The objectives of this longitudinal study were to examine the trajectory of breastfed infants’ growth in China to update growth standards for early childhood, and to compare these updated Chinese growth standards with the growth standards recommended by the World Health Organization (WHO) in 2006.This longitudinal cohort study enrolled 1,840 healthy breastfed infants living in an "optimal" environment favorable to growth and followed up until one year of age from 2007 to 2010. The study subjects were recruited from 60 communities in twelve cities in China. A participating infant’s birth weight was measured within the first hour of the infant’s life, and birth length and head circumference within 24 hours after birth. Repeated weekly and monthly anthropometric measurements were also taken. Multilevel (ML) modelling via MLwiN2.25 was fitted to estimate the growth curves of weight-for-age (WFA), length-for-age (LFA), and head circumference-for-age (HFA) for the study sample as a whole and by child sex, controlling for mode of delivery, the gravidity and parity of the mother, infant’s physical measurements at birth, infant’s daily food intaking frequency per day, infant’s medical conditions, the season when the infant’s physical measurement was taken, parents’ ages, heights, and attained education, and family structure and income per month. During the first four weeks after birth, breastfed infants showed an increase in weight, length, and head circumference of 1110g, 4.9 cm, and 3.2 cm, respectively, among boys, and 980 g, 4.4 cm, and 2.8 cm, respectively, among girls. Throughout infancy, the total growth for these three was 6930 g, 26.4 cm, and 12.5 cm, respectively, among boys, and 6480 g, 25.5 cm, and 11.7 cm, respectively, among girls. As expected, there was a significant sex difference in growth during the first year. In comparison with the WHO growth standards, breastfed children in our study were heavier in weight, longer in length, and bigger in head circumference, with the exception of a few age points during the first two to four months on the upper two percentile curves.Our data suggested the growth curves for breastfed infants in China were significantly different in comparison with those based on the WHO standards. The adoption of the WHO infant growth standards among Chinese infants, as well as the methods used in the development of such growth standards in China, need careful and coordinated consideration. Attained growth is the most widely used means to assess childhood growth patterns . The impIn recent years, growth standards developed from healthy breastfed infants have been used as a normative standard to assess the nutritional and health status of children, which has important public health implications for obesity prevention . From 19Nevertheless, questions emerged regarding the applicability of the WHO standards in various countries and regions –14. RoelSince the 1970s, many studies on children’s physical development have been conducted in China. For instance, national growth surveys of children under seven years were conducted in nine cities every decade . The curTherefore, the objectives of this longitudinal study are: a) to describe the trajectory of breastfed infant growth for infants in China to develop a more updated growth standard for early childhood; b) to delineate the determinants of the growth trajectory; c) to understand the growth characteristics of breastfed infants in China compared to the WHO standards. The methodology of this study was largely based on the MGRS protocol –21.This study was a population-based longitudinal study from Jan. 2007 to Jan. 2010. Field implementation was undertaken by the Maternal and Child Health Care Surveillance System (MCHCSS) in China, which provides postnatal care including immunization, physical examinations, and growth monitoring at no cost to infants and children from birth to seven years. The inclusion criteria for study sites were made to match the WHO criteria for living in an "optimal" environment favorable for growth . The detThe selection of study sites was based on a three-stage sampling at the provincial, municipal, and community levels, separately. At the provincial level, in addition to the inclusion ctieria described above, the selection decision was made on the basis of geographic distribution , the integrity and technical capacity of MCHCSS, and participation interest. As a result, 11 provinces were selected among those that meet the inclusion criteria.At the municipal level, one city per study province was chosen, often being the provincial capital city. Jingmen of Hubei was additionally selected, as it expressed high interest in this study and met the screening standards. Thus, 12 cities were enrolled. At the communities level, different numbers of representative communities from each city were selected to ensure sufficient eligible samples. Thus, a total of 60 communities from 12 cities were finally included in this study.The subjects included in this study were healthy breastfed infants living in an "optimal"environment favorable for growth. The inclusion criteria for families consisted of no health-related, environmental, or economic constraints on infant growth; non-consanguineous marriage for parents; both or one of the parents having urban registration; the mother-infant pair living together, routine well-baby checkups at least up to age one; and parents willing to participate in one-year follow-up and signing of the consent form. Eligible mothers were of singleton pregnancy, a maternal age between 20 and 35 years old, had a height ≥ 1.5 m, without pregnancy complications , nonsmokers, and willing to deliver at designated hospitals. Only infants born to the eligible mothers with gestational age ≥ 37 completed weeks and < 42 weeks, birth weight ≥ 2500 g and < 4000 g, free of serious or life-threatening conditions at birth , without significant morbidity during the first year after birth , and mothers’ compliance with feeding recommendations for this study were included.The feeding recommendations of this study were defined as follows: 1) exclusive or predominant breastfeeding for a minimum of four months and optimally for six months of age, as suggested by national policies; 2) introduction of complementary foods between four and six months of age; and 3) continuing partial breastfeeding up to at least 12 months of age. The definitions of exclusive or predominant breastfeeding and partial breastfeeding refer to the Breastfeeding Counseling Training Course released by the WHO, as well as lactation counseling and infant feeding guidelines provided to the mothers . At baseThis study was IRB approved by the Ethical Review Committee of the National Center for Women and Children's Health Chinese Center for Disease Control and Prevention. The approval number was FY2007-002. We also obtained written informed consent from parents on behalf of the children enrolled in our study.Infants’ anthropometric measurements included three indices: weight, length, and head circumference, and were taken at birth, weekly during the first month after delivery, and monthly from two to twelve months of age. Birth weight was measured within the first hour of the infant’s life. Birth length and head circumference were measured within 24 hours after birth. Weekly and monthly measurements were taken within 23 hours of the designated infant’s age. Replicate measures were obtained for each measurement and the average values were used. The differences in replicate measures were limited to 100 g for weight and 5 mm for length and head circumference. All anthropometrists underwent the systemic standardized anthropometric training before and during the study and followed the unified anthropometry manuals. All study sites used the same high-precision measuring equipment. Regular calibration for all tools was performed daily at the community level, quarterly by urban anthropometric experts appointed for the study, and bi-annually by the National Center for Women and Children Health (NCWCH), that was in charge of this study.Quality control procedures were critical to data quality and included the following: a) construction of a longitudinal study network from national to community level, consisting of data coordination, collection, and management; quality control; and advisory for groups; b) prior to implementation, the staff from each work group underwent thorough training to standardize the protocol for anthropometric measurements, feeding methods recommendations, and questionnaire survey administration; c) pilot testing of study protocol; d) regular visits to study sites  for the calibration of measurement tools, technology capacity assessment for filed researchers who conducted the anthropometric measurements, and home and telephone interviews; e) data quality assurance through calibration of equipment, standardization of measurements and surveys, and standardizing and auditing data entry; f) routine coordination of meetings, retraining, and staff exchanges at different levels.http://www.bristol.ac.uk/cmm/software/mlwin/download/) to estimate the growth curves of weight-for-age (WFA), length-for-age (LFA), and head circumference-for-age (HFA) for the study sample as a whole and by child sex. This study had 16 waves of assessment data on successive cohorts from multi-communities. The community comprised the level 3 unit, subjects comprised the level 2 unit, and visiting time comprised the level 1 unit. Since the infant’s physical measurements at birth were also an important factor for the infant’s growth after birth, the multi-level model was constructed separately for the anthropometric measures taken at birth and after birth. Based on the hierarchical structure of data, a basic 2-level multivariate model with level 2 comprised of the community and level 1 comprised of the subjects, was applied to construct the Birth Physical Model (BPM), while a 3-level polynomial model was applied to construct the Growth Prediction Model After Birth (GPMAB), starting with a simple 2nd degree model on age. Both the BPM and GPMAB were adjusted for the variables potentially affecting the infant’s physical growth, including the infant’s sex, mode of delivery, the gravidity and parity of mother, parents’ age, height, and attained education, and family structure and income per month, respectively. In addition to the aforementioned covariates, GPMAB also included the infant’s physical measurements at birth, infant’s daily food intaking frequency per day, infant’s medical conditions and the season when the infant’s physical measurement was taken. The final adjusted predicted model was developed based on the following criteria [To accommodate the multistage-sampling and the repeated measurement data collected in this study, we conducted multilevel (ML) growth curve modelling  via MLwicriteria : a) estiFrom July 1, 2007 to July 30, 2008, 1840 (67.4%: 905 boys and 935 girls) eligible mother-infant pairs were enrolled in the study, among which 1513 (82.2%) subjects completed the one-year follow-up, and 327 (17.7%) subjects had at least one missing assessment due to scheduling conflicts (54.7%), moving out of the study sites (33.3%), child illnesses (6.1%), mother illnesses (0.9%), and unknown or other reasons (4.9%).Mother’s attained education value<0.001, P Father’s attained education value<0.001).The demographic characteristics of 1840 subjects were presented in It is noteworthy that on 97th percentile curve of WFA, the difference between the raw data and the predicted value became larger after 10 months old, which may be due to the degree of dispersion in our data. It appears that the older age a child was, the greater the degree of dispersion was observed for WFA, compared to LFA or HFA.child sex value <0.0001, Tables child age x sex value<0.0001, We further stratified our analysis by child sex. As expected, there was a significant sex difference in growth during the first year  for Chinese infants were higher when based