Publications by authors named "Matthew L Workentine"

39 Publications

Associations Between the Gut Microbiota and Internalizing Behaviors in Preschool Children.

Psychosom Med 2021 Oct 12. Epub 2021 Oct 12.

Department of Pediatrics, University of Calgary, Calgary, Alberta, Canada Department of Psychology, University of Calgary, Calgary, Alberta, Canada Alberta Children's Hospital Research Institute (ACHRI), Calgary, Alberta, Canada Department of Community Health Sciences, University of Calgary, Calgary, Alberta, Canada UCVM Bioinformatics, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada Faculty of Kinesiology, University of Calgary, Calgary, Alberta, Canada Department of Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary, Alberta, Canada.

Objective: Emerging evidence points toward a connection between mental health and the gut microbiota and its metabolites (e.g., short-chain fatty acids - SCFAs). It is unknown whether gut microbiota levels are associated with the development of mental health problems (e.g., internalizing or externalizing behaviors) in preschool children. The objective of this study was to evaluate associations between the gut microbiota and internalizing and externalizing behaviors in preschool-aged children.

Methods: A community sample of 248 typically developing children (3-5 years) provided a stool sample for gut microbiota and SCFA analysis. Parents reported child internalizing and externalizing behaviors using the Child Behavior Checklist. Associations between child behaviors and gut microbiota measures were analyzed using Spearman correlations followed by an adjustment for multiple testing, with subanalysis conducted in children clinically "at risk" for behavioral problems compared to those who were not using Mann-Whitney U tests.

Results: There was a correlation between Shannon alpha diversity with internalizing behaviors (rs = -0.134, p = 0.035), and its subscale somatic complaints (rs = -0.144, p = 0.023), while children clinically "at risk" for internalizing problems had decreased alpha diversity (U = 551, p = 0.017). Internalizing behaviors correlated with valerate and isobutyrate (rs = -0.147, p = 0.021; rs = -0.140, p = 0.028, respectively), while the somatic complaints subscale additionally correlated with acetate and butyrate (rs = -0.219, p = 0.001; rs = -0.241, p < 0.001, respectively). These findings were also present in children "at risk" for internalizing problems (U = 569, p = 0.026; U = 571, p = 0.028), and somatic complaints (U = 164, p = 0.004; U = 145, p = 0.001).

Conclusions: These analyses reveal novel associations between internalizing behaviors and the gut microbiota in preschool children. Furthermore, a relationship between somatic complaints and acetate and butyrate was identified, indicating that interventions that increase SCFA production warrant future investigation.
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http://dx.doi.org/10.1097/PSY.0000000000001026DOI Listing
October 2021

Characterization of the β-tubulin gene family in Ascaris lumbricoides and Ascaris suum and its implication for the molecular detection of benzimidazole resistance.

PLoS Negl Trop Dis 2021 09 27;15(9):e0009777. Epub 2021 Sep 27.

Department of Virology, Parasitology and Immunology, Ghent University, Merelbeke, Belgium.

Background: The treatment coverage of control programs providing benzimidazole (BZ) drugs to eliminate the morbidity caused by soil-transmitted helminths (STHs) is unprecedently high. This high drug pressure may result in the development of BZ resistance in STHs and so there is an urgent need for surveillance systems detecting molecular markers associated with BZ resistance. A critical prerequisite to develop such systems is an understanding of the gene family encoding β-tubulin proteins, the principal targets of BZ drugs.

Methodology And Principal Findings: First, the β-tubulin gene families of Ascaris lumbricoides and Ascaris suum were characterized through the analysis of published genomes. Second, RNA-seq and RT-PCR analyses on cDNA were applied to determine the transcription profiles of the different gene family members. The results revealed that Ascaris species have at least seven different β-tubulin genes of which two are highly expressed during the entire lifecycle. Third, deep amplicon sequencing was performed on these two genes in more than 200 adult A. lumbricoides (Ethiopia and Tanzania) and A. suum (Belgium) worms, to investigate the intra- and inter-species genetic diversity and the presence of single nucleotide polymorphisms (SNPs) that are associated with BZ resistance in other helminth species; F167Y (TTC>TAC or TTT>TAT), E198A (GAA>GCA or GAG>GCG), E198L (GAA>TTA) and F200Y (TTC>TAC or TTT>TAT). These particular SNPs were absent in the two investigated genes in all three Ascaris populations.

Significance: This study demonstrated the presence of at least seven β-tubulin genes in Ascaris worms. A new nomenclature was proposed and prioritization of genes for future BZ resistance research was discussed. This is the first comprehensive description of the β-tubulin gene family in Ascaris and provides a framework to investigate the prevalence and potential role of β-tubulin sequence polymorphisms in BZ resistance in a more systematic manner than previously possible.
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http://dx.doi.org/10.1371/journal.pntd.0009777DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8496844PMC
September 2021

Characterization of microglial transcriptomes in the brain and spinal cord of mice in early and late experimental autoimmune encephalomyelitis using a RiboTag strategy.

Sci Rep 2021 07 12;11(1):14319. Epub 2021 Jul 12.

Hotchkiss Brain Institute, Snyder Institute for Chronic Diseases, Department of Physiology and Pharmacology, Cumming School of Medicine, University of Calgary, 3330 Hospital Drive NW, Calgary, AB, T2N 4N1, Canada.

Microglia play an important role in the pathogenesis of multiple sclerosis and the mouse model of MS, experimental autoimmune encephalomyelitis (EAE). To more fully understand the role of microglia in EAE we characterized microglial transcriptomes before the onset of motor symptoms (pre-onset) and during symptomatic EAE. We compared the transcriptome in brain, where behavioral changes are initiated, and spinal cord, where damage is revealed as motor and sensory deficits. We used a RiboTag strategy to characterize ribosome-bound mRNA only in microglia without incurring possible transcriptional changes after cell isolation. Brain and spinal cord samples clustered separately at both stages of EAE, indicating regional heterogeneity. Differences in gene expression were observed in the brain and spinal cord of pre-onset and symptomatic animals with most profound effects in the spinal cord of symptomatic animals. Canonical pathway analysis revealed changes in neuroinflammatory pathways, immune functions and enhanced cell division in both pre-onset and symptomatic brain and spinal cord. We also observed a continuum of many pathways at pre-onset stage that continue into the symptomatic stage of EAE. Our results provide additional evidence of regional and temporal heterogeneity in microglial gene expression patterns that may help in understanding mechanisms underlying various symptomology in MS.
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http://dx.doi.org/10.1038/s41598-021-93590-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8275680PMC
July 2021

A repeatable and quantitative DNA metabarcoding assay to characterize mixed strongyle infections in horses.

