Epidemiology and species distribution of anaerobic Gram-negative cocci: a 10-year retrospective survey (2008-2017)

Introduction: The group of anaerobic Gram-negative cocci (AGNC) includes the genera Veillonella, Megasphaera, Anaeroglobus, Negativicoccus and Acidaminococcus. These bacteria are an integral part of the microbiome of humans but may be causative agents in various infectious processes. The available data on the epidemiology and significance of AGNCs is scarce. Aims: To assess and compare the prevalence of different species of AGNCs among inpatients and outpatients at the Albert Szent-Györgyi Clinical Center retrospectively, during a 10-year study period. Methods: Isolates containing AGNC were identified retrospectively by reviewing the online microbiology records of the Institute of Clinical Microbiology. Results: The median age of affected patients overall was 52 years (range: 1–90 years), with a male dominance. 59.79% of samples originated from inpatients. 572 individual AGNCs isolates were recovered from clinical samples, most of the isolated GNACs were Veillonella spp. (95.28%), Megasphaera and Acidaminococcus species accounted for a minority of isolates (2.79% and 1.93%, respectively), while Anaeroglobus and Negativicoccus species were not isolated. In the second half of the study period (2013-2017), 91.31% of isolates were identified on the species level (p<0.001) using MALDI-TOF MS. Conclusion: The current study represents a long-term surveillance study on the isolation frequency and trends among anaerobic Gram-negative cocci (AGNCs), isolated in the Southern Great Plain of Hungary, highlighting the beneficial effect of MALDI-TOF MS on the diagnostic efficacy of the laboratorie

A Szülői Konfliktusok Észlelését Mérő Skála hazai alkalmazásával szerzett tapasztalatok = Psychometric evaluation of the Hungarian version of the Children’s Perception of Interparental Conflict Scale

Elméleti háttér: A Szülői Konfliktusok Észlelését Mérő Skála (Children’s Perception of Interparental Conflict Scale; CPIC) egy széles körben használt kérdőív, amely a szülői konfliktusok gyermekek által észlelt természetét és a gyermekeknek e konfliktusokra adott reakcióit vizsgálja. Cél: A kutatás célja a CPIC magyar nyelvre adaptálása és pszichometriai elemzése volt. Módszer: Keresztmetszeti, kérdőíves vizsgálatunkban 127 szülő-gyermek pár vett részt. A gyermekek (átlagéletkor = 10,8 év; szórás = 1,05 év; terjedelem: 9—12 év) az alábbi kérdőíveket töltötték ki: CPIC, Vonásszorongás Skála, Gyermek Depresszió Kérdőív. A szülőkkel a Rövidített Házastársi Stressz Skálát vettük fel. Eredmények: A konfirmatív faktoranalízis nem támasztotta alá a CPIC elméleti, nyolcfaktoros faktorstruktúráját (χ2(1052) = 1355,0, p90: 0,040—0,055, p = 0,698). A Trianguláció alskála három tételének törlésével az illeszkedés elfogadhatóvá vált (χ2(917) = 1113,4, p90: 0,032—0,049, p = 0,960). A négyfaktoros alternatív elméleti modell szignifikánsan rosszabbul illeszkedett az adatokra, mint a nyolcfaktoros modell (Δχ2 = 66,5, Δdf = 22, p<0,001). A kérdőív alskáláinak belső megbízhatósága a Trianguláció alskála (Cronbach-alfa: 0,40) kivételével elfogadható (Cronbach-alfa: 0,63—0,81). A mérőeszköz konstruktumvaliditását támogatja, hogy az alskálák zöme a várakozásnak megfelelő irányú és erősségű lineáris kapcsolatot mutat a depresszióval, a vonásszorongással, valamint a szülő által észlelt házastársi stresszel. Következtetések: Az alacsony mintaelemszám ellenére összességében elmondhatjuk, hogy a CPIC magyar változatának pszichometriai mutatói megfelelőek. Javasoljuk a mérőeszköz hazai kutatásba történő bevezetését és további vizsgálatát. | Background: The Children’s Perception of Interparental Conflict Scale is a widely used measure for assessing perceived interparental conflicts and children’s subsequent adjustment. Aim: The aims of this study were to prepare the Hungarian adaptation and evaluate the psychometric properties of the Hungarian version of Children’s Perception of Interparental Conflict Scale. Method: 143 child-parent pairs participated in this cross-sectional questionnaire study. Children between the ages of 9—12 years (mean of age 10.8 years, SD = 1.05 years, range: 9—12 years) completed the CPIC, anxiety (STAI-C) and depression (CDI) scales, whereas the parent’s battery of tests contained the short version of the Marital Stress Scale (MSS). Results: The results of the confirmatory factor analysis did not support the theoretical eight-factor structure of the CPIC (χ2 = 1355.0, DF = 1052, p90: .040—.055, p = .698). The model fit indices became acceptable after the deletion of three items of the Triangulation subscale: (χ2(917) = 1113.4, p90: .032—.049, p = .960). The four-factor alternative theoretical model showed significantly worse fit than the eight-factor model (Δχ2 = 66.5, Δdf = 22, p<.001). The internal consistency of the CPIC was acceptable (Cronbach-alpha: .63—.81) except the Triangulation subscale (Cronbach-alpha: .40). Construct validity was supported by the expected association in the case of six subscales with depression, anxiety and the self-reported marital stress. Conclusions: Although the number of participants in the present study was suboptimal, the Hungarian version of the Children’s Perception of Interparental Conflict Scale seemed to have adequate psychometric properties. We recommend the introduction of this scale to the Hungarian research and its further investigation

