In vivo analysis of the cellular interactions during taste sensory organ assembly in zebrafish Marina Soulika
To cite this version:
Marina Soulika. In vivo analysis of the cellular interactions during taste sensory organ assembly in zebrafish. Development Biology. Université Pierre et Marie Curie - Paris VI, 2014. English. NNT: 2014PA066633. tel-01149429
HAL Id: tel-01149429 https://tel.archives-ouvertes.fr/tel-01149429 Submitted on 7 May 2015
HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés.
Université Pierre et Marie Curie
ED515 - Complexité du vivant Laboratoire INSERM 1024-CNRS 8197 / Du développement au comportement
In vivo analysis of the cellular interactions during taste sensory organ assembly in zebrafish
Par Marina Soulika
Thèse de doctorat de Biologie du développement
Dirigée par Frédéric Rosa
Présentée et soutenue publiquement le 16 décembre 2014
Devant un jury composé de: Prof. Muriel Umbhauer Université Pierre et Marie Curie Président Prof. Alan Carleton Univ. de Genève, Neuroscience Center Rapporteur Dr. Elise Cau Univ. de Toulouse 3, CBD Rapporteur Dr. Marcel Tawk INSERM - Institut Baulieu, Examinateur Dr. Frédéric Rosa IBENS Examinateur Dr. Marika Kapsimali (encadrante) IBENS Examinateur
! "!
Στον πατέρα μου Μιχάλη για την αφοσίωση και την υπομονή του
Στη μητέρα μου Μαρία για τη δύναμη και την επιμονή της
Στα αδέρφια μου Δημήτρη, Αθηνά, Κυριακή, Άντα, Ντύλαν
για τη συνεχή τους φροντίδα και στήριξη
Στα ανίψια μου Μιχάλη, Μαρία, Αλέξανδρο και Μιχάλη
για την ανιδιοτέλεια και το θάρρος τους
2
«Το φάρμακό μας είναι η τροφή μας»
Περί Αισθήσεως και Αισθητών
Αριστοτέλης
«Let thy food be thy medicine»
On sense and the sensible
Aristotle
A rose by any other name would scent as sweet
William Shakespeare
Sequi naturam
3
I would like to express my gratidude to:
Marika Kapsimali for entrusting to this project, for the exceptional cooperation, constant support and everyday availability during the PhD.
Frederic Rosa for the warm welcome in his team and his support.
The members of the jury, Muriel Umbhauer, Alan Carleton, Elise Cau and Marcel Tawk for their participation, consideration and evaluation of my work.
Sylvie Schneider-Maunoury, Michel Gho and Jean-Pierre Levraud for their guidance, advice and support during my thesis.
Sylvain Ernest, Aline Bonnet, Florence Giger, Nicolas David, Raquel de Amaro Lourenço, Francois-Xavier Dutriex, Julien Dumortier, Brigitte Boyer, Guillaume Lambert, Benjamin Mathieu, Astou Tangara, Patrick Charnay, Pascale Gilardi, Piotr Topilko, Alexandre Jourdon, Fanny Coulpier, Yassine Bouchoucha, Elodie Thierion, Patricia Gongal, Renata Santos, Carole Desmarquet, Aurelie Gresset, Firas Bouallague, Aurelie Jubeau, Graziela Matesic, Carmen-Lucia Le Moal and Rouben Nagalingum for their for their patience, assistance, caring, encouragement, technical advice and good humor over the past three years.
Virginie Panayotopoulos and family, Emma Intiba and familly, Sideri and Evangelia Araka, Patera Panagioti Xenitelli, Maria Fesatidou, Viki Ntinoka, Maria Stampoulou, Kyriaki Saltidou, Odyssea Patouna, Kosta Poala with my deepest appreciation to whose love and support helped to go through this long road.
My family for their love…for everything
4 CONTENTS
PROLEGOMENON 9
I) INTRODUCTION 13
I.1) THE ORGANISATION, ANATOMY AND FUNCTION OF THE MAMMALIAN
TASTE SYSTEM 15
I.1.1) The mammalian taste buds 15
I.1.1.1) General morphology of the mouse taste bud 15
I.1.1.2) The main taste bud cell types and their function 16
I.1.1.2a) Type I (Support Cells) 16
I.1.1.2b) Type II (Taste receptor cells) 17
I.1.1.2bi) Taste specific receptors 18
I.1.1.2bii) Taste transduction in Type II cells 20
I.1.1.2biii) ATP, the Type II cell transmitter 20
I.1.1.2b.iv) L-glutamate, the ‘unknown’ in Type II
signal taste bud transmission 21
I.1.1.2c) Type III cells (Presynaptic) 24
THE MAMMALIAN TASTE BUDS IN SUMMARY 26
I.1.2) From the periphery to the cns: how is taste information
transmitted and encoded in mammals 27
I.1.2.1) Mammalian taste sensory ganglia 27
I.1.2.2) Mammalian brain taste-related nuclei 28
I.2) ANATOMY AND FUNCTION OF THE TELEOST FISH TASTE SYSTEM 31
I.2.1) Teleost fish taste buds 31
I.2.2) Taste innervation and brain nuclei in teleost fish 33
I.3) DEVELOPMENT OF TASTE SENSORY ORGANS 36
I.3.1) The embryonic origin of taste bud cells 36
5 I.3.2) Taste bud placode : the first morphological evidence
of a mammalian taste bud 37
I.3.3) The role of mesenchyme and innervation during taste bud development 37
I.3.4) Taste bud progenitor cells 38
I.3.5) Molecular signaling during taste bud development 40
I.3.5.1) Wnt/beta-catenin: key signal for taste placode formation 40
I.3.5.2) Taste placode patterning is regulated by BMP and Follistatin 41
I.3.5.3) Shh and taste placode formation 41
I.3.6) Key signals for the differentiation of taste bud cell types 43
I.3.6.1) FGF pathway for the differentiation of taste receptor cells 43
I.3.6.2) Notch signaling pathway during taste bud cell differentiation 44
I.3.6.3) Mir200 promotes taste bud cell formation 45
I.4) Project Goals 46
II) MATERIALS AND METHODS 48
II.1) Zebrafish 49
II.2) Phenotypic analysis 49
II.2.1) In situ hybridization 49
II.2.2) Immunohistochemistry 50
II.3) Fixed tissue image acquisition and time-lapse imaging 50
II.4) Time-Lapse Statistical analysis 51
II.5) Two-photon laser ablation 51
III) RESULTS 53
III.1) The fgf8a.enhancer:gfp enhancer is expressed in all developing taste
bud cell types 54
6 III.2) tg(fgf8a.enhancer:gfp) expressing cells form taste buds and generate 5-HT cells 54
III.3) tg(fgf8a.enhancer:gfp) expressing cells are motile, join and quit developing
taste bud organs 55
III.4) The 5-HT cell is required for the maintenance of tg(fgf8a.enhancer:gfp)
expressing cells into taste bud organs 56
Figures 58
IV) DISCUSSION 76
IV.1) tg(fgf8a.enhancer:gfp); tg(tph1b:mCherry) : an appropriate tool
to analyse taste bud development in vivo 77
IV.2) What is the functional significance of cell motility during taste bud
organ formation? 78
IV.2.1) Are the forming taste bud organs patterned during early
development? 78
IV.2.2) Is the cell motility negatively correlated with the functional activity
of the taste bud cell? 79
IV.2.3) What is the molecular mechanism underlying cell contact and adhesion
during taste bud formation? 80
IV.2.4) Is cell motility a common phenomenon during sensory organ
formation? 81
IV.3) ANNEXE 82
IV.3.1) Regeneration in adult taste buds 82
IV.3.2) The Drosophila taste system 83
IV.3.2.1) Sweet bitter symphony in flies 85
7 IV.1.3.2.2) Bitter transduction 86
IV.1.3.2.3) Sweet transduction 86
IV.1.3.2.3) Umami transduction 87
IV.1.3.2.3) Salty transduction 87
V) BIBLIOGRAPHY 88
8
hrough the centuries, numerous definitions have been given for taste from a physiological, sociological, chemical and gastronomic point of view. A physiologist would describe taste as one of the five senses, which defines the ability to distinguish the quality of a substance placed in the oral cavity, including the upper surface of the tongue and the epiglottis. Taste is produced after stimulation of the taste receptors located on taste sensory organs by flavoured substances. Along with olfaction and trigeminal nerve stimulation (registering texture, pain, and temperature), taste determines and interprets food quality.
A sociologist would say that taste is a personal and cultural pattern of choice and individual preference in food. According to Littré dictionary taste comes from Latin “gustus”, meaning « act of tasting, degustation; savour ». Gustus may also derive from Greek and the Greek term of Sanskrit “gush” meaning « like, good». Social and cultural phenomena concerning taste are closely associated to environmental resources and social relations between people; therefore culture and food are closely related.
A romantic would admit that the sense of taste is capable of the development of "an extreme delicacy” of the palate, which allows the appreciation, in tasting of a full range of flavors far beyond the register of basic sensation. Such sentiments are repeated and embroidered many times in the rich literature of gastronomy that blossomed in the ensuing decades [1, 2].
According to the great gastronome Brillat-Savarin, taste is a chemical process that occurs in a liquid phase where sapid molecules dissolve in any fluid. These molecules are consequently absorbed by the taste-related organs, the taste buds that coat the interior of the taste apparatus. For instance, pure water itself, as a neutral liquid, does not cause any gustatory effect on the taste buds but once a grain of salt or some drops of vinegar are diluted in it the result is really appreciated [3].
Gastronomes use the term "taste" to refer to the multi-sensory experience of the flavour and texture of food and drink. Taste however does not function alone outside the laboratory as it is strongly related to olfaction and vision. Texture and colour of food are among the basic reasons for feeding behaviour and curiously, who would choose to eat a blue apple?
9 Contrasting though the olfactory system of an average person that is capable to recognize approximately 50 different smells, of a chemist’s that can distinguish more than 500 and a perfumer’s up to 5000 scents, their taste system is limited to only 5 principal groups of savouriness: sweet, umami, bitter, sour and salty.
Sweet: As soon as we are born, we are attracted to sweet nutrients (initially to the breast milk in which 6,89% is sugars) indicating us the importance of this energy (and not only) rich nutrient. When sugars are consumed they transform into glucose in the body. Glucose is then used for immediate energy or stored in the liver and muscles in order to be used for energy at a later time, depending on body needs. Simple sugars, also known as simple carbohydrates, get digested quickly and provoke a fast burst of energy. The sweetness of sugars in fact represents a neuronal burst in the brain related to the pleasant access of these caloric-rich sources of metabolic energy.
Umami: A Japanese word is used to describe the internationally recognised taste produced by L-amino acids and more precisely L-glutamate and aspartate in human. Umami in general can be related to the taste of meat and protein-rich nutrients.
Bitter: While sweet and umami function in order to satisfy our papillae by determining energy rich-nutrients, bitter represents the repugnant but concurrently protective role to eliminate every noxious substance that could have fatal results, simply by aversion. Bitter can be bearable when weak, as indicated by the fact that many people enjoy a cup of coffee but can be really aversive when strong. In contrast to the three parsimonious taste receptors for sweet and umami, nature was quite generous for bitter detecting receptors, as they are quite numerous and reaches up to 30.
Sour: Although small amounts of low concentrated acid substances are quite pleasant but also useful e.g lemon juice (rich in vitamin C), the importance of sour taste lays on the protection of the organism by low-pH compounds which tend to be unpleasant and sometimes poisonous for our body. Sour is also necessary to regulate multiple functions of our body fluids especially within the brain. Although the transduction channels and/or
10 receptors involved in sour detection has been quite enigmatic, there are still some indications about molecules involved in the process.
Salty: The source of life according to the ancient Greek *αλάτι, * ἅλς (háls) Proto-
Indo-European *séh₂l, *séh₂l. Salty taste governs intake of sodium and other salts, essential for maintaining the body’s water balance and blood circulation. Salts in our body function as catalysts.
Nevertheless, it could be due to the importance, albeit neglected in function sense of taste that eating is a pleasurable experience. Furthermore, the human body homeostasis highly depends on this sense, as the ability to taste and smell food properly is of vital meaning. Apart for its nutritional aspect, taste is related with others emotions. People that lose their ability to taste properly tend to lose weight, become depressed or enrich their food with strong tastants such as salt or acid to obtain a taste sensation. This incapacity may lead to poor nutritional status, impaired immunity, and/or aggravation of several diseases. So what the world would have been without taste?
11
12 !
INTRODUCTION
! "$! INTRODUCTION
.4 '$5%'")+6#.&6$."#'"9'$5%'$6&$%'&8&$%,''
<.97+S!1,+!16!73+!6=T+!9+,9+9!2/.59!73+!-1/+!16!.!9+>8-=75!48.-;!61-!73+!1-4.,=90! :5!-+48/.7=,4!18-!6++;=,4!:+3.T=18-( '7 +T./8.7+9 73+!,87-=7=189!+/+0+,79!16!611; 73.7 O=//!9.7=.7+!73+!:1;5!O=73!+,+-45!.,;!2-+T+,79!=,4+97=1,!16!71W=>!98:97.,>+9!73.7!>18/;! :+!217+,7=.//5!3.-068/(!<3+!9+,9+!16!7.97+!=9!=,7+-2-+7+;!:5!73+!7.97+!9597+0S!O3=>3!=9! >10219+;!16!73+!7.97+!9+,91-5!1-4.,9 >.//+;!7.97+!:8;9S!73+!7.97+!9+,91-5!4.,4/=.!.,;! 7.97+H92+>=6=>! :-.=,! ,8>/+=! @N=4(! 'H")(! <3+! 7.97+! 9597+0! 3.9! :++,! +W7+,9=T+/5! 978;=+;! =,! g.O+;! T+-7+:-.7+9S! @+92+>=.//5! =,! 0.00./9! :87! ./91! =,! 6=93! .,;! .023=:=.)! .,;! L12+2.;'7*(! ! <1 =,7-1;8>+ 73+! 2-+9+,7! -+9+.->3! O1-L 1,! 73+! 7.97+! :8;9S! 9+/+>7+;! L+5! +/+0+,79!16!=)!73+!1-4.,=9.7=1,!.,;!==)!;+T+/120+,7!16!73+!7.97+!9597+0!=,!0189+!.,;! Q+:-.6=93!.-+!;+9>-=:+;!=,!B3.27+-9!'("S!'(#!.,;!'($!-+92+>7=T+/5(!
!
!
9@TEH<' .]2(! n-4.,=9.7=1,! 16! 73+! 380.,! 7.97+! 9597+0(! <3+! ;-.O=,4! 931O9! 73+! -+/.7=1,93=2! :+7O++,! 73+! 7.97+!9+,91-5!1-4.,9!=,!73+!1-./!>.T=75!.,;!73+!73-++!7.97+!9+,91-5!4.,4/=.!@o''S!'pS!p)!.,;!7.97+H-+/.7+;! ,8>/+=!=,!73+!:-.=,!`%a(!
! "%! INTRODUCTION
' .42/'6#6$",8'6#0'91#!$."#'"9'$5%',6,,6*.6#'$6&$%'&8&$%,'
.4242/'$;<'=>==>?@>A'B>CB<'DEFC' '
.424242/'+
! ',!0.00./9S!73+!7.97+!:8;9!.-+!;=97-=:87+;!O=73=,!73+!1-123.-5,4+./!+2=73+/=80! =,!.!2.77+-,+;!0.,,+-S!./1,4!73+!.,7+-1H2197+-=1-!.W=9!16!73+!71,48+S!2./.7+S!+2=4/177=9S! 23.-5,W! .,;! /.-5,W(! A+T+-./! 38,;-+;9! 16! 7.97+! :8;9! .-+! 618,;! =,! 73+! 0189+! 1-123.-5,4+./!>.T=75(!m.>3!1,+!16!73+0!=9!.,!1,=1,H93.2+;!97-8>78-+!>1,7.=,=,4!&EH"EE! >+//9! `&a! 73.7! 2199+99! 0=>-1T=//=! 1,! 73+=-! .2=>./! +,;! .,;! >1,7.>7! 7.97+! 9+,91-5! 4.,4/=.! 2-1>+99+9!1,!73+=-!:.9./!2.-7(!<3+!0=>-1T=//=!2-1g+>7!=,71!73+!,*+,-!.21-S!.!90.//!12+,=,4! 1,! 73+! 98-6.>+! 16! 73+! 71,48+! 2+-0=77=,4! 73+! 7.97+! >+//9! 71! :+! +W219+;! 71! .,;! 4+7! =,! >1,7.>7!O=73!73+!>1,7+,79!16!73+!01873!`?S!Ca(!
n,! 73+! 98-6.>+! 16! 73+! 0.00./=.,! 71,48+S! 73+! 7.97+! :8;9! -+9=;+! =,! 2.2=//.+S! q21>L+79c!61-0+;!6-10!73+!+2=73+/=./!>+//9!73.7!98--18,;!73+!7.97+!:8;9(!<3-++!;=66+-+,7! 752+9!16!/=,48./!2.2=//.+!.-+!;=97=,48=93+;!@N=4(!'H#)S!.>>1-;=,4!71!73+!71,48+!-+4=1,K!!
?!W53@'021
?W27'*,-X!73+5!3.T+!.,!+/1,4.7+;!61/;!-+2-+9+,7=,4!.!/+.6!1,!73+!+;4+!.,;!73+5! .-+! >/897+-+;! =,71! 7O1! 4-1829! 1,+! 1,! +.>3! 9=;+! 16! 73+! 2197+-=1-! /=,48./! +2=73+/=80(! <3+-+!.-+!.:187!%H&!=,!0.00./9!.,;!73+=-!9=Q+!.,;!93.2+!T.-=+9(!!
!HY'1/5<=*77*,-X!73+5!.-+!=,T+-7+;!oH93.2+;!97-8>78-+9S!9=78.7+;!1,!73+!2197+-=1-! 98-6.>+!16!73+!71,48+(!<3+-+!=9!.!9=,4/+!1,+!=,!0=>+!:87!DH"E!>=->80T.//.7+!=,!380.,(!
',! .;;=7=1,S! 6=/=61-0! 2.2=//.+! .-+! /1>.7+;! 1,! 73+! 71,48+]! 73+9+! .-+! 73+! 0197! ,80+-189S!:87!,17!.991>=.7+;!O=73!7.97+!:8;9(!<3+5!3.T+!.!0+>3.,=>./!.,;!2-17+>7=T+! 68,>7=1,!61-!73+!71,48+!9+-T=,4!.9!.!;+6+,9=T+!>1.7!>102/+0+,7+;!:5!73+!.,7=:.>7+-=./! .>7=1,!16!>1021,+,79!16!73+!9./=T. `Ra(
'
'
! "&! INTRODUCTION
' .42424G/'$;<'=>@A'B>CB<'DEF'LAF'B;<@H'OEALB@IA'
<1!8,;+-97.,;!31O!7.97+!:8;9!68,>7=1,S!O+!9+7!71!;+/=,+.7+!73+!>+//!752+9!73.7! >10219+! 73+! 7.97+! :8;9! .,;! 31O! 73+9+! >+//9! 7-.,9;8>+! 73+! >3+0=>./! 9=4,./9! 821,! 97=08/.7=1,!6-10!7.97.,79(!U!68,>7=1,./!7.97+!:8;!=9!>1,9=97+;!16!73-++!01-231/14=>.//5! ;=97=,>7! >+//! 752+9! 73.7! ;+7+>7! 7.97+! 97=08/=]! <52+! '! >+//9S! ./91! >.//+;! 98221-7S! 73+5! .-+! 4/=./H/=L+!>+//9S <52+ ''!>+//9S!1-!7.97+!-+>+271-!>+//9S 73+5 ;+7+>7!:=77+-S!9O++7!.,;!80.0=! .,;!<52+!'''!>+//9!1-!2-+95,.27=>!>+//9S!O3=>3!9+,9+!918-!.,;!>.-:1,.7+;!98:97.,>+9(!!
.42424G>/'$KJ<'.'b&EJJIHB'!
<3+!<52+!'!@98221-7)!>+//9!-+2-+9+,7!73+!4-+.7+97!,80:+-!16!>+//9 @./0197!3./6!16! .//!7.97+!:8;!>+//9)!=,!0.00./=.,!7.97+!:8;9(!<3+5!.-+!+/1,4.7+;!>+//9!O=73!>5712/.90=>! 2-17-89=1,9!.,;!+W7+,9=T+!/.0+//.-!2-1>+99+9!.//1O=,4!O-.22=,4!.-18,;!.,;!2-17+>7=,4! 173+-!>+//!752+9(!<52+!'!>+//9!;1!,17!2199+99!T1/7.4+H4.7+;!>./>=80!>3.,,+/9!.,;!;1!,17! 61-0!95,.29+9(!<3+5!+W2-+99!7-.,921-7+-9!61-!4/87.0.7+!@*XUA.--=+-!$U)S!.9!O+//!.9!P
<3+!68,>7=1,!16!73+!98221-7!>+//9!-+0.=,9!211-/5!8,;+-9711;(!'7!=9!:+/=+T+;!73.7! 73+5! ;+7+>7! P.B/! :87! ./91! 173+-! 9./79! 73.7! 2-1T1L+! ,17! ,+>+99.-=/5! 9./75! :87! :=77+-S! 0+7.//=>!1-!.97-=,4+,7!7.97+(!A./79!87=/=Q+!.2=>.//5!/1>.7+;!=1,!>3.,,+/9!61-!7-.,9;8>7=1,! @6=4('H?)(!U!98:9+7!16!<52+!'!>+//9!=,!73+!.,7+-=1-H71,48+!7.97+!:8;9!+W2-+99!73+!.0=/1-=;+H 9+,9=7=T+S! +2=73+/=./! 91;=80! >3.,,+/! @mP.B)! `""S! "#a! .,;! 0197! 7.97+! :8;9! >1,7.=,! 73+! 73-++!16!=79!98:8,=79!sS!tS!.,;!u(!<3+!mP.B!98:8,=79!.-+!+W2-+99+;!=,!.!9=0=/.-!0.,,+-!1,! 73+! .,7+-=1-! .,;! 2197+-=1-! /=,48./! +2=73+/=80! =,! 0.00./9! :87! =,! 4-+.7+-! +W7+,7! =,! 68,4=61-0!73.,!=,!>=->80T.//.7+!2.2=//.+(!sHmP.B!./1,+!>.,!2-1;8>+!.0=/1-=;+H9+,9=7=T+! 91;=80!>8--+,79S! .,;! >1H+W2-+99=1,! O=73! tHmP.B! .,;! uHmP.B! +,3.,>+9! 73+9+! >8--+,79!
! "?! INTRODUCTION
@6=4('HC)(!r+92=7+!73+!+W2-+99=1,!16!mP.B!=,!<52+!'!>+//9!.,;!=79!>.2.>=75!71!;+7+>7!91;=80S! .!>/+.-!/=,L!:+7O++,!9./75!7.97+!;+7+>7=1,!.,;!<52+!'!>+//9!3.9!,17!:++,!+97.:/=93+;!5+7! `"$a( U,173+-! 217+,7=.//5! 7.97+H-+/.7+;! 01/+>8/+! >18/;! :+! 73+ -+,./! 187+-! 0+;8//.-5! 217.99=80!>3.,,+/!@vnGw)S!O3=>3!=9!.,!U
!
