Microtiter assay for detecting Campylobacter spp. and Helicobacter pylori with surface gangliosides which bind cholera toxin.

Campylobacter jejuni with Gm1 ganglioside in the core of its lipopolysaccharide has been associated with Guillain-Barré syndrome. Since this epitope may be of considerable pathophysiologic importance and since this ganglioside binds cholera toxin, a rapid screening assay to detect bacteria that bind cholera toxin as an indication of Gm1 on their surfaces was developed. In the assay, bacterial lawns were grown on agar plates, harvested with phosphate-buffered saline, boiled, and incubated with a standard concentration of cholera B subunit. Preparations from strains with Gm1 were observed to inhibit the binding of cholera B subunit to Gm1 in a microtiter enzyme-linked immunosorbent assay. By using this assay with two groups of strains, 37 positive strains were detected among the 197 tested. Species with positive isolates included C. jejuni, Campylobacter coli, and Helicobacter pylori. The assay is capable of testing large numbers of isolates and should prove useful in future clinical and epidemiological studies of bacteria with this epitope.

Detection of Toxoplasma gondii tachyzoites and bradyzoites in blood, urine, and brains of infected mice.

Different techniques for identifying Toxoplasma gondii were compared. PCR was used to amplify part of the major surface antigen P30 gene of T. gondii. Amplified-DNA detection with the DNA enzyme immunoassay (PCR-DEIA) was more sensitive than ethidium bromide staining after agarose gel electrophoresis and as sensitive as nested PCR. PCR-DEIA, using common enzyme-linked immunosorbent assay (ELISA) methods, avoids agarose gel electrophoresis for the identification of amplified products. T. gondii can also be detected with equal sensitivity in infected fibroblasts, but only after at least 8 days of cell culture. PCR-DEIA is thus recommended because of its sensitivity and convenience for detecting early parasitemia in the surveillance of toxoplasmosis among pregnant women and immunocompromised hosts. The courses of infection in mice infected with two strains of T. gondii were compared. Tachyzoites of the virulent strain T. gondii RH, killing the host in 4 days, were identified in urine specimens and blood samples of mice 24 to 94 h after inoculation but not in brains, but no antibodies were detected. After intraperitoneal inoculation with cysts of the low-level virulence Beverley strain of T. gondii, parasites were identified in blood samples 4 days later and up to 17 days (but not in urine specimens) and in the brain from day 6 through day 525. By ELISA, high antibody titers were found from day 11 to day 525, with parasitemia preceding the appearance of antibodies. The usefulness of PCR-DEIA tests in conjunction with the search for circulating antibodies for the early diagnosis of toxoplasmosis in humans is discussed.

Molecular characterization of Vibrio cholerae O139 isolates from Asia.

In 1992, a serologically novel clone of Vibrio cholerae, designated O139, caused large epidemics of diarrhea in India and Bangladesh. To determine the extent of the spread of V. cholerae O139 worldwide, 484 V. cholerae non-O1 strains isolated from different patients with diarrhea in Thailand, Indonesia, the Philippines, and Peru in 1993 were tested for agglutination in O139 antisera. One hundred fifty-one of these 484 isolates were examined for genes encoding cholera toxin, zonula occlulans toxin, the repetitive sequence 1, and the toxin coregulated pilin A (the V. cholerae virulence gene complex). Thirty-three percent (122 of 364) of V. cholerae non-O1 strains isolated from different patients with diarrhea in Thailand agglutinated in O139 antisera. Ninety-eight percent (120 of 122) of V. cholerae O139 contained the V. cholerae virulence gene complex. None of the 104 V. cholerae non-O1 strains isolated from patients with diarrhea in Indonesia or the 14 strains from patients with diarrhea in the Philippines were serotype O139. Four different ribotypes were found in V. cholerae O139 isolated in Asia. Twenty-three (47%) of 49 Thai O139 strains examined were of different ribotypes than isolates from India and Bangladesh; V. cholerae strains that were not O1 or O139 that were isolated from flies and water in Thailand 11 years previously in 1981 contained the same V. cholerae virulence gene complex found in V. cholerae O1 and O139. This suggests that other unidentified virulence determinants are involved in V. cholerae O139 pathogenesis.

