With regard to tuberculosis, although paradoxical IRIS associated with http://www.selleckchem.com/products/ink128.html Mycobacterium tuberculosis occurs in over 45% of individuals, CNS complications are presumed to be much lower [37]. In our cohort, just one case

of IRIS was related to tuberculosis. In two recent studies of 80 and 144 patients coinfected with tuberculosis and HIV, no cases of CNS-associated complications were reported [38]. In our study, none of the patients who presented a paradoxical IRIS had a previous history of any AIDS-defining opportunistic infection. Previous studies had suggested that patients previously undiagnosed with HIV infection presented a higher risk for the development of paradoxical IRIS if a CNS opportunistic infection was the AIDS-defining illness [4]. We did not find any other clinical or immunological parameter at baseline that predicted the development of IRIS after initiation of HAART. As previously described, in our cohort baseline CD4 cell count was not predictive of developing paradoxical IRIS [4, 39, 40]. However, patients who developed paradoxical IRIS had a more rapid immune restoration in response to HAART than patients who did not. This is consistent with findings from both retrospective and prospective analyses which revealed

a greater CD4 response in patients developing IRIS [7, 9, 40-42]. As previously indicated, no differences in the risk of developing IRIS were observed when protease inhibitor-containing regimens were compared with other regimens Gemcitabine in vitro [11, 39]. In our cohort, initiation of HAART during the first 2 weeks after the diagnosis of a neurological infection was not associated with a higher risk for the development of paradoxical IRIS. However, the retrospective design of our study and the low number of patients with IRIS limit the significance of this observation. Optimal timing for initiation of HAART is still a controversial issue in patients

with CNS opportunistic infections. Nowadays we have consistent data that indicate the benefits of early HAART in patients with opportunistic infections, even in those with tuberculosis [43]. However, in none of these studies were patients with CNS infections sufficiently represented. In some retrospective studies of patients with cryptococcal meningitis, Galeterone beginning HAART within 30–60 days of the treatment of meningitis has been associated with a higher risk of paradoxical IRIS and a higher mortality rate [4, 40]. In contrast, a prospective multicentre study did not identify an association between the timing of HAART and the development of IRIS [36]. Finally, a recent prospective study performed in sub-Saharian Africa showed a risk of mortality 3 times greater in HIV-infected patients who had begun ART within 72 h after cryptococcal meningitis diagnosis than in those in whom HAART was delayed for 10 weeks [13]. Development of IRIS seems not to worsen prognosis in patients with CNS opportunistic infections.

The comparison of both primer systems was conducted based on all available genera described in the list of prokaryotic names in the nomenclature (http://www.bacterio.cict.fr/index.html).

Subsequently, a simple matching coefficient (Jaccard coefficient) was calculated as described below: To revise the in vitro primer specificity, 16S RNA gene fragments were GSK3235025 concentration amplified by Com2xf/Ac1186r-PCR, using DNA from plaster and compost materials and both bioaerosol samples. The genomic DNA from the 18 different building material samples was used to verify the new primer system Com2xf/Ac1186r for screening of 16S rRNA gene clone libraries, generated using 27f-1492r primers (Lane, 1991). For screening of these clone libraries, the Actinobacteria-specific primer system www.selleckchem.com/products/Adriamycin.html developed by Stach et al. (2003) was used to compare the detectable Actinobacteria species. All PCR amplification reactions were performed in a final volume of 25 μL, containing 10 mM Taq buffer (+KCl), each primer at 200 nM, each dNTP at 0.2 mM, 2.0 mM MgCl2 and 0.02–0.025 U of Taq. Primers, cycling parameters and concentrations for PCR contents are shown in Tables 2 and 3. Amplification programs were started by an initial denaturation step at 95 °C for 3 min

and finished with a final extension step at 72 °C for 15 min and 30 min for add-on cloning analyses. Reactions were performed in a Thermocycler (My Cycler™, BioRad, Munich, Germany). A control PCR using only PCR reagents was always carried out. PCR comprised 25 cycles, except primer system 27f/1492r (35 cycles). PCR reactions contained 4 mg mL−1 bovine serum albumin. Negative PCR controls containing nuclease-free water instead of DNA were always carried out. either The PCR products were visually evaluated by 1% agarose gel electrophoresis with ethidium bromide staining. PCR was purified using a QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) and quantified photometrically (Ultrospec 4000, Amersham Biosciences, Freiburg, Germany).

