Genetic and Molecular Basis of Encapsulation and Capsule Diversity in Kingella kingae

Kingella kingae is a bacterial pathogen that is increasingly recognized as an etiology of septic arthritis, osteomyelitis, bacteremia, and endocarditis in young children. The pathogenesis of K. kingae disease starts with bacterial adherence to the respiratory epithelium of the posterior pharynx. Previous work has identified type IV pili and a trimeric autotransporter protein called Knh (Kingella NhhA homolog) as critical factors for adherence to human epithelial cells. Additional studies established that the presence of a polysaccharide capsule interferes with Knh-mediated adherence. Given the inhibitory role of capsule during adherence we sought to uncover the genes involved in capsule expression to understand how capsule is elaborated on the cell surface. Additionally, this work aimed to further characterize capsule diversity among K. kingae clinical isolates and to investigate the relationship between capsule type and site of isolation.

We first set out to identify the carbohydrates present in the K. kingae capsule present in the prototype strain 269-492. Glycosyl composition and NMR analysis of surface extractable polysaccharides demonstrated two distinct polysaccharides, one consisting of GalNAc and Kdo with the structure →3)-β-GalpNAc-(1→5)-β-Kdop-(2→ and the other containing galactose alone with the structure →5)-β-Galf-(1→.

To discern the two polysaccharides we disrupted the ctrA gene required for surface localization of the K. kingae polysaccharide capsule and observed a loss of GalNAc and Kdo but no effect on the presence of Gal in bacterial surface extracts. In contrast, deletion of the pamABCDE locus involved in production of a reported galactan exopolysaccharide eliminated Gal but had no effect on the presence of GalNAc and Kdo in surface extracts. These results established that K. kingae strain KK01 produces a polysaccharide capsule with the structure →3)-β-GalpNAc-(1→5)-β-Kdop-(2→ and a separate exopolysaccharide with the structure →5)-β-Galf-(1→.

Having established that K. kingae produces a capsule comprised of GalNAc and Kdo, we next set out to identify the genetic determinants of capsule through a transposon mutagenesis screen. In addition to the previously identified ctrABCD operon, lipA, lipB, and a putative glycosyltransferase termed csaA (capsule synthesis region A gene A) were found to be essential for the production of surface-localized capsule. The ctr operon, lipA, lipB, and csaA were found to be present at unlinked locations throughout the genome, which is atypical for gram-negative organisms that elaborate a capsule dependent on an ABC-type transporter for surface localization. Through examining capsule localization in the ctrA, lipA, lipB, and csaA mutant strains, we determined that the ctrABCD, lipA/lipB, and csaA gene products respectively function in capsule export, assembly, and synthesis, respectively. The GalNAc transferase and Kdo transferase domains found in CsaA further support its role in catalyzing the synthesis of the GalNAc-Kdo capsule in the K. kingae prototype strain.

To investigate the capsule diversity that exists in K. kingae we screened a panel of strains isolated from patients with invasive disease or healthy carriers for the csaA capsule synthesis locus. We discovered that Kingella kingae expresses one of 4 capsule synthesis loci (csa, csb, csc, or csd) associated with a capsule consisting of Kdo and GalNAc (type a), Kdo and GlcNAc (type b), Kdo and ribose (type c), and GlcNAc and galactose (type d), respectively. Cloning of the csa, csb, csc, or csd locus into the empty flanking gene region in a non-encapsulated mutant (creation of an isogenic capsule swap) was sufficient to produce either the type a, type b, or type c capsule, respectively, further supporting the role of these loci in expression of a specific polysaccharide linkage. Capsule type a and capsule type b accounted for 96% of invasive strains. Conversely, capsule type c and capsule type d were found disproportionately among carrier isolates, suggesting that capsule type is important in promoting invasion and dissemination.

