Recent evidence has suggested that appendix plays a pivotal role in the development and preservation of intestinal immune system. The aim of this study is to examine whether prior appendectomy is associated with an increased risk for the development of antibiotic-resistant bacteria in bacteremia from biliary tract infection (BTI). Charts from 174 consecutive cases of bacteremia derived from BTI were retrospectively reviewed. Using multivariate analysis, independent risk factors for development of antibiotic-resistant bacteria were identified among the clinical parameters, including a history of appendectomy.p = 0.026), antibiotic use within the preceding three months , and bilioenteric anastomosis or sphincterotomy  were independent risk factors for antibiotic-resistant bacteria. In total, 221 bacteria strains were identified from 174 BTI events. Of those, 42 antibiotic-resistant bacteria were identified in 34 patients. Multivariate analysis revealed that prior appendectomy (Odds ratio (OR), 3.02; 95% confidence interval (CI), 1.15–7.87;  Prior appendectomy was an independent risk factor for the development of antibiotic-resistant bacteria in bacteremia from BTI. Biliary tract infection (BTI) with bacteremia represents 8–20% of community-acquired bacteremia in the elderly population and it is the second most common cause of sepsis in those subjects, leading to 10–20% of mortality [Bacteremia in BTI is caused by a wide spectrum of pathogens. In recent years, the prevalence of antibiotic-resistant pathogens in BTIs has risen steadily , 2. Infe Clostridium difficile infection [Previously, the human appendix has been regarded as a rudimentary part of the intestine. In recent years, however, several studies have suggested its immunological importance for the development and preservation of the intestinal immune system . In addinfection , 9 and rnfection . In thisThe focus of the current study is to analyze whether prior appendectomy is associated with an increased risk of the development of antibiotic-resistant bacteria in bacteremic BTI. Staphylococci, Corynebacterium, Propionibacterium, and Bacillus species) and (ii) bacteremia due to a non-BTI.This retrospective study was conducted at the Wakayama Rosai Hospital in Wakayama, Japan. Charts from 174 consecutive cases of bacteremia derived from BTI treated in the hospital between June 2005 and May 2016 were retrospectively reviewed. Patients with the following criteria were excluded from the study: (i) positive culture for probable skin contaminants , cause of infection, underlying diseases, medication, history of appendectomy, previous antibiotic use (within 90 days), presence of indwelling biliary device, previous hospitalization (within 30 days), and history of bilioenteric anastomosis or sphincterotomy. In addition, 30-day mortality of the patients was examined.This study was approved by the institutional review board at the Wakayama Rosai Hospital and conforms to the provisions of the Declaration of Helsinki .Cholecystitis was diagnosed on the basis of clinical presentation of fever, right upper quadrant pain, and findings of ultrasonography or computed tomography. Cholangitis was diagnosed based on the following criteria: (a) presence of fever with upper quadrant pain; (b) radiological (sonographic or computed tomographic) or endoscopic evidence of biliary tract obstruction due to stones or stricture from benign or malignant origin; and (c) laboratory findings of hyperbilirubinemia and an elevated serum alkaline phosphatase level.Oral temperature: two points for a temperature of ≤35°C or ≥40°C or one point for a temperature between 35.1 and 36.0°C or between 39.0 and 39.9°CHypotension: two points for an acute hypotensive event with decreases in systolic (>30 mmHg) and diastolic (>20 mmHg) blood pressures, the use of intravenous vasopressor agents, or systolic blood pressure of <90 mmHgReceiving mechanical ventilation: two pointsSuffering cardiac arrest: four pointsMental status: being alert, zero points; being disoriented, one point; being stuporous, two points; and being comatose, four points The Pitt bacteremia score was used to assess the severities of bacteremic BTIs and was calculated as follows:β-lactamase- (ESBL-) producing bacteria, ampicillin-resistant Enterococci and SPACE , in this study according to the previous reports [Antibiotic-resistant bacteria were defined as extended-spectrum  reports –14.Two sets of two 8–10 mL blood samples were taken from patients who presented with ≥38°C or who showed clinical signs or symptoms associated with bacteremia. Blood samples were processed in the Oxoid Signal Blood Culture System  between June 2005 and October 2011 and processed in an automated BacT/ALERT 3D System  between November 2011 and May 2016. The incubation period of blood samples was five days. Positive blood samples were subcultured onto blood agar plates or Brucella blood agar plates. Identification of aerobic and anaerobic bacteria was performed by the WalkAway System (Beckman Coulter) and RapID ANA II , respectively.t-test as appropriate. Independent risk factors for the detection of antibiotic-resistant bacteria were identified using multivariate logistic regression analysis that included parameters with p < 0.1 in univariate analysis. p value < 0.05 was considered statistically significant and odds ratios (ORs) with 95% confidence intervals (CIs) were determined. Data were statistically analyzed using JMP version 9 software  [Differences between patients with and without antibiotic-resistant bacteria were determined using the chi-square test and Student's NC, USA) .The demographic and clinical features of analyzed patients with bacteremia are summarized in  Klebsiella species were the most common isolates , followed by Escherichia coli , Enterococcus species , Streptococcus species , and Pseudomonas aeruginosa .The bacteria strains isolated from the blood cultures are listed in  Pseudomonas aeruginosa (24%), 8 Enterobacter species (19%), seven ampicillin-resistant Enterococcus faecium (17%), 6 ESBL-producing Klebsiella species (14%), and 5 ESBL-producing Escherichia coli (12%). Of these, 23 bacteria (55%) were categorized into the SPACE group.Forty-two antibiotic-resistant bacteria strains were identified in 34 patients , includiDifferences between patients with and without antibiotic-resistant bacteria are shown in p = 0.026), antibiotic use within the preceding three months , and bilioenteric anastomosis or sphincterotomy  were the independent risk factors for antibiotic-resistant bacteria (p = 0.039), suggesting that appendectomy contributes to the development of antibiotic-resistant bacteria in relatively younger subjects.Multivariate analysis revealed that prior appendectomy  of which had a history of appendectomy, whereas only 20 (14%) of the remaining 140 subjects had the history. Because the lifetime risk of appendectomy has been reported to be 17.6% in the national hospital discharge survey , our coh% of whic+ T cells activation [The immunological functions of the appendix may work for prevention of prevalence of antibiotic-resistant bacteria. The human appendix is associated with the highest concentration of gut-associated lymphoid tissue (GALT) in the gut . GALT istivation . Moreovetivation . In thistivation . In additivation . Thus, lOther functions of the appendix associated with intestinal flora may also be correlated with the results of the current study. The human appendix contains a wealth of microbes, including members of 15 phyla, which constitute more than 98% of the normal colonic microbiome . This inThe present study indicates that appendectomy contributes to the development of antibiotic-resistant bacteria in relatively younger subjects with BTIs. Older subjects generally have more comorbid diseases and are relatively immunocompromised. Therefore, appendectomy may no longer affect their immunological defense. Alternatively, the functions of the appendix may attenuate in older subjects because the number of lymphoid follicles and length and diameter of the appendix are known to degenerate with age . In clinConsidering these facts, the appendix should no longer be regarded as a rudimentary organ and we should carefully decide whether appendectomy should always be indicated for appendicitis. Efficacy of conservative treatment with antibiotics on appendicitis is still controversial. A meta-analysis of randomized controlled trials showed that antibiotics were both effective and safe as primary treatment for patients with uncomplicated acute appendicitis . The lar Pseudomonas aeruginosa, ampicillin-resistant Enterococcus, and ESBL-producing pathogens which were most frequently identified antibiotic-resistant pathogens in this study.Based on the results of the current study, the recommended first choice of antibiotics for patients with bacteremia from BTI could be based on the presence or absence of identified risk factors including a history of appendectomy. In the Tokyo Guidelines , broad-sWe demonstrated that history of appendectomy is a risk of development of antibiotic-resistant bacteria in patients with bacteremia from BTI. The majority of BTI patients, however, showed negative results in blood culture. The causative pathogens in these blood culture-negative BTI patients may also be different according to the risk factors including history of appendectomy. Although it may be difficult to verify the hypothesis due to difficulty in bacteria detection, bile culture could be an alternative, particularly in cases of cholangitis. Moreover, it is potentially useful to investigate whether causative microorganisms in a variety of infectious diseases other than BTI differ between patients with and without a history of appendectomy. Further studies are expected for these clinically relevant issues.There are limitations to this study. First, the number of the examined patients was relatively small and all subjects were Japanese. Reproducibility should be confirmed with larger cohorts including other ethnic populations. Another drawback, due to the study being retrospective, is the possibility of missing relevant clinical parameters and inclusion of bias caused by missed blood culture at appropriate timing.In conclusion, prior appendectomy was identified as an independent risk factor for antibiotic-resistant bacteria in BTIs. Before the choice of antibiotics, interviews regarding history of appendectomy are mandatory as well as other risk factors, including antibiotic use within the preceding three months and bilioenteric anastomosis or sphincterotomy.