Int J Parasitol 2021 02 23;51(2-3):183-192. Epub 2020 Nov 23.

Department of Comparative Biology and Experimental Medicine, Host-Parasite Interactions (HPI) Program, University of Calgary, 3280 Hospital Drive, Calgary, AB T2N 4Z6, Canada.

Horses are ubiquitously infected by a diversity of gastro-intestinal parasitic helminths. Of particular importance are nematodes of the family Strongylidae, which can significantly impact horse health and performance. However, knowledge about equine strongyles remains limited due to our inability to identify most species non-invasively using traditional morphological techniques. We developed a new internal transcribed spacer 2 (ITS2) DNA metabarcoding 'nemabiome' assay to characterise mixed strongyle infections in horses and assessed its performance by applying it to pools of infective larvae from fecal samples from an experimental herd in Kentucky, USA and two feral horse populations from Sable Island and Alberta, Canada. In addition to reporting the detection of 33 different species with high confidence, we illustrate the assay's repeatability by comparing results generated from aliquots from the same fecal samples and from individual horses sampled repeatedly over multiple days or months. We also validate the quantitative potential of the assay by demonstrating that the proportion of amplicon reads assigned to different species scales linearly with the number of larvae present. This new tool significantly improves equine strongyle diagnostics, presenting opportunities for research on species-specific anthelmintic resistance and the causes and consequences of variation in mixed infections.
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http://dx.doi.org/10.1016/j.ijpara.2020.09.003DOI Listing
February 2021

Crohn's Disease Pathobiont Adherent-Invasive E coli Disrupts Epithelial Mitochondrial Networks With Implications for Gut Permeability.

Cell Mol Gastroenterol Hepatol 2021 28;11(2):551-571. Epub 2020 Sep 28.

Gastrointestinal Research Group and Inflammation Research Network, Department of Physiology and Pharmacology, Calvin, Joan and Phoebe Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Alberta, Canada. Electronic address:

Background & Aims: Adherent-invasive Escherichia coli are implicated in inflammatory bowel disease, and mitochondrial dysfunction has been observed in biopsy specimens from patients with inflammatory bowel disease. As a novel aspect of adherent-invasive E coli-epithelial interaction, we hypothesized that E coli (strain LF82) would elicit substantial disruption of epithelial mitochondrial form and function.

Methods: Monolayers of human colon-derived epithelial cell lines were exposed to E coli-LF82 or commensal E coli and RNA sequence analysis, mitochondrial function (adenosine triphosphate synthesis) and dynamics (mitochondrial network imaging, immunoblotting for fission and fusion proteins), and epithelial permeability (transepithelial resistance, flux of fluorescein isothiocyanate-dextran and bacteria) were assessed.

Results: E coli-LF82 significantly affected epithelial expression of ∼8600 genes, many relating to mitochondrial function. E coli-LF82-infected epithelia showed swollen mitochondria, reduced mitochondrial membrane potential and adenosine triphosphate, and fragmentation of the mitochondrial network: events not observed with dead E coli-LF82, medium from bacterial cultures, or control E coli. Treatment with Mitochondrial Division Inhibitor 1 (Mdivi1, inhibits dynamin-related peptide 1, guanosine triphosphatase principally responsible for mitochondrial fission) or P110 (prevents dynamin-related peptide 1 binding to mitochondrial fission 1 protein) partially reduced E coli-LF82-induced mitochondrial fragmentation in the short term. E coli-LF82-infected epithelia showed loss of the long isoform of optic atrophy factor 1, which mediates mitochondrial fusion. Mitochondrial Division Inhibitor 1 reduced the magnitude of E coli-LF82-induced increased transepithelial flux of fluorescein isothiocyanate dextran. By 8 hours after infection, increased cytosolic cytochrome C and DNA fragmentation were apparent without evidence of caspase-3 or apoptosis inducing factor activation.

Conclusions: Epithelial mitochondrial fragmentation caused by E coli-LF82 could be targeted to maintain cellular homeostasis and mitigate infection-induced loss of epithelial barrier function. Data have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO series accession numbers GSE154121 and GSE154122 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE154121).
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http://dx.doi.org/10.1016/j.jcmgh.2020.09.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7797367PMC
September 2020

A database for ITS2 sequences from nematodes.

BMC Genet 2020 07 10;21(1):74. Epub 2020 Jul 10.

Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, Calgary, Canada.

Background: Marker gene surveys have a wide variety of applications in species identification, population genetics, and molecular epidemiology. As these methods expand to new types of organisms and additional markers beyond 16S and 18S rRNA genes, comprehensive databases are a critical requirement for proper analysis of these data.

Results: Here we present an ITS2 rDNA database for marker gene surveys of both free-living and parasitic nematode populations and the software used to build the database. This is currently the most complete and up-to-date ITS2 database for nematodes and is able to reproduce previous analysis that used a smaller database.

Conclusions: This database is an important resource for researchers working on nematodes and also provides a tool to create ITS2 databases for any given taxonomy.
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http://dx.doi.org/10.1186/s12863-020-00880-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350610PMC
July 2020

Human interleukin-4-treated regulatory macrophages promote epithelial wound healing and reduce colitis in a mouse model.

Sci Adv 2020 Jun 5;6(23):eaba4376. Epub 2020 Jun 5.

Gastrointestinal Research Group and Inflammation Research Network, Department of Physiology and Pharmacology, Calvin, Joan and Phoebe Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Alberta, Canada.

Murine alternatively activated macrophages can exert anti-inflammatory effects. We sought to determine if IL-4-treated human macrophages [i.e., hM(IL4)] would promote epithelial wound repair and can serve as a cell transfer treatment for inflammatory bowel disease (IBD). Blood monocytes from healthy volunteers and patients with active and inactive IBD were converted to hM(IL4)s. IL-4 treatment of blood-derived macrophages from healthy volunteers and patients with inactive IBD resulted in a characteristic CD206CCL18CD14 phenotype (RNA-seq revealed IL-4 affected expression of 996 genes). Conditioned media from freshly generated or cryopreserved hM(IL4)s promoted epithelial wound healing in part by TGF, and reduced cytokine-driven loss of epithelial barrier function in vitro. Systemic delivery of hM(IL4) to dinitrobenzene sulphonic acid (DNBS)-treated mice significantly reduced disease. These findings from in vitro and in vivo analyses provide proof-of-concept support for the development of autologous M(IL4) transfer as a cellular immunotherapy for IBD.
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http://dx.doi.org/10.1126/sciadv.aba4376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7274799PMC
June 2020

Cystic Fibrosis Patients Infected With Epidemic Strains Have Unique Microbial Communities.

Front Cell Infect Microbiol 2020 24;10:173. Epub 2020 Apr 24.

Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, AB, Canada.

is the archetypal cystic fibrosis (CF) pathogen. However, the clinical course experienced by infected individuals varies markedly. Understanding these differences is imperative if further improvements in outcomes are to be achieved. Multiple studies have found that patients infected with epidemic (ePA) strains may have a worse clinical prognosis than those infected with unique, non-clonal strains. Additionally, the traditionally uncultured CF lung bacterial community (i.e., CF microbiome) may further influence the outcome. We sought to identify if these two important variables, not identified through routine culture, associate and together may contribute to disease pathogenesis. Patients were classified as being infected with Prairie Epidemic ePA (PES) or a non-clonal strain, unique PA strains (uPA), through a retrospective assessment of a comprehensive strain biobank using a combination of PFGE and PES-specific PCR. Patients were matched to age, sex, time-period controls and sputum samples from equivalent time periods were identified from a sputum biobank. Bacterial 16S rRNA gene profiling and qPCR was used to characterize the respiratory microbiome. We identified 31 patients infected with PES and matched them with uPA controls. Patients infected with PES at baseline have lower microbial diversity (P = 0.02) and higher relative abundance ( < 0.005). Microbial community structure was found to cluster by PA strain type, although it was not the main determinant of community structure as additional factors were also found to be drivers of CF community structure. Communities from PES infected individuals were enriched with and OTUs. The disproportionate disease experienced by ePA infected CF patients may be mediated through a combination of pathogen-pathogen factors as opposed to strictly enhanced virulence of infecting strains.
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http://dx.doi.org/10.3389/fcimb.2020.00173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7212370PMC
June 2021

Th1-Th2 Cross-Regulation Controls Early Leishmania Infection in the Skin by Modulating the Size of the Permissive Monocytic Host Cell Reservoir.

Cell Host Microbe 2020 05 15;27(5):752-768.e7. Epub 2020 Apr 15.

Snyder Institute for Chronic Diseases, University of Calgary, Calgary, AB T2N 4Z6, Canada; Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, AB T2N 4Z6, Canada; Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB T2N 4Z6, Canada. Electronic address:

The impact of T helper (Th) 1 versus Th2 immunity on intracellular infections is attributed to classical versus alternative activation of macrophages leading to resistance or susceptibility. However, observations in multiple infectious settings demonstrate deficiencies in mediators of Th1-Th2 immunity, which have paradoxical or no impact. We report that prior to influencing activation, Th1/Th2 immunity first controls the size of the permissive host cell reservoir. During early Leishmania infection of the skin, IFN-γ- or STAT6-mediated changes in phagocyte activation were counteracted by changes in IFN-γ-mediated recruitment of permissive CCR2 monocytes. Monocytes were required for early parasite expansion and acquired an alternatively activated phenotype despite the Th1 dermal environment required for their recruitment. Surprisingly, STAT6 did not enhance intracellular parasite proliferation, but rather modulated the size and permissiveness of the monocytic host cell reservoir via regulation of IFN-γ and IL-10. These observations expand our understanding of the Th1-Th2 paradigm during infection.
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http://dx.doi.org/10.1016/j.chom.2020.03.011DOI Listing
May 2020

Benchmarking hybrid assemblies of Giardia and prediction of widespread intra-isolate structural variation.

Parasit Vectors 2020 Feb 28;13(1):108. Epub 2020 Feb 28.

Department of Ecosystem and Public Health, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada.

Background: Currently available short read genome assemblies of the tetraploid protozoan parasite Giardia intestinalis are highly fragmented, highlighting the need for improved genome assemblies at a reasonable cost. Long nanopore reads are well suited to resolve repetitive genomic regions resulting in better quality assemblies of eukaryotic genomes. Subsequent addition of highly accurate short reads to long-read assemblies further improves assembly quality. Using this hybrid approach, we assembled genomes for three Giardia isolates, two with published assemblies and one novel, to evaluate the improvement in genome quality gained from long reads. We then used the long reads to predict structural variants to examine this previously unexplored source of genetic variation in Giardia.

Methods: With MinION reads for each isolate, we assembled genomes using several assemblers specializing in long reads. Assembly metrics, gene finding, and whole genome alignments to the reference genomes enabled direct comparison to evaluate the performance of the nanopore reads. Further improvements from adding Illumina reads to the long-read assemblies were evaluated using gene finding. Structural variants were predicted from alignments of the long reads to the best hybrid genome for each isolate and enrichment of key genes was analyzed using random genome sampling and calculation of percentiles to find thresholds of significance.

Results: Our hybrid assembly method generated reference quality genomes for each isolate. Consistent with previous findings based on SNPs, examination of heterozygosity using the structural variants found that Giardia BGS was considerably more heterozygous than the other isolates that are from Assemblage A. Further, each isolate was shown to contain structural variant regions enriched for variant-specific surface proteins, a key class of virulence factor in Giardia.

Conclusions: The ability to generate reference quality genomes from a single MinION run and a multiplexed MiSeq run enables future large-scale comparative genomic studies within the genus Giardia. Further, prediction of structural variants from long reads allows for more in-depth analyses of major sources of genetic variation within and between Giardia isolates that could have effects on both pathogenicity and host range.
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http://dx.doi.org/10.1186/s13071-020-3968-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7048089PMC
February 2020

Complex patterns of cell growth in the placenta in normal pregnancy and as adaptations to maternal diet restriction.

PLoS One 2020 9;15(1):e0226735. Epub 2020 Jan 9.

Department of Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary, Calgary Alberta.

The major milestones in mouse placental development are well described, but our understanding is limited to how the placenta can adapt to damage or changes in the environment. By using stereology and expression of cell cycle markers, we found that the placenta grows under normal conditions not just by hyperplasia of trophoblast cells but also through extensive polyploidy and cell hypertrophy. In response to feeding a low protein diet to mothers prior to and during pregnancy, to mimic chronic malnutrition, we found that this normal program was altered and that it was influenced by the sex of the conceptus. Male fetuses showed intrauterine growth restriction (IUGR) by embryonic day (E) 18.5, just before term, whereas female fetuses showed IUGR as early as E16.5. This difference was correlated with differences in the size of the labyrinth layer of the placenta, the site of nutrient and gas exchange. Functional changes were implied based on up-regulation of nutrient transporter genes. The junctional zone was also affected, with a reduction in both glycogen trophoblast and spongiotrophoblast cells. These changes were associated with increased expression of Phlda2 and reduced expression of Egfr. Polyploidy, which results from endoreduplication, is a normal feature of trophoblast giant cells (TGC) but also spongiotrophoblast cells. Ploidy was increased in sinusoidal-TGCs and spongiotrophoblast cells, but not parietal-TGCs, in low protein placentas. These results indicate that the placenta undergoes a range of changes in development and function in response to poor maternal diet, many of which we interpret are aimed at mitigating the impacts on fetal and maternal health.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0226735PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952106PMC
April 2020

foxc1 is required for embryonic head vascular smooth muscle differentiation in zebrafish.

