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Publications (176)
Febrile illnesses contribute significantly to morbidity and mortality in sub-Saharan Africa, but the lack of diagnostic facilities and the broad spectrum of pathogens can lead to inadequate clinical management. The timely and reliable identification of the causative pathogens in febrile patients is the basis for the administration of optimal treatm...
The clinical presentation of COVID-19 and the specific antibody responses associated with SARS-CoV-2 variants have not been investigated during the emergence of Omicron variants in Bangladesh. The Delta and Omicron variants were identified by post-PCR melting curve analysis of the spike (S) protein receptor binding domain amplicons. Anti-S-protein...
Objectives:
Human and animal studies suggest an important role of autophagy in the pathogenesis of pancreatitis. ATG16L1 (autophagy-related 16 like 1) is part of a protein complex that is involved in the formation of autophagosomes. The c.898A > G (p.T300A) variant of ATG16L1 is associated with Crohn disease. In this study, we analyzed ATG16L1 c.8...
Novel SARS-CoV-2 variants are emerging at an alarming rate. The delta variant and other variants of concern (VoC) carry spike (S)-protein mutations, which have the potential to evade protective immunity, to trigger breakthrough infections after COVID-19 vaccination, and to propagate future waves of COVID-19 pandemic. To identify SARS CoV-2 variants...
Intense transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Africa might promote emergence of variants. We describe 10 SARS-CoV-2 lineages in Benin during early 2021 that harbored mutations associated with variants of concern. Benin-derived SARS-CoV-2 strains were more efficiently neutralized by antibodies derived from v...
In March 2020, the SARS-CoV-2 virus outbreak was declared as a world pandemic by the World Health Organization (WHO). The only measures for controlling the outbreak are testing and isolation of infected cases. Molecular real-time polymerase chain reaction (PCR) assays are very sensitive but require highly equipped laboratories and well-trained pers...
Currently, testing for coronavirus is performed with time and personnel consuming PCR assays. The aim of this study was to evaluate the sensitivity, specificity and capacity of a fully automated, random access high-throughput real-time PCR-based diagnostic platform for the detection of SARS-CoV-2. The NeuMoDx N96 system displayed an equal or better...
Background
The ongoing outbreak of the recently emerged novel coronavirus (2019-nCoV) poses a challenge for public health laboratories as virus isolates are unavailable while there is growing evidence that the outbreak is more widespread than initially thought, and international spread through travellers does already occur.AimWe aimed to develop an...
The analysis of circulating cell-free DNA (cfDNA) extracted from peripheral blood can serve as a minimally invasive alternative to tumor tissue biopsies in cases with impaired access to tissue. Its clinical utility has been well demonstrated for EGFR T790M testing in lung cancer patients suffering progress after tyrosine kinase inhibitor treatment....
Additional information about yellow fever virus surveillance, Brazil, 2017.
The current yellow fever outbreak in Brazil prompted widespread yellow fever virus (YFV) vaccination campaigns, imposing a responsibility to distinguish between vaccine- and wild-type YFV-associated disease. We developed novel multiplex real-time reverse transcription PCRs that differentiate between vaccine and American wild-type YFV. We validated...
Objective
Alcohol-related pancreatitis is associated with a disproportionately large number of hospitalisations among GI disorders. Despite its clinical importance, genetic susceptibility to alcoholic chronic pancreatitis (CP) is poorly characterised. To identify risk genes for alcoholic CP and to evaluate their relevance in non-alcoholic CP, we pe...
Background: Currently the detection of Zika virus (ZIKV) in patient samples is done by real-time RT-PCR. Samples collected from rural area are sent to highly equipped laboratories for screening. A rapid point-of-care test is needed to detect the virus, especially at low resource settings.
Methodology/Principal Findings: In this report, we describe...
Background
Currently the detection of Zika virus (ZIKV) in patient samples is done by real-time RT-PCR. Samples collected from rural area are sent to highly equipped laboratories for screening. A rapid point-of-care test is needed to detect the virus, especially at low resource settings.
