Annotation of Metagenomes: from Jigsaws to Taxa

8/2/2019 Annotation of Metagenomes: from Jigsaws to Taxa

1/52

I n t r o d u c t i o n t o M e t a g e n o m i c s T a x o n o m i c A n n o t a t i o n M T R d e s c r i p t i o n R e s u l t s C o n c l u s i o n

A n n o t a t i o n o f M e t a g e n o m e s : f r o m J i g s a w s t o T a x a

F a b i o G o r i

1

G i a n l u i g i F o l i n o

2

M i k e S . M . J e t t e n

3

E l e n a

M a r c h i o r i

1

1

R a d b o u d U n i v e r s i t y N i j m e g e n , I n s t i t u t e f o r C o m p u t i n g a n d I n f o r m a t i o n S c i e n c e s ,

T h e N e t h e r l a n d s

3

R a d b o u d U n i v e r s i t y N i j m e g e n , D e p a r t m e n t o f M i c r o b i o l o g y , T h e N e t h e r l a n d s

2

I C A R - C N R , R e n d e , I t a l y

N i j m e g e n , 1 0 S e p t e m b e r 2 0 1 0

g o r i @ s c i e n c e . r u . n l


2/52


T a b l e o f C o n t e n t s

I n t r o d u c t i o n t o M e t a g e n o m i c s

T a x o n o m i c A n n o t a t i o n

M T R d e s c r i p t i o n

R e s u l t s

C o n c l u s i o n


3/52


W h a t i s M e t a g e n o m i c s ?

M e t a g e n o m i c s :

s t u d y o f m i c r o b i a l

c o m m u n i t i e s a n a l y s i n g

t h e i r g e n e t i c m a t e r i a l

W h y ?

M o s t m i c r o b e s

c a n n o t b e c u l t u r e d

U n d e r s t a n d o r g a n i s m s

i n t e r a c t i o n s


4/52







W h y ?






5/52







W h y ?






6/52


H o w ? D N A S e q u e n c i n g T e c h n o l o g y

E n v i r o n m e n t a l

S a m p l e

D N A s

S m a l l - I n s e r t L i b r a r y C l o n i n g

A m e t a g e n o m i c d a t a s e t i s m a d e b y w o r d s i n 4 - l e t t e r a l p h a b e t

{A , C , G , T }


7/52


W h a t k i n d o f d a t a ? A m e t a . . . j i g s a w - p u z z l e

F r a g m e n t s o f D N A s

P i e c e s a r e s i m i l a r

O r i g i n a l p i c t u r e s a r e

u n k n o w n

M i s s i n g P i e c e s


8/52


A s s e m b l y o f m e t a g e n o m i c s f r a g m e n t s i s o f t e n n o t p o s s i b l e

( a ) L o w - d i v e r s i t y c o m m u n i t i e s ( b ) H i g h - d i v e r s i t y c o m m u n i t i e s


9/52


S o . . . W h a t c a n M e t a g e n o m i c s d o ?


10/52


E x a m p l e s o f m e t a g e n o m i c s s t u d i e s

S t u d y o f f u n c t i o n a l c o n t e n t o f a m e t a g e n o m e

[ S . D e m a n e c h e e t a l . , J o u r n a l o f M i c r o b i o l o g i c a l M e t h o d s , 2 0 0 9 ]



11/52

S t u d y o f t a x o n o m i c c o n t e n t o f a m e t a g e n o m e

[ J . F . B i d d l e e t a l . , P N A S , 2 0 0 8 ]



12/52




R e s u l t s

C o n c l u s i o n



13/52

T a x o n o m y : a b i o l o g i c a l c l a s s i c a t i o n

L i n n e a n t a x o n o m y :

F o r m a l s y s t e m f o r c l a s s i f y i n g a n d n a m i n g

l i v i n g t h i n g s

B a s e d o n a s i m p l e h i e r a r c h i c a l s t r u c t u r e

S i m i l a r e l e m e n t s a r e g r o u p e d t o g e t h e r

R a n k : l e v e l i n t h e h i e r a r c h y ( l e f t )

