Marcia Su | Penn State University (original) (raw)

Papers by Marcia Su

Food Microbiology

This study aimed to determine whether glutamine deamidation improves acid resistance of Lactobaci... more This study aimed to determine whether glutamine deamidation improves acid resistance of Lactobacillus reuteri, and to assess whether arginine, glutamine, and glutamate-mediated acid resistance are redundant or complementary mechanisms of acid resistance. Three putative glutaminase genes, gls1, gls2, and gls3, were identified in L. reuteri 100-23. All three genes were expressed during growth in mMRS and wheat sourdough. L. reuteri consistently over-expressed gls3 and the glutamate decarboxylase gadB. L. reuteri 100-23ΔgadB over-expressed gls3 and the arginine deiminase gene adi. Analysis of the survival of L. reuteri in acidic conditions revealed that arginine conversion is effective at pH of 3.5 while glutamine or glutamate conversion were effective at pH of 2.5. Arginine conversion increased the pHin but not ΔΨ; glutamate decarboxylation had only a minor effect on the pHin but increased the ΔΨ. This study demonstrates that glutamine deamidation increases the acid resistance of L. r...

Genome research, Jan 30, 2015

Short Tandem Repeats (STRs) are implicated in dozens of human genetic diseases and contribute sig... more Short Tandem Repeats (STRs) are implicated in dozens of human genetic diseases and contribute significantly to genome variation and instability. Yet profiling STRs from short-read sequencing data is challenging because of their high sequencing error rates. Here we developed STR-FM, Short Tandem Repeat profiling using Flank-based Mapping, a computational pipeline that can detect the full spectrum of STR alleles from short-read data, can adapt to emerging read-mapping algorithms, and can be applied to heterogeneous genetic samples (e.g., tumors, viruses, and genomes of organelles). We used STR-FM to study STR error rates and patterns in publicly available human, and in-house generated ultra-deep plasmid, sequencing datasets. We discovered that STRs sequenced with a PCR-free protocol have up to 9-fold fewer errors than those sequenced with a PCR-containing protocol. We constructed an error correction model for genotyping STRs that can distinguish heterozygous alleles containing STRs wi...

BioTechniques, 2014

Polymorphism discovery is a routine application of next-generation sequencing technology where mu... more Polymorphism discovery is a routine application of next-generation sequencing technology where multiple samples are sent to a service provider for library preparation, subsequent sequencing, and bioinformatic analyses. The decreasing cost and advances in multiplexing approaches have made it possible to analyze hundreds of samples at a reasonable cost. However, because of the manual steps involved in the initial processing of samples and handling of sequencing equipment, cross-contamination remains a significant challenge. It is especially problematic in cases where polymorphism frequencies do not adhere to diploid expectation, for example, heterogeneous tumor samples, organellar genomes, as well as during bacterial and viral sequencing. In these instances, low levels of contamination may be readily mistaken for polymorphisms, leading to false results. Here we describe practical steps designed to reliably detect contamination and uncover its origin, and also provide new, Galaxy-based...

Microbiology, 2014

This study characterized the two-component regulatory systems encoded by bfrKRT and cemAKR, and a... more This study characterized the two-component regulatory systems encoded by bfrKRT and cemAKR, and assessed their influence on biofilm formation by Lactobacillus reuteri 100-23. A method for deletion of multiple genes was employed to disrupt the genetic loci of two-component systems. The operons bfrKRT and cemAKR showed complementary organization. Genes bfrKRT encode a histidine kinase, a response regulator and an ATP-binding cassette-type transporter with a bacteriocin-processing peptidase domain, respectively. Genes cemAKR code for a signal peptide, a histidine kinase and a response regulator, respectively. Deletion of single or multiple genes in the operons bfrKRT and cemAKR did not affect cell morphology, growth or the sensitivity to various stressors. However, gene disruption affected biofilm formation; this effect was dependent on the carbon source. Deletion of bfrK or cemA increased sucrose-dependent biofilm formation in vitro. Glucose-dependent biofilm formation was particularly increased by deletion of cemK. The expression of cemK and cemR was altered by deletion of bfrK, indicating cross-talk between these two regulatory systems. These results may contribute to our understanding of the genetic factors related to the biofilm formation and competitiveness of L. reuteri in intestinal ecosystems.

Microbial Cell Factories, 2011

Background: Acid stress impacts the persistence of lactobacilli in industrial sourdough fermentat... more Background: Acid stress impacts the persistence of lactobacilli in industrial sourdough fermentations, and in intestinal ecosystems. However, the contribution of glutamate to acid resistance in lactobacilli has not been demonstrated experimentally, and evidence for the contribution of acid resistance to the competitiveness of lactobacilli in sourdough is lacking. It was therefore the aim of this study to investigate the ecological role of glutamate decarboxylase in L. reuteri. Results: A gene coding for a putative glutamate decarboxylase, gadB, was identified in the genome of L. reuteri 100-23. Different from the organization of genetic loci coding for glutamate decarboxylase in other lactic acid bacteria, gadB was located adjacent to a putative glutaminase gene, gls3. An isogenic deletion mutant, L. reuteri ΔgadB, was generated by a double crossover method. L. reuteri 100-23 but not L. reuteri ΔgadB converted glutamate to γ-aminobutyrate (GABA) in phosphate butter (pH 2.5). In sourdough, both strains converted glutamine to glutamate but only L. reuteri 100-23 accumulated GABA. Glutamate addition to phosphate buffer, pH 2.5, improved survival of L. reuteri 100-23 100-fold. However, survival of L. reuteri ΔgadB remained essentially unchanged. The disruption of gadB did not affect growth of L. reuteri in mMRS or in sourdough. However, the wild type strain L. reuteri 100-23 displaced L. reuteri ΔgadB after 5 cycles of fermentation in back-slopped sourdough fermentations. Conclusions: The conversion of glutamate to GABA by L. reuteri 100-23 contributes to acid resistance and to competitiveness in industrial sourdough fermentations. The organization of the gene cluster for glutamate conversion, and the availability of amino acids in cereals imply that glutamine rather than glutamate functions as the substrate for GABA formation. The exceptional coupling of glutamine deamidation to glutamate decarboxylation in L.Â reuteri likely reflects adaptation to cereal substrates.

