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Friday, 31 August 2018
Major challenges of integrating agriculture into climate change mitigation policy frameworks.
Mitig Adapt Strateg Glob Chang. 2018;23(3):451-468. doi: 10.1007/s11027-017-9743-2. Epub 2017 Apr 12. Fellmann T1, Witzke P2, Weiss F3, Van Doorslaer B1, Drabik D4, Huck I1, Salputra G1, Jansson T5, Leip A3. Author information 1 European Commission, Joint Research Centre, Directorate Sustainable Resources, Seville, Spain. 2 EuroCARE, Bonn, Germany. 3 European Commission, Joint Research Centre, Directorate Sustainable Resources, Ispra, Italy. 4 4Agricultural Economics and Rural Policy Group, Wageningen University, Wageningen, The Netherlands. 5 5Department of Economics, Swedish University of Agricultural Sciences, Uppsala, Sweden. Abstract Taking the European Union (EU) as a case study, we simulate the application of non-uniform national mitigation targets to achieve a sectoral reduction in agricultural non-carbon dioxide (CO2) greenhouse gas (GHG) emissions. Scenario results show substantial impacts on EU agricultural production, in particular, the livestock sector. Significant increases in imports and decreases in exports result in rather moderate domestic consumption impacts but induce production increases in non-EU countries that are associated with considerable emission leakage effects. The results underline four major challenges for the general integration of agriculture into national and global climate change mitigation policy frameworks and strategies, as they strengthen requests for (1) a targeted but flexible implementation of mitigation obligations at national and global level and (2) the need for a wider consideration of technological mitigation options. The results also indicate that a globally effective reduction in agricultural emissions requires (3) multilateral commitments for agriculture to limit emission leakage and may have to (4) consider options that tackle the reduction in GHG emissions from the consumption side. KEYWORDS: Agriculture; Climate change; Emissions; Mitigation; Policy PMID: 30093833 PMCID: PMC6054014 DOI: 10.1007/s11027-017-9743-2 Free PMC Article
I Worked With Avital Ronell. I Believe Her Accuser.
THE CHRONICLE REVIEW https://www.chronicle.com/article/I-Worked-With-Avital-Ronell-I/244415 Avital Ronell By Andrea Long Chu AUGUST 30, 2018 The humanities are ablaze. This month The New York Times reported that the Title IX office had found Avital Ronell, a professor of German and comparative literature at New York University and a superstar in literary studies, responsible for sexually harassing a former student, Nimrod Reitman, now a visiting fellow at Harvard. A lawsuit filed by Reitman fills in the details. Leading feminist and queer scholars like Judith Butler, Lisa Duggan, and Jack Halberstam have defended her — or at least deflected criticism. I believe the allegations. Last year I worked as a teaching assistant for Avital Ronell. I hadn't sought out the appointment; I am a doctoral student in comparative literature at NYU, and that semester I was, per the handbook, guaranteed a teaching job. A few months before the position began, I received an email from one of my professors informing me that Ronell's other teaching assistants were "all taking her class and working hard to familiarize themselves with her particular methodologies, texts, style, and so on." I was "encouraged" to do the same. I was told this was "an important part of the process with Prof. Ronell." After all, there were other students eager to replace me. This was not abusive, obviously, only irritating. The lightly mobbish tone of the email — "this is a nice job you got here, shame if something happened to it" — was jarring. In theory, at least, teaching assistants are junior colleagues, not employees, and I had thought that my position was guaranteed. Then again, given the things I'd heard about Avital from other graduate students over the years, I wasn't all that surprised. (Except on formal occasions, she always went by that one name, "Avital," like Plato, or Cher.) It is simply no secret to anyone within a mile of the German or comp-lit departments at NYU that Avital is abusive. Eventually I kissed the ring. I attended a session of Avital's seminar and visited her in office hours. Her manner in that meeting was odd, wounded. "I just wanted to make sure you and I are OK," she told me with a concerned look, as if we were recovering from a nasty fight. "Of course we are!" I exclaimed, tripping over myself to reassure her. Avital softened. She had just wanted to be coaxed, like a deer to a salt lick. She smiled. I smiled. This was the process. The course was called "Outrageous Texts." Like most purportedly edgy things, it was less edgy than it imagined. In practice, outrageous mostly meant some dead white dudes with weird sexual hang-ups. Sometimes we mixed it up; the dudes were still alive. When we did read women (four of the 15 writers assigned), Avital still mostly talked about men. Her lecture on Valerie Solanas's SCUM Manifesto, like the introduction she wrote for Verso's edition of that book, focused on Nietzsche and Derrida. It is not illegal to read men. Avital is a Germanist and a deconstructionist who has made no serious contribution to feminist scholarship. That's fine. But when news media report that she is a feminist — "What Happens to #MeToo When a Feminist Is the Accused?" read the Times headline — they are factually mistaken. This is a professional distinction, not a political