Biomedical subjects

H Schmidt

Publications and source records attributed to H Schmidt.

At least 433 records · Page 24Linked to original sources

PubMed1996

Human central neurocytoma cells show neuronal physiological properties in vitro.

Central neurocytoma is a rare brain tumor composed of small round synaptophysin-positive cells, suggesting a neuronal origin of these tumor cells. Glial properties are inferred, however, from the observation that the tumor cells exhibit a strong morphological similarity to oligodendroglioma cells and show an astrocytic differentiation in vitro. To test for neuronal or glial physiological properties, we studied cultured neurocytoma cells derived from a surgical specimen from a 44-year-old man, employing the patch-clamp technique. Early primary cultures were composed of morphologically unique bi- or multipolar cells which were positive for synaptophysin and negative for the astrocyte marker glial fibrillary acidic protein. In the majority of these cells, whole-cell membrane current recordings revealed physiological properties of neurons, i.e., a high density of Na+ currents, the capacity to generate action potentials, and the expression of inotropic neurotransmitter receptors. Metabotropic neurotransmitter receptors could be demonstrated by Ca2+ imaging techniques. The remaining bi- or multipolar cells and almost all cells in later culture stages and in vitro passage lacked these neuronal properties and showed physiological features characteristic of glial cells. We conclude that the major population of neurocytoma cells shows physiological properties of neurons and that with time in culture this population is replaced by electrically passive cells.

Action Potentials↗

PubMed1996

Caffeine-evoked contractures in single slow (tonic) muscle fibres of the frog (Rana temporaria and R. esculenta).

Single slow (tonic) muscle fibres were dissected from cruralis muscles of Rana temporaria and R. esculenta. Increasing concentrations of caffeine were applied in Ringer solution, and contractures were measured isometrically. Sigmoid caffeine concentration-response curves were obtained, the threshold value being near 1.2 mmol/l, and maximum contractures being obtained with 10 to 20 mmol/l concentrations of caffeine. Contracture solutions were modified by varying the Ca2+ concentration or by replacing Ca2+ with 1.8 mmol/l Mg2+, Ni2+, Co2+ or with 0.1-5.0 mmol/l La3+. The effects of low pH (5.3), K+ (6,10 and 95 mmol/l), adenosine (10 mmol/l) and gallopamil (D600; 30 micromol/l) were examined too. The caffeine threshold was lowered by Mg2+, K+, 0 .1 mmol/l La3+ and D600, while all other substances including 0.5-5.0 mmol/l La3+ increased it. The amplitude of contractures evoked by high caffeine concentrations was unaffected. Caffeine (1-40 mmol/l) was also pressure injected into slow fibres. The composition of the solution was modified in a number of ways, but a contractile response was not observed or measured. Extracellular application of caffeine from the same pipettes evoked local contractures. Similar injection experiments in twitch fibres revealed the same results. These observations suggest that an extracellular binding site seems to be involved in the initiation of caffeine-evoked contractures in intact frog muscle fibres. Possible reasons for the ineffectiveness of intracellular caffeine are discussed.

Adenosine↗

PubMed1996

The control of ichthyophthiriasis by a medicated food containing quinine: efficacy tests and ultrastructure investigations.

The present report demonstrates that a medicated food containing quinine kills the skin-inhabiting trophozoite stage of Ichthyophthirius multifiliis in ornamental fish. Artificially infected swordtails. (Xiphophorus helleri), black mollies (Poecilia sphenops), and black neons (Hyphessobrycon herbertaxelrodi) were used in the trials. The fish were maintained in groups of 10 or 20 inside aquaria (20 or 60 l) at 25 degrees C. Ultrastructure investigations by means of transmission electron microscopy revealed clear deleterious effects of quinine on the trophozoite stages. Following the initial application the outer limiting membrane of the trophozoite was broken at places. Within the nephridial plasma the plasma bridges were broken in part. After 2 days of treatment the lumen of the alveolar sac became enlarged. The food vacuoles in treated trophozoites were more electron-dense than those in the untreated controls. Numerous lipid droplets were found close to the vacuoles. The degree of damage in the nephridial plasma was intensified. When feeding was prolonged for 3 or more days, all kinds of damage became more extensive as seen in the trophozoites after 1 or 2 days of treatment. In addition food vacuoles in the final stages of digestion were no longer detectable. In long-term feeding tests, when typical ornamental fish species were fed three times daily ad libitum with the medicated food over a 12-week period, the animals showed no adverse clinical symptom. From the toxicological as well as the ecological and economical point of view the feeding of flakes containing quinine has considerable advantages as compared with conventional bath treatment.