In conclusion, we discovered that Kingella kingae expresses a polysaccharide capsule and an exopolysaccharide on its surface that require distinct genetic loci for surface localization. Further investigation into genetic determinants of encapsulation revealed the loci ctrABCD, lipA/lipB, and a putative glycosyltransferase are required for capsule expression, with the gene products having roles in capsule export, assembly, and synthesis, respectively. The putative glycosyltransferase CsaA was determined to be a bifunctional enzyme with both GalNAc-transferase and Kdo-transferase activity. Furthermore, we discovered a total of 4 capsule types expressed in clinical isolates of K. kingae, each with a distinct capsule synthesis locus. The variation in the proportion of capsule types found between invasive strains and carriage strains suggest that capsule type is important in promoting invasion and dissemination. Taken together, this work expands our knowledge of the capsule types expressed among K. kingae carrier and invasive isolates and provides insights into the common genetic determinants of capsule expression. These contributions may lead to selecting clinically relevant capsule types to develop into a capsule based vaccine to prevent K. kingae colonization.

Socialization of Early Prosocial Behavior: Parents' Talk about Emotions is Associated with Sharing and Helping in Toddlers.

What role does socialization play in the origins of prosocial behavior? We examined one potential socialization mechanism, parents' discourse about others' emotions with very young children in whom prosocial behavior is still nascent. Two studies are reported, one of sharing in 18- and 24-month-olds (n = 29), and one of instrumental and empathy-based helping in 18- and 30-month-olds (n = 62). In both studies, parents read age-appropriate picture books to their children and the content and structure of their emotion-related and internal state discourse were coded. Results showed that children who helped and shared more quickly and more often, especially in tasks that required more complex emotion understanding, had parents who more often asked them to label and explain the emotions depicted in the books. Moreover, it was parents' elicitation of children's talk about emotions rather than parents' own production of emotion labels and explanations that explained children's prosocial behavior, even after controlling for age. Thus, it is the quality, not the quantity, of parents' talk about emotions with their toddlers that matters for early prosocial behavior.

Iron status in early life and associations with developmental outcomes

Infants and young children are at particular risk of iron deficiency and its associated consequences for growth and development. The main objectives of this thesis were to quantify iron intakes, status and determinants of status in two year olds; explore determinants of neonatal iron stores; investigate associations between iron status at birth and two years with neurodevelopmental outcomes at two years and explore the influence of growth on iron status in early childhood, using data from the Cork BASELINE (Babies after SCOPE: Evaluating Longitudinal Impact using Neurological and Nutritional Endpoints) Birth Cohort Study (n=2137). Participants were followed prospectively with interviewer-led questionnaires and clinical assessments at day 2 and at 2, 6, 12 and 24 months. At two years, there was a low prevalence of iron deficiency and iron deficiency anaemia in this cohort, representing the largest study of iron status in toddlers in Europe, to date. The increased consumption of iron-fortified products and compliance with recommendations to limit unmodified cows’ milk intakes in toddlers has contributed to the observed improvements in status. Low serum ferritin concentrations at birth, which reflect neonatal iron stores, were shown to track through to two years of age; delivery by Caesarean section, being born small-for-gestational age and maternal obesity and smoking in pregnancy were all associated with significantly lower neonatal iron stores. Despite a low prevalence of iron deficiency in this cohort, both a mean corpuscular volume <74fl and ferritin concentrations <20μg/l were associated with lower neurodevelopmental outcomes at two years. An inverse association between growth in the second year of life and iron status at two years was also observed. This thesis has presented data from one of the largest, extensively-characterised cohorts of young children, to date, to explore iron and its associations with growth and development.

Hearing the Child's Voice: Their Lived Experience in the Pediatric Intensive Care Unit

Background: More than 200,000 children are admitted annually to Pediatric Intensive Care Units (PICUs) in the US. Research has shown young children can provide insight into their hospitalization experiences; child reports rather than parental reports are critical to understanding the child’s experience. Information relating to children’s perceptions while still in the PICU is scarce. Aims: The purpose of this qualitative study was to investigate school age children’s and adolescents’ perceptions of PICU while in the PICU; changes in perceptions after transfer to the General Care Unit (GCU); differences in perceptions of school age children/adolescents and those with more invasive procedures. Methods: Interviews were conducted in PICU within 24-48 hours of admission and 24-48 hours after transfer to GCU. Data on demographics, clinical care and number/types of procedures were obtained. Results: Participants were 7 school age children, 13 adolescents; 10 Hispanic; 13 males. Five overarching themes: Coping Strategies, Environmental Factors, Stressors, Procedures/Medications, and Information. Children emphasized the importance of peer support and visitation; adolescents relied strongly on social media and texting. Parent visits sometimes were more stressful than peer visits. Video games, TV, visitors, and eating were diversional activities. In the PICU, they wanted windows to see outside and interesting things to see on the ceiling above them. Children expressed anticipatory fear of shots and procedures, frustration with lab work, and overwhelming PICU equipment. Number of child responses was higher in PICU (927) than GCU (593); the largest difference was in Environmental Factors. Variations between school age children and adolescents were primarily in Coping Strategies, especially in social support. Number of GCU procedures were the same (8 children) or greater (2 children) than PICU procedures. Discussion: Admission to PICU is a very stressful event. Perceptions from children while still in PICU found information not previously found in the literature. Longitudinal studies to identify children’s perceptions regarding PICU hospitalization and post-discharge outcomes are needed.

