Nutrient sensor O-GlcNAc transferase controls cancer lipid metabolism via SREBP-1 regulation.

Elevated O-GlcNAcylation is associated with disease states such as diabetes and cancer. O-GlcNAc transferase (OGT) is elevated in multiple cancers and inhibition of this enzyme genetically or pharmacologically inhibits oncogenesis. Here we show that O-GlcNAcylation modulates lipid metabolism in cancer cells. OGT regulates expression of the master lipid regulator the transcription factor sterol regulatory element binding protein 1 (SREBP-1) and its transcriptional targets both in cancer and lipogenic tissue. OGT regulates SREBP-1 protein expression via AMP-activated protein kinase (AMPK). SREBP-1 is critical for OGT-mediated regulation of cell survival and of lipid synthesis, as overexpression of SREBP-1 rescues lipogenic defects associated with OGT suppression, and tumor growth in vitro and in vivo. These results unravel a previously unidentified link between O-GlcNAcylation, lipid metabolism and the regulation of SREBP-1 in cancer and suggests a crucial role for O-GlcNAc signaling in transducing nutritional state to regulate lipid metabolism.

The relationship between meteorological variables and sporadic cases of Legionnaires' disease in residents of England and Wales.

We studied the timing of occurrence of 1676 sporadic, community-acquired cases of Legionnaires' disease in England and Wales between 1993 and 2008, in relation to temperature, relative humidity, rainfall, windspeed and ultraviolet light using a fixed-stratum case-crossover approach. The analysis was conducted using conditional logistic regression, with consideration of appropriate lag periods. There was evidence of an association between the risk of Legionnaires' disease and temperature with an apparently long time lag of 1-9 weeks [odds of disease at 95th vs. 75th centiles: 3·91, 95% confidence interval (CI) 2·06-7·40], and with rainfall at short time lags (of 2-10 days) (odds of disease at 75th vs. 50th centiles: 1·78, 95% CI 1·50-2·13). There was some evidence that the risk of disease in relation to high temperatures was greater at high relative humidities. A higher risk of Legionnaires' disease may be indicated by preceding periods of warmer wetter weather.

An international trial of quantitative PCR for monitoring Legionella in artificial water systems.

AIMS: To perform an international trial to derive alert and action levels for the use of quantitative PCR (qPCR) in the monitoring of Legionella to determine the effectiveness of control measures against legionellae. METHODS AND RESULTS: Laboratories (7) participated from six countries. Legionellae were determined by culture and qPCR methods with comparable detection limits. Systems were monitored over ≥10 weeks. For cooling towers (232 samples), there was a significant difference between the log mean difference between qPCR (GU l(-1) ) and culture (CFU l(-1) ) for Legionella pneumophila (0·71) and for Legionella spp. (2·03). In hot and cold water (506 samples), the differences were less, 0·62 for Leg. pneumophila and 1·05 for Legionella spp. Results for individual systems depended on the nature of the system and its treatment. In cooling towers, Legionella spp. GU l(-1) always exceeded CFU l(-1) , and usually Legionella spp. were detected by qPCR when absent by culture. The pattern of results by qPCR for Leg. pneumophila followed the culture trend. In hot and cold water, culture and qPCR gave similar results, particularly for Leg. pneumophila. There were some marked exceptions with temperatures ≥50°C, or in the presence of supplementary biocides. Action and alert levels for qPCR were derived that gave results comparable to the application of the European Guidelines based on culture. Algorithms are proposed for the use of qPCR for routine monitoring. CONCLUSIONS: Action and alert levels for qPCR can be adjusted to ensure public health is protected with the benefit that remedial actions can be validated earlier with only a small increase in the frequency of action being required. SIGNIFICANCE AND IMPACT OF THE STUDY: This study confirms it is possible to derive guidelines on the use of qPCR for monitoring the control of legionellae with consequent improvement to response and public health protection.

Linkage scan of nicotine dependence in the University of California, San Francisco (UCSF) Family Alcoholism Study.

