QQ-SNV: single nucleotide variant detection at low frequency by comparing the quality quantiles

Background: Next generation sequencing enables studying heterogeneous populations of viral infections. When the sequencing is done at high coverage depth ("deep sequencing"), low frequency variants can be detected. Here we present QQ-SNV (http://sourceforge.net/projects/qqsnv), a logistic regression classifier model developed for the Illumina sequencing platforms that uses the quantiles of the quality scores, to distinguish true single nucleotide variants from sequencing errors based on the estimated SNV probability. To train the model, we created a dataset of an in silico mixture of five HIV-1 plasmids. Testing of our method in comparison to the existing methods LoFreq, ShoRAH, and V-Phaser 2 was performed on two HIV and four HCV plasmid mixture datasets and one influenza H1N1 clinical dataset. Results: For default application of QQ-SNV, variants were called using a SNV probability cutoff of 0.5 (QQ-SNVD). To improve the sensitivity we used a SNV probability cutoff of 0.0001 (QQ-SNVHS). To also increase specificity, SNVs called were overruled when their frequency was below the 80th percentile calculated on the distribution of error frequencies (QQ-SNVHS-P80). When comparing QQ-SNV versus the other methods on the plasmid mixture test sets, QQ-SNVD performed similarly to the existing approaches. QQ-SNVHS was more sensitive on all test sets but with more false positives. QQ-SNVHS-P80 was found to be the most accurate method over all test sets by balancing sensitivity and specificity. When applied to a paired-end HCV sequencing study, with lowest spiked-in true frequency of 0.5 %, QQ-SNVHS-P80 revealed a sensitivity of 100 % (vs. 40-60 % for the existing methods) and a specificity of 100 % (vs. 98.0-99.7 % for the existing methods). In addition, QQ-SNV required the least overall computation time to process the test sets. Finally, when testing on a clinical sample, four putative true variants with frequency below 0.5 % were consistently detected by QQ-SNVHS-P80 from different generations of Illumina sequencers. Conclusions: We developed and successfully evaluated a novel method, called QQ-SNV, for highly efficient single nucleotide variant calling on Illumina deep sequencing virology data

Molecular characterization of Salmonella Enteritidis : comparison of an optimized multi-locus variable-number of tandem repeat analysis (MLVA) and pulsed-field gel electrophoresis

Salmonella Enteritidis (SE) is a genetically homogenous serovar, which makes optimal subtype discrimination crucial for epidemiological research. This study describes the development and evaluation of an optimized multiple-locus variable number tandem-repeat assay (MLVA) for characterization of SE. The typeability and discriminatory power of this MLVA was determined on a selected collection of 60 SE isolates and compared with pulsed-field gel electrophoresis (PFGE) using restriction enzymes XbaI, NotI, or SfiI. In addition, the estimated Wallace coefficient (W) was calculated to assess the congruence of the typing methods. Selection of epidemiologically unrelated isolates and more related isolates (originating from layer farms) was also based on the given phage type (PT). When targeting six loci, MLVA generated 16 profiles, while PFGE produced 10, 9, and 16 pulsotypes using XbaI, NotI, and SfiI, respectively, for the entire strain collection. For the epidemiologically unrelated isolates, MLVA had the highest discriminatory power and showed good discrimination between isolates from different layer farms and among isolates from the same layer farm. MLVA performed together with PT showed higher discriminatory power compared to PFGE using one restriction enzyme together with PT. Results showed that combining PT with the optimized MLVA presented here provides a rapid typing tool with good discriminatory power for characterizing SE isolates of various origins and isolates originating from the same layer farm

Nasopharyngeal S. pneumoniae carriage and density in Belgian infants after 9 years of pneumococcal conjugate vaccine programme