Los más pequeños toman la palabra: La Escuela Infantil que a ellos les gustaría

El derecho a ser escuchado y participar en aquellos asuntos que afectan a los niños es un derecho fundamental de la infancia escasamente respetado por el adulto, quien tiende a silenciar su voz, infravalorando su visión y posible aportación. Ante esta realidad, la Escuela Infantil se erige como un marco privilegiado para la escucha y la participación de la infancia en aquellos temas que les afectan, contribuyendo a la formación de ciudadanos democráticos. A partir de un marco de escucha donde se concibe al pequeño como un sujeto activo, competente y experto en su propia vida, se ha llevado a cabo una investigación que ha adoptado el formato de estudio de caso, en la que se ha recogido a través de las técnicas del dibujo y de las conversaciones con niños, la visión que poseen los más pequeños sobre la Escuela Infantil, analizando el bienestar del niño en el contexto escolar, el sentido y finalidad que los niños otorgan a la escuela; el grado de satisfacción con la misma; sus gustos y preferencias en el contexto escolar; la autonomía y libertad que dicen poseer en la escuela; su visión sobre los aspectos organizativos que rigen el funcionamiento del centro y su perspectiva sobre cómo es y cómo les gustaría que fuera la participación de las familias en la escuela. Finalmente, se recogen sus propuestas de mejora para hacer de la Escuela Infantil a la que asisten su escuela ideal. Los resultados del estudio ponen de manifiesto la demanda de los niños de una Escuela Infantil diferente a la que conocen y la existencia de un pensamiento divergente del pensamiento adulto que es necesario escuchar y tener en consideración, siendo sus aportaciones de gran valía para poder iniciar procesos de innovación y mejora educativa.

Individual differences in speech and language ability profiles in areas of high deprivation

Speech and language ability is not a unitary concept; rather, it is made up of multiple abilities such as grammar, articulation and vocabulary. Young children from socio-economically deprived areas are more likely to experience language difficulties than those living in more affluent areas. However, less is known about individual differences in language difficulties amongst young children from socio-economically deprived backgrounds. The present research examined 172 four-year-old children from socio-economically deprived areas on standardised measures of core language, receptive vocabulary, articulation, information conveyed and grammar. Of the total sample, 26% had difficulty in at least one area of language. While most children with speech and language difficulty had generally low performance in all areas, around one in 10 displayed more uneven language abilities. For example, some children had generally good speech and language ability, but had specific difficulty with grammar. In such cases their difficulty is masked somewhat by good overall performance on language tests but they could still benefit from intervention in a specific area. The analysis also identified a number of typically achieving children who were identified as having borderline speech and language difficulty and should be closely monitored

Maternal history of early adversity and child emotional development : investigating intervening factors