Administration, Oral↗

PubMed1996

Autotransfusion after coronary artery bypass grafting halves the number of patients needing blood transfusion.

BACKGROUND: Several randomized studies about autotransfusion of shed mediastinal blood in patients undergoing coronary artery bypass grafting have resulted in divergent findings concerning reduction of the need for homologous blood transfusions. Most of these studies used less strict criteria for homologous blood transfusion than applied in daily clinical practice. METHODS: A prospective, randomized, controlled study involving 120 patients having elective, uncomplicated coronary artery bypass grafting was performed. The autotransfusion group received transfusion of shed mediastinal blood for 18 hours. Criteria for homologous blood transfusion were hemoglobin concentration less than 5.0 mmol/L in the intensive care unit and less than 5.5 mmol/L during the rest of the hospital stay. RESULTS: Twenty-eight percent of patients in the autotransfusion group received homologous blood transfusion versus 55% in the control group (p = 0.007). Ninety-five percent of the shed mediastinal blood was transfused. In the autotransfusion group, a total of 26 units of homologous blood was used versus 78 units in the control group (p < 0.001). CONCLUSIONS: Autotransfusion of shed mediastinal blood in patients undergoing elective, uncomplicated coronary artery bypass grafting halves the number of patients needing homologous blood and reduces the amount of homologous blood given.

Aged↗

PubMed1996

Autotransfused shed mediastinal blood has normal erythrocyte survival.

BACKGROUND: Autotransfusion of shed mediastinal blood may reduce the need for homologous blood transfusions in cardiac surgery. In an earlier study we have shown that the red blood cells (RBCs) of shed mediastinal blood have a normal membrane stability (osmotic fragility) compared with circulating RBCs after coronary artery bypass grafting and better than stored RBCs. This indicates that RBCs in shed mediastinal blood are not damaged further during salvage. It remains to be determined how autotransfusion affects the survival of RBCs from shed mediastinal blood. METHODS: We performed a prospective, randomized, and controlled study involving 26 patients having elective, uncomplicated coronary artery bypass grafting. Dual-isotope labeling technique (chromium 51 and technetium 99m) was used to investigate the 24-hour survival of RBCs from shed mediastinal blood and RBCs from circulating blood, and to estimate the mean survival time of RBCs. RESULTS: There was no significant difference between the 24-hour survival of shed mediastinal RBCs and circulating RBCs. The estimated mean cell lifespan was 20.5 days (range, 11.6 to 29.0 days) for shed mediastinal RBCs and 22.7 days (range, 14.4 to 36.2 days) for circulating RBCs. CONCLUSIONS: The survival of RBCs from shed mediastinal blood after autotransfusion is comparable with the survival of RBCs in the patients' circulating blood.

Blood Loss, Surgical↗

PubMed1996

Diarrhea in young children associated with Escherichia coli non-O157 organisms that produce Shiga-like toxin.

OBJECTIVE: To assess the clinical manifestations and incidence of infection associated with Shiga-like toxin-producing Escherichia coli (SLTEC). STUDY DESIGN: Children with diarrhea within a defined geographic area during a 12-month period were examined for the presence of SLTEC in their stools by polymerase chain reaction with the use of primers that were complementary to sequences of Shiga-like toxins types I and II and to other virulence factors. RESULTS: There were 13 SLTEC infections among 468 children with diarrhea. Besides Shiga-like toxin sequences, the virulence genes eae and EHEC-hly were found in 10 isolates; these isolates were categorized as enterohemorrhagic E. coli (EHEC). Only 2 of 13 isolates were of the O157 strain. All reported cases occurred in summer (June through September) with the exception of one case in April. The infections were sporadic, and the infected children lived in rural and urban areas. Three infections in children with disabilities were hospital acquired. The majority of children had watery diarrhea, two had bloody diarrhea, and one had mild hemolytic-uremic syndrome. The overall incidence of SLTEC infection was 12.5 hospitalized children per 100,000 children less than 16 years of age. CONCLUSIONS: The most frequent clinical manifestation of SLTEC infection was watery diarrhea indistinguishable from other forms of infectious diarrhea. The shift from the O157 strain toward non-O157 SLTEC strains associated with diarrhea, also observed in German patients with hemolytic-uremic syndrome, points to a change in the epidemiologic features of SLTEC-associated disease. Testing for non-O157 SLTEC should be considered in children with diarrhea without a recognized cause.

