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1.  A PAS domain-containing regulator controls flagella-flagella interactions in Campylobacter jejuni 
The bipolar flagella of the foodborne bacterial pathogen Campylobacter jejuni confer motility, which is essential for virulence. The flagella of C. jejuni are post-translationally modified, but how this process is controlled is not well understood. In this work, we have identified a novel PAS-domain containing regulatory system, which modulates flagella-flagella interactions in C. jejuni. Inactivation of the cj1387c gene, encoding a YheO-like PAS6 domain linked to a helix-turn-helix domain, resulted in the generation of a tightly associated “cell-train” morphotype, where up to four cells were connected by their flagella. The morphotype was fully motile, resistant to vortexing, accompanied by increased autoagglutination, and was not observed in aflagellated cells. The Δcj1387c mutant displayed increased expression of the adjacent Cj1388 protein, which comprises of a single endoribonuclease L-PSP domain. Comparative genomics showed that cj1387c (yheO) orthologs in bacterial genomes are commonly linked to an adjacent cj1388 ortholog, with some bacteria, including C. jejuni, containing another cj1388-like gene (cj0327). Inactivation of the cj1388 and cj0327 genes resulted in decreased autoagglutination in Tween-20-supplemented media. The Δcj1388 and Δcj0327 mutants were also attenuated in a Galleria larvae-based infection model. Finally, substituting the sole cysteine in Cj1388 for serine prevented Cj1388 dimerization in non-reducing conditions, and resulted in decreased autoagglutination in the presence of Tween-20. We hypothesize that Cj1388 and Cj0327 modulate post-translational modification of the flagella through yet unidentified mechanisms, and propose naming Cj1387 the Campylobacter Flagella Interaction Regulator CfiR, and the Cj1388 and Cj0327 protein as CfiP and CfiQ, respectively.
PMCID: PMC4519771  PMID: 26284050
Campylobacter; flagella; transcriptional repression; PAS domains
2.  A cross-sectional analysis of perinatal depressive symptoms among Punjabi-speaking women: are they at risk? 
Depression is the leading cause of disability for childbearing women. We examined three specific research questions among Punjabi-speaking women residing in the Fraser Health Authority: 1) What are the prevalence rates of prenatal depressive symptoms? 2) Do Punjabi-speaking women have a higher likelihood of reporting depressive symptoms compared to English-speaking women after controlling for age, level of education and financial worries, and 3) Given the same level of exposure to level of education and financial worries, do Punjabi-speaking women have the same likelihood of reporting depressive symptoms?
Data originated from the Fraser Health Authority prenatal registration database consisting of pregnant women (n = 9684) who completed a prenatal registration form between June 2009 and August 2010; 9.1 % indicated speaking Punjabi. The Whooley Depression Screen measured depressive symptoms. Chi-square tests and logistic multiple regression were used to examine the rates of reporting depressive symptoms among Punjabi-speaking women compared to English-speaking women.
Punjabi-speaking women are at a higher risk for perinatal depressive symptoms. Women needing an interpreter were more likely to report prenatal depressive symptoms compared to English-speaking women. All registrants who reported financial worries had four and a half times the odds of reporting depressive symptoms. The impact of financial worries was significantly greater in the English-speaking women compared to the Punjabi-speaking women needing an interpreter.
Using an established screening device, Punjabi-speaking women were found to be at higher risk for prenatal depressive symptoms.
PMCID: PMC4510901  PMID: 26197818
Pregnancy; South Asian; Ethnicity; Health disparity; Immigrant; Canada primary health care, public health
3.  Campylobacter jejuni biofilms contain extracellular DNA and are sensitive to DNase I treatment 
Biofilms make an important contribution to survival and transmission of bacterial pathogens in the food chain. The human pathogen Campylobacter jejuni is known to form biofilms in vitro in food chain-relevant conditions, but the exact roles and composition of the extracellular matrix are still not clear. Extracellular DNA has been found in many bacterial biofilms and can be a major component of the extracellular matrix. Here we show that extracellular DNA is also an important component of the C. jejuni biofilm when attached to stainless steel surfaces, in aerobic conditions and on conditioned surfaces. Degradation of extracellular DNA by exogenous addition of DNase I led to rapid biofilm removal, without loss of C. jejuni viability. Following treatment of a surface with DNase I, C. jejuni was unable to re-establish a biofilm population within 48 h. Similar results were obtained by digesting extracellular DNA with restriction enzymes, suggesting the need for high molecular weight DNA. Addition of C. jejuni genomic DNA containing an antibiotic resistance marker resulted in transfer of the antibiotic resistance marker to susceptible cells in the biofilm, presumably by natural transformation. Taken together, this suggest that eDNA is not only an important component of C. jejuni biofilms and subsequent food chain survival of C. jejuni, but may also contribute to the spread of antimicrobial resistance in C. jejuni. The degradation of extracellular DNA with enzymes such as DNase I is a rapid method to remove C. jejuni biofilms, and is likely to potentiate the activity of antimicrobial treatments and thus synergistically aid disinfection treatments.
