persica 16-FTT0376a, 17-FTT0523a, 20-ISFtu2b and 28-pdpDb.  Amplifies only F. tularensis (only when including the probe). 16-FTT0376a and 17-FTT0523a  Amplifies F. tularensis subsp. mediasiatica, F. tularensis subsp.holarctica and 6/7 F. tularensis subsp. novicida. 28-pdpDb  Amplifies isolates from all clade 1 species as well as W. persica. 20-ISFtu2b Marker with missing sequences as well as mismatches in almost all subspecies represented. 21-ISFtu2a Navitoclax manufacturer Successful amplification was defined as having a primer score below two in both the forward

and reverse primers. a Have associated TaqMan probe which is not considered here. bDetection by variable-length amplicon which is not considered here. cScore of F.noatunensis subsp orientalis <2. Evaluation of sample-sequencing approaches for phylogenetic analyses In the phylogenetic comparison analysis, we focused not only on the entire Francisella genus, but also selleck chemicals llc analysed clades 1 and 2 separately. These sub-populations exhibit different lifestyles and environmental niches and are therefore of interest to different scientific fields [3, 7, 18]. The differences between the poorest and best resolved single marker topologies of the entire genus compared to the whole-genome reference topology (Figure 2) are highlighted in Figure 3A-C. All topologies are shown in Additional File 2. The parameter estimates of the phylogenetic

analysis are summarised in Additional File 3. In general for the analysis of the entire genus, the optimal substitution model was parameter rich, i.e. typically the generalised Thiamine-diphosphate kinase time-reversible (GTR) [31] or Hasegawa-Kishino-Yano (HKY85) [32] models with either invariant sites parameter (α) or rate heterogeneity over sites (Г). Moderate or even low parameter-rich substitution models were favoured in the separate clade analyses, in particular for clade 1, where Jukes-Cantor (JC) [33] or HKY85 models were found to be the optimal choice without α or Г. For clade 2, it was important to include the proportion of invariant sites parameter in the analyses, because of detected recombination events [3].

Figure 2 Whole-genome SNP phylogeny. The whole-genome phylogeny for 37 Francisella strains obtained with model averaging Alpelisib mouse implemented in jModelTest using PhyML software. The removed part of the branches connecting clade 1 and 2 covers a genetic distance of 0.03. Figure 3 Single-marker phylogenies. Single-marker phylogeny of the Francisella genus: (A) highest ranked marker 08-fabH, (B) lowest ranked marker 33-rpoB, and (C) whole-genome phylogeny. Rank is based on difference in resolution between alternative and whole-genome topology. Throughout the study, to facilitate the phylogeny comparisons, we made use of two metrics: degree of incongruence (inc) and difference in resolution (res). The two topologies compared were the reference topology, obtained from whole genome data, and the single-sequence or the concatenated marker sequences topology.

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“Erratum to: Infect Dis Ther (2013) 2:27–36 DOI 10.1007/s40121-013-0006-6 The editors of Infectious Diseases and Therapy would like to make the following addition to the Acknowledgments section of the above-mentioned paper. This required wording was unintentionally missed off the original version of the manuscript. “Compliance with Ethics Guidelines: All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2000 and 2008. Informed consent was obtained from all patients for being included in the study.

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“Introduction

In Sweden, the maternal age in both primi- and multipara mothers has steadily increased during the last three decades. In this period, the mean age of mothers giving birth, both primi- and multipara included, increased from 26.0 to 30.3 years of age. For primiparous women only, the age has increased from 23.8 to 28.4 years of age during the same period. In urban areas in Sweden, the age of mothers giving birth to their first born increased even more, from 24.8 years in 1973 to 30.1 years in 2005 [1]. It has been previously reported that advancing maternal age increases the risk of fetal death [2, 3], but also of other morbidities in the offspring, such as chromosome abnormalities and childhood cancers like leukemia and retinoblastoma [4, 5]. The maternal age has also been associated with the development of diabetes mellitus type 1 and schizophrenia in the offspring, but these associations were also found to be dependent on paternal age [6, 7].

