Tissue gene expression profiles and communication networks inform candidate blood biomarker identification in psoriasis and atopic dermatitis.

Overlapping clinical and pathomechanistic features can complicate the diagnosis and treatment of inflammatory skin diseases, including psoriasis and atopic dermatitis (AD). Spatial transcriptomics allows the identification of disease- and cell-specific molecular signatures that may advance biomarker development and future treatments. This study identified transcriptional signatures in keratinocytes and sub-basal CD4+ and CD8+ T lymphocytes from patients with psoriasis and AD. In silico prediction of ligand:receptor interactions delivered key signalling pathways (interferon, effector T cells, stroma cell and matrix biology, neuronal development, etc.). Targeted validation of selected transcripts, including CCL22, RELB, and JUND, in peripheral blood T cells suggests the chosen approach as a promising tool also in other inflammatory diseases. Psoriasis and AD are characterized by transcriptional dysregulation in T cells and keratinocytes that may be targeted therapeutically. Spatial transcriptomics is a valuable tool in the search for molecular signatures that can be used as biomarkers and/or therapeutic targets.

Biodegradable and adjustable sol-gel glass based hybrid scaffolds from multi-armed oligomeric building blocks.

Biodegradability is a crucial characteristic to improve the clinical potential of sol-gel-derived glass materials. To this end, a set of degradable organic/inorganic class II hybrids from a tetraethoxysilane(TEOS)-derived silica sol and oligovalent cross-linker oligomers containing oligo(d,l-lactide) domains was developed and characterized. A series of 18 oligomers (Mn: 1100-3200Da) with different degrees of ethoxylation and varying length of oligoester units was established and chemical composition was determined. Applicability of an established indirect rapid prototyping method enabled fabrication of a total of 85 different hybrid scaffold formulations from 3-isocyanatopropyltriethoxysilane-functionalized macromers. In vitro degradation was analyzed over 12months and a continuous linear weight loss (0.2-0.5wt%/d) combined with only moderate material swelling was detected which was controlled by oligo(lactide) content and matrix hydrophilicity. Compressive strength (2-30MPa) and compressive modulus (44-716MPa) were determined and total content, oligo(ethylene oxide) content, oligo(lactide) content and molecular weight of the oligomeric cross-linkers as well as material porosity were identified as the main factors determining hybrid mechanics. Cytocompatibility was assessed by cell culture experiments with human adipose tissue-derived stem cells (hASC). Cell migration into the entire scaffold pore network was indicated and continuous proliferation over 14days was found. ALP activity linearly increased over 2weeks indicating osteogenic differentiation. The presented glass-based hybrid concept with precisely adjustable material properties holds promise for regenerative purposes. STATEMENT OF SIGNIFICANCE: Adaption of degradation kinetics toward physiological relevance is still an unmet challenge of (bio-)glass engineering. We therefore present a glass-derived hybrid material with adjustable degradation. A flexible design concept based on degradable multi-armed oligomers was combined with an established indirect rapid prototyping method to produce a systematic set of porous sol-gel-derived class II hybrid scaffolds. Mechanical properties in the range of cancellous bone were narrowly controlled by hybrid composition. The oligoester introduction resulted in significantly increased compressive moduli. Cytocompatible hybrids degraded in physiologically relevant time frames and a promising linear and controllable weight loss profile was found. To our knowledge, our degradation study represents the most extensive long-term investigation of sol-gel-derived class II hybrids. Due to the broad adjustability of material properties, our concept offers potential for engineering of biodegradable hybrid materials for versatile applications.

Indirect rapid prototyping of sol-gel hybrid glass scaffolds for bone regeneration - Effects of organic crosslinker valence, content and molecular weight on mechanical properties.