PMCID: PMC4498105  PMID: 26217328
Campylobacter jejuni; biofilm; food safety; extracellular matrix; extracellular DNA; antibiotic resistance
4.  Strain-Dependent Cellular Immune Responses in Cattle following Escherichia coli O157:H7 Colonization 
Infection and Immunity  2014;82(12):5117-5131.
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 causes hemorrhagic diarrhea and potentially fatal renal failure in humans. Ruminants are considered to be the primary reservoir for human infection. Vaccines that reduce shedding in cattle are only partially protective, and their underlying protective mechanisms are unknown. Studies investigating the response of cattle to colonization generally focus on humoral immunity, leaving the role of cellular immunity unclear. To inform future vaccine development, we studied the cellular immune responses of cattle during EHEC O157:H7 colonization. Calves were challenged either with a phage type 21/28 (PT21/28) strain possessing the Shiga toxin 2a (Stx2a) and Stx2c genes or with a PT32 strain possessing the Stx2c gene only. T-helper cell-associated transcripts at the terminal rectum were analyzed by reverse transcription-quantitative PCR (RT-qPCR). Induction of gamma interferon (IFN-γ) and T-bet was observed with peak expression of both genes at 7 days in PT32-challenged calves, while upregulation was delayed, peaking at 21 days, in PT21/28-challenged calves. Cells isolated from gastrointestinal lymph nodes demonstrated antigen-specific proliferation and IFN-γ release in response to type III secreted proteins (T3SPs); however, responsiveness was suppressed in cells isolated from PT32-challenged calves. Lymph node cells showed increased expression of the proliferation marker Ki67 in CD4+ T cells from PT21/28-challenged calves, NK cells from PT32-challenged calves, and CD8+ and γδ T cells from both PT21/28- and PT32-challenged calves following ex vivo restimulation with T3SPs. This study demonstrates that cattle mount cellular immune responses during colonization with EHEC O157:H7, the temporality of which is strain dependent, with further evidence of strain-specific immunomodulation.
PMCID: PMC4249286  PMID: 25267838
5.  Local food environment interventions to improve healthy food choice in adults: a systematic review and realist synthesis protocol 
BMJ Open  2015;5(4):e007161.
Local food environments have been linked with dietary intake and obesity in adults. However, overall evidence remains mixed with calls for increased theoretical and conceptual clarity related to how availability of neighbourhood food outlets, and within-outlet food options, influence food purchasing and consumption. The purpose of this work is to develop a programme theory of food availability, supported by empirical evidence from a range of local food environment interventions.
Methods and analysis
A systematic search of the literature will be followed by duplicate screening and quality assessment (using the Effective Public Health Practice Project tool). Realist synthesis will then be conducted according to the Realist And Meta-narrative Evidence Syntheses: Evolving Standards (RAMESES) publication standards, including transparent appraisal, synthesis and drawing conclusions via consensus.
The final synthesis will propose an evidence-based programme theory of food availability, including evidence mapping to demonstrate contextual factors, pathways of influence and potential mechanisms. With the paucity of empirically supported programme theories used in current local food environment interventions to improve food availability, this synthesis may be used to understand how and why interventions work, and thus inform the development of theory-driven, evidence-based interventions to improve healthy food choice and future empirical work.
Trial registration number
PROSPERO CRD42014009808.
PMCID: PMC4420941  PMID: 25941180
6.  Modifiable barriers to leisure-time physical activity during pregnancy: a qualitative study investigating first time mother’s views and experiences 
Evidence suggests physical activity often declines during pregnancy, however explanations for the decline are not well understood. The aim of this study was to identify modifiable barriers to leisure-time physical activity among women who did not meet physical activity guidelines during pregnancy.
Analyses were based on data from 133 mothers (~3-months postpartum) who were recruited from the Melbourne InFANT Extend study (2012/2013). Women completed a self-report survey at baseline in which they reported their leisure-time physical activity levels during pregnancy as well provided an open-ended written response regarding the key barriers that they perceived prevented them from meeting the physical activity guidelines during their pregnancy. Thematic analyses were conducted to identify key themes.
The qualitative data revealed six themes relating to the barriers of leisure-time physical activity during pregnancy. These included work-related factors (most commonly reported), tiredness, pregnancy-related symptoms, being active but not meeting the guidelines, lack of motivation, and a lack of knowledge of recommendations.
Considering work-related barriers were suggested to be key factors to preventing women from meeting the physical activity guidelines during pregnancy, workplace interventions aimed at providing time management skills along with supporting physical activity programs for pregnant workers should be considered. Such interventions should also incorporate knowledge and education components, providing advice for undertaking leisure-time physical activity during pregnancy.