Bacterial Toxins↗

PubMed1996

Mutational analysis of the p15 and p16 genes in acute leukaemias.

Recently the p16 and related p15 genes have been described as candidate tumour suppressors at chromosome 9p21. These genes have been found to be homozygously deleted at a high frequency in several types of solid tumours and also in acute lymphoid leukaemias. In order to determine whether these genes are more widely involved in haematological malignancies, we have investigated a total of 84 samples that did not have homozygous p16 or p15 deletions from patients with acute lymphoid leukaemia (n=13), acute myeloid leukaemia (n=24) and chronic myeloid leukaemia in blast crisis (n=43) as well as four haemopoietic cell lines. p15 and p16 exon 1 and exon 2 were amplified by polymerase chain reaction (PCR), analysed by single-stranded conformation polymorphism (SSCP) and subsequently by sequencing. Within the p16 gene, a G-->A polymorphism at nucleotide 436 was found in 3/80 (4%) leukaemias and the cell line HL60. This cell line had also a C-->T mutation at nucleotide 232 which causes a premature stop codon. Analysis of the p15 gene revealed a C-->A mutation within the noncoding sequence 27 nucleotides upstream of exon 2 in 10/80 (13%) cases. These data show that inactivation of either the p15 or p16 gene by point mutation is a very uncommon event in acute leukaemias.

Base Sequence↗

PubMed1996

Prospective cross-over comparisons of three low-density lipoprotein (LDL)-apheresis methods in patients with familial hypercholesterolaemia.

We prospectively compared effectiveness, selectivity and biocompatibility of three LDL-apheresis methods, immunoadsorption (IMAL), dextran sulphate adsorption (DSAL) and heparin-induced extracorporeal LDL precipitation (HELP). Seven patients with familial hypercholesterolaemia were treated twice with each method in random sequence. Reduction in atherogenic lipoproteins was without significant difference: LDL -60% to -75%, VLDL -20% to -30%, triglycerides -20% to -42%. High-density lipoprotein (HDL)-cholesterol was reduced by IMAL only (-27%, P < 0.05); DSAL and HELP did not decrease HDL. Total plasma protein reduction was 13-15% with each method, indicating unselectivity. Albumin was significantly decreased by IMAL (-15%, P < 0.05) but not by the other methods. DSAL and HELP reduced fibrinogen (-40%, -58%, P < 0.0001) and other clotting factors. IMAL had almost no effect on coagulation. The white blood cell count did not change. C3 and C4 complement were decreased (-20% to -46%) by all methods. C5a complement did not increase in systemic blood, but was increased in the extracorporeal circulation of IMAL (+200%) and HELP (+150%). Plasma PMN elastase rose in all methods (+200%) indicating neutrophile degranulation. In conclusion, in this short-term study of a small patient population, effectiveness of the three LDL-apheresis methods was similar, but selectivity and biocompatibility were different. The therapeutic relevance of these differences for long-term treatment remains to be elucidated.

Blood Component Removal↗

PubMed1996

[Surgically managed postoperative hemorrhage after tonsillectomy].

