Thesis (MScMedSc (Pathology. Medical Virology))--University of Stellenbosch, 2009. / Submitted in fulfilment for the degree MSc in BioMedical Science at Stellenbosch University. / ENGLISH ABSTRACT: HIV-1 was first diagnosed within South Africa in 1982. In the 1980’s homosexual transmission dominated the HIV-1 epidemic within the country. In the late 1980’s the second HIV-1 epidemic was recognized amongst heterosexual individuals. Today heterosexual transmission of HIV-1 dominates the epidemic in South Africa. Subtype C HIV-1 is responsible for the overwhelming majority of heterosexual infections. An estimated 95% of all infections in the country are thought to be subtype C related. To date only a few papers have been published on non-subtype C HIV within the country. This study characterized subgenomic and near full-length sequences of non-subtype C HIV-1 viruses from the Cape Town area. The gag p24, pol-integrase, and env gp41 regions of 11 of the 12 samples were characterized by amplification and direct sequencing. Phylogenetic analysis of the sequenced data, with online subtyping tools (REGA and jpHMM) and the drawing of NJ-trees revealed the presence of subtype A1, B, F1 and recombinant viral forms such as AD, AG and AC. One of the isolates was classified as a subtype C and was included for control purposes. Near full-length characterization of four of the samples were attempted, through full genome PCR amplification and sequencing. Analysis of sequenced data with the use of subtyping-, recombination identification, and tree drawing tools revealed a subtype B, and A1 isolate. The other two isolates were identified as possible AC and AD recombinants. The data that was generated will greatly improve our knowledge of non-subtype C isolates circulating within South Africa. Due to the possible impact that the high degree of genetic variation that HIV may have on vaccine design and development and ARV treatment and HIV diagnosis, ongoing research of the epidemiology and spread of HIV within South Africa are needed. / AFRIKAANSE OPSOMMING: MIV was in 1982 vir die eerste keer in Suid Afrika gediagnoseer en was hoofsaaklik deur homoseksuele kontak oorgedra. Aan die begin van die 1990’s is `n tweede MIV epidemie gewaar onder heteroseksuele individue. Heteroseksuele oordrag van die virus domineer tans die MIV epidemie in Suid Afrika en is meestal subtipe C verwant. Subtipe C, MIV-1 is verantwoordelik vir 95 persent van alle infeksies in die land. Tot hede is slegs `n paar publikasies oor die nie-subtipe C epidemie in die land gepubliseer. Die huidige studie was gemik op die karakterisering van subgenomiese en vollengte genome van nie-subtipe C MIV isolate van die Kaapstad omgewing. Die gag p24, pol-integrase en env gp41 subgenomiese fragmente van 12 monsters was gekarakteriseer deur amplifikasie en DNS nukleotied volgorde bepaling. Filogenetiese analise deur middel van subtipering (REGA en jpHMM aanlyn subtiperings programme) asook NJ-filogenetiese bome van die data het die teenwoordigheid van subtipe A1, B, en F1, asook verskeie rekombinante viruse insluitende AG, AD en AC vorme aangedui. Een van die isolate was geklassifiseer as `n subtipe C maar is in die studie ingevoeg vir kontrole doeleindes. Vollengte karakterisering van 4 uit die 12 isolate was ook gedoen deur vollengte genoom amplifikasie en DNS nukleotied volgorde bepaling. Tydens die analisering van die DNS volgorde data, deur middel van aanlyn subtipering, rekombinasie identifikasie (Simplot en RIP), en filogenetiese boom konstruksie programme is twee isolate geidentifiseer as subtipe B en A1 MIV-1 viruse. Die ander twee isolate was as moontlike AC en AD rekombinante geklassifiseer. Die data van nie-subtipe C MIV isolate sal ons kennis van die nie-subtipe C epidemie in Suid Afrika versterk. As gevolg van die impak wat die hoë graad van genetisie variasie van MIV op die ontwikkeling van entstowwe, sowel as die diagnose en behandeling van pasiente kan hê, is verdere navorsing in die epidemiologie van die MI-virus in Suid Afrika nodig.

