Conjugal transfer to L. mesenteroides M7-1 was only obtained with pRE25, albeit at very low frequency (Table 4). Gene transfer from RE25 to L. mesenteroides M7-1 has been observed before at low frequencies (Devirgiliis et al., 2009), and so the unsuccessful transfer of pRE25* from E. faecalis to L. mesenteroides is probably due to the naturally occurring low efficiency of gene transfer between these species No transconjugants were obtained with E. faecalis Lapatinib ic50 1528, Lactobacillus fermentum ROT1, and Staphylococcus aureus VG1 as recipients (Table 1), most probably due to plasmids incompatible to pRE25 present in those strains. The comparison of pRE25* with its parental plasmid pRE25 Selumetinib solubility dmso revealed

that the inserted 2.7-kb sequence did not affect the copy number of pRE25*, nor did it have a major impact on its conjugational potential. Furthermore, both pRE25* and the gfp marker were stable, showing that E. faecalis CG110/gfp/pRE25* is suitable as a marker tool to examine horizontal ABR gene transfer

in complex microbial communities using elevated experimental durations. After construction and characterization of E. faecalis CG110/gfp/pRE25*, the tool was tested in a complex microbial background for its functionality. Fresh overnight cultures of the donor strain E. faecalis CG110/gfp/pRE25*, the recipient strain L. monocytogenes 10403S, and the transconjugant L. monocytogenes 10403S/pRE25* (Table 1) were mixed at different transconjugants to donor ratios ranging from 0.2 : 1 to 2000 : 1 in complex microbiota background. The composition of this microbiota was determined by qPCR and consistent with the main groups usually encountered in infant feces (Laboratory of Food Biotechnology, ETH Zurich, unpublished data). Subsequently, donor and transconjugants were quantified by real-time PCR and plate counts. The

ratio of pRE25* to gfp quantified by real-time PCR was plotted against the ratio calculated from plate counts and showed linear correlation coefficient (R2 of >0.99) over a pRE25*/gfp ratio of more than three orders of magnitude (Fig. 2). Furthermore, differences as crotamiton low as 0.2 transconjugants per donor were detectable by qPCR, thereby elaborating the detection limit of the method. This demonstrates that the genetic markers of E. faecalis CG110/gfp/pRE25* can be quantified in complex backgrounds by qPCR and that E. faecalis CG110/gfp/pRE25* is indeed a suitable tool for quantification of HGT. Even though new technologies, for example metagenomic sequencing, yield a deep insight into the human microbiome (Qin et al., 2010), general links between DNA sequences and their transmission route within the microbiota cannot be established using such methods, making use of tagged strains and genes insurmountable for mechanistic studies. The novel strain E.

Second, we evaluated the stimulus-independent hemispheric balance thought to indicate higher level cortical processing. The results dissociate three, partly overlapping, time intervals: SB203580 solubility dmso the P1m (45–85 ms) was evoked by missed and detected target tones alike. Subsequent negative activity was only observed when listeners indicated awareness of the target stream inside the multi-tone masker. In the N1m time interval (75–175 ms), hemispheric balance of the ARN and N1m was modulated by stimulus lateralization. In the subsequent time interval (175–275 ms), auditory-cortex activity was generally right-lateralized in silence and balanced under informational masking, but was not modulated by

stimulus lateralization.

These results suggest that the BTK high throughput screening same auditory-cortex activity that varies with perceptual awareness also shows sensory response features. This is in accordance with models for visual perception, suggesting that sensory competition determines whether midlevel visual responses occur automatically or vary with perceptual state. “
“High-fat diet (HFD) consumption has been demonstrated to cause peripheral and neuronal insulin resistance, and brain mitochondrial dysfunction in rats. Although the dipeptidyl peptidase-4 inhibitor, vildagliptin, is known to improve peripheral insulin sensitivity, its effects on neuronal insulin resistance and brain mitochondrial dysfunction caused by a HFD are unknown. We tested the hypothesis that vildagliptin prevents neuronal insulin resistance, brain mitochondrial dysfunction, learning and memory deficit caused by HFD. Male rats were divided into two groups to receive either a HFD or normal diet (ND) for 12 weeks, after which rats in each group were fed with either