STUDIES ON THE EFFECT OF PROPIONIBACTERIUM ACNES ON THE BARRIER PROPERTIES OF HUMAN IN VITRO CULTURED KERATINOCYTES

The human skin is heavily colonized by a specialized microbial community called the microbiome, which plays a complex role in the protection from the attack of external pathogens. This microbial flora can interact with the cells in the healthy skin and play a role in the maintenance of skin homeostasis, but also known to contribute to the pathogenesis of different diseases. Our aim was to analyze whether the Propionibacterium acnes (P. acnes) bacterium, a member of the skin microbiome, or the bacterium induced pro-inflammatory mediator, TNF has any effect on the barrier properties of our epidermis. For that, a confluent monolayer of in vitro cultured human immortalized keratinocytes (HPV-KER cells) were treated with different P.acnes strains and external TNF in different doses, and changes in the barrier properties were analyzed in real time using the xCELLIgence system. We also analyzed the effect of the bacterium on the mRNA expression changes of tight junction proteins claudin 1, 2, 4 (CLDN1, 2, 4), ocludin1 (OCL1) and ZO1 in these cultures using real-time RT-PCR. Our results suggest that the bacterium induced an elevation, followed by a drop of the measured impedance values in the keratinocyte monolayers, possibly due to dynamic alterations of the barrier properties. The extent of these changes depended on the used P. acnes strain and the applied doses. Addition of TNF (1, 5, 10 ng/ml), a cytokine that is a know mediator of the P. acnes-induced innate immune and inflammatory events in keratinocytes also lead to a marked decrease of the measured impedance of the HPV-KER monolayers. Real-time RT-PCR analysis of tight junction genes suggested that CLDN2 and 4 mRNAs were not present in these cells. However, the expression of CLDN1 decreased, whereas ZO1 and OCL1 mRNA levels increased in response to the bacterial treatment. Our results suggest that our microbiome can modulate the barrier properties of the epidermis. It is possibly achieved, in one hand, through the direct regulation of genes playing a key role in the formation of cell-to-cell contacts. On the other hand, secreted factors, such as the TNFα pro-inflammatory mediator, may also have a direct effect and can loosen the epidermal barrier, possibly leading to the easier penetration of keratinocyte- as well as bacterial-derived factors to deeper tissue compartments. These findings strengthen the importance of a balanced interaction among the epidermal cells and our microbiome for the maintenance of healthy skin functions

PROPIONIC ACID SECRETED BY PROPIONIBACTERIUM ACNES MAY MODIFY THE CELLULAR PROPERTIES OF KERATINOCYTES

Propionibacterium acnes (P. acnes) bacterium is a member of the skin microflora, but may also serve as an opportunistic pathogen contributing to the pathogenesis of acne vulgaris. Earlier we have shown that various P. acnes strains (889, 6609, ATCC 11828) belonging to different phylogroups differentially affect the cellular properties of cultured human keratinocytes in a strain-specific and dose-dependent manner. High doses of the pathogenic 889 and ATCC 11828 strains also resulted characteristic morphological changes and membrane damage, which lead to the cytotoxicity of human in vitro cultures keratinocytes (HPV-KER). Our aim was to further analyze the interaction of human in vitro cultured keratinocyes and identify bacterially-derived factors that may mediate the previously observed effects. In order to systematically quantify the P. acnes-induced cytotoxicity we performed spectrophotometric lactate dehydrogenase (LDH) and hemoglobin (HgB) assays using supernatant samples of bacterial treated HPV-KER cells and erythrocytes. The amount of released free LDH and HgB exhibited strain- and dose-dependent differences. We also noted the differential acidification of the pH in the culture supernatants. P. acnes is known to secrete propionic acid (PA), a characteristic, acidic end-product of bacterial fermentation in these species. In order to analyze whether P. acnes-derived PA has any role in the observed cellular changes we treated HPV KER cells with the acid and analyzed the cell morphology. Microscopic analysis of the PA treated cultures revealed cells with similar irregular membrane morphologies observed earlier upon high dose P. acnes 889 and ATCC 11828 treatments. Finally, we measured the amount of secreted short chain fatty acids (SCFA) in the P. acnes 889, 6609 and ATCC 11828-treated HPV-KER supernatant samples by mass spectrometry. These studies revealed marked differences in the amount of secreted PA; high dose treatment of the 889 and ATCC 11828 strains leading to higher levels. P. acnes-induced cellular changes depend on the type and amount of the applied bacterial strains. The observed differences may be due to variations of the amount of a secreted metabolic end-product, PA. Together with other bacterially-derived molecules it may be an active contributor of the P. acnes-induced cellular changes

Identification and antimicrobial susceptibility testing of anaerobic bacteria: Rubik's cube of clinical microbiology?

Anaerobic bacteria have pivotal roles in the microbiota of humans and they are significant infectious agents involved in many pathological processes, both in immunocompetent and immunocompromised individuals. Their isolation, cultivation and correct identification differs significantly from the workup of aerobic species, although the use of new technologies (e.g., matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, whole genome sequencing) changed anaerobic diagnostics dramatically. In the past, antimicrobial susceptibility of these microorganisms showed predictable patterns and empirical therapy could be safely administered but recently a steady and clear increase in the resistance for several important drugs (beta-lactams, clindamycin) has been observed worldwide. For this reason, antimicrobial susceptibility testing of anaerobic isolates for surveillance purposes or otherwise is of paramount importance but the availability of these testing methods is usually limited. In this present review, our aim was to give an overview of the methods currently available for the identification (using phenotypic characteristics, biochemical testing, gas-liquid chromatography, MALDI-TOF MS and WGS) and antimicrobial susceptibility testing (agar dilution, broth microdilution, disk diffusion, gradient tests, automated systems, phenotypic and molecular resistance detection techniques) of anaerobes, when should these methods be used and what are the recent developments in resistance patterns of anaerobic bacteria