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Das Erscheinungsbild bzw. In den Untersuchungen von Flodmark et al. Pape, Wigglesworth Casteels ; s.

Shuman, Selednik ; Roland et al. Uggetti et al. Flodmark et al. So war auch in den Untersuchungen von Groenveld et al.

Hoyt, Walsh ; Weinberger et al. Casteels et al. Haas, Souner Griffith, Dodge ; Drymalski Auch Palinopsien 26 konnten beobachtet werden vgl.

Singh Der Sehverlust nach weniger schweren Traumata kann sich ca. Han, Wilkinson Tabelle 7. Hier manifestiert sich die Blindheit sofort oder innerhalb einiger Minuten nach dem Unfall.

Das adoleszente Muster, bei dem die Betroffenen zwischen 8 bis zu 14 Jahren alt sind, wird von Greenblatt als das Paradigma des Syndroms verstanden.

Gjerris und Mellemgaard betonen, dass Erwachsene wesentlich mehr verschiedene und schwerere Symptome haben.

Es kann sich dabei iktal Zung, Margalith ; Joseph, Louis oder postiktal vgl. Skolik et al. Kosnik et al.

Barry et al. Verdile, Verdile ; Chew ; Lau et al. Die Mehrheit der Betroffenen war zehn oder zwanzig Jahre alt, und der Sehverlust dauerte einige Minuten bis zu einer halben Stunde.

Der Sehverlust manifestierte sich entweder iktal oder postikatal. Postiktale Blindheit kann u. Olurin ; Fiume Aldrich, Vanderzant Daher ist nach Good et al.

Helmchen et al. Kinn, Breisblatt ; Kermode et al. Horwitz, Wener ; Good et al. Wesentlich seltener wird es nach Aortographie vgl.

Johnson, Moss 32 und nach Myelographie vgl. Smirniotopoulos et al. Kinn, Breisblatt ; Rama et al. Studdard et al.

Manifestation Der Sehverlust als Folge der Infusion von Kontrastmitteln manifestiert sich innerhalb von ein paar Minuten bis zu 12 Stunden nach einer Angiographie.

Fast alle Betroffenen regenerieren sich nach und nach innerhalb von zwei bis zu sieben Tagen vgl. Brewster et al.

Lawrence et al. Kabra et al. Waldron, Stallworth 36 oder als Komplikation einer Nephritis vgl. Devathasan et al. Drymalski Ackroyd Tepperberg et al.

Gado et al. Katafuchi et al. Es lassen sich auch Berichte von Neurotoxikosen nach einem Kobrabiss vgl. Berger, Brook , einer Quecksilbervergiftung vgl.

Davis et al. Gospe oder einer Toxoplasmose vgl. Wilson et al. Cyclosporin-A, Interleukin 2 vgl. Cisplatin berichtet vgl.

Higley et al. Das Kokain diffundiert sehr schnell durch die Plazentaschranke. Dixon , S. Kupferschmidt et al. Craigen et al. Brown et al.

Amano et al. Servidei et al. Kormguth et al. Connolly et al. Hagerty et al. Mathe et al. Durand et al. Tsutsumi et al.

Burke et al. Marsh, Hurst ; Hayashi et al. Goldberg, Custis 52, Leigh-Syndrom vgl. Morris, Harbord ; Donovan ; Oldfors et al. MacDonald ; Markowitz et al.

Schonlein-Henoch Syndrom vgl. Benhamou et al. Schimmelpfennig-Feuerstein-Mims-Syndrom vgl. Hager 56, Creutzfeld-Jakob Krankheit Pinto et al.

Carney, Anderson a,b 58, Mumps vgl. Ohama et al. Hahn et al. Fanton et al. Payton, Jones 61, Multiple Sclerose vgl.

Castaigne et al. Constantinou et al. Assadi et al. Opitz et al. Ikeda et al. Elner et al. Sahota et al. Es kann aber auch verschiedene neurodegenerative Erkrankungen z.

Estbe ; Jin et al. Krimmel, Reinert ; Okamoto et al. Hydrozephalus kann akute, aber auch chronische CVI bedingen.

Corbett Sekhar et al. Hochwald, Sahar ; Saher Osher et al. Keane ; Drymalski , aber auch schwerer bilateraler Sehverlust kann beobachtet werden vgl.

Keen ; Wybar ; Jan et al. Der Sehverlust kann abrupt auftreten und von einer Reihe visueller Symptome begleitet werden vgl. Hayreh und Okkipitalinfarkte aufgrund der Kompression der hinteren Zerebralarterie vgl.

Arroyo et al. Moore, Sterm Erstmals von zerebraler Blindheit in Folge einer Bestrahlung haben Pomeranz et al. In dieser haben sie drei Kinder untersucht.

Das Absterben eines Zwillings geschieht teilweise unbemerkt, so dass Good et al. Angeborene Hirnfehlbildungen s.

Bei 14 Kindern waren die zerebralen Anomalien unspezifisch. Barkovich, Norman a und der Abwesenheit des Septum Pellukidum vgl.

Barkovich, Norman b verbunden. Barkovich et al. Fantz Teller et al. Birch und Bane , S. McDonald et al. Good ; Jan et al.

Allen et al. Meienberg et al. Das Suchmuster kann dann als grobes Muster benutzt werden, um das visuelle Feld zu messen vgl.

Van Hof-van Duin, Mohn Manchmal erlaubt auch ein Goldmann Perimeter ein genaues Messen des Gesichtsfeldes s. Akiyama et al.

Danach werden differente Elektroden, die in Kontaktlinsen eingearbeitet sind, auf die Hornhaut gesetzt. Nickel, Hoyt Bei dieser Methode werden an der Kopfhaut der Patientinnen und Patienten Kontaktelektroden angebracht, die die Potentialschwankungen des Gehirns registrieren.

Der Alpharhythmus entwickelt sich im Alter von drei Monaten und stabilisiert sich mit dem ersten Lebensjahr. Er wird durch psychosensorische Reizung z.

Jan, Wong Ackroyd , Tepperberg et al. Bodis- Wollner et al. Reim , S. Kupersmith, Nelson Jan, Wong ; Whiting et al. So stellten auch Whiting et al.

Bowerman et al. Mohn et al. Von pathologischen Strukturen wie z. Weindling et al. PVL ist daher in der subakuten Phase mit Sonographie nur schwer zu diagnostizieren.

Bosley et al. Gottlob , S. Die visuellen Funktionen sollen sich bei diesen Kindern erst zwischen dem zweiten und vierten Lebensjahr verbessern.

Innerhalb der ersten zwei Jahre soll es bei dieser Gruppe zu einer Verbesserung des Visus kommen, selbst wenn die zugrundeliegende Augenkrankheit nicht behandelt werden kann.

Sie zeigen keine visuelle Aufmerksamkeit und fixieren oder folgen auch keinen hellen Objekten vgl.

Lancet Fielder, Mayer ; Wieser Vielmehr konnten auch visuelle Antworten bei Kindern ohne visuellen Kortex beobachtet werden vgl.

Aylward et al. So beschreiben Aykward et al. Lambert Nach Flodmark et al. Smythies , S. Griffith, Dodge Drymalski ; Ramani ; Beck et al.

Faust Viele Kinder haben Schwierigkeiten mit ganz spezifischen visuellen Stimuli vgl. Morse , die sich z. Daher kann nach Groenveld et al.

Diese Faktoren werden daher mit bestimmten Prognoseaussichten verbunden. Dabei gehen Flodmark et al. Auch Lambert et al.

Wong ; Foley, Gordon ; Hoyt ; Wunderlich et al. Erwachsene mit erworbenem CVI vgl. Wong, ; Foley, Gordon , Wong ; Casteels et al. Kennard ; Stewart, Riesen Janowsky, Finley ; Pilar et al.

Blakemore Wunderlich et al. Der These von Tresidder et al. Wong ; Foley, Gordon Wong So stellen van Hof- van Duin , S.

Beneveto, Fallon ; Mauguiere, Ballydier Arnott ; Braddick et al. Islands of preserved residual vision Andere Autorinnen und Autoren vgl.

Kasten et al. Chow et al. Provis et al. Frotscher Internet Adresse. Die Autorinnen und Autoren vermuten daher, ebenso wie Rausch et al.

Baskin, Hosobuchi Die Patientinnen und Patienten werden hierbei aufgefordert einen roten Lichtpunkt in der Mitte eines homogen beleuchteten Gebietes zu fixieren.

Dieser rote Lichtpunkt wird ca. Balliet et al. Auch Kasten et al. Unter diesen Kindern waren auch neun, bei denen cortical visual impairment diagnostiziert worden ist.

Sonksen et al. Auf einem Computermonitor werden verschiedene visuelle Stimuli dargeboten, die individuell an den entsprechenden Patienten angepasst werden.

Am Ende der Therapie werden die verschiedenen Stimuli gemischt angeboten. Diese Verbesserungen sollen sich sowohl in den Testsituationen als auch im Alltag gezeigt haben.

Tegenthoff Hoyt ; Foley, Gordon Wong und Hoyt konnten z. Ferner werden die Prognoseaussichten infolge von Traumata von Drymalski et al.

Fitzhardinge et al. Vielmehr beurteilen u. Dubowitz et al. So kamen Balliet et al. Durch die physiologische Reifung der Zapfen der Netzhaut, des Sehnerven, des Thalamus, des visuellen Kortex sowie der visuellen Koordinationszentren s.

