Ten years of external quality assessment (EQA) of Neisseria gonorrhoeae antimicrobial susceptibility testing in Europe elucidate high reliability of data.

BACKGROUND: Confidence in any diagnostic and antimicrobial susceptibility testing data is provided by appropriate and regular quality assurance (QA) procedures. In Europe, the European Gonococcal Antimicrobial Susceptibility Programme (Euro-GASP) has been monitoring the antimicrobial susceptibility in Neisseria gonorrhoeae since 2004. Euro-GASP includes an external quality assessment (EQA) scheme as an essential component for a quality-assured laboratory-based surveillance programme. Participation in the EQA scheme enables any problems with the performed antimicrobial susceptibility testing to be identified and addressed, feeds into the curricula of laboratory training organised by the Euro-GASP network, and assesses the capacity of individual laboratories to detect emerging new, rare and increasing antimicrobial resistance phenotypes. Participant performance in the Euro-GASP EQA scheme over a 10 year period (2007 to 2016, no EQA in 2013) was evaluated. METHODS: Antimicrobial susceptibility category and MIC results from the first 5 years (2007-2011) of the Euro-GASP EQA were compared with the latter 5 years (2012-2016). These time periods were selected to assess the impact of the 2012 European Union case definitions for the reporting of antimicrobial susceptibility. RESULTS: Antimicrobial susceptibility category agreement in each year was ≥91%. Discrepancies in susceptibility categories were generally because the MICs for EQA panel isolates were on or very close to the susceptibility or resistance breakpoints. A high proportion of isolates tested over the 10 years were within one (≥90%) or two (≥97%) MIC log2 dilutions of the modal MIC, respectively. The most common method used was Etest on GC agar base. There was a shift to using breakpoints published by the European Committee on Antimicrobial Susceptibility Testing (EUCAST) in the latter 5 years, however overall impact on the validity of results was limited, as the percentage categorical agreement and MIC concordance changed very little between the two five-year periods. CONCLUSIONS: The high level of comparability of results in this EQA scheme indicates that high quality data are produced by the Euro-GASP participants and gives confidence in susceptibility and resistance data generated by laboratories performing decentralised testing.

Emergence of the lymphogranuloma venereum L2c genovariant, Hungary, 2012 to 2016.

In eastern Europe, few countries have so far reported laboratory-confirmed cases of lymphogranuloma venereum (LGV). Here we describe 22 LGV cases in men who have sex with men (MSM) detected in Hungary from November 2012 to July 2016. Sequence analyses show that 16 of these 22 cases were affected by the L2c genovariant, with from 2012 to 2014, one LGV L2c case detected per year, followed by seven cases in 2015 and six up to July 2016. Of the 16 total L2c LGV cases, 10 had severe haemorrhagic proctitis. These findings are concerning as cases with this new genovariant among MSM have not been frequently reported in Europe to date. More research is needed to assess the spread of the L2c genovariant and its potential association with virulence and severe clinical manifestation.

[Lymphogranuloma venereum: the first Hungarian cases]. / Lymphogranuloma venereum: elso hazai esetek.

Lymphogranuloma venereum is a sexually transmitted infection caused by the Chlamydia trachomatis serovars L1-3. It has been found to be endemic in tropical countries. In the last decades several cases have been reported in Western Europe, particularly in men who have sex with men population infected with human immunodeficiency virus. The authors present three cases of lymphogranuloma venereum infections, observed at their department in 2013 and 2014. The three human immunodeficiency virus infected patients who belonged to men who have sex with men population had casual sexual contacts in Western Europe. The symptoms included urethral discharge, discomfort and inguinal lymphadenomegaly in two patients, and rectal pain, discharge and perianal ulceration in one patient. The diagnosis was confirmed by nucleic acid amplification test performed in samples obtained from urethral discharge and exudate of perianal ulcer; lymphogranuloma venereum 2b serovars were demonstrated in two patients and serovar 2 in one patient. Doxycyclin (daily dose of two times 100 mg for 21 days) resolved the symptoms in all cases. The authors conclude that lymphogranuloma venereum is a diagnostic challenge in Hungary, too. It is important to be aware of the altered clinical features of this disease to prevent complications and spreading.

