Stimulation and Enrichment of Two Microbial Polychlorinated Biphenyl Reductive Dechlorination...

8/22/2019 Stimulation and Enrichment of Two Microbial Polychlorinated Biphenyl Reductive Dechlorination Activities

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P e r g a m o nPII: S0045-6535(96)00360-8

Chemosphere, Vol . 34 , No . 3 , pp . 655- 66 9 , 1997Copy r igh t 1997 E lsev ier Sc i ence L tdPrinted in Great Br i tain. Al l r ights reserved0045-6535197 $17.00+0.00

S T I M U L A T I O N A N D E N R I C H M E N T O F T W O M I C R O B I A L P O L Y C H L O R I N A T E DB I P H E N Y L R E D U C T I V E D E C H L O R I N A T I O N A C T I V I T I E S

W i ll ia m A . W i ll ia m s

E n v i r o n m e n t a l L a b o r a t o r y , G E R e s e a r c h a n d D e v e l o p m e n t C e n t e r ,P . O . B o x 8 , S c h e n e c t a d y , N e w Y o r k 1 2 3 0 1 - 0 0 0 8( Rece ived in USA 22 June 1996; accep ted 16 Sep tember 1996)

A B S T R A C TI s o l a t i o n a n d c h a r a c t e r iz a t i o n o f t h e b a c t e r i u m o r b a c t e ri a l c o n s o r t i u m r e s p o n s i b l e f o r m i c r o b i a lp o ly c h lo r ina t e d b ip h e ny l ( P CB ) r e d u c t i v e d e c h lo r ina t a on wo u ld a l l ow a b e t t e r u nd e r s t a n d ing o f t h e m e c h a n i s mof t h i s a c t i v it y i n t h e e nv i r onm e n t . S t im u la t i on a nd e n r i c h m e n t o f a P CB d e c h lo r ina t l on a c t i v i ty i n a c u l t u re i sn e c e s s a r y f o r i so l a t i o n o f t h e m i c r o o r g a n i s m s t h a t c a u s e P C B d e c h l o r in a t i o n . T w o d i s t i n c t m i c r o b i a l PC Bd e c h l o r i n a t io n a c t i v it i es w e r e i n d e p e n d e n t l y s t im u l a t e d i n s l u r r ie s o f P C B - c o n t a m i n a t e d u p p e r H u d s o n R i v e rs e d im e n t b y a d d ing s p e c i f i c P CB c onge ne r s . Th e a d d i t i on o f 2 ,4 ,6 - t r i c h lo rob ip h e ny l s t im u la t e d a s p e c i f ic parad e c h lo r ina t i on a c t i v i t y on A r oc lo r 1 2 42 ( d e s igna t e d a P r oc e s s Q) . I n c on t r a s t , 2 ,3 ,6 - t r i c h lo r ob ip h e ny l a d d i t i ons t im u la t e d a s p e c i f i c meta d e c h lo r ina t i on a c t i v i t y ( d e s igna t e d a P r oc e s s M ) . E a c h d e c h lo r ina t i on a c t iv i t y wase n r i c h e d b y t r a ns f e r r i ng 1 0 p e r c e n t b y v o lu m e o f t h e s e d im e n t s l u r r y c on t a in ing t h e d e c h lo r ina t i on a c ti v i t y t oa n a u t o c l a v e d m e d i u m c o m p o s e d o f a n a n a e ro b i c m i n e r a l s o l u ti o n , h u m i c a c id s , a n d P C B s T h i s i s th e f ir str e p o rt o f a n e n r i c h m e n t c u l tu r e o f t h e m i c r o b i a l para d e c h lo r ina t i on a c t i v i t y ( t h e P r oc e s s Q) Bo th m ic r ob i a lP CB d e c h lo r ina t i on a c ti v i t i e s we r e i nh ib i t e d b y v a nc o m y c in ( 5 p p m ) o r n i s i n ( 5 0 0 p p m ) . Th e s e d a t a s u gge s tt h a t a p op u l a t i on o f g r a m - p o s i t i v e b a c t e r i a i s e s s e n t ia l f o r t h e s e P CB d e c h lo r ina t i on ac t i v it i es . 1 9 9 7 E l s e v i e r S c i e n c e L t d . A l l r ig h t s r e se r v e dK e y w o r d s : p o l y c h l o r i n a t e d b i p h e n y l s , P C B s , r e d u c t i v e d e c h l o r in a t i o n , a n a e r o b i c m i c r o b i o l o g y

I N T R O D U C T I O NP o l y c h l o r i n a t e d b ip h e n y l s ( P C B s ) a r e a g r o u p o f c o m p o u n d s t h a t w e r e b a n n e d f r o m p r o d u c t i o n in t h e

Un i t e d S ta t e s in 1 9 77 . Th e s e c om p ou n d s we r e m a nu f a c tu r e d as m ix tu r e s o f P CB c onge ne r s t h a t d i f fe r e d b y t h enu m b e r a nd c on f igu r a t i on o f c h lo r ine s on t h e b ip h e ny l nu c l e u s ( F igu r e 1 ); t h e m ix tu r e s we r e g iv e n t ra d e na m e ss u c h a s A r oc lo r 1 2 42 . W id e s p r e a d u s e o f P CB s r e s u l t e d i n t h e i r a c c u m u la t i on in th e o r ga n i c p o r t i ons o f so i ls ,

6 5 5


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6 5 6s e d im e n t s , a nd b io ta . A e r ob i c b iod e g r a d a t i on o f P CBs i s we l l c h a r a c t e ri z e d [ fo r a re v i e w , s e e re f e r e nc e 1 b u ti s g e n e r a l l y l i m i t e d t o t h e l i g h t ly c h l o r i n a t e d b i p h e n y l s . U n l i k e a e r o b ic P C B b i o d e g r a d a t i o n , a n a e ro b i cm ic r ob i a l r e d u c t i v e d e c h lo r ina t i on o f ne a r l y a l l t h e P CB h om ologs c a n oc c u r .

p a r a 4

m e tam e ta , ]e t a l, , m e t a 3 ~ Sr t h o ~ J o r t h o 2 6o r t h o ~ o r t h o 2 ' 6 '

3 ' 5 'p a r a 4F i g u r e I . T h e G r e e k a n d n u m e r i c a l n o m e n c l a t u r e sd e s c r i b ing t h e c on f igu r a t i on o f c h lo r ine s i n a P CB c onge ne r .

Br ow n a nd c o - wor ke r s [ 2 - 5] p r op o s e d t h a t th e d i s t i nc t i v e p a t t e m s o f P CB t r a ns f o r m a t ion s e e n m P CB-c on t a m ina t e d s e d im e n t s r e s u l t e d f r om s e v e r a l d i f f er e n t m ic r ob i a l d e c h lo r ina t i on a c t i v i ti e s t h at a r e d i s t ingu i s h e db y c on ge ne r s e l e c t iv i t y . Be d a r d a nd Qu e ns e n [ 6 ] p r ov id e d a d e t a i le d r e v i e w o f th e m ic r ob i a l P CB r e d u c t i v ed e c h lo r ina t i on a c t i v i ti e s ( a ls o t e r m e d , d e c h lo r ina t i on p r oc e s s e s ) a nd t h e d e c h lo r in a t i on p a t t e r ns , wh i c h h a v eb e e n g iv e n l e t t e r d e s igna t i ons . Th e s e i nv e s t i ga to r s no t e d t h a t c h lo r ine r e m ov a l ( i e . , a d e c h lo r ina t i on p roc es s)i s no t d e t e r m ine d s t r i c tl y b y t h e p os i t i on ( o r t h o , m e t a , o r p a r a ) of t h e c h lo r ine r e l a t iv e t o t h e op p o s i t e p h e ny lr i ng ; b u t a l s o b y t h e s u r r ou n d ing c h lo r ine c on f igu r a t i on , t h e c h lo r ine c on f igu r a t i on on t h e op p os i t e r i ng , a nd th eto t al nu m b e r o f c h lo r ine s on b o th r i ngs [ 4 , 6] .