We present a series of organic/inorganic hybrid sol-gel derived glasses, made from a tetraethoxysilane-derived silica sol (100% SiO2) and oligovalent organic crosslinkers functionalized with 3-isocyanatopropyltriethoxysilane. The material was susceptible to heat sterilization. The hybrids were processed into pore-interconnected scaffolds by an indirect rapid prototyping method, described here for the first time for sol-gel glass materials. A large panel of polyethylene oxide-derived 2- to 4-armed crosslinkers of molecular weights ranging between 170 and 8000Da were incorporated and their effect on scaffold mechanical properties was investigated. By multiple linear regression, 'organic content' and the 'content of ethylene oxide units in the hybrid' were identified as the main factors that determined compressive strength and modulus, respectively. In general, 3- and 4-armed crosslinkers performed better than linear molecules. Compression tests and cell culture experiments with osteoblast-like SaOS-2 cells showed that macroporous scaffolds can be produced with compressive strengths of up to 33±2MPa and with a pore structure that allows cells to grow deep into the scaffolds and form mineral deposits. Compressive moduli between 27±7MPa and 568±98MPa were obtained depending on the hybrid composition and problems associated with the inherent brittleness of sol-gel glass materials could be overcome. SaOS-2 cells showed cytocompatibility on hybrid glass scaffolds and mineral accumulation started as early as day 7. On day 14, we also found mineral accumulation on control hybrid glass scaffolds without cells, indicating a positive effect of the hybrid glass on mineral accumulation. STATEMENT OF SIGNIFICANCE: We produced a hybrid sol-gel glass material with significantly improved mechanical properties towards an application in bone regeneration and processed the material into macroporous scaffolds of controlled architecture by indirect rapid prototyping. We were able to produce macroporous materials of relevant porosity and pore size with compressive moduli, covering the range reported for cancellous bone while an even higher compressive strength was maintained. By multiple linear regression, we identified crosslinker parameters, namely organic content and the content of ethylene oxide units in the hybrids that predominantly determined the mechanics of the hybrid materials. The scaffolds proved to be cytocompatible and induced mineralization in SaOS-2 cells. This provides new insight on the critical parameters for the design of the organic components of covalent hybrid sol-gel glasses.

Changes in platelet morphology and function during 24 hours of storage.

For in vitro studies assessing the interaction of platelets with implant materials, common and standardized protocols for the preparation of platelet rich plasma (PRP) are lacking, which may lead to non-matching results due to the diversity of applied protocols. Particularly, the aging of platelets during prolonged preparation and storage times is discussed to lead to an underestimation of the material thrombogenicity. Here, we study the influence of whole blood- and PRP-storage times on changes in platelet morphology and function. Blood from apparently healthy subjects was collected according to a standardized protocol and examined immediately after blood collection, four hours and twenty four hours later. The capability of platelets to adhere and form stable aggregates (PFA100, closure time) was examined in sodium citrate anticoagulated whole blood (WB) using the agonists equine type I collagen and epinephrine bitartrate (collagen/epinephrine) as well as equine type I collagen and adenosine-5'-diphosphate (collagen/ADP). Circulating platelets were quantified at each time point. Morphology of platelets and platelet aggregates were visualized microscopically and measured using an electric field multi-channel counting system (CASY). The percentage of activated platelets was assessed by means of P-selectin (CD62P) expression of circulating platelets. Furthermore, platelet factor 4 (PF4) release was measured in platelet poor plasma (PPP) at each time point. Whole blood PFA100 closure times increased after stimulation with collagen/ADP and collagen/epinephrine. Twenty four hours after blood collection, both parameters were prolonged pathologically above the upper limit of the reference range. Numbers of circulating platelets, measured in PRP, decreased after four hours, but no longer after twenty four hours. Mean platelet volumes (MPV) and platelet large cell ratios (P-LCR, 12 fL - 40 fL) decreased over time. Immediately after blood collection, no debris or platelet aggregates could be visualized microscopically. After four hours, first debris and very small aggregates occurred. After 24 hours, platelet aggregates and also debris progressively increased. In accordance to this, the CASY system revealed an increase of platelet aggregates (up to 90 µm diameter) with increasing storage time. The percentage of CD62P positive platelets and PF4 increased significantly with storage time in resting PRP. When soluble ADP was added to stored PRP samples, the number of activatable platelets decreased significantly over storage time. The present study reveals the importance of a consequent standardization in the preparation of WB and PRP. Platelet morphology and function, particularly platelet reactivity to adherent or soluble agonists in their surrounding milieu, changed rapidly outside the vascular system. This knowledge is of crucial interest, particularly in the field of biomaterial development for cardiovascular applications, and may help to define common standards in the in vitro hemocompatibility testing of biomaterials.