Methodology/Principal Findings
In this report, we describe t...
Background. In spring 2014, a sudden rise in the number of notified MERS-Coronavirus infections occurred across Saudi Arabia with a focus in Jeddah. Hypotheses to explain the outbreak pattern include increased surveillance, increased zoonotic transmission, nosocomial transmission, changes in viral transmissibility, as well as diagnostic laboratory...
Activating epidermal growth factor receptor (EGFR) gene mutations can be successfully treated by EGFR tyrosine kinase inhibitors (EGFR-TKIs), but nearly 50% of all patients' exhibit progression of the disease until treatment because of T790M mutations. It is proposed that this is mostly caused by therapy-resistant tumor clones harboring a T790M mut...
Chronic pancreatitis is an inflammatory disorder of the pancreas. We analyzed CPA1, encoding carboxypeptidase A1, in subjects with nonalcoholic chronic pancreatitis (cases) and controls in a German discovery set and three replication sets. Functionally impaired variants were present in 29/944 (3.1%) German cases and 5/3,938 (0.1%) controls (odds ra...
In response to a recent outbreak in China, detection assays for a novel avian influenza A(H7N9) virus need to be implemented in a large number of public health laboratories. Here we present real-time reverse-transcription polymerase chain reaction (RT-PCR) assays for specific detection of this virus, along with clinical validation data and biologic...
Background
The genus Flavivirus includes several pathogenic agents that cause severe illness in humans. Re-emergence of West Nile virus in Europe and continuous spread of certain flaviviruses such as dengue, yellow fever and Japanese encephalitis viruses represent a global danger to public health. Therefore, a rapid and accurate molecular method is...
The acute disease antigen A (adaA) gene is believed to be associated with Coxiella burnetii strains causing acute Q fever. The detailed analysis of the adaA genomic region of 23 human- and 86 animal-derived C. burnetii isolates presented in this study reveals a much more polymorphic appearance and distribution of the adaA gene, resulting in a class...
We present a rigorously validated and highly sensitive confirmatory real-time RT-PCR assay (1A assay) that can be used in combination with the previously reported upE assay. Two additional RT-PCR assays for sequencing are described, targeting the RdRp gene (RdRpSeq assay) and N gene (NSeq assay), where an insertion/deletion polymorphism might exist...
We present a rigorously validated and highly sensitive confirmatory real-time RT-PCR assay (1A assay) that can be used in combination with the previously reported upE assay. Two additional RT-PCR assays for sequencing are described, targeting the RdRp gene (RdRpSeq assay) and N gene (NSeq assay), where an insertion/deletion polymorphism might exist...
COLD-PCR: A powerful tool in routine-diagnostic for cost-neutral detection of minor clones using real-time PCR or pyrosequencing quoted by the example of EGFR mutation analysis
We present two real-time reverse-transcription polymerase chain reaction assays for a novel human coronavirus (CoV), targeting regions upstream of the E gene (upE) or within open reading frame (ORF)1b, respectively. Sensitivity for upE is 3.4 copies per reaction (95% confidence interval (CI): 2.5–6.9 copies) or 291 copies/mL of sample. No cross-rea...
It is a matter of debate whether single nucleotide polymorphisms (SNP) in the promoter region of interleukin (IL)-13, an IgE regulator, and IL-18, an inducer of immune responses, modulating the respective protein expression, are accompanied by an increased risk of atopy, allergic asthma, and total IgE levels. The suspected associations were noted i...
Objective In chronic pancreatitis (CP), alterations in several genes have so far been described, but only small cohorts have been extensively investigated for all predisposing genes.
Design 660 patients with idiopathic or hereditary CP and up to 1758 controls were enrolled. PRSS1, SPINK1 and CTRC were analysed by DNA sequencing, and cystic fibrosis...
