Dynamic and coordinated epigenetic regulation of developmental transitions in the cardiac lineage.

Heart development is exquisitely sensitive to the precise temporal regulation of thousands of genes that govern developmental decisions during differentiation. However, we currently lack a detailed understanding of how chromatin and gene expression patterns are coordinated during developmental transitions in the cardiac lineage. Here, we interrogated the transcriptome and several histone modifications across the genome during defined stages of cardiac differentiation. We find distinct chromatin patterns that are coordinated with stage-specific expression of functionally related genes, including many human disease-associated genes. Moreover, we discover a novel preactivation chromatin pattern at the promoters of genes associated with heart development and cardiac function. We further identify stage-specific distal enhancer elements and find enriched DNA binding motifs within these regions that predict sets of transcription factors that orchestrate cardiac differentiation. Together, these findings form a basis for understanding developmentally regulated chromatin transitions during lineage commitment and the molecular etiology of congenital heart disease.

Comparing the Ecological Niches of Chlamydial and Gonococcal Infections in Winnipeg, Canada: 2007-2016.

BACKGROUND: Previous studies have shown substantial differences in geographic clustering of sexually transmitted infections (STI), such as chlamydia (CT) and gonorrhea (NG), conditional on epidemic phase. Chlamydia and NG have recently shown resurgent epidemiology in the northern hemisphere. This study describes the recent epidemiology of CT and NG in Winnipeg, Canada, combining traditional surveillance tools with place-based analyses, and comparing the ecological niches of CT and NG, in the context of their evolving epidemiology. METHODS: Data were collected as part of routine public health surveillance between 2007 and 2016. Secular trends for CT and NG, and CT/NG coinfection were examined. Gini coefficients and population attributable fractions explored the distribution, and concentration of infections over time and space. RESULTS: Rates of CT increased from 394.9/100,000 population to 476.2/100,000 population from 2007 to 2016. Gonorrhea rates increased from 78.0/100,000 population to 143.5/100,000 population during the same period. Each pathogen had its own ecological niche: CT was widespread geographically and socio-demographically, while NG was clustered in Winnipeg's inner-core. CT/NG co-infections had the narrowest space and age distribution. NG was shown to be undergoing a growth phase, with clear signs of geographic dispersion. The expansion of NG resembled the geographic distribution of CT. CONCLUSIONS: We demonstrated that NG was experiencing a growth phase, confirming theoretical predictions of geographic dispersion during a growth phase. During this phase, NG occupied similar geographic spaces as CT. Knowledge of different ecological niches could lead to better targeting of resources for subpopulations vulnerable to STIs.

His bundle pacing insights from electroanatomical mapping: Topography and pacing targets.

OBJECTIVES: To characterize 3D electroanatomical mapping (EAM) of the His bundle (HB) region. BACKGROUND: Visualization of selective (S) and nonselective (NS) HB capture areas by EAM has not been described and may help guide HB pacing (HBP). METHODS: EAM was performed via NavX system in 17 patients (pts) undergoing HBP. HB cloud, S-HB, NS-HB, and right bundle (RB) capture areas were mapped. RESULTS: S-HBP areas were identified in 11, NS-HBP in 14, and RB in 11 pts. Two NS-HBP areas (upper and lower) either separated by S-HBP (8 pts) or almost contiguous (5 pts) were observed. S-HBP area measured: 1.1 ± 0.9 cm2 , NS upper: -1.2 ± 0.9 cm2 , NS lower: -1.2 ± 0.9 cm2 , RB: -1.7 ± 1.3 cm2 , total His cloud: -4.1 ± 2.7 cm2 . Electrocardiogram (ECG) pacemaps were different between upper and lower NS-HBP areas in 13/14 pts (p = .006). ECG differences between NS clouds were present in inferior leads in 9 pts (more negative QRS complex from lower NS area) and in precordial leads in 5 pts. There was no correlation between HBP lead location and capture threshold. R-wave amplitude was higher at more distal locations on HB cloud (p = .02). CONCLUSION: (1) Pacemapping identifies distinct regions that may correspond to HB anatomy. (2) A linear S-HBP area is typically surrounded by two separate NS areas. (3) Pace-map ECGs from upper and lower NS-HBP areas have different morphologies. (4) These EAM features and pace-mapping may be helpful to the implanter.

Observational study of the populations accessing rapid point-of-care HIV testing in Winnipeg, Manitoba, Canada, through a retrospective chart review of site records.