T a x o n : u n i t o f t h e h i e r a r c h y

( g r o u p o f s i m i l a r l i v i n g t h i n g s )



14/52

A s i m p l i e d b r a n c h o f t h e t a x o n o m y

T h e s t r u c t u r e o f t h e t a x o n o m y c a n b e r e p r e s e n t e d a s a t r e e :

N o d e s a r e t a x a

L e a v e s a r e s p e c i e s



15/52

E x a m p l e s o f t a x o n o m i c c l a s s i c a t i o n s



16/52

T h e m e t a j i g s a w - p u z z l e

F r a g m e n t s o f D N A s



17/52

A n n o t a t i o n : d i s c o v e r i n g t h e o r i g i n a l p i c t u r e s o f t h e p u z z l e s

A s s i g n e a c h f r a g m e n t

t o a n o r g a n i s m o r t a x o n

( o f r a n k s p e c i e s , o r g e n u s . . . )



18/52

R e l a t e d w o r k s

C o m p o s i t i o n - b a s e d m e t h o d s : b a s e d o n n - g r a m s f r e q u e n c i e s

S i m i l a r i t y - b a s e d m e t h o d s : b a s e d o n c o m p a r i s o n w i t h

r e f e r e n c e s e q u e n c e s

O r g a n i s m - l e v e l c l a s s i c a t i o n :

B e s t H i t

( B H )

R a n k - l e v e l c l a s s i c a t i o n :

L o w e s t C o m m o n A n c e s t o r

( L C A ) ,

u s e d b y M E G A N a n d G a l a x y



19/52

C o m p a r e A l i g n : B L A S T s o f t w a r e

L o c a l a l i g n m e n t ( g a p s a l l o w e d )

N u c l e o t i d e a n d / o r P e p t i d e s e q u e n c e s

A l i g n m e n t S c o r e s

B i t S c o r e (SB

) = a l i g n m e n t q u a l i t y

E - v a l u e

P - v a l u e



20/52

S i m i l a r i t y - b a s e d m e t h o d s

W i t h v e r y s h o r t f r a g m e n t s ( 1 0 0 b p )

M e t h o d C o m p a r i s o n B H L C A

C l a s s i c a t i o n L e v e l O r g a n i s m R a n k s ( t o o h i g h )

Q u a n t i t y o f f r a g m e n t s A l l F i l t e r e d ( l o w )

A c c u r a c y N o t v e r y g o o d Q u i t e g o o d

G o a l : T a x o n o m i c A n n o t a t i o n o f S h o r t M e t a g e n o m i c s f r a g m e n t s

( r a n k - l e v e l ) s . t .

A s s i g n a s m a n y f r a g m e n t s a s p o s s i b l e

A s s i g n w i t h g o o d a c c u r a c y

A s s i g n t o t h e i r l o w e s t f e a s i b l e r a n k



21/52








( r a n k - l e v e l ) s . t .






22/52








( r a n k - l e v e l ) s . t .






23/52




R e s u l t s

C o n c l u s i o n



24/52

T h e m e t h o d : c o m p a r e a n d a s s i g n

M u l t i p l e T a x o n o m i c R a n k b a s e d c l u s t e r i n g


( r a n k - l e v e l )

1 A l i g n t h e f r a g m e n t s t o r e f e r e n c e s e q u e n c e s , s e l e c t i n g g o o d

m a t c h i n g s

2 A s s i g n e a c h f r a g m e n t t o a t a x o n o f o n e o f i t s b e s t m a t c h i n g

s e q u e n c e s

N E W

A s s i g n f r o m t h e h i g h e s t r a n k t o t h e l o w e s t f e a s i b l e r a n k

A s s i g n m e n t s o f f r a g m e n t s a r e d e p e n d e n t e a c h o t h e r



25/52

T h e m e t h o d : c o m p a r e a n d a s s i g n

M u l t i p l e T a x o n o m i c R a n k b a s e d c l u s t e r i n g


( r a n k - l e v e l )

