Thirty-five E. coli strains belonging to O-serogroups associated with enteroinvasive types of Escherichia coli (EIEC) isolated in Germany between 1989 and 1995 were investigated for invasivity-associated DNA sequences. Only 11 strains were positive for ipaH and thus confirmed as EIEC. All 11 EIEC isolates originated from human infections which were imported to Germany from Eastern Europe. EIEC O124 were most frequent and originated from asymptomatic Romanians arriving at Rostock, Germany in 1992 and 1993. In January 1993, EIEC O124 were isolated from faeces of a laboratory technician with diarrhoea working at the enteric pathogen department of the Institute of Hygiene in Rostock. By comparing her E. coli O124 isolate with recently imported O124 strains for Xba I restriction fragment length polymorphisms (RFLP) the probable source of infection could be determined. Four major RFLP patterns were found in the group of O124 strains. O124 strains with identical RFLP patterns were found in the group of O124 strains. O124 strains with identical RFLP patterns were isolated from people who were in close contact to each other.

To obtain insights into the pathogenic mechanisms involving Mycobacterium paratuberculosis in Crohn's disease (CD) we questioned if the strains of M. paratuberculosis isolated from CD are distinguishable from those involved in Johne's disease (JD), a chronic granulomatous enteritis in cattle. Accordingly we compared human and animal strains at the DNA level, both by the analysis of restriction fragment length polymorphism (RFLP) in and around the insertion sequence IS 900 and by the arbitrarily primed polymerase chain reaction (AP-PCR). Results are in favour of a common clonal origin for the 4 strains isolated from CD and for 8 of the 11 strains isolated from cattle and sheep JD.

This study assessed accuracy of (a) recording Vibrio vulnificus infection on death certificates and (b) International Classification of Disease (ICD)-9 codes for V. vulnificus. Patients with microbiologically confirmed V. vulnificus infection were identified as part of co-ordinated surveillance in four USA Gulf Coast states between 1989 and 1993. Of 60 deaths, 51 death certificates were reviewed and V. vulnificus was recorded as the immediate cause of death on 11 (22%). There was no ICD-9 code for V. vulnificus infection, thus no patients had an ICD-9 code indicating V. vulnificus infection. Of 23 certificates where V. vulnificus was recorded on the death certificate, only 5 (22%) were coded for Gram-negative, septicaemia. This study highlights the importance of teaching physicians how to provide epidemiologically meaningful data on death certificates and the need for accurate ICD mortality codes.

In this study, we investigated the frequency of co-existence of cerebral cysticercosis (CC) in Japanese encephalitis (JE) cases with special emphasis on its role in predicting the final clinical outcome. Amongst the 163 confirmed cases of JE, 37·42% (61/163) had co-existent CC. This was confirmed by antibody detection in the CSF of 45 cases, CT scan of the brain in 6 cases and at autopsy in 3 cases. In 2 cases confirmation was possible by CT scan as well as at autopsy, in 4, CSF antibody levels and CT scan were suggestive of CC while in 1, CSF antibodies and autopsy were suggestive of CC. The co-occurrence of Cysticercus cellulosae in the brain emerged as a prognosticator of poor outcome in JE cases (P<0·03).

Rapid, non-culture, serogroup determination of meningococcal infection is important in contact management where vaccination may be possible. The impending availability of polysaccharide–protein conjugate vaccines for serogroup C disease requires maximal case ascertainment, with serogroup determination, at a time when the number of culture confirmed meningococcal infections is decreasing. A polymerase chain reaction assay (PCR), based on a restriction fragment length polymorphism (RFLP) in the meningococcal serogroup B and C sialyltransferase (siaD) gene, was developed to combine the non-culture diagnosis of meningococcal infection from CSF, whole blood and serum with serogroup (B and C) identification. The PCR assay was adapted to an ELISA format incorporating hybridization with serogroup-specific B and C oligonucleotide probes. Specificity for CSFs was 100% and sensitivities were respectively 81, 63 and 30% for CSFs, whole blood and sera. The serogroup-specific PCR ELISA is a significant addition to currently available tests for non-culture diagnosis of meningococcal infection and outbreak investigation.