N = 750) spanning a spectrum of concreteness in a continuous manner from highly abstract to highly concrete. Participants (N = 328) rated each target word on a range of 14 cognitive dimensions . The dimensions reduced to three factors: Endogenous factor, Exogenous factor, and Magnitude factor. Concepts were plotted in a unified, multimodal space with concrete and abstract concepts along a continuous continuum. We discuss theoretical implications and practical applications of this dataset. These word norms are freely available for download and use at http://www.reilly-coglab.com/data/.Cognitive science has a longstanding interest in the ways that people acquire and use abstract vs. concrete words . One dominant theory holds that abstract and concrete words are subserved by two parallel semantic systems. We recently proposed an alternative account of abstract-concrete word representation premised upon a unitary, high dimensional semantic space wherein word meaning is nested. We hypothesize that a range of cognitive and perceptual dimensions  bound this space, forming a conceptual topography. Here we report a normative study where we examined the clustering properties of a sample of English words ( The word concreteness effect refers to a robust advantage that concrete words manifest over abstract words across numerous domains, including age-of-acquisition, spelling, reading, serial recall, speeded naming, and word recognition  proposes the dissociation between abstract and concrete concepts is due to differences in verbal context  has seen a lengthy and influential tenure as a dominant model of word concreteness Paivio, . DCT is DCT yields an explicit prediction about the effects of neurological damage on abstract-concrete word representation. That is, damage to the verbal semantic system should produce catastrophic loss of abstract concepts. In contrast, concrete concepts should show resilience to verbal semantic damage due to their redundant representation within a sensory-based semantic code. Conversely, selective damage to the sensory-based semantic code should impact abstract and concrete words uniformly, since both word types share coding within the verbal semantic system. Thus, DCT predicts one type of patient-based dissociation characterized by the loss of abstract words. While a majority of studies display this dissociation  were grouped together while INSTANCE and BANANA (low affective concepts) were very far from each other. It was also determined that the Magnitude factor was important for more finely grained distinctions between concepts. This finding can be understood through the concepts of HATE and DISLIKE. While HATE and DISLIKE describe similar feelings, their subtle differences can only truly be described through subtle differences in the scale of these feelings.We were able to plot the words in a three-dimensional space bounded by the three latent dimensions we created during factor reduction. Plotting the words in this manner allowed us to determine the distance between the concepts using the simple distance metric of Euclidian distance. In effect, we were able to create a semantic space where we could determine how close or far concepts were to each other, with distance in space indicating the relatedness of concepts. We compared the ability of our space to determine the relatedness of abstract concepts to that of Latent Semantic Analysis , followed a similar procedure to the ACF. Both approaches asked participants to rate concepts across a variety of dimensions on a seven point Likert scale. The CFR, however, included a greater number of dimensions that would aid in the specificity of the concrete space. We also increased the corpus of words to include not only a greater number of abstract and concrete concepts but also a number of words of middling concreteness which were not included in our previous work. This allowed us to create a continuous semantic space which is closer to the true nature of concepts as opposed to the bimodal space we had previously created.The greatest difference between the ACF and the CFR approach were the dimensions chosen to be included. We kept the previous dimensions of Emotion, Polarity, Social Interaction, Thought, Morality, Time, Space, and Quantity but also included the dimensions of Visual Form, Auditory, Tactile, Olfactory/Gustatory, Visual Color, and Self-Generated Motion.The overall polarity of concepts  was considered as a possible marker of the reward system  was considered because “survival depends upon effective social functioning”  and the motivation to act in accordance with certain social or group rules. Moral behavior has been hypothesized to reflect cognitive-emotional association complexes represented across a prefrontal cortex-temporo-limbic network  was selected in an effort to capture the demands that many terms that have multiple meanings or senses in different contexts may place upon skills such as planning, selection, inhibition, executive flexibility, and strategizing  were combined in this study. Both senses, in contrast to the visual, auditory and tactile senses, are not represented in the unimodal neocortex but in the limbic and paralimbic cortex  were recruited through the online crowd-sourcing program Mechanical Turk. Recent validation studies have shown that the Mechanical Turk data show comparable reliability to standard survey metrics, independent of the task or the amount of payment  included English nouns drawn from the Medical Research Council (MRC) Psycholinguistic Database  I can feel with my hands or body.”Smell/Taste “I relate this word to flavors and odors I can smell and/or taste.”Auditory “I relate this word to sounds, rhythms, etc. that I can hear.”Color “I relate this word to color.”Self-Motion “I relate this word to my own self-generated movement.”The 750 words were rated across 14 cognitive dimensions by participants. Participants signaled their rating for each word via mouse click in a horizontally arrayed series of survey bubbles, corresponding to a seven-point Likert Scale. This scale ranged from Strongly Disagree (1), through Neutral (4), to Strongly Agree (7). The specific wording we used for rating each cognitive dimension was as follows:Participants were instructed to use the entire scale and to work quickly but carefully. The order of presentation both between and within the cognitive dimensions was fully randomized.N = 750) to a hierarchical agglomerative cluster analysis. This clustering algorithm involves a bottom-up approach, aggregating clusters upward until attaining a stopping point. We used Ward's method of clustering . All dimensions had excellent inter-rater reliabilities Table  accordinFigure Using an Eigenvector >1 extraction criterion, SPSS-21 extracted three latent factors from the original 14 dimensions. Table Factor 1 (Endogenous): Emotion, Polarity, Social Interaction, Morality, Motion Self-Generated, ThoughtFactor 2 (Exogenous): Color, Olfactory/Gustatory, Tactile, Visual Form, AuditoryFactor 3 (Exogenous): Space, Quantity, TimeFactors aggregated as follows:We plotted the factor scores in order to create a visual representation of the semantic space. We created a three-dimensional scatterplot of the space along with two-dimensional scatterplots as it can be difficult to determine patterns in a three dimensional plan. We created two versions of each plot, one set of plots was colored based on the concreteness of the concepts Figures –6 and thWe evaluated various cluster sizes using Cohen's Kappa, and a 16-cluster solution (Kappa = 0.89) best fit the data. We then conducted a hierarchical cluster analysis, starting with 16 clusters, agglomerating upward through six distinct levels until achieving convergence. Figure In this experiment we applied the Conceptual Feature Rating (CFR) approach as a method to describe the features of both abstract and concrete concepts in a unified space. We improved upon our initial method by not only including dimensions from our previous work but also by expanding our approach to other dimensions suggested by feature listing approaches and the work on perceptual strength by Connel and Lyott .Figure Our cluster analysis revealed both an improvement in our overall method from our previous work but also some important theoretical findings. Through the inclusion of greater dimensionality, we were able to improve the specificity of our space for concrete concepts. Figure Further inspection of the cluster analysis revealed trends that were similar to those seen in the semantic space created by the ACF. Globally, abstract and concrete concepts split. Concrete concepts were more dominant on one side of the cluster analysis while abstract concepts were more dominant on the opposite side. It should also be noted that concrete concepts had higher Exogenous variable ratings while abstract concepts had higher Endogenous variable ratings overall.A closer inspection revealed, as with previous work, that the Endogenous variable had the ability to draw concepts high in the Exogenous variable toward the right side of the cluster analysis  and other abstract scientific conceptualizations .We also observed that even though polarity (at least not as described by Bradley and Lang) and dominance were not directly measured, items did cluster across these variables. Positive emotions such as LOVE, FREEDOM, and HOPE (C14) clustered separately from Negative emotions such as GRIEF, SADNESS, and MISERY (C16). Concepts low in dominance such as HELPLESSNESS, FEAR, and GUILT (C9) clustered as well.These findings have interesting implications for how we understand the relationship between abstract and concrete concepts. Our CFR space not only allows us to describe the features of just abstract and just concrete concepts but also to describe the relationship between abstract and concrete concepts. By offering a unimodal, multidimensional semantic space, we are able to describe concepts in a more naturalistic manner.There remains much controversy on the nature of concreteness in the literature. A large body of work suggests that abstract and concrete concepts are separate and unique entities with a bimodal distribution . Having older adults and persons with neurological disease perform a feature rating approach would be an excellent method for describing these differences. In this case, our CFR approach would likely be best with these groups due to the inherent difficulties in recruitment and limitations in time associated with them. Also with enough normative data with these groups, this approach could become a sensitive screening measure for neurological disease. We know that a variety of neurological disorders affect the semantic system differently. Therefore, the patterns in response might become an early indicator of a certain neurological disease. This would also bring us closer to an evaluation tool for abstract concept loss for which few options exist.Another important question that remains unanswered specific to the CFR space is how our space would map onto the brain space. It would appear to map well onto the brain space with each of the three factors being part of high level cognitive networks. Our work, however, cannot speak to the relationship between these brain areas and our latent factors. A study which invokes the use of functional connectivity analyses would be required in order to describe the highly distributed networks of these high level cognitive processes.If the CFR space does map onto the brain, then our CFR space could become a powerful computational model that would allow us to understand the way that concepts interact when certain brain areas are damaged. For example, Corticobasal degeneration (CBD) is a disorder which leads to lesioning of the parietal lobe with accompanying magnitude system deficits. This would be akin to lesioning a portion of the magnitude axis in of CFR space. By using the CFR space in this way, we would be able to understand better deficits in concept knowledge in a variety of disorders which would have huge implications for the assessment and treatment of conceptual knowledge.This study was carried out in accordance with the recommendations of the University of Florida, Institutional Review Board with written informed consent from all subjects. All subjects gave written informed consent in accordance with the Declaration of Helsinki. The protocol was approved by the University of Florida, Institutional Review Board.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.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.


