Health professionals in athletic training, chiropractic, osteopathy, and physiotherapy fields, require high-level knowledge and skills in their assessment and management of patients. This is important when communicating with patients and applying a range of manual procedures. Prior to embarking on professional practice, it is imperative to acquire optimal situation-specific levels of self-confidence for a beginner practitioner in these areas. In order to foster this professional self-confidence within the higher education context, it is necessary to have valid and reliable scales that can measure and track levels and how they change. This study reports on the development and psychometric analysis of two new scales, Patient Communication Confidence Scale (PCCS) and the Clinical Skills Confidence Scale (CSCS), to measure confidence in these two areas for students in manual medicine programs. The Rasch measurement model was used to guide the development of the scales and establish their psychometric properties.The responses to 269 returned questionnaires over two occasions were submitted to psychometric analysis, with various aspects of the scales examined including: item thresholds; item fit; Differential Item Functioning; targeting; item locations; item dependencies; and reliability. To provide further evidence of validity, scores were correlated with two existing valid scales.Analyses showed that the scales provided valid and reliable measures of confidence for this sample of persons. High Person Separation Indices  provided statistical evidence of reliability, meaning the scales are able to discriminate amongst persons with different levels of confidence. For the PCCS, item categories were operating as required, and for the CSCS only two items’ thresholds were slightly disordered. Three tests of fit revealed good fit to the model  and results of the correlations with two existing valid scales were consistent with expectations.The importance of confidence cannot be overlooked in health education because students learning new information and skills, and dealing with challenging situations can be negatively impacted by a lack of confidence which can result in students disengaging from placements or leaving a program. Valid and reliable instruments are essential in tracking change in levels of confidence in specific skills over time and the examination of the degree of congruence between confidence and competence. Analysis of responses to the two confidence scales established that they are valid and reliable instruments. Health professionals in athletic training, chiropractic, osteopathy, and physiotherapy fields, require high-level knowledge and skills in their assessment and management of patients. This is particularly important when communicating with patients and applying a range of manual procedures, such as physically assessing joints and soft-tissue, performing manipulative or soft-tissue procedures, carrying out rehabilitation protocols, and bracing and taping techniques. Prior to embarking on professional practice, it is imperative to acquire optimal situation-specific levels of self-confidence for a beginner practitioner in these areas. In order to purposely foster this professional self-confidence within the higher education context, it is advantageous to have valid and reliable scales that can measure and track levels and how they change, and that may be used in conjunction with objective measures of competence.Although findings on the congruence between self-confidence and competence in health education are mixed -5, what In an education context, both high and low achievement students are typically informed and guided by their beliefs and perceptions rather than by reality ; in otheGiven that professional confidence is important, health educators need to know what learning activities aid its development, and have appropriate tools to measure any change . The valHowever, the availability of psychometrically robust instruments to measure professional confidence in manual medicine education is limited. Therefore, this paper describes a research study which aimed to develop two new scales to measure confidence in clinical and patient communication skills for students in manual medicine programs, and to use the Rasch measurement model  to estabOverall, the aims of this study were to develop and validate measurement tools for use in health education and research. It is innovative in that it recognises the vital link between confidence in communication as well as clinical skills and patient outcomes, thus the development of two separate scales was deemed important. The use of the Rasch model adds to the novel aspects of the paper.The data garnered from various informal student group interviews and critical reviews of literature helped shape the content and objectives of the self-confidence scales and eventual confidence questionnaire. Items and their format were developed after careful review of related scales -30 and hAccordingly, a preliminary instrument of 52 items or statements was developed with a primary focus on patient communication and clinical skills, and  a secondary focus on supervising clinicians. A six-point Likert-style response format for each item was utilised. Response categories were coded from 1  to 6 (“very confident”). The questions reflected interactions and experiences with patients that students were likely to encounter, and ranged from discussing general health issues to performing basic and focused physical examination procedures. For example: “How confident are you in your ability to discuss personal and/or sensitive issues with new patients?”; and “How confident are you in your ability to perform basic physical examination procedures such as blood pressure, pulse and respiration rate on a patient?” Content validity was assessed by a panel of educators and researchers affiliated with education programs and chiropractic clinical education internship programs in Australia and the United States. Panel members were asked to review the scale and comment on each item and the overall format. They suggested minor alterations for a few items, and recommended the inclusion of a demographic section and a self-reflection section that invited the students to qualify their responses.The scale was divided into two parts. One part of the scale focussed on patient communication, labelled the Patient Communication Confidence Scale (PCCS). The other part focussed on clinical skills, labelled the Clinical Skills Confidence Scale (CSCS). The two scales represent different aspects of self-confidence, both of which are important, and more diagnostic information about these two different aspects could be gained by measuring them separately and thus being able to see whether levels differ. If levels on the two aspects were to differ, different teaching strategies could be aimed at developing each aspect. It is an empirical question whether the two scales could be conceived as representing the same construct and for some purposes a single score may be all that is required to make teaching and learning decisions. However, in this case, the research aimed to get information about each of the two aspects as they are each essential and are likely to require different strategies to address them.To assist with the validation process, two existing valid and reliable scales were incorporated, the Personal Report of Communication Apprehension (PRCA-24) and General Self-efficacy (GSE) scales. The PRCA-24 scale measures feelings about communicating with others. However, only one sub-category  was used in this study, as the other sub-categories are not typically encountered in clinical contexts. Prior research has demonstrated content, criterion, and construct validity of the PRCA-24 . The GSE1. General Self-efficacy Scale (GSE).2. Personal Report of Communication Apprehension Scale (PRCA-24), Interpersonal communication sub-scale.3. Patient Communication Confidence Scale (PCCS), 28 items, 6 response categories, no reverse items.4. Clinical Skills Confidence Scale (CSCS), 27 items, 6 response categories, no reverse items.5. Self-reflection section.6. Demographic section.The PCCS items covered nine aspects of patient communication such as encouraging behaviour change, history-taking, explaining, and being supportive. The CSCS addressed eight aspects of confidence in clinical skills such as manipulative, X-ray, and physical examination procedures.Participants included seven cohorts of chiropractic students (n = 269) enrolled in internships in tertiary institutions in Australia and the United States. All cohorts had comparable clinical curricula that provided similar professional experiences such as recording patient histories, and supervised assessment and treatment of patients. Human ethics approval and student consent were obtained. The CQ was administered at the beginning of students’ clinical internships and was repeated five months later  and again ten months later . Data from only the first and third occasions (beginning of the study and ten months later) were used to examine the validity and reliability of the PCCS and CSCS. Combining data in this way is an accepted procedure made possible by the Rasch model’s properties of invariant comparisons. The legitimacy of the procedure can be tested empirically using differential item functioning (DIF).Student responses to 269 returned questionnaires over two occasions were submitted to psychometric analysis using the polytomous Rasch model (PRM) ,38, throlinear scale; and second, the measures will be invariant . In addition, examination of differential item functioning will provide evidence of whether measures are