Thesis (PhD (Biomedical Sciences. Molecular Biology and Human Genetics))--University of Stellenbosch, 2009. / Dissertation presented for the degree of Doctor of Philosophy at Stellenbosch University. / ENGLISH ABSTRACT: This study describes the molecular epidemiology of Mycobacterium tuberculosis strains with the Beijing genotype. This genotype has received clinical prominence due to its global distribution and the hypothesis that these strains have acquired the ability to evade the protective effect of BCG vaccination, spread more readily, acquire drug resistance and cause severe forms of disease. Molecular biological techniques were used in a series of studies to elucidate the genetic evolutionary mechanisms underlying the success of this genotype in Cape Town, South Africa. Using a collection of 40 different markers it was possible for the first time to construct a phylogenetic history of Beijing genotype strains. This phylogeny was characterized by the consecutive evolution of 7 sublineages. Analysis of epidemiological data in relation to these sublineages showed an association between more recently evolved Beijing strains and an increased ability to transmit and cause disease. From these findings it was hypothesized that the pathogenic characteristics of the Beijing genotype were not conserved but rather that strains representative of the different sublineages had evolved unique properties. In order to determine whether these socalled unique properties were associated with either the host population or the genetic background of strains from sublineage 7, a meta-analysis of published Mycobacterial Interspersed Repetitive-Unit (MIRU) typing data (East Asia) was compared with MIRU typing data from the South African strains in the context of their phylogenetic histories. This study showed that Beijing genotype strains in South Africa originated in East Asia following their introduction during the early 18th century. A significant association was observed between the frequency of occurrence of strains from defined Beijing sublineages and the human population from whom they were cultured (p <0.0001). Based on these findings it was proposed that either the host population (South African) had selected for a particular Beijing sublineage (i.e. sublineage 7) or that strains from that sublineage had adapted to be more successful in the South African population. In a subsequent study, using the methodology developed in the above studies, it was shown that strains from the ancestrally positioned lineage (termed “atypical” Beijing genotype) were over-represented in drug resistant isolates in the Eastern Cape region. This contradicts current dogma which suggests that “atypical” Beijing genotype strains are attenuated in their ability to transmit. However, this phenomenon may be ameliorated in immune-compromised patients as review of the clinical records showed that transmission was associated with HIV co-infection. These findings highlight the need to improve tuberculosis control in vulnerable populations as strains which would normally not contribute significantly to the epidemic now become a cause for concern especially if they are associated with drug resistance. To improve our understanding of the evolution of the Beijing genotype, the genomic stability of an additional 27 polymorphic markers were analysed. These markers have recently been proposed as the new standard in molecular epidemiological studies and were based on MIRU-Variable Number Tandem-Repeats (VNTR) sequences. Superimposition of the MIRU-VNTR data onto the phylogenetic tree showed excellent concordance thereby demonstrating that these alleles were largely stable over time. It is currently not known how the alleles that do change could influence pathogenicity. The results of this study also demonstrated discordance between strains defined by IS6110 DNA fingerprinting and those defined by MIRU-VNTR typing thereby demonstrating that these markers evolve independently and at different rates. Furthermore, the MIRU-VNTR typing method was unable to predict transmission of drug resistant strains which contradict previous reports from low incidence settings. This has significant implications for the use of this typing method in high incidence settings. Using an improved PCR-based method it was possible for the first time, to identify the 5 most prominent phylogenetic lineages in primary cultures of adult tuberculosis patients resident in a high HIV/TB co-infection setting. The results of this study showed that 15% of the study population was infected with two or