Dimitrios Frangoulidis - Academia.edu (original) (raw)

Papers by Dimitrios Frangoulidis

Vaccine, 2007

The human anthrax vaccines currently licensed contain the protective antigen (PA) of Bacillus ant... more The human anthrax vaccines currently licensed contain the protective antigen (PA) of Bacillus anthracis as main antigen together with traces of some other bacillus components, e.g. lethal factor (LF). The present study aimed at monitoring the course of specific antibody titres against PA and LF by enzyme linked immunosorbent assays (ELISA), as well as the levels of toxin-neutralising antibodies, in 11 volunteers vaccinated with the human anthrax vaccine UK. After an initial seroconversion in all vaccinees, a significant reduction of both antibody titres against PA and LF, and of neutralising antibodies, was detected just prior to a vaccine boost 6 months after completion of the basic immunisation. Following the booster injection, titres increased again to levels comparable to those after the fourth immunisation. ELISA titres against PA correlated significantly with neutralising antibodies (r=0.816, p&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001). Therefore, the less work- and time-consuming ELISA should be favoured to monitor the efficacy of an anthrax vaccination.

Transactions of the Royal Society of Tropical Medicine and Hygiene, 2008

PLoS ONE, 2014

The case rate of Q fever in Europe has increased dramatically in recent years, mainly because of ... more The case rate of Q fever in Europe has increased dramatically in recent years, mainly because of an epidemic in the Netherlands in 2009. Consequently, there is a need for more extensive genetic characterization of the disease agent Coxiella burnetii in order to better understand the epidemiology and spread of this disease. Genome reference data are essential for this purpose, but only thirteen genome sequences are currently available. Current methods for typing C. burnetii are criticized for having problems in comparing results across laboratories, require the use of genomic control DNA, and/or rely on markers in highly variable regions. We developed in this work a method for single nucleotide polymorphism (SNP) typing of C. burnetii isolates and tissue samples based on new assays targeting ten phylogenetically stable synonymous canonical SNPs (canSNPs). These canSNPs represent previously known phylogenetic branches and were here identified from sequence comparisons of twenty-one C. burnetii genomes, eight of which were sequenced in this work. Importantly, synthetic control templates were developed, to make the method useful to laboratories lacking genomic control DNA. An analysis of twentyone C. burnetii genomes confirmed that the species exhibits high sequence identity. Most of its SNPs (7,493/7,559 shared by .1 genome) follow a clonal inheritance pattern and are therefore stable phylogenetic typing markers. The assays were validated using twenty-six genetically diverse C. burnetii isolates and three tissue samples from small ruminants infected during the epidemic in the Netherlands. Each sample was assigned to a clade. Synthetic controls (vector and PCR amplified) gave identical results compared to the corresponding genomic controls and are viable alternatives to genomic DNA. The results from the described method indicate that it could be useful for cheap and rapid disease source tracking at nonspecialized laboratories, which requires accurate genotyping, assay accessibility and inter-laboratory comparisons. Citation: Karlsson E, Macellaro A, Byströ m M, Forsman M, Frangoulidis D, et al. (2014) Eight New Genomes and Synthetic Controls Increase the Accessibility of Rapid Melt-MAMA SNP Typing of Coxiella burnetii. PLoS ONE 9(1): e85417.