9@TEH<'.]G4!@U)(!r=97-=:87=1,!16!7.97+!:8;9!1,!73+!0.00./=.,!71,48+!>1,>+,7-.7+;!O=73=,!90.//!:87!T=9=:/+! 2-1g+>7=1,9! >.//+;! .*.'77*-#! @M)(! F.2=//.+! .-+! 93.2+;! /=L+! -=;4+9S! 2=02/+9S! 1-! 0893-1109! +.>3! 16! .! 6+O! 0=//=0+7-+9!=,!;=.0+7+-!>1,9=97=,4!16!0=>-1T=//=S!.!7.97+!21-+!.,;!;=97=,>7!>+//!752+9(!@B)(!m.>3!2.2=//.!=,!78-,! 3.9!,80+-189!,*+,-!&54+(!m.>3!7.97+!:8;!=9!=,,+-T.7+;!:5!.66+-+,7!,+8-1,9!`Ra(! ! ' .42424GD/'$KJ<'..'b$>CB<'H <52+! ''! @7.97+! -+>+271-)! >+//9! 3.T+! /.-4+! 1T1=;! ,8>/+=! .,;! +/+>7-1,x/8>+,7! >5712/.90(! <3+5! +W2-+99! *! 2-17+=,H>182/+;! -+>+271-9! @*FBv9)! O=73! 9+T+,! 7-.,90+0:-.,+!;10.=,9!73.7!.-+!/1>.7+;!1,!73+=-!2/.90.!0+0:-.,+(!<3+5!3.T+!:++,! -+,.0+;! 7.97+! -+>+271-! >+//9! ;8+! 71! 73+! >.2.>=75! 16! 73+9+! *FBv9! 71! :=,;! /=4.,;9! 73.7! 2-1T1L+! 9O++7S! 80.0=! .,;! :=77+-! -+921,9+9(! h=971-=>.//5S! 73+! 6=-97! /1>89! =;+,7=6=+;! =,! 0=>+!73.7!;+7+-0=,+;!73+!;+7+>7=1,!16!9O++7!98:97.,>+9!O.9!I*//;*1'3!ZI*/(!`"?H#Ea(! I*/! +,>1;+9! 73+! *FBv! <.9"v$! .,;! =7! O.9! h11,! .,;! >1//+.48+9! 73.7! =;+,7=6=+;! 7O1! ;=97=,>7!6.0=/=+9!16!7.97+!*FBv9S!73+!7.97+!-+>+271-!752+!"!@<.9"v9!1-!<"v9)!;+;=>.7+;!71! ! "C! INTRODUCTION 9O++7!.,;!80.0=!-+921,9+9!.,;!73+!7.97+!-+>+271-!752+!#!@<.9#v9!1-!<#v9)!73.7!:=,;! :=77+-!>10218,;9!`#"a(! "#$#$#%&'( )*+,-!+.-/'0'/!1-/-.,21+ <3+! <.9"v9! 61-0! 3+7+-1;=0+-9S! <.9"v"J<.9"v$! -+921,;=,4! 71! .0=,1! .>=;9! @80.0=! 7.97+)! .,;! <.9"v#J<.9"v$! -+921,;=,4! 71! 9O++7(! <.9"v"J<.9"v#! -+>+271-9! -+921,;!71!.0=,1!.>=;9!98>3!.9!4/87.0.7+S!.92.-7.7+ .,; 4/8>=,+(!X=4.,;9!92+>=6=>!61-! <.9"v#J<.9"v$! -+>+271-9! .-+! 984.-9! @98>-19+S! 6-8>719+S! 4/8>19+S! /.>719+)! .,;! .-7=6=>=./! 9O++7+,+-9! @9.>>3.-=,S! 98>-./19+S! .92.-7.0+)(! ',! >1,7-.97S! 73+! <.#v9! ;1! ,17! ;=0+-=Q+! .,;! -+921,;! 71! :=77+-! 7.97.,79 98>3! .9! ;+,.71,=80S! >.66+=,+S! V8=,=,+S! 23+,573=18-+.! .,;! 3801/1,+(! <.9"v9! .-+! -+>+271-9! O=73! /1,4! P! 7+-0=,=! >1,7.=,=,4! 4/1:8/.-! +W7-.>+//8/.-!/=4.,;H:=,;=,4!;10.=,!O3+-+.9!<.9#v9!3.T+!931-7!+W7-.>+//8/.-!P!7+-0=,=! @6=4('HC)(!! ',.>7=T.7=1,!16!73+!,*+$1%!.,;!,*+$1[!4+,+9!=,!0=>+!-+T+./+;!73.7!73+9+!-+>+271-9! .-+! -+V8=-+;! =,! 9O++7! 7.97+! ;+7+>7=1,(! ,*+$1%?\?! 1-! ,*+$1[?\?! 0=>+! 2-+9+,7! O+.L! 2-+6+-+,>+! 71! 9O++7! 7.97.,79! 731843! ,*+$1%?\?],*+$1[?\?! +/=0=,.7+! >102/+7+/5! 9O++7! 7.97.,79! -+921,9+(! !A=0=/.-/5! ,*+$1$?\?! 21! ,*+$1[?\?! L,1>L187! 0=>+! 6.=/! >102/+7+/5! 71! -+>14,=9+!80.0=!7.97.,79!`##S!#$a(!G=>+!,8//!61-!92+>=6=>!,*+%1+!/19+!73+=-!9+,9=7=T=75!71! :=77+-! >10218,;9(! U;;=7=1,.//5S! 4+,+7=>.//5! +,4=,++-+;! 0=>+! +W2-+99=,4! 92+>=6=>.//5! 380.,! -+>+271-9! 61-! .! 98:9+7! 16! :=77+-! 98:97.,>+9! 98>3! .9! 9./=>=,! .,;! 23+,5/73=1>.-:.0=;+! 931O+;! .! -+28/9=T+! :+3.T=18-! 61-! 73+9+! 7O1! 98:97.,>+9(! N8-73+-01-+S!0=>+!+W2-+99=,4!H12=1=;!-+>+271-!@vUAAX]-+>+271-!.>7=T.7+;!91/+/5!:5!.! 95,73+7=>! /=4.,;) =,! <"v#y<"v$! +W2-+99=,4! >+//9! :+>10+! 92+>=6=>.//5! .77-.>7+;! 71! .! H 12=1=;!.,7.41,=97!92=-.;1/=,+S!.!,1-0.//5!7.97+/+99!>10218,;S!;+01,97-.7=,4!73.7!73+! .>7=T.7=1,! 16! 9O++7x-+>+271-! +W2-+99=,4! >+//9! @.,;! ,17! 73+! -+>+271-! =79+/6)! -+98/79! =,! 9O++7!2+->+27=1,(!A=0=/.-/5S!O3+,!73+!vUAAX!-+>+271-!O.9!+W2-+99+;!=,!:=77+-!+W2-+99=,4! >+//9!73+!4+,+7=>.//5!+,4=,++-+;!0=>+!931O+;!.T+-9=1,!61-!73+!7.97+/+99!92=-.;1/=,+!`#$S! #%a(! !! M87! 31O! 1,/5! 73+9+! 7O1! 6.0=/=+9! 16! -+>+271-9! @<.9"v9! .,;! <.9#v9)! .-+! .:/+! 71! ;=97=,48=93!73+!O=;+!T.-=+75!16!7.97.,79!+W219+;!=,!18-!+T+-5;.5!/=6+b!<3+!;=66+-+,>+9!=,! ! "D! INTRODUCTION 73+!9+V8+,>+9!16!<.9"v!.,;!<.9#v!.01,4!92+>=+9!-+6/+>7!73+=-!9+,9=7=T=75!.,;!9+/+>7=T=75! 16!9O++7!1-!:=77+-!>10218,;9!-+92+>7=T+/5!@6=4('H$)(!N1-!=,97.,>+S!.,!+/+4.,7!978;5!89+;! >3=0+-=>!380.,H-1;+,7!<.9"v#!.,;!<.9"v$!-+>+271-9!71!0.2 -+4=1,9 16 73+ <"v#J<"v$ ;=0+-! 73.7! .-+! -+V8=-+;! 61-! -+921,9=T+,+99! 71! 910+! 16! 73+9+! 9O++7+,+-9(! <3+9+! +W2+-=0+,79S! /.-4+/5! 2+-61-0+;! 89=,4! 3+7+-1/14189! >+//9S! -+T+./+;! 73.7! 73+! +W7-.>+//8/.- ;10.=, 16 380., <.9"v# =9 -+V8=-+; 61- -+>+271-! -+921,9+9 71 .92.-7.0+! .,;! ,+17.0+! `#&aS! 73+! >597+=,+H-=>3! ;10.=,! @Bvr)! 16! 380.,! <.9"v$! =9! -+V8=-+;! 61-! -+921,9+9! 71! 73+! 2-17+=,! :-.QQ+=,! `#?aS! .,;! 73+! 7-.,90+0:-.,+!;10.=,! @ ! ! 9@TEH<'.]3(!F35/14+,+7=>!-+/.7=1,93=29!16!T+-7+:-.7+!=,7.>7!<"v!4+,+9(!<3+!.--1O!21=,79!71!73+!7-++!-117S! O3=>3! =9! ;+7+-0=,+;! :5! 89=,4! T+-7+:-.7+! o#v! 4+,+9! .9! 1874-1829! `#Da(! <3+! <"v"! y! <"v$! 3+7+-10+-=>! -+>+271-! >.,! ;+7+>7! XH.0=,1! .>=;9! .,;! GA*S! O3=>3! =9! 73+! 7.97+! 16! 80.0=S! O3+-+.9! 73+! <"v#! y! <"v$! 3+7+-10+-=>!-+>+271-!>.,!-+921,;!:-1.;/5!71!9O++7!7.97.,79!`#Ra(! ! "R! INTRODUCTION "#$#$#%&''(!)*+,-!,1*3+45/,'23!'3!)6.-!""!/-77+! U01,4! 73+! *! 2-17+=,9S! 4897;8>=,! =9! >182/+;! 71! 73+! <.9v! =,! <52+! ''! >+//9! .,;! =,T1/T+; =,!-+921,9+9!71!9O++7S!:=77+-S!.,;!80.0=!>10218,;9!`$EH$$a(!',.>7=T.7=1,!16! 73+! *?@5+,45/'3! 4+,+! =,! 0=>+! -+98/79! =,! -+;8>+;! -+921,9+9! 71! :=77+-! @;+,.71,=80! :+,Q1.7+S! V8=,=,+! 98/6.7+)! 1-! 9O++7! >10218,;9! @98>-19+)(! I3+,! .! 7.97.,7! :=,;9! 71! .! 7.97+!-+>+271-!<.9vS!73+!*H2-17+=,!4897;8>=,!98:8,=79!*$!.,;!*"$!.-+!-+/+.9+;!.,;! .>7=T.7+! 73+! 2319231/=2.9+! :+7.#! @F/>t#)S! .! 0+0:-.,+H:18,;! +,Q50+! 92+>=6=>.//5! +W2-+99+;!=,!73+!<52+!''!>+//9(!I3+,!F/>t#!=9!.>7=T.7+;!=7!>.7./59+9!73+!2/.90.!0+0:-.,+! 2319231/=2=;9! =,71! 7O1! 2-1;8>79! ;=.>5/4/5>+-1/! @rU*)! .,;! "S%S&! =,19=71/! 7-=231923.7+! @'F$)! 73.7! 9+-T+! .9! 9+>1,;.-5! 0+99+,4+-9(! rU*! .>7=T.7+9! .! 2-17+=,! L=,.9+! BS! O3=>3! 2319231-5/.7+9!.,;!7389!.>7=T.7+9!.!4-+.7!T.-=+75!16!2-17+=,9(!'F$!-+48/.7+9!=,7-.>+//8/.-! /+T+/9! 16! >./>=80! :5! 12+,=,4! >./>=80! >3.,,+/9! 16! 73+! +,;12/.90=>! -+7=>8/80(! B5712/.90.7=>! >./>=80! -+/+.9+! >.,! -+48/.7+! 73+! 68,>7=1,! 16! 7.97+H-+/.7+;! >3.,,+/9(! <-20&S!.!0+0:+-!16!73+!7-.,9=+,7!-+>+271-!217+,7=./!@<-2)!>3.,,+/!6.0=/5S!=9!+W2-+99+;! 9+/+>7=T+/5!=,!<52+!''!>+//9!`"#S!##S!$%S!$&a(!m/+>7-12359=1/14=>./!978;=+9!16!3+7+-1/14189/5! +W2-+99+;! <-20&! 931O+;! 73.7! =7! =9! .! B.#yH.>7=T.7+;! >.7=1,! >3.,,+/! `$?H$Da! 9844+97=,4! 73.7! <-20&! 68,>7=1,9! O=73=,! 73+! <.9vH4897;8>=,HFXBt#H'F$! 2.73O.5! 71! ;+21/.-=Q+! 73+! <52+! ''! >+//9! .,;! /+.;! 71! ,+8-17-.,90=77+-! -+/+.9+(! <-20&HJH! 0=>+! 3.;! 0.-L+;/5! ;=0=,=93+;S! :87! ,17! .:1/=93+;S! -+921,9+9! 71! :=77+-S! 9O++7S! .,;! 80.0=! >10218,;9(!! A1;=80!=,6/8W!73-1843! "#$#$#%&'''(!8)9:!,;-!)6.-!""!/-77!,1*3+<',,-1! <3+!187>10+!16!F/>:#S!<-20&!.,;!91;=80!>3.,,+/!.>7=T.7=1,!=9!U ! #E! INTRODUCTION 0=>+! T1/7.4+H4.7+;! >8--+,79! .-+! 9+T+-+/5! -+;8>+;! =,! <52+! ''! >+//9S! U ',! .;;=7=1,S! U "#$#$#%&#'=(!>?@75,*<*,-:!,;-!A53B32C3D!'3!)6.-!""!+'@3*7!,*+,-!&54!,1*3+<'++'23! !r+92=7+!73+!;+:.7+!1,!73+!9=7+9!16!.>7=1,!16!U ! #"! INTRODUCTION >+//9! +W2-+99! ./91! =1,17-12=>! .,;! 0+7.:17-12=>! 4/87.0.7+! -+>+271-9! @6=4(! 'H%)! `&#H&&a(! A78;=+9!=,!=91/.7+;!7.97+!:8;9!:5!B.=>+;1!+7!./!931O+;!73.7!4/87.0.7+!7.-4+79!73+!7.97+! :8;9!.,;!,17 -+/+.9+;!:5!73+0 @6=4(!'H&)!`&?a(! ! 64! !!!!!!a(! ! 9@TEH<'.]U4!@U)!A5,.27=>S!2.-.>-=,+S!.871>-=,+!+66+-+,7!7-.,90=77+-9!.,;!73+=-!-+>+271-9!=;+,7=6=+;!=,!7.97+! :8;9!`&Ca(!<52+!''!@v+>+271-!>+//9)!-+921,;!.,;!7-.,9;8>+!9O++7S!:=77+-!1-!80.0=!97=08/=!:5!*FBv9!.,;! 9+>-+7+!U ! ! ! ##! INTRODUCTION ',!2.-7=>8/.-S!73+5!+W219+;!0189+!/=,48./!+2=73+/=80!71!4/87.0.7+!.,;!-+>1-;+;! 73+! -+921,9+9! 16! 73+! >+//9! O3+-+! 4/87.0.7+! >18/;! :+! ;=66+-+,7=.//5! ;=97-=:87+;! 1,! 73+! .2=>./! 2.-7! 16! 73+! 7.97+! :8;! @7=2)! 1- 73+! :.91/.7+-./! 2.-7! 73.7! >1--+921,;9! 71! 95,.27=>! -+4=1,(! <3+5! 618,;! 73.7! =1,17-12=>! 4/87.0.7+! -+>+271-9! ! @UGFUJw.=,.7+S! PGrU! -+>+271-9)!.-+!2-+9+,7!1,!73+!:.91/.7+-./!98-6.>+!16!7.97+!-+>+271-!>+//9S!9844+97=,4!73.7! 73+9+!.-+!219795,.27=>!9=7+9(!',!>1,7-.97S T+9=>8/.-!7-.,921-7+-9 61-!4/87.0.7+!@o*Xf<"S! #)!.-+!2-+9+,7!1,!73+!.66+-+,7!6=:+-9!73.7!+,;!82!.,;!=,,+-T.7+!7.97+!:8;9S!/+.;=,4!71! 73+! 92+>8/.7=1,! 73.7! 7.97+! 9+,91-5! 4.,4/=.! 0.5! -+/+.9+! XH4/87.0.7+! 73.7! :=,;9! 217+,7=.//5! :=,;9! 71! 7.97+! :8;H/1>.7+;! -+>+271-9 `&Ra(! '7! 0897! :+! -+0=,;+;S! 73.7! 73+! .22-12-=.7+! >1,>+,7-.7=1,! 71! .>7=T.7+! 95,.29+9! =9! 0=>-101/.-S! 12219=,4! 73+! >1,>+,7-.7=1,! ,++;+;! 61-! 7.97+! -+>+271-! 97=08/.7=1,S! O3=>3! =9! 0=//=01/.-(! h8.,4! +7! ./! 931O+;! 73.7! O3+,! 4/87.0.7+! =9! .22/=+;! 1,! =91/.7+;! 7.97+! :8;9! .7! 95,.27=>! >1,>+,7-.7=1,9S! 73+! 2-+95,.27=>! >+//9! 9+>-+7+! 9+-171,=,! `?EaS! 68-73+-! 98221-7=,4! .! 4/87.0.7+!95,.27=>H-+/.7+;!-1/+!`&Ra!`?Ea(!' ! ! 9@TEH<'.]:('1,17-12=>!.,;!0+7.:17-12=>!4/87.0.7+!-+>+271-9(!U>7=T.7=1,!16!=1,17-12=>!4/87.0.7+!-+>+271-9! /+.;!71!0+99.4+!7-.,90=99=1,!T=.!=1,!>8--+,79!O3+-+.9!.>7=T.7=1,!16!0+7.:17-12=>!4/87.0.7+!-+>+271-9! .>7=T.7+!9+>1,;!0+99+,4+-9(! ' ' ' ! #$! P. # INTRODUCTION ! ! ' 9@TEH<' .]S4! <3-++! 2-=,>=2./! >/.99+9! 16! 7.97+! >+//9(! <3=9! >.7+41-=Q.7=1,! =,>/8;+9! 8/7-.97-8>78-./! 6+.78-+9S! 2.77+-,9!16!4+,+!+W2-+99=1,!.,;!73+!68,>7=1,!16!+.>3!>+//!752+(!<52+!'!>+//9!@A8221-7)!=,!:/8+!;+4-.;+!1-! .:91-:!,+8-17-.,90=77+-9!.,;!0=437!+T.>8.7+!+W7-.>+//8/.-!wy!73.7!.>>808/.7+9!.67+-!.>7=1,!217+,7=./9!=,! <52+!''!>+//9!@<.97+!-+>+271-)!=,!5+//1O!.,;!<52+!'''!>+//9!@F-+95,.27=>)!=,!4-++,(!wy!0.5!:+!-+/+.9+;!73-1843! .2=>./! w! >3.,,+/9! 98>3! .9! vnGw(! A10+! <52+! '! >+//9! 0.5! 7-.,9;8>+! 9./75! 7.97+S! :87! 73=9! 97=//! -+0.=,9! 8,>+-7.=,(! AO++7S! :=77+-S! .,;! 80.0=! 7.97.,79! .>7=T.7+ -+>+271-! >+//9S! =,;8>=,4! 73+0! 71! -+/+.9+! U ! <3+!<52+!'''!@2-+95,.27=>)!>+//9!;+7+>7!918-!.,;!>.-:1,.7+;!7.97.,79(!',!4+,+-./S! .>=;9!>.,!:/1>LS!2+-0+.7+!1-!.>7=T.7+!=1,!>3.,,+/9!.,;!>3.,4+!=,7-.>+//8/.-!2h!:87!73+! 01/+>8/+9!=,T1/T+;!=,!73+!.>=;!7-.,9;8>7=1,!0.>3=,+-5!=,!7.97+!:8;9!.-+!,17!=;+,7=6=+;(! Fwr#X"!.,;!Fwr"X$!@61-!21/5>597=>!L=;,+5!;=9+.9+!#H/=L+!"!.,;!"H/=L+!$!-+92+>7=T+/5)!3.T+! :++,!2-1219+;!.9!>.,;=;.7+!7.97+!-+>+271-9!61-!918-!97=08/=!`?"H?$a(!<3+9+!>3.,,+/9!.-+! 9+,9=7=T+!=,!+W7-.>+//8/.-!2h!>3.,4+9!:87!,17!71!>5712/.90=>!2hS!O3=>3!=9!73+!+99+,7=./! 97=08/89!61-!918-!7-.,9;8>7=1,!`?%S!?&a!@6=4('HC)(!U;;=7=1,.//5S!Fwr"X$!L,1>LH187!0=>+!.-+! .:/+! 71! ;+7+>7! 918-! 97=08/=! =,;=>.7=,4! 73.7! 173+-! 01/+>8/+9! .-+! -+V8=-+;! 61-! 918-! ;+7+>7=1,!.7!73+!7.97+!:8;!/+T+/!`??a(! ! #%! INTRODUCTION ! <52+! '''! >+//9! 61-0! 95,.29+9! .,;! +W2-+99! ,+8-./H/=L+! 4+,+9! 98>3! .9! ,+8-./! .;3+9=1,!01/+>8/+!@PBUG)S!95,.271910./H.991>=.7+;!2-17+=,!#&!@APUFH#&)!+,Q50+9!61-! ,+8-17-.,90=77+-! 95,73+9=9S! 7-.,921-7+-9 .,;! T1/7.4+H4.7+;! >./>=80 61-! 73+! -+/+.9+! 16! ,+8-17-.,90=77+-9! `?CHCEa(! <3+5! ./91! +W2-+99! 91;=80! .,;! 217.99=80! >3.,,+/9! .,;! 4+,+-.7+!.>7=1,!217+,7=./9(!f21,!.>=;S!wB/!1-!U 9O++7S!:=77+-!1-!80.0=(!I3+,!*UMU!=9!-+/+.9+;S!=7!7.-4+79!*UMUU .,;!*UMUM -+>+271-9! ./91! /1>.7+;! =,! <52+! ''! >+//9! .,;! 2-1017+9! *UMU+-4=>! =,3=:=7=1,! 16! 7.97+H+T1L+;! 7-.,90=77+-9!@N=4(!'H%S?)(!! <52+!'''!+W2-+99!./91!>.-:1,=>!.,35;-.9+!%[email protected]%)S!.!0+0:+-!16!.!/.-4+!6.0=/5!16! +,Q50+9!=02/=>.7+;!=,!9+,9=,4S!.>7=,4!1,!.,;!-+921,;=,4!71!Bn#!=,!T.-=189!9597+09!`C%a(! B3.,;-.93+L.-!+7!./!;+T+/12+;!+,4=,++-+;!0=>+!=,!O3=>3!92+>=6=>!7.97+!>+//!752+9!O+-+! 4+,+7=>.//5! .:/.7+;! =(+(! 9O++7/+99S! 80.0=/+99! 1-! 918-/+99! 0=>+! .,;! 73+=-! -+921,9+9! 71! Bn#! O+-+! -+>1-;+;(! G=>+! O=73187! 73+=-! 918-H-+921,9=T+! >+//9! ;=;! ,17! -+921,;! 71! Bn#! +=73+-S!=,;=>.7=,4!73.7!73+9+!918-H-+921,9=T+!>+//9!.-+!+99+,7=./!61-!Bn#!;+7+>7=1,(!A=,>+! 73+!Bn#!-+921,9=T+!>+//9!.-+!=;+,7=6=+;S!73+!,+W7!97+2!O18/;!:+!71!=;+,7=65!73+!-+>+271-9! 1,!73+9+!>+//9!73.7!-+921,;!71!Bn#!.7!73+!7.97+!:8;!/+T+/(! 9@TEH<' .]P(! r=97=,>7! >/.99+9! 16! 0.00./=.,! 7.97+! -+>+271-9S! >+//! 752+9!.,;!/=4.,;9(!AO++7! .,;!80.0=!.-+!;+7+>7+;! :5! <"v! 3+7+-1;=0+-9! `"Ra! `&%a! `C&a! .,;! :=77+-! :5! <#v! -+>+271-9! `#%a! `C?a!`CCa(!A./7!=9!;+7+>7+;! :5! 91;=80! >3.,,+/! @mP.B)! `CDa(! Fwr#X"! =9! =,T1/T+;! =,! 918-! ;+7+>7=1,! `?"a! .,;! >.-:1,.7=1,! =9! 0+;=.7+;! :5! 0+0:-.,+H7+73+-+;! >.-:1,=>! .,35;-.9+! B.-%! `C%a(!' ! ! #&! INTRODUCTION ! )EF!G8GG8>"8H!)8I)F!JKLI!"H!IKGG8MN#! ',! 9800.-5S! 73+! 73-++! ;=97=,>7! >+//! 752+9! =,! 73+! 7.97+! :8;! .-+! ;+;=>.7+;! 71! ;+7+>7! ;=66+-+,7!7.97+!97=08/=(!!<3+9+!7.97+!:8;!>+//!752+9!-+/+.9+!9+T+-./!,+8-17-.,90=77+-9!@=(+(! 752+!''S!U ! ! 9@TEH<' .]V4! <3+! 2.-.>-=,+! 9=4,./=,4! =,! 7.97+! 7-.,9;8>7=1,(! ',! 7.97+! :8;9S! *H2-17+=,! >182/+;! -+>+271-9! @*FBv9)! 61-! :=77+-S! 9O++7! .,;! 80.0=! 7.97.,79! .-+! /1>.7+;! .7! 73+! .2=>./! 0+0:-.,+! 16! <52+! ''! >+//9(! U>7=T.7=1,!16!73+!*FBv!9=4,./=,4!2.73O.5!/+.;9!71!73+!2-1;8>7=1,!16!'F$!.,;!;=.>5/4/5>+-1/!@rU*)!73-1843! 2319231/=2.9+!Bt!@FXBt)H0+;=.7+;!>/+.T.4+!16!231923.7=;5/=,19=71/!%S&H:=9231923.7+!@F'F#)(!U,!=,>-+.9+! 16! `B.#ya=! :5! 'F$H=,;8>+;! B.#y -+/+.9+! 12+,9! 7-.,9=+,7! -+>+271-! 217+,7=./! >.7=1,! >3.,,+/! 98:6.0=/5! G! 0+0:+-!&!@ ! #?! INTRODUCTION ' .424G/'9HI='B;<'J ',!-1;+,79S!7.97+!:8;9!-+>+=T+!2-1>+99+9!6-10!29+8;1H8,=21/.-!,+8-1,9!/1>.7+;! =,! 73+! 6.>=./! @o')S! 73+! 4/199123.-5,4+./! @'p)! .,;! 73+! T.4./! @p)! >-.,=./! ,+-T+9(! <3=9! 1-4.,=9.7=1,!=9!>1,9+-T+;!=,!g.O+;!T+-7+:-.7+9!`CRa(!',!2.-7=>8/.-S!7.97+!:8;9!;=97-=:87+;! =,!73+!.,7+-=1-!71,48+!@68,4=61-0!2.2=//.+)!.,;!2./.7+!-+>+=T+!2-1>+99+9!16!73+!9+,91-5! ,+8-1,9! /1>.7+;! =,! 73+! 4+,=>8/.7+! 4.,4/=.! @o''73)(! <3+9+! 2-1>+99+9! =,T.;+! O=73=,! 73+! /;214*!,6<.*3'!,+-T+!.,;!73+!4-+.7+-!2+7-19./!,+-T+9S!-+92+>7=T+/5(!<.97+!:8;9!=,!73+! 2197+-=1-!1-./!>.T=75!@61/=.7+S!>=->80T.//.7+!2.2=//.+)!-+>+=T+!4/199123.-5,4+./!2-1>+99+9! 6-10!73+!2+7-19./!4.,4/=.!@'p73)!@6=4('HR)(!U!90.//!,80:+-!16!7.97+!:8;9!=,!73+!+2=4/177=9! -+>+=T+! 982+-=1-! /.-5,4+./! .66+-+,79! 6-10! 73+! ,1;19+! 4.,4/=.! @p73)! .,;! -+921,;! 01-+! 92+>=6=>.//5!71!73+!-+6/+W!16!-+g+>7=1,!1-!=,4+97=1,[email protected]!-+6/+W)!`DEa(!! ! F-1>+99+9! 6-10! $H"%! 9+,91-5! ,+8-1,9! -+.>3! +.>3! 7.97+! :8;! ;+2+,;=,4! 1,! 73+! 92+>=+9(!h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`D"a(! h1O+T+-! -+>1-;=,49! 6-10! 73+! /;214*! ,6<.*3'! ,+-T+! 6=:+-9! >.,! 931O! 08/7=2/+! 9+,9=7=T=75! 821,! 97=08/.7=1,! .,;! -+921,;! 71! 9+T+-./! 752+9! 16! 97=08/=S! 9844+97=,4! 73.7! 7.97+!V8./=7=+9!.-+!+,>1;+;!:5!.,!.>-199H6=:+-!2.77+-,(!',!.;;=7=1,S!O=73=,!73+!4.,4/=.S! ,+8-1,9!73.7!.-+!:-1.;/5!78,+;!3.T+!:++,!618,;(!`D#a(!U;;=7=1,./!=,7-=48=,4!.92+>79!.-+! 73.7!.!9=,4/+!<"v#J<"v$!1-!.!<#v!+W2-+99=,4!7.97+!:8;!>+//!>.,!-+>14,=9+!9+T+-./!9O++7! 1-!:=77+-!752+9!16!97=08/=S!-+92+>7=T+/5S!.,;!73.7!7.97+!:8;!>+//9!>1008,=>.7+!O=73!+.>3! 173+-]! 61-! =,97.,>+! <52+! '''! >+//9! .-+! =,;=-+>7/5! .>7=T.7+;! :5! <52+! ''! HU ! #C! INTRODUCTION .7! 73+! /+T+/! 16! 73+! 7.97+! :8;9S! 4.,4/=.! .,;! 73+! 2-=0.-5! 7.97+! >+,7+-! =,! 73+! 3=,;:-.=,! @,8>/+89!16!73+!91/=7.-5!7-.>7)!.,;!61-+:-.=,!7.97+H-+/.7+;!.-+.9!=,!127=>.//5!>/+.-!.,=0./9! 98>3! .9! Q+:-.6=93S! >18/;! 68-73+-! 93+;! /=437! =,! 73+! 0.,,+-! 73.7! 7.97+! =,61-0.7=1,! =9! +,>1;+;(! .424G4G/',>==>?@>A'DH>@A'B>CB<]HB <3+!4897.71-5!+66+-+,79!16!73+!o''S!'pS!p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`D%a(! ! f9=,4! .! >10:=,.7=1,! 16! 4+,+7=>! 7-.>=,4S! +W7-.>+//8/.-! -+>1-;=,49! .,;! >./>=80! =0.4=,4!71!.>3=+T+!9=,4/+!>+//!-+91/87=1,S =7!O.9!931O,!73.7!O=73!73+!+W>+27=1,!16!73+! 918-!7.97+S!+.>3!7.97+!V8./=75!=9!-+2-+9+,7+;!=,!.!9+2+-.7+!>1-7=>./!6=+/;S!.,;!7389!.!7.97+! 0.2!=9!2-+9+,7!=,!73+!:-.=,!