PMCID: PMC4409747  PMID: 25896111
Pregnancy; Antenatal; Exercise; Correlates; Influences
9.  Prevention of Biofilm Formation and Removal of Existing Biofilms by Extracellular DNases of Campylobacter jejuni 
PLoS ONE  2015;10(3):e0121680.
The fastidious nature of the foodborne bacterial pathogen Campylobacter jejuni contrasts with its ability to survive in the food chain. The formation of biofilms, or the integration into existing biofilms by C. jejuni, is thought to contribute to food chain survival. As extracellular DNA (eDNA) has previously been proposed to play a role in C. jejuni biofilms, we have investigated the role of extracellular DNases (eDNases) produced by C. jejuni in biofilm formation. A search of 2791 C. jejuni genomes highlighted that almost half of C. jejuni genomes contains at least one eDNase gene, but only a minority of isolates contains two or three of these eDNase genes, such as C. jejuni strain RM1221 which contains the cje0256, cje0566 and cje1441 eDNase genes. Strain RM1221 did not form biofilms, whereas the eDNase-negative strains NCTC 11168 and 81116 did. Incubation of pre-formed biofilms of NCTC 11168 with live C. jejuni RM1221 or with spent medium from a RM1221 culture resulted in removal of the biofilm. Inactivation of the cje1441 eDNase gene in strain RM1221 restored biofilm formation, and made the mutant unable to degrade biofilms of strain NCTC 11168. Finally, C. jejuni strain RM1221 was able to degrade genomic DNA from C. jejuni NCTC 11168, 81116 and RM1221, whereas strain NCTC 11168 and the RM1221 cje1441 mutant were unable to do so. This was mirrored by an absence of eDNA in overnight cultures of C. jejuni RM1221. This suggests that the activity of eDNases in C. jejuni affects biofilm formation and is not conducive to a biofilm lifestyle. These eDNases do however have a potential role in controlling biofilm formation by C. jejuni strains in food chain relevant environments.
PMCID: PMC4372405  PMID: 25803828
10.  Functional analysis of bovine TLR5 and association with IgA responses of cattle following systemic immunisation with H7 flagella 
Veterinary Research  2015;46:9.
Flagellin subunits are important inducers of host immune responses through activation of TLR5 when extracellular and the inflammasome if cytosolic. Our previous work demonstrated that systemic immunization of cattle with flagella generates systemic and mucosal IgA responses. The IgA response in mice is TLR5-dependent and TLR5 can impact on the general magnitude of the adaptive response. However, due to sequence differences between bovine and human/murine TLR5 sequences, it is not clear whether bovine TLR5 (bTLR5) is able to stimulate an inflammatory response following interaction with flagellin. To address this we have examined the innate responses of both human and bovine cells containing bTLR5 to H7 flagellin from E. coli O157:H7. Both HEK293 (human origin) and embryonic bovine lung (EBL) cells transfected with bTLR5 responded to addition of H7 flagellin compared to non-transfected controls. Responses were significantly reduced when mutations were introduced into the TLR5-binding regions of H7 flagellin, including an R90T substitution. In bovine primary macrophages, flagellin-stimulated CXCL8 mRNA and secreted protein levels were significantly reduced when TLR5 transcript levels were suppressed by specific siRNAs and stimulation was reduced with the R90T-H7 variant. While these results indicate that the bTLR5 sequence produces a functional flagellin-recognition receptor, cattle immunized with R90T-H7 flagella also demonstrated systemic IgA responses to the flagellin in comparison to adjuvant only controls. This presumably either reflects our findings that R90T-H7 still activates bTLR5, albeit with reduced efficiency compared to WT H7 flagellin, or that other flagellin recognition pathways may play a role in this mucosal response.
PMCID: PMC4333180  PMID: 25827709
11.  Structured Observations Reveal Slow HIV-1 CTL Escape 
PLoS Genetics  2015;11(2):e1004914.
The existence of viral variants that escape from the selection pressures imposed by cytotoxic T-lymphocytes (CTLs) in HIV-1 infection is well documented, but it is unclear when they arise, with reported measures of the time to escape in individuals ranging from days to years. A study of participants enrolled in the SPARTAC (Short Pulse Anti-Retroviral Therapy at HIV Seroconversion) clinical trial allowed direct observation of the evolution of CTL escape variants in 125 adults with primary HIV-1 infection observed for up to three years. Patient HLA-type, longitudinal CD8+ T-cell responses measured by IFN-γ ELISpot and longitudinal HIV-1 gag, pol, and nef sequence data were used to study the timing and prevalence of CTL escape in the participants whilst untreated. Results showed that sequence variation within CTL epitopes at the first time point (within six months of the estimated date of seroconversion) was consistent with most mutations being transmitted in the infecting viral strain rather than with escape arising within the first few weeks of infection. Escape arose throughout the first three years of infection, but slowly and steadily. Approximately one third of patients did not drive any new escape in an HLA-restricted epitope in just under two years. Patients driving several escape mutations during these two years were rare and the median and modal numbers of new escape events in each patient were one and zero respectively. Survival analysis of time to escape found that possession of a protective HLA type significantly reduced time to first escape in a patient (p = 0.01), and epitopes escaped faster in the face of a measurable CD8+ ELISpot response (p = 0.001). However, even in an HLA matched host who mounted a measurable, specific, CD8+ response the average time before the targeted epitope evolved an escape mutation was longer than two years.