M ic r ob i a l P CB d e c h lo r ina t i on p a t t e m s M a nd Q h a v e b e e n ob s e r v e d i n s e d im e n t s l u r r ie s o r m ic r oc os m sc o n t a i n i n g m i c r o o r g a n i s m s fr o m P C B - c o n t a m i n a t e d u p p e r H u d s o n R i v e r s e d i m e n t s [ 7 -1 4 ] . P a t te r n Md e c h lo r ina t i on a c t i v i t y i s c h a r a c t e r i z e d a s m e t a d e c h lo r ina t i on o f 2 ,3 ,4 - ; 2 ,3 ,6 - ; 2 ,3 - ; 2 ,5 - ; a nd 3 ,4 - c h lo rop h e ny lg r ou p s ; a nd p os s ib ly m e t a d e c h lo r ina t i on o f 3 - c h lo r op h e ny l g r ou p s [ 6 , 1 5 ] P a t t er n Q d e c h lo r ina t i on a c t i v it y i sc h a r a c t e r i z e d a s p a r a d e c h l o r i n a t i o n o f 2 , 4 , 5 - ; 2 , 4 -; 3 , 4 - ; a n d 4 - c h l o r o p h e n y l g r o u p s ; a n d p o s s i b l y p a r ad e c h lo r ina t i on o f 2 ,3 ,4 - c h lo r op h e n y l g r ou p s [ 6 , 1 5 ] . P a t t e rn Q d e c h lo r ina t i on a c t i v i ty i s a ls o c h a r a c t e r iz e d a sm e t a d e c h l o r i n a t i o n o f 2 , 3 - c h l o r o p h e n y l g r o u p s ; a n d p o s s i b l y m e t a d e c h l o r i n a t i o n o f 2 , 3 , 4 - a n d 2 , 3 , 6 -ch loro phen yl grou ps [6 , 15] .

A b e t t e r u n d e r s t a n d i n g o f m i c r o b i a l P C B d e c h l o r i n a t i o n c o u l d b e o b t a i n e d b y i s o l a t in g a n dc h a r a c t e r i z i n g t h e b a c t e r i u m o r b a c t e r i a l c o n s o r t i u m r e s p o n s i b l e f or t h e s e a c ti v i ti e s . E n r i c h m e n t o f ad e c h lo r ina t i on a c t i v i t y i s one s t ep i n i s o l a t ing m ic r oo r ga n i s m s th a t c a u s e P CB d e c h lo r ina t i on Th e r e h a v e b e e ns e v e r a l re p o r t s o f m ic r ob i a l p a t t e rn M d e c h lo r ina u o n a c t iv i t y e n r i c h m e n t c u l tu r e s [ 8 , 1 0 , 1 4 , 1 6 ], b u t no r e p o rto f a m ic r ob i a l p a t t e m Q d e c h lo r ina t i on a c ti v i t y e n r i c h m e n t c u lt u re .

S e l e c ti v e s t i m u l a t i o n o f a p a r t ic u l a r d e c h l o r i n a ti o n a c t iv i ty i s o n e m e t h o d o f o b t a i n i n g a n e n r i c h m e n tc u l t u re . S om e h a loge na t e d a r om a t i c c om p ou nd s , s u c h a s h a loge na t e d b ip h e ny l s o r b e nz o i c a c id s , h a v e b e e n


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6 5 7u se d t o s ti mu l a t e a nd e n r i c h sp e c i f i c m i c r ob i a l P C B d e c h l o r i na t i on a c t i v i t ie s [ 17 , 1 8 ]; b u t p a t te r n s M a nd Qa c t i v i t i e s h a v e no t b e e n s t i mu l a t e d a nd e n r i c h e d i n t h i s ma nne r . I n a p r e v i o u s s t u d y [ 1 9 ], I no t e d meta andpara, b u t n o t ortho d e c h l onna t i on o f 2 , 3 , 6 - tr i c h l o r ob i p h e ny l ( 2 , 3 , 6 - C B ) a nd 2 , 4 , 6 - t r i c h l o r ob i p h e ny l ( 2 , 4 , 6 - C B )b y mi c r oo r ga n i sms i n u p p e r H u d so n R i v e r s e d i me n t sl u r ri e s . T h e se r e su l ts [ 1 7 - 1 9 ] l e d t o t h e h y p o t h e s i s a ndsu b se qu e n t i nv e s t i ga t i on ( d e sc r i b e d h e r e i n ) t h a t 2 , 3 , 6 - C B a nd 2 , 4 , 6 - C B c ou l d b e u se d t o s t i mu l a t e a nd t h e ne n r i c h p a t t er n s M a nd Q d e c h l o r i na t i on a c t i v it i e s i n a P C B - c on t a m i na t e d u p p e r H u d son R i v e r s e d i m e n t

M A T E R I A L S A N D M E T H O D SS e d i m e n t C o l l e c t i o n S i t e . H u d son R i v e r s e d i me n t w a s c o l l e c t e d i n t h e r i v e r ne a r t h e e a s t b a nk a t S t i l l w a t e r ,N Y ( r i v e r m i l e 1 6 9) a nd s t o r e d a t 4 C . T h e s e d i me n t c on t a i ne d - 2 5 [ t g /g (d r y w t o f s e d i me n t ) meta- a nd para-d e c h l o r in a t e d A r o c l o r 1 24 2; - 6 0 % o f t h e P C B s w e r e 2 - c h l o r o b i p h e n y l ( 2 - C B ) ; 2 , 2 ' - C B ; 2 , 6 - C B ; a n d 2 , 6 , 2 ' -C B .S e d i m e n t S l u r r y P r e p a r a t io n . T h e s l u r r y s a mp l e s w e r e p r e p a r e d i n a n a na e r ob i c c h a mb e r ( C oy L a b or a t o r yP r od u c t s , A nn A r b o r , M I ) c o n t a i n i ng a n a t mosp h e r e o f _>9 8% N 2 a nd 14 h o u r s w i t h 6 m L o f d i e th y l e t h e r a n d - 0 . 3 m L m e r c u r yM e r c u r y w a s a d d e d t o p r e c i p it a t e t h e e le m e n t a l s u l f u r i n t h e s e d i m e n t A 9 0 % e x t r ac t io n e f f i c ie n c y o f th e


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6 5 8P C B s i n a s e d i m e n t s l u r ry w a s ob t a i ne d u s i ng t h is e x t r a c ti on p r oc e d u r e [ 1 3 , 1 9 ] . T h e e n r i c h me n t c u l t u r e s w e r esa m p l e d a nd t h e P C B s e x t r a c t e d i n t h e s a me f a sh i on a s t h e s e d i me n t s l u r r ie s a t 4, 9 , 1 4 , 1 8 , 2 6 , a nd 3 6 d a y s o fr oom t e mp e r a t u r e i nc u b a t i on .

T h e e t h e r e x t r a c t s w e r e a n a l y z e d i n a V a r i a n 3 5 0 0 g a s c h r o m a t o g r a p h ( G C ) u s i n g a f u s e d s i l i c ac a p i l l a r y c o l u mn ( 3 0 m b y 0 . 2 5 mm i nne r d i a me t e r ) c oa t e d w i t h D B - 1 ( p o l y d i me t h y l s i l oxa ne , J&W S c i e n t i f i c ,F o l som , C A ) a nd an e l e c tr on c a p t u r e d e te c t o r ( E C D ) a t 3 0 0 C . T h e c onc e n t r a t i ons o f s e le c t e d P C B c onge ne r si n a s a m p l e ( l i s t e d in T a b l e 1 ) w e r e d e t e r m i n e d f r o m a l i n e a r, t h r e e - p o i n t c a l i b r a t io n c u r v e o f k n o w nc o n c e n t r a ti o n s o f A r o c l o r 1 2 4 2 u s in g G C - E C D w i t h t h e D B - 1 c o l u m n f o l l o w i n g t h e p r o c e d u r e s p r e v i o u s l yd e sc r i b e d [ 2 1] . T h e p e a k nu m b e r i ng i n T a b l e 1 a nd F i gu r e s 2 a nd 3 is t h e 1 1 8 p ea k P C B p r o f i l e f r om G C - E C Du s i ng a D B - 1 c o l u m n d e sc r i b e d b y F r a me e t al . [2 1 ] . T h e c onc e n t r a t ions o f 2 , 3 , 6 - C B ; 2 , 4 , 6 - C B ; a nd 2 , 6 - C B i nt h e s a mp l e s w e r e a l so d e t e r mi ne d f r om a l i nea r , t h r e e - p o i n t c a li b r a t ion c u r v e o f know n c onc e n t ra t a ons o f th e sec onge ne r s . T h e P C B c o nge ne r s 2 , 3 , 6 - C B ; 2 , 4 , 6 - C B ; a nd 2 ,6 - C B ( e a c h a t >9 9 % p u r i ty ) ; a nd A r oc l o r 1 24 2 w e r ep u r c h a se d f r om A c c u s t a nd a r d , I nc. ( N e w H a v e n , C T ) .