[Bullous pemphigoid]. / Bullöses Pemphigoid.

Pemphigoid diseases are a group of autoimmune disorders characterized by subepidermal blistering and autoantibodies against structural proteins of the dermal-epidermal junction. In bullous pemphigoid, the most common subepidermal blistering autoimmune disease, antibodies are directed against the hemidesmosomal antigens BP180 (collagen type XVII) and BP230. Bullous pemphigoid typically presents with severe pruritus and tense blisters accompanied by erosions and crusts in elderly patients. Diagnostic landmarks are the detection of linear IgG and/or C3 deposits at the dermo-epidermal junction by direct immunofluorescence microscopy of a perilesional biopsy and the detection of serum autoantibodies by indirect immunofluorescence microscopy on human salt-split skin and ELISA employing recombinant immunodominant fragments of BP180 and BP230. Treatment options include topical (class IV) and/or systemic corticosteroids, frequently combined with immunomodulatory agents like dapsone and tetracyclines or immunosuppressants such as methotrexate and azathioprine.

Herd-level prevalence of Map infection in dairy herds of southern Chile determined by culture of environmental fecal samples and bulk-tank milk qPCR.

Paratuberculosis, an infectious disease of domestic and wild ruminants caused by Mycobacterium avium subsp. paratuberculosis (Map), is an economically important disease in dairy herds worldwide. In Chile the disease has been reported in domestic and wildlife animals. However, accurate and updated estimations of the herd-prevalence in cattle at national or regional level are not available. The objectives of this study were to determine the herd-level prevalence of dairy herds with Map infected animals of Southern Chile, based on two diagnostic tests: culture of environmental fecal samples and bulk-tank milk qPCR. Two composite environmental fecal samples and one bulk-tank milk sample were collected during September 2010 and September 2011 from 150 dairy farms in Southern Chile. Isolation of Map from environmental fecal samples was done by culture of decontaminated samples on a commercial Herrold's Egg Yolk Medium (HEYM) with and without mycobactin J. Suspicious colonies were confirmed to be Map by conventional IS900 PCR. Map detection in bulk-tank milk samples was done by real time IS900 PCR assay. PCR-confirmed Map was isolated from 58 (19.3%) of 300 environmental fecal samples. Holding pens and manure storage lagoons were the two more frequent sites found positive for Map, representing 35% and 33% of total positive samples, respectively. However, parlor exits and cow alleyways were the two sites with the highest proportion of positive samples (40% and 32%, respectively). Herd prevalence based on environmental fecal culture was 27% (true prevalence 44%) compared to 49% (true prevalence 87%) based on bulk-tank milk real time IS900 PC. In both cases herd prevalence was higher in large herds (>200 cows). These results confirm that Map infection is wide spread in dairy herds in Southern Chile with a rough herd-level prevalence of 28-100% depending on the herd size, and that IS900 PCR on bulk-tank milk samples is more sensitive than environmental fecal culture to detect Map-infected dairy herds.

[Value of one-stage surgical treatment of diffuse peritonitis (relaparotomy on demand): a single-center analysis]. / Stellenwert der einzeitigen chirurgischen Therapie bei diffuser Peritonitis (Relaparotomie "on demand"): eine unizentrische Analyse.

