GEOLOGICAL SURVEY CIRCULAR 848-E 

Microbiology of the 

Aquatic Environment 

Microbiology of the 

Aquatic Environment 

Edited by Phillip E. Greeson 

Briefing Papers on Water Quality 

GEOLOGICAL SURVEY CIRCULAR 848-E 

1981 
United States Department of the Interior 

JAMES G. WATT, Secretary 

Geological Survey 

Doyle G. Frederick, Acting Director 

Library of Congress Cataloging in Publication Data 

Main entry under title: 

Microbiology of the aquatic environment. 

(Briefing papers on water quality) (Geological Survey circular 848-E) 

Bibliography: p. 

Supt. of Docs. no.: I 19.412:848-E 

Contents: The ecological role(s) of aquatic micro-organisms in lakes and reservoirs I by Bruce L. KimmelEffects of bacteria on the chemical and physical state of iron I by Gail E. Mallard-Micro-organisms in 

stormwater I by Gail E. Mallard. 

1. Freshwater microbiology-Addresses, essays, lectures. 2. Microbial ecology-Addresses, essays, lectures. 3. Iron bacteria-Addresses, essays, lectures. 4. Runoff-Microbiology-Addresses, essays, 

lectures. I. Greeson, Phillip E. II. Kimmel, Bruce L. Ecological role(s) of aquatic micro-organisms in 

lakes and reservoirs. 1981. III. Mallard, Gail E. Effects of bacteria on the chemical and physical state of 

iron. 1981. IV. Mallard, Gail E. Micro-organisms in stormwater. 1981. V. Series. VI. Series: 

Geological Survey circular ; 848-E. 

QE75.C5 no. 848-E [QR105.5] 557.3s 81-607912 [576'.15'09169] AACR2 

Free on application to Distribution Branch, Text Products Section, 

U. S. Geological Survey, 604 South Pickett Street, Alexandria, VA 22304 
FOREWORD 

In August 1974, the Water Resources Division of the U.S. Geological Survey 

introduced the first of a series of briefing papers that were designed to increase 

the understanding of its employees of the significance of various aspects of 

water quality. Numerous briefing papers have been prepared under the direction 

of the Quality of Water Branch. Other papers will be prepared as the need arises. 

Each paper addresses a separate topic and is written in a nontechnical, easy-tounderstand manner for distribution within the organization. 

Because of the favorable reception that the papers have received and their apparent effectiveness in accomplishing the objective stated above, it would appear 

that their wider distribution would serve a useful purpose. It is hoped that a wide 

range of persons, including those interested in the quality of our Nation's water 

resources but who have little or no technical training, will find value in reading 

the papers. Furthermore, it is hoped that the papers will be suitable for supplemental reading in secondary education programs and in beginning collegelevel courses. 

The U.S. Geological Survey plans to publish several U.S. Geological Survey 

Circulars that contain compilations of briefing papers on particular aspects of 

water quality. This fifth Circular contains three papers describing the microbiology of the aquatic environment. 

III 

Philip Cohen 

Chief Hydrologist 

CONTENTS 

Page 

Foreword ------------------------------------- III Effects of bacteria on the chemical and physical state of 

Abstract 

The ecological role(s) of aquatic micro-organisms in 

lakes and reservoirs, by Bruce L. Kimmel ---------

E1 iron, by Gail E. Mallard------------------------

Micro-organisms in stormwater: A summary of recent 

3 investigations, by Gail E. Mallard----------------

ILLUSTRATIONS 

Page 

FIGURE 1. Flow diagram of the grazer and detritus pathways ------------------------ E4 

2. Formula showing the "equilibrium" relationship between autotrophic and 

heterotrophic processes ------------------------------------------- 5 

3. Flow diagram of processes involved in the decomposition and mineralization of 

organic matter in aquatic environments ------------------------------ 6 

4. Patterns of carbon cycling in lakes and reservoirs ------------------------- 8 

5. Patterns of nitrogen cycling in lakes and reservoirs ----------------------- 9 

6. Patterns of sulfur cycling in lakes and reservoirs -------------------------- 9 

7. Diagram of environmental phosphorus cycling---------------------------- 10 

8. Flow diagram of organic carbon conversions of potential importance in pelagic 

foodwebs ------------------------------------------------------ 10 9. A classical view of energy and nutrient flow in a lake or reservoir ecosystem ___ 11 

10. A more complex view of energy and nutrient flow in a lake or reservoir 

ecosystem ------------------------------------------------------ 11 

11. Graph showing the fields of stability for solid and dissolved forms of iron as a 

function of Eh and pH at 25 ° C and 1 atmosphere of pressure ------------- 14 

12. Diagram showing the direct and indirect changes in iron resulting from microbial 

activity -------------------------------------------------------- 16 

13. Hydrograph and graphs of densities of fecal coliform and fecal streptococci 

bacteria in a small stream near Houston, Texas, during a storm----------- 28 

14. Graphs of bacterial populations in runoff samples taken at stormwater outfall, 

Burlington, Ontario ---------------------------------------------- 29 

15. Graphs showing persistence of selected bacterial strains in stormwater ________ 32 

TABLES 

Page 

TABLE 1. Concentration of essential elements for plant growth in the living tissues of 

freshwater plants, in mean world river water, and the approximate ratio 

of elemental concentrations required to those available----------------- E6 

2. General categories of microbially mediated carbon transformations----------- 7 

3. General categories of microbially mediated nitrogen transformations --------- 7 

4. General categories of microbially mediated sulfur transformations ----------- 7 

5. Densities of indicator bacteria in stormwater ----------------------------- 27 

6. Densities of pathogens in storm water from Baltimore, Maryland ------------- 29 

v 

Page 

E13 

23 

BRIEFING PAPERS ON WATER QUALITY 

Microbiology of the Aquatic Environment 

Edited by Phillip E. Greeson 

ABSTRACT 

This is t~e fifth of several compilations of briefing papers on 

w~ter quahty prepared by the U.S. Geological Survey. Each 

bne:fing paper is prepared in a simple, nontechnical, easy-tounderstand manner. This U.S. Geological Survey Circular contains three papers on the microbiology of aquatic environment. 

Briefing papers are included on "The ecological role(s) of aquatic 

micro-organisms in lakes and reservoirs," "Effects of bacteria 

on the chemical and physical state of iron," and "Microorganisms in stormwater: A summary of recent investigations." 

El 

The Ecological Role(s) of Aquatic Micro-organisms 

in Lakes and Reservoirs 

By Bruce L. Kimmel 1 

ABSTRACT 

Micro-organisms form vital links in the structure of aquatic 

ecosystems by virtue of their roles in organic matter production 

and decomposition, nutrient uptake and regeneration, foodweb 

transfers, and biogeochemical transformations. By the nature 

of. their bi~logical activities, autotrophic and heterotrophic 

m1cro-orgamsms are sensitive indicators of the ecological and 

water-quality status of aquatic environments. However, only by 

thoroughly understanding how micro-organisms function in 

healthy aquatic ecosystems can we hope to recognize or accurately predict their response to water-quality changes. 

INTRODUCTION 

Much attention in the field of water quality is 

f?cused on micro-organisms as indicators of poilubon (for example, fecal coliform and fecal streptococci bacteria), threats to public health (for example, pathogenic bacteria and viruses), or sympto~s of water-quality deterioration (for example, 

nuisance algal blooms). Because characteristics 

common to water-quality problems are readily 

noticeable, the public often receives the mistaken 

impression that the presence of micro-organisms 

signals the demise of a body of water. As an example, the term "algae" may connote surface scums 

and foul odors to the layman, while "bacteria" implies disease-causing contamination. In fact, such 

~xtreme condi~ions are rare, and usually occur only 

In systems which have been altered significantly by 

excessive loading of nutrients, organic matter, 

and/or toxic contaminants. Under typical circumstances, micro-organisms such as algae and 

bacteria form vital links in the structure of aquatic 

ecosystems, and indeed are the operative factors in 

organic matter production, decomposition, and 

1 Biological Station and Department of Zoology, University of Oklahoma, Norman, 

Oklahoma. 

E3 

nutrient regeneration. As professionals in the field 

of water quality and communicators of waterquality information to other agencies and to the 

general public, it is important to maintain a 

perspective in regard to the ecological significance 

of aquatic micro-organisms. The purpose of this 

discussion, therefore, is to review the ecological 

roles of aquatic micro-organisms in lakes and 

reservoirs. 

AN OVERVIEW OF ECOSYSTEM STRUCTURE 

Before specifically addressing aquatic microorganisms and their ecological roles, let us briefly 

consider the basic structure of lentic (that is, lakelike or standing-water) ecosystems. Photosynthetic production usually forms the organic matter 

base of aquatic foodwebs. Organic matter formed 

photosynthetically by primary producers (for example, algae) is consumed by herbivores (for example, zooplankton), which are fed upon by higher 

consumers (carnivores; for example, fish). 

Predatory transfers, the ingestion and assimilation 

of living primary producers by herbivores and herbivores by carnivores, collectively are referred to 

as the "grazer pathway." All of these foodweb 

levels produce excreted or defecated material and 

dead organisms which are utilized by "decomposers" (for example, bacteria and fungi). Decomposers promote the breakdown of organic 

materials to inorganic compounds, and therefore, 

generally are assigned the role of nutrient 

regeneration. Nonpredatory transfers, which involve the use of nonliving organic matter by 

decomposers, collectively are termed the "detritus 

pathway" (fig. 1). 
GRAZER 

PATHWAY 

LIGHT ENERGY\ 

C0 ~ 

2 ~ PRIMARY 

0 2 ~PRODUCERS 

Phytoplankton and 

higher plants 

! 

C02 ~ PRIMARY 

02 ~CONSUMERS 

Zooplankton, and 

other herbivores 

whose nutrition is 

mediated by 

micro-organisms 

j 

DETRITUS 

PATHWAY 

DETRITUS 

Dead cells, 

carcasses, feces, 

excretion products 

1 

DECOMPOSERS ~~----~cg2 

Bacteria, fungi and 

detritus feeders 

C02 ~SECONDARY:;~::::::.::.::.::~----_-_-_-_:-_:' ________ CONSUMERS""'- C02 

02 ~CONSUMERS '""-.._. ---- 02 

Carnivores; 

e.g., fish 

I 

I 

I 

I 

t 

Organisms which prey 

on decomposers 

FIGURE 1.-The grazer and detritus pathways (modified from Campbell, 1977). 

Although the flow of energy and materials 

through both grazer and detritus pathways is 

recognized, the relative importance of the two 

pathways in various types of ecosystems appears 

not to be. It is often generalized that the grazer 

pathway dominates ecosystem energy flow in 

grasslands and pelagic (open-water) systems, while 

the detritus pathway is more important in forests, 

marshes, and streams. However, this generalization should not be accepted unquestioningly. 

Although the magnitude of detritus accumulation 

is obvious in some terrestrial and semiterrestrial 

systems in which litter (for example, fallen leaves 

and dead marsh grass) is readily visible, it is much 

less apparent in the open waters of lakes and reservoirs where the "litter" (for example, dead algal 

cells and zooplankton fecal pellets), as well as many 

of the producers and consumers, is of microscopic 

size. 

This discussion considers primarily those aquatic 

micro-organisms which fall into the "Producer" and 

"Decomposer" categories described above; 

E4 
however, as will be seen, some of these microorganisms also perform roles assignable to "Herbivores" and "Consumers." Our discussion will 

focus on two fundamental aspects of the ecological 

roles of micro-organisms in lakes and reservoirs: 

(1) Algae as primary producers, and bacteria as decomposers and nutrient regenerators; 

(2) Bacteria and fungi as particle producers and potential food sources for higher consumers. 

ORGANIC MATTER PRODUCTION AND 

DECOMPOSITION 

Algae and certain bacteria produce organic matter by the process of photosynthesis, which is the 

formation of organic compounds from carbon dioxide (C02) and other inorganic nutrients by way of 

light energy and the chlorophyll pigment in plants. 

Some bacteria produce organic compounds by 

chemosynthesis, in which energy-yielding chemical 

reactions, rather than sunlight, provide the energy 

necessary for C02 fixation. Both photosynthesis 

and chemosynthesis result in the formation of 

organic material from inorganic components, and, 

therefore, are considered to be autotrophic 

(primary production) processes. 2 Although some 

bacteria are autotrophic, most are heterotrophic 

and like other heterotrophs (for example, herbivores and higher consumers) they grow by 

assimilating organic materials previously synthesized by autotrophs. The metabolic processes of 

all organisms are energy consumptive, and require 

adenosine triphosphate (ATP, formed during 

cellular respiration) as their "energy currency." All 

living organisms, in order to continue living, must 

conduct cellular respiration, which is in some ways 

the reverse of photosynthetic carbon fixation (fig. 

2). Organic matter decomposition, a collective 

term describing the net conversion of organic 

material back to inorganic compounds (for example, C02, H20, and nutrients), occurs by virtue of 

the respiratory activities of those heterotrophic 

micro-organisms using the organic material for 

growth. 

