Adelges tsugae Annand, HWA) remains the single greatest threat to the health and sustainability of hemlock in the eastern USA. The loss of hemlock trees leads to further negative impacts on the diversity and stability of ecosystems in the eastern part of North America. It is, therefore, urgent to develop effective control measures to reduce HWA populations and promote overall hemlock health. Currently available individual and integrated approaches should continue to be evaluated in the laboratory and in the field along with the development of other new and innovative methods.Hemlock woolly adelgid ( Tsuga genus in the Pinaceae family. The Tsuga genus contains nine species -4,5-dihydroimidazol-2-yl} nitramide), a chloronicotinyl insecticide, is used for controlling sucking insects, soil insects, termites, and some chewing insects. It is one of the essential insecticides for HWA control in eastern hemlocks in the southern Appalachians [Current HWA control is largely focused on using either chemical or biological methods ,83. Preslachians ,94,95. Ilachians . As a melachians ,98. Imidlachians ,104,105.lachians , which alachians . Researclachians ,103,107.Benton et al. analysed the levels of imidacloprid and its metabolites in the foliage of hemlocks growing in the GSMNP to understand the longevity of imidacloprid treatments. Four to seven years post basal drench treatment, imidacloprid and olefin were still detectable in more than 65% of branchlets, although the levels were below the LC50 for HWA . While tIn 2013, the European Commission adopted a proposal to restrict the use of three insecticides, including imidacloprid, for two years due to the uncertainty about their chronic risk to ecological system . ImidaclChemical control as a stand-alone treatment is not a feasible option in a forest setting due to geographical constraints, or the height of the trees when bringing in equipment is inconvenient . In addiA. tsugae has no known parasitoids and no specific pathogens, so the search for natural enemies is limited to predators [Laricobius rubidus (LeConte), fed a diet of HWA completed development to the adult stage on A. tsugae as well as it did on its primary host Pineus strobi Hartig [L. rubidus has the potential to contribute to the biological control of A. tsugae in the eastern United States. Scymnus suturalis Thunberg moves among eastern white pine, Scotch pine (Pinus sylvestris L.) and eastern hemlock depending on the presence of adelgid prey; however, this predator is not found on eastern hemlock after July [S. suturalis for a source of adelgid eggs throughout the summer may limit its effectiveness as a predator of A. tsugae [The discovery and utilization of effective biological control agents is critical to help restore hemlock forests ,93. A. tredators . The fewredators . Evidenci Hartig . Therefoter July . The app. tsugae . Another. tsugae .Sasajiscymnus tsugae, a native predator of HWA in Japan was initially imported to and studied in New Jersey. Field evaluations demonstrated that S. tsugae could establish, locally disperse, and survive heat waves in southern states and severe winters in northern states [S. tsugae have been released at more than 400 sites on federal and non-federal lands from South Carolina to Maine [S. tsugae that recovered from the lower crown were relative low [S. tsugae on hemlock trees recovery.An effective biological-control agent should not disturb or disrupt the integrity of other aspects of its native habitat . The iden states . Since 1to Maine . Howevertive low . More reLaricobius nigrinus beetles, which are native to the northwestern United States and Canada, and feed exclusively on adelgids, have also been used for the biological control of HWA. After approval for release in 2000, L. nigrinus were distributed throughout the invasion range of HWA and isolated populations have established from the southern Appalachians to New England [S. tsugae, there is evidence that L. nigrinus can exert substantial mortality on HWA and its preference to feed on HWA over other adelgids, making it a viable biological control candidate [Laricobius osakensis, a species related to L. nigrinus, is a voracious predator of HWA widely spread in Japan. Recently, Arsenault, et al. have shown that L. osakensis, reared in the laboratory in North Carolina with little to no prior exposure to field environmental cues, responded preferentially to odours produced by eastern hemlock (regardless if it was infested by HWA or not) and moved promptly onto it [L. osakensis relies on volatiles produced by HWA’s host trees to locate potential prey. The phenology of L. osakensis is highly synchronous with the life cycle of HWA, making it a good candidate for biological control of HWA in North America [ England ,129,132.andidate . Laricob America ,134,135.L. nigrinus on hemlock has been reported between L. nigrinus, the predator introduced from western North America and L. rubidus [Laricobius species [L. osakensis and L. nigrinus will mate but produce only sterile eggs. This is fortunate because L. osakensis appears to be a better HWA predator than L. nigrinus [Recently evidence has emerged indicating imported predator beetles could hybridise in the field with native species of HWA predators. Widespread hybridisation with asymmetrical introgression towards  rubidus ,133,136. rubidus , and uni rubidus ,137,138; rubidus ; enhance rubidus ; decreas rubidus ,141,142  species . It was nigrinus ,144.S. tsugae and L. nigrinus. In the laboratory setting, S. tsugae showed increased activity during the day and at higher temperatures, while L. nigrinus showed increased activity during the night and in spring-like conditions. In a field study, it was found S. tsugae and L. nigrinus established two years after release onto eastern hemlocks indicating these two predators can survive and coexist. This is encouraging because they differ temporally in their occurrence and predation of HWA with L. nigrinus active from late fall to spring and S. tsugae from spring to early summer [Ideally, a combination of predators should be used to control HWA populations as opposed to one single species. Flowers and colleagues  studied y summer . These ry summer . More reA long-term, sustainable, and cost-effective approach to control HWA will have to integrate chemical control, biological control, and host-plant resistance methods into one comprehensive management programme ,146,147.Laricobius species; and the combination may result in suppression of HWA populations below damaging levels. Hemlock woolly adelgid remains the single greatest threat to the health and sustainability of hemlock in the eastern USA. Currently along with the development of other new and innovative methods, available individual and integrated approaches should continue to be evaluated not only in the laboratory but specific emphasis should also be focus on their application and effectiveness in the field.The introduction of the combination of multiple specialist predators is a promising biological intervention strategy. However, how to translate and test a successful laboratory story in the field on a much larger scale at various geographical locations, and how to effectively evaluate predators’ behaviour in the field, including their immigration and emigration propensity in response to prey abundance or intraspecific and interspecific predator cues, is still largely unaccomplished . Further


Only 22.8% of participants thought that Germany could accompany more URM. While few participants argued in support of immediate deportation of URM in general (38.6%) or of URM from the Middle East (35.3%), a majority advocated for immediate deportations of URM from the Balkan region (62%) or from Africa (51.1%). Difference in the variance regarding attitudes towards deportation was explained mostly by right-wing political attitudes as well as by islamophobic attitudes and general rejection of asylum seekers. High rates of approval were found for guaranteeing the same chances to schooling or apprenticeship for URM as to German children and for bestowing URM a right to permanent residence if they were able to complete school or apprenticeship. Education and qualification are key to integration. Studies about needs and wishes of URM consistently report a high motivation to learn the language of their new host country and attend school. At this point, hopes of URM and expectations of society meet, which underlines the importance of participation in education as key factor in integration.Germany saw an increase in numbers of refugees in 2015, with nearly a third being below the age of 18. Unaccompanied refugee minors (URMs) present an especially vulnerable group. In addition to pre-flight and flight stress, the acculturation process can work as potential stressor, and we wanted to explore attitudes towards URM. We conducted a study in a representative sample ( Among people seeking refuge, adolescents and young adults present a majority and it has been estimated that around 50% of refugees worldwide are below the age of 18 . Among tMany of these refugee minors have experienced potentially traumatic events in their countries of origin (pre-flight experiences) and have been exposed to life-threatening or dangerous situations throughout their migration (flight experiences) . In addiDue to the lack of a family support system, which could potentially buffer stress, as well as accumulation of traumatic experiences, URMs have to be considered as especially vulnerable group , 12.In determining the prevalence of mental health disorders among URM, studies using clinical interviews stated rates between 41 and 56%, with a high percentage of posttraumatic stress disorder (PTSD) 20–30%) [0% [3]. Tn = 75) reported rates of mental disorders to be 75% [The most recent study from Germany based on data from a child and adolescent psychiatric outpatient clinic  of potentially traumatizing events in URM, thus resulting in a high “trauma load” , with a Still, little is known about the long-term course of mental disorders in the group of URM. In a study of 75 URM in Norway, a 1.9 year follow-up was conducted, showing stability of symptoms of depression, anxiety, and externalizing problems over time . A studyDespite a high burden of potentially traumatizing life events, a great number of URM seem to be mentally stable and present themselves as “resilient”, not showing symptoms of a mental disorder, with rates between 44 and 58% in different studies .n = 1209), at a timepoint, when many refugees had entered Germany over the summer, rates of approval for hosting refugees in the region the participants lived in were as high as 54% [In a survey conducted by the Allensbach Institute for Demoscopic Research in May 2015 , 1453 inh as 54% . Still, h as 54% .While in 2015, in Germany, “welcome culture” was a political slogan accepted by large parts of the general population proud about this humanistic attitude, recently the debate shifted to issues, such as better border controls and potential islamistic terrorist threats. In general, accepted standards of youth welfare for URM have been questioned by high-ranking politicians. Most child welfare organizations and advocacy groups as well as the German scientific society of child and adolescent psychiatry psychosomatics and psychotherapy appealed against this proposal .Given that the acculturation process in a new country can act as a potential stressor, and daily stressors have a severe impact on URM’s mental health , we wantn = 2524) of the German population (ages 14 years or older) between January and March 2016. Using a random route method, participants were visited at home and informed about the study by 228 research assistants. After providing informed consent, a demographic interview was conducted and paper and pencil questionnaires were handed to the participants. These questionnaires were collected later on. Out of 4830 households that were approached, 1541 were not available (after four approaches) or did not respond, 14 persons were of ill health or not able to fill out the questionnaires, and 731 refused to participate. Out of 2544 collected questionnaires, 20 were not analyzable, thus resulting in 2524 participants (52.3%).This study was conducted to gather information about attitudes towards URM from a representative sample . This 18 item scale consists of six subscales  with three items/subscale. This scale has been used in bi-annual representative population surveys in Germany from the year 2002 to detect trends in right-wing extremism attitudes –26. FurtThe survey was in concordance with the Declaration of Helsinki met ethical guidelines of the international code of Marketing and Social Research practice by the International Chamber of Commerce and the European Society for Opinion and Marketing Research and was approved by the Institutional Review Board of the Medical Faculty of the University of Leipzig. Statistical analysis was performed using IBM SPSS, Vers. 21.