APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Sept. 1979, p. 499-5050099-2240/79/09-0499/07$02.00/0

Vol. 38, No. 3

Lytic Organisms and Photooxidative Effects: Influence on

Blue-Green Algae (Cyanobacteria) in Lake Mendota,Wisconsin

R. D. FALLON'* AND T. D. BROCK2

Environmental Transport Division, Savannah River Laboratories, E. I. du Pont Nemours & Co., Aiken,South Carolina 29801,1 and Department of Bacteriology, University of Wisconsin, Madison,

Wisconsin 537062

Received for publication 11 July 1979

The effects of exposure to high light intensities on blue-green algal (cyanobac-terial) populations were examined in Lake Mendota, Wis. The algal populationswere shown to be susceptible to inhibition of photosynthetic activity and pigmentbleaching as a result of exposure. These effects generally influence only a small

percentage of the lake population and thus are probably not important in causingmajor declines in chlorophyll a. Lytic organisms were shown to increase innumbers in the lake in response to the seasonal development of blue-green algae,reaching values of greater than 1,000 plaque-forming units per ml in midsummer.Both bacteria and protozoa were observed in plaque zones, but it could not bedetermined whether these lytic organisms had a major role in algal biomassdeclines.

In situations where lake phytoplankton isdominated by blue-green algae (cyanobacteria),the blue-green algal populations often undergoperiods of rapid decline in biomass (7; P. E.Sager, Ph.D. thesis, University of Wisconsin,Madison, 1967; R. E. Stauffer, Ph.D. thesis, Uni-versity of Wisconsin, Madison, 1974). These de-cline events are often associated with a changein the dominant genus of blue-green alga (12).Numerous suggestions have been made to ex-plain such decline events. Mechanisms such asallelopathic interactions, development of path-ogens, and photooxidation, along with manyother possibilities, have been suggested as pos-sible causes for these declines. Such events canhave severe consequences in small bodies ofwater. The death and subsequent decompositionof the algal blooms can result in reduced concen-trations of dissolved oxygen and thus increasedfish mortality.We have been studying the blue-green algal

populations in Lake Mendota during 1976 and1977. Periods of decline in blue-green algal bio-mass have been noted in the lake throughoutthe summer and fall (R. D. Fallon, Ph.D. thesis,University of Wisconsin, Madison, 1978) duringthe period of blue-green algal dominance inthese years. Declines in total lake chlorophyll aof 50% in 2 to 3 weeks were not unusual. Intrying to account for such declines, we haveexamined a number of factors. Here we considertwo of these: algal lytic organisms and photoox-

idation. Both of these factors have been sug-gested as possible causes for sudden declines inblue-green algal biomass in other studies.

Bacteria able to lyse blue-green algae havebeen shown to be very common in a number ofBritish lakes containing blue-green algae (4).The numbers of these bacteria increase duringthe period of blue-green algal growth. Lytic bac-teria have been shown to be quite effective inlysing various species of blue-green algae (15).Fungal and viral parasites, along with protozoanpredators, have also been observed in associa-tion with blue-green algal declines (3, 9, 11, 12).Work by Eloff et al. (6) has shown that photoox-idation and bleaching of pigments does occurunder natural conditions in Israeli fish ponds.They do caution, however, that natural popula-tions may be more resistant to high light expo-sure than many of the laboratory strains whichwere examined (5, 6). Thus, although both fac-tors have been shown to be potentially harmfulto blue-green populations, the exact nature oftheir effects under natural conditions is stillpoorly understood.