Carbon dioxide and oxygen are reactants in the 

photosynthesis-respiration equilibrium (fig. 2), and 

2 However, most chemosynthesis results from energy released in the oxidation of 

reduced comp_ounds (S- 2

' NH, +, CH4) derived from the decomposition of previously 

formed orgam? matter, and thus is actually a special type of secondary production. A 

notable exception, where chemosynthesis appears to be a true autotrophic process, is 

represented by the recently discovered Galapagos Rift hydrothermal vent 

~co~ystems. Here, chemosynthetic bacteria fix C02 with energy derived from the oxIdatiOn of geothermically reduced sulfur compounds emitted from the ocean-floor 

vents (see Karl and others, 1980). 

E5 

PRODUCTION & DECOMPOSITION 

C02 + H20 + Nutrients + Energy 

Autotrophic 

Production 

Respiration, 

Decomposition 

FIGURE 2.-The "equilibruim" relationship between autotrophic (photosynthesis, chemosynthesis) and 

heterotrophic (respiration, decomposition) processes. 

The photosynthetic conversion of light energy into 

chemical potential energy results in nonequilibrium concentrations of carbon, nitrogen, sulfur, and phosphorus 

in reduced organic compounds of high potential energy. 

Heterotrophic processes tend to restore the equilibrium 

by decomposing organic materials by way of energyyielding oxidation-reduction reactions. 

the balance or imbalance of these processes within 

a water body can have profound chemical and 

biological consequences for the entire system. The 

photosynthetic production of oxygen is restricted 

to the upper, lighted regions of the water column 

(the euphotic zone). Respiration is not directly 

light-dependent, but is dependent on temperature. 

Respiration and organic matter decomposition occur through the water column, and depending on 

the amount of organic matter present and the extent of wind-mixing, can deplete the dissolved oxygen supply in aphotic (unlighted) layers. The 

chemical and biological conditions associated with 

such oxygen depletion (for example, increased 

nutrient release from sediments, exclusion of 

oxygen-dependent fauna, and less complete 

decomposition of sedimented organic matter) are 

marked, and often characteristics of lake trophic 

status. In oligotrophic (nutrient-poor, unproductive) lakes, insufficient organic matter is produced 

for decomposition processes to affect available oxygen levels to any significant extent. In contrast, 

organic matter decomposition in eutrophic 

(nutrient-rich, productive) lakes can rapidly 

deplete dissolved oxygen in aphotic parts of the 

water column. 
MICROBIAL NUTRIENT 

TRANSFORMATIONS AND 

REGENERATION 

Thus far, organic matter production and decomposition have been discussed primarily in terms of 

carbon dioxide, water, and oxygen exchanges. 

However, living organisms are not composed entirely of C, H, and 0, but also contain small 

amounts of many other elements. Table 1 compares the elemental composition of living 

freshwater plants to the environmental availability 

of elements for plant growth (that is, demand-vssupply). Phosphorus, nitrogen, and carbon are the 

least available elements relative to plant growth 

requirements, and thus are most likely to limit 

biological productivity in aquatic systems. 

Although essential nutrients are incorporated into organic compounds during organic matter synthesis, they are regenerated as a result of excretion processes and organic matter decomposition. 

Organic matter decomposition (fig. 3) occurs by: 

(1) Hydrolysis of large (high molecular weight) 

organic polymers into smaller (low molecular 

weight) compounds; and 

(2) Mineralization of these smaller organic 

molecules to inorganic compounds (for example, 

H2S, NH4+, and P04- 3). Bacterial decomposition of 

organic materials accounts for a large fraction of 

the total nutrient regeneration which occurs; 

however, organisms such as phytoplankton, 

zooplankton, and fish excrete NH4 +, P04 - 3, and 

low molecular weight organic compounds, and 

therefore are also important nutrient 

regenerators. 

In an oxygenated environment, organic matter 

exists in a chemically reduced state (low Eh, high 

potential energy) relative to most of its inorganic 

components (fig. 2). The oxidation of certain 

mineralization products (for example, S-2-8°-

804-2, NH4=-N02--NOs, and CH4-C02) provides the energy for bacterial chemosyntheses. The 

basic types of microbially mediated transformations important in carbon, nitrogen, and sulfur 

cycling are summarized in tables 2, 3, and 4; and 

integrated into the overall nutrient cycling patterns for these elements in lake systems in figures 

4, 5, and 6. 

Phosphorus does not undergo the oxidationreduction transformations characteristic of C, N, 

and S cycles (fig. 7). Inorganic phosphorus occurs 

in aquatic environments primarily as orthophosphate (P04 - 3), and is incorporated directly by 

TABLE 1. -Concentrations of essential elements for plant growth 

in the living tissues of freshwater plants (demand), in mean 

world river water (supply), and the approximate ratio of 

elemental concentrations required to those available (that is, 

demand:supply ratio) (after Vallentyne, 1971,.) 

Element 

Oxygen ...... . Hydrogen .... . Carbon ...... . Silicon ...... . Nitrogen .... . Calcium ..... . Potassium ... . Phosphorus .. . 

Magnesium .. . Sulfur ....... . Chlorine ..... . Sodium ...... . Iron ........ . Boron ....... . Manganese ... . 

Zinc ........ . Copper ...... . Molybdenum .. Cobalt ....... . 

Symbol 

0 

H 

c 

Si 

N 

Ca 

K 

p 

Mg 

s 

Cl 

Na 

Fe 

B 

Mn 

Zn 

Cu 

Mo 

Co 

Demand by 

plants(%) 

80.5 

9.7 

6.5 

1.3 

.7 

.4 

.3 

.08 

.07 

.06 

.06 

.04 

.02 

.001 

.0007 

.0003 

.0001 

.00005 

.000002 

89 

11 

ORGANIC MATTER DECOMPOSITION 

Supply in water(%) 

.0012 

.00065 

.000023 

.0015 

.00023 

.000001 

.0004 

.0004 

.0008 

.0006 

.00007 

.00001 

.0000015 

.000001 

.000001 

.0000003 

.000000005 

Demand: 

Supply ratio 

1 

1 

5,000 

2,000 

30,000 

<1,000 

1,300 

80,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

<1,000 

ORGANISMS, 

FECES 

fLARGEl 

~ fSMALLl ... ~ 

~ ~ t 

Excretion 

Pmdocl•~ 

'----------.!~- INORGANIC NUTRIENTS ~ (

C02 , NH 3, P04 , H2S, CH4 ) 

S04 , N03 , etc. 

Jtt' ~ 

FIGURE 3. -Processes involved in the decomposition and 

mineralization of organic matter in aquatic environments. POM =particulate organic matter, 

DOM=dissolved organic matter, MW =molecular 

weight (modified from Sorokin and Kadota, 1972). 

algae and bacteria into phospholipids, nucleic 

acids, and ATP. Although phosphorus turnover 

time (the time required for phosphorus assimilation and regeneration) is often only a matter of 

minutes in surface waters during the summer 

(Lean, 1973), sedimentation of organic particles 

and precipitation of inorganic phosphorus with 

iron and manganese complexes can cause 

phosphorus depletion in the mixed layers of lakes 

E6 
TABLE 2.-General categories of microbially mediated carbon 

transformations 

[E=energy] 

MICROBIAL TRANSFORMATIONS: 

CARBON 

1. Inorganic C Assimilation (photosyn., chemosyn.) 

2. Respiration, Aerobic Decomposition 

3. Anaerobic Decomposition 

TABLE 3.-General categories of microbially mediated nitrogen 

transformations 

[E=energy] 

MICROBIAL TRANSFORMATIONS: 

NITROGEN 

1. Inorganic N Aaslmllatlon 

2. N2 Fixation 

3. Ammonification 

4. Nitrification 

02 

NH 3 ~ N02 

+E 

~ 

+E 

C02 (CH2 0) C0

~ 

2 (CH20) 

5. Denitrification 

N03 ---+ N2 , N20 + E 

(CH20) 

~ C02 + H20 

E7 

TABLE 4.-General categories of microbially mediated sulfur 

transformations 

[E=energy] 

MICROBIAL TRANSFORMATIONS: 

SULFUR 

1. Inorganic S Assimilation 

Sinorg. + E ~ (-SH-) 

2. Organic Matter Decomposition 

3. Inorganic S Reduction (anaerobic) 

4. Inorganic Sulfur Oxidation 

a. Chemosynthesis 

and reservoirs. Naturally low levels of available 

phosphorus in aquatic environments, the essentiality of phosphorus for plant growth, and the high 

phosphorus concentration in certain byproducts of 

our modern society (for example, fertilizers, detergents, and municipal sewage) make phosphorus an 

especially important element in regard to water 

quality. In short supply, phosphorus availability 

can severely limit biological productivity, while an 

overabundance of phosphorus can result in 

nuisance conditions concomitant with accelerated 
CARBON CYCLE IN A LAKE ECOSYSTEM 

Certain fungi 

and bacteriaCytophagaMethane.--

oxidizing 

bacteria 

Heterotrophic 

aerobic 

bacteria 

Heterotrophic 

/ aerobic bacterium 

Bact. alginovorum 

Hydrogen 

oxidizing bacteria 

Gases from 

sediment 

Methanobacterium 

omelianskii 

Methanobacterium -- sohngenii, 

Methanococcus 

mazei 

-- Anaerobic decompo-

-- sition of plant 

remains in the 

sediment by 

heterotrophic 

bacteria 

FIGURE 4.- Patterns of carbon cycling in lakes and reservoirs (from Brock, 1966, after 

Kuznetsov, 1959). 

eutrophication. 3 Since phosphorus is often the 

least available inorganic nutrient in aquatic environments (that is, relative to demand, see table 

1), extensive efforts have been undertaken in the 

s Eutrophication is a natural process involving the slow accumulation of nutrients 

and organic matter, and gradually increased productivity in an aquatic system. Accelerated eutrophication results from excessive nutrient loading from the inflow of 

municipal wastes and/or drainage from fertilized agricultural land, and often is 

associated with nuisance algal blooms, taste and odor problems, and severe O!Xygen 

depletion accompanied by fishkills. 

past decade to minimize phosphorus loading of 

natural waters. 

Although biogeochemical cycles often are 

discussed individually by element for the sake of 

simplicity, it should be realized that in nature they 

are inseparable. Carbon, nitrogen, sulfur, 

phosphorus, and other essential elements are 

assimilated simultaneously during organic matter 

synthesis, and similarly, are released simulE8 
NITROGEN CYCLE IN A LAKE ECOSYSTEM 

Sedimentation 

Phytoplankton 

and bacteria 

(inorganic 

nitrogen 

assimilation) 

FIGURE 5.-Patterns of nitrogen cycling in lakes and reservoirs (from Brock, 1966, after Kuznetsov, 1959). 

taneously during organic matter decomposition. 

Because the availability of certain elements (for 

example, P, Fe, and Mo) influences the uptake or 

transformations of others {C, N), the elemental 

cycles are highly interdependent. 

BACTERIA AS PARTICLE PRODUCERS AND 

POTENTIAL FOOD SOURCES FOR HIGHER 

TROPHIC LEVELS 

The photosynthetic conversion of inorganic compounds of organic matter is only the initial process 

in a sequence of trophic interactions collectively 

referred to as the "foodweb." The most familiar 

foodweb sequence (primary producer-grazerhigher consumer; the grazer pathway) may present 

an oversimplified view of trophic interactions in 

lakes and reservoirs by ignoring nonpredatory 

transfers (primary producer-detritus-bacteriagrazer-higher consumer; the detritus pathway). 

More specifically, the conversion of dissolved 

organic compounds (derived from algal extracellular excretion or cell lysis, zooplankton exE9 

Photosynthetic and/ 

chemosynthetic 

sulfur bacteria 

HYPOLIMNION 

SULFUR CYCLE IN A LAKE ECOSYSTEM 

Rainwater and other 

influents 

FIGURE 6.- Patterns of sulfur cycling in lakes and reservoirs 

(from Brock, 1966, after Kuznetsov, 1959). 

cretion, and/or allochthonous sources4) to organic 

particles by way of bacterial uptake and growth 

represents a potentially important mechanism for 

reintroducing otherwise unavailable (that is, 

unharvestable) dissolved organic matter to the 

foodweb (fig. 8). For pelagic marine systems, 

Pomeroy (197 4) indicated that the producerdetritus-bacteria-grazer sequence could provide at 

least 30 percent more energy to higher trophic 

levels than estimated from consideration of only a 

producer-grazer pathway. 

However, whether bacterial secondary production provides a significant source of particulate 

organic matter for planktonic grazers is a controversial question at present. Free-living (unattached) bacteria are probably too small ( < 1~tm) to 

be efficiently harvested by most zooplankton; 

however, bacteria colonizing suspended detritus or 

otherwise associated with aggregates of detritus, 

algal cells, and/or silt particles may be grazed 

readily by virtue of their greater effective particle 

size. Planktonic ciliates (a type of Protozoa) may 

help bridge the "particle-size gap" between freeliving bacteria and grazers by ingesting bacteria 

and then becoming prey for filter-feeding 

zooplankton (Porter and others, 1979). The occurrence of attached-vs-unattached bacteria in pelagic 

• The term "allochthonous" refers to organic matter originating from outside the 

lake (e.g., material from the watershed). In contrast, "autochthonous" material is that 

produced within the lake (for example, phytoplankton production). 
Deposition 

Industrial processing 

FIGURE 7.-Environmental phosphorus cycling 

(after Campbell, 1977). 

environments appears to be highly variable, both 

temporally and among systems, as numerous investigators have reported contradictory results 

and conclusions. 