L’objectif de cette thèse était de contribuer à l’avancement des connaissances quant aux circonstances permettant une transmission intergénérationnelle du risque émanant de l’adversité maternelle et aux mécanismes sous-tendant cette transmission, dans quatre articles empiriques. Le premier visait à explorer la relation entre un historique d’adversité maternelle, la sécurité d’attachement mère-enfant et le tempérament de l’enfant. Les mères ont complété une entrevue semi-structurée portant sur leurs représentations d’attachement avec leurs parents, à 6 mois, et ont évalué le tempérament de leur enfant à 2 ans. La sécurité d’attachement fut également évaluée à 2 ans. Les résultats ont démontré que les enfants dont les mères rapportaient des niveaux supérieurs d’adversité présentaient de moins bons niveaux d’activité comportementale, uniquement lorsqu’ils avaient un attachement sécurisant avec leur mère. Ces résultats suggèrent une transmission intergénérationnelle des effets d’un historique d’adversité maternelle sur le tempérament des enfants. Le deuxième article visait à investiguer si le transporteur de sérotonine (5-HTTLPR) module la transmission de risque intergénérationnelle de l’adversité maternelle sur le tempérament des enfants. L’historique d’adversité maternelle fut évalué en combinant deux mesures auto-rapportées. Les mères ont également évalué le tempérament de leur enfant à 18 et à 36 mois. Le génotype des enfants fut extrait à 36 mois. Les résultats ont révélé un effet d’interaction entre l’adversité maternelle et le génotype de l’enfant sur le tempérament, suggérant une transmission intergénérationnelle des effets de l’adversité maternelle sur le fonctionnement émotionnel des enfants. Le troisième article visait à explorer la relation entre les difficultés d’adaptation psychosociale des mères, la sensibilité maternelle et les symptômes intériorisés de leurs enfants. Les mères ont complété plusieurs questionnaires desquels un score composite de difficultés d’adaptation psychosociale fut extrait. La sensibilité maternelle fut observée à 12 mois. Les symptômes intériorisés des enfants furent évalués par les deux parents à 2 et à 3 ans. Les résultats ont démontré qu’une augmentation des difficultés maternelles d’adaptation psychosociale étaient associée à davantage de symptômes intériorisés chez les enfants, mais seulement chez ceux dont les mères étaient moins sensibles. Ces résultats ont été observés par les mères à 2 ans et par les deux parents à 3 ans. Ces résultats suggèrent que les enfants peuvent être différemment affectés par l’adaptation émotionnelle de leur mère tout en mettant l’emphase sur le rôle protecteur de la sensibilité maternelle. Le quatrième article visait à investiguer les rôles médiateurs de la dépression et de la sensibilité maternelle dans la relation entre un historique d’adversité maternelle et le tempérament de l’enfant. L’historique d’adversité maternelle fut évalué en combinant deux mesures auto-rapportées. Les mères ont également rapporté leurs symptômes dépressifs à 6 mois. La sensibilité maternelle fut évaluée de façon concomitante. Les mères ont évalué le tempérament de leur enfant à 36 mois. Les résultats ont révélé une transmission intergénérationnelle des effets d’un historique d’adversité maternelle à la génération suivante suivant une médiation séquentielle passant d’abord par la dépression maternelle et ensuite par la sensibilité maternelle. Finalement, les résultats des quatre articles ont été intégrés dans la conclusion générale.

Suporte, Stress e Solidão: um estudo sobre o apoio offline e online percecionado pelas mães portuguesas