Dev Biol 2019 09 11;453(1):34-47. Epub 2019 Jun 11.

Alberta Children's Hospital Research Institute, University of Calgary, Canada; Department of Biochemistry and Molecular Biology, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta, Canada, T2N 4N1. Electronic address:

Vascular smooth muscle of the head derives from neural crest, but developmental mechanisms and early transcriptional drivers of the vSMC lineage are not well characterized. We find that in early development, the transcription factor foxc1b is expressed in mesenchymal cells that associate with the vascular endothelium. Using timelapse imaging, we observe that foxc1b expressing mesenchymal cells differentiate into acta2 expressing vascular mural cells. We show that in zebrafish, while foxc1b is co-expressed in acta2 positive smooth muscle cells that associate with large diameter vessels, it is not co-expressed in capillaries where pdgfrβ positive pericytes are located. In addition to being an early marker of the lineage, foxc1 is essential for vSMC differentiation; we find that foxc1 loss of function mutants have defective vSMC differentiation and that early genetic ablation of foxc1b or acta2 expressing populations blocks vSMC differentiation. Furthermore, foxc1 is expressed upstream of acta2 and is required for acta2 expression in vSMCs. Using RNA-Seq we determine an enriched intersectional gene expression profile using dual expression of foxc1b and acta2 to identify novel vSMC markers. Taken together, our data suggests that foxc1 is a marker of vSMCs and plays a critical functional role in promoting their differentiation.
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http://dx.doi.org/10.1016/j.ydbio.2019.06.005DOI Listing
September 2019

The effects of cycled inhaled aztreonam on the cystic fibrosis (CF) lung microbiome.

J Cyst Fibros 2019 11 8;18(6):829-837. Epub 2019 Mar 8.

Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, AB, Canada; Department of Medicine, University of Calgary, Calgary, AB, Canada. Electronic address:

Background: To improve clinical outcomes, cystic fibrosis (CF) patients with chronic Pseudomonas aeruginosa infections are prescribed inhaled anti-pseudomonal antibiotics. Although, a diverse microbial community exists within CF airways, little is known about how the CF microbiota influences patient outcomes. We hypothesized that organisms within the CF microbiota are affected by inhaled-antibiotics and baseline microbiome may be used to predict therapeutic response.

Methods: Adults with chronic P. aeruginosa infection from four clinics were observed during a single 28-day on/off inhaled-aztreonam cycle. Patients performed serial sputum collection, CF-respiratory infection symptom scores (CRISS), and spirometry. Patients achieving a decrease of ≥2 CRISS by day 28 were categorized as subjective responders (SR). The airway microbiome was defined by Illumina MiSeq analysis of the 16S rRNA gene.

Results: Thirty-seven patients (median 37.4 years and FEV 44% predicted) were enrolled. No significant cohort-wide changes in the microbiome were observed between on/off AZLI cycles in either alpha- or beta-diversity metrics. However, at an individual level shifts were apparent. Twenty-one patients (57%) were SR and fourteen patients did not subjectively respond. While alpha-diversity metrics did not associate with response, patients who did not subjectively respond had a higher abundance of Staphylococcus and Streptococcus, and lower abundance of Haemophilus.

Conclusions: The CF microbiome is relatively resilient to AZLI perturbations. However, associated changes were observed at the individual patient level. The relative abundance of key "off-target" organisms associated with subjective improvements suggesting that the microbiome may be used as a tool to predict patient response - potentially improving outcomes.
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http://dx.doi.org/10.1016/j.jcf.2019.02.010DOI Listing
November 2019

Host genetics and diet composition interact to modulate gut microbiota and predisposition to metabolic syndrome in spontaneously hypertensive stroke-prone rats.

FASEB J 2019 06 1;33(6):6748-6766. Epub 2019 Mar 1.

Department of Production Animal Health, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada.

Metabolic syndrome encompasses obesity, glucose intolerance, hypertension, and dyslipidemia; however, the interactions between diet and host physiology that predispose to metabolic syndrome are incompletely understood. Here, we explored the effects of a high-fat diet (HFD) on energy balance, gut microbiota, and risk factors of metabolic syndrome in spontaneously hypertensive stroke-prone (SHRSP) and Wistar-Kyoto (WKY) rats. We found that the SHRSP rats were hypertensive, hyperphagic, less sensitive to hypophagic effects of exendin-4, and expended more energy with diminished sensitivity to sympathetic blockade compared to WKY rats. Notably, key thermogenic markers in brown and retroperitoneal adipose tissues and skeletal muscle were up-regulated in SHRSP than WKY rats. Although HFD promoted weight gain, adiposity, glucose intolerance, hypertriglyceridemia, hepatic lipidosis, and hyperleptinemia in both SHRSP and WKY rats, the SHRSP rats weighed less but had comparable percent adiposity to WKY rats, which supports the use of HFD-fed SHRSP rats as a unique model for studying the metabolically obese normal weight (MONW) phenotype in humans. Despite distinct strain differences in gut microbiota composition, diet had a preponderant impact on gut flora with some of the taxa being strongly associated with key metabolic parameters. Together, we provide evidence that interactions between host genetics and diet modulate gut microbiota and predispose SHRSP rats to develop metabolic syndrome.-Singh, A., Zapata, R. C., Pezeshki, A., Workentine, M. L., Chelikani, P. K. Host genetics and diet composition interact to modulate gut microbiota and predisposition to metabolic syndrome in spontaneously hypertensive stroke-prone rats.
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http://dx.doi.org/10.1096/fj.201801627RRRDOI Listing
June 2019

Macrophages treated with antigen from the tapeworm Hymenolepis diminuta condition CD25 T cells to suppress colitis.

FASEB J 2019 04 22;33(4):5676-5689. Epub 2019 Jan 22.

Department of Physiology and Pharmacology, Calvin, Joan, and Phoebe Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary Alberta, Canada.

