Circular RNA regulatory role in pathological cardiac remodelling.

Cardiac remodelling involves structural, cellular and molecular alterations in the heart after injury, resulting in progressive loss of heart function and ultimately leading to heart failure. Circular RNAs (circRNAs) are a recently rediscovered class of non-coding RNAs that play regulatory roles in the pathogenesis of cardiovascular diseases, including heart failure. Thus, a more comprehensive understanding of the role of circRNAs in the processes governing cardiac remodelling may set the ground for the development of circRNA-based diagnostic and therapeutic strategies. In this review, the current knowledge about circRNA origin, conservation, characteristics and function is summarized. Bioinformatics and wet-lab methods used in circRNA research are discussed. The regulatory function of circRNAs in cardiac remodelling mechanisms such as cell death, cardiomyocyte hypertrophy, inflammation, fibrosis and metabolism is highlighted. Finally, key challenges and opportunities in circRNA research are discussed, and orientations for future work to address the pharmacological potential of circRNAs in heart failure are proposed.

Workflow in transgenic mice for the study of specific cancers associated with the post-menopausal state.

Beta-hCG is associated with poor patient prognosis in several cancers, but the particular pathophysiology of beta-hCG in post-menopausal women remains unaddressed. Specific steps to culture Lewis lung carcinoma (LLC1) tumor cells are enumerated. Ovariectomy of syngeneic, beta-hCG transgenic mice is discussed, employing a protocol designed to ensure high survival. Implantation of LLC1 tumor cells in these mice is also described. The workflow can be easily adapted in the study of other cancers associated with the post-menopausal stratum. For complete details on the use and execution of this protocol, please refer to Sarkar et al. (2022).1.

Conjoined and non-conjoined coordination cages with palladium(II) vertices: structural diversity, solution dynamics, and intermolecular interactions.

Self-assembled coordination complexes prepared from a combination of Pd(II) components with one or more types of high-symmetry or low-symmetry bis/tris/tetrakis-monodentate ligands are considered in this review. The structures of these complexes are viewed in terms of the presence of a metallo-macromonocycle or conjoined metallo-macromonocycles/metallocages in the frameworks. Analysis of the typical molecular structures revealed an open truth that one or more units of metallo-macromonocycles can be conjoined to afford planar or non-planar systems. In the same line, the enveloping surface of a 3D cage can be considered as a multiple number of conjoined metallomacrocycles that embrace a 3D space from all directions. However, two or more units of cages are conjoined in a multi-3D-cavity cage system and such a system is considered as a conjoined cage. Construction of such conjoined cages having a finite but multiple number of 3D-cavities unified in a single molecular architecture is a challenging task when compared to that of single-3D-cavity based compounds. Conjoining of as many as four units of 3D cages is known so far. Single- as well as multi-cavity cages of lower symmetry have become a very recent trend in this regard where low-symmetry ligands or mixed ligand ensembles are crafted in the framework of the cages. Other structural diversities like helicity in cages, and supramolecular isomerism are also included in this assorted literature work. Although isomerism in classical coordination complexes is well known, it is very less studied in self-assembled coordination complexes. Ligand isomerism is one such feature that is reviewed here. The dynamic behavior of the cages results in interesting reactivity aspects. A large variety of dynamic processes are collected under an umbrella, i.e., "ligand exchange reactions" and described with examples. Intermolecular interaction among the already self-assembled molecules is possible in solution, solid, and gel-phases as discussed in the last part of this review. The understanding of intermolecular interaction is likely to influence different areas of research including crystal engineering, and materials chemistry.

Configurational ligand isomerism in conjoined-cages.

Double-decker shaped conjoined-cages of Pd3L4 formulation are prepared via self-assembly of Pd(II) with a set of three regioisomeric tridentate ligands. Alongside the targeted double-decker cage, unprecedented hour-glass shaped conjoined-cages of Pd3L4 formulation are also formed in two cases. The double-decker cage prepared from one ligand system and the hour-glass from another (but with a regioisomeric ligand) are structurally well suited to exemplify configurational ligand isomerism.

Progesterone limits the tumor-promoting effects of the beta-subunit of human chorionic gonadotropin via non-nuclear receptors.