vildagliptin (3 mg/kg/day) or vehicle for 21 days. The cognitive function was tested by the Morris Water Maze prior to brain removal for studying neuronal insulin receptor (IR) and brain mitochondrial function. In HFD rats, neuronal insulin resistance and brain mitochondrial dysfunction were demonstrated, with impaired learning and memory. Vildagliptin prevented neuronal insulin resistance by restoring insulin-induced long-term selleck chemicals llc depression and neuronal IR phosphorylation, IRS-1 phosphorylation and Akt/PKB-ser phosphorylation. It also improved brain mitochondrial dysfunction and cognitive function. Vildagliptin effectively restored neuronal IR function, increased glucagon-like-peptide 1 levels and prevented brain mitochondrial dysfunction, thus attenuating the impaired cognitive function caused by HFD. “
“Patent Examination Cooperation Center of the Patent Office, SIPO, Beijing, China The presence of minichromosomes is very common in haloarchaea, but little is known about the coordination of replication between the major and minor chromosomes.

0% and 16.9%, respectively).

There were also some discrepancies concerning the region of origin: in the cohort, German origin was more common (76.3% and 68.7%, respectively), while patients originating from sub-Saharan Africa and South and South-East Asia were particularly underrepresented. However, a good general correlation with national surveillance data (and hence representativeness at the national level) is the main strength of the ClinSurv HIV cohort compared with another HIV-infected cohort implemented in Germany in 2004, the patient cohort of the German Competence Network BIBF 1120 price for HIV/AIDS (KompNet) [24]. Although KompNet started data collection at 44 sites, because of reduced financing this number had to be reduced and is currently 25 sites. As patient enrolment in KompNet requires informed consent, comparison of the composition of this cohort with the composition of the national German HIV surveillance database reveals significant differences with regard to sex, age and transmission

group category [24]. However, the KompNet cohort collects more variables than ClinSurv HIV. The number of patients enrolled in KompNet HIV decreased from a total of 6817 new annual cases in 2005 to 1147 cases in 2007, while patient enrolment in ClinSurv HIV turned out to be very stable in the long term (Fig. 2). In Germany, PD0325901 mw a growing proportion of HIV-infected patients, especially at early stages of HIV infection, are treated by primary care physicians, who have special training in HIV treatment. They co-operate with the participating clinical

centres if their patients reach advanced disease stages. As the ClinSurv sites are very experienced in HIV treatment, the proportion of patients with advanced clinical stage disease or AIDS may be overrepresented in the cohort, explaining why the cohort is estimated to represent nearly one-third of all patients in HIV stage CDC-C, but only 20% of all PLWHA. In addition to the limited number of variables collected in ClinSurv HIV, another limitation of this cohort study is click here the unequal geographical distribution of sites, which are situated predominantly in the north, north-east and west of the country. However, the study population is surprisingly stable with regard to newly enrolled patients and loss to follow-up, in particular taking into consideration the open observational cohort design. Another advantage is that patients’ informed consent is not needed as the data collection remains under federal law regulations. This makes data collection more representative than in studies requiring informed consent, although the number of variables is more limited. The proportion of ∼11% of patients lost to follow-up seems rather high; however, this number reflects the German situation, where patients, including PLWHA, are free to choose their treating physician when they seek for medical care.

ZFF from Phytophthora nicotianae, Phytophthora sojae, and Pythium aphanidermatum triggered luminescence of the Vibrio harve7yi AI-2 reporter, indicating the presence of AI-2 in zoospore extracellular products and the potential of cross-kingdom communication between

oomycetes and bacteria. The production of AI-2 by zoospores was confirmed by chemical assays. These results Caspase activity assay provide a new insight into the physiology and ecology of oomycetes. Phytophthora and Pythium in Oomycota of Stramenopila are phylogenetically related to marine algae, but resemble fungi morphologically. Many species in these two genera are destructive pathogens that attack a broad range of economically important agricultural and ornamental crops as well as forest tree species. They produce asexual sporangia that release flagellate zoospores as their primary dispersal and infection agents (Deacon & Donaldson, 1993; Judelson & Blanco, 2005). Zoospores secrete a host of molecules during the homing process; however, with the exception of Ca2+ and an adhesive protein involved in aggregation, germination, and plant attachment (Deacon & Donaldson, 1993; Reid et al., 1995; Robold & Hardham, 2005), little is known of the presence of other products and their relevance to zoospore communication. Thiazovivin mw In