Cloning analyses from four environmental samples (plaster, mature compost and two bioaerosols) and subsequent sequencing of clone inserts was done by Agowa (Berlin, Germany) using the M13F primer (Invitrogen Corp., Carlsbad, CA). For construction of 16S rRNA gene clone libraries from building material samples, the cloning kit Promega p GEM-T® Vector Systems (Madison, WI) was used with 27f-1492r primers according to the manufacturer’s instructions. Here, 100 white colonies from each sample were picked and incubated overnight at 37 °C on Luria–Bertani (LB) agar containing ampicillin (100 μg mL−1), X-Gal (80 μg mL−1) and IPTG (100 mM) (Sambrook & Russel, 2001). Clone inserts were reamplified to screen all generated clones for affiliation to Actinobacteria with the Actinobacteria-specific primer systems (Com2xf/Ac1186r SC-Act-235aS20/SC-Act-878aA19).

, 2010). We used six different Pseudomonas strains, four of which produce well-characterized secondary metabolites that inhibit root-pathogenic fungi. Pseudomonas fluorescens DR54 produces viscosinamide: a membrane-bound cyclic lipopeptide with biosurfactant properties and broad antifungal activity (Nielsen et al., 1999; Thrane et al., selleck chemicals llc 2000). Pseudomonas fluorescens CHA0 produces various extracellular metabolites, two of them being DAPG (2,4-diacetylphloroglucinol), which causes membrane damage in fungi (Pythium) and inhibits zoospores, and pyoluteorin, which inhibits the fungal respiratory chain (Keel et al., 1992; Laville et al., 1992). Pseudomonas sp. DSS73 produces amphisin, an extracellular

cyclic lipopeptide with biosurfactant Palbociclib concentration properties and broad antifungal activity (Sørensen et al., 2001; Nielsen & Sørensen, 2003), and Pseudomonas chlororaphis MA342 produces DDR (2,3-de-epoxy-2,3-didehydro-rhizoxin), a membrane-bound compound that inhibits mitosis in eukaryotic cells (Hökeberg et al., 1997; Brendel et al., 2007). Two Pseudomonas strains, P. fluorescens type strain DSM50090T (Deutsche Sammlung von Mikroorganismen und Zellkulturen) and P. fluorescens ATCC43928 (American Type Culture Collection), produce no known antagonistic secondary metabolites. We further included the well-suited food bacterium Enterobacter aerogenes SC (Christensen

& Bonde, 1985) as a positive control, and a treatment only with phosphate buffer, but without bacteria, as a negative control. The bacteria for the protozoan growth experiments were pure cultures grown on tryptic soy broth (TSB) medium (3 g L−1, Difco Bacto, Detroit) at 22 °C for 24 h. Bacteria were then diluted 1/10 in weak phosphate buffer (‘Neff’s modified amoeba saline’; Page, 1988), which yields bacterial cultures with 5–10 × 107 cells mL−1. This approach yields

more reproducible results than if a fixed cell number (e.g. 5 × 107 cells mL−1) is used for standard comparison between cultures. This is because different bacterial cultures with similar cell numbers may vary considerably with regard to carbon content, because cell sizes differ (Lekfeldt & Rønn, 2008). Bacterial cell size depends on the growth medium. Here, all Rebamipide bacteria were cultivated on the same medium and microscopic evaluation demonstrated that differences between cell sizes were negligible. No biofilm formation was observed in the current set-up, even though both bacteria and protozoa settled at the bottom of the experimental units (data not shown). The protozoa used in the experiments belong to several very distantly related protozoan lineages (Adl et al., 2007). They included three amoeboid Rhizaria (Cercomonadida) Cercomonas longicauda (SCCAP C 1), Neocercomonas jutlandica (SCCAP C 161), and Heteromita globosa (SCCAP H 251), three non-amoeboid Excavata (Bodonidae) Bodo caudatus (SCCAP BC 330), Bodo designis (UJ), and B.

