Classically Salmonella enterica serovar Typhi (S. Typhi) is associated with typhoid, a major health problem in developing countries. However, in recent years S. Paratyphi A and Vi-negative variants of S. Typhi have emerged rapidly. We have developed a nested multiplex PCR targeting five different genes for differential diagnosis of typhoidal pathogens which has been optimized to be directly applicable on clinical blood samples. Of 42 multiplex PCR-positive blood samples, 26, nine, and two were Vi-positive S. Typhi, Vi-negative S. Typhi and S. Paratyphi A, respectively, and five patients were found to have mixed infection. Seventeen patients grew Salmonella from blood culture and the remaining 25 were positive in the Salmonella-specific PCR. Tests with several common pathogens confirmed the specificity of the assay. We conclude that the proposed multiplex PCR is rapid, sensitive and specific for the diagnosis of typhoidal pathogens directly from blood samples.

This study primarily aimed to estimate the association between influenza vaccination and the occurrence of hospitalization for acute respiratory or cardiovascular diseases, or all-cause death during the influenza season in an elderly population in South Africa. We conducted a nested case-control study using data from a cohort of 45 522 elderly members of a private medical funding organization during the moderate 2004 influenza season. In 1282 (2·8%) subjects the combined outcome occurred and the influenza vaccination rate in controls was 15·4%. After adjustments for measured confounders, vaccination was associated with a statistically significant reduction of 19% (95% confidence interval 3·1–32·9) in the combined outcome. Post-hoc sensitivity analysis of the potential impact of potential healthy user bias showed that confounding, if present, could have caused this finding. Our data were inconclusive regarding the benefits of influenza vaccination in elderly persons in South Africa and given the low vaccine uptake, long-term follow-up is warranted.

Salmonella infection in breeding pigs was the subject of a European survey in 2008. The prevalence of pig-breeding holdings infected with Salmonella was determined by microbiological culture of pooled pen faecal samples. The objective of this study was to estimate the sensitivity of pooled faecal sampling and to calculate the required sample sizes. To do this, individual and pooled faecal samples were collected from a sample of pens from nine farms. Bayesian methods were used to estimate the sensitivity of individual and pooled faecal sampling, and the degree of clustering of Salmonella at the pen level. Sample sizes were then calculated for various values of design prevalence, taking into account the clustering. Pooling was highly efficient compared to individual sampling, e.g. with 18 pooled samples required to detect a 10% prevalence with 95% certainty, compared to 35 individual rectal samples. We recommend that pooled sampling is used for detection of Salmonella in pigs. Results were influenced by the degree of clustering at pen level, and it is important to take this into account both in the estimation of appropriate sample sizes and the estimation of prevalence from pooled sample data.

Rift Valley fever virus (RVFV) is a mosquito-transmitted bunyavirus (genus Phlebovirus) associated with severe disease in livestock and fatal encephalitis or haemorrhagic fever in a proportion of infected humans. Although live attenuated and inactivated vaccines have been used in livestock, and on a limited scale in humans, there is a need for improved anti-RVFV vaccines. Towards this goal, Sindbis virus replicon vectors expressing the RVFV Gn and Gc glycoproteins, as well as the non-structural nsM protein, were constructed and evaluated for their ability to induce protective immune responses against RVFV. These replicon vectors were shown to produce the RVFV glycoproteins to high levels in vitro and to induce systemic anti-RVFV antibody responses in immunized mice, as determined by RVFV-specific ELISA, fluorescent antibody tests, and demonstration of a neutralizing antibody response. Replicon vaccination also provided 100% protection against lethal RVFV challenge by either the intraperitoneal or intranasal route. Furthermore, preliminary results indicate that the replicon vectors elicit RVFV-specific neutralizing antibody responses in vaccinated sheep. These results suggest that alphavirus-based replicon vectors can induce protective immunity against RVFV, and that this approach merits further investigation into its potential utility as a RVFV vaccine.