Macrophages play central roles in immunity as early effectors and modulating adaptive immune reponses; we implicated macrophages in the anticolitic effect of infection with the tapeworm Hymenolepis diminuta. Here, gene arrays revealed that H. diminuta antigen (HdAg) evoked a program in murine macrophages distinct from that elicited by IL-4. Further, HdAg suppressed LPS-evoked release of TNF-α and IL-1β from macrophages via autocrine IL-10 signaling. In assessing the ability of macrophages treated in vitro with an extract of H. diminuta [M] to affect disease, intravenous, but not peritoneal, injection of M protected wild-type but not RAG1 mice from dinitrobenzene sulphonic acid (DNBS)-induced colitis. Administration of splenic CD4 T cells from in vitro cocultures with M, but not those cocultured with M cells, inhibited DNBS-induced colitis; fractionation of the T-cell population indicated that the CD4CD25 T cells from cocultures with M drove the suppression of DNBS-induced colitis. Use of IL-4 or IL-10 CD4 T cells revealed that neither cytokine alone from the donor cells was essential for the anticolitic effect. These data illustrate that HdAg evokes a unique regulatory program in macrophages, identifies HdAg-evoked IL-10 suppression of macrophage activation, and reveals the ability of HdAg-treated macrophages to educate ( i.e., condition) and mobilize CD4CD25 T cells, which could be deployed to treat colonic inflammation.-Reyes, J. L., Lopes, F., Leung, G., Jayme, T. S., Matisz, C. E., Shute, A., Burkhard, R., Carneiro, M., Workentine, M. L., Wang, A., Petri, B., Beck, P. L., Geuking, M. B., McKay, D. M., Macrophages treated with antigen from the tapeworm Hymenolepis diminuta condition CD25 T cells to suppress colitis.
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http://dx.doi.org/10.1096/fj.201802160RDOI Listing
April 2019

An intact microbiota is required for the gastrointestinal toxicity of the immunosuppressant mycophenolate mofetil.

J Heart Lung Transplant 2018 09 23;37(9):1047-1059. Epub 2018 May 23.

Department of Physiology and Pharmacology, Cumming School of Medicine; Snyder Institute for Chronic Diseases; Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine.

Background: Mycophenolate mofetil (MMF) is commonly prescribed after transplantation and has major advantages over other immunosuppressive drugs, but frequent gastrointestinal (GI) side-effects limit its use. The mechanism(s) underlying MMF-related GI toxicity have yet to be elucidated.

Methods: To investigate MMF-related GI toxicity, experimental mice were fed chow containing MMF (0.563%) and multiple indices of toxicity, including weight loss and colonic inflammation, were measured. Changes in intestinal microbial composition were detected using 16S rRNA Illumina sequencing, and downstream PICRUSt analysis was used to predict metagenomic pathways involved. Germ-free (GF) mice and mice treated with orally administered broad-spectrum antibiotics (ABX) were utilized to interrogate the importance of the microbiota in MMF-induced GI toxicity.

Results: Mice treated with MMF exhibited significant weight loss, related to loss of body fat and muscle, and marked colonic inflammation. MMF exposure was associated with changes in gut microbial composition, as demonstrated by a loss of overall diversity, expansion of Proteobacteria (specifically Escherichia/Shigella), and enrichment of genes involved in lipopolysaccharide (LPS) biosynthesis, which paralleled increased levels of LPS in the feces and serum. MMF-related GI toxicity was dependent on the intestinal microbiota, as MMF did not induce weight loss or colonic inflammation in GF mice. Furthermore, ABX prevented and reversed MMF-induced weight loss and colonic inflammation.

Conclusions: An intact intestinal microbiota is required to initiate and sustain the GI toxicity of MMF. MMF treatment causes dynamic changes in the composition of the intestinal microbiota that may be a targetable driver of the GI side-effects of MMF.
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http://dx.doi.org/10.1016/j.healun.2018.05.002DOI Listing
September 2018

Effects of transportation to and co-mingling at an auction market on nasopharyngeal and tracheal bacterial communities of recently weaned beef cattle.

Vet Microbiol 2018 Sep 9;223:126-133. Epub 2018 Aug 9.

Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada; Simpson Ranch Chair in Beef Cattle Health and Wellness, University of Calgary, Calgary, AB, Canada; Feedlot Health Management Services, Okotoks, AB, Canada. Electronic address:

The objective was to study effects of transportation to and co-mingling at an auction market on nasopharyngeal and tracheal bacterial communities of feedlot cattle. Two groups of 30 Angus-cross heifers were studied from weaning to 28 d after arrival at a feedlot. For each group, half the heifers were either transported directly to a feedlot after weaning (RANC) or transported to and co-mingled at an auction market for 24 h before being placed in a feedlot (AUCT). Deep nasal swabs (DNS) and trans-tracheal aspirates (TTA) were collected at weaning (d0) and at on-arrival processing at the feedlot (d2). At 7 (d9) and 28 d (d30) after arrival, DNS were repeated. The DNA was extracted from DNS and TTA and the V4 region of the 16S rRNA gene sequenced (MiSeq). Alpha diversity analysis did not reveal differences between AUCT and RANC. However, bacterial diversity decreased over time in the nasopharynx, especially at d9. Although beta-diversity was not different between AUCT and RANC, interval after arrival and feedlot where heifers were placed affected composition of the nasopharyngeal bacterial communities. In both groups, a large increase in Mycoplasma was observed after arrival; in one group, Mycoplasma bovis was dominant at d9 and remained dominant until d30. However, in the other group, Mycoplasma dispar dominated at d9 and was replaced by Moraxella at d30. We concluded that transportation to and co-mingling at an auction market for 24 h did not significantly influence diversity or composition of nasopharyngeal or tracheal bacterial communities.
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http://dx.doi.org/10.1016/j.vetmic.2018.08.007DOI Listing
September 2018

Sputum microbiota is predictive of long-term clinical outcomes in young adults with cystic fibrosis.

Thorax 2018 11 22;73(11):1016-1025. Epub 2018 Aug 22.

Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada.

Background: Complex polymicrobial communities infect cystic fibrosis (CF) lower airways. Generally, communities with low diversity, dominated by classical CF pathogens, associate with worsened patient status at sample collection. However, it is not known if the microbiome can predict future outcomes. We sought to determine if the microbiome could be adapted as a biomarker for patient prognostication.

Methods: We retrospectively assessed prospectively collected sputum from a cohort of 104 individuals aged 18-22 to determine factors associated with progression to early end-stage lung disease (eESLD; death/transplantation <25 years) and rapid pulmonary function decline (>-3%/year FEV over the ensuing 5 years). Illumina MiSeq paired-end sequencing of the V3-V4 region of the 16S rRNA was used to define the airway microbiome.

Results: Based on the primary outcome analysed, 17 individuals (16%) subsequently progressed to eESLD. They were more likely to have sputum with low alpha diversity, dominated by specific pathogens including . Communities with abundant were observed to be protective. Microbial communities clustered together by baseline lung disease stage and subsequent progression to eESLD. Multivariable analysis identified baseline lung function and alpha diversity as independent predictors of eESLD. For the secondary outcomes, 58 and 47 patients were classified as rapid progressors based on absolute and relative definitions of lung function decline, respectively. Patients with low alpha diversity were similarly more likely to be classified as experiencing rapid lung function decline over the ensuing 5 years when adjusted for baseline lung function.