D B - 1 p e a k n u m b e r 5 c o n t a in i n g 2 , 2 ' - C B a n d 2 , 6 - C B w a s n o t i n c l u d e d in t h e d a ta o f F i g u r e s 2 an d 3 ,b e c a u se b o t h t h e a d d e d t r i c h l o r ob i p h e ny l s ( a d d e d i n l a r ge e xc e s s c omp a r e d t o t h e o t h e r P C B c onge ne r s ) w e r ed e c h l o r i na t e d to 2 , 6 - C B . I nc l u d i ng D B - 1 p e a k nu m b e r 5 i n t h e d a ta w o u l d s i gn i f i c a n t l y ob sc u r e t h e mo l ep e r c e n t P C B c o n g e n e r c a lc u l a t io n s . U n f o r t u n a t e l y , 2 , 2 ' - C B a n d 2 ,6 - C B w e r e t h e m a j o r d e c h l o r in a t i o np r od u c t s o f A r o c l o r 1 2 42 . E xc l u s i on o f t h e se c onge ne r s f r om t h e d a t a c a lc u l a t i ons r e su l te d i n ske w e d mo l ep e r c e n t d i f f e r e nc e h is t og r a ms . F o r P C B c onge ne r s th a t w e r e d e c h l o r i na t e d , th e a c tu a l a mou n t s w e r e mor e th a nt h e c a l c u l a te d v a l u e s d i sp l a y e d i n F i gu r e s 2 a nd 3. F o r P C B c onge ne r s t h a t w e r e d e c h l o r i na t i on p r od u c t s , th ea c t u al a m ou n t s w e r e l e s s t h a n t h e c a l c u l a te d v a l u e s d i sp l a y e d i n F i gu r e s 2 a nd 3 . A ny c a l c u l a t i ons o f r a te a nde x t e n t o f P C B d e c h l o r i n a t i o n b a s e d o n t h is d a t a w o u l d u n d e r e s t im a t e t h e tr u e v a lu e s . N e v e r t h e l e s s , t h emi c r ob i a l P C B d e c h l o r i na t i on p a t t e r n s a r e no t ob sc u r e d b y t h e d a t a c a l c u l a t i ons a nd t h e y c a n b e i d e n t i f i e d i nt h e h i s t og r a ms i n F i gu r e s 2 a nd 3 .E x t r a ct io n o f H u m i c A c i d s . A h u m i c a c id e x t ra c t w a s p r e p a r e d u s i ng u p p e r H u d son R i v e r s e d i me n t th a t d i dn o t c o n t ai n P C B s ( P C B - f r e e s e d i m e n t) . T h e e x t r a c ti o n p r o ce d u r e w a s m o d i f i e d f r o m t h e m e t h o d o f S te v e n s o n[ 22 ]. T h e s e d i m e n t w a s c o l l e c t e d u p s t r e a m f r om t h e P C B - c o n t a m i na t e d s e d i m e n t a t r i v e r m i l e 2 0 5. T h e w e ts e d i m e n t w a s e x t r ac t e d w i t h a n e q u a l v o l u m e o f 0 .5 M N a O H b y m e c h a n i c a l s t ir r in g f o r 1 4 h o u r s a t - 2 5 CT h e l i qu i d e x t r a c t w a s s e p a r a t e d f r om t h e so l i d s (s a nd a nd c l a y f r a c t ions ) b y g r a v i t y a nd c e n t r i f u g a t i on T h ep H o f t h e e x t r a c t ( - 1 1 . 5 ) w a s a d j u s t e d to - 6 b y t h e d r op - w i se a d d i t ion o f c onc e n t r a te d H C I w h i l e ma gne t i c a ll ys t i r ri ng th e so l u t ion . T h e p H a d j u s t me n t r e su l te d i n p r e c i p i t a ti on o f t h e h u m i c a c i d s , w h i c h w e r e c o l l e c t e d b yc e n t r i fu ga t i on . T h e h u m i c a c i d e x t r a c t w a s w a sh e d f ou r ti me s w i t h e i gh t t o t e n v o l u me s o f d i s ti l le d w a t e r ands t o r e d a s a ge l ( 9 % so l i d s b y w t ) a t 4 C T h e h u m i c a c i d e x t r a c t w a s 6 % b y w e i gh t o f t h e s e d i m e n tP r e p a r a t io n o f E n r i c h m e n t C u l t u r e s . E nr i c h m e n t c u l t u r es o f th e t w o m i c r ob i a l P C B d e c h l o r i na t i on a c t i v it i e sw e r e p r e p a r e d i n a me d i u m c on t a i n i ng t h e h u mi c a c i d e x t r a c t o f P C B - f r e e s e d ime n t . T h e h u m i c a c i d e x t r a ct


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6 5 9s t oc k w a s d il u t e d t o 3 % so l i d s b y w e i gh t w i t h r e d u c e d R A M M . A r o c l o r 1 2 4 2, a nd 2 , 3 ,6 - C B o r 2 ,4 , 6 - C B w e r ea d d e d t o ma ke f i na l c onc e n t r a t i ons o f 45 p p m ( v o l : v o l ) a nd 3 5 0 B M , re sp e c t i v e l y . E a c h s a m p l e w a s a u t oc l a v e df o r 3 0 m i n u t e s a n d a l l o w e d t o c o o l t o r o o m t e m p e r a t u r e , b e f o r e a d d i t io n o f a n u n h e a t ed , m e t h a n o g e n i cs e d i m e n t s l u r r y ( 1 0 % b y v o l ) i n w h i c h o n e o r t h e o t h e r m i c r o b i a l P C B d e c h l o r i n a t io n a c t i v i ty h a d o c c u r r ed .T w o b a c t e r i a l a n t i b i o t i c s , v a nc omy c i n a nd n i s i n , w e r e a d d e d a s s t oc k so l u t i ons ( 5 0 0 0 p p m e a c h i n r e d u c e dR A M M ) t o s e p a r a t e e n r i c h m e n t c u l t u r e s ; th e f i na l c onc e n t r a t ions o f e i t h e r a n t i b i o ti c w e r e 5 , 5 0 , o r 5 0 0 p p mF or e a c h o f th e t w o mi c r ob i a l P C B d e c h l o r i na t i on a c t i v i t ie s , a s e t o f d u p l i c a t e e n r i c h me n t c u l t u r e s w a s t r e a t edw i t h v a nc om y c i n o r n i s i n a t one o f t h r e e c onc e n t r a t i ons , o r u n t r e a te d ; t h e r e w e r e 1 4 e n r i c h m e n t c u l t u r e s fo re a c h a ct i v it y . B o t h a n t i b i o t ic s w e r e p u r c h a s e d f r o m S i g m a C h e m i c a l C o m p a n y ( S t . L o u i s , M O ) ; t h ev a n c o m y c i n w a s p u r c h a s e d a s t h e h y d r o c h l o r i d e s a l t a n d t h e n i s i n w a s p u r c h a s e d a s a 2 . 5 % s o l u t i o n i nd e na t u r e d mi l k so l id s . T h e 5 0 0 0 p p m s t oc k so l u ti on o f n i s in i n r e d u c e d R A M M w a s p r e p a r e d d i r e c t ly w i t h t h ep u r c h a se d ma t e r i a l ; th u s e n r i c h m e n t c u l t u r e s t re a t e d w i t h n i s in c on t a i ne d d e na t u r e d m i l k so li d s

R E S U L T SP C B D e c h l o r i n a t io n A c t i v it y R e s u l ti n g f r o m T r i c h l o r o b ip h e n y l T r e a t m e n t . T h e m i c r o b i a l P C Bd e c h l o r i n a t io n a c t i v i ti e s c a u s e d s i g n i f i c a n t c h a n g e s i n th e m o l e p e r c e n t o f s p e c i f i c P C B c o n g e n e r s i n t h es e d i m e n t s l u r r ie s a n d t h e e n r i c h m e n t c u l tu r e s. T h e h i s t o g r a m s i n F i g u r e s 2 a n d 3 s h o w t h e m o l e p e r c e n td i f f e r e nc e s o f P C B s i n t h e s e d i m e n t s l u r r i e s (i .e . , t h e m o l e p e r c e n t o f P C B c onge ne r s a t s e l e c t e d t ime p o i n t smi nu s t h e mo l e p e r c e n t o f P C B c onge n e r s a t t i me z e ro ) . D e c h l o r i na t i on p a t te r n s M a nd Q w e r e i d e n t i f i e d b yd e c r e a se s i n sp e c i f i c c onge ne r s ( t h e d e c h l o r i na t i on su b s t r a t es ) a nd i nc r e a se s i n t h e d e c h l o r i na t i on p r od u c t s ( 6 ,1 5 )