invariant  across designated groups for which the fit has been confirmed [The Rasch model can be used to examine data for flaws or problems indicated by a failure to fit the model . Showingonfirmed ,41,46.locations. More formally, the Rasch model provides measurements that are compatible with fundamental or additive conjoint measurement studied in mathematical psychology [In the Rasch model, the relevant statistic for any person is simply the total score across items where the scores are successive integers assigned to successive categories, which is the same statistic as that used traditionally. Some items may be dichotomous, and some may have more than two ordered categories. However, these scores are not themselves linear and should not generally be treated as measurements. In particular, they are affected by floor and ceiling effects so that a difference of a raw score of 2, say, at one part of the continuum of the construct does not represent the same difference as a score of 2 on another part of the continuum. The transformation of the raw scores using the Rasch model produces linearised scores for each person which can be treated as measurements and used in standard statistical analyses. These linearised scores are known as ychology .The Rasch model is a probabilistic one which provides an appropriate model for typical social science data. For polytomous items the equation takes the form:Xni, is the random variable of the response of person n to item i and where the value of this variable is an integer 0, 1, 2, 3, …, m, βn is the location of the person on the variable, (δik), k = 1, 2, 3, …, mi is a vector of thresholds of item i at which the probability of a response in adjacent categories is identical, and where (i) tatement ,50.The RUMM2030 software  providesAs mentioned in the introduction, data analysis addressed three primary aims, the first of which was to establish the internal consistency and reliability of each scale. In other words, do the sets of items each represent a single construct at this level of scale? If they do, then one is justified in adding scores to obtain a total score on each scale and then using those total scores  for other statistical tests such as comparisons of mean scores amongst groups or over time.The second aim was to determine whether the items of each scale have the same psychometric properties across different groups of participants: this is termed Differential Item Functioning (DIF) and it determines whether the items have similar psychometric properties across different groups of participants, that is, whether the items have invariant properties across groups. If items show DIF across groups, they should not be used to compare person performance, unless individuals are from the same group. In this study, the groups of interest were gender, age, experience with the profession, entry qualification (previous degree or not) and the occasion of administration.The third aim was to provide evidence of the convergent validity of the PCCS and CSCS by examining their statistical correlations with the established GSE and PRCA-24 scales which assess some aspects of confidence.To address the first aim, various aspects of the scales were examined. The first aspect was the operation of the response categories. The item thresholds  are required to be correctly ordered. The second aspect was the fit of each set of items to the Rasch model. If the items fit the model, which is evidence of internal consistency, they can be accepted as measuring a single variable at this level of scale. Two tests of fit - one statistical (the chi square) and one graphical  - were used to judge this. In the Rasch paradigm generally, no one test of fit is sufficient to make a decision about fit. A third aspect was the targeting of items and persons to each other: this is established by examining the joint distribution of item and person locations on the same continuum. A fourth aspect, item dependencies, was examined by inspection of the residual correlations between items. If items show dependency, then one item in each pair is redundant and retaining both artificially increases the reliability. Such dependencies may also indicate the presence of sub-scales which can be further examined through the principal component analysis of residuals. Lastly, reliability is gauged using the Person Separation Index (PSI), which is the Rasch equivalent of Cronbach’s alpha.To address the second aim – to establish whether the items operate relatively consistently across different groups, differential item functioning across the groups for Gender, Age, Prior Experience, Entry qualification and Occasion was examined.Lastly, to address the third aim to provide further evidence of validity , student scores on the scales were correlated with scores from the same students on two existing scales that measure constructs related to but different from confidence and whose validity has been established in the research literature, namely, the GSE and PRCA-24 .The results of these analyses provide information about the validity and reliability of the two scales. If these are satisfactory, the person locations (the linearised raw scores) can be used for further analyses as, for example, the comparison of mean scores (person locations) for the different groups of interest, and the investigation of changes in mean locations over time.Two hundred and sixty-nine students agreed to complete the questionnaire (269 matched questionnaires across two occasions) with the following demographics: 153 males and 116 females; age: 20–25 years (136), 26–35 years (106), 36+ years (27); possession of degree upon entry, 153 students, no degree upon entry, 116 students.Using complete information for each person and item, the scales were analysed using the polytomous Rasch model with 10 possible response categories. Table Analysis of the operation of the categories for the PCCS (28 items) and CSCS (27 items) indicated two items having disordered categories in the CSCS and none in the PCCS. There were also several significant residual correlations between pairs of items in both scales. The three tests of fit - two statistical  and one graphical  – revealed two misfitting items each in the PCCS and CSCS. No items in either scale showed Differential Item Functioning (DIF) according to any of the groups of interest. Although the Person Separation Indexes (PSI) were high at 0.962 (PCCS) and 0.933 (CSCS), these results are likely to have been inflated due to the high number of item dependencies as indicated by significant residual inter-item correlations.Upon review of the initial analysis, simultaneous examination of several aspects of each item was undertaken in order to achieve an improved psychometric properties for each scale. This examination included analysis of the operation of item thresholds (the cut points between adjacent categories), item fit, the item’s location, item dependencies , DIF, and the targeting between items and persons). Items that were flagged as anomalous by the statistics were examined for wording and content. If any plausible issue was detected with the item, it was removed from the scale.The following section addresses different aspects of the analytical review process and actions that were taken to improve them where necessary.For the PCCS, the six response categories operated as required, with all items having ordered thresholds. For the CSCS, two items (12 and 27) had slightly disordered thresholds, meaning that the categories were not operating correctly. The number of categories for item 12 was reduced to three , and those for item 27 reduced to four . Re-analysis then indicated that the categories for all items were ordered correctly.For each scale, two tests of fit were considered - the item-trait interaction (chi square) statistical test, and the Item Characteristic Curve (ICC) graphical test. These revealed generally good fit to the model. The results of the statistical tests of fit for the initial analyses with all items are shown in Table confidence in taking a medical history/interviewing a patient who is wheelchair bound) was the least well-fitting according to the chi square statistical test and the graphical test (ICC) of fit. The ICCs for this item and for an item with good fit (for comparison), are shown in Figures In the PCCS, item 20  showed some minor misfit. However, all items could be retained at present because Item 20 represents an important aspect of clinical practice, represents less than 5 percent of all items, and is unlikely to have a marked effect on person measures. Its performance should be monitored in future analyses. Item 16 , although appearing to be more chiropractic or osteopathic orientated, represents an important component which is discussed with patients who may undergo manipulative procedures by various manual health care professional such as physical therapists, and athletic trainers. Therefore it too should be retained for future evaluation.The ICCs show the theoretical probabilities (the continuous curve) of endorsing the item across the range of person locations on the whole set of PCCS items, and the obtained probabilities for six class-intervals of person locations (the dots). The dots should follow the theoretical curve closely if fit is good. The chi square statistic represents the deviations of the obtained dots from the theoretical curve. The operation of Item 20 is a little inconsistent and tends not to discriminate as well as the other items amongst persons with different total scores particularly at the highest and lowest person locations . This may be because it addresses a very specific situation which students