6.5 at 20 °C. The alkali tolerance of this strain extends the pH range of highly adaptable Fe(III)-reducing Serratia species from mildly acidic pH values associated with acid mine drainage conditions to alkali conditions representative of subsurface sediments stimulated for extensive denitrification and metal reduction. Dissimilatory Fe(III)-reducing

bacteria are widely distributed in freshwater and marine environments and have the ability to utilize a wide range of compounds as electron donors (Lovley et al., 2004; Weber et al., Selleckchem MK-2206 2006). Dissimilatory Fe(III) reduction has been shown to occur over a wide pH range from acid mine drainage sites to alkaline soda lakes (Johnson, 1995; Straub et al., 2001; Pollock et al., 2007). Although Fe(III) reduction at low (< pH 3) and circumneutral pH is well documented, few studies exist showing Fe(III) reduction above pH 9 (Gorlenko et al., 2004; Pollock et al., 2007), despite the potential significance of these reactions in a range of natural and engineered environments. Alkaline pH is challenging for microbial metabolism as microorganisms must maintain their optimum intracellular pH and possess a mechanism for

creating an electron motive force capable screening assay of driving solutes across the cell membrane against a proton counter gradient (Krulwich et al., 2001; Detkova & Pusheva, 2006; Stewart et al., 2010). It is suggested that in extreme alkaline environments, Na+ may replace H+ to create an electron motive force in some alkaliphilic microorganisms (Kevbrin et al., 1998; Krulwich et al., 2001; Detkova & Pusheva, 2006). Fe(III) reduction at a pH

> 9 has been observed by several species isolated from natural alkaline soda lakes, including Anaerobranca californiensis (Gorlenko et al., 2004), Alkaliphilus metaliredigens (Ye et al., 2004), Tindallia magadii (Kevbrin et al., 1998) and species most similar to (96%) Bacillus agaradhaerens (Pollock et al., 2007). In addition to natural high pH environments, such Adenosine triphosphate as soda lakes, there is interest in the biogeochemistry of engineered high pH sediments, for example those resulting from industrial contamination and the use of alkaline cements as a building material. Alkaline sediment geomicrobiology is of particular current interest to the nuclear industry owing to the proposed use of cement containment for deep geological disposal of radioactive wastes and for remediation scenarios for existing contaminated land (NDA, 2011). It is important to understand how changes in pH may affect the microbial community and therefore the biogeochemical processes occurring in the subsurface. Microbial processes are a key to predicting the mobility of problematic radionuclides in the subsurface (Lloyd, 2003).

18 mg, and the heme content

(mol mol−1 of protein) was estimated to 0.93, based on pyridine hemochrome analysis. The absorption maximum of the reduced-minus-oxidized difference spectrum of the pyridine hemochrome compound was 549.7 nm, supporting the notion of a c-type cytochrome. Figure 2 shows optical spectra of the purified protein in the oxidized and reduced states. The absorption maxima of reduced protein are 551 and 416 nm in the alpha and Soret bands, respectively, and 410 nm in the Soret band of the oxidized protein. To estimate the redox potential, optical spectra in the visible region were recorded from protein diluted into redox Lapatinib cell line buffer containing potassium hexacyanoferrate (II) and potassium hexacyanoferrate (III) in different proportions. Inset (b) in Fig. 2 shows the extent of reduction as a function of the redox potential of the buffer. A midpoint potential of 261 mV was obtained by curve fitting. The thermodynamics of chlorate reduction by the cytochrome depends on the difference between this potential and the potential of the chlorate/chlorite redox couple at pH=7. An estimate for the latter can be obtained from data given by Thompson (1986). The standard potential (pH=0) of the ClO3−/HClO2 redox couple is given as +1.16 V. From this, and a pKa value of 2 for the HClO2, a value of +0.708 V is obtained for the midpoint potential of the chlorate/chlorite couple at pH=7. This is considerably

higher than the potential found for Antiinfection Compound Library clinical trial the cytochrome, with the consequence that electron transfer from the cytochrome to chlorate is a thermodynamically favorable reaction. In a previous paper (Bäcklund et al., 2009), the chlorate-dependent reoxidation of reduced cytochrome c in periplasmic extract was demonstrated. In order to further investigate the reaction between the purified 9-kDa cytochrome