χ2: 5.19; p = 0.075). There was a tendency to a more positive attitude in younger participants with lower rates of approval in older participants. Furthermore, participants, who did not have a German citizenship, more often agreed to the statement that Germany has the potential to accept more URM . In participants with a lower monthly income after taxes, more negative attitudes were present . Having graduated from high school was associated with a more positive attitude   to the Balkans , Africa , and the Middle East   agreed that URM from the Balkan peninsula or from Africa should be sent back to their home countries. There was a gender effect with females being more clearly opposed to sending back URM to their home country in general  and the right to stay in Germany after finishing school or a job training .Looking into integration, questions regarding schooling/job training and housing were asked. There was a large majority both for granting URM the same schooling or apprenticeship possibilities as adolescents from Germany and for allowing URM with a finished school exam and job training to stay in Germany see Fig. . Female Asked in which place URM should live, most participants were in favor of youth welfare homes (44.6%), followed by refugee housings (29.2%) and foster care (18.5%) (elsewhere: 7.7%). This results in more than half of the participants approve of offering the same options to URM (youth welfare institutions and foster care) than to all other children in need in Germany.The variables gender, age, income, migration background, level of education, political attitudes (left- or right-orientation), right-wing extremism, islamophobia, and rejection of asylum seekers were entered into regression analyses.In a binary stepwise regression analysis, the question whether participants thought Germany was capable of taking on more URM (yes/no) was examined. A model, including the variables right-wing extremism, islamophobia, and rejection of asylum seekers, explained 44.3% of the variance. Islamophobia and rejection of asylum seekers predicted disagreement most strongly , followed by right-wing extremism . All other variables were excluded from the model due to insignificance. For further details, see Table In logistic stepwise regression analyses, the questions whether participants thought URMs should be sent back immediately, also in differentiation by origin countries of the URM, and questions regarding schooling or apprenticeship possibilities were analyzed.β = −0.4, T = −18.0, p < .001), right-wing extremism , rejection of asylum seekers , and political attitudes  significantly predicted attitudes towards this question.A model, including the variables political attitudes, right-wing extremism, islamophobia, and rejection of asylum seekers, explained 44.0% of the variance to the question whether URMs should be sent back immediately . Islamophobia , the same question regarding URMs from Africa was significantly predicted by islamophobia, rejection of asylum seekers, right-wing extremism, age and gender (model explaining 41.0% of the variance), and the same question regarding URMs from the Middle East was only significantly predicted by islamophobia, rejection of asylum seekers, and right-wing extremism (model explaining 42.0% of the variance). For further details, see Table The variables islamophobia, rejection of asylum seekers, right-wing extremism, political attitudes, and gender significantly predicted both questions whether URMs should have the same right to receive education as German youth (model explaining 17.0% of the variance) and whether URMs should have the right to stay in Germany if they completed their education in Germany (model explaining 15.7% of the variance). For further details, see Table Only a fifth 22.8%) of participants thought that Germany could accommodate more URM, whereas 45.6% responded negatively. This is quite a low rate, especially in comparison with the studies from Petersen  and Koec.8% of paWe found a differentiation for countries of origin when asking for immediate deportation as often claimed by right-wing populists. While fewer participants argued in support of immediate deportation of URM in general (38.6%) or of URM from the Middle East (35.3%), a majority advocated for immediate deportations of URM from the Balkan region (62%) or from Africa (51.1%). Of note, regression analysis revealed that attitudes to deportation were influenced by islamophobic tendencies and a general negative attitude towards asylum seekers. Gender and age only predicted attitudes towards deportation of youth from the Balkans or Africa, with generally more negative attitudes towards URM in male and in older participants. This might be due to the fact that there are no current war zones in the Balkans and these countries of origin have been declared as “safe” by the German administration. However, this is not true for a large number of countries or regions in Africa. It might be that our questions were to crude to differentiate between peaceful African nations and warzones, and it would be advisable to assess this distinction in future studies. It seems hardly understandable why islamophobic tendencies influenced tendencies towards deportation of URM from the Balkan peninsula, as only Bosnia and Hercegovina, and Albania and Kosovo have a muslim majority population. However, it might be that this result is due to over-generalization.Looking into explanations for attitudes towards deportation, we assessed both demographic variables as well as political attitudes. In doing so, we saw that political attitudes more leaning to the right spectrum, right-wing extremism, a general rejection of asylum seekers as well as islamophobic attitudes influenced the attitudes towards URM.The finding that right-wing political attitudes led to a lower acceptance of URM are in line with a study of 216 US students, also showing that a more conservative political attitude led to lower levels of acceptance regarding the intake of more refugees than a more liberal attitude .We saw that a higher level of education was associated with more positive attitudes towards URM, and it seems warranted to state that education holds the key to a more welcoming and accepting attitude towards URM. This is a finding often encountered in research on attitudes towards immigration per se .With regard to integration of URM already in Germany, high rates of approval where found for guaranteeing the same chances to schooling or apprenticeship for URM as to German children. This is in line with Article 22 of the Convention of children’s rights . In addiAlthough it has been shown that psychiatric disorders in URM seem to remain stable over time , some faIt seems that education and qualification hold the key to integration based on a large consensus in German society. Of note, studies about needs and wishes of URM consistently report a high motivation to learn the language of their new host country and attend school . At thisAlthough our study presents a representative sample of the German population, some limitations apply: (1) although answers were provided in questionnaire that was handed back in a sealed envelope, it might well be that we encountered a social desirability bias, shifting answers towards a more positive attitude towards URM. However, this process still seems preferable to—for example—telephone surveys, who are even more prone to this bias. (2) Due to the sampling method, some individuals could not be reached, especially those in institutions or homes and those not literate in German. These factors may have influenced representativity. (3) The evaluation was not based on a validated assessment scale. However, we decided to apply questions used in further large studies on the attitudes towards refugees in Germany , 18 to bOverall, to the best of our knowledge, this is the first study on attitudes towards drawing from a nationwide representative sample. Despite the low rates of approval for accepting more URM, the need for offering opportunities to URM and integrating them into society seems to be based on a broad consensus in the general population that contradicts political attempts to lower standards of youth welfare and to deny young refugees over the age of 18 any educational or apprenticeship perspective.


Palliative chemotherapy is the mainstay of treatment of advanced gastric carcinoma (GC). Monoclonal antibodies including trastuzumab, ramucirumab, and pembrolizumab have been shown to provide additional benefits. However, the clinical outcomes are often unpredictable and they can vary widely among patients. Currently, no biomarker is available for predicting treatment response in the individual patient except human epidermal growth factor receptor 2 (HER2) amplification and programmed death-ligand 1 (PD-L1) expression for effectiveness of trastuzumab and pembrolizumab, respectively. Multi-platform molecular analysis of cancer, including GC, may help identify predictive biomarkers to guide selection of therapeutic agents. Molecular classification of GC by The Cancer Genome Atlas Research Network and the Asian Cancer Research Group is expected to identify therapeutic targets and predictive biomarkers. Complementary to molecular characterization of GC is molecular profiling by expression analysis and genomic sequencing of tumor DNA. Initial analysis of patients with gastroesophageal carcinoma demonstrates that the ratio of progression-free survival (PFS) on molecular profile (MP)-based treatment to PFS on treatment prior to molecular profiling exceeds 1.3, suggesting the potential value of MP in guiding selection of individualized therapy. Future strategies aiming to integrate molecular classification and profiling of tumors with therapeutic agents for achieving the goal of personalized treatment of GC are indicated. Helicobacter pylori (H. pylori) and human papilloma virus (HPV) are involved in multi-step processes causing chronic gastritis, intestinal metaplasia, and invasive carcinoma [H. pylori and HPV would be a logical strategy for prevention of some types of GC, but no randomized trial to date has shown a clear benefit of this approach [With about a million