on the standard draw from the WHO MGRS, while underweight (with the 3rd percentile of weight-for-age as the cut-off point), and stunting (with the 3rd percentile of length-for-age as the cut-off point) were lower.Compared to the infant growth reference from the 2005 nine-city study (3), the growth levels reached by breastfed infants on the 97th percentile curve in our study were relatively lower both for weight and length. The differences between the two studies across the first year were 280–560 g and 1.1–1.4 cm for boys and 10–370 g and 0.7–1.5 cm for girls, respectively. However, conflicting results occurred on the 50th curve, as well as the lower two percentile curves. Especially on the 3rd percentile curve, the boys’ weights and lengths in our study were 120–630 g and 1.2–1.9 cm above the current national reference, which also presented in the girls group.The present study was the first longitudinal study of breastfed infants based on a large sample of Chinese children. Compared to Chinese growth standards developed 25 years ago by means of a longitudinal study design , a positThe considerable divergence in infant growth between our study and the 2005 nine-cities study, as noted above, likely resulted from the differences in study design and the characteristics of study samples, specifically from the differences in the types of feeding methods for the enrolled subjects. In the nine- cities study, 36.3% of the study sample reported partially breast-feeding during the first four months and 15.1% exclusively formula-feeding. Only 48.6% of infants were exclusively or predominantly breastfeeding , whereasAn updated infant growth standard based on data from longitudinal studies from a study sample meeting the recommended feeding practices is critical to provide an appropriate reference for the optimal growth and development of healthy Chinese children during their first year of life. In comparison with the growth curves of the WHO standards , 11, it Noteworthy, the weight and length values of Chinese infants from this study were shown to be slightly lower at birth (average differences of 220 g and 0.69 cm among boys and 190 g and 0.19 cm among girls) as compared to the reference issued in Sweden , but werIn view of these findings, the adoption of the WHO infant growth standards in China would result in an increased number of Chinese infants classified as overweight or obese but a decreased number of infants classified as underweight or stunted. With a vast territory, significant regional differences in child development exist in China. In urban areas, where obesity has become the main public health problem, the WHO standards might be helpful to avoid excess weight gain but probably will lead to inappropriate assessment concerning weight-for-age, resulting in needless parental worrying. By contrast, in rural areas, especially in Western China, malnutrition is still the most common health problem adversely affecting child growth. Quite a few malnourished infants would be classified as having normal development by the infant growth standard based on the WHO MGRS study and subsequently miss out on early interventions.Similar results were reported from Juliusson et al. on Norwegian and Belgian children that the number of Belgian and Norwegian children below -2 SD lines of the WHO standards was lower and above +2 SD higher than expected . Lately,Table 1showed, as compared to the non-compliant group, parents’ highest attained education was a bit higher among the compliant group. Our study findings may have overestimated the average infant growth of breastfeeding infants due to the missing anthropometric data of non-compliant samples [The strengths of the present study include its prospective design with repeated measures on breastfeeding infant growth. Most previous research on infant growth was limited by cross-sectional study design and without the clear definition of infant feeding methods ,35. Our  samples . HoweverIn conclusion, our study was a large, longitudinal-based study of a sample of healthy breastfed Chinese infants. This study showed that the growth curves for breastfed infants in China were significantly different in comparison with those based on the WHO MGRS study, as well as those from the current national growth standards based on the Chinese nine-cities study. The adoption of the WHO infant growth standards among Chinese infants, as well as the methods used in the development of such growth standards in China, need careful and coordinated consideration.


Enterococcus faecalis being the primary causative isolate. Infective endocarditis is a life-threatening infection of the endocardium that results in the formation of vegetations. Based on a literature review, this paper provides an overview of the virulence factors associated with E. faecalis infective endocarditis. Furthermore, it reports the effects of active or passive immunization against some of these involved factors.The enterococci are accountable for up to 20% of all cases of infective endocarditis, with E. faecalis infective endocarditis. Absence of these factors entailed attenuation of strains in both mixed- and mono-bacterial infection endocarditis models as well as in in vitro and ex vivo assays when compared to their virulence factor expressing parental strains.Nine virulence factors have in particular been associated with The virulence factors promote a broad spectrum of events that together allow for disease development and progression. The infection is initiated through bacterial binding to ligands present at the site of infection after which the colonization can be accelerated through inter-bacterial attachment and modulation of the host immune response. The formation and growth of the vegetation provide protection and promote growth. Controlled degeneration of the vegetation appears to increase the likelihood of embolization and dissemination, without exposing protected bacteria.In most cases, active and passive immunization against associated virulence factors provided partial protection.There is a need for further evaluation of the known virulence factors. Immunization against two or more virulence factors might be an effective prophylactic tool. E. faecalis being the causative isolate in >90% of the cases [Some of the first microbial colonizers of the infant gastrointestinal tract are the enterococci. They were once regarded as commensal intestinal organisms with minimal clinical importance, but since the mid to late 1970s, they have emerged as opportunistic pathogens capable of causing infective endocarditis (IE), bacteremia, intraabdominal infections, wound infections, and urinary tract infections, among others, especially in the hospital setting . Enteroche cases , 8.E. faecalis IE is a severe infection that still has an in hospital mortality of 15 to 20% and a one year mortality of approximately 25 to 30% [IE is an infection of the endocardium that results in the formation of lesions termed septic vegetations, which are clotted masses of platelets, fibrin, bacteria, and immune cells, that without effective therapy may lead to valvular destruction, congestive heart failure, and death , 9. E. f5 to 30% , 10.E. faecalis IE is initiated through bacteremia, caused by events such as surgical procedures, ascending urinary tract infections or intestinal bacterial translocation. This is followed by attachment of the bacteria either directly to the inner linings of the endothelium, to a valvular prosthesis or to a sterile vegetation, consisting of fibrin and platelets, formed due to prior valvular damage. Once initial attachment is achieved, the bacteria become encased in a matrix of platelets and fibrin that protects them from the recruited immune cells leading to the creation of a septic vegetation. The infective endocarditis vegetation is a biofilm process on the heart valves, which provides a protective barrier as well as a favorable growth environment, in which the bacteria are more resistant to antibiotics and immune mediated killing. Furthermore, the vegetation also potentially allows for an increased horizontal gene transfer as well as providing a vessel for dissemination of bacteria to other organs, when pieces of the vegetation break off [The course of reak off , 4, 5, 9E. faecalis IE and their individual contribution to the pathogenesis. Furthermore, it reports findings regarding the ability of active or passive immunization against involved virulence factors to provide protection against the disease.This mini review comprises an outline of the current knowledge regarding the virulence factors associated with 1). The search was not restricted to a certain timeframe, but the exclusion criteria seek to eliminate older redundant studies.Data was obtained through a PubMed literature search and a flowchart of the conducted literature search, including the inclusion and exclusion criteria, is shown in Fig. . So far, three different AS proteins  have been identified, each encoded by their own pheromone-responsive conjugative plasmid . Together they comprise a family of surface adhesins that are almost identical throughout most of the protein, the exception being a small variable segment with a shared identity of about 30 to 40% [AS is a large pheromone inducible surface protein of 0 to 40% .E. faecalis IE, with four included articles [Asc10 appears to be the most extensively studied AS protein in relation to articles , 12, 13 articles  investigprgB gene of pCF10 and several functional domains of the protein have been described, amongst these, an N-terminal and a central aggregation domain as well as an N- and C-terminal Arg-Gly-Asp (RGD) motif [Asc10 is encoded by the D) motif .et al. [E. faecalis IE. They found that not only did the combination of Asc10 and EBS expression increase lethality, but it also prevented pericardial inflammation [et al. [E. faecalis IE. Using a rabbit model of endocarditis, they found that a strain with mutations to both RGD motifs exhibited virulence that was essentially equivalent to that of a strain carrying the prgB null allele. Furthermore, mutations to either RGD motif alone, revealed that the N-terminal RGD motif seems to be more important for virulence than the C-terminal RGD motif. Disruption of either aggregation domain resulted in strains that were less virulent than the wild-type strain, but more virulent than the double RGD mutant [In 1998 Schlievert, et al. , used a ammation . Later,  [et al. , investiD mutant .et al., 2010 [ex vivo porcine heart valve model to evaluate the role of Asc10 in bacterial adherence and biofilm formation at early time points. They found that strains, regardless of Asc10 expression, were able to attach to and initiate biofilm development on the heart valves with similar colonization for the first two hours. However, after four hours, the Asc10- strain was found to colonize the valves at only 40% of the level of the Asc10+ strain. Furthermore, they observed that the micro colonies of the Asc10+ strain were large, containing hundreds of bacteria and abundant extracellular matrix (ECM), whereas the Asc10- strain produced smaller micro colonies, with less ECM. They also assessed the contribution of the aforementioned functional domains of Asc10 with regards to initial adherence and biofilm formation, and found that several of the mutations had less severe effects in their ex vivo valve model, compared to the in vivo endocarditis model used by Chuang, et al., 2008 [In a subsequent study, Chuang-Smith, l., 2010  used an l., 2008 et al., 1993 [+ Asa1+ group, a 15% mortality in the hemolysin– Asa1+ group and a 0% mortality in the hemolysin+ Asa1- group. Furthermore, they found that the Asa1+ strains produced vegetations with weights varying from 181-209 mg, whereas the Asa1- strains produced vegetations with weights varying from 81-92 mg [The enterococcal hemolysin is encoded on the pheromone-responsive plasmid pAD1 , the saml., 1993  used a r81-92 mg .E. faecalis strains with impaired or eliminated glycolipid