Altho~gh algal p~otosynthesis is the primary 

mechanism of organic matter production in most 

lakes and reservoirs, the potential significance of 

heterotrophic micro-organisms as both secondary 

producers (by conversion of dissolved organic matter to a particulate form) and consumers (by 

r~spiration) of organic matter should be recog-

~Ized. Indeed,. bacterial production may be of particular ecological importance in many reservoirs 

for the following reasons: 

(1) Reservoirs usually receive large quantities of allochthonous organic materials by tributary inflow from their watersheds. Some labile (biologically available) fraction of this allochthonous input may provide major supplements 

of organic substrates for bacterial production· 

(2) Erosion products from the watershed (for ex: 

ample, suspended silt and clay particles) are 

potential sites for adsorption of dissolved 

organic compounds and bacterial atC02 , INORGANIC 

~NUTRIENTS 

ALLOCHTHONOUS 

~SOURCES 

r---------.., 

I I 

-='"~~::.:..::..:.....:..:::::=~: DISSOLVED I ! ORGANICS i ._ ___________ J 

l 

G ? ? ? olf•rf''b',, 

CO ~--....:...--·--....:.--- »BACTERIA 

2 ";. .. ·~6 t 

~ ~C02 

FIGURE 8.-Organic carbon conversions of potential importance in pelagic foodwebs. Some photosyntheticproduced algal biomass is harvested directly by filterfeeding grazers. Dissolved oxygen compounds (present 

by virtue of allochthonous input, zooplankton excretion, 

algal excretion, or cell lysis) are unavailable to grazers, 

but may be returned to a particulate state by bacterial 

uptake. However, the relative availability of free-living 

bacteria and bacterial-detrital aggregates to pelagic 

grazers is uncertain. 

tachmentlaggregation, and thereby may constitute a harvestable source of organic matter 

for planktonic grazers. Alternatively, such 

microbial-detrital aggregates may sediment 

rapidly from the water column and result in 

the transport of organic matter and nutrients 

to benthic communities; 

(3) In turbid, well-mixed reservoirs, phytoplankton 

production is often limited more by available 

light than by nutrients. In contrast to 

photosynthesis, bacterial production is not 

directly light-dependent and, therefore, may 

form a predominant fraction of the particulate 

organic matter supply available to grazers in 

light-limited systems. Especially for reservoir 

foodwebs, in which bacteria may be of particular importance, an improved understanding of algal and bacterial production and 

respiration processes, and the relative 

availability of these primary and secondary 

particle producers (that is, phytoplankton and 

bacteria, respectively) to grazers is required. 

SUMMARY 

The classical trophic-dynamics approach 

(Lindeman, 1942) persists as an ecological 

E10 
paradigm even though the nondiscrete nature of 

trophic levels was recognized by Lindeman and is 

well known to most ecologists. Although useful as 

a conceptual framework, the trophic level approach may lead to a simplistic view of energy and 

materials flow in nature (Wetzel and others, 1972; 

Rigler, 1975; Goldman and Kimmel 1978). Most 

aquatic organisms can be considered as article producers (by either autotrophic or heterotrophic production of protoplasm), consumers (assimilation of 

dissolved or particulate organic compounds), and 

decomposers (by conversion of organic materials to 

inorganic materials by respiration and excretion). 

Nutrient regeneration, the classical role of 

bacteria and fungi, takes place at virtually every 

level of the food web. Figures 9 and 10 summarize 

contrasting views of energy and nutrient flow in 

lakes and reservoirs. Particularly for reservoirs, 

our present level of understandings of ecosystems 

dynamics is inadequate to determine the relative 

accuracy of either view. 

Micro-organisms, by virtue of their pivotal roles 

in organic matter production and decomposition, 

foodweb transfers by dissolved to particulate 

organic matter conversions, nutrient uptake and 

regeneration, and biogeochemical transformaPHYTOPLANKTON G./v ... ~, l ~ eAO"CUCA" HIGHER DETRITUS 

CONSUM'"S J 

MICROBIAL 

DEGRADATION 

t SEDIMENTATION 

DETRITIVORES BENTHIC 

~ 

FIGURE 9.-A classical view of energy and nutrient flow in a 

lake or reservoir ecosystem. As symbolized by the 

relative size of the arrows, most of the energy and 

nutrients flow through the grazer pathway, and the 

detritus pathway is assigned the role of nutrient 

regeneration (after Goldman and Kimmel, 1978). 

PARTICULATE ORGANIC 

MATTER SUPPLY 

(ALLOCHTHONOUS & AUTOCHTHONOUS) 

PLANKTONIC PARTICULATE 

FILTER-FEEDERS DETRiTUS 

(NUTRIENTS) J ~ ! (NUTRIENTS) 

'-. HIGHER MICROBIAL p ..........__ CONSU\MERS COLONIZtTION 

PARTICLE 

AGGREGATION 

SEDIMEtTATION 

BENTHIC/ 

~-FIGURE 10.-A more complex view of energy and nutrient 

flow in a lake or reservoir ecosystem. Allochthonous inputs of dissolved and particulate organic materials 

directly enter the detritus pathway, and support 

bacterial growth and detritivores (detritus-feeders). 

Much of the primary production may also enter the 

detritus pathway directly via phytoplankton mortality 

unrelated to grazing. Adsorption of dissolved organic 

compounds, microbial colonization, and particle aggregation produces microheterotroph-detrital aggregates 

which are, by virtue of their particle size, available to 

planktonic grazers. Sedimentation of detrital aggregates 

provides a major energy and nutrient source for benthic 

detritivores which become prey items for higher consumers. Nutrient regeneration occurs at virtually every 

foodweb level, and only a small fraction of the total 

organic matter supply accumulates as permanent sediment (after Goldman and Kimmel, 1978). 

tions, are essential to the ecological functioning of 

aquatic systems. By the nature of their biological 

activities, autotrophic and heterotrophic microorganisms are sensitive indicators of the ecological 

and water-quality status of aquatic environments. 

However, only by thoroughly understanding how 

micro-organisms function in healthy aquatic 

ecosystems, can we hope to recognize or accurately 

predict their responses to water-quality changes or 

other environmental disturbances. 

REFERENCES CITED 

Brock, T. D., 1966, Principles of microbial ecology: Englewood 

Cliffs, New Jersey, Prentice-Hall, 306 p. 

Campbell, R., 1977, Microbial ecology: London, Blackwell 

Scientific Publications, 148 p. 

Goldman, C. R., and Kimmel, B. L., 1978, Biological processes 

associated with suspended sediment and detritus in lakes 

and reservoirs, in Cairns, J., Benfield, E. F., and Webster, 

Ell 
J. R., eds., Current perspectives on river-reservoir 

ecosystems: North American Benthological Society, 

Publication 1, p. 19-44. 

Karl, D. M., Wirsen, C. 0., and Jannasch, H. W., 1980, Deep-sea 

primary production at the Galapagos Hydrothermal Vents: 

Science, v. 207, p. 1345-1347. 

Kuznetsov, S. 1., 1959, Die Rolleder Mikroorganismen im 

Stoffkreislauf der Seen: Berlin, VEB Deutscher Verlag der 

Wissenschaften, 301 p. 

Lean, D. R. S., 1973, Phosphorus dynamics in lake water: 

Science, v. 197, p. 678-680. 

Lindeman, R. L., 1942, The trophic-dynamic aspect of ecology: 

Ecology, v. 23, p. 399-418. 

Pomeroy, L. R., 1974, The ocean's food web: A changing 

paradigm: Bioscience, v. 24, p. 499-504. 

Porter, K., Pace, M., and Battey, J., 1979, Cilitate protozoans 

as links in freshwater planktonic food chains: Nature, v. 

277, p. 563-565. 

Rigler, F. H., 1975, The concept of energy flow and nutrient 

flow between trophic levels, in van Dobben, W. H., and 

Lowe-McConnell, R. H., eds., Wageningen, W. Junk B. V. 

Publ., eds., Unifying concepts in ecology, p. 15-26. 

Sorokin, Y. 1., and Kadota, H., eds., 1972, Techniques for the 

assessment of microbial production and decomposition in 

freshwaters: London, Blackwell Scientific Publications, IBP 

Handbook No. 23, 112 p. 

Vallentyne, J. R., 1974, The algal bowl: Lakes and man: Ottawa, 

Canada Department of the Environment, Miscellaneous 

Special Publication 22, 186 p. 

Wetzel, R. G., Rich, P. H., Miller, M. C., and Allen, H. L., 1972, 

Metabolism of dissolved and particulate detrital carbon in a 

temperature hard-water lake: Memorie dell'Istituto Italiano 

di Idrobiologia, 29 Suppl., p. 185-243. 

ADDITIONAL RECOMMENDED READING 

Deevey, E. S., 1970, Mineral cycles: Scientific American, v. 223, 

p. 148-158. 

Delwiche, C. C., 1970, The nitrogen cycle: Scientific American, 

v. 223, p. 136-146. 

Hutchinson, G. E., 1970, The biosphere: Scientific American, v. 

223, p. 44-53. 

E12 
Effects of Bacteria on the Chemical and 

Physical State of Iron 

By Gail E. Mallard 

ABSTRACT 

Bacterial activity can bring about transformations in the 

chemical and physical state of iron such as oxidation, reduction, 

solubilization, precipitation, and formation and decomposition 

of organic-iron complexes. A brief overview of the chemical and 

biological reactions of iron in soils and water is presented for the 

nonspecialist who has some familiarity with chemistry and 

microbiology. The physiology and ecology of certain bacteria 

that commonly affect iron are outlined and references for procedures used in the cultivation of these organisms are provided. 

INTRODUCTION 

This report presents, for the nonspecialist who 

has some familiarity with chemistry and 

microbiology, a brief overview of the chemical and 

biological reactions of iron in soils and water. It 

mentions some of the resulting problems and 

describes several of the principal organisms involved. Emphasis is placed on the bacteria that oxidize or reduce iron as an integral part of their 

metabolism, but organisms that cause reactions indirectly also are considered. 

In soils and water, iron and compounds of iron 

exist as dissolved ions, solids, and colloidal suspensions. Bacterial activity can bring about the following transformations in the chemical or physical 

state of iron: oxidation, reduction, solubilization, 

precipitation, and formation and decomposition of 

organic-iron complexes. These alterations may be 

enzyme-catalyzed, in which case they probably contribute to the cell's metabolism, or they may be due 

to nonspecific chemical reactions occurring in 

response to changes in the environment, which are 

in turn induced by microbial activity. In either 

case, the presence and activity of certain microorganisms will cause a change in the state of iron. 

Although the bacteria associated with iron in soils 

and water commonly are referred to as "iron 

E13 

bacteria," this is actually a general term applied to 

a diverse group of organisms having greatly differing metabolic capabilities and living in a wide 

variety of habitats. 

The two most serious problems associated with 

iron bacteria are the adverse effects on flow and 

water quality in water-supply systems, and 

acidification of water draining from mines ("acidmine drainage"). Water-supply systems typically 

are affected by the deposits of ferric hydroxide and 

the gelatinous growth of iron bacteria, both of 

which lead to reduced potability (unpleasant taste 

and odor), discoloration of water, clogged well 

screens, and a reduction in flow through pipes. In 

coal-mining operations, the bacterium, 

Thiobacillus ferrooxidans, produces large quantities of sulfuric acid as a waste product of its 

metabolism, and when this acid waste flows over 

land and enters a body of water, the land and 

receiving water become unfit for many terrestrial 

and aquatic organisms. 

CHEMICAL REACTIONS OF IRON 

Microbiological changes in suspended or dissolved iron may accompany naturally occurring 

chemical changes and in some case will compete 

with them. Additionally, alterations in the environment brought about by microbial activity may shift 

chemical equilibria and thereby change the dominant form of iron. For these reasons, the chemical 

reactions of iron, exclusive of biological activity, 

are described briefly below. The chemistry of iron 

in natural waters has been extensively reviewed 

and explained in numerous papers by Hem (1960, 

1961, 1970, 1972; and Hem and Cropper, 1959). 
In natural waters, iron occurs as ferrous (reduced, Fe+2) and ferric (oxidized, Fe+3) ions as well 

as various oxides and hydroxides. Some of the pertinent reactions of iron are listed below: 

Fe+2:;=Fe•3+e- (1) 

Fe+l+H20:;=FeOH•2+H•+e- (2) 

Fe+l+2Hl0:;=Fe(OH)l•+2H•+e- (3) 

Fe•2+3H20:;=Fe(OH)3+3H•+e- (4) 

FeOH•+2H20:;=Fe(OH)3+2H•+e- (5) 

Fe(OH)l + H20 :;= Fe(OH)3 + H• + e- (6) 

Fe•2+HC03-:;=FeC03+H• (7) 

Fe•3+HlO:;=FeOH•2+ H• (8) 

FeOH•2+HlO:;=Fe(OH)l•+H• (9) 

Fe(OH)2•+H20:;=Fe(OH)3+H• (10) 

Hydrogen ions, electrons, or both appear in all of 

the above equations; therefore, the final equilibrium concentrations of the iron compounds are, in 

part, dependent upon Eh and pH. The effect ofEh 

and pH on iron, and the ranges in which particular 

species of ions or solids will be stable at chemical 

equilibrium, may be depicted in stability-field 

diagrams as shown in figure 11. Such diagrams 

depict one or more solid components of known 

chemical composition in contact with water. The 

concentration of the dissolved ions will affect the 

location of the boundaries. It should be noted that 

in some cases, stability-field diagrams do not reflect the true situation because the equilibrium conditions assumed in their preparation may never be 

attained. 

