@article{bruno-barcena_azcarate-peril_hassan_2010, title={Role of Antioxidant Enzymes in Bacterial Resistance to Organic Acids}, volume={76}, ISSN={["1098-5336"]}, url={http://europepmc.org/abstract/med/20305033}, DOI={10.1128/aem.02718-09}, abstractNote={ABSTRACTGrowth in aerobic environments has been shown to generate reactive oxygen species (ROS) and to cause oxidative stress in most organisms. Antioxidant enzymes (i.e., superoxide dismutases and hydroperoxidases) and DNA repair mechanisms provide protection against ROS. Acid stress has been shown to be associated with the induction of Mn superoxide dismutase (MnSOD) inLactococcus lactisandStaphylococcus aureus. However, the relationship between acid stress and oxidative stress is not well understood. In the present study, we showed that mutations in the gene coding for MnSOD (sodA) increased the toxicity of lactic acid at pH 3.5 inStreptococcus thermophilus. The inclusion of the iron chelators 2,2′-dipyridyl (DIP), diethienetriamine-pentaacetic acid (DTPA), andO-phenanthroline (O-Phe) provided partial protection against 330 mM lactic acid at pH 3.5. The results suggested that acid stress triggers an iron-mediated oxidative stress that can be ameliorated by MnSOD and iron chelators. These findings were further validated inEscherichia colistrains lacking both MnSOD and iron SOD (FeSOD) but expressing a heterologous MnSOD fromS. thermophilus. We also found that, inE. coli, FeSOD did not provide the same protection afforded by MnSOD and that hydroperoxidases are equally important in protecting the cells against acid stress. These findings may explain the ability of some microorganisms to survive better in acidified environments, as in acid foods, during fermentation and accumulation of lactic acid or during passage through the low pH of the stomach.}, number={9}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Bruno-Barcena, Jose M. and Azcarate-Peril, M. Andrea and Hassan, Hosni M.}, year={2010}, month={May}, pages={2747–2753} }
 @misc{barrangou_azcarate-peril_altermann_duong_klaenhammer_2009, title={Compositions comprising promoter sequences and methods of use}, volume={7,495,092}, number={2009 Feb. 24}, author={Barrangou, R. and Azcarate-Peril, A. and Altermann, E. and Duong, T. and Klaenhammer, T. R.}, year={2009} }
 @article{goh_azcarate-peril_o'flaherty_durmaz_valence_jardin_lortal_klaenhammer_2009, title={Development and Application of a upp-Based Counterselective Gene Replacement System for the Study of the S-Layer Protein SlpX of Lactobacillus acidophilus NCFM}, volume={75}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.02502-08}, abstractNote={ABSTRACT
          
            In silico genome analysis of
            Lactobacillus acidophilus
            NCFM coupled with gene expression studies have identified putative genes and regulatory networks that are potentially important to this organism's survival, persistence, and activities in the gastrointestinal tract. Correlation of key genotypes to phenotypes requires an efficient gene replacement system. In this study, use of the
            upp
            -encoded uracil phosphoribosyltransferase (UPRTase) of
            L. acidophilus
            NCFM was explored as a counterselection marker to positively select for recombinants that have resolved from chromosomal integration of pORI-based plasmids. An isogenic mutant carrying a
            upp
            gene deletion was constructed and was resistant to 5-fluorouracil (5-FU), a toxic uracil analog that is also a substrate for UPRTase. A 3.0-kb pORI-based counterselectable integration vector bearing a
            upp
            expression cassette, pTRK935, was constructed and introduced into the Δ
            upp
            host harboring the pTRK669 helper plasmid. Extrachromosomal replication of pTRK935 complemented the mutated chromosomal
            upp
            allele and restored sensitivity to 5-FU. This host background provides a platform for a two-step plasmid integration and excision strategy that can select for plasmid-free recombinants with either the wild-type or mutated allele of the targeted gene in the presence of 5-FU. The efficacy of the system was demonstrated by in-frame deletion of the
            slpX
            gene (LBA0512) encoding a novel 51-kDa secreted protein associated with the S-layer complex of
            L. acidophilus
            . The resulting Δ
            slpX
            mutant exhibited lower growth rates, increased sensitivity to sodium dodecyl sulfate, and greater resistance to bile. Overall, this improved gene replacement system represents a valuable tool for investigating the mechanisms underlying the probiotic functionality of
            L. acidophilus
            .
