Effect of pH on the growth and activity of heterotrophic sediment microorganisms

Chemosphere,Vol.ll,No.lO,pp Printed in Great Britain

973-983,1982

0045-6535/82/100973-11503.00/0 © 1 9 8 2 Pergamon Press Ltd.

EFFECT OF pH ON THE GROWTH AND ACTIVITY OF HETEROTROPHIC SEDIMENT MICROORGANISMS

M. D. Baker, W. E. Inniss and C. I . M a y f i e l d * Department o f B i o l o g y , U n i v e r s i t y o f Waterloo, Waterloo, O n t a r i o , Canada N2L 3G1 and

P. T. S. Wong Great Lakes F i s h e r i e s Research Branch Canada Centre f o r Inland Waters B u r l i n g t o n , O n t a r i o , Canada L7R 4A6 ABSTRACT Low pH markedly reduced h e t e r o t r o p h i c m i c r o b i a l a c t i v i t y

in s e d i m e n t - l a k e water systems.

A

marked r e d u c t i o n in both the rate and the t o t a l amount o f oxygen consumed occurred as pH decreased.

In a d d i t i o n ,

low pH and a l s o low temperature (O°C) r e s u l t e d in longer t u r n o v e r

times (Tn) and a l s o s m a l l e r V I

values when the k i n e t i c s o f 14C-glucose, 14C-glycine, and

4C_g I utamic a c i d u t i l i z a t i o n max were examined. When the response o f the microorganisms to heavy

metals was examined, mercury was g e n e r a l l y more t o x i c than lead over the pH range o f 4.5 to 7.5 but both metals were more t o x i c a t pH 4.5 than a t h i g h e r pH l e v e l s . oxygen u t i l i z a t i o n

and 14C-glucose m i n e r a l i z a t i o n more than lead a t a l l

reduced m i c r o b i a l a c t i v i t y

at all

Mercury a l s o i n h i b i t e d pH l e v e l s .

Bisulfite

pH l e v e l s although i t was always more t o x i c to the sediment

microorganisms at pH values o f 4 and 5 than a t pH 7.

INTRODUCTION Acid p r e c i p i t a t i o n has decreased the pH o f many poorly buffered lakes, r i v e r s , and streams in North American and in Scandinavian countries ( I ) . uncommon (2).

In many lakes, pH values of 4 to 4.5 are not

I f the present trends o f a c i d i f i c a t i o n continue, or increase, many more waters

w i l l be seriously affected (3-5).

Although some of the biological aspects of a c i d i f i c a t i o n

973

974

have been s t u d i e d

(6, 7 ) ,

the e f f e c t s

o f low pH on microorganisms has received l i t t l e

I t has been r e p o r t e d t h a t o r g a n i c d e b r i s composed mainly o f coarse d e t r i t u s and mecrophytes) accumulates on the s u r f a c e - s e d i m e n t s o f a c i d i f i e d

attention.

( l e a v e s , Sphagnum

lakes (8).

Microorganisms are

thought to be o f fundamental importance w i t h regard to the decomposition of o r g a n i c m a t t e r and nutrient

cycling

in a c i d i c activity

in most lake ecosystems.

Thus, the apparent lack o f o r g a n i c m a t t e r decomposition

lakes may be the r e s u l t o f a r e d u c t i o n in m i c r o b i a l a c t i v i t y

(8, 9).

The growth and

o f the m i c r o b i a l p o p u l a t i o n o f lakes is a f f e c t e d by many f a c t o r s and t h e r e f o r e ,

difficult

t o d e t e r m i n e whether m i c r o b i a l a c t i v i t y

a l o n e , o r as the r e s u l t o f the i n d i r e c t the e f f e c t

is

in a c i d i c lakes is suppressed due to low pH

e f f e c t s o f pH.

o f pH, heavy m e t a l s , and b l s u l f l t e

it

Consequently, in the present i n v e s t i g a t i o n ,

on the growth and a c t i v i t y

of h e t e r o t r o p h i c

sediment m i c r o o r g a n i s m s was examined.

