Interactions between endocarditis-derived Streptococcus gallolyticus subsp. gallolyticus isolates and human endothelial cells

<p>Abstract</p> <p>Background</p> <p><it>Streptococcus gallolyticus </it>subsp. <it>gallolyticus </it>is an important causative agent of infective endocarditis (IE) but the knowledge on virulence factors is limited and the pathogenesis of the infection is poorly understood. In the present study, we established an experimental <it>in vitro </it>IE cell culture model using EA.hy926 and HUVEC cells to investigate the adhesion and invasion characteristics of 23 <it>Streptococcus gallolyticus </it>subsp. <it>gallolyticus </it>strains from different origins (human IE-derived isolates, other human clinical isolates, animal isolates). Adhesion to eight components of the extracellular matrix (ECM) and the ability to form biofilms <it>in vitro </it>was examined in order to reveal features of <it>S. gallolyticus </it>subsp. <it>gallolyticus </it>endothelial infection. In addition, the strains were analyzed for the presence of the three virulence factors <it>gtf</it>, <it>pilB</it>, and <it>fimB </it>by PCR.</p> <p>Results</p> <p>The adherence to and invasion characteristics of the examined <it>S. gallolyticus </it>subsp. <it>gallolyticus </it>strains to the endothelial cell line EA.hy926 differ significantly among themselves. In contrast, the usage of three different <it>in vitro </it>models (EA.hy926 cells, primary endothelial cells (HUVECs), mechanical stretched cells) revealed no differences regarding the adherence to and invasion characteristics of different strains. Adherence to the ECM proteins collagen I, II and IV revealed the highest values, followed by fibrinogen, tenascin and laminin. Moreover, a strong correlation was observed in binding to these proteins by the analyzed strains. All strains show the capability to adhere to polystyrole surfaces and form biofilms. We further confirmed the presence of the genes of two known virulence factors (<it>fimB</it>: all strains, <it>gtf</it>: 19 of 23 strains) and demonstrated the presence of the gene of one new putative virulence factor (<it>pilB</it>: 9 of 23 strains) by PCR.</p> <p>Conclusion</p> <p>Our study provides the first description of <it>S. gallolyticus </it>subsp. <it>gallolyticus </it>adhesion and invasion of human endothelial cells, revealing important initial information of strain variability, behaviour and characteristics of this as yet barely analyzed pathogen.</p

Optical Monitoring of the Didymos–Dimorphos Asteroid System with the Danish Telescope around the DART Mission Impact

The NASA’s Double-Asteroid Redirection Test (DART) was a unique planetary defence and technology test mission, the first of its kind. The main spacecraft of the DART mission impacted the target asteroid Dimorphos, a small moon orbiting the asteroid Didymos (65803), on 2022 September 26. The impact brought up a mass of ejecta which, together with the direct momentum transfer from the collision, caused an orbital period change of 33 ± 1 minutes, as measured by ground-based observations. We report here the outcome of the optical monitoring campaign of the Didymos system from the Danish 1.54 m telescope at La Silla around the time of impact. The observations contributed to the determination of the changes in the orbital parameters of the Didymos–Dimorphos system, as reported by Thomas et al., but in this paper we focus on the ejecta produced by the DART impact. We present photometric measurements from which we remove the contribution from the Didymos–Dimorphos system using an H–G photometric model. Using two photometric apertures we determine the fading rate of the ejecta to be 0.115 ± 0.003 mag day−1 (in a 2″ aperture) and 0.086 ± 0.003 mag day−1 (5″) over the first week postimpact. After about 8 days postimpact we note the fading slows down to 0.057 ± 0.003 mag day−1 (2″ aperture) and 0.068 ± 0.002 mag day−1 (5″). We include deep-stacked images of the system to illustrate the ejecta evolution during the first 18 days, noting the emergence of dust tails formed from ejecta pushed in the antisolar direction, and measuring the extent of the particles ejected Sunward to be at least 4000 km