J Microbiol Meth 2000, 2:175–179.CrossRef 48. Henriques M, Azeredo J, Oliveira R: Candida albicans and Candida dubliniensis : comparison of biofilm formation in terms of biomass and activity. Brit J Biomed Scien 2006, 63:5–11. 49. Silva S, Henriques M, Martins A, Oliveira R, Williams D, Azeredo J: Biofilms of non- Candida albicans Candida species: quantification, structure and matrix composition. Med Mycol 2009, selleck chemicals 20:1–9.CrossRef 50. Hiller E, Heine S, Brunner H, Rupp S: Candida albicans Sun41p, a putative glycosidase, is involved in morphogenesis, cell wall biogenesis, and biofilm formation. Eukaryot Cell 2007, 6:2056–2065.GDC-0068 concentration PubMedCrossRef 51. Nobile CJ, Mitchell AP: Genetics and genomics of Candida albicans

biofilm formation. Cell Microbiol 2006, 8:1382–1391.PubMedCrossRef 52. Selmecki A, Bergmann S, Berman J: Comparative genome hybridization reveals widespread aneuploidy in Candida albicans laboratory strains. Mol Microbiol 2005, 55:1553–1565.PubMedCrossRef 53. Brand A, MacCallum DM, Brown AJP, Gow NA, Odds FC: Ectopic expression of URA3 can infuence the virulence phenotypes and proteome of Candida albicans but can be overcome by targeted reintegration of URA3 at the RPS10 locus. Eukaryot Cell 2004, 3:900–909.PubMedCrossRef 54. Oelkers P, Tinkelenberg A, Erdeniz N, Cromley D, Billheimer J, Sturley S: A lecithin cholesterol acyltransferase-like gene mediates diacylglycerol esterification in yeast. J

Biol Chem 2000, 275:15609–15612.PubMedCrossRef 55. Silva L, Coutinho A, Fedorov A, Prieto M: Nystatin-induced lipid vesicles permeabilization see more is strongly dependent on sterol structure. Biochim Biophys Acta 2006, 1758:452–459.PubMedCrossRef 56. Klis FM, Selleck Staurosporine de Groot P, Hellingwerf

K: Molecular organization of the cell wall of Candida albicans . Med Mycol 2001, 39:1–8.PubMed 57. Klis FM, Mol P, Hellingwerf K, Brul S: Dynamics of cell wall structure in Saccharomyces cerevisiae . FEMS Microbiol Rev 2002, 26:239–253.PubMedCrossRef 58. Netea MG, Gow NA, Munro CA, Bates S, Collins C, Ferwerda G, Hobson RP, Bertram G, Hughes HB, Jansen T, Jacobs L, Buurman ET, Gijzen K, Williams DL, Torensma R, McKinnon A, MacCallum DM, Odds FC, van der Meer JW, Brown AJ, Kullberg BJ: Immune sensing of Candida albicans requires cooperative recognition of mannans and glucans by lectin and Toll-like receptors. J Clin Invest 2006, 116:1642–1650.PubMedCrossRef 59. Angiolella L, Micoci MM, D’Alessio S, Girolamo A, Maras B, Cassone A: Identification of major glucan-associated cell wall proteins of C. albicans and their role in fluconazole resistance. Antimicrob Agents Chemother 2002, 1688–1694. 60. Herrero AB, Magnelli P, Mansour MK, Levitz SM, Bussey H, Abeijon C: KRE5 gene null mutant strains of Candida albicans are a virulent and have altered cell wall composition and hyphae formation properties. Eukaryot Cell 2004, 3:1423–1431.PubMedCrossRef 61.

, 2003). **mean of quantification by oprL qPCR tested in duplicate. NA: not applicable. P. aeruginosa BI 2536 manufacturer isolation Ten μl of liquefied sputum pure and diluted into 1/1000, were inoculated and incubated onto several non selective and selective media for P. aeruginosa isolation, including Columbia blood agar supplemented with 5% defribinated horse blood (Oxoid, Dardilly, France), Columbia chocolate agar (Oxoid), and cetrimide agar (Oxoid).