PLoS ONE, 2013

The acute disease antigen A (adaA) gene is believed to be associated with Coxiella burnetii strai... more 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 classification of C. burnetii strains of better differentiation than previously anticipated. Three different genomic variants of the adaA gene were identified which could be detected in isolates from acute and chronic patients, rendering the association of adaA positive strains with acute Q fever disease disputable. In addition, all adaA positive strains in humans and animals showed the occurrence of the QpH1 plasmid. All adaA positive isolates of acute human patients except one showed a distinct SNP variation at position 431, also predominant in sheep strains, which correlates well with the observation that sheep are a major source of human infection. Furthermore, the phylogenetic analysis of the adaA gene revealed three deletion events and supported the hypothesis that strain Dugway 5J108-111 might be the ancestor of all known C. burnetii strains. Based on our findings, we could confirm the QpDV group and we were able to define a new genotypic cluster. The adaA gene polymorphisms shown here improve molecular typing of Q fever, and give new insights into microevolutionary adaption processes in C. burnetii. Citation: Frangoulidis D, Splettstoesser WD, Landt O, Dehnhardt J, Henning K, et al. (2013) Microevolution of the Chromosomal Region of Acute Disease Antigen A (adaA) in the Query (Q) Fever Agent Coxiella burnetii. PLoS ONE 8(1): e53440.

Molecular and Cellular Probes, 2008

Specific identification of Bacillus anthracis and differentiation from closely related Bacillus c... more Specific identification of Bacillus anthracis and differentiation from closely related Bacillus cereus and Bacillus thuringiensis strains is still a major diagnostic problem. Commercially available diagnostic kits targeting plasmid-markers cannot differentiate between B. anthracis, non-anthracis Bacillus species harbouring anthrax-specific virulence plasmids, and plasmidless B. anthracis strains. A TaqMan PCR assay was designed targeting sequences of gene locus BA_5345 of the B. anthracis strain Ames. Specificity was determined by using a panel of 328 Bacillus strains; sensitivity was determined by probit analysis. All B. anthracis isolates (n=92) were specifically detected by using the genomic TaqMan PCR assay whereas 236 strains belonging to 19 Bacillus species other than B. anthracis were PCR negative. The detection limit was determined to be 12.7 copies per reaction (95% confidence interval 10.2-17.5 copies). Here we present an extensively evaluated and - to our current knowledge - specific TaqMan PCR assay for the detection of B. anthracis based on a chromosomal marker.

FEMS Immunology & Medical Microbiology, 2003

The objective of the present study was to establish a system of real-time polymerase chain reacti... more The objective of the present study was to establish a system of real-time polymerase chain reactions (PCRs) for the specific detection of Yersinia pestis using the LightCycler1 (LC) instrument. Twenty-five strains of Y. pestis, 94 strains of other Yersinia species and 33 clinically relevant bacteria were investigated. Assays for the 16S rRNA gene target and the plasminogen activator gene (resides on the 9.5kb plasmid) and for the Y. pestis murine toxin gene and the fraction 1 antigen gene (both on the 100-kb plasmid) were combined for the use in two multiplex assays including an internal amplification control detecting bacteriophage V-DNA. Applying these multiplex assays, Y. pestis was selectively identified; other bacteria yielded no amplification products. The lower limit of detection was approximately 0.1 genome equivalent. Rat or flea DNA had no inhibitory effects on the detection of Y. pestis. The results obtained using the multiplex realtime assays showed 100% accuracy when compared with combinations of conventional PCR assays. We developed and evaluated a highly specific real-time PCR strategy for the detection of Y. pestis, obtaining results within 3 h including DNA preparation.

European Journal of Clinical Microbiology & Infectious Diseases, 2011

17 A collection of 40 Bacillus (B.) anthracis strains mostly isolated from soil in Bulgaria betwe... more 17 A collection of 40 Bacillus (B.) anthracis strains mostly isolated from soil in Bulgaria between 1960 18 and 1980 were investigated. All strains were proven to be B. anthracis by culture and amplification of 19 a B. anthracis-specific chromosomal marker. PCR demonstrated that in 9 strains both virulence 20 plasmids (pX01+/pX02+) and in 4 strains only one plasmid (pX02+) were present, whereas the 21 majority of strains (n=27) lacked both plasmids (pX01-/pX02-). Multi-Locus-Variable Number of 22 60 61 62 63 64 65