Genotyping of Coxiella burnetii from domestic ruminants and human in Hungary: indication of various genotypes.

BACKGROUND: Information about the genotypic characteristic of Coxiella burnetii from Hungary is lacking. The aim of this study is to describe the genetic diversity of C. burnetii in Hungary and compare genotypes with those found elsewhere. A total of 12 samples: (cattle, n = 6, sheep, n = 5 and human, n = 1) collected from across Hungary were studied by a 10-loci multispacer sequence typing (MST) and 6-loci multiple-locus variable-number of tandem repeat analysis (MLVA). Phylogenetic relationships among MST genotypes show how these Hungarian samples are related to others collected around the world. RESULTS: Three MST genotypes were identified: sequence type (ST) 20 has also been identified in ruminants from other European countries and the USA, ST28 was previously identified in Kazakhstan, and the proposed ST37 is novel. All MST genotypes yielded different MLVA genotypes and three different MLVA genotypes were identified within ST20 samples alone. Two novel MLVA types 0-9-5-5-6-2 (AG) and 0-8-4-5-6-2 (AF) (Ms23-Ms24-Ms27-Ms28-Ms33-Ms34) were defined in the ovine materials correlated with ST28 and ST37. Samples from different parts of the phylogenetic tree were associated with different hosts, suggesting host-specific adaptations. CONCLUSIONS: Even with the limited number of samples analysed, this study revealed high genetic diversity among C. burnetii in Hungary. Understanding the background genetic diversity will be essential in identifying and controlling outbreaks.

Industrial Dairy Cattle Farms in Hungary Source of Coxiella burnetii Infection in Humans.

Previous research has demonstrated a high prevalence of Coxiella burnetii in the bulk tank milk on large industrial dairy farms of the central and eastern European region. The aim of this survey was to estimate the prevalence of specific IgG antibodies to C. burnetii proving previous infection among dairy farm workers and to determine the possible risk factors. Serum samples from veterinarians, inseminators, animal caretakers, milking parlor workers, and herd managers working on dairy farms were tested for the presence of IgG to phase I and phase II of C. burnetii using an indirect microimmunofluorescence assay. Antibodies phase II to C. burnetii were detected in 59 out of 70 individuals tested (84.3%). All occupational groups are highly exposed to C. burnetii infection. Veterinarians, inseminators, and animal caretakers had 100% seropositivity rate of phase II, whereas the seropositivity rate found among herd managers and milking parlor workers was 71.4% and 47%, respectively. The findings of this survey suggest that the risk of C. burnetii infection is correlated with cattle density in the large dairy farms and also with occupational groups.

Ongoing evolution of Chlamydia trachomatis lymphogranuloma venereum: exploring the genomic diversity of circulating strains.

Lymphogranuloma venereum (LGV), the invasive infection of the sexually transmissible infection (STI) Chlamydia trachomatis, is caused by strains from the LGV biovar, most commonly represented by ompA-genotypes L2b and L2. We investigated the diversity in LGV samples across an international collection over seven years using typing and genome sequencing. LGV-positive samples (n=321) from eight countries collected between 2011 and 2017 (Spain n=97, Netherlands n=67, Switzerland n=64, Australia n=53, Sweden n=37, Hungary n=31, Czechia n=30, Slovenia n=10) were genotyped for pmpH and ompA variants. All were found to contain the 9 bp insertion in the pmpH gene, previously associated with ompA-genotype L2b. However, analysis of the ompA gene shows ompA-genotype L2b (n=83), ompA-genotype L2 (n=180) and several variants of these (n=52; 12 variant types), as well as other/mixed ompA-genotypes (n=6). To elucidate the genomic diversity, whole genome sequencing (WGS) was performed from selected samples using SureSelect target enrichment, resulting in 42 genomes, covering a diversity of ompA-genotypes and representing most of the countries sampled. A phylogeny of these data clearly shows that these ompA-genotypes derive from an ompA-genotype L2b ancestor, carrying up to eight SNPs per isolate. SNPs within ompA are overrepresented among genomic changes in these samples, each of which results in an amino acid change in the variable domains of OmpA (major outer membrane protein, MOMP). A reversion to ompA-genotype L2 with the L2b genomic backbone is commonly seen. The wide diversity of ompA-genotypes found in these recent LGV samples indicates that this gene is under immunological selection. Our results suggest that the ompA-genotype L2b genomic backbone is the dominant strain circulating and evolving particularly in men who have sex with men (MSM) populations.