An examination of salmonella isolates collected by the Scottish Agricultural College Veterinary Services Division from April 1994 to May 1995 was conducted to determine the extent to which Salmonella enterica serotype Typhimurium phage type 104 (DT104) occurred and to investigate the antimicrobial resistance patterns of isolates. Typhimurium DT104 was the predominant salmonella and was isolated from nine species of animal. All isolates of this phage type possessed resistance to at least one antimicrobial and 98% of the isolates were resistant to multiple antimicrobials with R-type ACTSp the predominant resistance pattern. Various other resistance patterns were identified and transferable resistance to the veterinary aminoglycoside antimicrobial apramycin was demonstrated in three strains. A retrospective study for gentamicin resistance in isolates from the Scottish Salmonella Reference Laboratory collection revealed a human isolate of Typhimurium DT104 resistant to gentamicin but sensitive to apramycin and a bovine isolate with apramycin and gentamicin resistance.

Sixty-eight isolates of Campylobacter jejuni and C. coli isolated from patients with diarrhoea (n=630) and controls (n=220) at Tikur Anbassa Hospital, Addis Ababa, Ethiopia were serotyped on the basis of the heat-labile (HL) and the heat-stable (HS) antigens, by using 16 and 34 antisera, respectively, for the two methods. With the antisera against heat labile antigens, 89–3% of the C. jejuni and 75% of the C. coli were typable. The HL serotypes 1, 2, 3, 4, 5, 6 and 7 were the most common among the C. jejuni while HL serotypes 1 and 2 were dominant among the C. coli isolates. These serotypes accounted for 63·2% of all isolates. For the heat-stable antigens, 60% of the C. jejuni and 83·3% of the C. coli isolates were typable. The HS serotypes 1, 3, 8, 26 and 34 were most common among the C. jejuni, while serotypes 3 and 8 were dominant among C. coli isolates. This study shows that the most common HL and HS antigens among campylobacter isolates from Ethiopia correspond to the most frequent antigenic types from other parts of the world. A limited number of antisera were sufficient to identify the majority of the isolates.

Escherichia coli O157 shedding in 14 cattle herds was determined by faecal culture at intervals of approximately 1 month for up to 13 months. The overall prevalence was 1·0% (113/10832 faecal samples) and 9 of the 14 herds were detected as positive. Herds positive 2 years previously (n=5) had a higher prevalence of positive cattle (median=1·9%) than herds which had been negative on a previous sampling (n=8, median=0·2%). Weaned heifers had a higher prevalence (1·8%) than did unweaned calves (0·9%) or adults (0·4%). For all herds the highest prevalence occurred in the summer months, which resulted in most of the positive faecal samples being collected on a minority of sampling visits.

The 16S-23S RNA gene intergenic spacers of isolates of Streptococcus equi (n=5), S. zooepidemicus (n=5), S. equisimilis (n=3) and S. dysgalactiae (n=2) were sequenced and compared. There were distinct regions within the spacer, arranged in the order 1–9 for all S. equi and one S. zooepidemicus isolate and 1,2 and 4–9 for the remaining isolates. Region 4 was identical to the tRNAala gene found in the 16S-23S intergenic spacers of other streptococci. Regions 1, 5, 6 and 7 had distinct variations, each conserved in different isolates. However, amongst the intergenic spacers there were different combinations of variant regions, suggesting a role for DNA recombination in their evolution. The intergenic spacer of all isolates of S. equi and one S. zooepidemicus isolate were almost identical. Primers derived from the variant sequences of regions 1 and 5 to 6 were used to group all S. zooepidemicus (n=17) and S. equi (n=5) into 1 of 8 types by polymerase chain reaction; three S. zooepidemicus isolates typed the same as S. equi. S. equi and S. zooepidemicus were clearly distinguishable from S. equisimilis and S. dysgalactiae which had shorter regions 5 and 6 and no region 7. Most homology for the group C sequences was found in previously published sequences for the 16S-23S intergenic spacers of S. anginosis, S. constellatus, S. intermedius, S. salivarius and S. agalactiae. A 75-90 nucleotide length shared with S. anginosus and S. intermedius in opposite orientations in the two main variants of region 6 supported the role for DNA recombination in the evolution of the spacer. The 16S-23S intergenic spacers indicate that S. zooepidemicus was the archetypal species for S. equi and that both are genetically more distant from S. equisimilis and S. dysgalactiae. The intergenic spacer can be used to identify specifically the group C. streptococci and as an epidemiological marker for S. zooepidemicus.

