Endotoxin levels in sera of elderly individuals.

The endotoxin levels in serum of 377 72-year-old individuals were quantitated. The study population was a representative sample of this age group and was participating in a general study of health and disease among the elderly in Göteborg, Sweden. The endotoxin levels in serum were quantified by the chromogenic Limulus amebocyte lysate assay and were correlated with the health status and laboratory findings for each individual. The mean endotoxin levels (+/- 1 standard deviation) in men and women, when excluding four outliers, were 6.6 +/- 3.8 and 6.9 +/- 3.8 pg/ml, respectively. All included, 21.5% of individuals had endotoxin levels equal to or above the sensitivity limit of 10 pg/ml. Strong positive correlations were found between endotoxin levels and plasma triglycerides (P > 0.995) and between endotoxin levels and serum protein (P > 0.9875). The endotoxin activity also correlated with mean corpuscular hemoglobin concentration (P < 0.005, negative correlation), body mass index (P > 0.9875), and decreased appetite (P > 0.9875). A high alcohol consumption was associated with increased endotoxin levels (P = 0.995). There are no previous studies which examine endotoxin levels in serum samples from individuals representative of the population. This study showed that elderly individuals had the same mean level of endotoxin as has been found in other age groups. The increased endotoxin levels seen in heavy drinkers may be explained by a decreased ability of the liver to remove endotoxin. The correlations found between endotoxin and triglycerides, protein, mean corpuscular hemoglobin concentration, decreased appetite, and body mass index are discussed.

Effects of anti-inflammatory agents on mucosal inflammation induced by infection with gram-negative bacteria.

Gram-negative bacterial infections of the urinary tract elicit a mucosal inflammatory response. Interleukin-6 is secreted into the urine, and polymorphonuclear leukocytes (PMNL) are recruited. In the present study we examined the effect of anti-inflammatory agents on these parameters and on bacterial clearance from the kidneys. Dexamethasone reduced interleukin-6 secretion, the PMNL response, and bacterial clearance. Diclofenac abolished the urinary interleukin-6 response but reduced the PMNL response and bacterial clearance only at the highest concentrations. Indomethacin drastically decreased bacterial clearance without the corresponding effect on interleukin-6 production or the PMNL response. The results demonstrate that the inhibition of inflammation impairs bacterial clearance from the kidneys. This is, however, not a direct function of inhibited interleukin-6 production or PMNL recruitment.

Glycosphingolipid receptors for Pseudomonas aeruginosa.

The binding of Pseudomonas aeruginosa to glycosphingolipids and to buccal and bronchial epithelial cells was analyzed. Three independently expressed specificities were found by bacterial binding to glycosphingolipids separated by thin-layer chromatography. All strains bound gangliotria- and gangliotetrasylceramide. All but one of the strains bound sialic acid-containing glycosphingolipids and lactosylceramide. The latter two specificities could be separated in that the lactosylceramide binding was retained and the sialic acid binding was suppressed when bovine serum albumin was used as a blocking agent in the thin-layer chromatography assay. The attachment to buccal epithelial cells, like the binding to sialylated compounds and lactosylceramide, was abolished by Formalin treatment of the bacteria, suggesting the importance of these specificities for cell adherence. In contrast, the binding to gangliotria- and gangliotetraosylceramide was retained by nonattaching Formalin-treated bacteria.

Outbreak of Haemophilus influenzae type b meningitis in a day care center.

Three cases of Haemophilus influenzae type b (Hib) meningitis in a Swedish day care center prompted the investigation of the nasopharyngeal carriage in attendees of that day care center (I), and among children in another day center (II) in the same city, but without meningitis cases. Because the evaluation of the spread of Haemophilus influenzae (Hi) isolates through a population, such as children in day care centers, requires stable identification markers for the bacteria, this study used multilocus enzyme electrophoresis to separate Hib carried by day care attendees from the disease-associated Hib clone. The three meningitis episodes were caused by the same clone of Hib (ET14). This and other Hib clones occurred in the healthy carriers. The frequency of H. influenzae carriage and composition of the H. influenzae flora differed between the two day care centers. Center I with the meningitis cases had a lower overall frequency of H. influenzae carriage, 11 of 40 (28%) compared with both the control day care center (57%) and previous studies. The main difference between the two day care centers was in the nontypable H. influenzae (HiNT) flora. The frequency of HiNT was higher in the center without Hib disease, and there was a higher degree of sharing of HiNT isolates among the children in Center II. This raised the question of an inverse relationship between carriage of HiNT and Hib infection. The results emphasized the importance of accurate identification of the disease isolate in order to estimate the risk of acquisition and dissemination to secondary cases.