Introduction: Common variants on human chromosome 8q24 were found to be associated with prostate cancer risk with different frequency and incidence among the investigated populations. We examined the effect of smoke on this type of cancer and its relationship with the risk variant rs6983267, located at region 3 of chromosome 8q24, in a prostate can...
Recently, several polymorphisms located on human chromosome 8q24 were found to be associated with prostate cancer risk with different frequency and incidence among the investigated populations. The authors conducted a prostate cancer case-control study in the Greek population to evaluate the association of the single nucleotide polymorphism (SNP) r...
CFTR mutations enhance susceptibility for idiopathic chronic pancreatitis (ICP) and congenital bilateral absence of the vas deferens (CBAVD); however, it is unknown why CFTR heterozygotes are at increased disease risk. We recently showed that common CFTR variants are associated with aberrantly spliced transcripts. Here, we genotyped for common CFTR...
We evaluated the performance of a laboratory-developed multiplex real-time reverse transcription-PCR assay (LDT rRT-PCR), the Centers for Disease Control and Prevention (CDC) 2009 H1N1 rRT-PCR protocol using the LightCycler 480 II, the multiplex xTAG Respiratory Virus Panel (xTAG RVP), and rapid immunodiagnostic testing (RIDT) using the BinaxNOW In...
A real-time PCR assay based on TaqMan probe chemistry was developed for the detection of Theileria parva DNA in blood samples. It uses a Theileria genus-specific PCR primer set and a T. parva-specific probe to amplify and hybridize with a species-specific part of the 18S rRNA gene of the parasite. The test was evaluated using positive and negative...
Nitric oxide (NO), produced by endothelial nitric oxide synthase (eNOS), plays a key role in the regulation of vascular tone. Endothelium-derived NO exerts vasoprotective effects by suppressing platelet aggregation, leukocyte adhesion and smooth muscle cell proliferation. The E298D polymorphic variant of eNOS has been associated with myocardial inf...
Clinically, both viral load and genotypes have been found to be major predictors of antiviral therapy outcome regarding chronic hepatitis C and they are, under normal circumstances, performed as separate assays.
In order to improve the diagnostic strategy and subsequently reduce the reagent costs we have developed and established the simultaneous q...
A single-tube multiprobe real-time PCR assay for simultaneous detection of Entamoeba histolytica and Entamoeba dispar was developed. One primer pair with 2 species-specific probes was designed based on new SSU RNA regions of the ribosomal DNA-containing episome. The sensitivity is 1 parasite per milliliter of feces and thus superior to the conventi...
Toll-like Receptor 8 (TLR8) plays an important role in the innate immune defense against various pathogens. The TLR8 genetic variation affects the course of infections with agents such as HIV, HCV, and mycobacteria. To assess the influence of single nucleotide polymorphisms (SNPs) large cohorts need to be studied; thus, rapid and cost-efficient pro...
Nitric oxide (NO), produced by endothelial nitric oxide synthase (eNOS), plays a key role in the regulation of vascular tone. The E298D polymorphic variant of eNOS has been associated with myocardial infarction (MI), but data relating to this variant are divergent in Greece. Accordingly, we examined a possible association between the E298D polymorp...
Recently, common variants on human chromosome 8q24 were found to be associated with prostate cancer risk. While conducting an association study at the Greek population to investigate the frequency and the susceptibility of one SNP, rs6983267, located at the region 3 of chromosome 8q24, to the prostate cancer, we found highly significant correlation...
KRAS mutation testing before anti-epidermal growth factor receptor therapy of metastatic colorectal cancer has become mandatory in Europe. However, considerable uncertainty exists as to which methods for detection can be applied in a reproducible and economically sound manner in the routine diagnostic setting. To answer this question, we examined 2...
Premature activation of pancreatic digestive enzymes is considered as a major factor in the pathogenesis of pancreatitis. Genetic alterations of different pancreatic zymogens or their inhibitors have been associated with chronic pancreatitis (CP).