Some of the characteristics of iron that are important in determining its solubility and availability were listed by Hem (1972, p. 443) and are 

quoted below: 

(1) Iron may occur in the ferric (oxidized) or ferrous 

(reduced) state, and oxidation or reduction is 

readily accomplished; 

(2) Oxidized forms tend to have low solubility, 

especially in alkaline solutions; 

(3) Reduced metals can form compounds with carbonate or sulfide that are low in solubility; 

( 4) Organic and inorganic solute complexes are 

formed with many ligands and this tends to 

increase the solubilities of iron species; 

(5) Colloidal dispersions of ferric hydroxide can 

occur. 

In general, when Eh and pH are low, iron solubility 

is high. "Water in contact with air will have an .Eh 

of approximately 0.35 to 0.50 volt, and usually its 

1.40 

1.20 

1.00 

0.80 

0.60 

0.40 

(/) 

1-

-' 0 

> 0.20 

~ 

.r:. 

w 

pH 

FIGURE 11.-Fields of stability for solid and dissolved forms 

of iron as a function of Eh and pH at 25°C and 1 atmosphere of pressure. Activity of sulfur species 96 mg/L 

as 804 - 2 , carbon dioxide species 1,000 mg/L as HCOs-, 

and dissolved iron 0.0056 mg/L (from Hem, 1970, p. 

118). 

pH is greater than 5." Under these conditions, "ferric hydroxide or ferric oxide is the stable form and 

iron solubility is low'' (Hem, 1961, p. 227). 

EFFECTS OF MICROBIAL ACTIVITY ON IRON 

The chemical and physical state of iron can be 

altered both directly and indirectly by microbial activity. Bacteria can oxidize and reduce iron by 

direct enzymatic mechanisms; they also can consume oxygen, produce and degrade organic compounds, and change the Eh and pH of the environment. Microbiologically mediated changes in iron, 

and environmental changes that affect iron, are illustrated in figure 12. Iron-oxidizing and ironreducing bacteria cause a direct, enzymatic change 

in iron that is important in energy generation for 

the cells. The other changes in iron illustrated in 

E14 
figure 12 are caused indirectly, by products of 

bacterial metabolism; the cells responsible may be 

unaffected by the presence or absence of iron. 

Some of the changes in iron and the bacteria that 

cause these changes are discussed in the following 

section. 

DECOMPOSITION AND FORMATION OF ORGANIC-IRON 

COMPOUNDS 

Sugars, simple organic acids, and highly 

polymerized humus constituents can combine with 

ferrous and ferric iron to form iron-organic complexes (Alexander, 1977). Because iron is more 

soluble and stable when complexed with organic 

compounds than when alone (Hem, 1960), 

microbial activity leading to the production or 

decomposition of such compounds may affect the 

chemical and physical state of iron. Bacterial activity may stabilize iron if the organic acids produced as waste products of microbial metabolism 

accumulate and combine with the iron. Conversely, 

if the organic part of an iron-organic complex is 

degraded by microbial metabolism, its stabilizing 

effect will be removed and iron will be subject to 

oxidation and (or) precipitation. For example, 

bacterial metabolism causes iron to precipitate 

from solutions of the citrate, lactate, acetate, 

malate, malonate, oxalate, and gallate salts of iron 

(Alexander, 1977). Some of the heterotrophic 

organisms that bring about the precipitation of 

iron from solution by using the organic part of an 

iron-organic complex are Enterobacter, Serratia, 

Bacillus, Klebsiella, Alcaligenes, Moraxella, Corynebacterium, Caulobacter, Mycobacterium, and 

Escherichia (Cullimore and McCann, 1977). 

IRON REDUCTION 

The reduction of iron requires an organic energy 

source and a low redox potential. The reduction 

may be either an indirect chemical process due to 

changes in Eh or a direct enzymatic reduction. Ottow (1968) demonstrated that bacteria capable of 

reducing iron were distributed through a column of 

soil and were present at levels of 104 to 105 per 

gram of dry soil. Some of the genera containing 

species able to reduce iron are Bacillus, 

Clostridium, Klebsiella, Pseudomonas, and Serratia (Alexander, 1977). 

Many heterotrophic organisms potentially are 

able to bring about a sequence of events that result 

in indirect iron reduction. As organic compounds 

are metabolized, oxygen is consumed and reducing 

compounds are produced; this lowers the Eh and 

shifts the equilibrium in the direction of ferrous 

iron. Another, more direct, mechanism is the 

transfer of electrons from a reduced carbon compound to ferric iron, possibly resulting in the 

generation of energy for the cells responsible. 

These bacteria are not highly specialized 

organisms; they seem to be using iron as one of 

several possible terminal electron acceptors. 

Evidence indicates that at least two different enzymes or enzyme systems are involved. Ottow 

(1968) investigated the physiology of iron reduction with pure cultures of Enterobacter aerogenes, 

Escherichia coli, and six species of Bacillus. 

Because nitrate suppressed iron reduction in 

cultures that had the enzyme nitrate reductase, Ottow suggested that ferric iron was acting as an 

alternative terminal electron acceptor for nitrate 

reductase. Later studies (Ottow and Munch, 1978) 

with a Clostridium species that did not possess the 

enzyme nitrate reductase, but was able to reduce 

iron, led to the suggestion that at least two different enzymes are involved in iron reduction- nitrate reductase and another enzyme able to 

reduce iron (and manganese), but unable to reduce 

nitrate. 

Iron reduction may be responsible for the production of gleys-sticky bluish-gray soils-in 

waterlogged areas (Alexander, 1977). The color of 

the gleyed zone is due to the ferrous sulfide produced when the end product of iron reduction 

(Fe+2) reacts with hydrogen sulfide, the end product of bacterial sulfate reduction. 

IRON OXIDATION AND PRECIPITATION OF IRON 

COMPOUNDS 

The oxidation of ferrous to ferric iron and the 

subsequent precipitation of insoluble ferric hydroxides commonly is associated with a heterogeneous 

group of bacteria. Thiobacillus ferrooxidans lives 

in environments of low pH, oxidizes iron enzymatically, and thereby derives its energy. 

Gallionella also obtains energy from iron oxidation, although it requires certain environmental 

conditions (neutral pH and the presence of oxygen) 

that force it to compete with the spontaneous 

chemical oxidation of ferrous iron. In addition, a 

variety of organisms living in aerobic environments at neutral pH deposit ferric hydroxide 

E15 
trj 

...... 

~ 

Iron precipitated 

Ferrous iron 

oxidized to 

ferric iron 

(Fe+2 Fe+3) 

Ferric iron 

reduced to 

ferrous iron 

(Fe+3 Fe+2) 

Electrons removed 

from ferrous iron 

Electron transport 

with Fe+J as 

terminal electron 

acceptor 

Iron-organic 

complexes 

formed 

Carbonate-

~-------------------------------------------~ bicarbonate 

system altered 

Hydrogen sulfide ~------1 

produced 

Oxygen consumed and(or) 

reducing compounds produced 

Organic and inorganic 

acids produced 

Organic compounds I>E 1 produced 

Carbon dioxide produced 

or consumed 

Organic part of ironorganic complex metabolized 

FIGURE 12.-Direct and indirect changes in iron resulting from microbial activity. 

Iron-oxidizing bacteria 

Sulfate-reducing bacteria 

Iron-reducing bacteria 

Various heterotrophic bacteria 
on the external surface of the cell (cell wall, capsule, or sheath). For these bacteria, it is likely that 

the oxidation of ferrous iron is nonenzymatic and 

that the bacteria derive no energy from the process, but merely serve as convenient sites for 

precipitation of the ferric compounds generated by 

spontaneous chemical reactions. The following sections summarize information about the ecology and 

physiology of bacteria that oxidize and (or) 

precipitate iron. 

Thiobacillus ferrooxidans 

Thiobacillus ferrooxidans, an acidophilic 

bacterium requiring a pH range of 2.5-5.8, is able 

to grow without organic compounds and derives 

energy from the oxidation of ferrous iron and cell 

carbon·from carbon dioxide. The ability to grow at 

low pH is significant for the cell's energygenerating metabolism because it is only below pH 

5 that ferrous iron is stable in the presence of the 

oxygen that is required by this organism. In 

nature, Thiobacillus ferrooxidans uses ferrous 

sulfate (FeS04), pyrite (FeS2), and other reduced 

iron compounds, and forms ferric oxides, hydroxides, sulfates, and phosphates as end products 

(Lundgren and others, 1972). It also is able to oxidize elemental sulfur, thiosulfate, and possibly 

other reduced sulfur compounds. 

Electrons removed from iron are transferred to 

oxygen, giving the following overall reaction: 

4 Fe+2+02+4 H+-4 Fe+3+2 H20 

The energy yield of this reaction is low [about 

10-11 kilocalories of energy per mole of iron (55.8 

g) oxidized], and energy requirements for cell synthesis are high [about 115 kilocalories of energy to 

fix 1 mole of carbon (Lundgren and others, 1972)]; 

therefore, Thiobacillus ferrooxidans must convert 

a large amount of substrate for a relatively low cell 

yield. Estimates of the ratio of iron oxidized to cell 

material formed a range as high as 500:1 (Wolfe, 

1964). 

In mining operations, the action of iron-oxidizing 

bacteria in degrading pyrite and other metal 

sulfide minerals, and the large amounts of waste 

products produced (sulfuric acid, ferrous iron, and 

other metal ions), leads to the major water-quality 

problem referred to as acid-mine drainage. This 

acid effluent has obvious detrimental effects on 

both the land over which it flows and upon the 

receiving waters. It has been estimated that 

"10,000 miles of streams and 29,000 surface acres 

of impoundments and reservoirs are seriously affected by surface mining operations and that deep 

mining (and other types of mining) add to these 

figures significantly." (Lundgren and others, 1972, 

p. 69). Alexander (1977, p. 370) cites an estimate 

that "the bituminous mines of western Pennsylvania alone add about a million tons of sulfuric 

acid each year to the Ohio River drainage area." 

The reactions leading to the production of acidmine waters are presented below with pyrite used 

as an example (Lundgren and others, 1972): 

2 FeS2 + 2 H20 + 7 02-2 FeS04 + 2 H2S04 (1) 

pyrite ferrous sulfuric 

sulfate acid 

4 FeS04+02+2 H2S04-2 Fe2(S04)3+2 H20 (2) 

ferrous ferric 

sulfate sulfate 

Fe2(S04)3+6 H20-2 Fe(OH)3+3 H2S04 (3) 

ferric ferric 

sulfate hydroxide 

The initial oxidation of pyrite or other metal 

sulfide may be biological or nonbiological. The slow 

nonbiological oxidation of pyrite is accelerated 

greatly by the addition of a small amount of acidmine water, which may contain as many as 106 

iron-oxidizing organisms per milliliter (Lundgren 

and others, 1972). The catalytic activity is attributed to the bacteria because it is eliminated by 

sterilization and is negligible at 0°C (Alexander, 

1977). In the absence of a catalyst, the oxidation of 

ferrous to ferric iron (reaction 2, above) proceeds 

very slowly below pH 4 and is the rate-limiting step 

in the oxidation of iron pyrite and the formation of 

acidity. Microbial activity has been shown to accelerate this reaction by a factor of larger than 106 

(Lundgren and others, 1972). Reaction 3 (above) is 

nonbiological. 

Pyrite oxidation also may occur by an alternative 

mechanism in which the ferric iron generated 

biologically in reaction 2 spontaneously oxidizes 

the iron sulfide as in the following reaction: 

FeS2+14 Fe+3+8 H20-

15 Fe+2+2 S042+16 H+ (4) 

The ferrous iron produced from this reaction may 

then be oxidized by the iron bacteria. 

Gallionella 

Gallionella is a unique stalked bacterium that 

lives at neutral pH and has been reported to obtain 

E17 
energy by oxidizing iron. In nature, these 

organisms are found in soil and water containing 

ferrous iron; they also may be found in watersupply systems, where the iron hydroxide deposits 

associated with this organism may lead to reduced 

flow in the lines, discolored water, and other problems. 

Gallionella is probably autotrophic. The . cells, 

which are found in waters low in organic carbon, 

oxidize ferrous to ferric iron and fix measurable 

quantities of 14C02 • Furthermore, Gallionella can 

grow on a simple medium (Wolfe, 1964) containing 

mineral salts, carbon dioxide, and ferrous sulfide 

as an energy source. The organisms are 

microaerophilic and grow well under low oxygen 

concentrations (0.1 to 0.2 mg/L of 0 2); higher 

levels (> 2. 75 mg/L of 0 2) inhibit growth (Cullimore 

and McCann, 1977). The optimum pH for growth is 

6 to 7. In laboratory culture, Gallionella exhibits 

zonal growth in an area of the test tube where the 

organisms can compete for ferrous iron as well as 

oxygen and in the same region where Fe+3 is 

deposited in uninoculated tubes (Wolfe, 1964). 