Epidemiology and Infection, 2009

Tularemia is a rare, notifiable zoonosis in Germany. Since November 2004, several lines of eviden... more Tularemia is a rare, notifiable zoonosis in Germany. Since November 2004, several lines of evidence including outbreaks in humans or animals and confirmed infections in indigenous hare and rodent populations have indicated a re-emergence of tularemia in different German federal states. Unfortunately, reliable basic information on the seroprevalence in different geographical regions, permitting the identification of risk factors, does not exist. Combining a sensitive screening assay with a highly specific confirmative immunoblot test, we performed a serological investigation on 2416 sera from a population-based, cross-sectional health survey of the city population of Leutkirch, Baden-Wuerttemberg. A total of 56 sera gave positive results indicating a seroprevalence of 2 . 32%. Thus, the seroprevalence is tenfold higher than that previously reported in a nationwide study in 2004. Francisella tularensis can cause a wide variety of clinical syndromes including severe, sometimes fatal disease. Missing epidemiological data on its spatial and temporal distribution in an endemic country complicate an appropriate risk assessment necessary for public health authorities to be prepared for an adequate outbreak management. This is of special concern regarding the extraordinary potential of F. tularensis as an agent of bioterrorism. Our investigation performed in a presumed low-risk area demonstrated that tularemia might be seriously underestimated in Germany and probably in other central European countries as well.

Epidemiology and Infection, 2013

Q fever is a notifiable disease in Germany. The majority of the reported cases are related to out... more Q fever is a notifiable disease in Germany. The majority of the reported cases are related to outbreaks. The objective of our study was to evaluate the general role of Q fever in community-acquired pneumonia (CAP). We investigated respiratory samples and sera from 255 patients with CAP, who were enrolled into a CAPNETZ cohort in summer 2005. Altogether, our data showed a significant prevalence of Q fever as CAP (3·5%). If a patient's condition leads to a diagnostic test for Chlamydophila sp., Mycoplasma sp. or Legionella sp., then a Q fever diagnostic test should also be included. In particular, ELISA as a first diagnostic step is easy to perform. PCR should be performed at an early stage of the disease if no antibodies are detectable. Because of our highly promising findings we suggest performing PCR in respiratory samples.

Emerging Infectious Diseases, 2014

Clinical Microbiology and Infection, 2009

BMC Microbiology, 2008

Background: Coxiella burnetii is the causative agent of Q-fever, a widespread zoonosis. Due to it... more Background: Coxiella burnetii is the causative agent of Q-fever, a widespread zoonosis. Due to its high environmental stability and infectivity it is regarded as a category B biological weapon agent. In domestic animals infection remains either asymptomatic or presents as infertility or abortion. Clinical presentation in humans can range from mild flu-like illness to acute pneumonia and hepatitis. Endocarditis represents the most common form of chronic Q-fever. In humans serology is the gold standard for diagnosis but is inadequate for early case detection. In order to serve as a diagnostic tool in an eventual biological weapon attack or in local epidemics we developed a realtime 5'nuclease based PCR assay with an internal control system. To facilitate high-throughput an automated extraction procedure was evaluated.

Annals of the New York Academy of Sciences, 2006

Four commercially available serological assays for the detection of IgM phase II antibodies in pa... more Four commercially available serological assays for the detection of IgM phase II antibodies in patients with acute Q fever infection were compared using a panel of 23 serum samples from patients with acute Q fever and 88 control sera from blood donors.

Standards in genomic sciences, 2014

We present the whole genome sequence and annotation of the Coxiella burnetii strain Namibia. This... more We present the whole genome sequence and annotation of the Coxiella burnetii strain Namibia. This strain was isolated from an aborting goat in 1991 in Windhoek, Namibia. The plasmid type QpRS was confirmed in our work. Further genomic typing placed the strain into a unique genomic group. The genome sequence is 2,101,438 bp long and contains 1,979 protein-coding and 51 RNA genes, including one rRNA operon. To overcome the poor yield from cell culture systems, an additional DNA enrichment with whole genome amplification (WGA) methods was applied. We describe a bioinformatics pipeline for improved genome assembly including several filters with a special focus on WGA characteristics.