OBJECTIVES: HIV point-of-care testing (POCT) has been available in Manitoba since 2008. This study evaluated the effectiveness of POCT at identifying individuals with previously unknown HIV status, its effects on clinical outcomes and the characteristics of the populations reached. METHODS: A retrospective database review was conducted for individuals who received HIV POCT from 2011 to 2014. Time to linkage to care and viral load suppression were compared between individuals who tested positive for HIV using POCT and controls identified as positive through standard screening. Testing outcomes for labouring women with undocumented HIV status accessing POCT during labour were also assessed. RESULTS: 3204 individuals received POCT (1055 females (32.9%) and 2149 males (67.1%)), being the first recorded HIV test for 2205 (68.8%). Males were more likely to be targeted with POCT as their first recorded HIV test (adjusted OR (AOR) 1.40). Between the two main test sites (Main Street Project (MSP) and Nine Circles Community Health Centre), MSP tested relatively fewer males (AOR 0.79) but a higher proportion of members of all age groups over 30 years old (AOR 1.83, 2.51 and 3.64 for age groups 30-39, 40-49 and >50, respectively). There was no difference in time to linkage to care (p=0.345) or viral load suppression (p=0.405) between the POCT and standard screening cohorts. Of 215 women presenting in labour with unknown HIV status, one was identified as HIV positive. CONCLUSIONS: POCT in Manitoba has been successful at identifying individuals with previously unknown HIV-positive status. Demographic differences between the two main testing sites support that this intervention is reaching unique populations. Given that we observed no significant difference in time to clinical outcomes, it is reasonable to continue using POCT as a targeted intervention. MESH TERMS: HIV infection; rapid HIV testing; vertical infectious disease transmission; community outreach; service delivery; marginalised populations.

A Comparison of Real-Time Polymerase Chain Reaction Assays for the Detection of Antimicrobial Resistance Markers and Sequence Typing From Clinical Nucleic Acid Amplification Test Samples and Matched Neisseria gonorrhoeae Culture.

Real-time polymerase chain reaction (PCR) assays to detect antimicrobial resistance-associated mutations were tested on Neisseria gonorrhoeae-positive clinical samples with matched isolates. Of the nucleic acid amplification tests/cultures, 87.7% (64/73), 98.6% (72/73), and 98.4% (62/63) predicted cephalosporin, ciprofloxacin, and azithromycin susceptibilities, respectively. N. gonorrhoeae multiantigen sequence type was correctly predicted for 98.7% (79/80), and 13 of 58 N. gonorrhoeae-negative specimens showed false-positive results.

Function-based identification of mammalian enhancers using site-specific integration.

The accurate and comprehensive identification of functional regulatory sequences in mammalian genomes remains a major challenge. Here we describe site-specific integration fluorescence-activated cell sorting followed by sequencing (SIF-seq), an unbiased, medium-throughput functional assay for the discovery of distant-acting enhancers. Targeted single-copy genomic integration into pluripotent cells, reporter assays and flow cytometry are coupled with high-throughput DNA sequencing to enable parallel screening of large numbers of DNA sequences. By functionally interrogating >500 kilobases (kb) of mouse and human sequence in mouse embryonic stem cells for enhancer activity we identified enhancers at pluripotency loci including NANOG. In in vitro-differentiated cardiomyocytes and neural progenitor cells, we identified cardiac enhancers and neuronal enhancers, respectively. SIF-seq is a powerful and flexible method for de novo functional identification of mammalian enhancers in a potentially wide variety of cell types.

Antibiotic susceptibility and molecular analysis of invasive Haemophilus influenzae in Canada, 2007 to 2014.

Background: Previously we studied the antibiotic susceptibility of invasive Haemophilus influenzae collected in Canada from 1990 to 2006 and characterized isolates by serotype, MLST and ftsI gene sequencing for significant PBP3 mutations. Objectives: To provide an update based on isolates collected from 2007 to 2014. Methods: A total of 882 case isolates were characterized by serotype using slide agglutination and PCR. MLST was carried out to determine ST. Isolates were tested for ß-lactamase production, presence of significant PBP3 mutations and antibiotic susceptibility by disc diffusion against 14 antibiotics. MIC values of three antibiotics were determined for 316 isolates using microbroth dilution. Results: Non-typeable H. influenzae accounted for 54.6% of the isolates and 45.4% were serotypeable, predominantly type a (23.1%), type b (8.3%) and type f (10.8%). The overall rate of ampicillin resistance due to ß-lactamase production was 16.4% and increased from 13.5% in 2007-10 to 19% in 2011-14. Significant PBP3 mutations were identified in 129 isolates (14.6%) with 23 (2.6%) also producing ß-lactamase. MLST identified related STs (ST-136, ST-14 and ST-367) associated exclusively with genetically ß-lactamase-negative, ampicillin-resistant isolates and confirmed previously reported associations between significant PBP3 mutations and ST. Conclusions: A significant increase in ß-lactamase-producing isolates was observed from 2007 to 2014; the rate of significant PBP3 mutations has increased since previously reported and 52.5% of non-typeable H. influenzae now show resistance markers. Resistance to trimethoprim/sulfamethoxazole was common and no resistance to fluoroquinolones or third-generation cephalosporins was found.