BACKGROUND: Bleeding after tonsillectomy is the most frequent and most dangerous complication. We compared the incidence of postoperative bleeding and secondary haemorrhage after tonsillectomy was examined using two methods of hemostasis (suture ligation versus electrocautery). METHODS: The study covered all tonsillectomies performed from 1992 to 1994. Data from about 2096 patients were assessed. RESULTS: The incidence of postoperative bleeding and hemorrhage was 5.4%. When suture ligation of vessels was used to obtain hemostasis during the operation it was evident that postoperative bleeding was rare but more difficult in comparison to the cases in which electrocautery was used alone. While primary bleeding for both methods was similar, secondary bleeding was more frequent after hemostasis achieved with electrocautery. DISCUSSION: Injury of the vessel by suture ligation is thought to be responsible for massive hemorrhage after tonsillectomy. Using electrocautery avoids this pathogenic mechanism. Since electrocautery of the tissue surrounding a vessel produces initial hemostasis, secondary bleeding is more frequent after the wound clot is opened. CONCLUSION: Use of electrocautery for hemostasis is justified since bleeding after primary hemostasis is usually easily managed.

Adolescent↗

PubMed1996

Effect of low-dose dopamine on intestinal villus microcirculation during normotensive endotoxaemia in rats.

Hypoperfusion of the gut mucosa is thought to be a factor in the development of gut barrier failure during sepsis and septic shock. Dopamine stimulates DA-1 receptors which mediate regional vasodilatation in the gut. Therefore, we have investigated the effect of low-dose dopamine (3 micrograms kg-1 min-1) on the intestinal villus microcirculation during endotoxaemia in a rat model of normotensive endotoxaemia, using in vivo videomicroscopy. Blood flow in and the diameters of central villus arterioles were measured before, immediately after and 60 min after a 1-h continuous infusion of endotoxin 1.5 mg/kg body weight. After baseline measurements were obtained, rats received either an infusion of 0.9% saline (group A; n = 7) or a volume-equivalent infusion of dopamine 3 micrograms kg-1 min-1 (group B; n = 7) throughout the study. Control animals (group C; n = 7) received no endotoxin or dopamine. In group A, villus blood flow (mean baseline 8.4 (SEM 0.9) nl min-1) decreased by 29.7 (8.9)% to 5.9 (0.9) nl min-1 immediately after endotoxin challenge and by a total of 43.1 (7.3)% to 4.7 (0.7) nl min-1 after another 60 min. Simultaneously, villus arteriolar diameters decreased from 7.8 (0.2) to 6.9 (0.3) microns and to 6.5 (0.3) microns, respectively. In group B, villus blood flow (baseline 8.7 (0.4) nl min-1) was unchanged immediately after the 1-h infusion of endotoxin (8.3 (0.4) nl min-1). However, another 60 min later blood flow decreased by 28.8 (8.0)% to 6.1 (0.7) nl min-1. In contrast with group A, the diameters of the central villus arterioles were unchanged despite administration of endotoxin (7.9 (0.2) microns; 8.1 (0.4) microns; 8.2 (0.5) microns). In group C, there were no changes in villus blood flow or arteriolar diameters throughout the study. Our results indicated that low-dose dopamine did not prevent, but delayed and attenuated, the decrease in intestinal villus blood during normotensive endotoxaemia.

Animals↗

PubMed1996

Retention of antibacterial activity and bacterial colonization of antiseptic-bonded central venous catheters.

We determined how long antiseptic impregnation with silver sulphadiazine and chlorhexidine (SCC) on polyurethane central venous double- or triple-lumen catheters is retained in vivo. A total of 116 antiseptic catheters were tested for antibacterial activity in an in-vitro bioassay after various periods of iv catheterization. Segments from the subcutaneous (sc) and intravenous (iv) portions of the catheters were cultured. The results of test antiseptic catheters were compared with those from 117 noncoated control (c) catheters. Retention of antibacterial activity followed an exponential curve and lasted for up to 520 h after catheter insertion. Significant differences (P = 0.0001) between SSC and C catheters were noticed with regard to the quantitative level of bacterial colonization (SSC-sc 87 +/- 34 vs C-sc 584 +/- 122; SSC-iv 52 +/- 17 vs C-iv 286 +/- 57; all values are given as mean cfu +/- S.E.M.), and the frequency of bacterial colonization (SSC-sc 20.7% vs C-sc 38.5%, P = 0.0047; SSC-iv 18.1% vs C-iv 30.8%, P = 0.0361). There was no significant difference between the incidence of catheter-related bacteraemia in the test (n = 0) and control groups (n = 3) (P = 0.2573). Further prospective studies are required to delineate the role of antiseptic catheters in preventing catheter-related infections.

