Persistence of sulfadoxine–pyrimethamine (SP) resistance has been described in an urban setting in Malawi where malaria transmission is relatively low. Higher malaria transmission is associated with greater genetic diversity and more frequent genetic recombination, which could lead to a more rapid re-emergence of SP-sensitive parasites, as well as more rapid degradation of selective sweeps. In this study, the impact of local variation in malaria transmission on the prevalence of SP-resistant haplotypes and selective sweep characteristics was investigated at an urban site with low parasite prevalence and two rural sites with moderate and high parasite prevalence.pfdhfr-ts and pfdhps and at microsatellites flanking these genes. Expected heterozygosity (He) was estimated to evaluate genetic diversity.Samples from three sites with different parasite prevalence were genotyped for resistance markers within e flanking pfdhfr-ts and pfdhps were seen between rural-moderate and the other sites, as well as some shared haplotypes between the rural-high and urban-low sites.No difference in the prevalence of highly resistant DHFR 51I/59R/108N and DHPS 437G/540E was found between sites. Small differences in HThe results do not show an effect of local variation in malaria transmission, as inferred from parasite prevalence, on SP-resistant haplotype prevalence.The online version of this article (doi:10.1186/s12936-015-0860-7) contains supplementary material, which is available to authorized users. Plasmodium falciparum, from Asia to Africa, has heightened interest in identifying factors affecting resistance allele dynamics, including the re-emergence of drug-sensitive malaria parasites. Chloroquine and sulfadoxine–pyrimethamine (SP) were both once used as safe and effective primary treatment for uncomplicated malaria. Successive waves of anti-malarial resistance from Southeast Asia prompted public health organizations to abandon chloroquine, and then SP, in favour of artemisinin-based combination therapy (ACT).The potential expansion of artemisinin-resistant After the removal of chloroquine drug pressure in Malawi in 1993, chloroquine-sensitive parasites re-expanded in the population, eventually outcompeting the chloroquine-resistant strains . This reIn late 2007, Malawi replaced SP with an ACT in response to failing SP efficacy. The persistence of the highly resistant haplotypes of dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS), DHFR 51I/59R/108N and DHPS 437G/540E, 5 years after the reduction of SP drug pressure, was recently demonstrated by the authors, with discussion of possible causes and implications of continued resistance [The effect of reduction of SP use on SP-resistant parasites has been variable in other regions. While the persistence of SP-resistant parasites in Venezuela was shown years after the removal of drug pressure , a studyMalaria parasites from areas of high transmission tend to be more diverse and undergo more recombination , 9, 10. P. falciparum parasites. Recent entomological inoculation rates (EIR) were not available for these sites, but the use of parasite prevalence as a surrogate for EIR and transmission intensity is supported in previous studies [Samples were collected as part of a malaria surveillance study conducted in health facilities in three districts in Malawi: a rural site near a river with high parasite prevalence (Chikwawa), a rural more arid site with moderate parasite prevalence (Thyolo) and an urban site with low parasite prevalence (Ndirande). Samples were collected from individuals presenting to the hospital with uncomplicated malaria. All samples were collected with informed consent according to an institutional review board approved protocol. Samples consisted of blood spots, collected on filter papers, representing a patient’s initial infection on the day they were admitted to the study, prior to treatment. Parasite prevalence from a community-based, cross-sectional survey was used as a surrogate measure of transmission intensity; 8.4, 14.3 and 29.6 % of individuals surveyed in the urban-low, rural-moderate and rural-high settings, respectively, were qPCR-positive for  studies .pfdhfr-ts and pfdhps genes were determined via pyrosequencing. Single nucleotide polymorphisms (SNPs) within codons 51, 59 and 108, of pfdhfr-ts and codons 437, 540 and 581 of pfdhps were genotyped for all samples using primers and amplification methods adapted from Zhou et al. [DNA was extracted from filter paper blood cards using a Qiagen BioRobot  following the Investigator Bloodcard Protocol. Parasite genotypes at polymorphic sites within u et al. . Pyrosequ et al. . An alleP. falciparum genome [To show that reduced heterozygosity around drug resistance genes was the result of selection rather than demographic processes, expected heterozygosity was measured in six unlinked neutral loci located throughout the m genome . These um genome . When mum genome , 17. Onlpfdhfr-ts were genotyped; four downstream , and four upstream  using previously described primers and protocols [pfdhps were genotyped; four downstream , and four upstream  of the gene, using primers described by Vinayak et al. [Eight polymorphic microsatellites flanking rotocols , 18. Eigk et al. . FragmenThe prevalence of each haplotype was estimated as the number of each haplotype observed among the successfully genotyped samples for each resistance gene, divided by the total number of genotyped samples. Haplotype frequency, the proportion of parasites with a given haplotype, could not be calculated as the number of parasite clones per infection was not estimated. A resistant haplotype was defined as containing any number or combination of resistance alleles at the genotyped codons within either of the resistance genes of interest. The sensitive haplotype was defined as parasites with sensitive alleles at all codons within both genes. Samples with mixed-genotype at two or more loci within the same gene were excluded because haplotype phase could not be determined. Chi squared tests with Yate’s correction were used, where appropriate.e), a measure of genetic diversity at each microsatellite locus, was calculated using the standard equations for He and variance: Expected heterozygosity (He (±1 standard deviation) was calculated for three groups: rural high, rural-moderate and urban-low sites, for both pfdhfr-ts and pfdhps genes. Samples without a predominant genotype or samples with missing data were excluded from expected heterozygosity calculations. Statistical significance was determined via permutation. Diversity ratios were calculated for Hurban-low/Hrural-high, Hurban-low/Hrural-moderate, and Hrural-moderate/Hrural-high. Calculations for He, standard deviation and permutations were conducted in R [The analysis focused on sweep characteristics flanking DHFR 51I/59R/108N and DHPS 437G/540E due to limited prevalence of other haplotypes in all three settings. Hted in R .pfdhfr-ts haplotypes were assembled for 549 samples from the urban-low site, 726 samples from the rural-moderate site, and 660 samples from the rural-high site , , rural-high site , , and urban-low site  and . Average He at unlinked microsatellites was 0.898. The proportion of samples scored as polyclonal based on unlinked microsatellites did not differ significantly between study sites (p = 0.611). None of the sites had He values that were consistently different from each other  though no differences between the rural-high and urban-low sites were found. Analysis of flanking microsatellites indicated the presence of a core haplotype flanking pfdhfr-ts and pfdhps, likely of Southeast Asian origin, in all three sites , n = 1, pfdhps5, ts, n ons Fig. .Fig. 2SuThis study estimates the effect of local differences in malaria transmission, as inferred from parasite prevalence, on the prevalence of SP-resistant haplotypes and the characteristics of associated selective sweeps within Malawi. Despite marked differences in the parasite prevalence between the rural-high and urban low sites, no difference in haplotype prevalence or sweep characteristics was observed. Statistically significant differences between the prevalence of DHFR double mutants in the rural-moderate and rural-high sites are possibly a statistical artifact due to large sample size and are not considered clinically relevant.This study also indicated similarity in sweep characteristics and some shared microsatellite haplotypes between the rural-high and urban-low sites, counter to what would be expected under the assumption that higher transmission would result in greater genetic diversity. These findings may indicate a shared parasite population between these locations. The divergence of some sweep characteristics and microsatellite haplotypes in the rural-moderate site compared to the other two sites, suggests that regional demographic events may affect sweep characteristics, without a significant effect on haplotype prevalence. Such events could have been the migration of SP-resistant haplotypes of diverse genetic background to the rural-moderate site (perhaps from neighbouring Mozambique).The similarity in the proportion of samples scored as polyclonal between study sites may indicate similarity in the level of malaria transmission, not reflected in the parasite prevalence observed at each site. Recent EIRs were unavailable for these study sites; therefore, parasite prevalence was used as a surrogate for transmission intensity. If parasite prevalence was an accurate estimation of transmission intensity in this study, a larger proportion of polyclonal samples in the higher transmission sites would have been expected. This discrepancy could indicate that parasite prevalence is not a good surrogate for transmission intensity in this study. The estimation of transmission intensity used here was based on average parasite prevalence from a community-based, district-wide, cross-sectional survey, while the data-set used in this study was based on a facility-based survey. The cross-sectional data provide a single estimate of parasite prevalence at a given point in time, and may not reflect historical differences in transmission intensity between the sites. It is possible that individuals reporting to the district health centres in this study may have been from areas with more similar malaria transmission than the district-wide average parasite prevalence suggests. It is also possible that the almost three-fold difference in parasite prevalence between these sites is not sufficient to produce a measurable difference in the proportion of polyclonal infections, haplotype prevalence, or sweep characteristics.pfdhfr-ts haplotype based on similarity in microsatellite repeat length between Malawian parasites and the V1S control strain. While, the Malawian pfdhps haplotype in this study differed at several markers from the V1S strain, core markers (those in close proximity to pfdhps) were similar to those published by Mita et al., and Alam et al. showing similar core haplotypes found in Malawi, Bangladesh, Thailand and Cambodia [pfdhps resistance allele in Malawi.The data also indicate a Southeast Asian origin of the Malawian Cambodia , 22. TheThis research presents evidence that after nearly two decades of high SP resistance in Malawi, differences in regional malaria transmission, as inferred from parasite prevalence, do not impact the prevalence of SP-resistant parasite haplotypes. The SP-resistant haplotypes, DHFR 51I/59R/108N and DHPS 437G/540E, have maintained their high prevalence long after the removal of SP as the first-line treatment in multiple settings in Malawi. Investigators working in epidemiologically similar settings can reasonably assume similarity in SP-resistant haplotype prevalence and sweep characteristics between local sites with different parasite prevalence.


