Cellular heterogeneity in red and melanized focal muscle changes in farmed Atlantic salmon (Salmo salar) visualized by spatial transcriptomics.

Spatial transcriptomics is a technique that provides insight into gene expression profiles in tissue sections while retaining structural information. We have employed this method to study the pathological conditions related to red and melanized focal changes in farmed Atlantic salmon (Salmo salar). Our findings support a model where similar molecular mechanisms are involved in both red and melanized filet discolorations and genes associated with several relevant pathways show distinct expression patterns in both sample types. Interestingly, there appears to be significant cellular heterogeneity in the foci investigated when looking at gene expression patterns. Some of the genes that show differential spatial expression are involved in cellular processes such as hypoxia and immune responses, providing new insight into the nature of muscle melanization in Atlantic salmon.

Salmon gill poxvirus, a recently characterized infectious agent of multifactorial gill disease in freshwater- and seawater-reared Atlantic salmon.

Gill diseases cause considerable losses in Norwegian salmon farming. In 2015, we characterized salmon gill poxvirus (SGPV) and associated gill disease. Using newly developed diagnostic tools, we show here that SGPV infection is more widely distributed than previously assumed. We present seven cases of complex gill disease in Atlantic salmon farmed in seawater and freshwater from different parts of Norway. Apoptosis, the hallmark of acute SGPV infection, was not easily observed in these cases, and qPCR analysis was critical for identification of the presence of SGPV. Several other agents including Costia-like parasites, gill amoebas, Saprolegnia spp. and bacteria were observed. The studied populations experienced significant mortalities, which increased to extreme levels when severe SGPV infections coincided with smoltification. SGPV infection appears to affect the smoltification process directly by affecting the gills and chloride cells in particular. SGPV may be considered a primary pathogen as it was often found prior to identification of complex gill disease. It is hypothesized that SGPV-induced gill damage may impair innate immunity and allow invasion of secondary invaders. The distinct possibility that SGPV has been widely overlooked as a primary pathogen calls for extended use of SGPV qPCR in Atlantic salmon gill health management.

Health-related quality of life after stroke a comprehensive review.

BACKGROUND AND PURPOSE: We performed a comprehensive review of all quality-of-life (QOL) estimates for stroke appearing in the peer-reviewed literature between 1985 and 2000. We examine variation in QOL weights and the rigor of methods used to assess QOL and discuss the implications for cost-utility assessment and resource allocation decisions. METHODS: Through a systematic search, we identified 67 articles that met our inclusion criteria. A team of trained researchers read each article and followed detailed guidelines to extract QOL weights and other parameters. This effort yielded 161 QOL estimates for stroke-related health states. All estimates were measured on a 0 to 1 scale, with 0 representing the worst outcome and 1 representing the best. RESULTS: QOL estimates range from -0.02 to 0.71 (n=67) for major stroke, from 0.12 to 0.81 (n=14) for moderate stroke, from 0.45 to 0.92 (n=38) for minor stroke, and from 0.29 to 0.903 (n=42) for general stroke. Although QOL should decrease with severity, there were many instances in which the QOL for major stroke as reported by one study exceeded the QOL for moderate stroke as reported by another. The same reversal was found for moderate and minor stroke, and it occurred even when both authors used similar assessment methods and subject populations. Authors of cost-utility and decision analyses rarely base their choice of QOL weights on their own primary data (19%). When obtaining weights from secondary sources, some authors (23%) chose QOL weights for a severity of stroke that did not match the severity for which they sought data. CONCLUSIONS: QOL estimates for stroke vary greatly and are not always estimated in sound fashion. This impedes the comparability and quality of the cost-effectiveness studies that use these QOL weights and hampers good resource allocation decisions.

Classification and identification of Pfiesteria and Pfiesteria-like species.

Dinoflagellates can be classified both botanically and zoologically; however, they are typically put in the botanical division Pyrrhophyta. As a group they appear most related to the protistan ciliates and apicomplexans at the ultrastructure level. Within the Pyrrhophyta are both unarmored and armored forms of the dominant, motile flagellated stage. Unarmored dinoflagellates do not have thecal or wall plates arranged in specific series, whereas armored species have plates that vary in thickness but are specific in number and arrangement. In armored dinoflagellates, the plate pattern and tabulation is a diagnostic character at the family, subfamily, and even genus levels. In most cases, the molecular characterization of dinoflagellates confirms the taxonomy on the basis of external morphology; this has been demonstrated for several groups. Together, both genetic and morphological criteria are becoming increasingly important for the characterization, separation, and identification of dinoflagellates species. Pfiesteria and Pfiesteria-like species are thinly armored forms with motile dinospore stages characterized by their distinct plate formulae. Pfiesteria piscicida is the best-known member of the genus; however, there is at least one other species. Other genetically and morphologically related genera, now grouped under the common names of "Lucy," "Shepherd's crook," and cryptoperidiniopsoid, are being studied and described in separate works. All these other heterotrophic dinoflagellate groups, many of which are thought to be benign, co-occur in estuarine waters where Pfiesteria has been found.

