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The epidermal growth factor receptor (EGFR) is commonly upregulated in multiple cancer types, including breast cancer. In the present study, evidence is provided in support of the premise that upregulation of the EGFR/MEK1/MAPK1/2 signaling axis during antiestrogen treatment facilitates the escape of breast cancer cells from BimEL‑dependent apoptosis, conferring resistance to therapy. This conclusion is based on the findings that ectopic BimEL cDNA overexpression and confocal imaging studies confirm the pro‑apoptotic role of BimEL in ERα expressing breast cancer cells and that upregulated EGFR/MEK1/MAPK1/2 signaling blocks BimEL pro‑apoptotic action in an antiestrogen‑resistant breast cancer cell model. In addition, the present study identified a pro‑survival role for autophagy in antiestrogen resistance while EGFR inhibitor studies demonstrated that a significant percentage of antiestrogen‑resistant breast cancer cells survive EGFR targeting by pro‑survival autophagy. These pre‑clinical studies establish the possibility that targeting both the MEK1/MAPK1/2 signaling axis and pro‑survival autophagy may be required to eradicate breast cancer cell survival and prevent the development of antiestrogen resistance following hormone treatments. The present study uniquely identified EGFR upregulation as one of the mechanisms breast cancer cells utilize to evade the cytotoxic effects of antiestrogens mediated through BimEL‑dependent apoptosis.

EGFR is upregulated commonly in multiple cancer types, including breast cancer. However, the multiple role(s) of EGFR in breast cancer are not fully elucidated. In this study, we characterized a unique antiestrogen resistant breast cancer subline and provide compelling data that upregulation of the EGFR/MEK1/MAPK1/2 signaling axis during antiestrogen treatment facilitated escape from senescence, autophagy, and BimEL-dependent apoptosis. Our studies were conducted with the antiestrogen resistant TR5 cell line established by our laboratory, using a step-wise drug selection with the antiestrogen 4-hydroxytamoxifen (4-OHT) of the estrogen receptor α (ER+) MCF-7 breast cancer cells. A preliminary microarray comparing mRNA expression between 4-OHT-selected TR5 cells and MCF-7 parent cells identified upregulation of the epidermal growth factor receptor (EGFR). Western blot showed a greater than 5-fold upregulation of EGFR expression and downregulation of estrogen receptor alpha (ERα) compared to the levels in MCF-7 and T47-D (antiestrogen sensitive). Selective inhibition of EGFR, phosphorylated at Tyr 1068, was achieved with erlotinib. Inhibition blocked MEK1/MAPK1/2 signaling, which was also upregulated in 4-OHT selected TR5 cells, with detectable increases in the pro-apoptotic BH3 protein BimEL, and a fifty percent reduction in cell viability. However, a high number of cells survived erlotinib therapy due to autophagy induction determined to be cytoprotective. Autophagy induction was identified by analyzing the expression/turnover of autophagy proteins LC3-II and p62, a standard approach to ascertain autophagy levels in cells. Further, erlotinib treatment induced a 2-fold increase in the number of senescent cells in the surviving population. Senescent cells were identified by β-galactosidase staining of erlotinib-treated cells utilizing confocal microscopy. Interestingly, blockade of autophagy with the lysosomotrophic compound chloroquine enhanced the senescence cell population, suggesting that either autophagy blocked senescence induction or lysosome impairment increased senescence. Mechanistically, we have identified phosphorylated AMP-activated protein kinase (pAMPK) as being elevated under conditions of erlotinib treatment, and reduced in 4-OHT-selected TR5 cells relative to MCF-7 parent cells. This correlation supports the hypothesis that pAMPK was downregulated during 4-OHT selection of TR5 cells by elevated EGFR/MEK/MAPK1/2 signaling to allow surviving cells escape senescence induction and/or anti-proliferative autophagy. Overall we propose that EGFR mediated downregulation of pAMPK is a major regulatory pathway in breast cancer cells surviving endocrine treatment. Our current studies are dissecting the role of EGFR-mediated AMPK regulation with somatic cell-based studies and pharmacological intervention. Citation Format: Mackenzie L. Hagan, Michael McGrath, Carol Joseph, Allison Lewis, John R. Barrett, Muthusamy Thangaraju, David Gewirtz, Patricia V. Schoenlein. EGFR upregulation in breast cancer cells: A key mechanism of escape from antiestrogen induced death, autophagy, and senescence [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB133.

Our long-term goal is to elucidate the role of JNK/Jun signaling in breast cancer cell autophagy induced by anti-estrogen treatment and/or chemotherapy in ERα expressing and TNBC cells, respectively. In this study, we analyzed the role of JNK/Jun signaling in regulating the high basal autophagy levels in TNBC. To do so, we utilized the highly metastatic murine 4T1 and the human MDA-MB-231 TNBC models. In addition, we performed autophagic flux assays after treating breast cancer cells with JNK-IN8, a non-reversible small molecule JNK1/2 inhibitor. Treatments were performed for various times in the presence and absence of chloroquine (CQ) to allow analysis of steady state levels of atg8 (LC3-II) and sequestosome-1 (p62). These LC3-II and p62 proteins are involved in autophagosome formation and function, and are catabolized during autolysosomal turnover. To impair this process, CQ is a lysosomotropic agent that raises the pH of the lysosome and blocks such turnover. We also determined the effect of JNK-IN-8 on 4T1 and MDA-MB-231 cell number (proliferation). These studies determined that JNK-IN8 was an effective inhibitor of autophagic flux in the TNBC cells when used at concentrations that effectively blocked cJun phosphorylation, the downstream readout of JNK-IN8 efficacy used for our experiments. Blockade of autophagic flux by JNK-1/2 inhibition in TNBC was in stark contrast to results obtained with JNK-IN-8 treatment of ERα expressing MCF-7 and T47-D breast cancer cells, in which JNK-IN-8 induced a robust autophagy response with elevations in the steady state levels and flux of LC3-II and p62. Induction of autophagy by JNK-IN-8 in ERα expressing breast cancer cells occurred concomitantly with a substantial reduction in cell proliferation but minimal induction of apoptosis. Overall, these pre-clinical in vitro studies support the following conclusions: (1) JNK1/2 targeting in TNBC effectively blocks autophagic flux and may be particularly effective when used in combination with chemotherapies that induce pro-survival autophagy which is a current focus of our studies; and (2) JNK1/2 targeting in ERα expressing breast cancers has the potential to mediate a pro-survival autophagic activity in response to endocrine therapy or chemotherapy and should be cautiously approached. Ongoing studies are aimed at understanding the underlying mechanisms of the differential effects of JNK/Jun signaling on autophagy in TNBC versus ERα expressing breast cancer. Such an understanding should provide important information to aid in the ongoing interest in utilizing JNK1/2 as a molecular target for improved treatment of TNBC. Citation Format: Michael K. McGrath, McKenzie L. Hagan, Timmothy Summers, Jesse Wayson, Carol Joseph, Amanda C. Barrett, John T. Barrett, Patricia V. Schoenlein. Effects of JNK/Jun blockade on breast cancer cell autophagy are dependent on breast cancer subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3006.

