The effects of deoxycholate and sodium dodecyl sulphate on the serological reactivity of antigens isolated from six Bacteroides reference strains

Abstract The detergents sodium dodecyl sulphate (SDS) and sodium deoxycholate (NaD) are frequently used as solvents for macromolecular polysaccharide complexes in immunochemical and serological techniques. The influence of the disaggregating surfactants on the serological reactivity of endotoxins isolated from six serotype specific reference strains of the Bacteroides fragilis group was investigated by comparing haemagglutinating and precipitating reactivities of antigen solutions in phosphate buffered saline (PBS), NaD and SDS. All antigens were phenol/water extracted endotoxins. Solutions of antigens isolated from serotypes A, B, C and D in PBS exhibited mainly serotype specificity and a few well known low-titer cross reactions; solutions in NaD showed additional cross reactivity, which was enhanced by solubilization of the antigens in SDS. In immunoelectrophoresis endotoxins isolated from serotypes A and C and dissolved in NaD or SDS showed additional precipitation lines compared to solutions of the same antigens in PBS. These changes in the serological reactivity are of relevance for investigations where the serological specificty of antigens is in question

Detection of Bacteroides fragilis endotoxin in amniotic fluid by counterimmunoelectrophoresis

The ability of counter immunoelectrophoresis (CIE) to detectBacteroides fragilis endotoxin in amniotic fluid in small concentrations was evaluated. A method was developed which, in combination with ultrafiltration, permits detection ofB. fragilis endotoxin in amniotic fluid in a concentration of 40 ng/ml or more. The sensitivity threshold was reduced to 2 ng/ml by using a highly reactive IgG-fraction isolated from rabbit anti-B. fragilis IPL E 323 antiserum

Bacteriological studies of blood, tissue fluid, lymph and lymph nodes in patients with acute dermatolymphangioadenitis (DLA) in course of ‘filarial’ lymphedema

Filarial lymphedema is complicated by frequent episodes of dermatolymphangioadenitis (DLA). Severe systemic symptoms during attacks of DLA resemble those of septicemia. The question we asked was whether bacterial isolates can be found in the peripheral blood of patients during the episodes of DLA. Out of 100 patients referred to us with ‘filarial’ lymphedema 14 displayed acute and five subacute symptoms of DLA. All were on admission blood microfilariae negative but had a positive test in the past. Blood bacterial isolates were found in nine cases, four acute (21%) and five subacute (26%). In 10 acute cases blood cultures were found negative. Six blood isolates belonged to Bacilli, four to Cocci and one was Sarcina. To identify the sites of origin of bacterial dissemination, swabs taken from the calf skin biopsy wounds and tissue fluid, lymph and lymph node specimens were cultured. Swabs from the calf skin biopsy wound contained isolates in nine (47%) cases. They were Bacilli in nine, Cocci in three, Acinetobacter and Erwinia in two cases. Tissue fluid was collected from 10 patients and contained Bacilli in four (40%) and Staphylococci in three (30%). Lymph was drained in four patients and contained isolates in all samples (100%). They were Staphylococcus epidermis, xylosus and aureus, Acinetobacter, Bacillus subtilis and Sarcina. Three lymph nodes were biopsied and contained Staphylococcus chromogenes, xylosus, Enterococcus and Bacillus cereus. In six cases the same phenotypically defined species of bacteria were found in blood and limb tissues or fluids. In the ‘control’ group of patients with lymphedema without acute or subacute changes all blood cultures were negative. Interestingly, swabs from biopsy wound of these patients contained isolates in 80%, tissue fluid in 68%, lymph in 70% and lymph nodes in 58% of cases. In healthy controls, tissue fluid did not contain bacteria, and lymph isolates were found only in 12% of cases. This study demonstrates that patients with acute episodes of DLA reveal bacteriemia in a high percentage of cases. Diversity of blood and tissue bacterial isolates in these patients points to a breakdown of the skin immune barrier in lymphedema and subsequently indiscriminate bacterial colonization of deep tissues and spread to an blood circulation. © 1999 Elsevier Science B.V. All rights reserved

Tumor necrosis factor-alpha in response to endotoxin administration in the pregnant guinea pig.