c-Id1 and chlorate reductase, the chlorate-dependent oxidation of the reduced cytochrome in the presence of purified chlorate reductase was studied. Fossariinae Figure 3 shows spectra obtained in the visible region up to 12 min after the addition of chlorate. The time course of the reaction was obtained by plotting the A552 nm as a function of time and is shown in the inset. The solid line in the inset shows the fit of a single exponential function to the time course, demonstrating that the reaction is first-order. Similar first-order kinetics were observed at all concentrations of cytochrome c investigated. The effect of the concentration of cytochrome c-Id1 on the initial rates obtained from the curve fits are shown in Fig. 4. In the concentration range investigated, initial rates appear to increase linearly with the substrate concentration, indicating a KM value substantially higher than the highest substrate concentration investigated (4 μM) under present conditions. Using the estimated concentration of chlorate reductase, a kcat/KM of 7 × 102 M−1 s−1 was calculated from the slope of the line in Fig. 4.

67 €/km, which was less expensive than PLX4032 regular air ambulance in the present study (7.49 €/km).6 The average cost per case was 12.992 €±11.445 € (1,458–114.078 €). A stretcher in a scheduled aircraft was significantly cheaper than in an air ambulance (p

< 0.0001). The AE is an established form of transportation for patients who fall ill abroad. An improvement in the epidemiological data on repatriation cases is desirable, as it is likely to improve the logistic, medical, and economic aspects of the planning process. Currently, epidemiological data on this form of air transportation are sparse, which is why this study was undertaken. In concordance with Chawla and colleagues, who investigated 100 stretcher cases in India, we found that trauma cases (femoral neck fracture) and stroke, together with myocardial infarction, were the most common diagnoses in our group of 504 aeromedical repatriation cases.7 A variety of private companies, aid agencies, and airlines have

specialized in aeromedical transportation. For example, Lufthansa, the largest German airline, has developed the PTC, which is an independent, fully enclosed intensive care module that is placed inside the cabin of a commercial aircraft. It can be installed in wide-bodied aircraft like the B 747-400, the A330-300, and the A 340-300/600 during routine time spent on the ground in Frankfurt am Main Airport, Germany. One of its advantages is the floor space inside the module,

which measures 6 m2 and offers normal standing height. RXDX-106 cost A flight attendant with training as an intensive care nurse or paramedic accompanies every PTC transport. Whether it has operative or medical advantages compared to other forms of air transportation cannot be evaluated due to the small number of cases in this study (n = 3; 0.6%). Metalloexopeptidase The costs of PTC transport were also not evaluated in the present study because of the small number of cases. Instead, we compared the previously published PTC transport costs (€/km) with the data in the present study. Compared to scheduled aircraft, which have an economic advantage, the benefit of an air ambulance is its high degree of availability and flexibility regarding patient transportation. Patients can be picked up at small airports that are often closer to the hospital of origin than larger airports operated by scheduled airlines. However, in cases of long-distance flights, their fueling stops can be more frequent compared to scheduled aircraft because air ambulances have a shorter range. Both PTC and air ambulances are capable of providing medical monitoring and treatment at the ICU level. Given the economic restraints on insurance companies and health-care systems, the economic aspects of AE need to be critically evaluated.

3a). Because gingipain activity can be regulated at the transcriptional and post-transcriptional levels (Tokuda et al., 1998), oligonucleotide primers, as described previously

(Vanterpool et al., 2005a), were used in RT-PCR analysis to determine whether these two sigma factors were involved in the transcriptional regulation of gingipain-encoding click here genes. As shown in Fig. 3b, the inactivation of PG0162 and PG1660 had no effect on the expression of rgpA, rgpB, or kgp at the transcriptional level. In FLL355 (PG1827∷ermF), the Kgp activity showed a 25% increase over the wild type. No change was observed in the transcription of the kgp gene in FLL355 (data not shown). Taken together, these results suggest that ECF sigma factors may be involved in the post-transcriptional regulation of gingipains. Post-transcriptional regulation of the gingipains in P. gingivalis is associated with its maturation pathway, which is linked to the biosynthesis AZD8055 of surface carbohydrates (Shoji et al., 2002; Paramonov et al., 2005) and several other proteins including the PorR (Shoji et al., 2002), PorT (Sato et al., 2005; Nguyen et al., 2009),