MATERIALS AND METHODS

Sediment Sediment was o b t a i n e d in O c t o b e r , 1979 from P l a s t i c Lake, a small a c i d i c , o l i g o t r o p h i c l o c a t e d near D o r s e t , O n t a r i o ( l a t i t u d e

45°!1'N;

l o n g l t u d e 78°50'W).

lake

The sediment was c o l l e c t e d

w i t h an Eckman grab sampler, packed and t r a n s p o r t e d in i c e , and stored in the l a b o r a t o r y a t 4°C. E f f e c t o f pH and heavy metals on oxygen uptake To d e t e r m i n e the e f f e c t lake w a t e r s l u r r y a magnetic s t i r r e r .

o f pH on the oxygen uptake o f the lake sediment, a 500-mL volume o f sediment-

(1.5 Darts o f wet weiQht s e d i m e n t : l p a r t lake w a t e r , pH 5 . 8 - 6 . 0 ) was mixed w i t h After equi'libration

5 . 5 , 6 . 5 , and 7.5 w i t h ! o r 2 N s u l f u r l c used since i t

at 20=C, the sediment was a d j u s t e d t o pH values o f 4 . 5 , a c i d and 1 o r 2 N sodium h y d r o x i d e .

is one o f the major a c i d s found in a c i d i c p r e c i p i t a t i o n

(10).

S u l f u r i c a c i d was A 5-mL volume o f

each p H - a d j u s t e d sediment was added to the main chamber o f a Warburg f l a s k and oxygen uptake was measured f o r 60 min, using standard manometric procedures. S i m i l a r s t u d i e s were a l s o conducted to determine the e f f e c t o f pH and heavy metals on the oxygen uptake o f the lake sediment.

Samples (4 mL) o f s e d i m e n t - l a k e w a t e r s l u r r i e s ,

pH values o f 4.5 and 7.5, were placed in Warburg f l a s k s , of either

a lead c h l o r i d e o r m e r c u r i c c h l o r i d e s o l u t i o n .

the sidearms o f which contained I mL After equilibration

a t 20°C f o r 10 min,

the metals were added to the sediments and the oxygen uptake was measured f o r 30 min. c o n c e n t r a t i o n o f each metal c a t i o n was 60 ug mL- I o f sediment s l u r r y . used f o r each t r e a t m e n t in a l l

a d j u s t e d to

oxygen uptake s t u d i e s .

E f f e c t o f pH and heavy metals on the growth o f sediment microorganisms

The f i n a l

Three r e p l i c a t e s were

975

The e f f e c t o f pH and heavy metals on the growth o f a mixed p o p u l a t i o n o f h e t e r o t r o p h i c sediment microorganisms was a l s o examined.

The mixed c u l t u r e , o r i g i n a l l y

sediment, was m a i n t a i n e d on a l i q u i d medium a t 20=C. ( D i f c o ) , 5 g; g l u c o s e , 1 g; yeast e x t r a c t ,

The medium c o n s i s t e d of n u t r i e n t

t g; d i s t i l l e d

replication)

c o n t a i n i n g 4.8 mL

l i q u i d medium which was a d j u s t e d to pH values o f 3.5, 4.5, 5 . 5 , 6 . 5 , and

7.5 by the a d d i t i o n o f 0.5 o r I N s u l f u r i c volume o f s t e r i l e

broth

w a t e r , 1000 mL.

Growth experiments were performed w i t h c u l t u r e tubes ( t r i p l i c a t e of filter-sterilized

o b t a i n e d from 1 g o f

a c i d and 0.5 o r 1N sodium hydroxide.

solution containing either

A O.l-mL

lead a c e t a t e o r m e r c u r i c c h l o r i d e was a s e p t i c a l l y

added to each o f the tubes, so t h a t c o n c e n t r a t i o n s o f 1, 10 and 100 ug mL- I o f the metal c a t i o n were o b t a i n e d .