1 A l i g n t h e f r a g m e n t s t o r e f e r e n c e s e q u e n c e s , s e l e c t i n g g o o d

m a t c h i n g s

2 A s s i g n e a c h f r a g m e n t t o a t a x o n o f o n e o f i t s b e s t m a t c h i n g

s e q u e n c e s

N E W

A s s i g n f r o m t h e h i g h e s t r a n k t o t h e l o w e s t f e a s i b l e r a n k

A s s i g n m e n t s o f f r a g m e n t s a r e d e p e n d e n t e a c h o t h e r



26/52

H o w c a n w e a l i g n s e q u e n c e s ? B L A S T

B l a s t X

W e s e l e c t t h e g o o d m a t c h i n g s a c c o r d i n g t o t h e s e i n d e x e s :

S

B

(r

,p

) =q u a l i t y o f a l i g n m e n t

E - v a l u e ( r , p ) P - v a l u e

P o s i t i o n o f p i n t h e l i s t o f h i t s o f r



27/52

O n a x e d r a n k , h o w d o w e a s s i g n t h e f r a g m e n t s ?

F o r e a c h r a n k j :

C l u s t e r o f f r a g m e n t s C

i

T a x o n t

i

o f r a n k j

C l u s t e r s e l e c t i o n = a s s i g n m e n t .

S e t C o v e r i n g P r o b l e m

m i n

I

{1,...,

n

}| I |, s u c h t h a t

i

I

C

i

=R

.( S C P )

R

:= {r1

, . . . , rm

}, f r a g m e n t s t h a t w e w a n t t o a s s i g n

C j := {C

i , . . . , C n }, c l u s t e r s o f f r a g m e n t s ( C

i R )

S C P i s N P - h a r d ; w e u s e d a f a s t g r e e d y a l g o r i t h m

[ C h v a t a l V , 1 9 7 9 ]



28/52

S e t C o v e r i n g P r o b l e m

C l u s t e r o f f r a g m e n t s C

i

T a x o n ti

o f r a n k j

S e l e c t c o l l e c t i o n o f c l u s t e r s ( t a x a ) s . t .

N o f r a g m e n t i s l e f t o u t s i d e

M i n i m a l n u m b e r o f s e l e c t e d c l u s t e r s

E x a m p l e :

C

1

C

2

C

3

C

4

C

5

C

6

r

1

r

2

r

3

r

4

r

5

r

6

r

7

r

8

r

9

r

1 0

C l u s t e r i n g S o l u t i o n :

C

1

C

2

C

3

C

4

C

5

C

6

r

1

r

2

r

3

r

4

r

5

r

6

r

7

r

8

r

9

r

1 0



29/52

A l g o r i t h m s c h e m e

1 C o m p a r e f r a g m e n t s R w i t h r e f e r e n c e p r o t e i n s ( N C B I - N R )

u s i n g B l a s t X a n d F i l t e r B l a s t X o u t p u t

2 F r o m t h e h i g h e s t t o t h e l o w e s t r a n k

( j=

k i n g d o m , . . . ,

s p e c i e s )

1 C r e a t e p o t e n t i a l c l u s t e r s c o l l e c t i o n Cj

, f o r r a n k j . ( C

i

R ) .

r

C

i

r h a s a B l a s t X - m a t c h w i t h a p r o t e i n o f t a x o n t

i

2 S e l e c t c l u s t e r s f r o m Cj

v i a S e t C o v e r i n g P r o b l e m

3 I d e n t i f y f r a g m e n t s w i t h i n c o h e r e n t t a x o n o m i c c l a s s i c a t i o n

( w . r . t . h i g h e r r a n k s c l a s s i c a t i o n s ) a n d r e m o v e t h e m , e v e n

f r o m R

3F r o m t h e l o w e s t t o t h e h i g h e s t r a n k

1 F o r t h i s r a n k , M a j o r i t y V o t e o n c l u s t e r s ' i n t e r s e c t i o n s

2 M a k e h i g h e r r a n k s c l a s s i c a t i o n s c o h e r e n t w i t h t h e M a j o r i t y

V o t e r e s u l t s



30/52





( j=

k i n g d o m , . . . ,

s p e c i e s )