more strains and Beijing genotype strains were over-represented in these mixed infections. Furthermore, drug susceptibility tests showed that one patient was co-infected with both a drug sensitive and a drug resistant strain. Since mixed infections have been implicated in treatment failure, these findings demonstrate the epidemiological importance of detecting mixed infections in vulnerable populations. This PCR-based method was further applied to cultures of paediatric tuberculosis patients to classify strains which spoligotyping was unable to define. The result of this study showed three mixed infections which otherwise would have been missed. In order to determine whether clinical disease presentation of patients infected with strains of the Beijing genotype were different from that of patients infected with non-Beijing genotype strains, clinical and demographic data of these two groups were analysed. This study showed that patients infected with strains of the Beijing genotype were highly infectious as defined by the increased bacterial load in sputum specimens. However, this finding could not be validated by lung pathology according to chest radiographs of infected patients. / AFRIKAANSE OPSOMMING: Hierdie studie beskryf die molekulêre epidemiologie van Mycobacterium tuberculosis rasse met die Beijing genotipe. Hierdie genotipe is van groot kliniese belang weens hul globale verspreiding en die hipotese dat hierdie rasse die vermoë ontwikkel het om die beskermende effek van BCG vaksinasie te vermy, om meer geredelik te versprei, middelweerstandigheid te ontwikkel en erger vorms van siekte te veroorsaak. Molekulêre biologiese tegnieke is gebruik in ‘n reeks studies om die genetiese evolusionêre meganismes onderliggend tot die sukses van hierdie genotipe in Kaapstad, Suid-Afrika te verklaar. Deur ‘n versameling van 40 verskillende merkers te gebruik, was dit moontlik om vir die eerste keer ‘n filogenetiese stamboom van die Beijing ras genotipe te skep. Hierdie filogenie word gekenmerk deur die opeenvolgende evolusie van 7 ras sublyne. Met die analise van epidemiologiese data in verhouding tot hierdie ras sublyne, is ‘n assosiasie tussen die mees onlangs ontwikkelde Beijing rasse en die verhoogde vermoë om te versprei en siekte te veroorsaak, getoon. Vanweë hierdie bevindinge, is ‘n hipotese daargestel dat die patogeniese kenmerke van die Beijing genotipe nie in alle raslyne voorkom nie, maar eerder dat verteenwoordigende rasse van die verskillende sublyne unieke eienskappe deur evolusie ontwikkel het. ‘n Metaanalise van gepubliseerde MIRU tipering data van Oos-Asië is vergelyk met MIRU tipering data van Suid-Afrikaanse rasse in die konteks van hul filogenetiese geskiedenis om te bepaal watter van hierdie sogenoemde unieke eienskappe geassosieer is met die gasheerpopulasie en watter eienskappe geassosieer is met die genetiese agtergrond van die sublyn 7 rasse. Hierdie studie het getoon dat die Beijing ras genotipe van Suid-Afrika hul oorsprong gekry het van Oos-Asië en vir die eerste keer waargeneem is in die vroeë 18de eeu. ‘n Betekenisvolle assosiasie is waargeneem tussen die frekwensie waarteen die rasse van ‘n bepaalde Beijing sublyn voorkom en die menslike populasie van wie hulle geïsoleer is (p < 0.0001). Gebaseer op hierdie bevindinge is dit voorgestel dat die menslike populasie (Suid-Afrikaners) vir ‘n spesifieke Beijing sublyn geselekteer het (bv. Sublyn 7) of dat rasse van hierdie sublyn aangepas het om meer suksesvol te wees in die Suid-Afrikaanse populasie In ‘n daaropvolgende studie is, deur gebruik te maak van die metodiek wat ontwikkel is vir die bogenoemde studies, getoon dat die voorouerlike sublyn (bekend as die“atipiese” Beijing genotipe) die mees verteenwoordigende sublyn was onder middelweerstandige isolate van die Oos-Kaap gebied. Dit is teenstrydig met die bestaande dogma wat bepaal dat die “atipiese” Beijing genotipe rasse hulle vermoë om te versprei verloor het. Hierdie verskynsel kan egter versterk word in immuun inkompetente pasiënte aangesien hersiening van die kliniese rekords aangedui het dat verspreiding geassosieer was met HIV ko-infeksie. Hierdie bevindinge bring die behoefte om TB beheer in vatbare populasies te verbeter, na vore, omrede rasse wat gewoonlik `n onbetekenisvolle bydrae tot die epidemie lewer, nou ‘n rede vir kommer is veral as hulle met middelweerstandigheid geassosieer is. Om ons insig rakende die evolusie van die Beijing genotipe te verbeter, is die genomiese stabiliteit van ‘n addisionele 27 polimorfiese merkers geanaliseer. Daar is onlangs voorgestel dat hierdie