The c-Jun N-terminal kinases, JNK1/2 are members of the mitogen activated protein kinase (MAPK) family that can mediate inflammatory responses, cell differentiation, cell proliferation, and/or cell death. In breast cancer, JNK signaling has been associated with increased proliferation, angiogenesis, and tumor metastasis. Also, inhibition of JNK has been shown to promote tumor cytostasis, but this inhibition also attenuates the cytotoxic effects of certain chemotherapeutics, calling into question the therapeutic potential of JNK inhibitors. In this study, we hypothesized that JNK1/2 inhibition induced autophagy, as a mechanism of protection against chemotherapy- and endocrine-induced apoptosis. Autophagy is a normal physiologic process required for the elimination of damaged mitochondria and misfolded proteins that can be induced in cancer cells under certain stresses such as hypoxia and drug insult to provide protection from death. To characterize the role of JNK1/2 in autophagy, we utilized two ERα expressing human breast cancer cell lines, T47-D and MCF-7. Autophagy flux experiments were performed in which the steady state levels of the autophagy protein LC3 II were analyzed in control cells versus cells undergoing JNK1/2 inhibition with the non-reversible JNK-IN8 inhibitor. JNK-IN8 was used as a single agent or in combination with estradiol and/or antiestrogen treatment. These studies reproducibly identified JNK1/2 inhibition as a mechanism of autophagy activation. The induction of autophagy correlated with sustained inhibition of proliferation as determined by cell counts, MTT, and clonogenic assays, and induction of senescence (in T47-D cells) as determined by the expression of β-galactosidase. In contrast, induction of apoptosis was not a major outcome of JNK1/2 inhibition. These pre-clinical studies provide strong evidence that JNK1/2 is an attractive molecular target to improve the treatment of endocrine responsive breast cancer, but they also emphasize that cells are surviving by autophagy and/or senescence. Because cell survival by autophagy and/or senescence could ultimately lead to breast cancer relapse, ongoing studies are addressing approaches to effectively kill these surviving cell populations. Citation Format: Kyler Herrington, Carol Joseph, Samar Abdulmoneim, Allison Lewis, Jingwen Cai, Hannah Youngblood, Yutao Liu, Patricia Schoenlein. Targeting JNK1/2 induces a potent cytostatic response in estrogen receptor expressing breast cancer cells, with induction of autophagy and senescence as measurable outcomes [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1231.

Once a year, countries that collaborate in the European Surveillance Scheme for Travel Associated Legionnaires’ Disease (EWGLINET) are requested to submit a dataset that provides epidemiological and microbiological information on cases of legionnaires’ disease (nosocomial (hospital-acquired), community and travel related) detected in their country for that year. This paper presents the data collected for 2003 and 2004. For this period, 9166 cases were reported to the dataset by 35 countries, of which 941 cases were associated with outbreaks. Fourteen countries reported a total of 218 detected outbreaks. National infection rates varied between countries from 28.7 to less than one case per million population. This information is valuable in that it allows countries to assess the effectiveness of their national surveillance schemes in detecting cases. Over the two year period, 748 cases were reported to have died, giving a case fatality rate of 8.2%. The lack of detailed epidemiological information on deaths from legionnaires’ disease is highlighted. The establishment of the European Centre for Disease Prevention and Control is seen as an opportunity to develop European collaborations more fully, and to increase further the protection of Europeans from outbreaks of legionnaires’ disease.

Influenza-associated bacterial and viral infections are responsible for high levels of morbidity and death during pandemic and seasonal influenza episodes. A review was undertaken to assess and evaluate the incidence, epidemiology, aetiology, clinical importance and impact of bacterial and viral co-infection and secondary infection associated with influenza. A review was carried out of published articles covering bacterial and viral infections associated with pandemic and seasonal influenza between 1918 and 2009 (and published through December 2011) to include both pulmonary and extra-pulmonary infections. While pneumococcal infection remains the predominant cause of bacterial pneumonia, the review highlights the importance of other co- and secondary bacterial and viral infections associated with influenza, and the emergence of newly identified dual infections associated with the 2009 H1N1 pandemic strain. Severe influenza-associated pneumonia is often bacterial and will necessitate antibiotic treatment. In addition to the well-known bacterial causes, less common bacteria such as Legionella pneumophila may also be associated with influenza when new influenza strains emerge. This review should provide clinicians with an overview of the range of bacterial and viral co- or secondary infections that could present with influenza illness.