may or may not have encountered. When this item was deleted and the PCCS re-analysed, only item 16  was the least well-fitting item, and was deleted from further analysis. It tended to over-discriminate—a typical result for items that are aimed at assessing a property in an overall or general sense. The ICCs for this item and for an item with good fit are shown in Figures confidence in ability to apply orthopaedic bracing or supports on a patient). It tended not to discriminate as well as other items, but this item was retained at this stage because it refers to a commonly used therapeutic procedure in the scale’s targeted population. Its performance should be reassessed in future analyses.In the CSCS, item 1 , and item 19 was the most difficult . For the CSCS, item 24 was the easiest , and item 27 was the most difficult . These results were as expected theoretically, in terms of the relative levels of confidence needed to agree with each item. This ordering of items provides evidence of the construct validity of the scales.Tables confidence in ability to start a conversation with a new patient and confidence in ability at taking a new patient’s current and past health medical history) is redundant. Accordingly, the PSIs for the PCCS (0.962) and CSCS (0.930) are likely to be inflated, but nevertheless suggest the scales can provide reliable measures.In the PCCS and CSCS, respectively, 18 and 20 pairs of items showed dependencies . This means that one item of each pair is not adding much information about levels of confidence, and, further, these dependencies are artificially inflating the reliabilities. For example, one of the two items 3 and 8  may be legitimately compared.Figure Table In summary, for the PCCS, all 28 items were retained with no alterations to response categories and a recommendation that item 20 be monitored in future research. The CSCS required a reduction in the number of response categories for two items, and a deletion of one item in order for the set of items to perform well as a scale to measure confidence in clinical skills. Additionally, it may be that the two scales could be combined into a single scale representing confidence in physical and person-oriented manual medicine skills. However, two measures are likely to be more useful than a single measure because they provide educators with a more focused guide when diagnosing where students most require help.This paper has described the development and psychometric analysis of two new scales aimed at measuring confidence in clinical skills and in patient communication skills for students undertaking internships in manual medicine programs, particularly focussing on chiropractic students. The Rasch model has not previously been applied for psychometric analysis in the manual medicine field, and thus, in this regard, the study breaks new ground.confidence in interviewing wheelchair bound patient) was the least well-fitting. Whilst this item was intended to suggest a potentially difficult scenario and challenge students’ confidence, it did not discriminate amongst persons across the range of total scores and thus is likely to be assessing something other than the construct represented by the majority of the items in the scale. Further qualitative analysis is required to establish why it did not fit the model as well as the rest of the items in its scale, but it may be because it predicted a scenario which some students may and other students may not have encountered. Item 16  also showed some misfit, but was retained at this stage. It’s content includes an important term in the chiropractic, physiotherapy and osteopathy professions, referring to a joint in the body not moving in a normal pattern. However, some chiropractic education programs may use the term ‘subluxation’ more often than ‘joint dysfunction’. Thus, for students unfamiliar with the term, the item measures knowledge of terminology. This highlights the need to carefully consider how each item is worded and whether the meaning mirrors the content of a particular program.The first aim was to establish the internal consistency and reliability of the PCCS and CSCS. The results showed that the scales, with some amendments, provided internally consistent and reliable measures of confidence for participants. With the exception of very few items, items fitted the model. For example, in the PCCS, item 20  was the least well-fitting. The conclusion was that it was too broad in focus. This was also the case for item 1 in the PCCS, which measured overall level of confidence in patient communication and was amongst the easiest items for participants to agree with. Items that measure confidence in a general way should be reconsidered. In the CSCS, item 13 (confidence in applying bracing/supports) also showed some misfit. It tended not to discriminate as well as other items, possibly due to some students having more experience working in athletic and sporting areas. However, it was retained because it refers to an important competence in most chiropractic, physiotherapy, osteopathy and athletic training programs. Its performance could be monitored in future research.In the CSCS, item 1 , and item dependencies indicate that only one of these items may be required to represent the concept of dealing with different categories of people.The second aim was to verify whether each scale had similar psychometric properties across different groups of participants. There was no evidence of differential item functioning in the PCCS or CSCS according to gender, age, prior experience, entry qualification, or occasion of administration, which means that the scales can be accepted as representing the same construct across different categories of persons  within each of these groups. Therefore, measures on each scale may legitimately be compared across different sub-groups. This is particularly important when changes over time are being investigated or when different curricula for different groups of students are being considered.The third aim was to investigate the convergent validity of the two new scales by examining the correlational association between the new scales and the established GSE and PRCA-24  scales. Results showing a positive correlation between confidence scores for clinical and patient communication skills and general self-efficacy, and a negative correlation between confidence scores for patient communication and interpersonal communication, indicated significant, though relatively small, associations between the new and existing scales, and add to the evidence of the validity of the new scales. Hence the new and old scales measure similar constructs to some degree, but the indications are also that the new scales provide measures of distinctive properties.In regard to validity, the Rasch analyses provide evidence of internal consistency which is an aspect of construct validity. Content validity was addressed by the assessment of a panel of educators and researchers, and convergent validity was assessed by the scores on the two new scales being correlated with scores from the same students on two existing scales. Consequential validity, a type of validity that addresses the intended and unintended consequences of test interpretation and use ,52 may bLimitations of the study include the focus on chiropractic students. The chiropractic field may not represent the full range of manual medicine programs, which also include osteopathy, physical therapy and athletic training fields. For example, osteopathy curricula can vary extensively, with many programs in the United States modelling medical education with minimal to no manual techniques, in comparison to programs in Australia, which have a strong focus on manual therapies. Physiotherapy  curricula especially emphasise pre- and post-surgical rehabilitation, and neuro-rehabilitation, such as post-stroke and post-spinal cord injury with chiropractic education covering this minimally. Athletic training specifically addresses on-field emergency procedures and pre-event functions such as taping, stretching of athletes and monitoring warm-ups. These noted differences in curricula, and thus professions, highlight the need to carefully consider the content of the items in the two new scales.Another limitation is the analysis of the relationship between confidence and competence, which was not undertaken in this research. These constructs are intrinsically related, and arguably this relationship needs to be better understood, in order to recognise self-confidence as a vital construct which requires thorough investigation in health education. According to Bandura ,15, the This is the first study to undertake rigorous development and psychometric analysis of two new scales that can be applied during student internships in manual medicine programs. The results indicate that the two scales can provide educators and researchers with sound measures of student confidence in clinical and patient communication skills, which may then be used to identify students who require additional help, and to guide curriculum development. The scales may also be used to measure changes in levels of confidence in specific skills over time, and to examine the degree of congruence between confidence and competence.There are no financial or non-financial competing interests  to declare in relation to this manuscript.MH developed the scales. IS performed the content analysis. SV assisted with content analysis and outline of the project and paper. All authors contributed substantially to the conception and design of the study, as well as to the critical revision of the paper. All authors approved the final manuscript.