          }, number={10}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Goh, Yong Jun and Azcarate-Peril, M. Andrea and O'Flaherty, Sarah and Durmaz, Evelyn and Valence, Florence and Jardin, Julien and Lortal, Sylvie and Klaenhammer, Todd R.}, year={2009}, month={May}, pages={3093–3105} }
 @misc{klaenhammer_russell_altermann_azcarate-peril_2009, title={Nucleic acid sequences encoding two-component sensing and regulatory proteins, antimicrobial proteins and uses therefor}, volume={7,550,576}, number={2009 Jun 23}, author={Klaenhammer, T. R. and Russell, W. M. and Altermann, E. and Azcarate-Peril, A.}, year={2009} }
 @article{buck_azcarate-peril_klaenhammer_2009, title={Role of autoinducer-2 on the adhesion ability of Lactobacillus acidophilus}, volume={107}, ISSN={["1365-2672"]}, DOI={10.1111/j.1365-2672.2009.04204.x}, abstractNote={Aims:  Lactobacilli adhere to the intestinal epithelium and this intimate association likely promotes retention in the gastrointestinal tract and communication with the immune system. The aim of this study was to investigate whether or not the quorum‐sensing signalling molecule, autoinducer (AI)‐2, was produced by Lactobacillus acidophilus and affected adherence to intestinal epithelial cells.}, number={1}, journal={JOURNAL OF APPLIED MICROBIOLOGY}, author={Buck, B. L. and Azcarate-Peril, M. A. and Klaenhammer, T. R.}, year={2009}, month={Jul}, pages={269–279} }
 @misc{klaenhammer_azcarate-peril_altermann_2008, title={Lactobacillus acidophilus nucleic acids and uses thereof}, volume={7,468,182}, number={2008 Dec. 23}, author={Klaenhammer, T. and Azcarate-Peril, A. and Altermann, E.}, year={2008} }
 @article{pfeiler_azcarate-peril_klaenhammer_2007, title={Characterization of a novel bile-inducible operon encoding a two-component regulatory system in Lactobacillus acidophilus}, volume={189}, ISSN={["1098-5530"]}, DOI={10.1128/JB.00337-07}, abstractNote={ABSTRACTLactobacillus acidophilusNCFM is an industrially important strain used extensively as a probiotic culture. Tolerance of the presence of bile is an attribute important to microbial survival in the intestinal tract. A whole-genome microarray was employed to examine the effects of bile on the global transcriptional profile of this strain, with the intention of elucidating genes contributing to bile tolerance. Genes involved in carbohydrate metabolism were generally induced, while genes involved in other aspects of cellular growth were mostly repressed. A 7-kb eight-gene operon encoding a two-component regulatory system (2CRS), a transporter, an oxidoreductase, and four hypothetical proteins was significantly upregulated in the presence of bile. Deletion mutations were constructed in six genes of the operon. Transcriptional analysis of the 2CRS mutants showed that mutation of the histidine protein kinase (HPK) had no effect on the induction of the operon, whereas the mutated response regulator (RR) showed enhanced induction when the cells were exposed to bile. These results indicate that the 2CRS plays a role in bile tolerance and that the operon it resides in is negatively controlled by the RR. Mutations in the transporter, the HPK, the RR, and a hypothetical protein each resulted in loss of tolerance of bile. Mutations in genes encoding another hypothetical protein and a putative oxidoreductase resulted in significant increases in bile tolerance. This functional analysis showed that the operon encoded proteins involved in both bile tolerance and bile sensitivity.}, number={13}, journal={JOURNAL OF BACTERIOLOGY}, author={Pfeiler, Erika A. and Azcarate-Peril, M. Andrea and Klaenhammer, Todd R.}, year={2007}, month={Jul}, pages={4624–4634} }
 @article{klaenhammer_azcarate-peril_altermann_barrangou_2007, title={Influence of the dairy environment on gene expression and substrate(1-3)}, volume={137}, ISSN={["0022-3166"]}, DOI={10.1093/jn/137.3.748s}, abstractNote={Lactic acid bacteria (LAB) are widely used for the industrial production of fermented dairy products and form a group of related low-GC-content gram-positive bacteria. The major species used in dairy manufacturing are Lactobacillus, Lactococcus, Streptococcus, and Leuconostoc. Traditionally most are applied as starter cultures for dairy fermentations or used as probiotic cultures, delivered in dairy vehicles. The appearance of the genomes of Lactococcus lactis, Bidifobacterium longum, Lactobacillus plantarum, L. johnsonii, L. acidophilus, 2 strains of Streptococcus thermophilus, and pending completion of many draft genomic sequences, is now promoting in-depth investigation into the comparative genetic content of LAB. Moreover, whole-genome transcriptional arrays are quickly revealing critical genes/operons that are coordinately expressed and the impact of environmental factors on expression of multiple gene sets. Comparative genomics between multiple genomes is providing insights into genes that are important in metabolic, physiological, and functional roles for different LAB in the environments they inhabit, ranging from the gastrointestinal tract to milk and acidified dairy products.}, number={3}, journal={JOURNAL OF NUTRITION}, author={Klaenhammer, Todd R. and Azcarate-Peril, M. Andrea and Altermann, Eric and Barrangou, Rodolphe}, year={2007}, month={Mar}, pages={748S–750S} }