synthesis, exhibit impaired biofilm formation, a process known to play a crucial role in IE. In E. faecalis, two glucosyltransferases, termed biofilm-associated glycolipid synthesis A and B , are responsible for the biosynthesis of the two main cell wall glycolipids, diglycosyl-diacylglycerol and its precursor monoglycosyl-diacylglycerol. BgsA deficiency has been shown to lead to a loss of diglycosyl-diacylglycerol in the cell membrane and to an accumulation of monoglycosyl-diacylglycerol, whilst bgsB deficiency leads to a complete loss of cell membrane glycolipids [Studies have shown that mutant colipids .et al. [E. faecalis IE. Using a rat model of endocarditis, they compared mutant strains, deficient for either bgsA or bgsB, with a parental wild-type strain. They found that both of the mutant strains were attenuated, compared to the wild-type strain, producing vegetations that weighed less and contained significantly less CFU per milliliter and per gram. The bgsB mutant exhibited the highest level of attenuation [In 2014 Haller, et al. , assesseenuation .E. faecalis during stress conditions such as glucose starvation, and exposure to cadmium chloride or bile salts [et al. [- strain, constituting 77% and 84% of the mean percentage of the total number of bacteria recovered from vegetations at 72 and 120 hours, respectively. The initial injection mixtures used for the experiments contained 36% and 43% OG1RF, respectively. The researchers also calculated the mean virulence index of the gls24- strain relative to OG1RF and found it to be 0.18 and 0.09 at 72 and 120 hours, respectively [Gls24 is a protein, synthesized in le salts . In 2005 [et al. , compareE. faecalis, and today it is known that their assembly is dependent on a three-gene EbpABC locus and an associated sortase gene, srtC. High titers of antibodies against the EbpABC proteins have been found to be present in the sera of patients suffering from E. faecalis IE, thus indicating that they are readily expressed and recognized by the host immune system during infection [Pili and fimbriae are fibrous protein organelles, anchored to the surface of the bacterium, that interact with the external environment. They are known to be used by some bacteria to accomplish adhesion to host cells , 9. In 1et al. [in vitro attachment assay revealed significant differences in initial attachment, with the wild-type parental strain (OG1RF) exhibiting a higher number of adherent cells per microscopic field, than any of the mutant strains. Lastly, the researchers used a mixed-infection rat model of endocarditis, in which they injected the rats with an equal mixture of a nonpiliated EbpA mutant and OG1RF. They found, that the mean percentage of the EbpA mutant in the total CFU of bacteria recovered from vegetations after 24 hours was 26.1% and calculated the mean virulence index of the EbpA mutant relative to OG1RF to be 0.06 [In 2006 Nallapareddy, et al. , assesse be 0.06 .E. faecalis, the other one being serine protease (SprE) [GelE is an extracellular zinc-metalloprotease that is thought to contribute to virulence through degradation of host substrates such as collagen, fibrinogen, fibrin, and complement components C3 and C3a among others. It is one of two proteases of e (SprE) .et al. [E. faecalis IE by comparing the vancomycin-resistant E. faecalis V583 with three mutant strains that lacked either GelE and/or SprE. They also assessed if any of the two proteases possessed proteolytic activity towards complement component C5a. Using a rabbit endocarditis model, they found no significant differences in bacterial burdens on the heart valves for any of the strains tested. However, an assessment of the mean CFU per gram of heart tissue, revealed a 14- and 7.2-fold reduction for the GelE- SprE+ and GelE- SprE- strains when compared to V583. Conversely, they observed a 6.5-fold increase in the mean CFU per gram of heart tissue for the GelE+ SprE- strain when compared to the GelE- SprE+ strain. Lastly, the researchers found that GelE is capable of hydrolyzing C5a [In 2010 Thurlow, et al. , sought E. faecalis IE as well as playing a vital role in the in vitro adherence of E. faecalis to immobilized collagen. The heart valves, the aortic tissue, as well as sterile vegetations are all known to be collagen-containing sites [Ace is a cell-wall anchored adhesin, shown to be antigenic during human ng sites .et al. [E. faecalis IE, using an in vitro adherence assay as well as mixed- and mono-infection rat endocarditis models. They tested four different strains; OG1RF , OG1RFΔace (Ace deletion mutant), pAT392::ace (Ace deletion mutant containing a shuttle vector with a cloned Ace gene) and pAT392 . The percentages of Ace-expressing cells in cultures of OG1RF, OG1RFΔace, pAT392 and pAT392::ace were >70%, <5%, <5% and >90%, respectively. In the adherence assay they found that both of the Ace- strains (OG1RFΔace and pAT392) were highly attenuated, compared to OG1RF, and even more so compared to pAT392::ace, in adherence to collagen type I and IV. To assess the importance of Ace in vivo, the researchers used a mixed-infection rat model of endocarditis, in which rats were injected with a 1:1 mixture of OG1RF and OG1RFΔace. After 72 hours, they recovered the bacteria from the valves and found the mean percentage of OG1RFΔace in the total CFU to be 18.5%, which corresponds to a mean virulence index of OG1RFΔace relative to OG1RF of 0.077. Lastly, they wanted to assess the role of Ace in the early attachment stage of valve colonization. To do so, they compared the results of two independent mono-infection models, in which rats were challenged with either pAT392 or pAT392::ace. They found that after four hours, valve vegetations of rats infected with pAT392::ace contained 1.4±0.6 log10 more CFU/geometric mean, than valve vegetations of rats infected with pAT392 [In 2010 Singh, et al. , assesseE. faecalis that confers adhesion to immobilized fibronectin. Fibronectin is a glycoprotein that, among other places, can be found in the ECM of the cardiac endothelium. It is exposed if the cardiac tissue gets damaged, whereupon it binds fibrin and platelets, thus contributing to the formation of a sterile vegetation [EfbA is an adhesin, localized on the outer surface of getation .et al. [E. faecalis IE. Using a microtiter plate-based assay, they demonstrated that recombinant EfbA (rEfbA) exhibits a concentration-dependent binding to immobilized human fibronectin as well as high affinity to collagen I and V. Furthermore, they showed that an EfbA deletion mutant (TX5707) showed a 44% decrease in whole cell binding to immobilized fibronectin compared to the wild-type parental strain (OG1RF). A derivative strain, in which the EfbA gene had been restored in its original chromosomal location (TX5708), showed whole cell binding to immobilized fibronectin similar to that of OG1RF. When examined for their ability to form biofilms, TX5707 was found to produce biofilms with a 15% reduced density compared to OG1RF and TX5708. To assess the contribution of EfbA to E. faecalis IE, the researchers used a mixed-infection rat model of endocarditis. First, they injected rats with a 1:1 mixture of OG1RF and TX5707 and after 48 hours, the mean percentage of TX5707 in the total CFU of bacteria recovered from the valves was found to be 25%. Next, they injected the rats with a mixture of 59% TX5707 and 41% TX5708 and after 48 hours, the mean percentage of TX5707 in the total CFU of bacteria recovered from the valves was found to be approximately 37% [In 2015 Singh, Eep is a membrane metalloprotease, that is known for its role in the proteolytic processing of sex pheromone peptides that, once matured, induce conjugation in a class of large pheromone-responsive conjugative plasmids, such as pCF10 and pAD1 .et al., 2011 [E. faecalis IE. They found that OG1RFΔeep exhibited a 4-log10 decrease in valve bacterial load compared to OG1RF. Furthermore, they observed that OG1RFΔeep produced small vegetations with a mean weight of 8.8 mg, whilst OG1RF produced large vegetations with a mean weight of 47.6 mg. The researchers also assessed a strain that had been complemented with the wild-type Eep locus in trans on a plasmid. They found that the complemented strain produced vegetations with a mean weight of 32.1 mg, despite the fact that only 2% of the cells retained the plasmid at the end of the experiment [Frank, l., 2011  used a rperiment .2.An overview of the assessed contribution of each virulence factor, to different steps of the disease process is available in Table The first step of IE, once bacteria have entered the bloodstream, is the initial attachment to and colonization of the inner linings of the heart, the valve prosthesis or the sterile vegetation.et al., 2008 [et al., 2010 [et al., 1993 [Asc10, Ace and EfbA were all found to mediate binding to substances that are either exposed once the endocardium is damaged or present in the following sterile vegetation -4. The rl., 2010 , who obsl., 2010 , it coull., 2010 . The AS l., 1993  observedet al., 2010 [in vitro adherence assay suggests, that the mechanism behind Ace mediated attachment is likely through binding to either collagen I or IV [The importance of Ace in the initial attachment process was highlighted by the results of the 4 hours endocarditis experiment conducted by Singh, l., 2010 , in whic I or IV .et al., 2015 [EfbA was found to mediate binding to fibronectin, collagen I and collagen V, and thus the observed attenuation of an EfbA mutant in an endocarditis model as observed by Singh, Two other factors, Eep and the Ebp pili, also seem to be important during the early stages of infection, but unlike Asc10, Ace and EfbA, the data obtained do not reveal any putative ligands for adherence.et al., 2011 [E. faecalis IE pathogenesis remain unknown and further studies are needed to elucidate this, but it seems clear that Eep plays a role that goes beyond the maturation of pheromone-responsive plasmids.Eep is known to be involved in the maturation of pheromone responsive plasmids that encode the AS proteins among others. However, since Frank, l., 2011  used a pl., 2011 . The speet al., 2006 [Likewise, the observations made by Nallapareddy, l., 2006  showing The IE vegetation is a biofilm process on the heart valves . A biofiE. faecalis IE. Asc10 was observed to contribute to the establishment of large microcolonies containing abundant ECM [Three identified virulence factors seem to be involved in the formation of biofilms during dant ECM , a non-pdant ECM  and an Edant ECM .et al., 2010 [The formation and accumulation of the vegetation is known to change both the growth as well as the physiological state of the bacteria, which in turn impedes the effectiveness of antibiotics and the host immune system and it is thus advantageous for the bacteria . Interesl., 2010 , who foul., 2010 .et al. [et al. [E. faecalis resistant to killing following ingestion by PMNs. In 2010 Chuang-Smith, et al. [ex vivo model compared to the in vivo results obtained by Chuang, et al., 2008 [ex vivo model, it is possible, that the observed differences are due to involvement of the RGD motifs in immune evasion.The data obtained from the included studies reveal that both Asc10 and GelE, might be involved in the process of immune modulation. The first indications of Asc10´s role in immune modulation came from results obtained in 1998 by Schlievert, et al. , who fou [et al. , who fou, et al. , observel., 2008 . Since tet al., 2010 [in vivo [GelE also seems to be involved in immune modulation, as Thurlow, l., 2010  found th[in vivo . DespiteE. faecalis IE, but it does not offer an exact explanation to its specific contribution.The existing data regarding gls24 reveal that it is important for virulence in E. faecalis are known to take part in biofilm formation [et al., 2014 [The cell-wall glycolipids of ormation , thus thl., 2014 , could bE. faecalis IE, revolves around antimicrobial therapy in the form of administration of a combination of antibiotics