The type species for the genus is Gallionella ferruginea. Other species may exist, but are not well 

described. The cells of Gallionella sp. are kidneyshaped or rounded and have a long stalk that is 

secreted from the side of the cell perpendicular to 

its long axis. The stalk, which is composed of 

bundles of helically twisted fibrils and covered with 

ferric hydroxide, usually is attached to a substrate. 

It elongates as the cell grows and may bifurcate at 

the time of cell division. 

Sphaeroti/us-Leptothrix GROUP 

The two genera, Sphaerotilus and Leptothrix, are 

frequently discussed together because they are 

similar in morphology and because there is some 

confusion in the literature as to their differentiation and classification. Although both are commonly referred to as "iron bacteria" and deposit 

ferric hydroxide, it is unlikely that either is able to 

obtain energy from iron oxidation. 

The genus, Sphaerotilus, contains only one 

species; the genus, Leptothrix, contains five. All of 

these organisms are rod-shaped and generally occur in chains encased within a sheath, but some occur as single cells that are motile. In the latest 

issue of Bergey's Manual of Determinative 

Bacteriology (Buchanan and Gibbons, 197 4), the 

recognized source of taxonomic information on 

bacteria, the two genera are differentiated on the 

basis of sheath coating: Leptothrix sheaths typically are encrusted with iron or manganese oxides, 

whereas Sphaerotilus sheaths rarely are encrusted 

with iron and never with manganese oxides. 

Although a complete discussion of the taxonomy of 

these organisms is beyond the scope of this paper, 

the uncertainty of differentiation between the two 

groups is indicated in the following statement by 

Mulder and van V een in their description of 

Sphaerotilus natans (in Buchanan and Gibbons, 

1974, p. 129): "S. natans may thus behave like an 

iron bacterium and according to some authors it is 

identical with Leptothrix ochracea * * *. However, the present authors have clearly shown that 

the two organisms are distinct * * * ." The taxonomy and classification of Sphaerotilus and Leptothrix have been reviewed by Wolfe (1964), 

Mulder (1964), and Dondero (1975), in addition to 

the discussion by Mulder and van V een (in 

Buchanan and Gibbons, 197 4). 

Although the sheath of Leptothrix, and rarely 

Sphaerotilus, may be encrusted with iron hydroxides, it is unlikely that the bacteria oxidize iron or 

derive energy from it. Because both organisms 

grow at the same pH, near neutrality, at which ferrous iron (Fe+2) is chemically oxidized to ferric iron 

(Fe+ a), it is probable that the sheaths serve only as 

a site for the deposition of the insoluble ferric 

hydroxides. Dondero (1975, p. 419) described 

studies by Cataldi, who was: 

* * * unable to grow 255 isolates of Leptothrix and 

Sphaerotilus on purely mineral media, although they grew on 

media containing 0.005% peptone and 0.01% manganese 

acetate. The absence of additional iron or manganese in the 

medium was not detrimental to growth, nor was the presence 

necessarily beneficial. In pure culture, iron was never oxidized * * *. Oxidation of the iron or manganese could be produced with dead cells, as well as with live cultures. 

Similarly, Wolfe (Heukelekian and Dondero, 1964, 

p. 108), in a discussion of Mulder's paper, reported 

that ferric hydroxide was deposited on the sheaths 

of Sphaerotilus natans cells that had been killed. It 

is, therefore, unlikely that the SphaerotilusLeptothrix group causes the oxidation of iron or 

obtains any energy from the process. 

Both Sphaerotilus and Leptothrix are found attached to submerged rocks, plants, and so on in 

slowly running waters. In this type of habitat, the 

flowing water provides the organisms with a continuous source of nutrients and also carries away 

their waste products. Leptothrix is found in unpolluted water containing iron, whereas 

E18 
Sphaerotilus is associated with water having a 

higher concentration of organic material. 

These organisms have some economic importance because "they grow in pipes along with 

deposits of iron or manganese and interfere with 

the passage of water in wells and filtration plants" 

(Dondero 1975, p. 408). Massive growth of 

Sphaerotilus in streams and sewage treatment 

plants is a common problem, although this 

phenomenon is not associated with iron deposition. 

OTHER IRON-DEPOSITING BACTERIA 

In addition to the bacteria described above, 

Bergey's Manual of Determinative Bacteriology 

(Buchanan and Gibbons, 1974) lists 14 other genera 

that become coated or encrusted with iron deposits 

(chiefly ferric hydroxide). Most of these organisms 

have not been cultivated in the laboratory or 

grown in pure culture; thus, details of their 

physiology and metabolism are unclear. However, 

because all of these bacteria require oxygen and 

live at near neutral pH, it is likely that the cells or 

associated sheaths or capsules serve only as a site 

for precipitation of chemically oxidized iron. It is 

unlikely that these organisms obtain energy from 

iron oxidation. 

A list grouping the iron-depositing bacteria into 

four major categories and giving some of their pertinent characteristics as in Bergey's Manual of 

Determinative Bacteriology (Buchanan and Gibbons, 197 4) is presented below: 

I. Gliding Bacteria 

Toxothrix: no pure cultures 

not cultivated in the laboratory 

iron not needed for growth 

optimum pH 6.5-7.2 

develops best at reduced oxygen tensions 

found in cold springs, bogs, ponds, and 

lakes containing ferrous iron. 

II. Sheathed Bacteria 

Lieskeella: no pure cultures 

deposits ferric hydroxide on capsule 

found in upper layers of mud in bodies of 

water. 

Crenothrix: not grown on artificial media in 

pure culture 

sheath may be encrusted with iron oxides 

found in stagnant and running waters containing organic matter and iron salts. 

Clonothrix: not grown in pure cultures on artificial media 

sheath may be encrusted with iron compounds giving yellowish brown color 

found attached to iron fittings in well 

water. 

III. Budding and (or) Appendaged Bacteria 

Pedomicrobium: pure cultures available 

ferric iron compounds deposited 

grow at neutral pH 

microaerophilic to aerobic 

widely distributed in soil and water. 

Seliberia: pure culture available 

deposits iron hydroxide in iron-containing 

media and in soils 

facultatively anaerobic 

common inhabitant of soils. 

Planctomyces: not grown on artificial media 

cells encrusted with ferric hydroxide 

aerobic 

found in surface layers of lakes. 

Metallogenium: have been grown in pure 

culture 

iron oxides deposited 

optimum pH 6.8-7.0 

found in soil and water. 

CaulococC'US: have not been cultured in the 

laboratory 

deposits ferric iron 

microaerophilic 

found in bottom mud of lakes. 

K usnezovia: not cultured in the laboratory 

oxidizes and deposits iron 

microaerophilic 

found in mud samples from lakes. 

IV. Gram-negative, Chemolithotrophic Bacteria 

Four genera are placed in the Family Siderocapsaceae. The main characteristic of this group is its 

ability to deposit iron and (or) manganese oxides 

on or in capsules when present, or on extracellular 

material. G. A. Zavarzin, writing about this family 

in Bergey's Manual of Determinative Bacteriology 

(Buchanan and Gibbons, 197 4, p. 464), makes the 

following observations: 

Few of the organisms in this family have been cultivated and the 

validity of a taxon based on deposition of iron and manganese 

oxides is doubtful, as it is now known that many other bacteria 

have this ability. The four genera recognized have been observed and described by numerous investigators and they are of 

geological importance. Differentiation is mainly on morphological features. 

E19 
The following editorial note also is included in the 

introductory material for the group (Buchanan and 

Gibbons, 1974, p. 464): 

The author has questioned the validity of the family and to the 

editors, its placement has presented a problem. 

The four genera included have only one feature in common-the ability to deposit or to cause the deposition of iron 

and/or manganese oxides. Most are organotrophs. Siderococcus 

reproduces by budding and could be placed in Part 4. Dr. 

Dubinina has recently obtained Siderocapsa in pure culture and 

found it indistinguishable from A rthrobacter globijormis. 

Ochrobium may be an alga. 

As it is impossible to place the family, as here constituted, in a 

more logical place it is included with the chemolithotrophs, even 

though it is known that in many instances the deposition of iron 

or manganese is the result of the metabolism of the organic ion 

of the compound. 

Siderocapsa: pure culture available 

capsule encrusted with iron compounds 

generally aerobic, but may grow at reduced oxygen tensions 

found in neutral to slightly alkaline conditions 

common in freshwater. 

N aumanniella: capsule coated with deposits of 

Iron 

complex organic compounds of iron are decomposed 

widely distributed in iron-bearing waters. 

Ochrobium: cells surrounded by capsule containing iron 

widely distributed in iron-bearing 

freshwater. 

Siderococcus: ferric hydroxide deposited in 

medium, cells not encrusted with iron 

compounds 

growth observed only in natural habitat 

and isolated mud samples 

found in mud habitats with low oxygen 

concentrations and neutral pH 

widely distributed in freshwaters and 

bottom deposits. 

CULTIVATION AND IDENTIFICATION OF 

IRON BACTERIA 

Bacteria which are able to bring about a change 

in the chemical or physical state of iron are a 

diverse group living in many different habitats and 

having greatly differing requirements for growth. 

Because it would be impossible to cultivate all of 

these organisms in the same medium and by the 

same procedures, a list of all media and procedures 

used is necessary, but beyond the scope of this 

report. Instead, references for techniques used to 

screen water samples for iron bacteria and additional references on methods to cultivate specific 

types of bacteria are presented. The best source of 

information for the identification of iron bacteria is 

Bergey's Manual of Determinative Bacteriology 

(Buchanan and Gibbons, 1974). 

Iron-reducing bacteria require an organic source 

of carbon and energy and may be grown on a 

general bacteriological medium with the addition 

of ferric iron to test for iron reduction. Ottow 

(1968) and Ottow and Munch (1978) describe some 

of the media and procedures used to detect ironreducing bacteria. 

General screening methods for iron bacteria are 

given by Rodina (1972) and Cullimore and McCann 

(1977). These methods include differential staining 

followed by microscopic observation, as well as 

several enrichment techniques in which visible color changes and (or) distinctive patterns of growth 

indicate the presence of iron bacteria. 

A medium suitable for the growth of . heterotrophic, iron-precipitating bacteria is given in 

Standard Methods (American Public Health 

Association and others, 1976). Rodina (1972) lists 

several media for the cultivation of iron-oxidizing 

and iron-precipitating bacteria; some of these 

media are evaluated by Cupimore and McCann 

(1977). Other references for media and procedures 

to culture specific types of iron bacteria are: 

(1.) Thiobacillus ferrooxidans 

a. Silverman and Lundgren (1959) 

b. Lundgren and others (1972) 

c. American Public Health Association and 

others (1976). 

(2.) Metallogenium sp. 

a. American Public Health Association and 

others (1976). 

(3) Gallionella sp. 

a. Wolfe (1958) 

b. American Public Health Association and 

others (1976). 

(4.) Budding Bacteria 

a. Ghiorse and Hirsch (1978). 

(5.) Sphaerotilus-Leptothrjx 

a. American Public Health Association and 

others (1976) 

b. Dondero (1975). 

E20 
SUMMARY 

Bacteria are able to bring about changes in both 

the chemical and physical state of iron in water and 

soils. Many of the changes result indirectly from 

microbial alterations of the environment; for example, iron increases and decreases in solubility as 

a result of Eh and pH changes associated with the 

growth of many heterotrophic bacteria. Iron can 

be affected more directly when iron-organic complexes are destroyed as the organic part is used by 

micro-organisms. Finally, some bacteria are able 

to bring about the direct enzymatic oxidation or 

reduction of iron as a part of their energy 

generating process. The physiology and ecology of 

certain bacteria that commonly affect iron are 

outlined in this paper. Emphasis is placed on those 

organisms that change iron directly as a significant 

part of their metabolism; although, indirect reactions also are described. References for procedures 

used in the cultivation of these organisms are provided. 

REFERENCES 

Alexander, Martin, 1977, Introduction to soil microbiology (2d 

ed.): New York, John Wiley and Sons, 467 p. 

American Public Health Association, American Water Works 

Association, and Water Pollution Control Federation, 1976, 

Standard methods for the examination of water and 

wastewater (14th ed.): Washington, American Public 

Health Association, Inc., 1193 p. 

Buchanan, R. E., and Gibbons, N. E., eds., 1974, Bergey's 

manual of determinative bacteriology (8th ed.): Baltimore, 

The Williams and Wilkins Company, 1268 p. 

Cullimore, D. R., and McCann, A. E., 1977, The identification, 

cultivation, and control of iron bacteria in ground water, in 

F. A. Skinner and J. M. Shewan, eds., Aquatic 

microbiology: New York, Academic Press, p. 219-261. 

Dondero, N. C., 1975, The Sphaerotilus-Leptothrix group: Annual Review of Microbiology, v. 29, p. 407-428. 