More than 2 years have passed since the proposal of the diagnostic criteria for IgG4-related kidney disease (IgG4-RKD). The aim of this study was to estimate the number of histological diagnosis for IgG4-RKD throughout Japan and to clarify the regional distribution of the development of this disease.A questionnaire was supplied to 140 research facilities registered in the Japan Renal Biopsy Registry (J-RBR). The items of the questionnaire were the total number of renal biopsies performed and the number of cases diagnosed as IgG4-RKD in 2012 and 2013 at each facility. Age, sex, and diagnosis category were also included for the IgG4-RKD cases. The geographic distribution of the disease development was evaluated using clinical case reports presented at the Eastern/Western regional meeting of the Japanese Society of Nephrology during the 15 years following 2001.Forty-seven facilities completed the questionnaire, resulting in a collection rate of 34 %. The total numbers of renal biopsies in 2012 and 2013 were 3387 and 3591, respectively. Forty-seven of these cases (24 in 2012 and 23 in 2013) were diagnosed as IgG4-RKD. The frequency of development of IgG4-RKD per one million over 40-year-old individuals during these 15 years varied between 0.9 and 3.1, depending on Japanese geographic region of Japan.The results of the present survey indicate that the number of diagnosis for IgG4-RKD is approximately 130 cases per year throughout Japan, and no regional differences in disease frequency appear to exist. The accumulation of clinical cases with extra-pancreatic lesions associated with autoimmune pancreatitis (AIP) led to the establishment of IgG4-related disease (IgG4-RD) as a new clinical disease. Although the renal disorder was first described in relation to AIP, it is now considered to be a characteristic condition within the category of IgG4-RD , 2. In fMore than 4 years have passed since the WG proposed the diagnostic criteria for IgG4-RKD, and, with the approval of the ethical board of the JSN , the WG has performed a multi-center retrospective study of clinicopathological features of IgG4-RKD. The present study aimed to determine how frequently the diagnosis of IgG4-RKD was made throughout Japan. To address this question, we designed a brief online frequency questionnaire to be administered to Japanese nephrologists.http://www.stat.go.jp/).The questionnaire was supplied to 140 research facilities registered as secondary facilities in the Japan Renal Biopsy Registry (J-RBR), which is a nationwide, web-based, prospective registry system designed to record pathological, clinical, and laboratory data about renal biopsies performed in Japan [t test or Chi square test. The frequency of IgG4-RKD in the 15-year material among the different regions was compared using one-way analysis of variance (ANOVA) and Kruskal–Wallis test. p values of < 0.05 (obtained by two-tailed testing) were considered to indicate statistical significance.Continuous variables are shown as mean values ± standard deviation (SD). Quantitative variables were compared using Student’s A total of 47 nephrologists (44 physicians and 3 pediatricians) affiliated with J-RBR-registered facilities completed the questionnaire (collection rate: 34 %). The total numbers of renal biopsies performed in these facilities in 2012 and 2013 were 3387 and 3591, respectively. Among these patients, 47 (24 in 2012 and 23 in 2013) were diagnosed with IgG4-RKD. This resulted in a rate of 6.7 IgG4-RKD cases per 1000 renal biopsies. Patient informations of age, sex, and diagnosis category were obtained besides one patient. The ages of the 46 patients ranged from 38 to 81 years, and the average and median values were 66.8 ± 9.3 and 67.5 years, respectively. The ratio of men to women was 44:2 .To determine the geographic distribution of the development of this disease in Japan, every case was assigned to a certain region based on the location of the corresponding hospital. The diagnosis frequencies per thousand renal biopsies using the 2-year material and the development frequencies per one million over 40-year-old individuals using the 15-year material were calculated in every district of Japan. Two cases younger than 40-year-old were excluded in this analysis to compare with the middle-age and elderly population size and the total number of cases was 156 in Table . From these data, the number of histological diagnosis for IgG4-RKD can be roughly estimated to be 130 (114–147) cases per year throughout Japan. Most patients with IgG4-RKD have IgG4-related extra-renal lesions, with salivary glands, lacrimal glands, lymph nodes, and the pancreas being most frequently affected [IgG4-RD is characterized by consistent pathological features, including tumefactive lesions and dense lymphoplasmacytic infiltrate rich in IgG4-positive plasma cells with fibrosis across a wide range of organ systems , 2. IgG4. From t30 114–14 cases peaffected . Accordi. The frequency of these diseases increased together with age, 3.9 % in the case of those 60-year- old and older and 7.2 % in the cases of those 80-year-old and older [It was reported that IgG4-RKD is strikingly more predominant in men, and average patient age is about 65 years , 7–10. T. The frnd older . TherefoThe geographical distribution of the development of IgG4-RKD was evaluated using both “2-year” and “15-year” materials (Table In summary, although IgG4-RKD was recently recognized as a clinical entity, and applicable diagnostic criteria and an algorithm were proposed in 2011, epidemiological data are still insufficient. Based on the J-RBR questionnaire utilized in the present study, the number of histological diagnosis of IgG4-RKD in Japan is estimated to be approximately 130 cases per year. There was no significant difference in diagnosis frequency per thousand renal biopsies between Eastern and Western Japan. In addition, the evaluation of available clinical cases reported within the past 15 years shows no regional variations in the development of this disease. Accordingly, we should pay attention to IgG4-RKD as one of the important kidney disease in the elderly distributed all over Japan.


P = 0.023), years of experience (P = 0.007), socioeconomic location of the pharmacy (P = 0.009) and number of clinics near pharmacy (P = 0.008). The results of this study confirmed that dispensing antibiotic without prescription is a common practice in Aleppo pharmacies despite being unlawful. The half of community pharmacists has a poor attitude and inadequate knowledge with regard to antibiotic resistance, reflecting the need for awareness-raising campaigns directed to community pharmacists to equip them for their main role in the community.Antibiotic resistance is a major public health concern. This study was conducted to evaluate the knowledge, attitudes of community pharmacists regarding antibiotic use and potential drug resistance besides assessing their behaviors about dispensing antibiotic without prescription and correlation of the outcomes with demographic variables. A cross-sectional survey was conducted on a random sample of 250 pharmacies in Aleppo, using validated self-administered questionnaire. The total scores of the pharmacistsʹ knowledge and attitudes were correlated with participant demographics using Chi-squared test. One-hundered-seventy-three pharmacies out of 250 agreed to participate in the study. The overall prevalence of dispensing antibiotic without prescription was 85.5%. Only 30.8% of participants exhibited good attitude and 37% had adequate knowledge about antibiotic resistance. Good attitude was strongly correlated with age ( The role of the pharmacist has been changing over the past two decades. He is not just a supplier of drugs and a distributor of medicinal products, but also a team member involved in the provision of healthcare , 2. CommThe major cause of antimicrobial resistance is the inappropriate use of antibiotics, which was found to be directly related to the tendency towards self-medication and the unnecessary use of antibiotics for viral disease .In this connection, WHO report 2014) emphasized on examination and, where necessary, the improvement of the role of the pharmacist as the main supplier and regulator of antibiotics . Most in emphasizPharmacistsʹ knowledge and attitude on antibiotics related issues can greatly influence the way this drug is used in. So, this study aimed at explore pharmacists’ knowledge, attitudes, and dispensing habits with respect to antibiotics and microbial resistance, in view of the potential link between these and the practice of dispensing of antibiotics without a medical prescription.Study designThis was a cross-sectional study conducted among community pharmacies from Aleppo, Syria. Pharmacies included in the study were identified from database of pharmacists syndicate. A pretested, revalidated, self-administered questionnaire was used to collect data.Study toolA random selection was performed of 250 pharmacy from a total of 1700 located in Aleppo city based on the geographical location to be representative of all pharmacists.A pharmacy student visited each selected pharmacy and invited community pharmacists to participate in the study after explaining the aims of the study, participants were told that all information provided was completely confidential and the results would be presented anonymously. During first visit, a self-administered questionnaire which was attached with the consent form was given to each participant. In the second visit, the filled questionnaires were collected from pharmacists who accepted to participate in this study. The questionnaire designed to assess the current knowledge, attitudes and towards bacterial resistance to antibiotic. The questionnaire comprised 21 questions. The first part consisted of two questions; one closed-ended and one open-ended, designed to investigate community pharmacistsʹ behavior toward dispensing antibiotics without prescription for tonsillitis (as example for infectious disease) and possible reasons behind it. The second part consisted of ten questions; five open-ended and five close-ended, which used a three-point scale . This part is about exploring the pharmacistʹs attitude toward providing guidance during dispensing antibiotics. In the third part, pharmacists’ knowledge about antibiotic resistance was measured with twelve three-point scale  questions. Finally, the fourth part is about pharmacist demographic characters. The questionnaire was designed in Arabic then further retranslated into English. The actual survey was conducted during a 2-month period from June to July 2012.Statistical analysisAfter completion of data collection, it was reviewed, organized, tabulated and entered into Statistical Package for Social Sciences  for windows version 18. Descriptive statistics were used to analyze the data . The knowledge scores were computed by