one. Personally, Avital may be a feminist, in the Taylor Swift sense of a woman who doesn't like being oppressed, but professionally, she is not a feminist scholar, any more than every person who believes that humans descended from apes is an evolutionary anthropologist. In class, Avital was waited on by her aide-de-camp, a graduate student who followed her around the Village like Tony Hale on HBO's Veep. If the energy in the room was not to her liking, she became frustrated. During one session, she abruptly stopped the lecture midthought, blaming her students for making her feel drained. It took a beat for anyone to realize she was serious. We were sent on a 15-minute break. That afternoon, quite without knowing it, like burrs attaching themselves to some passing animal, the students had been persecuting Avital. As far as I could tell (given that we had no prior relationship), I had done the same when I had failed, during my coursework years, to give much thought to her at all. It is simply no secret to anyone within a mile of the German or comp-lit departments at NYU that Avital is abusive. This is boring and socially agreed upon, like the weather. Stories about Avital's "process" are passed, like notes in class, from one student to the next: how she reprimanded her teaching assistants when they did not congratulate her for being invited to speak at a conference; how she requires that her students be available 24/7; how her preferred term for any graduate student who has fallen out of favor is "the skunk." Process: Wild things live in this word. These stories come from sources who strongly wished to remain anonymous, fearing that to have their names attached would threaten their chances in an already desiccated job market. But even if this was just gossip, I would believe it. When it comes to the American academy, I trust raw, red rumor over public statements any day of the week. Academic celebrity soaks up blood like a pair of Thinx. A letter to NYU's president, Andrew Hamilton, a draft of which leaked in June, argued that Avital's "brilliant scholarship" qualified her for special treatment. The 51 signatories included giants of feminist theory like Judith Butler and Gayatri Spivak, as well as my department chair — and the professor who emailed to "encourage" me to play nice with Avital. (Butler has since issued some tepid regrets.) Meanwhile, on social media and on their blog, the queer-studies scholars Lisa Duggan and Jack Halberstam dismissed the blowback against Avital as neoliberalism meets sex panic meets culture clash, straight people apparently being unable to decipher the coded queer intimacy of emails like "I tried to call you a number of times, unfortunately couldn't get through, would have liked to leave a msg" [sic]. That Avital's defenders are left-wing academic stars is not particularly surprising if you've spent much time in the academy. The institution has two choices when faced with political radicals: Ax them, especially if they are graduate students, or promote them. Make them successful, give them awards, power, enormous salaries. That way, when the next scandal comes along — and it will — they will have a vested interest in playing defense. This is how institutionality reproduces. Even the call to think critically about power becomes a clever smoke screen. There is a whole dissertation to be written on intellectuals using the word neoliberal to mean "rules I shouldn't have to follow." "If we focus on this one case, these details, this accuser and accused, we will miss the opportunity to think about the structural issues," wrote Duggan. This was code. It meant, "You can talk about structural issues all you want, so long as you don't use examples of people we know." You cannot have a cycle of abuse without actually existing abusers. In a milquetoast take for The New Yorker, Masha Gessen applauded Duggan as a model of "academics doing their job: engaging with things in great complexity." Of course power is messy. But there is no complexity in studying forests if you can't recognize a tree from a few feet away. This is not wisdom; it is an eye complaint. Structural problems are problems because real people hurt real people. You cannot have a cycle of abuse without actually existing abusers. That sounds simple, which is why so many academics hate it. When scholars defend Avital — or "complicate the narrative," as we like to say — in part this is because we cannot stand believing what most people believe. The need to feel smarter is deep. Intelligence is a hungry god. In this way, Avital's case has become a strange referendum on literary study. Generations of scholars have been suckled at the teat of interpretation: We spend our days parsing commas and decoding metaphors. We get high on finding meaning others can't. We hoard it, like dragons. We would be intellectually humiliated to learn that the truth was plain: that Avital quite simply sexually harassed her student, just as described. Sometimes analysis is simply denial with more words. Sometimes, as a frustrated student in a first-year literature course always mutters, the text just means what it says it means. My department has been largely silent since the news broke. Faculty members have said nothing to us. Upset and ashamed, my fellow graduate students and I speak with one another cautiously. We heal, or don't, alone. People I know are afraid to make any public comment, even on Facebook, where they are friends with older, richer scholars who might one day control their fates. Even I, who have by extraordinary