The post-menopausal state in women is associated with increased cancer incidence, the reasons for which remain obscure. Curiously, increased circulating levels of beta-hCG (human chorionic gonadotropin) (a hormonal subunit linked with tumors of several lineages) are also often observed post-menopause. This study describes a previously unidentified interplay between beta-hCG and progesterone in tumorigenesis. Progesterone mediated apoptosis in beta-hCG responsive tumor cells via non-nuclear receptors. The transgenic expression of beta-hCG, particularly in the absence of the ovaries (a mimic of the post-menopausal state) constituted a potent pro-tumorigenic signal. Significantly, the administration of progesterone had significant anti-tumor effects. RNA-seq profiling identified molecular signatures associated with these processes. TCGA analysis revealed correlates between the expression of several newly identified genes and poor prognosis in post-menopausal patients of lung adenocarcinoma, colon adenocarcinoma, and glioblastoma. Specifically in these women, the detection of intra-tumoral/extra-tumoral beta-hCG may serve as a useful prognostic indicator, and treatment with progesterone on its detection may prove beneficial.

Comprehensive characterization and molecular insights into the salt tolerance of a Cu, Zn-superoxide dismutase from an Indian Mangrove, Avicennia marina.

Superoxide dismutases are important group of antioxidant metallozyme and play important role in ROS homeostasis in salinity stress. The present study reports the biochemical properties of a salt-tolerant Cu, Zn-superoxide from Avicennia marina (Am_SOD). Am_SOD was purified from the leaf and identified by mass-spectrometry. Recombinant Am_SOD cDNA was bacterially expressed as a homodimeric protein. Enzyme kinetics revealed a high substrate affinity and specific activity of Am_SOD as compared to many earlier reported SODs. An electronic transition in 360-400 nm spectra of Am_SOD is indicative of Cu2+-binding. Am_SOD activity was potentially inhibited by diethyldithiocarbamate and H2O2, a characteristic of Cu, Zn-SOD. Am_SOD exhibited conformational and functional stability at high NaCl concentration as well in alkaline pH. Introgression of Am_SOD in E. coli conferred tolerance to oxidative stress under highly saline condition. Am_SOD was moderately thermostable and retained functional activity at ~ 60 °C. In-silico analyses revealed 5 solvent-accessible N-terminal residues of Am_SOD that were less hydrophobic than those at similar positions of non-halophilic SODs. Substituting these 5 residues with non-halophilic counterparts resulted in > 50% reduction in salt-tolerance of Am_SOD. This indicates a cumulative role of these residues in maintaining low surface hydrophobicity of Am_SOD and consequently high salt tolerance. The molecular information on antioxidant activity and salt-tolerance of Am_SOD may have potential application in biotechnology research. To our knowledge, this is the first report on salt-tolerant SOD from mangrove.

Occurrence and distribution of micro/nanoplastics in soils and their phytotoxic effects: A review.

Some recent studies have reviewed the occurrence and phytotoxicity of micro/nanoplastics, but their distribution in the soil environment, mechanisms of uptake by roots and the mode of action are unclear. Thus, this review comprehensively represents the relative abundance of micro/nanoplastics in different soil types and their toxicities in plants with insights into their partitioning to different soil matrices, uptake mechanisms, and the mode of action. Partitioning of micro/nanoplastics to different soil matrices (like-soil particles, naturally occurring soil organic matters, pore waters and soil fauna) could modify their bioavailability to plants. The small micro/nanoplastic particles can be taken up by roots through the apoplastic and symplastic pathways. In this regard, cellular endocytosis and aquaporin might play a significant role. The shape of the polymers can also regulate their uptake, and the polymers with spherical shapes are more easily absorbed by roots than the polymers with other shapes. Bioaccumulation of micro/nanoplastic induces oxidative stress, which, in turn, causes alterations of gene expressions and different metabolic pathways responsible for plant growth, biomass production and synthesis of secondary metabolites.

CNOT3 interacts with the Aurora B and MAPK/ERK kinases to promote survival of differentiating mesendodermal progenitor cells.