coli FC40 system under carbon

starvation conditions and for the emergence of tetracycline-resistant mutants in response to antibiotic (tetracycline) treatment. NusA, a modulator of RNA polymerase, was previously shown to interact with Pol IV (Cohen et al., 2009). Hence, a model is proposed that DNA replication initiated during DSBR could generate DNA substrates that will stall RNA polymerase and lead to the recruitment of Pol IV by NusA (Cohen & Walker, 2010). The LexA regulon of P. aeruginosa and P. putida is significantly smaller than that of E. coli (Courcelle et al., 2001; Cirz et al., Vismodegib nmr 2005; Abella et al., 2007). Among the specialized DNA polymerases, the transcription of the Pol II gene polB is not induced by DNA damage in these organisms. Although the Pol IV gene dinB promoter has a LexA-binding site and the level of transcription

from this promoter is slightly increased in Pseudomonas species in the presence of DNA-damaging agents (Tegova et al., 2004; Cirz et al., 2005; Abella et al., 2007), the extent of SOS induction is considerably smaller than that observed in E. coli. Compared with E. coli, pseudomonads seem to have evolved a regulatory system allowing a significantly high basal level of particular SOS regulon genes already in the absence of this website DNA damage. Notably, the promoters of P. putida dinB gene and rulAB genes (encoding the Pol V homologue on toluene catabolic plasmid), both of which contain the LexA-binding site, are highly inducible by DNA damage LY294002 in E. coli, whereas in P. putida, they express already at a considerably high basal level (Tegova et al., 2004; Tark et al., 2005). This indicates that the P. putida LexA repressor has evolved a lower affinity to its target sites compared with E. coli LexA. Importantly, the majority of bacteria, including pseudomonads, lack chromosomal Pol V genes umuD and umuC, but instead carry a

multiple gene cassette encoding a second copy of the α-subunit of DNA polymerase III and a protein related to Y-family DNA polymerases, DnaE2 and ImuB, respectively (Abella et al., 2004; Erill et al., 2006; Koorits et al., 2007). Similar to the Pol V genes, these genes are induced by DNA damage. In P. putida this gene cluster is negatively controlled by another LexA repressor, LexA2, which binds the DNA sequence GTACN4GTGC (Abella et al., 2004, 2007). The binding site of LexA2 differs completely from that recognized by E. coli-like LexA, which binds the classical CTGTN8ACAG box. Pseudomonas aeruginosa has only one LexA protein, which is related to E. coli LexA, and the imuB and dnaE2-containing gene cluster of P. aeruginosa is negatively controlled by this LexA (Cirz et al., 2005). The gene cluster similar to that identified in P. putida (Abella et al., 2004) or part of it has been identified in many families of Proteobacteria (Abella et al., 2004; Erill et al., 2006).

, 2007). These signaling pathways do not function independently but influence each other through a complex network of synergistic and antagonistic www.selleckchem.com/products/rxdx-106-cep-40783.html interactions (Koornneef & Pieterse, 2008). Trichokonins upregulated the expression of SA-responsive PR gene acidic NtPR1a, ethylene-responsive gene basic NtPR3 and the key player in activating the JA signaling pathway, NtCOI1 (Fig. 4b). These results suggested

that multiple defense pathways are involved in Trichokonin-induced resistance in tobacco against TMV. Likely, cross-talk between the different defense pathways occurs. In summary, we studied the antiviral effect of Trichokonins against TMV infection and the mechanism involved. Trichokonins from T. pseudokoningii STI571 nmr SMF2 can induce tobacco systemic resistance against TMV via activation of multiple plant defense pathways. The results imply the potential of peptaibols in plant viral disease control. This work was supported by Hi-Tech Research and Development program of China (2007AA091504),

National Natural Science Foundation of China (30870047) and Foundation of State Key Lab of Microbial Technology, Shandong University, China. Table S1. Primers used for RT-PCR analysis in tobacco plants. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding author for the article. “
“Berberine,

a natural isoquinoline alkaloid found in many medicinal herbs, has been shown to be active against a variety of microbial infections. To examine the potential effects of berberine on Shigella flexneri, a whole-genome DNA microarray was constructed and a transcriptome analysis of the cellular responses of S. flexneri when exposed to berberine chloride (BC) was performed. Our data revealed that BC upregulated a group of genes involved in DNA replication, repair and division. Intriguingly, the expression of many genes related to cell envelope biogenesis however was increased. In addition, many genes involved in cell secretion, nucleotide metabolism, translation, fatty acid metabolism and the virulence system were also induced by the drug. However, more genes from the functional classes of carbohydrate metabolism, energy production and conversion as well as amino acid metabolism were significantly repressed than were induced. These results provide a comprehensive view of the changes in gene expression when S. flexneri was exposed to BC, and shed light on its complicated effects on this pathogen. Shigella is a gram-negative, facultative, intracellular pathogen responsible for endemic shigellosis, which remains a major worldwide health problem, particularly in developing countries. The estimated annual incidence of this disease is 160 million individuals, most of whom are children, and the annual mortality is 1.1 million (Kotloff et al., 1999).