Roland , S. Dazu wurden mit Hilfe von Bildertechniken sowie elektrophysiologischer Verfahren s. Der WHO , S.

Es fehlen daher Aussagen, anhand deren beurteilt werden kann, welcher Vorgang als Sehvorgang interpretiert wird.

Engelhardt, Schipperges , S. Baillet , der Heidelberger Pupillograph vgl. Duchowny et al. Eken et al.

Teilweise war ein Elternteil anwesend, welches das Kind in den Situationen festhielt und passend positionierte vgl. Birch, Bane ; Porro et al.

He was reminded to avoid searching for the target beyond the first saccade, regardless of his success in localisation.

Verschiedene Autorinnen und Autoren vgl. Ross et al. Bronfenbrenner , S. Balgo , S. Desinteresse bedingen. Dementsprechend wird bei der Gegenstandskonstruktion der Untersuchung die Zielsetzung, der theoretische Bezugsrahmen, sowie das Untersuchungsdesign festgelegt.

Alle Dimensionen stehen in Wechselbeziehung mit Kontextfaktoren, die personenbezogene Faktoren und Umweltfaktoren umfassen.

Wenn das Behindernde erst im Umgang entsteht vgl. Simon , S. Werbik ; Eckensberger, Meacham Ihnen liegen bestimmte individuelle und subjektive Motive, Interessen und Bedeutungsdimensionen zugrunde, so dass sie als intentional verstanden werden vgl.

Groeben et al. Groeben , S. Den Ideen autopoietischer Prozesse zufolge ist Verhalten von dem operationalen Kontext bedingt, in dem es stattfindet.

Diesem Kontext entsprechend bildet das Individuum vor dem Hintergrund seiner eigenen Kriterien und Erfahrungen selbstbestimmt sinnvolle Verhaltensstrukturen s.

Aebli ; Alisch ; v. Cranach ; v. Cranach et al. Schimank , S. Der Begriff der Alltagswelt ist in der Entwicklungspsychologie und der Sozialisationsforschung relativ neu.

Engelbert ; Herlth, Schleimer ; Mundt Kinder nehmen ihre Umwelt selektiv wahr und greifen aktiv und sinnvoll in sie ein.

Das elementare Verstehen, d. Lamnek , S. In diesem Sinne halte ich das Sinnverstehen, d. Ferner gehe ich davon aus, dass sich die Diskrepanzen auf der Sinnebene durch die Anwendung hermeneutischer Zirkel reduzieren lassen.

Das Verstehen- Wollen der Verhaltensweisen basiert auf der prinzipiellen Annahme, dass diese Auseinandersetzung des Kindes mit seiner Umwelt sinnvoll und bedeutsam ist.

Bronfenbrenner ; Engelbert ; Geulen ; Hurrelmann et al. Entwicklung wird als Interaktion in sozialen Systemen, als Wechselwirkung zwischen dem Kind und seiner Umwelt verstanden Bronfenbrenner , so dass der Untersuchungsgegenstand weder das Individuum noch die Umwelt als solche ist, sondern vielmehr die Wechselwirkung zwischen beiden Schmidt- Denter , S.

Es betont die Notwendigkeit einer systematischen Untersuchung der unmittelbaren und mittelbaren Lebensbereiche, d. Walter Barker , Wright ; ; Wright ; Barker ; Dies kann z.

Mesosysteme beschreiben die wechselseitigen Beziehungen von Lebensbereichen, an denen die Person aktiv und direkt beteiligt ist.

Hiermit kann z. Kaminski , S. Kaminski Das Problem der Identifizierung und Beschreibung solcher Umwelten menschlichen Handelns wurde als ein empirisches betrachtet.

Wesentliche Elemente des Konzeptes stellen die Synomorphie, das Behavior Setting sowie das darin ablaufende Programm dar. Fuhrer , S.

Zwischen dem nonpsychologischem Milieu und dem Verhaltensmuster besteht in der Regel eine Art Passung matching. Gump, Ross Die Synomorphie stellt den Grundbaustein jedes Behavior Settings dar.

Einen solchen Kontext stellt z. Diese Kontexte werden als Behavior Settings vgl. Dieser Begriff wird auch in der deutschsprachigen Literatur unter nonpsychologischem Milieu verstehen Barker und Wright die Ort- Zeit- Konstellation, in die ein konstantes Verhalten eingebettet ist.

Unterschiedliche Standpunkte werden jedoch in Bezug auf das verhaltensdeterminierende Moment deutlich. Sie verstehen Handlungen nicht als kontextsondern als strukturdeterminiert.

Bei den Interaktionen zwischen dem Lebewesen und der Umgebung determinieren daher Pertubationen der Umgebung nicht, wie Barker es beschreibt, was dem Lebewesen geschieht, es ist vielmehr die Struktur 3 Gegenstandskonstruktion der Untersuchung des Lebewesens, die determiniert, zu welchem Wandel es infolge der Pertubationen kommt.

Miller , S. Wie bereits in Kapitel 2. Campbell, Stanley Willems, Raush Wie bereits in Kapitel 3.

Folgt man diesen Annahmen, kann eine Beobachterin bzw. Unter der Voraussetzung, dass Handlungen generell als zielorientiert und zweckgerichtet verstanden wird, zeigt das Konzept einen Weg auf, Unterscheidungen zu treffen.

Dadurch konnten die einzelnen Behavior Settings jedoch nur sehr grobrastig erfasst werden, d. Nur der Akteur oder weitere bzw.

Von Interesse sind in diesem Zusammenhang die Auseinandersetzung mit dem prototypischen Setting, den zu untersuchenden Kindern sowie der Teilnahme der Beobachterin.

Nach Miller , S. Zitzmann , S. Die Beobachtungen sollen daher in den Wohnungen der Familien stattfinden. Da ich davon ausgehe, dass zwischen den Handlungen der Kinder und den jeweiligen Kontexten wechselseitige Beziehungen bestehen s.

Lebensjahr, womit ich der Definition von Oerter und Montada , S. Lebensjahr betrachten. Atteslander , S.

In diesem Zusammenhang scheint es mir wichtig zu erfahren, ob es weitere Geschwister gibt, ob das Kind zuhause lebt oder im Internat, ob es zur Schule oder in den Kindergarten geht, ob bzw.

Brambring Anhand dieser Daten soll erfasst werden, in welchen Momenten und wie das Kind ein Element visuell fixiert, es sich einem visuellen Reiz zuwendet, diesen verfolgt, visuell abtastet oder ihn aktiv meidet bzw.

Hauptbezugspunkt bleibt dabei das Kind mit seinen Handlungen. Hall ; Zitzmann , sowie die dinghaften Elemente, d. Trudewind , S. Beschrieben werden soll daher welche Farbe bei einem Element dominiert, ob es bunt ist und ob es sich um Grund-, Misch- oder Pastellfarben handelt.

Festgehalten werden soll daher immer auch die Position und der Abstand, in welchem sich das Element zum Kind befindet.

Auf diese Weise kann einmal festgehalten werden, wie nah das Kind ein Element betrachtet, bzw. Die spezielle Handlungseinheit, die beschrieben wird, soll in ihrer Position innerhalb des Zeitgeschehens, sowohl zeitlich als auch situativ, eingeordnet werden.

Da ich die Handlungen der Kinder als sinnhaft, zielbezogen und bedeutungshaltig verstehe s. Newtson , S. Morse Es sollen daher die Einheiten erfasst werden, in denen sich Personen oder Objekte bewegen und diese von dem Kind verfolgt werden.

Eine solche Einheit beginnt, in dem Moment, in dem eine Person bzw. Flick , S. Freitag , S. Erickson Daher gehe ich davon aus, dass die Videoaufzeichnungen als Forschungsmethode relativ problemlos akzeptiert werden.

Die exakten farblichen Unterscheidungen der einzelnen Aktionen innerhalb dieser Kategorien lassen sich der folgenden Tabelle entnehmen: 3 Gegenstandskonstruktion der Untersuchung Tab.

Des weiteren soll die Position der an der Beobachtung teilnehmenden Personen s. Das auf diese Weise gewonnene Datenmaterial soll dann im Folgenden ausgewertet und interpretiert werden.

Gleichzeitig soll der Sinn der Gesamtsituation anhand der Interpretationen vertieft bzw. Wahl Verweigert das Erkenntnis- Objekt seine Zustimmung, erfolgt das sogenannte dialog- konsenstheoretische Wahrheitskriterium, so dass das Erkenntnis- Subjekt versuchen muss, ihre Verstehensleistungen zu verbessern.

Auf diese Weise soll der Versuch unternommen werden, den Sinn der Verhaltensweisen gemeinsam mit den unmittelbar Beteiligten zu verstehen s.

Ist eine 3 Gegenstandskonstruktion der Untersuchung solche Position gefunden und eine Kamera an dieser Stelle aufgebaut, werden beide Kameras gestartet und die Aufnahme begonnen.

Die Aufnahmen werden jederzeit unterbrochen bzw. Nachdem die Aufnahmen beendet sind, wird das Geschehen gemeinsam mit den an der Situation beteiligten Erwachsenen reflektiert s.

Dabei sollen vor allem auch aktuelle Befindlichkeiten, ob das Kind z. Besonders typische bzw. Diese letzte Fassung wird den Betreuungspersonen zugeschickt.

Die Beschreibungen von A. Erst im siebten Lebensmonat entwickelte sich bei A. Gegen die Anfallbereitschaft nimmt er Sirtal Retard.