Fractionation and characterization of two forms of peroxidase from Oryza sativa.

Peroxidase (E.C. 1.11.1.7., hydrogen donor oxidoreductase) is widely distributed and has been isolated from many higher plants (1). The wide distribution of the enzyme suggests that it could be of great biological importance. However the role that it plays in metabolism is not clear due to the large number of reactions it catalyzes and the considerable number of isozymic species (2). In tomato plants, Evans and Aldridge (3) separated out six isoperoxidases and in a later paper Evans reported 12 isoperoxidases from tomato shoots (4). A homogeneous tomato fruit peroxidase isozyme was obtained by Jen et al. (5) using hydrophobic chromatography. Isozymes were not detected in Euphorbia characias peroxidase (6), in Ipomoea batatas peroxidase (7) and in Hordeum vulgare peroxidase (8). The simultaneous presence of Cu (II) amine oxidase and peroxidase in cell walls suggests that the peroxide generated on oxidation of the amines could be utilized by the peroxidase (6,8,9). In the graminea Oryza sativa, widely distributed, an FAD amine oxidase is present that oxidizes diamines (10). In this plant we also found two isoperoxidases called perox I and II. Only perox I was purified to homogeneity and its enzymatic, physical and chemical properties have been studied.

Campylobacter diarrhea in Alexandria, Egypt.

The role of Campylobacter as a cause of bacterial diarrhea in young children in Alexandria, Egypt was investigated. Stools or rectal swabs were collected from 880 children (mean age 9.8 months) presenting to a hospital with the primary complaint of diarrhea and from 1,079 well children (mean age 8.8 months) attending a vaccination clinic. Isolation of Campylobacter was significantly (p<0.0002) more frequent from cases (17.2%) than from controls (6.4%). Campylobacter was isolated from children presenting with diarrhea more frequently than Salmonella (3% isolation rate), Shigella (2% isolation rate), or other bacterial pathogens (1% isolatoin rate). Isolation of Campylobacter was significantly more frequent during the rainy season (p<0.0012). These results implicate Campylobacter as a major bacterial cause of diarrhea for which young children are brought for medical attention in Alexandria, Egypt.

Persistent, recurring diarrhea in a colony of orangutans (Pongo pygmaeus) caused by multiple strains of Campylobacter spp.

A colony of 10 orangutans (Pongo pygmaeus) experienced persistent, recurring diarrhea caused by multiple infections with Campylobacter jejuni and C. coli. Infections appeared to have occurred through several mechanisms, including fecal-oral transmission between orangutans, and possibly transmission by houseflies contaminated with the organisms from nearby chicken feces. Among the 14 fecal and environmental C. jejuni isolates, 4 different antibiotic susceptibility profiles were detected; there were also 4 different profiles among the 8 isolates of C. coli. In 5 orangutans, there were back-to-back infections by different strains of C. jejuni, suggesting that a single C. jejuni infection may not confer protective immunity against heterologous strains circulating in the same vicinity. Transmission was effectively interrupted by environmental modifications and a 7-day course of oral erythromycin.

Oral challenge with Aeromonas in protein-malnourished mice.

The lack of an animal model for Aeromonas-associated diarrhoea has hindered progress toward understanding the pathogenesis of this potentially important enteric infection. Protein-malnourished mice were challenged orally with Aeromonas strains to determine if diminished levels of resistance would allow the induction of a diarrhoeal response. The 15 Aeromonas spp. faecal isolates used for challenge included 7 A. caviae, 4 A. hydrophila, 1 A. sobria bv. sobria, and 1 A. sobria bv. veronii from patients with diarrhoea, and 2 A. caviae from healthy volunteers. All had at least 1 known virulence marker, with the exception of 1 strain. Mice on the protein deficient diet had lost an average of 23% of their initial body weight at the time of challenge. Although mice consumed 10(8) cfu per day for a minimum of 4 days, none became ill due to Aeromonas spp. ingestion. Aeromonas spp. were isolated from 75% of faecal cultures obtained 7 days after initial challenge, indicating bowel colonization had occurred.