Mycoplasma genitalium Antibiotic Resistance-Mediating Mutations in Canadian Women With or Without Chlamydia Trachomatis Infection.

Testing remnant Aptima specimens from women infected with Chlamydia trachomatis detected 13.4% (53/396) with Mycoplasma genitalium compared with 5.4% (22/406) in matched C. trachomatis-negative women. Overall, 9.4% (provincial ranges of 3-20%) were infected with M. genitalium and resistance mediating mutations were found in 47.3% (26/55) to macrolides and 1.9% (1/53) to fluoroquinolones by sequencing.

Prior history of testing for syphilis, hepatitis B and hepatitis C among a population-based cohort of HIV-positive individuals and their HIV-negative controls.

Understanding patterns of serological testing for hepatitis B & C, and syphilis among HIV-positive individuals, prior to HIV diagnosis, can inform HIV diagnosis, engagement and prevention strategies. This was a population-based, retrospective analysis of prior serological testing among HIV-positive individuals in Manitoba, Canada. HIV cases were age-, sex- and region-matched to HIV-negative controls at a 1:5 ratio. Conditional logistic regression was used to examine previous serological tests and HIV status. Odds ratios (ORs) and their 95% confidence intervals (95% CI) were reported. A total of 193 cases and 965 controls were included. In the 5 years prior to diagnosis, 50% of cases had at least one test, compared to 26% of controls. Compared to those who did not have serological testing in the 5 years prior to HIV infection, those who had one serological test were at twice the odds of being HIV positive (OR: 1.9, 95% CI: 1.2-2.9), while those with 2 or more tests were at even higher odds (OR: 5.5, 95%CI: 3.7-8.4). HIV cases had higher serological testing rates. Interactions between public health and other healthcare providers should be strengthened.

What Goes Around: the process of building a community-based harm reduction research project.

BACKGROUND: Often, research takes place on underserved populations rather than with underserved populations. This approach can further isolate and stigmatize groups that are already made marginalized. What Goes Around is a community-based research project that was led by community members themselves (Peers). CASE PRESENTATION: This research aimed to implement a community-based research methodology grounded in the leadership and growing research capacity of community researchers and to investigate a topic which community members identified as important and meaningful. Chosen by community members, this project explored how safer sex and safer drug use information is shared informally among Peers. Seventeen community members actively engaged as both community researchers and research participants throughout all facets of the project: inception, implementation, analysis, and dissemination of results. Effective collaboration between community researchers, a community organization, and academics facilitated a research process in which community members actively guided the project from beginning to end. CONCLUSIONS: The methods used in What Goes Around demonstrated that it is not only possible, but advantageous, to draw from community members' involvement and direction in all stages of a community-based research project. This is particularly important when working with a historically underserved population. Purposeful and regular communication among collaborators, ongoing capacity building, and a commitment to respect the experience and expertise of community members were essential to the project's success. This project demonstrated that community members are highly invested in both informally sharing information about safer sex and safer drug use and taking leadership roles in directing research that prioritizes harm reduction in their communities.

Whole-Genome Sequencing Reveals the Origin and Rapid Evolution of an Emerging Outbreak Strain of Streptococcus pneumoniae 12F.