`D&a(! !!9@TEH<'.]2W4!U,.710=>./!1T+-T=+O!16!73+!>+,7-./!7.97+! 2.73O.59(!m/+>7-=>./!9=4,./9!6-10!>-.,=./!,+-T+9!o''S!'p! .,;! p! 73.7! >1,7.=,! =,61-0.7=1,! 1,! 73+! >3+0=>./! 2-12+-7=+9! 16! 7.97.,79! .-+! >1,T+5+;! 71! 73+! -197-./! ;=T=9=1,! 16! 73+! ,8>/+89! 16! 73+! 91/=7.-5! 7-.>7! @-P ! $E! INTRODUCTION ' .4G/'6#6$",8'6#0'91#!$."#'"9'$5%'$%*%"&$'9.&5'$6&$%'&8&$%,' .4G42/'$CB<'DEFC' <3+! 7.97+! 9597+0! =,! 7+/+1979! =9! 1-4.,=9+;! .7! 73-++! /+T+/9! .9! ;+9>-=:+;! =,! 0.00./9]!73+!7.97+!:8;9S!73+!7.97+!9+,91-5!4.,4/=.!.,;!73+!7.97+H-+/.7+;!:-.=,!,8>/+=(! `DCS!DDa(!<3+!Q+:-.6=93!7.97+!:8;9!.-+!2.77+-,+;!./1,4!73+!.,7+-12197+-=1-!.W=9!16!73+! 01873S!2./.7+!.,;!23.-5,4+./!.->3+9(!',!73+!.;8/7S!73+-+!.-+!./91!/1>.7+;!1,!73+!:.-:+/9! .,;!+2=;+-0=9(!!',!>1,7-.97!71!0.00./9S!73+5!.-+!/1>.7+;!;=-+>7/5!1,!73+!1-123.-5,4+./! +2=73+/=80S!.,;!,17!+,93+.73+;!=,71!2.2=//.(!! U>>1-;=,4!71!73+=-!+/+>7-1,H;+,9=75S!931O,!:5!+/+>7-1,!0=>-19>125!.,./59=9S!73+! Q+:-.6=93! 7.97+! >+//9! .-+! 9+2.-.7+;! =,! 73-++! ;=97=,>7! 4-1829]! 73+! ;.-L! >+//9S! O=73! 90.//! 0=>-1T=//=S! 73+! /=437! >+//9! O=73! 1,+! /.-4+! 0=>-1T=//89! 1,! 73+! .2=>./! +,;! .,;! 73+! =,7+-0+;=.7+!1-!:.9./!>+//9!1,!73+!:17710!16!73+!7.97+!:8;!>/897+-!.;g.>+,7!1,!73+!173+-! 7O1!752+9!`DRHRRa('U!>/+.-!>102.-=91,!:+7O++,!73+9+!01-231/14=>./!>3.-.>7+-=97=>9!.,;! 73+!0.00./=.,!<52+!'S!''!.,;!'''!>+//!752+9!3.9!,17!:++,!>/+.-/5!+97.:/=93+;(!h1O+T+-S! 9+T+-./!01/+>8/.-!>3.-.>7+-=97=>9!16!73+!Q+:-.6=93!7.97+!:8;!>+//!752+9!98221-7!73.7!73+! 97-8>78-+! .,;! 2199=:/5! 68,>7=1,! 16! 73+9+! 1-4.,9! =9! >1,9+-T+;! :+7O++,! 7+/+197! .,;! 0.00./9(!! N=-97!16!.//S!Q+:-.6=93!.,;!=,!4+,+-./!7+/+197!7.97+!:8;!>+//9!+W2-+99!7.97+!-+>+271-! 4+,+9!73.7!:+/1,4!71!73+!9.0+!6.0=/5!.9!73+=-!0.00./=.,! 1-731/149(! N=93!3.T+!9=,4/+! )*+$1$!.,;!)*+$1[!4+,+!1-731/149!.,;!9+T+-./!752+9!16!)*+$1%!4+,+9!=,!73+=-!4+,10+! `"EES!"E"a(!M87!;8+!71!73+!+W2.,9=1,!16!)*+$1%!4+,+9S!73+!+W2-+99=1,!2.77+-,9!16!<.9"-! 2-17+=,9!=,!6=93!7.97+!:8;9!.-+!;=T+-9+!>102.-+;!71!7319+!=,!-1;+,79!`"EES!"E"a(!! U,! =021-7.,7! ;=66+-+,>+! :+7O++,! -1;+,79! .,;! 6=93! =9! 73.7! 6=93! ;1! ,17! 9+,9+! 9O++7(!',;++;!6=93!7.97+H-+/.7+;!6.>=./!,+-T+9!;=;!,17!-+921,;!71!.,5!7.97+!98:97.,>+9! 73.7! 380.,9! 2+->+=T+! .9! 9O++7! `"E#a(! B1,9=97+,7/5S! >8/78-+;! >+//9! +W2-+99=,4! .,5! >10:=,.7=1,!16!<.9"-!2-17+=,9!6-10!Q+:-.6=93!.,;!0+;.L.!;1!,17!-+921,;!71!Z9O++7[! 98:97.,>+9! :87! .-+! .>7=T.7+;! :5! XH.0=,1! .>=;9! =,! 73+! 9.0+! O.5! .9! 0.00./=.,! )*+$1$J)*+$1[H+W2-+99=,4!80.0=!7.97+!>+//9!`"E#a(!! ! $"! INTRODUCTION <3+!/.>L!16!9O++7!7.97+!2+->+27=1,!=,!6=93!=9!-.73+-!,17!.,!+W>+27=1,(!,*+$1%!4+,+! =9!.!29+8;14+,+!=,!6+/=,+!4+,10+!@>.79!;1!,17!2-+6+-!984.-!+=73+-)!`"E$a!.,;!73+!>3=>L+,! 4+,10+! /.>L9! 73+! )*+$1% 4+,+! +,7=-+/5! `"E%a(! <14+73+-! O=73! 73+! 6.>7! 73.7! 6=93! 3.T+! 08/7=2/+!)*+$1%!4+,+9S!=7!=9!=,7-=48=,4/5!+T=;+,7!73.7!)*+$1%!4+,+9!.-+!6.-!01-+!;=T+-4+,7! 73.,! .-+! )*+$1$! .,;! )*+$1[! 4+,+9(! <.9"-H0+;=.7+;! 7.97+! 0.5! :+! 1-=4=,.//5! ;8+! 71! XH .0=,1!.>=;9 .,;!9O++7!7.97+!0.5!:+!.!,+O/5!.>V8=-+;!7.97+!=,!910+!0.00./=.,!92+>=+9! 73-1843!73+!+T1/87=1,!16!)*+$1%!`"E&a(!! ,*+%1H+W2-+99=,4!>+//9!=,!Q+:-.6=93!68,>7=1,!.9!;+7+>71-9!16!3.-068/!>3+0=>./9!.,;! 0.5! 7-=44+-! 9+/6H;+6+,9=T+! -+921,9+9! 98>3! .9! .T1=;.,>+(! <3+-+! .-+! &! <.9#-! 4+,+9! =,! Q+:-.6=93!:87!.9!73+!4+,10+!3.9!,17!:++,!68//5!.,,17.7+;S!173+-!4+,+9!0.5!:+/1,4!71! 73=9! 98:6.0=/5(! ',7-=48=,4/5S! )*+%1! 4+,+9! =,! 7+/+197! 6=93! .-+! 235/14+,+7=>.//5! ;=66+-+,7! 6-10!7+7-.21;!)*+%1!4+,+9S!.,;!73+!6=93!;+,.71,=80!-+>+271-9!Q6<#v&!.,;!06<#v"!.-+! ,17! 1-731/149! 16! 73+! 0189+! ;+,.71,=80! -+>+271-! 0<.9#-"ED! @0189+! <#vD)! `"E?a(! h1O+T+-S!Q+:-.6=93!.T1=;!+.7=,4!.!;=+7!73.7!>1,7.=,9!;+,.71,=80!`"E#a!9844+97=,4!73.7! <.9#-9! 16! 910+! 752+! .-+! =,T1/T+;! =,! .T1=;.,>+! 6++;=,4! :+3.T=1-9! =,! 6=93! .9! O+//! .9! =,! 0.00./9(! ',!Q+:-.6=93!7.97+!:8;9S!.!0=,1-!:87!9=4,=6=>.,7!2128/.7=1,!16!7.97+!>+//9!+W2-+99+9! :173!)*+$1+!.,;!)*+%1+!`"ECa(!B1,9=;+-=,4!73.7!Q+:-.6=93!2-+6+-9!.0=,1!.>=;9!73.7!.-+! -+>+=T+;! :5! T.-=189! <.9"-! 3+7+-11/=410+-9! `"E#a! 73+! .>7=T.7=1,! 16! 7.97+! >+//9! >1+W2-+99=,4! )*+$1+! .,;! )*+%1+! 9318/;! -+98/7! =,! .77-.>7=1,(! '7! =9! 8,>/+.-! O3+73+-! Q+:-.6=93! O18/;! /=L+! 1-! ;=9/=L+! 73+! 98:97.,>+9! ;+7+>7+;! :5! Q6<#v9! 173+-! 73.,! Q6<#v&(! ',7+-+97=,4/5S! 73+! +W2-+99=1,! 16! _0)%M`! =9! >102/+7+/5! 9+4-+4.7+;! 6-10! 73.7! 16! )*+$1+! `"E#a!91!73+!>+//9!+W2-+99=,4!_0)%M`S!>.89=,4!.T+-9+!-+921,9+9S!.-+!;=97=,>7!6-10!7319+! >.89=,4!.77-.>7=1,!-+921,9+9S!+T+,!=,!Q+:-.6=93!@6=4(!'H"")(!! U!9+>1,;!.-480+,7!=,!6.T18-!16!73+!>1,9+-T.7=1,!16!7.97+!:8;!<52+!''!>+//9!=,!6=93! =9! 73.7! .! 92+>=6=>! 98:9+7! 16! 6=93! 7.97+! :8;! >+//9! +W2-+99+9! F/>t#S! <-20&! .,;! >./>=80H :=,;=,4! 2-17+=,9! 9844+97=,4! 73.7! 73+! 7-.,9;8>7=1,! 0.>3=,+-5! =9! 9=0=/.-! 71! 0.00./9(! <3=-;S!,.;$&\&Hh+//9!3.T+!:++,!;+9>-=:+;!=,!Q+:-.6=93!:87!73+5!.-+!/1>.7+;!=,!.!:.9./! 219=7=1,!-+/.7=T+!71!73+!173+-!>+//!752+9!O=73=,!73+!7.97+!:8;!`"EDa(!<3=9!752+!16!7.97+!:8;! 1-4.,=9.7=1,! =9! ./91! 2-+9+,7! =,! .023=:=.,9(! <3+-+! .-+! ,1! 2359=1/14=>./! ;.7.! .T.=/.:/+! ! $#! INTRODUCTION .:187!73+!>+//!752+!73.7!;+7+>79!918-!=,!6=93(!P+T+-73+/+99S!73+-+!=9!.!>/+.-!-+921,9+!71! .>=;!.,;!2h!>3.,4+9!;+9>-=:+;!=,!>3.,,+/!6=93!/=,L+;!71!6++;=,4!:+3.T=18-(!<3+!e.2.,+9+! y 9+.!>.76=93!972,2+5+!a*.23'/5+ 9+,9+9!/1>./!2hH.991>=.7+;!=,>-+.9+9!=,!h JBn# +V8=T./+,7 71!.!;+>-+.9+!16!{E("!2h!8,=7!=,!.0:=+,7!9+.O.7+-!2-1T1L+;!:5!21/5>3.+7+!O1-09!.,;! ;+7+>79! 2-+5! =,! 73=9! O.5! `"ERa(! F+-3.29! 01-+! +,=40.7=>! =9! 73+! 2-+9+,>+! 16! <52+! '! @98221-7!>+//9)!=,!6=93!.9!*XUA ! 9@TEH<' .]224!r=T+-9+!.--.5!16!7.97+!>+//9!=,!0189+!.,;!Q+:-.6=93(!<52+9!16!7.97+!>+//9!.-+!=//897-.7+;!O=73! 92+>=6=>! 01/+>8/.-! 6+.78-+9! 16! 7.97+! -+>+271-9! .,;! >1--+921,;=,4! 0.-L+-9(! <3+! =;+,7=6=+;! /=4.,;9! .-+! =,;=>.7+;! 731843! 8,=;+,7=6=+;! -+>+271-9! .,;! /=4.,;9! .-+! 931O,! :5! V8+97=1,! 0.-L9(! <3+! 8,=;+,7=6=+;! -+>+271-9! =,! FXBHt#J U9!=,!0.00./9S!73-++!7.97+!9+,91-5!4.,4/=.!@o''S!'pS!p)!>1,T+5!7.97+!=,61-0.7=1,! =,!6=93(!',!2.-7=>8/.-S!7.97+!:8;9!1,!73+!:1;5!98-6.>+!.,;!1-./!-+4=1,9!.-+!=,,+-T.7+;!:5! 73+!6.>=./!@o''73)!,+-T+S!O3=/+!7.97+!:8;9!/1>.7+;!=,!73+!23.-5,41:-.,>3=./!-+4=1,9!.-+! =,,+-T.7+;! :5! 73+! 4/199123.-5,4+./! @o''73)! .,;! T.4./! @o73)! ,+-T+9! -+92+>7=,4! .,! .,7+-12197+-=1-!1-;+-!`"""a(!! ! $$! INTRODUCTION ! I=73=,! 73+! :-.=,S! 9=/8-=.,! .,;! >52-=,=;! 7+/+197! 6=93! @=,>/8;=,4! Q+:-.6=93)! 3.T+! +/.:1-.7+;! 92+>=./=Q+;! ,8>/+=! >.//+;! 6.>=./S! 4/199123.-5,4+./! .,;! T.4./! /1:+9! 73.7! .-+! 3101/14189! 71 73+! 2-=0.-5! 7.97+! >+,7+- =,! 73+! 0.00./=.,! 3=,;:-.=, O3=>3! =9! 73+! .,7+-=1-!2.-7!16!,8>/+89!16!73+!91/=7.-5!7-.>7(!<3+!7.97+H-+/.7+;!9+,91-5!6=:+-9!16!73+!6.>=./! ,+-T+! 7+-0=,.7+! =,! 73+! 6.>=./! /1:+S! 7319+! 16! 73+! 4/199123.-5,4+./! =,! 73+! 4/199123.-5,4+./!/1:+!.,;!7319+!16!73+!T.4./!,+-T+!=,71!73+!T.4./!/1:+(!',!910+!92+>=+9! 98>3!.9!41/;6=93S!73+!T.4./!/1:+!3.9!.,!+W7-+0+/5!+/.:1-.7+;!1-4.,=9.7=1,!=,!/.5+-9!!@/=L+! .! 0.00./=.,! >1-7+W)(! <3+! 9+,91-5! /.5+-9! @3101/14189! 71! 73+! 0.00./=.,! .,7+-=1-! ,8>/+89! 16! 73+! 91/=7.-5! 7-.>7)! .-+! >182/+;! 71! 0171-! /.5+-9! @3101/14189! 71! ,8>/+89! .0:=4889)!T=.!.!9=02/+!-+6/+W!.->!.,;!9+-T+!71!-.2=;!611;!91-7=,4!.,;!=,7.L+!`""#a(! <3+! 7.97+H-+/.7+;! 6=93! /1:+9! .-+! >1,,+>7+;! O=73! 01-+! -197-.//5! /1>.7+;! ,8>/+=! =,>/8;=,4!73+!9+>1,;.-5!4897.71-5!,8>/+89!=,!Q+:-.6=93!@T+,7-./!71!73+!>+-+:+//80)S!73+! 7+-7=.-5!2-+4/10+-8/.-!>102/+W!,8>/+89!@2199=:/5!3101/14189!71!7.97+H-+/.7+;!73./.0=>! -+/.5!=,!0.00./9)!.,;!73+!0+;=./!2.-7!16!73+!;1-9./!7+/+,>+23./1,!@r0)(!',!>.76=93S!73+! T.4./!/1:+!3.9!-+>=2-1>./!>1,,+>7=1,9!O=73!73+!=29=/.7+-./!/1:1:8/:.-!,8>/+89!.,;!6.>=./! /1:+(!h1O+T+-S!31O!7.97+!=,61-0.7=1,!=9!7-.,90=77+;!.,;!+,>1;+;!=,!73+!7+/+197!:-.=,!=9! 8,L,1O,(!! ! $%! INTRODUCTION ! ! ! 9@TEH<' .]2G(! b$/! A>3+0.7=>! ;=.4-.0! 16! .! 7.97+! :8;S! 7.97+! -+>+271-! >+//! .,;! .991>=.7+;! ,+8-1,9(! .)! '//897-.7=1,! 16! .! 7.97+! :8;! 73.7! =9! +0:+;;+;! =,! .,! +2=73+/=80(! <3+! ;=66+-+,7! 752+9! 16! <52+! ''! >+//9! .-+! =,;=>.7+;!:5!;=66+-+,7!>1/18-9!.9!73+5!>.,!>1,7.=,!;=66+-+,7!752+9!16!-+>+271-!.,;!=,7-.>+//8/.-!01;8/.71-(! <3+!4897.71-5!,+8-1,9!O=73!73+!.991>=.7+;!>1/18-9!73.7!0.7>3!73+!.991>=.7+;!<52+!''!>+//9!=,;=>.7+!73.7! 73+5!0=437!-+921,;!:+97!71!97=08/=!73.7!.>7=T.7+!73+9+! 2.-7=>8/.-!<52+! ''! >+//9(!<3+9+!2-=0.-5!4897.71-5! ,+8-1,9!2-1g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`""$a(! r+92=7+! 73+! ;=66+-+,7! 7=998+! 16! 1-=4=,! 1,/5! .! 6+O! ;=66+-+,>+9! .-+! 1:9+-T+;! :+7O++,! .,7+-=1-! .,;! 2197+-=1-! 7.97+! :8;9S! >1,>+-,=,4! 73+! +W2-+99=1,! 16! 2.-./14! 4+,+9! `""%S! ""&a(!<3+-+61-+S!=7!9++09!73.7!./731843!73+!7=998+!16!1-=4=,!.,;!2+-3.29!71!910+!+W7+,7! 73+!9=4,./=,4!01/+>8/+9!;=66+-!;8-=,4!73+!2-1>+99!16!7.97+!:8;!61-0.7=1,S!73+!187>10+!=9! -.73+-!9=0=/.-!=,!7+-09!16!1-4.,=Q.7=1,!.,;!68,>7=1,!16!73+9+!9+,91-5!1-4.,9(! ! <3+! +2=73+/=./! 1-=4=,! 16! 7.97+! :8;9! O.9! 6=-97/5! 98221-7+;! :5! 978;=+9! ;1,+! =,! .023=:=.,9!.,;!0=>+(!',!.023=:=.,9S!+>712=>!=91712=>!.,;!=91>3-1,=>!4-.679!16!2/.>1;./! +>71;+-0!1-!,+8-./!61/;9S!6.=/+;!71!4+,+-.7+!7.97+!:8;9(!',!>1,7-.97S!r='!/.:+//=,4!16!73+! 2-+98027=T+! 1-123.-5,4+./! +,;1;+-0! ;8-=,4! 4.97-8/.7=1,S! -+98/7+;! =,! r='! /.:+//=,4! 16! 23.-5,4+./! 7.97+! :8;9(! ! U;;=7=1,.//5S! A71,+! +7! ./! .,./59+;! 71,48+9! 16! p! >3-101910+H =,.>7=T.7=1,!019.=>!0=>+!71!+W.0=,+!O3+73+-!7.97+!:8;9!1-=4=,.7+!6-10!/1>./!+2=73+/=80! 1-! ,+8-14+,=>! +>71;+-0(! <3+! bH4./! 23+,1752+! 16! :173! 68,4=61-0! @.,7+-=1-)! .,;! >=->80T.//.7+! @2197+-=1-)! 7.97+! :8;9! 0.7>3+;! O=73! 73+! 98--18,;=,4! +2=73+/=80! 9844+97=,4! 73.7! :173! 7=998+9! 93.-+! 73+! 9.0+! 7.97+! 2-14+,=71-! >+//9! ;+-=T+;! 6-10! /1>./! +2=73+/=80! `""?S! ""Ca(! G1-+! -+>+,7/5S! v1731T.! +7! ./S! 89+;! A1W"CH#UH=B-+Jv19.#?! ! $?! INTRODUCTION -+21-7+-! 0=>+! 71! 61//1O! 73+! 6.7+! 16! 73+! +,;1;+-0! .,;! 618,;! 73.7! >=->80T.//.7+! .,;! 61/=.7+!2.2=//.+!7.97+!:8;9!.-+!16!+,;1;+-0./!1-=4=,(!U9!.//!7.97+!:8;9!.-+!;+-=T+;!6-10! +2=73+/=./! >+//9! .,;! 73+! .,7+-=1-! 71,48+! +2=73+/=80! =9! +>71;+-0./! ;+-=T.7=T+S =7! =9! .9980+;!73.7!73+!.,7+-=1-!7.97+!:8;9!.-+!16!+>71;+-0./!1-=4=,!`""Da(!! ! .434G/' $>CB<' DEF' J?>LIF<'M' B;<' O@HCB' =IHJ;I?IT@L>?' ! ',!0.00./9S!7.97+!:8;9!97.-7!71!;+T+/12!.7!m""(&!71!m"#(&!O3+-+!+2=73+/=./!>+//9! =,T.4=,.7+! .7! 2.-7=>8/.-! 9=7+9! 16! 73+! ;+T+/12=,4! 71,48+! .,;! 61-0! 73+! 91H>.//+;! q7.97+! 2/.>1;+9c(!F/.>1;+9 1-!2.2=//.!2/.>1;+9!-+6+-9!71!.!4-182!16!>+//9!16!73+!+.-/5!97.4+9!16! 2.2=//.!;+T+/120+,7!.,;!=,;+2+,;+,7!6-10!73+!,+8-14+,=>!2/.>1;+9(!<3+!2-+>=9+!97+29! 16!73+!01-2314+,+7=>!01T+0+,79!73.7!/+.;!71!73+!7.97+!:8;!61-0.7=1,!6-10!73+!=,=7=./! 2/.>1;+! .-+! 211-/5! 8,;+-9711;(! N-10! 6=W+;! 0189+! 7=998+S! =7! 9++09! 73.7! 73+! +2=73+/=./! 2/.>1;+! 73=>L+,9! .,;! 98--18,;9! 73+! 2-1W=0./! 0+9+,>350+(!!U;;=7=1,.//5S! .! >/897+-! 16! +2=73+/=./! >+//9! 2-+9+,79! ;=66+-+,7! 01-231/145! O=73! .! ;+,9+-! >5712/.90! .,;! 6+O+-! 0=>-1T=//=! .,;! 73=9! =9! >1,9=;+-+;! .9! .,! =,;=>.7=1,! 16! 7.97+! :8;! .,;! 98--18,;=,4! 2.2=//.! 61-0.7=1,! `""RS! "#Ea(! r8-=,4! 73+9+! +.-/5! ;+T+/120+,7./! 97+29S! 7.97+! 2/.>1;+9! .-+! =;+,7=6=.:/+! :5! 73+! +W2-+99=1,! 16! A33(! f9=,4! =,;8>=:/+! I;;>-+mv<#! 0=>+S! <3=-80.,4./.738!+7!./S!931O+;!73.7!A33H+W2-+99=,4!+0:-51,=>!7.97+!2/.>1;+9!.-+!7.97+! :8;!2-14+,=71-9S!O3=>3!4=T+!-=9+!71!.7!/+.97!7O1!;=66+-+,7!.;8/7!7.97+!>+//!752+9S!:87!;1! ,17!>1,7-=:87+!71!7.97+!2.2=//.+!`"#"S!"##a(! ! .4343/'$;<'HI?<'IO'= ! ',! 6=93! .,;! .023=:=.,9S! O3+-+! 7.97+! :8;9! /.5! O=73=,! 73+! +2=73+/=80S! .,;! ,17! =,>/8;+;! =,! 2.2=//.+S! 0+9+,>350+! .,;! ,+8-./! >-+97! ;+-=T.7=T+9! =,! 4+,+-./S! .-+! ,17! 7318437!71!>1,7-=:87+!71!7.97+!:8;!;+T+/120+,7!`"#$a(!',;++;S!91!6.-!.//!L,1O,!9=4,./=,4! 01/+>8/+9! -+V8=-+;! 61-! 7.97+! :8;! ;+T+/120+,7! .-+! 2-1T=;+;! :5! 73+! +2=73+/=80! =79+/6! `"EDS! "#%a(! ',! >1,7-.97! =,! 0.00./9S! 98:+2=73+/=./! 0+9+,>350+! 9+>-+79! 9+T+-./! 9=4,./9! 98>3!.9!N46!.,;!N1//=97.7=,!,+>+99.-5!61-!7.97+!:8;!;+T+/120+,7!.,;!73+-+61-+!./731843! =7!;1+9!,17!4=T+!-=9+!71!7.97+!:8;!>+//9S!=7!=9!-+V8=-+;!61-!73+=-!;+T+/120+,7(! ! $C! INTRODUCTION ! ',!6=93S!7.97+!9+,91-5!4.,4/=.!2-1>+99+9!-+.>3!73+!;+T+/12=,4!7.97+!:8;9!.67+-!73+! 61-0.7=1,! 16! <52+! '''! @&Hh<)! >+//9S! 73+! +.-/=+97! 61-0=,4! 7.97+! :8;! >+//9! 73.7! 3.T+! :++,! =;+,7=6=+;!91!6.-!@I257'B*!-,!*7#^ 53.5&7'+;-4!1-+57,+)(!<3=9!1:9+-T.7=1,!9844+979!73.7!7.97+! :8;! 61-0.7=1,! =,! 73+9+! 92+>=+9! =9! =,;+2+,;+,7! 16! 73+! 4.,4/=.(! ',! 73=9! -+92+>7S! +>712=>! +,;1;+-0./! 4-.679! =,! 73+! .W1/17/! 4=T+! -=9+! 71! 7.97+! :8;9! O=73187! 73+! 2-+9+,>+! 16! =,,+-T.7=1,! `"#&a(! ',! 0=>+S! =,7.>7! =,,+-T.7=1,! =9! ,17! -+V8=-+;! 61-! 7.97+! 2/.>1;+9! 61-0.7=1,!1-!7.97+!:8;!=,;8>7=1,!:87!=7!=9!,+>+99.-5!61-!7.97+!:8;!;=66+-+,7=.7=1,!;8-=,4! 73+!6=-97!2197,.7./!O++L(!',!2.-7=>8/.-S!=6!73+!,+-T+!=9!=,g8-+;!:87!.//1O+;!71!-+=,,+-T.7+! 73+!>=->80T.//.7+!2.2=//.S!73+!,80:+-!16!7.97+!:8;9!=9!2+-0.,+,7/5 ;=0=,=93+;`"#?H"#Da(! ! .434U/'$>CB<'DEF'JHIT ! <3+!7.97+!:8;!>+//9!=,!0.00./9!3.T+!.!T+-5!931-7!3./6H/=6+!=,!73+!/1>./!+2=73+/=80]! 61-!2-+95,.27=>!7.97+!>+//9!=9!.22-1W=0.7+/5!##!;.59!.,;!61-!7.97+!-+>+271-!>+//9S!D!;.59(! <3+-+61-+S! 73+! 61-0.7=1,! 16! 7.97+! :8;! >+//9! =9! .! -.73+-! -.2=;! 2-1>+99! 71! +66=>=+,7/5! >102+,9.7+!73+=-!/199(!G=717=>.//5!.>7=T+!>+//9!3.T+!:++,!1:9+-T+;!O=73=,!73+!:.9./!2.-7! 16!73+!+2=73+/=80S!=,!2-1W=0=75!1-!.;g.>+,7!71!73+!2.2=//.!.,;!71!.!/+99+-!+W7+,7!O=73=,! 73+!7.97+!:8;9!73+09+/T+9!`""$a(!!! *=T+,! 73+! ;=T+-9+! /1>./=9.7=1,! 16! 73+9+! >+//9S! 73+! 6=-97! V8+97=1,! -.=9+;! =9! O3+73+-! 73+5! >1,7-=:87+! 71! 7.97+! :8;! >+//9S! 71! 2.2=//.! >+//9! 1-! 71! L+-.7=,1>57+9! 16! 73+! 71,48+(! F+-+.HG.-7=,+Q! +7! ./! +T./8.7+;! 73+! =,>1-21-.7=1,! 16! 73+! ,+O/5! :1-,! >+//9! =,71! 73+! >=->80T.//.7+! 2.2=//.! 16! .;8/7! 0=>+! :5! 89=,4! &H+735/,=/H#cH;+1W58-=;=,+! @m;f)S! .! 3=43/5! 9+,9=7=T+!7350=;=,+!.,./148+(!U67+-!=,g+>7=1,S!0197!m;f!=,>1-21-.7+;!>+//9!O+-+!618,;!=,! 73+!+2=73+/=80!1879=;+!73+!7.97+!:8;9(!n,/5!.!6+O!m;f!>+//9!O+-+!618,;!O=73=,!73+!7.97+! :8;!@#E!>+//9!=,!##?!7.97+!:8;9S!%3!2197!=,g+>7=1,)(!<3+!0.g1-=75!16!m;f!>+//9!.22+.-+;! O=73=,!73+!7.97+!:8;!O=73!.!;+/.5!@"!;.5!1-!01-+)!.67+-!=,g+>7=1,(!<3+9+!;.7.!9844+97!73.7! 73+!0.g1-=75S!:87!,17!.//S!16!7.97+!:8;!2-14+,=71-9!.-+!/1>.7+;!=,!73+!+2=73+/=80!2-1W=0./! 71!73+!7.97+!:8;(`""$S!"#Ra!@6=4('H"$)(!! ! $D! INTRODUCTION ! 9@TEH<'.]234<.97+!2-14+,=71-!>+//9!/.:+//+;!O=73!m;f!@UHm)!1-!M-;f!@NS*)!1-!X4-&H462!@hS')!.-+!/1>.7+;!0.=,/5! 1879=;+!73+!7.97+!:8;(!! *+,+7=>!0.22=,4!.,./59=9!89=,4!.!7.01W=6+,!=,;8>=:/+!w+-.7=,H"%!B-+!-+>10:=,.9+! 931O+;! 73.7! :=217+,7=./! 2-14+,=71-9! /1>.7+;! .;g.>+,7! 71! 73+! 7.97+! :8;9! +W2-+99! A1W#! .,;!4+,+-.7+!:173!7.97+!:8;!.,;!L+-.7=,=Q+;!+2=73+/=./!>+//9(!I=73=,!73=9!2-14+,=71-!211/S! >+//9!16!73+!L+-.7=,1>57+!/=,+.4+!/19+!A1W#!+W2-+99=1,!O3+-+.9!>+//9!O=73!7.97+!:8;!6.7+! 82-+48/.7+!A1W#!`"$Ea(!B1,9=97+,7!O=73!73=9S!=,!73+!352101-23!A1W#m*NFJXF!+0:-519S! 6=/=61-0! 2.2=//.+! .-+! ,1-0./! :87! 73+! ,80:+-! 16! 68,4=61-0! .,;! 2./.7./! 7.97+! :8;9! =9! 4-.;8.//5!-+;8>+;!.,;!73+,!>102/+7+/5!.:9+,7!.7!:=-73S!=,;=>.7=,4!73.7!A1W#!.