parahaemolyticus possesses a full set of the exsACDE regulatory system, which is similar to that of P. aeruginosa and which regulates T3SS1-related gene expression. H-NS is a major component of the bacterial nucleoid and plays a crucial role in global gene regulation in enteric bacteria (Varshavsky et al., 1977; Hulton et al., 1990). H-NS affects the expression of many unrelated genes and several virulence genes in Salmonella Y-27632 molecular weight enterica serovar Typhimurium, Shigella sp. and Vibrio cholerae (Maurelli & Sansonetti, 1988; Hulton et al., 1990; Tobe et al., 1993; Harrison et al., 1994; O’Byrne & Dorman, 1994;

Nye et al., 2000). Therefore, we examined the possibility that T3SS1 genes are part of the H-NS regulon. As shown in Fig. 3a, the production of VscC1 and VepA proteins in a Δhns strain was considerably increased in both the bacterial pellet and the supernatant compared with that of the WT. A ΔhnsΔexsA mutant strain did not exhibit

increased production of these proteins (Fig. 3b), suggesting that exsA is necessary for overproduction of T3SS1-related proteins via hns gene deletion. We next examined the possibility that H-NS represses exsA expression using an exsA–lacZ transcriptional fusion reporter (Fig. 3c). Transcription of exsA–lacZ was dramatically increased in the hns deletion strain compared with that in the WT. The increase in exsA–lacZ transcription in the hns deletion strain was suppressed by Bcr-Abl inhibitor in trans complementation with the this website hns gene. These results

indicate that H-NS represses T3SS1-related gene expression by suppressing exsA gene expression. In summary, we identified VP1701 and VP1702 of V. parahaemolyticus as functional orthologues of P. aeruginosa ExsC and ExsE, respectively. As VP1701 has sequence similarity with its counterpart, it was not difficult to predict its function. Indeed, the production of T3SS1-related proteins was repressed in a Δvp1701 mutant and derepressed by complementation of the vp1701 gene (Fig. 1b and c). Unlike ExsA (VP1699), ExsD (VP1698) and ExsC (VP1701), sequence annotation of the T3SS1 region on the genome of V. parahaemolyticus did not reveal any CDSs predicted to encode the homologue of P. aeruginosa ExsE. However, we found that one hypothetical CDS (VP1702) was encoded next to the vp1701 (exsC) gene. Deletion of the vp1702 gene deregulated the production of T3SS1-related proteins. Furthermore, VP1702 itself was a substrate for the T3SS1 secretion system. These properties of VP1702 of V. parahaemolyticus conform with those of its counterpart in P. aeruginosa. In P. aeruginosa, the coupling of transcription to secretion is mediated by three interacting proteins (ExsC, ExsE and ExsD) that regulate ExsA transcription activity (Yahr & Wolfgang, 2006). Although it is still unknown whether ExsC, ExsE and ExsD of V.

“
“Institute of Food Research, Norwich, UK Norwich Medical School, University of East Anglia, Norwich, UK During bacterial infection, professional phagocytes are attracted to the site of infection, where they constitute a first line of host cell defense. Their function is to engulf and destroy the pathogens. Thus, bacteria must Roxadustat solubility dmso withstand the bactericidal activity of professional phagocytes, including macrophages to counteract the host immune system. Bacillus cereus infections are characterized by bacteremia despite the accumulation of inflammatory

cells at the site of infection. This implies that the bacteria have developed means of resisting the host immune system. Bacillus cereus spores survive, germinate, and multiply in contact with macrophages, eventually producing toxins that kill these cells. However, the exact mechanism by which B. cereus evades immune attack remains unclear. This review addresses the interaction between B. cereus and macrophages, highlighting, in particular, the ways in which the bacteria escape

the microbicidal activities of professional phagocytes. “
“In this review, we address some recent developments in the field of bacterial protein phosphorylation, focusing specifically on serine/threonine and tyrosine kinases. We present an overview of recent studies outlining the scope of physiological processes that are regulated by phosphorylation, ranging from cell cycle, growth, cell morphology, to metabolism, developmental phenomena, and virulence. Specific emphasis is placed on Mycobacterium tuberculosis Selleckchem Palbociclib as a showcase organism for serine/threonine kinases, and Bacillus subtilis to illustrate the importance of protein phosphorylation in developmental processes. We argue that bacterial serine/threonine and tyrosine kinases