Each tube was i n o c u l a t e d w i t h O.l mL (4.5 x lO6 c e l l s )

sediment microorganisms.

o f a 24-h c u l t u r e of the

Controls c o n t a i n i n g the i n o c u l a t e d , pH-adjusted medium w i t h o u t the

metal amendments were a l s o i n c l u d e d .

Incubation was at 20°C on a r o t a r y shaker, and growth was

measured s p e c t r o p h o t o m e t r l c a l l y at 650 nm (0D650). K i n e t i c s o f 14C-~lucose and 14C-amlno a c i d m i n e r a l i z a t i o n To o b t a i n more i n f o r m a t i o n on the e f f e c t o f pH on the m i c r o b i a l a c t i v i t y the a b i l i t y

and a l s o to determine

o f the sediment microorganisms to degrade o r g a n i c m a t t e r , methods s i m i l a r

o f Harrison e t al. ( l l )

were employed.

The e f f e c t o f pH and temperature on the m i n e r a l i z a t i o n

o f the u n i f o r m l y - l a b e l e d s u b s t r a t e s 14C-glucose ( s p e c i f i c a c t i v i t y 14C-glycine ( s p e c i f i c a c t i v i t y

to those

o f 260 mCi/rnmole),

o f 112 mCi/mmole), and 14C-glutamic a c i d ( s p e c i f i c a c t i v i t y

265 mCi/n~nole) was determined a f t e r

14 days o f i n c u b a t i o n .

All

radioactively-labeled

of

compounds

were o b t a i n e d from Amersham-Searle L t d . , O a k v i l l e , O n t a r i o . In the k i n e t i c

s t u d i e s , sediment-lake w a t e r systems were a d j u s t e d to pH values o f 4 and 7 by the

addition of 1 or 2 N sulfuric

a c i d and 1 o r 2 N sodium h y d r o x i d e , placed in aluminum f o i l -

covered beakers to minimize moisture l o s s , and incubated a t O=C and 20°C.

Each sediment s l u r r y

was maintained a t the a p p r o p r i a t e pH l e v e l throughout the e x p e r i m e n t a l p e r i o d . z a t i o n o f the r a d i o a c t i v e l y - l a b e l e d sediment microorganisms. control

The m i n e r a l i -

compounds to 14C02 measured the c a t a b o l i c a c t i v i t y

of the

D u p l i c a t e 50-mL h y p o - v l a l s were used f o r each t r e a t m e n t plus one

f o r each t r e a t m e n t .

Each h y p o - v i a l contained 0.1 pCi o f the a p p r o p r i a t e 14C-compound,

and c o n c e n t r a t i o n s o f unlabeled compound ranging from 0.05 to 5.0 Pg. t r e a t e d w i t h 1 mL o f 2 N s u l f u r i c

Control v i a l s were

a c i d to measure any blank r a d i o a c t i v i t y .

At each sampling

t i m e , 4 mL of the a p p r o p r i a t e sediment was added to each h y p o - v i a l , the v i a l s were sealed w i t h rubber serum caps which held a p l a s t i c N . J . ) , and incubated at e i t h e r O°C o r 20°C. Whatman #1 f i l t e r

cup and rod assembly (Kontes Glass Co., Vineland, The cup assembly contained a f l u t e d piece o f

paper (2.5 x 5.0 cm) suspended in the a i r space above the sediment.

ensure t h a t the system was a i r t i g h t sealed w i t h s i l i c o n e

rubber s e a l a n t .

To

and would prevent any leakage o f 14C02, each serum cap was M i n e r a l i z a t i o n was allowed to proceed f o r 10 min at 20°C,

w h i l e a 60-min i n c u b a t i o n was used a t O°C, a f t e r which the m i n e r a l i z a t i o n was stopped by injecting

1 mL o f 2 N s u l f u r i c

acid i n t o the sediment.