, f o r r a n k j . ( C

i

R ) .

r

C

i


i





f r o m R







31/52





( j=

k i n g d o m , . . . ,

s p e c i e s )


, f o r r a n k j . ( C

i

R ) .

r

C

i


i





f r o m R







32/52





( j=

k i n g d o m , . . . ,

s p e c i e s )


, f o r r a n k j . ( C

i

R ) .

r

C

i


i





f r o m R






33/52



34/52

S u m m a r y

M T R

G o a l : T a x o n o m i c A n n o t a t i o n o f S h o r t M e t a g e n o m i c s

f r a g m e n t s

M e t h o d :

C o m p a r i s o n w i t h r e f e r e n c e s e q u e n c e s

C o h e r e n t r a n k - b y - r a n k c l u s t e r i n g

M a j o r i t y v o t e o n i n t e r s e c t i o n s



35/52




R e s u l t s

C o n c l u s i o n



36/52

E x p e r i m e n t s - T h e D a t a

9 s i m u l a t e d d a t a s e t s , f r o m 3 s e t o f o r g a n i s m s a t 3 l e v e l o f c o v e r a g e

( 0 . 1 X , 1 X a n d 4 X )

L e n g t h :

1 0 0 b p

D a t a s e t M 1 a r t i c i a l l y g e n e r a t e d f r o m 9 g e n o m e p r o j e c t s

O r g a n i s m G e n o m e s i z e ( b p ) F r a g m e n t s s a m p l e d

C l o s t r i d i u m p h y t o f e r m e n t a n s I S D g 4 5 3 3 5 1 2 4 6 3 7 9

P r o c h l o r o c o c c u s m a r i n u s N A T L 2 A 1 8 4 2 8 9 9 1 8 6 8 1

L a c t o b a c i l l u s r e u t e r i 1 0 0 - 2 3 2 1 7 4 2 9 9 2 3 7 1 0

C a l d i c e l l u l o s i r u p t o r s a c c h a r o l y t i c u s D S M 8 9 0 3 2 9 7 0 2 7 5 2 9 4 9 6

C l o s t r i d i u m s p . O h I L A s 2 9 9 7 6 0 8 2 9 3 4 8

H e r p e t o s i p h o n a u r a n t i a c u s A T C C 2 3 7 7 9 6 6 0 5 1 5 1 6 9 3 8 7

B a c i l l u s w e i h e n s t e p h a n e n s i s K B A B 4 5 6 0 2 5 0 3 4 5 4 6 3

H a l o t h e r m o t h r i x o r e n i i H 1 6 8 2 5 7 8 1 4 6 2 6 9 8 0

C l o s t r i d i u m c e l l u l o l y t i c u m H 1 0 3 9 5 8 6 8 3 3 9 8 0 2

[ D . D a l e v i e t a l . , B i o i n f o r m a t i c s , 2 0 0 8 ]

P r o t e i n d a t a b a s e : N C B I - N R n o n - r e d u n d a n t p r o t e i n s e q u e n c e d a t a b a s e w i t h

e n t r i e s f r o m G e n P e p t , S w i s s p r o t , P I R , P D F , P D B , a n d N C B I R e f S e q



37/52

M a s k i n g a n d F i l t e r i n g

T o s i m u l a t e t h e p r e s e n c e o f U N K N O W N o r g a n i s m s

R e m o v e a l l t h e a l i g n m e n t s w i t h s p e c i e s c o n t a i n e d i n t h e d a t a s e t s

F i l t e r i n g B L A S T p a r a m e t e r s :

F i r s t 5 0 h i t s

S

B

>3 0

E - v a l u e

1 0

6



38/52

C o m p a r i s o n w i t h L C A

A c c u r a c y a n d N u m b e r o f f r a g m e n t s a s s i g n e d ( f o r e a c h r a n k )