Author Summary
The cytotoxic T-lymphocyte (CTL) arm of the immune response is thought to play a significant role in the control of HIV-1 infection. Mutations within the HIV-1 genome allow the virus to escape recognition by CTLs and so evade the immune response. These escape mutations have been well documented but observed waiting times to escape within an individual have ranged from days to years. Many studies describing CTL escape have taken a detailed look at a few patients. Our analysis is based on a cohort of 125 clinical trial participants with immunologic and viral sequence data taken at regular longitudinal time points within the first few years of infection. Results suggested that the majority of CTL-related mutations present early in infection had been transmitted in the infecting viral strain as opposed to arising in the new host due to selection pressure imposed by CTLs. Whilst the prevalence of CTL escape mutations in the dataset was high, the incidence of new escape was relatively low; around one third of patients did not drive an escape within the first two years. Patients possessing a ‘protective’ HLA genotype had a significantly shorter waiting time to first escape than those without.
PMCID: PMC4333731  PMID: 25642847
12.  A Health Impact Assessment Framework for Assessing Vulnerability and Adaptation Planning for Climate Change 
This paper presents a detailed description of an approach designed to investigate the application of the Health Impact Assessment (HIA) framework to assess the potential health impacts of climate change. A HIA framework has been combined with key climate change terminology and concepts. The fundamental premise of this framework is an understanding of the interactions between people, the environment and climate. The diversity and complexity of these interactions can hinder much needed action on the critical health issue of climate change. The objectives of the framework are to improve the methodology for understanding and assessing the risks associated with potential health impacts of climate change, and to provide decision-makers with information that can facilitate the development of effective adaptation plans. While the process presented here provides guidance with respect to this task it is not intended to be prescriptive. As such, aspects of the process can be amended to suit the scope and available resources of each project. A series of working tables has been developed to assist in the collation of evidence throughout the process. The framework has been tested in a number of locations including Western Australia, Solomon Islands, Vanuatu and Nauru.
PMCID: PMC4276652  PMID: 25514146
climate change; adaptation; health impact assessment; vulnerability
13.  Chicken Juice Enhances Surface Attachment and Biofilm Formation of Campylobacter jejuni 
Applied and Environmental Microbiology  2014;80(22):7053-7060.
The bacterial pathogen Campylobacter jejuni is primarily transmitted via the consumption of contaminated foodstuffs, especially poultry meat. In food processing environments, C. jejuni is required to survive a multitude of stresses and requires the use of specific survival mechanisms, such as biofilms. An initial step in biofilm formation is bacterial attachment to a surface. Here, we investigated the effects of a chicken meat exudate (chicken juice) on C. jejuni surface attachment and biofilm formation. Supplementation of brucella broth with ≥5% chicken juice resulted in increased biofilm formation on glass, polystyrene, and stainless steel surfaces with four C. jejuni isolates and one C. coli isolate in both microaerobic and aerobic conditions. When incubated with chicken juice, C. jejuni was both able to grow and form biofilms in static cultures in aerobic conditions. Electron microscopy showed that C. jejuni cells were associated with chicken juice particulates attached to the abiotic surface rather than the surface itself. This suggests that chicken juice contributes to C. jejuni biofilm formation by covering and conditioning the abiotic surface and is a source of nutrients. Chicken juice was able to complement the reduction in biofilm formation of an aflagellated mutant of C. jejuni, indicating that chicken juice may support food chain transmission of isolates with lowered motility. We provide here a useful model for studying the interaction of C. jejuni biofilms in food chain-relevant conditions and also show a possible mechanism for C. jejuni cell attachment and biofilm initiation on abiotic surfaces within the food chain.
PMCID: PMC4249011  PMID: 25192991
14.  Evidence of a high incidence of subclinically affected calves in a herd of cattle with fatal cases of Bovine Neonatal Pancytopenia (BNP) 
BMC Veterinary Research  2014;10:245.