MATERIALS AND METHODS

Lytic organisms. Water samples were collectedwith plastic Van Dorn bottles, placed in 60-ml bottles,and held at 6°C in the dark until processing. In 1976,the samples were collected at the east, central, andwest stations at the surface, 10 m and 2 m above thebottom. Two meters above the bottom equaled depths

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of 13 m, 22 m, and 18 m at the east, central, and weststations, respectively. In 1977, samples were takenfrom the central station only at depths of 0, 10, and 20m. To assay for lytic organisms, various dilutions oflake water were filtered through sterile 0.45-tAm mem-brane filters (Millipore Corp.), in combination with 2to 3 ml of an axenic culture ofAnacystis sp. The filterswere then incubated in the dark at 30°C on agar platescontaining Microcystis aeruginosa magic elixir me-dium, pH 8.5 (8). After 7 to 8 days, cleared zones(plaques) on the Anacystis lawn were counted. Allmembrane filters were soaked in 1% peptone brothbefore use, and the Anacystis cultures used had beenstored in the dark for 2 weeks at 6°C before use. Bothof these steps were included to hinder the developmentof viral plaques on the Anacystis lawns. The peptonetreatment removed charged sites from the Milliporefilter, decreasing electrostatic adsorption of viruses tothe filter. Prestorage and incubation in the dark lowersthe energy charge of the host cell (Anacystis), whichlowers the reproductive success of the viruses (10).

Photooxidation and high light exposure. Me-teorological data for wind speed, temperature, anddaily solar radiation were supplied by the NationalOceanic and Atmospheric Administration (NOAA)from monthly summaries of data collected at TruaxField, approximately 9 km east of the center of thelake. Light intensities were measured with a Li-Corquantum meter, model LI-185, equipped with a modelLI-192S quantum sensor or with a Whitney light meter(maximum sensitivity at 510 nm; Chipman Instru-ments, Madison, Wis.). Oxygen and temperature mea-surements were made with a YSI model 51A oxygenmeter equipped with a model 5700 oxygen temperatureprobe. Pigment measurements were made by themethod of Strickland and Parsons (16) by measuringthe optical density of an extract at 663 nm before andafter acidification. The solvent system used was di-methyl sulfoxide/acetone/water (10:9:1) (13). Data fortotal lake chlorophyll a are based on integrated chlo-rophyll profiles made weekly at four to eight stationsin the lake throughout the summer.A temperature-controlled aquarium was used for

laboratory incubations. The water temperature in theaquarium was kept to within 2 or 3°C of the initialtemperature of the sample. For these experiments,algal material was first concentrated by filtrationthrough 16 layers of nylon netting (150,tm). Algae soconcentrated were resuspended in lake water. To ex-amine the effects of exposure to high light intensities,2 to 3 ml of concentrated algal suspension was exposedfor prescribed periods. Upon removal from high-inten-sity exposure, an additional 8 ml of 0.45-,um filteredlake water was added to the samples. The sampleswere then incubated for 1 h in the presence of 1 ,uCi ofH'4C02 at a light intensity of 100 micro-Einstein units(j,E) m-2 s- supplied by two 40-W cool white fluores-cent bulbs. Radioactive samples were filtered throughWhatman GF/C glass fiber filters, and the filters weredried overnight. The dried filters were exposed to HCIfumes to remove any radioactivity present as carbon-ate precipitates. Radioactivity on the filters was thenmeasured by using a Packard 3375 liquid scintillationspectrometer. The scintillation cocktail consisted of1.475 g of PPO (2,5-diphenyloxazole) and 0.3898 g of

methyl-POPOP [1,4-bis-2-(4-methyl-5-phenyloxa-zole) benzene] in 3.8 liters of toluene. Quench correc-tions were made according to the channels ratiosmethod (2).The procedure of adding additional lake water to

the samples after high-light incubations insured stan-dard conditions for the period of low-light incubationwith H'4CO2. Thus, changes observed in CO2 uptakewere due to changes in the algal population, not tochanges induced in the incubation medium by theperiod of high light exposure. Photosynthetic activityin unconcentrated lake water samples was measuredin the same way.

RESULTSLytic organisms. Lytic organisms were

quantified as numbers of plaques appearing inthe Anacystis lawn. The plaque-forming units(PFU) in the water column increased to peakvalues of greater than 1,000 PFU ml-' by mid-summer in both years (Fig. 1). In both years,this represented a 1,000-fold or greater increaseover a 2-month period. This increase correlatedwith the development of blue-green algae in thelake (Fig. 2).