BACKGROUND: Despite modern surgical and intensive-care concepts, diffuse peritonitis remains a major source of high morbidity and mortality. The aim of this study was to critically evaluate the value of relaparotomy on demand (ROD) for patients with diffuse peritonitis. PATIENTS AND RESULTS: In a retrospective analysis, the clinical course of 231 patients with diffuse peritonitis was analysed. The mean Mannheim Peritonitis Index (MPI) was 25.3. Overall lethality in this cohort was 14.3 %. In 214 patients, source control was successful during the index operation, and these patients were treated according to an on-demand strategy. For 178 of these patients, there was no demand for a relaparotomy, whereas 36 of these patients required further surgical interventions. Lethality for these subgroups was 9 % (no relaparotomy) and 27 % (relaparotomy), respectively. CONCLUSIONS: This retrospective analysis confirms that an on-demand strategy is reasonable and feasible after successful source control and lavage. However, it still remains of clinical importance to identify parameters that may assist in selecting those patients who require a relaparotomy.

Acoustic radiation force impulse imaging for non-invasive assessment of liver fibrosis in chronic hepatitis B.

Acoustic radiation force impulse (ARFI) imaging is a novel ultrasound-based elastography method that is integrated in a conventional ultrasound machine. It might provide an alternative method to transient elastography for the noninvasive assessment of liver fibrosis. While previous studies have shown comparable diagnostic accuracy of ARFI to transient elastography in chronic hepatitis C, the aim of the present prospective multicenter study was to evaluate ARFI for the assessment of liver fibrosis in chronic hepatitis B. ARFI imaging involves the mechanical excitation of tissue using short-duration acoustic pulses to generate localized displacements in tissue. The displacements result in shear-wave propagation which is tracked using ultrasonic, correlation-based methods and recorded in m/s. In the present international prospective study, patients infected with chronic hepatitis B received ARFI imaging, blood tests and if available transient elastography. The results were compared to liver biopsy as reference method analysed by a central pathologist. In 92 of 114 patients, a comparison of ARFI with transient elastography was possible. ARFI imaging and transient elastography correlated significantly with histological fibrosis stage. The diagnostic accuracy expressed as areas under ROC curves for ARFI imaging and transient elastography was 0.75 and 0.83 for the diagnosis of significant fibrosis (F ≥ 2), 0.93 and 0.94 for the diagnosis of severe fibrosis (F ≥ 3), and 0.97 and 0.93 for the diagnosis of liver cirrhosis, respectively. No significant difference was found between ARFI and transient elastography. ARFI imaging is a reliable ultrasound-based method for the assessment of advanced stages of liver fibrosis in chronic hepatitis B.

[Extramural Gastral GIST Manifested by Spontaneous Perforation with Acute Intraabdominal Bleeding and Haemoperitoneum]. / Extramuraler GIST des Magens mit Manifestation durch Spontanruptur und akuter intraabdomineller Blutung mit Hämatoperitoneum.

We report on the case of a 38-year-old male patient with a huge extramural gastrointestinal stromal tumour (GIST) of the stomach, located in the left upper and middle abdominal cavity that was diagnosed on the basis of a spontaneous -rupture and consecutive haemoperitoneum. The lesion was resected completely in an emergency operation. The tumour was classified as a high-risk lesion for aggressive biological behaviour and with regard to tumour rupture with perforation of the serosa, an adjuvant systemic therapy was indicated.

Analysis of microsatellite instability in colorectal carcinoma by microfluidic-based chip electrophoresis.

Microsatellite analysis is an important tool in clinical research and molecular diagnostics because microsatellite instability (MSI) occurs frequently in various types of cancer. Approximately 10-15% of colorectal, gastric and endometrial carcinomas are associated with MSI, and this has an impact on clinical prognosis. The microsatellite loci Bat25, Bat26, D2S123, D5S346 and D17S250, recommended by the Bethesda guidelines, were analysed by microfluidic-based on-chip electrophoresis in 40 cases of colon carcinoma with known MSI status. In all cases, microfluidic separation of the PCR amplicons resulted in highly resolved, distinct patterns of each of the five microsatellite loci. Detection of MSI could be demonstrated by microsatellite-loci-associated, well-defined deviations in the electropherogram profiles of tumour and non-tumour material, and confirmed the classification of MSI cases performed by conventional technology. In conclusion, microfluidic chip technology is a simple and reliable approach for MSI detection that allows label-free and very fast analysis of microsatellite amplicons.