B o t h p a t t e r n s M a nd Q d e c h l o r i na t i on a c t i v i t i e s oc c u r r e d i n s l u r r i e s a me nd e d w i t h b o t h 2 , 3 , 6 - C B a nd2 , 4 , 6 - C B ( t op h i s t og r a m i n F i gu r e 2 ). P a t t e rn M a c t i v i ty oc c u r r e d f i r s t i n t h e se s l u r ri e s ; i t w a s e v i d e nc e d b ym e t a d e c h l o r i n a t i o n o f P C B s c o n t a i n i n g 2 , 5 - d i c h l o r o p h e n y l , s o m e 2 , 3 - d i c h l o r o p h e n y l , a n d 2 ,3 , 6-t r i c h lo r op h e ny l g r ou p s . B y t h e ne x t s a mp l i ng t ime , p a t t e m Q a c t i v i ty h a d oc c u r r e d w i t h p a r a d e c h l o r t na t ion o f2 , 4 - d i c h l o r o p h e n y l a n d 3 , 4 - d i c h l o r o p h e n y l g r o u p s , a n d p a r a d e c h l o r i n a t i o n o f s o m e 2 , 4 , 5 - a n d 2 , 4 , 6 -t r i ch l o r o p h e n y l g r o u p s . A t t h e la s t s a m p l i n g t im e , o n l y 1 0 % o f P C B s c o n t a i n i n g m e t a o r p a r a c h l o r i ne sr e ma i ne d . I n a s e p a r a t e e xp e r i me n t , p a t t er n M a c t i v it y b u t no t p at te r n Q a c t i v i ty w a s ob se r v e d i n s l u r r ie s o f th esa me se d i me n t to w h i c h A r oc l o r 1 2 4 2 b u t ne i t h e r 2 , 3 , 6 - C B no r 2 , 4 , 6 - C B w a s a d d e d ( u np u b l i sh e d d a t a )

I n s lu r r i e s a me nd e d w i t h on l y 2 , 4 , 6 - C B , on l y p at te r n Q d e c h l o r i na t ion a c t i v it y oc c u r r e d A n e xa m p l e o ft h e c h a nge s i n t h e mo l e p e r c e n t o f sp e c i f i c P C B c onge ne r s i nd i c a t i v e o f a p at t er n Q d e c h l o r i na t i on i s sh ow n i nt h e t op h i s tog r a m i n F i gu r e 3 . T h i s s e e m e d t o b e t h e on l y P C B d e c h l o r i na t i on a c t i v i t y t h a t oc c u r r e d i n t h e ses l u r ri e s , a s e v i d e nc e d b y t h e l o s s o f t h e p a r a c h l o r i ne f r om 2 , 4 ' - C B a nd s e v e r a l P C B s c on t a i n i ng 2 , 4 - a nd 3 , 4 -d i c h l o r op h e ny l , a nd 2 , 4 , 5 - t r ic h l o r op h e ny l g r ou p s ; a s w e l l a s l o ss o f t h e m e t a c h l o r i ne f r om P C B s c on t a i n i ng


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6 60

2 i l ( l l I l l I E 1 1 1 1 1 r l [ I t I i l F I I i l [ I l t l ! l l-5

4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 5S 58 61 64 67 70 73D B -1 P e a k N u m b e r

98

= 7Q)oco sm

~ - 3Q)- 6 2~ZI

4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 SS 5B 61 64 67 70 73D B - I P e ak N u m b e r

F i g u re 2 . M i c rob i a l r e d u c t i v e d e ch l o r i na ti on o f Aroc l o r 1 2 4 2 i n u nh e a t ed , m e t h a noge n i c H u d s on R i v e rsed imen t s lu rr ie s amen ded w~th 2 ,3 ,6-CB and 2 ,4 ,6-CB. The top h i s togram i s the average of dupl ica te samplesof the m ole pe rce nt d i f fe rences of PCB congeners be tween the se lec ted t ime point s and t ime ze ro The bottomhis togram i s a typ ica l mo le pe rcent PCB congener d i s t r ibu t ion in the sed im ent s lu rr ie s a t time zero A l i s t ing o fthe congeners in each peak i s shown in Table 1

2 ,3-d ich loroph enyl , and 2 ,3 ,4- and 2 ,3 ,6- t r i ch loropheny l groups . By the l a s t sampl ing, near ly a l l the para -chlor ine -co nta in ing congeners were dechlo r ina ted

Both pa t te rns M and Q dechlor ina t ion ac t iv i t i e s a l so occurred in s lurr ie s supplemen ted wi th only 2 ,3 ,6-CB (bot tom h i s togram in F ig ure 3) . The pa t te rn M d echlor ina t ion in these s lurr ie s was s im i la r to the pa tte rn Mdechlor ina t ion in s lurr ie s amend ed wi th both t r i ch lorobiphen yls ( top h i s togram in F igure 2). This pa t tern was


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661

2,4,6-CB Addit ions , ] i I i l l l l l I I I I ] i l I T T I I I I I I i l3 1 1 1 I i l l i l I I I ] l i l l l I 1 1 I I2 J l l l l l l l l l 1 l l i l l I l l l l l l l l l l ] l [ [ I I I [ I I, I I I L I I I I I l I I I I I I I i l I I I I I l l l I i 1 [ [ l i i l [ [ l l [ I i i l i [ I0 n

- 1 -2~ 3 I I I I l i i l l i l l l l [ , l l [ l lo ! I ~ l i l l l l i l I I I I I t l l i t l - t !' , ' " ; " ; ' ; 0 ;] ;~ ,9 2;'2~ 29 ;; , , ~, ~4 ,, ,6':,~ s2 'd s~ 6, [ 6, ~o 7~

DB-1 Peak NumberI F A2S DA Y S 1 4 1 D A Y S 9 2 D A Y S i

8c-O

~3o~ID-5

2,3,6-CB Ad dit ion

l l l N l l l l l l l l l l l l l l ' l l " i i~ J N I I I I I l l l l i l l l l l l l i l l l l i l l [ [ ! io I | I I I I L I I I I I I I I I I I ~ I I I t [ I I I . U I I [ L J I I [ I I L L L I [ t , I l l l i i l l l l [ I

s I I I I I l l l l l l l l l l l l l l l l l l l r l l l l t l l l l l l l l l l l l l l l l l l l i I I I I l l l l l l I ] l [1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 ss 58 61 64 67 70 73DB-1 P eak Number

m 2 S D AY S 4 1 D A YS 1 9 2 D AY S1F i g u r e 3 . M i c r o b i a l r e d u c t i v e d e c h l o r i n a t io n o f A r o c l o r 1 2 4 2 in u n h e a t e d , m e t h a n o g e n i c H u d s o n R i v e rs e d i m e n t s l u rr i es a m e n d e d w i th 2 , 3 , 6 -C B o r 2 , 4 , 6 - C B E a c h h i s t o g ra m i s t h e a v e r ag e o f d u p li c a te s a m p l e s o ft h e mo l e p e r c e n t d i f f e r e nc e s o f P C B c onge ne r s b e t w e e n t h e s e le c t e d ti me p o i n t s a nd t ime z e r o A l i s ti ng o f t h ec onge ne r s i n e a c h p e a k i s sh ow n i n T a b l e 1

f i r s t ob se r v e d a s meta d e c h l o r ina t a on o f P C B s c on t a i n i ng 2 , 5 - a nd 2 , 3 - d i c h l o r op h e ny l , a nd 2 , 3 , 5 - a nd 2 , 3 , 6 -t r i c h l o r o p h e n y l g r o u p s . A d d i t i o n a l m e t a d e c h l o r i n a t i o n o f 2 , 3 - an d 3 , 4 - d i c b l o r o p h e n y l , a n d 2 , 3 , 4 -t nc h l o r op h e ny l g r ou p s oc c u r r e d b y t h e ne x t s a mp l i ng t i me . P a t t er n Q a c t i v i t y oc c u r r e d a f t e r p a t te r n M a c t iv i t y ;i t w a s i d e n t i f i e d b y t h e p a r a d e c h l o r i na t i on o f s e v e r a l P C B c on ge ne r s , i nc l u d i ng 2 , 4 , 2 ' - C B ( p e a k 1 5 ) . B y t h el a s t s a mp l i ng p e r i od , meta and para d e c h l o r i na t ion o f ne a r l y a l l t h e P C B c onge ne r s oc c u r r e d .


