Wednesday, July 05, 2023

Invasive non-native species cost UK economy an estimated £4bn a year, new CABI-led study reveals


CABI scientists have carried out a study which reveals invasive non-native species (INNS) – such as the aquatic water weeds floating pennywort and Japanese knotweed as well as signal crayfish – cost the UK economy an estimated £4bn a year.

Peer-Reviewed Publication

CABI

Floating pennywort on the River Wey, Weybridge, UK 

IMAGE: FLOATING PENNYWORT ON THE RIVER WEY, WEYBRIDGE, UK view more 

CREDIT: DJAMI DJEDDOUR CABI



CABI scientists have carried out a study which reveals invasive non-native species (INNS) – such as the aquatic water weeds floating pennywort and Japanese knotweed as well as signal crayfish – cost the UK economy an estimated £4bn a year.

However, when species only covered by the GB Non-native Species Strategy are considered – for instance with fungi excluded from the estimate – the total cost was estimated to be £1.9bn.

Researchers working from CABI’s centres in Egham, UK, as well as Switzerland and Kenya, found a 135% increase in comparable costs since the last assessment was conducted in 2010. Annual estimated costs in 2021 were £3.02bn, £499m, £343m and £150m to England, Scotland, Wales and Northern Ireland respectively.

The cost to forestry increased eightfold, the cost to aquaculture and agriculture increased by 139.5% and 112.7%, respectively, and the cost of most of the other sectors increased roughly in line with inflation (47.6% for GB and 55.7% for Northern Ireland).

Agriculture is the industry affected the most with estimated costs for the UK put at £1.088bn followed by construction, development and infrastructure at £270m and tourism and recreation at £136m. The impact upon forestry is £123m.

The study, published in the journal Biological Invasions, updates the earlier assessment using the same methodology and the diversity of changes among sectors and species highlights the value of such a detailed approach.

There are currently around 2,000 INNS in the UK with 10-12 new species establishing themselves every year. The list includes well-known established species such as grey squirrel, killer shrimp, giant hogweed, mink and parakeets, as well as recently arrived, but highly impactful species such as the sea squirt Didemnum vexillum and ash dieback.

The fungus Hymenoscyphus fraxineus, which causes ash dieback disease has become the costliest species in the past decade in the UK at an estimated £883.5m followed by followed by Japanese knotweed (£246.5m), rabbits (£169.7m), rats and mice (£84.4m), cockroaches (£69.8m) and deer (£62.9m).

As a group, fungi were the costliest to the UK, accounting for 52.9% of the total estimated costs, followed by mammals, plants and terrestrial arthropods (21.9%, 15.5% and 7.5% of the total, respectively).

Dr Rene Eschen, lead author and Senior Scientist, Ecosystems Management, said, “Our research illustrates the usefulness of repeating economic cost assessments for INNS, as INNS are dynamic and their impacts vary.

“Repeat assessments like this one are important to maintain a focus on the impact of INNS, changes in impacts as a result of new or spreading species, as well as the identification of potential impacts of management or policies.”

The researchers recommend continued investment in sustainable, long-term solutions for widespread damaging species, such as classical biological control, which, they say, has been shown worldwide to be a cost-effective, safe and environmentally sensitive management option when other methods prove ineffective or are no longer feasible.

Dr Richard Shaw, co-author and Senior Regional Director, Europe and The Americas, said, “This assessment again shows the important costs of INNS to the UK economy. Few effects of INNS specific management efforts can be seen in these results. However, they highlight the need to continue prevention and early detection, followed by eradication of the highest-risk species prior to establishment.”

In February, the GB Invasive Non-native Species Strategy, which draws upon CABI’s research, was published to provide a strategic framework within which the actions of government departments, their related bodies and key stakeholders can be better co-ordinated.

Defra Head of GB Non-Native Secretariat, Niall Moore, said: “Invasive Non-Native species pose a serious threat to our natural environment and this Government is taking action through the recently launched GB Invasive Non-Native Species strategy, to protect our native animals and plants from INNS.

“CABI’s research, funded by Defra, reveals the significant financial impact of INNS. It is vital that we work together with researchers, scientists, and others, who are working to tackle INNS, to prevent their entry into and establishment in Great Britain and, when they do become established, to mitigate their negative impacts.”