Little is known about the individual factors that predict outcomes in Internet-administered psychological treatments. We hypothesized that greater cognitive flexibility  would provide a better foundation for learning and employing the cognitive restructuring techniques taught and exercised in therapy, leading to greater treatment gains. Participants in three trials featuring Internet-administered psychological treatments for depression (n = 36), social anxiety disorder (n = 115) and tinnitus (n = 53) completed the 64-card Wisconsin Card Sorting Test (WCST) prior to treatment. We found no significant associations between perseverative errors on the WCST and treatment gains in any group. We also found low accuracy in the classification of treatment responders. We conclude that lower cognitive flexibility, as captured by perseverative errors on the WCST, should not impede successful outcomes in Internet-delivered psychological treatments. Mounting evidence suggests that Internet-delivered psychological treatment programs based on cognitive behavioral therapy (CBT)  are effeCognitive flexibility (CF), an executive function, refers to the ability to simultaneously consider several concepts and tasks, and switch effortlessly between them in response to changes in environmental contingencies . The conNo previous study has investigated whether CF predicts outcomes in Internet-delivered CBT, which despite sharing many similarities with traditional CBT, has some inherent characteristics that we reasoned should put a greater demand on CF abilities. In most Internet interventions, psychoeducation, cognitive and behavioral exercises and other therapeutic components cannot be customized by a therapist according to the patient’s unique situation to the same degree as in traditional therapy. Few Internet interventions feature live therapist contact, meaning that it is not possible for the therapist and the patient to work together on a cognitive restructuring task. Hence, successfully completing Internet interventions likely requires a greater ability to not only comprehend the delivered therapeutic material, but also to identify emotional, cognitive and behavioral patterns entirely independently or with very little therapist assistance. Although cognitive restructuring has traditionally been considered a therapeutic technique specific to CBT, CF may also be important in psychodynamic therapy, where the goal is to explore, identify and change unconscious patterns of cognitions and emotions . Little The 64-card Wisconsin Card Sorting Test (WCST)  is an esData for the current study was collected as part of three randomized trials: therapist-guided CBT for social anxiety disorder (SAD) , psychodF = 35.18, p < .001), driven by SAD < MDD (p < .001) and SAD < tinnitus (p < .001) pair-wise differences (Bonferroni-corrected p < .05). Groups were not matched on sex (Fisher’s exact test p = .0014), but because there was no sex-difference in any WCST score , this was not considered a confounding variable. On raw WCST metrics, groups differed only on Total completion time; however, there were no differences between groups in age-adjusted WCST metrics .Because the aim of the current study was not to evaluate treatment efficacy, analyses were conducted per protocol: participants classified as treatment dropouts (including those with zero completed treatment modules) and/or had incomplete post-treatment data were not included in analyses. One additional participant in the SAD group with a WCST total completion time of 5170 s  was excluded due to presumed technical issues with the online WCST administration. Final sample sizes for the SAD group was n = 115 , n = 83 for the tinnitus group , and n = 73 for the MDD group (n = 36 received treatment). Pre- and post-treatment symptom scores on the primary outcome measure in each study, along with WCST metrics, age and sex for each subject, were compiled see . The totIn the MDD and tinnitus samples, WCST and pre-treatment (screening) data was collected prior to any treatment commencing, and post-treatment data soon afterwards. In the SAD sample, WCST data and pre-treatment symptom scores were collected after two weeks of initial attention bias modification, and post-treatment symptoms scores after a nine-week CBT program. The two forms of attention bias modification used produced similar results , and werAn online, in-house developed version of the standard, single-deck version of the WCST  featurinWe assessed the psychometric validity of online WCST by performing a principal component analysis (PCA) to estimate the latent structure of our results and comparing with past findings. A composite two-factor structure has been proposed, derived from a meta-analysis of ten factor analyses in four studies featuring neurological, senior and/or healthy samples . The proF = 15.03, p < .001), such that PSRMDD > PSRTinnitus > PSRSAD .The primary outcome measures in the studies were the Liebowitz Social Anxiety Scale—Self-Report version , the BecWe used two methods to test our hypothesis that cognitive flexibility is associated with treatment gains. First, age-adjusted PE was entered as a predictor of raw pre-treatment symptom scores in standard linear regression models, group-wise. Then, number of PE was used to predict PSR in the full sample and group-wise, and pre-post raw score difference group-wise. These analyses were repeated featuring percentage PE of total errors.Second, performance of the PE metric in classifying treatment responders vs. low-responders was investigated using receiver-operating characteristics (ROC). Symptom reductions of 25%, 50% and 75% were used as separate thresholds for treatment response. Analyses were conducted both group-wise and featuring the full sample. Bootstrapped 95% confidence intervals for area under the curve (AUC) for each classification curve were used to test whether classification accuracy differed from chance. All analyses were conducted in the R (3.2) statistical environment, using the pROC  and psycp = .065). However, inspection of the scatter plot revealed two PSR outliers more than three standard deviations from the group mean. If these were removed, the correlation was no longer approaching significance , there was only one instance when the AUC confidence interval did not cover 0.5. In this case, in the SAD group using the 75% PSR cut-off, only n = 2 scored above the PSR cut-off, leading to an irregular curve shape and a confidence interval ranging 0.51–0.81  should not be an obstacle in achieving symptom reduction after Internet-delivered psychological treatment, and that pre-treatment measurements of CF will not predict treatment outcomes. The lack of predictive power is congruent with there being no linear correlation between pre-treatment symptom measures and number of PE.In the context of traditional psychotherapy, findings on associations between CF and cognitive restructuring skill acquisition, considered important for improvement, have been mixed . We reasWhile lower CF is often reported in psychiatric samples, comparably little is known about the precise relationship between CF and psychiatric symptoms. It is noteworthy that even direct training of executive functions (including CF) appears to have little or no generalized effect beyond improved performance on the specific task trained . RecentlThe strengths of our study include a relatively large, well-powered and diverse sample drawn from three different studies on effective treatments of two common psychiatric disorders and one psychosomatic disorder. To control for differences in sample and treatment characteristics, we performed analyses using both the entire sample and the individual groups. Although originally designed to measure the cognitive effects of frontal lobe damage, the WCST is considered a valid and established measure of PE. We age-adjusted PE scores, and re-running analyses using self-normalized number of PE  did not change findings. Our study design also has some limitations. Primarily, only one measure of CF, number of perseverative errors on the WCST, was used for analyses. While we acknowledge that a composite measure calculated using results from several CF-indexing neuropsychological tests would have been preferable, past studies have reported associations between this particular measure (number of PE on the WCST) and cognitive restructuring abilities . Second,Contrary to our hypothesis, we found no generic or treatment-specific predicative power of cognitive flexibility (perseverative errors on the WCST) on patient improvement following Internet-delivered psychological treatment for social anxiety disorder, depression or tinnitus. Hence, comparably lower cognitive flexibility, as measured by the perseverative errors on the WCST, should not impede successful outcomes in Internet-delivered psychological treatments.10.7717/peerj.1934/supp-1Supplemental Information 1Click here for additional data file.10.7717/peerj.1934/supp-2Supplemental Information 2Click here for additional data file.