Antiestrogen resistance is still a major clinical challenge preventing the eradication of estrogen receptor positive (ER+) breast cancer. A major goal of our laboratory is to identify targeted therapies that can be combined with antiestrogen treatment to block the emergence of antiestrogen resistant breast cancer. Toward this goal, we previously determined that the catabolic process of autophagy facilitates the emergence of antiestrogen resistance (Sammadar et al., Molecular Cancer Ther. 9, 2008) and identified cathepsin L as a key lysosomal protease required for antiestrogen induced pro-survival autophagy ER+ MCF-7cells (Periyasamy-Thandavan et al., Autophagy. 6:19-35, 2010). Thus, we hypothesize that blocking cathepsin L expression / activity is one approach to targeting pro-survival autophagy during antiestrogen treatment of breast cancer. In support of this hypothesis, we now show that antiestrogen resistant TR5 cells show increased levels of active cathepsin L compared to the levels in the parent antiestrogen sensitive MCF-7 cells. The upregulation of cathepsin L expression is consistent with the fact that TR5 cells utilized autophagy to survive long-term 4-hydroxytamoxifen (4-OHT) selection. Withdrawal of 4-OHT selection during TR5 passage does not lead to a reduction in cathepsin L levels, suggesting that the increased expression of cathepsin L is a stable genetic or epigenetic alteration. Elevated expression of active (phosphorylated) MAPK1/2 is also present in TR5 cells and targeting MEK1/MAPK1/2 with the selective inhibitor U0126 consistently reduced the levels of active cathepsin L. We further established a key role for MEK1/MAPK1/2 in the regulation of cathepsin L in the parent MCF-7 cells by performing cathepsin L activity assays. Cells treated with 4-OHT showed increased cathepsin L activity compared to E2-treated control cells; whereas, cathepsin L activity in cells treated with 4-OHT + UO126 was significantly reduced compared to the levels in 4-OHT-treated cells. This reduction in cathepsin L by U0126-mediated blockade of MEK1/MAPK1/2 correlated directly to “impaired” autophagic flux and increased BimEL-dependent apoptosis in the antiestrogen-treated cell populations and was mediated, at least in part, via transcriptional regulation of cathepsin L as determined by quantitative PCR. Further, MEK1/MAPK1/2 up-regulation of cathepsin L is selective, as cathepsin B activity is not reduced by MEK1 targeting. These studies provide evidence that the targeting MEK1/MAPK1/2 in combination with antiestrogen treatment has the potential to reduce cathepsin-L mediated pro-survival autophagy in ER+ breast cancer. Citation Format: Annie Liu, Carol Joseph, Jesse Wayson, Timothy Summers, Haifeng Cai, Patricia V. Schoenlein. MEK1:MAPK1/2 targeting attenuates pro-survival autophagy and enhances antiestrogen-induced apoptosis in breast cancer cells via a cathepsin L-dependent mechanism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1337.

A source of infection is rarely identified for sporadic cases of Legionnaires' disease. We found that professional drivers are five times more commonly represented among community acquired sporadic cases in England and Wales than expected. We therefore investigated possible risk exposures in relation to driving or spending time in a motor vehicle. A case control study including all surviving community acquired sporadic cases in England and Wales with onset between 12 July 2008 and 9 March 2009 was carried out. Cases were contacted by phone and controls were consecutively recruited by sequential digital dialling matched by area code, sex and age group. Those who consented were sent a questionnaire asking questions on driving habits, potential sources in vehicles and known risk factors. The results were analysed using logistic regression. 75 cases and 67 controls were included in the study. Multivariable analysis identified two exposures linked to vehicle use associated with an increased risk of Legionnaires' disease: Driving through industrial areas (OR 7.2, 95%CI 1.5-33.7) and driving or being a passenger in a vehicle with windscreen wiper fluid not containing added screenwash (OR 47.2, 95%CI 3.7-603.6). Not adding screenwash to windscreen wiper fluid is a previously unidentified risk factor and appears to be strongly associated with community acquired sporadic cases of Legionnaires' disease. We estimated that around 20% of community acquired sporadic cases could be attributed to this exposure. A simple recommendation to use screenwash may mitigate transmission of Legionella bacteria to drivers and passengers.

Background Influenza incidence thresholds are used to help predict the likely impact of influenza and inform health professionals and the public of current activity. We evaluate the potential of syndromic data (calls to a UK health helpline NHS Direct) to provide early warning of national influenza outbreaks. Methods Time series of NHS Direct calls concerning ‘cold/flu’ and fever syndromes for England and Wales were compared against influenzalike-illness clinical incidence data and laboratory reports of influenza. Poisson regression models were used to derive NHS Direct thresholds. The early warning potential of thresholds was evaluated retrospectively for 2002‐06 and prospectively for winter 2006‐07. Results NHS Direct ‘cold/flu’ and fever calls generally rose and peaked at the same time as clinical and laboratory influenza data. We derived a national ‘cold/flu’ threshold of 1.2% of total calls and a fever (5‐14 years) threshold of 9%. An initial lower fever threshold of 7.7% was discarded as it produced false alarms. Thresholds provided 2 weeks advanced warning of seasonal influenza activity during three of the four winters studied retrospectively, and 6 days advance warning during prospective evaluation. Conclusion Syndromic thresholds based on NHS Direct data provide advance warning of influenza circulating in the community. We recommend that age-group specific thresholds be developed for other clinical influenza surveillance systems in the UK and elsewhere.

SUMMARYThis 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

The aim of this study was to investigate influenza immunisation rates in the United Kingdom over a 6-year period and examine trends in uptake by deprivation, ethnicity, rurality and risk group. Influenza immunisation rates were determined from 1999/2000 to 2004/2005 using a large general practice database (QRESEARCH). There was a relative increase of 59.5% in the overall influenza vaccination rate over the study period. In 2004/2005, 70.2% of all patients aged 65 and over were vaccinated, compared with 29.3% of patients in a clinical risk group aged less than 65. Males, patients from deprived areas and from areas with a higher proportion of non-White residents had slightly lower vaccination rates overall. This general practice based study suggests that substantial increases in influenza vaccination rates have occurred across all risk groups, but that increased focus should be given to immunising high-risk patients below the age of 65.