Bacteremia↗

PubMed1996

Dopexamine maintains intestinal villus blood flow during endotoxemia in rats.

OBJECTIVE: To determine the influence of dopexamine, a synthetic catecholamine ligand for dopaminergic and beta 2-adrenergic receptors, on alterations of the intestinal villus microcirculation in a model of normotensive endotoxemia. DESIGN: Randomized, controlled trial. SETTING: Experimental laboratory. SUBJECTS: Twenty-one male Wistar rats. INTERVENTIONS: Rats were treated with a continuous infusion of dopexamine (2.5 micrograms/kg/min; N = 7; group A) or 0.9% saline (n = 7; group B) during a study period of 120 mins. Both groups were given endotoxin (Escherichia coli lipopolysaccharide; 1.5 mg/kg Iv) over 60 mins. Animals in the control group (n = 7; group C) received a volume-equivalent infusion of 0.9% saline. Total volume substitution in all groups was 15 mL/kg/hr. MEASUREMENTS AND MAIN RESULTS: Blood flow in the intestinal villi of the distal ileum was determined using in vivo videomicroscopy at baseline, and 60 and 120 mins after the endotoxin challenge. These blood flow determinations were done by an observer who was unaware of the previous treatment of the animals. In addition, mean arterial pressure was monitored at baseline, and 15, 30, 45, 60, 75, 90, 105, and 120 mins later. The administration of 1.5 mg/kg endotoxin alone (group B) resulted in a reduction of the intestinal villus blood flow to 74.8 +/- 9.5% of baseline after 60 mins, and to 61.1 +/- 8.5% of baseline after 120 mins (baseline: 7.4 +/- 0.6 nL/min; 60 mins: 5.3 +/- 0.8 nL/min; 120 mins: 4.4 +/- 0.5 nL/min; p < .05). This reduction of blood flow was associated with a decrease in the arteriolar diameters by 13.8 +/- 2.5% after 60 mins, and by 17.1 +/- 4.3% after 120 mins (p < .05 vs. baseline). In contrast, villus blood flow in the dopexamine group (group A) did not show statistically significant changes during the entire study period, despite the administration of endotoxin (baseline: 8.2 +/- 0.6 nL/min; 60 mins: 7.3 +/- 0.8 nL/min; 120 mins: 7.8 +/- 0.5 nL/min). No vasoconstriction of the villus arterioles was noted in this group. In control animals (group C), the blood flow (baseline: 8.1 +/- 1.6 nL/min; 60 mins: 7.6 +/- 1.4 nL/min: 120 mins: 7.8 +/- 1.4 nL/min) and the arteriolar diameters remained unchanged throughout the observation period. Mean arterial pressure did not differ between groups: it remained unaltered in all groups during the entire study period. CONCLUSIONS: Dopexamine maintains intestinal villus arterial perfusion and prevents the vasoconstriction in villus arterioles during early normotensive endotoxemia. Therefore, further studies in critically ill patients will have to determine whether the early prophylactic use of dopexamine can limit ischemia and prevent the development of multiple organ failure.

Adrenergic beta-Agonists↗

PubMed1996

KatP, a novel catalase-peroxidase encoded by the large plasmid of enterohaemorrhagic Escherichia coli O157:H7.

A gene coding for a catalase-peroxidase activity was identified on a 9-7 kb Smal DNA fragment derived from the large plasmid pO157 of enterohaemorrhagic Escherichia coli (EHEC) O157:H7 strain EDL 933. Nucleotide sequencing revealed an ORF of 2208 bp and predicted a 736 amino acid polypeptide with a molecular mass of 81.8 kDa. This putative protein was found to be highly homologous to members of the bacterial bifunctional catalase-peroxidase family. Analysis of its amino acid sequence revealed the presence of characteristic peroxidase 1 and 2 motifs. In addition, an N-terminal signal sequence was found, suggesting that the catalase-peroxidase is transported through the cytoplasmic membrane. EHEC catalase-peroxidase activities were investigated in cytoplasmic and periplasmic crude extracts as well as in culture supernatants from wild-type and recombinant E. coli strains. EHEC-specific catalase-peroxidase activity was detected primarily in the periplasm in strain EDL 933. The newly discovered enzyme was designated KatP, to indicate its plasmid origin. PCR analysis of representative strains of all enteric E. coli pathogroups (i.e. enterohaemorrhagic, enterotoxigenic, enteropathogenic, enteroaggregative and enteroinvasive E. coli) revealed a close association between the occurrence of EHEC-haemolysin and the katP gene in Shiga-like-toxin-producing E. coli O157 strains.