Results from both years also showed a ten-dency for the numbers of PFU in the deep watersamples to be lower by a factor of between 10

o13 1976 -

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FIG. 1. Concentration ofPFU in Lake Mendota in(A) 1976 at the surface (0) and 10 m (A) and 2 mabove the bottom (U) and (B) in 1977 at the surface(0) and 10 m (A) and 20 m (U). Values in 1976 arethe arithmetic means of three stations in an east-westtransect of the lake. Values in 1977 are from thecentral station.

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FIG. 2. Total lake chlorophyll in Lake Mendota in1976 and 1977.

and 100 from those in the surface waters (0 and10 m), especially in midsummer (Fig. 1). Blue-green algal chlorophyll was also concentrated inthe upper 10 m of the water column. Samplestaken before the seasonal development of blue-green algae generally showed less than 1 PFU/ml. However, mud samples taken at varioustimes of the year from various parts of the lakeconsistently showed high numbers of PFU, con-taining from 50 to 200 PFU/0.05 ml of wet mud.

Microscopic examination of the plaques indi-cated the presence of both bacteria and proto-zoa. Bacteria present were commonly nonmotilerods, 0.5 to 1.5,um in diameter and 3 to 10 ,umlong. Occasionally, rapidly moving coccoid rodswere also observed. Unarmored amoebae werethe most commonly observed protozoans. Fun-gal hyphae were never observed. Two bacterialstrains were isolated from the plaques and par-tially characterized. They were gram-negativerods which were obligately aerobic and had atemperature optimnum for growth of around300C. Both isolates were strongly pigmented.The yellow isolate showed rapid gliding motilityon agar plates, whereas the orange isolate wasnonmotile. Axenic cultures of these two isolateswould form plaques on Anacystis lawns.Photooxidation and effects of high light

intensities. Field observations indicated thatconditions promoting photooxidation occurredin Lake Mendota during both summers. Obser-vations on calm, clear days showed that surfacelight intensities were 1,800 to 2,100 ,IE/m2 per s

during midday. Oxygen concentrations occasion-ally reached 200% saturation (15 mg of 02 perliter at 310C) in protected surface waters. Suchconditions were promoted by the developmentof a temporary, shallow thermocline due to heat-

ing of the surface water. On such occasions,surface concentrations of blue-green algae oftendeveloped. Although generally healthy in ap-pearance at 0800 h, small areas of bleachedcolonies appeared by 1030 to 1100 h, and bleach-ing of the surface scums was often quite exten-sive by 1300 h. The sequence of color changesduring bleaching indicated that chlorophyll waslost first. Small patches of blue colonies ap-peared by 1000 h, indicating the loss of chloro-phyll with the remaining phycocyanin pigmentscausing the blue coloration. Further exposure ofthese colonies to high light resulted in completeloss of color. Such colonies began to appear by1100 h. During this latter phase, the algal cellsbegan to lyse. Lysis was indicated by the pres-ence of a blue opalescent sheen in the surfacewaters, resulting from the release of phycocy-anin pigments and gas vesicles from the lysingalgae. Centrifugation of water from areas withthis appearance at 1,000 x g for 10 min yieldedan upper opalescent layer and a blue superna-tant. A similar result is obtained if one centri-fuges blue-green algal material which has beenground in a mortar and pestle. The opalescentsurface layer is found to consist of gas vesicles,and the blue pigment has an absorption maxi-mum of about 620 nm, indicating that it is prob-ably phycocyanin.Although dramatic in their occurrence, these