Global phylogeny of Treponema pallidum lineages reveals recent expansion and spread of contemporary syphilis.

Syphilis, which is caused by the sexually transmitted bacterium Treponema pallidum subsp. pallidum, has an estimated 6.3 million cases worldwide per annum. In the past ten years, the incidence of syphilis has increased by more than 150% in some high-income countries, but the evolution and epidemiology of the epidemic are poorly understood. To characterize the global population structure of T. pallidum, we assembled a geographically and temporally diverse collection of 726 genomes from 626 clinical and 100 laboratory samples collected in 23 countries. We applied phylogenetic analyses and clustering, and found that the global syphilis population comprises just two deeply branching lineages, Nichols and SS14. Both lineages are currently circulating in 12 of the 23 countries sampled. We subdivided T. p. pallidum into 17 distinct sublineages to provide further phylodynamic resolution. Importantly, two Nichols sublineages have expanded clonally across 9 countries contemporaneously with SS14. Moreover, pairwise genome analyses revealed examples of isolates collected within the last 20 years from 14 different countries that had genetically identical core genomes, which might indicate frequent exchange through international transmission. It is striking that most samples collected before 1983 are phylogenetically distinct from more recently isolated sublineages. Using Bayesian temporal analysis, we detected a population bottleneck occurring during the late 1990s, followed by rapid population expansion in the 2000s that was driven by the dominant T. pallidum sublineages circulating today. This expansion may be linked to changing epidemiology, immune evasion or fitness under antimicrobial selection pressure, since many of the contemporary syphilis lineages we have characterized are resistant to macrolides.

[Ornithosis--case report and current concerns]. / Ornithosis--aktualitások egy eset kapcsán.

The authors describe a severe case of psittacosis requiring intensive care, and summarize the potential means of diagnosis, as well as the epidemiologic and therapeutic aspects of the disease. Clinical signs of ornithosis do not allow a reliable differentiation from other atypical bacterial infections, thus, exploring the possible exposure to birds in the patient's history is most important in these cases. Knowledge of bird exposure in the history leads the clinician to the correct diagnosis that may result quick recovery due to the adequate therapy. This notifiable disease may presumably belong to the misdiagnosed or delayed diagnosed infections even today so the underrepresented case-reports do not necessarily reflect the actual epidemiological situation in Hungary. The aim of the authors was to call the attention to this sometimes fatal disease occurring not as rarely as supposed.

Genotyping of Chlamydia trachomatis from the endocervical specimens of high-risk women in Hungary.

The distribution of different Chlamydia trachomatis serovars in Hungary has not been reported previously. The objective of this study was to determine the distribution and prevalence of C. trachomatis serovars in a high-risk population by genotyping. The endocervical specimens of 484 female sex workers (FSWs) were screened for C. trachomatis by plasmid PCR. Genotyping was performed in all C. trachomatis-positive samples by PCR-based RFLP analysis of the omp1 gene. A total of 32 specimens (6.6 %) were positive for C. trachomatis. Age was an important risk factor for C. trachomatis infection in FSWs. The highest prevalence was detected in women under the age of 20 (18.8 %). All positive specimens were successfully genotyped and seven serovars were identified. The most prevalent was serovar D (34.4 %), followed by E (21.9 %), F (18.8 %), G (9.4 %), J (9.4 %), H (3.1 %) and I (3.1 %). A heterogeneous distribution of C. trachomatis serovars was observed in the study group, where the most common serovars were D, E and F comprising 75 % of the positive samples. This PCR-based RFLP method could be used in epidemiological studies on the prevalence of C. trachomatis infection to provide more information and to compare the serovar distribution among different cohorts.