Toxoplasma gondii can cause serious damage that can be diagnosed in utero or at birth, although most infants are asymptomatic at birth. Prenatal diagnosis of congenital toxoplasmosis considerably improves the prognosis and outcome for infected infants. For this reason, an assay for the quick, sensitive, and safe diagnosis of fetal toxoplasmosis is desirable.Congenital infection caused by To systematically review the performance of polymerase chain reaction (PCR) analysis of the amniotic fluid of pregnant women with recent serological toxoplasmosis diagnoses for the diagnosis of fetal toxoplasmosis.A systematic literature review was conducted via a search of electronic databases; the literature included primary studies of the diagnostic accuracy of PCR analysis of amniotic fluid from pregnant women who seroconverted during pregnancy. The PCR test was compared to a gold standard for diagnosis.A total of 1.269 summaries were obtained from the electronic database and reviewed, and 20 studies, comprising 4.171 samples, met the established inclusion criteria and were included in the review. The following results were obtained: studies about PCR assays for fetal toxoplasmosis are generally susceptible to bias; reports of the tests’ use lack critical information; the protocols varied among studies; the heterogeneity among studies was concentrated in the tests’ sensitivity; there was evidence that the sensitivity of the tests increases with time, as represented by the trimester; and there was more heterogeneity among studies in which there was more time between maternal diagnosis and fetal testing. The sensitivity of the method, if performed up to five weeks after maternal diagnosis, was 87% and specificity was 99%.2). The tests show low evidence of heterogeneity with a sensitivity of 87% and specificity of 99% when performed up to five weeks after maternal diagnosis. The test has a known performance and could be recommended for use up to five weeks after maternal diagnosis, when there is suspicion of fetal toxoplasmosis.The global sensitivity heterogeneity of the PCR test in this review was 66.5% (I Toxoplasma gondii, an obligate intracellular protozoan parasite. Its prevalence varies widely from place to place, especially when related to health conditions and socio-economic indices [Toxoplasmosis is a disease that is endemic globally and is caused by  indices ,2. In so indices .T. gondii infection is acquired primarily through ingestion of cysts in infected, undercooked meat or oocysts that may contaminate soil, water, and food [T. gondii is typically mild or subclinical in healthy humans. However, infection during pregnancy puts the fetus at risk of congenital infection, which can cause serious lesions that can be diagnosed in utero or at birth [and food . Infectiat birth . The risClassical signs, such as hydrocephalus, chorioretinitis, and intracranial calcifications (Sabin’s triad), may not be present at birth; however, sequelae resulting from undiagnosed disease appear in adolescence or adult life. Undiagnosed and untreated patients develop irreversible lesions, especially in the eyes and brain .Prevalence of congenital infection ranges from 0.1 to 0.3 per 1000 live births. The maternal–fetal transmission rate increases with gestational age at maternal seroconversion, from less than 15% at 13 weeks of gestation to over 70% at 36 weeks.The prenatal diagnosis of congenital toxoplasmosis has considerably improved the prognosis and outcome for infected children . MaternaIn some European countries, such as France, monthly serologic screening of susceptible pregnant women has been recommended since 1992. If a serologic exam indicates acute infection, maternal treatment with spiramycin is initiated in an attempt to prevent transmission to the fetus. If fetal infection is confirmed by polymerase chain reaction (PCR) analysis of the amniotic fluid, spiramycin is replaced by a combination of pyrimethamine, sulfadiazine, and folinic acid [A recent study  evaluateIn Brazil, prenatal screening is suggested but is a non-mandatory public policy with protocols that vary by region, thereby lacking uniformity . Fetal tThe interpretation of diagnostics for congenital toxoplasmosis using serological tests is not a trivial task due to the kinetics of emergence and the fact that transplacental transmission is highly variable ,13. The T. gondii has been the use of PCR analysis of amniotic fluid instead of umbilical cord blood samples [Toxoplasma was published in 1989 by Burg et al. [Perhaps the greatest advance in the prenatal diagnosis of fetal infection with  samples . The detg et al. . He descg et al. ,16.Against this background , the goal of this investigation was to systematically review the literature on PCR performance when using amniotic fluid for the antenatal diagnosis of congenital toxoplasmosis.T. gondii during pregnancy, (b) studies that evaluated the performance of PCR analysis of amniotic fluid, cord blood, or a pregnant woman’s blood, (c) studies that compared the PCR results with any “gold standard” or test accepted as better evidence, and (d) studies that reported the sensitivity and specificity or presented data to calculate these parameters. Summaries in any language were accepted for the initial classification. For full texts, the inclusion was restricted to the following languages: Portuguese, French, English, Spanish, and Italian. There was no limit on the paper’s disclosure date for inclusion.Studies on the use of PCR for fetal toxoplasmosis diagnosis were identified according to the following criteria: (a) original studies that included pregnant women with evidence of acute infection by The following exclusion criteria were applied: (a) studies that considered other populations  or ones in which testing for toxoplasmosis was not related to fetal disease, (b) reviews, (c) editorials, (d) consensuses, protocols, and guides for clinical practice, (e) case reports, (f) congress summaries, (g) validation of lab methods, (h) studies in which the PCR data were used only to assess the treatment outcome and (i) neonatal or postnatal diagnostic investigations.The initial search was conducted using the following databases: PUBMED, EMBASE, SCOPUS, WEB OF SCIENCE, and LILACS. The bibliographies of articles selected for full-text reading and of review articles were also checked to identify relevant works that had not been found in the electronic search. Our first search in the database was on January 25, 2012, and the last search on August 12, 2015. Initially, a search in MEDLINE with different combinations of terms, either in the title or in the summary and keywords, was conducted to identify terms capable of retrieving articles about the theme of interest that were already in the investigator’s possession. Details on the search strategy adopted to identify original studies are reported in After the application of search strategies for remote databases, the identified references were stored in bibliographical data management program for exclusion of replicas and to organize the summaries. The summaries were read independently by two reviewers, who met to resolve disagreements, selected the texts to be read in full, and extracted data. This last step was performed by three reviewers independently; one reviewer read all of the texts, whereas the other two read only parts of them. A second meeting was held to resolve disagreements regarding the inclusion and exclusion criteria, and the texts to include in the review were selected.A form for data capture was designed and tested specifically to be used during the reading of full texts, reported in The following were evaluated for each study: pregnancy age, nationality of the subjects, pregnancy stage at acute toxoplasmosis diagnosis, time between diagnosis of maternal infection and testing for fetal toxoplasmosis, proportion of pregnant women who received treatment for toxoplasmosis before fetal testing, time between the beginning of treatment and fetal testing, and whether there was evidence that the pregnant woman’s previous treatment influenced the result of the index test.The index test was defined as the PCR analysis of amniotic fluid of pregnant women acutely infected by T. gondii during pregnancy. Acute maternal infection was defined as change from negative to positive specific immunoglobulin G (IgG) antibodies or a significant increase in IgG with concomitant high immunoglobulin M (IgM) titers .Clinical follow-up of the child during its first year was considered the standard reference test. Diagnosis of congenital toxoplasmosis was confirmed when the specific antibodies  were detected in the newborn’s blood and persistence after the first year of life of specific IgG. Congenital toxoplasmosis absence was defined by negative IgG results in the absence of treatment at 12 months of age .Evaluation of the risk of bias in the full texts included in the review was conducted