A small national study was carried out in England in 2003/2004 to ascertain the views of primary care trusts (PCTs) and general practitioners (GPs) on whether influenza immunisation should be extended to all people aged 50-64 years from the current recommendation of 65 years or more. Results showed that as many primary care trusts would be in favour, as would not be in favour. A similarly divided view was expressed by general practitioners. Vaccine uptake rates for high-risk (HR) and low-risk (LR) adults aged 50-64 years in the study population were higher in those practices where the GP was in favour of a more inclusive policy of offering flu vaccine to all persons aged 50 years or more, compared with those that did not favour this policy (60% versus 54% HR (p=0.02) and 16% versus 11% LR (p=0.02)). Higher rates of vaccine uptake for low-risk patients aged 50-64 years were also reported from practices where GPs perceived a greater health benefit of immunisation for this age group. Although policy for recommending vaccine to all patients aged 50 years or more is established elsewhere, opinion on whether such a policy should be adopted in England is currently divided amongst those providing local health services.

Public health officials will normally take action at accommodation sites following an association with a cluster of cases of Legionnaires' disease. This paper seeks to determine the likelihood of such a cluster occurring at a site once it has been associated with a single case of the disease, and therefore whether more should be done at sites following individual cases.Information for UK residents reported to the EWGLINET system between 1993 and 2000 was included in a dataset. The size and country of hotel visited by the cases were divided into six country groups (France, Italy, Spain, Turkey, other Europe and other World), and eight size groups (20 rooms, 20-49, 50-99, 100-199, 200-299, 300-399, 400-499, 500+). The data were investigated using Cox proportional hazards regression to model the probability of at least one further case following the first.The dataset comprised 793 cases that had stayed at 605 sites in 51 countries between 1993 and the end of 2000. This included 605 cases that were the first case associated with a site, and 188 subsequent cases. Following the first case, 16.6% of sites were associated with at least one subsequent case during the period under study. The probability of a subsequent case occurring within 6 months of the first varied by country and size group, with some combinations returning a probability30%; the probability of a subsequent case occurring within 2 years of the first reached over 50% in some instances.There may be support for early intervention at some accommodation sites following a first case of Legionnaires' disease, in specific country and size groups.

SUMMARY Each year, countries that participate in the European Surveillance Scheme for Travel Associated Legionnaires’ Disease (EWGLINET) are requested to complete a set of standardized reporting forms that provide epidemiological and microbiological information on the total number of cases (non-travel as well as travel-related cases) detected in their country. Trends at the national and aggregated European level have been analysed for 2000–2002. For this period, 10 322 cases of Legionnaires’ disease were reported and national infection rates ranged from 0 to 34 . 1 cases per million population. A total of 189 outbreaks were associated with nosocomial infection, community exposure or travel. The upward trend in diagnosis through the urinary antigen detection test has resulted in a higher ascertainment of cases in many countries. However, the decline in diagnosis by culture of the organism is likely to severely hamper outbreak investigations in the future if fewer clinical isolates are available for matching with environmental isolates. This important data-set has been used for studying the effectiveness of surveillance and legionella control and prevention programmes within Europe.

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[ratio ]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.

Antiestrogen resistance is the major impediment to the eradication of estrogen receptor positive (ER+) breast cancer. Approximately 30% of ER+ breast tumors initially responsive to antiestrogen therapy will acquire resistance. One approach to reducing the occurrence of acquired antiestrogen resistance is to identify and target key signaling nodes that specifically attenuate the ability of antiestrogens to kill cancer cells. Toward this goal, we identified MEK/MAPK1/2 as a key molecular target based on the general inability of antiestrogens to effectively block MEK1/MAPK1/2-mediated phosphorylation of BimEL in breast cancer cells. BimEL is a pro-apoptotic member of the BH3 family of proteins that is inactivated (degraded by the proteasome) when phosphorylated by MAPK1/2. The combination of an antiestrogen and MEK1 inhibitor robustly-induced BimEL-dependent breast cancer cell apoptosis (Periyasamy-Thandavan et al., 2012). However, a subpopulation of breast cancer cells survive this combined treatment via an autophagy-dependent mechanism. Based on the key role of JNK in regulating autophagy in cancer cells via BimEL phosphorylation, we hypothesized that JNK was a key effector of autophagy in breast cancer cells surviving antiestrogen treatment when used as a single agent or in combination with a MEK inhibitor. To test this hypothesis, we utilized the selective JNK inhibitor SP600125 as a single agent or in combination with estradiol (E2), 4-hydroxytamoxifen (4-OHT), and/or U0126 (a selective MEK1 inhibitor). Treatments were conducted with the antiestrogen-sensitive cell lines MCF-7 and T-47D and the antiestrogen resistant cell line TR5, previously derived in our laboratory by a step-wise 4-OHT selection. Effects on cell death (determination of apoptosis) and autophagy (determination of autophagy levels and flux) were evaluated for the various hormonal treatments conducted in the presence or absence of JNK inhibition. These studies showed that: (1) targeting JNK in antiestrogen sensitive breast cancer cells induces apoptosis with caspase-dependent cleavage of PARP as a surrogate marker of apoptosis; (2) JNK activity is elevated in antiestrogen resistant TR5 cells and JNK inhibition induces TR5 cell death; and (3) antiestrogen sensitive and resistant breast cancer cells dying as a result of JNK inhibition can show extensive cytosolic vacuolization with the autophagy protein LC3 (ATG8) localized to the aberrant vacuoles. Overall, our results support the conclusion that JNK plays a key autophagy-dependent survival role in breast cancer cells. Ongoing studies aim to identify the specific JNK protein(s) that disrupt autophagy and enhance death of breast cancer cells toward the goal of identifying specific molecular targets to block autophagy in breast cancer cells. Citation Format: Rohit Munagala, Carol Joseph, Annie Liu, Kebin Liu, Muthusamy Thangaraju, Patricia V. Schoenlein. A critical role for c-Jun N-terminal kinase in autophagy and cell survival of breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3318. doi:10.1158/1538-7445.AM2017-3318