coli K-12 substrain MG1655 (Fig. 1; Rhodius et al., 2006). This sequence is MDV3100 supplier not present in strain BEN2908 due to the integration of a pathogenicity island at this site (Chouikha et al., 2006). A signal structure proposed by Laikova to be the XylR-binding site was also found between the putative ribosome-binding site and the σ70−10 promoter sequence, suggesting, as it was proposed, that the yicJI operon is a member of the xylose regulons (Fig. 2a; Laikova et al., 2001). This motif was not found in the yicI-ORF1frz intergenic region. Finally, a motif containing

a palindromic sequence was found in the identified promoter sequence of the yicJI operon and three nucleotides upstream of the σ70−35 promoter sequence of the frz operon (Fig. 2b). This motif is correctly spaced to be a binding site for a regulator involved in the transcription of the two operons. Works are in progress in our laboratory to determine whether it represents an FrzR-binding site. The identification of a common motif in the yicJI and frz intergenic regions prompted us to test whether the two operons are cotranscribed. We previously identified a hairpin structure containing a G+C-rich region in the yicI-ORF1frz intergenic region (294 nt)

Rucaparib purchase of strain BEN2908. This RNA secondary structure begins 13 nucleotides after the stop codon of yicI and is followed by a polyU sequence. These features indicate the presence of a rho-independent transcription terminator. We also identified Ergoloid σ70−10 and σ70−35 putative promoter sequences beginning 54 and 76 nucleotides upstream of ORF1frz, respectively (EMBL accession number FM253092). To determine whether the yicJI and the frz operons are cotranscribed, RT-PCR experiments were performed with a reverse primer localized at the 5′ end of ORF1frz and a forward primer localized at the 3′ end of the yicI gene. The functionality of the ORF1frz identified promoter was also tested by RT-PCR using a reverse primer localized at the 5′ end of ORF1frz and a forward primer localized between the identified σ70−10 ORF1frz promoter

sequence and the start codon of ORF1frz. As a control, transcription of the yicI gene was tested by RT-PCR with reverse and forward primers, both localized in the yicI gene. Figure 3 indicates that some transcripts of yicI pass through the transcriptional terminator identified in the yicI-ORF1frz intergenic region and form cotranscripts with frz genes (lanes 2 and 4). The stronger amplification of ORF1frz transcripts by RT-PCR with primers localized in ORF1frz and between its promoter and start codon than with primers localized in yicI and in ORF1frz suggests that the ORF1frz promoter identified is functional (Fig. 3, lanes 4 and 6). The identification of a similar DNA motif in the yicJI and frz promoter regions and of yicJI and frz cotranscripts suggests that these two operons could be involved in the same physiological pathway.

coli K-12 substrain MG1655 (Fig. 1; Rhodius et al., 2006). This sequence is this website not present in strain BEN2908 due to the integration of a pathogenicity island at this site (Chouikha et al., 2006). A signal structure proposed by Laikova to be the XylR-binding site was also found between the putative ribosome-binding site and the σ70−10 promoter sequence, suggesting, as it was proposed, that the yicJI operon is a member of the xylose regulons (Fig. 2a; Laikova et al., 2001). This motif was not found in the yicI-ORF1frz intergenic region. Finally, a motif containing

a palindromic sequence was found in the identified promoter sequence of the yicJI operon and three nucleotides upstream of the σ70−35 promoter sequence of the frz operon (Fig. 2b). This motif is correctly spaced to be a binding site for a regulator involved in the transcription of the two operons. Works are in progress in our laboratory to determine whether it represents an FrzR-binding site. The identification of a common motif in the yicJI and frz intergenic regions prompted us to test whether the two operons are cotranscribed. We previously identified a hairpin structure containing a G+C-rich region in the yicI-ORF1frz intergenic region (294 nt)