All media were incubated aerobically at 37°C for five days and monitored daily. All different morphotypes of bacterial colonies were identified phenotypically with conventional screening methods (Gram coloration, oxidase test) followed by mass spectrometry identification (MicroFlex LT, Bruker Daltonics, Germany) [33, 34]. Quantification was conducted based on the colony forming unit (CFU) counts and the dilution ratio of the plate. P. aeruginosa detection and quantification by quantitative PCR (qPCR) DNA extraction For each isolate of the bacterial EX527 collection, 1 ml of a 0.5 McFarland suspension was extracted. For each sputum sample, one of the two 1 ml-aliquots was treated by 5 min of sonication using a bath sonicator (Elamsonic

S10, Singen, Germany). After a 10 min-centrifugation (5000 g), the pellet was suspended in 200 μl of DNA free water. Ten μl of the IC2, an internal control provided in the DICO Extra r-gene™ kit (Argène, Verniolle, France), were added in each check details sample and, for each batch of extraction, in 200 μl of DNA free water as a negative control. DNA was extracted using the QIAamp DNA Minikit® (Qiagen, Courtaboeuf, France) according to the instructions of the manufacturer (“Tissue protocol”)

with elution volumes of 100 μl. oprL qPCR oprL qPCR was performed using primers OPRL-F and OPRL-R and hydrolysis probe ASK1 oprL-MGB, previously described by Joly et al. [30] (Table 2). The reaction mix comprised 12.5 μl of Qiagen Quantitect Probe Master Mix, 0.3 μM of each primer, 0.2 μM of hydrolysis probe and 4.5 μl of DNA extract, and was made up to a final reaction volume of 25 μl with water. A negative amplification control was used for each batch. For sputum samples, a standard curve provided a full concentration range of P. aeruginosa extending from 102 to 106 CFU/mL. Each qPCR assay was repeated twice, and the mean value of the quantification was calculated for each duplicate (Table 1). Cycling was performed on an ABI Prism 7300 Real Time PCR System (Applied Biosystem, Foster city, Californy), with an initial hold at 95°C for 15 min, followed by 50 cycles at 95°C for 15 s, and 60°C for 1 min. The oprL-MGB probe was labelled with carboxyfluorescein (FAM).

World J Surg 2004, 28:301–306.CrossRef 7. Wain J, Diep TS, Ho VA, Walsh AM, Hoa NTT, Parry CM: Quantitation of bacteria in blood of typhoid fever patients and relationship between counts and clinical features, transmissibility, and antibiotic resistance. J Clin Microbiol 1998, 36:1683–1687. 8. Stewart PS, Costerton JW: Antibiotic resistance of bacteria in biofilms. Lancet 2001, 358:135–138.CrossRef 9. Hetrick EM, Shin JH, Stasko NA, Johnson CB, Wespe DA, Holmuhamedov E, Schoenfisch MH: Bactericidal efficacy of

nitric oxide-releasing silica nanoparticles. ACS Nano 2008, 2:235–246.CrossRef 10. Diekema BV-6 DJ, Pfaller MA: Rapid detection of antibiotic-resistant organism carriage for infection prevention. Clin Infect Dis 2013, 56:1614–1620.CrossRef 11. Rai M, Yadav A, Gade A: Silver nanoparticles as a new generation of antimicrobials. Biotechnol Adv 2009, 27:76–83.CrossRef 12. Lusby PE, Coombes AL, Wilkinson JM: Bactericidal activity of different GANT61 honeys against pathogenic bacteria. Arch Med Res 2005, 36:464–467.CrossRef 13. Liu X, Wong KKY: Application of Nanomedicine in Wound Healing. New York: Springer; 2013. 14. Berndt S, Wesarg F, Wiegand C, Kralisch D, Müller FA: Antimicrobial porous hybrids consisting of bacterial nanocellulose and silver nanoparticles. Cellulose 2013, 20:771–783.CrossRef 15. Nablo BJ, Rothrock AR, Schoenfisch MH: Nitric oxide-releasing

sol-gels as antibacterial coatings for orthopedic implants. Biomaterials 2005, 26:917–924.CrossRef 16. Li L-L, Wang H: Enzyme-coated mesoporous silica nanoparticles as efficient antibacterial agents in vivo. Adv Healthcare Mater 2013, 2:1351–1360.CrossRef 17. Witte M, Barbul A: Role of nitric oxide in wound repair. Am J Surg 2002, 183:406–412.CrossRef 18. Friedman A, Friedman J: New biomaterials for the sustained release of nitric oxide: past, present and future. Expert Opin Drug Deliv 2009, 6:1113–1122.CrossRef 19. Ghaffari A, Miller