R a n k M T R ( # o f f r a g m e n t s ) L C A ( # o f f r a g m e n t s )

k i n g d o m 1 0 0 . 0 0

( 1 6 6 9 4 8 ) 9 9 . 9 9 ( 1 5 5 2 6 3 )

p h y l u m 9 9 . 8 6 ( 1 6 6 9 4 8 ) 9 9 . 9 3 ( 1 5 5 2 5 8 )

c l a s s 9 9 . 7 3 ( 1 6 6 9 3 6 ) 9 9 . 8 1 ( 1 4 1 8 2 9 )

o r d e r 9 7 . 6 7 ( 1 6 6 1 4 8 ) 9 8 . 1 4 ( 1 1 5 7 3 2 )

f a m i l y 9 7 . 6 2 ( 1 6 5 2 3 1 ) 9 8 . 0 4 ( 1 1 0 4 8 8 )

g e n u s 9 7 . 4 2 ( 1 4 0 4 7 6 ) 9 8 . 3 5 ( 1 1 0 1 3 9 )

T a b l e : D a t a n a m e : M 3 , C o v e r a g e 4 X , E v = 0 . 0 0 0 0 0 1 , T o t f r a g m e n t s : 1 , 3 8 5 , 0 2 8

R a n k M T R ( # o f f r a g m e n t s ) L C A ( # o f f r a g m e n t s )

k i n g d o m 9 5 . 0 7

( 8 8 5 3 7 ) 9 4 . 6 6 ( 7 3 1 7 6 )

p h y l u m 9 3 . 2 1

( 8 8 5 3 7 ) 9 2 . 5 7 ( 7 3 1 6 9 )

c l a s s 8 9 . 2 5

( 8 7 6 3 5 ) 8 8 . 9 8 ( 6 0 2 9 4 )

o r d e r 8 9 . 2 4

( 8 5 6 5 7 ) 8 8 . 4 4 ( 5 7 3 7 3 )

f a m i l y 7 7 . 3 5 ( 8 1 3 6 6 ) 8 1 . 8 4 ( 4 8 7 6 0 )

g e n u s 6 1 . 3 6 ( 7 7 3 0 7 ) 7 4 . 6 0 ( 4 0 8 2 3 )

T a b l e : D a t a n a m e : M 2 , C o v e r a g e 1 X , E v = 0 . 0 0 0 0 0 1 , T o t f r a g m e n t s : 2 8 8 , 7 3 0



39/52

P o p u l a t i o n D i s t r i b u t i o n s : M 2 , c o v 0 . 1 X , g e n u s



40/52




R e s u l t s

C o n c l u s i o n



41/52

C o n c l u s i o n s

G o a l : T a x o n o m i c A n n o t a t i o n o f S h o r t M e t a g e n o m i c s

f r a g m e n t s

S i m u l a t e d M e t a g e n o m e s w i t h 5 - 9 g e n o m e s

M T R a s s i g n s m o r e f r a g m e n t s t h a n L C A ( a l l r a n k s )

A c c u r a c y : l e s s a c c u r a t e t h a n L C A e s p . a t l o w r a n k s

T r a d e - o a c c u r a c y / q u a n t i t y ( c o n v e n i e n t )

A c c u r a c y : M T R i s v e r y c l o s e t o L C A , i f t h e r i g h t t a x a a r e i n

t h e d a t a b a s e

P o p u l a t i o n d i s t r i b u t i o n o f M T R t e n d s t o b e b e t t e r



42/52

F u t u r e D e v e l o p m e n t s

T e s t O t h e r f r a g m e n t s l e n g t h s

G o a l s

1 A n a l y z e A L L t h e f r a g m e n t s ( n o t o n l y t h e l t e r e d o n e s )

2 I n c r e a s e A c c u r a c y

P o s s i b l e A l g o r i t h m I m p r o v e m e n t s

I n c l u d e c o m p o s i t i o n - b a s e d m e a s u r e s

F o r G o a l 1 :

C h a n g e B L A S T l t e r i n g

A k i n d o f r e i t e r a t i o n o f t h e m e t h o d

F o r G o a l s 1 & 2 : C h a n g e c l u s t e r i n g m e t h o d

( W e i g h t e d S C P o r T r e e S C P i n s t e a d o f S C P )



43/52

T h a n k Y O U !