contrast, the identification of autoinducers or small hormone-like molecules has provided an unparalleled insight into cell-to-cell communication and its role in the physiology, ecology, evolution, and pathogenesis Florfenicol of bacteria and a few fungal species (Winans & Bassler, 2008). The vast majority of molecules, such as acyl-homoserine lactones or oligopeptides from bacteria (Waters & Bassler, 2005), and small primary alcohols from fungi (Hogan, 2006), are species specific and used for intraspecific communication. One signal molecule called autoinducer-2 (AI-2) can be produced by half of the known bacterial population (Sun et al., 2004) and by some eukaryotic plants (Gao et al., 2003; Hauck et al., 2003), although its production has not been reported in Fungi

and Stramenopila. This molecule facilitates interspecific communication among bacteria (Xavier & Bassler, 2005). AI-2 is a collective term for a group of signal molecules derived from 4,5-dihydroxy-2,3-pentanedione (DPD) and is used interchangeably with DPD because conversion of DPD to various forms of AI-2 is a spontaneous ring closure process (Miller et al., 2004). The well-known presence of bacteria in Phytophthora and Pythium cultures and stimulation of Phytophthora zoospore and oospore production by bacterial metabolites (Zentmyer, 1965; Malajczuk, 1983) led us to hypothesize that zoosporic pathogens may produce AI-2 to communicate with bacteria. To test this, we analyzed zoospore-free fluid (ZFF) from bacterium-free and nutrient-depleted zoospore suspensions for AI-2 activity using an AI-2 bacterial reporter strain (Bassler et al.

ZFF from Phytophthora nicotianae, Phytophthora sojae, and Pythium aphanidermatum triggered luminescence of the Vibrio harve7yi AI-2 reporter, indicating the presence of AI-2 in zoospore extracellular products and the potential of cross-kingdom communication between

oomycetes and bacteria. The production of AI-2 by zoospores was confirmed by chemical assays. These results GSK3 inhibitor provide a new insight into the physiology and ecology of oomycetes. Phytophthora and Pythium in Oomycota of Stramenopila are phylogenetically related to marine algae, but resemble fungi morphologically. Many species in these two genera are destructive pathogens that attack a broad range of economically important agricultural and ornamental crops as well as forest tree species. They produce asexual sporangia that release flagellate zoospores as their primary dispersal and infection agents (Deacon & Donaldson, 1993; Judelson & Blanco, 2005). Zoospores secrete a host of molecules during the homing process; however, with the exception of Ca2+ and an adhesive protein involved in aggregation, germination, and plant attachment (Deacon & Donaldson, 1993; Reid et al., 1995; Robold & Hardham, 2005), little is known of the presence of other products and their relevance to zoospore communication. INK 128 cost In

contrast, the identification of autoinducers or small hormone-like molecules has provided an unparalleled insight into cell-to-cell communication and its role in the physiology, ecology, evolution, and pathogenesis Oxalosuccinic acid of bacteria and a few fungal species (Winans & Bassler, 2008). The vast majority of molecules, such as acyl-homoserine lactones or oligopeptides from bacteria (Waters & Bassler, 2005), and small primary alcohols from fungi (Hogan, 2006), are species specific and used for intraspecific communication. One signal molecule called autoinducer-2 (AI-2) can be produced by half of the known bacterial population (Sun et al., 2004) and by some eukaryotic plants (Gao et al., 2003; Hauck et al., 2003), although its production has not been reported in Fungi

and Stramenopila. This molecule facilitates interspecific communication among bacteria (Xavier & Bassler, 2005). AI-2 is a collective term for a group of signal molecules derived from 4,5-dihydroxy-2,3-pentanedione (DPD) and is used interchangeably with DPD because conversion of DPD to various forms of AI-2 is a spontaneous ring closure process (Miller et al., 2004). The well-known presence of bacteria in Phytophthora and Pythium cultures and stimulation of Phytophthora zoospore and oospore production by bacterial metabolites (Zentmyer, 1965; Malajczuk, 1983) led us to hypothesize that zoosporic pathogens may produce AI-2 to communicate with bacteria. To test this, we analyzed zoospore-free fluid (ZFF) from bacterium-free and nutrient-depleted zoospore suspensions for AI-2 activity using an AI-2 bacterial reporter strain (Bassler et al.