>7=T=75!=9! ,+>+99.-5!61-!7.97+!:8;!61-0.7=1,!`"$"a(! !N=,.//5S! /=,+.4+H7-.>=,4! +W2+-=0+,79! 89=,4! .,! =,;8>=:/+! X4-&! @/+8>=,+H-=>3! -+2+.7H >1,7.=,=,4!*H2-17+=,H>182/+;!-+>+271-)!B-+!L,1>L=,4!.//+/+!=,!>10:=,.7=1,!O=73!v19.#?H X.>|!.,;!v19.#?H7;<10.71!-+21-7+-!97-.=,9!931O+;!73.7!X4-&H+W2-+99=,4!>+//9!4.T+!-=9+! ! $R! INTRODUCTION 71!7.97+!:8;S!2.2=//.!.,;!9+/6H-+,+O=,4!>+//9!.7!73+!:17710!16!7-+,>3!.-+.9!.7!73+!:.9+!16! Bo!.,;!61/=.7+!2.2=//.+(! <3+!9+>1,;!V8+97=1,!O.9 O3+73+-!7.97+!:8;!2-14+,=71-!>+//9!-+2-+9+,7!.!8,=V8+! /=,+.4+!1-!O=//!4=T+!-=9+!71!01-+!73.,!1,+!>+//!752+9(!!f9=,4!.!6.7+!0.22=,4!.22-1.>3! :.9+;! 1,! 73+! A33>-+mv! 0189+! /=,+S! z++! +7! ./S! 931O+;! 73.7! A33H+W2-+99=,4! +0:-51,=>! 7.97+!2/.>1;+9!.-+!7.97+ :8;!2-14+,=71-9S!O3=>3!4=T+!-=9+!71!.7!/+.97!7O1!;=66+-+,7!.;8/7! 7.97+!>+//!752+9!@<52+!'!.,;!'')(!X4-&H+W2-+99=,4!>+//9!>1H+W2-+99!0.-L+-9!16!<52+!'S!''!.,;! '''!@6=4(!'H"$)!`"$#a(!<3+-+61-+S!.!211/!16!2-14+,=71-!+2=73+/=./!>+//9!4+,+-.7+9!.//!7.97+!:8;! >+//!752+9!:87!73+=-!>/1,./!-+/.7=1,93=2S!=6!.,5S!-+0.=,9!8,L,1O,( .434:/',I? ' .434:42/'^ABcD]L>B ! <3+! I,7JtH>.7+,=,! 9=4,./=,4! 2.73O.5! -+48/.7+9! >+//! 6.7+S! >+//! 2-1/=6+-.7=1,! .,;! 9+/6H-+,+O./!16!97+0!.,;!2-14+,=71-!>+//9!=,!0.,5!7=998+9!`"$$a(!',!:-=+6S!=,!73+!.:9+,>+! 16! I,7! /=4.,;9S! tH>.7+,=,! =9! ;+4-.;+;! .67+-! 2319231-5/.7=1,! :5! L=,.9+! .>7=T=75! 16! >5719L+/+7./HL=,.9+!>102/+W!73.7!>102-=9+9!>.9+=,!L=,.9+!".!@Bw".)S!4/8>14+,!95,73.9+! L=,.9+!$!@*Aw$)S!UW=,!.,;!.;+,10.7189!21/5219=9!>1/=!@UFB)(!f21,!I,7!/=4.,;!:=,;=,4!71! N-=QQ/+;! -+>+271-! @9+T+,! 7-.,90+0:-.,+! ;10.=,H*BFv)S! 73+! ;+4-.;.7=1,! >102/+W! :+>10+9!=,.>7=T+S!tH>.7+,=,!=9!.>>808/.7+;!=,!73+!,8>/+89S!:=,;9!1,!/50231=;!+,3.,>+-H :=,;=,4!6.>71-!@XmN)!.,;! ! *+,+7=>! .:/.7=1,! 16! tH>.7+,=,! =,! 73+! /=,48./! +2=73+/=80! -+98/79! =,! /199! 16! 7.97+! 2/.>1;+9(! ',! >1,7-.97S! O3+,! tH>.7+,=,! =9! >1,;=7=1,.//5! .>7=T.7+;S! 73+! +,7=-+! 71,48+! 98-6.>+! =9! 7-.,961-0+;! =,71! .,! .--.5! 16! 7.97+! :8;! 2-+>8-91-9! @A33y)(! <3+-+61-+S! I,7J:+7.H>.7+,=,! .>7=T=75! =9! ,+>+99.-5! .,;! 9866=>=+,7! 61-! 7.97+! 2/.>1;+! 61-0.7=1,(! <3+! ,.78-+! 16! I,7! /=4.,;9! 73.7! -+48/.7+! 73=9! 2-1>+99! -+0.=,9! 8,>/+.-(! I,7"E:S! ./731843! +W2-+99+;!=,!7.97+!:8;!2/.>1;+9S!=9!,17!-+V8=-+;!61-!73+=-!61-0.7=1,S!9=,>+!2/.>1;+9!.-+! ! %E! INTRODUCTION =,7.>7!=,!I,7"E:HJH!0=>+S!9844+97=,4!73.7!173+-!/=4.,;9!9318/;!:+!=,T1/T+;(!',!.;;=7=1,! 71!+;;:!I,7JtH>.7+,=,!9=4,./=,4!2-1017+9!+2]%!+W2-+99=1,!=,!73+!/=,48./!+2=73+/=80S!.! 7-.,9>-=27=1,!6.>71-!+W2-+99+;!=,!7.97+!:8;!2-14+,=71-!>+//9!`""RS!"$%S!"$&a( ! .434:4G/'$>CB<'J?>LIF<'J>BB ! MGF9!.-+!.!/.-4+!98:4-182!16!9+>-+7+;!6.>71-9!16!73+!<*Nt!982+-6.0=/5!-+48/.7=,4! >+//!2-1/=6+-.7=1,S!;=66+-+,7=.7=1,S!.212719=9!.,;!0=4-.7=1,!`"$?H"$Da(!MGF9!.-+!L,1O,! 71!.>7!=,!1-4.,!2.77+-,=,4S!61-!+W.02/+S!.9!=,3=:=71-9!16!6+.73+-!2/.>1;+!61-0.7=1,!73.7! >1,7-=:87+! 71! 92.7=./! ;=97-=:87=1,! 16! 6+.73+-! :8;9! `"$RS! "%Ea(! MGF9! #S! %S! .,;! C! .-+! +W2-+99+;!=,!73+!;+T+/12=,4!71,48+!+2=73+/=80!=,!.!92.7=17+021-./!2.77+-,!>1,9=97+,7! O=73! .! -1/+! =,! 7.97+! 2/.>1;+! =,;8>7=1,! .,;! 2.77+-,=,4(! <3+=-! =,3=:=71-S! 61//=97.7=,! =9! 9+>-+7+;! :5! 73+! .;g.>+,7! 0+9+,>350+! `"%"H"%%a(! U01,4! 73+! MGF! .,7.41,=979! .-+! ,144=,!.,;!61//=97.7=,(!P144=,!:=,;9!MGF#S!%!.,;!C!O=73!3=43!.66=,=75!O3+-+.9!61//=97.7=,! :=,;9!.>7=T=,!O=73!3=43!.66=,=75!.,;!MGF9!O=73!/1O+-!.66=,=75!`"$?S!"$Ca(!<3+!=,3=:=71-5! 0+>3.,=90! 16! ,144=,! =9! ;=66+-+,7! 6-10! 73.7! 16! 61//=97.7=,S! 1-! >31-;=,S! .,173+-! MGF! .,7.41,=97!`"$?S!"%&a(!! ! ',!697HJH!0=>+S!73+!92.7=./!2.77+-,=,4!16!7.97+!2.2=//.+!=9!/197!.9!-+T+./+;!:5!./7+-+;! A33!+W2-+99=1,(!<3+!/199!16!73+!.,7.41,=97!0+,!=,!73+!0+9+,>350+!-+98/79!=,!=,>-+.9+;! MGFC! +W2-+99=1,! .,;! 9=4,./=,4! O=73=,! 73+! +2=73+/=80(! <3+-+61-+S! 73+! =,7+-.>7=1,! :+7O++,!+2=73+/=./!MGFC!.,;!0+9+,>350./!NA1--+>7!2.77+-,=,4!16! 7.97+!:8;!2/.>1;+9(!<3=9!=,7+-.>7=1,!0897!:+!1>>8--=,4!;8-=,4!.!2.-7=>8/.-!7=0+!O=,;1O! 16! 7.97+! :8;! 2/.>1;+! =,;8>7=1,(! ',;++;S! =,>8:.7=1,! 1-! :+.;9! O=73! MGF9! 1,! 71,48+! +W2/.,79!-+98/79!=,!=,>-+.9+;!1-!;+>-+.9+;!,80:+-!16!2/.>1;+9S!;+2+,;=,4!O3+73+-!73+! 7-+.70+,7! 1>>8--+;! :+61-+! 1-! .67+-! 73+! 97.4+! 16! 2/.>1;+! =,;8>7=1,(! ',! 73+! 9.0+! /=,+S! 61//=97.7=,!71!>8/78-+;!71,48+!+W2/.,79!;1+9!,17!-+2-+99!7.97+!2/.>1;+!;+T+/120+,7!`"$&S! "%?a(!! .434:43/'&;;'>AF'B>CB<'J?>LIF<'OIH=>B@IA! ! A33!+W2-+99=1,!=9!=,=7=.//5!:-1.;!73-1843187!73+!;+T+/12=,4!71,48+!+2=73+/=80S! :87!=79!+W2-+99=1,!>1,91/=;.7+9!=,!7.97+!2/.>1;+9!.9!73+5!61-0(!A33!=9!.01,4!73+!+.-/=+97! 0.-L+-9!+W2-+99+;!=,!7.97+!:8;!2/.>1;+9(!F7>3"S!.!A33!-+>+271-!.,;!7.-4+7S!.,;!*/="S!.! ! %"! INTRODUCTION ;1O,97-+.0! 7.-4+7! 4+,+S! .-+! /=L+O=9+! +W2-+99+;! +.-/5! =,! :173! +2=73+/=./! .,;! 0+9+,>350./!>+//9!16!73+!71,48+(!I3+,!71,48+!+W2/.,79!.-+!7.L+,!2-=1-!71!61-0.7=1,!16! 7.97+!2/.>1;+9 .,;!>8/78-+;!=,!73+!2-+9+,>+!16!.!68,>7=1,H:/1>L=,4!.,7=:1;5!.4.=,97!A33! `"%$a!1-!O=73!.!A33!2.73O.5!=,3=:=71-!`"%CaS!68,4=61-0!7.97+!2/.>1;+9!=,>-+.9+!=,!:173! 9=Q+!.,;!,80:+-(!G1-+1T+-S!O3+,!A33!9=4,./=,4!=9!=,3=:=7+;S!7.97+!2/.>1;+9!61-0!=,!.! 2197+-=1-!;10.=,!16!73+!71,48+!98-6.>+S!O3=>3S!=,!>1,7-1/!7=998+!.,;!=,7.>7!.,=0./9S!=9! ./O.59! ;+T1=;! 16! 7.97+! :8;9(! B1,9=97+,7! O=73! 73=9S! -+2-+99=T+! 68,>7=1,! 16! A33S! O3+,! 71,48+! +W2/.,79! .-+! >8/78-+;! O=73! +W>+99! A33! /=4.,;S! -+98/79! =,! ;-.0.7=>.//5! -+;8>+;! 7.97+!2/.>1;+9!`"%Da(! ! 9@TEH<' .]2U4' I,7JtH>.7+,=,! 9=4,./=,4! =9! -+V8=-+;! 8297-+.0! 16! A33! .,;! M02%! 61-! 7.97+! 2/.>1;+! 92+>=6=>.7=1,(!A33!-+2-+99+9!I,7!.,;!M02%S!.9!O+//!.9!=79!1O,!+W2-+99=1,S!O3=/+!M02%S!.7!/+.97!+.-/5!1,S!=9! .! 219=7=T+! -+48/.71-! 16! A33(! M/.>L! .--1O9! }! 219=7=T+! -+48/.7=1,]! -+;! /=,+9! }! ,+4.7=T+! -+48/.7=1,(! r.93+;! /=,+9!=,;=>.7+!73.7!73+!I,7!2.73O.5!2-1017+9S!O3=/+!A33!=,3=:=79!7.97+!2/.>1;+!61-0.7=1,(! ! ',7+-+97=,4/5S!O3+,!A33!68,>7=1,!=9!=,3=:=7+;S!A33!+W2-+99=1,!=9!+W2.,;+;!=,!73+! 01-+!,80+-189!.,;!+,/.-4+;!7.97+!2/.>1;+9!`"%$aS!=,;=>.7=,4!73.7!A33c9!=02.>7!1,!=79! 1O,! +W2-+99=1,! =9! =,;=-+>7(! n,+! >.,;=;.7+! 61-! 73=9! =,7+-0+;=.-5! =9! 73+! I,7JtH>.7+,=,! 2.73O.5(! A2+>=6=>.//5S! tH>.7+,=,! =9! -+V8=-+;! 61-! A33! +W2-+99=1,! :5! 7.97+! 2/.>1;+! >+//9S! 2/.>=,4!A33!;1O,97-+.0!16!I,7!`"%Ra(!h1O+T+-S!I,7!9=4,./=,4!=,!7.97+!2/.>1;+9!=9!./91! ,+4.7=T+/5! -+48/.7+;! :5! A33]! =,! T=7-1! =,3=:=7=1,! 16! A33! -+98/79! =,! +W2.,;+;! 2/.>1;+H 92+>=6=>!I,7!.>7=T=75S!=,!.,!=,>-+.9+;!,80:+-!16!7.97+!2/.>1;+9S!O3+-+.9!+W>+99!16!A33! .>7=T=75!3.9!73+!12219=7+!+66+>7!@6=4(!'H"%)!`"%Da(!v+0.-L.:/5S!A33!2/.59!.,!12219=7+!-1/+! =,!7.97+!:8;!61-0.7=1,!=,!73+!.;8/7(! '7!O.9!-+>+,7/5!931O,!73.7!0=9+W2-+99=1,!16!Ahh! 7-.,961-09!.;8/7!/=,48./!+2=73+/=./!>+//!6.7+S!98>3!73.7!;.8437+-!>+//9!16!/=,48./!+2=73+/=./! ! %#! INTRODUCTION 2-14+,=71-9! 61-0! >+//! 752+H-+2/+7+S! 1,=1,H93.2+;! 7.97+! :8;9S! -.73+-! 73.,! ,1,H7.97+S! 29+8;197-.7=6=+;!+2=73+/=80(!<3+9+!AhhH=,;8>+;!+>712=>!7.97+!:8;9!>1,7.=,!.//!>+//!752+9! .,;!.-+!618,;!=,!-+4=1,9!16!73+!.;8/7!71,48+!2-+T=189/5!7318437!=,>.2.:/+!16!4+,+-.7=,4! 7.97+!1-4.,9!`"%Ra(!! 9@TEH<' .]2:4! <.97+! :8;! =,;8>7=1,! =,! 0.00./9(! N46! .,;! =79! 921875! =,3=:=71-9! >1,7-1/! 73+! ,80:+-! 16! 73+! 2-14+,=71-!>+//9(!A1W#!=9!.!L+5!01/+>8/+!61-!=,;8>=,4!73+!+2=73+/=80!=,71!.!7.97+!:8;!6.7+(!f297-+.0!16! A1W#!=9!I,7JtH>.7+,=,(!I,7!9=4,./=,4!=9!=,3=:=7+;!:5!MGF!.,;!A33!.,;!7389!2.77+-,!73+!7.97+!:8;9!=,!73+! /=,48./!+2=73+/=80(! ! ' .434S/'X ' .434S42/'9+9'J>B;N>K'A ' N46!9=4,./=,4!=9!.>7=T.7+;!O3+,!7O1!3+2.-.,!98/6.7+H>1,,+>7+;!N46!/=4.,;9!:=,;!71! +W7-.>+//8/.-!;10.=,9!16!75-19=,+!L=,.9+!-+>+271-9!`"&Ea!.,;!.>7=T.7+!0.,5!=,7-.>+//8/.-! >.9>.;+9!98>3!.9!v.9Jv.6Jm-L"J#S!FXB4.00.JFwB!.,;!F'$wJUw<(!U01,4!73+!N46!/=4.,;9S! N46"E! =9! +W2-+99+;! =,! 73+! 0189+! 71,48+! 0+9+,>350+! .,;! N46D.! =,! 73+! Q+:-.6=93! 1-123.-5,4+./!+2=73+/=80!.,;!7.97+!:8;!>+//9(!U01,4!73+!-+>+271-9S!N46-#:!.,;!N46-">! .-+!+W2-+99+;!=,!73+!0189+!71,48+!.7!m""!71!m"$!.,;!0@01$:!0@017$*:!0@017$&!.,;!0@01%!.-+! +W2-+99+;!=,!73+!Q+:-.6=93!23.-5,4+./!.->3!+2=73+/=80!.7!#;26!`"&"H"&&a(! ! %$! INTRODUCTION N46!=,;8>+!73+!+W2-+99=1,!16!618-!A2-1875!2-17+=,9!73.7!:/1>L!73+!75-19=,+!L=,.9+! .>7=T=75! 16! 73+! N46! -+>+271-9(! ',! A2-5#HJH! 0=>+! 73+! ,80:+-! 16! .,7+-=1-! 7.97+! 2/.>1;+9! @68,4=61-0)!=9!-+;8>+;!O3=/+!=,!73+!2197+-=1-!2/.>1;+!@>=->80T.//.7+)!=9!:=44+-!.,;!73+! ,80:+-!16!7.97+!2.2=//.+!=9!=,>-+.9+;(!B1,T+-9+/5S!=,!646"EHJH!0=>+!73+!2197+-=1-!2/.>1;+! =9! .:9+,7! .,;! =,! 92-5#HJH]646"EHJH! 0=>+! 73+! 2197+-=1-! 7.97+! :8;! 2/.>1;+! 61-0.7=1,! =9! -+971-+;( <3+-+61-+S!N46!9=4,./=,4!-+48/.7+9!73+!61-0.7=1,!16!7.97+!:8;9!:87!;=66+-+,7=.//5! ./1,4!73+!.,7+-12197+-=1-!.W=9!`"&?a(!! !',! .;;=7=1,! 71! 73=9! +.-/5! -+V8=-+0+,7! 61-! N46! 9=4,./=,4! ;8-=,4! 7.97+! :8;! 61-0.7=1,S! N46! -+48/.7+9! 73+! >+//! 6.7+! 16! 7.97+! :8;! >+//9(! <=0+H>1,7-1//+;S! 7.-4+7+;! =,.>7=T.7=1,!16!N46!-+>+271-!9=4,./=,4!O=73=,!73+!23.-5,4+./!+2=73+/=80!16!73+!Q+:-.6=93! /.-T.!-+98/79!=,!9+T+-+!-+;8>7=1,!16!<52+!''!>+//9!931O=,4!73.7!N46!9=4,./=,4!=9!-+V8=-+;!61-! 73+!61-0.7=1,!16!73=9!>+//!752+!`"EDa(! .434S4G/'#IBL;'C@TA>?@AT'J>B;N>K'FEH@AT'B>CB<'DEF'LB@IA' f21,!/=4.,;!:=,;=,4!1,!P17>3!-+>+271-S!73=9!7-.,90+0:-.,+!2-17+=,!=9!>/+.T+;! :5! 4.00.! 9+>-+7.9+S! .,;! 73+! =,7-.>+//8/.-! 2.-7! 16! =7! =9! 7-.,9/1>.7+;! 71! 73+! ,8>/+89S! .>7=T.7+9! 73+! BAX! 7-.,9>-=27=1,! 6.>71-! .,;! -+48/.7+9! 7.-4+7! 4+,+! +W2-+99=1,(! P17>3! 9=4,./=,4!.>7=T.7+9!O=73=,!73+!>+//!73.7!+W2-+99+9!73=9!2-17+=,S!73+!h.=-5!+,3.,>+-!16!92/=7! @h+9)! 7-.,9>-=27=1,! 6.>71-9S! O3=>3! =,3=:=7! 2-1,+8-./! 4+,+9! 98>3! .9! .>3.+7+H9>87+! 1-731/149! ! @G.93! =,! 0189+S! U9>/! =,! 6=93)! 73.7! 2-1017+! ,+8-1,./! 6.7+! =,! 73+! >+//9! 73.7! +W2-+99! P17>3! /=4.,;9! `"&CS! "&Da(! P17>3! /=4.,;9! .-+! 8:=V8=7=,.7+;! :5! v'P*H752+! m$! /=4.9+9!=,>/8;=,4!G=,;:10:!.,;!4+7!+,;1>5719+;#!!! ! <3+! -1/+! 16! P17>3! 9=4,./=,4! ;8-=,4! 7.97+! :8;! ;+T+/120+,7! =9! ,17! >102/+7+/5! +/8>=;.7+;(! U01,4! P17>3! 2.73O.5! 0+0:+-9S! h+9"! :=,;9! 71! 2-1017+-9! 16! :173! F/>t#! .,;!'2$-$!@:173!+W2-+99+;!:5!<52+!''!7.97+!-+>+271-!>+//9)S!O3=/+!h+9"!2-17+=,!=9!/1>./=Q+;! =,!73+!,8>/+89!16!8,;=66+-+,7=.7+;!7.97+!:8;!>+//9(!<3+-+61-+S!n7.!.,;!>1//+.48+9!`"&Ra! 2-1219+;! 73.7! h+9"! 0=437! -+2-+99! +W2-+99=1,! 16! 7.97+! >+//! 92+>=6=>! 01/+>8/+9! =,! 2-+>8-91-!>+//9(!',;++;S!=,!h+9"H,8//!0=>+S!73+!,80:+-!16!<52+!''J7.97+!-+>+271-!>+//9!=9! =,>-+.9+;!&H61/;!>102.-+;!71!>1,7-1/9!9844+97=,4!73.7!=,!73+!.:9+,>+!16!h+9"S!.:+--.,7! 7-.,9>-=27=1,!16!7.97+!-+>+271-H92+>=6=>!4+,+9!7.L+9!2/.>+!=,!2-+>8-91-!>+//9(! ! %%! INTRODUCTION ! I3+,! P17>3! .>7=T=75! =9! :/1>L+;! =,! G=,;:10:! Q+:-.6=93! 087.,79S! +W>+99! A+-171,=,H+W2-+99=,4!7.97+!>+//9!;+T+/12S!73+!7.97+!2-14+,=71-!>+//!211/!=9!-+;8>+;!.,;! 73+! ,80:+-! 16! B./-+7=,=,H+W2-+99=,4! 7.97+! -+>+271-! >+//9! =9! ;+>-+.9+;! `"EDa(! ',! .! >102/+0+,7.-5! 0.,,+-S! P17>3! .>7=T.7=1,! g897! 2-=1-! 71! >+//! ;=66+-+,7=.7=1,! 3.9! 73+! 12219=7+! +66+>7]! A+-171,=,H+W2-+99=,4! >+//9! .-+! .:9+,7! O3=/+! 73+! B./-+7=,=,H+W2-+99=,4! 7.97+!-+>+271-!>+//!2128/.7=1,!=9!0.=,7.=,+;!:87!,17!=,>-+.9+;(!<3+9+!-+98/79!98221-7!73+! 352173+9=9! 73.7! >+//9! O=73! .>7=T.7+;! P17>3! 9=4,./=,4! 0.=,7.=,! .! 2-14+,=71-! 97.7+! .9! 2-+;=>7+;!:5!73+!0189+!01;+/!=,!O3=>3!h+9"!.,;!73+-+61-+!=79!P17>3H-+2-+99=,4!.>7=T=75! =9! /197! `"&Ra(! B1,9=97+,7! O=73! 73+! -1/+! 16! P17>3! =,! 73+! 61-0.7=1,J0.=,7+,.,>+! 16! .! 2-14+,=71-!211/!61-!7.97+!:8;!>+//9S!73+!9+T+-+!;+>-+.9+!16!7.97+!-+>+271-!>+//9!2-1T1L+;! :5!>102-10=9+;!N46!9=4,./=,4!=,!6=93!=9!-+971-+;!:5!P17>3!.>7=T.7=1,!`"EDa(!! ',! 0189+! .,;! Q+:-.6=93S! G*+;$! .,;! *+/7$.! .-+! 92+>=6=>.//5! +W2-+99+;! =,! &Hh+//9S!-+92+>7=T+/5(!G.93!,8//!0=>+!.,;!Q+:-.6=93!.9>/".HJH!087.,79!.-+!;+T1=;!16!&Hh+//9(!U/731843!=,!0=>+S!73+!&Hh+//!/199!=9!,17!.>>102.,=+;!O=73!.,5!2.-7=>8/.-!7.97+! :8;! 23+,1752+S! =,! Q+:-.6=93S! 73+-+! =9! 9/=437! ;=91-4.,=9.7=1,! .,;! =,>-+.9+! 16! 7.97+! :8;! <52+!''!>+//9!`"?Ea(! .434S43/',@HGWW'JHI=IB ! G=>-1vPU9! @0=vPU9)! .-+! 90.//S! ,1,H>1;=,4! vPU! 01/+>8/+9! ! @.:187! #EH##! ,8>/+17=;+9)!73.7!-+48/.7+!!4+,+!+W2-+99=1,!2197H7-.,9>-=27=1,.//5(!G1-+!6-+V8+,7/5S!73+5! :=,;!71!>102/+0+,7.-5!9+V8+,>+!1,!73+!$cf ! %&! INTRODUCTION ! 9@TEH<' .]2S(! @U)! A>3+0.7=>! -+2-+9+,7.7=1,9! 16! 73+! L+5! 9=4,./9! 73.7! .-+! -+V8=-+;! 61-! 73+! /.7+! 97+29! 16! ;+T+/120+,7!16!7.97+!-+>+271-!.,;!A+-171,=,H+W2-+99=,4!>+//9!16!73+!Q+:-.6=93!7.97+!:8;9(!N46!-+>+271-! @N46v)S!0=vH#EE!.,;!U9>/".!.>7=T=75!.-+!-+V8=-+;!O=73=,!23.-5,4+./!+2=73+/=./!>+//9!71!;-=T+!7.97+!-+>+271-! .,;!A+-171,=,!>+//!61-0.7=1,S!-+92+>7=T+/5(!U>7=T.7=1,!16!P17>3!9=4,.//=,4!=9!,+>+99.-5!61-!7.97+!-+>+271-! >+//!61-0.7=1,!O3+-+.9!=,3=:=7=1,!16!73=9!9=4,./!=9!-+V8=-+;!61-!A+-171,=,!>+//!61-0.7=1,(!P17>3!/=L+/5!3.9! .;;=7=1,./!68,>7=1,9!=,!7.97+!:8;!;+T+/120+,7!O=73=,!73+!23.-5,4+./!+2=73+/=80(!@M)!U!#~0!127=>./!9+>7=1,! 73-1843! .! O=/;H752+! Q+:-.6=93! 7.97+! :8;! -+T+./9! 7.97+! -+>+271-! >+//9! >1H+W2-+99=,4! B./-+7=,=,! @B./-)! .,;! 2/>t#(! ! .4U/'(HIY ! I1-L!1T+-!73+!/.97!7O1!;+>.;+9!3.9!93+;!/=437!=,!73+!01/+>8/.-!=,7+-.>7=1,9!73.7! .-+!-+V8=-+;!61-!7.97+!2/.>1;+!=,;8>7=1,!O=73=,!73+!1-123.-5,4+./!+2=73+/=80(!<3+-+!=9! ./91! =,>-+.9=,4! +T=;+,>+! .:187! 73+! 01/+>8/.-! =;+,7=75! 16! 2-14+,=71-! >+//9! .,;! ,17.:/5! A33! .,;! A1W#! .-+! >1,9=;+-+;! .9! 6.=7368/! 0.-L+-9! 16! +.-/5! ;+T+/12=,4! 7.97+! :8;! >+//9(! h1O+T+-S!=7!=9!8,>/+.-!O3+73+-!73+!+W2-+99=1,!16!73+9+!4+,+9!=9!0.=,7.=,+;!73-1843187! 73+! ;=66+-+,7=.7=1,! 7.97+! :8;! 2-1>+99(! ',;++;S! 73+! 01/+>8/+9! 73.7! -+48/.7+! 92+>=6=>.//5! 73+! ;=66+-+,7=.7=1,! 16! +.>3! 16! 73+! 73-++! 7.97+! :8;! >+//! 752+9S! 3.9! 1,/5! 97.-7+;! 71! :+! .;;-+99+;(! U,173+-! =,7-=48=,4! .92+>7! 16! 7.97+! :8;! ;+T+/120+,7! =9! 73+! >+//8/.-! =,7+-.>7=1,9!73.7!.-+!-+V8=-+;!71!.99+0:/+!;=66+-+,7=.7=,4!>+//9!=,71!.,!1-4.,(!G197!16! 73+!2-14+,=71-!>+//9!.-+!:.9./!.,;J1-!+W7+-,./!71!73+!61-0=,4!7.97+!:8;!1-4.,(!A=,>+!73+! ! %?! INTRODUCTION 7.97+!:8;!1-4.,!=9!.!68,>7=1,./!0=>-1>=->8=7S!1,+!O18/;!+W2+>7!73.7!>+//!219=7=1,!O=73=,! 73+!1-4.,!=9!,17!-.,;10(!',97+.;S!;=66+-+,7=.7=,4!>+//9!1,>+!:1-,S!73+5!0897!-+.>3!73+! 61-0=,4! 1-4.,! .,;! 73-1843! =,7+-.>7=1,9! O=73! 73+! 173+-! >+//9! 61-0! .,! .,.710=>./! .,;! 68,>7=1,./!8,=7(! ! r8-=,4!05 F3r!'!.;;-+99+;!2-+>=9+/5!73=9!V8+97=1,K!31O!;1!;=66+-+,7=.7=,4!7.97+! :8;!>+//9S!;+-=T+;!6-10!73+!1-123.-5,4+./!+2=73+/=80!.99+0:/+!=,71!.!68,>7=1,./!1-4.,(! <1! .>3=+T+! 73=9! 41./S! '! 9+/+>7+;! 6/81-+9>+,7! 7-.,94+,=>! /=,+9! O=73! 73+! 217+,7=./! 71! :+! +W2-+99+;!=,!+.-/5!;=66+-+,7=.7=,4!7.97+!:8;!>+//9(!M.9+;!1,!73+!1:9+-T.7=1,!16!73+!=,!T=T1! .,./59=9S!!'!>10:=,+;!73+9+!/=,+9!O=73!>+//8/.-!.,;!01/+>8/.-!/199!16!68,>7=1,!.22-1.>3+9S! 71!68-73+-!;=99+>7!73+!-1/+!16!73+!<52+!'''!>+//!752+!;8-=,4!7.97+!:8;!1-4.,!61-0.7=1,(! ! %C! ! MATERIALS and METHODS ! %D! MATERIALS AND METHODS ''..42/'Z ''<3+!6=93!O+-+!-.=9+;!=,!6-+93!O.7+-!=,!.!>1,97.,7!7+02+-.78-+!16!#DB!O=73!.!/=437! >5>/+!16!""3 16!;.-L,+99!.,;!"$3!16!/=437(!<3+!2-1;8>+;!+0:-519!O+-+ -.=9+;!.7!#DH$EB! =,!+0:-51!0+;=80!.>>1-;=,4!71!73+!;+9>-=:+;!>1,;=7=1,9!=,!73+!Z|+:-.6=93!:11L[!`"?$a(! <3+!7-.,94+,=>!/=,+9!89+;!O+-+!>1,97-8>7+;!=,!73+!/.:!1-!1:7.=,+;!73-1843!>1//.:1-.7=1,9(! ,@Z,.;$& X ' ..4G/'(; ' ''(#(")!',!9=78!35:-=;=Q.7=1,! I31/+!