BACKGROUND: Nicotine dependence has been shown to represent a heritable condition, and several research groups have performed linkage analysis to identify genomic regions influencing this disorder though only a limited number of the findings have been replicated. METHOD: In the present study, a genome-wide linkage scan for nicotine dependence was conducted in a community sample of 950 probands and 1204 relatives recruited through the University of California, San Francisco (UCSF) Family Alcoholism Study. A modified version of the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA) with additional questions that probe nicotine use was used to derive DSM-IV nicotine dependence diagnoses. RESULTS: A locus on chromosome 2q31.1 at 184 centiMorgans nearest to marker D2S2188 yielded a logarithm (base 10) of odds (LOD) score of 3.54 (point-wise empirical p=0.000012). Additional peaks of interest were identified on chromosomes 2q13, 4p15.33-31, 11q25 and 12p11.23-21. Follow-up analyses were conducted examining the contributions of individual nicotine dependence symptoms to the chromosome 2q31.1 linkage peak as well as examining the relationship of this chromosomal region to alcohol dependence. CONCLUSIONS: The present report suggests that chromosome 2q31.1 confers risk to the development of nicotine dependence and that this region influences a broad range of nicotine dependence symptoms rather than a specific facet of the disorder. Further, the results show that this region is not linked to alcohol dependence in this population, and thus may influence nicotine dependence specifically.

Distribution of Legionella pneumophila serogroups, monoclonal antibody subgroups and DNA sequence types in recent clinical and environmental isolates from England and Wales (2000-2008).

Clinical isolates of Legionella pneumophila, obtained from 167 patients, who acquired their illness in the community in England and Wales between January 2000 and March 2008, were compared with 276 environmental isolates of L. pneumophila obtained over the same period as part of the routine sampling of 'managed' water systems. The 443 isolates were typed by monoclonal antibody (mAb) subgrouping and the internationally standardised, seven-gene loci, sequence-based typing (SBT) scheme of the European Working Group for Legionella Infections (EWGLI). Of the clinical isolates, 97.6% were L. pneumophila serogroup (sgp) 1, compared with only 55.8% of environmental isolates (P = 0.0002); 91.6% were subgrouped as mAb3/1+ve, compared with only 8.3% of environmental isolates (P < 0.0001). The isolates were very diverse, with SBT identifying 111 sequence types (STs) (index of diversity [IOD] 0.954). Among the clinical isolates, 42 ST were seen, with one (ST47) accounting for 25.7% and three (ST47, ST37 and ST62) accounting for 46.1% of all isolates. Eighty-two STs were identified among the environmental isolates, with two (ST1 and ST79) accounting for 34.1% of these. Comparison of the STs seen among clinical and environmental isolates showed that there was very little overlap between the two populations (P < 0.0001), with common clinical strains found in the environment very infrequently: 0.4, 0.7 and 0% (ST47, ST37 and ST62, respectively), and common environmental strains rarely causing disease: 4.8 and 1.2% (ST1 and ST79, respectively). Combining phenotypic and genotypic data identified 144 phenons (IOD 0.970); 52 among clinical isolates and 101 among environmental isolates. The most abundant clinical strain, mAb 'Allentown' ST47, accounted for 22.8% of cases, but was only found once in the environment. Conversely, mAb 'Oxford/OLDA' ST1 was the most common environmental strain (17.0%), but only caused two infections. A review of the published data shows that mAb 'Allentown' ST47 is also an important cause of infection in France and possibly in the Netherlands. However, it was not found in a large study of German clinical isolates. This study confirms previous work showing that just a few strains of L. pneumophila cause the majority of community-acquired Legionella infection in England and Wales, and that these clinically significant strains are only rarely found in managed water systems. These data suggest that knowing which particular strain is present in an environment might be at least as important as knowing the quantity in which legionellae are present.

Weather patterns and Legionnaires' disease: a meteorological study.

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 chi2=30.59, 3 d.f., P<0.0001). These findings have implications for future case numbers and resource requirements.

Public health and economic costs of investigating a suspected outbreak of Legionnaires' disease.

This paper provides one of the first assessments of the burden of both the public health investigation and the economic costs associated with an apparent outbreak of Legionnaires' disease (LD) in South East London. In addition to epidemiological, microbiological and environmental investigations, we collected data on the staff time and resources committed by the 11 main organizations responsible for managing the outbreak. Of the overall estimated costs of 455,856 pounds, only 14% (64,264 pounds) was spent on investigation and control of the outbreak compared with 86% (391,592 pounds) spent on the hospital treatment of the patients. The time and money spent on public health services in this investigation appear to represent good value for money considering the potential costs of a major outbreak, including the high case-fatality rate in LD generally and the high health-care costs. Further research is needed to determine optimum strategies for the cost-effective use of health system resources in investigations of LD. Whether the threshold for investigation of cases should be based on observed incidence rates or the cost-effectiveness of investigations, or both, should be debated further.