Bundesverband der pharmazeutischen Industrie , S. Zur Lebenssituation A. Bis auf das Zimmer seiner Schwester darf A.

Seit September geht A. Er versteht einfache Handlungsaufforderungen und agiert bei der Nennung seines eigenen Namens.

Frau K. Meist spricht er eine Bezugsperson direkt an. Nach Angabe der Klassenlehrerin Frau S. Er wendet sich von Dingen ab, die ihn nicht ansprechen und sucht aktiv die Situationen auf, die ihn interessieren.

Ablehnung auch durch Kneifen, Schreien und Weinen deutlich machen. Auch gestisch und mimisch zeigt A. Als ihm sein Vater z. Gefallen an einer Sache kann A.

In Situationen, die A. Dabei geht er eine bestimmte Strecke auf und ab und singt. Dieser Ablauf kann unterschiedlich lange dauern.

Danach wirkt A. Sozialverhalten A. Manchmal geht er auch auf Kinder zu und ist bereit mit ihnen zu spielen, wenn sich diese auf seine Art des Spielens einlassen.

Abends spielt er z. Dabei verteilt er seine Spielsachen meist chaotisch im gesamten Raum. In 4 Ergebnisse der Untersuchung Situationen wie dem Kindergarten, in dem seine Gruppe aus 15 Kindern bestand, wurde dementsprechend beobachtet, dass sich A.

In bekannten Umgebungen kann er sich ohne Hilfe sicher bewegen. Auch Treppen bereiten ihm keinerlei Schwierigkeiten.

Interessen und Vorlieben A. Des Weiteren zeigt er generell ein besonderes Interesse an der Exploration von Materialien. Beobachten konnte ich auch, dass A.

In Schul- oder Therapiesituationen werden diese Spiele daher meist nicht von ihm, sondern von anderen beendet. Seitdem besucht 4 Ergebnisse der Untersuchung Frau K.

Frau und Herr W. Nach verschiedenen Telefonaten mit Herrn W. Vormittags wird A. Dadurch wird der Raum etwas dunkel.

Danach singt die Mutter das Lied mit den verbesserten Passagen und A. Frau W. Obwohl A. Herr W. Vor ihm steht die Kugelbahn, an dessen anderem Ende A.

Die Standkamera steht in der Ecke vor den Fenstern und dem Schreibtischregal. Durch die Fenster, deren Jalousien inzwischen hochgezogen worden sind, dringt Tageslicht, so dass der Raum hell beleuchtet ist.

Wenn die Kugeln auf eine tiefere Ebene rollen, erzeugen sie einen hellen Glockenton. Kapitel 4.

Die teilnehmenden Personen sind A. Irritiert zeigt A. Dies zeigt sich, indem er verschiedentlich nachfragt, ob heute Montag und ob heute Unterricht oder Wochenende sei.

Dies scheint A. Die Beobachterin filmt die Episode links neben Frau W. Vor der beobachteten Episode wollte Frau W. Der Bereich des Kinderzimmers, in dem sich A.

Wenn A. Die Kugel, nach der A. Deshalb vermute ich, dass er die Hand seiner Mutter bzw. Ich gehe davon aus, dass A. Es scheint mir ferner, dass A.

Nach ca. Auch hier scheint A. Festzuhalten bleibt daher, dass A. Als A. In diesem Moment tastet er sich mit seiner rechten Hand an der Hand seiner Mutter weiter, bis er die Kugel gefunden hat.

Dieses kann, wie auch in zahlreichen anderen Situationen, sicherlich als visuelles Interesse gedeutet werden. Ich gehe daher davon aus, dass A.

Auch in anderen Situationen konnte vermehrt beobachtet werden, dass A. Fotoalben anschaut. Ich neige daher zu der These, dass A.

Vielmehr hat Frau W. Interessant bei der zweiten Beobachtung ist vor allem, dass A. Tasterfahrungen vertrauter sein, da sich A.

Beginnend von dem Moment, in dem A. Er besteht aus hartem und glattem Plastik. Der Duplostein ist nicht komplex gestaltet.

Ich vermute jedoch, dass A. Oh, du Unmittelbar nachdem A. Da zu dieser Zeit beide Augen nasal gerichtet sind, gehe ich davon aus, dass A.

Als Frau W. Scheinbar erkennt A. Schwierigkeiten hat zwischen Vorder- und Hintergrundinformationen zu unterschieden.

Diese Handlungen sind erstaunlich, wenn man bedenkt, dass A. Sie lassen sich aber vielleicht verstehen, wenn man davon ausgeht, dass A. Offensichtlich bekundet der andauernde Blickkontakt, der auch nach der Identifikation des Objektes aufrecht gehalten wird, ein Interesse an den visuellen Eigenschaften des schwarzen Duplosteins.

Vielleicht konzentriert sich A. Auch in anderen Situationen betastet gelber Duplostein , beklopft blaue Gymnastikmatte, blauer Gymnastikball, Igelball, Klangkugel und bewegt bzw.

Die Beobachtungen legen ferner die Vermutung nahe, dass A. So zeigt A. Diese Bewegungen konnten auch in anderen Situationen beobachtet werden.

Auch in dieser Szene entsteht aufgrund der Handlungsabfolgen der Eindruck, dass A. So lassen sich die meisten Augenbewegungen beobachten, nachdem A.

Diese Sequenz, die von dem Moment, in dem Herr W. Vor dieser Episode wurde A. Die Beobachterin filmt A. Der Raumbereich hinter der Kugelbahn sowie der Bereich links von A.

Die Kugel, deren Bewegung A. Als Herr W. Da sich jedoch von dem Zeitpunkt, in dem A. Als hervorhebenswert erachte ich die Tatsache, dass A.

Dass die Kugel jedoch rot ist, scheint A. Als ihm Herr W. Damit zeigt er erneut, dass er offensichtlich eine Idee hat, wohin er sich wenden muss.

Erst ca. Zwar scheint A. Nachdem die Kugel die erste Bahn verlassen hat, wendet A. Geht man davon aus, dass A. Bewegt sich ein Objekt dagegen auf A.

Auch den weiteren Lauf der Kugel scheint A. Er wendet sich jedoch nach links und folgt damit der Richtung, in die sich die Kugel bewegt.

Diese Beobachtung impliziert die Annahme, dass A. Am linken Ende der Kugelbahn hebt A. Dadurch blickt er direkt zur roten Kugel, die er auch sofort sieht, obwohl sie sich ca.

Hier scheint es erneut so, dass es A. Elemente, die sich auf seiner rechten Seite befinden bzw. Nach einem unkomplizierten Schwangerschaftsverlauf wurde die Geburt nach der Schwangerschaftswoche aufgrund einer Thrombosegefahr von Frau H.

Dabei trat eine Asphyxie auf. Nachdem B. Nach vier Wochen konnte B. Seit Oktober wird B. Dies geschieht in einem bestimmten Rhythmus: zwei Tage das rechte, einen Tag das linke Auge.

Auch B. Zur Lebenssituation B. In dem Einfamilienhaus ist B. In der ersten Etage befindet sich B. In der Woche wird B. Frau H.

Auch mit ihrer Gestik und Mimik kann B. Ist B. Lehnt sie dagegen etwas ab, macht B. Sozialverhalten B. Diese Unterscheidung begann nach Angaben von Frau H.

Vielmehr nimmt B. Im Spiel sucht sich B. Befinden sich in B. Nach Angaben der Eltern bevorzugt B. Dann vergewissert sie sich jedoch z. Auf diese Weise kann sich B.

Wenn B. Ihr Bewegungsablauf ist insgesamt sehr langsam. Interessen und Vorlieben B. Aber auch bei Dosen und Schachteln nimmt sie immer wieder den Deckel ab, um ihn dann wieder auf die Dose zu setzen.

Bei den Versteckspielen verbirgt B. Sind diese jedoch z. Frau P. Sie besucht Familie H. Zwischenzeitlich nutzt sie den Karton, um dahinter ihr Gesicht zu verstecken.

Das Spielmaterial, das B. Umgang und Dauer der Auseinandersetzung damit bestimmt B. Da sich B. Auch die Kameras scheinen B.

Vielmehr scheint B. Die Aufnahmen werden beendet, als sich B. Nur einmal reicht sie weiteres Spielzeug an, von dem sich Frau H.

Zu Beginn greife ich dabei einmal direkt in das Geschehen ein, indem ich Frau H. Bisher hat B. Beginnend von dem Moment, in dem sich die Beobachterin auf das Sofa setzt und B.

Der Bereich des Wohnzimmers, in dem sich B. Die Beobachterin, die B. Als B. Damit schaut sie direkt in die Richtung, in der ich mich gerade auf das Sofa setze.

Eine knappe Sekunde, nachdem B. Stattdessen wendet und neigt sie ihren Kopf leicht nach links. Danach richtet B. In dieser Position, also mit leicht nach links geneigtem Kopf und mit leicht nasal gerichtetem linken und leicht mittig gestelltem rechtem 4 Ergebnisse der Untersuchung Auge, vermute ich, dass B.

Auch Frau H. Wahrscheinlich fixiert B. Von dem Zeitpunkt an, zu dem ich vermute, dass B. Eine solche Bewegungsabfolge, bei der B.

Lang sucht B. Bis sie die entsprechende Augenposition gefunden hat und auf mich zeigt, vergehen weitere 12 Sek.

Die motorische Kontrolle ihrer Augenbewegungen fallen ihr deutlich schwerer als die des Kopfes.