Bovine Neonatal Pancytopenia (BNP) is a disease of calves characterised by bone marrow trilineage hypoplasia, mediated by ingestion of alloantibodies in colostrum. Suspected subclinical forms of BNP have been reported, suggesting that observed clinical cases may not represent the full extent of the disease. However to date there are no objective data available on the incidence of subclinical disease or its temporal distribution. This study aimed to 1) ascertain whether subclinical BNP occurs and, if so, to determine the incidence on an affected farm and 2) determine whether there is evidence of temporal clustering of BNP cases on this farm. To achieve these aims, haematological screening of calves born on the farm during one calving season was carried out, utilising blood samples collected at defined ages. These data were then analysed in comparison to data from both known BNP-free control animals and histopathologically confirmed BNP cases. An ordinal logistic regression model was used to create a composite haematology score to predict the probabilities of calves being normal, based on their haematology measurements at 10–14 days old.
This study revealed that 15% (21 of 139) of the clinically normal calves on this farm had profoundly abnormal haematology (<5% chance of being normal) and could be defined as affected by subclinical BNP. Together with clinical BNP cases, this gave the study farm a BNP incidence of 18%. Calves with BNP were found to be distributed throughout the calving period, with no clustering, and no significant differences in the date of birth of cases or subclinical cases were found compared to the rest of the calves. This study did not find any evidence of increased mortality or increased time from birth to sale in subclinical BNP calves but, as the study only involved a single farm and adverse effects may be determined by other inter-current diseases it remains possible that subclinical BNP has a detrimental impact on the health and productivity of calves under certain circumstances.
Subclinical BNP was found to occur at a high incidence in a herd of cattle with fatal cases of BNP.
PMCID: PMC4216910  PMID: 25358526
Bovine Neonatal Pancytopenia (BNP); Haematology; Subclinical; Alloantibody
15.  HIV-1 DNA predicts disease progression and post-treatment virological control 
eLife  2014;3:e03821.
In HIV-1 infection, a population of latently infected cells facilitates viral persistence despite antiretroviral therapy (ART). With the aim of identifying individuals in whom ART might induce a period of viraemic control on stopping therapy, we hypothesised that quantification of the pool of latently infected cells in primary HIV-1 infection (PHI) would predict clinical progression and viral replication following ART. We measured HIV-1 DNA in a highly characterised randomised population of individuals with PHI. We explored associations between HIV-1 DNA and immunological and virological markers of clinical progression, including viral rebound in those interrupting therapy. In multivariable analyses, HIV-1 DNA was more predictive of disease progression than plasma viral load and, at treatment interruption, predicted time to plasma virus rebound. HIV-1 DNA may help identify individuals who could safely interrupt ART in future HIV-1 eradication trials.
eLife digest
HIV is a virus that can hide in, and hijack, the cells of the immune system and force them to make new copies of the virus. This eventually destroys the infected cells and weakens the ability of a person with HIV to fight off infections and disease. If diagnosed early and treated, most people with HIV now live long and healthy lives and do not develop AIDS—the last stage of HIV infection when previously harmless, opportunistic infections can become life-threatening. However, there are still numerous hurdles and challenges that must be overcome before a cure for HIV/AIDS can be developed.
Treatment with drugs called antiretrovirals can reduce the amount of the HIV virus circulating in an infected person's bloodstream to undetectable levels. However, when HIV infects a cell, the virus inserts a copy of its genetic material into the cell's DNA—and, for most patients, antiretroviral treatment does not tackle these ‘hidden viruses’. As such, and in spite of their side-effects, antiretroviral drugs have to be taken for life in case the hidden viruses re-emerge.
As research into a cure for HIV/AIDS gathers momentum, patients who might be candidates for new experimental treatments will need to be identified. Although it is not recommended as part of standard clinical care, the only way to test if a patient's viral levels would remain suppressed without the drugs would be to temporarily stop the treatment under the close supervision of a physician. As such, a new method is needed to identify if there are patients who might benefit from stopping antiretroviral therapy, and more importantly, those who might not.
Williams, Hurst et al. have now tested whether measuring the levels of HIV DNA directly might help to predict if, and when, the virus might re-emerge (or rebound). In a group of HIV patients participating in a clinical trial, those with higher levels of HIV DNA at the point that the treatment was stopped were found to experience faster viral rebound than those with lower levels of HIV DNA. This method could therefore identify those patients who are at the greatest risk of HIV viral rebound, and are therefore unlikely to benefit if their treatment is interrupted.
Williams, Hurst et al. also found that measuring the levels of HIV DNA could help to predict how the disease would progress in treated and untreated patients. Furthermore, these predictions were more accurate than those based on measuring the amount of the virus circulating in a patient's body.
The next challenge is to identify other methods to distinguish patients who may remain ‘virus-free’ for a period without treatment, from those who would not. With this achieved, it might be possible to identify the mechanisms that determine why the virus comes back and so develop new treatments to stop this happening. This would make developing a cure for HIV/AIDS a much more tangible prospect.