have a distinctive feature of phosphorylating multiple substrates and might thus represent integration nodes in the signaling network. Some open questions regarding Y-27632 2HCl the evolutionary benefits of relaxed substrate selectivity of these kinases are treated, as well as the notion of nonfunctional ‘background’ phosphorylation of cellular proteins. We also argue that phosphorylation events for which an immediate regulatory effect is not clearly established should not be dismissed as unimportant, as they may have a role in cross-talk with other post-translational modifications. Finally, recently developed methods for studying protein phosphorylation networks in bacteria are briefly discussed. “
“Multicellular organisms limit the availability of free iron to prevent the utilization of this essential nutrient by microbial pathogens. As such, bacterial pathogens possess a variety of mechanisms for obtaining iron from their hosts, including a number of examples of vertebrate pathogens that obtain iron directly from host proteins. Recently, two novel members of the colicin M bacteriocin family were discovered in Pectobacterium that suggest that this phytopathogen possesses such a system.

All participants were followed for a minimum of 6 months after KS diagnosis, until death or discharge or until 31 August 2008. Study physicians at Tororo District Hospital collected clinical information during screening, using standardized instruments. Demographic and socioeconomic characteristics and Tamoxifen manufacturer past medical history were obtained by interviewing patients and reviewing available medical records. All HIV-infected participants received a clinical assessment by a study physician including VL and CD4 cell count measurements as well as tests for liver and renal function. During

follow-up, field officers completed weekly client monitoring forms that included information on client symptoms, problems with taking medication or other information which might impact the participant’s health. Seriously ill patients were encouraged to come to the study clinic/hospital for treatment by study staff. Initial diagnoses of KS were made by study physicians based on clinical presentation and were confirmed histologically by pathologists at Makerere University Medical School in Kampala. We categorized KS based on the extent of the disease. Localized KS was defined as lesions that were

confined to the skin and/or lymph node and/or minimal oral ICG-001 disease. Visceral KS involved an internal lesion (e.g. oral, gastrointestinal or lung). Diagnosis of visceral KS was supported by chest radiographs and sonography where applicable. All proposed KS diagnoses were discussed and approved by the medical staff during a weekly medical case conference meeting. We identified participants diagnosed with KS at baseline (prevalent KS) or on follow-up (incident KS) through the database and abstracted further information from their medical

charts. Specific anti-neoplastic therapy was not available in Tororo; however, some participants were able to access chemotherapy at Mulago Hospital, the national referral hospital in Kampala. We defined participants as having completed chemotherapy if they received at least three courses of three agents (vincristine, vinblastine and adriamycin). Subjects were defined as having had partial chemotherapy if they started but did not Leukotriene-A4 hydrolase complete three courses of the three anticancer drugs. We defined complete resolution of KS lesions as the absence of any detectable KS disease including tumour-associated oedema, persisting for at least 4 weeks. For pulmonary KS, improvement of radiological findings was also required. We determined KS-related mortality by reviewing post mortem and case management conference forms. We calculated the incidence of KS in the participants, who were considered to be at risk from the day of enrolment in the study, if they had not been diagnosed with KS at baseline. Subjects were followed until they developed KS (the event), or until they died.

, 2008, 2009, 2011; Lovejoy & Krauzlis, 2010). We collected data from two (J and M) adult, male rhesus macaque monkeys (Macaca mulatta) that were 10–15 years of age and weighed 12–15 kg. The monkeys were prepared with standard surgical techniques that have been described Adriamycin purchase in detail

previously, and all experimental protocols for the monkeys were approved by the Institutional Animal Care and Use Committee (of the Salk Institute) and complied with US Public Health Service policy on the humane care and use of laboratory animals. Note that monkey J was referred to as monkey F in Lovejoy & Krauzlis (2010). Monkeys performed the selective attention tasks described in Lovejoy & Krauzlis (2010) and Hafed et al. (2011) (see also Fig. 1A). Briefly, every trial began with the onset of a small white fixation spot (9 × 9 min arc dimensions) similar to that in Hafed et al. (2009) and presented on a CRT display. Monkeys were allowed 500 ms to bring their gaze to within ~1–1.5° around this spot, after which four rings appeared in each visual quadrant in the periphery, alongside the fixation spot. Each ring was 4.4° in radius, with its center being at an eccentricity of 8.2° relative to the central spot. The rings were 0.25°