Ciclosporin-dependent, nu-independent, mucosal interleukin 6 response to gram-negative bacteria.

Regulation of the mucosal inflammatory response to Gram-negative bacteria was analysed. The interleukin 6 (IL-6) secretion, influx of polymorphonuclear leucocytes into urine, and bacterial clearance from the kidneys were compared between Balb/c (nu/nu) and nu/+/- mice, with and without ciclosporin (CsA) treatment. There was no significant influence of the nu genotype on any of the host responses measured. CsA pretreatment significantly decreased Il-6 secretion in both nu/nu and nu/+/- mice, but did not affect bacterial clearance or the leucocyte response in any mouse strain tested. Tissue damage, in addition to bacterial infection, resulted in significantly higher levels of IL-6 than bacterial infection alone. Tissue-damaged mice were significantly less likely to clear the bacterial infection than their non-damaged counterparts, but there was no significant difference in the leucocyte response. CsA pretreatment did not significantly reduce the levels of IL-6 in the tissue-damaged mice. These results demonstrate that the mucosal inflammatory response to Gram-negative infection, including IL-6 secretion, is nu-independent, and that bacterial infection alone or in combination with tissue damage induce IL-6 secretion by two different pathways.

Frequency and organization of pap homologous DNA in relation to clinical origin of uropathogenic Escherichia coli.

The pap operon encodes the gal alpha 1-4gal beta specific adhesins of Escherichia coli. The presence and organization of pap homologous DNA was determined using two probes specific for pap in 217 uropathogenic E. coli samples by dot blot and Southern blot analysis. The frequency of pap homologous DNA was 76% in pyelonephritis, 69% in cystitis, and 52% in an asymptomatic bacteriuria group. Further, the gal alpha 1-4gal beta binding phenotype among the pap-positive strains was expressed more often in acute pyelonephritis (91%) than the cystitis (60%) or asymptomatic bacteriuria (52%) strains. This was explained in part by difference in organization of pap homologous DNA between the genotypically positive pyelonephritis and asymptomatic bacteriuria strains. The pyelonephritis isolates contained three copies of pap significantly more often than the asymptomatic bacteriuria strains, and the pyelonephritogenic O-antigen types had a general increase in pap copy number. The difference in expression of gal alpha 1-4gal beta adhesins between pyelonephritis and asymptomatic bacteriuria isolates was thus not only a function of the frequency of pap homologous DNA but also of phenotypic expression among genotypically pap-positive strains.

Follicular cystitis in girls with untreated asymptomatic or covert bacteriuria.

Nodular changes of the bladder mucosa, that is cystitis follicularis or cystitis cystica, are found in 2 to 9% of all children with urinary tract infections. The nodules are composed of lymphoid aggregates, resembling Peyer's patches in the intestine. Children with this finding are considered to have a poor short-term prognosis with a marked tendency for recurrent infections. Screening programs for bacteriuria have revealed that a few per cent of the female population are bacteriuric without symptoms of overt disease. Of 59 girls followed with untreated asymptomatic bacteriuria 52 had nodular changes of the bladder mucosa at cystoscopy. Biopsy was performed in 22 girls, and revealed lymphocytic infiltration in 19 and follicular formation in 11. The nodular changes persisted when bacteriuria continued but disappeared in patients who became abacteriuric. This finding demonstrates the reversibility of the changes, and supports the assumption that they are secondary to the presence of bacteria and not a primary lesion.

Selection and evolution of virulence in bacteria: an ecumenical excursion and modest suggestion.

Why do parasites kill their hosts? During this past decade, research in three different areas; evolutionary ecology, medical microbiology, and population genetics has provided theory and data that address this and related questions of selection and the evolution and maintenance of parasite virulence. A general theory of parasite-host coevolution and the conditions for selection to favour parasite virulence has been put forth. Considerable advances have been made in elucidating the mechanisms of pathogenicity and inheritance of virulence in bacteria. The population genetic structure and the relationship between pathogenic and non-pathogenic forms has been determined for a number of species of bacteria. We critically review these developments and their implications for questions of selection and the evolution and maintenance of virulence in bacteria. We postulate how selection may operate on specific types of bacterial virulence and present a general protocol to experimentally test hypotheses concerning selection and the evolution of virulence in bacteria.