The a d d i t i o n o f the s u l f u r i c

lowered the pH o f the samples to a p p r o x i m a t e l y I and released a l l

acid

14CO2. A f t e r 1 h, O.15 mL o f

976

B-phenethylamine was i n j e c t e d onto the f i l t e r the released 14CO2. The r a d i o a c t i v i t y scintillation

cocktail

scintillation

counter (Model LS81OO).

and a l l

paper and l e f t

f o r an a d d i t i o n a l

1 h to trap a l l

due to the 14CO2 was measured in 10 mL o f PCS l i q u i d

(Amersham-Searle L t d . , O a k v i l l e , O n t a r i o ) ,

using a Beckman l i q u i d

Quenching was corrected f o r by the c h a n n e l s - r a t i o method

samples were corrected for c o n t r o l a c t i v i t y .

The 14C02 t r a p p i n g e f f i c i e n c y

of the

B-phenethylamine had been p r e v i o u s l y determined by the generation o f a known amount of 14C02 from NaHI4co 3 by treatment w i t h 2 N s u l f u r i c

acid, and was between 95 and 100~.

The Lineweaver-Burk equation modified by Wright and Hobble (12, 13) was employed in these studies to c a l c u l a t e the m i n e r a l i z a t i o n turnover time (Tn) at the natural substrate c o n c e n t r a t i o n , and also the maximum v e l o c i t y o f m i n e r a l i z a t i o n (Vmax) , expressed as ~g 14 c - l a b e l e d substrate mineralized g-| sediment dry weight h -1. E f f e c t o f pH,, b i s u l f i t e

and heavy metals on 14C-~lucose m i n e r a l i z a t i o n

Sedlment-lake water systems were amendedwith sodium b i s u l f i t e (NaHSO3) so that f i n a l concen-I t r a t i o n s o f I, lO, IO0, 1000, and lO000 ug b l s u l f i t e mL of sediment s l u r r y were obtained. Following t h l s , the pH of the sediment s l u r r i e s w a s a d j u s t e d to values of 4, 5, and 7 wlth I or 2 N hydrochloric acid and l or 2 N sodium hydroxide.

Hydrochloric acid rather than s u l f u r i c acld

was used to lower the pH of the sediments to avoid any possible interference between b l s u l f i t e and s u l f a t e .

Unamendedcontrols were also included f o r each pH l e v e l .

The beakers containing

the sediment systems were covered with aluminum f o l l to minimize moisture loss, and placed at 20°C. A f t e r 7 days, 4 mL o f sediment were added to r e p l i c a t e 50-mL hypo-vials containing 0.1 ~CI of 14C-glucose.

The v i a l s were sealed and mineraIization was allowed to proceed f o r 5 min

at 20°C. Three r e p l i c a t e v i a l s were used for each treatment and one other v i a l was treated with 2 N s u l f u r i c acid to serve as a control.

All other procedures were the same as described in

the k i n e t i c experiments above. Similar methods were also used to determine the e f f e c t o f pH and heavy metals on the mineralization o f 14C-glucose.

Sediment-lake water s l u r r i e s were adjusted to pH values of 4 and 7 by

the use of I or 2 N s u l f u r i c acid and I or 2 N sodium hydroxide.

The sediment-lake water

systems were amendedwith e i t h e r lead chloride or mercuric chloride so that a f i n a l concen-I t r a t l o n o f I00 ug mL o f metal cation was obtained. Unamendedcontrols at each pH level were also included.

The sediment s l u r r i e s were incubated at 20°C and a f t e r 7 days the e f f e c t o f

these treatments on glucose m i n e r a l i z a t i o n was determined according to the methods prevlously described.

RESULTS

Oxygen uptake studies

977

100,

Fig. I. bJ

uptake in the Plastic Lake sediment.