Six Outbursts of Comet 46P/Wirtanen

Cometary activity is a manifestation of sublimation-driven processes at the surface of nuclei. However, cometary outbursts may arise from other processes that are not necessarily driven by volatiles. In order to fully understand nuclear surfaces and their evolution, we must identify the causes of cometary outbursts. In that context, we present a study of mini-outbursts of comet 46P/Wirtanen. Six events are found in our long-term lightcurve of the comet around its perihelion passage in 2018. The apparent strengths range from −0.2 to −1.6 mag in a 5″ radius aperture and correspond to dust masses between ∼104 and 106 kg, but with large uncertainties due to the unknown grain size distributions. However, the nominal mass estimates are on the same order of magnitude as the mini-outbursts at comet 9P/Tempel 1 and 67P/Churyumov-Gerasimenko, events that were notably lacking at comet 103P/Hartley 2. We compare the frequency of outbursts at the four comets, and suggest that the surface of 46P has large-scale (∼10–100 m) roughness that is intermediate to that of 67P and 103P, if not similar to the latter. The strength of the outbursts appear to be correlated with time since the last event, but a physical interpretation with respect to solar insolation is lacking. We also examine Hubble Space Telescope images taken about two days following a near-perihelion outburst. No evidence for macroscopic ejecta was found in the image, with a limiting radius of about 2 m

Molekulargenetische Charakterisierung von Streptococcus gallolyticus subsp. gallolyticus : vergleichende Genomanalyse und Evaluation von Stammtypisierungsmethoden