diagnosed cases and over 700,000 deaths recorded annually, gastric carcinoma (GC) is the third most common cause of cancer deaths worldwide . While 8arcinoma . Populatapproach . Until aSystemic chemotherapy and targeted therapy play important roles in the multi-disciplinary management of GC. With the exception of GC diagnosed at T1 stage, chemotherapy is employed in the neoadjuvant and adjuvant settings, or concurrent with radiation therapy. Palliative combination chemotherapy and targeted therapy are the only treatment options for patients with advanced or metastatic GC. Selection of chemotherapeutic drugs is typically based on performance status, medical comorbidities, and medical oncologist’s experience or preference. There are no valid biomarkers predictive of treatment response of GC to therapeutic agents. Exceptions are, amplification of human epidermal growth factor receptor 2 (HER2) and expression of programmed death-ligand 1 (PD-L1), for which trastuzumab and pembrolizumab, respectively, have been demonstrated to produce clinical benefit ,12. PrelThe genomic characterization of GC has led to the development of two new classifications of GC by The Cancer Genome Atlas (TCGA) Research Network  and the The goal of this article is to provide a critical review of the molecular characterization of GC, and elaborate on the molecular features that can be translated into therapeutic biomarkers and targets for clinical use. First, we provide an overview of the conventionally used systemic chemotherapy and targeted therapeutics of GC. The data on molecular classification of GC by TCGA and ACRG as well as molecular profiling of GC are examined. The potential of translating the molecular classification and profiling of GC into therapeutic targets and predictive biomarkers are discussed. We hope that this article will help identify the opportunity and challenge of developing strategies towards the goal of precision medicine in GC by improving therapeutic efficacy and minimizing treatment-related toxicity.Systemic chemotherapy is employed for treatment of patients with localized GC as well as for those with advanced GC. Surgical resection with pre- and post-operative chemotherapy and/or radiation therapy represents the primary curative treatment of early-stage GC with 5-year survival rate of less than 30% ,19,20. FChemotherapeutic regimens currently being used for GC consist of anthracycline, fluoropyrimidine, taxane, and platinum-based agents. For advanced or metastatic GC, first-line combinations such as EOX  and DCF  have produced limited survival benefits, with median survival not exceeding one year . Second-Despite the clinical heterogeneity and molecular complexity of GC, targeted therapeutics directed against the genetic mutations and signaling pathways that drive tumor growth and invasion have been developed and clinically investigated. Targeted therapies currently in clinical use include trastuzumab, ramucirumab, and pembrolizumab. Other targeted therapeutics directed against the signaling pathways of mitogenesis, angiogenesis, and immune checkpoints are under clinical investigation for treatment of GC.Human epidermal growth factor receptor  is a transmembrane tyrosine kinase receptor of the ErbB family. The ErbB members play important roles in regulation of cellular functions including proliferation, growth, survival, adhesion, migration, and differentiation. HER2 acts by heterodimerization with other ErbB family receptors leading to activation of the RAS-MAPK (mitogen-activated protein kinase) and PI3K-AKT (phosphatidylinositol 3-kinase—AKT) pathways. HER2 has been found to be over-expressed in 20% to 30% of GC depending on the tumor subtype and location.p < 0.01) as well as prolonged progression-free survival  and overall survival  [Several HER2-targeting agents have been developed and evaluated in phase III trials for patients with advanced HER2-positive gastroesophageal junction carcinoma (GEJC)/GC. Trastuzumab is a humanized IgG1 monoclonal antibody (mAb) directed against the extracellular domain of HER2, and it prevents dimerization of the HER2 receptors. This triggers receptor internalization and mediates antibody-dependent cell-mediated cytotoxicity (ADCC), resulting in inhibition of tumor growth . In the  < 0.01) . FurtherSeveral signaling pathways are involved in tumor-associated angiogenesis, such as those activated by vascular endothelial growth factor (VEGF) , angiopoVEGF is a growth factor secreted by the tumor to stimulate formation of new blood vessels in response to hypoxia and nutrient depletion. When it binds to VEGFR, a complex cascade of downstream signaling pathways is activated, resulting in neovascularization, vasodilation, and increased vascular permeability . Blockadp = 0.0473) and PFS  in patients previously treated with first-line platinum- or fluoro-pyrimidine-based therapy [p = 0.0169), PFS , and ORR  compared with those treated with paclitaxel alone [Ramucirumab is a recombinant humanized monoclonal antibody (mAb) that binds to VEGF-R2 and prevents its activation by VEGF. In contrast to bevacizumab (anti-VEGFA mAb), it has shown clinical efficacy as a single agent  and in combination with paclitaxel . Based on the results of these studies, ramucirumab either alone or in combination with paclitaxel has become standard second-line treatments for advanced GEJC/GC. In the REGARD trial, ramucirumab was associated with a significant improvement in OS  and 2 (PD-L2) are normally expressed on antigen-presenting cells (APC) and also on tumor cells. Binding of PD-L1 and PD-L2 to their receptors (PD-1) on activated T cells leads to downregulation of cytotoxic T-cell activity and causing immunosuppression. PD-L1 is expressed in 15% to 70% of GCs, and they are associated with poor prognosis . Pembrolp = 0.102), PFS (p = 0.162), and OS (p = 0.124) was observed. In a phase II study (KEYNOTE-059) of 259 patients, pembrolizumab monotherapy showed clinical efficacy in previously treated advanced GEJC/GC [In the phase Ib KEYNOTE-012 trial, the activity and safety of pembrolizumab were evaluated in a cohort of 39 patients with advanced GEJC/GC. Pembrolizumab produced an ORR of 22.2%, 6-month PFS rate 24%, and OS rate 69% ,41. An aAdvances in next-generation sequencing (NGS) technologies and improved understanding of cancer biology have unlocked opportunities to characterize the genomic landscape of cancer including GC. Using multi-platform analyses, molecular profiling of GC has enabled The Cancer Genome Atlas (TCGA) Research Network and the Asian Cancer Research Group (ACRG) to classify GC into subtypes. The new molecular classification of GC is complementary to the conventionally used system of subtyping GC based on histopathology. Importantly, molecular classification of GC helps identify molecular alterations that may be targeted for therapy. Furthermore, molecular profiling of GC collected from individual patients using a multi-platform approach has offered new opportunity to identify biomarkers that may be predictive of tumor response to treatment ,43,44.Molecular classification of GC by the TCGA Research Network utilized six distinct platforms, including exome sequencing, copy number analysis, methylation, expression of miRNA and mRNA. Based on TCGA molecular data, GC were divided into four groups: Epstein–Barr virus-positive , microsatellite instability , chromosomal instability , genomically stable  . Each ofPIK3CA mutation, recurrent JAK2 and ERBB2 amplifications, interleukin-12 (IL-12) mediated signaling, and PD-L1/2 overexpression. The presence of viral antigens such as EBV  has been shown to result in increased neo-epitope presentation [PIK3CA (80%) suggests that inhibition of PI3K warrants further evaluation in EBV-positive GC.The EBV-positive tumors were found to be mainly located in the gastric fundus or body. They exhibited higher prevalence of DNA promoter hypermethylation, A to C transversions, entation , which mentation ,41. In aMLH1 promoter region (leading to MLH1 silencing), and targetable hotspot mutations in PIK3CA, ERBB3, ERBB2, and EGFR. Of note, the V600EBRAF mutation commonly seen in MSI colorectal cancer was absent. However, gastric MSI tumors had a high rate of PD-L1 expression. In particular, recent evidence shows that enhanced anti-PD-1 responsiveness of mismatch repair-deficient tumors is related to the high number of mutation-associated neoantigens [The MSI tumors, characterized by genomic instability due to a deficient DNA mismatch repair system, lacked targetable amplifications. This subtype of tumors was noted to have hypermethylation in the antigens .TP53 mutations (71%), as well as genomic amplifications of RTKs and cell cycle mediators. Phosphorylation of EGFR was significantly elevated. Recurrent amplification of the gene encoding ligand VEGFA was also notable. Additionally, frequent amplifications of cell cycle mediators  were present. Alterations of these genes have been confirmed in a cohort of 116 advanced/metastatic GC cases [The CIN tumors were more frequent in the gastro-esophageal junction/cardia. They were noted to have the highest frequency of GC cases .CDH1 encoding the cell adhesion molecule E-cadherin in hereditary diffuse gastric cancer [RHOA  and fusion of CLDN18-ARHGAP6 or 26 (30% of cases). RHOA modulates programmed cell death and actomyosin-dependent cell contractility and motility [CLDN18 and ARHGAP6 are involved in intercellular tight junction structure and Rho signaling activation, respectively. Thus, alterations in either RHOA or CLDN18-ARHGAP6 might contribute to lack of cellular cohesion, dispersed growth, and resistance to programmed cell death.The GS tumors, which lack either chromosomal alteration or microsatellite instability, exhibited elevated expression of molecules in the cell adhesion and angiogenesis-related pathways. Previous studies had demonstrated loss of the tumor-suppressor gene c cancer . The TCGmotility ,47,48, wRHOA and CLDN18 gene products could be exploited to develop new therapeutic strategies in the genomically stable subtype.The TCGA data indicate that each of the four defined molecular subtypes displays distinct but overlapping candidate therapeutic targets. These suggest the potential of targeted therapeutics in each subtype of GC . The dis+ (mutated) 26%, and MSS/TP53− (wild-type) 36%  p53-activity signature. Complementary to the TCGA data, the ACRG proposed a classification of GC that correlates four molecular subtypes with distinct patterns of molecular alterations, disease progression and prognosis. The molecular analyses include principal component analysis (PCA)  of expreype) 36% . TP53 isKRAS (23.3%), the PI3K-PTEN-mTOR pathway (42%), ALK (16.3%) and ARID1A (44.2%) [The MSI tumors, as in the TCGA cohort, were found to be hypermutated ,58 intes (44.2%) .+ phenotype is associated with a better prognosis, and a higher prevalence of mutations in APC, ARID1A, KRAS, PIK3CA and SMAD4, compared to the MSS/TP53− phenotype. Consistent with these observations, mutations in TP53 (54%), APC (10%), SMAD4 (5.9%), KRAS (5.9%), and PIK3CA (5.1%) were present at a high rate in a large cohort of 666 specimens of GC [− phenotype exhibited the highest prevalence of TP53 and RHOA mutations, as well as recurrent focal amplifications in ERBB2, CCNE1 and CCND1.The MSS/TP53ns of GC . The MSSCDH1 expression.The MSS/EMT phenotype includes tumors of the diffuse-subtype. It is associated with the worst prognosis, tendency to occur at an earlier age, and the highest recurrence frequency (63%) of the four subtypes. The MSS/EMT subtype also includes a large set of signet ring cell carcinomas and showed loss of The ACRG subtyping of GC could be complementary to the TCGA system for molecular classification of GC. The ACRG data are potentially important for generating prognostic biomarkers in GC. The validity of these biomarkers for prognosis of patients with GC will need to be investigated in prospective clinical studies.