Use of molecular probes to assess geographic distribution of Pfiesteria species.

We have developed multiple polymerase chain reaction (PCR)-based methods for the detection of Pfiesteria sp. in cultures and environmental samples. More than 2,100 water and sediment samples from estuarine sites of the U.S. Atlantic and gulf coasts were assayed for the presence of Pfiesteria piscicida Steidinger & Burkholder and Pfiesteria shumwayae Glasgow & Burkholder by PCR probing of extracted DNA. Positive results were found in about 3% of samples derived from routine monitoring of coastal waters and about 8% of sediments. The geographic range of both species was the same, ranging from New York to Texas. Pfiesteria spp. are likely common and generally benign inhabitants of coastal areas, but their presence maintains a potential for fish and human health problems.

Testing for the BRCA1 and BRCA2 breast-ovarian cancer susceptibility genes: a decision analysis.

OBJECTIVE: The authors developed a Markov decision model to evaluate the health implications of testing for mutations in the BRCA1 and BRCA2 breast-ovarian cancer susceptibility genes. Prophylactic measures considered included various combinations of immediate and delayed bilateral mastectomy and oophorectomy or taking no action. METHODS: The model incorporated the likelihood of developing breast and/or ovarian cancer, survival, and quality of life. Parameter values were taken from public databases, the published literature, and a survey of cancer experts. Outcomes considered were additional life expectancy and quality-adjusted life years (QALYs). Results are reported for 30-year-old cancer-free women at various levels of hereditary risk. RESULTS AND CONCLUSIONS: The vast majority of women will not benefit from testing because their pre-test risks are low and surgical prophylaxis is undesirable. However, women who have family histories of early breast and/or ovarian cancer may gain up to 2 QALYs by allowing genetic testing to inform their decisions.

Oregon's Medicaid ranking and cost-effectiveness: is there any relationship?

The authors examine whether Oregon's 1990, 1991, 1992, and 1993 prioritized lists were ranked in a manner consistent with cost-effectiveness. Two sets of cost-effectiveness data are used: data from economic analyses and Oregon's own cost-effectiveness data. Comparing the ranks of Oregon's lists with the ranks of cost-effectiveness estimates from the literature reveals Spearman correlations of -0.08 for the 1990 list, +0.39 for the 1991 list, +0.25 for the 1992 list, and +0.24 for the 1993 list. Comparing Oregon's lists with Oregon's own cost-effectiveness data reveals rank correlations of +0.99 for the 1990 list, +0.06 for the 1991 list, -0.05 for the 1992 list, and -0.03 for the 1993 list. Thus, there appear to be essentially no relationship between the 1990 list and cost-effectiveness estimates from the economic literature and modest positive relationships between the 1991-93 lists and the literature. In addition, there is virtually no relationship between the 1991-93 lists and Oregon's own cost-effectiveness data. Further, the correlations are very different from +1.0, suggesting that other factors are at play. For example, the 1993 list that is currently being implemented was ranked primarily by improvement in five-year survival and human judgment, not cost-effectiveness.

Francisella philomiragia subsp. noatunensis subsp. nov., isolated from farmed Atlantic cod (Gadus morhua L.).

Seven bacterial isolates from farmed Atlantic cod displaying chronic granulomatous disease were characterized by phenotypic and molecular taxonomic methods. The isolates were Gram-negative, facultatively intracellular, non-motile, strictly aerobic coccobacilli which produced H(2)S from cysteine-supplemented media and are therefore phenotypically consistent with members of the genus Francisella. Comparison of 16S rRNA gene sequences and six partial housekeeping gene sequences (groEL, shdA, rpoB, rpoA, pgm and atpA) confirmed the organism as a member of the genus Francisella, with Francisella philomiragia as its closest relative (99.3 % 16S rRNA gene sequence similarity, 92.2-99.0 % housekeeping gene sequence similarity). Despite the close relationship with F. philomiragia, isolates from Atlantic cod could be readily distinguished phenotypically and genetically from F. philomiragia ATCC 25015(T). DNA-DNA hybridization studies revealed a mean reassociation value of 68 %. Thus, on the basis of phenotypic and molecular genetic evidence, we propose that the strains isolated from Atlantic cod should be recognized as Francisella philomiragia subsp. noatunensis subsp. nov. with the type strain 2005/50/F292-6C(T) (=NCIMB 14265(T)=LMG 23800(T)). Francisella philomiragia ATCC 25015(T) (=DSM 735(T)) is reclassified as Francisella philomiragia subsp. philomiragia subsp. nov.

