PUBLISHED VERSION

Background: The duck-billed platypus (Ornithorhynchus anatinus) belongs to the mammalian subclass Prototheria, which diverged from the Theria line early in mammalian evolution. The platypus genome sequence provides a unique opportunity to illuminate some aspects of the biology and evolution of these animals. Results: We show that several genes implicated in food digestion in the stomach have been deleted or inactivated in platypus. Comparison with other vertebrate genomes revealed that the main genes implicated in the formation and activity of gastric juice have been lost in platypus. These include the aspartyl proteases pepsinogen A and pepsinogens B/C, the hydrochloric acid secretion stimulatory hormone gastrin, and the α subunit of the gastric H+/K+-ATPase. Other genes implicated in gastric functions, such as the β subunit of the H+/K+-ATPase and the aspartyl protease cathepsin E, have been inactivated because of the acquisition of loss-of-function mutations. All of these genes are highly conserved in vertebrates, reflecting a unique pattern of evolution in the platypus genome not previously seen in other mammalian genomes. Conclusion: The observed loss of genes involved in gastric functions might be responsible for the anatomical and physiological differences in gastrointestinal tract between monotremes and other vertebrates, including small size, lack of glands, and high pH of the monotreme stomach. This study contributes to a better understanding of the mechanisms that underlie the evolution of the platypus genome, might extend the less-is-more evolutionary model to monotremes, and provides novel insights into the importance of gene loss events during mammalian evolution. Published: 15 May 2008 Genome Biology 2008, 9:R81 (doi:10.1186/gb-2008-9-5-r81) Received: 16 December 2007 Revised: 4 April 2008 Accepted: 15 May 2008 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2008/9/5/R81 Genome Biology 2008, 9:R81 http://genomebiology.com/2008/9/5/R81 Genome Biology 2008, Volume 9, Issue 5, Article R81 Ordoñez et al. R81.2 Background A major goal in the sequencing of different genomes is to identify the genetic changes that are responsible for the physiological differences between these organisms. In this regard, the comparison between human and rodent genomes has identified an expansion in rodents of genes that are implicated in fertilization and sperm maturation, host defense, odor perception, or detoxification [1-3], confirming at the genetic level the physiological differences in these processes between humans and rodents. Additionally, the development of specific biological processes during evolution, for example the production of milk in mammals, has been accompanied by the appearance of novel genes that are implicated in these novel functions, such as casein and α-lactalbumin [4]. Therefore, it appears that the acquisition of novel physiological functions during vertebrate evolution has been driven by the generation of novel genes adapted to these newer functions. However, although gene gains constitute an intuitive mechanism for the development of novel biological functions, gene losses have also been important during evolution, both quantitatively and qualitatively [5-9]. The recent availability of numerous vertebrate genomes has opened the possibility to perform large-scale evolutionary analysis in order to identify differential genes responsible for the specific differences in particular biological processes. The duck-billed platypus (Ornithorhynchus anatinus) represents a valuable resource for unraveling the molecular mechanisms that have been active during mammalian evolution, due both to its phylogenetic position and to the presence of unique biological characteristics [10]. Together with the echidnas, platypus constitutes the Monotremata subclass (prototherians); this is one of the two subclasses into which mammals are divided, together with therians, which are further subdivided into marsupials (metatherians) and placental mammals (eutherians) [11]. The appearance of mammal-specific characteristics such as homeothermy, presence of fur, and mammary glands makes this organism a key element in elucidating the genetic factors that are implicated in the appearance of these biological functions. Nevertheless, since the last mammalian common ancestor, more than 166 million years ago (MYA) [12,13], other characteristics have emerged, such as the presence of venom glands or electroreception, and some vertebrate characteristics have been lost, resulting in the absence of adult teeth or a functional stomach [14,15]. In this work, we show that there has been a selective deletion and inactivation in the platypus genome of several genes that are implicated in the activity of the stomach, including all genes encoding pepsin proteases, which are involved in the initial digestion of proteins in the acidic pH of the stomach, as well as the genes required for the secretion of acid in this organ (Figure 1). The loss and inactivation of these genes provide a molecular basis for understanding the mechanisms that are responsible for the absence in platypus of a functional stomach, and expand our knowledge of the evolution of mam-


Introduction
Primary prevention of heart disease and health promotion has been in focus for decades. Health care in the primary and secondary care sector has evolved to strengthen the citizen's opportunities to engage in prevention and health promotion. 1 Half of patients with hypertension are unaware of their condition. [2][3][4][5][6][7] The prevalence of hypertension in the adult Danish population is estimated to be 22.3%. The prevalence spans from 1% in those aged 20-29 years to 69% among those aged 80-89 years. 8 Among Danish patients with known hypertension, only 40%-50% are treated to the guideline recommended blood pressure of less than 140/90 mm Hg. [7][8][9] The situation is far worse in less-developed countries. For patients with coronary heart disease or stroke, the use of blood pressure-lowering drugs decreases from 69% in high-income countries to 16% in low-income countries. 10 There is a genetic

