Shift in bacterial etiology from the CAPNETZ cohort in patients with community-acquired pneumonia: data over more than a decade.

To determine the most relevant pathogens for CAP in Germany, patients with radiologically confirmed pulmonary infiltrates and at least one clinical sign of lung infection were prospectively recruited within the CAPNETZ cohort from 2004 until 2016. In 990 out of 4.672 patients (21%) receiving complete diagnostics the most prominent change of pathogens was a decrease of S. pneumoniae (58% in 2004 to 37.5% in 2016; p ≤ 0.001, ρ = - 0.148) and an increase of H. influenzae (12.2% to 20.8%; p = 0.001, ρ = 0.104).

Prevalence and characterization of azole-resistant Aspergillus fumigatus in patients with cystic fibrosis: a prospective multicentre study in Germany.

Objectives: Aspergillus fumigatus is the most prevalent filamentous fungus in the respiratory tract of patients with cystic fibrosis (CF). The aim of this prospective multicentre study was to investigate the prevalence of azole-resistant A. fumigatus (ARAF) in respiratory secretions from CF patients across Germany and to characterize ARAF isolates by phenotypic and molecular methods. Methods: Twelve tertiary care centres from Germany participated in the study. In total, 2888 A. fumigatus isolates from 961 CF patients were screened for ARAF by using azole-containing agar plates. Antifungal susceptibility testing of isolates was performed by broth microdilution according to EUCAST guidelines. Analysis of mutations mediating resistance was performed using PCR and sequencing of the cyp51A gene. Furthermore, genotyping by microsatellite PCR was performed. Results: Of a total of 2888 A. fumigatus isolates, 101 isolates from 51 CF patients were found to be azole resistant (prevalence per patient 5.3%). The Essen centre had the highest prevalence (9.1%) followed by Munich (7.8%), Münster (6.0%) and Hannover (5.2%). Most ARAF isolates (n = 89) carried the TR34/L98H mutation followed by eight G54E/R, one TR46/Y121F/T289A and one F219S mutation. In two isolates no mutation was found. Genotyping results showed no major clustering. Forty-five percent of CF patients with ARAF had previously received azole therapy. Conclusions: This is the first multicentre study analysing the prevalence of ARAF isolates in German CF patients. Because of a resistance rate of up to 9%, susceptibility testing of A. fumigatus isolates from CF patients receiving antifungal treatment should be part of standard diagnostic work-up.

Difficulties in species identification within the genus Haemophilus - A pilot study addressing a significant problem for routine diagnostics.

Diagnostic misidentifications of commensalic Haemophilus haemolyticus as pathogenic Haemophilus influenzae are frequent. This pilot study evaluates whether isolations of H. haemolyticus are frequent enough in Germany to cause a relevant diagnostic problem, considering the fact that even H. influenzae is a mere colonizer in about 30% of isolations. In microbiological laboratories of two hospitals located in Northern and Southern Germany, the distribution of Haemophilus spp. was analyzed during a six-month-period. Site of infection, sex, and age of the patients was taken into consideration. A total of 77 Haemophilus spp. isolates was acquired and discriminated on species level, comprising: 48 H. influenzae, 25 Haemophilus parainfluenzae, 3 H. haemolyticus, and 1 Haemophilus parahaemolyticus. The proportion of H. haemolyticus was calculated to range between 1.2% and 16.2 % within the 95% confidence limits. Commensalic Haemophilus spp. were isolated from oropharynx-associated sites only. H. influenzae, in contrast, was detected in clinically relevant materials like lower respiratory materials and conjunctiva swabs. Altogether, there was a low proportion of clinical H. haemolyticus isolates. Accordingly, the problem of unnecessary antibiotic therapies due to misidentifications of H. haemolyticus as H. influenzae is quantitatively negligible compared with the risk of confusing H. influenzae colonizations with infections.

Electron tomography and cryo-SEM characterization reveals novel ultrastructural features of host-parasite interaction during Chlamydia abortus infection.