Sov (Saiki & Konishi, 2010), and VimA (Vanterpool et al., 2006). It is unclear how these factors are modulated by the ECF sigma factors and is an active area of further exploration in the laboratory. The correlation between gingipain activity and hemagglutination in P. gingivalis (Lewis et al., 1999; Shi et al., 1999; Vanterpool et al., 2005a) is related to the similar adhesion domains encoded by the hagA, rgpA, and kgp genes (Chen & Duncan, 2004). The hemagglutination potential of ECF sigma factor-defective mutants was assessed. In comparison with the wild-type strain, there was a decrease Avelestat (AZD9668) in the hemagglutination activity in all the mutants. In FLL350, the level of hemagglutination activity was comparable

to the negative control. This is in contrast to FLL354, which showed the greatest reduction in gingipain activity, but a higher hemagglutination activity. RT-PCR using hagA-specific primers indicated no change in the expression of that gene in FLL350 (Fig. 4c). While gingipains have been observed to have hemolytic activity (Shah & Gharbia, 1989; Lewis et al., 1999), hemolysin can be independent of their catalytic association (Deshpande & Khan, 1999). Several putative hemolysin genes have been identified in the P. gingivalis genome (Nelson et al., 2003) and cloned in E. coli (Karunakaran & Holt, 1993). The hemolysins produced by P. gingivalis provide the bacterium with heme-containing molecules that are required for their in vivo survival. Hemolytic activities of all the ECF-defective mutants in this study were similar to those of the wild type, except for FLL350 (Fig. 4d). The FLL350 mutant showed a 50% reduction in those activities compared with the parent strain.

3a). Because gingipain activity can be regulated at the transcriptional and post-transcriptional levels (Tokuda et al., 1998), oligonucleotide primers, as described previously

(Vanterpool et al., 2005a), were used in RT-PCR analysis to determine whether these two sigma factors were involved in the transcriptional regulation of gingipain-encoding Ferrostatin-1 supplier genes. As shown in Fig. 3b, the inactivation of PG0162 and PG1660 had no effect on the expression of rgpA, rgpB, or kgp at the transcriptional level. In FLL355 (PG1827∷ermF), the Kgp activity showed a 25% increase over the wild type. No change was observed in the transcription of the kgp gene in FLL355 (data not shown). Taken together, these results suggest that ECF sigma factors may be involved in the post-transcriptional regulation of gingipains. Post-transcriptional regulation of the gingipains in P. gingivalis is associated with its maturation pathway, which is linked to the biosynthesis Daporinad molecular weight of surface carbohydrates (Shoji et al., 2002; Paramonov et al., 2005) and several other proteins including the PorR (Shoji et al., 2002), PorT (Sato et al., 2005; Nguyen et al., 2009),

Sov (Saiki & Konishi, 2010), and VimA (Vanterpool et al., 2006). It is unclear how these factors are modulated by the ECF sigma factors and is an active area of further exploration in the laboratory. The correlation between gingipain activity and hemagglutination in P. gingivalis (Lewis et al., 1999; Shi et al., 1999; Vanterpool et al., 2005a) is related to the similar adhesion domains encoded by the hagA, rgpA, and kgp genes (Chen & Duncan, 2004). The hemagglutination potential of ECF sigma factor-defective mutants was assessed. In comparison with the wild-type strain, there was a decrease Benzatropine in the hemagglutination activity in all the mutants. In FLL350, the level of hemagglutination activity was comparable

to the negative control. This is in contrast to FLL354, which showed the greatest reduction in gingipain activity, but a higher hemagglutination activity. RT-PCR using hagA-specific primers indicated no change in the expression of that gene in FLL350 (Fig. 4c). While gingipains have been observed to have hemolytic activity (Shah & Gharbia, 1989; Lewis et al., 1999), hemolysin can be independent of their catalytic association (Deshpande & Khan, 1999). Several putative hemolysin genes have been identified in the P. gingivalis genome (Nelson et al., 2003) and cloned in E. coli (Karunakaran & Holt, 1993). The hemolysins produced by P. gingivalis provide the bacterium with heme-containing molecules that are required for their in vivo survival. Hemolytic activities of all the ECF-defective mutants in this study were similar to those of the wild type, except for FLL350 (Fig. 4d). The FLL350 mutant showed a 50% reduction in those activities compared with the parent strain.

, 1999). BIME-1 and BIME-2 correspond to SMAG TT and HH dimers. However, HH dimers are about 10 times more abundant than TT dimers. In contrast, BIME-1 (74 repeats) are three times more abundant than BIME-2 (24 repeats).