thick, and their luminance was 25 cd/m2. Background luminance selleck products was 14 cd/m2, and the white fixation spot was of luminance 50 cd/m2. One of the rings was a different color from the remaining three, serving as the cue to attend to the ring’s quadrant, but it had the same luminance as the other three rings. Random dot motion patches (0% coherence) appeared inside each ring after trial onset (radius of the motion patches, 4.25°), and, after some random delay, a brief coherent motion pulse appeared in the cued quadrant as well as in the diametrically

opposite one (called the ‘foil’). The monkeys’ task was to Fossariinae indicate the direction of the brief motion pulse in the cued quadrant, irrespective of the direction of the distracting motion pulse that appeared simultaneously in the diametrically opposite quadrant. In one variant of the task, the monkeys generated a saccade in the direction of the cued motion pulse to indicate their response; in the other variant, they pressed one of four buttons arranged spatially in the four possible directions of motion in the cued pulse. We inactivated the intermediate and deep layers of the SC, as described in detail in Lovejoy & Krauzlis (2010). Briefly, we injected the GABA agonist muscimol (0.3–0.5 μL, 5 μg/μL) into the intermediate and deep layers of the SC with an injection cannula like that described in Chen et al. (2001); supplementary Table 1 of Lovejoy & Krauzlis (2010) provides a complete list of injection volumes for each experiment. We aimed the cannula in the SC retinotopic map such that we could inactivate a population of neurons representing one of the visual quadrants used in the behavioral task of Fig. 1.

However, Che1-dependent signaling is shown to contribute indirectly to surface attachment, indicating that distinct mechanisms are likely underlying flocculation and attachment to surfaces in A. brasilense. Chemotaxis is a widespread function in motile soil bacteria as it affords cells with the ability to sense and to Alisertib supplier navigate toward the most favorable niches

for growth (Wadhams & Armitage, 2004). At the molecular level, the chemotaxis pathway is the dedicated chemosensory signal transduction system that allows cells to couple detection of physicochemical changes in their surroundings to changes in the swimming pattern (i.e. chemotaxis). Chemotaxis signal transduction has been best studied in Escherichia coli and experimental evidence indicates that this prototypical enteric model is conserved and functions similarly (with some variations on the theme) in phylogenetically diverse motile bacteria. In addition to regulating chemotaxis responses in motile bacteria, chemotaxis-like signal transduction pathways were shown to regulate cellular behaviors other than flagellar rotation in several other bacterial species (Kirby, 2009), including the alphaproteobacterium Azospirillum brasilense, a soil diazotroph (Bible et al., 2008).

In only a few cases, however, have the molecular targets of these chemotaxis-like pathways been identified. Selleckchem CHIR-99021 The A. brasilense Che1 chemotaxis-like pathway has been shown to have a minor,

and likely indirect, function in regulating chemotaxis behavior in this species (Hauwaerts et al., 2002; Bible et al., 2008; Edwards et al., 2011). Experimental evidence indicates that Che1 functions to modulate changes in adhesive cell surface properties which impact the propensity for cell-to-cell aggregation and flocculation (Bible et al., 2008). Deletions of cheA1 or cheY1, which each code for central proteins controlling the response output of the signal transduction pathway, yield cells that aggregate and flocculate more than the wild-type strain (Bible et al., 2008). A mutant strain deleted for all of the genes encoded within the che1 gene cluster has a phenotype similar to the strains lacking only CheA1 or CheY1, consistent with a role for Che1 this website in regulating the ability of cells to flocculate. A strain carrying a mutation that disrupts the function of both CheB1 and CheR1 is severely impaired in flocculation, consistent with CheB1 and CheR1 functioning in a signaling feedback loop that controls chemosensory adaptation (Stephens et al., 2006; Bible et al., 2008). Other possible roles that Che1 may have on functions such as adhesion to surfaces or root colonization, have been previously proposed to be related to flocculation (Burdman et al., 2000a, b) but have not yet been investigated. The purpose of the present study was to determine the conditions under which A.