Additional information

Main image: Floating pennywort is one invasive non-native species of concern. The aquatic weed causes dense mats that cover the water’s surface – such as here on this water course on the River Wey, Weybridge, UK (Credit: Djami Djeddour).

Full paper reference

RenĂ© Eschen, Mariam Kadzamira, Sonja Stutz, Adewale Ogunmodede, Djami Djeddour, Richard Shaw, Corin Pratt, Sonal Varia, Kate Constantine and Frances Williams, ‘An updated assessment of the direct costs of invasive non-native species to the United Kingdom,’ 6 July 2023, Biological Invasions, DOI: 10.1007/s10530-023-03107-2

The paper can be read in full open access from 1am UK Time 6 July 2022 here: https://link.springer.com/article/10.1007/s10530-023-03107-2

 

Acknowledgement

This work was funded by the UK Department for Environment Food and Rural Affairs (Defra) under contract #34247. CABI is an international intergovernmental organisation, and we gratefully acknowledge the core financial support from our member countries (and lead agencies) including the United Kingdom (Foreign, Commonwealth & Development Office), China (Chinese Ministry of Agriculture and Rural Affairs), Australia (Australian Centre for International Agricultural Research), Canada (Agriculture and Agri-Food Canada), Netherlands (Directorate-General for International Cooperation), and Switzerland (Swiss Agency for Development and Cooperation). See https://www.cabi.org/about-cabi/who-we-work-with/key-donors/ for full details.

 

New genetic technology developed to halt malaria-spreading mosquitoes


As envisioned, first-of-its-kind African mosquito suppression system would reduce child mortality and aid economic development

Peer-Reviewed Publication

UNIVERSITY OF CALIFORNIA - SAN DIEGO

Malaria Suppression System 

IMAGE: UC SAN DIEGO RESEARCHERS HAVE DEVELOPED A NEW TECHNOLOGY TO SUPPRESS ANOPHELES GAMBIAE, THE MOSQUITOES THAT PRIMARILY SPREAD MALARIA IN AFRICA AND CONTRIBUTE TO ECONOMIC POVERTY IN AFFECTED REGIONS. view more 

CREDIT: AKBARI LAB, UC SAN DIEGO




Malaria remains one of the world’s deadliest diseases. Each year malaria infections result in hundreds of thousands of deaths, with the majority of fatalities occurring in children under five. The Centers for Disease Control and Prevention recently announced that five cases of mosquito-borne malaria were detected in the United States, the first reported spread in the country in two decades.

Fortunately, scientists are developing safe technologies to stop the transmission of malaria by genetically editing mosquitoes that spread the parasite that causes the disease. Researchers at the University of California San Diego led by Professor Omar Akbari’s laboratory have engineered a new way to genetically suppress populations of Anopheles gambiae, the mosquitoes that primarily spread malaria in Africa and contribute to economic poverty in affected regions. The new system targets and kills females of the A. gambiae population since they bite and spread the disease.

Publishing July 5 in the journal Science Advances, first-author Andrea Smidler, a postdoctoral scholar in the UC San Diego School of Biological Sciences, along with former master’s students and co-first authors James Pai and Reema Apte, created a system called Ifegenia, an acronym for “inherited female elimination by genetically encoded nucleases to interrupt alleles.” The technique leverages the CRISPR technology to disrupt a gene known as femaleless (fle) that controls sexual development in A. gambiae mosquitoes.

Scientists at UC Berkeley and the California Institute of Technology contributed to the research effort.

Ifegenia works by genetically encoding the two main elements of CRISPR within African mosquitoes. These include a Cas9 nuclease, the molecular “scissors” that make the cuts and a guide RNA that directs the system to the target through a technique developed in these mosquitoes in Akbari’s lab. They genetically modified two mosquito families to separately express Cas9 and the fle-targeting guide RNA.

“We crossed them together and in the offspring it killed all the female mosquitoes,” said Smidler, “it was extraordinary.” Meanwhile, A. gambiae male mosquitoes inherit Ifegenia but the genetic edit doesn’t impact their reproduction. They remain reproductively fit to mate and spread Ifegenia. Parasite spread eventually is halted since females are removed and the population reaches a reproductive dead end. The new system, the authors note, circumvents certain genetic resistance roadblocks and control issues faced by other systems such as gene drives since the Cas9 and guide RNA components are kept separate until the population is ready to be suppressed.