PMCID: PMC4199415  PMID: 25217531
HIV-1; reservoir; antiretroviral therapy; cure; primary infection; human
16.  Health Consequence Scales for Use in Health Impact Assessments of Climate Change 
While health impact assessment (HIA) has typically been applied to projects, plans or policies, it has significant potential with regard to strategic considerations of major health issues facing society such as climate change. Given the complexity of climate change, assessing health impacts presents new challenges that may require different approaches compared to traditional applications of HIA. This research focuses on the development of health consequence scales suited to assessing and comparing health effects associated with climate change and applied within a HIA framework. This assists in setting priorities for adaptation plans to minimize the public health impacts of climate change. The scales presented in this paper were initially developed for a HIA of climate change in Perth in 2050, but they can be applied across spatial and temporal scales. The design is based on a health effects pyramid with health measures expressed in orders of magnitude and linked to baseline population and health data. The health consequence measures are combined with a measure of likelihood to determine the level of risk associated with each health potential health impact. In addition, a simple visual framework that can be used to collate, compare and communicate the level of health risks associated with climate change has been developed.
PMCID: PMC4199038  PMID: 25229697
climate adaptation; health impact assessment; health consequence scales
17.  Family-based interventions to increase physical activity in children: a meta-analysis and realist synthesis protocol 
BMJ Open  2014;4(8):e005439.
Despite the established relationship between physical activity and health, data suggest that many children are insufficiently active, and that levels decline into adolescence. Engaging the family in interventions may increase and maintain children's physical activity levels at the critical juncture before secondary school. Synthesis of existing evidence will inform future studies, but the heterogeneity in target populations recruited, behaviour change techniques and intervention strategies employed, and measurement conducted, may require a multifaceted review method. The primary objective of this work will therefore be to synthesis evidence from intervention studies that explicitly engage the family unit to increase children's physical activity using an innovative dual meta-analysis and realist approach.
Methods and analysis
Peer-reviewed studies will be independently screened by two authors for inclusion based on (1) including ‘healthy’ participants aged 5–12 years; (2) having a substantive intervention aim of increasing physical activity, by engaging the family and (3) reporting on physical activity. Duplicate data extraction and quality assessment will be conducted using a specially designed proforma and the Effective Public Health Practice Project Quality Assessment Tool respectively. STATA software will be used to compute effect sizes for meta-analyses, with subgroup analyses conducted to identify moderating characteristics. Realist syntheses will be conducted according to RAMESES quality and publication guidelines, including development of a programme theory and evidence mapping.
This review will be the first to use the framework of a traditional review to conduct a dual meta-analysis and realist synthesis, examining interventions that engage the family to increase physical activity in children. The results will be disseminated through peer-reviewed publications, conferences, formal presentations to policy makers and practitioners and informal meetings. Evidence generated from this synthesis will also be used to inform the development of theory-driven, evidence-based interventions aimed at engaging the family to increase physical activity levels in children.
Protocol registration
International Prospective Register for Systematic Reviews (PROSPERO): number CRD42013005780.
PMCID: PMC4127934  PMID: 25099934
19.  An expression atlas of human primary cells: inference of gene function from coexpression networks 
BMC Genomics  2013;14:632.
The specialisation of mammalian cells in time and space requires genes associated with specific pathways and functions to be co-ordinately expressed. Here we have combined a large number of publically available microarray datasets derived from human primary cells and analysed large correlation graphs of these data.
Using the network analysis tool BioLayout Express3D we identify robust co-associations of genes expressed in a wide variety of cell lineages. We discuss the biological significance of a number of these associations, in particular the coexpression of key transcription factors with the genes that they are likely to control.
We consider the regulation of genes in human primary cells and specifically in the human mononuclear phagocyte system. Of particular note is the fact that these data do not support the identity of putative markers of antigen-presenting dendritic cells, nor classification of M1 and M2 activation states, a current subject of debate within immunological field. We have provided this data resource on the BioGPS web site ( and on (
PMCID: PMC3849585  PMID: 24053356
Clustering; Meta-analysis; Human; Primary cells; Dendritic cell; Macrophage; Microarray; Transcriptomics
20.  Myocardial infarction incidence and survival by ethnic group: Scottish Health and Ethnicity Linkage retrospective cohort study 
BMJ Open  2013;3(9):e003415.
Inequalities in coronary heart disease mortality by country of birth are large and poorly understood. However, these data misclassify UK-born minority ethnic groups and provide little detail on whether excess risk is due to increased incidence, poorer survival or both.
Retrospective cohort study.
General resident population of Scotland.
All those residing in Scotland during the 2001 Census were eligible for inclusion: 2 972 120 people were included in the analysis. The number still residing in Scotland at the end of the study in 2008 is not known.