+, and CIN subtypes were enriched in ACRG MSS/EMT, MSS/TP53+, and TP53− subtypes, respectively. However, CDH1 and RHOA mutations were highly prevalent in the TCGA GS subtype but infrequent in the ACRG MSS/EMT subtype, these two subtypes were deemed not equivalent. Similarly, MSS/TP53 did not overlap with the TCGA EBV subtype, as EBV+ tumors represented a small proportion of samples in the MSS/TP53+ subtype.TCGA and ACRG integrated the results of a wide scale molecular analysis into two different and partially overlapping models encompassing four molecular subtypes with distinct salient genomic features . They boPossible reasons for the partial overlap of these two classifications include differences related to the patient population (Korea in ACRG vs. USA and Western Europe in TCGA), tumor sampling , and technological platforms . Despite these differences, these two classification schemes not only clarified and simplified the genomic and epigenomic heterogeneity of GC, but also revealed distinct salient genomic features among gastric cancer subtypes linked to clinical outcomes. These molecular classification systems of GC lay the groundwork for targeted therapies, patient stratification for clinical trials and treatment, and improved prognostication.Complementary to molecular classification of GC, analysis of molecular profiles of tumors collected from individual patients using a multi-platform approach has led to identification of targets for therapy as well as biomarkers that may be predictive of tumor response to treatment.O-6-methylguanine-DNA-methyltransferase (MGMT), phosphatase and tensin homolog deleted on chromosome ten (PTEN), topoisomerase (TOP), thymidine synthase (TS), and excision repair cross-complementing 1 (ERCC1). Molecular profiling of tumors including GC has been employed with the hope of identifying actionable and predictive biomarkers. In one study, 666 specimens of GC were analyzed by immunohistochemistry, in-situ hybridization, and genomic DNA sequencing . Some ofNegative expression of three non-NCCN  compendium actionable targets including RRM1 (62%), MGMT (45%), and PTEN (58%) was identified in more than 40% of the tumor specimens. These data suggest potential sensitivity to gemcitabine, temozolomide, or PI3K inhibitors, respectively. Negative RRM1 expression is associated with higher response rates to gemcitabine-based chemotherapy regimens ,60,61. TIn the HER2-positive cohort, co-expression of TOP2A occurred most frequently (93%), suggesting potential sensitivity to a combined anthracycline/trastuzumab approach to treatment. Furthermore, 50% of patients demonstrated possible benefit from a combination of trastuzumab with 5FU/capecitabine based on concurrent low TS, 53% with irinotecan (high TOPO1), 63% with cisplatin (low ERCC1) and 55% with gemcitabine (low RRM1). ® service . These included eleven GC, nine EGJC, and seven esophageal carcinoma (EC) [TP53 (33%), APC (7.4%), SMAD4 (7.4%), and PIK3CA (7.4%), were consistent with those in a cohort of 666 specimens of GC [The potential of these biochemical markers to predict treatment response of tumors to chemotherapy was examined. Molecular profiling of tumor specimens from 27 patients with gastroesophageal carcinoma was conducted by the Caris Molecular Intelligenceoma (EC) . The frens of GC . In several cases, the PFS based on tumor molecular profile (MP) was compared to that on therapy prior to molecular profiling. A ratio of PFS-MP to PFS prior to MP greater than 1.3 is considered clinically significant. As shown in the three cases in Systemic treatment plays important roles in the multi-disciplinary management of gastric carcinoma. Cytotoxic drugs, targeted agents, and immunotherapeutics have been shown to provide clinical benefit though to a limited extent. With the exception of trastuzumab for HER2-amplified and PD-L1-expressing GC, a clinical tool to predict the treatment response and outcomes of the currently used systemic therapy is lacking. Moreover, tumor heterogeneity and molecular evolution of tumor during treatment contribute to therapeutic resistance. Clinically tested and validated biomarkers for predicting tumor response to systemic treatment will be needed for patients to derive maximal benefit and avoid unnecessary toxic side effects.Molecular classification and profiling of GC generate potential targets for therapy as well as prognostic and predictive biomarkers. The TCGA and ACRG data have not only revealed the molecular and etiologic differences across the various subtypes of GC, but also yielded many potentially targetable genomic changes. In addition, molecular profiling of GC by analysis of proteomics ,64,65 anDevelopment of therapeutic agents targeting some of those molecular alterations as defined to the subtyping and profiling of GC are undergoing pre-clinical and clinical investigation. Identification and validation of prognostic and predictive biomarkers by correlation of molecular profiles of tumors with clinical outcomes such as tumor response, progression-free survival, and overall survival are indicated. Future studies aiming to identify and validate predictive tumor biomarkers through molecular profiling in large data sets are indicated. Results of these studies are expected to facilitate selection of optimal chemotherapy regimen individualized for the patients, and the development of novel targeted therapies.While the molecular data brings the possible hope of developing precision therapies, many challenges must be overcome to fully understand and realize their clinical impact. First, it is imperative to design and implement clinical trials that take into account the molecular heterogeneity across the various subtypes of GC and develop protocols specific for each of these entities. The Personalized Antibodies from Gastro-Esophageal Adenocarcinoma (PANGEA) “umbrella trial” is one such innovative trial in which patients are assigned to different treatment arms by matching the molecular characteristics of a single tumor type to a specific drug ,69. ConsSecondly, targeted therapy guided by molecular profiling will need to be tested in patient-derived tumor xenografts (PDX) and genetically-engineered mouse (GEM) models. The development and characterization of these realistic model systems represent the complex molecular heterogeneity of GC. They will be helpful for validating the genomic alterations in the molecular subtypes of GC and facilitating drug and biomarker development. Finally, development of novel therapies combining immunotherapeutics, cytotoxic chemotherapeutic agents, and molecularly-targeted therapeutics is expected to offer durable clinical benefits and maximize survival in patients with GC.By integrating the various molecular and clinical data, we hope to develop strategies that will enable clinicians and scientists to better characterize and classify these tumors, develop targeted therapies, and identify prognostic and predictive biomarkers for achieving the goal of precision therapy in patients with this malignant disease.


In general, media interest and reporting peaks and wanes during the course of an outbreak. In this study, we quantify the extent to which media interest during infectious disease outbreaks is indicative of trends of reported incidence. We introduce an approach that uses supervised temporal topic models to transform large corpora of news articles into temporal topic trends. The key advantages of this approach include: applicability to a wide range of diseases and ability to capture disease dynamics, including seasonality, abrupt peaks and troughs. We evaluated the method using data from multiple infectious disease outbreaks reported in the United States of America (U.S.), China, and India. We demonstrate that temporal topic trends extracted from disease-related news reports successfully capture the dynamics of multiple outbreaks such as whooping cough in U.S. (2012), dengue outbreaks in India (2013) and China (2014). Our observations also suggest that, when news coverage is uniform, efficient modeling of temporal topic trends using time-series regression techniques can estimate disease case counts with increased precision before official reports by health organizations.In retrospective assessments, internet news reports have been shown to capture early reports of  Infectious diseases are a threat to public health and economic stability of many countries. Open source indicators  are published with a considerable delay of weeks, months or even a year. Therefore, traditional surveillance systems are not always effective at real-time monitoring of emerging public health threats. Unlike traditional surveillance data, informal digital sources, such as news media, blogs, and micro-blogging sites (Twitter) are typically available in (near) real-time. Proper mining of signals from these digital sources can effectively help in minimizing the time lag between an outbreak start and formal recognition of an outbreak, allowing for an accelerated response to public health threats. The gains in supplementing traditional surveillance using digital sources have been discussed in Nsoesie Our key contributions are as follows. (i) We introduce EpiNews, a generic temporal framework for analyzing disease-related news reports using a supervised topic model. The supervised topic model discovers multiple disease topics of interest and their associated temporal trends of prominence in news media. (ii) EpiNews captures trends in disease progression, such as periodicity, peaks and troughs via temporal trends of disease topics in news media. (iii) When news coverage is adequate, EpiNews also estimates disease incidence before official reports by health agencies using time-series regression models interposed over the temporal trends of disease topics.We validated our method against disease case count reports, as available from public health agencies in U.S., China, and India. Disease-related news articles were provided by HealthMap21In this section, we discuss the data sources used to analyze the infectious disease outbreaks. We first describe the case count reports collected from public health agencies and complete our discussion about the HealthMap data used in this study.For each country, we collected case count data corresponding to multiple diseases over a certain time period. In United States, Florida) after reverse geocoding. Each HealthMap article was passed through a series of preprocessing steps. For China, majority (87.94%) of the articles were published in either Traditional Chinese or Simplified Chinese. We translated the textual content of these articles to English for ease of analysis using Google translate (https://translate.google.com/). Because of the unavailability of ground truth for these articles, we couldn’t validate the performance of Google translate in this context. However, Google translate is one of the state-of-the-art commercial machine translations used today. Recent advances in deep learning and neural machine translation have made it a reliable tool for Chinese-to-English translation. For more details, see http://www.androidauthority.com/google-translate-machine-learning-chinese-718813/. Prior research2526et