The prevalence of chronic Chlamydia pneumoniae infection was assessed in 54 patients with established chronic obstructive pulmonary disease (COPD), 41 of these with severe COPD (group I), 13 with mild to moderate COPD (group II), and in 23 patients with community-acquired pneumonia (controls, group III). Specific IgG and IgA antibody levels and circulating immune complexes (ICs) were measured in paired sera, and specific secretory IgA (sIgA) levels in sputum specimens. A polymerase chain reaction (PCR) test was used for the detection of C. pneumoniae in sputum. According to our definite diagnosis criterion, 65% of the COPD patients showed evidence of suspected chronic C. pneumoniae infection and the prevalence was still higher (71%) in patients with severe disease. The occurrence of specific markers of infection was invariably highest in patients with severe COPD, next-highest in patients with mild to moderate COPD and lowest in pneumonia patients. The association between COPD and C. pneumoniae infection persisted after controlling for the potential confounding factors.

Previous reports in the literature suggest that Burkholderia pseudomallei strains can be differentiated on the basis of animal virulence. Twenty environmentally and clinically derived isolates of Burkholderia pseudomallei were examined for the production of exoenzymes, morphological and biochemical phenotypes and virulence for Syrian golden hamsters. The partial sequence of the 16S ribosomal RNA [rRNA] genes from a number of these strains was also determined. Based upon these observations, it is suggested that highly virulent Burkholderia pseudomallei strains are true Burkholderia pseudomallei strains. The DNA sequences of the 16S rRNA genes of the true Burkholderia pseudomallei strains were identical to the published sequences for Burkholderia pseudomallei while differences were revealed between the published sequences and those of the lowly virulent strains. Thus, these latter strains have been designated as Burkholderia pseudomallei-like organisms since they demonstrate significant differences in exoenzyme production, hamster virulence and 16S rRNA gene sequences.

Cattle persistently infected with foot-and-mouth disease virus were treated with dexamethasone to suppress the immune system in an attempt to influence the level of virus recovery from oesophageal–pharyngeal (probang) samples. Twelve carrier cattle were assigned to one of three groups: control; 0·1 mg/kg dexamethasone; and 0·5 mg/kg dexamethasone. Groups 2 and 3 were injected intramuscularly three times weekly for 3 weeks with dexamethasone between days 33 and 56 post-infection with foot-and-mouth disease virus (FMDV). Cattle in both groups developed a leucocytosis, neutrophilia and lymphopenia. The secretory IgA response to FMDV infection was inhibited following, but not during, dexamethasone treatment between days 70 and 98 post-infection (P<0·05). FMDV recovery from probang samples was reduced between days 40 and 64 post-infection (P<0·05) during treatment with either 0·1 or 0·5 mg/kg dexamethasone. Following cessation of dosing with dexamethasone virus recovery returned to control levels. These observations suggest dexamethasone inhibits shedding of FMDV in a reversible manner which may be related to its immunosuppressive, anti-inflammatory or physiological actions.

Bovine tuberculosis remains a significant problem in some parts of Great Britain and Ireland largely because of a reservoir of infection in badgers. Little is currently known about the immunopathology of Mycobacterium bovis infection in the badger. Badgers, from 31 social groups, in a study area of the Cotswold escarpment, have been trapped and sampled from 1981 to 1995. Serum antibody responses directed against the 25 kDa antigen (MPB83) of M. bovis have been studied in detail in a selected social group (JM) which has endemic infection. Sequential sera from 44 badgers were studied and results compared with culture from faeces, urine, tracheal aspirates, bite wound swabs and at post mortem. The results indicate that some badgers (about 10%) remain uninfected despite exposure to endemic M. bovis infection within the social group. In culture-positive animals active excretion of organisms is not necessarily concomitant with seropositivity. Conversely, seropositivity is not an indicator that culture positivity is present or imminent. This is particularly true in cubs when a transient seropositivity can occur within the first 6–8 months of life but these animals can remain culture-negative for up to 5 years. Western blotting confirms that at least some of these antibodies, detectable by ELISA in the culture-negative cubs, are directed against the 25 kDa M. bovis antigen. In contrast antibodies detectable in the culture-positive animals do not Western blot prior to a positive culture. Thus, differential reactivity in Western blotting may distinguish between serum antibodies indicative of potentially culture-positive animals and animals which will remain culture-negative.