scoring 1 for each correct answer and 0 for an incorrect answer with ʹunsureʹ added into incorrect. The attitude scores were computed by scoring 1 for each good behavior and 0 for poor behavior with ʹsometimesʹ added into poor behavior. These scores were then summed up and divided by the total number of test items. A score greater than 70% of the possible maximum score was considered as adequate and good and less than 50% as inadequate or poor for knowledge and behavior respectively, whereas a score between 50 and 70% was considered as moderate. P = 0.05.Statistical associations of demographic variables  and total score of knowledge and attitude were examined by pearson’s chi-squared test. Association was established at Of the 250 pharmacy selected, 27 (13.8%) were working without a pharmacist and 9 (4%) were not located after two attempts. Of the 214 pharmacists met, 41 (17.6%) refused to participate, giving a response rate of 80.8%. The majority (70.5%) of the community pharmacists were males predominantly graduated from Syrian universities. The mean age of the participated pharmacists was 39.8 ± 10.2 (range 25- 65) years old and the average years of pharmacy working experience were 14 ± 7.6 years (range 2-30 years). The other demographic data of participants were shown in Dispensing antibiotics without medical prescriptionP = 0.008) Participants were asked to answer the questions about speculating case of tonsillitis to determine the percentage of pharmacists who sell antibiotics without a medical prescription. The majority of pharmacists admitted that they dispense antibiotics for tonsillitis treatment without physician prescription , 10.7% oPharmacistsʹ attitude To evaluate pharmacist behavior and role in directing the customer to optimal use of antibiotics, five questions were designed; the pharmacistsʹ answers were summarized in with no or sometimes) explained their behavior as they try to prevent bacterial secondary infection. About side effects of antibiotics, (47.4%) of respondent inform the patients about the possible incidence of diarrhea and rash after taking some antibiotics and 58.8% direct the patient to perform hypersensitivity skin test when asking for β-lactam antibiotic injection. Notably, the majority of pharmacists (87.9%) guided the patients to take antibiotic oral capsule treatment instead of injections in the case of minor infections. The pharmacistsʹ attitudes were classified as good, moderate and poor depending on the scores they gained. The results showed that (30.8%) of pharmacists exhibit a good attitude toward guiding customers, whereas (49.1%) and (20.1%) showed moderate and poor attitudes respectively. Four variables  showed a significant association with the participants’ attitudes according to the chi-squared test Pharmacistsʹ KnowledgePharmacists’ knowledge about antibiotic resistance was evaluated by using twelve questions as shown in On the other hand, only 47% think that combination therapy with two antibiotics can reduce resistance development and 90% consider that discovering new antibiotics will solve the resistance problem. Unexpectedly, 71% of participating pharmacists were disagreeing about activation of law that prevents antibiotic sales without prescription. The pharmacistsʹ knowledge was classified as adequate, moderate and inadequate depending on the score they gained. The results showed that (37%) of pharmacists have adequate knowledge, whereas (57.8%) and (5.2%) showed moderate and inadequate knowledge respectively. No association has been found between demographic characters and pharmacists knowledge scores regarding antibiotic resistance according to the chi-squared test Irrational use of antibiotics and weak regulatory enforcement of drug sales are serious issues in developing countries that contribute significantly to bacterial resistance . KnowledThis is the first qualitative study to be conducted in Aleppo, Syria with the aim of exploring pharmacists’ knowledge of and perceptions about antibiotic use and microbial resistance. Aleppo is the largest city in northern Syria. It has about 2.2 million inhabitants. The average number of persons per physician is 790 and 1451 per pharmacist according to Central Bureau of Statistics 2011 . The present findings would be the first step in providing a baseline quantitative data of antibiotics dispensing pattern, knowledge, and attitudes regarding antibiotic resistance among Aleppo pharmacists.The current study revealed a high prevalence (85.5%) and easy availability of these agents over the counter despite being illegal as ministry of health passed a law  determining which drugs could be sold to individuals without medical prescription and that the antibiotics were clearly excluded , Saudi 9The results of the study showed that the knowledge of participants about antibiotics resistance ranged from poor to good. Overall, the theoretical knowledge of participants about antibiotic resistance is inadequate. Despite 58% of pharmacist apparently have a good score, it should be noted that a half of the participants agreed to use antibiotic for common cold infections. This suggests that the pharmacistʹs knowledge may be affected by the fact that the majority ofclinicians were prescribed antibiotics too often in such conditions . It is a well-known fact that resistance to antibiotics has been linked to the consumption of drugs with evidence of a cause-effect relationship -32. HoweNo difference was found between pharmacistsʹ total scores of knowledge other variable indicating that pharmacists in this study are concentrating their efforts to the work in a pharmacy, not to keep up with new information.Among pharmacies chosen for study, there were 27 (13.8%) pharmacy working without a pharmacist and this finding is caused by concern. The staff of these pharmacies contributed to drug misused by providing misinformation about drugs and selling antibiotics according to popular demand , 36. In One third of the pharmacists exhibited a good attitude toward counseling and directing patients although the information provide by pharmacist about is as important as the appropriateness of the medicines themselves. This was in agreement with Indian study which reported that 61% of participating pharmacists considered that patient counseling is their professional responsibility, but only few pharmacists were offering this service . HoweverThe surprising finding was that the majority of participating pharmacists declared disagreement about activation of law that restricts sales of antibiotics without prescription. This might be explained by the fact that activation the law would negatively affect the sales quantities at their pharmacies and reflects pharmacistsʹ commerce orientation even with the evidence of harm based practice. No association was found between sale of antibiotics without prescription conducted by participated pharmacists and their demographic characters, which reflect general behavior due to lack of strict control by the authorities.The current study has several limitations. First, this survey included community pharmacists’ from only one city which limits its generalizability, further research is required to get the views of pharmacists in Syria. Second, as with all self-administered questionnaire based studies, there is the possibility that participants may over-report desirable behaviors or under-report undesirable behaviors. In addition, it may be possible that some respondents reading recommended materials and thus acquired a greater knowledge prior to questioning. As a result, the level of knowledge found in this study may be likely to represent a more favorable picture than would be observed. Howerver, despite the limitations described above, this study considered the first step for evaluating the pharmacistsʹ knowledge and attitude about antibiotic resistance which provides basic information for further studies. This study reveals high rate prevalence of antibiotics dispensing without prescription which calls for imposing strict measures leading to restriction of antibiotic sales. The knowledge and attitudes of the pharmacists participate in this study indicate the urgent need for continuous awareness campaigns directed to pharmacists and public.Educating the pharmacy staff on the risks of antibiotic resistant and emphasis on their role in helping to reduce the prevalence of resistance is an essential issue.Pharmacists have to accept these challenges to ensure that antibiotics are used appropriately even during the self-medication. In addition, there is a need for improving pharmacy curricula in colleges, thereby building correct practice into the profession from the beginning and shifting from the concept of the pharmacy as a ‘shop’ and building a culture of pharmaceutical services.The study was conducted with in-house resources.


Trypanosoma brucei cardiolipin synthesis is essential for survival, indicating that the enzymes involved in cardiolipin production represent potential drug targets. T. brucei cardiolipin synthase (TbCLS) is unique as it belongs to the family of phospholipases D (PLD), harboring a prokaryotic-type cardiolipin synthase (CLS) active site domain. In contrast, most other eukaryotic CLS, including the yeast ortholog ScCrd1, are members of the CDP-alcohol phosphatidyltransferase family. To study if these mechanistically distinct CLS enzymes are able to catalyze cardiolipin production in a cell that normally expresses a different type of CLS, we expressed TbCLS and ScCrd1 in CLS-deficient yeast and trypanosome strains, respectively. Our results show that TbCLS complemented cardiolipin production in CRD1 knockout yeast and partly restored wild-type colony forming capability under stress conditions. Remarkably, CL remodeling appeared to be impaired in the transgenic construct, suggesting that CL production and remodeling are tightly coupled processes that may require a clustering of the involved proteins into specific CL-synthesizing domains. In contrast, no complementation was observed by heterologous expression of ScCrd1 in conditional TbCLS knockout trypanosomes, despite proper mitochondrial targeting of the protein.The glycerophospholipid cardiolipin is a unique constituent of bacterial and mitochondrial membranes. It is involved in forming and stabilizing high molecular mass membrane protein complexes and in maintaining membrane architecture. Absence of cardiolipin leads to reduced efficiency of the electron transport chain, decreased membrane potential, and, ultimately, impaired respiratory metabolism. For the protozoan parasite  Owing to its particular physicochemical properties, it plays an important role in the structural organization and the function of these membranes and strongly enhances the efficiency of the electron transport chain Streptomyces coelicolor and most likely other actinobacteria Trypanosoma brucei and all