Chlamydia (C.) abortus is a widely spread pathogen among ruminants that can be transmitted to women during pregnancy leading to severe systemic infection with consecutive abortion. As a member of the Chlamydiaceae, C. abortus shares the characteristic feature of an obligate intracellular biphasic developmental cycle with two morphological forms including elementary bodies (EBs) and reticulate bodies (RBs). In contrast to other chlamydial species, C. abortus ultrastructure has not been investigated yet. To do so, samples were fixed by high-pressure freezing and processed by different electron microscopic methods. Freeze-substituted samples were analysed by transmission electron microscopy, scanning transmission electron microscopical tomography and immuno-electron microscopy, and freeze-fractured samples were analysed by cryo-scanning electron microscopy. Here, we present three ultrastructural features of C. abortus that have not been reported up to now. Firstly, the morphological evidence that C. abortus is equipped with the type three secretion system. Secondly, the accumulation and even coating of whole inclusion bodies by membrane complexes consisting of multiple closely adjacent membranes which seems to be a C. abortus specific feature. Thirdly, the formation of small vesicles in the periplasmic space of RBs in the second half of the developmental cycle. Concerning the time point of their formation and the fact that they harbour chlamydial components, these vesicles might be morphological correlates of an intermediate step during the process of redifferentiation of RBs into EBs. As this feature has also been shown for C. trachomatis and C. pneumoniae, it might be a common characteristic of the family of Chlamydiaceae.

Chlamydia trachomatis prevalence, genotype distribution and identification of the new Swedish variant in Southern Germany.

PURPOSE: In Germany, reliable data about the prevalence of urogenital Chlamydia trachomatis infections, causative genotypes, as well as corresponding clinical, demographic and behavioural information are sparse. We, therefore, performed a prospective prevalence study including 1,003 sexually active volunteers of a Southern German city. METHODS: Study participants completed a standardised questionnaire and provided first void urine samples for analysis. Our screening strategy included the performance of two nucleic acid amplification tests with different target genes, enabling the detection of the new Swedish variant of C. trachomatis (nvCT). Direct genotyping of positive specimens was performed by sequence analysis of the ompA gene. RESULTS AND CONCLUSION: The overall prevalence of C. trachomatis infection was 4.2 % in women and 4.6 % in men. A relatively high prevalence of 8.3 % was found in men older than 25 years. Never using condoms was an independent risk factor for infection. The most common symptom was discharge; however, 64.5 % of infected females and all of the infected men were asymptomatic, supporting the need for screening programmes. The most frequently encountered genotypes were E (46.5 %), F (20.9 %) and K (14.0 %). Since the nvCT was detected in one female student, this is one of the rare studies that reports on the molecular identification of nvCT apart from Sweden.

Performance of a novel microarray multiplex PCR for the detection of 23 respiratory pathogens (SYMP-ARI study).

Symptoms of acute febrile respiratory tract infection are often unspecific, but the rapid identification of pathogens allows optimised patient management. The objective of this study was to evaluate a novel multiplex polymerase chain reaction (PCR) suspension microarray which detects 19 viral and four atypical bacterial targets. A comprehensive set of sensitive monoplex real-time PCR assays was used for each pathogen as the gold standard. A panel of archived as well as 300 prospectively collected clinical samples was analysed by both methods. At least one target was detected in 165/300 (55 %) samples by monoplex PCR and in 140/300 (46 %) samples by multiplex PCR, respectively. The positivity rate was significantly higher in paediatric patients compared to adults [126/154 (82 %) vs. 39/146 (27 %) by monoplex and 114/154 (74 %) vs. 26/146 (18 %) by multiplex PCR, respectively]. Among all samples, 17/300 (5.6 %) were positive for atypical bacteria by monoplex and 8/300 (2.6 %) by multiplex PCR, respectively. Multiple detections were recorded in 35/300 (11.6 %) samples by monoplex and 26/300 (8.7 %) by multiplex PCR. For the most common pathogens, the sensitivity ranged from 57 to 93 % and the specificity ranged from 95 to 100 %. The overall concordance between both methods was 77 % [95 % confidence interval (CI) 72-81 %]. False-negative results by multiplex PCR were mainly due to the low target concentration. Compared to monoplex PCR, the novel microarray assay proved its principle but displayed overall lower sensitivities, potentially restricting its use to paediatric patients. For some targets, only small numbers of positive samples were available, requiring larger studies to firmly assess the sensitivity and specificity.

[Interdisciplinary management of a large Legionella outbreak in Germany]. / Interdisziplinäres Management eines länderübergreifenden Legionellenausbruchs.

Between December 2009 and the end of January 2010, the largest hitherto known outbreak of Legionella in Germany took place in the cities of Ulm and Neu-Ulm. Of a total of 64 patients involved, 60 patients had to be hospitalized, and 5 patients died from the infection. This event was caused by a wet cooling tower of a large air conditioning system in the city center of Ulm. The search for the source of the Legionella emission was extremely difficult, since these plants are neither notifiable nor subject to authorization in Germany. We report about the search for the source and the measures to control the outbreak. We also discuss communication and coordination during these investigations. Regulatory measures as proposed by the World Health Organization (WHO) and the European Network for Legionellosis (EWGLI) and already implemented in numerous other European countries would be desirable to prevent such outbreaks in the future.