Uma “rede social” diz respeito a uma estrutura constituída por pessoas ou organizações que partilham interesses, motivações e valores comuns. Um ponto partilhado pelos diversos tipos de rede social é a troca de informações, conhecimentos, interesses e esforços na tentativa de atingir objectivos comuns, muitas vezes potencializando forças e recursos em situações de crise. Recentemente tem-se ouvido falar bastante deste conceito aplicado à internet. A maternidade constitui, por si só, um momento de crise na vida de cada família e de cada mulher, requerendo adaptações e mudanças. O estudo realizado teve o intuito de avaliar o suporte online e offline (denominando-se essas ligações de redes sociais), no caso específico de assuntos ligados à maternidade. Pretendeu-se verificar também se existia uma correlação entre estes tipos de suporte e os níveis de stress e solidão sentidos pelas mães. Por último, aferiu-se ainda a influência que a idade dos filhos teria nesta adesão a grupos de suporte à maternidade, no Facebook. Este estudo teve uma amostra constituída por 170 mulheres (mães), estando as suas idades compreendidas entre os 25 e os 62 anos. Os resultados revelaram que apesar de não haver diferenças estatisticamente significativas entre o apoio online e offline, as mulheres que pertencem a grupos do Facebook tendem a sentir mais suporte por parte das plataformas de suporte disponíveis. Constatou-se ainda que não existem diferenças significativas entre os níveis de solidão e stress, entre mães que pertencem a grupos e mães que não pertencem. Foi interessante verificar que as mulheres com filhos mais novos (faixa etária dos 0-4 anos) têm uma presença mais significativa em grupos de apoio à maternidade, no Facebook. Pensamos que isso se deve às dúvidas e ao stresse gerado por um primeiro filho ou pela introdução de mais um filho no núcleo familiar, nos primeiros anos em que tal ocorre, procurando por isso mais activamente grupos que ajudem como rede de suporte, para partilhas, apoio emocional e esclarecimento de dúvidas. / A “social network” refers to a structure formed by individuals or organisations that share interests, motivations and common values. A feature shared by several types of social network is the exchange of information, knowledge, interests and efforts, in an attempt to achieve common goals, often potentiating strengths and resources in crisis situations. Recently, we have heard of this concept applied to the internet. Motherhood is, by itself, a moment of crisis in the life of every family and every woman, requiring adaptations and changes. This study aimed to assess online and offline supports (being these connections themselves denominated social networks), in the specific case of motherhood-related issues. It was also intended to confirm whether there was a correlation between these types of support and stress and loneliness levels sensed by mothers. Finally, it was evaluated as well the influence that the children’s age would have on this adherence to motherhood support groups on Facebook. This study had a sample of 170 women (mothers), with ages between 25 and 62 years. The results showed that, although there are no statistically significant differences between the online and offline support, women who belong to Facebook groups tend to feel more support from the available support platforms. It was shown as well that there are no significant differences between loneliness and stress levels among mothers who belong to groups and mothers who do not belong to those groups. It was interesting to find that women with young children (aged 0-4 years old) have a more significant presence in motherhood support groups on Facebook. We think that this is due to the doubts and the stress generated by a first child or the introduction of another child in the household, therefore making sense that they seek, in the early years of the child’s life, groups that help as a support network, for shares, emotional support and answering questions.

Non-invasive diagnosis of pediatric tuberculosis

Thesis (Master's)--University of Washington, 2016-08

Evaluation of Tottenham Thinking Space Pilot: Final Report. Research Report 11.

This reports summarises research that began in March 2014 and was completed in October 2015 by an experienced inter-disciplinary research team from the Centre for Social Justice and Change and Psycho-Social Research Group, School of Social Sciences, the University of East London (UEL) and included Dr Yang Li from the Centre for Geo-Information Studies, UEL, for the first phase of the study. Tottenham ‘Thinking Space’ (TTS) was a pilot therapeutic initiative based in local communities and delivered by the Tavistock & Portman NHS Foundation Trust and funded by the London Borough of Haringey Directorate of Public Health. TTS aimed to improve mental health and enable and empower local communities. TTS was situated within a mental health agenda that was integral to Haringey’s Health and Wellbeing Strategy 2012-2015 and aimed to encourage people to help themselves and each other and develop confident communities. On the one hand TTS was well-suited to this agenda, but, on the other, participants were resistant to, and were trying to free themselves from labelling that implied ‘mental health difficulties’. A total of 243 meetings were held and 351 people attended 1,716 times. The majority of participants attended four times or less, and 33 people attended between 5 and 10 times and 39 people attended over 10 times. Attending a small number of times does not necessarily mean that the attendee was not helped. Attendees reflected the ethnic diversity of Tottenham; 29 different ethnic groups attended. The opportunity to meet with people from different cultural backgrounds in a safe space was highly valued by attendees. Similarly, participants valued the wide age range represented and felt that they benefited from listening to inter-generational experiences. The majority of participants were women (72%) and they were instrumental in initiating further Thinking Spaces, topic specific meetings, the summer programme of activities for mothers and young children and training to meet their needs. The community development worker had a key role in implementing the initiative and sustaining its growth throughout the pilot period. We observed that TTS attracted those whose life experiences were marked by personal struggle and trauma. Many participants felt safe enough to disclose mental health difficulties (85% of those who completed a questionnaire). Participants also came seeking a stronger sense of community in their local area. Key features of the meetings are that they are democratic, non-judgemental, respectful, and focussed on encouraging everyone to listen and to try to understand. We found that the therapeutic method was put in place by high quality facilitators and health and personal outcomes for participants were consistent with those predicted by the underpinning psychoanalytical and systemic theories. Outcomes included a reduction in anxieties and improved personal and social functioning; approximately two thirds of those who completed a questionnaire felt better understood, felt more motivated and more hopeful for the future. The overwhelming majority of survey respondents also felt good about contributing to their community, said that they were more able to cooperate with others and accepting of other cultures, and had made new friends. Participants typically had a better understanding of their current situation and how to take positive action; of those who completed a questionnaire, over half felt more confident to seek support for a personal issue and to contact services. Members of TTS supported each other and instilled hope and build community-mindedness that reduced social isolation.