Identification of the organism responsible for Legionnaires' disease was first associated with an outbreak in 1976 in Philadelphia, Pa., where 189 cases and 29 deaths occurred among American Legion veterans who had attended a hotel-based convention. European Working Group for Legionella Infections (EWGLI) provides expert epidemiological, microbiological, and environmental health advice to clinical and public health officials, tour operators, national and international government departments, and other relevant groups such as hoteliers and members of the public. This chapter mainly presents data from 1999, which was provided by 28 of the 31 countries that currently participate in the European scheme. It also compares trends and incidence of cases in Europe from 1993. Since 1993, the overall rate of infection per million residents has ranged from 3.35 to 4.46. In 1999, 32% of the overall cases were reported as community acquired, 9% were hospital acquired, and 21% were associated with travel either in their own country or abroad. The objectives of the December 1999 meeting of experts convened by the European Commission to review the Belgium and The Netherlands whirlpool spa outbreaks, were to advise the Commission on the need for European recommendations and guidelines for the control and prevention of future whirlpool spa outbreaks and to produce specific recommendations that could be addressed by the Commission. EWGLI is assisting data contribution for effective surveillance at the international level through its sharing of information on cases, outbreaks, sources of infection, and continued developments in epidemiological, microbiological, and environmental aspects of legionella infection.

Ten cases of indigenously acquired Legionnaires' disease were identified by routine laboratory surveillance in Wales, United Kingdom, over a period of 7 months. Epidemiological investigations started with a case-control study. Case definitions used were those of the European Working Group for Legionella Infections; controls were taken from people who accompanied the three patients who had visited the hotel as part of a group. A cohort study was designed with staff who worked during the weekend that two patients had visited the hotel. The hypothesis was that more seroconversions would have occurred among people working close to the source, so that the average titer in this group would be higher than in people not exposed to the source. Six patients had a lung infiltrate on chest X ray. The first patient visited the hotel in July 1999, patients 2 and 3 visited in December 1999, patients 4 and 5 visited in January 2000, and patient 6 visited in February 2000. Only one stayed overnight and dined in the hotel, one used the swimming pool and spa for a week, three attended a lunch, and one attended an evening meal. Three of six patients were active smokers; none were immunosuppressed. Neither the case-control study nor the cohort study indicated a possible source of infection. Public Health officials and local authorities were notified of the possible role of ultrasonic humidifiers in the transmission of legionellae.

The European Surveillance Scheme for Travel Associated Legionnaires’ Disease (EWGLINET) has helped strengthen European national surveillance systems and improve case detection and reporting in travel-associated cases. The guidance and legislation introduced in England and Wales over the past 25 years aims to ensure that all relevant systems are properly installed and maintained and that investigation of these systems (e.g., wet cooling towers) in an outbreak situation is as efficient as possible. These measures are aimed at reducing the absolute number of cases of Legionnaires’ disease. England and Wales’ annual case reports severely underrepresent the true annual incidence of Legionnaires’ disease. The urinary antigen test was introduced in England and Wales in the 1990s and is now the primary diagnostic method for 80% of English and Welsh cases of Legionnaires’ disease. The test is quick and easy to perform and might therefore be enabling countries to detect milder cases of the disease that would otherwise have gone undiagnosed. However, in England and Wales the number of cases has remained relatively constant despite the increasing use of the test. The simple trend of an increasing number of cases of Legionnaires’ disease in England and Wales over the past 25 years masks a more complicated picture. Reporting systems that suffer from underdiagnosis and underreporting are likely to register an increase in case reports (as ascertainment improves) before any actual decrease from improved control and prevention in case numbers is detected.

The European surveillance scheme for travel-associated Legionnaires’ disease (EWGLINET) supports ongoing microbiological research and development for improved laboratory detection of clinical and environmental sources of infection using standardized methods within European countries. Adherence to the EWGLI guidelines, introduced in 2002, for control and prevention of travel-associated cases of Legionnaires’ disease is leading to greater protection of travelers in all European countries. This review covers EWGLINET’s role in the collection of national legionella data that enables trends over time at the aggregate level to be studied as well as historical trends within and between European countries. It covers the 10-year period 1995 to 2004 and mainly focuses on trends in aggregate data. The European data set from contributing countries between 1995 and 2004 is composed of 27,244 cases of Legionnaires’ disease. Community-acquired cases comprise the largest proportion of cases in Europe, at 38% overall, but vary from year to year depending on the number and size of reported outbreaks. This short review of surveillance data on cases of Legionnaires’ disease in Europe has mainly focused on descriptive aggregated data for the period 1995 to 2004. Major differences between countries in number of cases, category of exposure, number and range of outbreaks, and methods of diagnosis can only be highlighted rather than examined in depth. All countries with data in this review acknowledge the importance of international collaborations since these also benefit microbiological and epidemiological developments at the national level.

SUMMARYWe 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 95thvs. 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 75thvs.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.