Rapid identification of Acinetobacter spp. by fluorescence in situ hybridization (FISH) from colony and blood culture material.

Multi-drug-resistant strains of the Acinetobacter baumannii complex cause nosocomial infections. Rapid identification of Acinetobacter spp. is desirable in order to facilitate therapeutic or hygiene decisions. We evaluated a newly designed DNA probe that can be used under standard conditions in both a microwave oven and a slide chamber for the rapid identification of Acinetobacter spp. by fluorescence in situ hybridization (FISH). Using FISH, the new probe correctly identified 81/81 Acinetobacter spp. isolates and excluded 109/109 tested non-target organisms from agar culture. Furthermore, the new probe correctly identified 7/7 Acinetobacter spp. in 214 blood cultures determined to contain Gram-negative bacteria by Gram staining. Using either the microwave oven or slide chamber technique, the new probe was able to identify Acinetobacter spp. in 100% of the samples tested. FISH used in conjunction with our newly designed probe provides an easy, cheap, precise, and rapid method for the preliminary identification of Acinetobacter spp., especially in laboratories where more sophisticated methods like matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) are not available.

Preliminary report: outbreak of Legionnaires disease in the cities of Ulm and Neu-Ulm in Germany, December 2009 - January 2010.

Currently an investigation is ongoing to explore and control an outbreak of Legionnaires disease, affecting 65 people as of 22 January 2010, in the cities of Ulm and Neu-Ulm, south-west Germany. A hitherto unidentified wet cooling system in these twin cities is considered as the most likely source of infection.

Identification and evaluation of a combination of chlamydial antigens to support the diagnosis of severe and invasive Chlamydia trachomatis infections.

Chlamydia trachomatis is the most common sexually transmitted organism in industrialized countries. Nucleic acid amplification testing, using non-invasively collected specimens, is considered to be the method of choice for diagnosis of chlamydial infections of the urethra and the lower genital tract. Serological testing has the potential to circumvent the problem of specimen sampling in invasive C. trachomatis infections of the upper genital tract. However, only a few defined chlamydial antigens have been used in a standardized diagnostic assay format. In this study, we used serological two-dimensional proteomic analysis to broaden the spectrum of diagnostically relevant C. trachomatis proteins. The genes encoding an assortment of already known chlamydial antigens, as well as immunogenic proteins that have not been described before, were cloned, and the recombinant proteins were purified in order to compare their diagnostic usefulness in parallel with a newly developed line immunoassay. With 189 sera collected from patients with and without C. trachomatis infection, recombinant major outer membrane protein (MOMP), chlamydial protease-like activity factor (CPAF), outer membrane protein 2 (OMP2), translocated actin-recruiting protein, and polymorphic membrane protein D (PmpD) showed the highest level of diagnostic sensitivity and specificity. In patients suffering from ascending and invasive C. trachomatis infections, such as pelvic inflammatory disease and lymphogranuloma venereum, the sensitivity reached with these proteins ranged between 71% (PmpD) and 94% (OMP2), and the specificity ranged between 82% (PmpD) and 100% (MOMP and OMP2). Recombinant thio-specific antioxidant peroxidase, ribosomal protein S1 (RpsA) and hypothetical protein 17 showed lower sensitivity but comparably high specificity, ranging from 94% to 100%. The novel line immunoassay based on defined recombinant antigens has promise for improved serodiagnosis in severe and invasive C. trachomatis infections.

[Cat-scratch disease as cause of Lymphadenitis colli]. / Katzenkratzkrankheit als Ursache einer Lymphadenitis colli.

Cat-scratch Disease as Cause of Lymphadenitis colli. Cat-scratch disease is a frequent cause of lymphadenitis colli. It mainly affects children and adolescents younger than 21 years. Since the clinical picture is not characteristic, a history of contact to cats or kittens is highly valuable for diagnosing the disease. Major aspects of the disease concerning epidemiology, diagnostic procedures, clinical presentation and therapy are discussed.

Nasal carriage of Staphylococcus aureus in house dust mite allergic patients and healthy controls.