+0:-519!.-+!6=W+;!=,!%d!2.-.61/0.;+35;+!=,!F31923.7+!M866+-!A1;=80!@FMA)!61-! %D!318-9(!U67+-!2+-0+.:=/=9.7=1,!=,!0+73.,1/S!73+!+0:-519!8,;+-41!;=4+97=1,!=,!2-17+=,.9+! w!@"E~4J0X)(!P+W7!71!2197H6=W.7=1,!73+!+0:-519!.-+!=,>8:.7+;!=,!2-+H35:-=;=Q.7=1,!:866+-! @hzM%!K! AAB! &WS! 61-0.0=;+! &Ed!S! 71-8/.! 5+.97! vPU! @9=40.)! &04J0XS! 3+2.-=,+! &E~4J0XS! ''(#(#)!'008,13=971>3+0=97-5!! I31/+!+0:-519!O+-+!6=W+;!=,!;=66+-+,7!97.4+9!16!;+T+/120+,7!=,!.!6=W.7=1,!91/87=1,! @2.-.61-0./;+35;+ %dHFMA!"W)!61-!"!Å!1-!#!318-9(!U67+-!$!O.93+9!=,!FM<7-!73+!+0:-519! O+-+!=,>8:.7+;!=,!73+!:/1>L=,4!91/87=1,(!<3+!2-=0.-5!.,7=:1;=+9!O+-+K!sH*NF!@<1--+5!F=,+9! M=1/.:9S!"!K"EEES!-.7)S!sH>./-+7=,=,!@AO.,7!"!K"EEES!-.::=7!1-!0189+)S!sH&HhL71,! r=>L=,91,S! " K&EES! -.::=7)S! sH>.92.9+$! .>7=T+! @M+>L71,! r=>L=,91,S! "!K&EES!-.::=7)(!U!9+>1,;.-5!.,7=:1;5!>182/+;!71!.!6/81-1>3-10+!U/+W.N/81-!U%DDS!U&?DS! U?%C!@',T=7-14+,!"!K&EE)!O.9!89+;!71!;+7+>7!73+!2-=0.-5!.,7=:1;5(!<3+,!73+!+0:-519!O+-+! 2197!6=W+;S!73+!23.-5,W!O.9!;=99+>7+;S!018,7+;!=,!4/5>+-1/!.,;!1:9+-T+;!=,!:=,1>8/.-!1-! >1,61>./!0=>-19>12+(! ..43/'9@[ N/81-+9>+,7! =0.4+9! O+-+! 1:7.=,+;! :5! AF&! >1,61>./! 0=>-19>12+! 89=,4! %EW' 1=/! =00+-9=1,! 1:g+>7=T+9(! A+-=+9! 16! =0.4+9! O+-+! .>V8=-+;! .7! E(&! 00! =,7+-T./9(! '0.4+e! @P'h)! O.9!89+;!71!.,./59+!97.>L9S!.,;!U;1:+!F31719312J'//897-.71-!71!.;g897!:-=437,+99J>1,7-.97! .,;!018,7!=0.4+9(!<3-++H;=0+,9=1,./!-+>1,97-8>7=1,9!.,;!>+//!>18,7=,4!O+-+!>.--=+;!187! 89=,4!'0.-=9!@M=72/.,+)(! '0.4=,4! 16! +0:-519c! 01873! O.9! 2+-61-0+;! 1,! .! P=L1,! =,T+-7+;! 92=,,=,4! ;=9L! +V8=22+;!O=73!.,!+T1/T+!>.0+-.!@F31710+7-=>9)!.,;!G+7.01-23!9167O.-+(!<3+!+0:-519! O+-+! 018,7+;! =,! .! 0=W! 16! 2/.90.H73-10:=,+! =,! +0:-51! 0+;=80! O=73! F &E! MATERIALS AND METHODS .7! 73=9! .4+! .-+! 8,.T.=/.:/+(! <3+-+61-+S! 73+! 7=0+! 2+-=1;! >.,,17! :+! 2-+>=9+/5! >102.-+;! :+7O++,!01T=+9(!! U//!73+ .,./59=9!O.9 2+-61-0+; 1,!7=0+/.29+!16!+0:-519!97.4+;!:+7O++,!?E326!.,;! CE326!1,!;.7.!1:7.=,+;!6-10!=0.4+9!16!73+!6=-97!61-0=,4!7.97+!:8;!1,!73+!0197!.,7+-=1-! 0+;=.,!2.-7!16!73+!01873!.,;!=79!,+=43:18-=,4!@01-+!/.7+-./)!7.97+!:8;9(!A=0=/.-!V8./=7.7=T+! 23+,1752+9!O+-+!1:9+-T+;!1,!2./.7+!.,;!.,7+-=1-!0.,;=:8/.-!.->3(!h1O+T+-S!7=0+/.29+ 1,! 73+9+!.-+.9!=9!01-+!;+/=>.7+!.9!,1,!7.97+H:8;!-+/.7+;!+2=73+/=80!=,7+-6+-+9!O=73!=0.4=,4!16! 73+!2./.7+!.,;!*NF!9=4,./!6-10!73+!3+.-7!16!73+!,@Z,.;$&X ..4U/'$@=JC<'CB>B@CB@L>?'>A>?KC@C' <=0+/.29+! QH97.>L9! O+-+! .,./59+;! =,! '0.-=9! @M=72/.,+)(! r.7.! 1,! >+//! 1-! >+//! 4-182! 219=7=1,! @>+,7+-! 16! 0.99)! .,;! 92++;! =,=7=.//5! 1:7.=,+;! 73-1843! '0.-=9! O+-+! 0.,8.//5! T+-=6=+;S! >10:=,+;! O=73! >+//! >1,7.>7! 1:9+-T.7=1,! .,;! 73+,! 2-1>+99+;! =,! v! 71! >./>8/.7+! m8>/=;+.,! ;=97.,>+(! U//! 97.7=97=>./! .,./59+9! O+-+! 2+-61-0+;! =,! v! @3772KJJOOO(-H 2-1g+>7(1-4J)(! ..4:/'$NI]J;IBIA'?>C U//! +W2+-=0+,79! O+-+! 2+-61-0+;! O=73! .! >89710H:8=/7! -.,;10H.>>+99! 7O1H23171,! /.9+-H9>.,,=,4!0=>-19>12+!`"??a(!',!73=9!=,97-80+,7S!:173!p!.,;!z!9>.,,=,4!.-+!12+-.7+;!:5! .>18971H127=>!;+6/+>71-9!@Unr9)(!<3+9+!>89710H:8=/7!,1,0+>3.,=>./!:+.0H97++-=,4!;+T=>+9! @UHU!n271Hm/+>7-1,=>)!>.,!-+;=-+>7!73+!/.9+-!:+.0!=,!"E!~9(!<1!12+-.7+!73+!Unr9!.,;!-8,! 73+! 9>.,,=,4! 2-1>+;8-+9S! .! >89710H0.;+! 89+-! =,7+-6.>+! O.9! 2-14-.00+;! =,! X.:o=+O! @P.7=1,./!',97-80+,79)(!<3+!Unr!.>1897=>!6-+V8+,>5!;-=T+!O.9!4+,+-.7+;!:5!.!r=-+>7!r=4=7./! A5,73+9=Q+-! .,;! .! 6.97! @"E! ,9)! 21O+-! .02/=6=+-! @UHU! n271+/+>7-1,=>9)(! ! <3+! 0=>-19>12+! O.9! +V8=22+;! O=73! .! #&W! XfGF/.,NXJ'v! 1:g+>7=T+! O=73! E(R&! ,80+-=>./! .2+-78-+! @X+=>.0=>-19597+09)(! N/81-+9>+,>+! 23171,9! O+-+! ;+7+>7+;! :5! >11/+;! U9*.F! h"EC?RFUx%E! 2317108/7=2/=+-9! =,! 73+! 7-.,96/81-+9>+,>+! .,;! +2=6/81-+9>+,>+! &"! MATERIALS AND METHODS 2.73O.59S! O=73! ?%"HC&,0! :.,;2.99! 6=/7+-! @9+0-1>L)! .,;! .! &"EHD%,0! :.,;2.99! 6=/7+-! @A+0-1>L)!61-!:173!2.73O.59(!<3+!.:/.7=1,!O.9!2-1;8>+!=,!>1,6=,+!127=>./!-+91/87=1,!9217! @W5!$EE,0S!Q!"~0)!O=73!&EE0OHJy!"Ed!/.9+-21O+-!.,;!#EE~9!;O+//!7=0+(!<3+!/=T+!97.>L9 O+-+!=0.4+;!O=73!$!71!%!~9!;O+//7=0+!9>.,,=,4!.,;!#E0I!/.9+-!21O+-(! ' &#! RESULTS RESULTS ' .../')%&1*$&' ! ...42/'$;<'!"!#$%&'($')&*'@C'<[JH <1!4+7!=,9=437!=,71!7.97+!:8;!1-4.,!61-0.7=1,!O+!9+.->3+;!61-!-+48/.71-5!+/+0+,79! 73.7!;-=T+!-+21-7+-!+W2-+99=1,!.7!+.-/5!97+29!16!73+=-!;+T+/120+,7(!U9!0@0d*!=9!+W2-+99+;!=,! 73+!;+T+/12=,4!1-123.-5,4+./!+2=73+/=80!.,;!7.97+!:8;!>+//9!.,;!>1,7-=:87+9!71!7.97+!:8;! 61-0.7=1,!`"EDaS!O+!+W.0=,+;!O3+73+-!.,5!16!73+!L,1O,!`"?%a 0@0d* -+48/.71-5!+/+0+,79! ;-=T+9!+W2-+99=1,!=,!7.97+!:8;9(!I+!618,;!73.7!6-10!&&326!1,O.-;9!H!73.7!=9!2-=1-!.,;!;8-=,4! 7.97+!:8;!>+//!;=66+-+,7=.7=1,!H!73+!+,3.,>+-!,@Z0@0d*#-3;*3/-1X@0.(!;-=T+9!*NF!+W2-+99=1,!=,! 1-123.-5,4+./! +2=73+/=./! >+//9! 73.7! +W2-+99! +2]%:! .! 7-.,9>-=27=1,! 6.>71-! +W2-+99+;! =,! 2-+98027=T+!7.97+!:8;!>+//9!.,;!.;g.>+,7!+2=73+/=80!`"?Ca!@9@T(2>]L)(!! <1!68-73+-!>3.-.>7+-=Q+!73+!=;+,7=75!16!,@Z0@0d*#-3;*3/-1X@0.(!>+//9S!O+!.,./59+;!7.97+! :8;! -+/.7+;! 4+,+! +W2-+99=1,! .7! /.7+-! 97.4+9(! N-10! &DH?E326! 1,O.-;9S! O=73=,! 73+! 1-./! +2=73+/=80S! 4-1829! 16! >+//9! >1H+W2-+99! ,@Z0@0d*#-3;*3/-1X@0.(! .,;! F-1W"! 0.-L+-! 16! +.-/5! ;=66+-+,7=.7=,4!7.97+!:8;!>+//9!@-+6)!@N=4(";SgS2)(!N-10!?E326!1,O.-;9S!&Hh ' ...4G/'."/!"!#$%&'($')&*+"!,0'<[JH ! *=T+,! 73+! +.-/5! .,;! O=;+! ,@Z0@0d*#-3;*3/-1X@0.(! +W2-+99=1,! =,! 73+! 1-./! +2=73+/=80! .,;!7.97+!:8;!>+//9S!O+!89+;!73=9!7-.,94+,=>!/=,+!71!61//1O!=,!T=T1!7.97+!:8;!1-4.,!61-0.7=1,(! <1!;=97=,48=93!:+7O++,!9=7+9!O3+-+!7.97+!:8;!1-4.,9!61-0!.,;!73+!-+97!16!73+!+2=73+/=80S!O+! >10:=,+;! ,@Z0@0d*#-3;*3/-1X@0.(! O=73! ,@Z,.;$&X ! &%! RESULTS ',!T=T1!=0.4=,4!931O+;!73.7!,@Z0@0d*#-3;*3/-1X@0.(!=9!+W2-+99+;!=,!1-./!+2=73+/=./!>+//9! =,!.!;5,.0=>!0.,,+-(!U7!&&326S!+2=73+/=./!>+//9S!/1>.7+;!.7!73+!0197!.,7+-=1-!0+;=.,!2.-7!16! 73+!01873S!+W2-+99!/1O!.018,79!16!,@Z0@0d*#-3;*3/-1X@0.( @9@T(2>)# M5!?E326S!.7!;=97=,>7!9=7+9! @-197-1H0+;=.,!.,;!-197-1H/.7+-./!2.-7!16!73+!01873)S!*NF!+W2-+99=1,!=9!=,7+,9+!=,!>+//9!73.7! >1,7.>7!+.>3!173+-!.,;!61-0!.22.-+,7!4-1829!@9@T(!2D7',IQ@<'2)(!! I=73=,! .! 61-0=,4! ,@Z0@0d*#-3;*3/-1X@0.(! >+//! 4-182S! 1,+! ,@Z0@0d*#-3;*3/-1X@0.( >+//! 97.-79! +W2-+99=,4! ,@Z,.;$&X ' ...43/' ."/!"!#$%&'($')&*+"!,0- <[JH ! U9! 1-./! +2=73+/=./! ;+T+/120+,7! 2-1>++;9S! .;;=7=1,./! 97-1,4/5! +W2-+99=,4! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! .22+.-! =,! :+7O++,! 73+! 7.97+H:8;! 4-1829(! <3+9+! ,@Z0@0d*#-3;*3/-1X@0.(!!>+//9!O=73187!.,5!>1,7.>7!O=73!73+!,@Z,.;$&X ! &&! RESULTS U/714+73+-!73+!.,./59=9!16!7.97+!:8;!61-0.7=1,!=,!.!/=T+!T+-7+:-.7+!931O9!61-!73+!6=-97! 7=0+!73.7!7.97+!:8;!>+//9!.-+!017=/+(!,@Z0@0d*#-3;*3/-1X@0.(!>+//9!61-0!7.97+H:8;!4-1829!:87! ./91! 61-0! 1879=;+! 73+0S! O=73=,! 73+! 2-1W=0./! 1-./! +2=73+/=80S! .,;! 01T+! 71! g1=,! 73+! ;+T+/12=,4!7.97+!:8;9(!,@Z0@0d*#-3;*3/-1X@0.(!>+//9!.-+!0.=,7.=,+;!O=73=,!1-!V8=7!73+!7.97+! :8;S! ;+2+,;=,4! 1,! 73+=-! >1,7.>7! O=73! .! 95,73+7=9=,4! &Hh+//S!-+92+>7=T+/5(! ...4U/' $;<' :]5$' L@ABAL<' IO' ."/!"!#$%&'($')&*+"!,0' <[JH <1!.;;-+99!O3+73+-!.!,@Z0@0d*#-3;*3/-1X@0.( 1-!,@Z,.;$&X ',! .! >102/+0+,7.-5! 0.,,+-S! O+! .,./59+;! 73+! :+3.T=18-! 16! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9!=,!73+!*+/7$*?\?!Q+:-.6=93!73.7!.-+!;+T1=;!16!&Hh+//9!@`"EDa!9@T4'-6)(!<3+!,80:+-!16! ,@Z0@0d*#-3;*3/-1X@0.(!>+//9!=9!,17!9=4,=6=>.,7/5!;=66+-+,7!=,!*+/7$*?\?!/.-T.+!>102.-+;!71!73+! O=/;!752+!9=:/=,49!@9@T(!-0]%)(!,@Z0@0d*#-3;*3/-1X@0.(!>+//9!+W2-+99=,4!T.-=.:/+!/+T+/9!16!*NF! >1,7.>7!+.>3!173+-!.,;!61-0!4-1829!=,!*+/7$*?\?!/.-T.+!@,IQ@<'S)(!h1O+T+-S!=,!>1,7-.97!71! 73+!O=/;H752+!@,}EJ"?!>+//!4-1829S!&!+0:-519)S!*+/7$*?\?!,@Z0@0d*#-3;*3/-1X@0.(!>+//!4-1829! >.,!92/=7!=,71!7O1!.22-1W=0.7+/5!3./6x9=Q+;!>+//!4-1829!@,IQ@<'S7'"R73!9+>!1,O.-;9S!>+,7-./! 4-182S!'9@T4'2W7!,}&J"#!4-1829S!$!*+/7$*?\?!+0:-519S!0=,=0./!.,;!0.W=0./!;=97.,>+!:+7O++,! >+//! 4-1829S! $E("Rƒ0! .,;! "&E("ƒ0S! -+92+>7=T+/5)(! n>>.9=1,.//5S! >+//! 4-1829! >1,7.>7! +.>3! 173+-! @9@T' 2W4! ,}$J"#S! $! +0:-519S! ,IQ@<' S7! $C73H$R73! 9+>)(! <1! +W.0=,+! O3+73+-! 73+! ,@Z0@0d*#-3;*3/-1X@0.(! >+//! 4-182! 6-.40+,7.7=1,! 3.9! .,5! >1,9+V8+,>+! =,! 7.97+! :8;! 1-4.,! 61-0.7=1,!=,!*+/7$*?\?!!/.-T.+S!O+!.,./59+;!7.97+!:8;!0.-L+-!+W2-+99=1,!.7!&;26(!U/731843! 73+! ,80:+-! 16! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! =9! ,17! 9=4,=6=>.,7/5! ;=66+-+,7! :+7O++,! *+/7$*?\?! .,;! O=/;H752+! 9=:/=,49! .7! &;26S! 73+-+! =9! .! -+/.7=T+/5! 90.//! :87! 9=4,=6=>.,7! =,>-+.9+! 16! 73+! ,80:+-!16!7.97+!:8;9!=,!73+!.9>/".HJH!087.,7!>102.-+;!71!73+!O=/;H752+!9=:/=,49!@N@T4'-6]%)(!! ! &?! RESULTS N=,.//5S! O+! 0+.98-+;! ;=97.,>+! .,;! >1,7.>7! 7=0+! 2+-=1;9! :+7O++,! .;g.>+,7! >+//9! =,! *+/7$*?\?!/.-T.+#!',!73+!O=/;H752+S!73+!,.;$&!>+//!=9!73+!1,+!.7!73+!931-7+97!;=97.,>+!6-10!73+! >+,7+-! 16! 0.99! 16! 73+! 4-182! @,}"#J"#! 4-1829)(! <3+-+61-+S! O+! 89+;! 73+! ,@Z0@0d*#-3;*3/-1X@0.(!>+//!=,!73+!>1--+921,;=,4!219=7=1,!.9!.!-+6+-+,>+!61-!+.>3!>+//!4-182!=,! .9>/".HJH! 61-0=,4! 7.97+! :8;9(! N=-97S! .9! =,! 73+! O=/;H752+S! O+! 618,;! .;g.>+,7! ,@Z0@0d*#-3;*3/-1X@0.( >+//9!73.7!-+0.=,!=,!97.:/+!>1,7.>7!1T+-!7=0+!@;=97.,>+{#R("ƒ0S!9@T4' 22>)S! V8=7! 1-! g1=,! 73+! 4-182! @9@T4' 22D)(! h1O+T+-S! =,! >1,7-.97! 71! 73+! O=/;H752+S! O+! ./91! 1:9+-T+;! 73.7! ,+=43:18-=,4! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! 97.5! =,! 2-1W=0=75! 71! +.>3! 173+-! O=73187! L++2=,4! .! 2+-0.,+,7! >1,7.>7! :+7O++,! 73+0! 1T+-7=0+! .,;! O=73187! V8=77=,4! 73+! 4-182!@9@T4'22L)(!',!73+!/.77+-!>.9+S!73+!7=0+!2+-=1;!73.7!,@Z0@0d*#-3;*3/-1X@0.(!>+//9!-+0.=,! =,!>1,7.>7!71!+.>3!173+-!=9!>102.-.:/+!71!73+!O=/;H752+!@9@T4':)(!!! ',! >1,>/89=1,S! ./714+73+-! 73+! -+98/79! 931O! 73.7! .)! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! 61-0! 7.97+!:8;!1-4.,9S!:)!>.,!;=66+-+,7=.7+!=,71!&Hh+//9!>)!;+T+/12=,4!7.97+!:8;!>+//9!.-+!017=/+! .,;!>.,!:+!;=92/.>+;!6-10!1,+!1-4.,!71!.,173+-!;)!,@Z0@0d*#-3;*3/-1X@0.(H&Hh+//!>1,7.>7! -+98/79! =,! 73+! 0.=,7+,.,>+! 16! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! =,71! 4-1829! 61-! /1,4+-! 7=0+! 2+-=1;9(! ! ! ! ! ! ! ! &C! RESULTS ! ! ! ! &D! RESULTS ! ! ! &R! RESULTS ! ?E! RESULTS ! ! ?"! RESULTS ! ! ?#! RESULTS ! ! ?$! RESULTS ! N=48-+!C(!#H23171,!.:/.7=1,!16!0@0d*#-3;*3/-1X@0.!>+//!931O,!=,!@:)!=,!! .! ,@@0@0d*#-3;*3/-1X@0.(^,@Z,.;$&X ! ?%! RESULTS ! ! ! ! ?&! RESULTS ! ' ! ??! RESULTS ' ! ?C! RESULTS ! ?D! RESULTS ' ' ' ! ?R! RESULTS ! ! ! ! CE! RESULTS ! ! ! ! C"! RESULTS ! ! ! ! C#! RESULTS ! ! ! ! C$! RESULTS ! ! ! ! C%! RESULTS ! ! ! ! C&! ! DISCUSSION ! C?! DISCUSSION ! N1-! 73+! 6=-97! 7=0+! 73+! V8+97=1,! 16! .99+0:/5! 16! +2=73+/=./! ;=66+-+,7=.7=,4! >+//9! =,71! .! 7.97+! :8;! 1-4.,! =9! .;;-+99+;! =,! 73=9! 978;5(! f9=,4! Q+:-.6=93! 6/81-+9>+,7! 7-.,94+,=>! .,;! 087.,7!/=,+9!.,;!7O1H23171,!/.9+-!.:/.7=1,S!O+!618,;!73.7!;+T+/12=,4!7.97+!:8;!>+//9!.-+! 017=/+! .,;! 01T+! 6-10! 1,+! ;+T+/12=,4! 1-4.,! 71! .,173+-(! n,+! 16! 73+! 6.>71-9! -+48/.7=,4! 017=/=75!=9!>+//H71H>+//!=,7+-.>7=1,!O=73=,!73+!1-4.,(!',!2.-7=>8/.-S!73+!>+//!>1,7.>7!O=73!73+! ,.;$&J&Hh+//!=9!-+V8=-+;!61-!73+!/1,4H7+-0 0.=,7+,.,>+!16!73+!;+T+/12=,4!>+//9!=,71!73+! 61-0=,4!7.97+!:8;(! n,+!16!73+!=998+9!16!73=9!978;5!=9!73+!V8.,7=6=>.7=1,!.,./59=9!16!73=9!23+,10+,1,(! M+>.89+! 16! =79! >102/+W=75! .,;! 73+! 8,+W2+>7+;! ,1,H97+-+1752+; +.-/5! 7.97+! :8;! ;+T+/120+,7! =,! 6=93S! 73+! 7-.>L=,4! 16! 73+! >+//! 219=7=1,! 3.9! 71! :+! T+-=6=+;! .,;! .;g897+;! 0.,8.//5(! N1-! 73=9! -+.91,S! 3+-+! .-+! 2-1T=;+;! V8.,7=7.7=T+! ;.7.! 61-! >+//! .,;! 7.97+! :8;! ;=97.,>+!6-10!&!187!16!"E!O=/;H752+!,@Z0@0d*#-3;*3/-1X@0.(^!,@Z,.;$&X ' .R42/' ."/!"!#$%&'($')&*+"!,01- ."/.,(23+45(&**60! ' M' >A' >JJHIJH@>B<' BII?' BI' >A>?KC<' B>CB<'DEF'F ! <3+!.99+0:/5!16!;=66+-+,7=.7=,4!>+//9!=,71!=,7.>7!7.97+!:8;!1-4.,9!3.9!:++,!8,+W2/1-+;! 0.=,/5! ;8+! 71! 73+! =,.>>+99=:=/=75! 16! 73+! 1-123.-5,W! ;8-=,4! 0189+! +0:-514+,+9=9(! <3+! Q+:-.6=93!01873!2-1T=;+9!.!411;!01;+/!71!.;;-+99!73=9!V8+97=1,!;8+!71!=79!.>>+99=:=/=75!.,;! 127=>./! >/.-=75(! h+-+S! 7O1! 7-.,94+,=>! /=,+9! ,@Z0@0d*#-3;*3/-1X@0.(! ! .,;! ,@Z,.;$&X ! CC! DISCUSSION +W2-+99=,4! >+//9! >.,! +W2-+99! 0.-L+-9! 16! .//! 7.97+! :8;! >+//! 752+9]! 73+9+! 0.-L+-9! .-+! ./91! +W2-+99+;! =,! >+//9! ;+T1=;! 16! ! ! ,@Z0@0d*#-3;*3/-1X@0.(! ! +W2-+99=1,! @=(+(! N=4(! "+S3)(! ! <3+! 9=02/+97! +W2/.,.7=1,! 61-! 73=9! 0=W+;! +W2-+99=1,! 2-16=/+! =9! 73.7! .9! ;+T+/120+,7! 2-1>++;9S! ,@Z0@0d*#-3;*3/-1X@0.(!!=9!;1O,-+48/.7+;!=,!7.97+!:8;!>+//9!.,;!1-123.-5,4+./!+2=73+/=80]! 73=9!=9!1:T=189!=,!73+!&Hh+//9!@G1T=+!")(!',!.;;=7=1,S!.7!&;26S!0@0d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n,+!>.T+.7!3+-+!=9!73.7!2-+>=9+!97.4=,4!16!73+!7.97+!:8;!.,;!Q+:-.6=93! ;+T+/120+,7!73.7!=9!,17!2199=:/+!O=73!73+!2-+9+,7/5!.T.=/.:/+!711/9!.,;!+0:-51,=>!0.29! `"?Da(!I+!.-+!8,>+-7.=,!61-!73+!9=78.7=1,!=,!0189+!@=(+!m"#Hm"&!6=4('oH")]!./731843!2197,.7./! .,;!.;8/7!7.97+!:8;9!.-+!2.77+-,+;!71!.!/.-4+!+W7+,7S!=7!=9!./91!;=66=>8/7!71!T+-=65!=6!73=9!=9!73+! >.9+!;8-=,4!73+=-!+.-/5!1,714+,5(!P+T+-73+/+99S!=,!73+!O=/;!752+!Q+:-.6=93S!,+O!1-4.,9!;1! ,17!;+T+/12!.;g.>+,7!71!+.>3!173+-!:87!.7!910+!./731843!T.-=.:/+!;=97.,>+!@N=4(!%)(!<3=9!=9! 1,+!16!73+!L+5!;=66+-+,>+9!O=73!73+!*+/7$*?\?!087.,7S!O3+-+!7.97+!:8;!1-4.,9!,17!1,/5!61-0! ;+!,1T1!O=73=,!73+!+2=73+/=80S!:87!./91!.-=9+!6-10!73+!92/=77=,4!16!2-+H+W=97=,4!1-4.,9(! F+-3.29! 1,+! 16! 73+! -1/+9! 16! >+//! 017=/=75! =9! =,;++;! 73+! 2.77+-,=,4! 16! 7.97+! :8;! 1-4.,9]!.!,+O/5!61-0=,4!+2=73+/=./!q7.97+H:8;H71!:+c!>+//!3.9!7O1!>31=>+9S!+=73+-!71!61-0!.! ! CD! DISCUSSION ,+O!1-4.,!.-18,;!=7!1-!71!g1=,!1,+!16!73+!2-+H+W=97=,4!1-4.,9(!!<3+!.,./59=9!16!73+!01/+>8/.-! 0+>3.,=90! 8,;+-/5=,4! 73+! 017=/=75! .,;! +T+,78./! .77-.>7=1,! 16! ;+T+/12=,4! +2=73+/=./! >+//9! =,71!73+!61-0=,4!7.97+!:8;9!>18/;!2+-3.29!4=T+!3=,79!.:187!73=9!352173+9=9( ! 9@TEH<' .R]24! mW2-+99=1,! 2.77+-,9! 16! 91,=>! 3+;4+314! @I;;)S! :1,+! 01-2314+,+7=>! 2-17+=,9! H#! .,;! H%! @J<.?%)S! @J<.?g)! .,;! 6=:-1:/.97! 4-1O73! 6.>71-! HD! @W@0?d)! 6-10! m"E(&! 71! m"&! 16! 0189+! 71,48+! ;+T+/120+,7! O=73! O31/+! 018,7! =,! 9=78! 35:-=;=Q.7=1,! @.,7S! .,7+-=1-]! 2197S! 2197+-=1-)(! U//! 618-! 4+,+9! .-+! +W2-+99+;! 8,=61-0/5! =,! 73+! 71,48+! +2=73+/=80! :+7O++,! m"E(&! .,;! ""(! U7! m"$S! :+61-+! 01-231/14=>.//5! ;+7+>7.:/+! 4897.71-5! 2.2=//.+!=,=7=.7=1,S!A33S!M02H#!.,;!M02H%! +W2-+99=1,!9+4-+4.7+9!=,71!;=9>-+7+!92179S! O3+-+.9S!N46HD!=9!;1O,-+48/.7+;(!U7!m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h+-+S!O+!-+T+./+;!73.7!,@Z0@0d*#-3;*3/-1X@0.(! >+//!>1,7.>7! O=73! 73+! &Hh+//! H73.7! /=L+/5! >1--+921,;9! 71! 73+! 2-+95,.27=>! <52+! '''! 0.00./=.,! >+//H!! ! CR! DISCUSSION +,3.,>+9! 73+! 0.=,7+,.,>+! 16! 73+! 017=/+! >+//! =,71! 73+! 4-182S! ! =,! >1,7-.97! 71! 73+! >1,7.>7! :+7O++,! ,@Z0@0d*#-3;*3/-1X@0.(! >+//9! @N=4(! &)(! n,+! 352173+9=9! >18/;! :+! 73.7! ,@Z0@0d*#-3;*3/-1X@0.( >+//9! 4+7! =,>1-21-.7+;! =,71! 73+! 7.97+! :8;! 1,>+! 73+=-! .>7=T=75! =9! >1--+/.7+;!O=73!173+-!7.97+!:8;!>+//9!98>3!.9!73+!&Hh+//(!<3=9!0+>3.,=90!=9!>1001,!61-! ,+O/5!:1-,!>+//9!=,!73+!BPA!`"?Ra(!r+9>-=27=1,!16!73+!921,7.,+189!.>7=T=75!16!73+!0=4-.7=,4! >+//9!.,;!12714+,+7=>!0.,=28/.7=1,!>18/;!93+;!/=437!=,!73+!68,>7=1,./!0+>3.,=90!/+.;=,4!71! 73+!=,>1-21-.7=1,!16!,@Z0@0d*#-3;*3/-1X@0.(!>+//9!