luck options outside of academia, fear what being vocal will bring. A culture of critics in name only, where genuine criticism is undertaken at the risk of ostracism, marginalization, retribution — this is where abuses like Avital's grow like moss, or mold. Graduate students know this intuitively; it is written on their bones. They've watched as their professors play favorites, as their colleagues get punished for citing an adviser's rival, as funding, jobs, and prestige are doled out to the most obedient and obsequious. The American university knows only the language of extortion. "Tell," it purrs, curling its fingers around your IV drip, "and we'll eat you alive." Avital conducts herself as if someone somewhere is always persecuting her. She learned this, I imagine, in graduate school. No woman escapes the relentless misogyny of the academy. The humanities are sadistic for most people, especially when you aren't a white man. This is understood to be normal. When students in my department asked for more advising, we were told we were being needy. "Graduate school should destroy you," one professor laughed. The irony is that those who survive this destruction often do so at the cost of inflicting the same trauma on their own students. Avital, now a grande dame of literary studies, who Reitman alleges bragged to him of a "mafia"-like ability to make or break the careers of others, still feels persecuted. She makes it the job of those around her to protect her from that persecution: to fawn, appease, coddle. The lawsuit against her reads as a portrait, not of a macho predator type, but of a desperately lonely person with the power to coerce others, on pain of professional and psychic obliteration, into being her friends, or worse. It's possible that Avital genuinely believed that her student loved her, that he wanted to protect her from the scary, hostile world. In that case, the alleged assaults would have literalized the romantic tone she required he use. "Hold me," they would have said. "Make me feel loved." There is a phrase for all of this: cultish subjection. It comes from a book called Complaint, released this year, in which the author writes of graduate school as a kind of indentured labor. "I was a painfully earnest baby scholar," she recalls, "dedicated, conditioned for every sort of servitude, understanding that doing time, whether in graduate school or as part of a teaching body, amounted to acts — or, rather, passivities — of cultish subjection." The author's name is Avital Ronell. Andrea Long Chu is a writer and critic living in Brooklyn. Her book, Females: A Concern, is forthcoming from Verso.
Synergistic effect of land-use and vegetation greenness on vulture nestling body condition in arid ecosystems.
Sci Rep. 2018 Aug 29;8(1):13027. doi: 10.1038/s41598-018-31344-2. Santangeli A1, Spiegel O2, Bridgeford P3, Girardello M4. Author information 1 The Helsinki Lab of Ornithology, Finnish Museum of Natural History, University of Helsinki, P.O. Box 17, Helsinki, FI-00014, Finland. email@example.com. 2 School of Zoology, Faculty of Life Sciences, Tel Aviv University, Tel Aviv, 69978, Israel. 3 Vultures Namibia, Walvis Bay, Namibia. 4 cE3c - Centre for Ecology, Evolution and Environmental Changes/Azorean Biodiversity Group and Universidade. dos Açores - Depto de Ciências e Engenharia do Ambiente, PT-9700-042, Angra do Heroísmo, Açores, Portugal. Abstract Climate-driven environmental change and land-use change often interact in their impact on biodiversity, but these interactions have received little scientific attention. Here we study the effects of climate-driven environmental variation (i.e. vegetation greenness) and land-use (protected versus unprotected areas) on body condition of vulture nestlings in savannah landscapes. We combine ringing data on nestling measurements of two vultures (lappet-faced and African white-backed vulture) with land-use and environmental variables. We show that body condition of white-backed vulture nestlings decreased through the study period and was lowest inside protected areas. For the lappet-faced vulture, nestling condition was improved during harsh years with lower than average vegetation greenness assumed to result in increased ungulate mortality, but only within protected areas. Such interaction was not tested for the white-backed vulture due to collinearity. The species-specific effects of land-use and vegetation greenness on nestling condition of the two sympatric vulture species likely stem from their different life-histories, diet preferences and foraging behaviour. While translation of current findings on nestling conditions to their possible influence on population demography and species persistence require further studies, our findings demonstrate how environmental change may trigger selective bottom-up ecosystem responses in arid environments under global change. PMID: 30158660 DOI: 10.1038/s41598-018-31344-2 Free full text
The apparent role of climate change in a recent anthrax outbreak in cattle