Ghiorse, W. C., and Hirsch, Peter, 1978, Iron and manganese 

deposition by budding bacteria, in W. E. Krumbein, ed., Environmental biogeochemistry and geomicrobiology, v. 3, 

Methods, metals, and assessment: Ann Arbor, Ann Arbor 

Science Publishers, Inc., p. 897-909. 

Hem, J. D., 1960, Complexes of ferrous iron with tannic acid: 

U.S. Geological Survey Water-Supply Paper 1459-D, 94 p. 

-- 1961, Stability field diagrams as aids in iron chemistry 

studies: Journal American Water Works Association, v. 53, 

p. 211-232. 

-- 1970, Study and interpretation of the chemical 

characteristics of natural water, (2d ed.): U.S. Geological 

Survey Water-Supply Paper 1473, 363 p. 

-- 1972, Chemical factors that influence the availability 

of iron and manganese in aqueous systems: Geological 

Society of America Bulletin, v. 83, p. 443-450. 

Hem, J. D., and Cropper, W. H., 1959, Survey of ferrous-ferric 

chemical equilibria and redox potentials: U.S. Geological 

Survey Water-Supply Paper 1459-A, 31 p. 

Heukelekian, H., and Dondero, N.C., eds., 1964, Principles and 

applications in aquatic microbiology: New York, John 

Wiley, p. 98-112. 

Lundgren, D. G., Vestal, J. R., and Tabita, F. R., 1972, The 

microbiology of mine drainage pollution, in Ralph Mitchell, 

ed., Water pollution microbiology: New York, WileyInterscience, p. 69-88. 

Mulder, E. G., 1964, Some observations of the SphaerotilusLeptothrix group, in H. Heukelekian and N. C. Dondero, 

eds., Principles and applications in aquatic microbiology: 

New York, John Wiley, p. 98-112. 

Ottow, J. C. G., 1968, Evaluation of iron-reducing bacteria in 

soil and the physiological mechanism of iron-reduction in 

Aerobacter aerogenes: Zeitschrift fur Allgemeine 

Mikrobiologie, v. 8, p. 441-443. 

Ottow, J. C. G., and Munch, J. C., 1978, Mechanisms of reductive transformations in the anaerobic microenvironment of 

hydromorphic soils, in W. E. Krumbein, ed., Environmental 

biogeochemistry and geomicrobiology, v. 2, The terrestrial 

environment: Ann Arbor, Ann Arbor Science Publishers 

Inc., p. 483-491. ' 

Rodina, A. G., 1972, Methods in aquatic microbiology: 

Baltimore, University Park Press, 461 p. 

Silverman, M. P., and Lundgren, D. G., 1959, Studies on the 

chemoautotrophic iron bacterium Ferrobacillus ferrooxidans. I. An improved medium and a harvesting procedure 

for securing high cell yields: Journal of Bacteriology, v. 77, 

p. 642-647. 

Wolfe, R. S., 1958, Cultivation, morphology, and classification 

of the iron bacteria: Journal American Water Works 

Association, v. 50, p. 1241-1249. 

--1964, Iron and manganese bacteria, in H. Heukelekian 

and N. C. Dondero, eds., Principles and applications in 

aquatic microbiology: New York, John Wiley, p. 82-97. 

E21 

Micro-organisms in Stormwater 

A Summary of Recent Investigations 

By Gail E. Mallard 

ABSTRACT 

The major concepts and considerations in microbiological 

analysis of storm runoff are summarized and documented to 

provide a basis for use by community planners, water 

managers, and others interested in studies of stormwater. 

Twenty-five published reports dating since 1964 are cited. 

All storm runoff contains a variety of bacteria, including total 

coliforms, fecal coliforms, and fecal streptococci, that are derived from the land over which the water flows. Most of the 

total coliforms are native soil organisms, whereas the fecal coliforms and fecal streptococci originate from the feces of wild 

and domestic animals. Urban runoff has been reported to contain pathogenic organisms, but this probably presents little 

direct threat to human health because the runoff is not ingested. 

Runoff water, however, can have other negative effects such as 

contamination of surface waters, which may result in beach 

closures, or contamination of shellfish. This type of contamination is generally of short duration because indicator bacteria 

and pathogens die rapidly in the aquatic environment. Similarly, bacteria and viruses deposited on soil by stormwater are 

inactivated by drying, competition from soil microflora, and a 

variety of other processes. 

Every storm producing runoff is unique in terms of the 

number and type of micro-organisms, for they vary from site to 

site, from storm to storm, and over time during any one storm 

event. Stormwater to be examined for micro-organisms must be 

collected in sterile containers and processed immediately. 

INTRODUCTION 

Any attempt to find micro-organisms in stormwater generally will meet with success. Bacteria 

and viruses are normal inhabitants of soil, water, 

human and animal skin and gut, plant surfaces, 

and indeed almost every place on earth. Only certain extremely hostile environments and the internal tissues of plants and animals are free of microorganisms. Stormwater running over the land surface inevitably will become contaminated, but this 

should not be a cause for concern, unless it 

threatens human health or well-being. Does stormwater present a serious threat? Unfortunately, no 

unequivocal answer can be given. Stormwater 

carrying micro-organisms may enter a drinkingwater source and cause an outbreak of disease, or 

it may lead only to the temporary shutdown of 

recreation facilities, such as beaches. Depending 

on local conditions, the study of microbial contamination of runoff may be of real importance to a 

community or it may be of only academic interest. 

PURPOSE AND SCOPE 

This summary presents general information 

gained from recent investigations and describes 

problems that may arise in future investigations. It 

is hoped that this information will be of value in 

planning studies that involve micro-organisms in 

storm water. 

BASIC CONCEPTS IN MICROBIOLOGY 

All scientific and technical fields have concepts, 

procedures, assumptions, and terminology that are 

unfamiliar to the nonspecialist. Although many of 

the terms and concepts of microbiology are 

familiar to the general public, the exact meanings 

and limitations may be poorly understood. For this 

reason, the main discussion of micro-organisms in 

storm runoff is preceded by a brief introduction to 

some of the principles of microbiology. A glossary 

of biological and microbiological terms is provided 

by Greeson and others (1977). 

INDICATOR ORGANISMS 

Water that has been polluted by human sewage 

is hazardous because several diseases are transE23 
mitted by way of the fecal-oral route, such as 

typoid fever, cholera, dysentery, and hepatitis. 

When presented with a water sample that is 

suspect, the microbiologist typically does not try to 

isolate and identify the pathogens, but instead 

determines numbers of "indicator" organismstotal coliforms, fecal coliforms, and fecal streptococci. These bacteria are known as indicator 

organisms because they indicate the presence of 

sewage and ideally are correlated with the number 

of pathogens in a water sample. 

The growth characteristics and metabolic reactions of the indicator organisms have been investigated for many years, and relatively easy and 

straightforward methods for their detection and 

enumeration are available (American Public 

Health Association, 1975; Greeson and others, 

1977). In contrast, methods for detection of 

pathogens are complicated, tedious, and time consuming. 

In addition to being easier to isolate and work 

with in the laboratory than pathogens, indicator 

bacteria are used because water contaminated by 

human waste typically will contain many more of 

these organisms than pathogens. Indicator 

organisms are present at relatively high levels in 

all members of the population and are being shed 

at all times. In contrast, only a few clinical cases 

and carriers will be shedding pathogens at any 

given time. 

Most of the characteristics of an ideal indicator 

organism are listed in Drinking Water and Health 

(National Research Council, 1977, p. 71) and are 

summarized as follows: 

(1) Applicable to all types of water; 

(2) Present in sewage and polluted waters, 

when pathogens are present; 

(3) Number is correlated with the 

amount of pollution; 

( 4) Present in greater numbers than pathogens; 

(5) No aftergrowth in water; 

(6) Greater survival time than pathogens; 

(7) Absent from unpolluted waters; 

(8) Easily detected by simple laboratory tests 

in the shortest time consistent with 

accurate results; 

(9) Has constant characteristics; 

(10) Harmless to man and animal. 

Although no organism or group of organisms is a 

perfect indicator, coliforms are ideal in most ways 

and have been used by water microbiologists and 

others concerned with the public health for many 

years. The U.S. Environmental Protection 

Agency's safe drinking-water standards are written in terms of coliforms, rather than pathogens 

(American Public Health Association, 1975), as are 

standards for recreational waters. As recently as 

1977, the use of coliforms as indicators of fecal contamination was endorsed by the Safe Drinking 

Water Committee of the National Research Council (1977). Yet, the popularity of coliforms as an indicator does not preclude critical reviews of their 

use (Dutka, 1973) or suggestions that other 

organisms or groups would provide more accurate 

determinations (Carberry and Stapleford, 1970). 

Additionally, continuing efforts are being made by 

many investigators to refine the methods used to 

detect indicator organisms. 

The effectiveness of total coliforms, fecal coliforms, and fecal streptococci as indicators of viral 

contamination is particularly suspect. Several 

reports indicate that bacterial indicators are inadequate predictors of viruses in ground water (Marzouk and others, 1979; Vaughn and others, 1978), 

as well as in fresh and marine surface water 

(Vaughn and others, 1979). This lack of a reliable 

indicator for enteroviruses presents a serious problem because detection of viruses in water samples 

requires expensive and time-consuming procedures. Studies of viruses in the environment, 

however, are becoming more commonplace, and 

methods for their detection and enumeration are 

being improved. 

The use of indicator organisms in stormwaterrunoff studies may pose a special problem. Most of 

the fecal coliforms and fecal streptococci in stormwater are derived from animal feces, so that the 

level of these indicator organisms may be high at 

all times, regardless of the number of pathogens in 

the water sample. Olivieri and others (1977) found 

little or no correlation between indicator and 

pathogenic bacteria in storm samples. Negative 

correlations, although not significant, were observed between indicator bacteria and enteric 

viruses. 

RATIOS OF FECAL COLIFORMS TO FECAL 

STREPTOCOCCI 

Aside from the question of the reliability of indicator organisms, a significant question remains 

unanswered- how is human fecal contamination 

differentiated from other fecal contamination? 

E24 
Since fecal material from many warmblooded 

animals contains coliforms and streptococci, the 

presence of these groups does not mean that 

human waste was the source of contamination. 

Geldreich and Kenner (1969) approached this problem by examining feces from various warmblooded 

animals (including humans), domestic wastewater, 

stormwater, food processing wastes, agricultural 

waters, and recreational waters. They concluded 

that the ratio of fecal coliforms to fecal streptococci (FC:FS) in human feces and in water polluted 

with human waste is always greater than 4.0, 

whereas the ratio of fecal coliforms to fecal streptococci in feces from farm animals, cats, dogs, and 

rodents, and in separate stormwater systems and 

farmland drainage, is less than 0.7. 

Although the ratio of fecal coliforms to fecal 

streptococci in a water sample gives some indication of the source of the contamination, these 

ratios must be interpreted with caution. First, the 

water samples must be taken near the source of 

contamination, because once the organisms have 

entered the receiving body of water, variables such 

as temperature, pH, metal concentration, nutrient 

availability, and other environmental factors will 

alter the interrelationship between the two indicator systems. McFeters and others (197 4) investigated the question of survival of indicator 

bacteria in natural waters and found that the 

initially high FC:FS ratio of human sewage decreased with time, whereas the initially low ratio 

for waste from domestic livestock increased. This 

reinforces the advice given by Geldreich and Kenner (1969, p. R349) that ''the use of a ratio relationship for stream samples would be valid only during 

the initial 24-hour travel downstream from point of 

pollution discharge into the receiving stream." 

Even if used correctly, the FC:FS ratio should not 

be regarded as a "magic number," especially for 

samples that contain water from a mixture of nonpoint sources, because the ratio of fecal coliforms 

to fecal streptococci does not reflect absolute 

numbers. For example, if most of the contamination in a sample were from nonhuman sources, a 

small amount of human sewage might not be sufficient to shift the overall ratio upward enough to 

cause concern. As a result, the presence of human 

pathogens in human sewage would be masked by 

the indicator ratio characteristic of animal waste, 

and a real danger would go undetected. 

MICRO-ORGANISMS IN STORMWATER 

BACTERIAL LOAD OF STORMWATER 

Rain contains very few bacteria before it reaches 

the ground. Geldreich and others (1968) collected 

rain from 49 storms in sterile collectors and tested 

for total coliforms, fecal coliforms, and fecal streptococci. In 42 of the samples, counts of these 

bacteria were less than one organism per 100 mL; 

in the remaining seven samples, total coliform densities ranged between 1 and 92 per 100 mL. When 

this "contaminated" rain was filtered and the filter 

examined microscopically, the authors observed 

soil particles and insect and vegetation fragments, 

which they believed were the source of the 

bacteria. Whenever rain comes in contact with 

land, it becomes contaminated with bacteria. 

Counts of total coliforms, fecal coliforms, and fecal 

streptococci in stormwater, compiled from several 

references (table 1), range as follows: 103-105 

total coliforms per 100 mL, 102-104 fecal coliforms 

per 100 mL, and 102-105 fecal streptococci per 100 

mL. 