BACKGROUND: Staphylococcal colonization may influence the course of allergic diseases such as atopic dermatitis or allergic rhinitis. The frequency of Staphylococcus aureus (SA) nasal carriage and its possible influence on persistent allergic rhinitis was investigated. METHODS: In nasal lavages from 22 patients with house dust mite allergy and 18 healthy controls, the number of SA colony forming units per ml were assessed and related to nasal symptom scores, the concentrations of three inflammatory cell activation markers, nasal total IgE and 17 cytokines in nasal secretions. RESULTS: SA was found in 15/22 allergic patients and 4/18 controls (P < 0.01). Comparing allergic SA carriers with allergic noncarriers, nasal symptom scores tended to be higher (P < 0.1), and the cell activation markers ECP (10(2.23+/-0.33)vs 10(1.45+/-0.50) ng/ml; P < 0.05) and elastase (10(2.70+/-0.21)vs 10(2.12+/-0.34) ng/ml; P < 0.01), and nasal total IgE-levels (10(1.66+/-0.38)vs 10(1.2+/-0.28) kU/ml; P < 0.05) were significantly higher in allergic SA carriers. Nasal SA carriers had a higher nasal IL-13/IFN-gamma ratio (P < 0.01), and this was correlated with higher nasal total IgE in allergic patients (r = 0.6, P < 0.05). CONCLUSION: Nasal SA carriage is frequent in patients with persistent allergic rhinitis. The data of this study suggest that they are not only secondary bystanders, but actively modulate the disease by promoting local IgE production.

[Fulminant pneumococcal sepsis in a 13 months old child with congenital asplenia]. / Fulminante Pneumokokkensepsis bei einem 13 Monate alten Kleinkind mit angeborener familiärer Asplenie.

Asplenia may predispose to fulminant invasive infections caused by encapsulated bacteria. We observed a 13 months old child with (so far unknown) congenital familiar asplenia, who died from pneumococcal sepsis. General vaccination of all infants with pneumococcal conjugate vaccine may prevent this disease, which is associated with a high rate of mortality in infants with asplenia.

Lymphogranuloma venereum - a rare cause of genital ulcers in central Europe.

Lymphogranuloma inguinale, caused by Chlamydia trachomatis serovar L1-L3 is rare in patients from western countries but needs yet to be considered in the differential diagnosis of genital ulcers. We report a case of a young male patient without any eventful travel history who presented with a genital ulcer at the sulcus coronarius and painfully enlarged lymph nodes in the right inguinal area. The typical clinical picture and serum IgM and IgG antibody titers of 1:16 and 1:512, respectively, against C. trachomatis were suggestive of infection with C. trachomatis serovar L1-L3. The diagnosis was confirmed by isolation of the organisms from the ulcer ground and subsequent sequence analysis of the omp1 gene which led to identification of C. trachomatis genotype L2 with 99% homology to a reference strain of C. trachomatis serovar L2. The lesion healed rapidly under treatment with doxycycline for 3 weeks, and the lymph nodes did not ulcerate. Thus, clinical suspicion was confirmed by genotyping of the isolated strain allowing timely diagnosis and treatment of lymphogranuloma inguinale.

Rapid and standardized detection of Chlamydia pneumoniae using LightCycler real-time fluorescence PCR.

In order to meet the need of many microbiological laboratories for a standardized system for detecting Chlamydia pneumoniae in respiratory specimens, a hybridization probe-based LightCycler (Roche Diagnostics, Germany) PCR assay was developed. The assay's analytical sensitivity and specificity were evaluated according to the recommendations of the Centers for Disease Control and Prevention (USA) and Laboratory Centre for Disease Control (Canada). Seventy-four bacterial species other than Chlamydia pneumoniae, including strains of Chlamydia trachomatis, Chlamydia psittaci, and Chlamydia pecorum, tested negative. Six of six representative Chlamydia pneumoniae strains tested positive. An analytical sensitivity of 1 inclusion forming unit per ml of bronchoalveolar lavage fluid, corresponding to 0.02 inclusion forming units per PCR reaction, was observed. The assay showed 100% specificity and sensitivity for Chlamydia pneumoniae when testing DNA preparations from 12 specimens of patients with known pulmonary Chlamydia pneumoniae infection and from 78 specimens of patients with respiratory tract disease of other origin. The newly developed LightCycler assay may contribute to the urgently needed standardization of laboratory diagnosis of Chlamydia pneumoniae.

Taxonomic reconsideration of the genus Chorioptes Gervais and van Beneden, 1859 (Acari: Psoroptidae).

