Discussions on the quality of antibodies are no reason to ban animal immunization

Debates about the source of antibodies and their use are confusing two different issues. A ban on life immunization would have no repercussions on the quality of antibodies. Subject Categories: S&S: Economics & Business, Methods & Resources, Chemical Biology

There is an ongoing debate on how antibodies are being generated, produced and used (Gray, 2020; Marx, 2020). Or rather, there are two debates, which are not necessarily related to each other. The first one concerns the quality of antibodies used in scientific research and the repercussions for the validity of results (Bradbury & Pluckthun, 2015). The second debate is about the use of animals to generate and produce antibodies. Although these are two different issues, we observe that the debates have become entangled with arguments for one topic incorrectly being used to motivate the other and vice versa. This is not helpful, and we should disentangle the knot.Polyclonal antibodies are being criticized because they suffer from cross‐reactivity, high background and batch‐to‐batch variation (Bradbury & Pluckthun, 2015). Monoclonal antibodies produced from hybridomas are criticized because they often lack specificity owing to genetic heterogeneity introduced during hybridoma generation that impairs the quality of the monoclonals (Bradbury et al, 2018). These are valid criticisms and producing antibodies in a recombinant manner will, indeed, help to improve quality and specificity. But a mediocre antibody will remain a mediocre antibody, no matter how it is produced. Recombinant methods will just produce a mediocre antibody more consistently.Getting a good antibody is not easy and much depends on the nature and complexity of the antigen. And low‐quality antibodies are often the result of poor screening, poor quality control, incomplete characterization and the lack of international standards. Nevertheless, the technologies to ensure good selection and to guarantee consistent quality are much more advanced than a decade ago, and scientists and antibody producers should implement these to deliver high‐quality antibodies. Whether antibodies are generated by animal immunization or from naïve or synthetic antibody libraries is less relevant; they can all be produced recombinantly, and screening and characterization are needed in all cases to determine quality, and if the antibody is fit for purpose.But criticisms on the quality of many antibodies and pleas for switching to recombinant production of antibodies cannot be mixed up with a call to ban animal immunization. The EU Reference Laboratory for Alternatives to Animal Testing (EURL ECVAM) recently published a recommendation to stop using animals for generating and producing antibodies for scientific, diagnostic and even therapeutic applications (EURL ECVAM, 2020). This recommendation is mainly supported by scientists who seem to be biased towards synthetic antibody technology for various reasons. Their main argument is that antibodies derived from naïve or synthetic libraries are a valid (and exclusive) alternative. But are they?One can certainly select antibodies from non‐immune libraries, and, depending on the antigen and the type of application, these antibodies can be fit for purpose. In fact, a few of such antibodies have made it to the market as therapeutics, Adalimumab (Humira®) being a well‐known example. But up to now, the vast majority of antibodies continues to come from animal immunization (Lu et al, 2020). And there is a good reason for that. It is generally possible to generate a few positive hits in a naïve/synthetic library; and the more diverse the library, the more hits one is likely to get. But many decades of experience with immunization of animals—especially when they are outbred—shows that they generate larger amounts of antibodies with superior properties. And the more complex your antigen is, the more the balance swings towards animal immunization if you want to have a guarantee for success.There are different factors at work here. First, the immune system of mammals has evolved over millions of years to efficiently produce excellent antibodies against a very diverse range of antigens. Second, presenting the antigen multiple times in its desired (native) conformation to the animal immune system exploits the natural maturation process to fine‐tune the immune response against particular qualities. Another factor is that in vivo maturation seems to select against negative properties such as self‐recognition and aggregation. It also helps to select for important properties that go beyond mere molecular recognition (Jain et al, 2017). In industrial parlance, antibodies from animal immunization are more “developable” and have favourable biophysical properties (Lonberg, 2005). Indeed, the failure rate for antibodies selected from naïve or synthetic libraries is significantly higher.Of course, the properties of synthetic antibodies selected from non‐immune libraries can be further matured in vitro, for example by light chain shuffling or targeted mutagenesis of the complementarity determining region (CDR). While this method has become more sophisticated over the years, it remains a very complex and iterative process without guarantee that it produces a high‐quality antibody.Antibodies are an ever more important tool in scientific research and a growing area in human and veterinary therapeutics. Major therapeutic breakthroughs in immunology and oncology in the past decades are based on antibodies (Lu et al, 2020). The vast majority of these therapeutic antibodies were derived from animals. An identical picture appears when you look at the antibodies in fast‐track development to combat the current COVID‐19 crisis: again, the vast majority are either derived from patients or from animal immunizations. The same holds true for antibodies that are used in diagnostics and epidemiologic studies for COVID‐19.It is for that reason that we need the tools and methods that guarantee antibodies of the highest quality and provide the best chance for success. The COVID‐19 pandemic is only one illustration of this need. If we block access to these tools, both scientific research and society at large will be negatively impacted. We therefore should not limit ourselves to naïve and synthetic libraries. Animal immunization remains an inevitable method that needs to stay. But we all agree that these immunizations must be performed under best practice to further reduce the harm to animals.     

Reply to Bronstein and Vinogradov

The response by the author. Subject Categories: S&S: Economics & Business, S&S: Ethics

I thank Michael Bronstein and Sophia Vinogradov for their interest and comments. I would like to respond to a few of their points.First, I agree with the authors that empirical studies should be conducted to validate any approaches to prevent the spread of misinformation before their implementation. Nonetheless, I think that the ideas I have proposed may be worth further discussion and inspire empirical studies to test their effectiveness.Second, the authors warn that informing about the imperfections of scientific research may undermine trust in science and scientists, which could result in higher vulnerability to online health misinformation (Roozenbeek et al, 2020; Bronstein & Vinogradov, 2021). I believe that transparency about limitations and problems in research does not necessarily have to diminish trust in science and scientists. On the contrary, as Veit et al put it, “such honesty… is a prerequisite for maintaining a trusting relationship between medical institutions (and practitioners) and the public” (Veit et al, 2021). Importantly, to give an honest picture of scientific research, information about its limitations should be put in adequate context. In particular, the public also should be aware that “good science” is being done by many researchers; we do have solid evidence of effectiveness of many medical interventions; and efforts are being taken to address the problems related to quality of research.Third, Bronstein and Vinogradov suggest that false and dangerous information should be censored. I agree with the authors that “[c]ensorship can prevent individuals from being exposed to false and potentially dangerous ideas” (Bronstein & Vinogradov, 2021). I also recognize that some information is false beyond any doubt and its spread may be harmful. What I am concerned about are, among others, the challenges related to defining what is dangerous and false information and limiting censorship only to this kind of information. For example, on what sources should decisions to censor be based and who should make such decisions? Anyone, whether an individual or an organization, with a responsibility to censor information will likely not only be prone to mistakes, but also to abuses of power to foster their interests. Do the benefits we want to achieve by censorship outweigh the potential risks?Fourth, we need rigorous empirical studies examining the actual impact of medical misinformation. What exactly are the harms we try to protect against and what is their scale? This information is necessary to choose proportionte and effective measures to reduce the harms. Bronstein and Vinogradov give an example of a harm which may be caused by misinformation—an increase in methanol poisoning in Iran. Yet, as noticed by the authors, misinformation is not the sole factor in this case; there are also cultural and other contexts (Arasteh et al, 2020; Bronstein & Vinogradov, 2021). Importantly, the methods of studies exploring the effects of misinformation should be carefully elaborated, especially when study participants are asked to self‐report. A recent study suggests that some claims about the prevalence of dangerous behaviors, such as drinking bleach, which may have been caused by misinformation are largely exaggerated due to the presence of problematic respondents in surveys (preprint: Litman et al, 2021).Last but not least, I would like to call attention to the importance of how veracity of information is determined in empirical studies on misinformation. For example, in a study of Roozenbeek et al, cited by Bronstein and Vinogradov, the World Health Organization (WHO) was used as reliable source of information, which raises questions. For instance, Roozenbeek et al (2020) used a statement “the coronavirus was bioengineered in a military lab in Wuhan” as an example of false information, relying on the judgment of the WHO found on its “mythbusters” website (Roozenbeek et al, 2020). Yet, is there a solid evidence to claim that this statement is false? At present, at least some scientists declare that we cannot rule out that the virus was genetically manipulated in a laboratory (Relman, 2020; Segreto & Deigin, 2020). Interestingly, the WHO also no longer excludes such a possibility and has launched an investigation on this issue (https://www.who.int/health‐topics/coronavirus/origins‐of‐the‐virus, https://www.who.int/emergencies/diseases/novel‐coronavirus‐2019/media‐resources/science‐in‐5/episode‐21‐‐‐covid‐19‐‐‐origins‐of‐the‐sars‐cov‐2‐virus); the information about the laboratory origin of the virus being false is no longer present on the WHO “mythbusters” website (https://www.who.int/emergencies/diseases/novel‐coronavirus‐2019/advice‐for‐public/myth‐busters). Against this backdrop, some results of the study by Roozenbeek et al (2020) seem misleading. In particular, the perception of the reliability of the statement about bioengineered virus by study participants in Roozenbeek et al (2020) does not reflect the susceptibility to misinformation, as intended by the researchers, but rather how the respondents perceive reliability of uncertain information.I hope that discussion and research on these and related issues will continue.     

A CRISPR response to pandemics?: Exploring the ethics of genetically engineering the human immune system

Ethical challenges should be addressed before gene editing is made available to improve the immune response against emerging viruses. Subject Categories: S&S: Economics & Business, Genetics, Gene Therapy & Genetic Disease, Immunology

In 1881, Louis Pasteur proved the “germ theory of disease”, namely that microorganisms are responsible for causing a range of diseases. Following Pasteur’s and Robert Koch’s groundbreaking work on pathogens, further research during the 20^th century elucidated how the immune system fends off disease‐causing microorganisms from a molecular perspective.The COVID‐19 pandemic has again focused scientific and public attention on immunology not the least owing to the race of employing vaccines to halt the spread of the virus. Although most countries have now started vaccination programs to immunize a large part of the world''s population, the process will take time, vaccines may not be available to everyone, and a number of unresolved issues remain including the potential contagiousness of vaccinated individuals and the duration of protection (Polack et al, 2020).It would therefore be extremely helpful from a public health perspective—and indeed lifesaving for those with elevated risk of developing severe course of the disease—if we could boost the human immune system by other means to better fight off SARS‐CoV‐2 and possibly other viruses. Recent studies showing that some individuals may be less susceptible to contract severe COVID‐19 depending on their genetic status support such visions (COVID‐19 Host Genetics Initiative, 2020). This could eventually inspire research projects on gene therapy with the aim of generally enhancing immunity against viral infections.

