Suelene Brito do Nascimento Tavares

Comparison of the performance of rapid prescreening, 10% random review, and clinical risk criteria as methods of internal quality control in cervical cytopathology†

Rapid prescreening (RPS) is an internal quality‐control (IQC) method that is used both to reduce errors in the laboratory and to measure the sensitivity of routine screening (RS). Little direct comparison data are available comparing RPS with other more widely used IQC methods.

Deep Sequencing of the Trypanosoma cruzi GP63 Surface Proteases Reveals Diversity and Diversifying Selection among Chronic and Congenital Chagas Disease Patients

Background Chagas disease results from infection with the diploid protozoan parasite Trypanosoma cruzi. T. cruzi is highly genetically diverse, and multiclonal infections in individual hosts are common, but little studied. In this study, we explore T. cruzi infection multiclonality in the context of age, sex and clinical profile among a cohort of chronic patients, as well as paired congenital cases from Cochabamba, Bolivia and Goias, Brazil using amplicon deep sequencing technology. Methodology/ Principal Findings A 450bp fragment of the trypomastigote TcGP63I surface protease gene was amplified and sequenced across 70 chronic and 22 congenital cases on the Illumina MiSeq platform. In addition, a second, mitochondrial target—ND5—was sequenced across the same cohort of cases. Several million reads were generated, and sequencing read depths were normalized within patient cohorts (Goias chronic, n = 43, Goias congenital n = 2, Bolivia chronic, n = 27; Bolivia congenital, n = 20), Among chronic cases, analyses of variance indicated no clear correlation between intra-host sequence diversity and age, sex or symptoms, while principal coordinate analyses showed no clustering by symptoms between patients. Between congenital pairs, we found evidence for the transmission of multiple sequence types from mother to infant, as well as widespread instances of novel genotypes in infants. Finally, non-synonymous to synonymous (dn:ds) nucleotide substitution ratios among sequences of TcGP63Ia and TcGP63Ib subfamilies within each cohort provided powerful evidence of strong diversifying selection at this locus. Conclusions/Significance Our results shed light on the diversity of parasite DTUs within each patient, as well as the extent to which parasite strains pass between mother and foetus in congenital cases. Although we were unable to find any evidence that parasite diversity accumulates with age in our study cohorts, putative diversifying selection within members of the TcGP63I gene family suggests a link between genetic diversity within this gene family and survival in the mammalian host.

Congenital transmission of Trypanosoma cruzi in central Brazil. A study of 1,211 individuals born to infected mothers

Transmission of Trypanosoma cruzi during pregnancy is estimated to occur in less than 20% of infected mothers; however, the etiopathogenesis is not completely understood. The Centre for Studies on Chagas Disease provides confirmation of T. cruzi infection for individuals living in central Brazil. In this retrospective hospital-based study, all requests for diagnosis of T. cruzi infection in individuals less than 21 years old from 1994-2014 were searched. We end with 1,211 individuals and their respective infected mothers. Congenital transmission of infection was confirmed in 24 individuals (2%) in central Brazil, an area where the main T. cruzi lineage circulating in humans is TcII. This low prevalence of congenital Chagas disease is discussed in relation to recent findings in the south region of Brazil, where TcV is the main lineage and congenital transmission has a higher prevalence (approximately 5%), similar to frequencies reported in Argentina, Paraguay and Bolivia. This is the first report to show geographical differences in the rates of congenital transmission of T. cruzi and the relationship between the prevalence of congenital transmission and the type of Tc prevalent in each region.

Improvement in the Routine Screening of Cervical Smears A Study Using Rapid Prescreening and 100% Rapid Review as Internal Quality Control Methods

High rates of false‐negative results constitute a routine problem in cytology laboratories. Of currently available internal quality control methods, 10% random review is the least effective in detecting false‐negatives in routine screening. There is evidence that 100% rapid review and rapid prescreening perform well for this purpose. This study compared the performance of rapid prescreening and 100% rapid review as internal quality control methods for cervical cytology exams.

The Trypanosoma cruzi Satellite DNA OligoC-TesT and Trypanosoma cruzi Kinetoplast DNA OligoC-TesT for Diagnosis of Chagas Disease: A Multi-cohort Comparative Evaluation Study