al.http://babelfish.yahoo.com/).Disease-related news articles were found to be indicative of infectious disease outbreaksword(w), location(l), timepoint(t)}:count. The second component, referred to as temporal topic modeling, is used to extract temporal topic trends from In this section, we describe in details the components of our proposed framework EpiNews. In cold, contagious, nausea, blood, food-borne, waterborne, sanitation) as well as specific-  disease related terms. In The first component of EpiNews deals with the preprocessing of HealthMap articles through a series of preprocessing steps, such as removal of non-textual elements, tokenization2831et al.w, l, t}:count, where count is the number of news articles mentioning the word w associated with the location l and time point t in the tuple. For this transformation, we assumed that for each country, each processed article consists of words from a vocabulary V, corresponds to a discretized time window t ∈ {1, 2, …, T} and is geotagged with a location l from a set of locations L in the country. For China, disease case counts were available on a monthly granularity and as such each time point t represents a period of 1 month. However, for diseases in U.S. and India, case counts were obtained on a weekly basis and as such time point t represents a period of 1 week or more specifically, epidemiological week (hereafter referred to as epi week). For example, the tuple , 2013-10-06):9 denotes that the word salmonella was mentioned in 9 articles referring to the state of Kansas in U.S. over the epi week extending from 6th October 2013 to 12th October 2013. For each country, let Nl represent the collection of tuples for each location l ∈ L and Nl until time point T. This transformed set Nl and Following Rekatsinas Nl can be treated as a three-dimensional matrix of size V × L × T where the dimensions are represented by words (size V), locations (size L) and time points (size T). Each element xw,l,t in w (w ∈ V) referring to location l (l ∈ L) over the time point t . We assume that each entry in a non-zero element xw,l,t of The second component of EpiNews deals with the topic and pattern discovery problem. The set K disease topics and decomposes ξ: A K × T matrix where each row represents a discrete probability distribution over the time points  for a specific topic z ∈ 1, 2, 3, …, K. Each row of ξ (ξz) represents the temporal topic trends or distribution for the disease topic z ∈ 1, 2, 3, …, K.ϕs: A K × S matrix where each row represents a discrete probability distribution over the set S of seed words for a specific topic z ∈ 1, 2, 3, …, K. ϕs is hereafter referred to as the seed topic distribution.ϕr: A K × V matrix where each row represents a discrete probability distribution over the set of regular words for a specific topic z ∈ 1, 2, 3, …, K. The set of regular words refers to all the words in vocabulary V including the seed words. ϕr is hereafter referred to as the regular topic distribution.θ: A L × K matrix where each row represents a discrete probability distribution over K topics for a specific location l ∈ L.The supervised topic model takes ξ, ϕs, ϕr and θ, see θ, ϕr, ϕs and ξ in the supervised topic model given input observed data For more details on et al.To apply the model in practice, seed words for each disease topic were extracted by examining the content of a subset of news articles mentioning the disease. Additionally, following similar techniques as in Chakraborty ξ). Let D be the disease of interest. Without loss of generality, let the zth disease topic corresponds to D. Furthermore, let SD,T denotes case counts of D and ξz,T denotes temporal trend value for zth disease topic at a time point T. In general, reports of case counts published by health organizations are delayed  available till T′ and temporal topic trends (ξz) available till T. In general, disease case counts have a publication delay of 1 time point (T′ = T − 1) and hence, estimating SD,T at T is equivalent to 1-step ahead estimation.The final component of EpiNews is concerned with estimation of disease case counts using relevant information such as past case counts and temporal topic trends (ξz). We used Elastic NetFor 1-step ahead case count estimation, we used a regularized version of autoregressive model with external input variables (ARX) where external input variables are represented by the temporal topic trends 43λ1 equals to 0, λ2 equals to 0, where, Internal component (p): This component is an autoregressive model that captures the signal embedded in past case counts and thus describes a delayed model. p indicates the order of autoregression. (ii) External component : This component can also be thought of as an autoregressive component over the temporal topic trends (ξz) where q is the number of time points to look back. The temporal topic trends are subjected to two additional transformations as follows. (a) Shift indicator (s): Often, the incidence of news reports is not concurrent with the incidence of diseases, as recorded in the case counts. EpiNews-ARNet incorporates this information by shifting the temporal topic trend value ξz,T by s steps. The shift can be positive (indicating a lagging trend), negative (indicating a leading trend) or zero (indicating a co-incident trend). (b) Rolling transformation (r): Disease case counts (SD) do not follow a strictly linear relationship with temporal topic trends (ξz). One of the simplest methods is to detrend the signals using difference of trend values instead of absolute values. However, our experiments showed that such transformations using a single time point often lead to unstable estimates. As such, we define a rolling transformation g over a window length r given below in There are broadly two components to p, q, r, s) parameters.Essentially, such transformations aim to capture the changes in trend values over a period and were found to be more indicative than absolute values. We ran a cross-validation step to find the optimal (ξz) as the external input variables. It is to be noted that the disease case counts (SD) and the temporal topic distribution (ξz) are typically at different numerical scales since values in a distribution range from 0 to 1. To improve numerical stability we converted the temporal topic distributions to estimated case counts using multinomial samplingD) for disease D, the corresponding temporal distribution ξz for zth topic was used as the multinomial distribution and the total number of case counts available till T′ < T at T (due to delay in reporting of case counts) was used as the number of samples to be drawn from the distribution. See Algorithm (1) for more details.We described EpiNews-ARNet using the temporal topic trends or distribution (ξ) extracted by the supervised topic model are able to capture disease dynamics - including seasonality, abrupt peaks and troughs. Finally, we evaluated the quality of case counts estimated by EpiNews-ARNet against the actual disease case counts.In this section, we present an empirical evaluation of our proposed framework EpiNews. We first evaluated the disease topics discovered by the supervised topic model. Next, we analyzed whether the temporal topic trends (ϕs) and regular topic distributions (ϕr). Considering space limitations, we present the analysis of ϕs and ϕr in ϕs and ϕr were extracted from HealthMap data spanning over the entire time period shown in z), we show the seed words and their corresponding probabilities (sorted in descending order) in the seed topic distribution zth disease topic. For example, seed words such as food, salmonella, product, fda, drug, contamination serve as informative prior words for the discovery of salmonellosis topic in U.S. since they have higher probabilities in the seed topic distribution  for each disease topic (z) in a specific country (ξz) were extracted from HealthMap data spanning over the entire time period shown in We also examined the temporal distribution or trends , we also showed the corresponding sampled case counts (ΞD) generated via multinomial sampling (see Algorithm (1)) from ξz for a disease D in Along with the temporal topic trends  and temporal topic trends  available up to t = T. In As official reports of case counts by health agencies are usually lagged by a single time point (week or month), reliable early estimates of disease incidence can facilitate the allocation of public health resources to enable effective control measures. Therefore, we aim to perform 1-step ahead estimation of disease case counts starting from a particular time point. For the purpose of experimental validation, we used historical HealthMap data over a certain time period as the static training set in a specific country (referred to as the static training period) and progressively utilized the remaining time points as the evaluation period over which we evaluated the case count estimates of EpiNews-ARNet. To estimate case counts at a particular time point ξz). However, in case of EpiNews-ARMAX, we used an autoregressive—moving-average model with external input variables  Both EpiNews-ARNet and EpiNews-ARMAX perform better overall with sampled case counts as external variables than temporal topic trends. (iv) For none of the {country, disease} combinations, Casecount-ARMA is able to achieve lowest NRMSE values indicating the significance of incorporating temporal topic trends or sampled case counts as external variables for estimating case counts.We evaluated the case count estimates of each method over the evaluation period by comparing them against the actual case counts using normalized root-mean-square error (NRMSE). In In this paper, we studied the problem of monitoring and estimating outbreaks of multiple infectious diseases using disease-related online news reports obtained from HealthMap. We introduced EpiNews, a novel and generic temporal framework that combines supervised temporal topic models with time-series regression techniques to monitor and estimate disease incidence. Experimental results demonstrate that EpiNews is able to capture the time varying incidence of multiple diseases via temporal topic trends. Our experiments also illustrate that EpiNews can estimate disease incidence 1-step ahead with increased accuracy using information from temporal topic trends.EpiNews uses online news reports as the sole data source to capture disease dynamics during outbreaks. Therefore, it is generic in the sense that it is not tailored to a particular disease or class of diseases. Moreover, the set of diseases selected for each country represent a diversity of transmission pathways as shown in E. coli infection in U.S., periodic outbreaks of H7N9 and HFMD in China, dengue outbreaks in India (2013) and China (2014). However, there are certain deviations where temporal topic trends could not monitor the trends in disease outbreaks properly. We posit that such deviations are a factor of multiple effects as follows. (i) Firstly, news media coverage during disease outbreaks is driven by interest. News coverage for certain diseases can be inconsistent over time. For salmonellosis and E. coli infection outbreaks in 2010, 2013 and 2014  are characterized by sharp peaks during the outbreak period see  and 4. Ep) in For dengue and HFMD in China, EpiNews-ARNet shows considerable improvement on 1-step ahead estimation of disease incidence when compared to the baselines, specifically Casecount-ARMA see . In ordeet al.et al.The effectiveness of online sources  to monitor and forecast the emergence and/or spread of diseases is an ongoing topic of debate, as evidenced by the community response to the study of Lazer How to cite this article: Ghosh, S. et al. Temporal Topic Modeling to Assess Associations between News Trends and Infectious Disease Outbreaks. Sci. Rep.7, 40841; doi: 10.1038/srep40841 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