Amino Acid Sequence↗

PubMed1996

Apolipoprotein E e4 allele in the normal elderly: neuropsychologic and brain MRI correlates.

The presence of the apolipoprotein E e4 allele has been considered to be a risk factor for Alzheimer's disease and vascular dementia. We therefore used demanding neuropsychologic testing and brain MRI to determine if elderly normals with at least one e4 allele demonstrate subclinical changes in cognition and a higher frequency of brain atrophy or silent ischemic brain damage. The study population consisted of 214 randomly selected individuals aged 50 to 75 years without neuropsychiatric or general disease. There were 175 (81.8%) subjects without and 39 (18.2%) with at least one e4 allele. The two groups were comparable for age, length of education, verbal intelligence, mood and major vascular risk factors. Apolipoprotein E e4 carriers performed significantly worse than non-carriers when assessed for learning and memory abilities, while there were no differences in test results of conceptualization, attention, speed of mental processing and visuopractical skills. There were no between-group differences for thromboembolic and lacunar infarcts, white matter hyperintensity grading and the semiautomatically measured white matter hyperintensity area. The extent of sulcal and ventricular widening as well as hippocampal and parahippocampal volumes were also similar between the comparative subsets. We conclude that the apolipoprotein E e4 allele is associated with subtle learning and memory deficits in normal elderly persons and may therefore be suggested a marker for accelerated cognitive aging. In this group of subjects it was not associated with brain parenchymal changes as demonstrated by MRI.

Aged↗

PubMed1996

Analysis of the enterohemorrhagic Escherichia coli O157 DNA region containing lambdoid phage gene p and Shiga-like toxin structural genes.

In this study, we determined the nucleotide sequence of the p gene contained within a 5-kb EcoRI restriction fragment cloned from Shiga-like toxin II (SLT-II)-converting phage 933W of Escherichia coli O157:H7 strain EDL933. The p gene was 702 bp long and had 95.3% sequence similarity to the p gene of phage lambda. Multiple hybridization patterns were obtained when genomic DNA fragments were hybridized with both p and slt-I, slt-II, or slt-IIc sequences. All O157 isolates also possessed an analog of lambda gene p which was not linked with either slt-I or slt-II. Restriction fragment length polymorphism comparisons of clinical O157 isolates and derivates undergoing genotype turnover during infection were made, and loss of large DNA fragments that hybridized with slt-II and p sequences was observed. To further analyze the DNA region containing the p and slt genes, we amplified fragments by using a PCR with one primer complementary to p and the other complementary to either the slt-I or the slt-II gene. PCR analysis with enterohemorrhagic E. coli O157 and non-O157 strains yielded PCR products that varied in size between 5.1 and 7.8 kb. These results suggest that even within O157 isolates, the genomes of SLT-converting phages differ. The methods described here may assist in further investigation of SLT-encoding phages and their role in the epidemiology of infection with enterohemorrhagic E. coli.

Bacterial Toxins↗

PubMed1996

Specific antibody reactivity against a 110-kilodalton Actinobacillus actinomycetemcomitans protein in subjects with periodontitis.