Food Microbiology, 2014

Food Microbiology, 2013

Lactobacillus reuteri harbours alternative enzymes for sucrose metabolism, sucrose phosphorylase,... more Lactobacillus reuteri harbours alternative enzymes for sucrose metabolism, sucrose phosphorylase, fructansucrases, and glucansucrases. Sucrose phosphorylase and fructansucrases additionally contribute to raffinose metabolism. Glucansucrases and fructansucrases produce exopolysaccharides as alternative to sucrose hydrolysis. L. reuteri LTH5448 expresses a levansucrase (ftfA) and sucrose phosphorylase (scrP), both are inducible by sucrose. This study determined the contribution of scrP to sucrose and raffinose metabolism in L. reuteri LTH5448, and elucidated the role of scrR in regulation sucrose metabolism. Disruption of scrP and scrR was achieved by double crossover mutagenesis. L. reuteri LTH5448, LTH5448DscrP and LTH5448DscrR were characterized with respect to growth and metabolite formation with glucose, sucrose, or raffinose as sole carbon source. Inactivation of scrR led to constitutive transcription of scrP and ftfA, demonstrating that scrR is negative regulator. L. reuteri LTH5448 and the LTH5448DscrP or LTH5448DscrR mutant strains did not differ with respect to glucose, sucrose or raffinose utilization. However, L. reuteri LTH5448DscrP produced more levan, indicating that the lack of sucrose phosphorylase is compensated by an increased metabolic flux through levansucrase. In conclusion, the presence of alternate pathways for sucrose and raffinose metabolism and their regulation indicate that these substrates, which are abundant in plants, are preferred carbohydrate sources for L. reuteri.

Applied and Environmental Microbiology, 2012

Lactobacillus reuteri is both a gut symbiont and a stable member of sourdough microbiota. This st... more Lactobacillus reuteri is both a gut symbiont and a stable member of sourdough microbiota. This study employed multilocus sequence analysis and an analysis of host-specific physiological and genetic traits to assign five sourdough isolates to rodent-or human-specific lineages. Comparative genome hybridization revealed that the model sourdough isolate LTH2584 had a genome content very similar to that of the model rodent isolate 100-23. These results demonstrate that sourdough isolates of L. reuteri are of intestinal origin.

The manifestation of mitochondrial DNA (mtDNA) diseases depends on the frequency of heteroplasmy ... more The manifestation of mitochondrial DNA (mtDNA) diseases depends on the frequency of heteroplasmy (the presence of several alleles in an individual), yet its transmission across generations cannot be readily predicted owing to a lack of data on the size of the mtDNA bottleneck during oogenesis. For deleterious heteroplasmies, a severe bottleneck may abruptly transform a benign (low) frequency in a mother into a disease-causing (high) frequency in her child. Here we present a high-resolution study of heteroplasmy transmission conducted on blood and buccal mtDNA of 39 healthy
mother–child pairs of European ancestry (a total of 156 samples, each sequenced at ∼20,000× per site). On average, each individual carried one heteroplasmy, and one in eight individuals carried a disease-associated heteroplasmy, with minor allele frequency ≥1%. We observed frequent drastic heteroplasmy frequency shifts between generations and estimated the effective size of the germline mtDNA bottleneck at only ∼30–35 (interquartile range from 9 to 141). Accounting for heteroplasmies, we estimated the mtDNA germ-line mutation rate at 1.3 × 10−8 (interquartile range from 4.2 ×
10−9 to 4.1 × 10−8)mutations per site per year, an order of magnitude higher than for nuclear DNA. Notably,we found a positive association between the number of heteroplasmies in a child andmaternal age at fertilization, likely attributable to oocyte aging. This study also took advantage of droplet digital PCR (ddPCR) to validate heteroplasmies and confirm a de novomutation.Our results can be used to predict the
transmission of disease-causing mtDNA variants and illuminate evolutionary dynamics of the mitochondrial genome.

Microbiology-sgm, 2008

Escherichia coli O157:H7 tightly associates with host cells through the formation of a pedestal s... more Escherichia coli O157:H7 tightly associates with host cells through the formation of a pedestal structure in which cell cytoskeleton rearrangement has been observed. These pathogenic properties have been attributed to an island, known as the locus of enterocyte effacement (LEE), located on the bacterial chromosome. Gene l0017 is one of the LEE genes that has been less well characterized. To understand further the function of the gene, an l0017-deleted mutant was created. The mutant lost type III protein secretion (TTS) capacity. In terms of intracellular components, there was a substantial decrease in the level of EspA, but no apparent effect on Tir and EspB was observed. Fractionation of the bacterial proteins indicated that L0017 was part of the inner-membrane fraction. This association with the membrane is consistent with the hypothesis that L0017 may act as one of the TTS components. In addition, L0017 was found to affect regulation of EspA at a post-transcriptional level. The presence of L0017 readily stabilized EspA and the interaction between L0017 and EspA was demonstrated by their co-purification as well as by a bacterial two-hybrid system. Therefore, L0017 is a chaperone, the second chaperone identified in this system after CesAB, and escorts EspA, a protein with a great tendency to polymerize.