Dies wird noch deutlicher, wenn die Augen nach oben wegrollen. Im Gegensatz zu A. Um ihre Augen, auch nachdem ihre rechtes Auge weggerutscht ist, wieder in ihre Ausgangsposition zu bringen, scheint B.

In der optimalen Blickposition beruhigt sich sicherlich auch ihr Nystagmus. In allen anderen Situationen schaut B. Von dem Moment, in dem B.

Nach Ende dieser Sequenz wiederholt B. Erst 4 Ergebnisse der Untersuchung als B. Ich gehe daher davon aus, dass B.

Interpretation der Handlungseinheit B. Ich vermute stattdessen, dass die Bewegung zu schnell war, obwohl B.

Vielleicht besteht darin sogar der besondere Reiz, wenn B. Auch nach der verbalen Aufforderung ihrer Mutter kann B.

In allen Beispielen beginnt B. Auf diese Weise kann B. Die Beschreibungen von C. Wo ist denn der andere noch?

Lower respiratory tract infections Lower respiratory tract infections LRTIs were in identified as the 8 th leading cause of disability adjusted life years DALYs; a combined assessment of morbidity and mortality; refer to 1.

In the African region, 8. General Introduction - 12 - Table 3. Detection frequencies of the aetiologic agents of community acquired pneumonia Aetiologic agent Study of community acquired CAP Lieberman, 1 Lauderdale, Jokinen, Luna, Diagnosis of community acquired pneumonia CAP is based on clinical presentations, which should include cough and at least one other symptom of acute LRTI Scottish Intercollegiate Guidelines Network, , and identification of pulmonary infiltrate on a chest radiograph.

Young children and the elderly are the most commonly afflicted with pneumonia. Streptococcus pneumoniae is almost invariably the most frequently detected pathogen in aetiologic studies of CAP.

Differentiating viral and bacterial pneumonias is difficult based on presentation alone, one inevitable ramification being over-prescription of antibiotics.

Bronchitis, predominantly a condition of older adults - particularly males Ball, - is characterized by a cough of one to three weeks duration associated with URTI, where pneumonia has been excluded.

Bronchiolitis is characterized by wheezing, hypoxia and tachypnoea caused by obstruction of the small airways, and is predominantly seen in infants and young children.

Additional to symptoms of LRTI and URTI, infections with respiratory viruses may further be specifically associated with other clinical presentations and sequelae.

General Introduction - 15 - These sometimes involve the respiratory tract but may also be of neurological, systemic, gastrointestinal, ocular or other consequence.

Miscellaneous presentations during respiratory virus infections Pyrexia is frequently observed during acute respiratory tract illness Putto, Fever is incorporated in the case definition of influenza, and is exacerbated in children, in whom it may cause convulsions Brocklebank, ; Glezen, ; Price, Tissue tropism in virus infection is dictated by the receptor specificity of the infecting virus, and respiratory and gastrointestinal epithelia express many of the same cell surface molecules.

For example, abundant expression of sialic acids on the epithelia of both respiratory and gastrointestinal tracts would theoretically allow infection of both tissues by sialic acid binding respiratory viruses e.

Infection with influenza is sometimes associated with vomiting and diarrhoea, and the virus has been detected in stool samples from patients with concurrent gastrointestinal and respiratory symptoms Chan, Children have a higher incidence of gastrointestinal manifestations than adults Dingle, The degree of sequence similarity between these two viruses is sufficient for bi-directional serologic cross-reactivity Gerna, It is perhaps surprising, then, that evidence of GI General Introduction - 16 - presentations during respiratory coronavirus infections is sparse.

Clinical studies of SARS-CoV provided the strongest association of gastrointestinal presentations with respiratory coronavirus infection; diarrhoea and vomiting were reported in a quarter of cases Booth, ; Hsu, ; Lee, ; Tsang, In a longitudinal epidemiologic study of the four respiratory coronaviruses, four of 32 single coronavirus infections were associated with gastrointestinal symptoms Kuypers, In clinical studies of HCoVNL63, two of seven and four of twelve infections involved gastrointestinal presentations Leung, ; Wu, Adenoviruses are often associated with outbreaks of gastroenteritis, and a high incidence of gastrointestinal presentations during adenoviral RTI are reported Sterner, Summarily, that adenoviruses cause both respiratory and diarrhoeal presentations is established, but how extensively these conditions overlap is altogether unclear.

Cardiac involvement on HRSV infection has presented in a range of ways, including myocarditis, tachycardia, and arrhythmias Eisenhut, HRSV associated cardiac presentations have proven fatal in previously healthy patients.

Some enteroviruses which cause respiratory illness have been identified in patients with myocarditis, and death due to myocarditis preceded by upper respiratory tract infection in which the only pathogen detected was enterovirus has been documented Panagiotopoulos, During the course of influenza virus infection, onset of acute myopericarditis as respiratory symptoms wane has been described Adams, Additional to cardiac presentations, a generalized role for acute respiratory infection in the exacerbation of nephritic syndrome is suggested MacDonald, General Introduction - 17 - Table 5.

Similarly, apnoeas have been identified in PIV-3 infections have also been linked with apnoeas in infants under the age of six months Karron, The presentation of acute otitis media AOM is frequently attributed to acute respiratory virus infections, and reciprocally, acute respiratory virus infections are frequently accompanied by otitis media.

Case fatalities in immunocompromised populations have been observed on infection with most respiratory viruses, including but not limited to influenza Ljungman, ; Milne, ; Schnell, ; Simonsen, , adenovirus Halstead, ; Klinger, ; Murtagh, ; Wenman, , parainfluenza viruses Lewis, b , HRSV Englund, ; Whimbey, , HMPV Englund, ; Sivaprakasam, , and rhinoviruses Longtin, ; Nicholson, General Introduction - 19 - 1.

Respiratory sequelae Sequelae of respiratory virus infections are summarized Table 6. Perhaps the most widely studied sequela of ARTI is asthma, the most common chronic disease of childhood, characterized by recurrent attacks of wheezing and breathlessness.

There is strong epidemiologic evidence that early HRSV infection predisposes to asthma Sigurs, , but several studies of the association of other respiratory viruses with asthma onset provide less compelling results Martinez, In children under two years of age, asthma exacerbation is most often due to HRSV infection, whereas rhinoviruses are the predominant cause in older children and adults Johnston, ; Nicholson, ; Rakes, HRSV induces upregulation of the intracellular adhesion molecule ICAM-1, which supports adhesion and retention of infiltrating neutrophils and activated eosinophils, thereby contributing to inflammation Stark, Table 6.

Adenoviruses were first associated with CNS disease in the s Lelong, Reports of adeno-associated neuropathy intermittently since this time have been of sufficient rarity to prevent evaluation of epidemiologic trends or refined clinical definitions.

Coronavirus infections cause demyelination in primate models Murray, and virions have been isolated from and visualized in the brain of multiple sclerosis patients Burks, ; Tanaka, Detection of other coronaviruses in the nervous system of neurologically afflicted patients has not been reported; nevertheless, General Introduction - 21 - much speculation surrounds the association of coronaviruses with neurologic manifestations Dessau, ; Gilden, ; Lau, ; Severance, Despite extensive vaccination campaigns, poliovirus still causes epidemics of acute flaccid paralysis, and enterovirus 71 has been associated with poliovirus-like paralysis Chonmaitree, However, the cross-over between enterovirus types causing respiratory illness and the neurologic manifestations described is currently poorly defined.

Systemic sequelae In , Ernest Goodpasture published his observations on the histopathologic findings from post-mortem examinations of individuals who had died during the influenza pandemic Goodpasture, He rebuked suggestions that a bacterial pathogen was the cause of the pandemic, and inferred the disease was of an as yet unknown aetiology influenza A virus was discovered in Smith, The condition arises through an autoimmune response to Goodpasture antigen, expressed in the lung and kidney, and is extremely rare.

Adenovirus serotypes of species A 12, 18 and 31 are highly oncogenic Kuwano, The link between adenoviruses and oncogenesis was first suggested in the s Trentin, , when it was proposed that for malignancy due to adenovirus, integration of viral DNA into the host genome precedes transcription and translation of viral polyprotein with loss of infectious virus.

Studies conducted to detect adenovirus DNA, RNA and proteins in cancer patients have nevertheless yielded negative results Green, , and a report of adenoviral nucleic acid detection in neurogenic tumours General Introduction - 22 - Ibelgaufts, remains unconfirmed.

The mechanism by which adenoviruses cause cancer is therefore yet to be resolved. Adenoviruses have also long been associated with hepatitis Hartwell, , particularly in liver transplant patients Cames, Nevertheless, as for enteroviruses, the overlap of respiratory presentations due to adenovirus infection with the sequelae described is unclear.

Sudden unexplained death in infancy SUDI formerly sudden infant death syndrome SIDS is a poorly understood phenomenon, most likely caused in some cases by an immunodeficient response to respiratory infection.

Respiratory virus diagnostics in the clinical setting A lack of available therapeutics and influence of the diagnostic test result on patient management raises the question of the value of diagnosing respiratory infections.

Bonner et al Bonner, determined the differences in management of patients with influenza whose clinicians were aware of the diagnosis compared with patients whose clinicians were blinded to the diagnostic testing results.

For influenza patients managed by clinicians aware of the aetiology of illness, fewer tests were performed thereby saving money and resources , fewer antibiotics were prescribed, the length of stay in the accident and emergency department was shorter, and the number of influenza positive patients receiving antiviral therapy was significantly higher.