It would therefore be extremely helpful from a public health perspective […] if we could boost the human immune system by other means to better fight off SARS‐CoV‐2 …

The idea of genetically enhancing the human immune response is not new and spread from academic circles to policymakers and the general public even before the pandemic, when He Jiankui announced in November 2018 the birth of genetically edited twins who, he claimed, were resistant to HIV. The public outcry was massive, not only because He violated standards of methodological rigor and research ethics, but also because of fundamental doubts about the wisdom and legitimacy of human germline manipulation (Schleidgen et al, 2020).Somatic gene therapy has been met with a less categorical rejection, but it has also been confronted with skepticism when major setbacks or untoward events occurred, such as the death of Jesse Gelsinger during an early clinical trial for gene therapy in 1999. Nonetheless, given the drastic impact the current pandemic has on so many lives, there may be a motivation to put concerns aside. In fact, even if we managed to get rid of COVID‐19 owing to vaccines—or at least to keep its infectiousness and mortality low—another virus will appear sooner or later; an improved resistance to viral pathogens—including coronaviruses—would be an important asset.Interventions to boost the immune system could in fact make use of either germline gene editing, as has been the case of the Chinese twins, or through somatic gene editing. The first requires time and only the next generation would potentially benefit while the latter could be immediately applied and theoretically used to deal with the ongoing COVID‐19 pandemic.

Interventions to boost the immune system could in fact make use of either germline gene editing, as has been the case of the Chinese twins, or through somatic gene editing.

     

Strength is in engagement: The rise of an online scientific community during the COVID‐19 pandemic

Many scientists, confined to home office by COVID‐19, have been gathering in online communities, which could become viable alternatives to physical meetings and conferences. Subject Categories: S&S: Careers & Training, Methods & Resources, S&S: Ethics

As COVID‐19 has brought work and travel to a grinding halt, scientists explored new ways to connect with each other. For the gene regulation community, this started with a Tweet that quickly expanded into the “Fragile Nucleosome” online forum, a popular seminar series, and many intimate discussions connecting scientists all over the world. More than 2,500 people from over 45 countries have attended our seminars so far and our forum currently has ~ 1,000 members who have kick‐started discussion groups and mentorship opportunities. Here we discuss our experience with setting up the Fragile Nucleosome seminars and online discussion forum, and present the tools to enable others to do the same.Too often, we forget the importance of social interactions in science. Indeed, many creative ideas originated from impromptu and fortuitous encounters with peers, in passing, over lunch, or during a conference coffee break. Now, the ongoing COVID‐19 crisis means prolonged isolation, odd working hours, and less social interactions for most scientists confined to home. This motivated us to create the “Fragile Nucleosome” virtual community for our colleagues in the chromatin and gene regulation field.

… the ongoing COVID‐19 crisis means prolonged isolation, odd working hours and less social interactions for most scientists confined to home.

While the need to address the void created by the COVID‐19 pandemic triggered our actions, a large part of the international community already has had limited access to research networks in our field. Our initiative offered new opportunities though, in particular for those who have not benefited from extensive networks, showing how virtual communities can address disparities in accessibility. This should not be a stop‐gap measure during the pandemic: Once we come out from our isolation, we still need to address the drawbacks of in‐person scientific conferences/seminars, such as economic disparities, travel inaccessibility, and overlapping family responsibilities (Sarabipour, 2020). Our virtual community offers some solutions to the standing challenges (Levine & Rathmell, 2020), and we hope our commentary can help start conversations about the advantages of virtual communities in a post‐pandemic world.

… once we come out from our isolation we still need to address the drawbacks of in‐person scientific conferences/seminars, such as economic disparities, travel inaccessibility and overlapping family responsibilities…

     

When we increase diversity in academia,we all win

Increasing diversity in academia is not just a matter of fairness but also improves science. It is up to individual scientists and research organisations to support underrepresented minorities. Subject Categories: S&S: Economics & Business, S&S: Ethics

There has been a large body of research on diversity in the workplace—in both academic and non‐academic settings—that highlights the benefits of an inclusive workplace. This is perhaps most clearly visible in industry where the rewards are immediate: A study by McKinsey showed that companies with a more diverse workforce perform better financially and by substantial margins, compared to their respective national industry medians (https://www.mckinsey.com/business-functions/organization/our-insights/why-diversity-matters#).It is easy to measure success in financial terms, but since there is no similar binary metric for research performance (https://sfdora.org), it is harder to quantify the rewards of workplace diversity in academic research. However, research shows that diversity actually provides research groups with a competitive edge in other quantifiable terms, such as citation counts (Powell, 2018), and the scientific process obviously benefits from diversity in perspectives. Bringing together individuals with different ways of thinking will allow us to solve more challenging scientific problems and lead to better decision‐making and leadership. Conversely, there is a direct cost to bias in recruitment, tenure, and promotion processes. When such processes are affected by bias—whether explicit or implicit—the whole organization is losing by not tapping into the wider range of skills and assets that could otherwise have been brought to the workplace. Promoting diversity in academia is therefore not simply an issue of equity, which in itself is a sufficient reason, but also a very practical question: how do we create a better work environment for our organization, both in terms of collegiality and in terms of performance?Notwithstanding the fact that there is now substantial awareness of the importance of diversity and that significant work is being invested into addressing the issue, the statistics do not look good. Despite a substantial improvement at the undergraduate and graduate student levels in the EU, women remain significantly underrepresented in research at the more senior levels (Directorate‐General for Research and Innovation European Commission, 2019). In addition, the lion’s share of diversity efforts, at least in Sweden where I work, is frequently focused on gender. Gender is clearly important, but other diversity axes with problematic biases deserve the same attention. As one example, while statistics on ethnic diversity is readily available for US Universities (Davis & Fry, 2019), this information is much harder to find in Europe. While there is an increased awareness of diversity at the student level, this does not necessarily translate into initiatives to support faculty diversity (Aragon & Hoskins, 2017). There are examples of progress and concrete actions on these fronts, including the Athena Swan Charter (https://www.ecu.ac.uk/equality-charters/athena-swan/), the more recent Race Equality Charter (https://www.advance-he.ac.uk/charters/race-equality-charter), and the EMBO journals that regularly analyze their decisions for gender bias. However, progress remains frustratingly slow. In 2019, the World Economic Forum suggested that, at the current rate of progress, the global gender gap will take 108 years to close (https://www.weforum.org/reports/the-global-gender-gap-report-2018). I worry that it may take even longer for other diversity axes since these receive far less attention.It is clear that there is a problem, but what can we do to address it? Perhaps one of the single most important contributions we can make as faculty is to address the implicit (subconscious) biases we all carry. Implicit bias will manifest itself in many ways: gender, ethnicity, socioeconomic status, or disability, just to mention a few. These are the easily identifiable ones, but implicit bias also extends to, for example, professional titles (seniority level), institutional affiliation and even nationality. These partialities affect our decision‐making—for example, in recruitment, tenure, promotion, and evaluation committees—and how we interact with each other.The “Matilda effect” (Rossiter, 1993), which refers to the diminishment of the value of contributions made by female researchers, is now well recognized, and it is not unique to gender (Ross, 2014). When we diminish the contributions of our colleagues, it affects how we evaluate them in competitive scenarios, and whether we put them forward for grants, prizes, recruitment, tenure, and so on. In the hypercompetitive environment that is academia today, even small and subtle injuries can tremendously amplify their negative impact on success, given the current reward system that appears to favor “fighters” over “collaborators”. Consciously working to correct for this, stepping back to rethink our first assessment, is imperative.Women and other minorities also frequently suffer from imposter syndrome, which can impact self‐confidence and make members of these groups less likely to self‐promote in the pursuit of prestigious funding, awards, and competitive career opportunities. This effect is further amplified by a globally mobile academic workforce who, when moving to new cultural contexts (whether locally or internationally), can be unaware of the unwritten rules that guide a department’s work environment and decision‐making processes. Here, mentoring can play a tremendous role in reducing barriers to success; however, for such mentorship to be productive, mentors need to be aware of the specific challenges faced by minorities in academia, as well as their own implicit biases (Hinton et al, 2020).Other areas where we, as individual academics, can contribute to a more diverse work environment include meeting cultures and decision‐making. Making sure that the members of decision‐making bodies have diverse composition so that a variety of views are represented is an important first step. One complication to bear in mind though is that implicit biases are not limited to individuals outside the group: A new UN report shows that almost 90% of people—both men and women—carry biases against women, which in turn is what contributes to the glass‐ceiling effect (United Nations Development Program, 2020). However, equally important is inclusiveness in the meeting culture. Studies from the business world show that even high‐powered women often struggle to speak up and be heard at meetings, and the onus for solving this is often passed back onto themselves. The same holds true for other minority groups, and in an academic setting, it extends to seminars and conferences. The next time you plan a meeting, think about the setting and layout. Who gets to talk? Why? Is the distribution of time given to participants representative of the composition of the meeting participants? If not, why not?As a final example of personal action, we can take: language matters (Ås, 1978). Even without malicious intent, there can be a big gap between what we say and mean, and how it comes across to the recipient. Some examples of this are given by Harrison and Tanner (Harrison & Tanner, 2018), who discuss microagressions in an academic setting and the underlying message one might be unintentionally sending. Microaggressions, when built up over a long period of time, and coming from different people, can significantly impact someone’s confidence and sense of self‐worth. Taking a step back and thinking about why we choose the language, we do is a vital part of creating an inclusive work environment.Addressing diversity challenges in academia is a highly complex multi‐faceted topic that is impossible to do justice in a short opinion piece. This is, therefore, just a small set of examples: By paying attention to our own biases and thinking carefully about how we interact with those around us, both in terms of the language we use and the working environments we create, we can personally contribute to improving both recruitment and retention of a diverse academic workforce. In addition, it is crucial to break the culture of silence and to speak up when we see others committing micro‐ or not so microaggressions or otherwise contributing to a hostile environment. There is a substantial amount of work that needs to be done, at both the individual and organization levels, before we have a truly inclusive academic environment. However, this is not a reason to not do it, and if each of us contributes, we can accelerate this change to a better and more equitable future, while all winning from the benefits of diversity.     

The need for sustainable leadership in academia: A survey of German researchers reveals a widespread lack of training for leadership skills

A survey of academics in Germany shows a lack of and a great demand for training in leadership skills. Subject Categories: Careers, Science Policy & Publishing

Success and productivity in science is measured largely by the number of publications in scientific journals and the acquisition of third‐party funding to finance further research (Detsky, 2011). Consequently, as young researchers advance in their careers, they become highly trained in directly related skills, such as scientific writing, so as to increase their chances in securing publications and grants. Acquiring leadership skills, however, is often neglected as these do not contribute to the evaluation of scientific success (Detsky, 2011). Therefore, an early‐career researcher may become leader of a research group based on publication record and solicitation of third‐party funding, but without any training of leadership or team management skills (Lashuel, 2020). Leadership, in the context of academic research, requires a unique list of competencies, knowledge and skills in addition to “traditional” leadership skills (Anthony & Antony, 2017), such as managing change, adaptability, empathy, motivating individuals, and setting direction and vision among others. Academic leadership also requires promoting the research group’s reputation, networking, protecting staff autonomy, promoting academic credibility, and managing complexity (Anthony & Antony, 2017).     