We sequenced all 12 GP2 exons in 380 German CP patients and in 182 German control subjects. In additio...
IntroductionChip Design/Array DescriptionProtocol for a LCD Array ExperimentResults and DiscussionConclusions
Influenza A(H1N1)v virus was first identified in April 2009. A novel real-time RT-PCR for influenza A(H1N1)v virus was set up ad hoc and validated following industry-standard criteria. The lower limit of detection of the assay was 384 copies of viral RNA per ml of viral transport medium (95% confidence interval: 273-876 RNA copies/ml). Specificity...
A preformulated chikungunya virus real-time reverse transcription-PCR, quality-confirmed oligonucleotides, and noninfectious virus controls were distributed by the European Network for the Diagnosis of Imported Viral Diseases. An international proficiency study with 31 participants demonstrated that ad hoc implementation of molecular diagnostics wa...
5th International Conference for Rickettsiae and Rickettsial Diseases, in Marseille (France) on 18–20 May 2008, organized by the European Study Group for Coxiella, Anaplasma, Rickettsia and Bartonella (ESCAR) and by the American Society for Rickettsiology
The aim of this study was to determine the performance of real-time amplification based methods - NASBA, TaqMan, RT-FRET, and RT-PCR LUXtrade mark formats - for the detection of influenza A (H5N1) virus RNA. In an analysis of 54 samples obtained from a range of animal species in Thailand during the period 2003-2006, results showed that the NASBA (H...
Non-tuberculous mycobacteria (NTM) are emerging pathogens in immunocompromised patients. Therefore, it is important for hospitals to consider tap water as a source for these infections. Aim of the study was to establish highly sensitive and specific techniques to detect and identify NTM in hospital drinking water. A Mycobacterium genus-specific 16S...
Intercross studies in inbred mice susceptible or resistant to liver fibrosis revealed complement factor 5 as a quantitative trait gene associated with the development of fibrosis. In 277 patients with hepatitis C, two C5 SNPs, rs17611 and rs2300929, have been associated with advanced fibrosis.
We investigated the association of these C5 SNPs with a...
Acute pancreatitis (AP) is a disease whose pathogenesis remains largely obscure. Genetic research has focussed attention upon the role of the pancreatic protease/protease inhibitor system. The aim of this study was to investigate the prevalence of genetic variants of the trypsin inhibitor, SPINK1, in acute pancreatitis.
We genotyped 468 patients wi...
Current HIV-1 viral-load assays are too expensive and time-consuming for small sample quantity or resource-limited setting. In addition, some commercial assays have shown shortcomings in quantifying rare genotypes. We developed an internally controlled, two-step, reverse transcription-initiated real-time PCR protocol on the LightCycler instrument a...
Polymorphisms in cytokine genes can influence immune responses and inflammation and thereby affecting the outcome of hematopoietic stem-cell transplantation. We analyzed a single-nucleotide polymorphism in the gene for the interleukin-23 receptor (IL-23R) (1142G>A) in a cohort of 221 transplant recipients and their human leukocyte antigen (HLA)-ide...
A recent study reported that the c.30T>A (p.Cys10Ter; rs2043211) variant, in the CARD8 (TUCAN) gene, is associated with Crohn's disease (CD). The aim of this study was to analyze the frequency of p.C10X in 3 independent European (IBD) cohorts from Germany, Hungary, and the Netherlands.
We included a European IBD cohort of 921 patients and compared...
The Cys 2-Cys 10 disulfide bond in ribonuclease T1 was broken by substituting Cys 2 and Cys 10 by Ser and Asn, respectively, as present in ribonuclease F1. This C2S/C10N variant resembles the wild-type protein in structure and in catalytic activity. Minor structural changes were observed by 2-dimensional NMR in the local environment of the substitu...