“We show that Ifegenia males remain reproductively viable, and can load both fle mutations and CRISPR machinery to induce fle mutations in subsequent generations, resulting in sustained population suppression,” the authors note in the paper. “Through modeling, we demonstrate that iterative releases of non-biting Ifegenia males can act as an effective, confinable, controllable and safe population suppression and elimination system.”

Traditional methods to combat malaria spread such as bed nets and insecticides increasingly have been proven ineffective in stopping the disease’s spread. Insecticides are still heavily used across the globe, primarily in an effort to stop malaria, which increases health and ecological risks to areas in Africa and Asia.

Smidler, who earned a PhD (biological sciences of public health) from Harvard University before joining UC San Diego in 2019, is applying her expertise in genetic technology development to address the spread of the disease and the economic harm that comes with it. Once she and her colleagues developed Ifegenia, she was surprised by how effective the technology worked as a suppression system.

“This technology has the potential to be the safe, controllable and scalable solution the world urgently needs to eliminate malaria once and for all,” said Akbari, a professor in the Department of Cell and Developmental Biology. “Now we need to transition our efforts to seek social acceptance, regulatory use authorizations and funding opportunities to put this system to its ultimate test of suppressing wild malaria-transmitting mosquito populations. We are on the cusp of making a major impact in the world and won’t stop until that’s achieved.”   

The researchers note that the technology behind Ifegenia could be adapted to other species that spread deadly diseases, such as mosquitoes known to transmit dengue (break-bone fever), chikungunya and yellow fever viruses.

The full author list includes Andrea Smidler, James Pai, Reema Apte, Hector Sanchez C., Rodrigo Corder, Eileen Jeffrey Gutierrez, Neha Thakre, Igor Antoshechkin, John Marshall and Omar Akbari.

Fossils reveal how ancient birds molted their feathers— which could help explain why ancestors of modern birds survived when all the other dinosaurs died



Peer-Reviewed Publication

FIELD MUSEUM

Feathers in amber 

IMAGE: FEATHERS FROM A BABY BIRD THAT LIVED 99 MILLION YEARS AGO, PRESERVED IN AMBER. view more 

CREDIT: PHOTO BY SHUNDONG BI.




Every bird you’ve ever seen— every robin, every pigeon, every penguin at the zoo— is a living dinosaur. Birds are the only group of dinosaurs that survived the asteroid-induced mass extinction 66 million years ago. But not all the birds alive at the time made it. Why the ancestors of modern birds lived while so many of their relatives died has been a mystery that paleontologists have been trying to solve for decades. Two new studies point to one possible factor: the differences between how modern birds and their ancient cousins molt their feathers.

Feathers are one of the key traits that all birds share. They're made of a protein called keratin, the same material as our fingernails and hair, and birds rely on them to fly, swim, camouflage, attract mates, stay warm, and protect against the sun’s rays. But feathers are complex structures that can’t be repaired, so as a means of keeping them in good shape, birds shed their feathers and grow replacements in a process called molting. Baby birds molt in order to lose their baby feathers and grow adult ones; mature birds continue to molt about once a year.

“Molt is something that I don't think a lot of people think about, but it is fundamentally such an important process to birds, because feathers are involved in so many different functions,” says Jingmai O’Connor, associate curator of fossil reptiles at Chicago’s Field Museum. “We want to know, how did this process evolve? How did it differ across groups of birds? And how has that shaped bird evolution, shaped the survivability of all these different clades?” Two of O’Connor’s recent papers examine the molting process in prehistoric birds. 

A paper in the journal Cretaceous Research published in May 2023 detailed the discovery of a cluster of feathers preserved in amber from a baby bird that lived 99 million years ago.

Today, baby birds are on a spectrum in terms of how developed they are when they're born and how much help they need from their parents. Altricial birds hatch naked and helpless; their lack of feathers means that their parents can more efficiently transmit body heat directly to the babies’ skin. Precocial species, on the other hand, are born with feathers and are fairly self-sufficient. 

All baby birds go through successive molts— periods when they lose the feathers they have and grow in a new set of feathers, before eventually reaching their adult plumage. Molting takes a lot of energy, and losing a lot of feathers at once can make it hard for a bird to keep itself warm. As a result, precocial chicks tend to molt slowly, so that they keep a steady supply of feathers, while altricial chicks that can rely on their parents for food and warmth undergo a “simultaneous molt,” losing all their feathers at roughly the same time.