Bacterial attachment as a predictor of renal abnormalities in boys with urinary tract infection.

The development of renal scarring was analyzed prospectively in 241 boys with their first known episode of symptomatic urinary tract infection (140 acute pyelonephritis, 61 acute cystitis, and 40 nonspecific). Of 197 boys undergoing urography, 22 (11%) had scars; 20 were in the pyelonephritis group. Vesicoureteral reflux occurred in 81% of those with scarring, compared with 20% of those without scarring. The bacteria causing the first episode of urinary tract infection in each patient were saved, and Escherichia coli organisms were characterized for the expression of both galactose-alpha (1----4)galactose-beta (Gal-Gal)-specific adhesins and pap homologous DNA. Scarring occurred in 41% and other renal abnormalities in 11% of boys infected with bacteria that did not bind Gal-Gal (Gal-Gal negative), compared with 5% and 1%, respectively, in those infected with Gal-Gal-binding strains (Gal-Gal-positive) (relative risk 8.3; 95% confidence limits 3.3 to 20.4; p less than 0.001). That boys infected with Gal-Gal-negative strains more often had reflux did not explain the increased risk for renal scarring in this group. The possibility that the phenotypically negative strains could be induced to express Gal-Gal adhesions in vivo was excluded by dot blot analysis, which showed the absence of pap homologous DNA in all but one of the Gal-Gal-negative strains. The results suggest that the absence of Gal-Gal-specific adhesins in E. coli can be used as an indicator of risk for renal scarring and the need for radiologic examination.

Binding specificities of wild-type and cloned Escherichia coli strains that recognize globo-A.

In this study we compared the specificity for the globoseries of glycolipids of Escherichia coli expressing the O-negative, A-positive (ONAP) adhesin and clones transformed with the pap-like (prs or pap-2) gene cluster. Receptor-active glycolipids were identified by the ability of radiolabeled bacteria to bind to the glycolipids on thin-layer chromatogram plates. The ONAP adhesin and pap-like clones bound with high affinity to the globo-A and Forssman glycolipids. The ONAP strains did not recognize other glycolipids of the globoseries. In contrast, the pap-like clones also showed weak binding to globotriaosylceramide and reacted weakly with Gal alpha 1----4 Gal beta-latex beads. We suggest that the pap-like and ONAP adhesins recognize an epitope shared by the globo-A and Forssman structures, e.g., terminal GalNAc alpha 1----3 bound to Gal alpha 1----4Gal beta-containing glycolipids.

Interleukin-6 induced at mucosal surfaces by gram-negative bacterial infection.

Interleukin-6 (IL-6) was produced in response to mucosal and systemic infection of mice with gram-negative bacteria. The IL-6 response was controlled by the lipopolysaccharide gene, Lps; in C3H/HeN mice (Lpsn/Lpsn), the urinary IL-6 levels increased within 30 min after challenge with Escherichia coli, but no response occurred in C3H/HeJ mice (Lpsd/Lpsd). In lipopolysaccharide-responder mice, the levels of local and systemic IL-6 were related to the degree of infection. The urinary response dominated after intravesical challenge, and the serum response dominated after intraperitoneal challenge. The results demonstrate that IL-6 is activated as part of lipopolysaccharide-induced mucosal and systemic responses to gram-negative bacterial infections.

Turnover of nontypable Haemophilus influenzae in the nasopharynges of healthy children.

The nasopharyngeal Haemophilus influenzae flora of healthy children in a day care center was analyzed by repeated sampling during 4 winter months. The average carrier rate was 39%, but 74% of the children became colonized at some time during the study. The H. influenzae isolates were identified by capsular type, biotype, and multilocus enzyme electrophoresis. The turnover of the flora in individual children and in the day-care group was characterized. Four patterns of colonization were defined among the 38 children and 49 H. influenzae strains. Depending on the persistence in the group, the strains were designated as endemic, i.e., shared between several children on several occasions, or as epidemic, i.e., occurring only once but in several children. The individual hosts had two patterns; resident strains persisted for 2 weeks or more, and transient strains showed no evidence of persistence. The results suggest that there is considerable sharing of certain nontypable H. influenzae among healthy children in day care, whereas other strains remain restricted to a single host. The properties of host and bacteria determining these patterns remain to be defined.