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6 6 2T A B L E 1. A l i s t in g o f th e p r i m a r y P C B c o n g e n e r ( s ) q u a n t i f i e d f o r e a c h

~ e a k ~ s h ow n i n F i g u r e s 2 a n d 3D B - 1 c o n g e n e r ( s ) i n p e a k D B - 1 c o n g e n e r ( s ) i n p e a kp e a k n u m b e ra p e a k n u m b e ra

2 2 -6 2,4- ; ' :2 ,5-7 2 , 3 ' -8 2 , 4 ' - ; 2 , 3 -

1 0 2 , 6 , 2 ' -1 4 2 , 5 , 2 ' - ; 4 , 4 ' -1 5 2 , 4 , 2 ' -16 b 2 , 6 , 3 ' - ; 2 , 3 , 6 -1 7 2 , 3 , 2 ' - ; 2 , 6 , 4 ' -21 2 , 5 , 3" -22 2 , 4 , 3 ' -2 3 2 , 5 , 4 ' -2 4 2 , 4 , 4 ' -2 5 2 , 3 ' , 4 ' ; 2 , 3 , 3 ' - ; 2 , 5 , 2 ' , 6 ' -2 6 2 , 3 , 4 ' -; 2 , 4 , 2 ' , 6 ' -2 7 2 , 3 , 6 , 2 ' -2 9 2 , 3 , 2 ' , 6 ' -3 1 2 , 5 , 2 ' , 5 ' -3 2 2 , 4 , 2 ' , 5 ' -3 3 2 , 4 , 2 ' , 4 ' -

3 4 2 , 4 , 5 , 2 ' - ; 2 , 4 , 6 , 4 ' -3 7 2 , 3 , 2 ' , 5 ' -3 8 2 , 3 , 2 ' , 4 ' - ; 3 , 4 , 4 ' - ; 2 , 3 , 6 , 3 ' -3 9 2 , 3 , 4 , 2 ' - ; 2 , 3 . 6 , 4 ' -4 2 2 , 3 , 2 ' , 3 ' -4 6 2 , 4 , 5 , 4 ' -4 7 2 , 5 , 3 ' , 4 ' -4 8 2 , 4 , 3 ' , 4 ' - ; 2 , 3 , 6 , 2 ' , 5 ' -4 9 2 , 3 , 6 , 2 ' , 4 ' -5 0 2 , 3 ,3 ' , 4 ' - ; 2 , 3 , 4 , 4 ' -5 1 2 , 3 , 6 , 2 ' ,3 ' - ; 2 , 3 , 5 , 2 ' , 5 ' -5 3 2 , 3 , 5 , 2 ' , 4 ' - ; 2 , 4 , 5 , 2 ' , 5 ' -5 4 2 , 4 , 5 , 2 ' , 4 ' -5 7 2 , 4 , 5 , 2 ' , 3 ' -5 8 2 , 3 , 4 , 2 " , 5 ' - ; 2 , 3 , 4 , 6 , 4 ' -5 9 2 , 3 , 4 , 2 ' , 4 ' -6 1 2 , 3 , 6 , 3 ' , 4 ' -63 2 , 3 , 4 , 2 ' , 3 ' -6 9 2 , 4 , 5 , 3 ' , 4 '- ; 2 , 3 , 6 , 2 ' , 4 ' , 5 ' -74 2 , 3 , 4 , 3 ' , 4" - ; 2 , 3 , 4 , 2 ' , 3 ' , 6" -

a This list includes only the peaks shown in Figure s and 3. The peak numbers were assigned according to thesequence of 118 PCB peaks detected by DB -1 GC-ECD" the com plete table was d escribed by Frame el a1.[21].b B ecause 2,6,3"-CB co-migrates with 2.3,6-CB, peak number 16 was not included in the d ata of slurries towhich 2,3,6-CB was added (Figure 2 and the bottom chart of Figure 3).A c c l i m a t i o n T i m e s f o r P C B D e c h l o r in a t io n . T h e a c c l i m a t i o n t i m e f o r a p a r t i c u l a r m i c r o b i a l P C Bd e c h l o r i n a t i o n a c t i v i ty i s a n i m p o r t a n t i n d i c a t o r o f s t i m u l a t i o n a n d e n r i c h m e n t o f t h e a c t i v it y . A n a c c l i m a t i o nt i m e w a s r e c o r d e d a s t h e f i r st s a m p l i n g t i m e w h e n t h e m o l e p e r c e n t o f a s p e c i f i c P C B c o n g e n e r h a d d e c r e a s e d( T a b l e 2 ). S i n c e P C B s a m p l i n g w a s d o n e a t s p e c i f i c t i m e p o i n t s , t h e t i m e a t w h i c h d e c h l o r i n a t i o n o c c u r re db e t w e e n t w o t e s t i n g in t e r v a l s c o u l d n o t b e d e t e r m i n e d . T h e a c c l i m a t i o n t i m e f o r p a t t e m M d e c h l o r i n a t i o na c t i v it y w a s t h e s a m p l i n g t i m e a t w h i c h meta d e c h l o r i n a t i o n o f 2 , 5 , 2 ' - C B ( p e a k 1 4 ) w a s f i r s t o b s e r v e d [ 6 , 1 5 ];a t t h a t t i m e > 6 0 % o f t h e c o n g e n e r w a s d e c h l o r i n a t e d T h e s a m p l i n g ti m e a t w h i c h para d e c h l o r i n a t i o n o f2 , 4 , 2 ' - C B ( p e a k 1 5 ) w a s f i r s t o b s e r v e d w a s t h e a c c l i m a t i o n t i m e f o r p a t t e r n Q d e c h l o r i n a t i o n a c t i v i t y [ 6 , 1 5 ] ; a t


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663that time _>45% of the cong ener was dec hlor inated Both 2,3,6-CB and 2,4,6-CB were _>80% dechlorinated to2,6-CB at their acclimation times.

TABLE 2. Observed acclimation times for PCB dechlorinat ion inthe Hudson River sediment slurries

PCB dechlorinationapattem M c pattem Qc

congener pasteurizationb and andaddition 2,3,6-CB-->2,6-CB 2,4,6-CB---)2,6-CBC I Yes 14 days 41, 69 daysd

Cl Cl No 25 days 69 days d2,3,6-CBCle l ~ Yes not observed 25 days

Cl No not observed 41 days2,4,6-CB2,3,6-CB & Yes 14 days 25 days

2,4,6-CB No 25 days 36 daysa No PCB dechlorinationwas observed n the autoclaved sedimentslurries.b Pasteurization nhibitedmethanogenesis,which was observed n the unheated slurries.c Patterns describedby Brown [15], and Bedard and Quensen [6].d Pattem Q was observed n one sample at 41 days and in the other samplesat 69 days. N~2,4,6-CB was added o these slurries.