Moreover, both BIME-1 and BIME-2 are invariably comprised of elements from different subfamilies (Bachellier et al., 1999; see also http://www.pasteur.fr/recherche/unites/pmtg/repet/index.html). The predominance buy PLX3397 of TT over HH dimers, and the composite nature of dimers, is also a distinctive feature of the abundant REP families found in Pseudomonas putida (Aranda-Olmedo et al., 2002) and P. syringae (Feil et al., 2005). It has been hypothesized that REPs are mobilized by a VX 809 transposase of the IS200/IS605 family, and the corresponding genes have been shown to be flanked by REPs in many species (Nunvar et al., 2010). Four genes encoding this transposase were identified in K279a DNA (ORFs 1101,

1152, 2816 and 4509), but only ORFs 1101 and 2816 are flanked by SMAGs. We believe that REPs are an ancient component of the genomes of Proteobacteria, which have been actively mobilized by transposition only early in their history. According to this view, REPs disappeared in time from most species, their dissemination being plausibly detrimental to the cell, and have been maintained only in species in which they could no longer transpose. This hypothesis is supported by the observation that SMAG sequences were found in none of the 41 species-specific GEIs, plausibly acquired by lateral gene transfer, which account for >10% of the K279a chromosome (Rocco et al., 2009). REPs are similarly restricted to core genome regions in P. syringae (Tobes & Pareja, 2005). In contrast to what was observed for REPs in other species (Tobes & Pareja, 2006), SMAGs are not targeted by mobile DNA. However, it is worth noting that a K279a GEI encoding type 1 pili (Rocco et al., 2009) is flanked by SMAG-2 dimers. FER About 1/7 of the ORFs of the K279a strain are flanked by SMAGs in a distance range that makes the presence of promoter or terminator

sequences unlikely. It is plausible that most of these elements are transcribed into mRNA, and that their folding into RNA hairpins may influence the level of expression of flanking genes. The number of genes potentially controlled at the post-transcriptional level by SMAGs may be higher than estimated, because many repeats are inserted either upstream (17 elements) or downstream (150 elements) or within (44 elements) known or putative operons. We analyzed genes transcribed in the same direction intermingled with SMAG sequences, and found that the repeats influence the segmental mRNA stability. Both monomers and dimers function as stabilizers of upstream transcripts, and work with comparable efficiency when embedded in the same RNA context (Fig. 5).

It is important to note that not all HIV pregnancies are reported to the APR, as reporting is voluntary. A web and literature search reveals two case reports of myelomeningocoele associated with first-trimester efavirenz exposure [[7],[8]]. Data from the IeDEA West Africa and ANRS Databases, Abidjan, Cote d’Ivoire, found no significant Kinase Inhibitor Library increased risk of unfavourable pregnancy outcome in women with first trimester exposure to efavirenz

(n = 213) compared with nevirapine (n = 131) apart from termination, which was more common with efavirenz [9]. In 2010, a systematic review and meta-analysis of observational cohorts reported birth outcomes among women exposed to efavirenz during the first trimester [10]. The primary endpoint was a birth defect of any kind with secondary outcomes,

including rates of spontaneous abortions, termination of pregnancy, stillbirths and PTD. GPCR Compound Library in vitro Sixteen studies met the inclusion criteria, 11 prospective and five retrospective. Nine prospective studies reported on birth defects among infants born to women with efavirenz exposure (1132 live births) and non-efavirenz-containing regimens (7163 live births). The analysis found no increased risk of overall birth defects among women exposed to efavirenz during first trimester compared with exposure to other ARV drugs. There was low heterogeneity between studies and only one neural tube defect was observed with first-trimester efavirenz exposure, giving a prevalence of 0.08%. Furthermore, the prevalence of overall birth defects with first-trimester efavirenz exposure was similar to the ranges reported in the general population. This Tau-protein kinase meta-analysis, which included the data from the APR and the IeDEA and ANRS databases, has been updated to include published data to 1 July 2011. The addition of 181 live births reported from five studies together with the updated report from the APR resulted in a revised incidence of neural tube defects in infants exposed to efavirenz during the first trimester of 0.07% (95% CI 0.002–0.39) [11]. Two publications have reported higher rates of congenital birth defects associated with efavirenz, Brogly et al. (15.6%) [12] and Knapp

et al. (12.8%) [13]. The Writing Group considers these rates to be inflated. Recruitment occurred prenatally but also up to 12 months of age, which could confer recruitment bias. Although the overall study numbers were large, the number of efavirenz exposures used as the denominator in the final analyses of first-trimester exposure was small, 32 and 47, respectively. There was no difference in the anomaly rate found with no exposure vs. any exposure in first/second/third trimester. In addition, no pattern of anomalies specific to efavirenz was described by these studies: patent foramen ovale (n = 1); gastroschisis (n = 1); polydactyly (n = 1); spina bifida cystica (n = 1); plagiocephaly (n = 1); Arnold Chiari malformation (n = 1); and talipes (n = 1).

