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a n o m r e a k c i j o m p o l i m e r a z o m ( P C R ) , d e t e k t i r a n i D N A A n a l y z e r s e r v i s o m ( M a c r o g e n , S e o u l , K o r e j a ) . O d 1 1 i s t r a ~e n i h l o k u s a , p e t d o o s a m j e b i l o p o l i m o r f n o u r a z n i m p o p u l a c i j a m a , d o k s u u p o p u l a c i j i i z P o ~e g e p r o n a e n a d v a n o v a a l e l a , v j e r o j a t n o n a s t a l a s p o n t a n o m m u t a c i j o m . U d e v e t t e s t i r a n i h p o p u l a c i j a p r o n a e n o j e u k u p n o 6 6 r a z l i
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j a H r v a t s k e i S l o v e n i j e d o al o d o m n o g o m i j e aa n j a m e u p o p u l a c i j a m a k a o i r e k o m b i n a c i j a m e u j e d i n k a m a . T o j e p o s l j e d i c a s p o l n o g r a z n o ~a v a n j a , at o s e o d r a z i l o n a u g l a v n o m v e l i k i b r o j p r o n a e n i h S C A R h a p l o t i p o v a k a o i n a n e d o s t a t a k k o r e l c i j e t i p o v a v e g e t a t i v n e k o m p a t i b i l n o s t i ( v c ) i S C A R h a p l o t i p o v a . B a y e z i j a n s k o g r u p i r a n j e j e u k a z a l o n a p o s t o j a n j e t r i j u s k u p i n a k o j e s u i n d i c i r a l e d v a i z v o r a o d a k l e j e C . p a r a s i t i c a u n e ae n a u H r v a t s k u i S l o v e n i j u . J e d n a g r u p a j e d o m i n a n t n a u S l o v e n i j i , j e d n a u H r v a t s k o j , a j e d n a u p o p u l a c i j a m a u z H r v a t s k o - S l o v e n s k u g r a n i c u . R e z u l t a t i u k a z u j u d a j e p o
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A s c o m y c e t e C r y p h o n e c t r i a p a r a s i t i c a ( M u r r i l l ) B a r r . i s a n a g g r e s s i v e p a t h o g e n w h i c h l i v e s a s a p a r a s i t e o n s w e e t c h e s t n u t C a s t a n e a s a t i v a M i l l . a n d c a u s e s s i g n i f i c a n t e c o n o m i c d a m a g e . I n o r d e r t o g a i n b e t t e r i n s i g h t i n p o p u l a t i o n s t r u c t u r e o f f o u r C r o a t i a n a n d f i v e S l o v e n i a n p o p u l a t i o n s o f t h i s f u n g u s , 1 8 0 s a m p l e s w e r e g e n o t y p e d u s i n g 1 1 S C A R ( s e q u e n c e c h a r a c t e r i z e d a m p l i f i e d r e g i o n s ) l o c i . F o r w a r d p r i m e r s w e r e f l u o r e s c e n t l y l a b e l e d a n d a m p l i c o n s , o b t a i n e d w i t h p o l y m e r a s e c h a i n r e a c t i o n ( P C R ) , w e r e d e t e c t e d w i t h D N A A n a l y z e r s e r v i c e ( M a c r o g e n , S e o u l , K o r e a ) . O u t o f 1 1 l o c i t e s t e d , f i v e t o 8 w e r e f o u n d p o l y m o r p h i c i n d i f f e r e n t p o p u l a t i o n s a n d i n p o p u l a t i o n P o ~e g a , t w o n e w a l l e l e s , t h a t p r o b a b l y a r o s e t h r o u g h s p o n t a n e o u s m u t a t i o n s , w e r e d e t e c t e d . I n n i n e t e s t e d p o p u l a t i o n s a t o t a l o f 6 6 d i f f e r e n t S C A R h a p l o t y p e s w e r e f o u n d . T h a t w a s r e f l e c t e d i n h i g h S h a n n o n d i v e r s i t y i n d e x , r a g i n g f r o m 1 , 5 4 i n p o p u l a t i o n O s t r o v i c a t o 2 , 9 7 i n p o p u l a t i o n H r v a t s k a K o s t a j n i c a . E v e n n e s s w a s a l s o v e r y h i g h , f r o m 0 , 7 9 t o 0 , 9 8 . T w e l v e o u t o f 2 8 p a i r s o f l o c i w e r e f o u n d i n l i n k a g e d i s e q u i l i b r i u m . I n d e x o f m u l t i l o c u s l i n k a g e d i s e q u i l i b r i u m ( r d ) w a s a l s o p o i n t i n g t o p r e d o m i n a n t l y s e x u a l r e p r o d u c t i o n a n d m a n y r e c o m b i n a t i o n s o f C . p a r a s i t i c a i n t e s t e d p o p u l a t i o n s ; r d e s t i m a t e r a n g e d f r o m 0 , 0 2 4 t o 0 , 2 4 1 . N o c l e a r c o r r e l a t i o n b e t w e e n g e n e t i c d i s t a n c e o f s t u d i e d p o p u l a t i o n s a n d g e o g r a p h i c a l d i s t a n c e b e t w e e n l o c a t i o n s w h e r e f u n g a l i s o l a t e s w e r e c o l l e c t e d w a s o b s e r v e d . T h e r e f o r e w e t h i n k t h a t f r o m i n i t i a l i n t r o d u c t i o n o f t h i s p a t h o g e n i c f u n g u s i n C r o a t i a a n d S l o v e n i a , m a n y m i x i n g o f p o p u l a t i o n s a s w e l l a s r e c o m b i n a t i o n s b e t w e e n i n d i v i d u a l s h a s o c c u r r e d . T h i s i s a c o n s e q u e n c e o f s e x u a l r e p r o d u c t i o n w h i c h i s r e s p o n s i b l e f o r l a r g e n u m b e r o f S C A R h a p l o t y p e s o b s e r v e d a n d l a c k o f c o r r e l a t i o n b e t w e e n v e g e t a t i v e c o m p a t i b i l i t y ( v c ) t y p e s a n d S C A R h a p l o t y p e s . B a y e s i a n c l u s t e r i n g i n d i c a t e d e x i s t a n c e o f t h r e e c l u s t e r s w h i c h s u g g e s t e d t w o s e p a r a t e s o u r c e s o f C . p a r a s i t i c a i n t r o d u c t i o n i n C r o a t i a a n d S l o v e n i a . O n e c l u s t e r i s d o m i n a n t i n S l o v e n i a , o n e i n C r o a t i a , a n d j e t a n o t h e r a l o n g C r o a t i a n - S l o v e n i a n b o r d e r . R e s u l t s r e v e a l t h a t w e s t e r n E u r o p e ( I t a l y ) i s t h e f i r s t s o u r c e o f i n f e c t i o n , w h i l e t h e s e c o n d i n t r o d u c t i o n c a m e s u b s e q u e n t l y f r o m s o u t h e a s t e r n E u r o p e a n d c o n t r i b u t e d t o d i v e r s i t y o f o u r C . p a r a s i t i c a p o p u l a t i o n s .
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