Some of the acclimation times for the dechlorination activities were the same within a sediment slurry,and betwee n the different ly treated slurries (Table 2). Meta dechlorination of 2,3,6-CB and meta dechlorinationactivity on Aroclor 1242 (pattern M) had the same acclimation time within a sediment slurry, irrespective ofwhether or not the slurries were treated with both trichlor obiphenyls or only 2,3,6-CB. Likewise, paradechlorination of 2,4,6-CB and para dechlorination activity on Aroclor 1242 (pattern Q) had the sameacclimation time within a sediment slurry; these dechlorinations also bad the same acclimation time whencomparing pasteuriz ed slurries amended with 2,4,6-CB, with and without 2,3,6-CB addition. In slurries of thesame sedime nt to which Aroclor 1242 but neither 2,3,6-CB nor 2,4,6-CB was added, pattern M activity wasobserved by 36 days and pattern Q was not observed after 120 days (separate experiment, unpublished data)

The acclimation times for patterns M and Q dechlorination activities differe d relative to each other, andrelative to the pasteurization or the trichlorobiphen yl treatments (Table 2). Pattern M activity occurred soonerthan pattern Q activity within the same slurry and between the differently treated slurries. Pasteurization of theslurries caused shorter acclimation times for all the dechlorination activities relative to their acclimation timesin the unheated slurries. The pattern Q activity occurred sooner in the unheated slurries to which both


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6 6 4t r i c h l o r ob i p h e ny l s w e r e a d d e d , t h a n i n t h e u nh e a t e d s l u r r i e s to w h i c h on l y 2 , 4 , 6 - C B w a s a d d e d . P a t t e rn Qa c t i v i t y a l so oc c u r r e d i n s l u r r i e s su p p l e me n t e d w i t h on l y 2 , 3 , 6 - C B ( b o t t om h i s t og r a m i n F i gu r e 3 ) , b u t l a t e rt h a n i n s lu r r i e s c on t a i n i ng 2 , 4 , 6 - C B . T h e s i gn i f i c a nc e o f th e se a c c l i ma t i on t i me d i f f e r e nc e s a nd s i m i l a r i t ie s isd i s c u s se d b e l ow .E n r i c h m e n t o f M i c r o b i a l P C B D e c h l o r in a t i o n A c t iv i ti e s. E n r i c h m e n t c u l t u r e s o f e a c h d e c h l o r i n a t i o na c t i v i t y w e r e p r e p a r e d b y i n o c u l a t i n g a c u l t u r e m e d i u m w i t h t h e 2 , 3 , 6 - C B - a m e n d e d o r 2 , 4 , 6 - C B - a m e n d e d ,me t h a noge n i c s e d i me n t s l u r r i e s a t d a y 4 3 o f t h e e xp e r i m e n t ( w h e n M o r Q w a s t h e on l y d e c h l o r i na t i on p a tt e rnseen in each s lu r ry) . Af t e r 9 days , meta d e c h l o r i na t i on o f 2 , 3 ,6 - C B a nd p a t t er n M d e c h l o r i na t i on o f t h e A r oc l o r1 2 4 2 w a s ob se r v e d i n t h e 2 , 3 , 6 - C B - a me nd e d e n r i c h me n t c u l t u r e s t h a t w e r e i noc u l a t e d w i t h t h e s e d i me n t s l u r r yi n w h i c h p a t t e m M w a s ob se r v e d . A f t e r 1 4 d a y s , para d e c h l o r i na t i on o f 2 , 4 , 6 - C B a nd p a t t e rn Q d e c h l o r i na t iono f t h e A r oc l o r 1 2 4 2 w a s ob se r v e d i n t h e 2 , 4 , 6 - C B - a me n d e d e n r i c h m e n t c u l t u r e s t h a t w e r e i noc u l a t e d v a t h th ese d i m e n t s l u r ry i n w h i c h p a t t e r n Q w a s ob se r v e d . B y t h e l a st s a m p l i ng t i me ( 3 6 d a y s ) , on l y t h e mi c r ob i a ld e c h l o r i na t i on a c t i v i t y t h a t w a s t r a ns f e r r e d oc c u r r e d i n e a c h e n r i c h m e n t c u l t u re . P a t t e r n M a c t i v i t y w a sma i n t a i ne d t h r ou gh f i v e s e ri a l tr a ns fe r s . H o w e v e r , p a t te r n Q a c t i v i t y w a s no t ob t a i ne d a f t e r t h e t h ir d s e ri a lt ransfer .

T h e e n r i c h me n t c u l t u r e me d i a d i f f e r e d f r om t h e s e d i me n t s l u rr y me d i a , i n th a t c omp l e t e H u d son R i v e rse d i me n t w a s u se d i n t h e sl u r ry m e d i a , w h e r e a s a h u mi c a c i d e x tr a c t o f P C B - f r e e u p p e r H u d son R i v e r s e d i me n tw a s u se d i n th e e n r i c h m e n t c u l t u r e me d i a .

A n t i b i o t i c t r e a tm e n t o f a m i c r o b i a l m i c r o c o s m c a n b e u s e d t o i d e n ti f y o r s e l e c t s p e c i f i c m i c r o b ia lp op u l a t i ons . T h e e n r i c h me n t c u l t u r e s o f t h e mi c r ob i a l P C B d e c h l o r i na t i on a c t i v i t i e s w e r e tr e a t e d w i t h t w oa n t i b io t i c s , v a n c om y c i n a nd n i s in , th a t a r e know n t o i nh i b i t g r a m - p os i t i v e mi c r oo r ga n i sm s . V a n c om y c i n i s ag l y c op e p t i d e t h a t i nh i b i ts c r o s s l i nk i ng o f t h e p e p t i d og l y c a n su b u n i t s on t h e b a c t e r i al c e l l me m b r a ne ; t h e l a r ges i z e o f t h e v a n c om y c i n mo l e c u l e ( 1 4 88 d a l t ons ) s e v e r e l y i mp a i r s i ts e f f e c t i v e ne s s a ga in s t mo s t g r a m- ne ga t i v eb a c t e r ia , w h i c h h a v e s e v e r a l ou t e r l a y e r s su r rou nd i ng t h e c e l lu l a r me mb r a ne [ 23 ]. N i s i n i s a sma l l p o l y p e p t id et h at i n h i b i ts g r o w t h o f a b r o a d s p e c t ru m o f g r a m - p o s i t iv e b a c t e r ia a n d t h e o u t g r o w t h o f g e r m i n a t e d Clostridiaspores [24 , 25]. No PC B de ch lor in a t ion was obser ved in any enr i chm ent cu l tu re t rea t ed wi th 5 , 50 , or 500 ppmv a nc o my c i n ; o r 5 0 0 p p m n i s in . P C B d e c h l o r i na t ion w a s ob se r v e d i n t h e e n r i c h me n t c u l t u r e s tr e a t e d w i th 5 and5 0 p p m n i s i n . T h e se r e su l ts sh o w t h a t a g r a m - p os i t i v e mi c r ob i a l p op u l a ti on i s e s s e n ti a l f o r b o t h m i c r ob ia l P C Bdechlor ina t ion ac t iv i t i e s .

D I S C U S S I O NT r i c h l o r o b i p h e n y l S t i m u l a t i o n o f P C B D e c h l o r i n a t io n A c t i v it i es . T h e a d d i t i ons o f 2 , 3 , 6 - C B a nd 2 , 4 , 6 - C Bt o P C B - c o n t a m i n a t e d u p p e r H u d s o n R i v e r s e d i m e n t s l u r r ie s s t i m u l a te t h e p r o d u c t io n o f r e d u c ti v ed e c h l o r i na t i on p a t t e r n s M a nd Q , r e sp e c t i v e l y . T h e s e c onc l u s i ons a r e su p p or t e d b y t h e f o l l ow i ng e v i d e nc e .


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6 6 5F i r s t , p a t t e ms M a nd Q w e r e ob se r v e d a t t h e s a me sa mp l i ng t i me a s m e t a a n d p a r a d e c h l o r in a t l o n o f 2 , 3 , 6- C Ba nd 2 , 4 , 6 - C B , r e sp e c t i v e l y ( T a b l e 2 ). E xa c t a c c l i ma t i on t i me s a nd d e c h l o r i na t i on r a t e s i n t h e h e t e r oge ne ou sse d i m e n t s l u rr i e s c ou l d no t b e me a su r e d . H o w e v e r , t h e d e c h l o r i na t i on p a t t er n s w e r e i d e n t i f i e d f r om t h e P C Bc o n g e n e r d i s t r i b u t i o n i n t h e s e d i m e n t s lu r r i es . F o r e x a m p l e , in s l u r r i e s s u p p l e m e n t e d w i t h b o t ht r i c h l o r ob i p h e ny l s , p a t t er n M d e c h l o r i na t i on a nd m e t a d e c h l o r i na t i on o f 2 , 3 , 6 - C B w e r e c l e a r l y ob se r v e d b e f o r ep a t t er n Q d e c h l o r i na t i on a nd p a r a d e c h l o r i na t i on o f 2 , 4 , 6 - C B . S e c ond , p a t te r n Q oc c u r r e d s i gn i f i c a n t ly soone ri n t h e 2 , 4 , 6 - C B - a m e nd e d s l u r r i e s th a n i n t h e 2 , 3 , 6 - C B - a me nd e d s l u rr i es . T h i r d , p a t te r n Q b u t no t p a t te r n Mw a s ob se r v e d i n s l u r ri e s a me nd e d w i t h on l y 2 , 4 , 6 -C B . F ou r t h , t h e a c c li ma t i on t i me f o r p a t t er n M w a s l onge r ins l u r r i e s w i t h no t r i c h l o r ob i p h e ny l t r e a t me n t t h a n i n s l u r r i e s t r e a t e d w i t h 2 , 3 , 6 - C B ( 3 6 d a y s c omp a r e d t o 2 5d a y s ) ; p a tt e r n Q w a s no t ob se r v e d i n s l u r r i e s w i t h no t ri c h l o r ob i p h e ny l a d d i ti on . F i f t h , s e p a r a t e e n r i c h me n tc u l tu r e s o f e a c h m i c r o b i a l d e c h l o r in a t i o n a c t i v it y w e r e p r e p a r e d w i th a m e n d m e n t s o f 2 , 3 ,6 - C B o r 2 , 4 , 6 - C BT h e se d a t a d e mons t r a t e t h a t e a c h t r i c h l o r ob i p h e ny l a me nd me n t i s a s soc i a t e d w i t h a d i s t i nc t m i c r ob i a l P C Bd e c h l o r i na t i on p a t te r n , i m p l y i ng a s t i mu l a t o r y r o l e o f t h e t w o c ong e ne r s i n th e s e d i me n t s l u r r i e s