Rev Sci Tech. 2017 Dec;36(3):959-963. doi: 10.20506/rst.36.3.2727. . Maksimovic Z, Cornwell MS, Semren O, Rifatbegovic M. Abstractin English, French, Spanish An anthrax outbreak recently occurred in cattle in a region that had previously been free of the disease for more than two decades. This event followed heavy springtime rains that had caused flooding, and a hot, dry summer. These temporally connected events may indicate a new link between climate change and an increased incidence of bacterial diseases with environmental reservoirs. Récemment, un foyer de fièvre charbonneuse a affecté les bovins d'une région précédemment indemne de cette maladie depuis au moins deux décennies. De fortes précipitations printanières accompagnées d'inondations et suivies d'un été chaud et sec ont précédé cet événement. La relation chronologique entre ces événements semble indiquer un nouveau lien entre le changement climatique et l'incidence en hausse des maladies bactériennes ayant des réservoirs dans l'environnement naturel. En fechas recientes se produjo un brote de carbunco bacteridiano que afectó al ganado vacuno de una región que hasta entonces llevaba más de dos décadas libre de la enfermedad. Ese episodio ocurrió después de intensas lluvias primaverales, que causaron inundaciones, y de un verano seco y caluroso. Estos fenómenos conectados en el tiempo podrían ser indicativos de la existencia de un nuevo vínculo entre el cambio climático y una mayor incidencia de enfermedades bacterianas que disponen de reservorios en el medio natural. KEYWORDS: Anthrax outbreak; Cattle; Climate change; Vaccination; Weather conditions
Using herbaria to study global environmental change.
New Phytol. 2018 Aug 30. doi: 10.1111/nph.15401. [Epub ahead of print] Lang PLM1, Willems FM2, Scheepens JF2, Burbano HA1, Bossdorf O2. Author information 1 Research Group for Ancient Genomics and Evolution, Max Planck Institute for Developmental Biology, 72076, Tübingen, Germany. 2 Plant Evolutionary Ecology, Institute of Evolution and Ecology, University of Tübingen, 72076, Tübingen, Germany. Abstract During the last centuries, humans have transformed global ecosystems. With their temporal dimension, herbaria provide the otherwise scarce long-term data crucial for tracking ecological and evolutionary changes over this period of intense global change. The sheer size of herbaria, together with their increasing digitization and the possibility of sequencing DNA from the preserved plant material, makes them invaluable resources for understanding ecological and evolutionary species' responses to global environmental change. Following the chronology of global change, we highlight how herbaria can inform about long-term effects on plants of at least four of the main drivers of global change: pollution, habitat change, climate change and invasive species. We summarize how herbarium specimens so far have been used in global change research, discuss future opportunities and challenges posed by the nature of these data, and advocate for an intensified use of these 'windows into the past' for global change research and beyond. KEYWORDS: ancient DNA; biological invasions; climate change; habitat change; herbarium; phenology; pollution PMID: 30160314 DOI: 10.1111/nph.15401 Share on FacebookShare on TwitterShare on Google+
A study on diversity of medicinal plant usage by folk medicinal practitioners in different villages of Dhunat Upazila, Bogra district, Bangladesh
January 2017 Md Ronzu AhmmedSaleh AhmedAntora KarShow all 12 authorsShoaib Mahmud Mahmud Recommend Download Abstract Medicinal plants are invaluable natural resources of Bangladesh. Several studies were conducted in different region of Bangladesh about the medicinal plants and its benefit. In this study, data were collected from the 24 different kavirajes of Dhunat Upazila of Bogra district in the division of Rajshahi about the traditional medicinal plant which they used in the treatment of many ailments. Practitioners of this region use more-than 74 plants for their traditional medicinal practice that are distributed into 51 families. The objective of our present study was to show the importance of medicinal plants, there uses and the importance of documentation of the information as well as to help the regional people to get knowledge about these types of plants which are available in their surroundings nature for using these as primary treatment. Among these 74 plants, 14 plants were used for treatment of gastrointestinal disorders, 21 plants were used for male and female sexual problems; 11 were used in asthma and skin diseases. Beside these, 7 plants were used to cure kidney diseases, 5 plants were used to overcome mental depression, 4 plants were used for hypertension and diabetes and three plants were used for ailment of heart disease. Additionally, these plants were used to treat many other diseases. For the country like Bangladesh medicinal plants is a vital asset and has significant role in people health care system. So the awareness towards the conventional use of medicinal plants needs to be increased among the local people. Proper and developed cultivation process must be needed for improving and maintaining the quality and growth of these indigenous medicinal plants. Additionally, proper research should be conducted for using these medicinal plants in new drug design and many other pharmaceutical benefits
Thursday, 30 August 2018
How the killer whale became the Achilles heel of Trans Mountain pipeline approval