other trypanosomatids Dictyostelium and in many genera of the group of alveolates, including Plasmodium and Toxoplasma 5Cardiolipin  was identified as the donor of the second phosphatidyl moiety Xanthomonas campestris, which contains six putative CLS genes, a PLD-type bifunctional PE/CL synthase was identified and proposed to synthesize PE from CDP-DAG and ethanolamine as well as CL from CDP-DAG and PG While it has been shown for several bacterial CLS, including T. brucei is endemic in sub-Saharan Africa where it causes sleeping sickness in humans and nagana in livestock. Unlike E. coli or S. cerevisiae, which are viable without a functional CLS, T. brucei fails to survive in absence of TbCLS in culture S. cerevisiae CLS (ScCrd1) are able to functionally complement each other by introducing the genes into the respective  knock-out strains. Cross-species complementation of the two mechanistically different types of CLS has not been reported before.The protozoan parasite S. cerevisiae, we used the commercially available crd1Δ strain Y03840, which is viable under standard growth conditions but exhibits a growth defect when cultured at elevated temperature or on non-fermentable media S. cerevisiae, we fused the first 69 amino acids from the Fo ATPase subunit 9 from Neurospora crassa (in this study abbreviated with Su9) 13TbCLS and a C-terminally tagged form (Su9-TbCLS-HA) were then expressed on an episomal plasmid in the CRD1-deleted strain (crd1Δ). As positive and negative controls for the phenotypic complementation analysis, crd1Δ cells were also transfected with the native ScCRD1 gene and the empty expression vector.To assess the ability of TbCLS to complement ScCrd1-deficient crd1Δ cells transfected with either the empty vector or Su9-TbCLS-HA by immunoblotting. As shown in Figure 2A, a specific band at about 70 kDa was detected in both total lysate and mitochondria-enriched fraction of Su9-TbCLS-HA expressing cells. The apparent molecular mass corresponds to the calculated molecular mass of TbCLS-HA of 74 kDa after cleavage of the Su9 targeting peptide.To confirm expression and mitochondrial targeting of Su9-TbCLS, we analyzed protein extracts from whole cell lysates and isolated mitochondria from crd1Δ by TbCLS, we first performed serial dilutions on SD-agar plates at 30°C and 37°C to compare growth and colony-forming capabilities of wild-type, crd1Δ, crd1[CRD1] and crd1[Su9-TbCLS] strains. As shown in Fig. 2B, at 30°C all transformants grew equally well. In contrast, yeast cells lacking ScCrd1 (crd1Δ) and mutants transfected with the empty vector were unable to grow at 37°C. Interestingly, expression of Su9-TbCLS in crd1Δ yeast was able to restore proliferation and colony formation on plates at 37°C at least partly. In contrast, despite proper expression and mitochondrial localization , Su9-TbCLS-HA was unable to restore growth and colony formation of crd1Δ[Su9-TbCLS-HA] transformants at 37°C (data not shown), suggesting that the HA-tag interfered with the function of TbCLS.To test functional complementation of crd1Δ strains on a molecular level, we isolated lipids from spheroplasts and crude mitochondrial preparations and subjected them to MALDI-TOF MS (Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry) analysis. Figure 2C summarizes the observed CL molecular species distribution in extracts from wild-type, crd1Δ, crd1Δ[CRD1], and crd1Δ[Su9-TbCLS] strains from two independent experiments. Representative spectra from wild-type, crd1Δ, and crd1Δ[Su9-TbCLS] strains are shown in Fig. 2D. The results show that wild-type yeast produced the full spectrum of CL molecular species reported before 1516crd1Δ cells . In contrast, in Su9-TbCls-expressing crd1Δ cells, the CL molecular species with fatty acyl chains with 64, 66, and 68 carbon atoms were present , demonstrating that CL synthesis was restored by Su9-TbCls in these cells. However, the masses of the CL molecular species differed by 2 dalton between wild-type and Su9-TbCls-expressing crd1Δ cells. In wild-type yeast, the most prominent species were at m/z 1372.2 (representing CL(66:4)) and 1400.3 (representing CL(68:4)) while in crd1Δ cells complemented with Su9-TbCls the major species were at m/z 1374.7 (representing CL(66:3)) and 1402.8 (representing CL(68:3)). The abundance of CL molecular species relative to other phospholipids was substantially lower in Su9-TbCls complemented cells compared to wild-type or crd1Δ[CRD1] cells, suggesting that the expression level or activity of Su9-TbCls was lower than that of ScCrd1.To confirm CL synthesis by Su9-TbCls-expressing cld1Δ and, to some extent, taz1Δ mutants, in which remodeling of premature CL species is impaired due to the absence of the CL-specific deacylase ScCld1 1717The prevalence of triple-unsaturated CL species is reminiscent of the CL molecular species composition of the TbCLS knockout cell line of T. brucei procyclic forms, where the expression of the ectopic TbCLS recovery gene can be switched off by removing tetracycline from the growth medium ScCRD1 or cMyc-tagged ScCRD1 were stably integrated into the conditional TbCLS knockout genome along with the selectable marker gene SAT1 (conferring nourseothricin resistance) and under the control of a constitutively active procyclin promoter. Transformed cells were selected in the presence of the antibiotic nourseothricin after limiting dilution in 24 well plates and genotyped by PCR.In contrast to yeast, CL synthesis in trypanosomes is essential for cell proliferation and survival ScCRD1-cMyc . The subcellular localization of ScCrd1-cMyc in T. brucei was analyzed by immunofluorescence microscopy using antibodies against the cMyc epitope and against a protein of the archaic translocase of the outer mitochondrial membrane (ATOM) as mitochondrial marker Transcription of untagged ScCrd1 was examined by RT-PCR, confirming the presence of ScCrd1 mRNA. Expression of cMyc-tagged ScCrd1 on a protein level was confirmed by SDS-PAGE and immunoblotting of protein extracts from cells transformed with TbCLS conditional knock out cells, proliferation in absence of tetracycline ceased after 48 h, followed by a continuous decrease in cell number and death of the culture. A similar observation was made for trypanosomes expressing cMyc-tagged ScCrd1 (not shown). Together, these data show that both untagged and tagged ScCrd1 are unable to complement growth of T. brucei procyclic forms after ablation of TbCLS, despite proper mitochondrial localization.To test viability of the transformants in absence of TbCLS, tetracycline was removed from the culture medium to ablate TbCLS expression T. brucei are conceivable. Firstly, failure to cleave the mitochondrial targeting peptide in T. brucei may impair enzyme activity. As predicted by TargetP 1.1 21T. brucei parasites and may not be (fully) active.Several reasons for the inability of ScCrd1 to complement TbCLS in S. cerevisiae is embedded in a protein complex and that its activity is substantially reduced upon purification T. brucei or may not associate with ScCrd1. Conversely, since TbCLS is also part of a larger mitochondrial complex T. brucei.Secondly, the finding that ScCrd1 in E. coli and mammals. While the active site of E. coli CLS is most likely directed towards the periplasm T. brucei may be impaired. In addition, no mitochondrial CDP-DAG synthase has been identified in T. brucei T. brucei. Thus, it is possible that ScCRD1 may not be active since it has no access to its substrate CDP-DAG. Note that bacterial-type CLS do not depend on CDP-DAG as substrate for CL synthesis, i.e. a lack of mitochondrial CDP-DAG may not hamper CL formation by TbCLS in T. brucei.Thirdly, Crd1 may not have access to its substrates, CDP-DAG and PG. It is known that the orientation of CLS differs between CRD1 knockout cells under stress conditions, whereas CAP-type ScCrd1 fails to substitute the essential function of TbCLS in trypanosomes.In this study we investigated the ability of two structurally non-related forms of eukaryotic CL synthases to functionally complement each other in cells lacking the native CLS. We found that PLD-type TbCLS is functional in yeast, alleviating the proliferation deficiency of S. cerevisiae. A similar shift to a higher degree of saturation also occurs in CL from yeast strains with deficient CL-remodeling. The apparent lack of CL maturation in TbCLS-complemented crd1Δ cells suggests that biosynthesis and remodeling are tightly coupled processes, probably depending on a specific microenvironment and proper targeting of the involved proteins to specialized membrane domains. The non-homologous TbCLS may lack the property to localize correctly to such a microdomain or physically interact with the remodeling machinery.We observed that CL produced by TbCLS in yeast contains less unsaturated fatty acids compared to natural mature CL of E. coli XL-1 were used for molecular cloning. Growth media were prepared in the lab except for SDM79, for which a customized medium powder mix (Amimed®SDM79) was purchased from BioConcept .All reagents were of analytical grade and, unless otherwise stated, purchased from Merck-Sigma Aldrich . Kits for plasmid DNA extractions and PCR purification, Pfu polymerase and T4 DNA ligase were from Promega . Restriction enzymes were purchased from ThermoFisher Scientific . Home-made chemically competent TbCLS conditional knockout cell line containing a tetracycline-inducible copy of C-terminally hemagglutinin-tagged TbCLS has been described before The procyclic form ScCRD1 or ScCRD1-cMyc into the TbCLS conditional knock-out cell line, plasmid pG-EGFPΔLIIβ NEO) resistance marker by the nourseothricin resistance gene SAT1. pKOS SAT1. The triple-cMyc tag with flexible linker sequence to generate cMyc-tagged CRD1 was PCR-amplified from the vector pJM1 .For stable integration of TbCLS conditional knockout parasites were cultured at 27°C in SDM79 supplemented with 15% (vol/vol) heat-inactivated FBS , 160 µM hemin, 90 µM folic acid, 15 µg/ml G418, 2 µg/ml puromycin, 5 µg/ml blasticidin, 0.2 µg/ml phleomycin, and 1 µg/ml tetracycline to induce the expression of ectopic TbCLS. ScCRD1-complemented TbCLS conditional knockout cells were cultured in the same medium with an additional 150 µg/ml nourseothricin .crd1Δ (ORF YDL142c) strain Y03840  created in the course of the Saccharomyces Genome Deletion Project was purchased from Euroscarf and was used to test functional complementation with TbCLS.Designer deletion strain BY4741, referred to as wild type (WT) strain in this study, was used as control strain. The haploid URA3 selection marker had been replaced by HIS3, was used for complementation of strain Y03840 with TbCLS. Another high copy number YEp, pDR197  pVT-H, a modified version of expression vector pVT-U S. cerevisiae strains. For selections and large cultures of auxotrophic transformants, synthetic defined drop-out media (SD/DO) were