A p a t te r n M a nd t h e n p a t te r n Q d e c h l o r i na t i on s e qu e nc e w a s ob se r v e d i n s e d i m e n t s l u r r ie s t r e a te d w i t hb o t h t r i c h l o r ob i p h e ny l s ( F i gu r e 2 ) a nd i n s e d i me n t s l u r r i e s a me nd e d w i t h on l y 2 , 3 , 6 - C B ( b o t t om h i s t og r a m i nF i gu r e 3 ) . I n t h e l a tt e r c ase , p a t te r n Q w a s ob se r v e d s i gn i f i c a n t l y l a t e r t h a n p a t t e r n M a nd a l so l a t e r t h a np a t t er n Q i n th e 2 , 4 , 6 - C B - a m e nd e d s l u r r ie s ; i t i s u nc l e a r i f p a t te r n Q w a s s t i mu l a t e d b y t h e 2 , 3 , 6 - C B a d d i ti on ,t h e p a t t e r n M d e c h l o r i na t i on p r od u c t s , o r a no t h e r f a c t o r ( s ) w i t h i n t h e 2 , 3 , 6 - C B - a me nd e d s l u r r i e s ( d i s c u s se df u r t h e r b e l ow ) . T h e r e h a v e b e e n s e v e r a l r e p o r ts t h a t th e s e qu e n t ia l p a t te r n M a nd t h e n p a tt e r n Q d e c h l onna t i ona c t i v i ti e s oc c u r r e d i n P C B - c on t a mi na t e d u p p e r H u d son R i v e r s e d i me n t s l u r r i es o r m i c r oc o sms [ 6, 9 , 1 1 , 1 3 -1 5 ]I t was proposed [6 , 15] t ha t t oge ther t hese ac t iv i t i e s cause a pa t t e rn C dech lor ina t ion , which i s an env i ronmenta lP C B d e c h l o r i na t i on p a tt e rn f ou nd i n u p p e r H u d son R i v e r s e d ime n t s [ 2 - 5 ]

I t i s p os s i b l e t h a t t h e su c c e s s i v e , m i c r ob i a l p a t t e r n s M a nd t h e n Q d e c h l o r i na t i on a c t i v me s d o no tf u nc t i on i nd e p e n d e n t l y T h i s p os s i b i l it y is su gge s t e d m t w o w a y s . F i r st , i n s l u r r ie s t o w h i c h on l y 2 ,3 , 6 - C B w a sa d d e d , p a t t e r n M w a s s t i m u l a t e d b y t h e t r i c h l o r o b i p h e n y l a d d i t i o n b u t p a t t e r n Q a l s o o c c u r r e d , a l b e i ts i gn i f i c a n t l y la t e r. S e c on d , i n t h e u nh e a t e d s l u r r i e s to w h i c h 2 , 4 , 6 - C B w a s a d d e d t h e r e w a s a sh o r t e ra c c l i ma t i on t i m e f o r p a t te r n Q a c t i v i ty i f p a t t e rn M a c t i v i ty h a d oc c u r r e d ; t h e a c c l i ma t i on t i me w a s 3 6 d a y s i ns l u r r i e s a me nd e d w i t h b o t h t r i c h l o r ob i p h e ny l s c omp a r e d t o 4 1 d a y s i n s l u r r i e s a me nd e d w i t h on l y 2 , 4 , 6 - C BT h e s e d a t a su gge s t th a t t h e p a t t e r n M a c t i v i t y m i g h t h a v e a f f e c t e d s t imu l a t i on o f p a t t e r n Q a c t i v i t y T h ed i f f e r e nc e i n t h e a c c l i ma t i on t i me s f o r p a tt e r n Q a c t i v i t y in u nh e a t e d s l u r r i e s a me nd e d w i t h 2 , 4 , 6 - C B w a s no tob se r v e d i n p a s t e u r i z e d s l u r ri e s , i mp l y i ng t h a t p a s te u r i z a ti on e l i m i na t e s some i n t e ra c t i on b e t w e e n t h e t w o P C Bd e c h l o r i na t i on a c t i v i t i e s . T h e se d a t a c ou l d i nd i c a t e t h a t p a t t er n C d e c h l o r i na t i on ( t h e r e su lt o f c om b i ne dpa t t e rns M and Q) i s cause d by a mic ro b ia l consor t ium. She l ton and Tied je [26] i so l a t ed an anaerob ic , bac te r i a lc onso r t i u m t h a t c ou l d d e c h l o r i na t e a nd mi ne r a l i z e 3 - c h l o r ob e nz o i c a c i d


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6 6 6T h e c u r r e n t r e se a r c h sh ow s t h a t p a s t e u r i z a ti on c a n b e u se d i n c on j u nc t i on w i t h 2 , 3 , 6 - C B o r 2 , 4 ,6 - C B t o

s t i m u l a t e m i c r o b i a l p at te r n s M a n d Q d e c h l o r i n a t io n a c t i v i t ie s in P C B - c o n t a m i n a t e d u p p e r H u d s o n R i v e rs e d i m e n t . T h i s c o n c l u s i o n i s s u p p o r t e d b y t h e o b s e r v a t i o n t h a t p a s t e u r i z a ti o n c a u s e d a d e c r e a s e in th ea c c l i ma t i on t i me s f o r b o t h d e c h l o r i na t i on a c t i v i t i e s w h e n c omp a r e d t o t h e i r a c c l i ma t i on t i me s i n t h e u nh e a t e dsa mp l e s ( T a b l e 2 ).

S t i m u l a t i o n a n d e n r i c h m e n t o f tw o o t h e r para d e c h l o r i na t i on a c t i v i t i e s , p r oc e s se s P a nd H , h a v er e c e n t l y b e e n r e p o r t e d [ 1 7, 2 7 ] T h e c o nge n e r sp e c i f i c i t y o f p r oc e s s Q is d i s t i nc t l y d i f f e r e n t f r om t h e o t h e rpara d e c h l o r i na t i on a c t i v i ti e s . P a t te r n Q d e c h l o r i na t i on r e su l t s i n r e mo v a l o f a l l para c h l o r i ne s f r om mono- , d i- ,a nd t r i c h l o r op h e ny l g r ou p s a nd i s no t l i m i t e d t o r e mov a l o f f l a nke d para c h l o r i ne s l i ke p r oc e s se s P a nd H [ 6] .B e d a r d a n d c o - w o r k e r s [ 17 ] r e p o r t e d s t i m u l a ti o n o f p a t te m P d e c h l o r i n a ti o n o n P C B s i n H o u s a t o n i c R i v e rse d i me n t s l u r r ie s b y t h e a d d i t i on o f 2 ,5 ,3 ' , 4 ' - C B . Y e e t a l. [ 2 7 ] p r ov i d e d e v i d e nc e t h a t p a t t e m H d e c h l o r i na ti oni s c a u se d b y me t h a no ge n i c b a c t e r ia . T h e r e su l ts o f th e p a s t e u r iz a t i on a nd a n t i b i o t ic t re a t me n t s i n t h e c u r re n !r e p o r t, a nd o f Y e e t al . [2 7 ] su gg e s t t h a t p a t te r n s H a nd Q a r e ca u se d b y t w o d i f f e r e n t m i c r ob i a l p op u l at ions .E n r i c h m e n t o f M i c r o b i a l P C B D e c h l o r i n a t i o n A c t i v it i e s . A 1 0 % by v o l u m e i n o cu l a ti o n o f a c u lt u reme d i u m w i t h a s e d i me n t s l u r r y c on t a i n i ng e i t h e r P C B d e c h l o r i na t i on a c t i v i t y r e su l t e d i n a d e c r e a se i n t h ea c c l i m a t i o n t i m e f o r d e c h l o r i n a t i o n r e l a t i v e t o w h a t w a s o b s e r v e d i n th e p r i m a r y c u l t u re s ( t h e s e d i m e n ts l u rr i es ) . T h e s e r e su lt s d e mons t r a t e e n r i c h m e n t o f e a c h mi c r ob i a l a c t i v i t y F o r e xa mp l e , t h e p a t t er n Q a c t i v i tyh a d a 4 1 - d a y a c c l i ma t i on t i me i n t h e u nh e a t e d s e d i me n t s l u r r i e s , b u t on l y a 1 4 - d a y a c c l i ma t i on t i me i n t h eenr i ch m ent cu l tu res . Th i s i s t he f ir s t repor t of a mic rob ia l pa t t e rn Q dech lor ina t ion a c t iv i ty enr i ch me nt cu ltu re .B o t h M or r i s e t a l. [ 16 ] a nd Y e e t al . [ 1 4 ] r e p o r t e d l o s s o f p a t t e r n Q a c t i v i t y in e n r i c h m e n t c u l t u r e s p r e p a r e dw i t h a me t h od s i m i l a r t o t h e c u r re n t e xp e r i me n t s .