CC, McMullin B, Ghaharya A: Potential application of gaseous nitric oxide as a topical antimicrobial agent. Nitric Oxide 2006, 14:21–29.CrossRef 20. Marxer SM, Rothrock AR, Nablo BJ, Robbins ME, Schoenfisch MH: Preparation of nitric oxide (NO)-releasing sol - gels for BIX 1294 supplier biomaterial applications. Chem Mater 2003, 15:4193–4199.CrossRef 21. Carpenter AW, Slomberg DL, Rao KS, Schoenfisch MH: Influence CYTH4 of scaffold size on bactericidal activity of nitric oxide-releasing silica nanoparticles. ACS Nano 2011, 5:7235–7244.CrossRef 22. Hetrick EM, Shin JH, Paul HS, Schoenfisch MH: Anti-biofilm efficacy of nitric oxide-releasing silica nanoparticles. Biomaterials 2009, 30:2782–2789.CrossRef 23. Friedman AJ, Han G, Navati MS, Chacko M, Gunther L, Alfieri A, Friedman JM: Sustained release nitric oxide releasing nanoparticles: characterization of a novel delivery platform based on nitrite containing hydrogel/glass composites. Nitric Oxide 2008, 19:12–20.CrossRef 24.

Viral protein epitopes

are pivotal in the pathogenesis of virus infection and in the development of effective vaccines [33, 34]. Therefore, the identification of B-cell epitopes for DENV prM antibodies can provide important information for the understanding of the pathogenesis of DENV infection 3-Methyladenine and contribute to the development of dengue vaccine. In the case of DENV, many efforts have been made into mapping the epitopes of E protein [35–39], but only a few epitopes have been identified on prM protein [40, 41]. Consequently, the precise antigenic structures of prM and their functions in the immune response and infection pathogenesis remain poorly selleck chemicals llc studied. In the present study, the epitope recognized by prM mAb 4D10 was identified using a phage-displayed peptide library and comprehensive bioinformatic analysis. We investigated

the neutralizing versus enhancing capacity of the mAb 4D10 and antisera of epitope peptide PL10 towards Entinostat price standard DENV1-4 particles and imDENV particles. We found that 4D10 and antibody against epitope peptide PL10 showed broad cross-reactivity and poor neutralizing acvitity with the four standard DENV serotypes but significantly enhanced the infectious properties. In addition, these antibodies remained susceptible to partially neutralizing imDENV and indeed rendered virtually non-infectious imDENV highly infectious in Fc receptor-bearing cells. Taken together, we identified a novel infection-enhancing epitope on prM protein. These results may provide some important implications for a better understanding of the pathogenesis of DENV infection and advance the development of dengue vaccine. Methods Cells C6/36 cells derived from Aedes albopictus were maintained PAK6 in Modified Essential Medium (GIBCO) supplemented with 10%

fetal bovine serum (FBS) at 28°C, 5%CO2. Baby Hamster Kidney-21 (BHK-21) cells derived from the kidney of Mesocricetus auratus and Human adenocarcinoma LoVo cells derived from left supraclavicular region metastasis were cultured in Dulbecco’s Modified Eagle’s Medium (GIBCO) supplemented with 10% FBS at 37°C, 5% CO2. Human erythroleukemic K562 cells derived from bone marrow were maintained in Iscove’s Modified Dulbecco’s Medium (GIBCO) supplemented with 10% FBS at 37°C, 5% CO2. The media were supplemented with 2 mM L-glutamine, 10mM HEPES, penicillin (100 U/ml) and streptomycin (100 U/ml). All cells were purchased from ATCC. Viruses DENV1 strain Hawaii (GenBank: EU848545), DENV2 strain New Guinea C (NGC) (GenBank: AF038403), DENV3 strain H87 (GenBank: M93130), DENV4 strain H241 (GenBank: AY947539) and JEV (GenBank: AF315119) were propagated on C6/36 cells. Briefly, monolayer of C6/36 cells was infected with DENV at multiplicity of infection (MOI) of 1. The virus supernatants were harvested at 72 hours post-infection (hpi), cleared from cellular debris by low-speed centrifugation, purified by PEG 8000 precipitation.