Demographic and clinical risk factors are important in guiding vaccination policy for pneumococcal pneumonia. We present data on these variables from a population-based surveillance network covering adult bacteraemic pneumococcal pneumonia (BPP) in the Delaware Valley region from 2002 to 2004. Surveillance data were used with U.S. Census data and a community health survey to calculate stratified incidence rates. Missing data were handled using multiple imputation. Overall rates of adult BPP were 10·6 cases/100 000 person-years. Elevated rates were seen in the elderly (>65 years), Native Americans, African Americans, the less-educated (less than high-school education), the poor, smokers, and individuals with histories of asthma, cancer, or diabetes. Multivariable modelling suggested that income was more robustly associated with risk than African American race. Of methodological interest, this association was not apparent if census block-group median income was used as a proxy for self-reported income. Further research on socioeconomic risk factors for BPP is needed.

To investigate two clusters of diarrhoea cases observed in our geriatric hospital wards, the faecal specimens were analysed. Reversed passive latex agglutination assay revealed that 63·2% and 41·7% of the faecal specimens from each cluster were positive for Clostridium perfringens enterotoxin. PCR assay revealed that 71·4% and 68·8% of C. perfringens isolates from each cluster were positive for the enterotoxin gene (cpe). These observations suggested that both the clusters were outbreaks caused by enterotoxigenic C. perfringens. Subsequent pulsed-field gel electrophoresis analysis revealed that the two outbreaks were caused by different C. perfringens isolates. However, these outbreak isolates as well as other sporadic diarrhoea isolates shared a 75-kb plasmid on which the cpe gene and the tcp locus were located. The 75-kb plasmid had horizontally spread to various C. perfringens isolates and had caused outbreaks and sporadic infections. However, the site and time of the plasmid transfer are unclear.

We evaluated the effectiveness of a measles vaccine campaign in rural Kenya, based on oral-fluid surveys and mixture-modelling analysis. Specimens were collected from 886 children aged 9 months to 14 years pre-campaign and from a comparison sample of 598 children aged 6 months post-campaign. Quantitative measles-specific antibody data were obtained by commercial kit. The estimated proportions of measles-specific antibody negative in children aged 0–4, 5–9 and 10–14 years were 51%, 42% and 27%, respectively, pre- campaign and 18%, 14% and 6%, respectively, post-campaign. We estimate a reduction in the proportion susceptible of 65–78%, with ~85% of the population recorded to have received vaccine. The proportion of ‘weak’ positive individuals rose from 35% pre-campaign to 54% post-campaign. Our results confirm the effectiveness of the campaign in reducing susceptibility to measles and demonstrate the potential of oral-fluid studies to monitor the impact of measles vaccination campaigns.

Thermotolerant Campylobacter spp. have been the most common bacterial cause of human gastrointestinal disease in Denmark since 1999. In 2003, the Danish voluntary strategy to control Campylobacter was intensified. The focus was on biosecurity, allocation of meat from Campylobacter-negative broilers to the production of chilled products, and consumer information campaigns. From 2002 to 2007, the percentage of Campylobacter-positive broiler flocks at slaughter decreased from 43% to 27%. After processing, Campylobacter-positive samples of chilled broiler meat fell from 18% in 2004 to 8% in 2007. Furthermore, the number of registered human Campylobacter cases decreased by 12%; from 4379 cases in 2002 to 3865 cases in 2007. We believe that the observed decrease in the occurrence of Campylobacter in broilers and broiler meat and the coincidental fall in the number of registered human cases is, in part, a result of the implemented control strategy.

In this prospective cluster randomized controlled trial we evaluated the impact of short-term provision of enhanced infection control support on infection control practice in nursing homes in South London. Twelve nursing homes were recruited, six each in intervention (300 residents) and control (265 residents) groups. Baseline observations of hand hygiene facilities, environmental cleanliness and safe disposal of clinical waste showed poor compliance in both groups. Post-intervention observations showed improvement in both groups. There was no statistical difference between the two groups in the compliance for hand hygiene facilities (P=0·69); environmental cleanliness (P=0·43) and safe disposal of clinical waste (P=0·96). In both groups, greatest improvement was in compliance with safe disposal of clinical waste and the least improvement was in hand hygiene facilities. Since infection control practice improved in intervention and control groups, we could not demonstrate that provision of short-term, enhanced, infection control support in nursing homes had a significant impact in infection control practice.