SOURCES OF BACTERIA IN STORMWATER 

Geldreich and others (1968) examined 843 stormwater samples and observed that fecal coliforms 

constituted an average of 8.6 percent of the median total coliforms present. After examining over 

7, 000 separate strains of coliforms and determining their physiological type, they concluded that 

the remaining 91.4 percent of the total coliforms 

came from the soil. The distribution of strains was 

found to be the same in soil, surface waters, and 

stormwater. This exhaustive analysis leaves little 

doubt that most total coliforms in stormwater are 

native soil organisms that are washed off soil particles by water running over the land surface. 

If most of the total coliforms come from the soil, 

what is the source of the other two pollution indicators, fecal coliforms, and fecal streptococci? By 

definition these are contributed by warmblooded 

animals. Because the ratio of fecal coliforms to 

fecal streptococci in urban stormwater is usually 

much less than 1 (see table 5), it is assumed that 

most of the bacteria in stormwater are of 

nonhuman origin. In an urban environment, these 

bacteria probably originate from fecal material of 

dogs, cats, rodents, and other small animals; 

whereas in rural areas, larger domestic animals 

E25 
would make a significant contribution. In either 

case, the amount of waste generated by animals 

should not be underestimated. For example, a 

large city such as New York could easily have half 

a million dogs, and these dogs will deposit about 

150,000 pounds of feces and 90,000 gallons of urine 

each day on streets, sidewalks, and park areas. 

Even a small number of farm animals can have a 

significant impact because one cow will generate 

as much manure as 16.4 humans, a pig will produce 

as much as 1.9 humans, and 12 chickens as much as 

one human (Geldreich, 1976). Pollution from wild 

animals also must be considered. A beach in 

Madison, Wisconsin, was closed to swimming 

because of high bacterial counts in 1978. An investigation revealed that fecal coliforms deposited 

by wild ducks had multiplied in the beach sands and 

were carried into the water by storm runoff 

(Standridge and others, 1979). 

VARIATION IN NUMBER OF MICRO-ORGANISMS 

IN STORMWATER 

The microbiological character of runoff from any 

given storm and any given area is likely to be 

unique. This presents obvious problems, if one is to 

compare runoff between areas or within the same 

area over time. First, the land use and topography 

of the drainage area will have a significant effect 

on quality of the runoff. For example, urban stormwater and runoff from a feedlot will be very different from each other. Even within the general 

category of urban runoff, water from residential 

areas is likely to differ considerably from water 

from commercial areas in terms of microorganisms. 

If runoff from only one area is considered, time is 

the most significant variable because bacterial 

counts vary from storm to storm (Davis and 

others, 1977) and with season (see table 5). 

However, the amount of time elapsed between 

storms does not seem to affect microbiological 

quality, as indicated by Olivieri and others (1977, p. 

88), who, after studying storms in Baltimore, 

Maryland, for a 12-month period, concluded that 

"the levels of fecal coliforms observed in the storm 

runoff appear to be independent of the time between storms." Upon first consideration, this conclusion may seem unlikely because waste material 

will accumulate between storms. However, fecal 

organisms begin to die as soon as they leave their 

normal habitat, the gastrointestinal tract of warmblooded animals, so that only freshly deposited 

fecal bacteria are likely to affect the counts in 

runoff. 

Changes in bacterial counts during a storm have 

been documented by Davis and others (1977), who 

studied an area near Houston, Texas, that was being developed into a planned community. Figure 

13, which· shows a hydrograph and bacteriological 

data from one storm at a small stream site, indicates that the densities of both fecal coliforms 

and fecal streptococci rise sharply at the beginning 

of the storm and gradually decay over the next 60 

hours. The time from 20-50 hours represents the 

bulk of the hydrograph. During this period, the 

fecal coliform density decreases by slightly less 

than 1log unit (base 10), while the fecal streptococcal density decreases by just over 1 log unit. 

Qureshi and Dutka (1979) conducted a similiar 

study of stormwater draining a residential area in 

Burlington; Ontario, and emptying into a small 

creek. In that study, samples of runoff were collected at the outfall before entering the stream. 

Changes in bacterial counts at this site 

(arithmetic mean of 12 storms) are shown in figure 

14. Data for individual storms show no predictable 

pattern for maximum bacterial populations 

(Dutka, 1977; Qureshi and Dutka, 1979). In a 

report on the microbiological quality of stormwater from residential and commercial areas (3 

sites), Qureshi and Dutka (1979) concluded: 

"There appeared to be little relationship between the duration, 

intensity, and amount of rainfall and the occurrence of peak 

microbial populations. As a result, no typical pattern of timerelated distribution of indicator and pathogenic bacteria could 

be established in this investigation." 

The amount of time micro-organisms spend in 

the stormwater before collection and analysis has a 

significant effect on the final counts obtained. In 

general, storm runoff is a hostile environment for 

fecal indicator organisms and pathogens because 

these organisms require high nutrient levels and 

warm temperatures for growth. Furthermore, 

salts, organics, and other chemicals that may be in 

stormwater will have an adverse effect on microorganisms, so that the number of organisms in 

stormwater will decline with time. In addition, the 

relative concentrations will change with time 

because the strains die at differing rates. A more 

complete discussion of the importance of prompt 

analysis is presented in the section "Methods and 

Procedures." 

E26 
TABLE 5.-Densities of indicator bacteria in stormwater 

Source Total colifonns Fecal colifonns Fecal strep FC:FS Reference 

of stonrwater per 100 rnL per 100 rnL per 100 rnL Ratio 

Detroit - urban street VEibel and 

catch basins, 1949 25,000 - 930,000 not measured not measured not measured others, 1964 

Detroit - urban street 

catch basins, 1950 2,300 - 430,000 !h. !h. !h. Ih. 

Seattle - street gutters 

1959-1960 up to 16,100 !h. !h. !h. Ih. 

Stockholm, S\.Jeden - streets and parks, median: 4,000 

1945-1948 high: 200,000 !h. !h. !h. IX>. 

Pretoria, South Africa - residential, park, school, 

sports ground areas 240,000 !h. !h. !h. IX>. 

Pretoria, South Africa - business and flat area 230,000 [b. !h. [b. IX>. 

Cincinnati, Ohio - residential; 1962-1963 median 58, 000 median 1 0, 900 median 20, 500 0.53 ~. 

Cincinnati, Ohio - 1962-1964 Celdreich and 

Wooded hillside others, 1968 

Spring 2,400 190 940 0.20 

Suntner 79,000 1, 900 27,000 0.70 

fruturm 180,000 430 13,000 0.03 

Winter 260 20 950 0.20 

Street gutters 

Spring 1,400 230 3,100 0.07 

Suntner 90,000 6,400 150,000 0.04 Ih. 

Autum 290,000 47,000 140,000 0.34 

Winter 1,600 so 2,200 0.02 

Business district Ih. 

Spring 22,000 2,500 13,000 0.19 

Suntner 172,000 13,000 51,000 0.26 

Auturm 190,000 40,000 56,000 0.71 

Winter 46,000 4,300 28,000 0.15 

Rural Ih. 

Spring 4,400 55 3,600 0.02 

SUITI'!Er 29,000 2,700 58,000 o.os 

frutum 18,000 210 2,100 0.10 

Winter 58,000 9,000 790,000 0.01 

Baltimore - 907o less 

Inner city, residential than 4.0; 

coomercial - Pop. density mean of 24 s torrrs mean of 24 stonns mean of 24 stonns 80% less Olivieri and 

92.5/acre, 1974-1975 380,000 83,000 560,000 than 1.0 others, 1977. 

Baltimore - 90% less 

Residential/shopping than 4.0; 

center - Pop. density mean of 21 storms mean of 21 s tonns mean of 21 stonns 807o less Ih. 

26.6/acre, 1974-1975 38,000 6,900 50,000 than 1.0 

Business district not measured 13,000 51,000 0.26 Celdreich and 

Kenner, 1969 

Residential !h. 6,500 150,000 0.04 Ih. 

Rural !h. 2,700 58,000 o.os Ih. 

Burlington, Ontario - single-fanily 

residential 10,000 4,000 4,000 1.0 Q..treshi and 

J)ltka, 1979 

E27 
X 

(J) 10 LL 

u 8 ~ 

~ 6 

a: 

~ 4 u 5 2 

5 

40 60 

... ~ FECAL COLIFORMS 

I

' ........ •• • z_ ' • -------.- 1 • .----....... ~ 3 I ••'-.... z .... ,. • '-..! 6 2 ..... _...... • 

u 

80 

60 80 

g 4 ...J 

·~- FECAL STREPTOCOCCI .......................... . -.... , .. ............ __ ..... ______ ~ 

• 

TIME, IN HOURS 

FIGURE 13.-Hydrograph and graphs of densities of fecal coliforms and fecal streptococci bacteria in a small stream near 

Houston, Texas, during a storm (modified from Davis and 

others, 1977; reproduced with permission of Water Resources Bulletin). 

PATHOGENS IN STORMWATER 

Olivieri and others (1977) investigated the occurrence of pathogens in urban stormwater in 

Baltimore. An analysis of stormwater samples 

from a site not known to have any sewage 

overflows gave the results presented in table 6. 

It is obvious that urban stormwater can contain 

pathogens, although at relatively low levels. Could 

this represent a threat to human health? One way 

to begin to answer this question is to consider the 

number of bacteria and viruses necessary to 

establish infection. Unfortunately, some strains 

are more virulent than others. The susceptibility of 

a human population is also variable and depends on 

such considerations as age, general health, and 

degree of immunity. Nevertheless, rough estimates can be made from available data. A dose of 

105 Salmonella may be necessary to establish infection, but in the case of some Shigella strains, 102 or 

103 viable organisms is enough to infect (National 

Research Council, 1977). The situation with viruses 

is not as well documented; but, in general, a conservative approach is taken and one virus particle 

is considered sufficient to establish infection. It 

should be pointed out, however, that infection does 

not always lead to disease. 

By combining the known densities of pathogens 

in stormwater with the infective doses just cited, 

one might be able to make an estimate of the direct 

health hazard of urban runoff. From this line of 

reasoning, the amount of stormwater an individual 

would have to consume to acquire a Salmonella infection would be on the order of hundreds of 

gallons. The danger from virus infection may be 

greater because dilution will not destroy infectivity. It is unlikely that any single individual will 

consume enough stormwater to insure virus infection; but, if many individuals drink slightly contaminiated water, it is possible that one or a few 

may become infected . 

Stormwater does not need to be ingested to pose 

a health hazard. It is, perhaps, ironic that contamination of saltwater, which is not consumed, 

may be more dangerous than contamination of 

drinking water. For example, oysters and clams 

are known to concentrate viruses, and several 

well-documented outbreaks of hepatitis have been 

caused by virus-contaminated shellfish (Vaughn 

and Landry, 1977). Also, Pseudomonas aeruginosa 

and Staphylococcus au reus are associated with eye, 

ear, and skin diseases and might become a problem 

in areas where stormwater or surface waters, 

heavily contaminated with stormwater, are used 

for recreational activities. 

All of the pathogens mentioned above are of 

human origin and, as expected, are present in urban stormwater in relatively low numbers. The 

report on the Baltimore study discusses the issue 

at some length and concludes that urban stormwater probably represents little threat to public 

health (Olivieri and others, 1977). However, Van 

Donsel and others (1967, p. 1362) cite a U.S. Public 

Health Service analysis of waterborne-disease outE28 
5 Fecal Streptococcus 

6 .A Fecal Coliform 

X P. aeruginosa 

(!) 

0 4 ...J 5 

~ 

z 

::J 

0 

u 

~ 3 a: 4 w 

.... 

u 

<( 

co 

EXPLANATION 

3 • Coliform 2 

• Heterotroph 

2 3 4 2 3 4 

TIME OF COLLECTION, IN HOURS 

FIGURE 14.-Bacterial populations in runoff samples taken at stormwater outfall, Burlington, Ontario. Arithmetic mean of 

bacteriological data, based on 12 storms (from Qureshi and Dutka, 1979, reproduced from Water and Sewage Works, vol. 23, 

no. 3, March 1979, with permission). 

breaks occurring in' the United States and Puerto 

Rico from 1946 to 1960: 

"At least 29 outbreaks involving 9,233 cases were associated 

with stormwater runoff, caused either by rainfall washing 

human and animal feces or sewege into wells, springs, streams, 

reserviors, and open-water mains, or by wide-spread flooding of 

individual and public water systems." 

FATE OF MICRO-ORGANISMS IN 

STORMWATER 

As long as stormwater containing bacteria and 

viruses is confined to gutters, ditches, and storm 

sewers, it poses little threat to humans; but when it 

enters the receiving body, the viruses and bacteria 

may become a problem. Fortunately, bacteria that 

are adapted to live in or on a human host 

(pathogens and fecal indicators) are not well 

adapted for competition and survival in the aquatic 

ecosystem. McFeters and others (197 4) placed enteric bacteria from well water (temperature of 

9.5°C to 12.5°C) inside membrane dialysis 

chambers. The average time required for a 50 percent reduction in the initial population (half-time) 

was 17 hours for fecal coliforms and 22 hours for 

enterococci. Among a group of pathogens studied, 

TABLE 6. -Densities of pathogens in stormwater from 

Baltimore, Maryland1 

Salmonella species _______ _ 

Pseudomonas aeruginosa __ _ 

Staphylococcus aureus ____ _ 

Animal virus -------------

Polio virus --------------- Coxsackie Virus B --------- Echovirus 

Other unideritified~~s-----

Enterovirus --------------

' Data from Olivieri and others (1977). 