The European Surveillance Scheme for Travel Associated Legionnaires’ Disease was set up by the European Working Group on Legionella Infections (EWGLI) in 1987 to identify cases of legionella infection in returning travellers and to detect outbreaks and cl

SUMMARYCalls to a UK national telephone health helpline (NHS Direct) have been used for syndromic surveillance, aiming to provide early warning of rises in community morbidity. We investigated whether self-sampling by NHS Direct callers could provide viable samples for influenza culture. We recruited 294 NHS Direct callers and sent them self-sampling kits. Callers were asked to take a swab from each nostril and post them to the laboratory. Forty-two per cent of the samples were returned, 16·2% were positive on PCR for influenza (16 influenza A(H3N2), three influenza A (H1N1), four influenza B) and eight for RSV (5·6%). The mean time between the NHS Direct call and laboratory analysis was 7·4 days. These samples provided amongst the earliest influenza reports of the season, detected multiple influenza strains, and augmented a national syndromic surveillance system. Self-sampling is a feasible method of enhancing community-based surveillance programmes for detection of influenza.

Five cases of legionnaires disease and one death were associated with four members of a tour group from the United Kingdom (UK) and one French tourist who all visited Spain in the spring of 1993. The UK group stayed at four hotels, one of which was also used by the French tourist. Phenotypic and genotypic comparison of isolates of Legionella pneumophila obtained from one of the UK cases and the French patient demonstrated that they were indistinguishable from each other and from environmental isolates obtained from the water supply of the hotel at which all five cases had stayed. A cohort study of the UK tour group was carried out to determine the extent of the outbreak and showed that three further members of the group had respiratory illness but were serologically negative to legionella infection. International participation in this investigation has highlighted the value of a European surveillance scheme and the benefit of microbiological collaboration between legionella reference laboratories in Europe.

Outbreaks of Legionnaires' disease create high levels of public anxiety and media interest and inevitably consume a great deal of public health resources. Investigations should begin as early as possible in order to rapidly identify suspected sources of infection, control the outbreak and prevent further cases occurring. The investigations should be coordinated by an outbreak control team who work collaboratively within local/national/international public health guidelines and with clear terms of reference. The actions carried out by epidemiologists when investigating community-, hospital-, or travel-associated outbreaks are comprehensively outlined in this chapter. The microbiological and environmental actions that complement this work are discussed in the accompanying chapters.

Outbreaks of Legionnaires' disease create high levels of public anxiety and media interest and inevitably consume a great deal of public health resources. Investigations should begin as early as possible in order to rapidly identify suspected sources of infection, control the outbreak and prevent further cases occurring. The investigations should be coordinated by an outbreak control team who work collaboratively within local/national/international public health guidelines and with clear terms of reference. The actions carried out by epidemiologists when investigating community-, hospital-, or travel-associated outbreaks are comprehensively outlined in this chapter. The microbiological and environmental actions that complement this work are discussed in the accompanying chapters.

SUMMARYTwo hundred and eighteen nosocomial cases of Legionnaires' disease with 68 deaths were reported to the National Surveillance Scheme for Legionnaires Disease between 1980 and 1992, representing 15% of the reported infections acquired in England and Wales. Twenty–two nosocomial outbreaks accounted for 135 (62%) of these cases, the remainder occurring as single cases either in hospitals where other single cases or outbreaks had been reported in different years or as ‘sporadic’ cases in hospitals from which no other cases were reported. A clinical history prior to onset of Legionnaires' disease was available for 124 patients, 61 of whom had undergone recent transplant therapy or were immunosuppressed for other reasons.Sixty cases (27%) were diagnosed by culture of the organism and isolates from 56 patients were typed; 25 (42%) were nonL. pneumophilaserogroup 1 infections.Methods for prevention and control of nosocomial outbreaks are discussed, in particular the susceptibility to Legionnaires' disease of certain groups of hospital patients.

Background The source of infection for most sporadic cases of Legionnaires9 disease remains unknown. This study aims quantify the relationship between cases and wet cooling systems (WCS), a potential source of aerosolised legionella bacteria. Methods The study analysed data on 1163 sporadic, community-acquired cases of Legionnaires9 disease in England and Wales with onset between 1996 and 2006, and 11630 postcode controls randomly sampled in proportion to population size and matched on region, age group and sex. The relationship between risk of Legionnaires9 disease and distance from a WCS was analysed by conditional logistic regression. Results Cases and controls had a mean age of 56.3 years; 79.3% were male. Cases lived appreciably closer to WCS than their controls (mean distance of cases=2.11 km, controls=2.58 km; mean difference 0.47 km (95% CI 0.28 to 0.65)). The OR for disease within 1 km of a WCS compared with over 6 km (a distance taken to reflect background rates of Legionnaires9 disease) was 1.59 (95% CI 1.26 to 2.01) when adjusted for socio-economic deprivation, and 1.33 (95% CI 1.04 to 1.71) when additionally adjusted for population density. The results suggest that residential proximity to a WCS may account for 19.6% of sporadic community-acquired cases. Conclusions WCS may be an important source of sporadic, community-acquired cases of Legionnaires9 disease, an observation that has important implications for health protection, especially given the likely increase in such systems as a component of strategies to improve energy efficiency in buildings.

The Portland Veterans Affairs Medical Center (PVAMC) participated in a research-based National Demonstration and Evaluation Study of Hospital at Home Care for Elderly Patients. PVAMC continued hospital at home care in a modified form based on the results of that research phase and feedback from patients, families, and staff. The modified clinical program (referred to as Program @ Home) provided care for the same diagnoses (exacerbation of congestive heart failure, exacerbation of chronic obstructive pulmonary disease, community-acquired pneumonia, cellulitis) but differed from the research-based demonstration project in that it accepted patients of all ages, accepted early-discharge patients from the hospital, and provided a less-intensive physician and nursing model. In the first 42 months, 290 patients were admitted; 23% came from the emergency room, 54% were early hospital discharge, and the remainder came from an outpatient clinic or home care. Average length of stay was 3.2 days, and 37% were younger than 65. The results describe how a home hospital program has been integrated into the clinical care offerings of a managed care health system and how it supports inpatient, primary, emergency, and home care programs.