In industrialized countries enterotoxigenic Escherichia coli (ETEC) is mainly diagnosed as a cause of travellers' diarrhoea, but it is also known to cause foodborne outbreaks. We report an outbreak of acute gastroenteritis caused by ETEC serotypes O92:H− and O153:H2 as well as Salmonella Anatum, which affected around 200 students and teachers after a high-school dinner in Greater Copenhagen, Denmark, November 2006. A retrospective cohort study showed that consumption of pasta salad with pesto was associated with an increased risk of illness (attack rate 59·4%; risk ratio 2·6, 95% confidence interval 1·2–5·7). Imported fresh basil used for preparation of the pesto was the most likely source of contamination. Although ETEC is associated with travellers' diarrhoea in Denmark, this outbreak suggests that a proportion of sporadic ETEC infections might be caused by contaminated imported foodstuffs. To improve food safety further, it is important to target this poorly regulated and researched area.

Salmonella infections in Antarctic wildlife were first reported in 1970 and in a search for evidence linking isolations with exposure to human activities, a comparison was made of serovars reported from marine fauna in the Antarctic region from 1982–2004 with those from marine mammals in the Northern hemisphere. This revealed that 10 (83%) Salmonella enterica serovars isolated from Antarctic penguins and seals were classifiable in high-frequency (HF) quotients for serovars prevalent in humans and domesticated animals. In Australia, 16 (90%) HF serovars were isolated from marine birds and mammals compared with 12 (86%) HF serovars reported from marine mammals in the Northern hemisphere. In Western Australia, HF serovars from marine species were also recorded in humans, livestock, mussels, effluents and island populations of wildlife in urban coastal areas. Low-frequency S. enterica serovars were rarely detected in humans and not detected in seagulls or marine species. The isolation of S. Enteritidis phage type 4 (PT4), PT8 and PT23 strains from Adélie penguins and a diversity of HF serovars reported from marine fauna in the Antarctic region and coastal areas of Australia, signal the possibility of transient serovars and endemic Salmonella strains recycling back to humans from southern latitudes in marine foodstuffs and feed ingredients.

This study examined the impact of meteorological conditions on sporadic, community-acquired cases of Legionnaires' disease in England and Wales (2003–2006), with reference to the 2006 increase in cases. A case-crossover methodology compared each case with self-controlled data using a conditional logistic regression analysis. Effect modification by quarter and year was explored. In total, 674 cases were entered into the dataset and two meteorological variables were selected for study based on preliminary analyses: relative humidity during a case's incubation period, and temperature during the 10–14 weeks preceding onset. For the quarter July–September there was strong evidence to suggest a year, humidity and temperature interaction (Wald χ2=30·59, 3 d.f., P<0·0001). These findings have implications for future case numbers and resource requirements.

Legionnaires' disease (LD) is a major cause of severe community-acquired pneumonia but the source and mode of transmission are not always apparent, especially in sporadic cases. We hypothesized that LD can be acquired from the air-conditioning systems of motor cars. Swabs were taken from the evaporator compartments of the air-conditioning system of scrapped cars. Healthy subjects who were mainly employees of regional transportation companies were tested for antibody to Legionella pneumophila serogroups 1–6; they also completed a questionnaire. Legionella species were detected in 11/22 scrapped cars by the loop-mediated isothermal amplification method. The prevalence of microplate agglutination titres ⩾1:32 was significantly higher in subjects who sometimes used car air-conditioning systems. Although we did not prove a direct link between Legionella spp. in the car evaporator and LD, our findings point to a potential risk of car air-conditioning systems in LD, which needs further investigation.

The feasibility of a rubella screening and vaccination programme for unvaccinated young women was assessed after the 2004/2005 epidemic in The Netherlands. All 640 young women in two villages with low vaccination coverage were invited for a rubella seroprevalence test. Information on vaccination status was gathered by written questionnaire. Women testing seronegative were offered free rubella vaccination. The feasibility of the programme was evaluated in terms of participation, rubella susceptibility, and acceptance of the vaccination offer by seronegative women. The participation rate was 48% [95% confidence interval (CI) 44–52] with 108 unvaccinated participants. Eleven per cent (95% CI 6–19) of the women were identified as susceptible to rubella, of whom 17% (95% CI 2–48) accepted the vaccination offer. In the end only 0·9% (95% CI 0·1–2·5) of the target population was given protection by the programme. Under the present conditions this programme proved to be an inefficient strategy for rubella protection.