Background: Neurological complications are common after orthotopic liver transplantation (OLT). We aimed to characterize the risk factors associated with neurological complications and mortality among patients who underwent OLT in the post-model for end-stage liver disease (MELD) era.Methods: In a retrospective review, we evaluated 227 consecutive patients at the Keck Hospital of the University of Southern California before and after OLT to define the type and frequency of and risk factors for neurological complications and mortality.Results: Neurological complications were common (n = 98), with encephalopathy being most frequent (56.8%), followed by tremor (26.5%), hallucinations (11.2%), and seizure (8.2%). Factors associated with neurological complications after OLT included preoperative dialysis, hepatorenal syndrome, renal insufficiency, intra-operative dialysis, preoperative encephalopathy, preoperative mechanical ventilation, and infection. Preoperative infection was an independent predictor of neurological complications . One-month mortality was 8.8% and was independently associated with urgent re-transplant, preoperative intubation, and intra-operative arrhythmia.Conclusion: Neurological complications are common in patients undergoing OLT in the post-MELD era, with encephalopathy being most frequent. An improved understanding of the risk factors related to both neurological complications and one-month mortality post-transplantation can better guide perioperative care and help improve outcomes among OLT patients. Orthotopic liver transplantation (OLT) is the only definitive treatment for end-stage liver disease and is being performed with increasing frequency in the United States . Post-opDefining the risk factors for neurological complications and one-month mortality after OLT can help identify high-risk patients and guide perioperative care with the intention to minimize neurological complications, morbidity, and mortality. Identifying primary risk factors is particularly relevant given the high acuity recipients in the post-model for end-stage liver disease (MELD) era, as the MELD system prioritizes the allocation of organs to patients with the most severe liver disease . StudiesWhile complications after OLT have been described previously , risk faWe reviewed the hospital records of 227 consecutive cases of adults who underwent OLT between July 18, 2003 and August 19, 2006 at the Keck Hospital of the University of Southern California. A standardized form was used to collect demographic, historical, surgical, laboratory and radiological data. Data was collected for the period beginning one week prior to transplant and extending to one month post-transplant. The Institutional Review Board approved the study.Transplants were characterized as partial living donor or cadaveric and as first transplant, urgent re-transplant , non-urgent re-transplant , transfusion free, and simultaneous liver-kidney transplant. MELD scores were calculated on the day of transplant and included exception points where applicable in cases of hepatocellular carcinoma. Operating room records were reviewed for intra-operative events, surgery duration, pressors used, and hypotension (defined as systolic blood pressure < 90 mmHg). For re-transplants, preoperative data, labs, and intra-operative records from the first transplant were recorded. Neurological complications were identified by chart review by the study’s investigators, including resident physicians and attending physicians in the Department of Neurology. These complications were then confirmed by the study’s multiple board certified neurologist investigators. These complications were defined as new neurological problems or a worsening of known neurological disorders in one month after OLT. Neurological complications were further characterized by pre-specified type and date of onset relative to OLT.p-value <0.05 threshold for significance. Using a p <0.2 threshold for significance, multivariate logistic regression analysis was performed controlling for potential risk factors for neurologic complications after OLT; these included medical co-morbidities such as dialysis and renal insufficiency, preoperative conditions such as encephalopathy and infection, and intra-operative events such as dialysis. Six cases were excluded from this analysis due to intra-operative deaths. A similar multivariate regression analysis was performed controlling for potential risk factors for one-month mortality following OLT, such as race, transplant type, medical co-morbidities, preoperative conditions and intra-operative events, again using a p < 0.2 threshold.An analysis was done to determine whether neurological complications post-OLT were associated with one-month mortality post-OLT. Univariate analyses of mortality and neurological complications after OLT were performed using Fisher’s Exact Test and Wilcoxon Two Sample Test where appropriate, using a We identified 228 consecutive adult patients who underwent OLT. One patient record could not be found, thus 227 patients were included in the study. Demographic factors are listed in Table The one-month incidence of neurological complications was 44.3% 98/221). Sixty-six patients had one neurological complication, 27 had two complications, and five had three complications. The most common complication was encephalopathy (53.06%), followed by tremor (27.55%) and seizure (8.16%). Neurological complications among patients during the first post-operative month following OLT are summarized in Table 21. Sixtyp = 0.06). Medical co-morbidities, such as hepatorenal syndrome , renal insufficiency , and intra-operative dialysis  were also risk factors associated with a greater rate of neurological complications. Furthermore, preoperative use of pressors , intubation , and encephalopathy  were also factors associated with increased neurological complications based on the results of the univariate analysis.Univariate analysis of risk factors for neurological complications is summarized in Tables On multivariate analysis, preoperative infection was the only independent predictor of neurological complications . There were a total of 50 patients who had preoperative infection, of which 32 (64%) had neurological complications. The most common neurological complications in this group were encephalopathy (56.25%) and tremor (21.88%). Exclusion of intra-operative deaths did not change the study results.p = 0.181). Of the 20 deaths, six occurred intraoperatively (30%). Characteristics of the 14 mortalities in the study population are listed in Table Overall, one-month mortality was 8.8%. The MELD score was greater in the mortality group, but not statistically significant , preoperative intubation , and intraoperative arrhythmia .The incidence of post-operative neurological complications after OLT in this study 44%) was comparable to the incidence reported in the literature of approximately 10–47% . Previou4% was coOur results revealed high rates of neurological complications and mortality after OLT in the post-MELD era. These high rates of morbidity and mortality may reflect the selection of candidates with more advanced liver disease. The study population had a mean MELD score of 22, consistent with the reported mean MELD of 24 in the United Network for Organ Sharing (UNOS) MELD 2003–2006 database .With improved survival after OLT due to better perioperative care, operative techniques, and immune suppressive therapy, there has been increasing awareness of neurological complications . The maiThe primary aim of this investigation was to evaluate hypothesis-generating risk factors associated with post-operative neurological complications. We found that pre-operative infection was an independent predictor of such complications. Our study documented any infection in the week prior to surgery, including bacterial peritonitis and urinary tract infections. None of the patients who underwent OLT were septic at the time of surgery. Therefore, it appears that even mild or focal infections may be associated with an increased susceptibility to neurological events. This association is biologically plausible, as bacteria, viruses, and fungi can spread to the central nervous system just as they affect the abdomen, blood stream, respiratory tract, and other organ systems . IncreasIn our analyses of hypothesis-generating risk factors for one-month mortality after OLT, re-transplantation within the first month was independently associated with one-month mortality. Urgent liver re-transplant is necessary when complications such as primary non-function or vascular thrombosis cause the allograft to fail . Our morPre-operative intubation also was an independent predictor of one-month mortality, a finding that has not been reported in prior studies of complications after OLT. One study has demonstrated that among patients who underwent emergency or elective living-donor liver transplantation (LDLT), those who underwent emergency LDLT had higher rates of pre-operative intubation, suggesting greater pre-transplant acuity and possibly poorer post-transplant outcome . FurtherWhile prior study populations were predominantly Caucasian, our study population was predominantly Hispanic. Studies have demonstrated poorer outcomes in minority populations, particularly African Americans . Some reThe main limitation of this investigation is its retrospective design. It is possible that not all neurological findings were documented in the patient charts, resulting in an underestimation of some results. Our one-month follow-up may have missed neurological complications that occurred later in the post-operative course. Nevertheless, the findings contribute to a growing body of literature that examines neurological complications after OLT , 19, 28.In summary, neurological complications after OLT are common in the post-MELD era, with encephalopathy being the most common complication. Infection was independently associated with the development of post-operative neurological complications, suggesting that these complications may be reduced by treating infection during the preoperative period. In addition, urgent re-transplant, preoperative intubation, and intraoperative arrhythmias were independent predictors of one-month mortality after OLT. Identifying hypothesis-generating risk factors for both the development of neurological complications and one-month mortality supports a better understanding of the population most at risk for adverse outcomes after transplantation.Future studies might investigate the role of infection in neurological complications to better determine the pathophysiology involved in conditions such as encephalopathy and ischemic stroke among OLT patients. Similarly, factors this investigation revealed to be associated with one-month mortality after OLT, such as intraoperative arrhythmias, have been less well-characterized in the literature and warrant further investigation to clarify their impact on outcomes. An improved understanding of these hypothesis-generating risk factors for neurological complications and one-month mortality after liver transplantation ultimately may reduce adverse outcomes and improve graft and patient survival.