Human 8-oxoguanine DNA glycosylase 1 (hOGG1) plays an important role in the repair of 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodGuo), one of the major constituents in DNA damage. A recent in vitro study showed that the hOGG1 326Cys polymorphism (rs1052133) exhibits reduced 8-oxodGuo repair activity. This study aimed to develop a LightCycler (LC)...
SNP309 T/G (rs2279744) causes higher levels of MDM2, the most important negative regulator of the p53 tumor suppressor. SNP72 G/C (rs1042522) gives rise to a p53 protein with a greatly reduced capacity to induce apoptosis. Both polymorphisms have been implicated in cancer. The SNP309 G-allele has recently been reported to accelerate diffuse large B...
Chronic pancreatitis is a persistent inflammatory disease of the pancreas, in which the digestive protease trypsin has a fundamental pathogenetic role. Here we have analyzed the gene encoding the trypsin-degrading enzyme chymotrypsin C (CTRC) in German subjects with idiopathic or hereditary chronic pancreatitis. Two alterations in this gene, p.R254...
UDP-glucuronosyltransferases (UGT) catalyze the glucuronidation of various compounds and thus inactivate toxic substrates. Genetic variations reducing the activity of UGT1A7 have been associated with various gastrointestinal cancers. Most recently, the UGT1A7*3 allele has been reported as a significant risk factor for pancreatic disorders, but we c...
To determine whether the -308 A/G tumor necrosis factor alpha (TNFalpha) gene polymorphism can predict the outcome of etanercept therapy in 86 patients with rheumatoid arthritis (RA), as already observed in patients treated with infliximab.
Eighty-six RA patients treated with etanercept were genotyped for -308 A/G TNFalpha gene polymorphism by poly...
A recent study reported that a nonsynonymous SNP rs2241880 (c.898A>G, p.Thr300Ala) within ATG16L1 confers susceptibility to Crohn's disease (CD). We analyzed ATG16L1 c.898A>G in three independent European inflammatory bowel disease (IBD) cohorts from Germany, Hungary and the Netherlands.
In total, we included 910 European IBD patients and compared...
A recent study reported that a non-synonymous single nucleotide polymorphism (rs11209026, p.Arg381Gln) located in the IL23R gene is a protective marker for inflammatory bowel disease.
To analyse the frequency of p.Arg381Gln in three independent European inflammatory bowel disease cohorts and to evaluate how this variant influences disease behaviour...
The aim of the study was to examine diagnostic possibilities of LNA (locked nucleic acids) -clamped PCR for detection of somatic mutations in the K-ras gene in 133 samples of human malignant colorectal tumors. Selective real-time PCR amplification of mutant alleles of the K-ras gene (codon 12 and 13) revealed the presence of mutations in 45 samples...
The melanocortin-4 receptor (MC4R) gene pathogenic mutations are the most prevalent forms of monogenic obesity, responsible for ∼2% of obesity cases, but its
role in common obesity is still elusive. We analyzed the contribution of non-synonymous mutations V103I (rs2229616, c.307G
> A) and I251L (no rs, c.751A > C) to obesity in 16 797 individuals o...
Tularemia is a plague-like infection caused by Francisella (F.) tularensis classified as a biological warfare agent. F. tularensis subsp. tularensis is the most virulent subspecies demanding rapid diagnosis. Typing systems for this fastidious bacterium to the subspecies level are laborious and time consuming. Therefore, the aim of this study was to...
There are more than a dozen formats available for the fluorescent detection of amplified DNA in kinetic (real-time) PCR. These chemistries are adaptable to most real-time PCR instruments and may offer benefits over the usual manufacturer-recommended chemistries for the instrument. The most popular chemistries are the generic dye, SYBR Green I, TaqM...
A single amino acid substitution, from histidine to tyrosine at position 274 of the neuraminidase gene has converted Oseltamivir sensitive H5N1 influenza A virus into a resistant strain. Currently, Oseltamivir is being stockpiled in many countries potentially affected by the influenza A virus subtype H5N1 epidemic. To identify this change in Oselta...