To create a mathematical model to investigate the treatment impact and economic implications of introducing an antimicrobial resistance point-of-care test (AMR POCT) for gonorrhoea as a way of extending the life of current last-line treatments.Modelling study.England.Patients accessing sexual health services.Incremental impact of introducing a hypothetical AMR POCT that could detect susceptibility to previous first-line antibiotics, for example, ciprofloxacin or penicillin, so that patients are given more tailored treatment, compared with the current situation where all patients are given therapy with ceftriaxone and azithromycin. The hypothetical intervention was assessed using a mathematical model developed in Excel. The model included initial and follow-up attendances, loss to follow-up, use of standard or tailored treatment, time taken to treatment and the costs of testing and treatment.Number of doses of ceftriaxone saved, mean time to most appropriate treatment, mean number of visits per (infected) patient, number of patients lost to follow-up and total cost of testing.In the current situation, an estimated 33 431 ceftriaxone treatments are administered annually and 792 gonococcal infections remain untreated due to loss to follow-up. The use of an AMR POCT for ciprofloxacin could reduce these ceftriaxone treatments by 66%, and for an AMR POCT for penicillin by 79%. The mean time for patients receiving an antibiotic treatment is reduced by 2 days in scenarios including POCT and no positive patients remain untreated through eliminating loss to follow-up. Such POCTs are estimated to add £34 million to testing costs, but this does not take into account reductions in costs of repeat attendances and the reuse of older, cheaper antimicrobials.The introduction of AMR POCT could allow clinicians to discern between the majority of gonorrhoea-positive patients with strains that could be treated with older, previously abandoned first-line treatments, and those requiring our current last-line dual therapy. Such tests could extend the useful life of dual ceftriaxone and azithromycin therapy, thus pushing back the time when gonorrhoea may become untreatable. This study uses a simple framework to evaluate the potential impact of point-of-care tests to diagnose antimicrobial-resistant or antimicrobial-sensitive gonorrhoea infections.This study is parameterised with contemporary UK data on diagnoses, treatment and levels of antimicrobial resistance.This study uses a static model, so not possible to extrapolate future population effects.Neisseria gonorrhoeae, is the second most commonly diagnosed bacterial sexually transmitted infection (STI) in England. N gonorrhoeae has evolved resistance to all major drug classes and has been recognised as a bacterium of international concern by WHOIncreasing antimicrobial-resistant gonorrhoea represents a significant and urgent public health problem. Gonorrhoea, caused by Diagnoses have more than doubled from 16 839 in 2010 to 41 193 in 2015, mainly due to increased diagnoses in men who have sex with men (MSM), accounting for 70% of male infections in 2015, illustrated in In the UK, the Gonococcal Resistance to Antimicrobial Surveillance Program (GRASP) has performed sentinel antibiotic susceptibility testing of gonorrhoea since 2000.There are two main challenges to the management of gonorrhoea which contribute to the problem of resistance, illustrated in Strategies are required to extend the life of existing antimicrobials for the successful treatment of gonorrhoea. Most infections diagnosed in the UK are susceptible to cefixime, ciprofloxacin and even penicillin.We developed a decision tree model in Excel to consider the impact of a hypothetical new AMR POCT on testing, diagnosis and treatment of gonorrhoea in sexual health clinics in England  comparedThe model was based on an existing pathway model used to investigate the impact of introducing a dual POCT for gonorrhoea and chlamydia in a GUM setting,10.1136/bmjopen-2016-015447.supp1Hypothetical cohorts of patients were followed through the pathway . Individuals could either receive same day management or delayed management  under cuFull model parameters are provided in the online Current management: Clinicians have no knowledge of the resistance profile of gonorrhoea at the point of initial treatment and consequently all patients are treated with ceftriaxone and azithromycin. Some patients are managed on the same day, either due to symptoms and positive microscopy or as contacts of infected individuals, others wait for lab results, resulting in some unnecessary treatment and some delays to treatment or loss to follow-up .Simple POCT management: All patients tested and managed same day but all treated as if resistant to older antibiotics .assuming current ciprofloxacin resistance prevalenceassuming current penicillin resistance prevalence.AMR POCT management: All patients tested with AMR POCT for gonorrhoea that could identify infections that do not need to be treated with ceftriaxoneWe considered the following scenarios for each of the three patient groups .The primary outcomes were the number of doses of ceftriaxone saved and the mean time to appropriate treatment. In addition, we calculated the average number of visits per person and per infected person, the total cost of testing and the number of patients lost to follow-up. In each case, we compared the incremental benefit of an AMR POCT with current testing practice. Analyses were undertaken from the National Health Service perspective with costs measured in pounds sterling at 2014 prices.We modelled a snapshot of GUM attendance, gonorrhoea diagnosis and prevalence of resistance to ciprofloxacin and penicillin based on the situation in England, 2014The results for AMR POCT (strategies 3 and 4) and POCT (strategy 2) only differ by the choice of treatment regimen. If an AMR POCT for ciprofloxacin resistance were available (strategy 3a), we estimate its use could prevent 22 054 treatments of ceftriaxone annually (a 66% reduction) assuming the current levels of resistance to ciprofloxacin  could be achieved by using a simple POCT, as previously considered.Our model used recent published data on AMR levels, gonococcal incidence and current treatment and considered the impact of additional AMR POCT in distinct population groups, namely heterosexual men, MSM and females. The simplified model structure, which is available freely online, enables the parameters to be easily updated and the impact of different scenarios, in different settings, to be considered. We made the simplifying assumption that the cost of an AMR POCT would add £25 to the current tariff cost; however, in reality other current activities might be reduced or discontinued if an AMR POCT was available, such as testing, microscopy, culture and physical exams or reattendances, as well as reduced costs associated with reusing cheaper oral antibiotics. New DNA-based POCT technologies may be able to be combined to produce a multiplexed test, which may be more economically viable than the separate specific AMR tests we modelled here. Our cost estimates are therefore likely to be higher than in practice. New technologies are emerging that may be able to rapidly determine the bacterial response to a panel of potential antibiotics which would enable highly tailored therapy without the need to continuously monitor the efficacy of a test for resistance based on detecting DNA sequence, but for this preliminary exploration we selected a hypothetical AMR POCT test which could integrate with existing POCT technologies based on nucleic acid amplification.The model did not capture the indirect effects of reduced transmission to partners or progression to complications, such as pelvic inflammatory disease and epididymitis. It also did not consider the longer-term effects of changing treatment strategy on the evolution of drug resistance over time in gonorrhoea infections.et al previously showed that a dual chlamydia/gonorrhoea point-of-care NAAT diagnostic test pathway could be cost-neutral or cost-saving compared with existing methods even though the test kit itself is more expensive.Neisseria meningitidis.To our knowledge, no one has specifically addressed the question of the added value of an AMR POCT to discriminate between susceptible and resistant strains to guide initial treatment decisions for gonorrhoea. Others have considered in detail the relative benefits of POCTs, balancing the need for fast results against cost and test performance.Staphylococcus aureus,The main weakness of our study is that it did not address the population-level impact of the introduction of such tests, but only considered a static situation.The important next questions arising from this study are: how much time does the reduction in use of ceftriaxone buy in terms of slowing or preventing the emergence of clinically relevant gonorrhoea resistant to ceftriaxone and, second, what are the population-level benefits of improved gonorrhoea control?The major benefit of POCTs for gonorrhoea is increasing the proportion of patients treated appropriately on the same day as the test, which is likely to improve outcomes by reducing infectious duration, reducing loss to follow-up and potentially improving partner notification efficacy. A definitive diagnosis on the day of first presentation also prevents unnecessary treatment of those not infected with gonorrhoea. The main benefit of an AMR POCT that can discriminate between susceptible and resistant infections is in enabling the reintroduction of abandoned first-line therapies. Reducing the use of antibiotics, especially of last-line therapies, is a key aim of the UK national strategy on AMR. For heterosexual men and MSM, a relatively large proportion of infections are already treated on the same day as testing, based on epidemiological, clinical or microbiological evidence (microscopy). However, this proportion is lower for women due to the higher percentage of asymptomatic infections and from poorer sensitivity of detection of gonorrhoea in endocervical and urethral smears. Although new POCTs are likely to be more expensive than existing tests, this would to some extent be offset by the reduction in further attendances and in the ability to reuse older, cheaper drugs. Given the low prevalence of gonorrhoea even in high-risk GUM attendees, the cost of treatment and reattendances is small in comparison with the cost of attendances for testing and diagnosis. If a new discriminatory AMR POCT test were prohibitively expensive for routine use, a combination of a standard point-of-care NAAT  could be considered in conjunction with a more specialised gonorrhoea AMR test, although the time implications of this for patients and clinicians would have to be carefully considered.This estimation of the potential reduction in ceftriaxone use is the first step towards evaluating the long-term effects of such a reduction. Future research investigating how much the useful lifespan of ceftriaxone as a therapy for gonorrhoea is extended with particular reductions in ceftriaxone use would be valuable. In the context of the often slow and expensive new drug pipeline, there is also a question to be answered around the value placed on each additional year of ceftriaxone availability.