It is suggested that rhinovirus infections may be responsive to corticosteroid therapy, whereas this is not recommended for treatment of HRSV infection Templeton, General Introduction - 23 - and so in a child admitted to hospital with bronchiolitis a viral diagnosis can guide patient management.

Gonzalez et al Gonzales, b used bacterial prevalence rates in clinical presentations of otitis media, sinusitis, pharyngitis, bronchitis and URTI to determine the number of antibiotic prescriptions that should have been made for these diagnoses in the USA in Of 41 million prescriptions, The prognosis during mixed infections with viruses, and dual infections with viruses and bacteria is worse Hament, ; Richard, McCullers showed that treatment of influenza infection with neuraminidase inhibitors improved survival during secondary bacterial infections in mice McCullers, ; secondary bacterial pneumonia is widely acknowledged as a cause of influenza associated death Scadding, In the last few years there has been a shift in respiratory virus diagnostics from use of traditional cell culture methods to an overwhelming preference for molecular detection techniques, most commonly real-time polymerase chain reaction PCR.

Together with the near-patient test, these are now currently used exclusively for respiratory virus detection in Scottish diagnostic laboratories.

To prevent spread of respiratory infections in hospitals, patients with the same infection can be admitted to a designated cohort area.

However, laboratory testing for respiratory viruses by real-time PCR has a turnaround time of hours, which is too General Introduction - 24 - inefficient to guide cohorting procedures.

Patients admitted with respiratory symptoms may be allocated to an isolation unit if one is available until virus testing results are returned.

To improve the turnaround time for a diagnosis of viruses for which cohorting strategies are implemented, near patient NPT or point of care testing for respiratory viruses, with a turnaround time of just fifteen minutes, offers a sufficiently rapid diagnostic result to inform cohorting decisions at the time of admission.

NPTs have been successful in reducing nosocomial transmission events of HRSV Mackie, , although rapid tests are of reduced sensitivity and specificity compared with diagnostic PCR.

Commercially produced NPTs are available for most of the respiratory pathogens listed in Table 3 and Table 4, sometimes in multiplex form.

The sample swab is dipped into a vial of solution and rotated, then pressed against the side of the vial to release the HRSV into the solution.

The solution containing virus is then pipetted into the test well and incubated for fifteen minutes, during which time the HRSV fusion antigen binds to anti-fusion antibody adsorbed on the test strip.

Adhesion of the fusion antigen to the test strip induces a colour change to pink-purple. The suitability of near-patient testing for influenza virus infection is also established, though as for near-patient testing for HRSV infection, a negative result should be treated with caution due to the potential for false-negative results Harnden, ; Ruest, Regardless of the results of the near patient test, respiratory samples are always taken in parallel for routine diagnostics.

The first stage of routine diagnostic testing is extraction of nucleic acids from respiratory samples, prior to amplification by PCR for virus detection.

This can be undertaken using an automated or manual procedure, though increasingly in the diagnostic setting automated extraction is undertaken.

Studies General Introduction - 25 - comparing manual and automated extraction found no difference in sensitivity between the two methods Espy, ; Gobbers, ; Knepp, The automated machines used for extraction are large and so space is also a consideration.

Commercially available automated nucleic acid extraction machines are reviewed elsewhere Espy, The BioRobot MDx Qiagen used for extraction of nucleic acids throughout this work captures nucleic acids from diagnostic specimens by vacuum pull-through onto magnetic silica particles.

The throughput time for extraction of 96 samples quoted by the manufacturer is 2. Nevertheless, for large scale epidemiologic study, automated extraction is the fastest, easiest, and most highly sensitive, standardised, and most reliable method of extraction currently available.

Real-time PCR operates in a closed system to give semi-quantified results without the need for an additional step for detection of amplified product such as gel electrophoresis , improving turnaround time and reducing the risk of contamination compared with conventional PCR.

Several manufacturers produce thermocycling instruments for multiplex real-time PCR reviewed in Espy, Some of the most advanced technology on the open market manufactured by General Introduction - 26 - Cepheid, USA is a fully integrated system GeneXpert Infinity, which allows detection of up to 50 pathogens directly from diagnostic samples with the first results produced in under an hour, and boasts a theoretical throughput of over samples in 24 hours.

Such technology probably gives an indication of what the future holds for diagnostic laboratories. Nucleic acid amplified during a real-time PCR reaction is detected by a fluorescent signal produced when fluorophores are released during polymerization.

ABI machines support TaqMan probes and molecular beacons for nucleic acid detection. Multiplex PCR exploits this by use of probes which absorb different wavelengths for specific detection of multiple target sequences.

Theoretically, the yield of target sequence doubles each PCR cycle, and the strength of the fluorescence emitted by the reporter probe is semi-quantitative of the amount of target sequence in the sample.

The cycle threshold Ct value is the number of PCR cycles after which the sample is identified as positive.

A lower Ct value reflects a greater the concentration of target nucleic acid in the sample. As an alternative to real-time PCR for multiplex detection of respiratory pathogens, sophisticated microarray assays are now under development.

Arrays have been used for the simultaneous detection of nine pathogens with a turnaround time comparable to that for results produced by PCR detection, although they have the potential to detect all pathogens that are known to be causative of a specific clinical outcome simultaneously reviewed in Raoult, A clinical sample can be tested directly using microarray, which enables combination of purification, amplification and detection of pathogen into one step, reducing time, cost and risk of contamination.

General Introduction - 27 - Figure 2. Fluorescent probe hybridization to amplification product and release from sequestration by Taq polymerase As the polymerase progresses down the DNA strand, it displaces the TaqMan probe through its endonuclease activity.

This number varies substantially between different viruses, determined by a range of factors such as duration of infectivity, degree of immunity from re-infection, viral persistence, and so forth.

Low density populations can support some viruses; in primitive indigenous tribes still in existence in the most isolated parts of the world, hepatitis B, Epstein-Barr, and cytomegalovirus circulate, having survived through inter-generational transmission probably over millennia Black, General Introduction - 28 - No modelling has been undertaken to determine the threshold population size needed to maintain viruses associated with respiratory presentations.

The most relevant analogy can be drawn from comparison with measles virus, which probably emerged at least two thousand years ago Sharp, To sustain itself, measles virus requires a population size of which supports a maintained susceptible pool of Black, Critically, measles is transmitted by the respiratory route, causes non-persistent infections and does not survive for any length of time in the environment.

Measles infection induces life long immunity, which is likely the result of the systemic nature of MV infections and its antigenically monotypic nature, contrasting with the plethora of respiratory virus strains generated through antigenic drift.

It follows that the population size required for maintenance of respiratory viruses such as HRSV may be smaller than that needed for measles, and so other respiratory viruses may have been circulating for substantially longer.

Recombination events create new viruses by capturing genes or gene segments from other viruses or from the host cell genome , can occur in both segmented and non-segmented viruses, and can be homologous or heterologous.

Homologous recombination occurs between parental RNAs of the same virus species with crossovers occurring at homologous sites in the genome, and does not necessarily occur during virus replication.

Coronavirus progeny of the same infection have been demonstrated to undergo homologous recombination Keck, Heterologous recombination may occur between different virus species via the copy choice mechanism first proposed for polio virus Cooper, , wherein during replication the viral RNA-dependent RNA polymerase switches templates to create a hybrid virus.

Heterologous recombination occurs between strains of the same species General Introduction - 29 - e.

Reassortment involves the mixing of virus genome segments between strains of multipartite viruses during virion assembly to yield a novel virus, and is most widely associated with influenza virus discussed in more detail subsequently.

Independent circulation in humans To illustrate, rabies reaches stage two, causing sporadic human cases; ebola virus causes limited outbreaks in people, attaining stage three; dengue causes long outbreaks in humans before returning to the sylvatic host, thereby reaching stage four; and HIV-1 clade M has successfully established itself in humans to attain stage five.

The ability of respiratory viruses to re-infect hosts is widely thought to be a product of immune-driven adaptive evolution, facilitated by the RNA-dependent RNA polymerase which does not, unlike DNA-dependent RNA polymerase, have proofreading activity.

Frequent errors during RNA replication result inevitably in the production of some progeny viruses with deleterious mutations, which are removed from the population by purifying selection.

Recombinant and zoonotic respiratory viruses Historically, rhinoviruses and coronaviruses have been considered separately from other respiratory viruses, conceptually segregated as common cold viruses, which is in part attributable to the excellent epidemiologic studies of respiratory viruses conducted in the mid twentieth century Table 7.

Much of the understanding of respiratory virus epidemiology today is drawn from the results of these classic studies.

Rhinoviruses are the most common cause of acute infectious illness in humans. The mean infection rate in the Seattle study was 0.

Young adult females experience more rhinovirus infections and illnesses than their male peers, due to their increased contact with young children, though this is redressed in later years when illness rates are higher among males Gwaltney Jr, General Introduction - 31 - Table 7.

Nasopharyngeal swabs on respiratory and COPD in later life presentation General Introduction May - Jun Families observed for one year - 32 - Rhinovirus immunogenicity is poor, and varies greatly with serotype Fox, ; an increasing number of serotypes are being reported, with over of species A and B, and 61 of species C to date.

It is this diversity which permits re-infection with rhinoviruses year after year, and on which the presumption that there will never be a vaccine for the common cold is based.

Rhinovirus and enterovirus species group within the Enterovirus genus of the Picornaviridae, and undertake recombination as a mechanism to produce novel strains McIntyre, ; McWilliam Leitch, , ; Santti, ; Savolainen, Enteroviruses undergo periodical turnover and replacement of contemporary clades by a rapidly disseminating recombinant form with striking regularity.