P r e v i ou s l a b o r a t o r y i nv e s t i ga t i ons o f m i c r ob i a l P C B d e c h l o r i na t i on h a v e u se d s e d i m e n t b o t h a s a sou rc eo f m i c r oo r ga n i sm s a nd a g r ow t h m a t r i x [6 , 8 , 10 , 1 6 , 1 4 ] . W h i l e iso l a t i on o f a b a c t e r i u m c a p a b l e o f P C Bd e c h l o r i na t i on h a s no t b e e n su c c e s s f u l , M a y a nd c o - w o r ke r s [ 1 0 ] w e r e su c c e s s f u l i n su b c u l t u r i ng a P C B -d e c h l o r i n a ti n g , a n a e r o b i c e n r i c h m e n t o n so l i d m e d i a H o w e v e r , th e i r b a c t e ri a l c o l o n y c o u l d o n l y c a u sed e c h l o r i na t i on i n a s e d i me n t s l u r ry . A d e f i ne d c u l t u re me d i u m i s ne e d e d f o r i so l at i on o f t h e mi c r oo r ga n i sm o rc onso r t i u m t h a t c a u se s P C B d e c h l o r i na t i on a nd fo r u nd e r s ta nd i ng h ow P C B d e c h l o r i na t i on oc c u rs . T o t h i s e nd ,t h e c u r r e n t e xp e r i me n t s i nd i c a t e t h a t th e h u m i c a c i d f r a c t ion o f r i v e r s e d i me n t , s e p a r a t e f r om t h e s a nd a nd t h ec l a y f r a c t ions , su p p o r t s m i c r ob i a l P C B d e c h l o r ina t i on . T h e p h y s i c a l o r c h e mi c a l me c h a n i sm s b y w h i c h h u m i ca c i d s su p p or t P C B d e c h l o r i na t i on a r e no t know n .

V e r y l i tt l e i s k n o w n a b o u t w h a t g e n e r a o f b a c te r i a m i g h t c a u s e P C B r e d u c t i v e d e c h l o r i n a ti o n .P a s t e u r i z a t i on e xp e r i m e n t s h a v e sh ow n t h a t sp o r e - f o r me r s c a u se p a t te r n s M a nd Q d e c h l o r i na t i on [ 1 3 , 1 4 , a ndt h e c u r r e n t s t u d y ] . B e d a r d a nd Q u e nse n [ 6] p r op ose d t h a t p a t t e r n M d e c h l o r i na t i on c ou l d b e c a u se d b y asu l f a t e - r e d u c i ng , sp o r e - f o r mi ng b a c t e r i u m su c h a s a sp e c i e s o f Desul/btomaculum, w h i c h i s t h e on l y know ng e n u s o f s u l fa t e -r e d u c i n g s p o r e - f o r m e r s Desul~tomac~dllm a r e g r a m- ne ga t i v e b a c t e r i a ; h e nc e t h e y a r e l i ke l y


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667to be resistant to vancomycin or nisin treatments. On the other hand, Clostridia are gram-positive, anaerobic,spore-forming bacteria; but sulfate reduction is uncommon in species of this genus. Further study of theenrichment' cultures of each microbial PCB dechlorination activity should provide insights about the type ofbacteria that cause PCB reductive dechlorination.

R E F E R E N C E S1 D.A . Abramowicz, Aerobic and anaerobic biodegradation of PCBs: a review, In Critical Reviews in

Biotechnology (Edited by G. G Steward and I. Russell), Vol. 10, pp. 241-251. CRC Press, Inc., Boca Raton,FL (1990).

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3 J. F. Brown, Jr. and R E. Wagner, PCB movement, dechlorination, and detoxication in the AcushnetEstuary, Environ. ToxicoL Chem. 9, 1215-1233 (1990).

4. J. F. Brown, Jr., R E Wagner, H. Feng, D. L. Bedard, M. J. Brennan, J. C. Camahan and R J. May,Environmental dechlorination of PCBs, Environ. 7bxicol. Chem. 6, 579-593 (1987).

5. J.F. Brown, Jr., R E Wagner, H. Feng, D. L Bedard, M. J Brennan, J C Camahan, R J. May and T JTofflemire, PCB transformation in upper Hudson River sediments, Northeast. Environ Sci. 3, 167-179(1984).

6 D. L Bedard and Quensen, J. F., III, Microbial reductive dechlorination of polychlorinated biphenyls, InMicrobial Transformation and Degradation o]" Toxic Organic Chemicals (Edited by L Y Young and CCerniglia), pp. 127-216. Wiley-Liss Division, John Wiley & Sons, Inc, New York (1995)

7 A C. Alder, M. M. Haggblom, S. R Oppenheimer and L Y Young, Reductive dechlorination ofpolychlorinated biphenyls in anaerobic sediments, Environ. Sci Technol. 27, 530-538 (1993)

8. A W. Boyle, C. K Blake, W. A. Price, II and It. D. May, Effects of polychlorinated biphenyl congenerconcentration and sediment supplementation on rates of methanogenesis and 2,3,6-trichlorobiphenyldechlorination in an anaerobic enrichment., Appl. Environ. Microbiol. 59, 3027-3031 (1993)

9. K M. Fish and J. M. Principe, Biotransformation of Aroclor 1242 in Hudson River test tube microcosms,Appl. Environ. MicrobioL 60, 4289-4296 (1994).

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66 811. J. F . Q uensen, ~ I , S . A, ,Boyd and J . M. Tied je , Dechlor ina t ion of four com mercia l polyc hlor ina ted b iphenyl

mixtures (Aro clors ) by anaerobic m icroorganism s from sed iments, Appl. Environ Microbiol. 56, 2360-2369(1990).

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1 3. W . A . W i l l i a ms , Mi c ro b i a l rd d u c t i v e d e ch l o r i na t ion o f p o l y ch l o r i na t e d b i p h e ny l s i n h e a t t r e a te d a ndbrom oethanesu l fona te t rea ted an aerobic sed im ent s lurr ie s , Chemo~t~here28, 2269-2284 (1994).

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16. P . J . M orr i s , W . W. M ohn, J . F . Quen sen, I I I , J . M. Tied je and S. A. Bo yd, Es tab l i sh m ent of ap o l y c h l o r i na t e d b i p h e n y l -d e g ra d i ng e n r i ch me n t cu l t u re w i th p re d om i na n t l y meta dechlor ina t ion , Appl.Environ. Microbiol. 58, 3088-3094 (1992).

1 7. D . L . B e d a rd , S . C . B u nn e l l a nd L . A . Sm u l l e n , S t i mu l a t i on o f mi c rob i a l para-dechlorination o tp o l y ch l o r i na t e d b i p h e ny l s th a t h a v e p e r s i s te d i n H ou s a t on i c R i v e r s e d i me n t fo r d e ca d e s , Environ. SciTechnol. 30, 687-694 (1996).

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