//.~'/"

(3 bJ it) J=k Ld ,,r

Effect of pH on the microbial oxygen Treatments: pH 4.5 (o); pH 5.5 (A); pH 6.5 ( 4 ) ; pH 7.5 r e p r e s e n t s the mean

(e).

Each p o i n t

of three

replicates.

~<4o I!

i

: / ,,1 , ` /

ii

,I"

~1/ .tl/ / ~A.-"J '~~- Oo~ - - - o ~ ° I 1~/i,.5-O"~

oV~-° •

0

10

•

!

20

I

I

30 40 MINUTES

50

•

60

1OO,

F i g . 2.

BO

Effect of pH and heavy metals on the microbial oxygen uptake in the Plastic

LU

O

Lake sediment.

Treatments: pH 4.5

(m); pH 4.5 with lead (o); pH 4.5 with

~so

mercury (&); pH 7.5 ( o ) ; pH 7.5 w i t h lead ( o ) ; pH 7.5 w i t h mercury (A).

°/

Each p o i n t r e p r e s e n t s the mean o f three r e p l i c a t e s .

____Ida. ~ I.~71 0

i

,----

10

i l

I

.

20 MINUTES

.• .

3O

978

Oxygen uptake was r e a d i l y a f f e c t e d by sediment pH (Fig.

I).

As the sediment pH decreased, a

corresponding decrease in the r a t e and a l s o the e x t e n t o f oxygen u t i l i z a t i o n the f i r s t

uptake r a t e s were 2 . 4 , respectively.

the t o t a l ,

During

1.8, 1.1, and 0 . 8 UL o f oxygen consumed g - ] sediment dry w e i g h t min - i ,

T o t a l oxygen consumption over the 60-mln i n c u b a t i o n a ] s o decreased w i t h

decreasing pH. the t o t a l

occurred.

I0 mln o f i n c u b a t i o n at sediment pH values o f 7 . 5 , 6 . 5 , 5 . 5 , and 4 . 5 , the oxygen

I f the t o t a l amount o f oxygen consumed a t pH 7.5 represented 100~ (60 UL), then

amounts o f oxygen taken up a t pH values o f 6 . 5 , 5 . 5 , and 4.5 were 62, 45 and 33t o f respectively.

Lead had l i t t l e ,

i f any, i n h i b i t o r y

e f f e c t on the a e r o b i c r e s p i r a t i o n which was in c o n t r a s t t o

the r e s u l t s o b t a i n e d w i t h mercury ( F i g . 2 ) . oxygen uptake was s l i g h t l y the i n h i b i t i o n

At pH 4.5, the i n h i b i t o r y

more pronounced than a t pH 7.5.

o f oxygen uptake at pH 7.5 was h i t

effect

o f mercury on

At the end o f the 30-min i n c u b a t i o n ,

(mercury) and 6~ ( l e a d ) , compared t o 57~ and

5~, r e s p e c t i v e l y , at pH 4.5. Growth Studies The e f f e c t o f pH and various c o n c e n t r a t i o n s o f lead and mercury on b a c t e r i a l tested.

The growth o f the mixed b a c t e r l a l

growth was a l s o

p o p u ] a t i o n was a f f e c t e d by not only the pH o f the

medium, but a l s o by the presence o f the heavy metals and by v a r i o u s combinations o f pH w i t h heavy metals ( F i g . 3). the t o t a l

bacterial

Analysis o f the e f f e c t s o f pH alone showed t h a t low pH g r e a t l y reduced

growth w i t h no growth o c c u r r i n g a t pH 3.5.

90~ compared to pH 7.5.

3~

At pH 4 . 5 , growth was reduced

As the pH increased above 4.5, a corresponding increase in the growth

8O-

Fig. 3.

0nr

E f f e c t o f pH and heavy metals on the growth o f a mixed m i c r o b i a l

o

Z

_o

~

i

L

'

~

.

.

5

6

kin T

.

7

.

8

populatlon.

Inset - E f f e c t o f pH

on growth.