merkers, wat gebaseer is op MIRU-VNTR volgordes,die nuwe standaard vir molekulêre studies is. Die MIRU-VNTR data is op die filogenetiese boom geplaas en het uitstekende ooreenstemming getoon wat die allele se stabiliteit oor tyd gedemonstreer het. Dit is tans nie duidelik hoe van die allele wat wel verander, die patogenisiteit beïnvloed nie. Die resultate van die studie wys ook onenigheid tussen rasse wat deur IS6110 DNA tipering gedefinieer is en dié wat deur MIRU-VNTR tipering gedefinieer is. Dit impliseer dus dat die evolusie van merkers onafhanklik van mekaar plaasvind en teen verskillende tempos. Verder was die MIRU-VNTR tipering metode nie in staat om verspreiding van middelweerstandige rasse te voorspel nie, wat teenstrydig is met vorige verslae waar lae insidensie omgewings bestudeer is. Dit het noemenswaardige implikasies vir die gebruik van hierdie tipering metode in hoë insidensie omgewings. ‘n Verbeterde PKR-gebaseerde metode is vir die eerste keer gebruik om die 5 mees prominente filogenetiese sublyne in primêre kulture van volwasse tuberkulose pasiënte van ‘n hoë MIV/TB ko-infeksie omgewing, te identifiseer. Die resultate van hierdie studie het gewys dat 15% van die studiepopulasie geïnfekteer is met twee of meer rasse en dat die Beijing genotipe ras die meeste voorgekom het in gemengde infeksies. Verder het middelweerstandige toetse gewys dat een pasiënt geïnfekteer was met beide ‘n middelsensitiewe en ‘n middelweerstandige ras. Gemengde infeksies is al vantevore gekoppel aan onsuksesvolle behandeling en dus demonstreer hierdie bevindinge die epidemiologiese belang van die opsporing van gemengde infeksies in vatbare populasies. Hierdie PKR-gebaseerde metode is verder gebruik om rasse wat voorkom in kulture van pediatriese pasiënte, wat spoligotipering nie kon klassifiseer nie, te klassifiseer. Die resultate het drie gemengde infeksies gewys wat sonder die PKR-gebaseerde metode, nie geïdentifiseer sou gewees het. Om te bepaal of die kliniese beeld van pasiënte wat geïnfekteer is met rasse van die Beijing genotipe verskil van dié van pasiënte wat geïnfekteer is met rasse van die nie-Beijing genotipe, is die kliniese en demografiese data van die twee groepe pasiënte geanaliseer. Hierdie studie wys dat pasiënte wat geïnfekteer is met rasse van die Beijing genotipe hoogs aansteeklik is (gedefinieer op grond van hoë bakteriële lading in sputum monsters). Hierdie bevindinge kon egter nie met behulp van long patologie op borskas X-strale bevestig word nie.

DNA repair is a vital process of a living organism. Inherited or acquired defects in DNA repair systems and cellular surveillance mechanisms are expected to be important, if not crucial factors in the development of human cancers. DNA repair is a multigene and multifactorial process which is most comprehensively characterized by the phenotypic evaluation of DNA repair capacity (DRC). DRC represents a complex marker with high informative value, as it comprises all genetic, epigenetic and non-genetic factors, by which it is modulated. Accordingly, DRC reflects the actual capability of the cell, tissue or organism to protect its DNA integrity. The present PhD study was focused on investigating DRC, which specifically involves base and nucleotide excision repair pathways, in human populations with different characteristics. The main aim was to answer substantial questions on the possible use of DRC as biomarkers in epidemiological studies. The study was in fact designed to understand the extent of physiological variability of DRC in a population, its modulation by genetic and non-genetic factors, tentative adaptability to high genotoxic stress and, finally, its involvement in cancer aetiology. In order to explore these issues, DRC, in respect to genetic and environmental variability, was investigated...

As doenças virais de peixes causadas por vírus das famílias Iridoviridae e Alloherpesviridae estão distribuídas mundialmente e representam uma ameaça real a expansão do sistema aquícola mundial. Desse modo, tornam-se necessários o aprimoramento e aplicação das técnicas de diagnóstico existentes a fim de se controlar ou impedir o avanço dessas viroses entre países. Vírus do gênero Megalocytivirus (MV) e o Cyprinid herpesvirus (CyHV), tipos 1, 2 e 3, induzem enfermidades em peixes com elevada taxa de mortalidade. Já vírus do gênero Lymphocystivirus (LCDV), apesar da menor patogenicidade, causam graves perdas econômicas ao reduzirem o valor comercial dos animais infectados. Devido à escassez de dados disponíveis sobre a circulação desses vírus no Brasil, o