Experts in science communication: A shift from neutral encyclopedia to equal participant in dialogue

Even if the predominant model of science communication with the public is now based on dialogue, many experts still adhere to the outdated deficit model of informing the public. Subject Categories: Genetics, Gene Therapy & Genetic Disease, S&S: History & Philosophy of Science, S&S: Ethics

During the past decades, public communication of science has undergone profound changes: from policy‐driven to policy‐informing, from promoting science to interpreting science, and from dissemination to interaction (Burgess, 2014). These shifts in communication paradigms have an impact on what is expected from scientists who engage in public communication: they should be seen as fellow citizens rather than experts whose task is to increase scientific literacy of the lay public. Many scientists engage in science communication, because they see this as their responsibility toward society (Loroño‐Leturiondo & Davies, 2018). Yet, a significant proportion of researchers still “view public engagement as an activity of talking to rather than with the public” (Hamlyn et al, 2015). The highly criticized “deficit model” that sees the role of experts as educating the public to mitigate skepticism still persists (Simis et al, 2016; Suldovsky, 2016).Indeed, a survey we conducted among experts in training seems to corroborate the persistence of the deficit model even among younger scientists. Based on these results and our own experience with organizing public dialogues about human germline gene editing (Box 1), we discuss the implications of this outdated science communication model and an alternative model of public engagement, that aims to align science with the needs and values of the public.Box 1

The DNA‐dialogue project

The Dutch DNA‐dialogue project invited citizens to discuss and form opinions about human germline gene editing. During 2019 and 2020, this project organized twenty‐seven dialogues with professionals, such as embryologists and midwives, and various lay audiences. Different scenarios of a world in 2039 (https://www.rathenau.nl/en/making‐perfect‐lives/discussing‐modification‐heritable‐dna‐embryos) served as the starting point. Participants expressed their initial reactions to these scenarios with emotion‐cards and thereby explored the values they themselves and other participants deemed important as they elaborated further. Starting each dialogue in this way provides a context that enables everyone to participate in dialogue about complex topics such as human germline gene editing and demonstrates that scientific knowledge should not be a prerequisite to participate.An important example of “different” relevant knowledge surfaced during a dialogue with children between 8 and 12 years in the Sophia Children’s Hospital in Rotterdam (Fig 1). Most adults in the DNA‐dialogues accepted human germline gene modification for severe genetic diseases, as they wished the best possible care and outcome for their children. The children at Sophia, however, stated that they would find it terrible if their parents had altered something about them before they had been born; their parents would not even have known them. Some children went so far to say they would no longer be themselves without their genetic condition, and that their condition had also given them experiences they would rather not have missed.Open in a separate windowFigure 1 Children participating in a DNA‐dialogue meeting. Photographed by Levien Willemse.     

Biodiversity 2030: a road paved with good intentions: The new EU Commission's biodiversity Strategy risks to remain an empty husk without proper implementation

The EU''s Biodiversity Strategy for 2030 makes great promises about halting the decline of biodiversity but it offers little in terms of implementation. Subject Categories: S&S: Economics & Business, Ecology, S&S: Ethics

Earth is teeming with a stunning variety of life forms. Despite hundreds of years of exploration and taxonomic research, and with 1.2 million species classified, we still have no clear picture of the real extent of global biodiversity, with estimates ranging from 3 to 100 million species. A highly quoted—although not universally accepted—study predicted some 8.7 million species, of which about 2.2 million are marine (Mora et al, 2011). Although nearly any niche on the surface of Earth has been colonized by life, species richness is all but evenly distributed. A large share of the known species is concentrated in relatively small areas, especially in the tropics (Fig 1). Ultimately, it is the network of the interactions among life forms and the physical environment that make up the global ecosystem we call biosphere and that supports life itself.Open in a separate windowFigure 1Biological hotspots of the worldA total of 36 currently recognized hotspots make up < 3% of the planet''s land area but harbor half of the world''s endemic plant species and 42% of all terrestrial vertebrates. Overall, hotspots have lost more than 80% of their original extension. Credit: Richard J. Weller, Claire Hoch, and Chieh Huang, 2017, Atlas for the End of the World, http://atlas‐for‐the‐end‐of‐the‐world.com/. Reproduced with permission.Driven by a range of complex and interwoven causes–such as changes in land and sea use, habitat destruction, overexploitation of organisms, climate change, pollution, and invasive species–biodiversity is declining at an alarming pace. A report by the Intergovernmental Science‐Policy Platform on Biodiversity and Ecosystem Services (IPBES) issued a clear warning: “An average of around 25 per cent of species in assessed animal and plant groups are threatened, suggesting that around 1 million species already face extinction, many within decades, unless action is taken to reduce the intensity of drivers of biodiversity loss. Without such action, there will be a further acceleration in the global rate of species extinction, which is already at least tens to hundreds of times higher than it has averaged over the past 10 million years” (IPBES, 2019) (Fig 2). Although focused on a smaller set of organisms, a more recent assessment by WWF has reached similar conclusions. Their Living Planet Index, that tracks the abundance of thousands of populations of mammals, birds, fish, reptiles, and amphibians around the world, shows a stark decline in monitored populations (WWF, 2020). As expected, the trend of biodiversity decline is not homogeneous with tropical areas paying a disproportionately high price, mostly because of unrestrained deforestation and exploitation of natural resources.Open in a separate windowFigure 2The global, rapid decline of biodiversity(A) Percentage of species threatened with extinction in taxonomic groups that have been assessed comprehensively, or through a “sampled” approach, or for which selected subsets have been assessed by the IUCN Red List of Threatened Species. Groups are ordered according to the best estimate, assuming that data‐deficient species are as threatened as non‐data deficient species. (B) Extinctions since 1500 for vertebrate groups. (C) Red List Index of species survival for taxonomic groups that have been assessed for the IUCN Red List at least twice. A value of 1 is equivalent to all species being categorized as Least Concern; a value of zero is equivalent to all species being classified as Extinct. Data for all panels from www.iucnredlist.org. Reproduced from (IPBES, 2019), with permission.

Driven by a range of complex and interwoven causes […] biodiversity is declining at an alarming pace.

Against this dire background, the EU has drafted a Biodiversity Strategy 2030, an ambitious framework aimed to tackling the key reasons behind biodiversity loss. The plan hinges around a few main elements, such as the establishment of protected areas for at least 30% of Europe''s lands and seas (Fig 3); a significant increase of biodiversity‐rich landscape features on agricultural land by establishing buffer zones like hedges and fallow fields; halting and reversing the decline of pollinators; and planting 3 billion trees by 2030 (https://ec.europa.eu/info/strategy/priorities‐2019‐2024/european‐green‐deal/actions‐being‐taken‐eu/eu‐biodiversity‐strategy‐2030_en). The budget for implementing these measures was set at €20 billion per year.Open in a separate windowFigure 3Natura 2000, the EU''s network of protected areasIn 2019, 18% of land in the EU was protected as Natura 2000, with the lowest share of protected land in Denmark (8%) and the highest in Slovenia (38%). In 2019, the largest national network of terrestrial Natura 2000 sites was located in Spain, covering 138,111 km², followed by France (70,875 km²) and Poland (61,168 km²). Reproduced from Eurostat: https://ec.europa.eu/eurostat/statistics‐explained/index.php?title=Main_Page “Nature is vital for our physical and mental wellbeing, it filters our air and water, it regulates the climate and it pollinates our crops. But we are acting as if it didn''t matter, and losing it at an unprecedented rate”, said Virginijus Sinkevičius, Commissioner for the Environment, Oceans and Fisheries, at the press launch of the new EU action (https://ec.europa.eu/commission/presscorner/detail/en/ip_20_884). “This new Biodiversity Strategy builds on what has worked in the past, and adds new tools that will set us on a path to true sustainability, with benefits for all. The EU''s aim is to protect and restore nature, to contribute to economic recovery from the current crisis, and to lead the way for an ambitious global framework to protect biodiversity around the planet”.Environmental groups and other stakeholders have welcomed the EU''s pledge in principle. “This is a unique opportunity to shape a new society in harmony with nature”, applauded Wetlands International. “We must not forget that the biodiversity and climate crisis is a much bigger and persistent challenge for humanity than COVID‐19”, (https://europe.wetlands.org/news/welcoming‐the‐eu‐biodiversity‐strategy‐for‐2030/). EuroNatur, a foundation focused on conservation, stated that the goals set out by the new strategy provide a strong basis for improving the state of nature in the EU (www.euronatur.org).Alongside the voices of praise, however, many have expressed concerns that the strategy could turn into a little more than a wish list. “The big issue of the strategy is that while setting a goal for financial funds, the EU does not specify where the money is supposed to come from. It only says it should include ‘EU funds and national and private funding’”, commented the European Wilderness Society, an environmental advocacy non‐profit organization headquartered in Tamsweg, Austria. “Goals are important, but do not create change without an organized and sustainable implementation. It''s a good and ambitious document, but what is also obvious is the lack of strategy of how to implement it, and a lack of discussion of why previous documents of this type failed” (https://wilderness‐society.org/ambitious‐eu‐biodiversity‐strategy‐2030/).

Alongside the voices of praise, however, many have expressed concerns that the strategy could turn into a little more than a wish list.

The Institute for European Environmental Policy (IEEP) is on the same page. The sustainability think‐tank based in Brussels and London noted that the outgoing EU 2020 biodiversity strategy showed major implementation problems, especially because of lack of engagement at national level and of ad hoc legislation supporting the meeting of key targets. Therefore, “[it] can be argued that a legally binding approach to the biodiversity governance framework is urgently needed unless Member States and other key stakeholders can show greater intrinsic ownership to deliver on agreed objectives”, (https://ieep.eu/news/first‐impressions‐of‐the‐eu‐biodiversity‐strategy‐to‐2030). In addition, IEEP remarked that money is an issue, since the €20 billion figure appears more as an estimate than a certified obligation.“The intentions of the Commission are good and the strategy contains a number of measures and targets that can really make a difference. However, implementation depends critically on the member states and experiences with the Common Agricultural Policy the past decade or so have taught us that many of them are more interested in short‐term economic objectives than in safeguarding the natural wealth of their country for future generations”, commented David Kleijn, an ecologist and nature conservation expert at the Wageningen University, the Netherlands. “I think it is important that we now have an ambitious Biodiversity Strategy but at the same time I have little hope that we will be able to achieve its objectives”.

I think it is important that we now have an ambitious Biodiversity Strategy but at the same time I have little hope that we will be able to achieve its objectives.

There is further criticism against specific measures, such as the proposal of planting 3 billion trees. “To have lots of trees planted in an area does not necessarily translate into an increase of biodiversity. Biodiverse ecosystems are the result of million years of complex multi‐species interactions and evolutionary processes, which are not as easy to restore”, explained plant ecologist Susana Gómez‐González, from the University of Cádiz, Spain. Planting a large number of trees is a too simplistic approach for saving European forests from the combined effects of excessive anthropic pressure and climate change, and could even have detrimental effects (see Box 1). More emphasis should be placed instead in reducing tree harvesting in sensitive areas and in promoting natural forest renewal processes (Gómez‐González et al, 2020). “For a biodiversity strategy, increasing the number of trees, or even increasing the forest area, should not be an objective; priority should be given to the conservation and restoration of natural ecosystems, forests and non‐forests”, Gómez‐González said.In other cases, it could be difficult, if not impossible, to reach some of the goals because of lack of information. For example, one of the roadmap''s targets is to restore at least 25,000 km of Europe''s rivers back to free‐flowing state. However, the number of barriers dispersed along European rivers will probably prevent even getting close to the mark. An international research team has collected detailed information on existing instream barriers for 147 rivers in 36 European countries, coming up with the impressive figure of over 1.2 million obstacles that inevitably impact on river ecosystems, affecting the transport and dispersion of aquatic organisms, nutrients, and sediments (Belletti et al, 2020). Existing inventories mainly focused on dams and other large barriers, while, in fact, a large number of artificial structures are much smaller, such like weirs, locks, ramps, and fords. As a result, river fragmentation has been largely underestimated, and the models used to plan flow restoration might be seriously flawed. “To avoid ‘death by a thousand cuts’, a paradigm shift is necessary: to recognize that although large dams may draw most of the attention, it is the small barriers that collectively do most of the damage. Small is not beautiful”, concluded the authors (Belletti et al, 2020).