The purpose of the present study was to determine the serum immunoglobulin A (IgA), IgM, and IgG reactivities against proteins of Actinobacillus actinomycetemcomitans in patients with periodontitis. Serum samples from 20 patients with early-onset periodontitis, 20 patients with adult periodontitis, and 20 age- and sex-matched healthy controls were assessed by immunoblot analysis. IgG antibody reactivity against a sarcosyl-insoluble 110-kDa protein of A. actinomycetemcomitans was detected in 65 and 45% of patients with early-onset periodontitis and adult peritonitis, respectively, and IgA antibodies against this protein were found in 70 and 55% of these patients, respectively. However, control subjects showed no IgG reactivity, and IgA antibodies against the sarcosyl-insoluble 110-kDa protein were detected in only 5% of the patients (P < 0.05). There was no IgM antibody reactivity against this protein in any of the diseased or healthy subjects. The sensitivity and specificity of serum IgA antibody reactivity against the 110-kDa protein in detecting subgingival A. actinomycetemcomitans infection, as determined by PCR, were 77 and 66%, respectively. The results of the study indicated that the sarcosyl-insoluble 110-kDa protein is a potential candidate for use in the serodiagnosis of periodontal disease.

Adult↗

PubMed1996

Enterohemolytic phenotypes and genotypes of shiga toxin-producing Escherichia coli O111 strains from patients with diarrhea and hemolytic-uremic syndrome.

Thirty-six Shiga toxin-producing Escherichia coli (STEC) O111:H- strains, 18 of which were isolated from patients with hemolytic-uremic syndrome (HUS) and 18 from patients suffering from diarrhea, were investigated for their enterohemolytic phenotypes and genotypes. Twenty-two strains were EHEC hemolysin (EHEC Hly) positive by probe hybridization and by PCR with sequences complementary to the EHEC hlyA gene of E. coli O157:H7, but only 20 of these were hemolytic on blood agar plates. The remaining 14 strains were EHEC Hly negative according to DNA-based methods and did not express the enterohemolytic phenotype. The enterohemolytic phenotype was observed in 16 of 18 (88%) strains from patients with HUS but only in 4 of 18 (22.2%) of the STEC O111:H- strains from patients with diarrhea. All STEC O111:H- strains carried large plasmids, as shown by plasmid analysis, but only plasmids of EHEC Hly probe-positive strains hybridized with the CVD419 probe. A BamHI fragment of approximately 12 kb was cloned from the large plasmid of the E. coli O111:H- strain 78/92 and shown to mediate hemolytic activity when transformed into the E. coli laboratory strain HB101. The EHEC O111 hlyA gene was sequenced completely and shown to have 99.4% sequence identity to the corresponding EHEC O157 hlyA gene of the E. coli O157:H7 strain EDL 933. Our results indicate that detection of EHEC Hly either by DNA-based methods or by investigation of the enterohemolytic phenotype on blood agar alone is insufficient for screening STEC O111 strains. However, the high incidence of EHEC Hly in isolates from patients with HUS and its rare occurrence in isolates from patients with diarrhea may indicate that STEC O111 strains have a distinct pathogenic potential for humans and that the presence of EHEC Hly increases the ability of an STEC O111 strain to cause extraintestinal complications in humans.

Bacterial Toxins↗

PubMed1996

Serodiagnosis of Porphyromonas gingivalis infection by immunoblot analysis with recombinant collagenase.

The Porphyromonas gingivalis collagenase-specific serum immunoglobulin A (IgA), IgM, and IgG responses from 20 patients with early-onset periodontitis (EOP), 20 patients with adult periodontitis, (AP), and 20 age- and sex-matched healthy controls were examined by immunoblot analysis. A recombinant collagenase antigen used for the immunoblot analysis was produced by using the plasmid pGEX-2T, which allows the fusion between the collagenase and glutathione S-transferase. There was no significant difference in collagenase-specific IgG antibody detection between samples from the EOP, AP, and control groups. In contrast, 85% of AP and EOP sera had collagenase-specific IgA antibodies, whereas only 20% of control sera showed collagenase-specific IgA reactivity. Plaque samples from all groups were assessed by PCR with primers complementary to the collagenase-encoding gene prtC. The results indicated that 90% of AP and EOP plaque samples and 10% of control samples were positive for P. gingivalis. All patients with collagenase-specific IgA antibodies were PCR positive. The results of the study indicate a nearly complete concordance (k = 0.856) between the presence of collagenase-specific IgA antibodies and PCR detection of P. gingivalis. By using PCR as the "gold standard," the sensitivity and specificity of the IgA immunoblot test were 94.7 and 90.9%, respectively. Therefore, the recombinant collagenase is a potential candidate for use in the serodiagnosis of periodontitis.

Adult↗