presente estudo objetivou a prospecção, incluindo a detecção e caracterização por meio da utilização de técnicas moleculares, de MV, LCDV, CyHV-1, CyHV-2 e CyHV-3 em peixes ornamentais oriundos do município de São Paulo, bem como em espécies de peixes de vida livre e comerciais provenientes de 4 estados brasileiros. Para tanto, foram coletadas e caracterizadas amostras de 306 peixes de 47 espécies diferentes, sendo que todas foram processadas para CyHV-3 e 81 foram submetidas ao diagnóstico para os demais vírus, obtendo-se uma positividade de 47% para MV, 1,2% para LCDV e 2% para CyHV-3. Todas as amostras positivas eram oriundas de peixes ornamentais do município de São Paulo. A sequência nucleotídica de LCDV obtida agrupou-se filogeneticamente em clado do genótipo VII, com similaridade de 44,8-91,1% para com outras sequências homólogas. As sequências de MV agruparam-se em clado com outras sequências de origem asiática, exibindo similaridade de 47,2-99,2% para com outras sequências. Para CyHV-3, a similaridade entre as sequências obtidas e para com outras homólogas foi igual ou superior a 97,4%, agruparando-se em 2 clados distintos. Análises evolutivas indicaram seleção negativa fraca ou relaxada para todas as sequências dos 3 grupos virais estudados. Nesse sentido, destaca-se o ineditismo dos resultados obtidos para a melhor compreensão da epidemiologia molecular desses vírus no Brasil. / Viral fish disease caused by viruses of Iridoviridae and Alloherpesviridae families are distributed globally and represent a real threat to the expansion of world aquaculture system. Thus, improvement and application of current diagnostic techniques in order to control or stop the spread of these infections among countries become necessary. Virus of Megalocytivirus (MV) gender and the Cyprinid herpesvirus (CyHV), types 1, 2 and 3, induce diseases in fish with high mortality rate. Virus of Lymphocystivirus (LCDV) gender, despite their lower pathogenicity, cause severe economic losses by reducing the commercial value of infected animals. Due to the paucity of data available on the circulation of these viruses in Brazil, this study aimed to prospecting, including the detection and characterization through the use of molecular techniques, MV, LCDV, CyHV-1, CyHV-2 and CyHV-3 in ornamental fish come from the city of São Paulo, as well as in fish species of free and commercial life from 5 Brazilian states. For this purpose, 306 samples of 43 different species of fish were collected and characterized, all of which were processed to CyHV-3 and 81 were subjected to the diagnosis of other viruses, yielding a positivity rate of 47% for MV, 1.2% for LCDV and 2% for CyHV-3. All positive samples were from ornamental fish of São Paulo municipality. The only LCDV-nucleotide sequence obtained was grouped phylogenetically in the genotype VII clade, showing 44.8-91.1% similarity to other homologous sequences. Sequences of MV grouped into clade with Asian sequences, displaying 47.2-99.2% similarity to other homologous sequences. For CyHV-3, the similarity among the sequences obtained in this study and the similarity to homologous sequences was equal or superior to 97.4%, grouping into 2 distinct clades. Evolutionary analysis indicated weak or relaxed negative selection for all the sequences from the 3 studied virus groups. In this sense, it can highlight the novelty of the obtained results for a better understanding of the molecular epidemiology of these viruses in Brazil.

ABSTRACT Extended-spectrum beta-lactamase (ESBL)-producing Salmonella enterica serotype Isangi has emerged as a common Salmonella serotype affecting mainly children in hospitals throughout South Africa. Between 2000 and 2002, 279 S. Isangi isolates from single infection episodes were referred from 21 hospitals in 5 provinces to the Enteric Diseases Reference Unit of the National Institute for Communicable Diseases of South Africa. All isolates were subjected to antibiotic susceptibility testing and three disk-diffusion methods confirmed ESBL-production in 273 isolates. PCR and nucleotide sequencing of 101 isolates identified TEM-1 (2%), TEM-63 (91%), a novel TEM-131 (7%), and SHV-5 (2%), but CTX-M was not found. Plasmid profiling produced types with 1 to 6 plasmids, 7.4kb to 166kb in size, which were neither serotype nor ESBL-type specific. Pulsed-field gel electrophoresis revealed four major clusters while sub-clusters with identical, or near identical banding patterns suggested extensive intra-hospital transmission and clonal spread between hospitals and provinces in South Africa.