[Chlamydia trachomatis infections in neonates--overview of current laboratory diagnostics]. / Ujszülöttkori Chlamydia trachomatis-fertôzések--az aktuális laboratóriumi diagnosztikai módszerek áttekintése.

Based on recent literature and their own laboratory experiences the author summarizes the clinical manifestations, diagnostic possibilities and the tools of prevention of neonatal Chlamydia trachomatis infection, where a potential or a verified role of this pathogen is present. Definitive diagnosis is complicated by the common phenomenon of asymptomatic, undiagnosed maternal infection, as well as by the need for adequate sample-collection and targetted screening techniques. There are only estimated data regarding the incidence of neonatal Chlamydia trachomatis infection; however we do have the diagnostic tests to identify the affected neonates, who may have generally only a mild-moderate manifestation of infection in case of an early treatment. Focussed screening efforts should be made to reduce the number of infected pregnant women and thereby the vertical rate of transmission.

Two fatal cases of psittacosis caused by Chlamydophila psittaci.

Two fatal cases of psittacosis are described in two poultry-processing-plant employees presenting with pneumonia and respiratory failure. Diagnosis was confirmed by serological and PCR methods. Psittacosis due to Chlamydophila psittaci infection usually has a good recovery rate, although diagnostic delay and mistreatment can lead to severe complications and even death. This report emphasizes the need for rapid differential diagnosis and management of suspected cases of atypical pneumonia to prevent fatal outcomes.

Features of syphilis seropositive pregnant women raising alarms in Hungary, 2013-2016.

OBJECTIVES: The incidence of syphilis has been on the rise in Hungary over the last decades. We aimed to assess the syphilis seroprevalence in pregnant women during 2013-2016. The secondary aims were to describe seropositivity by age and gestational age, to estimate infectivity rates in different age groups, and to compare the efficacy of mandatory prenatal screening versus individual venereological testing in revealing syphilis. STUDY DESIGN: During the above mentioned period the reactive serum samples (N = 527) of 49,965 pregnant women undergoing routine screening were submitted for syphilis verification to the Bacterial STI Reference Laboratory, National Center for Epidemiology, Budapest, Hungary. The confirmation process included titrated RPR and TPHA tests performed simultaneously. The tested women were considered seronegative if both tests gave negative results. When any of these tests proved indeterminate or positive, anti-Treponema pallidum IgG and IgM ELISA tests were performed. Patients confirmed for the presence for specific IgG were judged seropositive. Further evaluation of potential infectivity of seropositive patients was carried out on RPR reactivity. RESULTS: Syphilis seropositivity was detected in overall 2.9‰ (N = 148) of the cases. RPR-negative cases, i.e. past infections were confirmed in 36% (53/148); weak-reactive RPR (titres≤8) cases, i. e. past/early acute infections in 37% (55/148); strong-reactive RPR (titres>8) cases suggesting recent syphilis in 27% (40/148). Half of the infectious syphilis cases (20/40) belonged to the 15-24 age group. The gestational age at screening was available of 123 seropositive women, out of whom 27 (22%) were diagnosed late, in the third trimester. Nineteen (13%) out of all seropositives were detected via individual venereological testing before/instead of general prenatal screening. CONCLUSIONS: The majority of infected pregnants may remain undetected due to the lack of mandatory general prenatal screening. The seropositivity for syphilis in pregnancy of 2.9/1000, of which one quarter were recent and infectious, the late diagnosis of syphilis in one in five, and the low pick-up rate of individualised instead of generalized screening are alarming signals and call for more effective prevention strategies, focusing on the most vulnerable adolescents, as well as on the first trimester of pregnancy.

Seroprevalence of anti-Chlamydia trachomatis IgM in neonatal respiratory tract infections in Hungary.