Box 1: Why many trees don''t (always) make a forestForests are cathedrals of biodiversity. They host by far the largest number of species on land, which provide food and essential resources for hundreds of millions of people worldwide. However, forests are disappearing and degrading at an alarming pace. The loss of these crucial ecosystems has given new impulses to a variety of projects aimed at stopping this devastation and possibly reversing the trend.Once it is gone, can you rebuild a forest? Many believe the answer is yes, and the obvious solution is to plant trees. Several countries have thus launched massive tree‐planting programs, notably India and Ethiopia, where 350 million trees have been planted in single day (https://www.unenvironment.org/news‐and‐stories/story/ethiopia‐plants‐over‐350‐million‐trees‐day‐setting‐new‐world‐record). The World Economic Forum has set up its own One Trillion Tree initiative (https://www.1t.org/) “to conserve, restore, and grow one trillion trees by 2030”. Launched in January last year at Davos, 1t.org was conceived as a platform for governments, companies and NGOs/civil society groups to support the UN Decade on Ecosystem Restoration (2021–2030). The initiative has been christened by renowned naturalist Jane Goodall, who commented: “1t.org offers innovative technologies which will serve to connect tens of thousands of small and large groups around the world that are engaged in tree planting and forest restoration”, (https://www.weforum.org/agenda/2020/01/one‐trillion‐trees‐world‐economic‐forum‐launches‐plan‐to‐help‐nature‐and‐the‐climate/).However, things are way more complicated than they appear: large‐scale tree planting schemes are rarely a viable solution and can even be harmful. “[A] large body of literature shows that even the best planned restoration projects rarely fully recover the biodiversity of intact forests, owing to a lack of sources of forest‐dependent flora and fauna in deforested landscapes, as well as degraded abiotic conditions resulting from anthropogenic activities”, commented Karen Holl from the University of Caliornia, Santa Cruz, and Pedro Brancalion from the University of São Paulo (Holl & Brancalion, 2020). A common problem of tree plantations, for example, is the low survival rate of seedlings, mostly because the wrong tree species are selected and due to poor maintenance after planting. Moreover, grasslands and savannas, which are often targeted for establishing new forests, are themselves treasure troves of biodiversity. Ending indiscriminate deforestation, improving the protection of existing forests, and promoting their restoration would therefore be a more efficient strategy to preserve biodiversity in the shorter term. If tree planting is indeed necessary, it should be well planned by selecting the right areas for reforestation, using suitable tree species that can maximize biodiversity, and involving local populations to maintain the plantations, Holl and Brancalion argue (Holl & Brancalion, 2020).

…even the best planned restoration projects rarely fully recover the biodiversity of intact forests, owing to a lack of sources of forest‐dependent flora and fauna in deforested landscapes…

The health of soil, where a high proportion of biodiversity is hosted, is another problem the new strategy should address in a more focused manner. “In my opinion, the EU Biodiversity Strategy is already a leap forward in terms of policy interest in soils in general and in soil biodiversity in particular. Compared with other nations/regions of the world, Europe is by far in the forefront regarding this issue”, commented Carlos António Guerra at the German Centre for Integrative Biodiversity Research (iDiv) in Leipzig, Germany, and Co‐leader of the Global Soil Biodiversity Observation Network (https://geobon.org/bons/thematic‐bon/soil‐bon/). “Nevertheless, the connection between soil biodiversity and ecological functions needs further commitments. Soils allow for horizontal integration of several policy agendas, from climate to agriculture and, very importantly, nature conservation. This is not explicit in the EU Biodiversity Strategy in regard to soils”. It remains to be seen if EU restoration plan will emphasize soil biodiversity, or consider it as a mere side effect of other initiatives, Guerra added. “A soil nature conservation plan should be proposed”, he said. “Only such a plan, that implies that current and future protected areas have to consider, describe and protect their soil biodiversity would make a significant push to help protect such a valuable resource”.More generally, research shows that the current paradigm of protection must be shifted to prevent further losses to biodiversity. In fact, an analysis of LIFE projects—a cornerstone of EU nature protection—found that conservation efforts are extremely polarized and strongly taxonomically biased (Mammola et al, 2020). From 1992 to 2018, investment in vertebrates was sixfold higher than that for invertebrates, with birds and mammals alone accounting for 72% of the targeted species and 75% of the total budget. In relative terms, investment per species for vertebrates has been 468 times higher than for invertebrates (Fig 4). There is no sound scientific reasoning behind this uneven conservation attention, but just popularity. “[T]he species covered by a greater number of LIFE projects were also those which attracted the most interest online, suggesting that conservation in the EU is largely driven by species charisma, rather than objective features”, the researchers wrote (Mammola et al, 2020).Open in a separate windowFigure 4Taxonomic bias in EU fauna protection effortsBreakdown of the number of projects (A) and budget allocation (B) across main animal groups covered by the LIFE projects (n = 835). (C) The most covered 30 species of vertebrates (out of 410) and invertebrates (out of 78) in the LIFE projects analyzed (n = 835). The vertical bar represents monetary investment and the blue scatter line the number of LIFE projects devoted to each species. Reproduced from (Mammola et al, 2020), with permission.     

Portugal’s prominence in malaria research: How a small country became a key player in European research on one of the world’s deadliest diseases

Despite its limited resources, Portugal has gained a prominent position in research on malaria. Several historical and personal factors have contributed to this achievement. Subject Categories: S&S: Economics & Business, S&S: History & Philosophy of Science, Microbiology, Virology & Host Pathogen Interaction

Despite a significant increase that started during the 1990s, Portugal’s scientific production remains rather modest compared with the overall research output in the European Union (EU). However, the country’s achievements in malaria research are truly remarkable and, in relative terms, far above its EU neighbors in most relevant accounts. The factors to explain this accomplishment include the fact that malaria was autochthonous in Portugal until 1973; the country’s colonial history and its close ties with its former colonies; and several outstanding scientists who each inspired generations of malariologists.For most of the 20th century, research in Portugal was underfunded, and the country’s overall contribution to science was modest at best. This started to change when Portugal joined the European Union (then the European Economic Community) in 1985 and gained further momentum in the 1990s with the creation of a dedicated Ministry of Science. As a consequence, the Portuguese scientific production increased significantly in terms of the number of scientific articles published. Nevertheless, public funding for research has remained well below that of many other EU countries, and far from the target of 3% of the country’s GDP, which limits Portugal’s overall scientific output. Yet, there is one field of research where Portugal has been making significant contributions, even long before 1985: malaria.

… there is one field of research where Portugal has been making significant contributions, even long before 1985: malaria.

Among many other achievements, Portuguese laboratories have delivered important contributions to malaria research in areas as diverse as drug development, discovery and repurposing, genetic diversity of Plasmodium parasites, mechanisms of drug resistance, co‐infection between Plasmodium and other parasites, host–Plasmodium interactions, nutrient sensing and acquisition by malaria parasites, modulation of Plasmodium liver infection, immune and inflammatory responses to Plasmodium infection, diagnosis, vaccines, the role of microbiota on malaria transmission, pathogenesis of placental and cerebral malaria and acute lung injury, mechanisms of tolerance to malaria, malaria epidemiology, and vector genetics (see Further Reading for examples). Portugal’s percentage of scientific papers published in the field of malaria during the past decade relative to the total number of published articles is the highest in the EU (Fig 1A). Naturally, Portugal cannot compete with larger or more affluent countries in terms of the absolute numbers of articles published on malaria. Yet, the country ranks 5^th in this regard, closely following the Netherlands, Belgium, Sweden, and Denmark, four countries that have been investing much more and much longer in scientific research (Fig 1B). In fact, if one takes into account the funding for R&D in the EU nations, Portugal ranks ahead of every other country in terms of the number of malaria papers published relative to the investment made in science at the national level (Fig 1C).Open in a separate windowFigure 1Malaria research in Portugal and in the EU(A) Percentage of papers on the subject of malaria relative to the total number of papers from each of the indicated countries from 2009 to present. (B) Number of malaria research articles per 1,000 researchers in each of the indicated countries. (C) Number of malaria research articles per 100,000 Euros of gross domestic expenditure on R&D in each of the indicated countries. Total R&D personnel and intramural R&D expenditure data are from 2017. Papers were quantified through searches of PubMed for articles with affiliation to each of the indicated countries, published from 2009 to present, by use of the terms “malaria” or “Plasmodium”. Data on R&D investments from Eurostat.This raises the question of why Portugal, a rather small country with only a few decades of research history and an overall moderate scientific performance, fares relatively so well when it comes to research on malaria. I argue that there are three independent, albeit interrelated factors to explain this feat.A lasting reality demanding an appropriate responseThe first factor was the presence of autochthonous malaria in Portugal until the second half of the 20th century and the establishment of research institutions largely dedicated to studying and fighting the disease. Until the end of the World War II, malaria was endemic throughout much of Southern Europe; Italy, Greece, and Portugal were particularly affected. From 1955 to 1969, the WHO conducted its Global Malaria Eradication Programme, which successfully eliminated malaria in several regions of the world, including Southern Europe. The specific history of malaria eradication in Portugal is described in great detail by Bruce‐Chwatt (Bruce‐Chwatt, 1977) and highlights the intense efforts by multiple state‐sponsored institutions dedicated to studying and combating the disease.Even before the war, in 1931, the Malaria Research Station (Estação Experimental de Combate ao Sezonismo, EECS) was created in Benavente, the goals of which included the collection and analysis of blood samples from infected individuals, treatment of malaria patients, identification of mosquito populations, and malaria prophylaxis. In 1938, the Malaria Institute (Instituto de Malariologia, iMal) was founded in Águas de Moura to investigate the epidemiology of the disease, promoting adequate treatment and implementing vector control measures (Saavedra, 2010). Nonetheless, it was not until 1973 that malaria was eventually eliminated in Portugal, three years after Italy, and only one year before Greece.

… it was not until 1973 that malaria was eventually eliminated in Portugal, three years after Italy, and only one year before Greece.

