Tracking and using genes in the environment - Environmental Science

TRACKING AND USING GENES The application of molecular techniques to under-

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environment has entered a renaissance period over the past five years. It is now possible to follow the fate of genetic determinants and their ability to function in the environment itself. The development of extraction techniques that isolate RNA and DNA from the environment, coupled with increasing availability of gene probes, has resulted in an explosion of information that affects microbial ecology, bioremediation and bioprocessing, public health indicators of microbial contamination, and genetic ecology. Further, the construction of genetically enhanced organisms is allowing advances in food and agricultural production, biocontrol, metal and mineral leaching and waste treatment ( I ) . Tables 1 and 2 contain examples of the types and nature of gene probes that are available. Gene probes detect specific genes or parts of genes in bacteria, viruses, fungi, or protozoa. Importantly, conserved regions of ribosomal gene sequences can be used to allow the identification of a generic group and regions of variation to i d e n t i f y specific subspecies. Many probes used for identification of pathogens have been developed in medical research. However, a n increasing number of probes to genes of catabolic and detoxification pathways has also been developed. The number of probes available to detect genes encoding for enzymes in degradative pathways is likely to continue to increase because of the ability to sequence the N-terminus of enzymes and construct cDNA from the code as well as the increasing ease of sequencing nucleic acid fragments. However, the mere occurrence of genes that allow the identification of an organism or an organism’s ability to degrade or detoxify a compound does not indicate if the organism is viable or if the system is

€44 Environ. Sci. Technal., Val. 25. NO.4, 1991

Betty H. Olson University of California Imine, CA 9271 7 being expressed by the production of enzymes that carry out the specific cellular processes. In order to study and detect specific mRNA messages, scientists have developed techniques to extract mRNA directly from cells and environmental samples (2,3).mRNA is transcribed from DNA and used as the template to formulate the polypeptides that comprise enzymes. It is far easier to measure mRNA than go through the abundance of lengthy extraction procedures for a variety of different enzymes that may be involved in a degradative pathway. Isolation of bacterial mRNA directly from environmental samples has been an elusive task because mRNA represents less than 5% of the total cellular RNA pool and has a half-life of 1-2 minutes. In addition to measuring mRNA directly, molecular reporters can be used to assess expression. Reporter systems utilize the lux gene cassette inserted behind a promoter gene for a function of interest or the luciferase gene fused to a promoter (4, 5). When the genes are “turned on” the lux genes are responsible for a light-emitting reaction. This system is highly sensitive and allows the determination of substrate availability in situ. Reporters may come to play a significant role in bioremediation. Thus far the system has been developed only for naphthalene (4). One additional technological advance just beginning to be applied to the environment is the polymerase chain reaction (PCR) for the amplification of specific DNA sequences. In this process a sequence of DNA or RNA can be repeatedly amplified in order to produce a measurable signal (Figure 1). The

technology is relatively new with the h u m a n B-globin

g ; l ) $ g ~ ~ ~ ~ ~ ( 6 ) . Primer DNA that flanks the region of interest is identified and must be unique to the system under investigation or false-positive results will be obtained. The process is carried out by repeated cycles in w h i c h t h e DNA is d e n a t u r e d through heating, the primers ann e a l e d to t h e c o m p l e m e n t a r y strands, and the primers extended with DNA polymerase. This technique allows the detection of genes that occur in very low frequency in the environment and is being applied to drinking water and foods. Microbial ecology The advent of nucleic acid probes has made it possible to track genes and genomes in soil, water, and sediments. The majority of studies undertaken thus far have focused on bacteria. The presence of genes and their relative significance from various ecological perspectives has been examined. One example of major interest is the use of gene probe technology in studying nitrogen fixation and as a means of understanding how to enhance the process. The density of catabolic degradative genes has been determined for a variety of compounds such as toluene and naphthalene and the detoxification of mercury (7,8, 9). In density assessments of genes in the environment, it is important to determine the precision of cell lysis as well as the internal and external variation of sample material. Many polluted sites contain gram negative bacteria, and the efficiency of lysis is approximately 85%. A number of gram-positive and gram-variable bacteria are more difficult to lyse. In certain polluted environments the external variation (variation between samples taken from the same location) can be

+ 100%.