Background The Trypanosoma cruzi satellite DNA (satDNA) OligoC-TesT is a standardised PCR format for diagnosis of Chagas disease. The sensitivity of the test is lower for discrete typing unit (DTU) TcI than for TcII-VI and the test has not been evaluated in chronic Chagas disease patients. Methodology/Principal Findings We developed a new prototype of the OligoC-TesT based on kinetoplast DNA (kDNA) detection. We evaluated the satDNA and kDNA OligoC-TesTs in a multi-cohort study with 187 chronic Chagas patients and 88 healthy endemic controls recruited in Argentina, Chile and Spain and 26 diseased non-endemic controls from D.R. Congo and Sudan. All specimens were tested in duplicate. The overall specificity in the controls was 99.1% (95% CI 95.2%–99.8%) for the satDNA OligoC-TesT and 97.4% (95% CI 92.6%–99.1%) for the kDNA OligoC-TesT. The overall sensitivity in the patients was 67.9% (95% CI 60.9%–74.2%) for the satDNA OligoC-TesT and 79.1% (95% CI 72.8%–84.4%) for the kDNA OligoC-Test. Conclusions/Significance Specificities of the two T. cruzi OligoC-TesT prototypes are high on non-endemic and endemic controls. Sensitivities are moderate but significantly (p = 0.0004) higher for the kDNA OligoC-TesT compared to the satDNA OligoC-TesT.

Internal quality control for cervical cytopathology: comparison of potential false-negatives detected at rapid prescreening and at 100% rapid review.

Objective: To evaluate the performance of rapid prescreening (RPS) and 100% rapid review (RR-100%) as internal quality control (IQC) methods assessed by outcome at colposcopy, histopathology and repeat cytopathology for cases with false-negative results on cervical cytopathology at routine screening (RS). Study Design: Out of 12,208 cytology smears analyzed, 900 were abnormal. Of these, 656 were identified at RS, and 244 were false-negative, with 90.2% identified at RPS and 57.4% at RR-100%. Of the 900 abnormal cases, 436 were submitted for additional testing. Results: Of the 244 women with cytopathological abnormalities identified only by the IQC methods, 114 had supplementary examinations: 35 were submitted for colposcopy, 22 for biopsy and 99 for repeat cytopathology. The sensitivity of RPS for the detection of abnormalities identified on colposcopy, histopathology and repeat cytopathology was 87.5% (95% CI 67.6-97.3), 82.4% (95% CI 56.6-96.2) and 95.7% (95% CI 85.2-99.5), respectively. The sensitivity of RR-100% was 54.2% (95% CI 32.8-74.4), 52.9% (95% CI 27.8-77.0) and 47.8% (95% CI 32.9-63.1), respectively. RPS was more sensitive than RR-100% when compared to the findings on colposcopy (p = 0.011) and repeat cytopathology (p = 0.000). When compared to colposcopy, histopathology and repeat cytopathology, the sensitivity of RS was 83.2% (95% CI 76.1-88.9), 85.7% (95% CI 78.1-91.5) and 73.3% (95% CI 66.0-79.7), respectively. Conclusion: RPS performed better than RR-100% when compared to the results of colposcopy and repeat cytopathology.

Mulheres com atipias, lesões precursoras e invasivas do colo do útero: condutas segundo as recomendações do Ministério da Saúde

PURPOSE: To verify whether women with atypias of undetermined significance and precursor lesions or invasive cervical outcomes were referred to Medium Complexity Units (MCU) following the guidelines recommended by the Brazilian Ministry of Health. METHODS: Retrospective study based on the cytopathological outcomes of users of the Unified Health System, seen at Basic Health Assistance Units (BHAU) and referred to MCUs in the municipality of Goiânia, state of Goias, from 2005 to 2006. We assessed 832 records according to the recommendations of the Brazilian Ministry of Health, as established by the Brazilian Nomenclature for Cervical Cytopathologic Outcomes and Recommended Clinical Practice. To check the distribution of variables such as reasons for referral, results of colposcopy and histopathology and clinical procedures we calculated absolute and relative frequencies, mean, minimum and maximum values. RESULTS: We understood 72.7% of the referrals were not in accordance with the recommendations of the Ministry of Health. There were 605 women with test results classified as atypical squamous cells of undetermined significance, possibly non-neoplasms, and squamous intraepithelial lesion of low level which were sent to MCU, and of these 71.8% were submitted to colposcopy, and 64.7% had histopathological examination which results were classified as 31.0% with non-neoplasms and 44.6% as NIC I. Out of 211 women with results classified as more severe squamous lesions, 86.3% were submitted to colposcopy and 68.7% of these had histopathological examinations. CONCLUSIONS: The results of this study revealed high rates of inappropriate referrals to MCU, which required a high percentage of unnecessary procedures. The recommendations of the Ministry of Health were followed by BHAU and the majority of women received counseling/treatment as recommended.PURPOSE To verify whether women with atypias of undetermined significance and precursor lesions or invasive cervical outcomes were referred to Medium Complexity Units (MCU) following the guidelines recommended by the Brazilian Ministry of Health. METHODS Retrospective study based on the cytopathological outcomes of users of the Unified Health System, seen at Basic Health Assistance Units (BHAU) and referred to MCUs in the municipality of Goiânia, state of Goiás, from 2005 to 2006. We assessed 832 records according to the recommendations of the Brazilian Ministry of Health, as established by the Brazilian Nomenclature for Cervical Cytopathologic Outcomes and Recommended Clinical Practice. To check the distribution of variables such as reasons for referral, results of colposcopy and histopathology and clinical procedures we calculated absolute and relative frequencies, mean, minimum and maximum values. RESULTS We understood 72.7% of the referrals were not in accordance with the recommendations of the Ministry of Health. There were 605 women with test results classified as atypical squamous cells of undetermined significance, possibly non-neoplasms, and squamous intraepithelial lesion of low level which were sent to MCU, and of these 71.8% were submitted to colposcopy, and 64.7% had histopathological examination which results were classified as 31.0% with non-neoplasms and 44.6% as NIC I. Out of 211 women with results classified as more severe squamous lesions, 86.3% were submitted to colposcopy and 68.7% of these had histopathological examinations. CONCLUSIONS The results of this study revealed high rates of inappropriate referrals to MCU, which required a high percentage of unnecessary procedures. The recommendations of the Ministry of Health were followed by BHAU and the majority of women received counseling/treatment as recommended.