Approximately 10 L of surface sediments (depth 5–10 cm) from each site were collected in 2008, which were transferred to 20-L aquaria and overlaid with lake water (microcosms) in a laboratory. The microcosms were loosely covered and stored in dim light at room temperature without disturbance. MTB in the sediment were magnetically enriched using a double-ended open magnetic separation apparatus (MTB trap),

which could simultaneously collect both north- Natural Product Library in vitro and south-seeking MTB (Jogler et al., 2009). Specifically, about 200 mL of surface sediments from each microcosm were scratched and directly transferred to the ‘MTB trap’ (500 mL in volume). A homogeneous magnetic field, about seven times that of the Earth’s magnetic field, was applied for cell enrichment for 6 h. The retrieved MTB cells were then washed with sterile-distilled water twice and stored at−20 °C until further processing. For the microcosm MY8, MTB were collected in 2009 on 26 February (MY8a), 18 March (MY8b) and 23 April (MY8c), respectively; for the microcosm MY11, MTB were collected in 2009 on 25 February

(MY11a), 18 March (MY11b) and 24 April (MY11c), respectively. The oxygen concentrations of surface sediments in microcosms were determined using an HQ40d Oxygen Meter (HACH). Pore water was separated from the surface sediments by centrifugation at 1000 g for 20 min as described previously (Liu et al., 2003). The pH of pore water was measured using a Mettler Toledo Delta 320 pH meter. Physical–chemical analyses of various learn more anions and major cations were conducted at the Analytical Laboratory Beijing Research Institute of Uranium Geology, using a Dionex-500 chromatograph (BioPortfolio) and 785 DMP Titrino (Metrohm AG). The concentrations of total iron of pore water were measured using HR-ICP-MS (Finnigan MAT). PCR amplifications of nearly

complete 16S rRNA genes of MTB were carried out using bacterial universal primers 27F (5′-AGAGTTTGATCCTGGCTCAG-3′) and 1492R (5′-GGTTACCTTGTTACGACTT-3′) based on the previous report (Lin et al., 2008). The PCR amplification program Cetuximab manufacturer consisted of 5 min at 95 °C, 30 cycles of 1.5 min at 92 °C, 1 min at 50 °C and 2 min at 72 °C; the final extension was carried out at 72 °C for 10 min. To avoid potential sample biases, duplicate PCR products for each sample were pooled and then purified by 0.8% (w/v) agarose gel electrophoresis. PCR controls with no template were negative. Purified PCR products were cloned into the pMD19-T vector and chemically DH5α competent cells (TaKaRa) according to the manufacturer’s instruction. A total of six 16S rRNA gene clone libraries (MY8a, MY8b, MY8c, MY11a, MY11b and MY11c) were constructed. Thirty positive clones from each library were randomly selected. The cloned inserts were amplified by PCR with the primers specific for the pMD19-T vector. The PCR products were analyzed by electrophoresis in 0.8% (w/v) agarose.

We have shown previously that MH-S cells are not capable

of killing M. pulmonis unless the mycoplasma is first bound by the macrophages. Killing is dependent on phagocytosis (Shaw et al., 2012). Vsa proteins act as a shield that reduces binding to macrophages when the proteins Bleomycin supplier are long with many tandem repeats. We show here that the EPS-I polysaccharide of M. pulmonis is a second shielding factor that inhibits binding to macrophages and hence is antiphagocytic. Both long Vsa protein and EPS-I have a role in protection from complement and inhibit biofilm formation (Bolland & Dybvig, 2012; Bolland et al., 2012). The amount of EPS-I that is associated with the mycoplasma cell is about the same regardless of the length of the Vsa protein being produced (Bolland et al., 2012). It OSI-906 nmr is unknown whether the Vsa proteins and EPS-I interact directly, but it is apparent that full shielding from host defences requires both a long Vsa protein and EPS-I. The shield primarily protects mycoplasmas from macrophages by inhibiting binding, but there are indications that a maximal shield may also inhibit phagocytosis of the bound mycoplasmas. Mycoplasmas bound to MH-S cells are not phagocytosed efficiently when they produce a very long VsaA protein (Shaw et al., 2012). The relative resistance of CTG1701-C