Yet, the threat of malaria reemergence meant ongoing vigilance, and iMal paved the way for the creation of the Centre for the Study of Malaria and Parasitology (Centro de Estudos de Malária e Parasitologia), in 1973, later to become the Centre for the Study of Zoonoses (Centro de Estudos de Zoonoses) in 1987, and the Centre for Vector and Infectious Disease Studies (Centro de Estudos de Vetores e Doenças Infeciosas) in 1993. In addition, the Portuguese School of Tropical Medicine (later called National School of Public Health and Tropical Medicine, ENSPMT, now the Institute of Tropical Medicine and Hygiene, IHMT), founded in 1902, was one of only four institution of its kind in the world (Amaral, 2008). Since its inception, its mission has been the teaching and research in tropical medicine, biomedical sciences, and international health and, to this day, a significant part of its research continues to focus on malaria.A close bond with AfricaAnother major factor for Portugal’s prominent position in malaria research is its colonial past and the country’s close ties with its former colonies. During its period of maritime expansion in the 15^th and 16^th centuries, Portugal colonized many territories from Asia to the Americas and Africa. Most, if not all, of these territories were, and for a large part still are, endemic for malaria. Former colonies, such as Brazil or the Portuguese territories in India, gained their independence during the 19^th century, but maintained close ties with Portugal.However, several African countries, specifically Angola, Cape Verde, Guinea‐Bissau, Mozambique, and S. Tomé & Príncipe, remained under Portuguese rule until well into the second half of the 20^th century (Miller, 1975). In fact, while most African nations gained their independence from European countries during the 1950s and 1960s, Portugal’s dictatorship held on to and suppressed its African overseas territories, which led to armed uprisings in Angola and Guinea‐Bissau in 1961, and in Mozambique in 1964 (Miller, 1975). During the ensuing colonial wars, thousands of Portuguese soldiers were sent to these countries, where they were exposed not only to the horrors of war, but also to malaria (Campos, 2017). The Portuguese military actions in Africa finally came to an end in 1974 after the peaceful Carnation Revolution, which established democracy in Portugal and ended the colonization of all Portuguese‐held African territories.Over the next few years, hundreds of thousands of military personnel and former residents of the ex‐colonies, known as “retornados”, moved back to Portugal, leading to an increase in the number of imported malaria cases (Bruce‐Chwatt, 1977). Since then, these numbers have subsided, but the close ties that Portugal maintains with its former colonies mean that travel to and from malaria‐endemic regions remains high, contributing to the prevalence of imported malaria cases (Piperaki, 2018). It also means that malaria is not such a distant threat for most Portuguese; even today, many younger people have direct contact with family members or friends who have experienced malaria, bringing the reality of this scourge closer to home than in many other EU countries.

… even today, many younger people have direct contact with family members or friends who have experienced malaria, bringing the reality of this scourge closer to home than in many other EU countries.

Remarkable and inspiring figuresThe third and final factor is the enormous and lasting influence of various uniquely inspiring figures from several generations of malaria researchers. Indeed, the history of Portuguese malaria research is rich in prominent scientists who shaped the national research landscape. Attempting to highlight specific names among the many doctors, epidemiologists, and scientists from the past and present is a naturally risky exercise that runs the risk of overlooking important figures. Nevertheless, the crucial contribution of a few representatives of four generations of Portuguese scientists is beyond dispute.Ricardo Jorge (1858–1939) was a renowned epidemiologist responsible for the 1899–1901 National Sanitary Plan, which marked the introduction of modern sanitary concepts in Portugal and changed national public health. In 1903, Jorge was the first to collect reliable and extensive data on the incidence of malaria and its seasonal distribution (JORGE, 1903). He was Portugal’s Health Inspector‐General from 1899 to 1926, succeeded by José Alberto de Faria (1888–1958), another key figure who, with the support of the Rockefeller Foundation (Saavedra, 2014), created the EECS in Benavente, the first step for advancing knowledge about malaria in Portugal (Bruce‐Chwatt, 1977).Well within the 20^th century, Francisco Cambournac (1903–1994) and Fausto Landeiro (1896–1949) were arguably the most important contributors to Portuguese malariology during that period. Following extensive training in some of the most reputed parasitology schools in Europe, Cambournac became Director of Benavente’s EECS in 1933, and Landeiro occupied that position from 1938 to 1949. Cambournac founded the iMal in Águas de Moura, serving as its Director from 1939 to 1954, and became Director of the WHO’s African region from 1954 to 1964 (Lobo, 2012).Cambournac and Landeiro published extensively on the epidemiology, entomology, and control of malaria during the 1930s and 1940s, and gave a comprehensive account of the status of the disease in Portugal during that period. Cambournac’s 237‐page long review (Cambournac, 1942) provided all the epidemiological and other data needed for future planning of control and eradication of malaria in the country, the success of which is widely acknowledged to his immense work (Bruce‐Chwatt, 1977).During the 1960s and early 1970s, the National School of Public Health and Tropical Medicine, ENSPMT, now the Institute of Tropical Medicine and Hygiene, IHMT, played an important role not only in Portuguese research on malaria and other tropical diseases, but also in the cooperation with Portugal’s overseas territories at the time. The 1974 revolution and the decolonization in Africa led to a reshaping of this cooperation, which became increasingly centered on reinforcing the newly independent countries’ health systems, on their capacity to carry out research on endemic diseases, and on training programs in tropical and preventive medicine (Havik, 2015). Virgílio do Rosário, professor at the IHMT and, later, head of the Institute’s Centre for Malaria and Other Tropical Diseases (CMDT), played a pivotal role in this process. Do Rosário was the founder of several national and international networks for studying malaria and neglected diseases in various regions around the world. He inspired a whole generation of future malaria researchers, making him an inescapable figure among Portuguese malariologists in the second half of the 20^th century.At the dawn of the 21^st century, many Portuguese scientists, who had benefitted from the country’s investment in science in the 1980s and 1990s to acquire international training, came back home to set up their own research groups. Among them was Maria Mota, who returned from New York University to Portugal in 2002 to become a group leader, initially at the Instituto Gulbenkian de Ciência (IGC), and subsequently at the Instituto de Medicina Molecular (iMM). Mota’s research on the liver stage of infection by Plasmodium parasites has had an enormous impact and yielded a plethora of outstanding publications. She became Director of iMM in 2014, and commonly features among the most influential women in Portugal. Mota is also a gifted and engaging communicator, who has helped to garner public attention to malaria research and to the fight against the disease. As a great scientist and public advocate for malaria research, Mota has inspired numerous scientists, several of whom have become independent malaria researchers themselves, both in Portugal and internationally.

As a great scientist and public advocate for malaria research, Mota has inspired numerous scientists, several of whom have become independent malaria researchers themselves…