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Qvist et al predisposition for hypertension, and men develop hypertension earlier than women. Obesity, 11,12 high salt intake, 13,14 physical inactivity, 15,16 and excess alcohol consumption 17 increase the risk of hypertension. Proven interventions to prevent the development of hypertension include reduction of weight and alcohol intake; increase in leisure time exercise; healthy diet, with low saturated fat; and reduced salt intake. 11,12,[15][16][17][18][19][20] Health campaigns have been addressed to entire populations, but a systematic patient education program for hypertensive patients does not exist. In this context, it is surprising that there is little information about elderly people's knowledge and awareness of their blood pressure.
In this study we examined self-reported hypertension knowledge in a population-based random sample of people aged 60-74 years. We hypothesized that knowledge and awareness about blood pressure would be better among individuals with self-reported hypertension compared with subjects without self-reported hypertension. Individuals with "self-reported hypertension" were defined as those who reported use of antihypertensive drugs.

Study population and methods source population
The source population consisted of 6,803 men and 6,923 women aged 60-74 years who were living in the municipality of Silkeborg, Denmark. In the year 2010, the population of the municipality of Silkeborg comprised 42,396 inhabitants. The mean income per inhabitant in the municipality of Silkeborg was DKK 282,000 vs DKK 283,000 in the general Danish population. The employment rate in the municipality of Silkeborg was 74.9% vs 72.2% in the general Danish population. The sample population was randomly drawn from the Central Person Registry of Denmark by the research department at the Danish Health and Medicines Authority. The age interval of 60-74 years was chosen because of a high prevalence of hypertension in this age group. To increase the internal validity of our study, we did not include subjects older than 74 years because we anticipated a low response rate among people older than 75 years.
The study was approved by the Danish Data Protection Agency. There was no requirement for approval from an ethical committee because the responses from the participants were anonymous.

The questionnaire
The questionnaire was developed by the investigator in cooperation with nurses and doctors working in cardiology wards, dialysis units, outpatient clinics, and general practices, based on relevant and useful questions that emerged from the literature 21 and clinical experience.
The questionnaire asked for information on sex, age, civil status, number of children, and education. There were additional questions about height, weight, cholesterol, and hypertension risk factors. The questions on the knowledge of hypertension symptoms were in multiple-choice format and focused on the following areas: a) factors that influence blood pressure, b) the symptoms of hypertension, c) the consequences of hypertension, and d) awareness of one's own blood pressure.
The questionnaire was tested in a pilot study of 31 subjects aged 60-74 years and adjusted accordingly. The questionnaire was then mailed to the study population, together with a letter explaining the purpose of the study. We did not validate the questionnaire by repeating it because we assumed participants might seek information between examinations.

statistics
We used the two-sample Student's t-test, chi-square test, and Wilcoxon rank sum test, where appropriate. Odds ratios (ORs) were calculated with 95% confidence intervals (CIs). We used STATA 9 statistical software (StataCorp, College Station, TX, USA).

Results
The sample population consisted of 493 men and 507 women. Of those, 727 subjects responded. Seven of the returned questionnaires were excluded because they were received after the response deadline. Two respondents were excluded due to missing data. Thus, the final study population (ie, the responders) consisted of 718 subjects, which corresponds to a 72% response rate. Table 1 shows characteristics of the responders and nonresponders. These two groups were comparable with respect to sex and age, but fewer of those living alone responded (OR=0.38 [95% CI: 0.28-0.52]). Table 2 shows the demographic, social, anthropometric, and self-assessed general health characteristics, in participants with and without self-reported hypertension. A total of 307 participants (43%) reported having hypertension, and 411 (57%) reported not having hypertension. Those with self-reported hypertension were more often men, were older, were overweight or obese, and had a lower educational level. Those with self-reported hypertension more often had a family history of hypertension, increased cholesterol levels, and a lower self-estimated general health condition than did those with no self-reported hypertension.