Evaluation of Community-led Complementary Feeding and Learning Sessions

Thesis (Master's)--University of Washington, 2016-08

Parental mediation, YouTube’s networked public, and the baby-iPad encounter: mobilizing digital dexterity.

This study collected a sample of YouTube videos in which parents recorded their young children utilizing mobile touchscreen devices. Focusing on the more frequently viewed and highly-discussed videos, the paper analyzes the ways in which babies’ ‘digital dexterity’ is coded and understood in terms of contested notions of ‘naturalness’, and how the display of these capabilities is produced for a networked public. This reading of the ‘baby-iPad encounter’ helps expand existing scholarly concepts such as parental mediation and technology domestication. Recruiting several theoretical frameworks, the paper seeks to go beyond concerns of mobile devices and immobile children by analyzing children’s digital dexterity not just as a kind of mobility, but also as a set of reciprocal mobilizations that work across domestic, virtual and publically networked spaces.

RELATIONAL MEMORY IN EARLY CHILDHOOD: DOES UNITIZATION HELP?

Young children often experience relational memory failures, which are thought to be due to underdeveloped recollection processes. Manipulations with adults, however, have suggested that relational memory tasks can be accomplished with familiarity, a processes that is fully developed during early childhood. The goal of the present study was to determine if relational memory performance could be improved in early childhood by teaching children a memory strategy (i.e., unitization) shown to increase familiarity in adults. Six- and 8-year old children were taught to use visualization strategies that either unitized or did not unitize pictures and colored borders. Analysis revealed inconclusive results regarding differences in familiarity between the two conditions, suggesting that the unitization memory strategy did not improve the contribution of familiarity as it has been shown to do in adults. Based on these findings, it cannot be concluded that unitization strategies increase the contribution of familiarity in childhood.

Quichua-Spanish language contact in Salcedo, Ecuador: revisiting media lengua syncretic language practices

