Erik Ekwall

Enteropathogens in Adult Patients with Diarrhea and Healthy Control Subjects: A 1-Year Prospective Study in a Swedish Clinic for Infectious Diseases

A 1-year prospective study was conducted to identify enteropathogens in adults with diarrhea (n=851) and in healthy control subjects (n=203) by use of conventional laboratory methods. Virulence factor genes for diarrheagenic Escherichia coli were detected by polymerase chain reaction. Enteropathogens were identified in 56% of patients and 16% of control subjects. The isolation rate was 65% for patients with symptoms for <1 week and for travelers; >1 pathogen was found in 11% of patients. The most frequent enteropathogens were Campylobacter (13% of patients), Clostridium difficile (13%), enterotoxigenic Escherichia coli (8%), Salmonella (7%), Shigella (4%), Blastocystis hominis (4%), calicivirus (3%), rotavirus (3%), enteroaggregative E. coli (2%), Aeromonas (2%), Giardia intestinalis (2%), Cryptosporidium (2%), and astrovirus (2%). Less frequently isolated (< or =1% of patients) were verotoxigenic E. coli, enteropathogenic E. coli, enteroinvasive E. coli, Entamoeba histolytica/Entamoeba dispar, microsporidia, and adenovirus. Fifty percent of the patients were hospitalized, and 43% needed intravenous fluids. The median duration of diarrhea was 14 days. Clinical features were not helpful for predicting the etiology of diarrhea.

Aetiology and epidemiology of acute gastro-enteritis in Swedish children

In a prospective 1-year study, 144 children attending or admitted to hospital and 272 children outside hospital with acute gastro-enteritis and 200 controls were investigated by a broad panel of diagnostic methods for enteropathogenic agents in the faeces and for related antibody responses. Enteropathogens were identified in 77% of the inpatients, 63% of the outpatients and 8% of the controls. Rotavirus and Yersinia enterocolitica were detected significantly more often among inpatients. Altogether, viral, bacterial and parasitic agents were found in 58%, 14% and 1% of diarrhoeal patients, respectively. The isolation of more than one pathogenic agent was uncommon (6.5%). Rotavirus (45%) and enteric adenoviruses 40 and 41 (7.9%) predominated among the viruses, while Campylobacter jejuni (4.8%) was most common among the bacteria. Clostridium difficile and/or its cytotoxin, which were found in 14% of the children with gastroenteritis and in 15% of the controls, were significantly associated with antibiotic therapy but not with gastro-intestinal illness. Diarrhoeal infections of unknown aetiology exhibited a seasonal peak in the autumn. The duration of excretion of enteropathogens was investigated. Rotavirus particles were detectable by solid-phase immune electron microscopy for 14-25 days after the diarrhoea had ceased. Transmission of rotavirus and bacterial pathogens within families was studied also.

Shigella flexneri O-antigen-specific enzyme immunoassay: class-specific antibody titres against lipopolysaccharide antigens in healthy Vietnamese and Swedish populations

Sera collected from 397 healthy Vietnamese individuals, living in two districts in the surroundings of Hanoi, and 62 healthy Swedish individuals, living in the greater Stockholm area were analysed in enzyme-immunoassays (EIA) for their class-specific immunoglobulin titres against a panel of 10 O-antigen containing lipopolysaccharides (LPS) from Shigella flexneri, Shigella dysenteriae serotype I, Shigella sonnei and Salmonella serogroups A and B. The Vietnamese population was divided into age groups: 0·1–5 year, 1–2, 3–4, 5–9, 10–14 years, etc. Young Vietnamese children had low anti-S. flexneri LPS IgA, IgG and IgM titres which reached adult levels for IgA in the 10- to 14-year-old group, and for IgG and IgM in the 3- to-4-year-old group. The IgA, IgG and IgM titres were significantly higher (P < 0·001) in sera from Vietnamese individuals than in sera from Swedish individuals. The EIA titres against S dysenteriae serotype 1 and S. sonnei LPSs were higher in the Vietnamese than in Swedish population, but to a lesser extent than seen for S. flexneri. The titres against Salmonella serogroups A and B LPS antigens did not differ significantly in the study populations. These results suggest that the incidence of S. flexneri infections is high in Vietnam, followed in frequency by S. dysenteriae serotype I and S. sonnei infections. On the contrary the incidence of salmonellosis appears to be low.