The incidence of dengue infection, a vector-borne disease transmitted by the mosquito Aedes aegypti, shows clear dependence on seasonal variation. Based on the quantification method that furnishes the size of the A. aegypti population in terms of the estimated entomological parameters for different temperatures, we assessed the risk of dengue outbreaks. The persistence and severity of epidemics can be assessed by the basic reproduction number R0, which varies with temperature. The expression for R0 obtained from ‘true’ and ‘pseudo’ mass action laws for dengue infection is discussed.

Most studies reporting pneumonia morbidity are restricted to hospitalized patients, although only a minority of pneumonia patients are admitted to hospital. To get a better understanding of the burden of disease in the general population, we conducted a population-based retrospective study to examine trends in pneumonia incidence in general practice, hospitalization, and mortality due to pneumonia in The Netherlands between 1997 and 2007. Between 2001/2002 and 2006/2007 there was an adjusted yearly increase of 12% in the clinical diagnosis of pneumonia in patients consulting general practitioners. Hospitalizations increased 5% per year between 1999/2000 and 2006/2007, while mortality annually decreased by 2% between 1997/1998 and 2006/2007. Our study suggests that the morbidity of pneumonia in the Dutch population increased considerably over this period, especially in primary-care settings, and that focusing only on hospitalization might underestimate the increasing public health burden of pneumonia.

Fourteen strains of S. Typhi (n=13) and S. Paratyphi A (n=1) resistant to ciprofloxacin were compared with 30 ciprofloxacin decreased-susceptibility strains on the basis of qnr plasmid analysis, and nucleotide substitutions at gyrA, gyrB, parC and parE. In ciprofloxacin-resistant strains, five S. Typhi and a single S. Paratyphi A showed triple mutations in gyrA (Ser83→Phe, Asp87→Asn, Glu133→Gly) and a novel mutation outside the quinolone resistance determining region (QRDR) (Met52→Leu). Novel mutations were also discovered in an isolate (minimum inhibitory concentration 8 μg/ml) in gyrA gene Asp76→Asn and outside the QRDR Leu44→Ile. Out of 30 isolates with reduced susceptibility, single mutation was found in 12 strains only. Genes encoding qnr plasmid (qnr A, qnr B, AAC1-F) were not detected in ciprofloxacin-resistant or decreased-susceptibility strains. Antimicrobial surveillance coupled with molecular analysis of fluoroquinolone resistance is warranted for reconfirming novel and established molecular patterns of resistance, which is quintessential for reappraisal of enteric fever therapeutics.

Multiple gastroenteritis outbreaks occurred between 25 and 31 July 2006 in 10 workplace canteens in south-western Finland. One vegetable processing plant provided raw vegetables to all the canteens. We conducted cohort studies in the three most visited canteens and environmental investigations in the kitchens and the plant. Patients' stools, food, water and environmental samples were tested for enteric bacteria and viruses. Of the three canteens, 150/273 respondents (response rate 82%) had gastroenteritis. Consumption of mixed raw vegetables was significantly associated with the illness but no single vegetable explains the outbreak. An identical norovirus GII.1 genotype was detected from all genotyped patient samples. Water, food, and environmental samples were negative for norovirus. The facilities had appropriate hygienic conditions and no staff member had gastroenteritis prior to the outbreak. Tracing back the vegetables to the farm level proved unsuccessful. This was the largest foodborne norovirus outbreak in Finland.

Since 2003, Connecticut laboratories have reported Gram-positive rod (GPR) isolates detected within 32 h of inoculation from blood or cerebrospinal fluid. The objectives were to rapidly identify inhalational anthrax and unusual Clostridium spp. infections, and to establish round-the-clock laboratory reporting of potential indicators of bioterrorism. From 2003 to 2006, Connecticut's GPR surveillance system identified 1134 isolates, including 657 Bacillus spp. (none B. anthracis) and 241 Clostridium spp. Reporting completeness and timeliness improved to 93% and 92%, respectively. Baseline rates of Bacillus spp., Clostridium spp. and other GPR findings have been established and are stable. Thus far, no cases of anthrax and no unusual clusters of Clostridium spp. have been detected by the GPR surveillance system. This system would probably have confirmed the inhalational anthrax case in Pennsylvania in 2006 3 days sooner than traditional reporting. Using audits and ongoing evaluation, the system has evolved into a highly functional 24/7 laboratory telephone reporting system with almost complete reporting.