Q u e s t i o n s ?

g o r i @ s c i e n c e . r u . n l



44/52

T a b l e : R e a l - l i f e d a t a s e t s : n u m b e r o f r e a d s a s s i g n e d u p t o a r a n k .

R e a l L i f e S a l t e r n C o r a l C h i c k e n

M T R

k i n g d o m 1 5 8 1 2 4 5 2 2 1 1 1 6 5 5

p h y l u m 1 5 7 6 2 3 0 2 7 1 1 1 6 5 0

c l a s s 1 5 3 0 2 1 9 2 0 1 0 9 9 8 6

o r d e r 1 3 1 7 2 1 0 1 9 1 0 8 1 0 0

f a m i l y 1 0 3 5 1 5 5 8 3 1 0 0 6 7 6

g e n u s 9 7 9 1 1 4 2 2 9 4 5 0 7

s p e c i e s 9 3 7 9 5 6 0 8 9 8 1 8

L C A

k i n g d o m 1 2 1 7 2 1 2 8 7 9 3 4 1 6

p h y l u m 1 2 0 8 1 6 5 2 6 9 3 3 9 9

c l a s s 1 0 5 1 1 2 3 0 1 8 7 9 1 7

o r d e r 8 0 7 6 8 4 1 8 7 1 4 6

f a m i l y 6 9 1 5 0 4 5 7 0 3 7 6

g e n u s 6 3 5 4 6 8 5 6 9 6 3 6

s p e c i e s 3 1 1 4 3 4 0 2 9 1 6 0



45/52

F i g u r e : P o p u l a t i o n d i s t r i b u t i o n s ( r a n k g e n u s ) o f c o r a l d a t a s e t b y M T R

( l e f t ) a n d L C A ( r i g h t )



46/52

T a b l e : A c c u r a c y a n d n u m b e r o f a s s i g n e d r e a d s o n M 1 d a t a s e t s

M 1 0 . 1 x 1 x 4 x

M T R

k i n g d o m 1 0 0 . 0 0 ( 5 6 6 9 ) 9 9 . 9 3 ( 5 6 3 4 8 ) 9 9 . 9 3 ( 1 7 3 5 4 1 )

p h y l u m 9 2 . 5 0 ( 5 6 6 9 ) 9 2 . 5 9 ( 5 6 3 2 5 ) 9 3 . 3 9 ( 1 7 3 5 2 1 )

c l a s s 8 4 . 0 4 ( 5 5 5 6 ) 8 5 . 4 4 ( 5 4 3 4 1 ) 8 7 . 1 5 ( 1 6 7 5 4 6 )

o r d e r 6 4 . 9 3 ( 5 3 6 6 ) 6 6 . 2 3 ( 5 3 3 9 5 ) 6 6 . 6 9 ( 1 6 3 8 4 0 )

f a m i l y 6 4 . 8 7 ( 4 9 0 4 ) 6 3 . 6 7 ( 5 0 5 8 7 ) 6 3 . 2 2 ( 1 5 4 1 3 4 )

g e n u s 6 3 . 6 6 ( 4 6 2 8 ) 6 2 . 5 8 ( 4 8 2 4 4 ) 6 0 . 5 0 ( 1 4 4 4 7 5 )