Southeast Asia, a vibrant region that has recently undergone unprecedented economic development, is regarded as a global hotspot for the emergence and spread of antimicrobial resistance (AMR). Understanding AMR in Southeast Asia is crucial for assessing how to control AMR on an international scale. Here we (i) describe the current AMR situation in Southeast Asia, (ii) explore the mechanisms that make Southeast Asia a focal region for the emergence of AMR, and (iii) propose ways in which Southeast Asia could contribute to a global solution. In 1928 Alexander Fleming initiated a medical revolution by discovering penicillin, which was developed as a medication and cured previously life-threatening infections. In 1945, Fleming foretold the risks of antimicrobial resistance (AMR): ‘The time may come when penicillin can be bought by anyone in the shops. Then there is the danger that the ignorant man may easily under dose himself and by exposing his microbes to non-lethal quantities of the drug make them resistant.’,Southeast Asia is a highly dynamic region characterized by rapid (yet uneven) economic development and has been proposed as an epicentre for emerging infectious diseases and AMR.AMR is a vast and complex problem, and rather than presenting a meta-analysis of AMR we aim to summarize and assess the key themes in regard to AMR in Southeast Asia. AMR in animals is inherently linked to AMR in humans, but will not be covered here as it has been recently been the subject of a review in Southeast Asia.Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, Streptococcus pneumoniae, non-typhoidal Salmonella, Shigella spp. and Neisseria gonorrhoeae).The WHO recently combined available worldwide data on AMR in seven common bacterial pathogens ; no data were available for Myanmar, Cambodia and Lao PDR.,,AMU in primary healthcare is an important driver of regional AMR,It has been estimated that veterinary AMU for food production will increase globally by 67% by 2030, and that one-third of this increase will be caused by a shift towards intensive livestock production in middle-income countries. Some of the largest increases in AMU (>200%) between 2010 and 2030 are predicted to arise in Myanmar and Indonesia.http://www.who.int/antimicrobial-resistance/national-action-plans/library/en/). Notably, there is no universally accepted definition of AMS, but the term refers to the strategies, policies, guidelines and tools used to promote and increase the appropriate use of antimicrobials.In 2011, the members of the WHO South East Asia Region  signed the Jaipur Declaration, in which they recognized the seriousness of the AMR problem and committed to act in order to safeguard the efficacy of antimicrobial drugs. A year after signing this antimicrobial stewardship (AMS) initiative, 42% of the countries surveyed in Asia had national AMS standards (the worldwide proportion is 52%),In Thailand, the Antimicrobial Smart Use (ASU) initiative was introduced in 2007 to promote the rational use of antimicrobials; this included a direct ban on the use of antimicrobials as growth promoters in animal food.The drivers that are likely to have an impact on AMR emergence and transmission in Southeast Asia are outlined below and displayed in Figure ,,,,,Southeast Asia has recently experienced tremendous socioeconomic development and population growth Table .6 The ra,Antimicrobials are amongst the most frequently prescribed drugs in human medicine worldwide.,,,,,A common theme across Southeast Asia is self-medication (without prescription): antimicrobials are available ‘over the counter’ in all of the selected countries, and self-treatment is generally cheaper and more convenient than visiting a healthcare facility.,,,The risk of inappropriate AMU through self-medication, unregulated access and irrational prescription is magnified by a sub-optimal understanding of antimicrobials and their use amongst the public, drug sellers and doctors in Southeast Asia.A lack of knowledge among the public stimulates irrational demand for antimicrobials and/or treatment non-adherence, which are two key drivers of AMR. A study from Malaysia found that over half of the patients discontinued their antimicrobial treatment once symptoms disappeared.In a recent survey of doctors from the Lao PDR,The aforementioned study in Lao PDR also confirmed the difficulty of prescribing an appropriate antimicrobial (recognized by 72.5% of the doctors).A further study conducted in Malaysia indicated that despite the existence of national antimicrobial guidelines, antimicrobial overprescribing is common in primary healthcare facilities, particularly in private clinics.Antimicrobial overuse, misuse and a lack of awareness are also common in animal production across the region; for a summary of available data on AMU in animal production in Southeast Asia readers are directed to a recent review.,,In Vietnam, antimicrobials for animal production are available over the counter, and veterinary pharmacists are a major source of advice for farmers.,,,,,Studies in Vietnam have found that antimicrobials are predominantly used to prevent rather than to treat infections in poultry and pig production systems;,,,,,In Southeast Asia, the use of antimicrobials is also normal in intensive aquaculture, as well as integrated agriculture–aquaculture systems where humans, vegetable/rice fields, livestock and aquaculture ponds are in close proximity.,,,E. coli isolated from the Matang mangrove estuaries in Malaysia were resistant to at least 1 of 15 antimicrobials tested, and 34% were resistant to three or more antimicrobial classes.AMR organisms and genetic elements encoding for resistance generated and maintained on farms may subsequently transmit to humans via direct contact with animals, consumption of foods of animal origin and/or their dissemination through animal waste.Access to antimicrobials has improved regionally; this has both positive (treating illnesses) and negative  consequences. It is worth noting that lack of timely access to good-quality antimicrobials remains a reality for many.It is generally difficult to ascertain the overall quality of antimicrobials available in Southeast Asia. According to the WHO, up to 10% of all drugs worldwide are counterfeit. Counterfeit drugs include drugs that are incorrectly dosed, contain the wrong active ingredient, contain no active ingredient, are of sub-standard quality or are wrongly packaged. Antimalarials and antibacterials are thought to be the most frequently counterfeited drugs.While it is difficult to quantify precisely the relative contribution of each driver to the development and spread of AMR throughout Southeast Asia, acting locally on defined drivers and finding region-specific solutions to mitigate AMR should be a priority. Members of the public, of the scientific and medical communities, local and national authorities as well as the international community can and should contribute to the solution. The United Nations has recognized the need to act and, in a resolution adopted in October 2016, reaffirmed that ‘the blueprint for tackling AMR is the World Health Organization Global Action Plan on AMR’.Considerable surveillance infrastructure, capacity and know-how already exist in Southeast Asia, but are highly variable across countries and sectors. In general, surveillance capacity occurs in many hospitals, but the animal production sector lags behind and knowledge gaps in the community remain. Collaboration and coordination between existing surveillance networks could develop synergies, avoid redundancy and promote standardized collection and analysis of samples. Sample collection and analysis (particularly determination of AMR profile) and the dissemination of results should follow the recommendations of WHO’s Global Antimicrobial Resistance Surveillance System (GLASS),Sample collection, analysis and result dissemination should remain compatible with the implementation of new technologies  for future research projects, such as elucidating gene flow between species and understanding the sinks and sources of AMR genes and organisms. PCR amplification of AMR genes and WGS are emerging into mainstream diagnostic laboratories in high- and middle-income countries. Whilst currently expensive and technically demanding, WGS of sentinel organisms within a Southeast Asia surveillance programme will become a key tool for understanding how AMR emerges and spreads across the region and beyond. In order to create a comprehensive picture of AMR, longitudinal sample collection for surveillance should extend to healthcare settings, the community, animal production and agriculture.,,The availability of a rapid diagnostic test that can differentiate between viral and bacterial infection in primary point-of-care facilities in Southeast Asia would not only improve clinical management of undifferentiated fever, but also help to optimize antimicrobial prescription, thereby limiting irrational AMU. Recent studies in Lao PDR, Cambodia, Thailand and Vietnam have demonstrated that rapid C-reactive protein (CRP) tests were able to differentiate between viral and bacterial aetiology in patients with non-specific fever, and were a comparatively inexpensive way to reduce inappropriate AMU in basic healthcare facilities without a laboratory.Diagnostic capacity  and/or rapid testing  should be improved in Southeast Asia. Capacity building, staff training and education would increase the prevalence of testing, thereby causing a shift from (mostly) empirical treatment towards evidence-based prescribing. In addition, there is a critical need to advertise broadly the importance of testing as a tool for rational diagnostics, and to understand the local, national and regional barriers to better diagnostic testing.,Studies suggest that education targeted at providers and consumers can contribute to reducing antimicrobial overuse.AMS is a key component of the fight against AMR. More AMS initiatives should be developed in Southeast Asia, but additional research is necessary to understand which elements of a stewardship programme are the most effective, as this remains a matter of considerable debate.AMU and AMR have a large behavioural component. Antimicrobials are seen as ‘wonder drugs’, but their true positive and negative potentials are misunderstood. Social research would generate invaluable insight for understanding behaviours underlying irrational prescribing by healthcare professionals, irrational demand by the public and irrational use by farmers. Therefore, social research is an essential priority for achieving the changes necessary to optimize AMU. Similarly, understanding why antimicrobial prescribing is often performed without diagnostic testing will begin to restrict the inclination to use antimicrobials indiscriminately. There is an urgent need to assess what combinations of cost, reagent accessibility, time and training are restricting better diagnostic approaches. Lastly, understanding the reasons for self-medication, treatment non-adherence and the reasons why antimicrobials are sold without prescription is crucial.Beyond morbidity, mortality and economic cost, qualitative research will be particularly important to estimate the wider societal cost of AMR, including the loss of efficacy of antimicrobials (and the effect of this on modern medicine), as well as deleterious effects of AMR on human capital, labour force, gross domestic product and economic growth.Finally, research will prove key to understanding and estimating the relative contribution of the different drivers of the development of AMR. Additional research will be necessary to design effective interventions and estimate their ability to curb AMR. This novel information will be vital to galvanize political will, convince regulators to modify policy, and persuade people to adapt their behaviour.AMU and AMR are increasing in Southeast Asia, driven by rapid intensification of food-production systems, loosely regulated access to antimicrobials, poor awareness with respect to antimicrobials , widespread irrational prescribing and self-medication, and an abundance of low-quality or counterfeit drugs. Combined with a high prevalence of infectious disease and weak diagnostic capacity, particularly in primary healthcare settings, Southeast Asia is a global hub for AMR, and contributes to the global spread of AMR as bacteria are readily transported to other parts of the world by international travellers, and by international trade of animals and goods. Across the region, there is generally good technical capacity to conduct laboratory testing and bacterial diagnostics. However, further work is needed to reach a consensus as to how to move forward with surveillance systems both for AMU and AMR in humans and in animal production systems.While concerted global action across multiple sectors is required to tackle the spread of AMR, we suggest that Southeast Asia should be a crucial location for generating a global solution by: (i) improving, coordinating and developing surveillance capacity to detect outbreaks and monitor trends; (ii) promoting the widespread use of diagnostic tests to support evidence-based prescribing; (iii) developing stewardship initiatives to raise awareness, educate and optimize AMU; and (iv) performing social research to understand the human drivers of irrational prescribing and irrational demand for antimicrobials. All these activities would contribute to optimize and reduce AMU, curb AMR spread and ultimately improve population health while preserving the efficacy of existing antimicrobial drugs.