Also of great interest is the occurrence of transposons in genomes that allow genetic material to move

0013-936)(/9110925-604502.50/00 1991 American Chemical Society

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within a cell and can also be important in the transfer of genetic information among microorganisms. Several degradative and detoxification systems have been linked with transposons such as naphthalene, narrow spectrum mercury resistance, and most recently phenol (IO, 11, 12J.The association of these genetic determinants with transposons increases the likelihood that they will be amplified within a cell and may become located on a conjugative plasmid. Location on the latter replicon aids i n interspecies transfer. A few studies have used nucleic acid probes to examine the density of occurrence of transpositional elements within the environment (13, 14, 15). These studies found that narrow spectrum mercury resistance was more closely associated with Tn21-like elements than Tn501-like elements. However, based on the molecular model one would have predicted Tn501 to predominate because mercury induces transposition within this molecular element (12). Further, studies conducted 01 bacterial isolates from the medical environment indicated that no Pseudomonos were isolated that contained TnZ1-like elements, whereas soil and sediment studiec found these transpositional units t commonly occur i n this genus Thus, molecular models should be viewed with caution when extrapolating from the field of molecular hi ology to the environment, and a VE riety of environments should b tested before conclusions are draw regarding the occurrence or lack c particular elements within a genu: Another important factor in az sessing the distribution of genes c bacterial origin in nature is the df g e e to which evolutionary divei gence occurs amongst nucleic acid! The question of divergence an how it affects detection of specifi DNA sequences in all environmer tal microorganisms is not well dc lineated; this is especially so for v ruses a n d protozoa. Howevei divergence is demonstrated in sex era1 medical models for bacteri and recently in symbiotic cyanobac teria (IS). Divergence can be detec ed by altering stringency or th number of mismatches allowed in nucleic acid hybridization reactio1 Hybridization is the process b which probe nucleic acids ar matched to a target sequence of ni cleic acids. DNA-DNA or DNA RNA hybridizations are carried 01 to minimize the number of mi: €46 Environ. Sci. Technol., VoI. 25, No. 4, 1991

Examples of nucleic acid probes for transformation c degradation of environmentally important compound Slruclural Compound -.

genes

Arsenic Cadmium

fes

Chromium

Naphthalene Salicylate Toluene Trichloroethylen Methylxylene Mercury Methylmerci Phenol Polychlonnated biphenyl Xylene Zlnc

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TABLE 2

Nucleic acid probes available for athogenic and indicator microorganismsof public health mportance

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minants as probes. Currently, if the conserved regions in viruses are used divergence is not thought to be a problem. Evidence that a new generation of investigation is beginning in microbial ecology is based on direct extraction techniques from waters, soils, and sediments 117, IS, 19). These techniques when completely successful will all but eliminate the use of culture media. Rapid tecbniques exist already for the extraction of bacterial DNA, RNA, and even mRNA from a variety of environmental substrates (3,20,21). Other proof that nucleic acid technologies will be commonplace in all aspects of environmental microbiology is the development of labels for nucleic acid probes other than radiolabeled isotopes. These include biotin and chemiluminescent dyes. The advantage of both colorimetric products is the long shelf life and the safety of nonradioactive markers. Biotin has not been widely used in environmental research because of its low sensitivity, tendency to react nonspecifically, and difficulty in reading positive results. For many organisms such as viruses and protozoa that occur in low densities in the environment, sample concentration is usually required and has been applied for years in the detection of these organisms. Although culture systems do exist for human viruses and are simple for bacteriophage, there are no similar methods available for parasitic protozoa. In the past both parasitic protozoa and human viruses have required elaborate testing procedures such as tissue culture or live animal infection. The problem with many of the extraction techniques developed is the time required, the labor intensiveness of the procedures, and the low yield of nucleic acid recovered. However, a second generation of procedures including polymerase chain reaction appears to be improving the procedures on all fronts. The development of mRNA metbods allows the assessment of bacterial function in situ. Information from molecular and physiological research suggests that many bacteria that possess a wide variety of genes do not normally express these genes in the environment. The exact reasons for this are unclear, but gene occurrence is not equivalent to the ability to express genes (221. Thus, the term "genetic potential" is more appropriate to use when re-