The present review of the literature on mites of the genus Chorioptes Gervais and Van Beneden, 1859 argues for a support of the validity of C. bovis (Hering, 1845) and C. texanus Hirst, 1924 based on biological, morphological and molecular genetic studies. However, the validity of three further species. C. crewei Lavoipierre, 1958, C. mydaus Fain, 1975 and C. panda Fain and Leclerc, 1975, is regarded as questionable because discriminations of mites, which were described as isolated cases only, were based on morphological features while transfer or cross-breeding studies were not done.

Survival of Chlamydia pneumoniae-infected Mono Mac 6 cells is dependent on NF-kappaB binding activity.

The respiratory tract pathogen Chlamydia pneumoniae has been associated with atherosclerosis. Monocytes are supposed to serve as a vehicle for systemic dissemination of intracellular C. pneumoniae from the lung to the artery vessel wall. We were therefore interested in pathogen-induced cellular events associated with NF-kappaB, a crucial transcription factor for both inflammatory cytokines and antiapoptotic molecules. In this study we demonstrate by electrophoretic mobility shift assay that C. pneumoniae infection of the human monocytic cell line Mono Mac 6 induces activation of NF-kappaB over 48 h, with a maximum level at 1 h postinfection. As shown by supershift assay, the activated NF-kappaB complex consists of the subunits RelA (p65) and NF-kappaB1 (p50). Apoptotic host cells were not detected during the early stages of the infection when maximal activation of NF-kappaB was detected. Pretreatment of Mono Mac 6 with the antioxidant and NF-kappaB inhibitor PDTC (pyrrolidine dithiocarbamate) induced activation of caspase-3 and led to apoptotic cell death. The C. pneumoniae-induced activation of the NF-kappaB complex was reduced by PDTC, which in parallel resulted in an increased apoptosis, as quantified by annexin V labeling and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling reaction. In the complete absence of activated NF-kappaB, when Mono Mac 6 cells were pretreated with the more potent NF-kappaB inhibitors MG-132 and parthenolide a C. pneumoniae-mediated rescue of cells from induced apoptosis could not be achieved. Our results indicate that activation of NF-kappaB in C. pneumoniae-infected Mono Mac 6 cells is associated with protection of Mono Mac 6 cells against apoptosis and might thereby contribute to systemic spread of the pathogen.

Value of Candida antigen and antibody assays for the diagnosis of invasive candidosis in surgical intensive care patients.

During a 3-year period, a clinical diagnosis of invasive candidosis was made in 8 out of 2054 consecutive surgical intensive care unit (ICU) patients. These patients were retrospectively matched with 16 control patients who underwent similar surgical procedures and had a similar clinical course except for negative Candida cultures. In all patients, Candida antigen (Ramco CandTec serum antigen test) and antibody serology (Candida HA test) were determined at least once a week during their stay. The antigen test was positive in 1/8 patients and 4/16 controls and thus did not differentiate patients with candidosis from non-infected controls. The HA antibody titer results fulfilled the manufacturer's criteria for positivity in 7/8 patients with candidosis and 2/16 control patients. Thus, the Candida HA antibody test, but not the Ramco antigen test, can be recommended to confirm a clinical diagnosis of invasive candidosis.

Species of the genus Psoroptes (Acari: Psoroptidae): a taxonomic consideration.

The biosystematic status of mite species belonging to the genus Psoroptes Gervais, 1841 is difficult to determine by phenotypic methods and has been subject to taxonomic revisions and ongoing debate. At present, the existence of five species, P cuniculi (Delafond, 1859), P. ovis (Hering, 1838). P. equi (Hering, 1838), P. cervinus Ward, 1915 and P. natalensis Hirst, 1919, is generally accepted. This classification is based mainly on the host species, the localization of the mites on their hosts and morphological characters of male mites. However, a critical review of the literature indicates that the features used to discriminate between the five species are not unequivocal: (a) the localization of mite populations on host animals is not completely strict, (b) the lengths of the outer opisthosomal setae of male mites, which are the main morphological features used for species discrimination, overlap between the five postulated species, and (c) host specificity cannot be deduced from results of transfer experiments. Rather, conspecificity of the members of the genus Psoroptes has to be presumed which is supported by molecular genetic analyses. On these grounds and on rules of priority P. cervinus Ward, 1915, P. cuniculi (Delafond, 1859), P. natalensis Hirst, 1919 and P. ovis (Hering, 1838) are seen as synonyms of P. equi (Hering, 1838).