Hinse D. Molekulargenetische Charakterisierung von Streptococcus gallolyticus subsp. gallolyticus : vergleichende Genomanalyse und Evaluation von Stammtypisierungsmethoden. Bielefeld: Universität Bielefeld; 2012.Streptococcus gallolyticus subsp. gallolyticus ist ein fakultativ pathogenes Bakterium, das schwerwiegende Erkrankungen wie die infektiöse Endokarditis, Meningitis und Sepsis in Mensch und Tier verursachen kann. Der Organismus wird im humanen Gastrointestinaltrakt als putativer Kommensale nachgewiesen, zeigt jedoch eine starke Assoziation mit kolorektalen Karzinomen. Unklar ist, ob eine Transmission zwischen Mensch und Tier möglich ist und inwiefern Tierpopulationen ein Reservoir für dieses Pathogen darstellen. Im Rahmen dieser Arbeit wurden zunächst alle in einer Streptokokken-Stammsammlung vorhandenen Isolate tierischer und humaner Herkunft, mittels sodA DNA-Sequenzierung und MALDI-TOF-MS, identifiziert. Die erhaltenen Daten zeigten eine ausgezeichnete Korrelation und wurden für eine phylogenetische Analyse der Stämme verwendet. Die bestehende Taxonomie wurde bestätigt und es konnten 18 Stämme aus verschiedenen Stammsammlungen als taxonomisch falsch hinterlegt identifiziert werden. Weiterhin wurde die Voraussetzung geschaffen, um eine zuverlässige und kosteneffiziente Identifizierung des S. bovis / S. equinus Komplexes mittels der in der Routinediagnostik verwendeten MALDI-TOF-MS Methode zu ermöglichen. Ferner wurde in dieser Arbeit das Genom eines hochvirulenten humanen S. gallolyticus subsp. gallolyticus Isolats sequenziert und analysiert. Es wurden entscheidende, für die bakterielle Pathogenität verantwortliche, Virulenzfaktoren identifiziert und in silico mit anderen S. gallolyticus subsp. gallolyticus Genomen verglichen. Darunter befinden sich putative Hämolysine, Kollagenadhäsine und Pilusproteine, die alle maßgeblich an der Infektionskaskade beteiligt sind. Erstmals konnte das Plasmid pSGG1 beschrieben werden, welches Träger einer Tetracyclinresistenz ist. Diese Informationen tragen zum Verständnis der Pathogenese von S. gallolyticus subsp. gallolyticus bei und dienten als fundamentale Grundlage für weitere durchgeführte Forschungsvorhaben. Ausgehend von den gesammelten Genominformationen wurde in weiteren Arbeiten ein S. gallolyticus subsp. gallolyticus spezifischer Microarray entwickelt. Dieser sogenannte "GalloChip" detektiert 203 für Virulenz- und Antibiotikaresistenzproteine kodierende Gene und wurde erfolgreich für die Charakterisierung des vorhandenen Stammkollektivs eingesetzt. Die ermittelten Daten lassen unter anderem Rückschlüsse auf die Kollagenbindungsfähigkeit und damit die putative Virulenz zu. Erstmalig wurde eine real-time PCR basierte Methode zum Nachweis von S. gallolyticus subsp. gallolyticus in Stuhlproben etabliert und angewendet. Erste Daten weisen auf eine hohe Kolonisierungsrate des humanen Gastrointestinaltraktes hin und unterstreichen den Bedarf einer umfassenden Studie zur Prävalenz von S. gallolyticus subsp. gallolyticus. Durch die Etablierung einer ERIC-PCR, einer rep-PCR und der erstmaligen Entwicklung eines S. gallolyticus subsp. gallolyticus spezifischen MLST-Schemas konnten Stämme in charakteristische Cluster eingeordnet werden. Es wurde gezeigt, dass keine signifikanten wirtsspezifischen Populationen nachweisbar sind. Damit konnte die Grundlage geschaffen werden, um epidemiologische Zusammenhänge zu analysieren und Infektionsketten zu erkennen.The facultative pathogen Streptococcus gallolyticus subsp. gallolyticus is able to cause several serious diseases, including infective endocarditis, meningitis and sepsis in animals and humans. It can be identified as a commensal in humans, but equally has a strong association with colorectal cancer. A transfer from animals to humans is not proven up to now and whether infected animals might be a potential reservoir for this pathogen is still elusive. Within this work the identification of an existing collection of Streptococcus strains with human and animal origin, by use of sodA DNA-sequencing and MALDI-TOF-MS, was performed. The respective data show an excellent correlation and were used for phylogenetic analysis. The current taxonomy was confirmed and, furthermore, 18 strains from different collections were updated because of a previous false taxonomy classification. In addition, the preconditions for reliable and cost-efficient MALDI-TOF-MS routine diagnostic identification for the S. bovis / S. equinus complex were created. Moreover, the genome of a high virulent human S. gallolyticus subsp. gallolyticus isolate was sequenced and analyzed. Important virulence factors for bacterial pathogenicity could be identified, including putative genes coding for hemolysins, collagen-adhesins and pilusproteins, which are known to mediate infection processes. For the first time the presence of the plasmid pSGG1 was shown, carrying genes for tetracycline resistance. These results will lead to a better understanding of the pathogenesis of S. gallolyticus subsp. gallolyticus and provides fundamental information for further projects. Based on acquired genetic data, a S. gallolyticus subsp. gallolyticus specific qualitative microarray has been established. The "GalloChip" is able to detect 203 virulence and antibiotic resistance genes and was used for typing the established strain collection. The obtained data show correlation to collagen binding ability and therefore lead to estimation of putative virulence. A real-time PCR based method for detecting S. gallolyticus subsp. gallolyticus in human feces was established, too. First results of screening experiments present a high prevalence in human gastrointestinal tracts and underline the need for an extensive study of prevalence of S. gallolyticus subsp. gallolyticus. With establishing an ERIC-PCR, a rep-PCR and for the first time a S. gallolyticus subsp. gallolyticus specific MLST-scheme, strains could be characterized and grouped in clusters. No evidence for host- or geographic related specific clustering of strains occurred. The development of these methods provides the basics for analyzing the epidemiological connections and infection chains

The Pan Genera Detection Immunoassay: a Novel Point-of-Issue Method for Detection of Bacterial Contamination in Platelet Concentrates ▿

