ronment will not be valid. The consequences of impeded or inhibited microbial degradation of the components of a mixed-pollutant system are serious and should be investigated.
Literature Cited (1) Nelson, J. D., Jr., Colwell, R. R., Microb. Ecol., 1, 191-218 (1975). (2) Sayler, G. S., Shon, M., Colwell, R. R., ibid., submitted for publication. ( 3 ) Walker, J. D., Colwell, R. R., Appl. Microbiol., 27, 1053-60 (1974). (4) Hartung, R., Klingler, G. W., Enuiron. Sci. Technol., 4, 407-10 (1970).
(5) Seba, D. B., Corcoran, E. F., Pestic. Monit. J., 3, 190-93
(1969). (6) Walker, J. D., Colwell, R. R., Appl. Microbiol., 27, 285-87
(1974).
(7) Kenega, E. E., Proc. Intl. Conf. Transport Persist. Chem. Aquatic Ecosys., Vol 11, pp 19-22, Ottawa, Canada, 1974. (8) Larson, J. O., Tandeski, E. V., Anal. Chem., 47, 1159-61 (1975). (9) Sniegoski, P. J., Water Res., 9,421-23 (1975). (10) Alberts, J. A., Schindler, J. E., Miller, R. W., Nutter, D. E., Jr., Science, 184,895-97 (1974). (11) Walker, J. D., Colwell, R. R., Enuiron. Sci. Technol., 10, 1145
(1976).
Received for review September 2,1975. Accepted May 12,1976. Work supported by Sea Grant Project No 04-5-15811, National Oceanic and Atmospheric Administration, Washington, D.C.
Oil, Mercury, and Bacterial Interactions John D. Walker‘ and Rita R. Colwell” Department of Microbiology, University of Maryland, College Park, Md. 20742 Several strains of bacteria were examined for ability t o utilize oil with and without mercury added t o the culture medium. The majority of the oil-utilizing bacterial strains had been isolated from a n oil-contaminated creek in Baltimore Harbor of the Chesapeake Bay. Relatively low concentrations of mercury inhibited the utilization of oil by the strains of bacteria examined in this study. Concentration of environmental pollutants, viz., mercury and chlorinated pesticides, in oil has been reported by several investigators (1-3). In a study reported elsewhere ( 4 ) , partitioning of mercury and polychlorinated biphenyls in water and oil and on diatomacems earth was examined. If mercury, pesticides, and other pollutants concentrate in oil, effects of such combinations on the microbial degradation of each of the pollutants must be investigated since the potential consequences are of great environmental significance. Hence, as an initial step toward the understanding of the microbial degradation of combinations of selected pollutants, the effect of mercury concentrated in oil on the microbial biodegradation of oil was examined.
Materials a n d M e t h o d s Cultures. Procedures employed for the isolation and identification of cultures used in this study have been published previously (5,6). Bacterial strains examined are listed in Table I. A total of 20 strains of bacteria were screened for mercury resistance when grown in heterotrophic and in oil media. The strains were isolated from water, plankton, and sediment samples collected in several areas of Chesapeake Bay: Colgate Creek, a n oil-polluted site located in Baltimore Harbor: Poole’s Island, a dredge spoil disposal site in Chesapeake Bay; and Eastern Bay, a shellfish-harvesting area of the Bay. American Type Culture Collection (ATCC) strains were also examined as reference cultures. C u l t u r e Systems. The strains were cultured in a basal I Present address, Environmental Technology Center, Martin hlarietta Corp., 1430 South Rolling Road, Baltimore, Md. 21227.
Table I. Strains of Bacteria Used in This Study Genus and species name0
Lab no.
Strain no.
Source
Coryneform
4
...
P s e u d o m o n a s sp.
9s
...
P s e u d o m o n a s sp.
125
P s e u d o m o n a s sp.
215
Coryneform
232
V i b r i o sp.
254
...
A c i n e t o b a c t e r sp.
263
...