often alongside with surgery [E. faecalis that are resistant towards the most commonly used antibiotics are complicating the treatment and has created the need for alternative ways of dealing with the infection [The current management of  surgery . Strainsnfection , 3. One et al., 2010 [et al., 2010 [et al., 2014 [et al., 2015 [Experiments assessing the ability of active and/or passive immunization against the virulence factors AS, Ace, Ebp pili and EfbA, to prevent or reduce the severity of the infection in animal models have been conducted by Schlievert, l., 2010 , Singh, l., 2010 , PinkstoE. faecalis in vegetations by more than two log CFUs [in vitro dose-dependent inhibition on biofilm formation, resulted in 3/10 rats developing E. faecalis IE compared to 9/10 rats in the control group [10 CFU bacteria in the vegetations from 4.8 to 2.1 [Their collective results revealed that immunization with IgG antibodies against AS enhanced vegetation formation and caused extensive lung congestion in test animals. Conversely, immunization with IgG antigen binding fragments (Fabs) against AS, reduced both the mean weight of vegetations from 40 mg to 10 mg as well as the number of log CFUs . Both aclog CFUs . Passive3.An overview of the obtained immunization results and their corresponding p-values is available in Table E. faecalis IE, in animal models, through active and passive immunization against factors that are putatively involved in the initial attachment process. However, it must be taken into account that immunization against an individual virulence factor provides the host with a factor-specific immune response that might only be effective against strains expressing that particular factor. Thus, targeting a frequently expressed virulence factor could prove effective against many genetically different isolates.These results indicate that it is possible to achieve partial protection against E. faecalis IE have antibodies against Ace in their sera. The Ace gene is also ubiquitously present in E. faecalis isolates and its amino acid sequence is highly conserved [E. faecalis isolates produce AS [E. faecalis IE, it could be necessary to target multiple factors in order to achieve a protection that is more robust. Both the Ace gene and the Ebp locus are ubiquitously present in E. faecalis isolates [Studies have shown that 90% of patients with prior onserved , indicatoduce AS . Furtherisolates , 9, thusLastly, it must be taken into account, that the people who are at the greatest risk of getting IE, are those who are either immunocompromised due to chemotherapy or weakened by age, underlying diseases or surgical treatments . This paE. faecalis IE. Furthermore, this study has not addressed the activation and transfer of the genes encoding the virulence factors, nor has it addressed how E. faecalis manages to enter the bloodstream. Future reviews elucidating these areas might provide even further insight into the understanding of the disease, as well as new possible ways to prevent it. Lastly, the emergence of multi-resistant E. faecalis strains, complicating the treatment, means that it is important to search for and identify new treatment strategies. Immunization against two or more virulence factors might prove to be an effective prophylactic tool, but further studies are needed to elucidate this. For treatment of patients suffering from E. faecalis IE, the best course of action may include attacking the vegetation, in order to expose the bacteria and make them vulnerable to antibiotics and the host immune system, but bearing the results from the GelE experiments in mind, such an approach could also end up increasing the dissemination of bacteria.The review of the included articles has revealed that there is still a need for further studies evaluating the known virulence factors associated with


As predicted, in paintings the measures of hedonic tone were less strongly correlated , and when controlling for familiarity, the association with complexity was significantly positive for beauty (rs = 0.26), weakly negative for pleasantness (rs = -0.16) and not present for liking. Experiment 2 followed a similar approach and 77 female participants, all non-musicians, rated 92 musical excerpts (15 s) in three conditions of hedonic tone . Results indicated a strong relationship between complexity and arousal . When controlling for familiarity effects, the relationship between complexity and beauty followed an inverted-U curve, whereas the relationship between complexity and pleasantness was negative (rs = -0.26) and the one between complexity and liking positive (rs = 0.29). We relate our results to Berlyne’s theory and the latest findings in neuroaesthetics, proposing that future studies need to acknowledge the multifaceted nature of hedonic tone in esthetic experiences of artforms.In his seminal book on esthetics,  Esthetic experiences are phenomena that are inherently challenging to study due to a myriad of underlying factors. Therefore, it is not surprising that the field of empirical esthetics has generated competing accounts for why humans show preferences for certain objects over others or why they consider some objects as more beautiful . For exaNew Experimental Esthetics have been very influential in the field , evaluative scales , internal-state scales  as well as, to a lesser degree, stylistic scales (referring to stylistic features of an artwork). Factor analysis conducted on these variables often showed pleasure to be correlated with other evaluative scales   . AlthougThe main goal of the current study was to clarify how verbal reports of different measures of hedonic tone relate with visual and musical complexity, aiming to contribute to the elucidation of Berlyne’s theory . Recent Concepts such as complexity, arousal, pleasantness, liking, and beauty are also essential features of more recent models of esthetic experiences , thus thSeveral neuroimaging reports on the esthetic experience of art have also confirmed activation of different brain areas constituting the reward circuit , which mThere is a recent interest in comparing esthetic experiences across object classes and sensory domains  the nature of hedonic tone and its relationship with visual complexity as defined by the number and diversity of elements, and (ii) the nature of hedonic tone and its relationship with musical complexity. We studied three evaluative scales frequently used as measures of hedonic tone, namely pleasantness, beauty and liking. In Experiment 1, we collected subjective reports on affective environmental scenes (IAPS pictures) and representational paintings , in whicWe decided to employ a between-subjects design in which participants rated pictures or musical excerpts for familiarity, complexity, and arousal as well as for only one of the three measures of hedonic tone. This ensured that participants could not guess the aim of the experiment, and further, this design was also closer to what other researchers mostly employed in their designs, i.e., to use only one measure of hedonic tone. Last, we decided to restrict our sample to females because gender effects regarding these picture sets have been reported earlier . We alsoFor Experiment 1 we hypothesized that, in line with For Experiment 2, our hypotheses were less concrete because much less is known about the underlying dimensions of musical complexity. However, as objective measures of musical complexity have indicated in Two-hundred thirty female psychology students at the University of Vienna participated in the experiment in exchange for course credit. All participants were unfamiliar with the three picture sets and did not participate in previous studies by Table 1).The participants of the nine groups were screened for outliers with respect to their mood prior to the experiment (participants with very low mood scores were removed), their art interest as well as the trait emotional intelligence, empathy and stress reactivity scales . Twenty-four participants were removed for these reasons by investigating the respective boxplots (an outlier was defined as a data point more than 1.5 interquartile ranges below the first quartile or above the third quartile). The remaining 206 participants in the nine groups did not significantly differ regarding their age, mood prior to the experiment and their scores on the trait EI, empathy quotient and SRS. We also ensured that art interest was similar, and generally low, in the six groups rating cartoons and paintings . The first step involved the following processing stages: Filter-Artistic-Poster Edges . Next, the Cutout filter was applied from the Photoshop filter gallery and the settings for number of levels, edge simplicity and edge fidelity were individually adjusted to obtain a picture whose semantic content could be easily deduced. In other words, the level of abstraction was not very high. Last, the brightness and contrast settings were adjusted .The 96 affective environmental scenes , with stronger relationships for cartoons than IAPS pictures. The relationships between familiarity and measures of hedonic tone were positive, strong, and constant across the three measures of hedonic tone (rs > 0.6), and comparable in IAPS pictures and cartoons. A similar pattern of results was observed for paintings; however, there was more variation across the three measures of hedonic tone in terms of effect size.Table 4 shows how complexity, arousal and the three measures of hedonic tone were related. In line with Berlyne’s prediction, we generally observed small to moderate associations between complexity and arousal for both IAPS pictures and cartoons. In paintings, this association was somewhat stronger (rs ∼0.5), nearly reaching the benchmark of a large effect  show these relationships including a regression line based on results of curve-fitting analyses conducted in SigmaPlot 13.0. By not controlling for familiarity effects, the multifaceted nature of hedonic tone became visually apparent in cartoons and paintings but not in IAPS pictures. The relationship between complexity and pleasantness was negative in cartoons, whereas there were no relationships for the beauty and liking conditions. In paintings, the relationships differed to a much larger extent and the graphs seemed to indicate a positive association between complexity and beauty, a negative association between complexity and pleasantness, and no relationship in the liking condition.Of particular interest was the comparison of the relationships between complexity and the respective measures of hedonic tone across the three picture sets. Table 4). The relationship between complexity and beauty was positive and significant only for paintings (rs = 0.26). Next, we predicted no relationship between complexity and pleasantness, which held true for IAPS pictures. For cartoons and paintings we observed indications of a weak, non-significant negative relationship. The relationships between complexity and liking ratings were absent in IAPS pictures and paintings, but weakly non-significantly positive in cartoons. We also compared the correlation coefficients expressing the relationship between complexity and hedonic tone by means of a Fisher r-to-z transformation in paintings. The coefficients of the beauty-complexity and pleasantness-complexity relationships significantly differed from each other (p = 0.004), whereas the beauty-complexity and liking-complexity relationships (p = 0.097) as well as the pleasantness-complexity and liking-complexity relationships (p = 0.223) were not significantly different form each other. Taken together, these results show that the relationship between complexity and hedonic tone depends on the specification of the latter concept , which became mostly apparent in the study of paintings.Turning to the results of partial correlations controlling for familiarity effects, we predicted a positive relationship for beauty based on results by Table 4): we observed negative relationships for pleasantness (rs = -0.56) and liking (rs = -0.23) but not for beauty, providing more evidence for the multifaceted nature of hedonic tone in the perception of paintings.Due to the pre-selection of the stimuli, the relationships between the respective measures of hedonic tone and arousal were quadratic in IAPS pictures see , 2016 anN = 96), the correlation strengths between measures of hedonic tone were very high: rbeauty-pleasantness = 0.91, rbeauty-liking = 0.95, and rliking-pleasantness = 0.92. In cartoons (N = 96), these relationships were similar, but the beauty-pleasantness relationship was slightly lower: rbeauty-pleasantness = 0.85, rbeauty-liking = 0.90, and rliking-pleasantness = 0.92. However, in paintings (N = 94), the three measures correlated to a lesser extent with each other: rbeauty-pleasantness = 0.73, rbeauty-liking = 0.91, and rliking-pleasantness = 0.77. Overall the pattern of results suggests that the relationship between beauty and pleasantness as well as the one between liking and pleasantness were weaker for paintings than for the other two stimulus sets.If hedonic tone is not a uniform concept when considering the perception of paintings, intercorrelations between the different measures of hedonic tone should be lower for ratings of paintings than for IAPS pictures and cartoons. In IAPS pictures : IAPS pictures: Mbeauty = 3.87, SDbeauty = 0.77, Mpleasantness = 3.65, SDpleasantness = 0.77, Mliking = 3.55, SDliking = 0.74; cartoons: Mbeauty = 3.76, SDbeauty = 0.61, Mpleasantness = 4.02, SDpleasantness = 0.63, Mliking = 3.66, SDliking = 0.61; paintings: Mbeauty = 4.05, SDbeauty = 0.63, Mpleasantness = 4.31, SDpleasantness = 0.68, Mliking = 3.90, SDliking = 0.78.The mean complexity ratings of all pictures per type of category and condition were as follows: IAPS pictures: Table 5) showed that participants’ mean familiarity ratings were generally low . Furthermore, participants’ liking ratings were neutral on average, which was to be expected given that pictures of different emotional contents were shown. In all conditions, participants reported to have felt feelings , which were of an average intensity. Moreover, participants also reported moderate emotional involvement with the picture content in all conditions. The difficulty to judge complexity was neutral across conditions. Ratings of the artistic status of cartoons showed that participants rated them above average . A comparison across the three picture types by means of a series of Kruskal–Wallis tests revealed that participants neither preferred one picture type to the other nor did they report different levels of emotional involvement. All picture types induced emotions of similar intensity. However, the data suggest that reporting complexity judgments was somewhat easier for paintings than for the other two picture types, H(2) = 5.70, p = 0.058. General familiarity ratings were higher for cartoons than for the other two picture sets, H(2) = 6.35, p = 0.042, which was due to higher familiarity ratings in the condition in which participants reported liking of cartoons.The analysis of the post-experimental questionnaire  between complexity and pleasantness observed for paintings. However, rs = 0.22), which was not supported by the current data, and also not by Our findings are in line with previous studies employing more than one measure of hedonic tone, suggesting that people differentiate more between concepts such as beauty, pleasantness and liking during the perception of paintings  than durThe current results regarding the comparison across different picture sets are relevant to neuroimaging studies reporting brain areas being specifically active during the perception of art. Several neuroimaging studies have recently followed a comparative approach and investigated brain regions active during the perception of art and non-art images matched for semantic contents . Di Dio These previous studies comparing the neural correlates of different visual stimulus categories have largely ignored the affective contents of the stimuli and mostly focused on the matching of the semantic contents between categories. If one decided to follow an affective approach to the study of visual art, it would be crucial to incorporate pleasantness and arousal in the research design and stimulus selection, given that distinct neural networks related to the processing of pleasant and unpleasant environmental scenes (IAPS pictures) have been described . The proTable 6).Participants were 92 female psychology students of the University of Vienna, all non-musicians . Participants were randomly assigned to one of three groups differing in the type of hedonic tone to be rated . Participants were screened for outliers with respect to their mood prior to the experiment (participants with very low mood scores were removed), their musical sophistication as well as the trait emotional intelligence, emotional self-efficacy and SRSs. Fifteen participants were removed for these reasons by investigating the respective boxplots. The remaining 77 participants in the three groups did not significantly differ regarding their age, mood prior to the experiment and their scores on the trait EI, emotional self-efficacy and SRS. We also ensured that musical sophistication was similar and generally low . All participants read and signed an informed consent form.Participants filled in the short form A of the multidimensional mood questionnaire prior to the experiment. The experimental setting was identical to the one described in Experiment 1 except that musical stimuli were played through an external soundcard  and participants were wearing Sennheißer HD 380 pro headphones. The volume was fixed to approximately 72 dB SPL (A-weighted) as measured during the presentation of the second practice trial see . SittingThe musical excerpts were blocked according to emotional contents . The order of the four blocks as well as the order of the 23 excerpts within each block were randomized. The participants were told that the blocks would vary in their emotional content, and they were allowed to take breaks between blocks. After the experiment, participants filled in the short form B of the mood questionnaire, and then the other questionnaires in one of two different orders . The experimental session lasted around 90 min, after which participants were debriefed, thanked and dismissed.Table 7 shows that the ICCs were generally high , which justifies averaging across participants and considering excerpts as unit of analysis. Next, outliers were determined by examining boxplots for each type of rating for each condition. There were no outliers regarding familiarity, complexity and arousal ratings. One excerpt was removed due to very low pleasantness ratings, four excerpts due to very low beauty ratings, and one due to very low liking ratings. Since most distributions were not normal, non-parametric analyses were employed to investigate the interrelationships between the four types of ratings in each condition.The analysis of the data follows the one presented in Experiment 1. The subjective ratings were averaged across participants for each of the musical excerpt in each condition. Then we assessed the inter-rater reliability by calculating the average intra-class correlation coefficient (ICC) using a two-factor random effects model and type consistency . Table 7Table 8 depicts the interrelationships between the four types of ratings, separately given for each of the three conditions. All relationships were visually inspected to ensure that they were monotonous before Spearman’s rank correlations were computed. Familiarity was only mildly negatively associated with complexity and arousal, whereas the relationships between familiarity and measures of hedonic tone were positive, ranging from medium (rs ∼0.53) to strong effect sizes (rs ∼0.75). The relationship between complexity and arousal was strong and of equal strength in all three conditions of hedonic tone (rs ∼0.85). The relationship between complexity and beauty was not linear . The relationship followed an inverted-U curve and curve-fitting analysis confirmed its quadratic nature, F = 5.24, p = 0.007, 2R = 0.11, y = 0.9 + 1.58x–0.18x2. Complexity and pleasantness were significantly negatively associated (rs ∼-0.25), and the data revealed no relationship between complexity and liking.Table 9 shows how complexity, arousal and the three measures of hedonic tone were associated. The strong and positive association between complexity and arousal is in line with Berlyne’s theory. Three different relationships between complexity and the respective measures of hedonic tone emerged: complexity and pleasantness were negatively associated (rs ∼-0.26), complexity and liking positively (rs ∼0.29), and finally, complexity and beauty ratings followed an inverted-U curve as predicted by Berlyne. The two linear relationships significantly differed from each other, z = -3.72, p = 0.0002. Next, a hierarchical linear regression analysis was performed to control for effects of familiarity on beauty ratings and to demonstrate the quadratic nature of the relationship. Familiarity was entered in the first block, followed by adding the linear term for complexity ratings, and finally, the quadratic term for complexity was entered in the third step. The results of the final model were significant, F = 17.31, p < 0.001, adjusted 2R = 0.36, and revealed the following significant predictors: familiarity , complexity  and complexity squared .Controlling for familiarity effects, Table 9 also shows that the relationships between arousal and measures of hedonic tone differed from each other. The relationship between arousal and beauty was of a non-monotonic quadratic nature, though curve-fitting analysis did not reveal a significant trend. The relationship between pleasantness and arousal was negative (rs ∼-0.44), and the one between arousal and liking positive (rs ∼0.24). In general, the correlations between arousal and measures of hedonic tone reflect the nature of the complexity-hedonic tone relationship.N = 88), with the lowest correlation between liking and pleasantness ratings: rbeauty-pleasantness = 0.70, rbeauty-liking = 0.76, and rliking-pleasantness = 0.59. Intercorrelations between the three complexity ratings of the three conditions were generally very high: rbeauty-pleasantness = 0.96, rbeauty-liking = 0.96, and rliking-pleasantness = 0.96. For arousal ratings, these intercorrelations across the three conditions were of similar strength: rbeauty-pleasantness = 0.94, rbeauty-liking = 0.95, and rliking-pleasantness = 0.95. The mean complexity ratings of the three conditions were as follows: Mbeauty = 4.35, SDbeauty = 0.88, Mpleasantness = 4.27, SDpleasantness = 0.91, Mliking = 4.49, SDliking = 0.94.Next, intercorrelations between measures of hedonic tone were computed (Table 10). The three groups did not differ regarding their general liking of the musical excerpts, their familiarity with them, the frequency and strength of reported feelings as well as the difficulty of judging musical complexity. Significant group differences were observed for the role of music in participants’ life, H(2) = 8.64, p = 0.013, as well as for the frequency of listening to classical music, H(2) = 7.73, p = 0.021. Participants in the beauty group reported a higher role of music in their lives and a lower frequency of listening to music in comparison to the other two groups.Participants also answered several questions referring to the experiment  in a highly controlled participant sample and between-subject design. The same stimulus set was rated for familiarity, complexity, arousal and one of the three measures of hedonic tone. We observed that the nature of the relationship differed across groups when controlling for familiarity effects. Not supporting our hypotheses, but in line with Berlyne’s theory , our datTable 9), lending more support to Berlyne’s theory regarding the crucial role of arousal in esthetic experiences. This finding was also