Detection of oncogenic mutations in the EGFR gene in lung adenocarcinoma with differential sensitivity to EGFR tyrosine kinase inhibitors.

The complete sequencing of the human genome and the development of molecularly targeted cancer therapy have promoted efforts to identify systematically the genetic alterations in human cancer. By high-throughput sequencing of tyrosine kinase genes in human non-small-cell lung cancer, we identified somatic mutations in the kinase domain of the epidermal growth factor receptor tyrosine kinase gene (EGFR) that are correlated with clinical response to EGFR tyrosine kinase inhibitors (TKIs). We have shown that these mutant forms of EGFR induce oncogenic transformation in different cellular systems. Cells whose growth depends on EGFR with mutations in exons 19 and 21 are sensitive to EGFR-TKIs, whereas cells expressing insertion mutations in exon 20 or the T790M point mutant, found in tumor biopsies from patients that relapsed after an initial response to EGFR-TKIs, are resistant. Furthermore, by applying a novel, massively parallel sequencing technology, we have shown that clinically relevant oncogene mutations can be detected in clinical specimens with very low tumor content, thereby enabling optimal patient selection for mutation-directed therapy. In summary, by applying high-throughput genomic resequencing, we have identified a novel therapeutic target, mutant EGFR, in lung cancer and evaluated its role in predicting response to targeted therapy.

One thousand health-related quality-of-life estimates.

OBJECTIVE: Analysts performing cost-effectiveness analyses often do not have the resources to gather original quality-of-life (QOL) weights. Furthermore, variability in QOL for the same health state hampers the comparability of cost-effectiveness analyses. For these reasons, opinion leaders such as the Panel on Cost-Effectiveness in Health and Medicine have called for a national repository of QOL weights. Some authors have responded to the call by performing large primary studies of QOL. We take a different approach, amassing existing data with the hope that it will be combined responsibly in meta-analytic fashion. Toward the goal of developing a national repository of QOL weights to aid cost-effectiveness analysts, 1,000 health-related QOL estimates were gathered from publicly available source documents. METHODS: To identify documents, we searched databases and reviewed the bibliographies of articles, books, and government reports. From each document, we extracted information on the health state, QOL weight, assessment method, respondents, and upper and lower bounds of the QOL scale. Detailed guidelines were followed to ensure consistency in data extraction. RESULTS: We identified 154 documents yielding 1,000 original QOL weights. There was considerable variation in the weights assessed by different authors for the same health state. Methods also varied: 51% of authors used direct elicitation (standard gamble, time tradeoff, or rating scale), 32% estimated QOL based on their own expertise or that of others, and 17% used health status instruments. CONCLUSIONS: This comprehensive review of QOL data should lead to more consistent use of QOL weights and thus more comparable cost-effectiveness analyses.

The link between smoking and impotence: two decades of evidence.

BACKGROUND: Anti-tobacco advertisements now feature the risk of impotence as a reason to avoid or cease tobacco use. The scientific evidence, however, is mixed. To shed light on the controversy, we compiled 2 decades of published data on the link between smoking and impotence. METHODS: We searched MEDLINE from 1980 to the present for studies that reported smoking prevalence among impotent male subjects. For each study we recorded the age of subjects, their state of residence, the percentage that were current smokers, and the time period over which study data were collected. For comparison purposes, we estimated age-, state-, and year-specific smoking rates in the general male population using the Behavioral Risk Factors Surveillance System. We performed a meta-analysis using a random effects model. RESULTS: Among the 1008 journal articles examined, we identified 19 studies that reported the smoking habits of 3819 impotent men. Of these 19 studies, 16 indicated a smoking prevalence exceeding that of the general population. The 6 largest studies all revealed a higher prevalence of smoking among impotent men. Meta-analysis reveals that 40% of impotent men were current smokers compared with 28% of men in the general population. CONCLUSIONSBased on almost 2 decades of evidence, tobacco use is an important risk factor for impotence. Anti-tobacco advertisements featuring impotence as a reason to avoid or cease tobacco use are well grounded in scientific fact.

Public health impact of changes in smoking behavior: results from the Tobacco Policy Model.

OBJECTIVES: The relative magnitude of the public health gains from preventing smoking initiation versus encouraging cessation or avoiding relapse in different ages and genders is estimated and compared. METHODS: Health gains are defined as the predicted increase in Quality-Adjusted Life-Years (QALYs) to the US population during a century. To estimate QALYs, we developed the Tobacco Policy Model. The model simulates a 10% reduction in the annual probability of initiation versus a 10% increase in cessation versus a 10% reduction in relapse in males and females in six age groups: 10 to 19, 20 to 29, 30 to 39, 40 to 49, 50 to 59 and 60 to 69. RESULTS: Among youth and young adults, reducing initiation yields far more QALYs than encouraging cessation or averting relapse. In middle-aged adults, cessation yields the most QALYs, followed by averting relapse and reducing initiation. In the oldest age group, averting relapse yields the most QALYs followed by cessation and reducing initiation. In general, increasing cessation and reducing relapse is more beneficial in males than in females whereas reducing initiation is more beneficial in females. CONCLUSIONS: The relative value of preventing initiation, encouraging cessation, and averting relapse differs by age and gender. Reducing initiation in youth is likely to offer the largest public health impact during the next century.