Diluted dried blood drops on filter paper were compared with serum samples as a specimen source for qualitative anti-HAV antibody determination by ELISA. A total of 298 serum samples and dried blood drops were collected from a population of healthy adolescents (15·3±1·2 years old). The prevalence of anti-HAV antibody obtained by testing serum samples was 7·7% (95% CI: 4·8–10·1). Compared with serum sampling the sensitivity and specificity of diluted dried blood drops were 91·3 and 99·3%. The positive and negative predictive values were 91·3 and 99·3%, respectively, and the likelihood ratios of positive and negative results were 91 and 0·09. It is proposed that this test represents a reliable procedure for anti-HAV antibody testing.

A clinical trial was conducted in Argentina to determine the efficacy of clarithromycin plus lansoprazole for the treatment of Helicobacter pylori in duodenal ulcers and non-ulcer dyspepsia. PCR–RFLP was conducted on an 820-bp amplified product of the ureC gene of H. pylori to determine the genetic heterogeneity of 83 pretreatment and 21 post-treatment isolates. Twelve different restriction patterns were observed when digested with Sau 3A or Hha I, resulting in 40 different RFLP types. Comparison of isolates before treatment to after treatment showed that 20 of 20 patients had the same RFLP type. In addition, the presence of the cytotoxin-associated gene (cagA) and the vacuolating gene (vacA) were determined. All pretreatment isolates were positive for vacA whereas 75% of the pretreatment isolates were positive for cagA. The results of this study indicate that a high degree of heterogeneity exists among H. pylori and that infection is not limited to a small number of RFLP types.

Pulsed field gel electrophoresis (PFGE) of the genomic DNA of penicillin resistant serotype 19B Streptococcus pneumoniae was carried out. Thirteen strains form the Nagasaki area and 12 strains from other areas in Japan were examined. Twenty-three strains were resistant to erythromycin, tetracycline and trimethoprim/sulfamethoxazole but susceptible to chloramphenicol. Eight strains were resistant to ceftriaxone. All strains were multiply resistant. Five strains isolated from Nagasaki were indistinguishable from each other by using restriction enzymes Apa I and Sma I. Two strains isolated from other areas were indistinguishable from the above five strains. We could classify 13 Nagasaki strains into 3 groups and the total of 25 Japanese strains into 6 groups. These results suggest that the increasing prevalence of multiply drug resistant S. pneumoniae serotyped 19B in Japan is not due to a single clone, but at least one clone has spread widely in Japan.

Ten antisera were produced in rabbits by two or three intravenous injections of inactivated whole influenza type A virions. All contained haemagglutination-inhibition (HI) antibody directed predominantly to an epitope in antigenic site B and, in addition, various amounts of antibodies to an epitope in site A and in site D. The ability of untreated antisera to select neutralization escape mutants was investigated by incubating virus possessing the homologous haemagglutinin with antiserum adjusted to contain anti-B epitope HI titres of 100, 1000 and 10000 HIU/ml. Virus-antiserum mixtures were inoculated into embryonated hen's eggs, and progeny virus examined without further selection. Forty percent of the antisera at a titre of 1000 HIU/ml selected neutralizing antibody escape mutants as defined by their lack of reactivity to Mab HC10 (site B), and unchanged reactivity to other Mabs to site A and site D epitopes. All escape mutant-selecting antisera had a ratio of anti-site B (HC10)-epitope antibody[ratio ]other antibodies of [ges ]2·0[ratio ]1. The antiserum with the highest ratio (7·4[ratio ]1) selected escape mutants in all eggs tested in four different experiments. No antiserum used at a titre of 10000 HIU/ml allowed multiplication of any virus. All antisera used at a titre of 100 HIU/ml permitted virus growth, but this was wild-type (wt) virus. We conclude that a predominant epitope-specific antibody response, a titre of [ges ]1000 HIU/ml, and a low absolute titre of other antibodies ([les ]500 HIU/ml) are three requirements for the selection of escape mutants. None of the antisera in this study could have selected escape mutants without an appropriate dilution factor, so the occurrence of an escape mutant-selecting antiserum in nature is likely to be a rare event.