Mesendoderm cells are key intermediate progenitors that form at the early primitive streak (PrS) and give rise to mesoderm and endoderm in the gastrulating embryo. We have identified an interaction between CNOT3 and the cell cycle kinase Aurora B that requires sequences in the NOT box domain of CNOT3 and regulates MAPK/ERK signaling during mesendoderm differentiation. Aurora B phosphorylates CNOT3 at two sites located close to a nuclear localization signal and promotes localization of CNOT3 to the nuclei of mouse embryonic stem cells (ESCs) and metastatic lung cancer cells. ESCs that have both sites mutated give rise to embryoid bodies that are largely devoid of mesoderm and endoderm and are composed mainly of cells with ectodermal characteristics. The mutant ESCs are also compromised in their ability to differentiate into mesendoderm in response to FGF2, BMP4, and Wnt3 due to reduced survival and proliferation of differentiating mesendoderm cells. We also show that the double mutation alters the balance of interaction of CNOT3 with Aurora B and with ERK and reduces phosphorylation of ERK in response to FGF2. Our results identify a potential adaptor function for CNOT3 that regulates the Ras/MEK/ERK pathway during embryogenesis.

Cloning and immunobiochemical analyses on recombinant chymopapain allergen Cari p 2 showing pollen-fruit cross-reaction.

Papaya is reported to trigger food and respiratory allergy. Here, we identified chymopapain Cari p 2 as an allergen that can sensitize atopic individuals through fruit consumption followed by respiratory hazards through pollen exposure. Recombinant Cari p 2 displayed IgE-reactivity with 78% of papaya allergic sera. rCari p 2 also displayed allergenic activity through basophil degranulation. rCari p 2 is correctly folded and showed irreversible denaturation in the melting curve. rCari p 2 displayed IgE-cross-reactivity with homologous cysteine proteases from kiwi and pineapple. Cari p 2 transcript was also detected in papaya pulps. rCari p 2 was resistant to pepsin digestion and retained IgE-reactivity after 60 minutes of pepsin digestion. In mouse model, rCari p 2 was found to elicit inflammatory responses in the lung and gastrointestinal epithelium. Hence, Cari p 2 is a newly characterized allergen with diagnostic and immunotherapeutic potential for managing allergic disorders in papaya sensitized individuals.

Tree parity machine guided patients' privileged based secure sharing of electronic medical record: cybersecurity for telehealth during COVID-19.

In the COVID-19 pandemic, telehealth plays a significant role in the e-healthcare. E-health security risks have also risen significantly with the rise in the use of telehealth. This paper addresses one of e-health's key concerns, namely security. Secret sharing is a cryptographic method to ensure reliable and secure access to information. To eliminate the constraint that in the existing secret sharing schemes, this paper presents Tree Parity Machine (TPM) guided patients' privileged based secure sharing. This is a new secret sharing technique that generates the shares using a simple mask based operation. This work considers addressing the challenges presents in the original secret sharing scheme. This proposed technique enhances the security of the existing scheme. This research introduces a concept of privileged share in which among k number of shares one share should come from a specific recipient (patient) to whom a special privilege is given to recreate the original information. In the absence of this privileged share, the original information cannot be reconstructed. This technique also offers TPM based exchange of secret shares to prevent Man-In-The-Middle-Attack (MITM). Here, two neural networks receive common inputs and exchange their outputs. In some steps, it leads to full synchronization by setting the discrete weights according to the specific rule of learning. This synchronized weight is used as a common secret session key for transmitting the secret shares. The proposed method has been found to produce attractive results that show that the scheme achieves a great degree of protection, reliability, and efficiency and also comparable to the existing secret sharing scheme.

The transgenic expression of the ß-subunit of human chorionic gonadotropin influences the growth of implanted tumor cells.

The beta subunit of human chorionic gonadotropin (ßhCG) is secreted by various tumors, and its presence associated with poor prognosis. Though exogenous hCG elicits the synthesis of molecules associated with angiogenesis, invasion, immune suppression and chemoresistance from responsive tumor cells in vitro, the influence of cell-extrinsic ßhCG on tumorigenesis in vivo has not been adequately explored. Female C57BL/6-/- × FVBßhCG/- F1 transgenic mice demonstrated ovarian hyperplasia and pituitary adenomas; transcripts of hCG-driven, tumor-associated molecules were heightened in the pituitary. Upon the implantation of Lewis Lung Carcinoma cells (murine lung tumor cells derived from C57BL/6 mice) in transgenic mice, tumor incidence and volume were enhanced, and increased transcription and expression of hCG-driven, tumor-associated molecules was observed in excised tumors. While treatment of these mice with Cabergoline (a potent dopamine receptor agonist) had no significant effects, ovariectomy resulted in a reduction in the lag phase, accompanied by an increase in tumor incidence and volume upon Lewis Lung Carcinoma cell implantation. In tumors derived from Lewis Lung Carcinoma cell-implanted ovariectomized, transgenic mice, the transcription and expression of hCG-driven, tumor-associated molecules remained elevated and enhanced animal mortality was observed. Cell-extrinsic ßhCG can therefore induce pro-tumorigenic effects in vivo (even on tumor lineages not part of the reproductive axis), with ovarian products mediating an ameliorating influence.