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Selenium (Se) and sulfur (S) are closely related elements that exhibit similar chemical properties. Some genes related to S metabolism are also involved in Se utilization in many organisms. However, the evolutionary relationship between the two utilization traits is unclear.In this study, we conducted a comparative analysis of the selenophosphate synthetase (SelD) family, a key protein for all known Se utilization traits, in all sequenced archaea. Our search showed a very limited distribution of SelD and Se utilization in this kingdom. Interestingly, a SelD-like protein was detected in two orders of Crenarchaeota: Sulfolobales and Thermoproteales. Sequence and phylogenetic analyses revealed that SelD-like protein contains the same domain and conserved functional residues as those of SelD, and might be involved in S metabolism in these S-reducing organisms. Further genome-wide analysis of patterns of gene occurrence in different thermoproteales suggested that several genes, including SirA-like, Prx-like and adenylylsulfate reductase, were strongly related to SelD-like gene. Based on these findings, we proposed a simple model wherein SelD-like may play an important role in the biosynthesis of certain thiophosphate compound.Our data suggest novel genes involved in S metabolism in hyperthermophilic S-reducing archaea, and may provide a new window for understanding the complex relationship between Se and S metabolism in archaea.The online version of this article (doi:10.1186/1471-2164-15-908) contains supplementary material, which is available to authorized users. Escherichia coli[[Ser]Sec (SelC) and selenophosphate synthetase (SelD). In archaea and eukaryotes, substitute/additional components are needed for Sec biosynthesis, such as O-phosphoseryl-tRNA[Ser]Sec kinase (PSTK), archaeal/eukaryotic Sec synthase (SecS), SECIS-binding protein 2 (SBP2) and ribosomal protein L30 [Selenium (Se) is an important micronutrient in all three domains of life . The majhia coli, 8–10. Stein L30 , 11, 12.tein L30 –16. Spectein L30  and SelDAmong known Se-related genes, SelD is considered as a key gene which is essential for all known Se utilization traits. It is a member of a small ATP-binding superfamily that also includes the purine biosynthetic enzymes PurM (phosphoribosylaminoimidazole synthetase) and PurL (formylglycinamide ribonucleotide amidotransferase), thiamine-monophosphate kinase (ThiL) and nickel-iron hydrogenase maturation protein HypE . The funOn the other hand, Se and sulfur (S) are two closely related elements utilized for a vast array of biochemical reactions . It has In this study, we carried out a comparative and phylogenetic analysis of SelD family in archaea based on the rapidly increased genomes sequenced in this kingdom. Occurrence of SelDs and their homologs were identified, which revealed a limited utilization of Se in archaea. In addition, we found a new SelD-like family in several hyperthermophilic S-utilizing archaea, which contains the same catalytic Cys residue as those in SelD proteins. Phylogenetic and genome context analyses of SelD-like genes revealed that they are more likely to share a common ancestor with SelD genes and may participate in S metabolism, probably involved in the biosynthesis of certain thiophosphate compound in these organisms. Further bioinformatics approaches were used to analyze patterns of gene occurrence, and several genes that may co-occur with SelD-like gene were identified, implying functional links among them. Thus, our results provide new insights into the complex and dynamic relationship between Se and S metabolism in S-utilizing archaea.In the recent decade, comparative genomics has become an important strategy for understanding the evolutionary dynamics of a variety of cellular pathways in different organisms. Several comparative studies on Se have been carried out to examine Se utilization traits and identify new genes involved in Se metabolism in prokaryotes and eukaryotes , 26–28. In this study, we analyzed the distribution of both SelD gene and the three known Se utilization pathways in different archaeal taxa , was found to have an orphan SelD gene and lack components of any known Se utilization trait. The SelD protein in this organism is a typical SelD which is highly similar to other orthologs  and contains a conserved Cys residue in the active site. In a separate study, orphan SelD genes were also found in some newly sequenced bacteria, implying an additional but unknown SelD-dependent use of Se (unpublished data).Surprisingly, a newly sequenced archaeon, M. jannaschii SelD and Sulfolobus acidocaldarius SelD-like). However, these SelD-like proteins contain the same domain as SelD (COG0709) and the conserved Cys residue corresponding to the catalytic Sec/Cys in the active site of SelD, both of which are not detected in other families of the ATP-binding superfamily that contains SelD  were exclusively found in two orders of the phylum Crenarchaeota: Sulfolobales and Thermoproteales, in which all sequenced organisms do not have any known Se utilization trait  residues may coordinate metal ions  to bind the phosphate groups of ATP [E. coli SelD also confirmed that these Asp residues which are involved in the binding of magnesium at the active site to allow the interaction with ATP, and a conserved asparagine (Asn87 in E. coli SelD), play an essential role for catalysis in E. coli SelD [Recently, the crystal structures of SelD have been solved in several bacteria, which provide important basis for understanding the catalytic mechanism of SelD-mediated selenophosphate synthesis. For example, based on the crystal structures of s of ATP . In addioli SelD . InteresConsidering that SelD belongs to an ATP-binding superfamily which also includes PurM, PurL, ThiL and HypE families, we collected proteins from representative organisms for each of them. Phylogenetic analysis revealed that SelD-like proteins are different from SelD family and form a separate branch of this superfamily Figure . The evoAs mentioned above, genes encoding SelD-like proteins were only detected in Sulfolobales and Thermoproteales; however, Desulfurococcales, the third order of Crenarchaeota, lack this gene. This may suggest that SelD-like protein evolved in the ancestor of crenarchaeota and lost in desulfurococcales. To further study the possible function of SelD-like proteins, we examined genes that are located within 10,000 nucleotides upstream and downstream of SelD-like gene in all organisms containing this gene. It has been known for some time that many clustered genes, especially those organized in one operon, may be functionally related . Thus, iS. solfataricus and Pyrococcus horikoshii have revealed that normally folded acylphosphatases retain a significant tendency to form amyloid fibrils through a direct assembly of monomers in their native-like topology [First, all SelD-like proteins in Thermoproteales were found to be fused with an acylphosphatase-like domain. Acylphosphatase belongs to a large protein family (COG1254) that specifically catalyzes the hydrolysis of the carboxyl-phosphate bond of acylphosphate substrates such as carbamoylphosphate and 1,3-biphosphoglycerate, and results in the production of carboxylate and phosphate . It is wtopology , 35. TheA second interesting finding is that several common genes were found to be localized very close to SelD-like genes in both sulfolobales and thermoproteales, including genes encoding SirA-like (pfam01206) and peroxiredoxin(Prx)-like (COG2210) proteins. The SirA-like protein is considered as a S transfer protein that belongs to a predicted redox protein superfamily and has been found to be associated with SelD . HomologAlthough there is no clear evidence with regard to the function of SelD-like, we could always find that genes associated with S metabolic pathways are located nearby. Besides SirA-like and Prx-like, additional S-related genes were also observed in almost all sequenced crenarchaeota containing SelD-like, such as rhodanese-related sulfurtransferase PspE, dissimilatory sulfite reductase (DSR) and Fe-S oxidoreductases  reductase, was found to be strongly related to SelD-like gene. This enzyme contains two subunits (AprA and AprB), which participates in S metabolism and catalyzes the reduction of APS to sulfite and AMP . HoweverAnother related gene is a predicted transporter that contains three transmembrane regions. It belongs to a large and diverse group of transporters , which includes uniporters, symporters, and antiporters of a variety of substrates such as ions, sugar phosphates, drugs, nucleosides, amino acids, and peptides. This gene is found in all sulfolobales and many other prokaryotes. However, its function is unclear. Further studies are needed to verify its biological role and the relationship with SelD-like gene.It has been reported that many S-reducing archaea carries out sulfate reduction via the pathway originally proposed for bacterial species , 39. AftBased on the results described above, the SelD-like protein may play a key role in the biosynthesis of thiophosphate (P=S) compound in sulfolobales and thermoproteales. In this study, two possible models were suggested Figure . In ModeCys biosynthesis based on a pathway similar to Sec biosynthesis. It has been known that cysteinyl-tRNACys is typically synthesized by cysteinyl-tRNA synthetase (CysS). In methanogenic archaea that lack the CysS gene, cysteinyl-tRNACys is synthesized by O-phosphoseryl-tRNA synthetase (SepS) that acylates tRNACys with phosphoserine (Sep), and Sep-tRNA: Cys-tRNA synthase (SepCysS) that converts Sep-tRNACys to Cys-tRNACys[An interesting hypothesis is that SelD-like protein might be involved in cysteinyl-tRNA-tRNACys. One posIn this study, we carried out comparative and phylogenetic studies of SelD family in all sequenced archaea. Our data revealed restricted occurrence of SelD as well as limited utilization of Se in this kingdom. In addition, a distant homolog of SelD (SelD-like) was found to form a separate family in hyperthermophilic S-utilizing Crenarchaeota: Sulfolobales and Thermoproteales. The SelD-like protein has the same catalytic Cys as well as other functional residues as those in SelD proteins. Further sequence and genome context analysis of SelD-like genes revealed that they might be involved in S metabolism in these organisms. We also analyzed patterns of gene occurrence in different thermoproteales at a genome-wide level, and identified several genes that may co-occur with SelD-like gene. Finally, a model for the possible function of SelD-like protein in these organisms was proposed, which may provide insights into understanding the relationship between Se and S utilization in biology.M. jannaschii SelD protein (gi: 190352215) was used as query sequence to search for SelD homologs in archaea. TBLASTN program was used to identify genes coding for homologs with the cutoff of E-value ≤0.1. In addition, other key components of different Se utilization traits  from representative