The amber-preserved feathers in this study are the first definitive fossil evidence of juvenile molting, and they reveal a baby bird whose life history doesn’t match any birds alive today. “This specimen shows a totally bizarre combination of precocial and altricial characteristics,” says O’Connor, who was the first author of the paper alongside senior author Shundong Bi of the Indiana University of Pennsylvania. “All the body feathers are basically at the exact same stage in development, so this means that all the feathers started growing simultaneously, or near simultaneously.” However, this bird was almost certainly part of a now-extinct group called the Enantiornithines, which O’Connor’s previous work has shown were highly precocial. 

O’Connor hypothesizes that the pressures of being a precocial baby bird that had to keep itself warm, while undergoing a rapid molt, might have been a factor in the ultimate doom of the Enantiornithines. “Enantiornithines were the most diverse group of birds in the Cretaceous, but they went extinct along with all the other non-avian dinosaurs,” says O’Connor. “When the asteroid hit, global temperatures would have plummeted and resources would have become scarce, so not only would these birds have even higher energy demands to stay warm, but they didn’t have the resources to meet them.” 

Meanwhile, an additional study published July 3 in Communications Biology by O’Connor and Field Museum postdoctoral researcher Yosef Kiat examines molting patterns in modern birds to better understand how the process first evolved. 

In modern adult birds, molting usually happens once a year in a sequential process, in which they replace just a few of their feathers at a time over the course of a few weeks. That way, they're still able to fly throughout the molting process. Simultaneous molts in adult birds, in which all the flight feathers fall out at the same time and regrow within a couple weeks, are rarer and tend to show up in aquatic birds like ducks that don’t absolutely need to fly in order to find food and avoid predators. 

It’s very rare to find evidence of molting in fossil birds and other feathered dinosaurs, and O’Connor and Kiat wanted to know why. “We had this hypothesis that birds with simultaneous molts, which occur in a shorter duration of time, will be less represented in the fossil record,” says O’Connor— less time spent molting means fewer opportunities to die during your molt and become a fossil showing signs of molting. To test their hypothesis, the researchers delved into the Field Museum’s collection of modern birds. 

“We tested more than 600 skins of modern birds stored in the ornithology collection of the Field Museum to look for evidence of active molting,” says Kiat, the first author of the study. “Among the sequentially molting birds, we found dozens of specimens in an active molt, but among the simultaneous molters, we found hardly any.”

While these are modern birds, not fossils, they provide a useful proxy. “In paleontology, we have to get creative, since we don’t have complete data sets. Here, we used statistical analysis of a random sample to infer what the absence of something is actually telling us,” says O’Connor. In this case, the absence of molting fossil birds, despite active molting being so prevalent in the sample of modern bird specimens, suggests that fossil birds simply weren’t molting as often as most modern birds. They may have undergone a simultaneous molt, or they may not have molted on a yearly basis the way most birds today do. 

Both the amber specimen and the study of molting in modern birds point to a common theme: prehistoric birds and feathered dinosaurs, especially ones from groups that didn’t survive the mass extinction, molted differently from today’s birds. 

“All the differences that you can find between crown birds and stem birds, essentially, become hypotheses about why one group survived and the rest didn’t,” said O’Connor. “I don't think there's any one particular reason why the crown birds, the group that includes modern birds, survived. I think it's a combination of characteristics. But I think it's becoming clear that molt may have been a significant factor in which dinosaurs were able to survive.”

  

Illustration of what a newly hatched Enantiornithine bird may have looked like.

CREDIT

Image by Yu Chen and Shundong Bi.

An illustration of a more mature juvenile Enantiornithine.

Tracking ships’ icy paths amidst climate change


Peer-Reviewed Publication

MICHIGAN STATE UNIVERSITY

Breaking through Arctic ice 

IMAGE: RV SIKULIAQ, A RESEARCH VESSEL OWNED BY THE NATIONAL SCIENCE FOUNDATION, BREAKS THROUGH ARCTIC ICE. view more 

CREDIT: PHOTO BY CHRISTINA GOETHEL OF THE UNIVERSITY OF MARYLAND CENTER FOR ENVIRONMENTAL SCIENCES.