Conclusions: We observed that the diversity of microbial communities in CF airways is predictive of progression to eESLD and disproportionate lung function decline and may therefore represent a novel biomarker.
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http://dx.doi.org/10.1136/thoraxjnl-2018-211510DOI Listing
November 2018

The effects of inhaled aztreonam on the cystic fibrosis lung microbiome.

Microbiome 2017 05 5;5(1):51. Epub 2017 May 5.

Department of Microbiology, Immunology and Infectious Diseases, The University of Calgary, Calgary, AB, Canada.

Background: Aztreonam lysine for inhalation (AZLI) is an inhaled antibiotic used to treat chronic Pseudomonas aeruginosa infection in CF. AZLI improves lung function and quality of life, and reduces exacerbations-improvements attributed to its antipseudomonal activity. Given the extremely high aztreonam concentrations achieved in the lower airways by nebulization, we speculate this may extend its spectrum of activity to other organisms. As such, we sought to determine if AZLI affects the CF lung microbiome and whether community constituents can be used to predict treatment responsiveness.

Methods: Patients were included if they had chronic P. aeruginosa infection and repeated sputum samples collected before and after AZLI. Sputum DNA was extracted, and the V3-hypervariable region of the 16S ribosomal RNA (rRNA) gene amplified and sequenced.

Results: Twenty-four patients naïve to AZLI contributed 162 samples. The cohort had a median age of 37.1 years, and a  median FEV of 44% predicted. Fourteen patients were a priori defined as responders for achieving ≥3% FEV improvement following initiation. No significant changes in alpha diversity were noted following AZLI. Furthermore, beta diversity demonstrated clustering with respect to patients, but had no association with AZLI use. However, we did observe a decline in the relative abundance of several individual operational taxonomic units (OTUs) following AZLI initiation suggesting that specific sub-populations of organisms may be impacted. Patients with higher abundance of Staphylococcus and anaerobic organisms including Prevotella and Fusobacterium were less likely to respond to therapy.

Conclusions: Results from our study suggest potential alternate/additional mechanisms by which AZLI functions. Moreover, our study suggests that the CF microbiota may be used as a biomarker to predict patient responsiveness to therapy suggesting the microbiome may be harnessed for the personalization of therapies.
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http://dx.doi.org/10.1186/s40168-017-0265-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5420135PMC
May 2017

Changes in Composition of the Gut Bacterial Microbiome after Fecal Microbiota Transplantation for Recurrent Infection in a Pediatric Heart Transplant Patient.

Front Cardiovasc Med 2017 4;4:17. Epub 2017 Apr 4.

Department of Paediatrics, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada.

The microbiome is increasingly recognized as an important influence on human health and many of the comorbidities that affect patients after solid organ transplantation (SOT) have been shown to involve changes in gut bacterial populations. Thus, microbiome changes in an individual patient may have important health implications after SOT but this area remains understudied. We describe changes in the composition of the fecal microbiome from a pediatric heart transplant recipient before and >2.5 years after he underwent repeated fecal microbiota transplantation (FMT) for recurrent infection (CDI). With both documented episodes of CDI, there was marked loss of bacterial diversity with overgrowth of Proteobacteria (>98.9% of phyla identified) associated with symptomatic colitis that was corrected after FMT. We hypothesize that a second CDI occurring after FMT was related to incomplete restoration of normal bowel flora post-FMT with relative deficiencies of the phyla Firmicutes and Bacteroidetes and the families and . Following the second FMT, there was a gradual shift in gut bacterial composition coincident with the recipient developing lymphonodular hyperplasia of the colon and painless hematochezia that resolved with discontinuation of mycophenolate mofetil (MMF). This case documents dynamic changes in the bacterial microbiome after FMT and suggests that MMF may influence the gut microbiome with consequences for the patient.
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http://dx.doi.org/10.3389/fcvm.2017.00017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5378704PMC
April 2017

Giardia duodenalis induces pathogenic dysbiosis of human intestinal microbiota biofilms.

Int J Parasitol 2017 05 22;47(6):311-326. Epub 2017 Feb 22.

Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, Alberta T2N 4N1, Canada; Department of Physiology & Pharmacology, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta T2N 4N1, Canada. Electronic address:

Giardia duodenalis is a prevalent cause of acute diarrheal disease worldwide. However, recent outbreaks in Italy and Norway have revealed a link between giardiasis and the subsequent development of chronic post-infectious irritable bowel syndrome. While the mechanisms underlying the causation of post-infectious irritable bowel syndrome remain obscure, recent findings suggest that alterations in gut microbiota communities are linked to the pathophysiology of irritable bowel syndrome. In the present study, we use a laboratory biofilm system to culture and enrich mucosal microbiota from human intestinal biopsies. Subsequently, we show that co-culture with Giardia induces disturbances in biofilm species composition and biofilm structure resulting in microbiota communities that are intrinsically dysbiotic - even after the clearance of Giardia. These microbiota abnormalities were mediated in part by secretory-excretory Giardia cysteine proteases. Using in vitro cell culture and germ-free murine infection models, we show that Giardia-induced disruptions of microbiota promote bacterial invasion, resulting in epithelial apoptosis, tight junctional disruption, and bacterial translocation across an intestinal epithelial barrier. Additionally, these dysbiotic microbiota communities resulted in increased activation of the Toll-like receptor 4 signalling pathway, and overproduction of the pro-inflammatory cytokine IL-1beta in humanized germ-free mice. Previous studies that have sought explanations and risk factors for the development of post-infectious irritable bowel syndrome have focused on features of enteropathogens and attributes of the infected host. We propose that polymicrobial interactions involving Giardia and gut microbiota may cause persistent dysbiosis, offering a new interpretation of the reasons why those afflicted with giardiasis are predisposed to gastrointestinal disorders post-infection.
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http://dx.doi.org/10.1016/j.ijpara.2016.11.010DOI Listing
May 2017

Genomic analysis of the multi-host pathogen Erysipelothrix rhusiopathiae reveals extensive recombination as well as the existence of three generalist clades with wide geographic distribution.

BMC Genomics 2016 06 14;17:461. Epub 2016 Jun 14.

Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada.