These historical, epidemiological, and humane factors have made Portugal an important player in malaria research, from the basic science of the parasite to the pathology of the disease, and from epidemiology to clinical research and drug development. However, these great achievements, and the role played by individual inspiring scientists, should not be taken for granted, but rather serve as an argument for nurturing and supporting research on malaria by future generations of scientists and political decision‐makers. A small country with fairly limited financial and human resources cannot reasonably aspire to excel in every area of research, but it can efficiently direct and focus its investment on those that are more likely to generate success. The history of Portuguese malaria research clearly demonstrates this and warrants its continued support as a top priority for national science policies.Further ReadingImportant contributions to malaria research by Portuguese laboratories during the past decade Drug development, discovery and repurposing Oliveira R, Guedes RC, Meireles P, Albuquerque IS, Goncalves LM, Pires E, Bronze MR, Gut J, Rosenthal PJ, Prudencio M, Moreira R, O''Neill PM, Lopes F (2014) Tetraoxane‐pyrimidine nitrile hybrids as dual stage antimalarials. J Med Chem 57: 4916–4923da Cruz FP, Martin C, Buchholz K, Lafuente‐Monasterio MJ, Rodrigues T, Sonnichsen B, Moreira R, Gamo FJ, Marti M, Mota MM, Hannus M, Prudencio M (2012) Drug screen targeted at Plasmodium liver stages identifies a potent multistage antimalarial drug. J Infect Dis 205: 1278–1286Hanson KK, Ressurreicao AS, Buchholz K, Prudencio M, Herman‐Ornelas JD, Rebelo M, Beatty WL, Wirth DF, Hanscheid T, Moreira R, Marti M, Mota MM (2013) Torins are potent antimalarials that block replenishment of Plasmodium liver stage parasitophorous vacuole membrane proteins. Proc Natl Acad Sci USA 110: E2838–E2847Machado M, Sanches‐Vaz M, Cruz JP, Mendes AM, Prudencio M (2017) Inhibition of Plasmodium Hepatic Infection by Antiretroviral Compounds. Front Cell Infect Microbiol 7: 329 Genetic diversity of Plasmodium parasites Guerra M, Neres R, Salgueiro P, Mendes C, Ndong‐Mabale N, Berzosa P, de Sousa B, Arez AP (2017) Plasmodium falciparum Genetic Diversity in Continental Equatorial Guinea before and after Introduction of Artemisinin‐Based Combination Therapy. Antimicrob Agents Chemother 61Mendes C, Salgueiro P, Gonzalez V, Berzosa P, Benito A, do Rosario VE, de Sousa B, Cano J, Arez AP (2013) Genetic diversity and signatures of selection of drug resistance in Plasmodium populations from both human and mosquito hosts in continental Equatorial Guinea. Malar J 12: 114 Mechanisms of drug resistance Escobar C, Pateira S, Lobo E, Lobo L, Teodosio R, Dias F, Fernandes N, Arez AP, Varandas L, Nogueira F (2015) Polymorphisms in Plasmodium falciparum K13‐propeller in Angola and Mozambique after the introduction of the ACTs. PLoS One 10: e0119215Ferreira A, Marguti I, Bechmann I, Jeney V, Chora A, Palha NR, Rebelo S, Henri A, Beuzard Y, Soares MP (2011) Sickle hemoglobin confers tolerance to Plasmodium infection. Cell 145: 398–409Veiga MI, Osorio NS, Ferreira PE, Franzen O, Dahlstrom S, Lum JK, Nosten F, Gil JP (2014) Complex polymorphisms in the Plasmodium falciparum multidrug resistance protein 2 gene and its contribution to antimalarial response. Antimicrob Agents Chemother 58: 7390–7397 Host‐Plasmodium interactions Portugal S, Carret C, Recker M, Armitage AE, Goncalves LA, Epiphanio S, Sullivan D, Roy C, Newbold CI, Drakesmith H, Mota MM (2011) Host‐mediated regulation of superinfection in malaria. Nat Med 17: 732–737Real E, Rodrigues L, Cabal GG, Enguita FJ, Mancio‐Silva L, Mello‐Vieira J, Beatty W, Vera IM, Zuzarte‐Luis V, Figueira TN, Mair GR, Mota MM (2018) Plasmodium UIS3 sequesters host LC3 to avoid elimination by autophagy in hepatocytes. Nat Microbiol 3: 17–25Sa ECC, Nyboer B, Heiss K, Sanches‐Vaz M, Fontinha D, Wiedtke E, Grimm D, Przyborski JM, Mota MM, Prudencio M, Mueller AK (2017) Plasmodium berghei EXP‐1 interacts with host Apolipoprotein H during Plasmodium liver‐stage development. Proc Natl Acad Sci USA 114: E1138–E1147 Nutrient sensing and acquisition Itoe MA, Sampaio JL, Cabal GG, Real E, Zuzarte‐Luis V, March S, Bhatia SN, Frischknecht F, Thiele C, Shevchenko A, Mota MM (2014) Host cell phosphatidylcholine is a key mediator of malaria parasite survival during liver stage infection. Cell Host Microbe 16: 778–786Mancio‐Silva L, Slavic K, Grilo Ruivo MT, Grosso AR, Modrzynska KK, Vera IM, Sales‐Dias J, Gomes AR, MacPherson CR, Crozet P, Adamo M, Baena‐Gonzalez E, Tewari R, Llinas M, Billker O, Mota MM (2017) Nutrient sensing modulates malaria parasite virulence. Nature 547: 213–216Meireles P, Mendes AM, Aroeira RI, Mounce BC, Vignuzzi M, Staines HM, Prudencio M (2017) Uptake and metabolism of arginine impact Plasmodium development in the liver. Sci Rep 7: 4072 Modulation of Plasmodium liver infection Ruivo MTG, Vera IM, Sales‐Dias J, Meireles P, Gural N, Bhatia SN, Mota MM, Mancio‐Silva L (2016) Host AMPK Is a Modulator of Plasmodium Liver Infection. Cell Rep 16: 2539–2545Zuzarte‐Luis V, Mello‐Vieira J, Marreiros IM, Liehl P, Chora AF, Carret CK, Carvalho T, Mota MM (2017) Dietary alterations modulate susceptibility to Plasmodium infection. Nat Microbiol 2: 1600–1607 Immune and inflammatory responses to Plasmodium infection Liehl P, Zuzarte‐Luis V, Chan J, Zillinger T, Baptista F, Carapau D, Konert M, Hanson KK, Carret C, Lassnig C, Muller M, Kalinke U, Saeed M, Chora AF, Golenbock DT, Strobl B, Prudencio M, Coelho LP, Kappe SH, Superti‐Furga G et al (2014) Host‐cell sensors for Plasmodium activate innate immunity against liver‐stage infection. Nat Med 20: 47–53Munoz‐Ruiz M, Ribot JC, Grosso AR, Goncalves‐Sousa N, Pamplona A, Pennington DJ, Regueiro JR, Fernandez‐Malave E, Silva‐Santos B (2016) TCR signal strength controls thymic differentiation of discrete proinflammatory gammadelta T cell subsets. Nat Immunol 17: 721–727Seixas E, Gozzelino R, Chora A, Ferreira A, Silva G, Larsen R, Rebelo S, Penido C, Smith NR, Coutinho A, Soares MP (2009) Heme oxygenase‐1 affords protection against noncerebral forms of severe malaria. Proc Natl Acad Sci USA 106: 15837–15842 Diagnosis Frita R, Rebelo M, Pamplona A, Vigario AM, Mota MM, Grobusch MP, Hanscheid T (2011) Simple flow cytometric detection of haemozoin containing leukocytes and erythrocytes for research on diagnosis, immunology and drug sensitivity testing. Malar J 10: 74 Vaccines Reuling IJ, Mendes AM, de Jong GM, Fabra‐Garcia A, Nunes‐Cabaco H, van Gemert GJ, Graumans W, Coffeng LE, de Vlas SJ, Yang ASP, Lee C, Wu Y, Birkett AJ, Ockenhouse CF, Koelewijn R, van Hellemond JJ, van Genderen PJJ, Sauerwein RW, Prudencio M (2020) An open‐label phase 1/2a trial of a genetically modified rodent malaria parasite for immunization against Plasmodium falciparum malaria. Sci Transl Med 12 Pathogenesis of placental and cerebral malaria de Moraes LV, Tadokoro CE, Gomez‐Conde I, Olivieri DN, Penha‐Goncalves C (2013) Intravital placenta imaging reveals microcirculatory dynamics impact on sequestration and phagocytosis of Plasmodium‐infected erythrocytes. PLoS Pathog 9: e1003154Ribot JC, Neres R, Zuzarte‐Luis V, Gomes AQ, Mancio‐Silva L, Mensurado S, Pinto‐Neves D, Santos MM, Carvalho T, Landry JJM, Rolo EA, Malik A, Silva DV, Mota MM, Silva‐Santos B, Pamplona A (2019) gammadelta‐T cells promote IFN‐gamma‐dependent Plasmodium pathogenesis upon liver‐stage infection. Proc Natl Acad Sci USA 116: 9979–9988 Mechanisms of tolerance to malaria Gozzelino R, Andrade BB, Larsen R, Luz NF, Vanoaica L, Seixas E, Coutinho A, Cardoso S, Rebelo S, Poli M, Barral‐Netto M, Darshan D, Kuhn LC, Soares MP (2012) Metabolic adaptation to tissue iron overload confers tolerance to malaria. Cell Host Microbe 12: 693–704Jeney V, Ramos S, Bergman ML, Bechmann I, Tischer J, Ferreira A, Oliveira‐Marques V, Janse CJ, Rebelo S, Cardoso S, Soares MP (2014) Control of disease tolerance to malaria by nitric oxide and carbon monoxide. Cell Rep 8: 126–136 Epidemiology Corder RM, Ferreira MU, Gomes MGM (2020) Modelling the epidemiology of residual Plasmodium vivax malaria in a heterogeneous host population: A case study in the Amazon Basin. PLoS Comput Biol 16: e1007377 Vector genetics Salgueiro P, Moreno M, Simard F, O''Brochta D, Pinto J (2013) New insights into the population structure of Anopheles gambiae s.s. in the Gulf of Guinea Islands revealed by Herves transposable elements. PLoS One 8: e62964Vicente JL, Sousa CA, Alten B, Caglar SS, Falcuta E, Latorre JM, Toty C, Barre H, Demirci B, Di Luca M, Toma L, Alves R, Salgueiro P, Silva TL, Bargues MD, Mas‐Coma S, Boccolini D, Romi R, Nicolescu G, do Rosario VE et al (2011) Genetic and phenotypic variation of the malaria vector Anopheles atroparvus in southern Europe. Malar J 10: 5Early Portuguese institutions dedicated to malaria investigation and researchLandeiro F (1932) Relatório do primeiro ano de luta antisezonática na estação de BenaventeLandeiro F (1934) Organização do Serviço Antisezonático em Portugal     

USP26: a genetic risk factor for sperm X‐Y aneuploidy

Segregation of the largely non‐homologous X and Y sex chromosomes during male meiosis is not a trivial task, because their pairing, synapsis, and crossover formation are restricted to a tiny region of homology, the pseudoautosomal region. In humans, meiotic X‐Y missegregation can lead to 47, XXY offspring, also known as Klinefelter syndrome, but to what extent genetic factors predispose to paternal sex chromosome aneuploidy has remained elusive. In this issue, Liu et al (2021) provide evidence that deleterious mutations in the USP26 gene constitute one such factor.Subject Categories: Cell Cycle, Development & Differentiation, Molecular Biology of Disease

Analyses of Klinefelter syndrome patients and Usp26‐deficient mice have revealed a genetic influence on age‐dependent sex chromosome missegregation during male meiosis.

Multilayered mechanisms have evolved to ensure successful X‐Y recombination, as a prerequisite for subsequent normal chromosome segregation. These include a distinct chromatin structure as well as specialized proteins on the pseudoautosomal region (Kauppi et al, 2011; Acquaviva et al, 2020). Even so, X‐Y recombination fails fairly often, especially in the face of even modest meiotic perturbations. It is perhaps not surprising then that X‐Y aneuploidy—but not autosomal aneuploidy—in sperm increases with age (Lowe et al, 2001; Arnedo et al, 2006), as does the risk of fathering sons with Klinefelter syndrome (De Souza & Morris, 2010).Klinefelter syndrome is one of the most common aneuploidies in liveborn individuals (Thomas & Hassold, 2003). While most human trisomies result from errors in maternal chromosome segregation, this is not the case for Klinefelter syndrome, where the extra X chromosome is equally likely to be of maternal or paternal origin (Thomas & Hassold, 2003; Arnedo et al, 2006). Little is known about genetic factors in humans that predispose to paternal XY aneuploidy, i.e., that increase the risk of fathering Klinefelter syndrome offspring. The general notion has been that paternally derived Klinefelter syndrome arises stochastically. However, fathers of Klinefelter syndrome patients have elevated rates of XY aneuploid sperm (Lowe et al, 2001; Arnedo et al, 2006), implying a persistent defect in spermatogenesis in these individuals rather than a one‐off meiotic error.To identify possible genetic factors contributing to Klinefelter syndrome risk, Liu et al (2021) performed whole‐exome sequencing in a discovery cohort of > 100 Klinefelter syndrome patients, followed by targeted sequencing in a much larger cohort of patients and controls, as well as Klinefelter syndrome family trios. The authors homed in on a mutational cluster (“mutated haplotype”) in ubiquitin‐specific protease 26 (USP26), a testis‐expressed gene located on the X chromosome. Effects of this gene’s loss of function (Usp26‐deficient mice) on spermatogenesis have recently been independently reported by several laboratories and ranged from no detectable fertility phenotype (Felipe‐Medina et al, 2019) to subfertility/sterility associated with both meiotic and spermiogenic defects (Sakai et al, 2019; Tian et al, 2019). With their Klinefelter syndrome cohort findings, Liu et al (2021) also turned to Usp26 null mice, paying particular attention to X‐Y chromosome behavior and—unlike earlier mouse studies—including older mice in their analyses. They found that Usp26‐deficient animals often failed to achieve stable pairing and synapsis of X‐Y chromosomes in spermatocytes, produced XY aneuploid sperm at an abnormally high frequency, and sometimes also sired XXY offspring. Importantly, these phenotypes only occurred at an advanced age: XY aneuploidy was seen in six‐month‐old, but not two‐month‐old Usp26‐deficient males. Moreover, levels of spindle assembly checkpoint (SAC) proteins also reduced in six‐month‐old males. Thus, in older Usp26 null mice, the combination of less efficient X‐Y pairing and less stringent SAC‐mediated surveillance of faithful chromosome segregation allows for sperm aneuploidy, providing another example of SAC leakiness in males (see Lane & Kauppi, 2019 for discussion).Liu et al’s analyses shed some light on what molecular mechanisms may be responsible for the reduced efficiency of X‐Y pairing and synapsis in Usp26‐deficient spermatocytes. USP26 codes for a deubiquitinating enzyme that has several substrates in the testis. Because USP26 prevents degradation of these substrates, their levels should be downregulated in Usp26 null testes. Liu et al (2021) show that USP26 interacts with TEX11, a protein required for stable pairing and normal segregation of the X and Y chromosomes in mouse meiosis (Adelman & Petrini, 2008). USP26 can de‐ubiquitinate TEX11 in vitro, and in Usp26 null testes, TEX11 was almost undetectable. It is worth noting that USP26 has several other known substrates, including the androgen receptor (AR), and therefore, USP26 disruption likely contributes to compromised spermatogenesis via multiple mechanisms. For example, AR signaling‐dependent hormone levels are misregulated in Usp26 null mice (Tian et al, 2019).The sex chromosome phenotypes observed in Usp26 null mice predict that men with USP26 mutations may be fertile, but producing XY aneuploid sperm at an abnormally high frequency, and that spermatogenic defects should increase with age (Fig 1). These predictions were testable, because the mutated USP26 haplotype, present in 13% of Klinefelter syndrome patients, was reasonably common also in fertile men (7–10%). Indeed, sperm XY aneuploidy was substantially higher in fertile men with the mutated USP26 haplotype than in those without USP26 mutations. Some mutation carriers produced > 4% aneuploid sperm. Moreover, age‐dependent oligospermia was also found associated with the mutated USP26 haplotype.Open in a separate windowFigure 1Mutated USP26 as genetic risk factor for age‐dependent X‐Y defects in spermatogenesisMouse genetics demonstrate that deleterious USP26 mutations lead to less‐efficient X‐Y pairing and recombination with advancing age. Concomitant decrease of spindle assembly checkpoint (SAC) protein levels leads to less‐efficient elimination of metaphase I spermatocytes that contain misaligned X and Y chromosomes. This allows for the formation of XY aneuploid sperm in older individuals and subsequently increased age‐dependent risk for fathering Klinefelter syndrome (KS) offspring, two correlates also observed in human USP26 mutation carriers. At the same time, oligospermia/subfertility also increases with advanced age in both Usp26‐deficient mice and USP26 mutation‐carrying men, tempering Klinefelter syndrome offspring risk but also decreasing fecundity.As indicated by its prevalence in the normal control population, the USP26 mutated haplotype is not selected against in the human population. With > 95% of sperm in USP26 mutation carriers having normal haploid chromosomal composition, the risk of producing (infertile) Klinefelter syndrome offspring remains modest, likely explaining why USP26 mutant alleles are not eliminated. Given that full Usp26 disruption barely affects fertility of male mice during their prime reproductive age (Felipe‐Medina et al, 2019; Tian et al, 2019; Liu et al, 2021), there is little reason to assume strong negative selection against USP26 variants in humans. USP26 as the first‐ever genetic risk factor predisposing to sperm X‐Y aneuploidy and paternal origin Klinefelter syndrome offspring in humans, as uncovered by Liu et al, may be just one of many. 90% of Liu et al’s Klinefelter syndrome cases were not associated with USP26 mutations. But even in the age of genomics, discovery of Klinefelter syndrome risk factors is not straightforward, since most sperm of risk mutation carriers will not be XY aneuploid and thus not give rise to Klinefelter syndrome offspring. In addition, as Usp26 null mice demonstrate, both genetic and non‐genetic modifiers impact on penetrance of the XY aneuploidy phenotype: Spermatogenesis in the absence of Usp26 was impaired in the DBA/2 but not the C57BL/6 mouse strain background (Sakai et al, 2019), and in older mice, there was substantial inter‐individual variation in the severity of the X‐Y defect (Liu et al, 2021). In human cohorts, genetic and non‐genetic modifiers are expected to blur the picture even more.Future identification of sex chromosome aneuploidy risk factors has human health implications beyond Klinefelter syndrome. Firstly, XXY incidence is not only relevant for Klinefelter syndrome livebirths—it also contributes to stillbirths and spontaneous abortions, at a 4‐fold higher rate than to livebirths (Thomas & Hassold, 2003). Secondly, persistent meiotic X‐Y defects can, over time, result in oligospermia and even infertility. Since the mean age of first‐time fathers is steadily rising and currently well over 30 years in many Western countries, age‐dependent spermatogenic defects will be of ever‐increasing clinical relevance.     