Pseudomonas aeruginosa R h i z o b i u m m elilo t i Rhizobium leguminosarum Nocardia otitidis caviaru m Leucothrix mucor Nocardia corallina No cardia asteroides
492
1014s-1
Colgate Creek sed i men t Eastern Bay water Colgate Creek sediment Eastern Bay water Colgate Creek sediment Colgate Creek water Poole’s Island sediment Colgate Creek water Colgate Creek plankton Colgate Creek sediment Eastern Bay sed i men t Colgate Creek water Eastern Bay sediment ATCC f J
495 496
4399 10004
ATCC ATCC
497
14629
ATCC
498 50 1 5 10
25107 4273 14759
ATCC ATCC ATCC
Coryneform
53
P s e u d o m o n a s sp.
1o s
Coryneform
76
...
P s e u d o m o n a s p.
11s
...
P s e u d o m o n a s sp.
101
P s e u d o m o n a s sp.
12s
...
O w h e r e t h e c o m m o n e p i t h e t , i.e., “ C o r y n e f o r m ” i s used o r speCieS g i v e n as “sp,!’, i d e n t i f i c a t i o n is p r e s u m p t i v e a n d f u r t h e r S t u d y o f t h e c l a s s i f i c a t i o n o f t h e s t r a i n i s i n progress. b o b t a i n e d f r o m t h e A m e r i c a n T y p e C u l t u r e Collection. 1 2 3 0 1 P a r k l a w n . R o c k v i l l e , Md.
Volume 10, Number 12, November 1976
1145
Table II.Growth of Chesapeake Bay Bacteria in Heterotrophic (HI and Oil (OFMediaa at Selected Concentrations of Mercury Med in which HgCI, tested ppm
H 0 H 0 H 0 H
O
4
9s
53
.
.
.
.
0
-
I
.
1
l
Laboratory no. designating culture tested
5 5 10
+
+
-
-
10s
.
+
+
-
76
.
+ +
-
11s
.
+ +
-
+
+ -
-
-
-
+
-
-
+
+
215
-
+
-
254
.
-
+
-
+ -
-
+
-
+ -
232
.
+
-
-
-
+ +
-
+
+
.
+ +
-
125
.
+
-
+ + + + O - - - - - - -
12s
.
+ +
-
101
.
+
-
-
+
-
-
+
-
-
.
+
-
+
-
495
.
+
-
+
-
. +
-
-
+
2 6 3 492
-
498
.
.
.
-
-
-
497
-
-
-
496
-
-
- - - - -
-
-
+
-
+
-
-
-
501
510
.
.
-
-
+
-
-
-
-
aSee Materials and Methods section.
Table 1 1 1 . Percent Isolates Examined and Found Resistant to Selected Concentrations of MgCI, % resistant HgCI,. mgll.
0 1
5 10 20 30 40 50
ATCC ref strains
100 43 0 0 0 0 0 0
Chesapeake Bay isolates
100 100 69 62 23 15 8 0
Total isolates
100 80 45 40 15 10
5 0
medium designed for heterotrophic bacteria ( 5 ) or in an oil salts medium supplemented with 1.0% (v/v) oil (7). Five-ml quantities of the appropriate medium in 16 X 150 mm test tubes were used. The medium was inoculated with l o 2cells/ml of each isolate. Inorganic mercury, HgC12, was added a t a final concentration indicated in the experiments described (vide infra). Inoculated media were incubated quiescently for 30 days a t 25 "C. Growth was measured by examining turbidity in liquid media or in the oil phase. Uninoculated controls were always included in the experiments.