Background: Knowledge about how bacterial populations are structured is an important prerequisite for studying their ecology and evolutionary history and facilitates inquiry into host specificity, pathogenicity, geographic dispersal and molecular epidemiology. Erysipelothrix rhusiopathiae is an opportunistic pathogen that is currently reemerging in both the swine and poultry industries globally. This bacterium sporadically causes mortalities in captive marine mammals, and has recently been implicated in large-scale wildlife die-offs. However, despite its economic relevance and broad geographic and host distribution, including zoonotic potential, the global diversity, recombination rates, and population structure of this bacterium remain poorly characterized. In this study, we conducted a broad-scale genomic comparison of E. rhusiopathiae based on a diverse collection of isolates in order to address these knowledge gaps.

Results: Eighty-three E. rhusiopathiae isolates from a range of host species and geographic origins, isolated between 1958 and 2014, were sequenced and assembled using both reference-based mapping and de novo assembly. We found that a high proportion of the core genome (58 %) had undergone recombination. Therefore, we used three independent methods robust to the presence of recombination to define the population structure of this species: a phylogenetic tree based on a set of conserved protein sequences, in silico chromosome painting, and network analysis. All three methods were broadly concordant and supported the existence of three distinct clades within the species E. rhusiopathiae. Although we found some evidence of host and geographical clustering, each clade included isolates from diverse host species and from multiple continents.

Conclusions: Using whole genome sequence data, we confirm recent suggestions that E. rhusiopathiae is a weakly clonal species that has been shaped extensively by homologous recombination. Despite frequent recombination, we can reliably identify three distinct clades that do not clearly segregate by host species or geographic origin. Our results provide an essential baseline for future molecular epidemiological, ecological and evolutionary studies of E. rhusiopathiae and facilitate comparisons to other recombinogenic, multi-host bacteria.
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http://dx.doi.org/10.1186/s12864-016-2643-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4906694PMC
June 2016

Cyanide Toxicity to Burkholderia cenocepacia Is Modulated by Polymicrobial Communities and Environmental Factors.

Front Microbiol 2016 18;7:725. Epub 2016 May 18.

Department of Medicine, Faculty of Health Sciences, Farncombe Family Digestive Health Research Institute, McMaster UniversityHamilton, ON, Canada; Department of Biochemistry and Biomedical Sciences, Faculty of Health Sciences, McMaster UniversityHamilton, ON, Canada.

Microbes within polymicrobial communities can establish positive and negative interactions that have the potential to influence the overall behavior of the community. Pseudomonas aeruginosa and species of the Burkholderia cepacia complex (Bcc) can co-exist in the lower airways, however several studies have shown that P. aeruginosa can effectively kill the Bcc in vitro, for which hydrogen cyanide (HCN) was recently proposed to play a critical role. Here we show that modification of the environment (i.e., culture medium), long-term genetic adaptation of P. aeruginosa to the cystic fibrosis (CF) lung, or the addition of another bacterial species to the community can alter the sensitivity of Burkholderia cenocepacia to P. aeruginosa toxins. We specifically demonstrate that undefined rich media leads to higher susceptibility of B. cenocepacia to P. aeruginosa toxins like cyanide as compared to a synthetic medium (SCFM), that mimics the CF lung nutritional content. Overall, our study shows that the polymicrobial environment can have profound effects on negative interactions mediated by P. aeruginosa against B. cenocepacia. In fact, evolved P. aeruginosa or the presence of other species such as Staphylococcus aureus can directly abolish the direct competition mediated by cyanide and consequently maintaining a higher level of species diversity within the community.
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http://dx.doi.org/10.3389/fmicb.2016.00725DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4870242PMC
May 2016

The Challenge and Potential of Metagenomics in the Clinic.

Front Immunol 2016 3;7:29. Epub 2016 Feb 3.

Faculty of Veterinary Medicine, University of Calgary , Calgary, AB , Canada.

The bacteria, fungi, and viruses that live on and in us have a tremendous impact on our day-to-day health and are often linked to many diseases, including autoimmune disorders and infections. Diagnosing and treating these disorders relies on accurate identification and characterization of the microbial community. Current sequencing technologies allow the sequencing of the entire nucleic acid complement of a sample providing an accurate snapshot of the community members present in addition to the full genetic potential of that microbial community. There are a number of clinical applications that stand to benefit from these data sets, such as the rapid identification of pathogens present in a sample. Other applications include the identification of antibiotic-resistance genes, diagnosis and treatment of gastrointestinal disorders, and many other diseases associated with bacterial, viral, and fungal microbiomes. Metagenomics also allows the physician to probe more complex phenotypes such as microbial dysbiosis with intestinal disorders and disruptions of the skin microbiome that may be associated with skin disorders. Many of these disorders are not associated with a single pathogen but emerge as a result of complex ecological interactions within microbiota. Currently, we understand very little about these complex phenotypes, yet clearly they are important and in some cases, as with fecal microbiota transplants in Clostridium difficile infections, treating the microbiome of the patient is effective. Here, we give an overview of metagenomics and discuss a number of areas where metagenomics is applicable in the clinic, and progress being made in these areas. This includes (1) the identification of unknown pathogens, and those pathogens particularly hard to culture, (2) utilizing functional information and gene content to understand complex infections such as Clostridium difficile, and (3) predicting antimicrobial resistance of the community using genetic determinants of resistance identified from the sequencing data. All of these applications rely on sophisticated computational tools, and we also discuss the importance of skilled bioinformatic support for the implementation and use of metagenomics in the clinic.
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http://dx.doi.org/10.3389/fimmu.2016.00029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4737888PMC
February 2016

Hydrogen sulfide protects from colitis and restores intestinal microbiota biofilm and mucus production.

Inflamm Bowel Dis 2015 May;21(5):1006-17

Departments of *Biological Sciences, and †Physiology and Pharmacology, University of Calgary, Calgary, AB, Canada; ‡Department of Medicine, McMaster University, Hamilton, ON, Canada; §Centre of Neurosciences and Cell Biology, Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal; and ‖Department of Biology, Lakehead University, Thunder Bay, ON, Canada.

Background: Microbiota dysbiosis and impaired barrier function are among the most prominent features of inflammatory bowel disease. In the gastrointestinal tract, hydrogen sulfide (H(2)S) is an important regulator of mucosal homeostasis. We hypothesized that H(2)S promotes resolution of colonic inflammation through actions on microbiota biofilm and the mucus barrier.

Methods: We used mice genetically deficient for a key enzyme for H(2)S production (cystathionine γ-lyase) and pharmacologically inhibited that enzyme during colitis in wild-type mice. We tested the effects of administering an H(2)S donor (diallyl disulfide) to rodents during hapten-induced colitis. Colonic microbiota biofilm was visualized by fluorescent in situ hybridization, and mucus granules were quantified with periodic acid-alcian blue staining. We exposed human microbiota biofilms and planktonic bacteria to H(2)S donors ex vivo to determine changes in their growth, viability, and biomass.