marcescens (~5

μM). To examine if this could be due to the fact that the two bacteria were treated with the same dose despite their very different MIC values, we determined their dose response curves. For both bacteria a minimum chimera dose of 500 μg/mL (i.e. 145-180 μM) was needed to obtain the maximum immediate response (data not shown) ruling out that the rapid release of ATP from S. aureus seen in Figure 3A is due to a higher concentration/MIC ratio than employed for S. marcescens. Figure 3 Chimera-induced ATP leakage in S. aureus (A) and S. marcescens (B) after treatment with 1000 μg/mL chimera. The assays were performed in two independent experiments. Mean (SEM) intracellular (IC, solid line) and extracellular (EC, punctuated line) ATP concentration MX69 molecular weight for S. aureus cells (figure A, grey lines) and S. marcescens cells (figure B, grey lines) treated with chimera 1 compared to MilliQ-treated control (black lines). To investigate if 4SC-202 cost the degree of ATP leakage from the bacterial cell corresponded to the simultaneous decrease in the number of viable cells (i.e. if S. marcescens cells on the basis of their elevated MIC were in fact able to survive even after a moderate ATP leakage) we determined time-kill under exactly the same conditions as the ATP bioluminescence assay had been performed. Irrespective of which of the three chimeras that were used, both bacteria were reduced 2-3 log from an selective HDAC inhibitors initial value of log ~9.5 per mL within the first 20

minutes before the ATP leakage tailored off and no further decrease in viable count was seen for up to 60 minutes (not shown). This indicates that the degree of ATP leakage from the two bacteria (i.e. the concentration of the extracellular ATP) does not reflect differences in viability. No reduction in the number of viable

Baricitinib bacteria was seen for the control (not shown), and the intracellular concentration of ATP did not change (Figure 3A and 3B). Although there was no systematic difference in the MIC values between Gram-positive and -negative bacteria, we speculated that the Gram-negative outer membrane could act as a barrier to the penetration of AMPs, since polymyxin B resistance in S. marcescens has been linked to induced changes in the amount and composition of lipopolysaccharide (LPS) in the outer membrane [33]. Moreover, similar resistance-conferring membrane alterations have also been seen for other bacteria in response to polymyxin B treatment [34–36]. Accordingly, we studied how a membrane-destabilizing pre-treatment of S. marcescens, E. coli and S. aureus with the divalent metal cation-chelating agent EDTA would affect the killing caused by chimera 1. In these experiments we used a non-lethal 0.5 mM concentration of EDTA together with the non-lethal 1.5 μM concentration of the tested AMP analogue. A slight reduction in the number of viable cells corresponding to 0.5 log was seen for S. aureus when treated with chimera 1 alone while E. coli and S. marcescens were reduced with 1.

Conflicts of interest Jean-Yves Reginster on behalf of the Department of Public Health, Epidemiology and Health Economics of the University of Liège, Liège, Belgium. Consulting fees or paid advisory boards: Servier, Novartis, Negma, Lilly, Wyeth, Amgen, GlaxoSmithKline, Selumetinib cost Roche, Merckle, Nycomed, NPS, and Theramex. Lecture fees when speaking at the invitation of

a commercial sponsor: Merck Sharp and Dohme, Lilly, Rottapharm, IBSA, Genevrier, Novartis, Servier, Roche, GlaxoSmithKline, Teijin, Teva, Ebewee Pharma, Zodiac, Analis, Theramex, Nycomed, and Novo-Nordisk. Grant support from industry: Bristol Myers Squibb, Merck Sharp & Dohme, Rottapharm, Teva, Lilly, Novartis, Roche, GlaxoSmithKline, Amgen, and Servier. Jean-Jacques Body has received speakers and CP673451 cell line consultant fees from Amgen and Novartis, and

research support from Merck Sharp & Dohme, Novartis, Procter & Gamble, Servier, and Roche. Yves Boutsen has received speakers and/or consultant fees and/or research support from Procter & Gamble, Eli-Lilly, Daiichi-Sankyo, Merck Sharp & Dohme, Novartis, Servier, and Roche. Jean-Marc Kaufman has received speakers and/or consultant fees and/or research support from Amgen, Daiichi-Sankyo, Glaxo Smith Kline, Meck Sharp & Dohme, Novartis, Nycomed, Servier, and Roche. Stephan Goemaere has received speakers fees and/or research support from Amgen, Eli Lilly, Glaxo Smith Kline, Merck Sharp & Dohme, Novartis, Nycomed, Proctor & Gamble, Sanofi-Aventis, Servier,