[Arthroplasty of the knee using total condylar prosthesis. Long term results (10 to 17 years) and survival analysis]. / Arthroplastie du genou par prothèse Total Condylar. Résultats à long terme (10 à 17 ans) et analyse de survie.

INTRODUCTION: This study refers to the first 76 Total Condylar knee prostheses that were implanted in 66 patients, with a minimum follow-up of 10 years. MATERIALS AND METHODS: There were 54 women and 12 men with an average age of 65 years (range 45-81). The diagnosis was osteoarthrosis in 63 knees and rheumatoid arthritis in 13. We were able to review 54 prostheses with an average follow-up of 13 years (10-17) using the rating system of the Hospital for Special Surgery and the Survivorship analysis. RESULTS: The results were excellent and good in 72.5 per cent knees and fair and poor in 22 per cent, 3 knees (5.5 per cent) had been revised for deep infection in 2 cases and for aseptic loosening in 1. Pain at rest was absent or mild in all but one patient; walking pain was absent in 32 knees (63 per cent), mild in 12 (23 per cent), moderate in 5 (10 per cent) and severe in 2 (4 per cent). Average range of motion was 89 degrees, ranging from 40 degrees to 120 degrees. A flexion contracture greater 5 degrees was seen in 8 knees (16 per cent). Instability in the frontal plane tested in full extension was more than 5 degrees in 10 patients (19.5 per cent). A significant deterioration of the results occurred in the last 4-5 years, since in 1988 excellent and good results were still 82 per cent. A loss of postoperative alignment of 5 degrees or more was observed in 11 knees: aseptic loosening was identified in 2 of these cases and deformation of the tibial component was seen in 1 knee. In the remaining 8 knees we observed increased lateral instability, compared to the postoperative evaluation. In this group of 8 knees we measured in the AP view the thickness of the tibial component on the lateral and medial side. We found polyethylene wear on the medial side in 4 cases. Ten of the 11 knees with loss of alignment had an early postoperative femoro-tibial axis less than 5 degrees. We observed radiolucent lines in 18 patellar and 27 tibial components. A width greater than 2 mm, correlated to an extension to 5 or more zones, was seen in 3 tibial components with aseptic loosening. Survivorship analysis using aseptic loosening, mechanical failure of the polyethylene and deep infections as end point gave a cumulative success rate at 10 years of 92 per cent% with 95 per cent confidence interval ranging from 85.1 to 98. DISCUSSION: Our study reports a lower percentage of excellent and good results if compared to other series of Total Condylar prosthesis with a comparable follow-up. A significant deterioration of the results occurred in the last years, but this was often not related to the knee itself but to poor general conditions in some patients. In this series there is a relevant number of knees with a post-operative alignment that today we don't consider correct and all the aseptic loosenings occurred in knees with a tibial component positioned in varus. CONCLUSION: Survivorship analysis gave a cumulative success rate comparable with those reported in other studies and confirmed the durability and longevity of this model of prosthesis.

Modified CAMP test for biogrouping Vibrio cholerae O1 strains and distinguishing them from strains of V. cholerae non-O1.

A modified CAMP test was used to identify 973 Vibrio cholerae isolates by phenotype. Eltor and non-O1 strains were CAMP positive; classical strains were CAMP negative. Sausage-shaped zones of hemolysis of eltor strains were easily distinguished from narrower bands of non-O1 isolates. For O1 isolates, there was 100% agreement between the CAMP test and inhibition by polymyxin B.