Special virulence of the Escherichia coli O1:K1:H7 clone in acute pyelonephritis.

The relationship between acute-phase responses and bacterial properties was studied in a population of 88 children with their first known episode of acute pyelonephritis. One strain from each patient was included in the study. Eighty-four of the patients were infected with Escherichia coli, which was assigned a clonotype according to the O:K:H stereotype; 55 patients carried one of the 12 multiply occurring clones. Globotetraosylceramide-specific (globo+) adhesion was present in 90% of these 12 clones, compared with 62% in the remaining 29 singly occurring clones. The patients infected with globo+ strains had significantly increased inflammatory reactions compared with patients with globo- strains. The O1:K1:H7 strain was the single most frequent clone (n = 14) that always expressed globo+ adhesins. Patients infected with O1:K1:H7 had an inflammatory response similar to that of other globo+ infections, but had a shorter duration of symptoms before diagnosis, higher fever, and higher peripheral leukocyte count. These results demonstrate special virulence of the O1:K1:H7 clone, reflected by the acuteness of onset of infection.

Secretor state and renal scarring in girls with recurrent pyelonephritis.

The non-secretor phenotype was significantly associated with the occurrence of renal scarring among patients with recurrent pyelonephritis. Girls (n = 77) with recurrent pyelonephritis were followed from the first known episode of infection for up to twelve years with repeated radiological investigations. They were divided into two categories: those with renal scars (n = 35) and those who did not develop scars (n = 42). There was a significant over-representation of non-secretors among the patients with scarring, (14/35, 40%) compared to the healthy controls (21.8%, P less than 0.05). The frequency of non-secretors among the girls who did not develop scars in spite of repeated episodes of acute pyelonephritis was not significantly different from the healthy controls (9/42, 21% n.s.). This study provides a basis for analysis of the influence of secretor state on host-parasite interaction in the urinary tract.

Selective use of excretory urography in women with acute pyelonephritis.

The outcome of excretory urography was analyzed in 103 nonpregnant women followed prospectively after community acquired acute pyelonephritis. Radiological abnormality was found in 40 per cent of the patients (17 per cent had major abnormalities, including renal scarring, calculi and obstruction). All 5 women with surgically correctable lesions had rapid bacteriological relapse or recurrent acute pyelonephritis. Neither a history of urinary tract infection, the acute inflammatory response nor infection due to Escherichia coli with or without adhesins specific for Gal alpha 1----4Gal beta-containing receptors was efficient in predicting major radiographic lesions or the outcome of treatment. Bacteremia was detected in 27 per cent of the patients but in the absence of obstruction. These results suggest that excretory urography is dispensable in most women with acute pyelonephritis, and that those needing such investigation may be identified by failure to respond to antibiotic treatment or by the recurrence pattern.

Distribution of the P-associated-pilus (pap) region among Escherichia coli from natural sources: evidence for horizontal gene transfer.

Variation in chromosomal DNA in Escherichia coli was studied with probes specific for the P-associated-pilus (pap) region. The presence of DNA homologous to pap was determined by dot blots. Variation in the number of copies of pap and in the organization of internal and flanking sequences was determined by Southern blot hybridization. The 229 strains studied were also classified by O:K:H serotyping and multilocus enzyme electrophoresis. There was considerable heterogeneity in the presence of pap and distribution of pap-homologous DNA in these E. coli strains from natural sources. In general, there was less variation in pap among strains of the same specific O:K:H serotype and enzyme electrophoretic type than among random isolates. There were, however, E. coli strains identified as members of the same clone by O:K:H serotyping and enzyme electrophoresis that were pap positive and pap negative or had different Southern blot patterns for the pap probes (pap type). There were also isolates of the same pap type that differed in two of three O:K:H serotype antigens and the majority of enzymes that determined their enzyme electrophoretic type. These latter two observations were interpreted as evidence for the horizontal (infectious) transfer of the pap-homologous sequences among clones of E. coli.