=,71!73+!7.97+!:8;(! ' '''.R4G43/' ^;>B' @C' B;<' =I? <3+!>+//!017=/=75!1:9+-T+;!;8-=,4!7.97+!:8;!61-0.7=1,!=9!-+0=,=9>+,7!16!73+!2-1>+99! 16! +2=73+/=./H0+9+,>350./! 7-.,9=7=1,! @mG<)(! r8-=,4! mG ! ! <3+-+61-+! 0=vH#EE! .,;! PBUG! .-+! 411;! >.,;=;.7+! 01/+>8/+9! 71! :+! =,T1/T+;! =,! 73+! 2-1>+99!16!7.97+!:8;!>+//!017=/=75!.,;!1-4.,!61-0.7=1,(! ! DE! DISCUSSION ' .R4G4U/'.C'L'LI==IA'J; ! F+-3.29!73+!0197!.22-12-=.7+!9+,91-5!1-4.,9!71!>102.-+!O=73!73+!7.97+!:8;9!.-+!73+! r-19123=/.! 100.7=;=.(! <3+! r-19123=/.! >10218,;! +5+! =9! .! -+0.-L.:/5! 2-+>=9+! >+//8/.-! 2.77+-,(! '7! ;+-=T+9! 6-10! .! 01,1/.5+-! +2=73+/=80! >.//+;! +5+H.,7+,,./! ;=9>! `"C#H"C%a! .,;! >1,9=979! 16! .22-1W=0.7+/5! DEE! 8,=7! +5+9! .99+0:/=,4! =,71! .! 3+W.41,! O=73! .! -+0.-L.:/+! 2-+>=9=1,(!<3+-+!.-+!$#H$%!T+-7=>./!>1/80,9!16!8,=7!+5+9(!<3+!2.77+-,=,4!16!73+!>1-+!16!73+! 100.7=;=80!=,!73+!.;8/7!>1,7.=,9!+=437!23171-+>+271-!,+8-1,9S!618-!/+,9!9+>-+7=,4!>1,+! >+//9!.,;!7O1!2=40+,7!>+//9(!! <3+!2.77+-,=,4!16!73+!;-19123=/.!>10218,;!+5+!1-=4=,.7+9!6-10!73+!2197+-=1-!0.-4=,! 16! 73+! +5+! ;=9>! .,;! .;T.,>+9! .,7+-=1-/5! O3+-+! 8,;=66+-+,7=.7+;! >+//9! .99+0:/+! =,71! 100.7=;=./!>/897+-9(!',!73+!6=-97!97.4+9!16!+0:-51,=>!;+T+/120+,7S!73+-+!.-+!DH"E!>+//9!=,! 73+!+5+H.,7+,,./!;=9>9!O319+!,80:+-!4-+.7/5!=,>-+.9+9!=,!73+!6=-97!7O1!/.-T./!=,97.-9!`"C&a(! U7! 73+! /.7+! 9+>1,;J73=-;! /.-T./! 7-.,9=7=1,! ;=66+-+,7=.7+;! >+//9! 61-0! 73+! 01-2314+,+7=>! 68--1O!O3=>3!2.77+-,9!73+!+5+!;=9>(!B/897+-9!73.7!.-=9+!O=73=,!73+!01-2314+,+7=>!68--1O! @>3.,,+/)! 61-0! 73+! 100.7=;=80(! f,;=66+-+,7=.7+;! >+//9! =,7+4-.7+! 73+! 01-2314+,+7=>! 68--1O! 61-0! 2-+>/897+-9! >.//+;! -19+77+9(! A8:9+7! 16! 73+! 2-+>/897+-+;! >+//9! 61-0! 23171-+>+271-! ,+8-1,9! `"C?a(! <3+! >+//9! 73.7! ;1! ,17! =,7+4-.7+! 73+! 2-+>/897+-9! 8,;+-41! 0=719=9!.,;!73+!,+O/5!:1-,!>+//9!4+,+-.7+!0.78-+!100.7=;=.!`"C?a!`"CCa!`"CDa(!<3+!/+,9! 9+>-+7=,4!>+//9!.-+!73+!/.97!71!=,>1-21-.7+!73+!100.7=;=80(!<3+!>+//9!73.7!.-+!,17!92+>=6=+;! ,+=73+-!.9!23171-+>+271-9!,1-!/+,9!O=//!:+>10+!2=40+,7!>+//9(!<3+!+5+!6=+/;!4+,+-.7+9!01-+! >+//9!73.,!-+V8=-+;S!7389!73+9+!+W7-.!>+//9!.-+!+/=0=,.7+;!:5!.212719=9!`"CRa!`"DEH"D#a(!! N-10!73+!.:1T+!;+9>-=27=1,!=7!=9!+T=;+,7!73.7!>+//!-+.--.,4+0+,7!;1+9!1>>8-!71!910+! +W7+,7! 71! 4+,+-.7+! 73+! 2-+>=9+S! 97+-+1752+;! >+//! >1,7+,7! .,;! 219=7=1,! O=73=,! 73+! 100.7=;=.(!h1O+T+-S!=,!.,5!>.9+!=7!3.9!,17!:++,!;+9>-=:+;!>+//!01T+0+,7!6-10!1,+!1-4.,! 71!.,173+-!.9!O+!618,;!;8-=,4!7.97+!:8;!61-0.7=1,(!<3=9!=9!./91!73+!>.9+!61-!9+,91-5!1-4.,9! 98>3!.9!73+!6=93!/.7+-./!/=,+!O3+-+!2-=01-;=80H>+//!0=4-.7=1,!.>3=+T+9!.22-12-=.7+!219=7=1,! 16! ,+8-10.979! ./1,4! 73+! .,7+-1H2197+-=1-! .W=9! 16! 73+! +0:-51(! B+//! +W>3.,4+! :+7O++,! ,+=43:1-=,4!/.7+-./!/=,+!1-4.,9!3.9!,17!:++,!;+9>-=:+;(!mW2/1-=,4!73+!68,>7=1,./!9=4,=6=>.,>+! 16!7.97+!:8;!>+//!017=/=75!.,;!=,>1-21-.7=1,!O=//!:+!3+/268/!=,!1-;+-!71!:+77+-!>102.-+!=7! O=73!173+-!>.9+9!=,!73+!,+-T189!9597+0(! ! D"! DISCUSSION ! .R43/!6##%`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w)!"%y!`"D$a(! <3=9!w"%y!>+//9!4+,+-.7+!21970=717=>!>+//9!@<52+!'o)!73.7!+,7+-!73+!7.97+!:8;!.,;!-+2/.>+!73+! ;.0.4+;! 1-! .212717=>! >+//! :5! ;=66+-+,7=.7=,4! =,71! 7.97+! -+>+271-! >+//9! `"#RS! "$ES! "D$H"D&a(! G=8-.!+7!./!3.T+!931O,!=,!.!-+>+,7!978;5!73.7!<52+!'o!>+//9!.-+!1T1=;!.,;!9=78.7+;!:.9.//5!=,! +.>3! 7.97+! :8;S! .,;! 0.,5! =6! ,17! .//! 16! 73+9+! :.9./! >+//9! +W2-+99! 73+! 01-2314+,! 91,=>! 3+;4+314! @A33)! `R?a(! M5! 89=,4! 4+,+7=>! /=,+.4+! 7-.>=,4S! 73+5! ;+01,97-.7+;! 73.7 A33y! :.9./! >+//9! .-+! =00+;=.7+! 2-+>8-91-9! 61-! .//! 7.97+! >+//! 752+9! O=73=,! 7.97+! :8;9! `"D?a(! ',! >8/78-+;! +0:-51,=>!71,48+9S!73+5!931O+;!73.7!+W14+,189!Ahh!-+2-+99+9!7.97+!2/.>1;+!;+T+/120+,7! `"%DaS!O3+-+.9!23.-0.>1/14=>./!:/1>L!16!Ahh!9=4,./=,4!-+98/79!=,!/.-4+-!7.97+!2/.>1;+9!=,!9=Q+! .,;! ,80:+-S! .,;! ./91! 61-0+;! =,! -+4=1,9! 16! 73+! 71,48+! 752=>.//5! .:9+,7! 16! 2/.>1;+9! `"%$S! "%Ca(!'7!=9!./-+.;5!L,1O,!73.7!A33!=9!+W2-+99+;!:5!:.9./!>+//9!'3+'4-!7.97+!:8;9!O3=/+!73+!Ahh! 7.-4+7!4+,+9!F7>3"!.,;!*/="!.-+!+W2-+99+;!:5!w"%y!2-14+,=71-!>+//9!.;g.>+,7!71!:8;9!`"DCS! "DDa! 9844+97=,4! 73.7! Ahh! 3.9! .! 2-=,>=2./! -1/+! =,! 73+! -+48/.7=1,! 16! 7.97+! :8;! -+4+,+-.7=1,(! F-+T=189!978;=+9!3.T+!931O,!73.7!A33S!:5!>+//!2-1/=6+-.7=1,S!-+48/.7+9!3.=-!4-1O73S!6+.73+-! 61-0.7=1,! .,;! 9L=,! 310+197.9=9! `"DRS! "REa! 97-8>78-+9! 73.7! .-+! +>71;+-0! ;+-=T.7=T+9! /=L+! /=,48./!7.97+!:8;9(!G=8-.!+7!./!618,;!73.7!+>712=>!7.97+!:8;9!.-=9+!6-10!w"%y!2-14+,=71-9!.,;! +W2-+99!4+,+9!-+V8=-+;!61-!73+!;=66+-+,7=.7=1,!16!+,;14+,189!7.97+!>+//9S!>102-=9+!.//!73-++! ;=66+-+,7=.7+;! 7.97+! >+//! 752+9S! =,>-+.9+! =,! ,80:+-! .,;! :+>10+! 219=7=1,+;! 2-14-+99=T+/5! 01-+! 982+-6=>=.//5! =,! 73+! /=,48./! +2=73+/=80S! .,;! Ahh! =,;8>7=1,! 16! +>712=>! 7.97+! :8;9! =9! =,;+2+,;+,7!16!7.97+!.,;!910.719+,91-5!=,,+-T.7=1,!`"R"a(! ! D#! DISCUSSION ',!380.,S!7.97+!:8;!-+4+,+-.7=1,!=9!+W7-+0+/5!=021-7.,7!=,!2.7=+,79!O=73!3+.;!.,;! ,+>L! >.,>+-S! O31! -+>+=T+! -.;=173+-.25! ;.=/5! 61-! .7! /+.97! ?HC! O++L9(! <3=9! 73+-.25! 7.-4+79! 2-1/=6+-.7=,4! 2128/.7=1,! 16! 7801-! >+//9S! 31O+T+-S .66+>79! ;-.0.7=>.//5! 73+! 611;! =,7.L+! -+98/7=,4!=,!O+=437!/199(!v+2+.7+;!=--.;=.7=1,!.66+>79!2-14+,=71-!>+//9!O3=>3!.-+!2-14-+99=T+/5! /197(! N8-73+-01-+S! 2.7=+,79! 73.7! -+>+=T+! 7-+.70+,7! 61-! ,1,H! 3+.;! .,;! ,+>L! >.,>+-9! -+21-7! 7.97+!;5968,>7=1,(!n,>+!73+!;-849!7.-4+7!73+!;=T=;=,4!>+//9S!2-14+,=71-!>+//9!.-+!=--+T+-9=:/5! .66+>7+;(! M.>7+-=./! .,;! T=-./! =,6+>7=1,9! 3.T+! :++,! -+21-7+;! 71! .66+>7! 7.97+! 2-1:.:/5! ;8+! 71! =,6/.00.7=1,! 2-1T1L+;! :5! 73+9+! 2.7314+,9(! I.,4! +7! ./! 931O+;! 73.7! 73+9+! 752+9! 16! =,6+>7=1,9!>.89+!.!-+921,9+!16!=,7+-6+-1,!9=4,.//=,4!2.73O.59S!O3=>3!/+.;9!71!7.97+!>+//!;+.73! `"R#S!"R$a(! .R434G/'$;<'7*898,(:;$'B>CB<'CKCB<=' B1,>+-,=,4! 73+! 1-4.,=9.7=1,! 16! 73+! 7.97+! 9597+0S! 7.97+! 1-4.,9! =,! 6/=+9! .,;! =,! 0.,5! 173+-! =,9+>79! .-+! ;=97-=:87+;! =,! 9+T+-./! :1;5! 2.-79! O3+-+.9! =,! 0.00./9! .-+! /1>.7+;! =,! -+97-=>7+;!-+4=1,9!16!73+!01873!.,;!23.-5,W(!N/=+9!.-+!90.//+-!73.,!0.00./9S!91!73+5!>.,! O./L! .,;! -.0:/+! 1,! 73+! 98-6.>+! 16! 73+=-! 611;]! =7c9! 61-! 73=9! -+.91,! O35! 6/=+9! 2199+99! 7.97+! -+>+271-! >+//9! ,17! 1,/5! =,! 73+=-! 01873! -+4=1,! :87! ./91! 1,! 73+=-! 2-1:19>=9S! /+49S! O=,49! .,;! 1T=21971-(!! <1! =,4+97! 73+=-! 611;S! 6/=+9! 89+! 2-1:19>=9S! .! 97-8>78-+! 73.7! >18/;! :+! .! >102.-.:/+! +V8=T./+,7!71!73+!0.00./=.,!71,48+(!F-1:19>=9S!9+7!.7!73+!:.9+!16!73+!3+.;S!=9!.!089>8/.-! 78:+!>1,,+>7+;!71!73+!23.-5,W!.,;!>102-=9+;!16!+W7+-,./!.,;!=,7+-,./!7.97+!1-4.,9!@N=48-+! "r)(!<3+!+W7+-,./!7.97+!1-4.,!>1,9=979!16!7O1!/.:+//.!@1-!/.:=./!2./29)!73.7!.-+!689+;!.7!73+! +,;!16!73+!2-1:19>=9!@N=48-+9!"r!.,;!"m)(!m.>3!/.:+//80!;=92/.59!$"!3.=-H/=L+!7.97+!:-=97/+9! @9+,9=//.)! 1,! 187+-! 98-6.>+! .,;! $E! =,! =,,+-! 7.97+! 2+49S! O3=>3! .-+! +W219+;! O3+,! 6/=+9! 97.-7! .>7=T+!6++;=,4(!r-19123=/.!/.-T.+!./91!2199+99!9+,9=//.!=,!;1-9./!7+-0=,./!.,;!T+,7-./!1-4.,9! 16!73+!3+.;!-+4=1,!.,;!23.-5,W(! U//!1T+-!73+!2-1:19>=9S!/+49!.,;!.,7+-=1-!O=,49!73+-+!.-+!4897.71-5!-+>+271-!,+8-1,9! @*vP9)! 3189+;! O=73=,! #EEH$EE! 9+,91-5! 1-4.,9! >.//+;! 9+,9=//.! .,;! 9+-T+! .9! 4.7+L++2+-9! .//1O=,4!1-!,17!73+!611;!71!2-1>++;!71!73+!;=4+97=T+!78:+(!I3+,!97=08/.7+;S!+T+,!:5!/+4H!1-! ! D$! DISCUSSION /.:=./!7.97+!,+8-1,9S!73+!2-1:19>=9!=9!+W7+,;+;!.,;!2+->+=T+9!,87-=+,79!1-!-+7-.>7+;!71!.T1=;! -+28/9=T+!98:97.,>+9(!N/5!*vP9!.-+!:=21/.-!,+8-1,9!73.7!+W7+,;!;+,;-=7+9!=,71!73+!:-=97/+9! .,;!2-1g+>7!.W1,9!71!73+!>+,7-./!7.97+!>+,7+- =,!73+!98:+9123.4+./!4.,4/=1,!@An*)(! M.9+;! 1,! 73+=-! /+,473S! 73+! 9+,9=//.! .-+! >.7+41-=Q+;! =,! /1,4! @X)S! =,7+-0+;=.7+! @')! .,;! 931-7!@A)(!m.>3!16!73+!XH!.,;!AH!>1,7.=,!618-!4897.71-5!-+>+271-!,+8-1,9!.,;!73+!'H!7O1(!<3+! 7.97+!:-=97/+9!1,!/+49!.,;!2-1:19>=9!>1,7.=,!/=L+O=9+!/1,4S!=,7+-0+;=.7+!.,;!931-7!*vP9(!m.>3! :-=97/+!2199+99+9!1,+!0+>3.,19+,91-5!.,;!+=73+-!7O1!@=,7+-0+;=.7+)!1-!618-!@931-7!.,;!/1,4)! 4897.71-5! ,+8-1,9S! O3=>3! +W7+,;! .! ;+,;-=7+! .7! 73+! 7=2! 16! 73+! :-=97/+! .9! O+//! .9! .,! .W1,./! 2-1>+99!73.7S!+,;9 =,!73+!98:+9123.4+./!4.,4/=1, `"R%a(! <3+! L12+2.;'7*! 4897.71-5! :-=97/+9! -+921,;! 71! +W7-.>+//8/.-! 9=4,./9! .,;! 3.T+! 73+! >.2.>=75!71!;=9>-=0=,.7+!73+!;=66+-+,7!-+921,9+9(!<3+!931-7!.,;!/1,4!:-=97/+9!>1,7.=,!.!9=,4/+! ,+8-1,!73.7!-+921,;9!71!984.-9S!1,+!71!O.7+-S!1,+!71!9./7!.,;!1,+!71!:=77+-!.,;!3=43!9./7H >1,>+,7-.7+;! 98:97.,>+9(! <3+! =,7+-0+;=.7+! :-=97/+9! 3.T+! 7O1! >3+019+,91-5! >+//9S! 1,+! -+921,;9! 71! :173! /1O! 9./7! .,;! 984.-9! .,;! 73+! 173+-! 71! 3=43! 9./7! .,;! :=77+-! 7.97.,79(! U9! .! >1,9+V8+,>+! 73+! 9.0+! >+//! .,;! 9=,4/+! ,+8-1,9! 9+,9+! ;=66+-+,7! 7.97+! V8./=7=+9! 73.7! 3.T+! 731843!73+!9.0+!T./+,>+!91!O+!>18/;!9844+97!73.7!73+!48=;+/=,+!16!7.97+!=,!6/=+9!=9!>/.99=6=+;! =,! .! \!T./+,>+!^! /.:+//+;! /=,+! 01;+/! 16! >1;=,4(! '7! 3.9! :++,! 931O,! 73.7! 910+! *vP9! .-+! .>7=T.7+;!:5!984.-9!.,;!6.775!.>=;9!.,;!7389!2-1017+!6++;=,4!O3=/+!.,173+-!9+7!16!*vP9!61-! :=77+-!.,;!3=43!>1,>+,7-.7=1,9!16!9./7!/+.;!71!.,!.T+-9=T+!:+3.T=18-!`"R&a(!! $>D?<24!P80:+-!.,;!;=97-=:87=1,!16!9+,9=//.!.,;!*vP9!73.7!68,>7=1,!=,!>1,7.>7!>3+019+,9.7=1,!`"R?aS!!`"RCa! ! D%! DISCUSSION ! ! 9@TEH<'.R]G(!<3+!<.97+!n-4.,9!=,! G.00./9S!A8>3!.9!G=>+!.,;! h80.,9S!.,;!=,!N/=+9! @U)! <3+! -1;+,7! 71,48+! >1,7.=,9! 7.97+!:8;9!/1>.7+;!=,!73-++!;=97=,>7! -+4=1,9(! @M)! <3+! 7.97+! :8;! =9! >10219+;! 16! &Ex"EE! 01;=6=+;! +2=73+/=./! >+//9! 73.7! +W7+,;! .! 2-1>+99! 71! 73+! 7.97+! 21-+S! O3+-+! 73+5! >10+! =,71! >1,7.>7! O=73! =,4+97+;! >3+0=>./9(! U7! /+.97! 6=T+! 752+9! 16! 9+,91-5! >+//9! @;+2=>7+;! =,! ;=66+-+,7! >1/1-9)! .-+! 618,;! =,! 73+! 7.97+!:8;S!>1--+921,;=,4!71!73+!6=T+! >.,1,=>./! 7.97+9(! @B)! *-++,! >=->/+9! =,;=>.7+! /1>.7=1,9! 16! +W7+-,./! 4897.71-5! 1-4.,9! ;=97-=:87+;! 1,! .,! .;8/7! L12+2.;'7*! 6+0./+(! @r)! <3+! r-19123=/.! 2-1:19>=9(! A31O,! 3+-+! 73+! /.:+//80! .,;! 73-++! =,7+-,./! 7.97+! 1-4.,9! =,;=>.7+;! =,! :/8+K! 73+! /.:-./! 9+,9+! 1-4.,9! @XAn9)S! 73+! ;1-9./!>=:.-=./!9+,9+!1-4.,!@rBAn)S! .,;!73+!T+,7-./!>=:.-=./!9+,9+!1-4.,! @oBAn)(! @m)! r=97-=:87=1,! 16! 73+! XHS'H S.,;! AH752+! 9+,9=//.! 1,! .! 6/5! /.:+//80(! @N)! U,! AH! 1-! XH752+! 9+,9=//80!>1,7.=,=,4!618-!*vP9(!@*)! U! L12+2.;'7*! /.-T.(! <3+! +W7+-,./! >3+019+,91-5! 1-4.,9! .-+! /1>.7+;! .7!73+!.,7+-=1-(!@h)!U,7+-=1-!+,;!16! .!/.-T.(!<3+!/1>.7=1,9!16!73+!;1-9./! 1-4.,! @rn)S! 73+! 7+-0=,./! 1-4.,! @ .R434G42/'&N< N-8=7!6/=+9!/=L+!0.00./9!.-+!.77-.>7+;!:5!9O++7H7.97=,4!>10218,;9S!/1O!>1,>+,7-.7+;! 9./=+,7!98:97.,>+9!.,;!.T+-9+!:=77+-!>10218,;9!:87!.-+!=,;=66+-+,7!71!XH.0=,1!.>=;[email protected]=)! 1-! /1O! 2h! @918-)! ;=+79! O3=>3! >18/;! +W2/.=,! 73+=-! 2-+6+-+,>+! =,! 921=/+;! 6-8=79(! *897.71-5! -+>+271-9!=,!6/=+9!=9!.!4-182!16!?D!;=97.,7/5!-+/.7+;!4+,+9!73.7!+,>1;+!287.7=T+!3+27.3+/=>./! 7-.,90+0:-.,+!2-17+=,9S!1-=4=,.//5!7318437!71!68,>7=1,!.9!*FBv9!:87!97=//!=7!=9!,17!>/+.-!=6!73+! 9=4,./!7-.,90=99=1,!=9!*!2-17+=,!;+2+,;+,7!1-!/=4.,;H4.7+;!=1,!>3.,,+/9(!! .R42434G4G/' a@BB ! D&! DISCUSSION 4897.71-5! -+>+271-! ,+8-1,9! +W2-+99! >+-7.=,! -+>+271-9! 61-! +W.02/+! 7-+3./19+! @*-&.)! =9! .! 9O++7!7.97+!-+>+271-!+W2-+99+;!=,!.!2128/.7=1,!16!4897.71-5!,+8-1,9!+=73+-!73.,!73+!:=77+-! 7.97+!-+>+271-!@*-??.)!`"RRa `#EEa `#E"a(!m.>3!16!73+9+!-+>+271-9!=9!2-+9+,7!=,!1,/5!1,+!*vP! =,!./0197!.//!:-=97/+9(!F-1g+>7=1,9!16!73+!*vP!+,;!=,!;=66+-+,7!.-+.9(!',!6/=+9S!.9!=,!0.00./9! ;=97=,>7!2128/.7=1,9!.-+!;+T17+;!71!92+>=6=>!6++;=,4!:+3.T=18-!16!.77-.>7=1,!1-!.T+-9=1,(!*-&.! /.:+//+;!>+//9!-+2-+9+,7!.!/.:+//+;!/=,+!61-!9O++7!7.97.,79!O3+-+.9!*-??.!=9!.!-+>+271-!73.7! >1--+921,;9! 71! .! /.:+//+;! /=,+! 61-! :=77+-! `#EEH#E#a(! *-$#.S! *-$$.! .,;! *-??.! .-+! -+>+271-9! ,+>+99.-5! 61-! 73+! ;+7+>7=1,! 16! :=77+-! >3+0=>./9! .,;! 73+5! .-+! +W2-+99+;! =,! .//! :=77+-! -+921,9=T+!*vP9(!r=66+-+,7!>10:=,.7=1,9!16!73+9+!90.//!7+02/.7+!16!-+>+271-9!>18/;!2199=:/5! +W2/.=,!73+=-!>.2.>=75!71!;+7+>7!.!O=;+!-.,4+!16!97-8>78-.//5!;=T+-9+!:=77+-!98:97.,>+9(!! U2.-7! 73+! *v9S! 73-++! 7-.,9=+,7! -+>+271-! 217+,7=./! >3.,,+/9! @ .R42434G43/' &N< .R42434G4U/'1=>=@'BH>ACFELB@IAM!<3+!-+921,9+!1,!.0=,1!.>=;9!=,!6/=+9!;+2+,;9!1,! 73+=-!9+W]!6+0./+9!2-+6+-!23+,5/./.,=,+S!>597+=,+S!75-19=,+!.,;!73-+1,=,+!-=>3!611;!731843! 0./+9! 931O! 97-1,4+-! 2-+6+-+,>+! 61-! /+8>=,+! .,;! 3=97=;=,+! -=>3! >10218,;9(! <3+! -+>+271-9! ! D?! DISCUSSION L,1O,! 61-! 9O++7H7.97+! 7-.,9;8>7=1,! .-+! ,17! =,T1/T+;! =,! 80.0=! 7-.,9;8>7=1,! -.=9=,4! 73+! 2199=:=/=75!73.7!7.97+!2+49!;+7+>7!.0=,1!.>=;9(! .R42434G4:/'&>?BK'BH>ACFELB@IA !N/=+9! -+921,;! 71! 9./75! 9+,9=,4! >10218,;9! 9=0=/.-! 71! 380.,S! 61-! +W.02/+! 73+5! -+921,;! O=73! 2/+.98-+! 71! .! /1OH9./7! 611;! .,;! .T+-9+! >10218,;9! >1,7.=,=,4! .! 3=43! >1,>+,7-.7=1,!16!9./7(!X.-T.+!;+7+>7!9./7!T=.!mP.B!>3.,,+/!0+0:+-9!..B$$ .,; ..B$h!:87! 73+9+!>3.,,+/9!;1!,17!.22+.-!=,!.;8/7!6/=+9(!!U,!=1,17-12=>!4/87.0.7+!-+>+271-!@'v)!=9!-+V8=-+;! 61-!/1OH9./7!9+,9=,4!.,;!73+5!.-+!+W2-+99+;!=,!*vP9(!G1-+!2-+>=9+/5!'vC?:!=9!+W2-+99+;!=,! *vP9H ;=66+-+,7!73.,!73+!1,+9!73.7!-+921,;!71!9O++7!.,;!:=77+-H .,;!+W7+,;!73+=-!2-1g+>7=1,9! =,71!.!9=,4/+!-+4=1,!=,!73+!An*(!U;8/7!6/=+9!;+7+>7!/1OH9./7!>10218,;9!O=73!*vP9!=,!XH752+! 9+,9=//.! .,;! .T+-9=T+/5! 73+! 3=43H9./7! >10218,;9! O=73! AH752+! 9+,9=//.S! O3=>3! >1,9+V8+,7/5! 9822-+99!6++;=,4(!<3+-+61-+S!73+!9./7!>1,>+,7-.7=1,!=9!=,!.!>102+7=7=1,!:+7O++,!73+9+!7O1! 9+,9=//.(! I3+,! 73+! P.y! >1,>+,7-.7=1,! =9! -+.//5! =,>-+.9+;S! 73+! P.y! 6/8W! 73-1843! mP.B! .,;! 'vC?:!;+21/.-=9+!73+!7.97+!-+>+271-!>+//9S!91!6/=+9!0.,=28/.7+!73+9+!P.y!/+.L!>3.,,+/9!.,;! :+77+-!-+921,;!71!9./7c9!V8.,7=75(! ' ! ! ! ! ! ! ! ! DC! BIBLIOGRAPHY 1. Weiss, A.S., Feast and Folly: Cuisine, Intoxication and the poetics of the Sublime. Suny press, 2002. 2. Korsmeyer, C., Romantic gastronomies: Tastes and Pleasures. Romantic circles SUNY press, 2006. 3. Brillat-Savarin, J.-A., Physiologie du goût. Sautelet, 1825. 1. 4. Purves, D., Neuroscience 2nd ed. 2001, Singapore: Sinauer Associates. 759. 5. Lindemann, B., Taste reception. Physiological reviews, 1996. 76(3): p. 718-66. 6. Lalonde, E.R. and J.A. Eglitis, Number and distribution of taste buds on the epiglottis, pharynx, larynx, soft palate and uvula in a human newborn. The Anatomical record, 1961. 140: p. 91-5. 7. Miller, I.J., Jr., Variation in human fungiform taste bud densities among regions and subjects. The Anatomical record, 1986. 216(4): p. 474-82. 8. Miller, I.J., Jr. and A.J. Preslar, Spatial distribution of rat fungiform papillae. The Anatomical record, 1975. 181(3): p. 679-84. 9. Boron W, B.E., Medical physiology. Elsevier, 2011. 10. Vandenbeuch, A., et al., Role of the ectonucleotidase NTPDase2 in taste bud function. Proceedings of the National Academy of Sciences of the United States of America, 2013. 110(36): p. 14789-94. 11. Vandenbeuch, A., T.R. Clapp, and S.C. Kinnamon, Amiloride-sensitive channels in type I fungiform taste cells in mouse. BMC neuroscience, 2008. 9: p. 1. 12. Clapp, T.R., et al., Immunocytochemical evidence for co-expression of Type III IP3 receptor with signaling components of bitter taste transduction. BMC neuroscience, 2001. 2: p. 6. 13. Baquero, A.F. and T.A. Gilbertson, Insulin activates epithelial sodium channel (ENaC) via phosphoinositide 3-kinase in mammalian taste receptor cells. Am J Physiol Cell Physiol, 2011. 