Contains fulltext : 86685.pdf (publisher's version ) (Open Access)OBJECTIVE: Our purpose was to test the hypothesis that an intramuscular endotoxin challenge induces production of tumor necrosis factor-alpha in the pregnant guinea pig and to investigate some of the metabolic effects. STUDY DESIGN: Twelve randomly selected guinea pigs at 33 days' gestation with a sampling catheter in the carotid artery received an intramuscular injection of a solution of endotoxin isolated from Bacteroides fragilis (n = 6) or of solvent alone (n = 6). Plasma values of tumor necrosis factor-alpha, hematocrit, and 6-keto-prostaglandin F1 alpha were determined before and several hours after injection. RESULTS: Tumor necrosis factor-alpha was detected in five of six guinea pigs, but it could not be demonstrated in five of six placebo animals. The hematocrit was significantly decreased, and prostaglandin F1 alpha significantly increased 24 to 48 hours after endotoxin injection. CONCLUSION: In pregnant guinea pigs an intramuscular endotoxin challenge induces the release of tumor necrosis factor-alpha, followed by a reduced hematocrit and an increased prostacyclin concentration. These effects could be involved in the pathogenesis of endotoxin-induced fetal growth retardation.1 juli 199

Immune Response to Endotoxin Isolated from <i>Bacteroides fragilis</i> in the Pregnant Guinea Pig

The humoral immune response to endotoxin isolated from &lt;i&gt;Bacteroides fragilis was &lt;/i&gt;analyzed in the pregnant guinea pig by means of passive hemagglutination, passive hemolysis, a modified Coombs test, and by crossed immunoelectrophoresis. Pregnant animals were immunized with endotoxin on day 30 of gestation, and antibodies were determined on day 61 in maternal and fetal sera, and in amniotic fluid. The IgG and IgM responses in maternal sera were of the same magnitude as in sera of nonpregnant animals. Fetal sera contained IgG and sometimes IgM, and a higher percentage of incomplete antibodies against endotoxin than maternal sera. Low-titer anti-endotoxin antibodies, partially sensitive to dithiothreitol, were found in amniotic fluid. A statistically significant reduction in the growth of fetuses from endotoxin-immunized females was observed.</jats:p

Genotyping of Clostridium difficile isolates from a hospital in Warsaw: A preliminary study

AbstractBackground: Toxigenic Clostridium difficile is the etiologic agent of C. difficile-associated diarrhea. It is believed that patient-to-patient transmission and environmental contamination are risk factors for the spread of this disease in hospitals. The development of precise typing schemes has provided some understanding of the epidemiology of this nosocomial infection.Objective: The purpose of this study was to identify strain types of C. difficile present in the environment of a maternity ward and surgical wards of a hospital in Warsaw, and to correlate these types with those isolated from neonates and surgical patients.Methods: Arbitrarily primed polymerase chain reaction (APPCR), with the arbitrary primer PG-05, was used to genotype 14 isolates of C. difficile obtained from different sources in a hospital in Warsaw. These included clinical isolates from neonates and surgical patients, as well as environmental isolates from the maternity and surgical wards.Results: A predominant toxigenic isolate was found in the maternity environment, and this strain type was also isolated from the stools of two of five newborns. Unique and different DNA banding patterns were observed in the remaining three isolates from neonates. In addition, isolates from an infant with diarrhea also showed a unique AP-PCR type. The isolates from three surgical patients analyzed had the same AP-PCR profile. This type was also found in the surgery ward environment.Conclusion: These data support environmental contamination as an important factor in the etiology of C. difficile-associated diarrhea in this hospital

PCR ribotyping and arbitrarily primed PCR for typing strains of Clostridium difficile from a Polish maternity hospital

Detection of the source of Clostridium difficile strains is of importance for the control of the nosocomial spread of this microorganism. For this purpose, vaginal and rectal swabs from 183 mothers, duplicate fecal samples (taken on days 1 and 4 after birth) from 183 neonates, and 94 environmental samples were cultured for C. difficile. The microorganism was never detected in the meconium obtained on day 1 after birth. On the other hand, an incidence of 17% C. difficile positivity was noted in the fecal samples obtained on day 4 after birth. Forty-two percent of the 31 colonized neonates had been delivered with complications. The bacteria were never encountered in the rectal swabs of the mothers, and C. difficile was identified in only one vaginal swab. In contrast, 13% of the environmental samples were positive for C. difficile. No major difference was encountered between patient and environmental isolates with respect to toxigenicity (58 to 65% toxigenic isolates). All strains were subsequently typed by PCR amplification of the 16S-23S ribosomal intergenic spacer regions and by arbitrarily primed PCR (AP-PCR) with different primers and combinations thereof. All environmental isolates and 11 of 31 neonatal strains were of a single type. The vaginal strain was unique, and among the maternity ward- and neonate-related isolates, only two additional AP-PCR types were identified. When a collection of C. difficile strains from patients hospitalized in other institutions and suffering from antibiotic-associated diarrhea or pseudomembranous colitis was analyzed in a similar manner, it appeared that the strain from the maternity ward was unique. The other strain commonly encountered among the neonates was also identified frequently among the isolates from patients with antibiotic-associated diarrhea or pseudomembranous colitis, indicating its general occurrence. On the basis of both epidemiological studies and PCR-mediated genotyping, it was shown that the environment and not the birth canal is the major source of C. difficile acquisition by neonates in this maternity hospital setting. Furthermore, AP-PCR appears to be a fast and useful method for epidemiologically relevant typing of C. difficile isolates.</jats:p