Safety, immunogenicity, and transmissibility of single-dose live oral cholera vaccine strain CVD 103-HgR in 24- to 59-month-old Indonesian children.

Recombinant A-B+ Vibrio cholerae O1 strain CVD 103-HgR is a safe, highly immunogenic, single-dose live oral vaccine in adults in industrialized countries. Safety, excretion, immunogenicity, vaccine transmissibility, and environmental introduction of CVD 103-HgR were investigated among 24- to 59-month-old children in Jakarta. In 81 households, 1 child was randomly allocated a single dose of vaccine (5 x 10(9) cfu) and another, placebo. Additionally, 139 unpaired children were randomly allocated vaccine or placebo. During 9 days of follow-up, diarrhea or vomiting did not occur more often among vaccines than controls. Vaccine was minimally excreted and was isolated from no controls and from 1 (0.6%) of 177 unvaccinated family contacts. A 4-fold or higher rise in serum vibriocidal antibody was observed in 75% of vaccines (10-fold rise in geometric mean titer over baseline). Of 135 paired placebo recipients or household contacts, 5 had vibriocidal seroconversions. Moore swabs placed in sewers and latrines near 97 households failed to detect vaccine. These observations pave the way for a large-scale field trial of efficacy.

Enterotoxigenic Escherichia coli and other causes of infectious pediatric diarrheas in Jakarta, Indonesia.

A hospital stool survey of Indonesian children less than 5 years of age determined the prevalence of diarrhea caused by enterotoxigenic Escherichia coli (ETEC) and other bacterial enteropathogens, compared to non-diarrheic control patients. ETEC were the second most frequent cause of diarrhea, isolated from 16 of 194 (8.2%) of patient's stools compared to 2 of 97 (2.1%) of control stools. The highest prevalence was in infants 12 to 23 months of age (17.9%).

Use of vaginal tampons in sewer surveys for non-O1. Vibrio cholerae.

Vaginal tampons were shown to be a practical alternative to conventional Moore swabs for isolating Vibrio cholerae from sewage. Associated laboratory investigations demonstrated improved isolation of V. cholerae by using 12- or 18-h enrichments in alkaline peptone water, in comparison with 6-h enrichments, when cultures were incubated at ambient temperatures.

Detection of immunoglobulin A in urine specimens from children with Campylobacter-associated diarrhea by a chemiluminescent indicator-based western immunoblot assay.

A Western blot (immunoblot) assay was used to detect Campylobacter-specific immunoglobulin A in urine. Acute-phase urine samples from six children with Campylobacter diarrhea had titers ranging from 2 to 8. The highest titer was detected 4 days postonset. Campylobacter-specific immunoglobulin A was undetectable in the paired convalescent-phase specimens and urine samples from three control children.

[Comparison of Gunther, Filcard and LGM definitive caval filters. Our experience]. / Filtri cavali definitivi Gunther, Filcard e LGM a confronto. Nostra esperienza.

The authors report their experience with 30 caval filters placed throughout 3 years, and compare it with the data from the literature. In 8 patients (26.5%) Gunther filters were used, in 16 cases (53.5%) Filcard filters (4 DF-01 and 12 DF-04) and finally the last 6 patients (20%) were treated with LGM filters. In 18 cases (60%) the filters were positioned through right common femoral vein, whereas in the remaining 12 patients (40%) they were placed through the right internal jugular vein. Early complications were 1 mispositioning (3.3%) with a LGM filter, 2 incomplete opening (6.6%) of Filcard DF-01 filters, 3 significant tiltings (10%) with Filcard DF-04 filters. One patient (3.3%) died one week after the placement of a Gunther filter. The radiologic follow-up, which included controls with conventional radiology, US, cavography, MR and CT revealed no later complications. All the filters exhibited both advantages and disadvantages. Although all the filters commonly in use are effective to prevent pulmonary embolism, further experience is necessary to find out the "ideal" filter.

