Integrated Place-Based Primary Interventions: Levers and Tensions Related to Multilevel Governance for Community Integrated Pathways, A Multiple Case Study.

Integrated Place-Based Primary Interventions (IPPIs) are considered an innovative response to the challenges and complex issues faced in disadvantaged areas where traditional institutional services have difficulty reaching people in vulnerable situations. IPPIs are an innovative approach to the delivery of in services, conceived as an original community-based local care and service pathways. However, these intervention practices require adaptive modes of governance. In this article, we explore how and to what extent the mode of governance of IPPIs influences the performance of community-integrated pathways. To this end, using a qualitative exploratory multiple-case study design (observation and semi-structured interviews), we describe 4 IPPIs in 3 territories in Quebec. This includes an examination of the levers of action and tensions related to their governance and the performance levels of the community-integrated pathways. We conclude that collaborative and shared multilevel governance, despite its demanding nature, appears to contribute to the longevity of the actions and benefits of IPPIs and could prevent their relevance from being questioned.

[The analysis of complex interventions in public health: the case of the prevention of sexually transmitted infections and blood-borne infections in Montreal]. / L'analyse des interventions complexes en santé publique : le cas de la prévention des ITSS à Montréal.

OBJECTIVES: Based on a theory of intervention as a complex action system, analyze collaboration among partners in Montréal's sexually transmitted and blood-borne infections (STBBI) prevention program to identify main operations problems and possible scenarios for change to achieve better outcomes. METHODS: A descriptive study was conducted using three data sources - public policies and programs, system management documents, and interviews with three types of partners. The results were validated with stakeholders. RESULTS: Five main operations problems affecting the capacity of the system to provide expected services were identified, as well as strategies the partners use to address these. CONCLUSION: Two scenarios for system change to increase its effectiveness in achieving program goals are discussed.

Novel, versatile, and tightly regulated expression system for Escherichia coli strains.

A novel tightly regulated gene expression system was developed for Escherichia coli by applying the regulatory elements of the Pseudomonas putida F1 cym and cmt operons to control target gene expression at the transcriptional level by using p-isopropylbenzoate (cumate) as an inducer. This novel expression system, referred to as the cumate gene switch, includes a specific expression vector, pNEW, that contains a partial T5 phage promoter combined with the Pseudomonas-based synthetic operator and the cymR repressor protein-encoding gene designed to express constitutively in the host strain. The induction of transcription relies on the addition of the exogenous inducer (cumate), which is nontoxic to the culture, water soluble, and inexpensive. The characteristics and potential of the expression system were determined. Using flow cytometry and fed-batch fermentations, we have shown that, with the newly developed cumate-regulated system, (i) higher recombinant product yields can be obtained than with the pET (isopropyl-beta-D-thiogalactopyranoside [IPTG])-induced expression system, (ii) expression is tightly regulated, (iii) addition of cumate quickly results in a fully induced and homogenous protein-expressing population in contrast to the bimodal expression profile of an IPTG-induced population, (iv) expression can be modulated by varying the cumate concentration, and (v) the cumate-induced population remains induced and fully expressing even at 8 h following induction, resulting in high yields of the target protein Furthermore, the cumate gene switch described in this article is applicable to a wide range of E. coli strains.

Heat-health warning systems: a comparison of the predictive capacity of different approaches to identifying dangerously hot days.

OBJECTIVES: We compared the ability of several heat-health warning systems to predict days of heat-associated mortality using common data sets. METHODS: Heat-health warning systems initiate emergency public health interventions once forecasts have identified weather conditions to breach predetermined trigger levels. We examined 4 commonly used trigger-setting approaches: (1) synoptic classification, (2) epidemiologic assessment of the temperature-mortality relationship, (3) temperature-humidity index, and (4) physiologic classification. We applied each approach in Chicago, Illinois; London, United Kingdom; Madrid, Spain; and Montreal, Canada, to identify days expected to be associated with the highest heat-related mortality. RESULTS: We found little agreement across the approaches in which days were identified as most dangerous. In general, days identified by temperature-mortality assessment were associated with the highest excess mortality. CONCLUSIONS: Triggering of alert days and ultimately the initiation of emergency responses by a heat-health warning system varies significantly across approaches adopted to establish triggers.