This remarkably high frequency of recombination provides some insight into the tremendous success of rhinoviruses to circumvent population immunity and re-infect individuals frequently throughout life.

Coronaviruses have a longer incubation period than rhinoviruses, though the duration of illness is generally shorter and symptoms are indistinguishable.

The classic epidemiologic studies of coronaviruses were of course limited to HCoVE and HCoV-OC43, though recent studies of the four coronaviruses together including the results presented here do not imply any deviating trends for the newly discovered coronaviruses2.

Antibody to coronaviruses appears in early childhood and increases in prevalence rapidly with age McIntosh, , with seroconversion in most occurring for HCoVE and HCoV-NL63 by the age of 3 and a half Dijkman, The development of a cell culture system for HCoV-HKU1 is only very recently reported Pyrc and serosurveillance was undertaken using a baculovirus expression system incorporating nucleocapsid gene.

HCoV-HKU1 comprises three genotypes Woo , the serologic determinants of which are poorly defined, and so this result is highly subjective in light of current understanding, especially when one considers the epidemiologic traits of other coronaviruses and other respiratory viruses in general.

General Introduction - 33 - rhinovirus antibody Callow, Recombination has played an intrinsic role in the evolution of the coronaviruses.

Group two, but not group one coronaviruses express a haemagglutinin esterase HE protein, acquired through a historical recombination event with an ancestral influenza C virus.

This remarkable acquisition provided the group two coronaviruses with a protein which mediates a reversible attachment to O-acetylated sialic acids Zeng, , through activity as a receptor destroying enzyme.

The emergence of SARS-CoV provides an excellent illustrative example of zoonotic transfer of a virus from an animal host into man, giving rise to a novel virus in humans.

The virus emerged in southern China with the first case reports in late and spread as far as Canada, altogether detected in 37 countries.

The species from which SARS-CoV was transferred to man will remain unknown; it is thought that frequent and close contact between humans and any of the species in which SARS-CoV infection has been demonstrated in live animal markets may have been instrumental in the zoonotic transfer of the virus.

Influenza viruses are the only viruses to repeatedly cause pandemics of respiratory disease, and a great deal of interest has been taken in the evolutionary mechanisms pervading to this feature.

Influenza viruses can circulate in a range of hosts, including human, avian, and swine species. The host specificity of the influenza virus is determined by the conformation of the host sialic acid receptor, which in the upper respiratory tract General Introduction - 34 - of human and avian hosts is distinct.

However, pigs abundantly express sialic acids of both conformations, meaning they can be infected with influenza viruses of both human and avian origin.

If a pig is infected with a human and avian virus simultaneously, the two strains are free to reassort, producing a novel virus with genomic regions of both human and avian origin.

If the haemagglutinin attachment protein is specific for the human sialic acid, a novel influenza variant with human infectivity is born.

This circumstance is the usual origin of pandemic influenza strains arising every thirty or forty years.

The reassortant pandemic strain of influenza which emerged in in Mexico incorporated genes of swine, human and avian influenza origin Novel Swine-Origin Influenza A H1N1 Virus Investigation Team, The pandemic strain has now ceased to circulate in humans due to rapidly acquired global herd immunity through both natural and artificial means.

The pandemic potential of a novel influenza variant is of course dependent on successful zoonotic transfer of viruses. While this is frequently observed for influenza viruses rendering this event seemingly unremarkable, other respiratory viruses are very rarely seen to have this ability, usually limited to occasional reports of transfer between humans and chimps.

Just eight years ago, influenza would have been the only example of a contemporary zoonotic respiratory virus; the emergence of SARS-CoV serves as a reminder that respiratory viruses are not so predictable, and the inter-continental dissemination of the virus touches on the effects of changing human demography on the potential for virus emergence.

Seasonal influenza A epidemics are caused by viruses with novel genomic and immunologic traits generated by host-population driven selection acting on the haemagglutinin and neuraminidase surface proteins.

The influenza epidemic peak can be identified by an increase in school absenteeism soon followed by an excess mortality in the infected population.

The General Introduction - 35 - characteristic winter seasonality of influenza virus arises through the optimal air humidity at this time of year, which facilitates virus spread as virus respiratory droplets containing virions do not be come so dense as to settle Hemmes, ; Schaffer, , and at lower temperatures respiratory mucosa are thicker, so viruses stick in the respiratory tract better Palese, Humans are the sole host of influenza B viruses, which provides the likely explanation for the lack of pandemic strains.

Influenza B comprises several co-circulating lineages Bao-Lan, ; Oxford, which are not susceptible to immunologically driven evolutionary selection pressures Air, ; Oxford, , but do undergo recombination.

An evolutionary rate for influenza B was estimated at 1. Influenza C causes mild respiratory infections in children and the elderly Katagiri, , though the epidemiologic traits of the virus are less widely defined than for other influenza viruses.

The classic studies of respiratory virus epidemiology Table 7 did not report on the characteristics of the virus for a variety of reasons.

The Cleveland study was undertaken prior to the discovery of the virus, the Seattle virus watch did not recover the virus Kim, , and the Tecumseh study was limited to retrospective determination of seroconversion rates, which were highest in the age bracket of years Troisi, Influenza C viruses, like the other influenza viruses, comprise several cocirculating lineages which undergo reassortment Buonagurio, a; Peng, , and are, like influenza B viruses, not responsive to immunologic selection pressures - the nonstructural gene of two strains isolated 19 years apart showed no nucleotide differences Buonagurio, a , and the haemagglutinin genes of two strains isolated 31 years apart differed only by two nucleotides Buonagurio, Influenza C evolves more slowly than influenza viruses A and B Buonagurio, a , and naturally infects swine Guo, General Introduction - 36 - 1.

Infection rates with parainfluenza viruses are estimated at Maternal antibodies elicit protective immunity to PIV-1 and PIV-2 in the first four months of life, but despite the presence of PIV-3 maternal antibody in this age group, the protective effect is less.

PIV-3 generally infects all individuals in a semi-closed population e. Host immunity to parainfluenza viruses is dependent on high serum antibody, and so when humoral immunologic responses wane, the host is once again susceptible to infection Ray, PIV-4 also circulates at highest frequency in late autumn or early winter, but no cyclicity is apparent Laurichesse, During this study conducted over 22 years, The incubation period for parainfluenza viruses is between two and six days Chanock, ; Kapikian, ; Smith, During primary infection with PIV-3, virus is shed for three to ten days, and shedding is of shorter duration during subsequent infections Chanock, Prolonged shedding is observed in immunocompromised individuals Gross, Parainfluenza viruses do not persist long in the environment Ansari, and transmission occurs primarily via respiratory droplet rather than through fomites.

Both viruses comprise two genogroups, A and B, distinguishable genetically and serologically Anderson, ; Mufson, ; van den Hoogen, which co-circulate with fluctuating frequencies.

The two HRSV genogroups are referred to as subgroups; these comprise genotypes distinguished on the basis of antibody cross reactivity McGill, or phylogeny Venter, Each of the two HMPV genogroups are referred to, somewhat paradoxically, as genotypes, and each genotype comprises two sub-genotypes A1, A2, B1 and B2 ; genotypes are distinguishable serologically and sub-genotypes are differentiated phylogenetically van den Hoogen, HRSV subtype A is more prevalent than subtype B Zlateva, , and whether clinical differences between the two subtypes exist is still debated, though HRSV-A has been associated with higher morbidity outcomes Taylor, Re-infections with HRSV occur throughout life, and many re-infections with the same subtype are observed, even in infancy Mufson, Re-infections in all ages are usually symptomatic Hall, HRSV epidemics occur every winter, and sporadically a particularly bad season arises Taylor, ; the reasons underlying this are unknown.

Stockton et al reported a 2. HMPV can cause asymptomatic infections Falsey, ; Williams, , though how often these occur is not established.

The only epidemiologic study of sub-clinical infection with HMPV in the community determined a 4.

General Introduction - 39 - Reports contradict as to whether mixed infection with these two viruses causes exacerbated disease Caracciolo, ; Greensill, ; McNamara, ; van Woensel, ; Wilkesmann, Nonstructural proteins common to both viruses are invariably highly conserved and include the matrix protein M , the second matrix protein M2 , the phosphoprotein P , the polymerase L , and the nucleocapsid N , which tightly encapsidates genomic RNA.

The fusion F protein mediates fusion of viral and cell membranes and is highly conserved. Anti-HRSV antibody directed against F protein is cross-reactive for strains of both subtypes Anderson, ; Hendry, ; Mufson, ; Olmsted, , and studies on HMPV using human sera and animal models have indicated similar antibody reactivity patterns Endo, ; Herfst, ; Skiadopoulos, ; van den Hoogen, General Introduction - 40 - The attachment G glycoprotein of pneumoviruses conversely exhibits several characteristics expressly for immune evasion.

In both viruses the G protein is extensively glycosylated with both N- and O-linked sugars and a high proportion of proline residues Ishiguro, ; Johnson, b; Zlateva, , thought to reduce ordered secondary structure of the protein.