Legionnaires' disease outbreak associated with a cruise liner, August 2003: epidemiological and microbiological findings.

Eight cases of Legionnaires' disease were identified among the 215 German passengers after a cruise to the Nordic Sea in August 2003. An unmatched case-control study was conducted to identify risk factors and the source of infection. In total, eight passengers fulfilled the case definition, one of these died. Forty-two passengers served as controls. The attack rate was 4%. The mean age was 60 years for cases and 62 years for controls. Prolonged exposure to the spa pool seemed to be a risk factor of infection (OR 4.85, P=0.09). Legionella pneumophila serogroup 1, monoclonal antibody (mAb) subgroup 'Knoxville' was isolated from clinical and environmental samples. DNA sequence-based typing revealed that these isolates were indistinguishable from each other. The investigation showed the importance of an interdisciplinary approach of microbiology and epidemiology as not all sites on the ship that tested positive for L. pneumophila actually posed a relevant risk for the passengers.

Culture of Helicobacter pylori from domestic water samples--the impact of strain variation on growth on solid and in liquid media.

Helicobacter pylori is an important global human pathogen and there is growing evidence from PCR assays that contaminated drinking water might be a possible source of infection in some circumstances. There are no validated protocols for direct isolation but various culture media have been developed for possible environmental sampling. Our aim here was to investigate how inter-strain variation might affect the interpretation of results with such media. Two laboratory adapted reference strains and four recent clinical isolates were tested on four solid media and in ten liquid media. Considerable variation was found between strains in their ability to recover on the different media after stress exposure (suspension in sterile tap water). Generally, clinical isolates were less robust than the laboratory-adapted strains and, overall, the former required longer recovery times. Our findings highlighted the importance of using a range of isolates for evaluations, as examination of laboratory-adapted strains alone did not provide an accurate representation of the utility of media that may be used to recover H. pylori from water.

Legionnaires' disease: when an 'outbreak' is not an outbreak.

During August 2006, there was a large increase in non-travel related legionella cases throughout England and in the Netherlands.

Detection of Helicobacter pylori by PCR but not culture in water and biofilm samples from drinking water distribution systems in England.

AIMS: To investigate treated water distribution systems in England as a source of Helicobacter pylori. METHODS AND RESULTS: Water and biofilms were obtained from 11 domestic and seven educational properties and from hydrants, reservoirs and water meters supplied by three water utilities. Samples were cultured on nonselective and antibiotic containing media combined with immunomagnetic separation concentration. Viable helicobacters were not detected in any of the 151 samples but Helicobacter-specific PCR assays detected DNA in 26% of samples from domestic properties, schools and hydrants with the highest frequency in biofilms (42%). Direct sequencing of six selected amplicons confirmed >95% sequence homology to H. pylori. CONCLUSIONS: While viable helicobacters were not isolated, evidence was obtained for the presence of Helicobacter DNA, including that of H. pylori. Biofilms on surfaces within water distribution systems may act either as sites for the passive accumulation of helicobacters or as potentially important reservoirs of infection. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings strengthen evidence that H. pylori may be transmitted through drinking water. However, there is currently no evidence that viable cells can survive the disinfection levels used in UK mains supplies and the health risk from this source remains unclear.

Comparison of agar-based methods for the isolation and enumeration of heterotrophic bacteria with the new multidose IDEXX SimPlate method.

Pour and spread plates are the conventional methods of choice for the isolation and enumeration of heterotrophic microorganisms in treated water supplies. The tests are performed at 22 degrees C and 37 degrees C for 72 h and 48 h respectively. Counts at 22 degrees C are associated with pollution of water systems from external sources, while counts at 37 degrees C are used as an indication of treatment plant performance and the deterioration of the general quality of water. Conventional methods using Yeast Extract Agar for a pour plate and R2A agar for a spread plate were compared with the multidose IDEXX SimPlate method for the isolation and enumeration of heterotrophic bacteria in water. SimPlate gave a significantly higher count on average than the conventional methods. The R2A method showed the next highest count, being significantly higher than Yeast Extract Agar. In addition, unlike the pour and spread plate methods, SimPlate was easier to use, reduced labour, and the test results were far easier to read.

A comparison of the International Standards Organisation reference method for the detection of coliforms and Escherichia coli in water with a defined substrate procedure.