Results a n d Discussion Fifty-five percent of the strains examined grew on the oil medium, and over half had been isolated from samples collected in oil-contaminated Colgate Creek (Table 11). Eighty percent of the total isolates were resistant to 1 ppm HgC12, when cultured in heterotrophic medium (Table 111). The percentage of resistant isolates decreased as the concentration of HgC12 added to the medium was increased (Table 111).The decrease was not exponential with increasing HgC12 concentrations, but instead exhibited threshold tendencies a t concentrations of 1-10 ppm HgC12, especially in the case of the Chesapeake Bay isolates. Of the ATCC strains examined, only two were resistant to the concentrations of mercury tested. Therefore, when results for these strains were included in the calculations, the percentage of the total set of isolates that were mercury-resistant was lower (Table 111). Sixty-two per1146
Environmental Science & Technology
cent of the strains isolated from the Chesapeake Bay samples were resistant to mercury a t concentrations of 5 10 ppm. All of these strains had been isolated from Colgate Creek in Baltimore Harbor. When mercury was added to the oil medium used in this study, none of the strains grew, even a t 1ppm HgC12. Analysis of a medium containing ~ Woil O showed that 85%of the HgC12 was found in the oil phase ( 4 ) .Assuming that the concentration of HgC12 in the oil was saturating for a 10% oil-water mixture, a 1.0%oil-water mixture, therefore, can be calculated to contain a t least 85% of the Hg added. The 5 pg of HgC12 added to 5 ml medium, yielding 1ppm mercury, would provide a maximum of 100 ppm mercury for the volume of oil used (0.05 ml). If 85% of the mercury is concentrated in the oil, the concentration of mercury in the oil, Le., a 1ppm 5-ml mixture, would be 85 ppm, an amount far greater than that observed to be inhibitory for growth of mercury-resistant bacteria in oil. When the concentration of mercury was adjusted relative to final concentration in the oil phase, two strains 12s and 125, both of which had been isolated from Colgate Creek, were able to grow in the oil at a concentration of 1-5 ppm mercury in the oil. Prior exposure of these strains to mercury-oil combinations in Colgate Creek water and sediment (3), probably induced resistance to mercury in the oil, enabling degradation of the oil to proceed in the presence of relatively low concentrations of mercury. However, oil in which mercury is heavily concentrated is concluded to be recalcitrant to degradation since the petroleum-degrading isolates examined in this study could not tolerate extremely high concentrations of mercury in oil, and many of the petroleum degraders were unable to grow in oil in which low levels of mercury were present. Since oil degradation in the marine environment is a relatively slow process, even under optimum conditions, interference or inhibition of microbial degradation caused by combinations of pollutants, as shown in this case for oil-mercury concentrations, must be considered as a serious environmental hazard at sites where mixtures of effluents entering estuaries, coastal waters, or the ocean result in antagonism toward microbial degradation.
Acknowledgment The excellent technical assistance of P. A. Seesman is gratefully acknowledged.
Literature Cited (1) Hartung, R., Klingler, G . W., &hiron. sei. Technol., 4,407-10 (1970). ( 2 ) Seba, D. B., Corcoran, E. F., Pestic. Monit. J., 3, 190-3 (1969). (3) Walker, J. D., Colwell, R. R., A .P.P ~Microbiol., . 27. 285-7 (1974). (4) Sayler, G. S., Colwell, R. R., Enuiron. Sci. Technol., 10, 1142 (1976). (.i) Walker, J. D., Colwell, R. R., Microb. Ecol., 1,63295 (1974).
(6) Walker, J. D., Austin, H. F., Colwell, R. R., J . Gen. Appl. Microb i d , 21, 27-39 (1975). (7) Walker, J. D., Colwell, R. R., A p p l . Microbiol., 31, 198-207 (1976).
Receiued for recieu: September 2,1975. Accepted May 12, 1976. Work supported by Contract N00014-75-C-0340 between the Office of Nacal Research and the Uniuersity of Maryland and National Science Foundation Grant No. B M S 72-02227-AO2.