Concentration o f metal

cation

(ug mL-1): Pb ] ( o ) ; Pb IO

(A); Pb ]00 ( o ) ; Hg I ( e ) ; Hg lO

z 4o.

(&); Hg lO0 (m).

Each p o i n t

represents the mean o f three replicates.

2O" O

0 4

,

fl

D

~ pH

~

979

was apparent.

In this study, the effects of heavy metals alone, and als~ the combined effects

of pH and heavy metals could be observed.

Assuming a control pH of 7.5, the toxic effects

due to the metals alone showed that lead concentrations of I, 10, and 100 ~g mL-I had r e l a t i v e l y l i t t l e e f f e c t , i n h i b i t i n g the bacterial growth by only 2, O, and 13%, respectlvely. c o n c e n t r a t i o n s , mercury i n h i b i t e d growth by 31, 97, and lOOt, r e s p e c t i v e l y . pH 4 . 5 , a combined e f f e c t

due to both pH and heavy metals was noted.

At the same

I n t e r e s t i n g l y , r at

For example, lead at

c o n c e n t r a t i o n s o f 1, 10, and 100 pg mL"1 i n h i b i t e d growth by O, 87, and 100~, r e s p e c t i v e l y , w h i l e mercury a t the same c o n c e n t r a t i o n s c o m p l e t e l y i n h i b i t e d growth. microbial

cells

were much more s e n s i t i v e

Thus, a t pH 4 . 5 ,

the

to the heavy metals.

K i n e t i c s o f 14C-~lucose and 14C-amlno a c i d m i n e r a l i z a t i o n The e f f e c t o f pH and temperature on the m i n e r a l i z a t l o n o f u n i f o r m l y r a d i o a c t i v e l y - l a b e l e d compounds is presented in Table 1. parameters o f m i n e r a l l z a t i o n .

14C-

Both pH and temperature markedly a f f e c t e d the k i n e t i c

At 20°C in sediments at pH 4, the t u r n o v e r times were longer

w h i l e the V values were s m a l l e r f o r a l l o f the 14C-labeled s u b s t r a t e s examined compared to max the same k i n e t i c parameters measured a t pH 7. S i m i l a r r e s u l t s were o b t a i n e d a t both pH l e v e l s when the sediments were incubated at O°C, although a large r e d u c t i o n in a c t i v i t y at the lower temperature.

r a p i d l y , w h i l e both 14C-glucose and 14C-glycine were u t i l i z e d

less e f f i c i e n t l y .

Table I , E f f e c t of pH and temlxerature on the mineralization of ieC-uniformly labeled organic ~nds tn I lake water-sedlment system

14C-labeled

pH

compound

Temperature ('C)

Kinetics of mineralization Turnover time

Vmaa

(hours)

14C-glucose

14C-glycine

14C-glutam;c

was observed

On a comparative b a s i s , 14C-glutamlc a c i d was m i n e r a l l z e d the most

4

0 2O

10.30 1.5g

0.19 4 .BJ

7

0 20

7.83 0.67

2 .&9 8.76

4

0 20

12.22 1.21

0.;0 2.20

7

0 20

3.07 0.89

O.&2 7,00

4

0

6.09

0.92

7

20 0

0.65 1.15

IO.98 2.81

20

0.15

15.97

acid

* IJg Ikc-labeled substrJte mlnerallzed g-I sediment dry weight h "1

980

E f f e c t o f pH~ b i s u l f i t e

and heavy metals on

14C-91ucose

mineralization

Bisulfite

and pH e x e r t e d i n h i b i t o r y e f f e c t s on m i n e r a l i z a t i o n ( F i g . 4). At b i s u l f i t e -1 c o n c e n t r a t i o n s o f I and I0 gg mL , glucose m i n e r a l i z a t i o n was o n l y s l i g h t l y i n h i b i t e d . c o n c e n t r a t i o n s o f lO0 and 1000 pg mL-1 the i n h i b i t o r y c o n c e n t r a t i o n o f i0000 pg mL- l o f b i s u l f i t e , At each pH l e v e l , controls.