bleaching events are usually very localized, con-fined to areas protected from the wind. Onlyunder very calm, clear conditions was bleachingwidespread. Normally, wind-induced turbulenceappeared to prevent the algae from remaining atthe surface long enough to become bleached.Based on observations made in 1976 and 1977,estimates were made for critical wind speed andsolar radiation values required for widespreadbleaching to occur. Average wind speeds for theday had to be less than about 3 m s-' and dailysolar radiation had to be greater than 3.4 x 10+3to 3.9 x 10+3 E m-2 day-'. The effects of turbu-lence on the algal colonies was also influencedby the colony size. Algae which formed largerclumps were less influenced by turbulence andthus more prone to exposure. Solar radiationless than the critical value apparently did notallow a long enough midday period of high lightto promote the bleaching sequence. Based onNOAA data for the June through August periodin both years, only 24 days in 1976 and 11 daysin 1977 had wind and light conditions suitablefor lake-wide bleaching to occur. The value for1976 was unusually high, based on NOAA datafor previous years.To examine the effects of exposure to high

light more closely, a number ofexperiments were

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carried out using natural populations undermore controlled conditions. Examination ofchlorophyll a and carotenoid bleaching (Fig. 3)showed that, at intensities ranging from 900 to2,100 ,uE m-2 s-1, a loss of 50% of initial chloro-phyll a concentrations required from 2.5 to 6 h.Changes occurring in carotenoids (optical den-sity, 480 nm) during the same were less, rangingfrom no change to a 30% loss in 4 h at a lightintensity of 1,800 to 2,100 ILE m-2 s-I sunlight.

In contrast to the rather slow declines in chlo-rophyll a during high light exposures, photosyn-thetic activity in the same algal populationsdeclined rapidly. Although 2.5 h was requiredfor a 50% decline in chlorophyll a, carbon fixa-tion declined to about 15% of the initial valueafter only 40 min of exposure (Fig. 4). Exposureto sunlight at less than 100% will cause similardeclines in photosynthetic activity (Fig. 5). Ex-periments performed on consecutive days withsimilar algal populations showed that at inten-sities of 75% or greater of full sunlight, the timecourse of inhibition was quite similar. However,at an intensity equal to only 50% of full sunlight,no inhibition was observed.

Laboratory experiments with natural popula-tions indicated that exposure to intense sunlightshould result in a more rapid loss of photosyn-

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FIG. 3. Changes in chlorophyll a (0) and opticaldensity at 480 nm (carotenoids) (A) in (A) an Aphan-izomenon-dominated population, June 1976, exposedto 900 pE m2 s1 from an incandescent lamp and in(B) an Anabaena-Aphanizomenon-dominated pop'u-lation, June 1977, exposed to 1,800 to 2,100,E m-2s ' of sunlight.

TIME (MINUTES)

FIG. 4. Changes in (A) photosynthetic activity (car-bon fixation) (0) and assimilation number (A) andin (B) photosynthetic pigments (chlorophyll a) (0)and optical density at 480 nm (carotenoids) (A) in anAnabaena-Aphanizomenon population exposed tosunlight intensities of 1,800 to 2,100 ,uE m-2 s-', July1977. Each bar indicates one standard deviation.

thetic activity than of chlorophyll a. Such aneffect would not be expected to occur in popu-lations exposed to less than 50% of full sunlight.If such high light effects were occurring in thelake, one might expect to see a more rapiddecrease in the assimilation ratio (CO2 uptake tochlorophyll a) in the surface populations versusthose at depths of reduced light intensity. On anumber of days with suitable conditions,changes in such populations were observed andcompared under natural conditions. Samples ofnatural populations from suitably protectedareas of the lake were collected over half-daycycles at three different depths. The assimilationratios were measured at each depth over theperiod and consistently failed to show the ex-pected differences between surface populationsand those at depth (Fig. 6).Relationships to decline events. Table 1

and Fig. 2 show the periods of major decline intotal lake chlorophyll a. Also presented in Table1 are the dates during June, July, and August inboth years when meteorological conditions weresuitable for lake-wide bleaching of the algal pop-ulations. Field observations on such daysshowed that only a portion of the algae in theupper centimeter of the water column was ex-posed to light intensities high enough to cause