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matches between the base pairs of the probe and target sequence. DNA is denatured at high temperatures and reannealed by lowering the temperature. By making the reannealing temperature very close to the denaturation temperature of the nucleic acid sequence, the number of mismatches is minimized. Mismatches are also decreased by maintaining high stringency of wash conditions where extraneous DNA is removed from the filter-containing hybrid target and probe DNA. Generally the higher the wash temperature the higher the stringency of hybridization. Interpretation of the correct stringency to be used in environmental studies is problematic if divergence is common in bacterial genetic material from the environment. Some

examples for mercury indicate that relaxing stringency from 88-72% increased positive hybridization reactions from 55-92%. If stringency is allowed to drop too low then false positives will result, but if it is kept too high many organisms that possess the genes will be missed. A rule of thumb is to determine the distribution of the correlating phenotypes and then examine what striugency will produce accurate results for the specific system. Certain studies have shown that 70-80% stringency is appropriate for environmental studies, compared to >go% for medical research. Of course, certain genes or regions of genes are highly conserved, and as our knowledge increases the problem may be alleviated by selecting conserved regions of genetic deter-

Environ. Sci. Technol., Vol. 25. No. 4.

1991 €47

porting on gene density in the environment that has not been determined to show maximum expression in situ. Further, there appear to be three categories of gene expression in the environmental isolates measured by laboratory assessments. These are genotypes that can express immediately upon isolation from the environment, those that can express following subculture on nonselective media, and those that never express the genes they contain. For example, toluene biodegradation is well documented and toluene-degrading populations are commonly found in gasoline-contaminated environments. Bacteria isolated from a gasoline-contaminated site demonstrate the above phenomena as do bacteria isolated from chlorinated biphenyl hydrocarbons, mercury, and methylmercury-contaminated locations (23, 24). The environmental significance of these findings is as yet unclear, but they suggest that in situ bioremediation may be greatly enhanced. Additionally, reporter systems should be verified in concert with mRNA production in situ to determine if the natural populations are reacting in the same manner as the reporter system. Viruses and protozoa pose a different dilemma. Viruses cannot replicate outside the host, and protozoa of parasitic origin usually are inactive also. Therefore, it is difficult to ascertain viability and infectivity of these organisms without complex host assay systems. Recently activation of viral enzyme systems suggests an approach to determine ability to function. The data base is so small that no information exists regarding gene occurrence and function in protozoa. Bioremediation, bioprocessing Bioremediation has gained renewed acceptance because of the success of its application to several oil spills during the past three years. This technology stimulates the natural capacity of the microorganisms to degrade pollutants through the addition of nutrients. However, add i t i o n of n u t r i e n t s s t i m u l a t e s growth of all bacteria o r fungi present and does not select for those with special degradative abilities. Bioremediation and bioprocessing can be carried out by microorganisms using several biochemical mechanisms: oxidation, reduction, dehalogenation, and hydrolytic and condensation reactions. Recently attention was focused 608 Environ. Sci. Technol., Vol. 25, No. 4,1991