Recently, a sensational study on endospore formation in Mycobacterium

marinum has been published (Ghosh et al., 2009); however, this claim was not confirmed in a later study (Traag et al., 2010). According to WHO, find more one-third of the world’s population is latently infected with Mycobacterium tuberculosis (MTB) (Inge & Wilson, 2008), which likely persist as dormant cells in the human organisms, posing a significant problem due to resistance to chemotherapy (Mitchison, 1980). Although dormancy is the commonly accepted explanation of latent mycobacterial infection (Young et al., 2005), limited information has been available about persisting bacterial forms and molecular mechanisms behind their stability and resistance to stressful factors. Among the known mechanisms responsible for the adoption of stress resistance MG-132 datasheet of bacterial cells, it is worth considering the role of histone-like proteins, which bind DNA, changing its topology (Dorman & Deighan, 2003)

and making it more stable against damage caused, for example, by γ or UV radiation (Boubrik & Rouvière-Yaniv, 1995). In Escherichia coli, histone-like proteins HU, H-NS, FIS also play an important role in transcription, recombination and replication (Thanbichler et al., 2005 and references therein). Histone-like protein, Hlp, is present in Mycobacterium smegmatis and contains the N-terminal domain, homologous to HU and the C-terminal domain with the mycobacterial specific PAKKA motif (Mukherjee et al., 2008). Regarding the physiological function of Hlp, it is worthwhile to note the significant increase in its level during transition of M. smegmatis cells to a nonreplicating state under microaerophilic conditions in the Wayne dormancy model. However, the viability

of cells of M. smegmatis strain with inactivated hlp gene was not clearly distinct from that of wild-type strain (Lee et al., 1998) in the same dormancy model. We may reason that Hlp has no significant RANTES role in the transition to dormancy in the relatively short-term Wayne model but may be essential for developing dormancy in nonreplicating cells at later stages. Indeed, many genes, different from those expressed in cells undergoing starvation in the Wayne model, are upregulated at late stages (>24 h) in M. tuberculosis cells subjected to hypoxia (enduring response) (Rustad et al., 2008). The objective of the present study is to clarify the role of Hlp in dormancy in M. smegmatis cells obtained in two experimental models after incubation in a prolonged stationary phase. We found that Hlp was essential for survival of NC cells or for a greater stability of specialized dormant forms, likely due to DNA condensation. Strains and plasmids used in this study are listed in Table 1. Mycobacterium smegmatis strain MC2 155 was routinely maintained on solid (1.

In addition, in the remaining PF–Purkinje cell synapses, the postsynaptic densities are disproportionally longer than the presynaptic active zones. These unique morphological phenotypes and Ca2+-resistant binding of the

NRX/Cbln1/GluD2 complex is consistent with the function of the complex as synaptic glue, connecting pre- and postsynaptic elements. The second unique feature of the NRX/Cbln1/GluD2 complex is that the secreted Cbln1 works by being sandwiched between presynaptic NRX and postsynaptic GluD2. In central nervous system synapses, synaptic organizers are classified into two categories: cell adhesion molecules that directly link pre- and postsynaptic elements and soluble factors. Most soluble synaptic organizers in the central nervous system, such as neuronal pentraxins (Xu et al.,

2003), fibroblast Napabucasin order growth factors (Terauchi et al., 2010) and Wnt-7a (Hall et al., 2000), work on either the pre- or postsynaptic site, depending on the location of their receptors (Johnson-Venkatesh & Umemori, 2010). Thus, the sandwich-type signaling by the NRX/Cbln1/GluD2 complex is unique in that secreted Cbln1 serves as a bidirectional synaptic organizer. For Cbln1 to bind to pre- and postsynaptic receptors simultaneously, Cbln1 needs to have at least two binding sites. This could have been achieved by the presence of multiple binding sites within single Cbln1 monomers or by the presentation of single binding sites in different