reported in The present results further show that the relationship between arousal and measures of hedonic tone is comparable to the respective relationships between complexity and hedonic tone , possibly due to the fact that cartoons were not considered as having a high esthetic quality or artistic status by participants.With respect to It is difficult to speculate why hedonic measures are not similarly associated with complexity, although they are themselves highly correlated in all experimental conditions and across stimulus sets. Esthetic experiences are currently being understood as interplay between emotion and cognition as well as interplay between bottom–up and top–down processes. For example, it is possible that beauty, pleasantness and liking differ in their affective connotations, with pleasantness presumably most closely linked to emotion. It can be surmised that the relationship between complexity and cognitive processing is partly determined by the affective content of the hedonic tone measures. Another related explanation refers to the different types of associations we observed between arousal and measures of hedonic tone in both paintings and music. If Berlyne was right to assume that arousal has a key function in esthetic experiences, then the nature of the arousal-hedonic tone relationship may play a role in the relationship between complexity and hedonic tone. Furthermore, it remains to be seen whether measures of hedonic tone contribute to one type of subjective experience in the perception of visual arts and music, which could be labeled as hedonic tone, or whether these concepts contribute to distinct experiences that are somewhat correlated and part of an esthetic experience, which may comprise other aspects than hedonic tone. This approach would also lead to a better understanding of whether and how visually and musically induced esthetic experiences differ.To the best of our knowledge, no neuroimaging study has compared the activation patterns related to different hedonic concepts and complexity for the same picture set, let alone examined functional connectivity patterns. Our current results thus allow for the formulation of concrete hypotheses for future neuroimaging studies. For instance, since we present evidence that hedonic tone has a multifaceted nature in the experience of visual art and music, the respective underlying neural substrates may slightly differ and perhaps be dissimilarly connected to other brain regions involved in esthetic experiences, particularly those involved in structural feature processing. To be specific, paradigms used by The role of arousal during esthetic processing has largely been ignored in neuroaesthetics, not only in the musical domain, as pointed out by The present study also sheds more light on the role of familiarity in esthetic experiences and clearly indicates that it is necessary to account for familiarity effects even though participants may generally be unfamiliar with the stimuli, and as it was the case here, were non-experts in visual arts and non-musicians. In both the visual and musical domains, we observed on average moderate positive correlations between familiarity and measures of hedonic tone, and small negative correlations between familiarity and arousal as well as familiarity and complexity. Consequently, it is crucial to account for this effect by statistical techniques if one is interested in the nature of the complexity-hedonic tone relationship across stimulus categories and different participant groups.We are aware of several limitations of the current study impeding broader generalizations. In both experiments, as well as in our previous work , 2016, wTo conclude, this systematic study demonstrates that discrepancies regarding Berlyne’s psychobiological model might partly be rooted in the largely ignored multifaceted nature of hedonic tone. Future research, especially in the growing field of neuroaesthetics, may thus explore these nuanced aspects in greater depth.MM designed the experiment and created the stimulus library. AL, MW, and MM collected the data. MM analyzed and interpreted the data. MM drafted the work and revised it after feedback from AL, MW, and HL. All authors provided final approval and agree to be accountable for all aspects of the work.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.


Metabolic syndrome (MS) is considered to be a cluster of interrelated risk factors for metabolism, which may increase arterial stiffness and cardiovascular morbidity. The cardio-ankle vascular index (CAVI) is a reliable indicator of arterial stiffness and early arteriosclerosis. The main objective of this study is to evaluate the gender-specific relationship between MS and CAVI in the general Chinese population.A total of 1,301 subjects aged 20 to 60 years participated in this study. CAVI was measured noninvasively using a Vasera VS-1000 device. Blood samples and waist circumference were examined to identify metabolic syndrome according to the criteria set forth in the 2009 Joint Scientific Statement.The prevalence of MS in the study subjects was 17.4% . CAVI values were significantly higher in MS subjects than in non-MS subjects and increased linearly as the number of MS components increased in females, but not in males. Using multiple regression analysis, we found that BMI was correlated with CAVI in the overall population and in both genders, and that high-density lipoprotein cholesterol (HDL-C) was associated with CAVI in males, while the number of MS components was related to CAVI in females. CAVI values increased linearly with age in both genders (P-trend < 0.001 for both), and this correlation was stronger in males than in females.There are gender-specific differences in the association of MS and CAVI. First, the effects of the number of MS components on CAVI are stronger in females than in males. Second, the effect of each MS component on arterial stiffness may differ in relation to gender. In addition, aging affects arterial stiffness more severely in males, and the increase in arterial stiffness tends to occur at a younger age in males than in females. Larger samples and longitudinal studies are needed to further confirm our results in the future. Metabolic syndrome (MS) is considered to be a cluster of risk factors that are interrelated with metabolism, including obesity, hypertension, dyslipidemia, and dysglycemia, which could lead to diabetes mellitus and cardiovascular disease , 2. CurrIt has been reported that there are some changes in arterial elasticity in MS patients before the occurrence by target organ damage of cardiovascular disease , while iAs a noninvasive and effective tool, pulse wave velocity (PWV) has been used widely in the examination of early atherosclerosis. However, it is strongly dependent on blood pressure, which leads to poor reproducibility . In contAs mentioned above, dyslipidemia is one of the components of MS, which is also a significant risk factor for atherosclerosis and coronary heart disease , 11. RecMetabolic syndrome is related to an increase in arterial stiffness, which could be detected by CAVI. Many studies have discussed the relationship between MS and arterial stiffness in other countries –20. HoweFrom June 2016 to October 2016, 1301 subjects  who underwent health check-ups at the First Affiliated Hospital of Jilin University (China) were recruited.This study was reviewed and approved by the Ethics Committee of the Jilin University School of Public Health (No. 2015-12-08), and written informed consent was obtained from all subjects before they participated in the study.Subjects were required to avoid drinking alcohol and consuming high-fat or high-sugar foods the day before the physical assessment. Fasting blood samples were collected between 7 and 8 AM. The samples were centrifuged and analyzed for total cholesterol (TC), triglycerides (TG), HDL-C and fasting plasma glucose (FPG). AC and AIP were calculated by the following formulas , 22:AC= HDL − C .)AIP=Lo2). Blood pressure was measured using an automatic blood pressure monitor, with each subject sitting for at least 5 minutes.Waist circumference (WC) was measured at the midsection between the lowest rib and the iliac crest. Height and weight were measured to obtain the body mass index (BMI): BMI = weight (kg) / height .We used a Vasera VS-1000 system  to noninvasively measure CAVI. CAVI values were obtained by substituting the stiffness parameter β into the following equation , 9:CVAIThe subjects were required to rest in the supine position, and the cuffs were wrapped around both their upper arms and ankles. Electrocardiographic electrodes were fixed at the wrists, and microphones were placed on the sternal angle to record an electrocardiogram and a phonocardiogram. After approximately 5 minutes, we obtained the values of CAVI and other waveforms from the Vasera VS-1000 system’s embedded printer.According to the Joint Scientific Statement of the International Diabetes Federation; the National Heart, Lung, and Blood Institute; the American Heart Association and three other international institutions in 2009 , MS shouabdominal obesity: the WC threshold for abdominal obesity in the Chinese population is 85 cm and above in males, or 80 cm and above in females;a high TG level (≥ 1.7 mmol/L) or specific treatment for this dyslipidemia;a low HDL-C level , or specific treatment for this dyslipidemia;raised blood pressure (SBP ≥ 130 mmHg and/or DBP ≥ 85 mmHg) or drug treatment for hypertension;elevated FPG level (≥ 5.6 mmol/L) or drug treatment for elevated glucose.χ2-test. One-way analysis of variance (ANOVA) was performed to detect significant difference among the groups satisfying different numbers of MS components. Post hoc analysis was conducted by Fisher’s protected least significant difference (PLSD). The relationships between MS components and CAVI were assessed by Pearson’s correlation analysis. In the multivariate analysis, multiple linear regression models were conducted (ENTER method) using CAVI values as the dependent variable and MS and its components as independent variables, using age, smoking and drinking status, high-stress life, BMI, non-HDL, coronary heart disease, and stroke as adjusting variables. P < 0.05 was considered statistically significant.SPSS Statistics for Windows, version 20.0 , was used to perform the statistical analyses. The data were presented as the mean ± standard deviation (SD) or median (interquartile range). Differences in quantitative variables between two groups were evaluated using Student’s unpaired t-test, while differences in percentage of subjects were determined by P = 0.074). The overall percentage of MS in the subjects was 17.4% . CAVI value and the other variables were significantly higher in males than in females, with the exception of HDL-C and the percentage of low HDL-C.The characteristics of the 1301 participants are shown in P-trend < 0.001 for both). Further, CAVI values in males were more influenced by age than those in females .Although it is not a component of MS, age has long been considered one of the uncontrollable factors affecting arterial stiffness. Our results showed that the mean of CAVI values among different age groups was statistically significant in both sexes. The changes of CAVI with age for the two sexes are presented in P-trend < 0.05 for both).Figs P-trend < 0.001), while this trend did not exist in males (P-trend = 0.427).Pearson’s correlation analysis showed that BMI and all MS components were associated with CAVI, except for WC and HDL-C. Among males, BMI, DBP, FPG, and WC were associated with CAVI, while SBP, DBP, and FPG were associated with CAVI in females . In the Old age, obesity, hypertension, dyslipidemia and dysglycemia are risk factors of coronary artery disease . In addiCAVI, the measurement of which is noninvasive, quick and convenient and has the advantage of not being affected by blood pressure, is a reliable predictor of arterial stiffness and early arteriosclerosis. Therefore, we