on the Exxon Valdez oil spill in Alaska where the application of bioremediation greatly enhanced degradation of oil residues on beaches. Since that time this technology has been applied to several oil spills with marked success. Factors that affect bioremediation are redox potential, pH, temperature, salinity, nutrient levels, concentration of the contaminant, effect of mixed contaminants, and microbial constraints. Bioremediation has its greatest potential when the pollutants are easily accessible such as in spills in aquatic environments or on soil surfaces. Major drawbacks in bioremediation are the substrate-chemical interaction, the ability to get nutrients to bacteria in complex media, and the potential of microbial growth to restrict aquifer flow. The ability of a substrate to adsorb or absorb to particles in soil and sediment systems may limit the bioavailability of a compound to microorganisms and can seriously limit the feasibility of bioremediation. The development of reporter systems will aid in determining whether chemicals are bioavailable in situ and will give an instantaneous result for use in field operations. However, the questions of in situ expression when substrate is available will also need to be addressed before the technology can be fully implemented, as so many bacteria in the environment possess the genetic capability to degrade a compound but do not express it even in the presence of substrate. Pump-and-treat technologies have been developed to remove chemicals from a sludge or water phase, and then return the liquid to the ground. Anaerobic degradation (reduction) has gained considerable attention for dehalogenation and degrad a t i o n of s u c h c o m p o u n d s as chlorophenol, trichlorethylene (TCE), and polychlorinated biphenyls (25, 26) because it is difficult to deliver oxygen in many contaminated soils. This approach has several drawbacks because it is slower than aerobic degradation and in certain instances can lead to more toxic intermediates or end products such as TCE being converted to vinyl chloride. Innovative combinations of approaches such as aerobic and anaerobic schemes allowing the in situ degradation of trichlorethylene are appealing. However, these processes can be slow and may not achieve complete mineralization.

In situ leaching involves the pumping of Thiobacillus or related microorganisms through drilled passages where ore remains in its original location. Usually the rock has to be loosened through blasting prior to this process. Both uranium and copper can be obtained through biological leaching using bacteria (27). Fungi are also used to solubilize low-rank coals, and recently the metabolite responsible for solubilization from T. versicolor has been isolated (28). Lastly, bioreactor technologies are improving through the development of genetically engineered bacteria that can be used in contained systems, the development of innovative substrate materials to support mixed culture growth for degrading more complex chemical combinations (29), and a better understanding of how to direct “cometabolism. ”

Public health pathogens, indicators Although the vast majority of researchers have been slow to apply molecular techniques to the water field, certain individuals have pioneered the effort. Nucleic acid probe technology was first applied to viruses and subsequently to bacteria and protozoa in waters and wastewaters. The investigators used traditional concentration techniques developed over the past several decades coupled first with direct hybridizations and later with PCR-enhanced hybridization detection (30). The incorporation of PCR greatly enhanced the sensitivity of detection. Low-level transmission of viruses and protozoa by water or food may be significant in the spread of these agents among communities. Even if only a few individuals become infected, a disease foci may be established (31). Currently, rota, hepatitis, and enteroviruses can be detected in waste water and fresh and marine waters using reverse transcriptase. In a reverse transcriptase reaction polymerase produces cDNA from RNA and then the cDNA is amplified by PCR. Thus, RNA viruses or a very low signal of mRNA from bacteria or protozoa can be detected. The detection of coliforms by PCR in water has involved the amplification of a target DNA sequence followed by gene probe analysis. To detect coliforms a region of lacZ gene was selected, because most coliforms can ferment lactose (32). To date two genes have been used in the identification of E. coli: malB,

the gene that encodes for maltose transport across the cell membrane, and lamB, which encodes for a surface protein recognized only by the E. coli-specific bacteriophage lambda. However, the researchers found that both Salmonella and Shigella contained portions of the lamB gene, although Salmonella could be excluded from detection by increasing the primer annealing temperature (31). Thus an E. coli-specific generic probe has not yet been developed for use in the environment. All of these systems use hybridization as a means of verification after amplification. Further, the detection of Legionella has been described by PCR. This study, as in the majority of those using PCR, relies on amplification and verification by hybridization. It has not been used to identify Legionella from any water systems (33). A panspecific probe for enteroviruses has been developed. It allows the detection of a family of viruses important in waterborne disease (34). Caution must always be used in developing systems such as those described above because of the possibility of shared DNA or RNA homology. Another perplexing issue facing public health microbiology is the concepts of bacterial injury and viable but nonculturable or dormant bacteria in water (35, 36). In the case of the former, special isolation techniques are required; in the latter case the organism can not be recovered by culture methods. Recently, in the case of L. pneumophila, viable but nonculturable bacteria were resuscitated through a heat shock procedure (37). In the case of Legionella the heat shock treatment also changes environmental serotypes to the pathogenic 01 serotype. These groups are significant to public health because (1)pathogenic organisms go undetected and (2) the bacteria are deemed no longer significant because these life stages are perceived as indicating probable organism death. Obviously, the data provided from the Legionella work suggest possible environmental stimuli for the conversion of nonpathogenic environmental strains to pathogenic strains. Further questions to be answered are the importance of heat shock as a means of reviving these bacteria from a sporelike stage and whether this is a phenomenon that affects all gramnegative bacteria. Importantly, the presence of heat shock genes and other pathogenic

