56 1.0 1.2 0 0

10/1 1 2.0 10.0 0 02 22.0 3.2 0 03 17.0 6.0 0 04 20.0 2.2 0 05 1.4 10.0 0 06 3.0 48.0 0 0

10/27 1 20.0 4.0 0 1.02 50.0 1.2 0 03 42.0 0 0 04 40.0 0 0 05 10.0 1.4 0 06 10.0 2.4 0 0.3

a Data for June to October 1980 are reported as number per gram (dry weight) of carbon. No significant datawere developed before June 1980.

b Microbial groups responsive to medium types: MacConkey agar, gram-negative bacteria (some Pseudomo-nas species and most Enterobacteriacae); actinomycete isolation agar, actinomycetes; Burk agar, nitrogen-fixingbacteria; Cooke rose bengal agar, fungi.

microbial populations in the warmer months and the GAC biomass was higher in the ozonatedshows that the highest levels occurred in the than the nonozonated columns; however, thissecond year (April to August 1981). During the difference had reversed itself by the end of theearly stages of the study, the number of CFU in study.

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B. MAR. MAY JUNE JULY AUG.

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b c b c b c b c b c b c b c

JAN, FEB MAR APR MAY JUNE JULY1981

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a b c a bc ab c a b c a b c ab c abc abc abc abcFEB. FEB. MAR. MAY JUNE JULY AUG. SEPT. OCT. NOV.

o t,t,, nt 1980o Influent* Effluent

Sets of Columns Receiving:a- 03(3 min) Waterb- 03 (40 min) Waterc- Non-03 Water

-Ib c

DEC.

b c b c b cJAN FEB. MAR.

b c b c b c b c

APR. MAY JUNE JULY1981

FIG. 4. Biomass as reflected in ATP and total plate counts for influent and effluent waters of GAC columns.

Total and fecal coliforms were monitored inthe final effluent waters from the ozonated andnonozonated sets of carbon columns (Fig. 7). Nofecal coliforms were detected in any of themonthly effluent samples from the columns.However, fecal coliforms were present in theraw influent waters to the pilot treatment facility(data not shown). Low numbers of total coli-forms (10 to 40/100 ml) were observed in efflu-ents from the nonozonated columns during July

through October 1980. Toward the end of thestudy (April, May, June, and July 1981) the totalcoliforms in the effluents from the ozonated andnonozonated sets of columns increased substan-tially (Fig. 7).

DISCUSSIONThe predominant microbial genera isolated

during the study were oxidative, saprophyticchemogranotrophs, otherwise characterized as

a

aa

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MICROBIAL BIOMASS ON GRANULAR ACTIVATED CARBON 413

1980

0 03 Water* Non-03 Water

Sets of Columns Receiving:a- 03 (3 min) Water (Columns 1, 2)b- 03 (40 min ) Water (Columns 3, 4)c- Non-03 Water (Columns 5,6)

3456 3456 3456

b c b c b cJAN. FEB. MAR.

3 4 5 6

b cAPR1981

3 4 5 6

b cMAY

FIG. 5. Microbial biomass on GAC as measured by ATP in columns receiving ozonated and nonozonatedwater.

(i) gram-negative, aerobic, nonfermentative ba-cilli (e.g., Pseudomonas, Chromobacterium,etc.) and (ii) gram-positive, aerobic, sporeformers (Bacillus spp.). Smaller numbers of ni-

trogen-fixing bacteria, actinomycetes, were alsofound in the GAC columns. McElhaney et al.(25) and Donlan and Yohe (13) found similartypes of organisms on GAC in a pilot water

500

400 -

0

a 3000

3'

5 200_0.4

100

IL3 4 5 6

b cJULY

3 45 6

b cJUNE

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v--Column No. (3 Min. Ozonated)_ _Column No. 3 (40 Min. Ozonoted)*--* Column No. 5 (Non-Ozonated)

Feb Mor Apr Moy Jun Jui Aug Sep Oct Nov 'Mar Apr May Jun Jul- 1980 1981

MONTH

FIG. 6. CFU on GAC particles for GAC columns 1, 3, and 5.

treatment facility operated by the city of Phila-delphia, Pa. Pseudomonads were found byHoehn et al. (paper presented) to be the domi-nant bacteria on GAC. It is clear that the micro-bial biomass on the carbon surfaces is dynamicand provides a niche for the normal microfloraof soil and water. Unfortunately, we did not lookfor autotrophs or for budding, gliding, orsheathed bacteria common to aquatic habitats.The effects of preozonation on organic materi-

als before microbial processing has been studiedby several investigators (15, 16, 50; Rice andMiller, paper presented). Most have found thatozonation of influent waters tends to enhancemicrobial growth on GAC particles. This en-hanced growth is credited with extending theperiod of time before the GAC must be regener-

- 10

9gEn2 8U)z 7o E

6

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ated. Our comparative study failed to showclearly enhancement of microbial growth byozonation. Generally, levels of ATP and CFU inozonated columns were similar to those found innonozonated columns. It is evident from thefluctuating data on ATP and CFU in influent andeffluent waters (Fig. 4) that microorganismswere growing and metabolizing in the GACcolumns. The effluents from all GAC columnswere often higher in CFU than the influent,suggesting intermittent sloughing of the GACbiomass. The carbon columns were not simplyacting as microbial filters.

All bacteria isolated from soil extract agar orfrom the selective agars inoculated with GACspecimens were found to be typical aquaticspecies. None belonged to the family Enterobac-

I -

FINAL EFFLUENTFROM NON-OZONATED COLUMNS

FINAL EFFLUENTFROM 40 MIN. OZONATED COLUMNS

I I o I I I 11___

III,

FIG. 7. Coliform bacteria in effluent waters from those GAC columns receiving nonozonated and ozonated(40 min) influents.

z9 _z

lma:

L) 8

cj 7

a:tD 6

z

0 5

tr:

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MICROBIAL BIOMASS ON GRANULAR ACTIVATED CARBON 415

teriaceae. Potential human pathogens encoun-tered included Pseudomonas spp., Alcaligenesspp., Acinetobacter spp., and Bacillus spp. Al-though no fecal coliforms were detected in theeffluents from the carbon columns, total coli-forms increased appreciably during the latterpart of the study (Fig. 7). The reason for thiscoliform breakthrough is not apparent. Whethercoliforms or other types of bacteria which ac-company them in the effluents from GAC col-umns present any health hazards would dependon the efficacy of chlorination after GAC filtra-tion. Further microbiological studies, includingefforts to detect animal viruses, are needed toestablish the public health significance of GACmicroflora.

ACKNOWLEDGMENTSWe thank James Wallace and Amiss Water Treatment Plant

(Shreveport, La.) laboratory personnel for their efforts inmaintaining the pilot water treatment facility and for providinglaboratory space and equipment. We extend special thanks toWilliam Glaze, University of Texas at Dallas, for his collabo-ration during the study.

This work was supported by contract CR-806157 to WilliamGlaze from the U.S. Environmental Protection Agency. Someof the microbiological efforts were supported by a grant fromthe Robert C. Brown Foundation, Dallas, Tex.

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