organisms were also used for identification of the occurrence of each Se trait in archaeal species. Orthologous proteins were defined using the conserved domain database  and bidirectional best hits [Sequenced archaeal genomes from the Entrez Microbial Genome Database at NCBI were used in this study . A totalest hits .Multiple sequence alignments were performed by Clustal Omega program (Version 1.2.0)  with numP. arsenaticum  were used as a query dataset. BLAST programs were used to search these sequences against all sequenced thermoproteales genomes using the following criteria: (i) e-value is less than 1e-07; (ii) the length of the alignment should be extending over half the length of the query sequence; and (iii) the alignment identity should be at least 30%. A simple Perl script was developed to parse the BLAST output and examine presence/absence of homologs in all analyzed genomes. Genes present in any of the thermoproteales lacking SelD-like were dismissed. The remaining genes were then searched against thermoproteales containing SelD-like to determine their occurrences. Top candidate genes were further analyzed for possible function. The occurrence of these candidates in SelD-like-containing sulfolobales was also analyzed.Initial searches for the occurrence of SelD-like genes revealed that only the order Thermoproteales  has both SelD-like-containing and SelD-like-lacking organisms . To identify genes associated with SelD-like, we adopted a modified strategy which had been used previously to identify the archaeal/eukaryotic SecS gene . ProteinThe data sets supporting the results of this article are included within the article and its additional files.Additional file 1: Table S1: This file contains Table S1, which shows the distribution of SelD, SelD-like and all known Se utilization traits in archaea. (XLS 48 KB)Figure S1. shows the multiple sequence alignment of SelD and SelD-like proteins. Figure S2. shows genomic context of SelD-like gene in completely sequenced genomes that contain this gene. (DOC 178 KB)Additional file 2: Figure S1: This file contains two supplementary figures: This file contains representative SelD and SelD-like protein sequences used for multiple sequence alignment .(TXT 6 KB)Additional file 3:


Intra-aortic balloon pumps (IABP) have generally been used for patients undergoing high-risk mechanical coronary revascularization. However, there is still insufficient evidence to determine whether they can improve outcomes in reperfusion therapy patients, mainly by percutaneous coronary intervention (PCI) with stenting or coronary artery bypass graft (CABG). This study was designed to determine the difference between high-risk mechanical coronary revascularization with and without IABPs on mortality, by performing a meta-analysis on randomized controlled trials of the current era.Pubmed and Embase databases were searched from inception to May 2015. Unpublished data were obtained from the investigators. Randomized clinical trials of IABP and non-IABP in high-risk coronary revascularization procedures (PCI or CABG) were included. In the case of PCI procedures, stents should be used in more than 80% of patients. Numbers of events at the short-term and long-term follow-up were extracted.A total of 12 randomized trials enrolling 2155 patients were included. IABPs did not significantly decrease short-term mortality (relative risk (RR) 0.66; 95% CI, 0.42–1.01), or long-term mortality , with low heterogeneity across the studies. The findings remained stable in patients with acute myocardial infarction with or without cardiogenic shock. But in high-risk CABG patients, IABP was associated with reduced mortality .In patients undergoing high-risk coronary revascularization, IABP did not significantly decrease mortality. But high-risk CABG patients may be benefit from IABP. Rigorous criteria should be applied to the use of IABPs. Nowadays, coronary artery bypass graft (CABG) and percutaneous coronary intervention (PCI) with stents are the most common techniques of coronary revascularization. Although tremendous advances have been made in stent types, adjunctive pharmacotherapy, and surgical techniques, mortality rates are still high in high-risk patients .As a circulatory-assist device, the intra-aortic balloon pump (IABP) has been shown to improve clinical outcomes and decrease the mortality rate in high-risk patients undergoing coronary revascularization . HoweverFor emergency high-risk patients, IABP has always been inserted before coronary revascularization. So it could be really important to decide whether or not to use an IABP before deciding using PCI or CABG. Therefore, we conducted this meta-analysis using rigorous inclusion criteria, and regardless of the type of revascularization, to evaluate the risks of IABPs on mortality in patients undergoing high-risk coronary revascularization in our current era of high arterial graft and stent use.A systematic search of all published studies in Pubmed and Embase, from inception to May 2015, was independently performed by two members of our team (YD.W and TW.S). Search terms included: percutaneous coronary intervention, stent, CABG, coronary bypass, CAD, IABP, and intra-aortic balloon counterpulsation. We also reviewed reference lists and recent reviews to identify other potentially eligible studies that had not been identified in our initial search. Details of the search strategy are summarized in Studies were required to be RCTs of revascularization procedures, as well as IABP vs. revascularization procedures alone, in high-risk CAD patients with the individual outcomes of death. In addition, to reflect contemporary interventions and medical practice, inclusion criteria also required the revascularization procedures to be PCI or CABG, and stent implantation in at least 80% of PCI procedures. Finally, all high-risk factors had to be documented for all patients prior to randomization. We excluded reperfusion therapy studies with thrombolytic therapy or drug therapy.Two members of our team (YD.W and ZQ.L) used a standardized data collection form to extract the following information from each included study: source, study design, number of IABP group cases, number of control group cases, population characteristics, definition of IABP and control group, IABP procedure and characteristics of the enrolled patients , prior MI, and stent placed). The supplementary files were also examined for data extraction. When necessary, we contacted authors of the included studies to obtain additional information. The quality of eligible RCTs was assessed using the Cochrane Collaboration's tool for assessing the risk of bias for RCTs .P < 0.10. Heterogeneity was also assessed by means of the I2 statistic, with significance being set at I2 > 50%. The random-effects model was used for statistical analyses due to the wide range of clinical and methodological variability across the trials. We further conducted subgroup analyses to explore possible explanations for heterogeneity. Sensitivity analyses were performed using the one-study-out method, and applying various exclusion criteria to test the robustness of the pooled estimate. Publication biases were evaluated using Funnel plots and Egger’s tests[P-value <0.05 was considered to be statistically significant.Relative risk (RR) was used as a common measure of the association between IABP and mortality from PCI or CABG, across studies. Heterogeneity across trials was assessed using a standard chi-squared test, with significance being set at r’s tests. StatistWe performed this meta-analysis in accordance with the PRISMA statement for repoThe characteristics of the trials included in this study are listed in I2 = 37.6%, P = 0.091). Similarly, in three studies[I2 = 41.6%, P = 0.180)  .We performed a subgroup analysis for short-term mortality according to the type of disease (AMI with or without cardiogenic shock) and the type of revascularization (PCI or CABG). The mortality was not significantly decreased in patients with IABP experiencing AMI with cardiogenic shock , AMI without cardiogenic shock . SimilarP = 0.13) and long-term mortality (P = 0.25), and the funnel plot was symmetric  did not make a significant difference, and the findings of our meta-analysis remained stable despite the exclusion of any single study . No evidymmetric .Our meta-analysis on 12 RCTs including 2155 patients suggests that IABPs did not significantly decrease short-term mortality and long-term mortality. In addition, The mortality of IABP group did not significantly decreased in MI patients with or without cardiogenic shock. But a survival advantage was found in high risk CABG patiens.IABPs have been widely used since the 1960s, and are still a common technique. However, its effects on mortality are still controversial. In a large National Registry of 23,180 patients with cardiogenic shock, a significant reduction in mortality rates was observed. HoweverAnother interesting finding supported the use of prophylactic IABPs in high-risk CABG patients to reduce mortality, and this was consistent with previous meta-analyses,29,35. OOur study showed that survival advantage was found in high risk CABG patients, but not PCI patients. This conflicting result may be explained by the following two reasons. First, The advancement in PCI technology and cardiac medications are one possible explanations. The PCI technology, stent material, and treatment concept advanced greatly, which result to decreased mortality in AMI patients. The loss of benefit of IABP could be remedied by the use of effective cardiac medications. Therefore, it might be hypothesized that the IABP might not show any beneficial effect due to improved care . AnotherOur findings may have several significant advantages. First, to reflect current clinical practice, the inclusion criteria required all revascularization procedures to be PCI or CABG, with stent implantation in at least 80% of PCI procedures. As a result, all studies about PCI with stenting were published in the last 5 years. Given that the first published RCTs about IABP in PCI patients were conducted as early as 1989 years by Ohman EM et al, when thSeveral limitations of this meta-analysis merit consideration. First, the time when the IABP was inserted and removed (before or after PCI and CABG), and the type of IABP device employed, were not strictly described in all trials. Second, there were no long-term mortality data available for some RCTs,19–26. TFor patients undergoing high-risk coronary revascularization with PCI, the routine use of IABPs does not reduce the risk of death compared to no IABPs. But high-risk CABG patients may be benifit from IABP. These findings underline the importance of rigorous criteria in using IABPs to avoid possible complications.S1 TableThe electronic search strategy without language limitation.(DOCX)Click here for additional data file.S2 TableThe risk of bias assessed with Cochrane Collaboration's tool.(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 TableChecklist of preferred reporting items for systematic reviews and meta-analyses.(DOC)Click here for additional data file.