Genomic and phenotypic changes of Campylobacter jejuni strains after passage of the chicken gut.

The ability to colonize the chicken gut was determined for 17 Campylobacter jejuni strains of human and bovine origin. The level of colonization varied according to the strain used for experimental infection. Two Campylobacter isolates from patients suffering from gastroenteritis were found in the group of non-colonizing strains, suggesting that other reservoirs as poultry are also important sources of human Campylobacter infections. Bovine Campylobacter isolates can also effective colonize the chicken intestine and may be a source for poultry infection. The invasion ability of the strains as determined in the cell culture model using Caco-2 cells correlates with their colonization capacity in the chicken gut. The genomic and phenotypic stability of the selected strains were evaluated by analysis of their pulsed-field gel electrophoresis (PFGE) patterns, flaA-typing and in vitro determination of motility, adhesion and invasion abilities after colonizing chickens for up to 21 days. Changes were identified in flaA-types of six isolates and three isolates from chicken showed different patterns by PFGE using SmaI or KpnI as restriction enzymes. One isolate showed phenotypic differences after in vivo passage which were seen in enhancement of adherence to eukaryotic cells, decrease of motility and changes in morphology. These phenotypic changes were not associated with the observed genomic instabilities.

Asymmetric dimethylarginine is associated with parameters of glucose metabolism in Caucasian but not in African women from South Africa.

AIM: Ethnic differences in obesity and obesity related disorders prompted us to search for possible contributors. The impact of the novel cardiovascular risk factor asymmetric dimethylarginine (ADMA) has been never determined in the African population. The present observational study aimed to compare ADMA levels between healthy African (102) and Caucasian women (115) from South Africa, and its impact on glucose metabolism. METHODS: All participants underwent an oral glucose tolerance test with measurements of glucose, insulin, C-peptide, proinsulin and free fatty acids before and after 30, 60, 90, 120 minutes. Fasting serum ADMA was measured by ELISA assay and obesity was determined by anthropometry. RESULTS: Serum ADMA did not differ between the ethnic groups. After stratification according to ADMA quartiles Caucasian women in the upper quartile had significantly higher body mass index and waist circumference as well as elevated insulin resistance, insulin, C-peptide and proinsulin levels with no differences in serum glucose compared to women in the lowest quartile. There was a significant stronger postchallenge insulin response in Caucasian women of the upper quartile. No differences were found in African women. CONCLUSIONS: Despite similar ADMA levels in both ethnic groups ADMA was positively correlated with parameters of glucose metabolism in the Caucasian but not in the African women from South Africa.

[Epithelial-mesenchymal transition of biliary epithelial cells in advanced liver fibrosis]. / Epitheliale-mesenchymale Transdifferenzierung von Gallengangsepithelzellen bei fortgeschrittener Leberfibrose.

UNLABELLED: The conversion of epithelial cells in a mesenchymal cell type is called "epithelial-mesenchymal-transition" (EMT). This process is defined by a loss of epithelial specific characteristics such as cell adhesion, polarity and a reorganization of cytoskeletal proteins. EMT has been shown to be involved in progression of cancer and in obstructive renal fibrosis. In this study we analyzed liver tissues in a bile-duct ligation model of rats and human liver biopsies with cholestatic fibrosis and chronic hepatitis c infection to determine if biliary epithelial cells undergo phenotypical and functional changes during chronic injury. METHODS: Liver tissue of rats and human patients was examined by immunohistochemistry using antibodies against epithelial and mesenchymal specific targets as well as molecules of potentially activated signaling pathways. To study contribution of biliary epithelial cells in extracellular matrix production we performed laser microdissection combined with real-time PCR. RESULTS: Bile duct ligation in rats induced a prominent biliary epithelial proliferation and a pronounced expression of vimentin was observed in biliary epithelial cells, whereas no vimentin expression was detectable in bile duct cells of sham operated rats. In human liver biopsies from patients with cholestatic fibrosis and chronic hepatitis c infection a prominent biliary expression of vimentin could be shown. Despite this, epithelial marker proteins were still detectable. Further, we observed collagen I mRNA expression in laser microdissected bile ducts. CONCLUSION: Biliary epithelial cells show cytoskeletal rearrangements during chronic liver injury towards a mesenchymal phenotype. The detection of collagen I mRNA in bile duct cells suggests that they might participate in extracellular matrix production.