Campylobacter-associated diarrhoea in Egyptian infants: epidemiology and clinical manifestations of disease and high frequency of concomitant infections.

A diarrhoeal disease survey in Alexandria, Egypt determined the prevalence, seasonality, and household risk factors for Campylobacter-associated diarrhoea in young children. The study population was 880 children (mean age = 9.8 months) presenting with diarrhoea at one of two hospitals. A control group consisted of 1,079 healthy children (mean age = 8.8 months) attending two nearby vaccination clinics. The overall isolation frequencies for Campylobacter spp. were 16.8% for cases and 6.4% for the comparison group (p = 1 x 10(-8)). Other enteropathogens detected in diarrhoeal stools were: rotavirus (28.6%), Giardia lamblia trophozoites (21.3%), enterotoxigenic Escherichia coli (8.7%), Entamoeba histolytica (3.9%), Salmonella spp. (2.7%), and Shigella spp. (1.8%). There were few or no isolates of Aeromonas spp., Vibrio spp., Yersinia spp., or Plesiomonas spp. Comparisons among cases showed that Campylobacter spp. isolations were more prevalent during the rainy season (p = 0.001) and positively associated with keeping fowl in the home (p = 0.003) or having an outdoor source of drinking water (p = 0.029). Among Campylobacter-positive diarrhoeal patients, 69.0% had faecal leukocytes present and 16.3% had bloody stools. Patients with Campylobacter-positive diarrhoeal stools were frequently co-infected with rotavirus (28.6%) or G. lamblia (24.5%).

[Outbreak of paratyphoid fever among Peruvian naval personnel]. / Brote de fiebre paratifoidea entre personal de la marina del Perú.

A retrospective epidemiological study was conducted in connection with an outbreak of paratyphoid fever at a Peruvian naval installation in Callao. The study sought to determine the magnitude of the outbreak, the source of infection, the attack rates, the persistence of bacilli excretion, and the clinical picture of the disease. The source of Salmonella paratyphi B infection had been a meal of chicken and rice served to around 400 members of the naval police. Over a period of three weeks, 21 persons were hospitalized and 52 received outpatient treatment at the naval hospital. In addition, through a questionnaire it was revealed that 86 unreported cases of diarrhea related to the outbreak had occurred. The most common clinical manifestations were fever, headache, weakness, anorexia, abdominal pain, and diarrhea. The general attack rate was 39.8%. In a follow-up survey carried out 37 days after exposure, fecal cultures indicated that 8.5% of the persons affected continued to excrete the microorganism. The high rates of attack and transmission of S. paratyphi B in this outbreak point up the considerable pathogenicity and virulence of some strains of the microorganism and their impact on public health. It is suggested that preventive measures be taken at naval and other similar installations, including the education of workers who handle and prepare food, in order to ensure proper hygiene.

The etiology of diarrhea among American adults living in Peru.

During 1984-1989, 655 diarrheic and 287 nondiarrheic stool specimens from adult U.S. citizens living in Lima, Peru were tested for presence of bacterial enteropathogens. Frequencies of isolation among diarrheic specimens were: Shigella 9.8%; Campylobacter 6.1%; enterotoxigenic Escherichia coli (ETEC) 6.0%; Plesiomonas 2.0%; Salmonella 1.4%; and Vibrio 0.6%. Isolates recovered from non-diarrheic stools were: Shigella 4.5%; Campylobacter 2.1%; Salmonella 1.0%; ETEC 0.7%; Plesiomonas 0.7%; and Vibrio 0.3%. Aeromonas, an unproven cause of diarrhea, was isolated from 9.2% of cases and 3.5% of controls. Disease occurrence was strongly associated with isolation of Shigella, ETEC, Campylobacter, or Aeromonas (p less than or equal to 0.01). During the 6-year period of study, shifts in the dominant phenotypes of Shigella and Campylobacter occurred which may have important implications for vaccine development and intervention strategies.