and Roche. Steven Boonen has received consulting fees and/or research support from Amgen, Merck, Novartis, Nycomed, Procter & Gamble Pharmaceuticals, and Sanofi-Aventis. Pierre Bergmann has no conflict of interest. Jean-Pierre Devogelaer participated in most of trials with antiosteoporotic drugs. Serge Rozenberg has no conflict of interest. Open Access This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited. References 1. Cummings SR, Black DM, Rubin SM (1989) Lifetime risks of hip, Colles’, or vertebral fracture Bumetanide and coronary heart disease among white postmenopausal women. Arch Intern Med 149:2445–2448PubMedCrossRef 2. Autier P, Haentjens P, Bentin J, Baillon JM, Grivegnee AR, Closon MC, Boonen S (2000) Costs induced by hip fractures: a prospective learn more controlled study in Belgium. Belgian Hip Fracture Study Group Osteoporos Int 11:373–380 3. Cranney A, Tugwell P, Wells G, Guyatt G (2002) Meta-analyses of therapies for postmenopausal osteoporosis. I. Systematic reviews of randomized trials in osteoporosis: introduction and methodology. Endocr Rev 23:496–507PubMedCrossRef 4.

While, PMF may be generated through PPi hydrolysis using a membrane 4-Hydroxytamoxifen datasheet bound proton-translocating pyrophosphatase (PPase), the directionality of this PPase is unknown, and may in fact use PMF for PPi synthesis. PPi is a by-product of various endergonic biosynthetic reactions, including poly-nucleic acid synthesis from (deoxy)nucleotide triphosphates and activation of amino acids, carbohydrates, and fatty acids for protein, polysaccharide, and lipid synthesis [21].

Thus, the effective removal of PPi improves the thermodynamic feasibility of these reactions. Concentrations as low as 2 mM PPi have shown to inhibit growth of some bacteria [94]. In addition to serving as a central energy carrier, PPi serves to regulate key enzymes in carbohydrate metabolism including LDH in Ca. saccharolyticus[21], malic enzyme in C. thermocellum (Taillefer and Sparling, unpublished), ATP-dependent PFK in T. maritima[95], and PTA in C. acidiurici[96]. As mentioned above, PPi can be utilized in the glycolytic direction by (i) PPi-dependent 6-P-fructokinase, (ii) PPDK, and (iii) acetate thiokinase. Alternatively, hydrolysis of PPi via a membrane-bound PPase (Cthe_1425) can be coupled to check details PMF generation that could

be utilized for transport of nutrients, motility, and ATP synthesis. The PPi-dependent enzymes used by C. thermocellum have remarkable similarities to that of parasitic protists (ie. Trichomonas foetus, Entamoeba histolytica; [75]) and other bacteria such as Ca. saccharolyticus[97]. PPi levels in Ca. saccharolyticus have been shown to be elevated (4 ± 2 mM) during exponential phase and lower during transition

to stationary phase [97], consistent with other organisms that do not contain a cystolic PPase (C. thermoaceticum and C. pasteuranum; [98]). Conversely, PPi levels in E. coli, which possesses a cystolic PPase, were low (0.3 mM) and did not fluctuate during growth [98]. We observed a 1.9-fold increase in membrane-bound PPase expression in stationary phase cells. Conclusions A unified understanding of how gene and gene-product expression, stability, and regulation, in conjunction with Selleckchem Alpelisib intracellular metabolic Glutathione peroxidase profiling and thermodynamics of product formation, are key elements for targeted metabolic engineering strategies and fermentation optimization for the economic feasibility of biofuels production via consolidated bioprocessing. Clostridium thermocellum, like many cellulolytic, fermentative, biofuel producing organisms, has multiple enzymes capable of catalyzing parallel reactions and branched product pathways. Measuring peptide spectral counts via shotgun proteomics has been shown to be a valid method for determining relative protein abundance profiles [57–60]. In turn, understanding protein expression profiles may provide genetic engineering strategies targeted at redirecting carbon and electron flux for the optimization of end-product production. Furthermore, responses of protein expression in response to physiological conditions (ie.