Results: Intestinal microbiota formed linear biofilms in the colon of healthy rodents. During colitis, microbiota biofilms were fragmented and mucus granule production decreased. Endogenous production of H(2)S had beneficial effects on establishment of microbiota biofilms and colonic mucus production. Therapeutic delivery of H(2)S into the colon reduced inflammation, restored the microbiota biofilm, and increased the production of mucus granules. In ex vivo human microbiota, H(2)S not only promoted biofilm formation but also reduced growth of planktonic bacteria.

Conclusions: Our results suggest that H(2)S donors could be used therapeutically during colitis, facilitating correction of microbiota biofilm dysbiosis and mucus layer reconstitution.
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http://dx.doi.org/10.1097/MIB.0000000000000345DOI Listing
May 2015

Draft Genome Sequence of Burkholderia dolosa PC543 Isolated from Cystic Fibrosis Airways.

Genome Announc 2014 Feb 13;2(1). Epub 2014 Feb 13.

Farncombe Family Digestive Health Research Institute, Faculty of Health Sciences, Department of Medicine, McMaster University, Hamilton, Ontario, Canada.

Burkholderia dolosa is a member of the Burkholderia cepacia complex, a group of opportunistic bacterial pathogens often associated with fatal chronic infections in the lungs of patients suffering from cystic fibrosis (CF). Here, we announce the draft genome sequence of B. dolosa PC543 (LMG 19468), a CF airway isolate.
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http://dx.doi.org/10.1128/genomeA.00043-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3924365PMC
February 2014

Twenty-five-year outbreak of Pseudomonas aeruginosa infecting individuals with cystic fibrosis: identification of the prairie epidemic strain.

J Clin Microbiol 2014 Apr 22;52(4):1127-35. Epub 2014 Jan 22.

The University of Calgary, Calgary, Alberta, Canada.

Transmissible strains of Pseudomonas aeruginosa have been described for cystic fibrosis (CF) and may be associated with a worse prognosis. Using a comprehensive strain biobank spanning 3 decades, we sought to determine the prevalence and stability of chronic P. aeruginosa infection in an adult population. P. aeruginosa isolates from sputum samples collected at initial enrollment in our adult clinic and at the most recent clinic visit were examined by a combination of pulsed-field gel electrophoresis and multilocus sequence typing and compared against a collection of established transmissible and local non-CF bronchiectasis (nCFB) isolates. A total of 372 isolates from 107 patients, spanning 674 patient-years, including 66 patients with matched isolates from initial and final encounters, were screened. A novel clone with increased antibacterial resistance, termed the prairie epidemic strain (PES), was found in 29% (31/107 patients) of chronically infected patients referred from multiple prairie-based CF centers. This isolate was not found in those diagnosed with CF as adults or in a control population with nCFB. While 90% (60/66 patients) of patients had stable infection over a mean of 10.8 years, five patients experienced strain displacement of unique isolates, with PES occurring within 2 years of transitioning to adult care. PES has been present in our cohort since at least 1987, is unique to CF, generally establishes chronic infection during childhood, and has been found in patients at the time of transition of patients from multiple prairie-based CF clinics, suggesting broad endemicity. Studies are under way to evaluate the clinical implications of PES infection.
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http://dx.doi.org/10.1128/JCM.03218-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3993468PMC
April 2014

Genetic signature of bacterial pathogen adaptation during chronic pulmonary infections.

Nat Genet 2014 Jan;46(1):5-6

Farncombe Family Digestive Health Research Institute, Faculty of Health Sciences, Department of Medicine, McMaster University, Hamilton, Ontario, Canada.

To establish and maintain chronic infections, many pathogens adapt in response to selective pressures within the host, leaving unique genetic signatures. A new study uses whole-genome and population sequencing approaches to identify evidence of adaptive evolution in Burkholderia dolosa genomes isolated from chronic infections in patients with cystic fibrosis.
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http://dx.doi.org/10.1038/ng.2859DOI Listing
January 2014

Spatial distributions of Pseudomonas fluorescens colony variants in mixed-culture biofilms.

BMC Microbiol 2013 Jul 28;13:175. Epub 2013 Jul 28.

Department of Biological Sciences, University of Calgary, Calgary, Canada.

Background: The emergence of colony morphology variants in structured environments is being recognized as important to both niche specialization and stress tolerance. Pseudomonas fluorescens demonstrates diversity in both its natural environment, the rhizosphere, and in laboratory grown biofilms. Sub-populations of these variants within a biofilm have been suggested as important contributors to antimicrobial stress tolerance given their altered susceptibility to various agents. As such it is of interest to determine how these variants might be distributed in the biofilm environment.

Results: Here we present an analysis of the spatial distribution of Pseudomonas fluorescens colony morphology variants in mixed-culture biofilms with the wildtype phenotype. These findings reveal that two variant colony morphotypes demonstrate a significant growth advantage over the wildtype morphotype in the biofilm environment. The two variant morphotypes out-grew the wildtype across the entire biofilm and this occurred within 24 h and was maintained through to 96 h. This competitive advantage was not observed in homogeneous broth culture.

Conclusions: The significant advantage that the variants demonstrate in biofilm colonization over the wildtype denotes the importance of this phenotype in structured environments.
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http://dx.doi.org/10.1186/1471-2180-13-175DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3734068PMC
July 2013

Phenotypic heterogeneity of Pseudomonas aeruginosa populations in a cystic fibrosis patient.

PLoS One 2013 3;8(4):e60225. Epub 2013 Apr 3.

Farncombe Family Digestive Health Research Institute, McMaster University, Hamilton, Ontario, Canada.

The opportunistic pathogen Pseudomonas aeruginosa chronically infects the lower airways of patients with cystic fibrosis. Throughout the course of infection this organism undergoes adaptations that contribute to its long-term persistence in the airways. While P. aeruginosa diversity has been documented, it is less clear to what extent within-patient diversity contributes to the overall population structure as most studies have been limited to the analysis of only a few isolates per patient per time point. To examine P. aeruginosa population structure in more detail we collected multiple isolates from individual sputum samples of a patient chronically colonized with P. aeruginosa. This strain collection, comprised of 169 clonal isolates and representing three pulmonary exacerbations as well as clinically stable periods, was assayed for a wide selection of phenotypes. These phenotypes included colony morphology, motility, quorum sensing, protease activity, auxotrophy, siderophore levels, antibiotic resistance, and growth profiles. Each phenotype displayed significant variation even within isolates of the same colony morphotype from the same sample. Isolates demonstrated a large degree of individuality across phenotypes, despite being part of a single clonal lineage, suggesting that the P. aeruginosa population in the cystic fibrosis airways is being significantly under-sampled.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0060225PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3616088PMC
October 2013
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