09; 95% CI 0.92 to 1.29; P=0.32]. However, the number of cases of arthralgia (RR 1.49; 95% CI 1.17–1.89; P=0.001) and oedema (RR 3.95; 95% CI 1.73 to 9.00; P=0.001) were higher in the GH axis drug arm than in the placebo arm. Despite the extraordinary progress that has been made in the treatment of HIV infection with HAART, metabolic derangements, click here including central fat accumulation and peripheral lipoatrophy, have become a serious concern for many patients. Treating HIV-associated lipodystrophy is important for a number of reasons. Loss of SAT, especially in the face, can

cause significant emotional distress and can lead to poor self-esteem [21]. Some patients with lipodystrophy become worried that their HIV status is easily apparent [22]. Potential interventions, particularly for abnormal fat deposition, include exercise, medical therapy and surgery. Unfortunately, medical therapeutic options in the treatment of HIV-associated lipodystrophy are Dorsomorphin limited. Patients with HIV-associated lipodystrophy have decreased secretion of GH, a hormone with lipolytic properties [20]. Therefore, we sought to investigate GH axis treatments. The results of our systematic review demonstrate that GH axis treatments significantly reduced VAT by an average of 20.20 cm2. GH and tesamorelin were particularly effective, reducing VAT by 35.61 and 22.65 cm2, respectively.

Our results also demonstrate that GH axis treatments significantly increased LBM. Tesamorelin was the most effective GH axis treatment for improving LBM, resulting in an increase of 1.35 kg relative to placebo. With a total of 610 participants in the treatment groups and 281 in the placebo groups, we feel that the results are a reliable measure of the efficacy of tesamorelin. GHRH was also effective at improving

LBM, resulting in an increase of 1.20 kg compared to placebo, but there was only one study [23] in our systematic review that Calpain compared GHRH with placebo, and there were only 14 participants in the treatment group and 15 in the placebo group. The overall effects of GH and IGF-1 vs. placebo in improving LBM were not statistically significant (P=0.09 and 0.06, respectively). Funnel plots were constructed for the primary outcomes to assess publication bias. A symmetric inverted funnel shape was obtained for change in LBM. There were insufficient points in the funnel plots for change in VAT or SAT to allow any meaningful conclusions to be drawn. Thus, no evidence for publication bias was detected. The overall effect of GH axis treatments on improving SAT was not significant. Although adipose is fundamentally the same tissue in the viscera and in subcutaneous locations, loss of SAT (lipoatrophy) and VAT accumulation (lipohypertrophy) appear to be separate processes controlled by different mechanisms [24], and our results support this hypothesis.

J.L. and J.-S.H. contributed equally to this work. “
“Paecilomyces lilacinus was described more than a century ago and is a commonly occurring fungus in soil. However, Enzalutamide nmr in the last decade this fungus has been increasingly found as the causal agent of infections in man and other vertebrates. Most cases of disease are described from patients

with compromised immune systems or intraocular lens implants. In this study, we compared clinical isolates with strains isolated from soil, insects and nematodes using 18S rRNA gene, internal transcribed spacer (ITS) and partial translation elongation factor 1-α (TEF) sequences. Our data show that P. lilacinus is not related to Paecilomyces, represented by the well-known thermophilic and often pathogenic Paecilomyces variotii. The new genus name Purpureocillium is proposed for P. lilacinus and the new combination Purpureocillium lilacinum is made here. Furthermore, the examined Purpureocillium lilacinum isolated grouped in two clades based on ITS and partial TEF sequences. The ITS and TEF sequences of the Purpureocillium lilacinum isolates used for biocontrol of nematode pests are identical to those causing infections Smoothened inhibitor in (immunocompromised) humans. The use of high concentrations of Purpureocillium lilacinum spores

for biocontrol poses a health risk in immunocompromised humans and more research is needed to determine the pathogenicity factors of Purpureocillium lilacinum. Paecilomyces lilacinus is a ubiquitous, saprobic filamentous fungus commonly isolated from soil, decaying vegetation, insects, nematodes and laboratory air (as contaminant), and is a cause of infection in man Endonuclease and other vertebrates. This species can colonize materials such as catheters and plastic implants and can contaminate antiseptic creams and lotions, causing infections in immunocompetent and immunocompromised patients (Castro et al., 1990; Orth et al., 1996; Itin et al., 1998). The prevalence of P. lilacinus in patients has increased recently (Carey et al., 2003; Rosmaninho et al., 2010). A review of 119 infections caused by P. lilacinus after 1964 showed that the most frequent