There has been much buzz about the warming planet’s melting Arctic region opening shipping routes and lengthening travel seasons in ocean passageways that ice once blocked. Expanded fishing, trade and tourism is envisioned.

Operative word: Envisioned.

Scientists at Michigan State University (MSU), University of Waterloo, and University of Alaska Fairbanks report in Climatic Change where vessels are traveling in the ice-covered waters of the Arctic between Alaska and Russia, and what those reports may mean for important wildlife and communities in the region.

“Even with climate change, sea ice is still a substantial barrier to Arctic vessel traffic,” said Kelly Kapsar, a research associate at MSU’s Center for Systems Integration and Sustainability (CSIS). “Sea ice also provides critical habitat for many endemic Arctic species and a hunting platform for Indigenous subsistence hunters. Understanding when and where ships are entering areas of sea ice can help us to better understand potential impacts of vessel traffic in the region.”

Whether its fishing vessels seeking better catches over a longer season, or Russian shipping companies eager for better ways to deliver oil and gas to Chinese customers, increased marine traffic is a given. Whether this traffic occurs only in the open water season, or also in times of ice cover is not.

But the researchers point out the difference between what ships could do as ice changes, and what they will do can be vastly different.

“Up until now projections have been about theoretical ships, such as noting certain vessel types can travel through up to 2 meters of ice,” Kapsar said. “But that’s like saying a car can drive up to 200 mph – just because it can doesn’t mean it will.”

Combining satellite pictures of ice cover with GPS vessel tracking data the team was able to analyze how the ships have been behaving as the shipping passages change. What they’ve found is that many ships are following the ice, fishing close to the edge of ice packs. The researchers also found marked overlap between areas with vessels traveling in sea ice and the overwintering areas for bowhead whales.

Previous research by another group has demonstrated that between 1990 and 2012, some 12% of bowhead whales harvested by Alaska Native subsistence hunters showed signs they had been tangled in fishing gear, and 2% had scars from being struck by vessels. The new analysis points to a growing threat to wildlife which also are using the receding ice as they travel and breed.

Noise from large boats also can disrupt marine mammals. Ships equipped to break ice potentially could strand both animals and people traveling across the frozen expanses. Increased traffic also raises fear of accidents and oil spills. The new pathways are far away from rescue or clean-up crews.

So far, Kapsar said, their work indicates ship travel reflects a certain caution, offering indications that capability is balanced by practical and economic realities. For now.

Kapsar and co-author Jianguo “Jack” Liu are members of MSU’s Ecology, Evolution, and Behavior Program. “Mapping vessel traffic patterns in the ice-covered waters of the Pacific Arctic” also was written by Lawson Brigham and Grant Gunn. The work is funded by the National Science Foundation.

Study shines light on why companies use a variety of dark money strategies

Peer-Reviewed Publication

UNIVERSITY OF TEXAS AT AUSTIN




AUSTIN, Texas — As public concerns mount over lack of transparency in political giving, a new study from researchers at The University of Texas at Austin is the first to illuminate how and why corporations choose to legally conceal their lobbying and campaign contributions.

U.S. companies are required to disclose the total amount they spend on political activity, but beyond that, the disclosure is incredibly vague, according to Tim Werner, associate professor of business, government and society at the McCombs School of Business.

“We don’t even know which individual members of Congress they meet with,” Werner said. “If you’re only looking at observable political activities of corporations, it’s like looking at the tip of the iceberg.”

Below the waterline, Werner and colleagues, Nan Jia of the University of Southern California and Stanislav Markus of the University of South Carolina, found that when corporations conceal their political activities, they may use a wide variety of strategies, including:

  • Lobbying people who are not obvious political players.
  • Secretly creating a “citizens coalition” to advocate for their position.
  • Using obfuscation to spin an issue in their favor.
  • Contributing to groups that are not legally required to report their donors, known as “dark money” groups.

The research is published in Academy of Management.

How do corporations decide when to use these strategies? The researchers developed a mathematical model of a company’s political decision-making to help predict which activities it’s most likely to hide. Their calculus finds:

  • The lower the costs of concealment or of being caught, the more likely companies are to attempt it.
  • Companies are more likely to conceal their activities on issues where they reap most of the benefits or bear most of the costs.
  • The harder it is to win a lawmaker’s support, the more a company will try to hide its spending to influence that lawmaker.