used, composed of 0.67% yeast nitrogen base without amino acids, 2% dextrose, and the appropriate amino acids according to the Yeast Protocols Handbook (Clontech Laboratories) Standard YPD growth medium YPD agar plates were used for general culture and maintenance of SAT1 was amplified by PCR from plasmid pKOS using primer 1 and 2 (see table S1), digested with NheI and ClaI (dam sensitive) and ligated into the equally digested vector pG EGFPΔLIIβ isolated from damE. coli strain GM48. The resulting vector was named pGS-EGFPΔLIIβ.Exchange of resistance marker of pG EGFPΔLIIβ ScCRD1 was amplified by PCR from isolated genomic DNA from strain BY4741 using primer 3 and 4 that were designed to replace EGFP from pGS-EGFPΔLIIβ by CRD1 flanked by new multiple cloning sites.CRD1-cMyc, CRD1 was first amplified using primer 5 and 6 from pGS-CRD1, digested with HindIII and BamHI and ligated into equally digested vector pJM1, fusing CRD1 to a C-terminal triple-cMyc-tag. The fusion-sequence was subsequently PCR-amplified again using primer 5 and 7 followed by digestion with HindIII and PstI. Ligation with HindIII/PstI digested pGS-EGFPΔLIIβ resulted in pGS-CRD1-cMyc.For the construction of CRD1(-cMyc) and SAT1 under the control of the constitutively active procyclin promoter and a modified procyclin 3’ UTR. Digestion with KpnI and NotI allows genomic integration of the construct via homologous recombination with a procyclin locus.The resulting vectors pGS-CRD1 and pGS-CRD1-cMyc contained TbCLS, was amplified from purified genomic DNA from T. brucei strain Lister 427 by PCR using primers 8 and 9 for the untagged copy and primer 10 for the construction of a C-terminally HA-tagged version. In both versions the thymidine base of the BamHI restriction site close to the end of the TbCLS ORF was mutated to cytidine.ORF Tb427.4.2560, encoding o ATPase subunit 9 from Neurospora crassa, in this study abbreviated with Su9, was PCR-amplified using primers 11 and 12 from template plasmid pGEM4 Su9(1-69)-DHFR Su9 and TbCLS-HA into the multiple cloning site of pDR197, resulting in the expression vector pDR-Su9TbCLS-HA.The FScCRD1 in crd1Δ, ORF YDL142c from S. cerevisiae was amplified by PCR from isolated genomic DNA from strain BY4741 using primers 13 and 14 and ligated into pDR197.For the expression of an addback copy of BamHI and XbaI and ligated into equally digested pVT-H.For the construction of untagged pVT H-Su9TbCLS constructs, the Su9 fusion sequences were PCR-amplified from pDR-Su9TbCLS-HA using primers 15 and 16, digested with 7 cells. Electroporation was performed in 100 µl Nucleocuvettes™ using a Lonza 4D Nucleofector™  as described in Trypanosomes were grown to mid-log phase, harvested by centrifugation and resuspended in trypanosome transformation buffer 7 cells/ml and then washed in sterile water. Aliquots of 108 cells were transferred to 1.5 ml tubes, pelleted and resuspended in 360 µl of transformation mixture containing 240 µl 50% PEG 3500 (w/v), 36 µl 1 M LiOAc, 50 µl 2.0 mg/ml single-stranded carrier DNA, 34 µl plasmid DNA (0.1 - 10 µg). Following incubation at 42°C for 40 min, cells were pelleted in a microcentrifuge, gently resuspended in 1 ml sterile water and plated onto selective medium.Transformation of yeast was performed according to the high efficiency transformation protocol by Gietz & Woods (2002) crd1Δ strain by TbCls was analyzed by making use of the temperature sensitive phenotype of the crd1Δ strain. Yeast WT, crd1Δ, crd1Δ[CRD1], crd1Δ[Su9-TbCLS], and crd1Δ [empty vector] strains were cultured in SD complete or the appropriate drop-out medium to an optical density (OD600) of 0.1, serially diluted by factors of ten and spotted on SD complete agar plates. Plates were incubated at 30°C or 37°C for 3 - 4 days and growth of the respective strains was compared between control (30°C) and stress (37°C) temperature.Functional complementation of the et al.600) of 0.8 - 2, washed 2 x with water and incubated in buffer containing 100 mM Tris/H2SO4 (pH 9.4) and 10 mM dithiothreitol [2 ml/g (wet weight) cells] for 20 min at 30°C with gentle shaking. Subsequently, cells were pelleted, washed, and resuspended in zymolyase buffer  [7 ml of buffer/g (wet weight) cells]. 2 mg/g (wet weight) cells of zymolyase 20-T  was added and the suspension was incubated with gentle shaking at 30°C until the OD600 reached 20% of the initial value (20 - 40 min). Isolation of crude mitochondria was done exactly following the protocol of Gregg et al.Yeast spheroplasts and mitochondria-enriched fractions were prepared according to Gregg g for 15 min. The organic phase was collected, dried under a stream of nitrogen and washed 1 x with a mixture of 0.1 N HCl:chloroform:methanol . The washed and dried organic phase was dissolved in 1 ml chloroform: methanol  and subjected to mass spectrometry.Lipids were extracted from washed spheroplasts (zymolyase buffer without zymolyase) or crude mitochondria using a modification of the protocol published by Bligh and Dyer et al. (2012) MALDI-TOF MS was performed according to Angelini 7 cells/ml were harvested by centrifugation, washed with Tris-buffered saline  and resuspended in SoTE buffer  to a concentration of 108 cells/ml. Digitonin was added to the cell suspension at a final concentration of 0.25 mg/ml to lyse the cells. After incubation on ice for 5 min, non-solubilized membranes (including mitochondria) and cell debris were pelleted by centrifugation at 7000 x g in a table-top centrifuge, resuspended in sample buffer containing 2.5% SDS and boiled at 95°C for 5 min.Trypanosomes grown to a concentration of 10Yeast proteins from whole cells were extracted according to Kushnirov (2000) Proteins were separated on 12% polyacrylamide gels and transferred to polyvinylidene difluoride membranes (Millipore) by semi-dry blotting (Bio-Rad). Tagged proteins were visualized on films using SuperSignal West Pico (Pierce) chemiluminescent substrate after incubation with tag-specific primary antibodies and horseradish peroxidase-conjugated secondary antibodies. Antibodies used in this study were: mouse monoclonal antibody anti-cMyc ; mouse monoclonal antibody anti-HA ; rat polyclonal anti-p20 antibody ® goat anti-mouse 488 and goat anti-rabbit 594  were added for another 45 min. Following washing with PBS, slides were dried and coverslips were mounted with Vectashield® containing 4’, 6-diamidino-2-phenylindole .Slides for immunofluorescence microscopy of trypanosomes were prepared as described in Fluorescence microscopy was performed with a Leica DM 16000 B inverted microscope using a 60 x immersion objective and pictures were acquired using a connected Leica DFC360 FX camera. Image deconvolution and further processing was performed using Leica LAS X software and Fiji (NIH), respectively.Click here for supplemental data file.http://microbialcell.com/researcharticles/cross-species-complementation-of-bacterial-and-eukaryotic-type-cardiolipin-synthases/.All supplemental data for this article are also available online at


Human activity recognition, tracking and classification is an essential trend in assisted living systems that can help support elderly people with their daily activities. Traditional activity recognition approaches depend on vision-based or sensor-based techniques. Nowadays, a novel promising technique has obtained more attention, namely device-free human activity recognition that neither requires the target object to wear or carry a device nor install cameras in a perceived area. The device-free technique for activity recognition uses only the signals of common wireless local area network (WLAN) devices available everywhere. In this paper, we present a novel elderly activities recognition system by leveraging the fluctuation of the wireless signals caused by human motion. We present an efficient method to select the correct data from the Channel State Information (CSI) streams that were neglected in previous approaches. We apply a Principle Component Analysis method that exposes the useful information from raw CSI. Thereafter, Forest Decision (FD) is adopted to classify the proposed activities and has gained a high accuracy rate. Extensive experiments have been conducted in an indoor environment to test the feasibility of the proposed system with a total of five volunteer users. The evaluation shows that the proposed system is applicable and robust to electromagnetic noise. For decades, human activity recognition has been an important area of human-computer interfaces (HCI). Recent advances in human activity recognition (HAR) and sensing technologies have presented various ideas aiming to enable novel applications in different areas such as healthcare, security, and entertainment ,2. A hugHowever, such approaches depend on a special device that may be considered as the main drawback of these systems, for instance: a vision-based system requires a camera to monitor human activity, which causes a privacy concern for the target people. Furthermore, the camera is not suitable in some special places such as a “bathroom”. In a bathroom, the activity monitoring is very important in special cases such as elderly fall detection because a bathroom is a potential place for falling. Moreover, the camera cannot traverse through walls. In addition, the camera requires good lighting conditions. The second type of traditional approach is a sensor-based activity recognition system. Such a type is inappropriate for human usage because sometimes elderly people may forget to carry or wear such sensors. Furthermore, all device-based activity recognition systems whether vision-based or sensor-based are not free and require expensive installation, maintenance, and more exhaustive work.The recent trend in HAR is wireless-based sensing technology or so-called device-free activity recognition that uses only the emitted signals from a wireless transmitter such as wireless local area network (WLAN) 811.b.n.c. Earlier, Woyach et al.  proved tMost recent, Channel State Information (CSI) of the physical layer of the radio system has been used in tracking human motion and activity. CSI has attracted more attention in recent years since CSI can be exported from commodity wireless Network Interface Controller (NIC) . CSI refWu et al.  presenteUnlike previous approaches, in this paper, we extract both amplitude and phase information of CSI. Moreover, we focus on extracting the CSI information in a realistic environment, which means in the existence of different radio device working in the test area at the same time of the testing experiments. The surrounding devices cause real noise to the CSI, and such noise cannot be removed by filtering because such noise affected some CSI streams and distorted the real trend of them. We observed that CSI streams have a distinctive sensitivity to the human motion. In some streams, the affection of human motion is less than others, and