Fahrenheit 101

Is there any convincing explanation for why mammals and birds maintain their body temperature close to 40°C? Subject Categories: Ecology, Metabolism,

Having recently become very interested in the way organisms balance heat production against “useful work”, I have stumbled into an old‐chestnut question in biology. That is, why do birds and mammals maintain their body temperature close to 40°C—at least during periods of normal circadian activity—regardless of their size or their local environment? This question comes into even sharper focus when realizing that the vast majority of organisms on our planet do not do anything like this, and still thrive at temperatures anywhere between below the freezing and above the boiling point of water. Warm‐blooded animals as a whole represent only a fraction of a percent of the total biomass on Earth. Most animals live at or very close to environmental temperature. Moreover, birds and mammals inhabit similar environments to all those other organisms, ranging from the cool depths of the oceans to the dramatic temperature fluctuations of the world''s deserts. The male emperor penguin (Aptenodytes forsteri) overwinters on the landward side of the Antarctic sea ice, remaining almost motionless for months, while incubating his egg at air temperatures some 70–80°C cooler than that of his own body, without even factoring in the additional chill of frequent gale‐force winds, and then goes off foraging in the comparatively warm waters at the ice edge and beyond, maintaining all the time his body temperature at the same 38–39°C. The only significant deviation from the 40°C rule is the special case of hibernation and torpor, although the extent and frequency of the associated body temperature decrease(s) vary greatly between species.Whenever I have posed this question to colleagues or students or just about anyone, I invariably get a similar answer: that this must be the optimal temperature for life, for the stability of proteins; for the functionality of enzymes; for just the right amount of membrane fluidity to facilitate cell–cell signaling, endocytosis, exocytosis, and charge separation; and for the highest fidelity of nucleic acid and protein synthesis. Even if these assertions are true—and I am not aware of compelling evidence that they are—this flies in the face of the fact that the vast bulk of non‐homeothermic organisms are not to be found in the very few environments on the planet that come close to the range of avian and mammalian body temperatures. Instead, life seems concentrated in the cool soil and oceans, and in great forests found in equatorial, temperate, and even sub‐polar climes alike. The most biologically productive zone of the world''s oceans is not even in the tropics, but at the Antarctic convergence, which is teeming with animal, plant, and bacterial life. Moreover, if 40°C were so favorable, wouldn''t homeothermy geared to maintaining that temperature have evolved countless times, in different organisms? Or, for that matter, wouldn''t the proteins and membranes of birds and mammals have evolved so as to function better at temperatures closer to the global average of around 10–15°C, instead of wasting all that energy to stay warm? After all, if the Antarctic sea urchin (Sterechinus neumayerii) has adapted over a few tens of millions of years to live and develop perfectly well at −1.5°C (Foo et al, 2016), just more slowly than its close relatives in temperate or tropical waters, what''s stopping us from having turned down the heat by the same amount?I anticipate that you are now expecting me to reveal a brilliant idea to explain everything. But all I have to offer are new meanderings and wrong turnings. The first is the idea that, like the traditional explanation for our cellular and blood salinity being close to that of the ancient ocean, maybe that ancient ocean was stable at 40°C for most of evolutionary time. Except that it was not. It was closer to 80°C throughout at least the first half of our planet''s history, when the major cellular forms evolved (Knauth, 2005), and after that, it cooled down to below the magic 40°C and remained there ever since, albeit with large fluctuations. During the period when warm‐blooded creatures are believed to have arisen, the oceans never rose to 40°C, and in any case, this evolutionary step is generally assumed to have occurred on land, not in water.Do our own observations that mitochondria are 10–12°C warmer than the cells in which they reside (Chrétien et al, 2018) have any bearing on the question? Is this again some kind of happy medium, whereby the maximum efficiency of oxidative phosphorylation dictates a certain heat output that naturally maintains the cell’s temperature at around 40°C? But if this were the case, other eukaryotes that did not maintain any kind of internal thermoregulation at the whole‐organism level would also tend to be at 40°C, with their mitochondria at 50°C. Yet mitochondria in Drosophila cells are again about 10°C warmer than their surroundings, but at much cooler ambient temperatures (M. Terzioglu & H. T. Jacobs, unpublished observations). And prolonged exposure to 40°C represents a lethal heat shock for wild‐type Drosophila (Stefanou & Alahiotis 1982). Poikilotherms simply lose excess heat to the environment, whereas homeotherms must use specific mechanisms both to generate and to retain, additional heat when needed, and radiate excess heat to the outside, so as to maintain a constant internal temperature.Is there some other defining feature of mammalian and avian biology that might default body temperature to the observed constant? After much reflection, I cannot think of one. But perhaps it is possible to construe an argument in the opposite direction that having evolved mechanisms to maintain a constant body temperature, birds and mammals have, as argued elsewhere (Grigg et al, 2004), been able to colonize extremely diverse habitats, remain active at night, and perhaps resist mass extinction events driven by climate change a bit better than other taxa. But this did not help the dinosaurs. And it does not explain why 40°C is any better than 80 or 20°C, or why it is so evolutionarily stable, in birds of paradise as in penguins, or kangaroos as in polar bears.Perhaps one can cobble some argument together by combining the adaptive range and mitochondrial arguments; plus the fact that it is probably easier to envisage single mutations that can shift the balance between metabolic heat production and useful work, to maintain 40°C, as opposed to the many mutations required to re‐optimize biological processes for a different temperature. But I am not very convinced. Evolution normally mirrors environmental change, rather than resists it.If an intelligent insect were writing this column, they would no doubt herald the virtues of the arthropod lifestyle in being able to go with the climactic flow and not waste so much energy keeping warm or cool like all those primitive furry and feathered creatures, yearning for their balmy but non‐existent Eden. Maybe some reader out there will come up with a cute idea that will prove experimentally testable and eventually seem self‐evident, yet has escaped me and all others who have pondered this question. But I am now going out to frolic in the snow. Over to you.     

Membrane protein biogenesis by the EMC

Recent cryo‐EM‐based models reveal how the ER membrane protein complex may accomplish insertion of protein transmembrane domains with limited hydrophobicity.