manifestation is keratitis, but other sites of the body were also affected (Pastor & Guarro, 2006). Keratitis caused by P. lilacinus typically occurs by external invasion. Common predisposing factors are chronic keratopathy, environmental trauma, implant surgery following lens and/or cornea replacements and extended use of contact lenses (Domniz et al., 2001; Yuan et al., 2009). Paecilomyces lilacinus infections are reported in patients taking immunosuppressant drugs for transplant surgery for liver, kidneys, bone marrow and heart (e.g. Castro et al., 1990; Orth et al., 1996; Lott et al., 2007; Schooneveld et al., 2007). Although commonly reported as a component of the soil mycobiota, the source of P. lilacinus infections in humans has rarely been traced. Exceptions are a catheter-related P. lilacinus fungemia in an immunocompromised child (Tan et al.

Clearly, this expands on previous studies on

the effect of ribosome inhibitors on tmRNA levels in other bacteria (Montero et al., 2006; Paleckova et al., 2006). To our knowledge, this is the first direct study of tmRNA in mycobacteria. Funding for this study was provided by National Institutes of Health (NIH)/National Institute of Allergy and Infectious Diseases (NIAID) grant RO1-AI052291 and the Department of Pathology and Laboratory Medicine, Children’s Hospital Los Angeles. Fig. S1. Changes in the level of pre-tmRNA (shaded bars) and tmRNA (open bars) in Mycobacterium bovis BCG following a 24-h incubation with streptomycin (STR) at 0, 4, 8, or 16 μg mL-1. 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. “
“Ethyl carbamate Saracatinib (EC) is a group 2A carcinogen generated from a few precursors in many fermented foods and alcoholic beverages. Citrulline, urea, carbamoyl phosphate, and learn more ethanol are common precursors detected in fermented foods. In this study, citrulline was proved to be the main EC precursor in soy sauce, which was found to be accumulated in moromi mash period and correlated with the utilization of arginine by koji bacteria. Six koji isolates belonging to three genera were identified to be able to accumulate citrulline via the arginine

deiminase (ADI) pathway. Among these strains, only Pediococcus acidilactici retained high activities in synthesis and accumulation of citrulline in the presence of high concentration of sodium chloride. These results suggested that P. acidilactici is responsible for the accumulation of citrulline, one of the EC precursors, in the process of soy sauce fermentation. “
“Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The

University of Tokyo, Japan Melittin is one of the best-studied antimicrobial peptides, and many studies have focused on the membrane underlying its membrane-disruptive activity. We previously showed that melittin could cause some hallmarks of apoptosis in Candida albicans. Here, we first report the exact mechanism of melittin-induced fantofarone fungal apoptosis. We first characterized the reactive oxygen species generated by melittin. The results showed that melittin strongly produced highly reactive hydroxyl radicals (˙OH), which contribute to cell death. Next, we showed that melittin also disrupted the mitochondrial membrane potential (ΔΨm) and induced the Ca2+ release from the endoplasmic reticulum and its remarkable accumulation in mitochondria. Finally, we investigated the role of caspase in the apoptotic pathway. The results showed that melittin activated metacaspase, which was mediated by cytochrome c release. To summarize, melittin is involved in the mitochondria- and caspase-dependent apoptotic pathway in C. albicans.