Purpose. To determine the seroprevalence of specific IgM indicative of respiratory tract infection (RTI) due to Chlamydia trachomatis (CT) among symptomatic infants.Methodology. A descriptive study was conducted on young infants up to 5 months old at the Bacterial Sexually Transmitted Infections Reference Laboratory, National Centre for Epidemiology, Budapest, covering the period 2008-2016. Serum samples from infants suffering from RTIs were screened with a micro-immunofluorescence test (Focus, Cypress, USA) for the presence of anti-Chlamydia trachomatis-specific IgM. A parallel Bordetella pertussis screening was performed by an indirect immunofluorescence test (Euroimmun, Lübeck, Germany) that detected specific IgM.Results.The CT-specific serum IgM was highly reactive in 50 (19.1 %) of the 262 neonates with RTIs, while all proved negative for Bordetella pertussis-specific IgM.Conclusion. Vertically transmitted C. trachomatis must be regarded as a common pathogen among symptomatic neonates with RTIs in Hungary. Routine screening and treatment of pregnant women could be one option to help prevent these conditions. Focused laboratory testing based on raised clinical awareness should enable early diagnosis and appropriate therapy for symptomatic infants.

Distribution of Chlamydia trachomatis genotypes in neonatal conjunctivitis in Hungary.

The objective of the present study was to determine the frequency and age distribution of different Chlamydia trachomatis (CT) genotypes causing ophthalmia neonatorum (ON) in Hungary. Using CT specific PCR, we tested 76 conjunctival samples from symptomatic infants up to 3 months old in the National Centre for Epidemiology, Budapest between 2008 and 2016. CT tested positive in 30 of 76 conjunctival samples (39.5 %). The sequencing of the positive samples was successful in every case but one, and resulted in 48 % dominance for genotype E (14/29), followed by 24 % for genotype G (7/29), 10 % for J (3/29), 6.9 % for K and F (2/29), and 3.4 % for H (1/29). CT must still be regarded as a common pathogen causing ON in Hungary. Routine screening and treatment of pregnant women can be recommended to prevent these conditions. Chronic ON cases can be reduced by early diagnosis. Further research is needed to explain the dominance of genotypes E and G.

Chlamydial conjunctivitis: prevalence and serovar distribution of Chlamydia trachomatis in adults.

The extragenital manifestation of Chlamydia trachomatis infection frequently results in non-specific conjunctivitis among sexually active adults. The aims of the present study were to determine the prevalence of C. trachomatis, to describe the distribution of serovars among patients with conjunctivitis and to characterize the relationship between the prevalence and patient demographics such as age and gender. A total of 245 conjunctival specimens were screened for C. trachomatis DNA targeting the plasmid gene. Serovar determination of the C. trachomatis-positive specimens was carried out by an omp1 PCR-based RFLP analysis method. Statistical analysis was done using a generalized linear model. C. trachomatis was detected in 53 cases (21.6 %) of adult conjunctivitis. Molecular genotyping differentiated seven distinct urogenital serovars, the most prevalent being serovar E (16/53), followed by F (15/53), D (6/53), K (6/53), G (4/53), H (4/53) and J (2/53). Statistical analysis showed higher C. trachomatis prevalence in the younger age groups, and this peaked at younger age in women than in men. The high prevalence of this pathogen found in ocular samples should alert ophthalmologists to focus on the role of C. trachomatis in adult conjunctivitis. The serovar distribution indicated that ocular chlamydial infections usually have a genital source. Nevertheless, conjunctivitis might be the only sign of this sexually transmitted infection. Further comparative genotyping of C. trachomatis in ocular and genital specimens might give more detailed epidemiological information about the aetiology of the disease.

Streptococcus pneumoniae as an etiologic agent in infectious complications of pancreatic disease.

Systematic culturing of perioperative intra-abdominal samples allowed us to recognize the presence of Streptococcus pneumoniae in infectious complications of chronic pancreatitis in 9 male patients. In 8 of 10 of the intra- or peripancreatic samples, S. pneumoniae was the single isolate identified. All but one of the S. pneumoniae isolates were sensitive to penicillin. The patients had predisposing underlying conditions such as alcoholism and diabetes mellitus. All patients were cured due to adequate surgery and antibiotic treatment. Analysis of the case histories suggests that S. pneumoniae may have been a relevant organism causing the infectious complications of pancreatitis in these patients.