Insertion of strongly hydrophobic TMDs into the ER membrane is mediated by the Sec61 complex for co‐translational insertion and the GET complex for post‐translational insertion of tail‐anchors (Volkmar & Christianson, 2020). By contrast, the EMC inserts TMDs of limited hydrophobicity, frequently located at the N‐ or C‐termini of proteins, and is involved in biogenesis of multi‐spanning membrane proteins (Volkmar & Christianson, 2020).The EMC is highly conserved (Wideman, 2015). In vertebrates, ten subunits have been identified (EMC1‐10), two of which, EMC8 and EMC9, are homologous and the result of a vertebrate‐specific gene duplication (Wideman, 2015). In Saccharomyces cerevisiae, EMC8 has been lost (Wideman, 2015). Only EMC3 displays clear homology to other membrane protein insertases, the Oxa1 family (Wideman, 2015; Volkmar & Christianson, 2020). This family includes YidC, which inserts TMDs into the bacterial cytoplasmic membrane, usually in cooperation with the Sec61‐homologous SecYEG channel (Volkmar & Christianson, 2020). Their association, along with the SecDF ancillary complex, forms a holo‐translocon capable of protein secretion and TMD insertion, with striking similarities to the EMC complex (Martin et al, 2019).Recent work by Pleiner et al (2020) presented a 3.4 Å cryo‐EM structure of the human EMC purified via a GFP‐tag on EMC2 and incorporated into a phospholipid nanodisc. The complex is formed by nine proteins (EMC1‐8, EMC10) (Pleiner et al, 2020). EMC8 and EMC9 are structurally similar, and their association with EMC2 is mutually exclusive (O''Donnell et al, 2020). Of the 12 TMDs, nine constitute the pseudosymmetric central ordered core, with a basket‐shaped cytosolic vestibule formed primarily by alpha‐helices of the EMC3 and EMC6 TMDs and cytosolic EMC2 (Fig 1A; Pleiner et al, 2020). The L‐shaped lumenal domain of the EMC consists mostly of beta‐sheets (Fig 1A; Pleiner et al, 2020), flanked by a conspicuous and conserved amphipathic alpha‐helix of EMC1 sealing the vestibule at the interface between the membrane and the ER lumen, together with another smaller amphipathic helix contributed by EMC3 (Fig 1A; Pleiner et al, 2020). In the ER lumen, the two 8‐bladed propellers of EMC1 contact six of the eight other subunits and stabilize the entire complex (Fig 1A; Pleiner et al, 2020). Beta‐sandwiches of EMC7 and EMC10 are anchored to the EMC1 lumenal domain (Fig 1A; Pleiner et al, 2020). In the cytosol, the tetratricopeptide repeat (TPR) spiral of EMC2 forms a cup underneath the partially hydrophilic vestibule in the membrane between the TMDs of EMC3 and EMC6, bridging the cytosolic ends of TMDs of EMC1, 3 and 5 (Fig 1A; Pleiner et al, 2020). Cytosolic EMC8 is bound to the opposite face of EMC2 (Fig 1A).Open in a separate windowFigure 1Comparison of the structures of human and yeast EMC(A) Cryo‐EM 3D map of the human (emdb‐21929) and yeast (emdb‐21587) EMC, showing front and back views with individual subunits coloured. Membrane position, obtained from the OPM database, is shown by grey discs. (B) Close‐up view of the EMC cavity formed by EMC3 and EMC6. Left, shown in a hydrophobicity surface pattern. Right, surface representation overlapped with the TMDs of EMC3 and EMC6. EMC4, flexible and with a gate function at the substrate‐binding place, is shown in pink in the yeast representation. EMC4 is not visible at the atomic EMC human structure, although is observed as a weak density at the human model, accompanied by TMs of EMC7 and EMC10 (Pleiner et al, 2020). (C) The yeast EMC following > 5 µs of CG‐MD simulation. The protein is shown as surface and coloured as per Pleiner et al (2020). The computed densities of waters and phospholipid tails and phosphates are shown as blue, yellow and lime green densities, sliced to bisect the cavity for clarity. Right, inset of the EMC cavity. Methods: CG‐MD simulations were built using PDB 6WB9 in a solvated symmetric POPC/POPE/cholesterol membrane and run in the Martini forcefield as described in Martin et al (2019). 3 µs unrestrained simulations were run, followed by 2.5 µs backbone restrained simulation for density calculation, done using VolMap in VMD (Humphrey et al, 1996).The 3.0 Å cryo‐EM structure of the yeast EMC presented by Bai and colleagues shows a very similar overall organization (Bai et al, 2020). Here, purification was via a 3xFLAG‐tag on EMC5, and the structure of the 8‐subunit complex (without EMC8/9) was visualized in detergent solution (Bai et al, 2020). The yeast complex has twelve TMDs like the human EMC, but unlike the human structure, EMC4 in yeast has three TMDs that are clearly visible (Bai et al, 2020). They are angled in the membrane pointing away from the complex at the cytosolic end (Fig 1A), and Bai et al (2020) propose that TMDs of EMC4, EMC3 and EMC6 form a substrate‐binding pocket similar to that of YidC. As in the human EMC, there are two amphipathic helices (EMC1 and EMC3) at the membrane/lumen interface (Fig 1A; Bai et al, 2020). In the ER lumen, yeast EMC1 only has one 8‐bladed beta‐propeller, to which the beta‐sandwiches of EMC7 and EMC10 are anchored (Fig 1A; Bai et al, 2020). In the cytosol, EMC2 bridges EMC3, 4 and 5, and its TPR repeats form a cup underneath the vestibule similar to human EMC2 (Fig 1A; Bai et al, 2020).The authors propose that insertion of a partially hydrophilic TMD by the yeast EMC is mechanistically similar to insertion by bacterial YidC (Bai et al, 2020). Yeast EMC is proposed to bind substrate between TMD2 of EMC3 and TMD2 of EMC4 in a pocket with polar and positively charged amino acids at either end and hydrophobic amino acids in the centre (Fig 1B; Bai et al, 2020). Much has been made of a conserved positive region within the EMC complex here, present in an equivalent position also in YidC (Kumazaki et al, 2014): It is claimed to be important for the incorporation of more‐hydrophilic TMDs and perhaps responsible for the “positive‐inside” orientation rule (von Heijne, 1992). Yeast and human EMC3 contain a specific R31 and R26 residue, respectively, conserved also in YidC and important for function of the EMC, as well as for YidC in Gram‐positive, but interestingly not Gram‐negative, bacteria (Chen et al, 2014; Pleiner et al, 2020; Bai et al, 2020). Another interesting feature, also conserved with YidC, is the flexibility of the TMDs flanking the substrate‐binding pocket, critical for EMC entry of substrates (Bai et al, 2020).In the human EMC, methionine residues in a cytosolic loop of EMC3 act as a substrate bait (Pleiner et al, 2020). Polar and charged residues within the substrate‐binding groove guide the lumenal domain across the membrane, facilitated by local membrane thinning (Pleiner et al, 2020; Fig 1B). The positive charges within the substrate‐binding site exclude signal peptides and enforce the “positive‐inside rule” (von Heijne, 1992; Pleiner et al, 2020). Flexible TMDs of EMC4, EMC7 and EMC10 forming a “lateral gate” of the substrate‐binding groove allow sampling of the bilayer by the substrate TMD (Pleiner et al, 2020). As the shortened TMDs of EMC3 and EMC6 cannot stably bind the substrate TMD, they favour its release into the bilayer (Pleiner et al, 2020). The EMC1 beta‐propeller(s) may recruit additional protein maturation factors in the ER lumen (Pleiner et al, 2020; Bai et al, 2020) or bind the Sec61 channel to allow cooperation between the two insertases (Bai et al, 2020).Arguably, the most interesting feature of the EMC complex is the location of a large interior cavity with distinctive hydrophilic character, which likely aids TMD insertion (Fig 1B). We ran a coarse‐grained molecular dynamics (CG‐MD) simulation of the yeast EMC structure, which highlights a profound perturbation of the phospholipid bilayer in the EMC interior cavity (Fig 1C). Here, a deep gorge forms in the cytoplasmic leaflet of the bilayer, allowing the cavity to become flooded with water (Fig 1C). Note the location of the lipid head groups here (lime green), which presumably define the site of amphipathic TMD insertion. The incursion of phospholipids into the centre of the EMC complex is a feature shared by the bacterial holo‐translocon (Martin et al, 2019) and perhaps by all membrane protein insertases. The shape and character of the EMC cavity presumably dictate its predisposition for less hydrophobic TMDs; it would be interesting to see whether the cavities of different insertases are similarly tailored to suit their substrates.     

Towards best practices in research: Role of academic core facilities

Academic Core Facilities are optimally situated to improve the quality of preclinical research by implementing quality control measures and offering these to their users. Subject Categories: Methods & Resources, Science Policy & Publishing

During the past decade, the scientific community and outside observers have noted a concerning lack of rigor and transparency in preclinical research that led to talk of a “reproducibility crisis” in the life sciences (Baker, ²⁰¹⁶; Bespalov & Steckler, ²⁰¹⁸; Heddleston et al, ²⁰²¹). Various measures have been proposed to address the problem: from better training of scientists to more oversight to expanded publishing practices such as preregistration of studies. The recently published EQIPD (Enhancing Quality in Preclinical Data) System is, to date, the largest initiative that aims to establish a systematic approach for increasing the robustness and reliability of biomedical research (Bespalov et al, ²⁰²¹). However, promoting a cultural change in research practices warrants a broad adoption of the Quality System and its underlying philosophy. It is here that academic Core Facilities (CF), research service providers at universities and research institutions, can make a difference.It is fair to assume that a significant fraction of published data originated from experiments that were designed, run, or analyzed in CFs. These academic services play an important role in the research ecosystem by offering access to cutting‐edge equipment and by developing and testing novel techniques and methods that impact research in the academic and private sectors alike (Bikovski et al, ²⁰²⁰). Equipment and infrastructure are not the only value: CFs employ competent personnel with profound knowledge and practical experience of the specific field of interest: animal behavior, imaging, crystallography, genomics, and so on. Thus, CFs are optimally positioned to address concerns about the quality and robustness of preclinical research.     

From the freezer to the clinic: Antifreeze proteins in the preservation of cells,tissues, and organs

Understanding the mechanisms by which natural anti‐freeze proteins protect cells and tissues from cold could help to improve the availability of donor organs for transplantation.

The first successful organ transplant in humans was performed in 1954 by Joseph Murray, who used a patient’s twin as a kidney donor. Murrays’ breakthrough paved the way for organ transplantation and the number of transplanted organs has grown ever since. For example, in 2017, a total of 139.024 solid organs—mostly kidney, liver, heart, lung, pancreas, and small bowel—were transplanted (Fig 1A). But this number only reflects 10% of the worldwide need; many patients still die of end‐stage organ failure while on a waiting list. The limited number of donor organs contributes only partially to this shortage. Many donor organs are not transplanted eventually owing to inefficient preservation techniques that shorten their extracorporeal lifetime. In fact, the percentage of donor organs that are unused is estimated to range from around 25% for kidneys and livers up to 70–80% for hearts and lungs (Giwa et al, 2017); Fig 1B).Open in a separate windowFigure 1Organ transplantation and preservability statusStatistics show a positive correlation between the duration of ex vivo preservation and the number of organ transplants. Number of solid organs transplanted in 2017 (A). Percentage of organs failed to be transplanted (B). Duration of solid organ ex vivo preservation in static cold storage (C). Sources: Data from the Global Observatory on Donation and Transplantation and (Parsons et al, 2014), (Guibert et al, 2011) and (Editorial: Buying time for transplants (2017))

Many donor organs are not transplanted eventually owing to inefficient preservation techniques that shorten their extracorporeal lifetime.

To address the shortage of donor organs and decrease the number of organs that go to waste, biobanks could efficiently store viable tissues and organs until transplantation. Yet, the current standard for ex vivo preservation of donor organs is static cold storage (4–8°C) which, depending on the organ, ensures viable conservation for only some hours; hearts are typically viable for a maximum of only 4 h (Fig 1C). In addition, this approach leads to hypothermic damage and to ischemia/reperfusion injury.Hence, there is an urgent need for strategies that prolong the viable preservation of donor organs. Two main strategies have emerged for cryopreservation and subzero storage, both of which cool tissues below the freezing point. While subzero storage just below 0°C may suffice for short‐term preservation, cryopreservation at −80°C or even lower temperatures is required for long‐term storage in biobanks. A proof‐of‐principle study already demonstrated that subzero preservation extended the preservation of rat hearts up to 24 h after collection (Amir et al, 2004); cryopreservation of whole hearts is currently not possible. The main reason is that lowering the temperature below the freezing point of water leads to ice formation, which causes cell damage and destroys tissues. One of the main challenges in biomedical research for organ transplantation is therefore finding non‐toxic and biocompatible antifreeze compounds that enable subzero storage and cryopreservation without causing tissue damage. An additional benefit is a larger time window to perform evaluation in terms of organ size and human leukocyte antigens matching and preparing the recipient patient to increase the chance of a successful transplantation.