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LYTIC ORGANISMS AND PHOTOOXIDATIVE EFFECTS 503

bleaching. Algae below the upper centimeter areprotected from the light by shading in the highlyconcentrated surface scums. Estimates from in-tegrated water column chlorophyll a showedthat such exposed surface layers were likely torepresent 5% or less of the total lake chlorophylla. Because only a small percentage of the algalpopulation was exposed on any one day, it waslikely that a period of consecutive days withsuitable conditions would be necessary to causesignificant declines in chlorophyll a (Table 1).No such periods occurred in 1977. Four suitableperiods occurred in 1976, but only one of thosewas correlated with a chlorophyll decline: 1 to 6July. Although such a correlation is intriguing,the change was apparently due to a decline inPlanktosphaeria sp., a green alga. During thisperiod, the blue-green algal populations, mostlyAphanizomenon (which, based on our observa-tions, is very susceptible to high light damage),actually showed a slight increase. Thus the de-cline was not a result of photooxidative effects.Table 1 and Fig. 1 show that in both years

peak numbers of PFU occurred in late Julythrough mid August. In 1976, the peak in PFUoccurred in the 2 weeks after a 3-week decline inchlorophyll a (Table 1 and Fig. 2). The peak inPFU in 1977 occurred coincidentally with a 3-week period of decline in total lake chlorophylla (Table 1 and Fig. 2). In both years, algalpopulations were dominated by Microcystis sp.during the periods of peak PFU concentrationin the surface waters.

DISCUSSIONExposure to high light intensities, especially

under conditions of high P02 and low PCO2, willeventually lead to death as a result of photoox-idation. Accompanying this process, but appar-ently independent of it, is the phenomenon ofpigment bleaching (1, 6). In the strains tested byEloff et al. (6), 50% bleaching of the chlorophyllrequired 1 to 3.5 h. Lake Mendota populationswhich were tested under similar intensities re-quired 2.5 to 6 h of exposure to achieve a 50%reduction in chlorophyll a. Data from Eloff etal. (6) also showed that significant levels of celllysis, indicated by a 50% loss in turbidity, re-quired 2.5 to 4 h. In Lake Mendota, the appear-ance of opalescence and blue pigmentation inthe water, which indicated that a significantamount of lysis had occurred, was generally inthe afternoon, after 3 to 6 h of high light expo-sure. In the study by Eloff et al. (6), loss ofchlorophyll and decrease in turbidity were cor-related with loss of viability in the test strains.Because populations in Lake Mendota were ableto maintain chlorophyll a levels and cellular

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FIG. 5. Response of an Anabaena-Aphanizome-non-dominated population to exposure to varioushigh sunlight intensities, July 1977. No decline inchlorophyll was observed in the experiment.

integrity better than the laboratory strainstested by Eloff et al. (6) and Eloff (5), it is likelythat they are also more resistant to photooxi-dative death. Such a result is not unexpected.Eloff et al. (6) and Eloff (5) indicate that labo-ratory strains of blue-green algae appear to beless resistant than wild types because the resist-ance is lost by continued culturing under lowlight intensities. Although the Lake Mendotapopulations do appear to be resistant to pho-tooxidative death, experiments performed didshow that carbon fixation ability was rapidly lostby exposure to high light. This is in agreementwith observations made by Abeliovich and Shilo(1). The bleaching sequence observed in theLake Mendota populations differed from thatseen by Eloff et al. (6) in laboratory strains. Wealways observed a more rapid loss of chlorophylla than phycocyanin in the lake populations,whereas Eloff et al. showed bleaching sequences

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TIME OF DAY

FIG. 6. Response ofa Microcystis-Anabaena-dominatedpopulation in situ. Populations were monitored atthe surface (-----), at 0.25 m ( ), and at 1.0 m (. ). The inset shows the light, temperature, and oxygenconditions at each depth over the period. Surface light intensities (100%o) were 1,000 piE m2 s ' at 0820 h, 1,850,uE m-2 s- at 1120 h, and 1,900 ILE m-2 s- at 1420 h.