capabilities can be determined by nucleic acid hybridizations even in the absence of viable cells. Although more information is needed before the exact public health significance can be determined, the controversy can only be resolved by further molecular studies. Protozoa and viruses pose another question asked by public health officials regarding whether occurrence equates to infectivity. To date the only way of determining infectivity is by passage through tissue culture with a variety of tests to measure cytopathogenic and noncytopathogenic effects for viruses and direct animal infection andlor excystation studies in the case of protozoa, Recently, the activation of a RNA transcriptase in enteroviruses indicates that the nucleic acid of the virus is intact. However, this procedure does not answer the question of capsid (the outer coat of enteric viruses) integrity and its role in infection. It is likely that in the future, infectivity may well be determined by mRNA assays for protozoa in the cyst stage.

Genetic ecology This approach is newly conceived and is still early in its development (38). Genetic ecology has already demonstrated that in situ d e g r a d a t i o n c a n be e n h a n c e d through targeting and enhancing natural genetic capabilities in contaminated materials. The process augments naturally occurring organisms in situ through the addition of inducer compounds that result in increased amplification of the genes of interest in the contaminated site. Figure 2 indicates the means by which this technology is taken from the laboratory to the field for individual pollutants or combinations of pollutants. One promising mechanism for natural amplification within living bacterial cells that can be directed is hypothesized to be controlled by transpositional events as well as other types of gene amplification such as formation of tandem repeats. This system has been used to amplify genes (merA) encoding mercury reductase, the enzyme that transforms Hg** to Hg" and naphthalene degradation (nahAB) (39, 40). The amount of amplification appears to be more dependent on the size of the operon being amplified rather than specific bacterial genus or species. Thus, the technology should be applicable to a wide range of bacterial genera, but cer-

tainly has been demonstrated for the genus Pseudomonas. The importance of Pseudomonas in degradation of recalcitrant compounds is well established. Further, it has been shown that these increases in gene copy number are accompanied by increases in the production of messenger transcripts (2, 40). Another means of increasing degradative capacity in bacterial communities, especially those associated with aquatic environments, is through the stimulation of gene transfer. In Table 3 a number of conjugative genes are shown for the genus Pseudomonas. Conjugation can be stimulated by increased nutrients, optimization of temperature and pH, as well as several other manipulations of physical or chemical characteristics applicable to the natural environment. Recent studies indicate that in polluted environments as much as 100% of bacterial populations of the same genus can receive and express genetic information important in detoxification of pollutants. These data indicate that the occurrence of certain genetic determinants could be greatly increased within a specific polluted site (40). Research is currently under way to determine the effect of these amplifications on degradation and detoxification rates. Thus far data suggest that rates may be increased several fold (39, 41). This approach differs from seeding a site with genetically engineered bacteria because only those organisms that have the natural ability to amplify are affected by the treatment. Thus, in situ the degraders of interest may be targeted and stimulated without affecting the rest of the community. After the inducer is removed the community returns to normal distribution of populations. The process by which this is achieved is shown in Figure 2.