the i n h i b i t i o n

e f f e c t s were more pronounced, and at a

m i n e r a l i z a t i o n was almost c o m p l e t e l y i n h i b i t e d .

o f m i n e r a l i z a t i o n was c a l c u l a t e d r e l a t i v e

to the n o n - t r e a t e d

The pH o f the sediment systems a l s o had a marked e f f e c t on the t o x i c i t y

to the sediment microorganisms.

Bisulflte

s l u r r y was c o n s i s t e n t l y more i n h i b i t o r y

At

of blsulfite

a t c o n c e n t r a t i o n s o f 1 to 1000 pg mL"1 o f sediment

to glucose m i n e r a l i z a t i o n a t pH values o f 4 and5 than

at pH 7.

100~

• __

,,A

Z

o 80

Fig.

N q¢ (Z t~ Z m

=i

Percentage i n h i b i t i o n mineralization

o f 14C-glucose

in a lake w a t e r -

sediment system by pH and b l s u l f i t e . 60,

Concentration o f b i s u l f i t e sediment s l u r r y ) :

Z

o

CO -r

4.

1 (el;

100 (m); 1000 ( o ) ;

(~g mL-1 lO ( o ) ;

10000 (A).

Each

40,

p o i n t represents the mean o f t h r e e

z_

replicates. (.~ ¢:

20,

0 DH

When the t o x i c e f f e c t s o f pH, mercury, and lead on the m i n e r a l i z a t i o n o f glucose were examined, lead had l i t t l e

e f f e c t on m i n e r a l i z a t i o n ,

values o f 4 and 7, r e s p e c t i v e l y . at pH 4 and by 41~ a t pH 7.

producing i n h i b i t i o n

T h e r e f o r e , a t low pH, mercury was much more t o x i c to the sediment

microorganisms and markedly decreased the p o t e n t i a l sediment.

values o f o n l y I% and 4% a t pH

On the o t h e r hand, mercury i n h i b i t e d m i n e r a l i z a t i o n by 78% f o r glucose m i n e r a l i z a t i o n

in t h i s

981

DISCUSSION

Measurement o f the e f f e c t s o f pH on the sediment m i c r o b i a l a c t i v i t y

using both oxygen uptake

and s u b s t r a t e m i n e r a l i z a t i o n techniques, revealed t h a t both low pH and low temperature markedly reduced m i c r o b i a l a c t i v i t y activity

in the lake water-sediment systems.

A reduction in m i c r o b i a l

at low pH has a l s o been observed by o t h e r workers (14-18).

of relatively

Such reduced m i n e r a l i z a t i o n

simple o r g a n i c compounds suggests that the more complex and r e c a l c i t r a n t

commonly found in the o r g a n i c f r a c t i o n o f lake sediments would p e r s i s t o f time.

The observed decrease in m i c r o b i a l a c t i v i t y

materials

f o r even longer p e r i o d s

may e x p l a i n the increased c o n c e n t r a t i o n s

o f o r g a n i c d e b r i s commonly found in the sediments of a c i d i c lakes (6, 8). Lake a c i d i f i c a t i o n in p r e c i p i t a t i o n

is thought to occur p r i m a r i l y in the form o f s u l f u r l c

n i t r o g e n are the p r i n c i p a l

through the increased inputs o f hydrogen ions

and n i t r i c

acids (10).

The oxides o f s u l f u r and

sources o f these strong acids in p r e c i p i t a t i o n

(10, 19, 22).

However, these gases may a l s o reach a q u a t i c systems d i r e c t l y .

Depending on the pH, s u l f u r

d i o x i d e reacts w i t h w a t e r to form several s o l u b i l i t y

including, bisulfite

sulfite

(S03 =) (20).