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Knowledge and awareness about blood pressure and hypertension   In general, knowledge about the lifestyle factors that may influence blood pressure was good, irrespective of the self-reported hypertension status (Table 3). More than 80% of the respondents reported that blood pressure is influenced by stress and obesity. Table 3 shows the knowledge about hypertension symptoms, according to the self-reported hypertension status. In general, the knowledge about hypertension symptoms was sparse. For example, less than one-third reported that hypertension may not cause symptoms, and a minority knew that hypertension may cause nosebleeds. There were no systematic differences between the groups, although more people with self-reported hypertension knew that hypertension may not cause symptoms, and more people without self-reported hypertension knew that hypertension may cause headache and palpitations.
The knowledge about complications related to hypertension was good, irrespective of the self-reported hypertension status (Table 4). Eight out of ten subjects reported that hypertension may cause stroke. Table 5 shows the data on the awareness of blood pressure and blood pressure control, according to the self-reported blood pressure status. A total of 81% of those with selfreported hypertension reported that they knew their blood pressure value, while 55% of those without self-reported hypertension knew their blood pressure (OR=3.51 [95% CI: 2.46-5.04]). More subjects with self-reported hypertension planned to have their blood pressure measured within one year (OR=5.0 [95% CI: 2.47-10.0]).

Comparison with prior literature
Our finding of an association between age, weight, selfreported cholesterol level, a family history of hypertension, education level, and self-reported hypertension is in concordance with previous findings. [22][23][24] In comparison with our study, a survey in the USA among people older than 65 years found a similar knowledge about the lifestyle factors that may influence blood pressure but a higher prevalence of knowledge about the complications of untreated high blood pressure. 25 A survey in Canada among people older than 40 years found a limited knowledge of the lifestyle issues affecting hypertension and a poor understanding of the consequences of high blood pressure or hypertension. The Canadian study also showed that two-thirds of the respondents thought hypertension had clearly identifiable signs or symptoms. 26 We found that only 20% knew that hypertension can occur without symptoms, which is in line with the results of a study from Austria, among a sample aged 15 years and older. 27 It was not surprising that people with self-reported hypertension had better blood pressure-related knowledge because they are assumed to have blood pressure measured more frequently and to have more frequent contact with health care services. On the other hand they did not have more knowledge regarding lifestyle influences on blood pressure than those without self-reported hypertension. This finding could be explained by the fact that hypertensive patients were older and less educated. This accords with the Euroaspire III Clinical Epidemiology 2014:6 submit your manuscript | www.dovepress.com

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Knowledge and awareness about blood pressure and hypertension survey, which documented that patients with coronary heart disease and obesity, diabetes, or dyslipidemia need better management and control of their hypertension. 28

strengths and weaknesses
The study sample was drawn randomly and restricted to elderly people, a population with a high prevalence of hypertension. The response rate was good, making the risk of nongeneralizability of study results low. Our study findings may well be extended to the general Danish population because the population of the municipality of Silkeborg shares income and employment rate characteristics with the general Danish population. The study is unique because it was specifically designed for a population with a high prevalence of hypertension, in contrast to other studies that were aimed at the general public. 2,3,5,7,26,27 The multiple choice questionnaire was adjusted after a pilot study. Only a few questionnaires had missing data.
Among those who did not respond, the majority of people were living alone. It may be that the lack of support from a partner contributed to nonresponse. Less knowledge of the subject may also be a reason why almost 28% did not answer the questionnaire. It was not possible to examine for potential selection bias by education and income levels. However, it is well known that participation rates are lower among deprived people. 29 We consider it very likely that those with self-reported hypertension did indeed have hypertension. In cases of self-reported hypertension, part of the questionnaire would have been impossible to fill out if there was no use of antihypertensive drugs or contact with the health care system. Some of the participants without self-reported hypertension may have had hypertension.
In this age group it is likely that about 40%-50% of those who are unaware of their blood pressure do have high blood pressure. 2,3,[5][6][7] Studies that use self-reported hypertension underestimate the true prevalence of hypertension. 30, 31 We did not include questions on the effect of salt on blood pressure because a reduction in population intake of salt primarily is a public health issue that involves governments, regulatory authorities and the food industry. 32 The diagnosis and management of hypertension can be improved by opportunistic screening for hypertension, together

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Qvist et al with complex interventions in the health care system to support health care for those with hypertension. In that respect nurseled clinics have been recommended in the management of hypertension 33 and promising results have been reported. 34 Finally, the multiple choice format of the questionnaire is likely to cause "false positive" responses. This may have caused us to overestimate the knowledge about hypertension.

Future research
Much more knowledge about the efficacy of patienteducation strategies need to be gained to improve general knowledge, behavior, lifestyle and hopefully prognosis related to hypertension.

Conclusion
The knowledge about blood pressure and hypertension was reasonable in general. However, nearly half of the study population without self-reported hypertension was unaware of their blood pressure, and 80% of the respondents did not know that hypertension can occur without symptoms. There is still room for improvement in elderly people's knowledge and awareness about blood pressure.