L C A

k i n g d o m 1 0 0 . 0 0 ( 4 1 4 5 ) 9 9 . 9 2 ( 4 2 6 2 0 ) 9 9 . 9 1 ( 1 3 2 1 3 0 )

p h y l u m 9 5 . 0 8 ( 4 1 4 5 ) 9 4 . 8 1 ( 4 2 5 9 3 ) 9 5 . 0 2 ( 1 3 2 0 9 9 )

c l a s s 9 4 . 4 6 ( 3 7 3 9 ) 9 3 . 2 4 ( 3 8 9 7 0 ) 9 3 . 6 0 ( 1 2 1 9 8 0 )

o r d e r 7 5 . 2 9 ( 3 4 9 7 ) 7 4 . 1 8 ( 3 6 8 5 7 ) 7 2 . 4 3 ( 1 1 6 6 3 2 )

f a m i l y 7 1 . 9 4 ( 2 9 6 1 ) 6 9 . 9 4 ( 3 1 9 1 3 ) 6 9 . 0 7 ( 1 0 2 2 3 9 )

g e n u s 7 1 . 0 3 ( 2 6 8 6 ) 6 8 . 3 9 ( 2 9 3 6 0 ) 6 6 . 6 3 ( 9 4 3 4 6 )



47/52


M 2 0 . 1 x 1 x 4 x

M T R

k i n g d o m 9 5 . 2 7 ( 9 0 3 0 ) 9 5 . 0 7 ( 8 8 5 3 7 ) 9 1 . 4 1 ( 1 7 4 5 8 3 )

p h y l u m 9 3 . 8 3 ( 9 0 3 0 ) 9 3 . 2 1 ( 8 8 5 3 7 ) 8 8 . 7 5 ( 1 7 4 5 8 3 )

c l a s s 8 9 . 9 8 ( 9 0 1 2 ) 8 9 . 2 5 ( 8 7 6 3 5 ) 8 6 . 3 2 ( 1 6 8 8 5 4 )

o r d e r 9 0 . 4 4 ( 8 8 2 2 ) 8 9 . 2 4 ( 8 5 6 5 7 ) 8 6 . 1 4 ( 1 6 7 2 2 2 )

f a m i l y 8 0 . 5 6 ( 7 2 6 4 ) 7 7 . 3 5 ( 8 1 3 6 6 ) 7 3 . 0 1 ( 1 5 9 5 9 1 )

g e n u s 6 4 . 4 1 ( 6 4 8 0 ) 6 1 . 3 6 ( 7 7 3 0 7 ) 5 5 . 9 1 ( 1 4 7 1 3 9 )

L C A

k i n g d o m 9 4 . 8 2 ( 7 2 0 5 ) 9 4 . 6 6 ( 7 3 1 7 6 ) 9 0 . 7 6 ( 1 4 3 2 2 6 )

p h y l u m 9 3 . 2 1 ( 7 2 0 5 ) 9 2 . 5 7 ( 7 3 1 6 9 ) 8 7 . 8 0 ( 1 4 3 2 0 6 )

c l a s s 8 9 . 8 2 ( 5 9 4 1 ) 8 8 . 9 8 ( 6 0 2 9 4 ) 8 3 . 5 9 ( 1 1 7 8 8 1 )

o r d e r 8 9 . 9 0 ( 5 6 1 5 ) 8 8 . 4 4 ( 5 7 3 7 3 ) 8 3 . 0 1 ( 1 1 3 1 6 8 )

f a m i l y 8 3 . 7 7 ( 4 7 5 7 ) 8 1 . 8 4 ( 4 8 7 6 0 ) 7 7 . 6 1 ( 1 0 0 9 2 5 )

g e n u s 7 6 . 9 1 ( 3 9 0 7 ) 7 4 . 6 0 ( 4 0 8 2 3 ) 6 9 . 6 8 ( 8 2 8 0 5 )