<p>Characteristically, in infections that are caused by the zoonotic pathogen <i>Bartonella</i> naturally infected reservoir hosts are asymptomatic, where infected incidental, non-natural, hosts develop symptomatic disease. Cat-scratch disease (CSD) is a well known example. <i>Bartonella </i>infections in humans may be self-limiting or fulminant and affect different organ systems. </p><p>The objectives of the present thesis were to (1) identify and characterise <i>Bartonella </i>infection cases in Sweden, (2) to investigate certain human populations regarding <i>Bartonella </i>infections, and (3) compare natural populations of different <i>Bartonella </i>species.</p><p>Cases with typical and atypical CSD were recognised by using a combination of PCR and serology. Gene sequence comparisons of different genes in <i>B. henselae</i> isolates from the United States and Europe showed that<i> fts</i>Z gene variation is a useful tool for <i>Bartonella</i> genotyping. </p><p>Myocarditis was a common finding among Swedish elite orienteers succumbing to sudden unexpected cardiac death (SUCD). The natural cycle of <i>Bartonella</i> spp., the life style of orienteers, elevated antibody titres to <i>Bartonella</i> antigens, <i>Bartonella</i> DNA amplified from myocardium and the lack of another feasible explanation make <i>Bartonella</i> a plausible aetiological factor.</p><p>The first reported case of <i>Bartonella</i> endocarditis (<i>B. quintana</i>) was identified in an immunocompromised patient who underwent heart valve replacement. The patient had been body louse-infested during his childhood. It is hypothesised that a chronic <i>B. quintana</i> infection was activated by the immunosuppression.</p><p>There was no evidence of an ongoing trench fever (TF) epidemic in a Swedish homeless population, although an increased risk for exposure to <i>Bartonella</i> antigens was demonstrated. The lack of louse infestation might explain the absence of <i>B. quintana</i> bacteremia and low <i>B. quintana</i> antibody titres. </p><p>Comparisons of genetic loci and the whole genomes of environmental <i>B. grahamii</i> isolates from the Uppsala region, Sweden displayed variants that were not related to specific host species but to geographic locality. Natural boundaries seemed to restrict gene flow.</p>

Thesis (Ph.D.)--Ohio University, November, 2009. / Release of full electronic text on OhioLINK has been delayed until December 1, 2010. Title from PDF t.p. Includes bibliographical references.

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / Microbiology / Master / Master of Philosophy

Recently, progress was made in collecting, classifying, and characterizing the genetic diversity of type 2 porcine reproductive and respiratory syndrome virus (PRRSV) using all known and publically available sequencing information. Despite this voluminous attempt, these analyses were largely na?ve of the Canadian contribution to circulating viruses. This represented a vital omission in the study of molecular epidemiology due to the fact that Canada had recorded the earliest evidence of the existence of type 2 PRRSV. To this end, the genetic diversity and evolutionary aspects of PRRSVs distributed in the Province of Ontario in Canada were characterized to abridge this existing knowledge gap on type 2 PRRSV. Genotyping of type 2 strains is primarily based on either a phylogenetic or restriction fragment length polymorphism (RFLP) approach. Classification of Ontario PRRSV field isolates (n = 505) from 1999 to 2010, based on a global type 2 PRRSV ORF5 phylogenetic framework, revealed genetic diversity comparable to PRRSV in the USA, with sequences assigned to five of nine lineages (1, 2, 5, 8 and 9). A majority (~85%) of these isolates were typed to the first two lineages (1 and 2). Despite a relatively smaller sample size to the USA, the topology of the phylogenetic tree indicated Canadian origins of these two lineages. Mapping RFLP patterns of Ontario isolates onto the phylogenetic tree revealed numerous examples of different patterns located within the same phylogenetic cluster. Examples of the non-specificity of RFLP patterns to any particular lineage or sub-lineage were abound. Statistical analysis showed occurrences where similar RFLP patterns masked diverse genetic distances and instances of close genetic proximity with divergent RFLP patterns. An examination of the most abundant 15 RFLP patterns revealed that the discrepancy between RFLP typing and genetic distances was not attributable to a single or few patterns but was rather a permeating feature. Importantly, the tree topology also indicated a Canadian ancestry for the highly virulent MN184-related strains that first emerged in 2001 in the USA. Selective pressure analyses highlighted a handful of positively selected sites most of which were located in the ORF5 ectodomains of outbreak strains, implicating the host immune system as the possible selective agent. This was in contrast to the closely-related Ontario strains which were subject to strong purifying selection. A broader survey of transmission dynamics in North America unveiled a higher virus flow from Canada to the USA with the primary targets being the Lake States and Corn Belt. In turn, these regions served to disseminate viruses to other swine production regions in the USA. Virus flow from the USA to Canada occurred on a much smaller scale. Collectively, extensive genetic diversity prevails in type 2 PRRSV in one region of the North American swine industry and it is not described adequately by RFLP typing which might have some value in differentiating strains at the local farm level, instead. For diagnostic and research purposes, phylogenetic typing should be the preferred method. Finally, stronger surveillance needs to be adopted to minimize cross-border virus transmission. / published_or_final_version / Biological Sciences / Master / Master of Philosophy