BACKGROUND: Streptococcus pneumoniaeis a major cause of community-acquired pneumonia and septicemia in adults. The global drug-susceptible capsular serotype 12F, clonal complex 218 caused several outbreaks in the United States between 1989 and 2008, as well as a recent large outbreak in Manitoba, Canada, that resulted in 36 cases of septicemia and 3 deaths. The evolutionary origin of the Canadian outbreak strain and its relationship to the historical US outbreak strains are not known. METHODS: Whole-genome deep sequencing was performed on isolates from the Canadian outbreak (n = 36), the US outbreaks (n = 9), and nonoutbreak surveys (n = 21). Phylogenomic analysis and comparative genomics were used to assess evolutionary relationships and to detect gene content differences between the isolates. RESULTS: The Canadian outbreak was closely related to sporadic cases that occurred preoutbreak in cross-border geographic regions in Manitoba, North Dakota, and Iowa. The emerging Canadian strain differed from US strains by acquisition of a cell-surface protein and macrolide resistance determinants via incorporation of a 5.3-kb mega cassette harboringmsrDandmefE Furthermore, during 11 months of transmission, this clone evolved rapidly and acquired fluoroquinolone resistance through precise stepwise mutations in bothparCandgyrA, and putative compensatory mutations inuraAorIMPDHunder drug selection. Alarmingly, this drug-resistant clone appears to have spread quickly to other regions of Canada and the United States, and replaced drug-susceptible strains. CONCLUSIONS: Whole-genome sequencing revealed an independent emergence and secondary adaptation of a new virulent and drug-resistant pneumococcal epidemic clone. Ongoing molecular surveillance is required, and measures to prevent its spread should be developed.

Genomic Epidemiology and Molecular Resistance Mechanisms of Azithromycin-Resistant Neisseria gonorrhoeae in Canada from 1997 to 2014.

The emergence of Neisseria gonorrhoeae strains with decreased susceptibility to cephalosporins and azithromycin (AZM) resistance (AZM(r)) represents a public health threat of untreatable gonorrhea infections. Genomic epidemiology through whole-genome sequencing was used to describe the emergence, dissemination, and spread of AZM(r) strains. The genomes of 213 AZM(r) and 23 AZM-susceptible N. gonorrhoeae isolates collected in Canada from 1989 to 2014 were sequenced. Core single nucleotide polymorphism (SNP) phylogenomic analysis resolved 246 isolates into 13 lineages. High-level AZM(r) (MICs ≥ 256 µg/ml) was found in 5 phylogenetically diverse isolates, all of which possessed the A2059G mutation (Escherichia coli numbering) in all four 23S rRNA alleles. One isolate with high-level AZM(r) collected in 2009 concurrently had decreased susceptibility to ceftriaxone (MIC = 0.125 µg/ml). An increase in the number of 23S rRNA alleles with the C2611T mutations (E. coli numbering) conferred low to moderate levels of AZM(r) (MICs = 2 to 4 and 8 to 32 µg/ml, respectively). Low-level AZM(r) was also associated with mtrR promoter mutations, including the -35A deletion and the presence of Neisseria meningitidis-like sequences. Geographic and temporal phylogenetic clustering indicates that emergent AZM(r) strains arise independently and can then rapidly expand clonally in a region through local sexual networks.

Phenotypic H-Antigen Typing by Mass Spectrometry Combined with Genetic Typing of H Antigens, O Antigens, and Toxins by Whole-Genome Sequencing Enhances Identification of Escherichia coli Isolates.

Mass spectrometry-based phenotypic H-antigen typing (MS-H) combined with whole-genome-sequencing-based genetic identification of H antigens, O antigens, and toxins (WGS-HOT) was used to type 60 clinical Escherichia coli isolates, 43 of which were previously identified as nonmotile, H type undetermined, or O rough by serotyping or having shown discordant MS-H and serotyping results. Whole-genome sequencing confirmed that MS-H was able to provide more accurate data regarding H antigen expression than serotyping. Further, enhanced and more confident O antigen identification resulted from gene cluster based typing in combination with conventional typing based on the gene pair comprising wzx and wzy and that comprising wzm and wzt The O antigen was identified in 94.6% of the isolates when the two genetic O typing approaches (gene pair and gene cluster) were used in conjunction, in comparison to 78.6% when the gene pair database was used alone. In addition, 98.2% of the isolates showed the existence of genes for various toxins and/or virulence factors, among which verotoxins (Shiga toxin 1 and/or Shiga toxin 2) were 100% concordant with conventional PCR based testing results. With more applications of mass spectrometry and whole-genome sequencing in clinical microbiology laboratories, this combined phenotypic and genetic typing platform (MS-H plus WGS-HOT) should be ideal for pathogenic E. coli typing.

Interpretation of laboratory detection trends for Chlamydia trachomatis and Neisseria gonorrhoeae: Manitoba, Canada, 2000-2012.