Risk assessment of novel biological technologies EPA, the Food and Drug Administration, and the U.S. Department of Agriculture officially review the release of genetically engineered microorganisms into the environment. The role of EPA comes under the Federal Insecticide, Fungicide, and Rodenticide Act. A broad spectrum view of federal agency interaction in regulating biotechnology was published by the White House Office of Science and Technology (41). Public perception of risk from genetically engineered organisms has Einviron. Sci. Technol., Vol. 25,No. 4,1991 609

References ABLE 3

:atabolic and transforming plasmids from Pseudc...,. larmid

Substrate

AH AM ICT OL JP1 AC21 JP4 AC27

naphthalene camphor octane, hexane, decane p or mxylene, toluene

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fluoroacetate inorganic mercury phenol

154644

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2,4-dichlornphenoxyaceticacta

pchlorobiphenyl 2.4 D 3- and 4chlorobenzoate

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conjugative conjugative nonconjugative conjugative conjugative nonconjugative conjugative conjugative conjugative? conjugative conjugative

thus far not k : : Jverly enthusias tic. However, genetic engineering in relation to the environment has produced a great deal of information that is benefiting many of the above mentioned topics. Although the future of the application of genetically engineered microorganisms to the environment hangs in the balance, it is likely that, as our knowledge increases, it will come to be an acceptable and extremely useful technology. Environ. Sci. Technol., Vol.

778-81.

( 5 ) Braiser, A. R.: Tate, J. E.: Habner. J. F. Biotechnol. 1989, 7,1116-22. (61 Saiki, R. S. et al. Science 1985, 230, 1350-54. (71 Ogunseitan, 0.A. et al. I, Indust. Micmbiol. 1987.. l .. 311-17. (8)

Barkay, T. Appl. Envimn. Micmbiol. 1987,53,2725-32.

[continued on next page)

w w Refsrance 40

610

K. H. Crit. Rev. Biotechnol. 1988,8,85-97. (2) Tsai, Y. L.: Olson, B. H. Appl. Envimn. Microbiol. 1990,56, 3266-72. 131 . . Tsai. Y. L.: Parks. M.: Olson. B. H. Appl. Environ. Mi&ohiol., in ires,. (4) King, M. H. et al. Science 1990, 249, (1) Keeler,

25, No. 4, 1991

Obviously, the application of nucleic acid technologies to the environment will not solve all our local or global problems. These technologies will catapult this part of the field of environmental sciences as well as many others into the 21st century, causing rethinking of environmental engineering approaches to all aspects of the treatment of environmentally significant substances from drinking water to hazardous waste.

Betty H.Olson is a professor in the pro-

gram of social ecology, civil engineering, and environmental toxicology at the University of California, Imine.