the affection of surrounding noise is more than other streams. Therefore, the fluctuation of such streams due to the human motion is not clear. Previous approaches such as  extract Thereafter, we extract features from both time and frequency domains of principal components of CSI. Finally, our system uses Random Forest classification to recognize implemented activities. Experimentally, we find that the CSI-PCA method promotes the classification accuracy.We present a device-free activity recognition system, which requires only a ubiquitous Wi-Fi router, where the user is free and does not need to wear sensors or be monitored by a camera. The proposed system can classify seven daily activities in line-of-sight (LOS) and none-line-of-sight (NLOS) scenarios.Unlike previous CSI-based human activity recognition approaches, we extract both CSI amplitude and phase information, thus providing us with more information.We present an efficient CSI-PCA method to reduce dimensionality and to expose the real trend of the CSI stream that is caused by human motion.We adopt the Random Forest classification algorithm to classify the proposed activities.We present empirical results of various activities during exhaustive experiments conducted in a real-word environment and in different scenarios.Our main contributions can be summarized as follows:The reminder of this paper is organized as follows. In the next section, we describe the system architecture and methodology. This section describes the core part of the proposed system. In what follows, we give a detailed breakdown of our system architecture and implementation. One of the key points of elderly activity recognition systems based on CSI is to extract the feature of each motion from filtered raw CSI after applying the PCA algorithm across CSI streams to expose information that we need. Therefore, our novel system consists of the following modules: the CSI Preprocessing module, Feature extraction module and Classification module as shown in The CSI preprocessing module carries out some basic processes to extract amplitude and phase information that we use. An exponential filter is also implemented to filter out electromagnetic interference caused by 2.4 GHz devices around the testing environment. The main purpose of the PCA is to reduce dimensionality and to expose the real trend of CSI. Since the speed and duration of each motion are different, we can obtain a dynamic time window via the analysis of each motion for extracting features in the latter module. In the last module, we test the classification algorithm on the training and testing sets.Y and X represent the i is the number of OFDM subcarriers and N is the noise in each subcarrier. CSI can be represented as Equation (2):j is the number of CSI streams. The measurement of CSI depends on the amplitude and phase of CSI at a time point. If we have a number of transmitter antennas The proposed system collects radio signals from the fixed access point in the test environment. Let IWL 5300 . In thisMany methods can be applied to represent the collected CSI data. Here, to reduce computational complexity and reduce data dimensions, we use the mean of the 30 subcarriers of each stream as a vector that represents CSI. Thus, in our system, CSI is measured as six streams. The PCA  is a usep principle components, as shown in The fluctuation of CSI measurements caused by human motion occurs across all the streams and subcarriers in the wireless channel. The different streams have a distinctive sensitivity to the fluctuation of CSI due to human motion as shown in We apply a moving window across CSI principal components (CSI-PCs) to calculate the following features: (1) normalized standard deviation (STD); (2) median absolute deviation (MAD); (3) interquartile range (IR); (4) signal entropy; and (5) duration of human motion. Features (1), (2), (3) expose the differences of different motions in time domain distribution of CSI. Signal entropy reflects different amounts of information in different motions. Duration is an important and useful feature in our system, since it varies widely over different actions.We build our prototype with the Random Forest classification method. Random Forest was invented by Breiman , and canWe implement all the experiments in a laboratory building with a lot of devices working at 2.4 GHz. Multipath signals are very rich in this building. During the experiments, we set the period of each experiment to 30 s, and the motion happens beginning about 15 s. The packet rate is 100 Hz, so that we can get enough information of each motion. In order to detect fall from daily life motions, we test seven motions: walk, sit down, lie, stand up, squat down , fall, and crawl.We implemented our system using off-the-shelf devices. The system consists of two devices: one WiFi access point (AP) which serves as the transmitter and has three transmitter antennas, and one HP laptop equipped with an Intel WiFi link IWL 5300 802.11n chipset, which serves as a detection point (DP) and has three receiving antennas. The laptop installed Ubuntu 14.04 with a modified Intel NIC driver.The software that was used in our experiments is the open source CSI-Tools presented by Halperin et al. . After dWe conducted experiments with five different volunteer users aged between 22 to 33 years. Each volunteer user was asked to implement the activities described in LOS scenario: in this scenario, the target user, the detection point (DP) and the access point (AP) are placed in the same room as shown in NLOS scenario: in this scenario, the access point (AP) is fixed in a room where the target user and the detection point (DP) are placed in another room as shown in The proposed activities implemented in two scenarios:We collected 500 samples of each activity from five users divided into five groups. In each group, 100 samples of each activity were collected from five users as 20 samples of each activity per one user as described in Thereafter, we build our classifier in each group with 100 samples of each activity, and we measure the five-cross validation accuracy. Each user implements the intended activity individually close to the detection point as shown in Moreover, to evaluate the performance of the proposed system, precision, recall, and F1 score are used to analyze results of the experiments. Precision is the positive predictive value, recall is the sensitivity, and F1 score or the F-measure is the weighted average of both the precision and the recall. These three mathematics rules are used as expressed in the following expressions:To show the validity of the Principal Component Analysis (PCA) method, we measure the precision of each implemented activity in both LOS and NLOS scenarios with PCA and without applying the PCA method. It is very important to test the difference between both device-free sensing metrics, namely RSSI and CSI, since RSSI-based device-free activity recognition systems were adopted earlier. In order to also prove the superiority of CSI over RSSI, we experimentally compare the proposed seven activities in LOS scenarios based on RSSI and CSI.The main advantage of RSSI over CSI is that RSSI is available in wireless devices, whereas CSI still obtained only with specific NIC cards such as IWL 5300.We compare Random Forest with popular classification algorithms such as Support Vector Machine (SVM) and Naive Bayesian Classifier (NBC). We test the accuracy of the three classification algorithms of all the collected data in LOS scenarios. Overall results showed the validity and stability of our proposed system. The second stage of this system is to extend it to classify all potential human activities and to test multiple people activities at the same time. An important issue to be addressed in the future work is to test human activity in case of interference of moving objects in the test area.In this paper, the proposed human activity recognition system is motivated by a need for decreasing the installation complexity and development costs induced by the traditional device-based activity recognition systems. As described above, the proposed system can recognize seven daily activities by analyzing the channel state information of Wi-Fi signals with only a ubiquitous Wi-Fi router as an access point and a laptop as a detection point. As mentioned in the results section, the system has gained a high accuracy rate that justified the validity and stability of the proposed methodology.The presence of moving objects in the test area. This is a complex challenge since CSI is very sensitive to an object’s motion, especially if the moving object is a big object like a big pet .Since a device-free concept means no need for using special devices in the monitored area or on the perceived object body, it is a challenge to detect two or more human actions at the same time. The solution of such a problem depends on more improvement to the wireless sensing technique.CSI outperforms RSS in tracking human motion and activity classification since CSI has a unique capability to eliminate multipath effects and is more robust to environmental changes due to the fact that CSI has more information than RSS since CSI measured per OFDM subcarrier while RSS measured per packet. However, CSI can be obtained only on specific hardware such as IWL 5300 NICs. Thus, it is not appropriate to be utilized in many devices such as tablets and smart mobile devices. Such a limitation needs more development in receiver antenna hardware.The device-free CSI-based sensing technique is a promising technique, which may go far in the future. However, there are three main challenges for further study. Here, we highlight such challenges as the following:By solving such challenges, the device-free wireless-based sensing technology will be generalized and be available to track human motion and activity. We plan to solve the first and second challenges in our next stage of study, whereas the third challenge needs a manufacturing solution.In this paper, we have presented a device-free activity recognition system using the CSI of Wi-Fi signals. We have extracted the amplitude and phase information from the CSI of Wi-Fi signals. We have designed a low-pass filter to remove the electromagnetic noise to obtain the real trend of CSI caused by human motion. To reduce CSI dimensionality and to choose useful information across all CSI streams, we have used the PCA algorithm, and used PC vectors to be CSI representatives. Thereafter, we have extracted features from PCs that were labeled as inputs to the classification algorithm. The Random Forest algorithm has been adopted to classify the proposed activities. We have experimentally conducted our system in two different scenarios with several volunteers. The results have verified the validity of our system in both the LOS and NLOS scenarios as 95.43% and 91.4%, respectively.