independently by the reviewers during data extraction using the QUADAS 2 . This tohttp://www.r-project.org/ [2- and p-values from Cochrane’s Q test were obtained for all trimesters together, each trimester individually, and early programmed elements that eventually displayed significance in this analysis.The data were analyzed using the R-project statistical software, available at ect.org/  with theForest plot graphics were created for each group of data separated by the trimester of pregnancy when the diagnostic investigation occurred.The risk of reporting bias was explored by using funnel charts and Begg tests for each group defined by pregnancy trimester and variables found to be significant during an exploration of the data’s heterogeneity.The initial search returned 1.269 summaries and titles, 715 of which remained after the exclusion of duplicates. Applying the inclusion and exclusion criteria, 118 summaries were selected to have the full texts recovered. The full texts were evaluated independently by three reviewers. At the end of this phase, 20 articles were selected for inclusion in the review .The 20 works included ,12,19–35Test performance at first trimester at seroconversion was reported in six studies (30%); in one of these studies , two tesMaternal treatment before fetal investigation was reported in ten works, covering a total of 1.685 patients; 1.623 (96.3%) received treatment. Neither data nor sufficient discussions regarding the influence of maternal treatment on the sensitivity of PCR tests sing amniotic fluid were provided in these studies.Three studies proposed a model to limit the probability of fetal infection based on a test that amplifies T. gondii DNA. In a European multi-center study , the posIn all studies, the PCR test was an in-house assay; that is, it was not commercially available. Regarding the type of test, four studies (20%) employed quantitative real-time PCR and 16 (80%) used qualitative PCR, in two of these studies more than one assay was used ,35. No tAmong the included studies, 15 (75%) amplified the target DNA of the B1 gene. The primers used for the gene amplification process varied among the studies.All studies used clinical and serological assessments of the infant during the first year of life as a reference. Some works used additional tests for reference, such as inoculation of amniotic fluid in the peritoneum of mice , parasite isolation in cell culture , and histopathological exam of the placenta .The low risk of bias was found in the applicability of the test rather than how studies were conducted and reported .The flow of patients through the study was the domain most susceptible to bias, followed by the conduction of the index and the reference tests. The greatest rate of “unclear” results was in relation to the index test, followed by the reference test. .The lack of information regarding the risk of bias domains makes it difficult to truly appreciate the quality of research conduction reporting. Regarding the applicability, the risk of bias was lower and was similar in the three domains .2 = 66.5%). The global specificity of the test was 98.3%. The DOR for all trimesters together was 453.482, with moderate evidence of heterogeneity  the utility of PCR tests for the diagnosis of fetal toxoplasmosis is poorly studied, and reports on this topic lack information, which complicates the interpretation of the results; (b) the protocols vary among studies; (c) the heterogeneity is concentrated in the sensitivity; (d) there is evidence that the test sensitivity increases with progressive trimesters; and (e) there is evidence that the heterogeneity is concentrated in studies in which there was more time between maternal diagnosis and fetal investigation.In our review and evaluation by QUADAS 2, we found that the risk of bias was primarily related to the study methodology, particularly the choice of the index and reference tests. We observed no reduction in the risk of bias with publication time; the risk was similar between the older and more recent studies, reflecting the lack of significant advancements in study methodology and reporting in this field despite initiatives, such as Standards for Reporting of Diagnostic Accuracy (STARD) . VariabiThe existence of many protocols may be related to two fundamental aspects: (a) adaptations of an original protocol to situations that change over time and in different places according to available resources and (b) the perception that the tests can be improved through protocol modifications.We observed that the heterogeneity is concentrated in the sensitivity; the sensitivities vary so widely that they likely do not correspond to the same test. The statistical measure of heterogeneity can represent differences in tests, samples, or investigation procedures.Heterogeneity is often attributed to varying test protocols derived from the lack of a standardized method. The experience and technical ability, quality, and proficiency control of each laboratory are fundamental elements necessary to solve this problem .Although the heterogeneity in this investigation was not completely explained, there was a considerable reduction in the heterogeneity for sensitivity when the results were stratified by the time between maternal diagnosis and fetal investigation. This result indicates that beyond the differences among test protocols, the clinical protocols and disease development during this period may be primarily responsible for the observed differences in test performance.Relevant events that may occur between maternal diagnosis and fetal investigation include, for instance, variations in parasitaemia. It is reasonable to imagine an increasing initial parasitaemia followed by a peak and a period of decreasing parasitaemia. However, this behavior is based on anecdotes and thus speculative; rigorous evidence remains nonexistent. Tests performed on samples collected during these different periods of infection can simplify or complicate the DNA amplification of the parasite depending on the initial amount of parasite in the sample. Additionally, the encystment of the parasites can occur in unfavorable conditions, which in turn complicates DNA identification.It has previously been suggested that the treatment be instituted up to three weeks after maternal seroconversion, with the intent of reducing transmission of the maternal infection to the fetus . HoweverOur review provided evidence that the test’s sensitivity increases with pregnancy trimester, although there were no statistically significant differences. This observation is based on data in the literature . The lowEven though the possible explanations for this observation are speculative, if confirmed, this observation is relevant to the diagnosis of fetal toxoplasmosis; similar results from tests conducted during different stages of pregnancy could have different interpretations.Toxoplasma gondii infection at less than 18 weeks’ gestation and should be offered no less than 4 weeks after suspected acute maternal infection to lower the occurrence of false-negative results [Another result from our review is that heterogeneity tends to be concentrated in studies where there was more time between maternal diagnosis and fetal investigation. Test performance tended to be better for investigations performed up to five weeks after the diagnosis of acute maternal infection compared to investigations performed after this period. The literature recommends that amniocentesis should not be offered for the identification of  results ,44.Although there is a lack of evidence evaluating the effect of treatment on fetal infection, a possible interpretation is that maternal treatment may affect test performance; in other words, when the treatment is instituted before conducting a diagnostic investigation, the parasite load may be reduced such that the identification of the parasite is hindered. Authors seldom discuss possible interference of the treatment with the test when treatment is started before the diagnostic investigation, and the few authors who do comment on this matter did not observe this effect ,22,34. IA pertinent hypothesis suggested previously  is that A limitation of this study was the exclusion of 17 articles available in languages inaccessible to the reviewers –61 and tAs discussed above, our inability to fully explain the heterogeneity is a limitation of this work. The presence of heterogeneity means that the estimated sensitivity does not represent all included tests, which makes the test’s overall sensitivity unknown. Although we have found that heterogeneity decreases in studies in which the average of time between maternal diagnosis and fetal investigation was less than five weeks, these data were not available for many of the included studies. Additionally, we must consider that this information is aggregated by study and does not consider each patient individually. Besides