Primary and secondary outcome measures
As specified in the analysis plan, the primary outcome measures were first occurrence of admission or death due to myocardial infarction and time to event. There were no secondary outcome measures.
Acute myocardial infarction (AMI) incidence risk ratios (95% CIs) relative to white Scottish populations (100) were highest among Pakistani men (164.1 (142.2 to 189.2)) and women (153.7 (120.5, 196.1)) and lowest for men and women of Chinese (39.5 (27.1 to 57.6) and 59.1 (38.6 to 90.7)), other white British (77 (74.2 to 79.8) and 72.2 (69.0 to 75.5)) and other white (83.1 (75.9 to 91.0) and 79.9 (71.5 to 89.3)) ethnic groups. Adjustment for educational qualification did not eliminate these differences. Cardiac intervention uptake was similar across most ethnic groups. Compared to white Scottish, 28-day survival did not differ by ethnicity, except in Pakistanis where it was better, particularly in women (0.44 (0.25 to 0.78)), a difference not removed by adjustment for education, travel time to hospital or cardiac intervention uptake.
Pakistanis have the highest incidence of AMI in Scotland, a country renowned for internationally high cardiovascular disease rates. In contrast, survival is similar or better in minority ethnic groups. Clinical care and policy should focus on reducing incidence among Pakistanis through more aggressive prevention.
PMCID: PMC3773657  PMID: 24038009
Epidemiology; Public Health
21.  RNA-Seq Differentiates Tumour and Host mRNA Expression Changes Induced by Treatment of Human Tumour Xenografts with the VEGFR Tyrosine Kinase Inhibitor Cediranib 
PLoS ONE  2013;8(6):e66003.
Pre-clinical models of tumour biology often rely on propagating human tumour cells in a mouse. In order to gain insight into the alignment of these models to human disease segments or investigate the effects of different therapeutics, approaches such as PCR or array based expression profiling are often employed despite suffering from biased transcript coverage, and a requirement for specialist experimental protocols to separate tumour and host signals. Here, we describe a computational strategy to profile transcript expression in both the tumour and host compartments of pre-clinical xenograft models from the same RNA sample using RNA-Seq. Key to this strategy is a species-specific mapping approach that removes the need for manipulation of the RNA population, customised sequencing protocols, or prior knowledge of the species component ratio. The method demonstrates comparable performance to species-specific RT-qPCR and a standard microarray platform, and allowed us to quantify gene expression changes in both the tumour and host tissue following treatment with cediranib, a potent vascular endothelial growth factor receptor tyrosine kinase inhibitor, including the reduction of multiple murine transcripts associated with endothelium or vessels, and an increase in genes associated with the inflammatory response in response to cediranib. In the human compartment, we observed a robust induction of hypoxia genes and a reduction in cell cycle associated transcripts. In conclusion, the study establishes that RNA-Seq can be applied to pre-clinical models to gain deeper understanding of model characteristics and compound mechanism of action, and to identify both tumour and host biomarkers.
PMCID: PMC3686868  PMID: 23840389
22.  Examination of mid-intervention mediating effects on objectively assessed sedentary time among children in the Transform-Us! cluster-randomized controlled trial 
The optimal targets and strategies for effectively reducing sedentary behavior among young people are unknown. Intervention research that explores changes in mediated effects as well as in outcome behaviors is needed to help inform more effective interventions. Therefore, the purpose of this study was to examine the mid-intervention mediating effects on children’s objectively assessed classroom and total weekday sedentary time in the Transform-Us! intervention.
The results are based on 293 children, aged 7- to 9-years-old at baseline, from 20 schools in Melbourne, Australia. Each school was randomly allocated to one of four groups, which targeted reducing sedentary time in the school and family settings (SB; n = 74), increasing or maintaining moderate- to vigorous-intensity physical activity in the school and family settings (PA; n = 75), combined SB and PA (SB + PA; n = 80), or the current practice control (C; n = 64). Baseline and mid-intervention data (5–9 months) were collected in 2010 and analyzed in 2012. Classroom and total weekday sedentary time was objectively assessed using ActiGraph accelerometers. The hypothesized mediators including, child enjoyment, parent and teacher outcome expectancies, and child perceived access to standing opportunities in the classroom environment, were assessed by questionnaire.
The SB + PA group spent 13.3 min/day less in weekday sedentary time at mid-intervention compared to the control group. At mid-intervention, children in the SB group had higher enjoyment of standing in class (0.9 units; 5-unit scale) and all intervention groups had more positive perceptions of access to standing opportunities in the classroom environment (0.3-0.4 units; 3-unit scale), compared to the control group. However, none of the hypothesized mediator variables had an effect on sedentary time; thus, no mediating effects were observed.