Production of an insecticidal crystal protein from Bacillus thuringiensis by the methylotroph Methylobacterium extorquens.

The Cry1Aa protein from Bacillus thuringiensis is an insecticidal protein that is highly active against several species of Lepidoptera. We cloned and expressed the cry1Aa gene in a plant-colonizing methylotroph, Methylobacterium extorquens, under the control of the strong M. extorquens AM1 methanol dehydrogenase promoter, P(mxaF). Transmission electron microscopy revealed characteristic bipyramidal intracellular delta-endotoxin crystals similar to the crystalline inclusions formed by B. thuringiensis. Both the protoxin protein and the activated toxin were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western analysis. In single-dose assays of the recombinant against the silkworm, Bombyx mori, both whole cells and cell lysates caused rapid feeding inhibition followed by mortality. The biomass and growth rate of recombinant cells in shake flask culture were similar to those of the wild-type strain, indicating a lack of fitness cost to the recombinant under controlled culture conditions. Recombinant Cry1Aa was expressed at a level of 4.5% of total M. extorquens cell protein. The potential benefits of modifying M. extorquens to deliver insecticidal Cry proteins for crop and forest protection are discussed.

Bestowing inducibility on the cloned methanol dehydrogenase promoter (PmxaF) of Methylobacterium extorquens by applying regulatory elements of Pseudomonas putida F1.

PmxaF is a strong methanol-inducible promoter in Methylobacterium extorquens. When this promoter is cloned in expression vectors and used to drive heterologous gene expression, methanol inducibility is either greatly reduced or entirely lost. In order to bestow inducibility upon the cloned PmxaF promoter in expression vectors, we adopted combinational methods (regulatory elements of the Pseudomonas putida F1 cym and cmt operons and Tn7 transposon system) to control reporter gene expression at the transcriptional level in M. extorquens. An operator fragment (26 nucleotides) of the cmt operon was inserted downstream of the cloned PmxaF promoter in the broad-host-range expression vector (pCHOI3). The repressor gene (cymR) located upstream of the cym operon in P. putida F1 was amplified by PCR. To avoid cellular toxicity for M. extorquens caused by the overexpression of CymR, single and/or double copies of cymR were integrated into the chromosome of M. extorquens using the mini-Tn7 transposon system. Cultures containing the chromosomally integrated cymR gene were subsequently transformed with pCHOI3 containing modified PmxaF (i.e., PmxaF plus operator). In this construct, inducibility is afforded by cumate (p-isopropylbenzoate). In this report, we describe the inducible and tightly regulated expression of heterologous genes (bgl [for beta-galactosidase], est [for esterase], and gfp [for green fluorescent protein]) in M. extorquens. This is the first documented example of an inducible/regulated heterologous gene expression system in M. extorquens.

Diversity of soluble methane monooxygenase-containing methanotrophs isolated from polluted environments.

Methanotrophs were enriched and isolated from polluted environments in Canada and Germany. Enrichments in low copper media were designed to specifically encourage growth of soluble methane monooxygenase (sMMO) containing organisms. The 10 isolates were characterized physiologically and genetically with one type I and nine type II methanotrophs being identified. Three key genes: 16S rRNA; pmoA and mmoX, encoding for the particulate and soluble methane monooxygenases respectively, were cloned from the isolates and sequenced. Phylogenetic analysis of these sequences identified strains, which were closely related to Methylococcus capsulatus, Methylocystis sp., Methylosinus sporium and Methylosinus trichosporium. Diversity of sMMO-containing methanotrophs detected in this and previous studies was rather narrow, both genetically and physiologically, suggesting possible constraints on genetic diversity of sMMO due to essential conservation of enzyme function.