Serum Antibody Responses Against Shigella Lipopolysaccharides and Invasion Plasmid-Coded Antigens in Shigella Infected Swedish Patients

Serum antibody responses to shigella lipopolysaccharide (LPS) and invasion plasmid-coded antigens (Ipa) were studied in 74 Swedish patients with culture verified bacillary dysentery using class-specific enzyme immunoassay (EIA). Anti-LPS responses were found in 80% and 79% serum samples, respectively, from S. flexneri and S. sonnei infected patients and anti-Ipa responses in 60% and 43%, respectively. The mean anti-Ipa IgG antibody titres in S. flexneri infected patients remained high for 4-6 months after onset while the anti-LPS IgG antibody titres had dropped to normal levels. The specificity of EIA for shigella Ipa was 90% and for S. dysenteriae, S. flexneri and S. sonnei LPSs it varied between 84% and 90%. No close correlations between the anti-LPS and anti-Ipa antibody responses were observed indicating that they may be differently regulated. The dynamics of the serum antibody responses indicates that an anti-LPS response is a good indicator of a recent shigella infection and an anti-Ipa IgG response a good indicator of a previous infection.

Shigella flexneri O-antigen specific enzyme immunoassay: lipopolysaccharides and synthetic glycoconjugates as antigens

Abstract Sera were collected from either mice or rabbits immunized with heat-inactivated Shigella flexneri serotype 1b or 2a bacteria or from humans with bacillary dysentery caused by serotype 1b or 2a. The sera were assayed for their antibody response by an enzyme-immunoassay (EIA). The sensitivity and specificity of the assay was examined by a panel of phenol-water extracted lipopolysaccharides (LPS) from all S. flexneri serotypes, including a complete core rough mutant, and artificial glycoconjugates synthetic or chemically derived di-, tri-, tetra- and octasaccharides, which are elements of the O-antigenic S. flexneri polysaccharide chain, covalently linked to bovine serum albumin (BSA). In terms of sensitivity the LPS antigens were superior to the artificial glycoconjugate antigens. The chemically and immunochemically defined LPS antigens did not, however, permit a serotype-specific immunodiagnosis. The results indicated that besides antibodies against serotype-specific epitopes, antibodies also appeared to be formed against epitopes common to all serotypes. The artificial saccharide-BSA glycoconjugates were also unable to allow a serotype-specific diagnosis with one possible exception, the group-antigen 6-BSA glycoconjugate. The results suggest that S. flexneri species-specific serodiagnosis can be obtained using characterized LPS antigens.

Shigella flexneri O-antigen specific enzyme immunoassay: a prospective study of class-specific antibody titres against lipopolysaccharide antigens in Vietnamese children and adults with serotype 1b or 2a dysentery

A total of 278 sera were collected from 97 patients with a bacteriologically verified Shigella flexneri serotype 1b or 2a infection. Of the patients, 65 were children below the age of 5 years and admitted to the Department of Infectious Diseases at St. Pauls Hospital, Hanoi, Vietnam; 32 were adults, aged 20–24 years, and infected during a dysentery outbreak at Son Tay technical school, Hanoi. The sera were analysed for their specific immunoglobulin A (IgA), M (IgM) and G (IgG) titres against phenol-water-extracted and chemically defined lipopolysaccharide (LPS) antigens of S. flexneri and other shigellae by an enzyme immunoassay (EIA). The titres estimated in sera from patients were compared with titres seen in sera from age-matched healthy people living in the Tu Liem district, Hanoi. Positive titres were defined as greater than the mean titre+ 2 SD in sera from healthy persons. Children 1–2 years old and infected with S. flexneri serotype 1b responded with significantly elevated IgA titres up to 60 days after falling ill (P-values 0·06 to <0·001). The IgG titres against the homologous S. flexneri serotype 1b were significantly elevated in samples collected up to 6 months after children up to 5 years old had fallen ill (P-values ranging from 0·007 to <0·001). IgM titres were elevated, but not significantly higher than those seen in healthy individuals. The results suggested that children younger than 3 years who responded with IgA, IgG and, to a lesser extent, IgM increases were experiencing their first S. flexneri dysentery, whereas older children and adults who only showed IgG increases had had experience with previous S. flexneri infections and displayed an anamnestic response. In adults, significant titre increases were seen only in individuals with low pre-infection titres. The clinical data and laboratory analyses of the strains suggest that a virulent S. flexneri serotype 1b clone was prevalent in the Hanoi area in 1982–1983. Young children infected with S. flexneri serotype 2a also responded with elevated IgA and IgG titres, but there were too few children to permit statistical analyses.

Serum IgG Antibody Responses to Shigella Invasion Plasmid-coded Antigens Detected by Immunoblot

Serum IgG antibody responses to Shigella invasion plasmid-coded antigens (Ipa) from 58 Shigella flexneri, S. sonnei, and S. dysenteriae infected Swedish patients were investigated by immunoblot technique. Intense responses to most components of Ipa (Ipas A, B, C, D, and VirG-virulence determinant on SalI fragment G of the plasmid) were evident in sera from S. flexneri infected patients. The strongest response was to Ipa B and the weakest, to Ipa D. In contrast, there were weaker responses to Ipas A, B, C, and VirG but none at all to Ipa D in sera from S. sonnei infected patients. After absorption of the Ipa-positive sera by Ipa expressing strains of S. flexneri and S. sonnei, most IgG antibodies to components of Ipa were removed in sera absorbed by S. flexneri, but IgG antibodies to Ipas--especially to Ipa D--were only slightly reduced in sera absorbed by S. sonnei, suggesting that Ipa D in S. sonnei may not be exposed on the S. sonnei cell surface.