It is hypothesized that switching of the predominant circulating subtype of HRSV is brought about by short-lived subtype-specific herd immunity in a population generated over one or two seasons probably against the G protein, which favours dissemination of the alternate subtype in a subsequent season Botosso, ; Scott, ; White, ; this suggestion has been borrowed to explain HMPV genotype switching also Agapov, Adenoviruses Adenoviruses are somewhat of a misfit in the respiratory virus group, as they are predominantly transmitted by the faecal-oral route Fox, Initial spread may occur via the respiratory route, but prolonged carriage of adenoviruses in the gut makes it such that faecal oral transmission is more widely observed both during the acute phase of illness and during intermittent recurrences of viral shedding in persistent infections.

Adenoviruses can persist in healthy adults for a duration of 24 months Fox, , and documented survival of adenovirus for ten years in a tear film was documented in patients with recurrent conjunctivitis Kaye, Zoonotic transmission of bovine, simian and feline adenoviruses to humans has been demonstrated by seroconversion General Introduction - 41 - Afshar, ; Phan, ; Xiang, , though infections were asymptomatic.

Recently it has been demonstrated that adenoviruses are able to undergo recombination Halstead, Postinfection, the host usually develops type-specific antibodies which may protect against disease or re-infection, but do not resolve the carrier state.

Asymptomatic adenovirus infection is frequent Fox, , Adenoviruses may circulate in both endemic and epidemic forms.

Adenoviruses follow no seasonal trends, and are serotype specific in their predilections for the age-profile of the host infected Schmitz, Epidemiologic relationships between respiratory viruses That respiratory viruses might facilitate bacterial infections was first suggested in the literature in the s, when it was demonstrated that during URTI, H.

The predisposing factors which turn asymptomatic carriage of common bacteria such as S. Excess mortality during influenza pandemics has been attributed to S.

There are two mechanisms suggested by which prior virus infection might facilitate bacterial adhesion.

Physical damage to the respiratory epithelia, possibly including ciliostasis, may prevent host clearance of invading bacteria and so aid in establishment of infection.

Alternatively, viral glycoproteins expressed on the surface of General Introduction - 42 - an infected cell could act as receptors for invading bacteria.

For example, the HRSV G protein is inserted into the membrane of the infected cell, and this expression increases binding of N. Surprisingly, neither of these mechanisms has been put forward to suggest that primary infection with a virus may facilitate secondary viral infection, nor has it been suggested that HRSV might use commensal N.

Virus-virus interactions are documented in the form of antibody-dependent enhancement ADE , whereby viruses use heterologous anti-virus antibody to mediate attachment and entry into target cells.

ADE during HRSV infection in infants has been suggested in explanation of the exacerbated morbidity seen during infections in those under 6 months of age Osiowy, , though this idea has not been followed up in the literature.

Conversely, infection with one respiratory virus may protect from infection with other viruses.

It has been noted that rhinoviruses were less likely than expected to be found in mixed infections Greer, , and this relationship has been reported more specifically for HRSV and HRV-C Wisdom, a , when an unexpectedly low mixed infection frequency with these two viruses was observed.

It is suggested that the innate immune response to one of these infections is protective from a secondary infection with the alternate virus.

Another study which was undertaken during an interval of concurrent high circulation of both HRSV and rhinoviruses observed high mixed infection frequencies Richard, , and so these observations may have been a coincidental occurrence of differing virus seasonalities.

Nevertheless, the suggestion that innate immunity stimulated during one virus infection might be protective from infection with another respiratory virus is interesting.

General Introduction - 43 - 1. Analyses of virus evolution at the genomic level RNA viruses are the most rapidly evolving entities known; the lack of proofreading activity of the RNA polymerase augments the potential for transcriptional errors.

Nucleotide substitutions may or may not result in an amino acid change, and may or may not become fixed in the virus population. Some residues are functionally conserved and so substitutions at these sites produce non-selected changes, and the likelihood of detecting such changes in a sampled virus population is minimal.

Other changes are inconsequential; their occurrence in such a large virus population renders the chances of them coming to predominance in the virus population low.

These are neutral or nearly neutral changes. Some mutations may be advantageous, for example by increasing receptor affinity or subverting host immune responses, and so the change is selected for by the host population rather than the virus.

Such changes usually occur at specific residues, and convergence of the virus population at these residues occurs.

Statistical analyses of sequence datasets can be undertaken to detect residues which are under this positive selection.

Virus evolution can be captured by observing changes in residues at both neutral and positively selected sites, visualised by phylogenetic analyses.

Phylogenetic reconstruction models the diversification events in the history of a virus population which gave rise to the sample population, and it is possible to estimate when such events occurred.

The number of years since the root of a phylogenetic tree i. Evolutionary modelling of several respiratory viruses has been undertaken.

A well conducted analysis which identified positively selected sites and the evolutionary characteristics of HRSV was undertaken in two parts corresponding with the two subtypes A and B Zlateva, , The most recent common ancestor of HMPV and avian metapneumovirus, the most closely related virus to HMPV identified to date, was estimated to have existed de Graaf, or Yang, years ago; the former study analysed data from sequences spanning twelve years, whereas the latter study analysed sequence data generated from samples collected over twenty years.

Sensitivity of the evolutionary analyses to the input sequence data is demonstrated by the disparity in the estimates of divergence times indicated by the species level and inter-species level tMRCAs.

This exemplifies the risk inherent in modelling evolutionary events with sequence data for less than ten percent of the predictive period.

Disease burden of respiratory viruses A tremendous ethical debate ensues on publication of any method which attempts to quantify the burden of any disease, yet the value of such studies in providing indications of where limited resources should be diverted to maximize their return in terms of human life validates the dogged pursuit of acceptable methodologies.

QALYs were used to identify where resources should be diverted in order to maximize a social utility outcome, that is, where people perceive the greatest benefits to quality of General Introduction - 45 - life are to be gained, and are utilitarian in their aim to do the greatest good for the greatest number of people.

QALYs identify Health Related Quality of Life HRQL weightings on a scale between zero and one, where zero is death and one is full health, without specific linkages to diseases, conditions or disabilities, but rather directed by the values of individuals and how they perceive their own health state.

A variety of methods to standardise perceptions of health across ages, cultures and genders while including considerations of physical, psychological, and social parameters have been described.

One of the most common used is asking respondents what they would be willing to sacrifice in terms of risk of death or time in order to return from their current status to perfect health.

This is nevertheless inevitably subject to variability inherent between sampling populations.

Health professionals have been found to provide lower values than others when scoring illnesses. To overcome the practical difficulties associated with collecting comparable primary data on a global scale and the concerns raised over self-assessments of health by using QALYs, the need for a new way of quantifying disease burden was identified.

Quantification of the global burden of premature death, disease and injury in order to identify optimum strategies for improvement of health globally was undertaken in a collaborate effort by the World Bank and the World Health Organization, which in heralded the introduction of the Disability Adjusted Life Year DALY model.

This, for the first time, provided the means to assess whether one intervention or another for two different diseases would provide the greatest benefit e.

Reviews of this approach have nevertheless often been critical with inferences made that panelists were led in their scoring. The practical drawback of this approach in comparison with QALYs is the inability to account for co-morbidities.

In conclusion, both QALYs and DALYs strive to produce summary measures of population health which combine the effects of morbidity and mortality for simultaneous consideration.

Notwithstanding, such calculations currently represent the most sophisticated means with which to directly compare health outcomes, and without strategies for quantification of disease, policy makers are left in the dark when making decisions on the optimal way to allocate finite resources.

Quantification of the global burden of disease has been undertaken for HRSV using the DALY model Nair, , and the study concluded the tremendous value of General Introduction - 47 - development of vaccines, prophylaxes and treatments to reduce the morbidity and mortality caused by this virus.

HRSV is widely purported as the single greatest cause of viral ARTI regardless of sampled population, yet without some form of direct comparison between respiratory viruses the value of this study serves only to emphasise the importance of HRSV without making clear the need for investment into researching interventions for this virus rather than other respiratory viruses.

General Introduction - 48 - 2. Materials and Methods This chapter contains general descriptions of the materials, methods and software used during the course of study, and is divided into three sections.

The first describes the samples used, and the second outlines the laboratory methods and materials employed. Computational techniques are detailed in the third section.

Retained information recorded in an Excel database included patient age Figure 3 and sex, month and origin of sample collection Figure 4, Figure 5 , clinical information recorded on referral forms, and routine diagnostic screening results.

Figure 3. Patient ages were recorded on a banded basis in accordance with ethical restrictions, corresponding with the ranges indicated.

Methods - 50 - Samples collected between July and June were anonymised using sequential ascending integers3. A nucleic acid archive complementary to the sample archive comprising nucleic acids extracted from samples during diagnostic testing was also assembled.

Figure 4. Between 1 patients and 80 1 3 There were two good reasons for terminating epidemiologic studies from this time.

Primarily, the influenza pandemic changed the sampling frame from around this time, with GPs sending an increased proportion of samples and some samples were taken for screening of asymptomatic contacts.

Additionally, the ethical requirement for anonymisation of all samples and nucleic acids became overwhelming for one individual due to the increased influx of samples it took a day to re-label around a thousand samples or fifteen hundred nucleic acids.

Methods - 51 - patient samples were collected from each patient. Repeat samplings from the same patient arose through both re-sampling during one illness, and sampling from temporally discrete illnesses.

In all, referral sites were recorded. Most samples were hospital referred Figure 5. Origin of all samples collected between July and June total Methods - 52 - Figure 6.

The proportion of samples in each group in which at least one virus from the routine panel described in 2. Nasal secretions, nasal swab and perinasal swab were grouped under nasal.

Oral included oral secretions, saliva, and mouth swab. Tracheal consisted of tracheal aspirate, endotracheal tube, endotracheal secretions, tracheal secretions, throat swab, and tracheal ring.