evaluated the association of MS components with CAVI as the main marker of arterial stiffness in this study.Age has long been thought to be a significant risk factor for arterial stiffness, directly influencing the values of CAVI , 26. In In this cross-sectional study, the prevalence of MS and all its components, except for low HDL-C, were significantly higher in males than in females, and the values of BP, WC, FPG and TG were also higher in males than in females, which are findings that were in accord with the population-based study from Chinese Center For Disease Control and Prevention (CDC) of the prevalence of metabolic syndrome in Chinese adults . In the P = 0.074). However, the average age of male subjects with MS was younger than that of females with MS . This finding suggests that arterial stiffness develops gradually with age, and perhaps having MS at a relatively young age does not significantly influence the progression of atherosclerosis. Therefore, it is necessary to focus attention on early detection of and intervention for MS to reduce the impairment it causes to artery walls.Consistent with the results of previous studies , 29, theIn this study, we have investigated the association of MS and its components with CAVI. The results showed that FPG and HDL-C were related to CAVI overall. HDL-C was associated with CAVI in men, while the number of MS components was associated with CAVI in women. It has been confirmed that each MS component has an obvious sex-dependent effect on PWV , while tNam et al. reported that CAVI increases with increasing FPG . Other sHDL-C has traditionally been considered as a protective factor for arteriosclerosis and cardiovascular diseases. Conclusions of previous studies that CAVI significantly increased in low HDL-C groups support this view , 29, 38.In our study, SBP and DBP were associated with CAVI in univariate analysis, but not in multivariate analysis, which may be affected by the adjusting variables. Raised BP directly affects the arterial walls, resulting in the increase of vascular tension. Continuous elevation of BP will lead to structural and functional changes in the arterial walls, resulting in further decrease of large arterial elasticity, which reflects arteriosclerosis.WC reflects the amount of intra-abdominal adipose tissue, and it has been demonstrated to predict the risk of cardiovascular events and has been applied to clinical examinations . As the In addition, we have found a negative correlation between BMI and CAVI in the overall population and in both genders. Nagayama et al. found that an inverse correlation between BMI and CAVI in healthy middle-aged subjects, and they hypothesized that systemic accumulation of adipose tissue, per se, leads to reduced arterial stiffness . In anotMS components often exist simultaneously, interacting with each other, and they may increase the risk of arterial stiffness . ResearcIn this study, CAVI increased linearly as the number of MS components increased in females, but not in males. This result was not consistent with that reported by Scuteri et al. , who fouIn addition, among the 728 female subjects in this study, only 51 subjects (7.0%) had MS, which may have led to a false negative association because of sample size limitations. This incidence is lower than that of the general population in China . The difStrengths of this study include its gender-specific exploration of the relationship between MS and CAVI, as well as its application of two lipid ratios indicating arterial stiffness to some extent. However, there remain limitations in this study as well. First, our results do not establish causal relationships between MS and CAVI because of the cross-sectional study design. Second, since all subjects were recruited from Jilin Province in China, caution should be used when extrapolating these conclusions to other regions.In conclusion, our findings suggest that MS is significantly associated with CAVI, and there are gender-specific differences in this association. The effects of the number of MS components on CAVI are stronger in females than in males, and the association of each MS component with CAVI may differ in relation to gender. Furthermore, aging affects arterial stiffness more severely in males, and the increase in arterial stiffness tends to occur at a younger age in males. We should account for gender differences into consideration when interpreting study results. Larger samples and longitudinal studies are needed to further confirm our results in the future.S1 Database(SAV)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.


Streptococcus pneumoniae and its severe invasive infectious diseases has gained considerable interest. However, the target proteins to be included in a vaccine formulation have to accomplish several genetic and immunological characteristics, , in order to ensure its suitability and effectiveness. This study aimed to get comprehensive insights into the genomic organization, population distribution and genetic conservation of all pneumococcal surface-exposed proteins, genetic regulators and other virulence factors, whose important function and role in pathogenesis has been demonstrated or hypothesized.In recent years, the idea of a highly immunogenic protein-based vaccine to combat S. pneumoniae. A total of 25 S. pneumoniae strains, representing different pneumococcal phylogenetic lineages and serotypes, were considered. A set of 92 different genes and proteins were identified, classified and studied to construct a pan-genomic variability map (variome) for S. pneumoniae. Both, pneumococcal virulence factors and regulatory genes, were well-distributed in the pneumococcal genome and exhibited a conserved feature of genome organization, where replication and transcription are co-oriented. The analysis of the population distribution for each gene and protein showed that 49 of them are part of the core genome in pneumococci, while 43 belong to the accessory-genome. Estimating the genetic variability revealed that pneumolysin, enolase and Usp45 (SP_2216 in S. p. TIGR4) are the pneumococcal virulence factors with the highest conservation, while TCS08, TCS05, and TCS02 represent the most conserved pneumococcal genetic regulators.After retrieving the complete set of DNA and protein sequences reported in the databases GenBank, KEGG, VFDB, P2CS and Uniprot for pneumococcal strains whose genomes have been fully sequenced and annotated, a comprehensive bioinformatic analysis and systematic comparison has been performed for each virulence factor, stand-alone regulator and two-component regulatory system (TCS) encoded in the pan-genome of The results identified well-distributed and highly conserved pneumococcal virulence factors as well as regulators, representing promising candidates for a new generation of serotype-independent protein-based vaccine(s) to combat pneumococcal infections. Streptococcus pneumoniae, also known as the pneumococcus, is a Gram-positive, α-hemolytic and facultative aerobic bacterium. This microorganism is normally found as a harmless commensal in the upper respiratory tract of humans. Pneumococi have a great epidemiological importance due to their high impact on public health, causing more than one and a half million of deaths per year around the world [S. pneumoniae is the main etiologic agent of community-acquired pneumonia. However, this is not its only clinical manifestation, because other kind of diseases such as otitis media, sinusitis, septicemia and meningitis are also caused by this pathogen and associated with high mortality rates [he world . S. pneuty rates .Streptococcus pneumoniae , its identification process in the laboratory is relatively simple. Nevertheless, the great molecular, biochemical and immunological diversity of its capsule and other antigens such as choline-binding proteins make them one of the hardest bacterial pathogens to face because of its variability [Quellung Reaction”, developed over 100 years ago by Neufeld, allows the specifical and reliable identification of each one of the >94 serotypes that have been discovered up to date. The capsular polysaccharide is the sine qua non virulence factor, however the pathogenic potential of serotypes may vary and similarly, the frequencies or prevalence varies from one geographic region to the other [S. pneumoniae. In fact, the surface of the pneumococcus is decorated by various proteins, which have been already associated with its high pathogenic potential. In addition, their interaction level with the host cellular receptors has been proved, exhibiting crucial pathogenic functions such as adhesion, colonization, breaching tissue barriers and immune evasion [Given the particular biochemical and molecular features of iability , 4. The he other . Despite evasion .Streptococcus pneumoniae, 13 TCS and one orphan RR have been identified [An important group of regulatory proteins of great interest are the histidine kinases (HK), located in the bacterial surface and functioning as the sensors of two-component regulatory systems (TCS). The sensing of environmental signals via TCS, regulates the genetic expression of cellular processes that are of great importance such as natural competence, antibiotic resistance, adaptation to different environmental situations, surface proteins expression, and others , 8. In gentified .S. pneumoniae, (2) possessing a wide distribution among the circulating strains and clinical isolates, (3) exhibiting a major conservation at its genetic and protein sequence, (4) being inmunogenic, (5) demonstrating protectivity in experimental assays, and (6) having favorable physico-chemical properties for expression and purification of its recombinant products.The relevance of the cellular, physiological and pathogenic functions that these pneumococcal proteins fulfill, have aroused a great scientific and biotechnological interest, given their potential pharmaceutical applications as vaccine candidates . NowadayStreptococcus pneumoniae is a pathogen exhibiting a fratricide behavior and an enormous capacity for natural competence, acquiring foreign genetic material and integrating it into its genome [s genome . These ps genome , 13, gres genome , 15. This genome . In conss genome .Escherichia coli and Salmonella enterica, is providing such information for microorganisms. Remarkably, there are no open-source data of this nature for any Gram-positive bacterial genome. Hence, this study focused on the construction of the first S. pneumoniae Variome model, starting with the identification of all allellic and protein variants, a mutation and distribution analysis (presence and absence) of the virulence factors and regulators, among a set of pneumococcal strains that possess a fully sequenced and annotated genome.Due to these molecular events and their importance, there is a need to fully and globally understand the genetic heterogeneity and variability among the different pneumococcal strains/serotypes (variome), and to get a deeper and detailed molecular undestanding of the different physiological and pathogenic mechanisms that this microorganim uses to cause severe and life-threatening diseases. Definitely, obtaining this knowledge will allow to identify potential pharmaceutical targets for new antimicriobial therapies. By the recognizition of their conservation and distribution degree among pneumococcal strains, this will confirm protein candidates for vaccines. However, despite the availability of a high number of completely sequenced genomes and the importance to analyse the genetic differences among pneumococci, only a few studies have focused on studying its variability from a global perspective, similarly as the Human variome databases do . To dateStreptococcus pneumoniae strains for the analysis in this study was done using the microbial database of the “National Center for Biotechnology Information” NCBI (http://www.ncbi.nlm.nih.gov/genome) [S. pneumoniae genomes was carried out. In brief, DNA alignments, employing