Percent 

occurrence 

in collected 

samples 

52 

100 

82 

83 

42 

50 

33 

8 

Geometric 

mean 

densities 

5.7 per 10 L 

592 per 100 mL 

12 per 100 mL 

170 Plaque 

forming units per 

10 L 

the longest half-time was 26.8 hours (Shigella sp.) 

and the shortest, 2.4 hours (Salmonella sp.). The 

authors observed no growth of the bacteria 

studied. 

Although viruses will not grow outside of their 

hosts, they apparently do not disappear as rapidly 

as bacteria. Studies cited by Vaughn and others 

(1979) indicate that human viruses may persist in 

aquatic environments for long periods of time, as 

much as 154 days in one example. 

E29 
Van Donsel and others (1967) studied the survival of indicator bacteria in soil using tracer 

strains that could be identified. They found that in 

summer, the fecal coliforms survived slightly 

longer than the fecal streptococci (3.3 days as compared to 2. 7 days for 90 percent reduction); 

whereas in spring and winter, the fecal streptococci survived much longer than the fecal coliforms. In 

autumn, survival time was the same for both types, 

and the shortest survival time for both groups was 

at an exposed site in summer. This study produced 

evidence of aftergrowth of nonfecal coliforms in 

the soil as a result of temperature and rainfall 

variations, and Van Donsel and others (1967) 

pointed out that such aftergrowth may contribute 

to variations in the bacterial count of storm runoff 

and that these variations have no relation to the 

sanitary history of the drainage area. The problem 

of aftergrowth of fecal coliforms deposited by 

ducks on a beach in Wisconsin (Standridge and 

others, 1979) already has been described. 

Generally, soil is effective in removing bacteria 

and viruses. Gerba and others (1975) presented an 

extensive review of the topic. Factors that affect 

the survival of bacteria in soil are moisture content 

and moisture-holding capacity, temperature, pH, 

sunlight, availability of organic matter, and competition and antagonism from soil microflora. 

Movement of bacteria into the ground water is 

limited by the straining of bacteria at the soil surface and by adsorption to clays in the soil. 

Deterioration of ground-water quality from the 

downward movement of micro-organisms is likely 

to be a problem only where the water table is near 

land surface. 

A recent study (Yeager and O'Brien, 1979) concluded that drying is the most important factor in 

the inactivation of viruses in soils. Virus removal 

by soils depends mainly on adsorption by clays 

(Gerba and others, 1975). However, viruses that 

are adsorbed to clays can retain their infectivity 

(Schaub and Sagik, 1975). Binding of virus particles to clay depends on the net charge of the 

virus, which in turn is dependent on pH. Unfortunately, the viruses are not bound irreversibly to 

the soil particles and may be desorbed by rain (Landry and others, 1979). Wellings and others (1975) 

have isolated viruses from ground water under a 

recharge basin after a period of heavy rain. 

The movement of pathogens to ground water 

may be a problem in any area where storm runoff 

is stored in basins or used for artificial recharge. 

The largest source of information on this hazard is 

the literature on artificial recharge of ground 

water and land treatment of sewage. Both practices present a greater potential contamination 

problem than storm runoff, since pathogens likely 

will be applied to the same land area in greater 

numbers over a long period of time. Several 

studies indicate that ground-water contamination 

from artificial recharge is possible. For example, 

Vaughn and others (1978) found viruses and 

bacteria in ground water beneath recharge basins 

at two Long Island, New York, sewage treatment 

plants where the depth to water was 18 and 30 

feet, but found no viruses under basins that were 

80 feet above the water table. However, even 

where viruses and bacteria were found in the 

ground water, the numbers had been reduced considerably during passage through the soil. 

Although the numbers of bacteria and viruses are 

likely to be much lower in urban stormwater than 

in treated wastewater, Vaughn and Landry (1977) 

recovered viruses from ground water beneath a 

storm water-recharge basin on Long Island. The 

source of these viruses has not been definitely 

established; however, their presence suggests the 

possibility of ground-water contamination by 

stormwater. 

METHODS AND PROCEDURES 

A detailed discussion of the methods used to 

grow and enumerate micro-organisms in stormwater is beyond the scope of this paper. Other 

sources (American Public Health Association, 

1975; Greeson and others, 1977) give much information on the procedures used to cultivate indicator bacteria (fecal coliforms and fecal streptococci). Pathogenic organisms are more difficult 

to cultivate, and great care must be exercised to 

avoid laboratory-acquired infections. Viruses must 

be grown inside a living host cell, and virus concentration and propagation are complicated processes 

that should be left to specialists in virology. 

Setting aside any discussion of the methods used 

in processing a water sample, an important issue 

remains. If the sampling program is improperly 

designed or if the water sample is collected improperly, the data will be at best meaningless or at 

worst misleading. Some of the points that must be 

considered in sampling are addressed below. 

E30 
SAMPLE COLLECTION 

STORMWATER 

Water samples for microbiological analysis 

should be collected by hand in sterile containers by 

a trained individual (American Public Health 

Association, 1975). Two early reports (Burn and 

Vaughn, 1966; Weibel and others, 1964) mentioned 

allegedly sterile automatic-sampling devices for 

storm water; however, the descriptions of the 

devices are somewhat incomplete and leave doubt 

as to the integrity of the sample. Almost all recent 

papers indicate that stormwater samples were collected by hand in sterile containers. This is obviously expensive, time consuming, and some may 

feel unnecessary. It might be argued that since 

stormwater is already heavily contaminated, slight 

additional contamination from the sampling device 

will be negligible. However, this is circular reasoning, and the true level of contamination in the 

stormwater can never be established by this approach. Another argument might be that fecal coliforms and fecal streptococci are unlikely to be in 

the air or in washed bottles and automatic 

samplers. This argument may seem reasonable; 

nevertheless, microbiological data reported for 

samples collected under nonsterile conditions 

always will be suspect. 

RECEIVING WATERS 

Collecting stormwater at the outfall of a storm 

drain or sewer will give the best estimate of the 

microbiological quality of the runoff. However, to 

judge the impact of stormwater on some environments, it may be necessary to sample a 

stream or other receiving water. In lakes, reservoirs, deep rivers, and estuaries, bacterial abundance may vary transversely, with depth, and with 

time of day. The time variable was pointed out by 

Davis and others (1977) in their study of a stream 

receiving storm runoff. In their study, bacterial 

densities were not constant in stream waters over 

a period of 24 hours, even during constant low-flow 

conditions. Their data showed highly variable 

batch flows, and they concluded that grab samples 

reflected the condition and content solely at the 

time at which the sample was taken. A possible 

solution to this problem might be to collect samples 

upstream from the discharge point to serve as a 

control for each stormwater sample taken. 

STORAGE OF STORMWATER SAMPLES 

The rule when working with micro-organisms is 

that the more rapidly the samples are processed, 

the more accurate the results will be. Bacterial 

samples must be refrigerated if held more than 1 

hour before processing; the maximum transport 

time is 6 hours, and the samples must be processed 

within 2 hours after arrival at the lab (American 

Public Health Association, 1975; Greeson and 

others, 1977). 

A measure of the potential for survival of various 

bacterial groups in stormwater is presented in 

figure 15, taken from Geldreich (1976). In that 

study, urban storm water was collected, filtered to 

remove the "native" bacterial populations, and 

pure cultures were added to the water. The culture 

densities then were determined over a period of 2 

weeks. Problems with this approach are that the 

laboratory cultures might have declined at a different rate than those occurring naturally in 

stormwater and also that the added bacteria might 

have survived longer because they did not have to 

compete with the "native" population. 

As figure 15 indicated, the apparent number of 

bacteria decreased significantly with storage time 

and different types of bacteria died at different 

rates. In general, the bacteria survived longer 

when stored at 10°C (fig. 15A) than at 20°C (fig 

15B), but even when stored at 10°C (fig 15A), 50 

percent of the Salmonella typhimurium died 

within 2 days. Fecal coliforms survived slightly 

longer, with 50 percent surviving for 14 days, 

whereas Streptococcus bovis (a component of 

animal feces but not human feces) disappeared 

within the first day at 10°C and within 3 days at 

20°C. These data clearly support the general rule 

that rapid processing of microbiological samples is 

of utmost importance. 

DESIGN OF SAMPLING PROGRAM 

When designing a program to determine the 

microbiological quality of stormwater, one must 

consider the following points: 

(1) The number and type of bacteria in runoff will 

be influenced by the intensity of the storm, 

duration of the storm, season of the year, and 

land use of the area being drained. Therefore, 

great care must be taken in any extrapolation 

from one area to another or from one storm to 

another. 

E31 
....J 

<( 

> 

> 

a: 

::J 

50 

en 10 

1-

z 

w 

u 

a: 

~ 5 

....J 

<( 

> 

> a: 

~ 10 

1-

z 

w 

u 

ffi 5 a.. 

I 

I 

I 

I SB 

I 

I 

I 

A 

8 

10 12 

LENGTH OF TIME, IN DAYS 

........ 

. ...._ ........ '--...... SF 

--....... 

----------------------

10 12 

LENGTH OF TIME, IN DAYS 

FIGURE 15.-Persistence of selected bacterial strains in stormwater: A, stored at 10°C; B, stored at 20°C. SF=Streptococcus faecalis, FC =fecal coliform bacteria, 

ST = SalmoneUa typhimurium, SR =Streptococcus bovis 

(modified from Geldreich, 1976; reprinted with permission 

from Critical Reviews in Environmental Control, 

copyright by the Chemical Rubber Company, CRC Press, 

Inc.). 

(2) Provision must be made for a nearby laboratory 

to handle all samples. Samples must be processed within hours of collection. 

(3) No good predictor or indicator for the presence 

of pathogens in stormwater is known. 

Pathogenic bacteria and viruses seem to occur 

in a random fashion throughout the storm. 

SUMMARY AND CONCLUSIONS 

Information obtained from recent investigations 

of micro-organisms in stormwater has been reviewed and summarized with the intent of providing assistance to those planning studies of 

stormwater. The following conclusions are indicated by this review: 

(1) Most stormwater contains relatively large 

numbers of total coliforms, fecal coliforms, and 

fecal streptococci. 

(2) Most of the total coliforms in runoff are native 

soil organisms. 

(3) The source of most fecal coliforms and fecal 

streptococci in runoff is probably wild or 

domestic animals. 

( 4) A report from Baltimore, Maryland, has 

documented the occurrence of hazardous 

organisms in urban runoff. The source of the 

contamination is unknown . 

(5) Stormwater probably presents little direct 

threat to human health because it is not ingested, but it can produce other negative effects such as contamination of beaches or 

shellfish. 

(6) Contamination of surface waters by bacteria in 

stormwater is of short duration because indicators and pathogens die rapidly in the 

aquatic environment. Viruses apparently do 

not disappear as rapidly as bacteria and may be 

concentrated by shellfish. 

(7) Bacteria and viruses deposited on soil by stormwater are inactivated by drying, competition 

from soil microflora, and a variety of other processes. The movement of pathogens to ground 

water is likely to be a problem only where the 

water table is near land surface. 

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American Public Health Association, 1975, Standard methods 

for the examination of water and wastewater, 14th ed.: 

Washington, D.C., American Public Health Association, 

1193 p. 

Burn, R. J., and Vaughan, R. D., 1966, Bacteriological 

comparison between combined and separate sewer 

discharges in southwestern Michigan: Journal Water Pollution Control Federation, v. 38, p. 400-409. 

Carberry, J. B., and Stapleford, L. R., 1970, A consideration of 

microbial water quality analysis criteria: Journal American 

Water Works Association, v. 71, p. 213-218. 

Davis, E. M., Casserly, D. M., and Moore, J.D., 1977, Bacterial 

relationships in stormwaters: Water Resources Bulletin, v. 

13, p. 895-905. 

E32 
Dutka, B. J., 1973, Colifonns are an inadequate index of water 

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a 100% residential area in Canada: Canadian Research, v. 

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Geldreich, E. E., 1976, Fecal colifonn and fecal streptococcus 

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Geldreich, E. E., Best, L. C., Kenner, B. A.,and Van Donsel, 

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Geldreich, E. E., and Kenner, B. A., 1969, Concepts of fecal 

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C. A., 1979, Adsorption of enteroviruses to soil cores and 

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Environmental Microbiology, v. 38, p. 680-687. 

Marzouk, Y., Goyal, S.M., and Gerba, C. P., 1979, Prevalence of 

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McFeters, G. A., Bissonnette, G. K., Jezeski, J. J., Thomson, 

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-- 1979b, Microbiological studies on the quality of urban 

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212-222. 

Standridge, J. H., Delfino, J. J., Kleppe, L. B., and Butler, R., 

1979, Effect of waterfowl (Anas platyrhynchos) on indicator bacteria populations in a recreational lake in 

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Van Donsel, D. J., Geldreich, E. E., and Clarke, N. A., 1967, 

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E33