selleck screening library of strain BEN2908. This RNA secondary structure begins 13 nucleotides after the stop codon of yicI and is followed by a polyU sequence. These features indicate the presence of a rho-independent transcription terminator. We also identified Tryptophan synthase σ70−10 and σ70−35 putative promoter sequences beginning 54 and 76 nucleotides upstream of ORF1frz, respectively (EMBL accession number FM253092). To determine whether the yicJI and the frz operons are cotranscribed, RT-PCR experiments were performed with a reverse primer localized at the 5′ end of ORF1frz and a forward primer localized at the 3′ end of the yicI gene. The functionality of the ORF1frz identified promoter was also tested by RT-PCR using a reverse primer localized at the 5′ end of ORF1frz and a forward primer localized between the identified σ70−10 ORF1frz promoter

sequence and the start codon of ORF1frz. As a control, transcription of the yicI gene was tested by RT-PCR with reverse and forward primers, both localized in the yicI gene. Figure 3 indicates that some transcripts of yicI pass through the transcriptional terminator identified in the yicI-ORF1frz intergenic region and form cotranscripts with frz genes (lanes 2 and 4). The stronger amplification of ORF1frz transcripts by RT-PCR with primers localized in ORF1frz and between its promoter and start codon than with primers localized in yicI and in ORF1frz suggests that the ORF1frz promoter identified is functional (Fig. 3, lanes 4 and 6). The identification of a similar DNA motif in the yicJI and frz promoter regions and of yicJI and frz cotranscripts suggests that these two operons could be involved in the same physiological pathway.

Miniaturization of isothermal calorimeters provides an even wider range of possibilities.

The first isothermal calorimeter was devised and used in 1782–1783 by Lavoisier and Laplace to determine the heat produced during chemical changes. This was the ‘ice-calorimeter,’ in which a sufficient amount of ice was used to keep the temperature constant (Lavoisier & Laplace, 1780). These early scientists realized that the mass of liquid water produced by the melting ice was directly proportional to the heat X-396 mouse produced by the reaction taking place atop the ice. Many improvements have of course been made since the early 19th century. In addition, several other types of calorimeters have evolved besides the ones that operate isothermally – for example adiabatic, constant-volume, constant-pressure, heat loss and temperature (differential) scanning calorimeters (van Herwaarden, LY2606368 manufacturer 2000). Some of them can also be used in the isothermal mode. In the isothermal approach, isothermal titration calorimetry has emerged as the broadly used standard for thermodynamic characterization of relatively fast reactions between molecules – for example ligand binding – especially for molecules of interest in biology (Cooper, 2003).

However, because isothermal titration calorimetry is mostly a tool for molecular studies, it is not covered here. This review focuses on IMC in microbiology for a wide variety of L-NAME HCl purposes including microorganism detection and discrimination, evaluation of microbial processes and determining the performance of antimicrobial agents. The term IMC is used here to refer to measurements in the microwatt range under essentially isothermal conditions (Wadsö, 2001). The related instruments are often called isothermal microcalorimeters. Most isothermal microcalorimeters are heat conduction calorimeters in which heat produced in the reaction vessel is allowed to flow to a heat sink, usually made of aluminum. Therefore,

so-called isothermal microcalorimeters are not truly isothermal, but allow small variations of the sample temperature (up to 0.1 °C). Variation in the sample temperature mostly does not affect the heat sink temperature significantly because the heat sink has a much higher heat capacity than the reaction vessel and its contents (usually × 100). In addition, the heat sink is often placed in a thermostat, ensuring its temperature stability. The heat transfer between the vessel and the heat sink takes place through a thermopile, allowing measurements of the heat produced or consumed (Wadsö & Goldberg, 2001). In other isothermal microcalorimeters, thermoelectric compensation is preferred to maintain isothermal conditions. Heat produced is compensated using Pelletier elements, and similarly, heat consumed is compensated either by an electric heater or by reversing the polarity of the Peltier elements (van Herwaarden, 2000).