Multicopy integration and expression of heterologous genes in Methylobacterium extorquens ATCC 55366.

High-level expression of chromosomally integrated genes in Methylobacterium extorquens ATCC 55366 was achieved under the control of the strong M. extorquens AM1 methanol dehydrogenase promoter (PmxaF) using the mini-Tn7 transposon system. Stable maintenance and expression of the integrated genes were obtained in the absence of antibiotic selective pressure. Furthermore, using this technology, a multicopy integration protocol for M. extorquens was also developed. Chromosomal integration of one to five copies of the gene encoding the green fluorescent protein (gfp) was achieved. The multicopy-based expression system permitted expression of a preset number of gene copies. A unique specific Tn7 integration locus in the chromosome of M. extorquens, known as the Tn7 attachment site (attTn7 site), was identified. This single attTn7 site was identified in an intergenic region between glmS, which encodes the essential enzyme glucosamine-6-phosphate synthetase, and dhaT, which encodes 1,3-propanediol dehydrogenase. The fact that the integration event is site specific and the fact that the attTn7 site is a noncoding region of the chromosome make the mini-Tn7 transposon system very useful for insertion of target genes and subsequent expression. In all transformants tested, expression and segregation of the transforming gene were stable without generation of secondary mutations in the host. In this paper, we describe single and multicopy chromosome integration and stable expression of heterologous genes (bgl [beta-galactosidase], est [esterase], and gfp [green fluorescent protein]) in M. extorquens.

Heterologous extracellular production of enterocin P from Enterococcus faecium P13 in the methylotrophic bacterium Methylobacterium extorquens.

Enterocin P (EntP), a strong antilisterial pediocin-like bacteriocin from Enterococcus faecium P13, was produced by Methylobacterium extorquens. For heterologous expression of EntP in the methylotrophic bacterium M. extorquens, a recombinant plasmid was constructed. The gene encoding the EntP structural gene (entP) was cloned into the plasmid vector pCM80, under control of the methanol dehydrogenase promoter (P(mxaF)), to generate plasmid pS25. When M. extorquens ATCC 55366 was transformed with pS25, EntP was detected and quantified in supernatants of the recombinant M. extorquens S25 strain by using specific anti-EntP antibodies and a non-competitive indirect enzyme-linked immunosorbent assay (NCI-ELISA). Purification of EntP by hydrophobic adsorption and reverse-phase (RP-FPLC) chromatographies, permitted recovery of active EntP from the supernatants of M. extorquens S25 grown in a synthetic defined medium.

Frequency of emergency room visits for childhood asthma in Ottawa, Canada: the role of weather.

The aim of this study was to evaluate associations between meteorological conditions and the number of emergency department visits for asthma in a children's hospital in Ottawa, Canada. A case-crossover study design was used. Hospital emergency department visits for asthma between 1992 and 2000 were identified based on patients' presenting complaints. We obtained hourly measures for the following meteorological variables: wind speed, temperature, atmospheric pressure, relative humidity, and visibility. Particular emphasis was placed on exploring the association between asthma visits and fog, thunderstorms, snow, and liquid and freezing forms of precipitation. In total, there were 18,970 asthma visits among children between 2 and 15 years of age. The number of visits and weather characteristics were grouped into 6 h case and control intervals. The occurrence of fog or liquid precipitation was associated with an increased number of asthma visits, while snow was associated with a reduced number (P<0.05). Stratified analyses by season found no association in any of the four calendar intervals between the number of asthma visits and visibility, change in relative humidity and change in temperature. In contrast, summertime thunderstorm activity was associated with an odds ratio of 1.35 (95% CI=1.02-1.77) relative to summer periods with no activity. Models that incorporate calendar and meteorological data may help emergency departments to more efficiently allocate resources needed to treat children presenting with respiratory distress.