https://www.cbc.ca/news/canada/british-columbia/how-the-killer-whale-became-the-achilles-heel-of-trans-mountain-pipeline-approval-1.4804932 Court ruled the NEB's failure to assess impacts of marine shipping was a 'critical error' Michelle Ghoussoub · CBC News · Posted: Aug 30, 2018 3:07 PM PT | Last Updated: 3 minutes ago Southern resident killer whales are designated under the Species At Risk Act, which means federal prohibitions exist against anything that would harm them or habitat considered critical to their survival. (Valerie Shore/Shorelines Photography) It's been a summer of dramatic killer whale news — from a mother holding up her dead calf for 17 days in a gut-wrenching display of grief, to a boatload of scientists shooting a sick whale with a dart full of antibiotics. Now, B.C.'s ailing southern resident killer whale population is proving itself a wedge in one of the most headline-grabbing issues in the province. In the 200-page decision released by the Federal Court of Appeal Thursday morning, effectively quashing the government's approvals to build the Trans Mountain expansion project, B.C.'s southern resident killer whales are mentioned no fewer than 57 times. The court ruled that the National Energy Board (NEB) review failed to assess the impacts of marine shipping — saying it was so flawed, it should not have been relied on by the federal cabinet when it gave final approval to proceed in November 2016. Federal Court of Appeal quashes construction approvals for Trans Mountain, leaving project in limbo Activists, lawyers and academics say the decision demonstrates environmental corners cannot be cut when governments seek social license for major infrastructure projects — especially in a case where increased tanker traffic and vessel noise are known to be key threats to killer whales. "It's very clear from this decision that environmental assessment considerations and Species At Risk Act decisions aren't optional, and they need to be taken seriously," said Dyna Tuytel, a lawyer with Ecojustice, who represented conservation groups that filed a court challenge to the federal government's approval for a pipeline expansion. "There's a risk in taking shortcuts," said Eric Taylor, a professor of zoology at the University of B.C., and the chair of the Committee on the Status of Endangered Wildlife in Canada. "It's going to come back and bite you, as it's done here." Narrow reading of the law According to the ruling, the shortcut, or "critical error" made by the NEB, was to define the scope of the project as only the pipeline and the marine terminal for the purposes of its environmental assessment. So although the project looked at marine shipping during the review, it did not assess it to environmental standards, nor did it apply the Species At Risk Act to the effects of marine shipping on endangered species. B.C.'s southern resident killer whales are considered at risk because of their small population, low reproductive rate and threats including marine traffic and lack of food. (Dave Ellifrit/Center for Whale Research) "The NEB acknowledged that on the facts there were significant adverse effects of the project on southern resident killer whales — but the board, by defining the project narrowly, was able to say that the project was not likely to cause significant adverse effects," said Tuytel. Taylor called the decision to leave out the project-related tanker traffic in the review "unfathomable." "The oil is not going to sit there in barrels, it's got to move out by ships. And ship traffic has clearly been identified as a threat to this endangered species. So it's unconscionable that they ignored it," he said. Cutting corners Tuytel called the ruling "fairly unusual." But Taylor said he wasn't surprised, given the threats to southern resident killer whales have been clear for over a decade. "I think the court really had no other choice than to do this," he said. IN DEPTH | The killer whale crisis that a shot won't solve The whales, which are also threatened due to toxic contamination levels and low supplies of Chinook salmon, are designated under the Species At Risk Act, which means federal prohibitions exist against anything that would harm them or habitat considered critical to their survival. In June, the federal Department of Fisheries and Oceans pledged measures to slow down vessel traffic, noting the population was facing an imminent threat to survival. Fuel spills and underwater noise from tankers are just some of the threats that have endangered the southern resident killer whale population. (Michael Mcarthur/CBC) Since the time the NEB reviewed the project, B.C.'s southern resident population has declined from about 82 to 75. The project would increase capacity from five ships a month through Burrard Inlet to a maximum of 34 oil tankers capable of carrying 120,000 tonnes of diluted bitumen at a time. According to the NEB report, Trans Mountain acknowledged the additional noise the project would create, but argued that the shipping lanes "will continue to host marine vessel traffic with or without the project, and that the impacts to the southern resident killer whales will continue to exist with or without the project." Killer whales a 'flare' for other issues This is not the first time whales have played a role in halting a major Canadian infrastructure project. Last year, energy giant TransCanada scrapped plans for a port for its proposed Energy East pipeline after protesters raised alarms about impacts on the calving grounds of the vulnerable beluga population in the St. Lawrence Estuary. ANALYSIS | Big oil vs. big whale: Will pipeline trump iconic orca? Taylor said the pipeline and its associated infrastructure are likely to have impacts on many species, but because of the popularity of killer whales, they tend to act as a "flare" for many of the issues associated with the project. "If this was a lichen, many, many fewer people would be paying attention," he said. ABOUT THE AUTHOR Michelle Ghoussoub @MichelleGhsoub Michelle Ghoussoub is a journalist with CBC Vancouver. She previously reported in Lebanon and Chile.