this, we haven´t been able to identify if contamination in DNA assay could be responsible for results different of 100% in specificity before or after 5 weeks of maternal infection.It was possible to ensure that none of the diagnostic tests related a reproducibility measure. Noncommercial diagnostic tests and tests that are highly operator dependent typically have a higher variability in their measurements, that is, a lower reproducibility. A fundamental question of reproducibility is how variable the measures can be for the tests to be considered reproducible. This concept is not directly connected to test validity but does interfere with the interpretation of its results. Some authors have adopted more automated procedures ,35,30 anDespite the limitations regarding the diagnostic performance of PCR analysis of amniotic fluid, this method provides operational advantages compared to other methods, such as mouse inoculation. PCR analysis of amniotic fluid is the fastest method, yielding results within 4 h, and it is the safest method, with a fetal loss rate attributed to amniocentesis of 0.5–1% . DespiteClinical and serological follow-up of the infant is the most accepted and most widely used method of determining a definitive diagnosis of congenital toxoplasmosis and discarding false negative or false positive molecular diagnoses in clinical studies. However, it is reasonable to consider that such follow-up procedures can suffer from the effect of anti-parasite treatment, meaning that correct identification of the parasite followed by treatment can result in the absence of antibodies at the end of successful treatment, thus leading to improper classification of the reference test. The authors of the included texts did not discuss this issue as a possible limitation, although in theory, it would reduce the sensitivity and specificity of the test. Unfortunately, this issue is a possible limitation of any study that uses a follow-up as a reference.Despite these limitations, the available evidence demonstrates that the PCR test has a sensitivity of 87% and a specificity of 99% when performed up to five weeks after maternal diagnosis. Additionally, the test performance can be differentiated according to pregnancy trimester. Therefore, the test can be recommended in a period when there is increased suspicion of fetal toxoplasmosis. However, the absence of a Kit commercial widely available and reliable, will require health professionals to refer these patients to a research unit capable of executing the test or to treat the suspected infection empirically without diagnosis.Toxoplasma DNA; and (e) exploration of how the molecular test results can be combined with other clinical information, particularly the stage of pregnancy at which maternal infection occurred.The following aspects must be improved for future investigations of PCR performance for the diagnosis of fetal toxoplasmosis: (a) adherence of research reports to recommended standards; (b) discussion of the extent to which the therapeutic schemes administered before diagnostic investigation impact the test performance; (c) estimation of reliability, even with the use of automated methods; (d) exploration of new targets for amplification that are more repetitive in the For the diagnosis of fetal toxoplasmosis, having PCR as a tool is useful because molecular technologies are becoming less expensive and simpler to execute. Additionally, these technologies are operationally simpler than the alternative tests, and the results can modify the therapeutic regimes for pregnant women identified with acute infection.We find a lot of heterogeneity in our results. Considering the diversity of parasite concentration in amniotic liquid during the pregnancy, and amniocentesis as an invasive procedure, someone could argue that introduction of monthly prenatal serological screening would be enough to detect seroconversion in appropriate time and then help for a significant reduction in the rate of congenital infection. Maybe a monthly screening using serological and molecular tests could be more suitable for the Brazilian Health System. However this does not invalidate the need for fetal research and its importance for a proper fetal and neonatal approach.2 = 66.5% for sensitivity), which makes difficult to interpret these summary estimates. In the subgroup with an average time between maternal infection and fetal investigation of up to five weeks, there was moderate to low evidence of heterogeneity in sensitivity and specificity .This review found that the PCR test had a global sensitivity of 83% and specificity of 98.3%; however, these measurements display strong evidence of heterogeneity Click here for additional data file.S2 Text(PDF)Click here for additional data file.


Hospital Israelita Albert Einstein.To describe the flow and costs associated with the diagnosis and treatment of patients with breast cancer who come from the public healthcare system and were treated at Hospital Israelita Albert Einstein for diagnostic radiology, medical oncology, radiotherapy, and oncologic/ breast reconstruction surgery. The data were collected retrospectively from the hospital records, patient charts, pharmacy records, and from the hospital billing system.Between August 2009, and December 2011, 51 patients referred by the Unified Public Healthcare System (SUS) had access to The total sum spent for diagnosis and treatment of these 51 patients was US$ 1,457,500.00. This value encompassed expenses with a total of 85 hospitalizations, 2,875 outpatient visits, 16 emergency room visits, and all expenses associated with these stays at the hospital. The expenditure for treatment of each patient submitted to biopsy, breast conserving surgery, adjuvant chemotherapy without trastuzumab (a regime with taxane followed by anthracycline), radiotherapy, and 5 years of tamoxifen was approximately US$ 25,500.00.Strategies for cost-reduction of treatment in the private setting are necessary to enable future large-scale public-private partnerships in oncology. In the female Brazilian population, excluding non-melanoma skin cancer, breast cancer is the neoplasm with the highest incidence  and the primary cause of cancer-related mortality. On the other hand, the Unified Healthcare System [Sistema Único de Saúde] (SUS) has too limited a structure to be able to adequately diagnose and treat all these new cases, thus generating deficiencies in tracking and diagnosis, with consequent delay at various phases of treatment, leading to a negative impact on the prognosis of patients with cancer. Published data have demonstrated that in public health system there are delays between the first symptoms of the disease and the establishment of the diagnosis, between the diagnosis and treatment, and among the various phases of treatment,4. These delays can lead to worse outcomes, including in lower survival rates,6. Within this context, the structural deficiency of SUS could be partially compensated by the establishment of public-private partnerships,7(PPP) between governments and private hospitals, which could absorb part of the cases for immediate treatment until the public service is ready to care for all the demands with no delays. In order for a partnership such as this to be possible, it is necessary to know the details of the costs of breast cancer treatment.In Brazil, according to the estimates of the National Cancer Institute Hospital Israelita Albert Einstein (HIAE) initiated the Program for Oncologic Patients (PRPO). The program assisted SUS in the training of human resources related to the oncology screening, diagnosis and treatment, in addition to making available the HIAE structure for the diagnosis and treatment of breast cancer patients. These patients used HIAE from the diagnosis through the surgical intervention, radiotherapy and chemotherapy, until follow-up once treatment was ended. The program also had the objective of promoting research by investigating the costs of breast cancer treatment, following the best international standards(8), in order to collect this information to be used as a basis for future PPPs in this area.With the aim of better assisting SUS and establishing the actual costs of such a PPP, in 2009, To describe patient flow between SUS and HIAE and to evaluate the costs associated with diagnosis and treatment of patients with breast cancer seen by the PRPO, in order to allow planning of future social projects of public-private partnerships.Vila das Belezas and from the Specialties Outpatient Clinic in Pirajussara, both located in the Southern area of the city of São Paulo (SP), were referred to HIAE for breast biopsies, and then again referred to the public network after the procedures. Once the diagnosis of breast cancer was confirmed, patients selected as per the judgment of the original UBS were directed to the Oncology and Hematology Center of HIAE for treatment. For a 30-month