(9) Sayler, G. S. et al. Dev. Indust. Microbiol. 1987,27, 135-49. (10) Tsuda, M.; IIno, T.; Mol. Gen. Genet. 1990,223,33-39. (11) Kivisar, M. et al. Plasmid 1990, 24, 25-36. ( 1 2 ) Brown, N. L.; Lund, P. A; Nibhriani, N. In Genetics of Bacterial Diversity; Hopward, D. A.; Chater, K. F., Eds.; Academic Press: New York, 1989, pp. 175-95. Summers, A. 0 . Ann. Rev. Microbiol. 1986,40, 607-34. Olson et al. Bull. Contamin. Toxicol. 1990,in press. Olson, B. H. et al. WaterRes. 1989,21, 1209-1 7. Plazinski, J. et al. Appl. Environ. Microbiol. 1990,56, 1263-70. Holben, W. E. et al. Appl. Environ. Microbiol. 1988,54, 703-11. (18) Ogram, A,; Sayler, G. S.; Barkay, T. J. Microbiol. Methods 1987,7, 57-66. (19) Deflaun, M. F.; Paul, J. H.; Davis, D. Appl. Environ. Microbiol. 1986, 52, 654-59. (20) Somerville, C. C. et al. Appl. Environ. Microbiol. 1989,55,548-54. ( 2 1 ) Tsai, Y . L.; Olson, B. H. Appl. Environ. Microbiol., in press. (22) Rochelle, P.; Wetherbee, M.; Olson, B. Appl. Environ. Microbiol., i n press. (23) Ridgway, H. F. et al. Appl. Environ. Microbiol. 1990,56, 3565-75. (24) Walia, S.; Khan, A,; Rosenthal, N. Appl. Environ. Microbiol. 1990, 56, 254-59. (25) Criddle, C. S.;DeWitt, J. T.; McCarty, P.L. Appl. Environ. Microbiol. 1990, 56,3247-54. (26) Haggblom, M. M., Young, L. Y . Appl. Environ. Microbiol. 1990,56,3255-60. (27) Crueger, W.; Crueger, A. Biotechnology: A Textbook of Industrial Microbiology; Sinauer Assoc.: Sunderland, MA, 1989. (28) Cohen, M. et al. Appl. Environ. Microbiol. 1990,56, 3285-91. (29) Adriaens, P.; Focht, D. Environ. Sci. Technol. 1990,24,1042-49. (30) Innis, M. et al. PCR Protocols, A Guide to Methods and Applications; Academic Press: San Diego, 1990. (31) Fields, B.; Martin, M. A,; Kamely, D. “Genetically Altered Viruses and the Environment”; Banbury Report 2 2 ; Cold Spring Harbor Laboratory, Cold Spring Harbor: New York, 1983. (32) Bej, A. K. et al. Appl. Environ. Microbiol. 1990,56, 307-14. (33) Starnbach, M.N.; Falkow, S.; Tompkins, L.S. J. Clin. Microbiol. 1989,27, 1257-61, (34) Chapman, M. N. et al. J. Clin. Microbi01. 1990,28, 843-50. (35) Bissonnette, G. K. et al. Appl. Environ. Microbiol. 1975,29,186-94. (36) Rozak, D. B.; Colwell, R. R. Microbiol. Rev. 1987,51,365-79. (37) Colburne, J. S.; Dennis, P. J. J. Inst. Water Environ. Manag. 1989, 3, 345-50. (38) Olson, B. H.; Goldstein, R. A. Environ. Sci. Technol. 1988,22, 370-72. (39) Ogunseitan, 0. A. et al. Abstr. Amer. SOC.Microbiol. 1990,Q425, 30. (40) Ogunseitan, 0. A.; Olson, B. H. Abstr. Amer. SOC.Microbiol. 1990,Q239, 328. (41) Rochelle, P.; Acacio, B. Abstr. Amer. SOC.Microbiol. 1989,Q79.

Hazardous Waste

Management

ere is an insightful reference on developing technologies for treating and managing wastewater and solid residuals that are contaminated with hazardous chemicals. Written by leading authorities in the field, this volume’s 22-chapters cover such diverse topics as municipal soiid wastes. water purification by radiation (solar and UV).the isolation of organic species and inorganic radionuclides, and solvent recycling. Several chapters cover radiolysis chemistry in dilute aqueous media, solar treatment, chemical separations, the biological and chemical treatment of soils and sludges, and solids immobilization. This concise presentation draws from a variety of science and engineering disciplines with specific emphasis on physical, inorganic/ organic, and biological chemistry. It provides an across-the-board perspective of problems and proposed management technologies. This volume will serve as a useful introduction to hazardous waste treatment for the novice as well as a valuable reference for the technical expert. D, William Tedder, Editor, Georgia Institute of Technology Frederick G. Pohland, Editor, University of Pittsburgh

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The scope of the book goes slightly beyond the title and includes cyanophytes that occur in fresh or brackish waters. Also, an examination of the interactions between toxins and their primary sites of action is provided. Shewood Hall, Editor, US. Food and Drug Administration Gary Strichartz, &ditor, Harvard Medical School Developed from a symposium held under the auspices of the Commission on Food Chemistry,Applied Chemistry Division, International Union of Pure and Applied Chemistry. ACS Symposium Series No. 418 390 pages (1989) Clothbound ISBN 0-8412-1733-5 LC 89-18505 $74.95 American Chemical Society Distribution Office. Dept. 59 1155 Sixteenth St.. N.W. Washington,DC 20036 or CALL TOLL FREE

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