Wet cooling systems are often associated with large outbreaks of Legionnaires' disease. Several European countries have legislation for registering such systems. The authors aimed to obtain an overview of the situation in Europe. A questionnaire survey was sent to 35 of the countries that collaborate in the European Working Group for Legionella Infections. In two countries it was passed to a regional level (to three regions in both Belgium and the United Kingdom), so that 39 countries or regions were sent the survey; 37 responded. Nine countries stated having legislation for the registration of wet cooling systems. Separate legislation exists at a regional level for two regions in Belgium and all three regions in the UK, giving a total of twelve countries/regions with legislation. In nine of these countries/regions, the legislation has been introduced since 2001. All of these countries/regions require periodic microbiological monitoring between twice a year and weekly; in nine, the legislation requires periodic inspection of the systems. Regulations for the registration of wet cooling systems should be required by public health authorities. During an outbreak of legionellosis, a register of wet cooling systems can speed up the investigation process considerably. The authors believe that the European Centre for Disease Prevention and Control (ECDC) should take the initiative to propose European Community (EC) regulations for all Member States.

Background Currently, the monitoring of influenza vaccination uptake is mainly a national issue. As influenza infection easily crosses international borders, it is in the interest of all countries to have a high vaccine uptake in people who may be vulnerable when influenza spreads. A Europe-wide monitoring system can provide insight into the strengths and weaknesses of uptake rates in countries and, once sufficient levels are achieved, can safeguard the continuation of the achieved levels. Aims This paper aims to address the following issues: a) How is influenza vaccination uptake monitored in Europe? b) What methods to monitor vaccination uptake are available and what are their limitations? c) What steps should be taken to implement a European-wide influenza vaccination uptake monitoring system? Methods Based on existing literature and experiences in monitoring influenza vaccination uptake, an approach to set up a European-wide monitoring system is proposed. Results The following issues were identified as relevant for influenza vaccination uptake monitoring: a) Agreement on the population groups in which vaccination uptake should be monitored b) The frequency of data collection c) The importance of sharing experiences regarding existing influenza vaccination campaigns in order to learn from each other, and develop ‘best practices’ d) The need to publish uptake data in close relation with influenza surveillance data and other European efforts on dissemination of vaccination knowledge. Recommendations To stimulate the discussion on implementing a pan-European influenza uptake monitoring scheme the following recommendations were suggested: a) Develop a common set of variables b) Build on experience from individual countries c) Create a coordinating body d) Create or identify a platform to publish the data; e) Start small and expand rapidly.

Travel and tourism is an increasingly important economic activity throughout Europe and the rest of the world.

In England and Wales, clinical reports from primary care showed that influenza activity for the season 2005/06 only rose above the base line for four weeks during February 2006. However, outbreaks of influenza-like illness and/or gastrointestinal infection in schools began to be reported to the Health Protection Agency, Centre for Infections in early January 2006. To quantify the type, size and the spread of these outbreaks a reporting form was distributed to local Health Protection Units in England and to Wales for retrospective and prospective weekly completion. Between weeks 48/05 and 11/06, a total of 688 school outbreaks were reported, including 658 outbreaks of influenza-like illness with or without other symptoms. The remaining 30 outbreaks listed as gastrointestinal only were excluded from the present analysis. Influenza B was confirmed in 70 outbreaks where testing took place. 61% of the outbreaks were reported from primary schools for children aged 4-11 years. This large scale outbreak in school children with flu-like illness across England and Wales was not picked up by most of the routine surveillance schemes, therefore, we believe that a school absentee monitoring and reporting system may be needed to give an early warning of increased influenza activity, especially for the mild form of the disease caused by influenza B virus

On 27 July 2007, the Department of Communicable Disease Control and Prevention of Stockholm County Council (SCC), Sweden, was informed by one of the infectious diseases hospitals in Stockholm that three elderly women travelling on a cruise ship had been hospitalised due to severe pneumonia

Background: EWGLINET, the European surveillance scheme for travel associated Legionnaires' disease, was established in 1987 following the identification of the disease in 1976. In 1998, the European Commission's Decision 2119/98/EC provided a legal framework for EWGLINET's operation, and its aims and objectives were formalised. Since its inception, the scheme has encountered a number of challenges which have influenced its development as a Disease Specific Network. The solutions to these challenges, and their successes, may be of interest to similar schemes. Aim: This article traces the development of the scheme and its responses to the challenges it has encountered. Results: One especially significant document developed by the scheme is the European Guidelines for Control and Prevention of Travel Associated Legionnaires' Disease;[1][1] its history is explored. In addition, EWGLINET's relationship with collaborating centres and other groups such as tour operators is highlighted. Conclusions: Despite changing over time, the collaborations and partnerships have been maintained and continue to ensure a close cooperation, maximizing public health effects. [1]: #ref-1

A recent rise in the number of sporadic legionellosis cases has been observed in England and Wales and the Netherlands which may be associated with certain weather conditions.

Twelve cases of community acquired legionnaires’ disease have been associated with south east London in July and August 2005

On 1 July 2002, EWGLINET introduced European guidelines for the control and prevention of travel-associated legionnaires' disease. This paper presents the results gathered by the surveillance scheme during the first two and a half years of the operation of the guidelines (to the end of 2004). Two hundred and thirty-seven new clusters and 70 cluster updates were identified. Investigations at 146 sites returned positive samples for legionella, and the proportion of positive sites reached over 60% in 2004. Thirty-four cluster sites were reported to have been investigated satisfactorily, but have gone on to be associated with subsequent cases ('repeater sites'). Fifty-one sites were published on the European Working Group for Legionella Infections (EWGLI) website; the publication states that EWGLINET cannot be confident that the sites have adequate control measures in place. The operation of the guidelines is discussed, and the situation in Turkey highlighted as a particular success.