period, the PRPO offered the structure of HIAE so that these women could have access to the services of radiology, oncology, radiotherapy, oncologic surgery, and breast reconstruction, besides supplementary tests .Initially, women with radiological alterations of the breast (BIRADS≥4) coming from the Basic Healthcare Unit (UBS) in Once integrated into the program, the patients followed the flow described below.At the first visit, all the patients were evaluated by a breast surgeonnece and a nurse, who determined the necessary tests and the subsequent. Subsequently, all the cases discussed at a multidisciplinary meeting between medical oncologists, breast surgeons, pathologist, radiologists, radiotherapists, nurses, dieticians, psychologists, nursing students, and oncology medical residents. This weekly multidisciplinary discussion occurred before treatment in all cases except those in which the chemotherapeutical treatment needed to be started urgently, even before the weekly meeting. The discussions of cases by the multidisciplinary team allowed the establishment of an institutionally standardized plan, with a optimal chance of success. Standard staging consisted of an abdominal ultrasound and chest X-ray. In specific cases of patients with clinical complaint, or for those with tumors >5cm, a clinically axillary involvement or after confirmation of surgery of more than four compromised lymph nodes, staging was complemented by computed tomography of the abdomen and chest , and bone scan.. All the patients underwent sentinel lymph node biopsy (SLN), with axillary lymph node dissection in case there was presence of macrometastases in the SLN. In patients who received neo-adjuvant therapy, the SLN biopsy was performed at the time ofsurgery. For patients submitted to mastectomy, the surgical plan included immediate reconstruction, with planning made together by the breast surgeon and a plastic surgeon. Selected cases  were submitted to magnetic resonance imaging of the breast for surgical planning.All the patients with surgical indication were operated on after the multidisciplinary discussion. Breast conserving surgery was recommended for all the initial cases  and for those who presented with a response to neo-adjuvant therapy, with a resulting lesion that allowed the removal of the tumor surrounded by a margin of healthy tissue. The rare cases in which there was a deviation from these guidelines were a result of multidisciplinary discussion, and the consequence of limitations imposed by comorbidities or by characteristics detected in the geriatric assessment of the patients.Systemic treatment followed the protocol recommended internationally by the National Comprehensive Cancer Network (NCCN). For women who received neo-adjuvant systemic treatment, the indication for radiotherapy was dictated by the original clinical staging rather thanthe pathological staging.Radiation therapy was recommended for all the women submitted to conservative surgery, for those with tumors larger than 5cm and/or with four or more positive lymph nodes in the axillaAll the patients underwent evaluation and were subsequently followed by psychologist, nutrition specialist, and physical therapy & rehabilitation treatmentas needed. Patients older than 60 years of age were evaluated preoperatively by a geriatrician, and had a geriatric follow up during treatment. In cases with a family history for cancer or when clinical evaluation raised the suspicion of a hereditary syndrome of predisposition to cancer, the patients were evaluated by the Genetic Counseling Service at HIAE. After the end of adjuvant chemotherapy and radiation therapy, all the patients were followed every three months or more frequently, depending on clinical necessity.Once the patient was enrolled in this programe until the end of treatment, the dates of tests and procedures carried out were tabulated in real-time, allowing the measurement of time intervals between the various phases of care. The team from the Hospital Cancer Registrar, responsible for collecting the data from all the oncologic patients treated at the hospital, tabulated part of the data; the remaining data were obtained retrospectively from patient charts, pharmacy records, and from the billing system of the hospital. All the monetary values are in reference to costs in US Dollars, paid by the PRPO for every procedure. Collection of clinical data was allowed after thefrom the Ethics in Research Committee of HIAE, filed under protocol # CAAE 11902312.7.0000.0071.Between August 2009 and December 2011, 96 patients were referred by the public health system for investigation of an abnormal mammogram. All 96 patients were biopsied and then referred with the biopsy result back to the original UBS. Of those 96, 51 were redirected for treatment by PRPO. Mastectomies and breast conserving surgeries determined hospitalization with a median duration of 2 days (varying from 1 to 7 days). There was no death or life-threatening adverse event related to the toxicity of the systemic treatment or resulting from complications of the surgical treatment.Throughout 2009 to 2011, 72 multidisciplinary meetings were held to discuss the cases. During the same period, there were 85 hospitalizations (including those for surgery) and 2,875 visits . The total ammount spent by PRPO between 2009 and 2011 for the diagnosis and treatment of these 51 patients was US$ 1,457,500.00. Besides the items described on Another documented parameter was the evaluation of the intervals between the various phases of diagnosis and treatment, as shown in . PRPO was set up including one arm dedicated to clinical assistance, in which 51 women with breast cancer were treated at HIAE. Since they were patients coming from a Basic Health Unit Service and from a SUS Specialties Outpatient Clinic, they are a small, but representative sample of breast cancer in our society and they provide a reliable insight of the cost of treatment.The establishment of auxiliary programs to SUS through PPP, including those that are mostly clinical, may assist not only in training of staff from the public system, but also may help diminish the number of patients with cancer who await treatment, who reported that patients with a clinical suspicion of breast cancer waited for a mean interval of 6.5 months for diagnostic confirmation in the public network. It is possible that many of these cases could have been diagnosed in a more agile manner, if there were a flow of scheduling in the private network (via PPP) for each case in which the biopsy could not be scheduled in the public network within one month. Of the patients in our study, approximately one third presented with locally advanced or metastatic disease at the time of diagnosis, similar to what was found by Trufelli et al., with the consequent negative implication on prognosis. Once the pathological diagnosis was obtained, the median interval to the first treatment in our study was considered excessive  after surgery was a lot longer. Despite the fact that there is no consensus as to the ideal time between termination of neo-adjuvant treatment and surgical treatment, we considered an interval longer than 30 days excessive  in an integrated manner among the various specialties. This is the principle that determines the establishment of High Complexity Centers and Units in Oncology (CACONs and UNACONs) in the country, but their reach still does not cover all the demand. Therefore, in structuring a future PPP, the most important intervals to be monitored are clinical suspicion-mammogram-biopsy; biopsy-surgery; pathology report-adjuvant treatment.Although the most important result of any PPP is the reduction in cancer mortality, the documentation of this result would require a large number of women treated in this program and would take several years to complete. The most immediate benefit that can be determined is the adoption of quality treatments in the shortest time possible. When compared to treatment at a public institution. Therefore, the PRPO was fully financed by HIAE, after an agreement with the Municipal Government Núcleo de Planejamento Estratégico da Secretaria Municipal de Saúde (NUPES) and with the Coordenadoria Regional Sul de Saúde.From an economic point of view, since it contains a significant component of clinical assistance, the project was not considered by the Ministry of Health as adequate to fulfill the criteria of the “SUS Institutional Development Support Project” , it is probable that the value spent in diagnosis and treatment of the patients from our PRPO are higher than what is spent in the public healthcare network. As an example, using this table as a reference, SUS pays the public hospitals the value of US$ 54.16 for the a pathology report with hormone receptors status evaluation; US$ 391 for a mastectomy with lymphadenectomy; US$ 179 for a breast conserving surgery with lymphadenectomy; and