TABLE 1. Temporal relationships between decline periods and inhibitory factorsaNet decline

Decline period, total Dominant genera rate (% of Dates suitable for bleachinglake chlorophyll original

week-')1976

27 June-6 July Aphanizomenon 22 6-8, 20-22 June11 July-2 August Aphanizomenon, Anabaena 20 1-6, 8, 12, 13, 24, 25, and 27 July; 1-

3, 29 August1977

1-8 July Aphanizomenon, Anabaena 26 7, 14, 17, 25 June19-26 July Aphanizomenon, Anabaena 19 13, 22, 23 July1-23 August Anabaena, Microcystis 19 18, 24, 29 August

a Peak PFU periods for 1976 and 1977 were 26 July to 10 August and 2 to 14 August, respectively.

in which both pigments were lost at similarrates.

Besides the apparent resistance in the algalpopulations, physical factors also tend to limitwidespread exposure damage in the lake. Self-shading is important in all situations, since thesurface accumulations of blue-green algae willlimit light penetration. Wind turbulence is an-

other important physical factor in large lakes.Circulation of the epilimnion prevents long pe-

riods of surface exposure. In Lake Mendota, suchphysical factors in association with the relativelyresistant populations tend to minimize photoox-idative effects. This fact is supported by the lackof evidence for significant differences in thechanges of assimilation ratios in the surface pop-ulations versus deeper populations (Fig. 6).Organisms able to lyse the indicator Anacystis

sp. reached peak numbers in late July to mid-August. Lower numbers of PFU in the hypolim-nion (Fig. 1) may have been due to anaerobicconditions. Most of the known lytic bacteria areobligately aerobic and would have been unableto grow in the hypolimnion. Vertical profiles ofchlorophyll a during the summer showed that80% or more of the blue-green algal biomass wasconcentrated in the upper 10 of the water col-umn. This biomass distribution for the hostswould have also favored higher concentrationsof PFU in the epilimnion.The peak values of PFU in the present study,

1 x 103 to 5 x 103 PFU ml-', are about 10-foldhigher than values obtained by Daft et al. (4) insome English reservoirs and by J. A. McMillan(M. S. thesis, University of Wisconsin, Madison,1973) in Lake Mendota. This may be due to the

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different assay procedure used in the presentstudy. Because incubations for plaque develop-ment were done in the dark in the present study,the indicator organism may have been moresusceptible to lysis. The lack of any growth inthe indicator lawn also favors higher plaquenumbers. In the present study, we have alsonoted high numbers of PFU in the lake sedi-ments. In contrast, Daft et al. (4) normally ob-served values of 10 PFU mn-l or less in sediment.Once again, the difference in assay conditionsmay partially explain the results.The characteristics of the lytic bacterial iso-

lates were similar to those seen in Cytophagaspecies capable of lysing blue-green algae. Thecharacteristics of obligately aerobic growth witha temperature optimum around 300C have beencommonly observed in freshwater isolates.Strong pigmentation seen in the two isolatesalso appears to be common (Stauffer, Ph.D.thesis). It is interesting to note in relation topigmentation that Snow and Fred in 1926 (14)showed that the percentage of pigmented iso-lates in total bacterial counts of the lake in-creased during the summer. Thus, these workersmay have been observing a phenomenon relatedto the development of lytic bacterial species inthe lake during the summer. Based on totalbacterial counts by epifluorescence (C. Pedros-Alios, personal communication), total numbersof bacteria increased only 3- to 10-fold in bothyears during the same period when lytic countsincreased by 1,000-fold in the surface waters.Thus, there was a definite preferential growth ofthe lytic organisms in the surface waters in thepresence of the blue-green algae.The significance of the lytic activity in rela-