Biology as a science is beginning its third century with new genomic foundations. The challenge of combining experimental evolution with genomics, is being met by a growing number of researchers, including the contributors to the current Frontier Topics Issue volume on the Genomics of Experimental Evolution, who in part discuss emerging new strategies. Today, experimental evolutionary genomic studies have the promise and perhaps the possibility of showing causal connections within the vast assemblages of genomic data generated.E. coli  explores how in cell cultures experimental gene variant RNA viruses of the Mononegavirales order have gene order-dependent varying fitnesses within both immune compromised and non-compromised prostate cancer cell lines. Using a data re-analysis of published studies in microbial evolution as well as simulations, the second topic perspective paper, Couce and Tenaillon consider the basis for the recurring observation in microbial evolution that fitness adaptation rates decline with adaptation across bacterial, viral, and yeast evolution. This powerful observation stemming from experimental evolution may inform us at a fundamental level about the nature of the multi-dimensional adaptive Euclidean space models of adaptation. In our third topic paper, Matos et al. in their opinion piece discuss how the application of genome-wide techniques in one of the oldest experimental evolution model organisms, Drosophila, help us understand the contours and tendencies of evolution. In their original research, the fourth topic paper by Graves et al. explores a very different technical territory by introducing the implications of microbial life adaptation to the emerging nanotechnology of heavy metals and their oxides, engineered nanoparticles (eNPs)—specifically how rapidly Escherichia coli adapted to silver eNPs. Their findings have implications for the use of heavy metal eNPs as antimicrobials on targeted and natural populations of microbes. In the fifth topic paper, O'Rourke et al. consider operon-based gain-of-function mutations leading to colony morphology variation in biofilm and planktonic growth within the Burkholderia cenocepacia pathogen complex, and the implications for human and agricultural thriving. Deatherage et al. in the final topic paper, lay out the challenges to identifying structural variants (SVs) in microbial genome evolutionary studies, which are more technically difficult to detect than nucleotide polymorphisms (NPs) and insertion deletions (indels). They discuss the theory, sensitivity, and simulations in applying their breseq analysis pipeline to the detection of SVs.Our contributors have helped address this emerging field's challenges. The first topic paper (The author confirms being the sole contributor of this work and approved it for publication.The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Radiologic severity may predict adverse outcomes after lower respiratory tract infection (LRI). However, few studies have quantified radiologic severity of LRIs. We sought to evaluate whether a semi-quantitative scoring tool, the Radiologic Severity Index (RSI), predicted mortality after parainfluenza virus (PIV)-associated LRI.We conducted a retrospective review of consecutively-enrolled adult patients with hematologic malignancy or hematopoietic stem cell transplantation and with PIV detected in nasal wash who subsequently developed radiologically-confirmed LRI. We measured RSI (range 0–72) in each chest radiograph during the first 30 days after LRI diagnosis. We used extended Cox proportional hazards models to identify factors associated with mortality after onset of LRI with all-cause mortality as our failure event.After adjustment for patient characteristics, each 1-point increase in RSI was associated with an increased hazard of death . Baseline RSI was not predictive of death, but both peak RSI and the change from baseline to peak RSI (delta-RSI) predicted mortality . A delta-RSI of ≥19.5 was 89% sensitive and 91% specific in predicting 30-day mortality.We conclude that the RSI offers precise, informative and reliable assessments of LRI severity. Progression of RSI predicts 30-day mortality after LRI, but baseline RSI does not. Our results were derived from a cohort of patients with PIV-associated LRI, but can be applied in validated in other populations of patients with LRI. The use of mortality as a primary endpoint in clinical trials of antimicrobials has been hotly debated due to difficulty in the determination of attribution , 2. The Systematic quantification of radiological severity could allow for a valid and reliable score that can serve as a biomarker of mortality. While thoracic computed tomography (CT) and chest X-ray (CXR) are important clinical tools for evaluating pneumonia , few stuTo explore the value of quantitative assessments of radiologic severity, we sought to determine whether radiologic progression of pneumonia as quantified by a systematic scoring tool, the Radiologic Severity Index (RSI) , would pWe conducted a retrospective chart review of a previously-established cohort of consecutively-enrolled adult patients with hematologic malignancies or those who had undergone hematopoietic stem cell transplantation (HSCT) who developed symptomatic PIV upper respiratory tract infection (URI) between October 1, 2002 and November 30, 2007 . PIV wasPIV-associated LRI was defined as the detection of PIV from nasal wash with symptoms of a clinical pneumonia syndrome  in accordance with guidelines . We chosAll clinical and oncologic data were collected prospectively from a database of viral infections used in infection control. Data collected included demographic information , underlying malignancy, cancer status , PIV detection in BAL (if performed), presence of BAL co-pathogens, cytotoxic chemotherapy, and administration of antiviral therapy with ribavirin and/or intravenous immunoglobulin therapy. For patients who underwent HSCT, we collected type of HSCT)  prior to LRI, and corticosteroid administration within 30 days of infection.We applied a semi-quantitative scoring tool for LRI severity initially developed in patients with H7N9 influenza LRI  to our sTwo senior thoracic radiologists  reviewed the RSI literature with two pulmonary investigators  and scored sample CXRs and CTs from a training set to establish a consistent rule-based system for scoring images. This training set consisted of patients without PIV-associated lower respiratory tract infection (LRI) but with a broad range of severity of infiltrates. After training, the radiologists independently scored all CXR and CT images performed within 60 days of LRI in sequential order, and we used the mean of the two RSI scores for analyses. The radiologists were blinded to the primary outcome of death and to each other’s scoring. We defined baseline RSI as the RSI score on the day of LRI onset. We defined peak RSI as the highest RSI score on CXR or CT after LRI onset and delta-RSI as the change from baseline to peak RSI.2 method to determine which model was best [2 estimates in Cox models are often low due to censored data and do not reflect the robustness of models; however, this technique can compare different models.We used chi-square or Fisher’s exact test to analyze categorical data. We used a Student’s t-test or Wilcoxon rank sum to analyze continuous data. We used an extended Cox proportional hazards model to analyze the factors associated with time to death after onset of LRI . We incowas best . GeneralWe used univariate logistic regression models with baseline, peak and delta-RSI as separate predictors of 30-day mortality. We used Bland-Altman plots , paired In our primary extended Cox model, we incorporated 241 measurable RSI-CXR scores. Non-survivors underwent more radiologic tests within the study period . In univariate Cox regression models, each 1-point increase in RSI-CXR measured as a time-varying covariate was associated with a 13% increase in the hazard for death . The proportional hazards assumption was met. AML was associated with mortality in our univariate Cox regression model . After adjusting for age, presence of AML and gender, only the progression of RSI remained in the multivariate model and predicted mortality  [In separate univariate Cox models using all RSI-CXR and RSI-CT scores for 14-, 30- or 60-day outcomes, RSI remained predictive of mortality . In secondary univariate analyses, extended Cox models using the number of involved zones predicted mortality , but RSI was better at predicting mortality than the number of involved zones (RSI: R = 0.04) .Using univariate logistic regression models, we found that longitudinal assessments of RSI were informative, whereas baseline assessments were not. Mean delta-RSI scores were highly predictive of mortality , as were peak RSI scores  . HoweverReliability between readers was excellent for all RSI scores (ICC: 0.99). We present the Radiologic Severity Index (RSI), a systematic radiologic scoring tool for LRI severity previously used in H1N1 influenza LRI , and tesFew studies have attempted to systematically quantify LRI severity radiologically. Grieser et al. developed a semi-quantitative CT-scoring tool for the acute respiratory distress syndrome after H1N1 influenza LRI in 23 patients . This toOur study of RSI adds to the existing literature by showing that progression of RSI on longitudinal assessment predicts mortality after PIV-associated LRI. Moreover, delta-RSI had excellent discriminatory ability to predict 30-day mortality, while baseline RSI scores were not predictive of mortality after PIV-associated LRI. This demonstrates that it is the change in radiologic severity that is informative. Commonly used clinical risk assessment tools for community acquired pneumonia (CAP), such as the CURB-65, were primarily developed to predict outcomes using baseline data and therefore cannot serve as outcome measures of pneumonia –32. In aOur findings build on those of other investigators. A substantial proportion of patients have persistent infiltrates weeks after LRI onset. Mittl et al. found that in 81 patients hospitalized with CAP, 33% had delayed resolution of pneumonia , but thiIn addition, precise quantification of radiographic results likely improved our ability to detect associations of radiologic severity with mortality. Others have found that multilobar pneumonia, as determined on qualitative chest radiograph interpretation, is associated with death –41, treaThe high reliability of RSI among readers improves precision in the assessment of radiologic severity. In studies using qualitative interpretations of radiologic severity, inter-observer variability between readers of chest radiographs may have impacted the ability to find an association between radiologic severity and mortality. For example, El Solh et al found that when determining whether chest radiographs had consolidation, interstitial infiltrates, or a mixed pattern, the kappa statistic between two radiologists was 0.6.  Taylor eOthers have highlighted the discrepancy between CXR and CT when diagnosing LRI, with discordance between imaging modalities in the diagnosis of LRI in 27–48% of patients , 44, 45.Not all patients in this study had proven PIV LRI, which is defined as detection of PIV from a lower respiratory tract sample and has been well established as a criterion , 47, 48.Our study must be interpreted in the context of certain limitations. Since we tested RSI only in a cohort of patients with PIV-associated LRI, further validation is necessary in other cohorts with LRI such as CAP or RSV-associated LRI. The study was retrospective in nature, and measurements were not obtained systematically. Measurement bias may exist because sicker patients were likely to have more assessments, and patients who died may not have had radiologic assessment at peak severity. However, the lack of follow-up studies in healthier patients would likely bias our results towards the null hypothesis, since we do not capture resolution of pneumonia. Furthermore, the lack of radiologic studies in patients who died before follow-up studies could be obtained would also bias our results towards the null hypothesis, because we would miss the progression of severe pneumonia. Our method of imputation ensured that our model likely captured progression of clinical disease, since radiology is typically performed due to changes in clinical status and not performed when a patient’s clinical status is stable . RadioloIn summary, we show that progression of radiologic infiltrates as assessed by the RSI is predictive of mortality in a high-risk cohort of patients with LRI. Longitudinal assessments of RSI were predictive of mortality, while baseline assessments were not. RSI was superior to previous methods to quantify LRI severity due to improved precision and reliability. Substantial discrepancies exist between CT and CXR assessments of LRI severity. RSI was tested in a cohort of patients with PIV-associated LRI, and future studies are needed to validate RSI in other LRI populations and to correlate RSI with other important outcomes such as health care utilization and cost. RSI is a promising tool that offers granular, reliable, precise and informative assessments of LRI severity and is well suited for consideration as part of a composite endpoint of clinical treatment failure in therapeutic trials of pneumonia.S1 DiscussionThis section offers a brief discussion on the utility of RSI depending upon a study population’s baseline risk of death and based on the type of statistical analysis employed.(DOCX)Click here for additional data file.S2 DiscussionThis section offers a brief discussion on pitfalls of data sampling in retrospective and prospective cohort studies, with suggestions on optimal data sampling techniques for potential prospective studies using RSI as a biomarker of radiological pneumonia severity.(DOCX)Click here for additional data file.S1 Dataset(XLS)Click here for additional data file.