The concept of hybrid coronary revascularization (HCR) combines the left internal mammary artery (LIMA)–left anterior descending (LAD) graft and percutaneous coronary intervention (PCI) to non-LAD vessels. Multiple comparative studies have evaluated the safety and feasibility of HCR and coronary artery bypass grafting (CABG) for multivessel coronary artery disease (MCAD). However, the sample size of each study was small, and evidences based on single-institutional experience. The purpose of this meta-analysis was to compare the short-term outcomes of HCR with those of CABG for MCAD.PubMed, EMBASE and Cochrane Library databases, as well as conference proceedings, were searched for eligible studies published up to March 2014. We calculated summary odds ratios (OR) for primary endpoints ; target vessel revascularization (TVR); major adverse cardiac or cerebrovascular events (MACCEs)) and secondary endpoints ; renal failure; length of stay in the intensive care unit (LoS in ICU); length of stay in hospital ; red blood cell (RBC) transfusion). Data from 6176 participants were derived from ten cohort studies.HCR was non-inferior to CABG in terms of MACCEs during hospitalization (odds ratio (OR), 0.68, 95% confidence interval (CI), 0.34–1.33)and at one-year follow-up , and no significant difference was found between HCR and CABG groups in in-hospital and one-year follow-up outcomes of death, MI, stroke, the prevalence of AF and renal failure, whereas HCR was associated with a lower requirement of RBC transfusion and shorter LoS in ICU and LoS in hospital than CABG (weighted mean difference (WMD) –1.25, 95% CI, –1.62 to –0.88; –17.47, –31.01 to –3.93; –1.77, –3.07 to –0.46; respectively).Our meta-analysis indicates that HCR is feasible, safe and effective for the treatment of MCAD, with similar in-hospital and one-year follow-up outcome, significantly lower requirement of RBC transfusion, and faster recovery compared with CABG. Coronary artery bypass grafting  offers superior long-term advantages owing largely to the left internal mammary artery (LIMA) to the left anterior descending (LAD) artery graft ,2. Conve patency ,7. In co patency ,9. HowevThe concept of hybrid coronary revascularization (HCR), which combines the LIMA–LAD graft and percutaneous coronary intervention (PCI) to non-LAD vessels, was first introduced by Angelini and colleagues in 1996 . IntroduTwo independent reviewers  searched PubMed, Embase, Web of Science, and the Cochrane Library for randomized controlled trials (RCTs) and non-RCTs up to 1 March 2014 and compared HCR with CABG for MCAD without language or publication restrictions. The following medical subject heading terms and their variants were used in database searches: hybrid coronary revascularization; coronary artery bypass grafting (on-pump or off-pump); multivessel coronary artery disease. Reference lists within selected studies and abstracts published at major international conferences were also searched.The safety endpoints of this meta-analysis were death, stroke, myocardial infarction (MI) and major adverse cardiac or cerebrovascular events (MACCEs). The efficacy endpoint was revascularization. All the primary endpoints were measured in hospital and one year of follow-up. Death was defined as death from any cause. MI was diagnosed by symptoms, electrocardiography and changes in serum levels of cardiac biomarkers. Target vessel revascularization (TVR) was the need for repeated CABG or percutaneous coronary intervention (PCI). MACCEs were defined as a composite of death, MI, stroke or revascularization.Secondary outcomes were atrial fibrillation (AF), renal failure , length of stay in the intensive care unit (LoS in ICU), length of stay in hospital , and transfusion of red blood cells (RBCs).Five main criteria were used: (i) comparison of HCR with CABG for MCAD; (ii) studies reporting at least one of the outcomes mentioned above; (iii) studies documenting surgical procedures such as one-stop HCR or staged HCR, on-pump or off-pump CABG, and documenting surgical methods such as HCR or CABG; (iv) follow-up duration ≥30 days; (v) non-RCTs with a score >5 as assessed by the Newcastle–Ottawa Scale (NOS) -24.http://www.ohri.ca/programs/clinical_epidemiology/oxford.asp), which addressed three items: patient selection, comparability of groups, and outcome assessment  assessed the methodological quality of the included studies, and disagreement was resolved by consensus and discussion. Quality of non-RCTs was evaluated with the modified NOS . Continuous and dichotomous variables were assessed by weighted mean differences (WMDs) and odds ratios (OR), respectively. A 95% confidence interval (CI) was recorded. Heterogeneity among studies was quantified using the I2 statistic. According to Higgins’ method, IWe identified ten studies eligible for inclusion: nine non-randomized and one randomized Figure , All of P = 0.64; Figure Five studies reported on in-hospital mortality in 5770 patients. Pooled results showed no significant difference in mortality between the HCR group and CABG group <1 month .Patients treated with HCR did not display a significant reduction in risk for MI as compared with those who received CABG within hospital .Stroke was assessed in five studies reporting on 5793 patients. The prevalence of stroke was not significantly different between groups. .MACCEs occurred in 2.5% (10/408) of patients after HCR and 3.6% (181/5092) of patients with CABG.Six studies (5500 patients) provided data on the prevalence of MACCEs. Pooling of the outcomes of these studies revealed no significant differences in the prevalence of MACCEs between patients treated by HCR and those treated by CABG  or the prevalence of renal failure .There was no significant difference in the prevalence of AF between the two groups , LoS in hospital , and fewer instances of RBC transfusion   of patients after HCR and 11.8% (18/152) of patients with CABG at one year of follow-up. The ORs for MACCEs were not significantly different at one year of follow-up  or LoS in hospital  between the two groups.Inclusion of studies with ≥80 patients in each group did not substantially change the pooled point estimate except for LoS in ICU and LoS in hospital. Pooling the outcomes of these studies revealed no significant difference in the LoS in ICU  whereas HCR was associated with a reduced need for RBC transfusion as well as shorter LoS in ICU and LoS in hospital than CABG.We revealed no significant differences in the prevalence of in-hospital and one-year follow-up mortality, MI, stroke, TVR, MACCEs between the two groups, findings which were consistent with the early results of an ongoing RCT . In thatvs 9.6%) after a mean follow-up of 18 months. Shen et al. [P = 0.030).The prevalence of in-hospital one-year follow-up mortality, MI, stroke and MACCEs were similar between the two groups, however, we hypothesized that HCR may be superior to CABG in terms of long-term MACCEs. Hu et al.  reportedn et al.  also repAs duration of follow-up extends, HCR may be superior to CABG in terms of long-term MACCEs.Several reasons support this findings mentioned above. Firstly, avoiding aortic clamping is one of the unique advantages of the hybrid procedure. Aortic manipulation  is necessary during on-pump CABG . SecondlCardiac surgery and administration of contrast dyes during PCI tends to increase the risk of renal failure. Hence, we suggest that the prevalence of renal dysfunction might be higher in patients treated with HCR compared with those undergoing CABG alone. In this meta-analysis, however, no significant difference in the prevalence of renal failure was found between the two groups. The more stable hemodynamics and better urine output in the hybrid group may have resulted in a similar prevalence of renal failure in the present study ,32.In this analysis, the significantly lower requirement of RBC transfusion in the HCR group was attributed to the less invasive nature of minimally invasive direct coronary artery bypass grafting in the hybrid procedure despite continuous perioperative use of aspirin and perioperative administration of clopidogerl ,14,31.LoS in ICU and LoS in hospital were significantly shorter in the HCR group in this analysis, so the hybrid group had a shorter recovery with a substantial reduction in utilization of hospital resources . This phHeterogeneity among studies was observed for several continuous variables, including LoS in ICU and LoS in hospital. This heterogeneity may have resulted from variations in the surgeon’s caseload, the learning-curve effect, the HCR procedure, perioperative management, and standards regarding hospital discharge among the included studies.http://www.clinicaltrials.gov/ct2/show/NCT01699048 and http://www.clinicaltrials.gov/ct2/show/NCT01035567). Ideally, a meta-analysis should include RCTs only, but inclusion of high-quality non-RCTs can improve the statistical power while maintaining an acceptable level of evidence. Abrahama et al. [This meta-analysis had several limitations. The main limitation of our meta-analysis was the retrospective nature of the available data. No RCT has been published but two protocols have been registered (available at a et al.  reportedSecondly, the duration of clinical follow-up was limited to one year in most studies, whereas a meta-analysis of long-term outcomes was not possible due to insufficient data ,17,18,20Thirdly, the definition of endpoints such as MI, MACCEs, and renal failure in different studies varied to some degree, which may have weakened the evidences in our analysis.Finally, obvious heterogeneity was observed for several continuous variables. Therefore, the random-effects model was used.This meta-analysis suggested that HCR is feasible, safe and effective for treatment of MCAD, with similar in-hospital and one-year follow-up outcomes, significantly lower requirement for RBC transfusion, and faster recovery compared with CABG. It may provide a safe and effective alternative for treating selected patients with MCAD. However, to validate the long-term results of HCR for MCAD, more large-scale, multicenter, prospective RCTs are warranted.