directions by forming a multimeric Cbln1 complex find more (Iijima MycoClean Mycoplasma Removal Kit et al., 2007). Recently, glial-derived neurotrophic factor was also proposed to serve as a synaptic adhesion molecule being sandwiched by its receptor glial-derived neurotrophic factor family receptor (GFR)α1 located at pre- and postsynaptic neurons (Ledda et al., 2007). In addition, leucine-rich glioma inactivated 1 was recently shown to be secreted from neurons and to organize presynaptic potassium channels and postsynaptic AMPA receptors by binding to its pre- and postsynaptic receptors, a disintegrin and metalloproteinase (ADAM) 22 and ADAM23, respectively (Fukata et al., 2010). These recent findings indicate that the sandwich type constitutes the third category of synaptic organizers. Advantages of sandwich-type synaptic organizers may include an additional level of regulation of synapse formation and its functions. For example, the expression of cbln1 mRNA is completely shut down in granule cells when neuronal activity is increased for several hours (Iijima et al., 2009). Similarly, a sustained increase in neuronal activity causes the internalization of GluD2 from the postsynaptic site of cultured Purkinje cells (Hirai, 2001). As Cbln1 and NLs compete for NRXs, such activity-dependent regulation of Cbln1 and GluD2 might lead to switching between NRX/NL and NRX/Cbln1/GluD2 modes of synaptogenesis.

The ammonium shock was achieved Gefitinib solubility dmso by adding ammonium chloride to 1 mM of the solution. Cellular fractions were obtained as follows: 50 mL of the culture was withdrawn before or 5 min after the ammonium shock. The culture was cooled by immersion in liquid nitrogen and the cells were harvested by centrifugation (5000 g for 5 min at 4 °C). The cells were resuspended in 1 mL of SP buffer (40 mM K2HPO4, 22 mM KH2PO4, 150 mM NaCl, pH 7.2) and processed exactly as described (Huergo et al. 2006). Membrane preparations were suspended in 6 M urea, 2 M thiourea, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate 4% (w/v), Pharmalyte pH 4–7, 0.5% (v/v) and 20 mM dithiothreitol.

For isoelectrofocusing, 500 μg of protein was loaded onto a 13 cm, pH 4–7 Selleckchem Torin 1 linear IPG strip (GE Healthcare). After rehydration, the isoelectrofocusing run was performed until the accumulation of 56 kVh, following the manufacturer’s instructions (GE Healthcare). The second dimension was achieved by 11% sodium dodecyl sulfate polyacrylamide gel electrophoresis

(SDS-PAGE). Gels were stained with colloidal Coomassie blue and images were analyzed using imagemaster platinum 6.0. The signal of each spot was normalized using the total density of the gel image. The experiments reported here were reproduced in two different biological replicates and two technical 2D-PAGE repetitions from the same sample. The spots were excised from the gels (ammonium shock treatment) and destained for 1 h in a solution of acetonitrile

50% (v/v) and 20 mM ammonium bicarbonate. Resveratrol The gel piece was immersed in pure acetonitrile for 5 min, the acetonitrile removed and the gel dried under air. In-gel digestion was performed using 0.1 μg of sequencing-grade trypsin in acetonitrile 10% (v/v) and 20 mM ammonium bicarbonate. After overnight incubation at 37 °C, aliquots of each digested sample were mixed with a saturated matrix solution of α-cyano-4-hydroxycinnamic acid (in acetonitrile 50% v/v, TFA 0.1% v/v), spotted onto the MALDI target and allowed to dry. Mass spectra were acquired using a MALDI-TOF/TOF Autoflex II (Bruker Daltonics). MS analyses were performed in a positive ion reflection mode using an accelerating voltage of 20 kV. MS/MS analyses were performed in a positive ion LIFT reflection mode. Peak lists were created using flexanalysis 3.0 software (Bruker Daltonics). Trypsin autolysis signals (842.5 and 2211.1) were used as internal standards when present. A database search was performed using mascot 2.2. Mass lists were searched against a database of H. seropedicae predicted proteins. Carbamidomethylation of cysteines was set as fixed and oxidation of methionine as variable modifications. Error tolerance was 100 p.p.m. for peptide mass fingerprint (PMF) search and for MS/MS parent ion. MS/MS ion search error was set as 0.3 Da. signalp 3.0 (Bendtsen et al., 2004) was used for prediction of Sec signal peptides. tatp 1.0 (Bendtsen et al., 2005b) was used to predict TAT-dependent signal peptides.