to killing even after being bound by the macrophages (Figs 1b and 2d) suggests that the high level of EPS-I on CTG1701-C has the capability to inhibit phagocytosis. There are several possible explanations for CTG1701-C producing as much as five times more EPS-I than CTG38. CTG1701 was complemented to generate CTG1701-C by inserting the 2-gene operon containing MYPU_7410 and MYPU_7420 along with its native promoter into the mycoplasmal genome using transposon Tn4001C as the vector (Daubenspeck et al., 2009). One possibility for increased production of EPS-I is that sequences upstream of the complementing operon in CTG1701-C have promoter activity that enhances transcription above that of the native promoter alone. ADAMTS5 Alternatively, the complementing operon might be missing

regulatory sequences that are present in the native operon. Another possibility is the position of the complementing operon in the genome might enhance transcription, as has been shown for genes near the origin for DNA replication (Li et al., 2003; Manna et al., 2004). Killing of M. pulmonis by MH-S cells was only modest. Host factors absent in the in vitro assays may be required for efficient phagocytosis. We show that yeast extract enhances killing. We view it likely that the mannosylated proteins from the yeast cell wall are responsible, possibly through interactions with complement receptor 3 or the mannose receptor on the macrophages. Complement receptor 3 contains a lectin domain that is believed to bind polysaccharide and increase killing of iC3b-opsonized microorganisms (Todd, 1996).

1; Jaouen & Metzler-Nolte, 2010; Romao et al., 2012). In spite of the stable character of the CO molecule and of its binding ability being restricted to metals, CO-RMs exhibit vasodilatory, renoprotective, anti-inflammatory and anti-apoptotic properties. Moreover, CO-RM-based therapies for inflammation, sepsis, lung injury, cardiovascular

diseases, cancer and organ transplantation and preservation have been supported by preclinical studies in animals (Johnson et al., 2003; Motterlini et al., 2005; Foresti et al., 2008; Motterlini & Otterbein, 2010; Gullotta et al., 2012a). So far, those studies indicate that upon treatment with CO-RMs only a small part of the CO released is found bound to haemoglobin, as judged by the low levels of COHb present in blood (Foresti et al., 2008). Clearly, other proteins need to be targeted to Selleck Epacadostat support the action of CO-RMs; however, until they have been identified, their pharmacological usefulness is severely hindered. Studies of the effects of CO-RMs on bacteria, which will be described in the following sections, might provide a significant contribution to the implementation of CO-RMs as therapeutic drugs. In particular, studies in bacteria have already revealed how the metal affects the properties of

CO-RMs, a factor that cannot be neglected as it contributes to formation Proteasome cleavage of ROS (to be discussed). As in mammals, high concentrations of CO and CO-RMs cause the death of bacteria. These antimicrobial properties have been demonstrated for Gram-negative and Gram-positive bacteria such as Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa (Nobre et al., 2007; Desmard et al., 2009). The bactericidal concentration depends on the microorganism, its growth requirements Thymidine kinase for oxygen, and the metal present in the CO-releasing molecule. For example, for P. aeruginosa the ruthenium-based carbonyls CORM-2 and CORM-3 are more bactericidal than the manganese-containing CORM-371 (Desmard et al., 2011). A seminal study of the effect of CO

on bacteria demonstrated that both CO and CO-RMs strongly decreased the cell viability of the Gram-negative E. coli and Gram-positive S. aureus (Nobre et al., 2007). The effect observed was confirmed to be bactericidal and not simply bacteriostatic. In particular, CORM-2 and CORM-3 were demonstrated to be very efficient bacterial killers, as after 30 min of treatment between 50% and 80% of the bacteria were not viable. Furthermore, even 4 h after addition of those CO-RMs, cells were not able to resume growth (Nobre et al., 2007). ALF021 and ALF062, which contain manganese and molybdenum, respectively, also proved to reduce the viability of the two pathogens. In all cases, supplementation with haemoglobin, a CO scavenger, abolished the bactericidal effect.