OBJECTIVES: Increases in case numbers for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) have been noted on a global level. This study analysed 13 years of testing data to better understand case detection trends over time. METHODS: Data consisted of all nucleic acid probe and nucleic acid amplification diagnostic testing for CT and NG for the population of Manitoba, Canada (1.2 million); January 2000 to December 2012. Logistic regression models were used to analyse ORs associated with positive CT and NG tests by year. Included in the model as predictor variables were test type, specimen type, patient age and residence location. RESULTS: For both male and female CT results, unadjusted OR by year mimicked absolute case counts, reflecting a general increase over time in case counts. Adjustment for laboratory-related variables altered this relationship such that a general decline in the odds of identifying a CT case over time was evident. For both male and female NG results, adjustment for laboratory and demographic variables altered the OR associated with each year, but to a lesser extent than for CT. CONCLUSIONS: Temporal trends associated with CT case numbers should be interpreted after controlling, at a minimum, for the influence of laboratory-related variables. Interpretation of NG trends is feasible using only the number of reported NG cases.

Cooperative and antagonistic roles for Irx3 and Irx5 in cardiac morphogenesis and postnatal physiology.

The Iroquois homeobox (Irx) homeodomain transcription factors are important for several aspects of embryonic development. In the developing heart, individual Irx genes are important for certain postnatal cardiac functions, including cardiac repolarization (Irx5) and rapid ventricular conduction (Irx3). Irx genes are expressed in dynamic and partially overlapping patterns in the developing heart. Here we show in mice that Irx3 and Irx5 have redundant function in the endocardium to regulate atrioventricular canal morphogenesis and outflow tract formation. Our data suggest that direct transcriptional repression of Bmp10 by Irx3 and Irx5 in the endocardium is required for ventricular septation. A postnatal deletion of Irx3 and Irx5 in the myocardium leads to prolongation of atrioventricular conduction, due in part to activation of expression of the Na(+) channel protein Nav1.5. Surprisingly, combined postnatal loss of Irx3 and Irx5 results in a restoration of the repolarization gradient that is altered in Irx5 mutant hearts, suggesting that postnatal Irx3 activity can be repressed by Irx5. Our results have uncovered complex genetic interactions between Irx3 and Irx5 in embryonic cardiac development and postnatal physiology.

Whole-genome phylogenomic heterogeneity of Neisseria gonorrhoeae isolates with decreased cephalosporin susceptibility collected in Canada between 1989 and 2013.

A large-scale, whole-genome comparison of Canadian Neisseria gonorrhoeae isolates with high-level cephalosporin MICs was used to demonstrate a genomic epidemiology approach to investigate strain relatedness and dynamics. Although current typing methods have been very successful in tracing short-chain transmission of gonorrheal disease, investigating the temporal evolutionary relationships and geographical dissemination of highly clonal lineages requires enhanced resolution only available through whole-genome sequencing (WGS). Phylogenomic cluster analysis grouped 169 Canadian strains into 12 distinct clades. While some N. gonorrhoeae multiantigen sequence types (NG-MAST) agreed with specific phylogenomic clades or subclades, other sequence types (ST) and closely related groups of ST were widely distributed among clades. Decreased susceptibility to extended-spectrum cephalosporins (ESC-DS) emerged among a group of diverse strains in Canada during the 1990s with a variety of nonmosaic penA alleles, followed in 2000/2001 with the penA mosaic X allele and then in 2007 with ST1407 strains with the penA mosaic XXXIV allele. Five genetically distinct ESC-DS lineages were associated with penA mosaic X, XXXV, and XXXIV alleles and nonmosaic XII and XIII alleles. ESC-DS with coresistance to azithromycin was observed in 5 strains with 23S rRNA C2599T or A2143G mutations. As the costs associated with WGS decline and analysis tools are streamlined, WGS can provide a more thorough understanding of strain dynamics, facilitate epidemiological studies to better resolve social networks, and improve surveillance to optimize treatment for gonorrheal infections.