 @article{barrangou_azcarate-peril_duong_conners_kelly_klaenhammer_2006, title={Global analysis of carbohydrate utilization by Lactobacillus acidophilus using cDNA microarrays}, volume={103}, ISSN={["0027-8424"]}, DOI={10.1073/pnas.0511287103}, abstractNote={
            The transport and catabolic machinery involved in carbohydrate utilization by
            Lactobacillus acidophilus
            was characterized genetically by using whole-genome cDNA microarrays. Global transcriptional profiles were determined for growth on glucose, fructose, sucrose, lactose, galactose, trehalose, raffinose, and fructooligosaccharides. Hybridizations were carried out by using a round-robin design, and microarray data were analyzed with a two-stage mixed model ANOVA. Differentially expressed genes were visualized by hierarchical clustering, volcano plots, and contour plots. Overall, only 63 genes (3% of the genome) showed a >4-fold induction. Specifically, transporters of the phospho
            enol
            pyruvate:sugar transferase system were identified for uptake of glucose, fructose, sucrose, and trehalose, whereas ATP-binding cassette transporters were identified for uptake of raffinose and fructooligosaccharides. A member of the LacS subfamily of galactoside-pentose hexuronide translocators was identified for uptake of galactose and lactose. Saccharolytic enzymes likely involved in the metabolism of monosaccharides, disaccharides, and polysaccharides into substrates of glycolysis were also found, including enzymatic machinery of the Leloir pathway. The transcriptome appeared to be regulated by carbon catabolite repression. Although substrate-specific carbohydrate transporters and hydrolases were regulated at the transcriptional level, genes encoding regulatory proteins CcpA, Hpr, HprK/P, and EI were consistently highly expressed. Genes central to glycolysis were among the most highly expressed in the genome. Collectively, microarray data revealed that coordinated and regulated transcription of genes involved in sugar uptake and metabolism is based on the specific carbohydrate provided.
            L. acidophilus
            's adaptability to environmental conditions likely contributes to its competitive ability for limited carbohydrate sources available in the human gastrointestinal tract.
          }, number={10}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, publisher={Proceedings of the National Academy of Sciences}, author={Barrangou, R and Azcarate-Peril, MA and Duong, T and Conners, SB and Kelly, RM and Klaenhammer, TR}, year={2006}, month={Mar}, pages={3816–3821} }
 @article{klaenhammer_peril_barrangou_duong_altermann_2005, title={Genomic Perspectives on Probiotic Lactic Acid Bacteria}, volume={24}, ISBN={1342-1441}, DOI={10.12938/bifidus.24.31}, abstractNote={The lactic acid bacteria are Gram-positive fermentative microorganisms known primarily for their roles as starter cultures and probiotics. The food industry represents one of the largest manufacturing industries in the world and recent trends are rapidly expanding the use of probiotic cultures within functional foods. Understanding and control of lactic acid bacteria is now being revolutionized by genomic sciences and the appearance of the complete genome sequences for Bifidobacterium longum, Lactobacillus johnsonii, Lactobacillus plantarum, and draft sequences for Lactobacillus gasseri and Lactobacillus casei. This explosion of DNA sequence information, accompanied by the development of bioinformatic tools for nucleic acid and protein analysis, now allows rapid characterization of the lactic acid bacteria for their genomic content and expression profiles across the entire genome. Comparative genomics has already revealed important similarities and differences in strains, species, and genera and will likely identify key genetic features responsible for the beneficial properties ascribed to probiotic lactic acid bacteria. Practical genomics for the lactic acid bacteria promises to establish the genetic landscape, correlate genotypes with desirable phenotypes, establish genetic criteria for strain selection, improve culture stability by stress preconditioning, provide opportunities for metabolic engineering, and uncover a mechanistic basis for the beneficial activities of probiotics when delivered in various foods. This presentation will examine the genomic content of probiotic Lactobacillus cultures, compared to those lactic acid bacterial genomes that have appeared recently. In addition, expression profiling by whole genome microarrays will be used to illustrate how environmental conditions encountered during biomanufacturing, fermentation, and the gastrointestinal tract can impact gene expression and culture functionality.}, number={2}, journal={Bioscience and Microflora}, publisher={BMFH Press}, author={Klaenhammer, Todd R. and Peril, Andrea Azcarate and Barrangou, Rodolphe and Duong, Tri and Altermann, Eric}, year={2005}, pages={31–33} }