Attitudinal questions about the role of pharmacy in the provision of CAM advice revealed that whilst only 11% of respondents reported their pharmacist aware of their use of CAM, and 52% agreed it important their pharmacist knowledgeable about CAM, only 25% felt their pharmacist currently to be a useful source of information. However, 55% reported they would use their pharmacist as a preferred source of information about CAM if they felt them more knowledgeable. 49% thought their pharmacist ought to be the most reliable source of information about

safety of CAMs and interactions with medication. However, 45% used their family and friends as their primary source of information about CAM. The results concur with Australian and Canadian studies that report customers expect pharmacists to be knowledgeable about CAMs Stem Cell Compound Library nmr and provide an advisory role to help them assess information and communicate guidance about safety issues (2). However, whilst the study demonstrates many UK

customers http://www.selleckchem.com/products/BIBW2992.html expect their pharmacist to be knowledgeable about CAM and believe they should be a source of reliable safety information and advice regarding possible interactions with medications, they feel that there is a lack of understanding within the profession on the subject. This pilot investigation demonstrates the need for a larger scale study to better understand consumer’s more general and specific needs in greater detail together with a Forskolin ic50 parallel assessment of the requirements of community pharmacy to meet any identified demands in the context of an evidence based scenario. One place to begin to enhance the provision of good quality advice regarding CAM products may

be through the provision of CPD pharmacy training. 1. Cramer H. et al. Over the counter advice seeking about complementary and alternative medicines (CAM) in community pharmacies and health shops: an ethnographic study. Health and Social Care in the Community 2010; 18: 41–50. 2. Kwan D et al. Exploring consumer and pharmacist views on the professional role of the pharmacist with respect to natural health products: a study of focus groups. BMC Complement Altern Med 2008; 8: 40. Rebecca Dickinson1, DK Raynor1, Peter Knapp2, Jan MacDonald3 1University of Leeds, Leeds, UK, 2University of York, York, UK, 3Medicine and Healthcare products Regulatory Agency, London, UK This objective of this study was to explore whether patients use a headline section in a patient information leaflet to find key information about their medicines in a user-test. Quantitative findings showed the headline was used for 55/140 opportunities (39%), and qualitative findings suggested the headline was viewed as a positive inclusion. The headline section was only used just over a third of the time, but its inclusion was viewed as a valuable addition. European legislation requires a PIL be provided with each licensed medicine.

Recall bias may have also affected the responses since this is a retrospective study. 1. Latif A, Pollock

K, Boardman HF. The contribution of the Medicines Use Review (MUR) consultation to counseling practice in community pharmacies. Patient Education and Counseling. 2011; 83: 336–344. 2. Al-Nagar A, Constantine D, Thayaparan J, De-La-Mare N, Desborough J. Views and experiences of community pharmacists about consultation skills training: a national survey. International Journal of Pharmacy Practice 2012; 20 (Suppl. 2): 3–30. 3. Martin BA, Bruskiewitz RH, Chewning BA. Effect of a tobacco cessation continuing professional education program click here on pharmacists’ confidence, skills, and practice-change behaviors. Journal of the American Pharmacists Association: JAPhA 2010; 50: 9. Adam Todd1, Hamde Nazar2, Inga Andrew3, Lisa Baker3, selleck screening library Andy Husband1 1Durham University, Stockton-on-Tees, UK, 2University of Sunderland, Sunderland, UK, 3St. Benedict’s Hospice, Sunderland, UK Polypharmacy is common amongst patients with limited life expectancy; Prescribing of inappropriate medicines for patients with limited life expectancy can lead to multiple drug interactions of varying severity; Patients with limited life expectancy should have their medicines reviewed in line

with the original therapeutic goals. For patients with limited life expectancy – typically surviving for less than one year from diagnosis – polypharmacy is common as medication is prescribed to manage both life limiting illness and to treat

or prevent other long-term conditions. Consequently, there is an increased risk of developing drug-related toxicity resulting Immune system from drug-drug or drug-disease interactions. The aim of this work was to assess the prevalence of inappropriate medication and identify any potential theoretical drug-drug interactions in patients attending a specialist palliative care unit. This was a prospective study that examined medication and medical histories for patients attending a specialist palliative care day care centre from November 2012 until March 2013. Medication was assessed for appropriateness using a conceptual framework, which considers remaining life expectancy of the patient, time until benefit of the treatment, goals of care and treatment targets.1 Consensus was reached via Delphi methodology using a range of clinical pharmacists and consultants in palliative medicine; to reach consensus agreement was required from all panel members. Drug interactions were identified and assessed according to significance using the drug interaction recognition software, Proscript®. Drug interactions identified as significant were further sub-classified as moderate or severe based upon the potential to cause harm or hospitalisation, if they were reversible or irreversible and, if any treatment would be required to manage the outcome.