Development of a cell culture assay for the quantitative determination of vaccination-induced antibodies in rabbit sera against Clostridium perfringens epsilon toxin and Clostridium novyi alpha toxin.

Cell culture assays are possible alternatives to replace in vivo neutralization tests currently required for potency testing of clostridial vaccines. Cell culture assays based on the MDCK cell line and the Vero cell line which are sensitive to the Clostridium (C.) perfringens type D epsilon toxin and Clostridium novyi type B alpha toxin, respectively, were developed, and the test conditions were standardized. The antibody titres of vaccinated rabbits measured in vitro were compared with the results of current test procedures recommended by European Pharmacopoeia. The correlation coefficients calculated were significant for all sera tested. The cell culture assays proved to be sensitive, specific, reproducible and reliable. Therefore, these cell culture assays could be suitable in vitro alternatives to the in vivo mouse neutralization experiments required for potency tests of clostridial vaccines, but further validation studies are necessary.

PCR detection of virulence-associated genes in Campylobacter jejuni strains with differential ability to invade Caco-2 cells and to colonize the chick gut.

In this study, the presence of 20 putative virulence genes was examined in 11 Campylobacter jejuni isolates with different colonization and invasion abilities as determined in a chick colonization model and on Caco-2 cells, respectively. The majority of the genes were detected in all strains. Among them, there were genes of the flagellar secretion apparatus like flhA, flhB, flgB, flgE2, the flagellin genes flaA and flaB, invasion-associated genes like ciaB and iamA, the cytotoxin genes cdtA-C, the adhesion related gene cadF, and some genes involved in the colonization process (docA, docB). The plasmid gene virB11 could not be detected in any strain. Specific differences between the isolates were observed only in genes cgtB and wlaN involved in lipo-oligosaccharide (LOS) biosynthesis. The gene cgtB was only detectable in three of five strains with strong colonization and invasion abilities. Probably, wlaN can overcome the lack of cgtB in the two cgtB- isolates.

Determination of a reference value for N(G), N(G)-dimethyl-L-arginine in 500 subjects.

BACKGROUND: Asymmetric dimethylarginine (ADMA) acts as an endogenous inhibitor of NO-synthase. In the last years ADMA has emerged as a cardiovascular risk factor. The aim of this study was to determine a reference value for ADMA. METHODS: Plasma samples of 500 healthy subjects in the 19-75 year age group were analyzed. Exclusion criteria from this study were smoking, any known significant disease, body-mass-index (BMI) above 30 kg m(-2), elevated plasma lipid levels, impaired renal function, hypertension, and intake of any medication. The ADMA levels were determined by ELISA, (DLD Diagnostics, Hamburg, Germany). RESULTS: Mean ADMA plasma concentration of the total population was 0.69 micromol L(-1) (SD 0.20) and 95% of the measured values were in the range from 0.36 micromol L(-1) to 1.17 micromol L(-1). Women below 50 years of age had lower ADMA levels than men below 50 years of age [0.62 (0.17) micromol L(-1) vs. 0.69 (0.19) micromol L(-1); P = 0.001] and woman above 50 years of age had higher ADMA levels than men above 50 years of age [0.80 (0.22) micromol L(-1) vs. 0.73 (0.20) micromol L(-1); P = 0.036]. A regression analysis of ADMA levels and age was performed for each sex. The regression factor was r = 0.444 for women in a squared regression model (P < 0.001) and r = 0.212 for men in a linear regression model (P < 0.001). CONCLUSION: The study was able to define a reference value for ADMA plasma levels with 0.36-1.17 micromol L(-1) and found sex dependent correlations between ADMA and age. Women showed a significant increase in ADMA plasma levels with onset of menopause.