Usefulness of repeat cytology at the time of first colposcopy

The value of repeat cytology in the detection of high‐grade preinvasive or invasive disease was analyzed at the time of colposcopy in cases of patients with abnormal referral cytology. A series of 136 patients with squamous abnormalities in their referral cytology, who had undergone repeat cytology and for whom outcome had been established, were included in the study. Final diagnosis consisted of 27 negative results (20%), 37 cases of CIN1 (27%), 31 CIN2 (23%), 39 CIN3 (29%), and 2 (1%) cases of squamous cell carcinoma (SCC). Referral and repeat cytology suggestive of atypical squamous cells cannot exclude high‐grade squamous intraepithelial lesion (ASC‐H), high‐grade squamous intraepithelial lesion (HSIL) were findings significantly associated with a final diagnosis of CIN2 or worse. Sensitivity and positive predictive value of repeat cytology for the histological diagnosis of significant histological neoplasia (CIN2 or worse) were 81.9% and 77.6%, respectively. In women with referral cytology findings of atypical squamous cells of undetermined significance or low‐grade squamous intraepithelial lesion (ASC‐US/LSIL), repeat cytology was suggestive of HSIL in 20% of cases (11/54), and in 4% (2/54) biopsy was negative or CIN1, and the final diagnosis established by LLETZ was CIN3. Eight cases (10%) of HSIL in referral and repeat cytology in which biopsies were negative or insufficient were diagnosed as CIN3 or worse in the final diagnosis. Repeat cytology would have changed therapeutic management in 4 and 10%, respectively, of patients included as ASC‐US/LSIL and ASC‐H/HSIL in referral cytology. Diagn. Cytopathol. 2009.

Reproducibility of cervical cytopathology following an intervention by an external quality control laboratory

This study assessed the effects of a continued education program on the agreement between cervical cytopathology exams interpreted by local laboratories and interpretation made by an external quality control laboratory (LabMEQ).

Internal quality control indicators of cervical cytopathology exams performed in laboratories monitored by the External Quality Control Laboratory

PURPOSE To evaluate the impact of continued education provided by an external quality control laboratory on the indicators of internal quality control of cytopathology exams. METHODS The internal quality assurance indicators for cytopathology exams from 12 laboratories monitored by the External Quality Control Laboratory were evaluated. Overall, 185,194 exams were included, 98,133 of which referred to the period preceding implementation of a continued education program, while 87,061 referred to the period following this intervention. Data were obtained from the Cervical Cancer Database of the Brazilian National Health Service. RESULTS Following implementation of the continued education program, the positivity index (PI) remained within recommended limits in four laboratories. In another four laboratories, the PI progressed from below the limits to within the recommended standards. In one laboratory, the PI remained low, in two laboratories, it remained very low, and in one, it increased from very low to low. The percentage of exams compatible with a high-grade squamous intraepithelial lesion (HSIL) remained within the recommended limits in five laboratories, while in three laboratories it progressed from below the recommended levels to >0.4% of the total number of satisfactory exams, and in four laboratories it remained below the standard limit. Both the percentage of atypical squamous cells of undetermined significance (ASC-US) in relation to abnormal exams, and the ratio between ASC-US and intraepithelial lesions remained within recommended levels in all the laboratories investigated. CONCLUSION An improvement was found in the indicators represented by the positivity index and the percentage of exams compatible with a high-grade squamous intraepithelial lesion, showing that the role played by the external quality control laboratory in providing continued education contributed towards improving laboratory staff skills in detecting cervical cancer precursor lesions.