Samples for which the origin was not clear e. Breakdown of the physiological origin of samples collected over the three year study period is indicated Figure 6.

Diagnostic testing For respiratory virus diagnosis, respiratory samples are typically screened by molecular amplification methods such as the polymerase chain reaction PCR for a panel Methods - 53 - of viruses which varies between diagnostic laboratories.

Figure 7. Number of samples and proportion for which a virus included in the routine panel was detected is indicated at the head of each column.

Only a fraction of the samples were tested for these viruses, so the results could not be included in epidemiologic study. Methods - 54 - 2.

Clinical data Clinical information reported on referral forms was recorded in the respiratory database and divided into seven categories.

These were: 1. Included samples collected from patients with neoplasia, a known immunosuppressive disorder and transplant patients.

Samples collected from patients with chronic respiratory conditions such as asthma and cystic fibrosis.

Lower respiratory tract infection LRTI. Samples taken from patients with pneumonia, bronchiolitis, bronchitis, influenza-like illness ILI , tested for HRSV using a near-patient test NPT 5 Mackenzie, , chest infection, shortness of breath, atypical pneumonia, acute respiratory distress syndrome ARDS , difficulty breathing, community acquired pneumonia, cough, wheeze and respiratory failure if a sample number from the same patient preceding this clinical report was associated with URTI or LRTI.

Upper respiratory tract infection URTI. Tonsillitis, coryza, rhinorrhoea and sore throat were classified as URTI. Clinical evidence for respiratory tract infection was indeterminate e.

No data. No data was recorded for Clinical data relating to some samples could be assigned to more than one category, and so categories were treated in a hierarchical fashion with precedent given in the order as above.

The relative frequency of the described clinical categories is indicated Figure 7. Infants presenting with symptoms of lower respiratory tract infection requiring hospitalization are tested for HRSV using a point of care test to guide cohorting.

Methods: Laboratory based techniques Laboratory techniques were undertaken at two sites. Automated nucleic acid extraction 2.

At both research sites, a one way flow through laboratory system is operational to reduce risk of contamination.

These were replenished by sample re-extraction using the same protocol used for diagnostics of automated BioRobot MDx extraction for continuity.

Extraction of nucleic acids from specimens used in studies of molecular evolution or as positive controls where a standardised approach to extraction was not essential was undertaken using Qiagen Southampton, UK viral RNA mini kit following the protocol for purification of viral RNA from sample fluid by centrifugation, with a double elution step.

Methods - 56 - 2. Pooling of respiratory RNA To maximize the number of samples tested, screening of pooled nucleic acids was undertaken during epidemiologic studies.

Pooling has previously been demonstrated as an effective technique for efficient screening of respiratory samples for viruses circulating at sufficiently low frequency e.

Reverse transcription To enable amplification of viral nucleic acids by nested PCR, single nucleic acids and RNA pools were converted to cDNA by reverse transcription RT using Promega Southampton, UK A kit according to manufacturers instructions for use of random primers, modified to incorporate an extended elongation step of 55 minutes.

Successful amplification of the target region was confirmed by agarose gel electrophoresis 2. This enzyme metabolises X-Gal.

One culture was selected and plasmid was extracted using Qiagen Crawley, UK miniprep kit according to manufacturer instructions.

Polymerase Chain Reaction PCR techniques The polymerase chain reaction PCR amplifies nucleic acids by a cyclic chain of DNA denaturation, primer annealing and nucleotide extension events, each of which occur optimally at different temperatures.

PCR played a dual role in this work, used for virus detection and sequence analysis. Primer design Design of primers was undertaken by downloading available target region sequences from Genbank and aligning in Simmonics software www.

The primer pair was inputted into the search, and the results returned for products complementary to these primers were checked to ensure they matched the target product, and that no undesired targets were amplified.

The positive control was usually a clinical specimen. All PCR reactions comprised the same basic reagents. Methods - 60 - Figure 8.

The algorithmic process of PCR protocol design. PCR protocol variables are described in white boxes which follow the chain of events in setting up a PCR reaction.

Questions to determine the next stage of optimization are in grey boxes. Decisions are in black boxes. Methods - 61 - 2. Virus detection was by direct visualization of fluorescence emitted by virus-specific probes on ABI bioluminescence machines Applied Biosystems technologies.

Probes bound to a specific nucleotide sequence were cleaved on complementary DNA binding, releasing the probe to emit fluorescence, and so the more DNA amplified the stronger the fluorescence refer to 1.

Amplified nucleic acid was detected directly using Bioluminescence ABI machine. Negative control product from automated extraction was used as negative control.

Component samples of coronavirus positive pools in this group were screened using the 7 Dr Kate Templeton who supervised this project previously worked at Leiden University Medical Centre, Leiden, The Netherlands.

With a colleague there, Dr Eric Claas, Kate designed but did not publish this protocol. Methods - 62 - same four way multiplex assay to detect constituent positive samples.

Additionally, components of nine coronavirus negative pools were screened by the multiplex assay. Table 9.

Primers are described in Table 8. IS, inner sense; IAS, inner antisense. Synthesis of high copy number coronavirus RNA and multiplex assay sensitivity To assess the sensitivity of the multiplex assay, a standard curve was constructed using ten-fold serially diluted coronavirus RNA of known copy number, and the limit of detection was assumed to be the RNA copy number theoretically detectable after 50 PCR cycles.

PCR product was cloned into pCR2. Methods - 63 - transformation of electrocompetent E. Blue colonies were picked and success of transformation was confirmed by sequence analysis.

Table PCR product was quantified and checked for purity by spectrophotometry using nanodrop testing for absorbance at wavelengths of A and A These RNA standards were subject to the multiplex assay in triplicate to construct a standard curve.

Second round PCR was then undertaken as previously described. This method successfully amplified HRSV RNA in around half of cases and so the insufficiencies of the PCR techniques were attributed to RNA degradation, possibly 10 Platinum Taq polymerase is a recombinant Taq polymerase complexed with an activity blocking antibody at room temperature.

At high temperatures, antibody is denatured and polymerase activity is restored. This prevents non-specific annealing of the polymerase, thereby increasing sensitivity and reducing the need for PCR optimization.

Methods - 65 - arising due to the inevitable freeze-thaw associated with repeatedly moving RNA within and between sites.

Agarose gel electrophoresis PCR positive samples were detected by visualization of reaction product by agarose gel electrophoresis.

Loading dye adds colour and density to the sample for ease of loading and is negatively charged, so will migrate unidirectionally with DNA.

Gels were immersed in tanks containing 1xTAE solution at volts for minutes, and visualized using a UV transilluminator.

Sequencing reaction Nucleotide sequencing was undertaken using the Sanger method of a cyclic reaction wherein at each stage one fluorescently tagged nucleotide is knocked off the transcript and detected.

To prepare transcripts comprising fluorescent signals, DNA was amplified using fluorescently tagged nucleotides For each amplicon this reaction was undertaken twice, once in the sense and once in the antisense direction with inner primers used accordingly.

This removed free dNTPs and primers to improve accuracy of sequencing. Methods: Computational analyses 2. Sequence alignment Sequences generated during the course of this study were used for molecular epidemiologic and evolutionary analyses.

Nucleotide sequencing results were returned from the Genepool sequencing service by email, and were imported into the most contemporary version of the in-house sequence alignment and analysis software program Simmonics www.

Phylogenetic analyses Phylogenetic reconstruction produces a topography representative of modeled ancestral relationships for a taxonomic group.

There are several software packages available for phylogenetic reconstruction, which use a variety of techniques for establishing the tree which best fits input sequence data.

The most widely used models for phylogenetic reconstruction are briefly summarized Table Phylogenetic analyses were conducted by nearest neighbour joining from samplings of maximum composite likelihood Methods - 67 - MCL distances with pair-wise deletions for missing nucleotides and gamma distributed rates among sites to allow variability in substitution rates between nucleotide positions.

Neighbour joining trees are constructed iteratively whereby all branches are initially connected by one internal node to give a star-like tree.

A pairwise distance matrix is calculated representing the sum of the branch lengths across the tree if each pair of taxa was joined.

An internal branch is added to join the pair of taxa giving the smallest sum of branch lengths. Neighbour joining trees are computationally efficient and so can be used on large datasets, do not assume all lineages evolve at the same rate and produce an unrooted tree.

However NJ assumes that no backward or parallel substitutions occur. Neighbour joining trees are exposed to Long Branch Attraction LBA Felsenstein, , whereby differentially evolving lineages may be subject to multiple substitutions at specific sites, thereby increasing likelihood that the neighbour joining model will predict that two strains are more closely related than they actually are.

As such, NJ trees are not ideal for sequence sets under positive selection. Maximum composite likelihood is ideal for phylogenetic reconstruction of rapidly evolving sites as it corrects for multiple mutational events at the same site.

MCL modifies the estimate of evolutionary distances between sequences in a pairwise fashion by fitting common parameters for nucleotide substitution events to every sequence pair.

Different transition purine to purine or pyrimidine to pyrimidine substitutions and transversion purine to pyrimidine or pyrimidine to purine substitutions ratios are modeled, reflecting the observation that transitions occur more readily than transversions.

Estimation of pairwise evolutionary distances by maximum composite likelihood overcomes any exposure to LBA. Trees were rooted using an outlier e.

Methods - 68 - Table Summary of models commonly cited for phylogenetic tree construction. Detailed in 2.

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