Bacterial contamination of platelet concentrates (PCs) still represents an ongoing risk in transfusion-transmitted sepsis. Recently the Pan Genera Detection (PGD) system was developed and FDA licensed for screening of bacterial contamination of PCs directly prior to transfusion. The test principle is based on the immunological detection of lipopolysaccharide (for Gram-negative bacteria) or lipoteichoic acid (for Gram-positive bacteria). In the present study we analyzed the applicability of this method with regard to detection limit, practicability, implementation, and performance. PCs were spiked with Staphylococcus aureus, Bacillus subtilis, and five different Klebsiella pneumoniae strains, as well as eight different Escherichia coli strains. The presence of bacteria was assessed by the PGD immunoassay, and bacteria were enumerated by plating cultures. Application of the PGD immunoassay showed that it is a rapid test with a short hands-on time for sample processing and no demand for special technical equipment and instrument operation. The lower detection limits of the assay for Gram-positive bacteria showed a good agreement with the manufacturer's specifications (8.2 × 103 to 5.5 × 104 CFU/ml). For some strains of K. pneumoniae and E. coli, the PGD test showed analytical sensitivities (>106 CFU/ml) that were divergent from the designated values (K. pneumoniae, 2.0 × 104 CFU/ml; E. coli, 2.8 × 104 CFU/ml). Result interpretation is sometimes difficult due to very faint bands. In conclusion, our study demonstrates that the PGD immunoassay is an easy-to-perform bedside test for the detection of bacterial contamination in PCs. However, to date there are some shortcomings in the interpretation of results and in the detection limits for some strains of Gram-negative bacteria

Potential Transmission Pathways of Streptococcus gallolyticus subsp. gallolyticus.

Streptococcus gallolyticus subsp. gallolyticus (S. gallolyticus subsp. gallolyticus), a member of group D streptococci, is an inhabitant of the animal and human gastrointestinal tract. Furthermore, it is a facultative pathogen which causes e.g. endocarditis, septicemia and mastitis. S. gallolyticus subsp. gallolyticus may be transmitted either directly or indirectly between animals and humans. However, the transmission routes are an unsolved issue. In this study, we present systematic analyses of an S. gallolyticus subsp. gallolyticus isolate of an infective endocarditis patient in relation to isolates of his laying hen flock. Isolates from pooled droppings of laying hens, pooled dust samples and human blood culture were characterized by using multilocus sequence typing (MLST) and DNA fingerprinting. MLST revealed the same allelic profile of isolates from the human blood culture and from the droppings of laying hens. In addition, these isolates showed clonal identity regarding a similar DNA fingerprinting pattern. For the first time, we received a hint that transmission of S. gallolyticus subsp. gallolyticus between poultry and humans may occur. This raises the question about the zoonotic potential of isolates from poultry and should be considered in future studies

Organic Turkey Flocks: A Reservoir of Streptococcus gallolyticus subspecies gallolyticus.

Streptococcus gallolyticus subspecies gallolyticus (S. gallolyticus) can colonise the gastrointestinal tract of humans and animals and is known to cause similar infections in both humans and animals. Data about the spread or prevalence in farm animals are missing. In this study, Trypton Soya Agar was modified to a selective medium enabling the isolation and quantification of S. gallolyticus from faecal samples. The bacterium was observed in 82 out of 91 faecal samples obtained from 18 different organic turkey flocks. The prevalence of shedding birds was estimated by the number of positive fresh droppings and reached up to 100% on most farms. Furthermore, for the first time S. gallolyticus was quantified in faeces from poultry flocks. The median of colony forming units (CFU) per gramme faeces was 3.6 x 10(5) CFU/g. Typing of one isolate from each positive faecal sample by multilocus sequence typing delivered 24 sequence types (STs). Most of the isolates belonged to the clonal complex CC58. The same STs of this complex were detected in up to six different flocks. Partly, these flocks were located in various regions and stocked with varying breeding lines. Regarding the biochemical profiles of the same STs from different farms, the results did not contradict a spread of specific STs in the organic turkey production. Moreover, checking the pubMLST database revealed that STs found in this study were also found in other animal species and in humans. The high detection rate and the number of S. gallolyticus in turkey faeces indicate that this bacterium probably belongs to the common microbiota of the gastrointestinal tract of turkeys from organic flocks. Furthermore, the findings of this study support the suggestion of a possible interspecies transmission