AIMS: This study investigated the use of the International Standards Organisation (ISO) procedure for the comparison of microbiological methods. Using this procedure the ISO reference procedure for the detection of coliforms and Escherichia coli in water was compared with a defined substrate method (ColilertTM). METHODS AND RESULTS: A total of 20 laboratories from 13 European countries compared the use of Colilert/Quanti-TrayTM, a quantitative defined substrate test (DST) for the presence of coliforms and E. coli with the ISO reference procedure, which utilizes tergitol-TTC medium. Results of the study showed that DST detected significantly more coliforms and E. coli than did the reference procedure. In the case of E. coli the recoveries were also higher when using DST and the difference seen was statistically significant. The confirmation rate obtained when using the DST product suggested that no confirmation of wells positive for E. coli was necessary during routine use. CONCLUSIONS: Colilert is a suitable alternative to the ISO reference procedure for the detection of coliforms and E. coli in water. The methods used during the comparison study indicated that confirmation of all colonies/positive wells led to the most accurate information and it is recommended that for future comparison studies this should become standard practice. Confirmation of a small proportion of colonies led to misleading conclusions and should be avoided when comparing microbiological methods. SIGNIFICANCE AND IMPACT OF STUDY: It has been demonstrated that the ISO reference procedure fails to detect a significant proportion of coliforms and E. coli in drinking water. Colilert/QuantiTrayTM is a more suitable alternative.

Community-acquired Legionnaires' disease in Nottingham--too many cases?

The aim of this study was to compare the incidence of community-acquired Legionnaires' Disease in Nottingham with England and Wales and to explore reasons for any difference observed. Based on data from the National Surveillance Scheme for Legionnaires' Disease (1980-1999), the rate of infection in England and Wales was 1.3 per million/year compared with 6.6 per million/ year in Nottingham. Domestic water samples were obtained from 41 (95%) of 43 Nottingham cases between 1997 and 2000. In 16 (39%) cases, Legionella sp. were cultured in significant quantities. Proximity to a cooling tower was examined using a 1:4 case-controlled analysis. No significant difference in the mean distance between place of residence to the nearest cooling tower was noted (cases 2.7 km vs. controls 2.3 km; P = 0.5). These data suggest that Nottingham does have a higher rate of legionella infection compared to national figures and that home water systems are a source.

Guidelines for investigating single cases of Legionnaires' disease.

These guidelines for the investigation of single cases of legionnaires' disease have been updated from those produced in 1994 to take account of developments in microbiological and environmental diagnostic capabilities and the recognition that infection may be acquired from the patient's domestic water system. The new guidelines recommend that when a case of legionnaires' disease has been diagnosed, it should be investigated to determine whether it is part of an outbreak or cluster, work related, suspected to be a hospital acquired infection or is travel associated. If information concerning the patient's movements during the incubation period shows it to be none of these, then appropriate environmental investigations should be considered which might include the patient's domestic water system as a potential source of infection. The guidelines are designed to provide advice and information to all public health personnel involved in the control and prevention of legionnaires' disease.

Detection of Legionella pneumophila using a real-time PCR hybridization assay.

A real-time PCR hybridization assay for Legionella pneumophila is described; the assay uses LightCycler (Idaho Technology) methodology to specifically detect 2.5 CFU/reaction, equivalent to 1,000 CFU/liter of starting water sample. The assay, including DNA extraction and confirmation of product identity, is completed within 90 min of receipt of a sample.

Mixed genogroup SRSV infections among a party of canoeists exposed to contaminated recreational water.

Samples of faeces collected from a party of canoeists involved in a gastroenteritis outbreak were examined by electron microscopy and RT-PCR for evidence of infection with SRSVs. A broadly reactive primer pair was used to detect SRSVs followed by application of genogroup-specific primers to SRSV-positive specimens. Exposure data were collected by means of a questionnaire. SRSVs were detected in 1/4 specimens examined by EM and 3/4 by RT-PCR. Genogrouping, and sequencing of PCR products revealed two distinct strains: a genogroup I strain, related to the Desert Shield virus, and a genogroup II strain, related to the Lordsdale virus to be associated with the outbreak. Exposure data indicated that capsising and eating food before getting changed were associated with an increased risk of gastroenteritis and was consistent with infection following the consumption of contaminated water. This study confirms the greater sensitivity of RT-PCR for the diagnosis of SRSV infections and its utility, when incorporating genogroup-specific primers, in establishing more complex epidemiological data.