Background Routinely collected data from patients registered with general practices participating in the General Practice Research Database (GPRD) were used to analyse influenza vaccine uptake and distribution in England and Wales between 1989/90 and 1996/97. Major changes to influenza immunization policy were introduced in 1998 and 2000 when immunization of the elderly became age related rather than risk related. This new study examines trends in vaccine uptake for high- and low-risk patients and the impact of the policy changes on uptake in the elderly. Methods Between 0.5 and 2.7 million patients registered with practices participating in the GPRD from 1989 to 2004 were included. Data were examined by age group, medical risk group and evidence of vaccination per study year. Results Vaccine uptake among high-risk persons aged 65 or more increased from 36.7 per cent in 1989/90 to 72.1 per cent in 2003/04. For the same period, uptake rates for highrisk persons under 65 years increased from 10.8 to 24.3 per cent. For those at high risk, uptake by females was higher in all age groups up to 65 years. Of those that were vaccinated, a higher proportion of the 65 and over were vaccinated in October each year compared with the high risk under 65 (p < 0.001). Conclusions Coverage among high-risk patients in younger age groups continues to fall well below satisfactory levels, especially among the youngest groups. Government policy should now focus on ways to improve uptake in these patients.

Much data has been gathered by the EWGLINET scheme on the distribution of cases of travel-associated Legionnaires' disease (TALD) by country of infection, but less analysis has been carried out on the distribution of these cases within countries. Travel-associated cases with onset in 2002 linked to France, Italy, Spain and Turkey were mapped. Rates of Legionnaires' disease per 100000 tourists were calculated for internal and foreign visitors for the regions of each country, and mapped. Rates of 1·5 cases/100000 and 2 cases/100000 tourists were classified as ‘high’ and ‘very high’ respectively. Cases of TALD were concentrated in certain regions, but when rates were calculated using tourist data, the results were relatively constant throughout each country. Rates were higher among foreign visitors than internal visitors; three of the countries had at least one region with ‘high’ rates, whilst Turkey additionally had three regions with ‘very high’ rates.

The European Working Group for Legionella Infections (EWGLI) was set up in 1986 and introduced the European Surveillance Scheme for Travel Associated Legionnaires’ Disease in 1987. The microbiologists working in reference laborat-ories and the epidemiolog

The UK influenza season of 2001/2002 was characterized by widespread geographic circulation of a new subtype of influenza A (H1N2) virus throughout the duration of the season. Younger children were predominantly infected, suggesting primary infection. Despite this, the public health impact of this new virus was minimal. It remains to be seen whether influenza A (H1N2) virus will persist and co-circulate with the previously circulating subtypes of influenza A (H1N1 and H3N2), or whether it was a sporadic recombination event that will disappear. Ultimately, the emergence of a new strain highlights the importance of continual surveillance of circulating viruses.

The European Surveillance Scheme for Travel Associated Legionnaires' Disease (EWGLINET) was notified of 360 cases in 2000 and 481 cases in 2001, the highest number reported since 1987. This increase reflects enhanced surveillance activities, especially in the Netherlands, France and Italy, mainly through urinary antigen detection test (78% of cases in 2001). The median delay in reporting to the scheme fell to under 30 days, at the cost of some loss of information on the outcomes of illness. In 2000, 28 clusters were detected compared to 72 in 2001, most of this rise resulting from a change in the definition of clusters. In 2000 and 2001, 55 and 140 environmental investigations were reported respectively.

The European Surveillance Scheme for Travel Associated Legionnaires#39; Disease was established in 1987 to identify clusters and outbreaks of cases of the disease. Twenty-nine collaborating centres in 25 countries contribute case reports in a standard format

The European Surveillance Scheme for Travel Associated Legionnaires Disease was set up by the European Working Group on Legionella Infections (EWGLI) in 1987 to identify cases of legionella infection in returning travellers and to detect outbreaks and cl

Immunisation against influenza is an important means of reducing morbidity and mortality amongst high-risk groups, and especially the elderly. Although immunisation has been recommended for these groups for many years, no timely or comprehensive monitoring at the national level was carried out in England before the beginning of this century. Annual numbers of doses administered were available, but not to whom they were given. This paper describes the results of a national rapid reporting scheme for England that was introduced by the Department of Health (DH) and PHLS Communicable Disease Surveillance Centre (CDSC). The scheme collected monthly data on influenza immunisation uptake among people aged 65 or more during the winters of 2000/2001 and 2001/2002. Results showed that DH annual targets of 60 and 65%, respectively were both met, although considerable variation occurred at the local levels. The feasibility of this method of monitoring immunisation uptake in the elderly is discussed.

During influenza epidemics, school-aged children are amongst the first affected patients. They frequently then spread the virus within their families. Recognising influenza activity in schools may therefore be an important indicator of early activity in the wider community. During 2005/06, influenza B was associated with high levels of morbidity in school-children and over 600 schools outbreaks were reported to the Health Protection Agency by local Health Protection Units. While it is not possible to directly monitor influenza in schools, the feasibility and validity of using sentinel school absenteeism data, as a proxy for influenza in the community can be investigated. From week 02/07 to 20/07, eight primary and three secondary schools from five HPA regions were able, via the Department of Health-funded Health Protection Informatics website, to report daily electronic registration data, relating to absenteeism due to illness. Aggregated absenteeism data due to illness peaked the same week as indices for the age group comparable to that used by the Royal College for General Practitioners and NHS Direct schemes. When illness-defined absenteeism data was stratified into primary and secondary schools, absence in primary schools peaked one week before that in secondary schools and the established schemes for all ages. The start time of the study meant that initial increases in activity could not be measured. These encouraging results justify expanding this sentinel scheme to collect more rigorous evidence of the usefulness of absenteeism as a proxy for influenza activity and a tool to inform policy and trigger local responses.