The purpose of the current thesis is to develop a better understanding of the interaction between Spanish and Quichua in the Salcedo region and provide more information for the processes that might have given rise to Media Lengua, a ‘mixed’ language comprised of a Quichua grammar and Spanish lexicon. Muysken attributes the formation of Media Lengua to relexification, ruling out any influence from other bilingual phenomena. I argue that the only characteristic that distinguishes Media Lengua from other language contact varieties in central Ecuador is the quantity of the overall Spanish borrowings and not the type of processes that might have been employed by Quichua speakers during the genesis of Media Lengua. The results from the Salcedo data that I have collected show how processes such as adlexification, code-mixing, and structural convergence produce Media Lengua-type sentences, evidence that supports an alternative analysis to Muysken’s relexification hypothesis. Overall, this dissertation is developed around four main objectives: (1) to describe the variation of Spanish loanwords within a bilingual community in Salcedo; (2) to analyze some of the prominent and recent structural changes in Quichua and Spanish; (3) to determine whether Spanish loanword use can be explained by the relationship consultants have with particular social categories; and (4) to analyze the consultants’ language ideologies toward syncretic uses of Spanish and Quichua. Overall, 58% of the content words, 39% of the basic vocabulary, and 50% of the subject pronouns in the Salcedo corpus were derived from Spanish. When compared to Muysken’s description of highlander Quichua in the 1970’s, Spanish loanwords have more than doubled in each category. The overall level of Spanish loanwords in Salcedo Quichua has grown to a level between highlander Quichua in the 1970’s and Media Lengua. Similar to Spanish’s lexical influence in Media Lengua, the increase of Spanish borrowings in today’s rural Quichua can be seen in non-basic and basic vocabularies as well as the subject pronoun system. Significantly, most of the growth has occurred through forms of adlexification i.e., doublets, well-established borrowings, and cultural borrowings, suggesting that ‘ordinary’ lexical borrowing is also capable of producing Media Lengua-type sentences. I approach the second objective by investigating two separate phenomena related to structural convergence. The first examines the complex verbal constructions that have developed in Quichua through Spanish loan translations while the second describes the type of Quichua particles that are attached to Spanish lexemes while speaking Spanish. The calquing of the complex verbal constructions from Spanish were employed when speaking standard Quichua. Since this standard form is typically used by language purists, I argue that their use of calques is a strategy of exploiting the full range of expression from Spanish without incorporating any of the Spanish lexemes which would give the appearance of ‘contamination’. The use of Quichua particles in local varieties of Spanish is a defining characteristic of Quichuacized Spanish, spoken most frequently by women and young children in the community. Although the use of Quichua particles was probably not the main catalyst engendering Media Lengua, I argue that its contribution as a source language to other ‘mixed’ varieties, such as Media Lengua, needs to be accounted for in descriptions of BML genesis. Contrary to Muysken’s representation of relatively ‘unmixed’ Spanish and Quichua as the two source languages of Media Lengua, I propose that local varieties of Spanish might have already been ‘mixed’ to a large degree before Media Lengua was created. The third objective attempts to draw a relationship between particular social variables and the use of Spanish loanwords. Whisker Boxplots and ANOVAs were used to determine which social group, if any, have been introducing new Spanish borrowings into the bilingual communities in Salcedo. Specifically, I controlled for age, education, native language, urban migration, and gender. The results indicate that none of the groups in each of the five social variables indicate higher or lower loanword use. The implication of these results are twofold: (a) when lexical borrowing occurs, it is immediately adopted as the community-wide norm and spoken by members from different backgrounds and generations, or (b) this level of Spanish borrowing (58%) is not a recent phenomenon. The fourth and final objective draws on my ethnographic research that addresses the attitudes of syncretic language use. I observed that Quichuacized Spanish and Hispanicized Quichua are highly stigmatized varieties spoken by the country’s most marginalized populations and families, yet within the community, syncretic ways of speaking are in fact the norm. It was shown that there exists a range of different linguistic definitions for ‘Chaupi Lengua’ and other syncretic language practices as well as many contrasting connotations, most of which were negative. One theme that emerged from the interviews was that speaking syncretic varieties of Quichua weakened the consultant’s claim to an indigenous identity. The linguistic and social data presented in this dissertation supports an alternative view to Muysken’s relexification hypothesis, one that has the advantage of operating with well-precedented linguistic processes and which is actually observable in the present-day Salcedo area. The results from the study on lexical borrowing are significant because they demonstrate how a dynamic bilingual speech community has gradually diversified their Quichua lexicon under intense pressure to shift toward Spanish. They also show that Hispanicized Quichua (Quichua with heavy lexical borrowing) clearly arose from adlexification and prolonged lexical borrowing, and is one of at least six identifiable speech styles found in Salcedo. These results challenge particular interpretations of language contact outcomes, such as, ones that depict sources languages as discrete and ‘unmixed.’ The bilingual continuum presented in this thesis shows on the one hand, the range of speech styles that are accessible to different speakers, and on the other hand, the overlapping, syncretic features that are shared among the different registers and language varieties. It was observed that syncretic speech styles in Salcedo are employed by different consultants in varied interactional contexts, and in turn, produce different evaluations by other fellow community members. In the current dissertation, I challenge the claim that relexification and Media Lengua-type sentences develop in isolation and without the influence of other bilingual phenomena. Based on Muysken's Media Lengua example sentences and the speech styles from the Salcedo corpus, I argue that Media Lengua may have arisen as an institutionalized variant of the highly mixed "middle ground" within the range of the Salcedo bilingual continuum discussed above. Such syncretic forms of Spanish and Quichua strongly resemble Media Lengua sentences in Muysken’s research, and therefore demonstrate how its development could have occurred through several different language contact processes and not only through relexification.