US$ 240 for radiation therapy. The discrepancy among those values and our costs reflect, at least in part, the sub-standard values paid by the Ministry of Health to the service providers.Even though no studies were found for comparison, using as reference the table of fees for procedures paid for by SUS to hospitals of the public sector. For a patient submitted to biopsy, breast conserving surgery, chemotherapy without trastuzumab (regime with taxane followed by anthracycline), radiation therapy, and 5 years of tamoxifen, our approximate cost reached US$ 25,500.00. It is possible to consider that this would be the cost of treatment  for about 50% of the women diagnosed with breast cancer in Brazil . If, on the other hand, adjuvant chemotherapy without the addition of a taxane (and without trastuzumab) were used, increasing by a few percentage points the risk of relapses (with an increase in metastatic cases and their consequent expenses), the median cost of treatment  would be US$ 16,000.00. The addition of trastuzumab to the neoadjuvant or adjuvant treatment (during one year) would raise the cost to a total of US$ 95,000.00. Recently, trastuzumab was incorporated for use at SUS only in adjuvant or neo-adjuvant setting of breast cancer, though the indication for use in metastatic disease may still be approved. Considering the values negotiated by the Ministry of Health, one year with medication for a 60-kg patient would be approximately US$ 45,500.00, compared to the US$ 66,250.00 in our study  . Within the context of budget destined to healthcare (US$ 45.5 billion in 2012) and more specifically to the treatment of cancer (US$ 1.2 billion in 2012), we observe how the treatment of breast cancer, in following international standards of excellence, could consume vast resources of this budget.The data from our study do not discriminate the costs of treatment of metastatic disease, due to the small number of cases and the immense variation in costs, which depend on the clinical scenario of each patient, even though the costs were considered regarding the total amount spent on the project. In the United States, the mean cost per month of treatment for one patient with metastatic breast cancer is US$ 9,788.00, varying according to the use of systemic or oral chemotherapy, reaching a total value of US$ 250,000.00 per patient with a median survival of 2 yearsOne limitation of our results is that there were expenses that could not be assessed, such as expenditures with human resources , since these professionals were not hired for exclusive dedication to the project . Nevertheless, the study maintains its relevance, since it provides pioneer information on the main costs related to diagnosis and treatment of breast cancer in Brazil., this would be done within the structure of SUS. This would require an extremely rigorous reference and counter-reference flow chart so that all the benefit achieved through timely surgical treatment and radiation therapy is not lost in the interval of returning to SUS for systemic treatment.Based on what was observed at PRPO, a possible economic strategy would be that of limiting assistance strictly to what is offered by SUS, even if provided by private hospitals. For example, not offering trastuzumab to patients with metastatic Her-2 positive disease, not offering Aprepitant even in cases of highly emetogenic chemotherapy, avoiding placement of central venous catheters, and not proceeding with genetic tests, among others. A second strategy would be that of offering assistance in partnership only during the more critical phases of treatment and those most deficient at SUS, such as access to rapid diagnosis (making available core needle biopsies and mammotomiesat centers of excellence), and surgery within 30 days of the biopsy if the time schedule at SUS is not possible, besides radiation therapy. In systemic treatment, proportionally more expensive and with a greater availability at SUSThe expenses described provide relevant information about the costs associated with the diagnosis and treatment of patients with breast cancer, pointing to the need for cost reduction strategies in the private network in order to allow future large-scale public-private partnerships in oncology. Furthermore, the analysis of the flow we worked with enables the institution of priorities for future assistance in breast cancer, suggesting that patients' journeys to and from the private partner and the original public service should be avoided or optimized along the process of diagnosis and treatment, so that there are no delays with potential deleterious effects in the prognosis.


Fifteen years after the complete sequencing of the human genome, our understanding of how that sequence information is packaged within the cell nucleus and the significance of that packaging to the proper spatial and temporal regulation of gene expression is still poorly understood. While the breakthroughs in understanding the first level of packaging, the nucleosome, occurred at the end of the twentieth century, higher levels of interphase chromatin packaging in cell nuclei have remained unverifiable. Researchers had to rely on a combination of microscopic methods to either look anonymously at chromatin by electron or light microscopy or examine the spatial arrangement of a few specific loci by fluorescence in situ hybridization . MappingGenome Biology on the three dimensional organization of the nucleus highlights several new developments in 3D genome structure. A good overview of the field is provided in a high level review by Britta Bouwman and Wouter de Laat of the discoveries that have been made at various scales of analysis of chromatin interactions, from loops to topologically-associating domains, and sub-nuclear compartments [et al., systematically compare Hi-C results using different ligation modes [This special issue of artments . A revieartments . Ferhat artments . Also onon modes .et al. [et al. introduce a method to identify enhancer-promoter interactions by capturing the sequences from Hi-C libraries that are specific to promoters (HiCap) [Essential to the three dimensional organization of chromatin are the machineries that process the genetic information: replication, recombination, repair, transcription and RNA processing machineries. Understanding how the 3D organization of chromatin influences, and is influenced by, these machineries is likely central to our understanding of how cells access and interpret genetic information. In particular, with the wealth of new transcriptome and epigenomic data, an important goal has been to determine which putative functional elements interact with which outputs. For example, linking promoters to their regulatory enhancers is central to understanding transcriptional regulation and to link GWAS SNPs found in regulatory elements to the affected genes , 17. In et al.  introduc (HiCap) , focusinXist, the lncRNA mediating inactivation [et al. explore the bipartite structure of the inactivated X chromosome [et al. investigate the time course of inactivation of specific genes during differentiation [In female mammals, one of the two X chromosomes is almost completely inactivated to balance the double dose of the genes on this chromosome as compared to males. As part of this silencing process, the inactive X takes on a very unusual compact structure that is explored in a series of papers. Philip Avner and colleagues provide a review of what is known about tivation . Deng etromosome , and Marntiation . Togetheet al. report on estrogen-induced changes in chromatin architecture [et al. show how BORIS cooperates with its paralog CTCF to shape gene expression and chromatin architecture in cancer cells and germ cells [et al. introduce new high-throughput microscopy approach to detect chromosomal translocations resulting from aberrant repair of DNA breaks [The issue also contains a series of articles highlighting other aspects of the three dimensional ‘nucleome’. A pair of research articles illustrates how these three dimensional concepts are put into a functional context to regulate transcription. Rafique itecture , and Pugrm cells . UnderstA breaks . FinallyA breaks .Genome Biology is excited to publish this special issue now, and looks forward to publishing related articles in the future that build on the platform provided by these and other studies to refine our knowledge of the organization of the nucleus and its impact on genome function.It has long been known that the simple linear model of genes on a chromosome, activated by upstream promoters, is not the complete picture of gene control. The recent explosion of 3C and Hi-C data, and 3D FISH, is beginning to show us the complexity of how genes, regulatory elements and chromosomes interact. As ever, with a new technology, the excitement of the new results is tempered by the realization of its limitations and how little we still know.