Shigella flexneri O-antigen specific enzyme immunoassay: class-specific antibody titres against lipopolysaccharide antigen in adult Swedes with bacillary dysentery

Abstract Serum samples were collected from 14 healthy Swedes who fell ill due to bacteriologically verified bacillary dysentery caused by Shigella flexneri of various serotypes IgA, IgM and IgM serum antibody titres were estimated in enzyme immunoassays (EIA) with chemically defined S. Flexneri serotype Y lipopolysaccharide (LPS) as antigen. Serum samples collected from 35 healthy Swedes served as controls. Positive titres were defined as titres greater than the mean +2 SD in the control population. Eleven out of fourteen (79%) patients responded with positive IgA titres within the first 4 weeks after falling ill. IgM titres were positive for 4 14 (29%) in the same serum samples and IgG titres in 13 14 (93%) of the samples. All data suggest that patients showed a primary antibody response as a result of the shigella infection. One year post infection 56% showed positive IgA titres, 18% positive IgM and 82% positive IgG titres. The sensitivity of the assay was high, all patients showed one or more positive titres. In terms of specificity, the S. flexneri serotype Y LPS EIA was highly selective for IgG titres when tested on sera from 34 patients with non-flexneri shigellosis, salmonellosis, campylobacteriosis or yersiniosis diagnosed by faecal culture: 2 34 (6%) positive. IgA titres, although positive, were lower than titres seen in sera from patients with S. flexneri infection. We consider the S. flexneri serotype Y LPS EIA a useful diagnostic complement to faecal culture.

A Comparison of the Combination Pivmecillinam/Pivampicillin and Co-trimoxazole in the Treatment of Convalescent Carriers of Salmonella and Shigella

52 convalescent carriers of Salmonellae (n = 25) and Shigellae (n = 27) were treated with a 4-week course of either co-trimoxazole or the combination pivmecillinam/pivampicillin. 84% of the salmonella isolates and 89% of the shigella were resistant to one or more antibiotics. Sulphonamide resistance was observed in 52 and 58% of the strains, respectively. 12 and 44% of the isolates, respectively, were resistant to ampicillin. All were sensitive to mecillinam and all except 2 were sensitive to co-trimoxazole. In salmonella carriers, co-trimoxazole was successful in 54% of the subjects and pivmecillinam/pivampicillin in 58%. Co-trimoxazole cured 83% of the shigella carriers and pivmecillinam/pivampicillin 87%. Shigella carriers responded to therapy promptly. Concurrent biliary disease or diverticulosis adversely affected the prognosis in salmonella carriers.

Mecillinam and ampicillin separately or combined in gram-negative septicemia

SummaryOf 20 patients with gram-negative septicemia treated with mecillinam alone or in combination with ampicillin, successful therapeutic results were obtained in 16. In 11 patients treated with ampicillin alone, three failures responded successfully to a combination of mecillinam and ampicillin. Mecillinam MIC values of isolatedEnterobacteriaceae were 0.05–0.4 µg/ml. In patients receiving 5 mg/kg mecillinam intravenously every six hours, the mean 0.5 hour concentration was 11.0 µg/ml and in those given 10 mg/kg 23.3 µg/ml. No serious side effects were recorded. One patient on mecillinam developed an exanthema, as did three patients on combined therapy.ZusammenfassungBei 20 Patienten mit gramnegative Sepsis, die mit Mecillinam allein oder in Kombination mit Ampicillin behandelt wurden, war das Therapieergebnis in 16 Fällen erfolgreich. Bei 11 mit Ampicillin allein behandelten Patienten waren drei Therapieversager, die anschließend auf eine Kombination von Mecillinam und Ampicillin ansprachen. Die Mecillinam-MHK-Werte isolierterEnterobacteriaceae betrugen 0,05–0,4 µg/ml. Bei Patienten, die 5 mg/kg Mecillinam intravenös alle sechs Stunden erhielten, betrug die mittlere 0,5 h Konzentration 11,0 µg/ml und bei denen, die 10 mg/kg erhielten, 23,3 µg/ml. Ernsthafte Nebenwirkungen wurden nicht beobachtet. Ein Patient entwickelte unter Mecillinam-Therapie ein Exanthem, ebenso drei Patienten, die die Kombinationstherapie erhielten.