Applications of Artificial Neural Networks in integrated water management: fiction or future?

An Artificial Neural Network (ANN) is nowadays recognized as a very promising tool for relating input data to output data. It is said that the possibilities of artificial neural networks are unlimited. Here we focus on the potential role of neural networks in integrated water management. An Artificial Neural Network (ANN) is a mathematical methodology which describes relations between cause (input data) and effects (output data) irrespective of the process laying behind and without the need for making assumptions considering the nature of the relations. The applications are widespread and vary from optimization of measuring networks, operational water management, prediction of drinking water consumption, on-line steering of wastewater treatment plants and sewage systems, up to more specific applications such as establishing a relationship between the observed erosion of groyne field sediments and the characteristics of passing vessels on the river Rhine. Especially where processes are complex, neural networks can open new possibilities for understanding and modelling these kinds of complex processes. Besides explaining the method of ANN this paper shows different applications. Three examples have been worked out in more detail. An intelligent monitoring system is shown for the on-line prediction of water consumption, ANN are successfully used for sludge cost monitoring and optimizing wastewater treatment and the usage of ANN is shown in optimizing and monitoring water quality measuring networks. An ANN appears to be a multiuse and powerful tool for modelling complex processes.

Correlation between invasion of Caco-2 eukaryotic cells and colonization ability in the chick gut in Campylobacter jejuni.

In an in vitro cell culture model using Caco-2 cells the adhesion and invasion properties of 11 Campylobacter (C.) jejuni isolates of different origin were studied. Additionally, we investigated the colonization ability of the strains in a chick model. Virtually, all C. jejuni showed cell adherence in the in vitro assay, but there were large differences in the invasion frequencies among the Campylobacter isolates. The colonization ability in the chick gut also differed markedly and enabled the formation of three groups: non-colonizing, weak or delayed colonization and strong colonization ability. On this occasion, we found a putative correlation between invasion of Caco-2 cells and colonization in the chick gut. Non-colonizers are not invasive or only have small invasion indexes. Strains which colonize weakly or exhibit delayed colonization have a medium invasion index and strong colonizers show markedly higher values of this parameter. The characterization of the flagellin gene of the used C. jejuni strains resulted in eight flaA types. There was no association between flaA type and invasion or colonization ability in the chick gut.

[Identification and differentiation of Campylobacter fetus subspecies by PCR]. / Identifizierung und Differenzierung der Campylobacter-fetus-Subspezies mittels PCR.

The species Campylobacter (C.) fetus is divided into the subspecies venerealis and fetus, which differ in epidemiology and clinical importance. The differences between these subspecies make an accurate distinction essential. Differentiation of C. fetus by traditional microbiological methods is only based on two reactions (tolerance to glycin, Na selenite reduction), in which C. fetus ssp. venerealis reacts negatively. However, the value of both reactions is limited. We used a specific PCR-based assay for identifying and differentiating the two C. fetus subspecies, which was recently developed by HUM et al. (1997). In this assay, a 764 bp amplicon is produced using primers MG3F and MG4R for both subspecies of C. fetus. In contrast to HUM et al. (1997), this amplicon was approximately 200 bp smaller. This discrepancy can't be explained. Afterwards, the primers VenSF and VenSR are used for differentiation. The identification of the sub-species venerealis is based on the presence of a 142 bp amplicon, which is not formed with subspecies fetus. The type strains of both C. fetus subspecies were used as positive controls. Non-specific reactions were not observed. In this PCR assay, 73 field strains were investigated (among them 24 C. fetus ssp. veneralis, 26 C. fetus ssp. fetus). In these investigations, the method has proved its diagnostic suitability. The results of the traditional microbiological differentiation of the C. fetus field strains could be confirmed by the PCR assay. In future, the traditional phenotypic characterization of C. fetus subspecies remains indispensable, but this PCR assay constitutes a valuable method for the confirmation of these results.