There have been concerns about the recent private turn and re-emergence of philanthropies in world health, with many worrying about philanthropies’ perceived lack of transparency and accountability. In contrast, I argue that while the private turn might have led to a decline in democratic or public accountability, it did not bring an end to all forms of accountability. Specifically, I suggest that philanthropists’ involvement in global health has led to the spread of another, new form of accountability: epidemiological accountability. The latter is a combination of two regimes of expertise and practices hitherto kept separate: audit and epidemiology. To substantiate this argument, I draw on my research on the Bloomberg Initiative – a global effort to reduce tobacco use spearheaded by the Bloomberg and Gates foundations. In God we trust; everyone else bring data. et al., et al., The shift from international to global health at the turn of the twenty-first century has been characterized by a private turn and the re-emergence of philanthropic foundations as major actors in world health, a trend symbolized by the Gates Foundation . Compiled by the WHO every two years, the report monitors the worldwide progress of tobacco control policies by counting the number of countries that have implemented and the percentage of the globe's population covered by at least one of the MPOWER policies WHO, . The secEvaluation [is] a collaborative learning tool that provides us and our partners with feedback so we can learn, adjust and decide how best to achieve outcomes … [It] provides feedback on what is and is not working. (Gates Foundation, In addition, the regular assessments carried out by the two foundations also provide them with the necessary information to showcase their work to the global health community and beyond in annual reports, public presentations and media briefings (e.g. Bloomberg Philanthropies, It is standard policy for both the Bloomberg and Gates foundations to continually evaluate the projects in which they invest. When describing their ‘approach’ to giving in their publications, Bloomberg Philanthropies , p. 7 alWhen we are reporting back to Mike [Bloomberg] about our progress and our success in the Initiative … we talk about the training of journalists and how many stories have come out in support of tobacco control around the world; we look at all of the efforts you were involved in; and we count all of these on a regular basis.A good illustration of these counting practices can be found in a recent Bloomberg Philanthropies (see To date, 300,000 individuals … have completed the GATS … Funded partner organizations and in-country grantees [have]: … completed air quality monitoring studies in 34 countries; … developed a technical guide … [on] warning labels; launched 64 anti-tobacco … mass media campaigns in 18 countries, viewed by 643 million people; … conducted workshops with journalists from 28 countries on tobacco control issues; … provided legal expertise … [in] 55 countries; … [and] published 13 white papers on tobacco economics. In addition, … almost 300 senior-level government and civil society representatives from 34 countries have been trained through an annual intensive leadership program for tobacco control [and] more than 5,000 [individuals] from 172 countries participated in a free online training. (Bloomberg Philanthropies, The third and last category is ‘outcomes’ or ‘impacts’. As an epidemiologist working for the Initiative explained to me, the ‘ultimate impact is reducing deaths’. In the absence of reliable data on global smoking-attributable mortality, the Initiative uses computer simulation models, such as the ‘SimSmoke’ model elaborated by American economist David Levy and his colleagues, to estimate the number of lives saved through its investments and activities (e.g. Dubray et al., et al., et al., et al., We have made quite a bit of progress in the eight years since we started. We have had quite an impact. There has been a lot of success in recent years as the growing numbers of countries having adopted one MPOWER policy shows … And we estimated that the lives saved based on the policy progress we have made … We had expected some increases … but we were absolutely astounded … to see that all the policy progress made … had doubled our lives saved estimates, from 7 to 14 million.While the aim of this paper is not to expose the shortcomings of the quantification techniques used in the Bloomberg Initiative, it is still worth noting here that the methodology and computer simulation models underpinning ‘saved lives’ metrics are not without problems. Indeed, as McCoy, Jensen et al. (et al., et al., et al., et al., et al., Both the Bloomberg and the Gates foundations like to think of their evaluation protocols as quasi-scientific processes characterized by systematicity, objectivity and rigour. So, for example, in an interview with me, an expert at Bloomberg Philanthropies explained that the regular assessment of their projects was based on ‘very rigorous monitoring and surveillance’. Likewise, in a recent document on its evaluation policy, the Gates Foundation , p. 1 supies see  publicatThe values and practices developed by Bill Gates and Michael Bloomberg in their business ventures, their philanthropic work and, for Bloomberg, in public service offer insights into how audit and epidemiology, both alone and in combination, became central to the management of the Initiative. For both Gates and Bloomberg, information or data has been at the heart of their work at Microsoft Corporation and Bloomberg L.P., respectively. To start with, information and information technology was central to what their companies did. Writing in the 1990s, Gates , 2008b. Know your numbers is a basic business principle. You need to gather your business's data at every step of the way and in every interaction with your customers – and with your partners, too. Then you need to understand what the data means … If you do not understand what is happening in your business factually, and you are making decisions based on soft data or emotion, you will eventually pay a big price. Numbers give you the factual basis for the directions in which you take your products … They help you identify your highest priorities. (Gates, Bloomberg had very similar values and beliefs, repeatedly stressing that ‘following the data’ was the motto at the heart of his company and famously stating that: ‘In God we trust; everyone else bring data’ (Bloomberg Philanthropies, Moreover, echoing the cultures of audit deployed in the administration of commercial and corporate life in the twentieth century, Gates and Bloomberg also understood information as critical to good management and a successful business. Gates, in particular, was often viewed as ‘one of the high priests’ of this new culture of audit ‘built around the massive gathering and meticulous analysis of numerical data’, with managers at Microsoft expected to ‘crunch numbers for everything before they made decisions’ and ‘deliver quantifiable results for each dollar spent’ Smith , p. 148.The common sense of the business world, with its urgency and focus, has strong application in the philanthropic world. I am sure I will make mistakes in over-applying some elements from my previous experience and will need to adjust … However, I am equally confident that our maniacal focus on … measuring results will make a difference. Gates, , p. 2You can achieve amazing progress [in your philanthropic endeavours] if you set a clear goal and find a measure that will drive progress towards that goal … Given a goal, you decide on what key variable you need to change to achieve it – the same way a business picks objectives for inside the company … – and develop a plan for change and a way of measuring the change. You use the measurement as feedback to make adjustments. Gates, , pp. 2–3Unsurprisingly perhaps, Gates and Bloomberg brought their belief in data and audit that had been so central to their businesses over into their philanthropic work. Discussing the similarities and differences between his work at Microsoft and running a foundation, Gates declared that many of the lessons he had learned in business, and especially his concern with data and measurement, would apply to philanthropy:Data is the driving force behind my company's value to the financial services industry … Bloomberg Philanthropies is no different. It harnesses the power of data to assess opportunities, understand impact, evaluate results and improve performance. (Bloomberg Philanthropies, Of course, Gates and Bloomberg were not the only ones applying their belief in the power of data and audit acquired in the business world to their philanthropic work. Rather, as Matthew Bishop and Michael Green (Bloomberg was, if anything, even more confident about applying the insights he picked up in his business and, specifically, his belief in data and evaluation to his philanthropic work:el Green , pp. 1–2el Green .Unlike business, where profit is the bottom line, foundations … pick their own goals [and make that their bottom line]. Gates, , p. 2[A] way that running a foundation is not like running a business is that you don't have customers who beat you up when you get things wrong or competitors who work to take those customers away from you. You don't have a stock price that goes up and down to tell you how you are doing. This lack of natural feedback loop means that we as a foundation have to be even more careful in picking our goals and being honest with ourselves when we are not achieving them. We work hard to get … feedback. Gates, , p. 14This explains why the Gates Foundation has spent a lot of time and money in identifying clear, measurable alternative ‘bottom lines’ to assess its philanthropic investments. In the field of education, this has led it to get states in the United States to ‘agree to common rules on how to measure graduation rates and on what standards must be achieved to graduate’ (Bishop & Green, The metric of success is lives saved … Which is slightly different than units sold [or] profits achieved. But it's all very measurable [through epidemiology], and you can set ambitious goals and see how you do. (Gates cited in Smith, For Bloomberg, in contrast, his discovery and interest in epidemiology developed during his time as Mayor of New York City. As Julian Brash (Bloomberg is 100% about data. His entire business is built on data. At City Hall he implemented all sorts of data-driven initiatives. This is a person very interested in data. Similarly, I am a trained epidemiologist and my focus is also on data … We both just want to know what impact we are having. That is why we fund measurement strategies like the GATS.It is also because of Gates and Bloomberg's belief in data and measurement that epidemiological expertise and practices became so central to the Bloomberg Initiative. But here the precise pathways differ for the two philanthropists. For Gates, epidemiology was the response to his search for the ultimate yardstick of success or failure in his philanthropic work on global health. Gates quickly realized that while he could transfer the techniques of audit like reports, indicators and evaluations from his business to his foundation, he could not do the same with the notion of profit. Profit in business was the bottom-line, the definitive measure of whether what you did worked or not. The fact that it did not apply to philanthropic work was a problem for Gates in so far as he was left without any criteria to judge whether his efforts were making a difference or not. As he argued:an Brash  explainsContrary to what is often argued in the more critically minded literature on global health, I suggested that philanthropists’ growing influence in the organization and management of world health has not led to the disappearance of all forms of accountability. Specifically, and focusing on the Bloomberg Initiative spearheaded by the Bloomberg and Gates foundations, I have argued that the increasing involvement of philanthropists in global health has led to the development of a new form of accountability – epidemiological accountability – which combines the expertise and practices of audit with those of modern epidemiology. In these concluding remarks, I want to draw some of the broader implications that my argument might have for those of us in the social sciences and humanities who work on and write about the languages and cultures of accountability that permeate the fields of global health, international development and beyond.Postscript on the societies of control, the ‘continuous forms of control’ and the ‘perpetual training’ logic associated with the Initiative – the constant self-recording and reporting; the regular inspections during which accountants and management experts offer advice and counsel on how to organize and work; and the training courses in project management and accounting (see also Jensen & Winthereik, When faced with a new form of accountability like epidemiological accountability, one question that some might be tempted to ask is whether this new form of accountability works at all. This, I believe, would be asking the wrong question. Indeed, such a question fails to unpack the key ideas and practices – the desire to save lives; the drive for effectiveness; the trust in numbers; the monitoring systems and evaluation procedures – around which epidemiological accountability is articulated. Put differently, to ask such a question would mean evaluating whether this new form of accountability really makes saving lives and the management of world health more effective, thus very much operating from within the epidemiological accountability episteme and reproducing rather than interrogating its core assumptions. A more fruitful approach, I suggest, would be to ask what this new form of accountability produces. This question shifts our analytical gaze from epidemiological accountability's possible shortcomings to its performative power and the new forms of knowledge and subjectivity it generates (Desrosières, Another aspect of epidemiological accountability that I would like to emphasize is the way it has transformed audit from a profit-making tool of late-twentieth-century corporate capitalism into a biopolitical instrument of modern philanthropy. This transformation is particularly evident when one compares the culture of corporate accountability that Bill Gates put in place at Microsoft with the practices of epidemiological accountability permeating the Initiative. As I suggested in this paper, both share similar audit techniques and expertise. Indeed, like Bloomberg and other philanthropists, Bill Gates made very clear that he wanted to bring his business acumen and know-how – including and especially his belief in data and audit practices – into his philanthropic work. But, as he made clear himself, there is an important difference between the audit practices used at Microsoft and those at work within the Initiative: at Microsoft, audit practices are a tool for making profit; in the Initiative, these same practices are an instrument to save lives – an audit of saving lives, if you will. This shift from profit-making to saving lives is made possible by the combination of audit practices with epidemiological techniques that is so characteristic of epidemiological accountability. An excellent illustration is the computer simulation models mentioned in the previous paragraph, which associate the grantees’ activities captured through audit and the numbers of lives saved measured through epidemiological proxies like smoking prevalence. It is worth noting that the way epidemiological accountability de-couples audit practices from profit and re-couples them with life directly contradicts the claim made by anthropologist Susan Erikson  that theThe Lancet editor Richard Horton and Oxford professor David Stuckler – are prominent global health experts who have developed and published in leading journals sophisticated and powerful critiques of philanthropies like the Gates and Bloomberg foundations. Contrary to what they seem to think, it is them and their reasoned critiques – rather than an elusive world citizenry and a utopian global democratic system – that can and already have forced philanthropists to listen and change the way they work.To finish, I would like to reflect on the figure of the accountee in epidemiological accountability and, in this way, return to the body of literature critiquing philanthropies’ lack of accountability explored at the start of the paper.