300(4): p. C860-71. 14. Richter, T.A., et al., Acid-sensitive two-pore domain potassium (K2P) channels in mouse taste buds. Journal of neurophysiology, 2004. 92(3): p. 1928-36. 15. Richter, T.A., et al., Acid-sensing ion channel-2 is not necessary for sour taste in mice. The Journal of neuroscience : the official journal of the Society for Neuroscience, 2004. 24(16): p. 4088-91. 16. Kitagawa, M., et al., Molecular genetic identification of a candidate receptor gene for sweet taste. Biochemical and biophysical research communications, 2001. 283(1): p. 236-42. 17. Max, M., et al., Tas1r3, encoding a new candidate taste receptor, is allelic to the sweet responsiveness locus Sac. Nature genetics, 2001. 28(1): p. 58-63. 18. Montmayeur, J.P., et al., A candidate taste receptor gene near a sweet taste locus. Nature neuroscience, 2001. 4(5): p. 492-8. 19. Nelson, G., et al., Mammalian sweet taste receptors. Cell, 2001. 106(3): p. 381-90. 20. Sainz, E., et al., Identification of a novel member of the T1R family of putative taste receptors. Journal of neurochemistry, 2001. 77(3): p. 896-903. 21. Hoon, M.A., et al., Putative mammalian taste receptors: a class of taste-specific GPCRs with distinct topographic selectivity. Cell, 1999. 96(4): p. 541-51. 22. Zhang, Y., et al., Coding of sweet, bitter, and umami tastes: different receptor cells sharing similar signaling pathways. Cell, 2003. 112(3): p. 293-301. 23. Zhao, G.Q., et al., The receptors for mammalian sweet and umami taste. Cell, 2003. 115(3): p. 255-66. 89 BIBLIOGRAPHY #%(! G8+//+-S!w(X(S!+7!./(S!);-!1-/-.,21+!*34!/24'3@!72@'/!021!&',,-1!,*+,-#!P.78-+S!#EE&(!U3U@CE$E)K! 2(!##&HR(! #&(! p8S! h(S! +7! ./(S! L'00-1-3,!053/,'23*7!127-+!20!)$M!+5&53',+!'3!,;-!;-,-12<-1'/!,*+,-!1-/-.,21+#! F-1>++;=,49! 16! 73+! P.7=1,./! U>.;+05! 16! A>=+,>+9! 16! 73+! f,=7+;! A7.7+9! 16! U0+-=>.S! #EE%(! 2W2@$R)K!2(!"%#&DH?$(! #?(! e=.,4S! F(S! +7! ./(S! "4-3,'0'/*,'23! 20! ,;-! /6/7*<*,-! '3,-1*/,'23! +',-! C',;'3! ,;-! ,1*3+<-<&1*3-! 42<*'3!20!,;-!;5<*3!+C--,!,*+,-!1-/-.,21!+5&53',!)$M[# <3+!e18-,./!16!:=1/14=>./!>3+0=97-5S! #EE&(!GVW@%E)K!2(!$%#R?H$E&(! #C(! I=,,=4S!G(S!+7!./(S!);-!&'34'3@!+',-!021!3-2;-+.-1'4'3!4';6412/;*7/23-!*,!,;-!;5<*3!+C--,! ,*+,-!1-/-.,21#!MGB!97-8>78-./!:=1/145S!#EEC(!PK!2(!??(! #D(! z.,4S!h(S!+7!./(S!Y2<.2+','23!*34!-=275,'23!20!,;-!j%1!=2<-123*+*7!1-/-.,21!@-3-!1-.-1,2'1-!'3! <'/-!*34!1*,+#!*+,10=>9S!#EE&(!VS@$)K!2(!$E?H"&(! #R(! G+5+-316S! I(! .,;! A(! w1-9>3=,4S! Y;-<2+-3+216! +6+,-<+! '3! <*<<*7+:! 0'+;-+:! *34! '3+-/,+#! 91-0*/-#!v+98/79!.,;!2-1:/+09!=,!>+//!;=66+-+,7=.7=1,S!#EER(!UPK!2(!THW=(! $E(! G>X.843/=,S! A(w(S! F(e(! G>w=,,1,S! .,;! v(N(! G.-41/9L++S! f5+,45/'3! '+! *! ,*+,-?/-77?+.-/'0'/! f! .12,-'3!/72+-76!1-7*,-4!,2!,;-!,1*3+45/'3+#!P.78-+S!"RR#(!3:P@?$CR)K!2(!&?$HR(! $"(! I1,4S! *(<(S! w(A(! *.,,1,S! .,;! v(N(! G.-41/9L++S! )1*3+45/,'23! 20! &',,-1! *34! +C--,! ,*+,-! &6! @5+,45/'3#!P.78-+S!"RR?(!3V2@?&D&)K!2(!CR?HDEE(! $#(! v8=QHUT=/.S!X(S!+7!./(S!L2<'3*3,!72++!20!1-+.23+'=-3-++!,2!+C--,!*34!&',,-1!/2<.2534+!/*5+-4! &6!*!+'3@7-!<5,*,'23!'3!*7.;*!?@5+,45/'3#!F-1>++;=,49!16!73+!P.7=1,./!U>.;+05!16!A>=+,>+9!16! 73+!f,=7+;!A7.7+9!16!U0+-=>.S!#EE"(!-V@"&)K!2(!DD?DHC$(! $$(! h+S!I(S!+7!./(S!K<*<'!,*+,-!1-+.23+-+!*1-!<-4'*,-4!&6!*7.;*?,1*3+45/'3!*34!*7.;*?@5+,45/'3#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S! #EE%(! GU@$&)K!2(!C?C%HDE(! $%(! r.0.LS!A(S!+7!./(S!)1.<`!3577!<'/-!1-+.234!,2!&',,-1:!+C--,:!*34!5<*<'!/2<.2534+#!B3+0=>./! 9+,9+9S!#EE?(!32@$)K!2(!#&$H?%(! $&(! F+-+QS! B(U(S! +7! ./(S! 8! ,1*3+'-3,! 1-/-.,21! .2,-3,'*7! /;*33-7! -].1-++-4! '3! ,*+,-! 1-/-.,21! /-77+#! P.78-+!,+8-19>=+,>+S!#EE#(!:@"")K!2(!""?RHC?(! $?(! h1660.,S! X(B(! .,;! N(r(! G+//+77S! k5*7',6! /;*1*/,-1'+,'/+! 20! 72C! 0*,! 2+,1'/;! <-*,! .*,,'-+! 021<57*,-4! C',;! -',;-1! .21B! 7*14! 21! <24'0'-4! /213! +,*1/;:! +26*! '+27*,-! *34! C*,-1#! G+.7! 9>=+,>+S!#EE$(!S:@#)K!2(!D?RHC&(! $C(! X=8S! r(! .,;! m(v(! X=0.,S! "3,1*/-7757*1! Y*%l! *34! ,;-! .;2+.;27'.'4! 9"9%! 1-@57*,-! ,;-! ,*+,-! ,1*3+45/,'23! '23! /;*33-7! )M9G`#! F-1>++;=,49! 16! 73+! P.7=1,./! U>.;+05! 16! A>=+,>+9! 16! 73+! f,=7+;!A7.7+9!16!U0+-=>.S!#EE$(!2WW@#&)K!2(!"&"?EH&(! $D(! F-.O=77S!r(S!+7!./(S!)M9G`!'+!*!,1*3+'-3,!Y*%l?*/,'=*,-4!/*,'23!/;*33-7!1-+.234'3@!,2!1*.'4! /;*3@-+!'3!mY*%ln'#!F-1>++;=,49!16!73+!P.7=1,./!U>.;+05!16!A>=+,>+9!16!73+!f,=7+;!A7.7+9!16! U0+-=>.S!#EE$(!2WW@#&)K!2(!"&"??HC"(! $R(! I.,4S!P(S!+7!./(S!9*1*/1'3-!+'@3*7'3@!,;125@;!.7*+<*!<-<&1*3-!;-<'/;*33-7+#!M=1>3=0=>.!+7! :=12359=>.!.>7.S!#E"$(!2VGV@")K!2(!$&H&E(! %E(! <.-8,1S!U(S!+7!./(S!Y8>EG$!'23!/;*33-7!<-4'*,-+!.51'3-1@'/!3-512,1*3+<'++'23!20!+C--,:!&',,-1! *34!5<*<'!,*+,-+#!P.78-+S!#E"$(!U-:@C%%E)K!2(!##$H?(! %"(! M1S!p(S!+7!./(S!>2/*7'_*,'23!20!8)9?@*,-4!9%o%!*34!9%o[!1-/-.,21!'<<5321-*/,'=-!3-1=-+!'3!1*,! ,*+,-!&54+#!P+8-1-+21-7S!"RRR(!2W@&)K!2(!""ECH""(! ! RE! BIBLIOGRAPHY %#(! B3+8,4S!w(w(!.,;!*(!M8-,971>LS!>2/*7'_*,'23!20!9%o[!1-/-.,21+!*34!/2-].1-++'23!C',;!9%o%! 1-/-.,21+!451'3@!1*,!-<&1623'/!3-512@-3-+'+#!<3+!e18-,./!16!>102.-.7=T+!,+8-1/145S!#EE#(! UU3@%)K!2(!$?DHD#(! %$(! M8-,971>LS! *(S! Y511-3,! +,*,5+! 20! 9%o! 1-/-.,21+X! 4'+,1'&5,'23! *34! .*,;2.;6+'272@'/*7! 127-+#! F-1>++;=,49!16!73+!I+97+-,!F3.-0.>1/145!A1>=+75S!"RRR(!UGK!2(!""RH#"(! %%(! N=,4+-S!<(m(S!+7!./(S!8)9!+'@3*7'3@!'+!/15/'*7!021!/2<<53'/*,'23!012=+,>+S!#EE&(!32W@&C&$)K!2( "%R&HR( %&(! h8.,4S!z(U(S!+7!./(S!c32/B'3@!25,!9%o!1-/-.,21+!1-45/-+!,1*3+<',,-1!+-/1-,'23!'3!,*+,-!&54+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S! #E""(! 32@$D)K!2(!"$?&%H?"(! %?(! h8.,4S!z(U(S!v(!r.,;1S!.,;!A(r(!v12+-S!85,2/1'3-!*34!.*1*/1'3-!127-+!021!8)9!*34!+-12,23'3!'3! <25+-! ,*+,-! &54+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S!#EER(!G-@%%)K!2(!"$RERH"D(! %C(! r.,;1S!v(S!+7!./(S!84-32+'3-!-3;*3/-+!+C--,!,*+,-!,;125@;!8%J!1-/-.,21+!'3!,;-!,*+,-!&54#! <3+!e18-,./!16!,+8-19>=+,>+!K!73+!166=>=./!g18-,./!16!73+!A1>=+75!61-!P+8-19>=+,>+S!#E"#(!3G@")K! 2(!$##H$E(! %D(! w.7.1L.S! A(S! +7! ./(S! 8%JM! *4-32+'3-! 1-/-.,21! <2457*,-+! +C--,! ,*+,-! '3! /'1/5<=*77*,-! ,*+,-! &54+#!F/1A!1,+S!#E"#(!P@")K!2(!+$EE$#(! %R(! '93=;.S!z(S!+7!./(S!9%oZ%(?!*34!9%oZ[(?.2+','=-!0'&-1+!'3!053@'021102.-.7=T+! ,+8-1/145S!#EER(!:2U@#)K!2(!"$"H%%(! &E(! n3L8-=S! <(S! +7! ./(S! M-+'45*7!/;-<21-+.23+'=-3-++!,2!*/'4+!'3!,;-!+5.-1'21!7*163@-*7!3-1=-!'3! p,*+,-?&7'34p!Z9%o%\9%o[!425&7-?cq(!<'/-#!B3+0=>./!9+,9+9S!#E"#(!3P@?)K!2(!&#$H$#(! &"(! z.,4S! v(! .,;! M(<(! X=.,4S! Y*14'*/! 9%oZg(! 1-/-.,21+X! ,*1@-,+! '3! '+/;-<'*! *34! ;-*1,! 0*'751-r! B=->8/.7=1,!-+9+.->3S!#E"#(!222@%)K!2(!$RCH%E"(! &#(! B3.8;3.-=S! P(S! U(G(! X.,;=,S! .,;! A(r(! v12+-S! 8! <-,*&2,12.'/! @75,*<*,-! 1-/-.,21! =*1'*3,! 053/,'23+!*+!*!,*+,-!1-/-.,21#!P.78-+!,+8-19>=+,>+S!#EEE(!3@#)K!2(!""$HR(! &$(! A.,!*.:-=+/S!U(S!+7!./(S!G-,*&2,12.'/!@75,*<*,-!1-/-.,21!,6.-!$!'3!,*+,-!,'++5-#!<3+!U0+-=>.,! g18-,./!16!>/=,=>./!,87-=7=1,S!#EER(!-W@$)K!2(!C%$AHC%?A(! &%(! P+/91,S!*(S!+7!./(S!83!*<'32?*/'4!,*+,-!1-/-.,21#!P.78-+S!#EE#(!U2S@?DCC)K!2(!"RRH#E#(! &&(! X++S! A(M(S! +7! ./(S! )6.-! ""! *34! """! )*+,-! J54! Y-77+! 91-0-1-3,'*776! F].1-++-4! c*'3*,-! f75,*<*,-! M-/-.,21+!'3!M*,+#!<3+!w1-+.,!g18-,./!16!2359=1/145!…!23.-0.>1/145!K!166=>=./!g18-,./!16!73+! w1-+.,!F359=1/14=>./!A1>=+75!.,;!73+!w1-+.,!A1>=+75!16!F3.-0.>1/145S!#EER(!23@?)K!2(!%&&H?E(! &?(! B.=>+;1S! U(S! G(A(! e.6-=S! .,;! A(r(! v12+-S! "3! +',5! Y*%l! '<*@'3@! 1-=-*7+! 3-512,1*3+<',,-1! 1-/-.,21+! 021! @75,*<*,-! '3! ,*+,-! 1-/-.,21! /-77+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./!16!73+!A1>=+75!61-!P+8-19>=+,>+S!#EEE(!GW@#")K!2(!CRCDHD&(! &C(! h+-,+99S!A(!.,;!N(X(!|3.1S!);-!3-512.-.,'4-+!YYc!*34!H9N!*34!,;-!/;*3@'3@!='-C!20!/-77?,2? /-77!/2<<53'/*,'23!'3!,;-!,*+,-!&54#!F359=1/145!…!:+3.T=1-S!#EER(!-P@&)K!2(!&D"HR"(! &D(! h+-,+99S! A(S! +7! ./(S! 841-3-1@'/! +'@3*77'3@! &-,C--3! 1*,! ,*+,-! 1-/-.,21! /-77+#! <3+! e18-,./! 16! 2359=1/145S!#EE#(!:U3@F7!#)K!2(!?E"H"%(! &R(! o.,;+,:+8>3S!U(S!+7!./(S!F='4-3/-!021!*!127-!20!@75,*<*,-!*+!*3!-00-1-3,!,1*3+<',,-1!'3!,*+,-! &54+#!MGB!,+8-19>=+,>+S!#E"E(!22K!2(!CC(! ?E(! h8.,4S! z(U(S! e(! *-.,7S! .,;! A(! v12+-S! f75,*<*,-! <*6! &-! *3! -00-1-3,! ,1*3+<',,-1! ,;*,! -7'/',+! '3;'&','23!'3!<25+-!,*+,-!&54+#!F/1A!1,+S!#E"#(!P@")K!2(!+$E??#(! ! R"! BIBLIOGRAPHY ?"(! h8.,4S! U(X(S! +7! ./(S! );-! /-77+! *34! 72@'/! 021! <*<<*7'*3! +251! ,*+,-! 4-,-/,'23#! P.78-+S! #EE?(! UUG@C"E&)K!2(!R$%HD(! ?#(! '93=0.-8S!z(S!+7!./(S!)1*3+'-3,!1-/-.,21!.2,-3,'*7!0*<'76!<-<&-1+!9cL$>[!*34!9cL%>$!021++;=,49! 16! 73+! P.7=1,./! U>.;+05! 16! A>=+,>+9! 16! 73+! f,=7+;!A7.7+9!16!U0+-=>.S!#EE?(!2W3@$$)K!2(!"#&?RHC%(! ?$(! X12+Qe=0+,+QS! P(r(S! +7! ./(S! )C2! <-<&-1+! 20! ,;-! )M99! 0*<'76! 20! '23! /;*33-7+:! 9B4$7[! *34! 9B4%7$:!*1-!/2?-].1-++-4!'3!*!+5&+-,!20!,*+,-!1-/-.,21!/-77+# e18-,./!16!,+8-1>3+0=97-5S!#EE?(! -V@")K!2(!?DHCC(! ?%(! X5.//S!o(S!+7!./(S!L-/1-*+-!'3!1*,!,*+,-!1-/-.,21!/-77!'3,1*/-7757*1!.E!'+!,;-!.12]'<*,-!+,'<575+!'3! +251! ,*+,-! ,1*3+45/,'23#! U0+-=>.,! g18-,./! 16! 2359=1/145(! B+//! 2359=1/145S! #EE"(! GV2@$)K! 2(! B"EE&H"$(! ?&(! h8.,4S!z(U(S!+7!./(S!91-+63*.,'/!Z)6.-!"""(!/-77+!'3!<25+-!,*+,-!&54+!+-3+-!+251!Z*/'4(!,*+,-#!<3+! e18-,./!16!2359=1/145S!#EED(!:VS@F7!"#)K!2(!#RE$H"#(! ??(! P+/91,S! <(G(S! +7! ./(S! )*+,-! 053/,'23! '3! <'/-! C',;! *! ,*1@-,-4! <5,*,'23! 20! ,;-! .B4$7[! @-3-#! B3+0=>./!9+,9+9S!#E"E(!3:@C)K!2(!&?&HCC(! ?C(! r+N.Q=1S! v(U(S! +7! ./(S! I-.*1*,-! .2.57*,'23+! 20! 1-/-.,21! /-77+! *34! .1-+63*.,'/! /-77+! '3! <25+-! ,*+,-!&54+#!<3+!e18-,./!16!,+8-19>=+,>+!K!73+!166=>=./!g18-,./!16!73+!A1>=+75!61-!P+8-19>=+,>+S! #EE?(!GS@"&)K!2(!$RC"HDE(! ?D(! h8.,4S! z(U(S! z(! G.-85.0.S! .,;! A(r(! v12+-S! H21-.'3-.;1'3-! '+! /21-7-*+-4! C',;! +-12,23'3! '3! <25+-! ,*+,-! &54+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S!#EED(!GV@%R)K!2(!"$EDDHR$(! ?R(! rT1-5.,>3=L1TS!*(S!+7!./(S!f8J8:!',+!1-/-.,21+:!*34!f8J8-1@'/!'3;'&','23!'3!<25+-!,*+,-!&54+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S! #E""(! 32@"&)K!2(!&CD#HR"(! CE(! rT1-5.,>3=L1TS!*(S!A(G(!<10>3=LS!.,;!P(!B3.8;3.-=S!J'2@-3'/!*<'3-!+63,;-+'+!*34!5.,*B-!'3! 124-3,!,*+,-!&54+#!<3+!e18-,./!16!>102.-.7=T+!,+8-1/145S!#EEC(!:W:@$)K!2(!$E#H"$(! C"(! h8.,4S! z(e(S! +7! ./(S! );-! 127-! 20! .*33-]'3! $! ;-<'/;*33-7+! '3! 8)9! 1-7-*+-! *34! /-77?/-77! /2<<53'/*,'23!'3!<25+-!,*+,-!&54+#!F-1>++;=,49!16!73+!P.7=1,./!U>.;+05!16!A>=+,>+9!16!73+! f,=7+;!A7.7+9!16!U0+-=>.S!#EEC(!2WU@"&)K!2(!?%$?H%"(! C#(! h8.,4S! z(e(S! +7! ./(S! G25+-! ,*+,-! &54+! 5+-! +-12,23'3! *+! *! 3-512,1*3+<',,-1#! <3+! e18-,./! 16! ,+8-19>=+,>+!K!73+!166=>=./!g18-,./!16!73+!A1>=+75!61-!P+8-19>=+,>+S!#EE&(!G:@%)K!2(!D%$HC(! C$(! h8.,4S! z(U(S! m(! F+-+=-.S! .,;! A(r(! v12+-S! 8/'4! +,'<57*,'23! Z+251! ,*+,-(! -7'/',+! f8J8! *34! +-12,23'3!1-7-*+-!012=+,>+S!#EER(!3GS@&R&")K!2(!%%$H&(! C&(! X=S! p(S! +7! ./(S! E5<*3! 1-/-.,21+! 021! +C--,! *34! 5<*<'! ,*+,-#! F-1>++;=,49! 16! 73+! P.7=1,./! U>.;+05!16!A>=+,>+9!16!73+!f,=7+;!A7.7+9!16!U0+-=>.S!#EE#(!--@C)K!2(!%?R#H?(! C?(! B3.,;-.93+L.-S!e(S!+7!./(S!)%M+!053/,'23!*+!&',,-1!,*+,-!1-/-.,21+#!B+//S!#EEE(!2WW@?)K!2(!CE$H""(! CC(! G+5+-316S! I(S! F75/'4*,'23!20!<*<<*7'*3!&',,-1!,*+,-#! v+T=+O9!16!2359=1/145S!:=1>3+0=97-5! .,;!23.-0.>1/145S!#EE&(!2:UK!2(!$CHC#(! CD(! h+>LS! *(X(S! A(! G=+-91,S! .,;! e(U(! r+A=01,+S! I*7,! ,*+,-! ,1*3+45/,'23! 2//51+! ,;125@;! *3! *<'721'4-?+-3+','=-!+24'5=+,>+S!"RD%(!GG3@%?$%)K!2(!%E$H&(! CR(! G.7980171S! '(S! f5+,*,216!3-51*7!.*,;C*6+!1-=-*7-4!&6!@-3-,'/!,1*/'3@!012=+,>+S!:=17+>3,1/145S!.,;!:=1>3+0=97-5S!#E"$(!PP@C)K!2(!"$&RH?#(! ! R#! BIBLIOGRAPHY DE(! X8,;5S!v(N(S!e-(!.,;!v(!P1-4-+,S!8/,'=',6!'3!,;-!;6.2,;*7*<5+:!*<6@4*7*:!*34!/21,-]!@-3-1*,-+! &'7*,-1*7! *34! /23=-1@-3,! <2457*,'23! 20! .23,'3-! @5+,*,216! 3-5123+#! e18-,./! 16! ,+8-12359=1/145S!#EE%(!-2@$)K!2(!""%$H&C(! D"(! B3.,;-.93+L.-S! e(S! +7! ./(S! );-! 1-/-.,21+! *34! /-77+! 021! <*<<*7'*3! ,*+,-#! P.78-+S! #EE?(! UUU@C""C)K!2(!#DDHR%(! D#(! F6.660.,,S!B(S!I2<-+,;-+'+!*34!,;-!/;-<'/*7!+-3+-+#!U,,8./!-+T=+O!16!295>31/145S!"R&"(!GK!2(! CRHR%( D$(! P1-4-+,S!v(!.,;!*(!I1/6S!912a-/,'23+!20!,;*7*<'/!@5+,*,216!*34!7'3@5*7!*1-*+!'3!,;-!1*,#!M-.=,! -+9+.->3S!"RC&(!-G@")K!2(!"#$HR(! D%(! B.-/+71,S!U(S!v(!U>>1//.S!.,;!A(U(!A=01,S!Y24'3@!'3!,;-!<*<<*7'*3!@5+,*,216!+6+,-<#!<-+,;9! =,!,+8-19>=+,>+9S!#E"E(!33@C)K!2(!$#?H$%(! D&(! B3+,S!p(S!+7!./(S!8!@5+,2,2.'/!<*.!20!,*+,-!s5*7','-+!'3!,;-!<*<<*7'*3!&1*'3#!A>=+,>+S!#E""(! 333@?E%C)K!2(!"#?#H?(! D?(! A=01,S!A(U(S!+7!./(S!);-!3-51*7!<-/;*3'+<+!20!@5+,*,'23X!*!4'+,1'&5,-4!.12/-++'3@!/24-#!P.7! v+T!P+8-19>=S!#EE?(!P@"")K!2(!DREHRE"(! DC(! N=,4+-S! <(m(! .,;! z(! G1-=7.S! )C2! @5+,*,216! +6+,-<+X! 0*/'*7! *34! =*@*7! @5+,*,216! 35/7-'! ;*=-! 4'00-1-3,!&1*'3+,-=+,>+S!"RD&(!GGP@%?DD)K!2(!CC?HD(! DD(! G1-=7.S! z(! .,;! <(m(! N=,4+-S! )2.2@1*.;'/! *34! 7*<'3*1! 21@*3'_*,'23! 20! ,;-! =*@*7! @5+,*,216! +6+,-102.-.7=T+! ,+8-1/145S! "RD&(! G3V@#)K!2(!"DCH#E"(! DR(! B3.,;-.93+L.-S! e(S! +7! ./(S! );-! /-77+! *34! .-1'.;-1*7! 1-.1-+-3,*,'23! 20! +24'5+//! :=1/145S! #E"E(! 2-W@$)K!2(!#D&HR?(! R"(! r+/.5S!v(e(S!A(B(!w=,,.01,S!.,;!A(r(!v12+-S!I-12,23'3!<2457*,-+!=27,*@-?4-.-34-3,!/*7/'5102.-.7=T+!,+8-1/145S!"RR$(!33:@%)K!2(!?E?H"$(! R$(! N=,4+-S!<(m(S!Y-77!,6.-+!*34!7'3-*@-+!'3!,*+,-!&54+#!B3+0=>./!9+,9+9S!#EE&(!3W'&EJJ?'2K!2(!=&%H&(! R%(! h.,9+,S!U(S!w(!v+877+-S!.,;!m(!|+=9L+S!)*+,-!&54!4-=-72.<-3,!'3!,;-!_-&1*0'+;:!L*3'2!1-1'2#! r+T+/120+,7./!;5,.0=>9!K!.,!166=>=./!28:/=>.7=1,!16!73+!U0+-=>.,!U991>=.7=1,!16!U,.710=979S! #EE#(!GG3@%)K!2(!%D$HR?(! R&(! w-=00S! v(N(S! W*/,21+! ,;*,! 1-@57*,-! -<&1623'/! @5+,*,216! 4-=-72.<-3,#! MGB! ,+8-19>=+,>+S! #EEC(!V'&EJJ?'3K!2(!A%(! R?(! G=8-.S!h(S!z(!w89.L.:+S!.,;!A(!h.-.;.S!Y-77!7'3-*@-!*34!4'00-1-3,'*,'23!'3!,*+,-!&54+#!U->3=T+9! 16!3=971/145!.,;!>571/145S!#EE?(!S-@%)K!2(!#ERH#&(! RC(! P1-73>877S!v(*(S!)*+,-!&54+X!4-=-72.<-3,!*34!-=275,'23#!M-.=,S!:+3.T=1-!.,;!+T1/87=1,S!#EE%(! SU@$)K!2(!"RDH#E?(! RD(! v12+-S! A(r(S! I'@3*7! ,1*3+45/,'23! *34! '3021<*,'23! .12/-++'3@! '3! <*<<*7'*3! ,*+,-! &54+#! F6/84+-9!U->3=T!K!m8-12+.,!g18-,./!16!2359=1/145S!#EEC(!U:U@&)K!2(!C&RHC?(! RR(! |.>3.-S!F(B(!.,;!G(*(!e1,QS!Y2302/*7!'<*@'3@!20!G-1B-7?7'B-!&*+*7!/-77+!'3!,;-!,*+,-!&54+!20! _-&1*0'+;#!U>7.!3=971>3+0=>.S!#E"#(!22U@#)K!2(!"E"H"&(! "EE(! '93=0.-8S! z(S! +7! ./(S! )C2! 0*<'7'-+! 20! /*34'4*,-! ,*+,-! 1-/-.,21+! '3! 0'+;-+#! G+>3.,=909! 16! ;+T+/120+,7S!#EE&(!2GG@"#)K!2(!"$"EH#"(! ! R$! BIBLIOGRAPHY "E"(! A3=S!F(S!e(N(!h8.,4S!.,;!z(F(!|3.,4S!J',,-1!*34!+C--,\5<*<'!,*+,-!1-/-.,21+!C',;!4'00-1-3,76! -=275,'23*16!.*,;C*6+#!z=!>38.,!W8+!:.1!}!U>7.!4+,+7=>.!A=,=>.S!#EE&(!3G@%)K!2(!$%?H&$(! "E#(! n=L+S! h(S! +7! ./(S! Y;*1*/,-1'_*,'23! 20! 7'@*34+! 021! 0'+;! ,*+,-! 1-/-.,21+#! <3+! e18-,./! 16! ,+8-19>=+,>+!K!73+!166=>=./!g18-,./!16!73+!A1>=+75!61-!P+8-19>=+,>+S!#EEC(!GP@#")K!2(!&&D%HR#(! "E$(! X=S!p(S!+7!./(S!9+-542@-3'_*,'23!20!*!+C--,?1-/-.,21!@-3-!*//253,+!021!/*,+t!'34'00-1-3/-!,2C*14! +5@*1#!FX1A!4+,+7=>9S!#EE&(!2@")K!2(!#CH$&(! "E%( A3=S! F(! .,;! e(! |3.,4S! Y23,1*+,'3@! <24-+! 20! -=275,'23! &-,C--3! =-1,-&1*,-! +C--,\5<*<'! 1-/-.,21!@-3-+!*34!&',,-1!1-/-.,21!@-3-+#!G1/+>8/.-!:=1/145!.,;!+T1/87=1,S!#EE?(!G3@#)K!2(! #R#H$EE(! "E&(! G.7980171S!'(S!G(!n30171S!.,;!w(!U:+S!W53/,'23*7!4'=-1+'0'/*,'23!20!,*+,-!/-77+!'3!