Current evidence suggests that beta-blocker lower the risk of development of atrial fibrillation (AF) and in-hospital stroke after cardiac surgery. This study was to assess whether beta-blockers could decrease incidence of new-onset AF in patients with end stage renal disease (ESRD). We identified patients from a nation-wide database called Registry for Catastrophic Illness, which encompassed almost 100% of the patients receiving dialysis therapy in Taiwan from 1995 to 2008. Propensity score matching and Cox’s proportional hazards regression model were used to estimate hazard ratios (HRs) for new-onset AF. Among 100066 patients, 41.7% received beta-blockers. After a median follow-up of 1500 days, the incidence of new-onset AF significantly decreased in patients treated with beta-blockers . The prevention of new-onset AF was significantly better in patients taking longer duration of beta-blockers therapy (P for time trend <0.001). The AF prevention effect remains robust in subgroup analyses. In conclusion, beta-blockers seem effective in the primary prevention of AF in ESRD patients. Hence, beta-blockers may be the target about upstream treatment of AF. Atrial fibrillation (AF) is a result of continuous remodeling of the atrial, a dynamic interaction between a trigger and the substrate13479Beta-blockers administration is the most widely used prophylactic strategy of POAF based on numerous studies showing benefit, ease of use and cost consideration118Patients with chronic kidney disease and end-stage renal disease (ESRD) are more likely to develop cardiovascular disease, including myocardial infarction1720In the AFFIRM study, beta-blockers were the most effectively and commonly used drug class for rate control23There were 100066 patients who met the study inclusion criteria; 58382 (58.3%) did not use beta-blockers while 41684 (41.7%) used beta-blockers. Patients not receiving beta-blocker treatment were served as control group. The median follow-up time was 1500 days. The algorithm was listed in Clinical and demographic characteristics were listed in The median durations of follow-up were 1295 and 1741 days in control and in beta-blocker group. As demonstrated in The results of Cox’s regression analyses were demonstrated in The results of subgroup analyses were demonstrated in Currently, accepted primary prevention of AF was discouragingly limited. According to ESC 2012 guidelines, recent double-blind, placebo-controlled trials with ARBs and the majority of trials with polyunsaturated fatty acids failed to show convincing results2728AF, while being the most common arrhythmia among general population, is also the most common arrhythmia among patients with ESRD2DS2VASc score, but currently there is insufficient evidence to support application of these recommendations to patients with ESRD. From prior study, warfarin was associated with inconsistent results in patients with ESRD, and was even a risk factor for stroke38Both increased comorbid illnesses and non-traditional factors, such as inflammation and oxidative stress, contribute to the formation of substrate of AF in patients with ESRD3233In the field of AF primary prevention, namely upstream therapy, effective therapy was discouragingly limited. ACEI/ARB did show its preventive effect in patients with HF with reduced LV ejection fraction from some retrospective studies4041Several other pathophysiological processes, including sympathetic activation and oxidative stress, are associated with atrial remodeling that may predispose patients to AF, and could also be the targets of upstream therapyPatients with ERSD are also a special group in a milieu of high baseline sympathetic tone and oxidative stress3350There are several limitations needed to be addressed. First, the study is limited by its retrospective, nonrandomized nature, and the imbalance in risk factors among different antihyperglycemic agents’ users in the whole cohort. Also, treatment allocation was not randomized and treatment selection bias had to be taken into account. Although we adjusted the confounding factors, the result might be still confounded by other underlying disease we did not consider. Second, our primary end-point was the time to the occurrence of AF diagnosed via ICD-CM coding. The accuracy of the diagnoses, which are based on the administrative data reported by physicians, may be a concern. Patients prescribed with beta-blocker might benefit more from beta-blocker originally, for example, more HF with reduced EF, which cannot be discriminated from patients with HF with preserved EF from the database. There is a risk that much AF is underreported because of diagnostic modality being used. Third, although we adjusted confounding factors, the result might be still confounded by other underlying disease we did not consider. Finally, we did not distinguish between chronic and paroxysmal AF and the results might differ in these two conditions.In patients with ESRD receiving replacement therapy, beta-blocker usage is associated with appeared to be effective in the primary prevention of AF.This large-scale, longitudinal cohort study used integrated medical and pharmacy claims data from National Health Insurance Research Database (NHIRD) in Taiwan. The National Health Insurance program has provided compulsory universal health insurance in Taiwan since 1995. More than 98% of the total Taiwanese population of 23 million is covered by the program. The NHIRD contains nearly complete claims history of diagnosis and procedures, provided as the International Classification of Diseases Ninth Revision Clinical Modification (ICD-9-CM) codes, and drug dispensing for every beneficiary. The NHIRD established a registry system for “Catastrophic Illnesses”, including cancer, chronic mental illness, congenital illness and ESRD. In Taiwan, ESRD patients undergoing hemodialysis and peritoneal dialysis are categorized as “severe illness” and the national insurance covers almost all the medical fees. All the medications, procedures, every out-patient clinic visits and hospital admission covered by the insurance were recorded in the database. The Bureau of National Health Insurance performs routine validations of the diagnoses by reviewing the original medical charts of all of the patients who applied for catastrophic illness registration. To comply with data privacy regulations, personal identities were encrypted and all data were analyzed in a de-identified mannerWe investigated the database of NHIRD during year of 1995 to 2008. The index date for the study cohort was identified as the date of the first-time that had both a diagnosis of ESRD  and a procedure of hemodialysis (ICD9-CM: 39.93) or peritoneal dialysis (ICD9-CM: 54.98). These ESRD patients have already undergone dialysis treatment without receiving renal transplantation. We identified all patients who were above 18 years old. The exclusion criteria included the following: (1) diagnosis of valvular heart disease , (2) prior ambulatory visit for AF , (3) receiving beta-blockers less than 30 days. A flowchart for the identification of study subjects is shown in Patients were classified into use and non-use of beta-blockers. We had 12 kinds of beta-blockers (around 90 of generic drugs with different doses). To simplify the presentation, we used the total treatment duration of beta-blockers to demonstrate the dose-response effect. The majority of the treatment frequencies for beta-blockers were once daily  and twice a day .Comorbidity was defined by diagnoses at hospital discharge or in clinic records. For our study population, we searched the database to see if they had HTN(ICD-9-CM codes: 401.X-405.X), DM , dyslipidemia (272.X), ischemic stroke , hemorrhagic stroke , CAD , CHF hospitalization , PAD . Medications that were dispensed at time of index date, including beta-blockers, angiotensin converting enzyme inhibitors (ACEI), angiotensin receptor blockers (ARB), calcium channel blockers (CCBs), diuretics, statin, oral anti-diabetic drugs (OADs) and insulin were identified.The main purpose of the study was to compare long-term development of AF among patients with and without taking beta-blockers. Diagnosis of AF was based on ICD-9-CM coding  in any ambulatory visit and discharge diagnoses.For comparison of the baseline characteristics between two groups, Student’s t test was used for continuous variables and chi-square test was employed for categorical variables. Because patients were not randomly allocated to the two groups, we adjusted for age, gender, risk factors , comorbidities  and medication usage. In addition, the propensity score was applied to reduce the potential bias and to make the two groups more comparable. We derived a propensity score, which is the logit (probability) for receiving beta-blockers treatment from a logistic regression model by using all background covariates listed in For survival analyses, multivariate Cox’s proportional hazard regression analyses were used to derive the adjusted hazard ratios (HR) for developing AF by using patients without beta-blockers treatment as controls. Three models were used to adjust potential confounders. The model 1 was adjusted for age, gender, risk factors , comorbidities  and medication usage. The model 2 was similar to model 1 but included patients after propensity matching. In the model 3, the propensity score was entered as a covariate in the Cox’s proportional hazard regression model to adjust the potential selection bias. To test the consistency of the results, we also did subgroup analyses for different gender, age, presence of HTN, DM, cardiovascular disease (CVD) and CHF with adjustment for all confounders. The event-free survival curves of the two groups were illustrated by using the Kaplan-Meier method. The log-rank analysis was applied to test the differences in survival among groups.All of the analyses were conducted using the Statistical Package for the Social Sciences (SPSS) for Windows, Version 19.0 . A P value < 0.05 was considered statistically significant.How to cite this article: Lin, T.-T. et al. Primary prevention of atrial fibrillation with beta-blockers in patients with end-stage renal disease undergoing dialysis. Sci. Rep.5, 17731; doi: 10.1038/srep17731 (2015).