Cari p 1, a Novel Polygalacturonase Allergen From Papaya Acting as Respiratory and Food Sensitizer.

Papaya has been reported to elicit IgE-mediated hypersensitivity via pollen inhalation and fruit consumption. Certain papaya sensitive patients with food allergy were found to experience recurrent respiratory distresses even after quitting the consumption of fruits. This observation prompted us to investigate the allergens commonly present in fruits and pollen grains of papaya. A discovery approach consisting of immunoproteomic detection followed by molecular characterization led to the identification of a novel papaya allergen designated as Cari p 1. This allergen was detected as a 56 kDa IgE-reactive protein from pollen as well as fruit proteome through serological analysis. The protein was identified as an endopolygalacturonase by tandem mass spectrometry. Full length Cari p 1 cDNA was isolated from papaya pollen, cloned in expression vector, and purified as recombinant allergen. The recombinant protein was monomeric and displayed pectinolytic activity. Recombinant Cari p 1 reacted with IgE-antibodies of all the papaya sensitized patient sera. In addition to IgE-reactivity, rCari p 1 displayed allergenic activity by stimulating histamine release from IgE-sensitized granulocytes. CD-spectroscopy of rCari p 1 revealed the presence of predominantly ß-sheet characters. The melting curve of the allergen showed partial refolding from a fully denatured state indicating the possible presence of conformational IgE-epitopes characteristic of inhalant allergens in addition to the linear IgE-epitopes of food allergens. The expression of this allergen in papaya fruits was detected by immunoblot with anti-Cari p 1 rabbit IgG and reconfirmed by PCR. In an in vivo mouse model, rCari p 1 exhibited a comparable level of inflammatory responses in the lung and duodenum tissues explaining the dual role of Cari p 1 allergen in respiratory sensitization via pollen inhalation and sensitization of gut mucosa via fruit consumption. Purified rCari p 1 can be used a marker allergen for component-resolved molecular diagnosis. Further immunological studies on Cari p 1 are warranted to design immunotherapeutic vaccine for the clinical management of papaya allergy.

DEAD box RNA helicases: crucial regulators of gene expression and oncogenesis.

DEAD box protein family of RNA helicases are vital players of RNA metabolism, and constitute the largest family of RNA helicases. Members of this family share nine conserved motifs including an Asp-Glu-Ala-Asp motif, giving this family its characteristic name as DEAD box RNA helicases. These conserved motifs confer RNA binding and RNA unwinding properties. Besides functioning in RNA metabolism, emerging evidences suggests several DEAD box RNA helicases to possess potential roles in regulating gene expression by acting as a transcriptional co-activator. Many of them are deregulated in cancers, and are implicated in possessing oncogenic potential. On the contrary, each of them also possesses tumor suppressive property in a context dependent manner. In this review, we discuss the mechanistic insights of gene regulation by DEAD box RNA helicases, and their significance in cancers.

Exosome-mediated delivery of the intrinsic C-terminus domain of PTEN protects it from proteasomal degradation and ablates tumorigenesis.

PTEN mutation is a frequent feature across a plethora of human cancers, the hot-spot being its C-terminus (PTEN-CT) regulatory domain resulting in a much diminished protein expression. In this study, the presence of C-terminus mutations was confirmed through sequencing of different human tumor samples. The kinase CKII-mediated phosphorylation of PTEN at these sites makes it a loopy structure competing with the E3 ligases for binding to its lipid anchoring C2 domain. Accordingly, it was found that PTEN-CT expressing stable cell lines could inhibit tumorigenesis in syngenic breast tumor models. Therefore, we designed a novel exosome-mediated delivery of the intrinsic PTEN domain, PTEN-CT into different cancer cells and observed reduced proliferation, migration, and colony forming ability. The delivery of exosome containing PTEN-CT to breast tumor mice model was found to result in significant regression in tumor size with the tumor sections showing increased apoptosis. Here, we also report for the first time an active PTEN when its C2 domain is bound by PTEN-CT, probably rendering its anti-tumorigenic activities through the protein phosphatase activity. Therefore, therapeutic interventions that focus on PTEN E3 ligase inhibition through exosome-mediated PTEN-CT delivery can be a probable route in treating cancers with low PTEN expression.