the tool “Microbial Nucleotide BLAST” [http://blast.ncbi.nlm.nih.gov/Blast.cgi, were performed for all the currently reported draft or complete sequenced genomes. The comparative data was then employed to construct a DNA-based Phylogenetic Tree (dendrogram), by using the Genome Tree Report Tool of the NCBI (ncbi.nlm.nih.gov/genome/tree/176). Afterwards, the file containing the dendrogram, constructed for the 8290 strains, was downloaded from the NCBI database. Finally, the dendrogram file was viewed, analyzed and adapted in order to generate circular, slanted and/or rectangular cladograms, by using the online NCBI Tool “Tree Viewer 1.17.0”, which is available online at the website: ncbi.nlm.nih.gov/projects/treeview  . Likewise BLAST” , that caVirulence Factors DataBase - VFDB” [http://www.mgc.ac.cn/VFs. Aditionally, the virulence factors and proteins involved in interactions with the host were confirmed and completed by a systematic review of the literature [http://www.uniprot.org/, with the aim of obtaining the locus tag for S. pneumoniae TIGR4 genome/strain. In addition, the genes encoding the HK or RR of the pneumococcal TCS were identified by using the database Prokaryotic Two-Component Systems - P2CS [http://www.p2cs.org/index.php. Likewise, the corresponding locus tag for S. pneumoniae TIGR4 genome / strain, of each one of the histidine kinases genes (hk) and response regulator genes (rr), were also recovered from the same database.The search and selection of genes and proteins widely known as virulence factors or gene encoding factors possessing a proven interaction with the human host was done by an exhaustive bioinformatic screening in the database “ - VFDB” , availabterature , 25. Theterature , availabs - P2CS , availabS. pneumoniae TIGR4 and the construction of the genomic maps, in linear or circular representation, was done by using the software SnapGene® (GSL Biotech), available at http://www.snapgene.com. In brief, the studied genomes of S. pneumoniae were imported through its corresponding access code in GenBank (ie: NC_003028.3 for TIGR4). Then, the chromosomal location of each virulence factor gene, and the factors involved in the interaction with the host and the genes encodying for proteins of simple or two-component regulatory systems were identified. Finally, the lineal maps for the scale genomic localization for the virulence factors and the circular maps for the genomic periphery of the genes that form the two-component regulatory systems were constructed.The chromosomal location of all the genes in the genome of S. pneumoniae TIGR4 and/or R6 in the database Kyoto Encyclopedia of Genes and Genomes – KEGG [http://www.kegg.jp/kegg/. Once every gene of interest was established in the database, a series of comparisons (BLASTs) were performed using the GenomeNet [http://www.genome.jp/, using only the fully sequenced and annotated genomes of S. pneumoniae. For the nucleotide sequences the search was performed using the program BLASTN 2.2.29+, which uses nucleotide vs nucleotide alignments based on a punctuation matrix BLOSUM62 [S. pneumoniae they belong. All the DNA and protein sequences were downloaded and stored in an organized way using the fasta format.The identification of the genetic and protein sequences of interest to perform the comparative analysis was done, having as reference the codes (Locus Tag) in the genomes of s – KEGG , availabenomeNet , availabBLOSUM62 , 30. In BLOSUM62 , 32, thahttp://multalin.toulouse.inra.fr/multalin/, for which an identity matrix 1–0 was used to assign a penalty even for the slightest change in the nucleotides or amino acids sequences, covering substitution, deletions, insertions and variations in the length. From these analyses, the number of allelic and protein variants were determined for each gene according to the registry value assigned by the program to each sequence, where equal sequences have the same registry value, while different sequences possess different values. The results of the alignments were manually curated and stored for further analysis. Finally, the precise determination of the total mutations, synonymous and nonsynonymous was done using the software DnaSP V.5.1 [http://www.ub.edu/dnasp/. There, all the sequences found for a determined gene were introduced and the calculations were perfomed for the corresponding type of mutation as mentioned before.The multiple comparative alignments of pneumococcal sequences were done using the web tool MultAlin , availabSP V.5.1 , 35, avaS. pneumoniae strains and clinical isolates recovered from the nasopharynx, blood or cerebrospinal fluid (CSF) have been included up to date in genomic sequencing projects worldwide. However, pneumococcal strains, whose genomes are fully sequenced, annotated and publicly available, are the focus of this study. Therefore, a set of 25 pneumococcal strains were selected from the NCBI database, as population study, to perform the bioinformatic analysis needed to accomplish the construction of the variome of the virulence factors and two-component regulatory systems of Streptococcus pneumoniae  of those 8290 available S. pneumoniae genomes/strains was performed and a DNA-based Phylogenetic Tree was constructed by using the Genome Tree Report Tool of the NCBI. The topology of this phylogenetic tree (slanted cladogram) showed different pneumococcal lineages, where the selected set of 25 pneumococcal genomes/strains can be identified as external nodes (“well-distributed” key features highlighted in red), evidencing an optimal representation of the pneumococcal population  was determined using the database Prokaryotic 2-Component Systems - P2CS [An initial considerable number of pneumococcal virulence factor genes were identified, by employing the database VFDB . This daVFDB  did not cs genes  was detecs genes 7 was detThe genomes of 25 analyzed pneumococcal strains comprise genome sizes ranging from 2,024,476 bp in SPN034156 up to 2,245,615 bp in Hungary 19A-6. Likewise, the G + C content varies between 39.50% in CGSP14 and 39.90% in SPN034156. 670-6B is the strain with the highest number of genes (2430) and proteins (2352) and SPN034156 is the strain with the lowest number of genes (1956) and proteins (1799). Hence, the difference among genomes, regarding the number of genes and proteins can be up to 474 genes and 553 proteins, respectively. The overall number of genes for each pneumococcal genome evaluated here overmatches the overall number of proteins because the reported number of genes includes all the tRNA-, rRNA- and protein-encoding genes.oriC: 2.160.822–196). During the bidirectional replication of the genome, gene transcription must be simultaneous [S. pneumoniae, where only one operon, the tcs04 genes (TCS04), is not co-oriented with the replication fork  of the datasets and supplementary material reported by van Tonder and colleagues in 2017, only a few of the pneumococcal virulence factors  and regulators  were found in the pneumococcal “supercore” genomic list of 303 genes, based on the analysis of 3121 pneumococci recovered from healthy individuals from four different subsets of the global pneumococcal population [lytA gene, encoding the major pneumococcal autolysin, is found also in two and even three copies in 13, and 2 strains, respectively. This is most likely due to the multiple integration of prophages in the chromosomal DNA [gnd, encoding the enzyme 6-phophogluconate dehydrogenase, is duplicated and fused with a second copy of its downstream neighbor gene, which encodes the orphan response regulator (rr14). The remaining 39 of the 65 virulence factor genes were found to belong to the accesory genome, presenting different degrees of absence in the 25 strains. Thus, all these genes are not essential but are beneficial for fitness and pathogenesis. Striking examples are the genes encoding the Pilus-1 and Pilus-2 structures that have been identified to mediate adherence, contribute to virulence and promote invasion [pht genes). These gene products are associated with different processes of bacterial fitness and pathogenesis  and has an essential function in the metabolism [pcsB (Usp45) gene, which encodes for a 45-kDa secreted and immunogenic protein that is involved in cell division and stress response [hk05 and rr05 of ciaR/H (tcs05). The TCS CiaRH is involved in the resistance to cefotaxime, regulation of genetic competence and increase in pathogenicity in the respiratory tract in murine models [hk02 and rr02 , have been associated with resistance to erythromycin and are essential for the bacterial growth. Nevertheless, the latter was proven to be due to its regulon (pcsB), and was no longer essential upon ectopic expression of PcsB [rr05, rr02, rr06, and rr08 the most conserved. All the results obtained in this study support the global idea of a new generation of protein-based and serotype-independent vaccines for Streptococcus pneumoniae. The basis is the high degree of distribution and conservation of the virulence proteins in combination with the importance of their functions and immunogenic capacities. This probably makes them ideal pharmacological targets to treat the pneumococcus and its diseases. This might be an alternative to the immunization with the conjugated serotypes, or represent a strategy to combine immunogenic and highly conserved proteins with capsular polysaccharides to generate a serotype-independent immune response.The estimation of the variability for each individual virulence factor and pneumococcal regulator (at the DNA and protein level) allowed the construction of a partial variome for the analysed 25 pneumococcal strains. Briefly, the variome takes into consideration the estimation of (1) the presence, absence or the number of copies of genes in the different strains, (2) the number of total synonymous and nonsynonymous mutations, and (3) the number of allelic and protein variants explaining the variability for each factor. The results summarized in Tables umolysin , (2) thetabolism , but alsresponse . As for e models , 49, 50. of PcsB , 48. Pne of PcsB . The mutStreptococcus pneumoniae was achieved from 25 pneumococcal strains with fully sequenced and annotated genomes. According to the Molecular Phylogenetic Analysis performed on the NCBI website, this selected set of pneumococcal genomes ensured an optimal representation of the pneumococcal population (8290 strains) reported in the NCBI database up to date. Similarly, this study population set also represented an important group of highgly pathogenic pneumococcal serotypes , which have been also included in the current pneumococcal conjugate vaccine formulations (except serotypes 2 and 11A), used to prevent penumococal infections. A total of 92 different genes and proteins were identified, classified, and studied for the construction of the variome. The genes of the pneumococcal virulence factors and TCS, are distributed along the genome, and are located in such a manner that transcription is co-oriented with replication. The analysis of the gene distribution in this study population set showed that 26 of them were found in the 100% of the 25 pneumococcal genomes/strains (core genome), while 39 are part of the flexible genome. The estimation of the variability for each individual virulence factors, stand-alone regulator or TCS, indicated that the virulence factors with the lowest variability in the pneumococcus are pneumolysin, enolase and PcsB, while the regulators with the highest conservation are TCS05 (CiaR/H), TCS02 (VicR/K) and TCS08. Finally, all the results obtained here with the bioinformatic analysis performed, constitute the first model to compare, visualize and understand the future flood of new genomic data about the genetic variation (in terms of gene presence/absence or mutation) of pneumococcal virulence factors and regulators [Streptococcus pneumoniae and its disease burden.The construction of this “low-scale” Variome model for the virulence factors and regulators of gulators –53. The