Background: In Belgium, the infant pneumococcal conjugate vaccine (PCV) programme changed from PCV7 (2007-2011) to PCV13 (2011-2015) and to PCV10 (2015-2016). A 3-year nasopharyngeal carriage study was initiated during the programme switch in 2016. Main objective of the year 1 assessment was to obtain a baseline measurement of pneumococcal carriage prevalence, carriage density, serotype distribution and antibiotic resistance. Materials/methods: Two infant populations aged 6-30 months and without use of antibiotics in the seven days prior to sampling were approached: (1) attending one of 85 randomly selected day-care centres (DCC); (2) presenting with AOM at study-trained general practitioners and paediatricians. Demographic and clinical characteristics were documented and a single nasopharyngeal swab was taken. S. pneumoniae were cultured, screened for antibiotic resistance and serotyped, and quantitative Taqman real-time PCR (qRT-PCR) targeting LytA was performed. Results: Culture-based (DCC: 462/760; 60.8%- AOM: 27/39; 69.2%) and LytA-based (DCC: 603/753; 80.1% - AOM: 32/39; 82.1%) carriage prevalence was high. Average pneumococcal DNA load in LytA-positive day-care samples was 6.5 x 10(6) copies/mu l (95%Cl = 3.9-9.2 x 10(6), median = 3.5 x 10(5)); DNA load was positively associated with signs of common cold and negatively with previous antibiotic use. Culture based frequency of 13 pneumococcal vaccine (PCV) serotypes was 5.4% in DCC and 7.7% in AOM, with 19F and 14 being most frequent, and frequencies below 0.5% for serotypes 3, 6A, 19A in both populations. Predominant non-PCV serotypes were 23B and 23A in day-care and 11A in infants with AOM. In day-care, resistance to penicillin was rare (<0.5%) and absent against levofloxacin; 32.7% and 16.9% isolates were cotrimoxazole- and erythromycin-resistant respectively. Conclusion: Four years after PCV13 introduction in the vaccination programme, PCV13 serotype carriage was rare in infants throughout Belgium and penicillin resistance was rare. Continued surveillance in the context of a PCV programme switch is necessary

Een Romeinse waterput met een vlechtwerkmand uit de vicus van Oudenburg

Bij saneringswerken in het centrum van Oudenburg, net ten westen van het castellum, kwam begin 2010 een Romeinse houten waterput aan het licht. De inhoud bleek gekenmerkt door uitstekend bewaard organisch materiaal en was ook qua culturele vondsten opmerkelijk. Vooral de grotendeels bewaarde vlechtwerkmand in de vulling van de waterput maakte de vondst zeer bijzonder. Het voor onze contreien unieke karakter van deze vondst gaf aanleiding tot dit artikel. Door middel van natuurwetenschappelijk onderzoek en de studie van de gebruiksvoorwerpen uit de waterput werd geprobeerd om zoveel mogelijk ecologische en culturele informatie te verkrijgen. De afmetingen en de vlechtwerktechniek van de mand suggereren een functie als wan, waarmee het kaf van het koren werd gescheiden. De vlechtwerkmand, waarvan de bodem mogelijk opzettelijk was afgescheurd of afgesneden, en met een complete Noord-Menapische beker in haar vulling, is naar alle waarschijnlijkheid een rituele depositie na opgave van de waterput. Dit idee wordt zelfs nog meer versterkt vanwege de betekenis en de rol die de wan bekleedde in mysteriecultussen. Zowel literaire als iconografische bronnen verwijzen naar de rituele sfeer waarin de wan figureerde en die in relatie staat tot agrarische vruchtbaarheid. Deze waterput is een nieuw puzzelstukje voor de vicus van Oudenburg. De archeologische waarnemingen van de laatste decennia geven een idee van de maximale omvang van deze nederzetting tijdens de 3de eeuw. De waterput bevindt zich vrij centraal in de nederzetting die zich vanuit het westen naar het oosten lijkt te hebben uitgebreid. In het gebied ten westen van het castellum kon ook nooit eerder natuurwetenschappelijk onderzoek worden uitgevoerd, wat het belang van de detailstudie van de vulling van deze waterput nog onderstreept