Rapid, Sensitive, and Specific Escherichia coli H Antigen Typing by Matrix-Assisted Laser Desorption Ionization-Time of Flight-Based Peptide Mass Fingerprinting.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has gained popularity in recent years for rapid bacterial identification, mostly at the genus or species level. In this study, a rapid method to identify the Escherichia coli flagellar antigen (H antigen) at the subspecies level was developed using a MALDI-TOF MS platform with high specificity and sensitivity. Flagella were trapped on a filter membrane, and on-filter trypsin digestion was performed. The tryptic digests of each flagellin then were collected and analyzed by MALDI-TOF MS through peptide mass fingerprinting. Sixty-one reference strains containing all 53 H types and 85 clinical strains were tested and compared to serotyping designations. Whole-genome sequencing was used to resolve conflicting results between the two methods. It was found that DHB (2,5-dihydroxybenzoic acid) worked better than CHCA (α-cyano-4-hydroxycinnamic acid) as the matrix for MALDI-TOF MS, with higher confidence during protein identification. After method optimization, reference strains representing all 53 E. coli H types were identified correctly by MALDI-TOF MS. A custom E. coli flagellar/H antigen database was crucial for clearly identifying the E. coli H antigens. Of 85 clinical isolates tested by MALDI-TOF MS-H, 75 identified MS-H types (88.2%) matched results obtained from traditional serotyping. Among 10 isolates where the results of MALDI-TOF MS-H and serotyping did not agree, 60% of H types characterized by whole-genome sequencing agreed with those identified by MALDI-TOF MS-H, compared to only 20% by serotyping. This MALDI-TOF MS-H platform can be used for rapid and cost-effective E. coli H antigen identification, especially during E. coli outbreaks.

Multi-Province Listeriosis Outbreak Linked to Contaminated Deli Meat Consumed Primarily in Institutional Settings, Canada, 2008.

A multi-province outbreak of listeriosis occurred in Canada from June to November 2008. Fifty-seven persons were infected with 1 of 3 similar outbreak strains defined by pulsed-field gel electrophoresis, and 24 (42%) individuals died. Forty-one (72%) of 57 individuals were residents of long-term care facilities or hospital inpatients during their exposure period. Descriptive epidemiology, product traceback, and detection of the outbreak strains of Listeria monocytogenes in food samples and the plant environment confirmed delicatessen meat manufactured by one establishment and purchased primarily by institutions was the source of the outbreak. The food safety investigation identified a plant environment conducive to the introduction and proliferation of L. monocytogenes and persistently contaminated with Listeria spp. This outbreak demonstrated the need for improved listeriosis surveillance, strict control of L. monocytogenes in establishments producing ready-to-eat foods, and advice to vulnerable populations and institutions serving these populations regarding which high-risk foods to avoid.

Youth Perspectives on Sexual Health Workshops: Informing Future Practice.

Newcomer and street-involved youth provided their perspective on the design and content of a sexual education workshop. Following the workshop, focus group discussions were held with 80 youth from four youth-serving agencies. Youth expressed increased levels of confidence and empowerment, consistent with recent criticism that a focus on specific behaviors as intervention outcome measures may miss important psychosocial changes in participants. Some youth views on cultural adaptation of workshops were not consistent with current views expressed by some adult educators, highlighting the need to ensure a youth perspective is captured during intervention development. Finally, the dichotomous views that youth expressed regarding workshop activities emphasizes a research gap related to how best to adapt interventions to different cognitive capacities, literacy levels, and learning styles. Information of this kind is relevant in terms of knowledge translation from youth to program planners and educators.

Outbreak of invasive Streptococcus pneumoniae serotype 12F among a marginalized inner-city population in Winnipeg, Canada, 2009-2011.

BACKGROUND: In 2010, Winnipeg, Canada, experienced a doubling of invasive pneumococcal disease (IPD) rates, with a significant increase in the number of cases due to Streptococcus pneumoniae serotype 12F, which previously had accounted for very few cases each year. METHODS: All serotype 12F IPD cases reported between September 2009 and January 2011 were reviewed. Pulsed-field gel electrophoresis (PFGE) and multilocus variable number tandem repeat analysis (MLVA) were conducted on all isolates. PFGE and MLVA patterns identified several possible clusters. Additional interviews were conducted to obtain information on risk factors and outcomes. RESULTS: Between September 2009 and January 2011, 169 cases of IPD were identified. The number of IPD cases due to 12F serotype increased sharply from about 3-4 cases per year (6% of IPD cases) in 2007-2009 to 28 (29%) in 2010. All 12F isolates belonged to a single sequence type (ST218), and they were generally susceptible to penicillin and fluoroquinolones but not to erythromycin. Compared with cases caused by other serotypes, patients with serotype 12F were more likely to be homeless, reside in low-income inner-city communities, and engage in substance abuse, including intravenous and crack cocaine use. Subclusters identified using MLVA had even higher rates of homelessness and substance use. CONCLUSIONS: An immunization campaign targeting high-risk groups was undertaken with pneumococcal polysaccharide vaccine, and subsequently rates of serotype 12F decreased. To our knowledge, this is the largest documented community outbreak of serotype 12F IPD and the first report of an outbreak of IPD serotype 12F in a marginalized urban population in Canada.