Clearly, this expands on previous studies on

the effect of ribosome inhibitors on tmRNA levels in other bacteria (Montero et al., 2006; Paleckova et al., 2006). To our knowledge, this is the first direct study of tmRNA in mycobacteria. Funding for this study was provided by National Institutes of Health (NIH)/National Institute of Allergy and Infectious Diseases (NIAID) grant RO1-AI052291 and the Department of Pathology and Laboratory Medicine, Children’s Hospital Los Angeles. Fig. S1. Changes in the level of pre-tmRNA (shaded bars) and tmRNA (open bars) in Mycobacterium bovis BCG following a 24-h incubation with streptomycin (STR) at 0, 4, 8, or 16 μg mL-1. 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. “
“Ethyl carbamate selleck kinase inhibitor (EC) is a group 2A carcinogen generated from a few precursors in many fermented foods and alcoholic beverages. Citrulline, urea, carbamoyl phosphate, and selleck screening library ethanol are common precursors detected in fermented foods. In this study, citrulline was proved to be the main EC precursor in soy sauce, which was found to be accumulated in moromi mash period and correlated with the utilization of arginine by koji bacteria. Six koji isolates belonging to three genera were identified to be able to accumulate citrulline via the arginine

deiminase (ADI) pathway. Among these strains, only Pediococcus acidilactici retained high activities in synthesis and accumulation of citrulline in the presence of high concentration of sodium chloride. These results suggested that P. acidilactici is responsible for the accumulation of citrulline, one of the EC precursors, in the process of soy sauce fermentation. “
“Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The

University of Tokyo, Japan Melittin is one of the best-studied antimicrobial peptides, and many studies have focused on the membrane underlying its membrane-disruptive activity. We previously showed that melittin could cause some hallmarks of apoptosis in Candida albicans. Here, we first report the exact mechanism of melittin-induced FER fungal apoptosis. We first characterized the reactive oxygen species generated by melittin. The results showed that melittin strongly produced highly reactive hydroxyl radicals (˙OH), which contribute to cell death. Next, we showed that melittin also disrupted the mitochondrial membrane potential (ΔΨm) and induced the Ca2+ release from the endoplasmic reticulum and its remarkable accumulation in mitochondria. Finally, we investigated the role of caspase in the apoptotic pathway. The results showed that melittin activated metacaspase, which was mediated by cytochrome c release. To summarize, melittin is involved in the mitochondria- and caspase-dependent apoptotic pathway in C. albicans.

In 2011, MSM accounted for 54% of all new HIV diagnoses in Spain [1]. HIV

testing is an important part of HIV prevention activities, as it is required to diagnose HIV infection. Based on the results of HIV testing, prevention programmes focused on the HIV status of the person may be very appropriate to reduce acquisition and transmission of the infection. The advantage of being tested regularly for HIV is that early diagnosis is vital for timely access to treatment and to control the spread of the virus. Some studies have reported that, once people know they are HIV-positive, many of them reduce high-risk sexual behaviours compared with untested people [2]. Diagnosis is also desirable because it allows Barasertib early initiation of antiretroviral therapy, which reduces viral load, which in turn may reduce the risk of transmission Trichostatin A chemical structure of HIV. Serostatus awareness is beneficial at the individual and population levels, and is in line with the

‘test and treat’ approach to control the spread of HIV [3]. Undiagnosed HIV infection is a major potential source of the spread of infection. An important number of new infections are acquired from sexual partners whose infection is undiagnosed [4, 5]. Therefore, to monitor the epidemic among MSM, it is important to know why, when and where they are tested or, conversely, why individuals do not seek HIV testing or refuse it if it is offered. In view of the relatively limited knowledge regarding MSM who have never been tested for HIV in Spain, the aims of this study were to describe the sociodemographic profile of MSM who have never been tested for

HIV, and to analyse factors associated with never having been tested for HIV. A total of 13 753 participants completed the survey. The inclusion criteria were: being male; living in Spain; being at ifenprodil or over the age of sexual consent in Spain; having sexual attraction to men and/or having had sex with men; indicating having understood the nature and purpose of the study; and providing consent to take part in the study. After exclusion of individuals who did not fulfill the inclusion criteria or with inconsistent data, the final sample consisted of 13 111 men. The questionnaire was available in 25 European languages simultaneously and included core questions on sociodemographic characteristics, risk behaviours, history of diagnoses of HIV infection and other STIs, HIV prevention needs (information, access to condoms, etc.), and service uptake. The European MSM Internet Survey (EMIS) was approved by the Research Ethics Committee of the University of Portsmouth, UK (REC application number 08/09:21). This study had a collective approach, including public health, academic and nongovernmental organization (NGO) sectors, and social media. The EMIS was available online for completion over the course of 12 weeks in 2010. Promotion occurred mainly through national and transnational commercial and NGO websites, and social networking websites.