tionship to the midsummer chlorophyll declineevents is difficult to evaluate. Daft et al. (4)observed peaks of PFU associated with peaks inblue-green algal chlorophyll, and such peaks de-clined rapidly and coincidentally. The cycle gen-erally covered about a 1- or 2-week period. Inspite of these observed temporal correlations,Daft et al. (4) did not feel that any definiteconclusions could be made about the influenceof lytic activity on the blue-green algal popula-tions. In the present study, changes in total lakechlorophyll a and PFU were even less tempo-rally correlated. Thus, although lytic organisms

may play a primary role in causing declines, nodefinite conclusions can be made. The questionalso remains as to whether the lytic bacteria areacting as pathogens, and thus are a primarycause for declines, or are acting as saprophytes,decomposing dead algal material resulting fromother primary processes.

LITERATURE CITED

1. Abeliovich, A., and M. Shilo. 1972. Photooxidativedeath in blue-green algae. J. Bacteriol. 111:682-689.

2. Bush, E. T. 1963. General applicability of the channelratios method of measuring liquid scintillation countingefficiencies. Anal. Chem. 35:1024-1029.

3. Canter, H. M. 1972. A guide to the fungi occurring onplanktonic blue-green algae. In T. V. Desikachary (ed.),Taxonomy and biology of blue-green algae, p. 145-158.University of Madras Press, Madras, India.

4. Daft, M. J., S. B. McCord, and W. D. P. Stewart. 1975.Ecological studies on algal lysing bacteria in freshwa-ters. Freshwater Biol. 5:577-596.

5. Eloff, J. N. 1978. The photooxidation of laboratory cul-tures of Microcystis under low light intensities, p. 95-100. In W. E. Krumbein (ed.), Environmental biogeo-chemistry and geomicrobiology, vol. 1. The aquaticenvironment. Ann Arbor Science, Ann Arbor, Mich.

6. Eloff, J. N., Y. Steinitz, and M. Shilo. 1976. Photooxi-dation of cyanobacteria in natural conditions. Appl.Environ. Microbiol. 31:119-126.

7. Fogg, G. E. 1969. The physiology of an algal nuisance.Proc. R. Soc. London Ser. B 173:175-189.

8. Gerloff, G. C., G. P. Fitgerald, and F. Skoog. 1952.The mineral nutrition of Microcystis aeruginosa. Am.J. Bot. 39:26-32.

9. Granhall, U. 1972. Aphanizomenon flos-aquae: infectionby cyanophages. Physiol. Plant. 26:332-337.

10. Padan, E., and M. Shilo. 1973. Cyanophages-virusesattacking blue-green algae. Bacteriol. Rev. 37:343-370.

11. Redhead, K., and S. J. L. Wright. 1978. Isolation andproperties of fungi that lyse blue-green algae. Appl.Environ. Microbiol. 35:962-969.

12. Reynolds, C. S. 1975. Interrelationships of photosyn-thetic behavior and buoyancy regulation in a naturalpopulation of a blue-green alga. Freshwater Biol. 5:323-338.

13. Shoaf, W. T., and B. W. Lium. 1976. Improved extrac-tion of chlorophyll a and b from algae using dimethylsulfoxide. Limnol. Oceanogr. 21:926-928.

14. Snow, L. M., and E. B. Fred. 1926. Some characteristicsof the bacteria of Lake Mendota. Trans. Wisc. Acad.Sci. Arts Lett. 22:143-154.

15. Stewart, W. D. P., and M. J. Daft. 1976. Algal lysingagents of freshwater habitats, p. 63-90. In S. Skinnerand J. G. Carr (ed.), Symposium Series, Society forApplied Bacteriology, Microbiology in Agriculture,Fisheries, and Food. Academic Press Inc., New York.

16. Strickland, J. D. H., and T. R. Parsons. 1972. A prac-tical handbook of seawater analysis. Fish. Res. BoardCan. Bull. 167.

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