M & L Jaargang 2/5

RedactioneelMarcel Smets en Koen Verbruggen De werderopbouw van Dendermonde als constructie van een artificieel decor. [The post-war reconstruction of Dendermonde: erecting an artificial decor.]Herman De Laet De doorwerking van de verfraaiingsgedachte tijdens de wederopbouw: het plan Viérin voor Nieuwpoort (1906-1920). [The cortinuity of the Embellishment-idea during post-war reconstruction: the Plan Viérin for Nieuwpoort (1906-1920).]Ronny De Meyer Antwerpen, Lier en Mechelen: een debat over stedebouw en architectuur naar aanleiding van drie wederopbouwwedstrijden. [Antwerp, Lier and Mechelen: a debate concerning town planning and architecture on the occasion of three reconstruction contests.]Herman Stynen Architect Richard Acke (1873-1934) en de woningbouw in de frontstreek. [Architect Richard Acke (1873-1934) and house-building in the front area.]Pieter Uyttenhove Van stadsbouwkunst tot stedebouwkunde: een modernistische oefening van Raphaël Verwilghen. [From town architecture to town planning: a modernistic exercise by Raphaël Verwilghen.]SummaryM&L Binnenkran

A qPCR assay to detect and quantify Shiga toxin-producing E. coli (STEC) in cattle and on farms : a potential predictive tool for STEC culture-positive farms

Shiga toxin-producing E. coli (STEC), of various serogroups harboring the intimin gene, form a serious threat to human health. They are asymptomatically carried by cattle. In this study, a quantitative real-time PCR (qPCR) method was developed as a molecular method to detect and quantify Shiga toxin genes stx1 and stx2 and the intimin gene eae. Subsequently, 59 fecal samples from six farms were tested using qPCR and a culture method as a reference. Three farms had contaminated animals as demonstrated by the culture method. Culture-positive farms showed moderate significantly higher stx prevalences than culture-negative farms (p = 0.05). This is the first study which showed preliminary results that qPCR can predict STEC farm contamination, with a specificity of 77% and a sensitivity of 83%, as compared with the culture method. Furthermore, the presence or quantity of stx genes in feces was not correlated to the isolation of STEC from the individual animal. Quantitative data thus did not add value to the results. Finally, the detection of both stx and eae genes within the same fecal sample or farm using qPCR was not correlated with the isolation of an eae-harboring STEC strain from the respective sample or farm using the culture method

Activity limitations predict health care expenditures in the general population in Belgium.

BACKGROUND: Disability and chronic conditions both have an impact on health expenditures and although they are conceptually related, they present different dimensions of ill-health. Recent concepts of disability combine a biological understanding of impairment with the social dimension of activity limitation and resulted in the development of the Global Activity Limitation Indicator (GALI). This paper reports on the predictive value of the GALI on health care expenditures in relation to the presence of chronic conditions. METHODS: Data from the Belgian Health Interview Survey 2008 were linked with data from the compulsory national health insurance (n = 7,286). The effect of activity limitation on health care expenditures was assessed via cost ratios from multivariate linear regression models. To study the factors contributing to the difference in health expenditure between persons with and without activity limitations, the Blinder-Oaxaca decomposition method was used. RESULTS: Activity limitations are a strong determinant of health care expenditures. People with severe activity limitations (5.1%) accounted for 16.9% of the total health expenditure, whereas those without activity limitations (79.0%), were responsible for 51.5% of the total health expenditure. These observed differences in health care expenditures can to some extent be explained by chronic conditions, but activity limitations also contribute substantially to higher health care expenditures in the absence of chronic conditions (cost ratio 2.46; 95% CI 1.74-3.48 for moderate and 4.45; 95% CI 2.47-8.02 for severe activity limitations). The association between activity limitation and health care expenditures is stronger for reimbursed health care costs than for out-of-pocket payments. CONCLUSION: In the absence of chronic conditions, activity limitations appear to be an important determinant of health care expenditures. To make projections on health care expenditures, routine data on activity limitations are essential and complementary to data on chronic conditions