48/52


M 3 0 . 1 x 1 x 4 x

M T R

k i n g d o m 1 0 0 . 0 0 ( 1 1 7 9 2 ) 9 9 . 9 7 ( 1 1 6 8 6 9 ) 1 0 0 . 0 0 ( 1 6 6 9 4 8 )

p h y l u m 9 9 . 5 8 ( 1 1 7 9 2 ) 9 9 . 4 7 ( 1 1 6 8 6 9 ) 9 9 . 8 6 ( 1 6 6 9 4 8 )

c l a s s 9 6 . 9 7 ( 1 1 7 6 3 ) 9 7 . 0 7 ( 1 1 6 1 3 4 ) 9 9 . 7 3 ( 1 6 6 9 3 6 )

o r d e r 9 1 . 7 9 ( 1 1 6 0 6 ) 9 1 . 7 0 ( 1 1 5 0 3 4 ) 9 7 . 6 7 ( 1 6 6 1 4 8 )

f a m i l y 9 2 . 2 7 ( 1 1 1 1 7 ) 9 1 . 2 5 ( 1 1 1 5 6 0 ) 9 7 . 6 2 ( 1 6 5 2 3 1 )

g e n u s 9 4 . 0 6 ( 1 0 4 1 9 ) 9 2 . 1 9 ( 1 0 1 5 3 3 ) 9 7 . 4 2 ( 1 4 0 4 7 6 )

L C A

k i n g d o m 1 0 0 . 0 0 ( 1 0 3 3 3 ) 9 9 . 9 6 ( 1 0 2 8 2 4 ) 9 9 . 9 9 ( 1 5 5 2 6 3 )

p h y l u m 9 9 . 7 2 ( 1 0 3 3 3 ) 9 9 . 6 9 ( 1 0 2 8 1 3 ) 9 9 . 9 3 ( 1 5 5 2 5 8 )

c l a s s 9 8 . 8 6 ( 9 1 6 2 ) 9 8 . 8 2 ( 9 1 4 4 5 ) 9 9 . 8 1 ( 1 4 1 8 2 9 )

o r d e r 9 6 . 7 4 ( 7 7 8 8 ) 9 6 . 6 2 ( 7 7 8 2 2 ) 9 8 . 1 4 ( 1 1 5 7 3 2 )

f a m i l y 9 6 . 8 7 ( 7 5 4 5 ) 9 6 . 4 2 ( 7 5 6 1 6 ) 9 8 . 0 4 ( 1 1 0 4 8 8 )

g e n u s 9 7 . 6 1 ( 6 7 4 8 ) 9 6 . 0 1 ( 6 8 5 7 3 ) 9 8 . 3 5 ( 1 1 0 1 3 9 )


B e s t H i t ( B H )


49/52

F o r e a c h ( q u e r y ) f r a g m e n t

C o m p a r e w i t h r e f e r e n c e s e q u e n c e s u s i n g B L A S T

A s s i g n t h e f r a g m e n t t o t h e h i t w i t h t h e h i g h e s t S

B


B H : P r o s a n d C o n s


50/52

P r o s :

A s s i g n a l l t h e f r a g m e n t s

A s s i g n t o p r o t e i n s / o r g a n i s m s

C o n s ( s h o r t f r a g m e n t s ) :

L o w a c c u r a c y

U n k n o w n o r g a n i s m n o t r e l i a b l e


L o w e s t C o m m o n A n c e s t o r ( L C A )


51/52

F o r e a c h ( q u e r y ) f r a g m e n t

C o m p a r e w i t h r e f e r e n c e s e q u e n c e s u s i n g B L A S T

F i l t e r t h e h i t l i s t

A s s i g n t h e f r a g m e n t t o t h e l o w e s t c o m m o n t a x o n o f a l l

t h e s e h i t s

L C A

H

1

H

2

H

3

H

4

H

5

H

6

H

7

H

8

H

9

H

1 0

H

1 1

H

1 2


L C A : P r o s a n d C o n s


52/52

P r o s :

H i g h e r a c c u r a c y t h a n B H

A s s i g n t o t a x a i s m o r e r e a l i s t i c ( w i t h s h o r t

f r a g m e n t s )

C o n s :

F e w f r a g m e n t s a t l o w r a n k s

M a n y u n a s s i g n e d f r a g m e n t s

Documents

Annotation of Metagenomes: from Jigsaws to Taxa