While beneficial intervention effects were observed on some hypothesized mediating variables and total weekday sedentary time at mid-intervention, no significant mediating effects were found. Given the dearth of existing information, future intervention research is needed that explores mediated effects. More work is also needed on the development of reliable mediator measures that are sensitive to change overtime.
Trial registration
PMCID: PMC3681598  PMID: 23688180
Sedentary behavior; Intervention; Mediators; Children
23.  A systematic review of intervention effects on potential mediators of children’s physical activity 
BMC Public Health  2013;13:165.
Many interventions aiming to increase children’s physical activity have been developed and implemented in a variety of settings, and these interventions have previously been reviewed; however the focus of these reviews tends to be on the intervention effects on physical activity outcomes without consideration of the reasons and pathways leading to intervention success or otherwise.
To systematically review the efficacy of physical activity interventions targeting 5-12 year old children on potential mediators and, where possible, to calculate the size of the intervention effect on the potential mediator.
A systematic search identified intervention studies that reported outcomes on potential mediators of physical activity among 5-12 year old children. Original research articles published between 1985 and April 2012 were reviewed.
Eighteen potential mediators were identified from 31 studies. Positive effects on cognitive/psychological potential mediators were reported in 15 out of 31 studies. Positive effects on social environmental potential mediators were reported in three out of seven studies, and no effects on the physical environment were reported. Although no studies were identified that performed a mediating analysis, 33 positive intervention effects were found on targeted potential mediators (with effect sizes ranging from small to large) and 73% of the time a positive effect on the physical activity outcome was reported.
Many studies have reported null intervention effects on potential mediators of children’s physical activity; however, it is important that intervention studies statistically examine the mediating effects of interventions so the most effective strategies can be implemented in future programs.
PMCID: PMC3585884  PMID: 23433143
Mediator; Child; Physical activity promotion; Theory
24.  Help bring back the celebration of life: A community-based participatory study of rural Aboriginal women’s maternity experiences and outcomes 
Despite clear evidence regarding how social determinants of health and structural inequities shape health, Aboriginal women’s birth outcomes are not adequately understood as arising from the historical, economic and social circumstances of their lives. The purpose of this study was to understand rural Aboriginal women’s experiences of maternity care and factors shaping those experiences.
Aboriginal women from the Nuxalk, Haida and 'Namgis First Nations and academics from the University of British Columbia in nursing, medicine and counselling psychology used ethnographic methods within a participatory action research framework. We interviewed over 100 women, and involved additional community members through interviews and community meetings. Data were analyzed within each community and across communities.
Most participants described distressing experiences during pregnancy and birthing as they grappled with diminishing local maternity care choices, racism and challenging economic circumstances. Rural Aboriginal women’s birthing experiences are shaped by the intersections among rural circumstances, the effects of historical and ongoing colonization, and concurrent efforts toward self-determination and more vibrant cultures and communities.
Women’s experiences and birth outcomes could be significantly improved if health care providers learned about and accounted for Aboriginal people’s varied encounters with historical and ongoing colonization that unequivocally shapes health and health care. Practitioners who better understand Aboriginal women’s birth outcomes in context can better care in every interaction, particularly by enhancing women’s power, choice, and control over their experiences. Efforts to improve maternity care that account for the social and historical production of health inequities are crucial.
PMCID: PMC3577503  PMID: 23360168
Aboriginal; Rural; Maternity care; Outcomes; Colonialism; Critical ethnography
25.  Paired Ductal Carcinoma In Situ and Invasive Breast Cancer Lesions in the D-Loop of the Mitochondrial Genome Indicate a Cancerization Field Effect 
BioMed Research International  2012;2013:379438.
Alterations in the mitochondrial genome have been chronicled in most solid tumors, including breast cancer. The intent of this paper is to compare and document somatic mitochondrial D-loop mutations in paired samples of ductal carcinoma in situ (DCIS) and invasive breast cancer (IBC) indicating a potential breast ductal epithelial cancerization field effect. Paired samples of these histopathologies were laser-captured microdissected (LCM) from biopsy, lumpectomy, and mastectomy tissues. Blood samples were collected as germplasm control references. For each patient, hypervariable region 1 (HV1) in the D-loop portion of the mitochondrial genome (mtGenome) was sequenced for all 3 clinical samples. Specific parallel somatic heteroplasmic alterations between these histopathologies, particularly at sites 16189, 16223, 16224, 16270, and 16291, suggest the presence of an epithelial, mitochondrial cancerization field effect. These results indicate that further characterization of the mutational pathway of DCIS and IBC may help establish the invasive potential of DCIS. Moreover, this paper indicates that biofluids with low cellularity, such as nipple aspirate fluid and/or ductal lavage, warrant further investigation as early and minimally invasive detection mediums of a cancerization field effect within breast tissue.
PMCID: PMC3591154  PMID: 23509716

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