Werner said this line of research opens an important conversation.

“The more we understand about the reasons a business does or does not conceal its political involvement, the more informed our decision-making around policies related to transparency,” he said.

Read the McCombs Big Ideas story.

Health professions requiring advanced degrees have few Latinos


New study shows that Mexican Americans are the most underrepresented Latino group in occupations such as doctor and dentist

Peer-Reviewed Publication

GEORGE WASHINGTON UNIVERSITY

Hispanic Physician Current Workforce, 2010-2019 (National) 

IMAGE: HEALTH WORKFORCE DIVERSITY TRACKER DATA ON HISPANIC PHYSICIAN CURRENT WORKFORCE, 2010-2019 (NATIONAL) view more 

CREDIT: FITZHUGH MULLAN INSTITUTE FOR HEALTH WORKFORCE EQUITY. HEALTH WORKFORCE DIVERSITY TRACKER. WASHINGTON, DC: GEORGE WASHINGTON UNIVERSITY, 2021



WASHINGTON (July 5, 2023)--Although the situation is improving, Latinos and especially Mexican Americans, remain very underrepresented in U.S. health professions that require advanced degrees, according to a study published today in the journal Health Affairs. The study by George Washington University researchers is the first to examine the representation of the four largest Latino populations in the U.S. health workforce and the findings raise concerns about the lack of diversity in the U.S. health workforce.

The study revealed that Mexican Americans, despite being the largest Latino subpopulation in the United States, are greatly underrepresented in the health professions that require an advanced degree. One out of ten people in the United States are Mexican American yet in five of the eight professions requiring advanced education included in the study, Mexican Americans represent one-quarter or less of the professions. This study suggests they and other Latinos in the study face many barriers to occupations in the healthcare field that require an advanced degree such as doctor, nurse or pharmacist.

“Our analysis did not address why Latino representation was low in the health professions requiring advanced degrees,” Indira Islas, who conducted the study while a graduate student at the GW Milken Institute School of Public Health, said. “Other evidence suggests that the findings can be attributed to obstacles such as structural racism.”

Islas and her colleagues at the GW Fitzhugh Mullan Institute for Health Workforce Equity used data from the 2020 American Community Survey, which includes information on ethnicity collected by the Census Bureau. The team compared the representation of non-Latinos and four Latino subgroups, including Mexican Americans, Puerto Ricans, Cuban Americans and other Latinos.

The research showed a surprising finding for one subpopulation. Compared to other Latino groups, Cuban Americans are well-represented in the advanced degree health professions. For example their representation among physicians and dentists is greater than their numbers in the overall population.

The study also found that Mexican Americans, Puerto Ricans and other Latinos were overrepresented in most healthcare support occupations, low-wage jobs such as medical or dental assistants or home health aides.

Latino representation in the health workforce did show improvement during the time frame studied from 2016-2020: For example, Latino representation among recent graduates went from 6.7 to 13.5% for health professions requiring a bachelor’s degree. However, the researchers note that Latinos represented more than 21% of the population ages 20-35.

“There has been significant progress in getting more Latinos into the advanced health professions over the last few years,” Edward Salsberg, Co-Director of the Health Workforce Diversity Initiative at the Mullan Institute, said. “Yet as we can see from this study, much more needs to be done to strengthen and diversify the health workforce in the U.S. A lack of diversity among the health professions can exacerbate health disparities for Latinos and other minority populations.”

Islas, whose family came to the U.S. when she was a child, has experienced some of the obstacles to getting an advanced degree first hand. 

Her parents, who were doctors in Mexico, had to find work in factory jobs when the family moved to the U.S. “I remember in high school we didn’t have a lot of resources,” Islas said. But Islas had a dream of becoming a doctor like her parents and she ended up getting a scholarship and went to college. In May of last year, she graduated from GW with a Masters of Public Health. She is now doing a fellowship with the Congressional Hispanic Caucus Institute with plans to apply to medical school in the fall.

The paper, Documenting Latino Representation in the U.S. Health Workforce, appears in the July issue of Health Affairs. The research was funded by the California Endowment and the Josiah Macy Foundation.

For additional data on Latino representation in the health professions including by state and by graduates of individual schools, visit: The Health Workforce Diversity Tracker.

-GW-