The cost-effectiveness of intensive national school-based anti-tobacco education: results from the tobacco policy model.

OBJECTIVES: School-based anti-tobacco education using the "social influences" model is known to reduce smoking among youth by 5-56%. Program effectiveness, however, dissipates in 1-4 years. Consequently, opinion leaders have questioned whether a more intensive national educational effort would be economically efficient. To address this question, we evaluated the cost-effectiveness of enhanced nationwide school-based anti-tobacco education relative to the status quo. METHODS: To estimate cost-effectiveness, we created the Tobacco Policy Model, a system dynamics computer simulation model. The model relies on secondary data and is designed to calculate the expected costs and public health gains of any tobacco policy or intervention over any time frame. RESULTS: Over 50 years, cost-effectiveness is estimated to lie between $4,900 and $340,000 per quality-adjusted life-year (QALY), depending on the degree and longevity of program effectiveness. Assuming a 30% effectiveness that dissipates in 4 years, cost-effectiveness is $20,000/QALY. Sensitivity analysis reveals that cost-effectiveness varies with cost, survival, and quality-of-life estimates but cost-effectiveness ratios generally remain favorable. CONCLUSIONS: Although not cost saving, a much more intensive school-based anti-tobacco educational effort would be an economically efficient investment for the nation.

Minimizing the time to search visual displays: the role of highlighting.

Office computer users frequently need to search long lists of options for a particular target. Highlighting a subset of the options can shorten the average display search time. The goal of the current research was to identify the subset of options to highlight that minimizes users' average search time for any given display. To begin, a mathematical model of the visual search process was constructed which could be used to predict the average search time for a target in a display with or without highlighted options. The predictions of this model were then compared with the results of a visual search experiment that varied the type of highlighting (blocked, random, or no highlighting), the number of highlighted options, and the probability that a target was highlighted. Overall the predictions of the model were consistent with the results from the experiment. Finally, procedures based on the model were developed which could be used to find the optimal subset of options to highlight for a given display--that is, the subset of highlighted options that minimizes users' search times.

Genetic polymorphism in Gymnodinium galatheanum chloroplast DNA sequences and development of a molecular detection assay.

Nuclear and chloroplast-encoded small subunit ribosomal DNA sequences were obtained from several strains of the toxic dinoflagellate Gymnodinium galatheanum. Phylogenetic analyses and comparison of sequences indicate that the chloroplast sequences show a higher degree of sequence divergence than the nuclear homologue. The chloroplast sequences were chosen as targets for the development of a 5'--3' exonuclease assay for detection of the organism. The assay has a very high degree of specificity and has been used to screen environmental water samples from a fish farm where the presence of this dinoflagellate species has previously been associated with fish kills. Various hypotheses for the derived nature of the chloroplast sequences are discussed, as well as what is known about the toxicity of the species.

Development of real-time PCR assays for rapid detection of Pfiesteria piscicida and related dinoflagellates.

Pfiesteria complex species are heterotrophic and mixotrophic dinoflagellates that have been recognized as harmful algal bloom species associated with adverse fish and human health effects along the East Coast of North America, particularly in its largest (Chesapeake Bay in Maryland) and second largest (Albermarle-Pamlico Sound in North Carolina) estuaries. In response to impacts on human health and the economy, monitoring programs to detect the organism have been implemented in affected areas. However, until recently, specific identification of the two toxic species known thus far, Pfiesteria piscicida and P. shumwayae (sp. nov.), required scanning electron microscopy (SEM). SEM is a labor-intensive process in which a small number of cells can be analyzed, posing limitations when the method is applied to environmental estuarine water samples. To overcome these problems, we developed a real-time PCR-based assay that permits rapid and specific identification of these organisms in culture and heterogeneous environmental water samples. Various factors likely to be encountered when assessing environmental samples were addressed, and assay specificity was validated through screening of a comprehensive panel of cultures, including the two recognized Pfiesteria species, morphologically similar species, and a wide range of other estuarine dinoflagellates. Assay sensitivity and sample stability were established for both unpreserved and fixative (acidic Lugol's solution)-preserved samples. The effects of background DNA on organism detection and enumeration were also explored, and based on these results, we conclude that the assay may be utilized to derive quantitative data. This real-time PCR-based method will be useful for many other applications, including adaptation for field-based technology.