In Phytophthora-ID, the Sequence-ID application allows identification based on ITS or cox spacer sequences. Genotype-ID groups individuals into clonal lineages based on simple sequence repeat (SSR) markers for the two invasive plant pathogen species P. infestans and P. ramorum. All code is open source and available on github and CRAN. Instructions for installation and use are provided at https://github.com/grunwaldlab/Microbe-ID.Development of tools to identify species, genotypes, or novel strains of invasive organisms is critical for monitoring emergence and implementing rapid response measures. Molecular markers, although critical to identifying species or genotypes, require bioinformatic tools for analysis. However, user-friendly analytical tools for fast identification are not readily available. To address this need, we created a web-based set of applications called Microbe-ID that allow for customizing a toolbox for rapid species identification and strain genotyping using any genetic markers of choice. Two components of Microbe-ID, named Sequence-ID and Genotype-ID, implement species and genotype identification, respectively. Sequence-ID allows identification of species by using BLAST to query sequences for any locus of interest against a custom reference sequence database. Genotype-ID allows placement of an unknown multilocus marker in either a minimum spanning network or dendrogram with bootstrap support from a user-created reference database. Microbe-ID can be used for identification of any organism based on nucleotide sequences or any molecular marker type and several examples are provided. We created a public website for demonstration purposes called Microbe-ID ( Phytophthora ramorum causing sudden oak death  for the oomycete P. ramorum (Clavibacter michiganensis subsp. michiganensis (Aphanomyces euteiches (Phytophthora (Phytophthora-ID) with two sequence databases containing over 110 species that are mostly plant pathogens  , provided as a demonstration site (http://microbe-id.org/), and a working implementation of Sequence-ID and Genotype-ID that we use in our own work for the genus Phytophthora (phytophthora-id.org). Finally, a companion paper describes application of Microbe-ID for the implementation of a new website, Gall-ID, with novel tools for identification of gall-forming bacteria (Our objective here was to report the development of a toolbox for microbe identification (Microbe-ID) that can readily be customized for sequence based species identification (Sequence-ID) or molecular marker-based identification of genotypes (Genotype-ID) for any group of organisms. Our objectives were two-fold: (1) to implement Microbe-ID as a demonstration site that is customizable for any group of organisms and (2) to demonstrate a working implementation at sion 1.0 . Microbe ramorum , concateganensis , Fig. S3ganensis  for the uteiches . Moreoveathogens , and twotype-ID) . All of bacteria .http://microbe-id.org/) with two separate modules, Sequence-ID and Genotype-ID, for sequence-based species identification and genotyping, respectively. The website is written using bootstrap (http://getbootstrap.com/), a HTML, CSS, and JS framework for developing responsive, mobile first projects on the web. Specific instructions, code, and resources necessary for implementation of Microbe-ID are provided on the GitHub repository . The server currently hosting Microbe-ID is running Centos Linux release 6.6, NCBI BLAST 2.2.28+  the user will encounter a menu bar with links to Sequence-ID, Genotype-ID, and an about page. The home page has a general description of the functionality and components implemented in Microbe-ID, as well as links to the github site. Since input from the user is entered into forms as a query, each form implemented in Microbe-ID is encoded to check that the format of the data supplied by the user is supported. If the format is incorrect, the page will prompt the user with an error message, making the use of Microbe-ID more user-friendly.Navigating the demonstration site of Microbe-ID , multilocus genotype sequence types (MLST data), and AFLP, RFLP, SNP and other binary datasets (Binary (AFLP) data). The user supplies a query via the web framework that is read into R and queried against a curated dataset. Genetic distances  are calcP. ramorum  and number of bootstrap replicates. The user can also download results as a PDF or in NEWICK format. For the minimum spanning network, the user can adjust the grey scale for edge distances and download results as a PDF file. Implementations for other genetic data and different visualizations can be added.Microbe-ID contains implementations of each of the modules for different custom databases: the SSR implementation uses nine diploid SSR loci for the oomycete species  ramorum ; the MLShttp://Phytophthora-ID.org, a functional website for identifying samples from the genus Phytophthora, a group of economically important plant pathogens in the stramenopile branch of the tree of life . This particular version of the sequence identification tool was created in PERL-CGI, and permits the search of a FASTA sequence query against a curated database of Phytophthora species. The PERL-CGI for Phytophthora-ID version 2.0 was also redesigned to run directly on the server to make the web application more stable and faster. In contrast, in Phytophthora-ID 1.0, the web application was designed as a communication wrapper to an external cluster, thus rendering functionality dependent on the external server.The current version of cox spacer region spanning the cox1 and cox2 loci. Databases were created using sequences from published Phytophthora species descriptions  and mitochondrial riptions  or thoseas valid .cox spacer region, we created a database of 150 sequences representing 106 species. Laboratory protocols for preparing samples for sequence analysis are available on the website and in a previous publication  were genotyped at nine SSR loci  represented by a total population of 96 isolates were genotyped at 11 SSR loci, using protocols from P. ramorum and one sample of clonal lineages US-23 and US-8 clonal lineages for P. infestans. All samples grouped with the corresponding reference lineages with high support values . We implemented a demonstration available at http://Microbe-ID.org. New implementations for any organisms require reference databases, scripts, and web pages including choices from the set of computational tools shown in Phytophthora-ID (http://Phytophthora-ID.org/) and Gall-ID  of Clavibacter michiganensis. The query used is a concatenation of a WASH sample using all 8 genes. Queries are represented in blue. Note that all queries are correctly placed amongst samples of its presumptive clonal lineage while also representing relationships between lineages in the reference dataset.Click here for additional data file.10.7717/peerj.2279/supp-4Figure S4The queries used for this iteration of Binary-ID are two samples from a presumptive “Athena” origin. Queries are represented in red. Note that all queries were correctly placed amongst samples of its presumptive clonal lineage while also representing relationships between lineages in the reference dataset, indicating both queries are more closely related to the “Athena” population than the “Mt. Vernon” population.Click here for additional data file.