Bisulfite

exists

the predominant form above pH 7 (20). bisulfite all

More s i g n i f i c a n t l y ,

reduced 14C-glucose m i n e r a l i z a t i o n more at pH l e v e l s o f 4 and 5 than a t pH 7.

As the b i s u l f i t e

progressively higher levels of sulfite bisulfite

Bisulfite

is more

on the sediment

c o n c e n t r a t i o n increased, the r a t i o o f 14C-glucose m i n e r a l i -

z a t i o n at pH 4 and 5 compared to t h a t a t pH 7 decreased. were r e q u i r e d to i n h i b i t

bisulfite

(21), thus the decreased 14C-glucose m i n e r a l i z a t i o n at the

low pH values o f 4 and 5 was p r o b a b l y due to the t o x i c e f f e c t s o f b i s u l f i t e microorganisms.

is

glucose m i n e r a l i z a t i o n was markedly reduced at

c o n c e n t r a t i o n was increased.

t o x i c to microorganisms than s u l f i t e

(HS03-) and

In the s t u d i e s conducted to determine the e f f e c t s o f

on the sediment m i c r o b i a l a c t i v i t y ,

pH l e v e l s as the b i s u l f i t e

products,

in the pH range o f a p p r o x i m a t e l y 2 to 7 w h i l e s u l f i t e

This was presumably due to the

accumulating at pH 7.

Greater c o n c e n t r a t i o n s o f s u l f i t e

m i n e r a l i z a t i o n at pH 7 compared t o the r e l a t i v e l y

low l e v e l s o f

r e q u i r e d at pH values o f 4 and 5.

A c i d i c lakes o f t e n contain high c o n c e n t r a t i o n s o f heavy metals (22, 23) and the increased solubility

o f heavy metals in a c i d i c c o n d i t i o n s could r e s u l t in increased t o x i c e f f e c t s on

microbial activity. alone.

The growth o f a mixed b a c t e r i a l p o p u l a t i o n was markedly a f f e c t e d by pH

However, when the combined e f f e c t s o f pH and heavy metals were examined, lead and

mercury a t pH 4.5 were more t o x i c to b a c t e r i a l growth than e i t h e r the low pH o r the heavy metals alone.

In a c i d i c c o n d i t i o n s , heavy metals may e x i s t

their solubility.

The g r e a t e r t o x i c i t y

to the increased a v a i l a b i l i t y

in free c a t i o n i c form, thus i n c r e a s i n g

o f both lead and mercury a t low pH was probably due

of the heavy metals to the sediment microorganisms.

was a l s o found t o be more t o x i c to the microorganisms than lead a t a l l by oxygen u t i l i z a t i o n

and 14C-glucose m i n e r a l i z a t i o n .

n o n - t o x i c compared to i t s e f f e c t s

In these s t u d i e s ,

lead was r e l a t i v e l y

in the growth e x p e r i m e n t s , p o s s i b l y due to the g r e a t e r

b i n d i n g and a d s o r p t i o n c a p a c i t y o f the sediment. microorganisms at low pH.

Mercury

pH l e v e l s as measured

Mercury was again more t o x i c to the sediment

982

The present study has shown that microbial growth and a c t i v i t y may be s e r i o u s l y altered in a c i d i c lake sediments by the d i r e c t e f f e c t s of low pH, and also by the increased t o x i c i t y of heavy metals and b i s u l f i t e

in a c i d i c conditions.

b l o l o g i c a l e f f e c t s of a c i d i f i c a t i o n ,

Consequently, when studying the apparent

i t is important to consider not only the d i r e c t but also

the i n d i r e c t e f f e c t s of pH.

ACKNOWLEDGEHENTS

This work was supported by Natural Sciences and Engineering Research Council operating grants to W.E.I. and C.l.M. and a scholarship to M.D.B.

The f i n a n c i a l assistance of the Department

of Fisheries and Oceans is also g r a t e f u l l y acknowledged.

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