1). In situations where there is a high risk of short-term mortality the addition of rifabutin should be strongly considered, for instance in persons with: 1 Advanced immunosuppression (CD4+ RG7204 order T lymphocyte count <25 cells/μL); There are few supporting data for the use of other drugs such as a fluoroquinolones or parenteral amikacin [33]. These should therefore only be considered when rifabutin or other first-line drugs

cannot be used because of drug interactions, intolerance or treatment failure. Clofazimine should not be used in the treatment of MAC as it is associated with excessive toxicity and higher mortality rates [34]. In summary, the preferred regimen for disseminated MAC is clarithromycin (500 mg twice daily) or azithromycin (500 mg/day) plus ethambutol (15 mg/kg/day). If rifabutin (usually 300 mg/day) is included in the regimen a dose adjustment is necessary if concurrently administered with a ritonavir-boosted protease inhibitor (150 mg three times a week) or efavirenz (450 mg daily) (seeTable 8.1). 8.3.4.2 Length of treatment for DMAC. • Individuals receiving HAART with a virological response and a CD4 count >100 cells/μL for at least this website 3 months in whom there has been a clinical response to DMAC therapy for at least 3 months can discontinue

Tobramycin therapy (category 3 recommendation) Most studies of the treatment of DMAC were performed in the pre-HAART era. However, there is no doubt that one of the most effective treatments for DMAC is

HAART. HAART should be initiated simultaneously or within 1–2 weeks of initiation of antimycobacterial therapy for DMAC disease, based on the experience with a range of opportunistic infections including a small number of cases with MAC [35] (category IV recommendation). If patients are already on HAART at the time of DMAC diagnosis, HAART should be continued and/or adjusted to ensure the viral load is undetectable (<50 copies/mL HIV-1 RNA) (category IV recommendation). Successful initiation of HAART is a key determinant of the duration of DMAC therapy. The incidence of DMAC has dropped dramatically with the use of HAART. Prior to the HAART era, therapy for DMAC was life-long. It has become clear that immune reconstitution and CD4 cell recovery secondary to HAART enables successful withdrawal of MAC therapy in most cases. Whilst there are no randomized clinical trial data to strongly recommend duration of MAC therapy after initiation of HAART, prospective non-randomized studies [36,37] and cohort studies [38,39] would suggest DMAC therapy can be safely discontinued in patients responding to HAART.

Zumbrunn and colleagues, unpublished data) (Figure 1). For reasons of consistency, the experts assessed all risks separately for an average traveler

Selleck PF-2341066 to Africa, Latin America, and Asia/Pacific. The study was approved by the Ethics Committee of Basel. The visual psychometric measuring instrument applied to record the participants’ risk perception, pictorial representation of illness and self measure (PRISM), was developed in 1995 and validated for the assessment of the subjective burden of suffering in patients with chronic diseases.[15, 16] It consists of a white board in DINA4 format with a fixed yellow disc, symbolizing a subject’s “self” in her/his current life situation, and a movable disc, representing an illness, which is placed on the board by the subject (Figure 2). The distance between the “self” and the illness [self-illness separation (SIS)] is the primary outcome of PRISM and inversely proportional to the perceived importance of the illness. For this study, “life situation” was specified as the planned journey and “illness” replaced by nine

health risks. The primary outcome was adjusted to self-risk separation (SRS) as a proxy for the risk perception. According to the severity, frequency of occurrence, or estimated concern for travelers, the following risks were included: Opaganib research buy general risk (overall danger of a specific journey), mosquitoes, malaria, rabies, Immune system epidemic outbreaks, sexually transmitted infections (STIs), accidents, terrorist attacks, and vaccination-associated adverse events (VAEs). In 2008, pre-travel data collection was carried out by a computer application of PRISM[17] (T. Zumbrunn and colleagues, unpublished data). For technical reasons, pre-travel data in 2009, expert data, and all post-travel data were collected using hard copies of the computer application. The

forms were scanned and distances measured by means of a computer-aided design (CAD) program.[18] The CAD coordinates were converted to the original scale (cm). Differences between the median perceptions of travelers and experts with nonoverlapping confidence intervals (CIs) were considered as statistically significant. The CIs were calculated using a bootstrap re-sampling method with 500 replicates. Linear regression was applied to detect differences among traveler subgroups and the SRS log10-transformed prior to analysis. A two-sided p value < 0.05 was considered as statistically significant. No adjustments for multiple testing were made. All analyses were performed using PASW Statistics 18 and R version 10.