=-1,-&1*,-+#! A+0=,.-9!=,!>+//!…!;+T+/120+,7./!:=1/145S!#E"$(!GU@$)K!2(!#"EH%(! "E?(! *1S!z(S!912/--4'3@+!20!,;-!IGJF!)1'?H*,'23*7!N253@!"3=-+,'@*,21+t!u21B+;2.!%vv`#!>'3-*@-? +.-/'0'/! -].*3+'23+! *34! /23,1*/,'23+! 20! ,;-! &',,-1! ,*+,-! 1-/-.,21! @-3-! 1-.-1,2'1-! '3! =-1,-&1*,-+#!G1/+>8/.-!:=1/145!.,;!+T1/87=1,S!#EE?(!G3@&)K!2(!R?%HC#(! "EC(! n30171S! G(S! +7! ./(S! G5,5*776! -]/75+'=-! -].1-++'23! 20! f*7.;*'*! *34! f*7.;*$g! 1-=-*7+! 4'=-1+'0'/*,'23! 20! ,*+,-! 1-/-.,21! /-77+! '3! _-&1*0'+;#! <3+! e18-,./! 16! >102.-.7=T+! ,+8-1/145S! #E""(!:2-@D)K!2(!"?"?H#R(! "ED(! w.29=0./=S!G(S!+7!./(S!W@0!+'@3*7'3@!/23,127+!.;*163@-*7!,*+,-!&54!021<*,'23!,;125@;!<'M?%vv! *34!L-7,*?H2,/;!*/,'=',6#!r+T+/120+,7S!#E""(!23V@"?)K!2(!$%C$HD%(! "ER(! B.2-=1S! e(S! +7! ./(S! G*1'3-! ,-7-2+,! 72/*,-+! 7'=-! .1-6! ,;125@;! .E! +-3+'3@#! A>=+,>+S! #E"%(! 3UU@?"DD)K!2(!""&%H?(! ""E(! G.7980171S!'(S!G(!n30171S!.,;!w(!U:+S!W53/,'23*7!4'=-1+'0'/*,'23!20!,*+,-!/-77+!'3!=-1,-&1*,-+#! A+0=,!B+//!r+T!M=1/S!#E"$(!GU@$)K!2(!#"EH%(! """(!G1-=7.S!z(!.,;!<(m(!N=,4+-S!M-07-]!/233-/,'23+!20!,;-!0*/'*7!*34!=*@*7!@5+,*,216!+6+,-<+!'3!,;-! &1*'3+,-102.-.7=T+!,+8-1/145S! "RD&(!G32@%)K!2(!&%CH&D(! ""#(! 'L+,.4.S!<(S!<(!n48-.S!.,;!<(m(!N=,4+-S!j*@*7!@5+,*,216!1-07-]!/'1/5',+!021!'3,1*21*7!0224!+21,'3@! &-;*='21! '3! ,;-! @2740'+;X! /-7757*1! 21@*3'_*,'23! *34! 3-512,1*3+<',,-1+#! <3+! e18-,./! 16! >102.-.7=T+!,+8-1/145S!#EER(!:2S@$)K!2(!#"$H#&(! ""$(! F+-+.HG.-7=,+QS! '(S! <(! P.4.=S! .,;! P(! B3.8;3.-=S! W53/,'23*7! /-77! ,6.-+! '3! ,*+,-! &54+! ;*=-! 4'+,'3/,!723@-=','-+#!F/1A!1,+S!#E"$(!V@")K!2(!+&$$RR(! ""%(! w=,,.01,S! A(B(S! )*+,-! ,1*3+45/,'23#! 8! &',,-1?+C--,! &-@'33'3@#! P.78-+S! "RR?(! 3V2@?&D&)K! 2(! C$CHD(! ""&(! M.-/1OS!X(U(!.,;!v(*(!P1-73>877S!83*76+'+!20!,;-!-<&1623'/!7'3-*@-!20!=-1,-&1*,-!,*+,-!&54+#! B3+0=>./!9+,9+9S!"RR%(!2-@?)K!2(!C"&H#%(! ""?(! A71,+S!X(G(!.,;!<(m(!N=,4+-S!G2+*'/!*3*76+'+!20!,;-!-<&1623'/!21'@'3!20!,*+,-!&54+#!B3+0=>./! 9+,9+9S!"RR%(!2-@?)K!2(!C#&H$&(! ""C(! M.-/1OS!X(U(!.,;!v(*(!P1-73>877S!F<&1623'/!21'@'3!20!*<.;'&'*3!,*+,-!&54+#!r+T+/120+,7./! :=1/145S!"RR&(!2S-@")K!2(!#C$HD&(! ""D(! v1731T.S! G(S! +7! ./(S! >'3-*@-! ,1*/'3@! 20! ,;-! -3424-19!K!.,!166=>=./!28:/=>.7=1,!16!73+!U0+-=>.,!U991>=.7=1,!16!U,.710=979S! #E"#(!GU2@C)K!2(!""D$HR"(! ""R(! X=8S!N(S!+7!./(S!u3,?&-,*?/*,-3'3!+'@3*7'3@!'3','*,-+!,*+,-!.*.'77*!4-=-72.<-3,#!P.78-+!4+,+7=>9S! #EEC(!3-@")K!2(!"E?H"#(! ! R%! BIBLIOGRAPHY "#E(! G=97-+77.S!B(G(!.,;!h(p(!X=8S!L-=-72.<-3,!20!053@'0213=T+9!16!3=971/145!.,;!>571/145S!#EE?(!S-@%)K!2(!"RRH#ED(! "#"(! <3=-80.,4./.738S! A(! .,;! X(U(! M.-/1OS! "3! ='=2! 0*,-! ,1*/'3@! +,54'-+! 20! <*<<*7'*3! ,*+,-! /-77! .12@-3',21+#!U,,./9!16!73+!P+O!z1-L!U>.;+05!16!A>=+,>+9S!#EER(!22PWK!2(!$%HD(! "##(! <3=-80.,4./.738S!A(S!+7!./(S!W*,-!<*..'3@!20!<*<<*7'*3!-<&1623'/!,*+,-!&54!.12@-3',21+#! r+T+/120+,7S!#EER(!23S@R)K!2(!"&"RH#D(! "#$( M.-/1OS!X(U(S!I.-/'0'/*,'23!20!.;*163@-*7!-3424-1+//!…!;+T+/120+,7./! :=1/145S!#E"$(!GU@$)K!2(!#EEHR(! "#&(! M.-/1OS!X(U(S!B(M(!B3=+,S!.,;!v(*(!P1-73>877S!F<&1623'/!,*+,-!&54+!4-=-72.!'3!,;-!*&+-3/-!20! '33-1=*,'23#!r+T+/120+,7S!"RR?(!2GG@%)K!2(!""E$H""(! "#?(! h=//S!r(X(!.,;!B(G(!G=97-+77.S!L-=-72.<-3,*7!3-512&'272@6!20!+*7,!,*+,-!+-3+*,'23#!<-+,;9!=,! ,+8-19>=+,>+9S!"RRE(!23@&)K!2(!"DDHR&(! "#C(! G:=+,+S!e(F(!.,;!B(G(!G=97-+77.S!"3','*7!'33-1=*,'23!20!-<&1623'/!1*,!,23@5-!*34!4-=-72.'3@! ,*+,-! .*.'77*-X! 3-1=-+! 02772C! 4'+,'3/,'=-! *34! +.*,'*776! 1-+,1'/,-4! .*,;C*6+#! U>7.! .,.710=>.S! "RRC(!2SW@$)K!2(!"$RH&D(! "#D(! w-=00S! v(N(S! .,;! M.-/1OS! X(! U(! ! S! L-=-72.<-3,! 20! ,;-! ,*+,-! +6+,-<#! "3! );-! I-3+-+X! 8! Y2<.1-;-3+'=-!M-0-1-3/-#! n/6.>7=1,!.,;!<.97+!@+;(!A(!N=-+97+=,!.,;!*(!w(!M+.8>3.02)S!A.,! r=+41S!BUK!U>.;+0=>!F-+99(S!#EED('U'2(!"&CH"D#(! "#R(! A8//=T.,S!e(G(S!U(U(!M1-+>L=S!.,;!A(!n/+9L+T=>3S!I,-=+,>+S!#E"E(!32@R)K!2(!"&%RH?E(! "$E(! nL8:1S!<(S!B(!B/.-LS!.,;!M(X(!h14.,S!Y-77!7'3-*@-!<*..'3@!20!,*+,-!&54!/-77+!*34!B-1*,'32/6,-+! '3!,;-!<25+-!,23@5-!*34!+20,!.*7*,-#!A7+0!>+//9S!#EER(!GP@#)K!2(!%%#H&E(! "$"(! nL8:1S!<(S!X(h(!F+T,5S!.,;!M(X(!h14.,S!I2]%!'+!1-s5'1-4!021!4-=-72.<-3,!20!,*+,-!&54!+-3+216! /-77+#!*+,+9!…!;+T+/120+,7S!#EE?(!GW@"R)K!2(!#?&%HR(! "$#(! z++S!w(w(S!+7!./(S!>@1`?FfW9!<*1B+!,*+,-!&54!+,-<\.12@-3',21!/-77+!'3!.2+,-1'21!,23@5-#!A7+0! >+//9S!#E"$(!32@&)K!2(!RR#H"EEE(! "$$(! B/+T+-9S! h(S! w(G(! X13S! .,;! v(! P899+S! I,-=+,>+S!#E"%(!3US@?#E&)K!2(! "#%DE"#(! "$%(! U,4+-9S! A(! .,;! v(<(! G11,S! 912]'<*7! -=-3,+! '3! u3,! +'@3*7! ,1*3+45/,'23#! P.78-+! -+T=+O9(! G1/+>8/.-!>+//!:=1/145S!#EER(!2W@C)K!2(!%?DHCC(! "$&(! M+=7+9S!B(X(S!+7!./(S!W277'+,*,'3!<2457*,-+!*!JG9!*5,21-@57*,216!722.!,2!/23,127!,;-!+'_-!*34! .*,,-13'3@! 20! +-3+216! 42<*'3+! '3! ,;-! 4-=-72.'3@! ,23@5-#! r+T+/120+,7S! #EER(! 23S@"$)K! 2(! #"DCHRC(! "$?(! M./+0.,9S! I(! .,;! I(! o.,! h8/S! F],1*/-7757*1! 1-@57*,'23! 20! JG9! +'@3*7'3@! '3! =-1,-&1*,-+X! *! /2/B,*'7!20!<2457*,21+#!r+T+/120+,7./!:=1/145S!#EE#(!G:W@#)K!2(!#$"H&E(! "$C(! M17>3L.-+TS!o(U(!.,;!U(U(!A3.-1TS!JG9!+'@3*7'3@!'3!,;-!/23,127!20!+B'3!4-=-72.<-3,!*34!;*'1! 0277'/7-!@12C,;#!r=66+-+,7=.7=1,]!-+9+.->3!=,!:=1/14=>./!;=T+-9=75S!#EE%(!PG@RH"E)K!2(!&"#H#?(! "$D(! |3.,4S! e(! .,;! X(! X=S! JG9! +'@3*7'3@! *34! +,-2/*7! '3;'&',216! */,'23! 20! JG9+! *34! ,;-'1! 1-7*,'23+;'.+! C',;! */,'=*,21+! '3! 0-*,;-1!021<*,'23X!'<.7'/*,'23+!021!.-1'24'/!.*,,-13'3@#!r+T+/120+,7./!:=1/145S!"RRD(!2-S@")K! 2(!""H#$(! ! R&! BIBLIOGRAPHY "%E(! P1-.0/5S!A(!.,;!M(U(!G1-4.,S!JG9+!<-4'*,-!7*,-1*7!'3;'&','23!*,!+5//-++'=-!+,*@-+!'3!0-*,;-1! ,1*/,!4-=-72.<-3,#!r+T+/120+,7S!"RRD(!2G:@"R)K!2(!$CC&HDC(! "%"(! M=7411;S!G(e(!.,;!U(F(!G>G.31,S!E-4@-;2@!*34!J<.!@-3-+!*1-!/2-].1-++-4!*,!<*36!4'=-1+-! +',-+! 20! /-77?/-77! '3,-1*/,'23! '3! ,;-! <25+-! -<&162#! r+T+/120+,7./! :=1/145S! "RR&(! 2PG@")K! 2(! "#?H$D(! "%#(! e8,4S! h(A(S! o(! n-12+Q.S! .,;! '(! <3+9/+66S! I;;:! J<.?%:! J<.?g! *34! W@0?d! *1-! *++2/'*,-4! C',;! '3','*,'23!*34!.*,,-13'3@!20!<25+-!,23@5-!.*.'77*-# G+>3.,=909!16!;+T+/120+,7S!"RRR(!V2@"H #)K!2(!"CRHD#(! "%$(! h.//S!e(G(S!G(X(!M+//S!.,;!<(m(!N=,4+-S!L'+15.,'23!20!+23'/!;-4@-;2@!+'@3*7'3@!*7,-1+!@12C,;!*34! .*,,-13'3@!20!7'3@5*7!,*+,-!.*.'77*-#!r+T+/120+,7./!:=1/145S!#EE$(!G::@#)K!2(!#?$HCC(! "%%(! B3+,S! p(r(S! +7! ./(S! );-! +<*77! 7-5/'3-?1'/;! .12,-2@76/*3! &'@76/*3! <2457*,-+! JG9?g?'345/-4! 2+,-2&7*+,!4'00-1-3,'*,'23#!NUAmM!g18-,./!K!166=>=./!28:/=>.7=1,!16!73+!N+;+-.7=1,!16!U0+-=>.,! A1>=+7=+9!61-!mW2+-=0+,7./!M=1/145S!#EE%(!2V@R)K!2(!R%DH&D(! "%&(! '+08-.S!A(S!+7!./(S!L'1-/,!&'34'3@!20!0277'+,*,'3!,2!*!/2<.7-]!20!&23-?<21.;2@-3-,'/!.12,-'3!*34! ',+!1-/-.,21!'3;'&',+!=-3,1*7!*34!-.'4-1<*7!/-77!0*,-+!'3!-*176!o-32.5+!-<&162#!F-1>++;=,49!16! 73+!P.7=1,./!U>.;+05!16!A>=+,>+9!16!73+!f,=7+;!A7.7+9!16!U0+-=>.S!"RRD(!-:@"?)K!2(!R$$CH%#(! "%?(! |318S!z(S!h(p(!X=8S!.,;!B(G(!G=97-+77.S!J23-!<21.;2@-3-,'/!.12,-'3+!*34!32@@'3X!'3;'&','3@! *34!'345/'3@!053@'021++;=,49! 16! 73+! P.7=1,./! U>.;+05! 16! A>=+,>+9! 16! 73+! f,=7+;! A7.7+9! 16! U0+-=>.S! #EEC(! 2WU@C)K!2(!##&$HD(! "%R(! B.97=//1S! r(S! +7! ./(S! "345/,'23! 20! -/,2.'/! ,*+,-! &54+! &6! IEE! 1-=-*7+! ,;-! /2<.-,-3/6! *34! .7*+,'/',6!20!*457,!7'3@5*7!-.',;-7'5<#!r+T+/120+,7S!#E"%(!2U2@"&)K!2(!#RR$H$EE#(! "&E(! m9O.-.L80.-S!o(F(S!'(!X.WS!.,;!e(!A>3/+99=,4+-S!Y-7757*1!+'@3*7'3@!&6!0'&12&7*+,!@12C,;!0*/,21! 1-/-.,21+#!B571L=,+!…!4-1O73!6.>71-!-+T=+O9S!#EE&(!2S@#)K!2(!"$RH%R(! "&"(! P=+S!p(S!8.2.,2+'+:!.127'0-1*,'23!*34!@-3-!-].1-++'23!.*,,-13+!'3!<25+-!4-=-72.'3@!,23@5-#! U,.7105!.,;!+0:-51/145S!#EE&(!G2W@#)K!2(!"#&H$#(! "&#(! h.//S! B(S! +7! ./(S! 83! -++-3,'*7! 127-! 021! _-&1*0'+;! W@017$! 451'3@! @'77! /*1,'7*@-! 4-=-72.<-3,#! G+>3.,=909!16!;+T+/120+,7S!#EE?(!2G3@"#)K!2(!R#&H%E(! "&$(! <3=99+S!M(!.,;!B(!<3=99+S!W53/,'23+!*34!1-@57*,'23+!20!0'&12&7*+,!@12C,;!0*/,21!+'@3*7'3@!451'3@! -<&1623'/!4-=-72.<-3,#!r+T+/120+,7./!:=1/145S!#EE&(!GVP@#)K!2(!$REH%E#(! "&%(! <1,18HN8g=01-=S!P(S!+7!./(S!F].1-++'23!20!,;-!WfW!1-/-.,21!%!@-3-!Z0@01%(!451'3@!-<&162@-3-+'+! '3!,;-!_-&1*0'+;!L*3'2!1-1'2#!G+>3.,=909!16!;+T+/120+,7S!#EE#(!22-'&EJJ?'2K!2(!A"C$HD(! "&&(! B-802S!e(*(S!+7!./(S!83!-++-3,'*7!127-!021!W@0+!'3!-3424-1<*7!.25/;!021<*,'23!'3075-3/-+!7*,-1! /1*3'20*/'*7!+B-7-,*7!.*,,-13'3@#!r+T+/120+,7S!#EE%(!232@##)K!2(!&CE$H"?(! "&?(! F+7+-9+,S! B('(S! +7! ./(S! WfW! +'@3*7'3@! 1-@57*,-+! ,;-! 35<&-1! 20! .2+,-1'21! ,*+,-! .*.'77*-! &6! /23,1277'3@!.12@-3',21!0'-74!+'_-#!FX1A!*+,+7S!#E""(!P@?)K!2(!+"EE#ERD(! "&C(! *8-83.-93.S! w(*(S! G(I(! w.,L+/S! .,;! A(! U-7.T.,=9H<9.L1,.9S! );-! H2,/;! +'@3*77'3@! +6+,- ! R?! BIBLIOGRAPHY "&D(! z11,S!w(!.,;!P(!*.=.,1S!H2,/;!+'@3*7'3@!'3!,;-!<*<<*7'*3!/-3,1*7!3-1=25+!+6+,- ! RC! BIBLIOGRAPHY "CC(! v+.;5S! r(N(S! <(m(! h.,91,S! .,;! A(! M+,Q+-S! L-=-72.<-3,! 20! ,;-! L12+2.;'7*! 1-,'3*:! *! 3-512/16+,*77'3-!7*,,'/-#!r+T+/120+,7./!:=1/145S!"RC?(!:3@#)K!2(!#"CH%E(! "CD(! ;+! P11=gS! e(B(! .,;! '(w(! h.-=3.-.,S! K3/25.7'3@! /-77! 0*,-! 4-,-1<'3*,'23! 012=+,>+S!"RR&(!GPW@&#$D)K!2(!RD$H&(! "CR(! B.4.,S!v(X(!.,;!r(N(!v+.;5S!H2,/;!'+!1-s5'1-4!021!+5//-++'=-!/-77!4-/'+'23+!'3!,;-!4-=-72.'3@! L12+2.;'7*!1-,'3*#!*+,+9!…!;+T+/120+,7S!"RDR(!3@D)K!2(!"ERRH""#(! "DE( I1/66S! <(! .,;! r(N(! v+.;5S! Y-77! 4-*,;! '3! 321<*7! *34! 125@;! -6-! <5,*3,+! 20! L12+2.;'7*# r+T+/120+,7S!"RR"(!223@$)K!2(!D#&H$R(! "D"(! G=//+-S!r(<(!.,;!v(X(!B.4.,S!>2/*7!'345/,'23!20!.*,,-13'3@!*34!.12@1*<<-4!/-77!4-*,;!'3!,;-! 4-=-72.'3@!L12+2.;'7*!1-,'3*#!r+T+/120+,7S!"RRD(!2G:@"#)K!2(!#$#CH$&(! "D#(! X.-91,S! r(m(S! +7! ./(S! Y2<.5,-1!+'<57*,'23!20!/-7757*1!.*,,-13'3@!C',;'3! ,;-!L12+2.;'7*!.5.*7! -6-#!FX1A!>10287.7=1,./!:=1/145S!#E"E(!SK!2(!+"EEED%"(! "D$(! M+=;/+-S! X(G(! .,;! v(X(! A0.//0.,S! M-3-C*7! 20! /-77+! C',;'3! ,*+,-! &54+#! <3+! e18-,./! 16! >+//! :=1/145S!"R?&(!GP@#)K!2(!#?$HC#(! "D%(! N.-:0.,S!U('(S!M-3-C*7!20!,*+,-!&54!/-77+!'3!1*,!/'1/5<=*77*,-!.*.'77*-#!B+//!.,;!7=998+!L=,+7=>9S! "RDE(!23@%)K!2(!$%RH&C(! "D&(! G=8-.S!h(!.,;!X(U(!M.-/1OS!)*+,-!&54!1-@-3-1*,'23!*34!,;-!+-*1/;!021!,*+,-!.12@-3',21!/-77+#! U->3=T+9!=7./=+,,+9!;+!:=1/14=+S!#E"E(!2UV@#)K!2(!"ECH"D(! "D?(! G=8-.S!h(S!+7!./(S!I23'/!;-4@-;2@?-].1-++'3@!&*+*7!/-77+!*1-!@-3-1*7!.2+,?<',2,'/!.1-/51+21+!20! 053/,'23*7! ,*+,-! 1-/-.,21! /-77+#! r+T+/120+,7./! ;5,.0=>9! K! .,! 166=>=./! 28:/=>.7=1,! 16! 73+! U0+-=>.,!U991>=.7=1,!16!U,.710=979S!#E"%(!GU3@"E)K!2(!"#D?HRC(! "DC(! X=8S!h(p(S!+7!./(S!G57,'.7-!I;;!+'@3*7'3@!/-3,-1+!.*1,'/'.*,-!'3!053@'0213.,=909!16!;+T+/120+,7S!#EE"(!2WS@"H#)K!2(!"%$H&(! "DR(! U73.-S! G(S! +7! ./(S! E-4@-;2@! +'@3*77'3@! '3! +B'3! 4-=-72.<-3,! *34! /*3/-1#! mW2+-=0+,7./! ;+-0.71/145S!#EE?(!2:@R)K!2(!??CHCC(! "RE(! G>w=,,+//S!'(I(S!G(!<8-0.=,+S!.,;!w(!F.7+/S!I23'/!E-4@-;2@!053/,'23+!&6!72/*7'_'3@!,;-!1-@'23! 20!.127'0-1*,'23!'3!-*176!4-=-72.'3@!0-*,;-1!&54+#!r+T+/120+,7./!:=1/145S!#EE%(!GPG@")K!2(!C?H DD(! "R"(! G=8-.S!h(S!+7!./(S!8!+,123@!3-1=-!4-.-34-3/-!20!+23'/!;-4@-;2@!-].1-++'23!'3!&*+*7!/-77+!'3! <25+-!,*+,-!&54!*34!*3!*5,232<25+!,1*3+/1'.,'23*7!/23,127!20!@-3-+!'3!4'00-1-3,'*,-4!,*+,-! /-77+#!B3+0=>./!9+,9+9S!#EE%(!G-@R)K!2(!D#$H$"(! "R#(! I.,4S!h(S!+7!./(S!"307*<<*,'23!*/,'=*,-+!,;-!'3,-10-123!+'@3*7'3@!.*,;C*6+!'3!,*+,-!&54!/-77+#! <3+! e18-,./! 16! ,+8-19>=+,>+! K! 73+! 166=>=./! g18-,./! 16! 73+! A1>=+75! 61-! P+8-19>=+,>+S! #EEC(! GP@%E)K!2(!"ECE$H"$(! "R$(! I.,4S!h(S!+7!./(S!"307*<<*,'23!*34!,*+,-!4'+214-1+X!<-/;*3'+<+!'3!,*+,-!&54+#!U,,./9!16!73+! P+O!z1-L!U>.;+05!16!A>=+,>+9S!#EER(!22PWK!2(!&R?H?E$(! "R%(! A71>L+-S! v(N(S! );-! 21@*3'_*,'23! 20! ,;-! /;-<2+-3+216! +6+,-=S!#EE#(!2-@R)K!2(!"EERH"D(! "R?(! A71>L+-S! v(N(S! );-! 21@*3'_*,'23! 20! ,;-! /;-<2+-3+216! +6+,-3S!"RR%(!GP:@")K!2(!$H#?(! ! RD! BIBLIOGRAPHY "RC(! *+,;-+S!P(S!+7!./(S!"3,-@1*,'23!20!/2<.7-]!7*1=*7!/;-<2+-3+216!21@*3+!'3,2!,;-!*457,!3-1=25+! +6+,-=+,>+S!#EE"(!U@"#)K!2(!""D#H?(! #E%(! r.3.,8L.-S! U(S! +7! ./(S! )C2! f1! @-3-+! 534-17'-! +5@*1! 1-/-.,'23! '3! L12+2.;'7*#! P+8-1,S! #EEC(! :S@$)K!2(!&E$H"?(! #E&(! e=.1S!z(S!A(e(!G11,S!.,;!B(!G1,7+//S!8!L12+2.;'7*!@5+,*,216!1-/-.,21!1-s5'1-4!021!,;-!1-+.23+-+! ,2! +5/12+-:! @75/2+-:! *34! <*7,2+-! '4-3,'0'-4! &6! ! RR! Abstract Taste buds are the sensory organs of taste, located within the oropharyngeal cavity and conserved in jawed vertebrates. They derive from two epithelia, ectodermal rostrally and endodermal caudally, without showing any major physiological differences. They are multicellular organs, composed of three distinct cell types that detect different taste stimuli. Several signaling pathways, including Wnt/b-catenin, Shh, Bmp, Notch and Fgf are required in the induction and differentiation of taste bud cells. The current work aims at understanding how oropharyngeal epithelial cells that differentiate to taste bud cell types, do behave in vivo to form functional sensory organs. The zebrafish larva is used as model because of its genetics, optical clarity and accessibility of the oropharynx. Firstly, a new transgenic line tg(fgf8a.enhancer:GFP) is characterized with respect to taste bud development. It is expressed in epithelial and all differentiating taste bud cell types. Second, spinning disk timelapse shows that gfp+ cells form groups, one of them differentiates to the 5-HT cell and altogether correspond to a developing taste bud organ. Third, surprisingly, developing taste bud cells are motile between neighbouring taste bud organs. Finally, focal two-photon laser ablation of the 5-HT cell and analysis of ascl1a-/- that is devoid of 5-HT cells show that the 5-HT cell is required for the maintenance of gfp+ cells into the developing taste bud. This is the first evidence that taste bud formation is dynamic, developing cells are motile between taste buds and require one taste bud cell type (5-HT) for their maintenance into the organ. Resumé Les bourgeons gustatifs, les organes sensoriels du goût, sont localisés dans la cavité oropharyngéenne et ils sont conservés chez les vertébrés à mâchoire. Selon leur localisation, ils ont une origine épithéliale différente (rostralement, ectodermique et caudalement endodermique). Ils sont composés de trois types cellulaires distincts qui détectent les différentes qualités gustatives. Bien que l’induction et la différenciation cellulaire de ces organes sont bien étudiées, les mécanismes de la formation des organes intactes et fonctionnelles à partir de ces cellules restent inconnus. L’objectif de ce travail est de comprendre comment les cellules épithéliales se comportent in vivo afin de former des organes sensoriels gustatifs fonctionnels chez le poisson zèbre. En premier, une nouvelle lignée transgénique tg(fgf8a.enhancer:GFP) (gfp+) a été caractérisée. Elle est exprimée dans tous les types cellulaires du bourgeon au cours du développement. En deuxième, la vidéomicroscopie in vivo montre que les cellules gfp+ peuvent se différencier à des cellules sérotoninergiques (tph1b/5-HT) et elles se regroupent autour de ces dernières. En troisième, de façon surprenant, les cellules gfp+ sont motiles et elles peuvent se déplacer d’un bourgeon à l’autre. Enfin, l’ablation au laser multi-photon et l’analyse du mutant ascl1a-/- montrent que la cellule 5-HT est nécessaire au maintien des cellules gfp+ dans le bourgeon. Ce travail démontre pour la première fois que la formation des bourgeons gustatifs est un processus dynamique et que les cellules de cet organe sensoriel sont motiles. 100