Combined Process of Pyrolysis and Combustion for Sludge Disposal Nobuo Takeda" and Masakatsu Hiraoka Department of Sanitary Engineering, Faculty of Engineering, Kyoto University, Sakyo-ku, Kyoto, Japan 606
A combined process of pyrolysis and combustion of sewage sludge was examined by use of a double hearth incinerator. The effectiveness of the introduction of a pyrolysis stage to incineration was investigated and compared with the usual sludge incineration. The behavior of heavy metals and other toxic or unburnt gases was analyzed in connection with the pyrolysis temperature, showing the effectiveness of this process for the prevention of air pollution. Pyrolysis at low temperature minimized the vaporization of heavy metals contained in the sludge without sacrificing the reduction of bulk density. The analyses of heavy metals caught in the particulates and scrubbed water suggested that much of the vaporized metals escape as fumes. An adequate temperature control may suppress nitrogen oxide emission. In Japan, incineration is the prevalent method for sewage sludge disposal. As sludge production increases with the spread of sewer systems, new incineration plants are constantly being constructed. Indeed, the incineration method is excellent in the respect that perishable organic sludge can be converted into stable inerts, reduced in volume, and disposed under good sanitary conditions. But it is inevitably such a high temperature operation that the vaporization of some heavy metals and/or the production of toxic gases, e.g., sulfur oxides and nitrogen oxides, can cause serious air pollution problems. Moreover, the highly excess air necessary for incineration results in so much volume of flue gas that gas cleaning equipment becomes prohibitively expensive. A continuous sequence of drying, decomposition of organics, flame and char combustion is carried out simultaneously in a sludge incinerator. The lower limit of the combustion temperature is governed by the condition that no odorous matter is permitted to remain in the flue gas, and the higher limit that the residual ash does not melt. Hence, the combustion temperature is usually maintained between 700 and 1000 "C. T o prevent the heavy metals from vaporizing into the air, it is preferable to burn the sludge a t a temperature as low as possible. Also, a homogeneous gas-phase combustion is better than a heterogeneous gas-solid phase condition for reducing the flue gas volume. Thus, the authors planned a "two-stage combustion" in which the organics of the sludge were decomposed into gases in the first stage, and the destructed gases were burnt through a gas-phase reaction in the secondary stage. Some experimental results are reported in this paper, demonstrating the effectiveness of this process. Experimental
A double hearth incinerator, MHI-2SE (4 m$ X 5 m), with a secondary combustion furnace was used. T o maintain an
oxygen-free atmosphere, the double hearth incinerator was preliminarily improved to prevent any air leakage. Sludge was supplied to the top of the incinerator and thermally decomposed through the double hearth incinerator; the pyrolysis gases were burnt in the secondary combustion furnace. For the pyrolysis experiment, the air supply to the preheater burner was minimized to prevent the advancement of the combustion reaction in the double hearth incinerator. After burn out in the secondary combustion furnace, the flue gas was mixed with cooling air prior to conduction to the stack. Part of the flue gas was delivered to a venturi-type scrubber to examine particulate collection efficiency. The entire experimental equipment is shown in Figure 1. The temperature of the lower stage of the double hearth incinerator was varied from 450 "C (Run I), 500 "C (Run 2), 600 "C (Run 3) to 800 "C (Run 4), while the secondary combustion furnace was kept between 700 and 820 "C. Run 4 was carried out as a control experiment under usual incinerating conditions, namely, sufficient air was supplied to burn out the sludge within the double hearth incinerator. The sludge consisted of a filtrated cake of mixed primary and surplus activated sludge from a municipal sewage treatment plant. The properties of this sludge were typical of an activated sewage sludge. A certain amount of heavy metal chlorides (nitrate for lead) was added to the sludge to point up the behavior of the heavy metals more clearly. Since the moisture content of the received sludge cake was too high for pyrolysis at a low temperature in this incinerator, it was initially dried to about 40% moisture content. This, however, was not done for Run 4. The sludge feeding rate was 100 kg-wet sludge/h for Runs 1-3 and 240 kg-wet sludgeh for Run 4; this corresponded to about 60 kg-dry sludge/h. Flue gas, 1000 nm3/h, was delivered to the scrubber at 200 "C and was scrubbed with 7-15 1. of water/min. The scrubber
Figure 1. Schematic diagram of experimental equipment 1: Sludge feeder, 2: upper stage, 3: lower stage, 4: ash container, 5: preheater. 6: secondary combustion furnace, 7: fuel oil tank, 8: venturi scrubber, 9: water tank, 10: mist separator, 11: stack, FI: flow indicator, PI: pressure indicator, TR: temperature recorder, S: solid sampling point, G: gas sampling point, L: liquid sampling point
Volume
IO, Number 12, November 1976
1147