DEAD-box protein p68 is regulated by ß-catenin/transcription factor 4 to maintain a positive feedback loop in control of breast cancer progression.

INTRODUCTION: Nuclear accumulation of ß-catenin is important for cancer development and it is found to overlap with p68 (DDX5) immunoreactivity in most breast cancers, as indicated by both clinical investigations and studies in cell lines. In this study, we aim to investigate the regulation of p68 gene expression through ß-catenin/transcription factor 4 (TCF4) signaling in breast cancer. METHODS: Formalin-fixed paraffin-embedded sections derived from normal human breast and breast cancer samples were used for immunohistochemical analysis. Protein and mRNA expressions were determined by immunoblotting and quantitative RT-PCR respectively. Promoter activity of p68 was checked using luciferase assay. Occupancy of several factors on the p68 promoter was evaluated using chromatin immunoprecipitation. Finally, a syngeneic mouse model of breast cancer was used to assess physiological significance. RESULTS: We demonstrated that ß-catenin can directly induce transcription of p68 promoter or indirectly through regulation of c-Myc in both human and mouse breast cancer cells. Moreover, by chromatin immunoprecipitation assay, we have found that both ß-catenin and TCF4 occupy the endogenous p68 promoter, which is further enhanced by Wnt signaling. Furthermore, we have also established a positive feedback regulation for the expression of TCF4 by p68. To the best of our knowledge, this is the first report on ß-catenin/TCF4-mediated p68 gene regulation, which plays an important role in epithelial to mesenchymal transition, as shown in vitro in breast cancer cell lines and in vivo in an animal breast tumour model. CONCLUSIONS: Our findings indicate that Wnt/ß-catenin signaling plays an important role in breast cancer progression through p68 upregulation.

Screening and referral to identify children at risk for FASD: Search for new methods 2006-2013.

As part of the Canadian Association of Paediatric Health Centres Taskforce on FASD Screening commitment to further pilot, validate and evaluate the multiple components of the Canadian FASD Screening Tool Kit, it was deemed necessary that recent developments and/or improvements in FASD screening were identified and considered. In 2008 a literature review of methods for screening for FASD was published until 2006 and identified five tools which met pre-set criteria. A review of all new papers was published from the period January 2006 until July 1, 2013. Out of 1392 papers, two new screening methods met the inclusion criteria: Clarren et al's new norms for palpebral fissure length by age in Canada; and Breiner et al's extension of the Neurobehavioral Screening Test (NST) to age 4 years. Further work is needed to validate these methods in other settings. 

Risk factors for and epidemiology of community-onset vancomycin-resistant Enterococcus faecalis in southeast Michigan.

BACKGROUND: Given the known link between vancomycin-resistant Enterococcus faecalis (VREF) and vancomycin-resistant Staphylococcus aureus (VRSA), the recent increase in prevalence of VREF in southeast Michigan has raised concerns about the presence of a large "community" reservoir of VREF. Efforts to control its spread face some important challenges. METHODS: Patients with clinical isolates of community-onset (CO) VREF (cases) were compared with matched uninfected controls (study 1) and patients with hospital-onset (HO) VREF (study 2). Here, CO was defined as a hospital stay of ≤2 days before VRE isolation. RESULTS: Independent predictors for the isolation of CO-VREF compared with uninfected controls were nonhome residence; chronic skin ulcers; previous invasive procedures/surgery; exposure to cephalosporin, penicillin, and/or vancomycin; immunosuppressive status; and the presence of indwelling devices. Independent predictors for isolation of CO-VREF compared with HO-VREF included no stay in an intensive care unit in the previous 3 months and recent hospitalization. VREF isolation from wounds and aminoglycoside exposure were inversely associated with isolation of CO-VREF. CONCLUSIONS: Health care-related exposures and antimicrobial exposures are risk factors for the isolation of CO-VREF. Regional infection control practices are imperative in controlling CO-VREF, in addition to the emergence and spread of VRSA.

Epidemiology and risk factors for isolation of Escherichia coli producing CTX-M-type extended-spectrum ß-lactamase in a large U.S. Medical Center.