 @misc{klaenhammer_barrangou_buck_azcarate-peril_altermann_2005, title={Genomic features of lactic acid bacteria effecting bioprocessing and health}, volume={29}, ISSN={["1574-6976"]}, DOI={10.1016/j.femsre.2005.04.007}, abstractNote={The lactic acid bacteria are a functionally related group of organisms known primarily for their bioprocessing roles in food and beverages. More recently, selected members of the lactic acid bacteria have been implicated in a number of probiotic roles that impact general health and well-being. Genomic analyses of multiple members of the lactic acid bacteria, at the genus, species, and strain level, have now elucidated many genetic features that direct their fermentative and probiotic roles. This information is providing an important platform for understanding core mechanisms that control and regulate bacterial growth, survival, signaling, and fermentative processes and, in some cases, potentially underlying probiotic activities within complex microbial and host ecosystems.}, number={3}, journal={FEMS MICROBIOLOGY REVIEWS}, publisher={Wiley-Blackwell}, author={Klaenhammer, TR and Barrangou, R and Buck, BL and Azcarate-Peril, MA and Altermann, E}, year={2005}, month={Aug}, pages={393–409} }
 @article{bruno-barcena_azcarate-peril_klaenhammer_hassan_2005, title={Marker-free chromosomal integration of the manganese superoxide dismutase gene (sodA) from Streptococcus thermophilus into Lactobacillus gasseri}, volume={246}, ISSN={["1574-6968"]}, url={http://europepmc.org/abstract/med/15869967}, DOI={10.1016/j.femsle.2005.03.044}, abstractNote={A strategy for functional gene replacement in the chromosome of Lactobacillus gasseri is described. The phospho-beta-galactosidase II gene (lacII) was functionally replaced by the manganese superoxide dismutase (MnSOD) gene (sodA) from Streptococcus thermophilus, by adapting the insertional inactivation method described for lactobacilli [Russell, W.M. and Klaenhammer, T.R. 2001 Efficient system for directed integration into the Lactobacillus acidophilus and Lactobacillus gasseri chromosomes via homologous recombination. Appl. Environ. Microbiol. 67, 4361-4364]. L. gasseri carrying the heterologous sodA gene grew on lactose as efficiently as the wild-type parent. An active MnSOD was expressed in the transgenic strain, and the enzyme migrated on PAGE-SOD activity gels to the same position as that of MnSOD from S. thermophilus. The expression of MnSOD from a single copy of sodA integrated in the chromosome of L. gasseri provided enhanced tolerance to hydrogen peroxide, and extended the viability of carbon/energy starved cultures stored at 25 degrees C. This is the first report showing the successful utilization of the pORI plasmids system to generate marker-free gene integration in L. gasseri strains.}, number={1}, journal={FEMS MICROBIOLOGY LETTERS}, author={Bruno-Barcena, JM and Azcarate-Peril, MA and Klaenhammer, TR and Hassan, HM}, year={2005}, month={May}, pages={91–101} }
 @article{azcarate-peril_altermann_hoover-fitzula_cano_klaenhammer_2004, title={Identification and inactivation of genetic loci involved with Lactobacillus acidophilus acid tolerance}, volume={70}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.70.9.5315-5322.2004}, abstractNote={ABSTRACTAmino acid decarboxylation-antiporter reactions are one of the most important systems for maintaining intracellular pH between physiological limits under acid stress. We analyzed theLactobacillus acidophilusNCFM complete genome sequence and selected four open reading frames with similarities to genes involved with decarboxylation reactions involved in acid tolerance in several microorganisms. Putative genes encoding an ornithine decarboxylase, an amino acid permease, a glutamate γ-aminobutyrate antiporter, and a transcriptional regulator were disrupted by insertional inactivation. The ability ofL. acidophilusto survive low-pH conditions, such as those encountered in the stomach or fermented dairy foods, was investigated and compared to the abilities of early- and late-stationary-phase cells of the mutants by challenging them with a variety of acidic conditions. All of the integrants were more sensitive to low pH than the parental strain. Interestingly, each integrant also exhibited an adaptive acid response during logarithmic growth, indicating that multiple mechanisms are present and orchestrated inL. acidophilusin response to acid challenge.}, number={9}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Azcarate-Peril, MA and Altermann, E and Hoover-Fitzula, RL and Cano, RJ and Klaenhammer, TR}, year={2004}, month={Sep}, pages={5315–5322} }