A case-case-control study was conducted to identify independent risk factors for recovery of Escherichia coli strains producing CTX-M-type extended-spectrum ß-lactamases (CTX-M E. coli) within a large Southeastern Michigan medical center. Unique cases with isolation of ESBL-producing E. coli from February 2010 through July 2011 were analyzed by PCR for blaCTX-M, blaTEM, and blaSHV genes. Patients with CTX-M E. coli were compared to patients with E. coli strains not producing CTX-M-type ESBLs (non-CTX-M E. coli) and uninfected controls. Of 575 patients with ESBL-producing E. coli, 491 (85.4%) isolates contained a CTX-M ESBL gene. A total of 319 (84.6%) patients with CTX-M E. coli (282 [74.8%] CTX-M-15 type) were compared to 58 (15.4%) non-CTX-M E. coli patients and to uninfected controls. Independent risk factors for CTX-M E. coli isolation compared to non-CTX-M E. coli included male gender, impaired consciousness, H2 blocker use, immunosuppression, and exposure to penicillins and/or trimethoprim-sulfamethoxazole. Compared to uninfected controls, independent risk factors for isolation of CTX-M E. coli included presence of a urinary catheter, previous urinary tract infection, exposure to oxyimino-cephalosporins, dependent functional status, non-home residence, and multiple comorbid conditions. Within 48 h of admission, community-acquired CTX-M E. coli (n = 51 [16%]) and non-CTX-M E coli (n = 11 [19%]) strains were isolated from patients with no recent health care contacts. CTX-M E. coli strains were more resistant to multiple antibiotics than non-CTX-M E. coli strains. CTX-M-encoding genes, especially bla(CTX-M-15) type, represented the most common ESBL determinants from ESBL-producing E. coli, the majority of which were present upon admission. Septic patients with risk factors for isolation of CTX-M E. coli should be empirically treated with appropriate agents. Regional infection control efforts and judicious antibiotic use are needed to control the spread of these organisms.

2,2'-diphenyl-3,3'-diindolylmethane: a potent compound induces apoptosis in breast cancer cells by inhibiting EGFR pathway.

Despite recent advances in medicine, 30-40% of patients with breast cancer show recurrence underscoring the need for improved effective therapy. In this study, by in vitro screening we have selected a novel synthetic indole derivative 2,2'-diphenyl-3,3'-diindolylmethane (DPDIM) as a potential anti- breast cancer agent. DPDIM induces apoptosis both in vitro in breast cancer cells MCF7, MDA-MB 231 and MDA-MB 468 and in vivo in 7,12-dimethylbenz[α]anthracene (DMBA) induced Sprague-Dawley (SD) rat mammary tumor. Our in vitro studies show that DPDIM exerts apoptotic effect by negatively regulating the activity of EGFR and its downstream molecules like STAT3, AKT and ERK1/2 which are involved in the proliferation and survival of these cancer cells. In silico predictions also suggest that DPDIM may bind to EGFR at its ATP binding site. DPDIM furthermore inhibits EGF induced increased cell viability. We have also shown decreased expression of pro-survival factor Bcl-XL as well as increase in the level of pro-apoptotic proteins like Bax, Bad, Bim in DPDIM treated cells in vitro and in vivo. Our results further indicate that the DPDIM induced apoptosis is mediated through mitochondrial apoptotic pathway involving the caspase-cascade. To the best of our knowledge this is the first report of DPDIM for its anticancer activity. Altogether this report suggests that DPDIM could be an effective therapeutic agent for breast cancer.

The maternal drinking history guide: development of a national educational tool.

BACKGROUND: The National Taskforce for the development of screening tools for FASD has identified maternal drinking as a critical area that should be screened. We describe the steps of development and implementation of a knowledge translation program for health care providers. The slide presentation is attached in English and French to allow its maximal use. METHODS: In 2010, the National Taskforce for the development of screening tools for FASD identified maternal drinking as a critical area that should be screened. The systematic review and associated recommendations have been published and were included in the toolkit developed by the Canadian Association of Paediatric Health Centres with funding support from the Public Health Agency of Canada. Effective inquiry of maternal drinking can be conducted at three levels: Primary level, as part of practice-based screening; Level 2 use of structured questionnaires; and Level 3 laboratory-based screening. CONCLUSION: It was acknowledged that most physicians do not ask women of reproductive age questions regarding their drinking habits, and the Taskforce was seriously concerned that even an effective guide may not change practice at the primary level. To that end, the Taskforce developed a three phase Knowledge Translation plan, to ensure that the educational program developed will be optimally effective for Canadian healthcare providers.