Repellent and Lethal Activities of Extracts From Fruits of Chinaberry (Melia azedarach L., Meliaceae) Against Triatoma infestans
ORIGINAL RESEARCH ARTICLE Front. Vet. Sci., 26 July 2018 | https://doi.org/10.3389/fvets.2018.00158 Martín Dadé, Pedro Zeinsteger, Facundo Bozzolo and Nora Mestorino* Laboratory of Pharmacological and Toxicological Studies (LEFyT), Faculty of Veterinary Science, Universidad Nacional de La Plata, La Plata, Argentina Triatoma infestans is the principal vector of Trypanosoma cruzi, parasite responsible of Chagas's Disease transmission in Argentina. Pyrethroids have become common pesticides for the control of T. infestans but increasing resistance encourages the search of new alternatives and the use of natural products for biological control arises as a new strategy. Melia azedarach L. is originated from the Himalaya's region and several compounds are part of its rich phytochemistry. Folk medicine of the plant is due to its repellent and insecticidal activities. Aims of this work were to evaluate the repellent activity of methanolic and acetonic extracts from fruits of M. azedarach by means of the area preference method of fifth and first nymph stages as well as to test the acute lethal effect of the more repellent extract by means of direct application on cuticle on both stages. For repellence, qualitative filter papers were divided into two halves, one treated with methanolic (ME) or acetonic (AC) extract and the other without treatment. Controls were impregnated half with methanol or acetone and half without the solvents. One nymph was located in each Petri or well and repellence percentage was determined. For the lethal effect, fasted and fed to repletion 5th stage nymphs were topically administered with different concentrations of AC and deaths were registered after 24, 48, 72, 96, and 120 h. Phytochemical analysis of extracts was performed as well. AC demonstrated high repellent activity (100%, both stages), whereas ME extract activity was slight (10–21%). AC extract was selected for lethal assays due to early repellent activity. Fed to repletion nymphs were more sensitive to the lethal activity of the extract when compared to fasted nymphs (LD50: 11.5 vs. 23.1 μg/insect, respectively). Phytochemistry assays of extracts showed a higher concentration of flavonoids, alkaloids and triterpenes for AC. Considering these results, next assays will include the test of Melia azedarach extract on T. infestans that are resistant to pyrethroids for a possible synergism between AC and the pesticides. Introduction Chagas's Disease, also known as American Trypanosomiasis, is a zoonosis caused by the protozoan parasite Trypanosoma cruzi which needs a host body and a vector to complete its life cycle, being the latter the hematophagous insect Triatoma infestans ("kissing bug") distributed from Southern United States (1) to Argentina (2). The disease is endemic to Latin America and has been reported from Southern Argentina to Northern Mexico. It has also been diagnosed in people from non-endemic countries because of the increment of international migration during recent years (USA and countries from Europe, Asia and Oceania). According to the World Health Organization, up to 10 million humans are infected worldwide with more than 10,000 deaths in 2008 (3). Transmission of T. cruzi is not only due to T. infestans hematophagous activity but also a consequence of the ingestion of contaminated food with vector stools (4), congenital infection (5, 6), blood transfusion (7, 8) and organ transplantation (9, 10). Triatomines live in dark and warm cracks of poorly-constructed homes in both rural and suburban areas. They become active during the night when they feed on hot blooded species including man. An insect usually bites an exposed area of the skin and defecates while feeding close to the bite, this situation enhances infection as the bitten person smears the feces into the bite or into the eyes, mouth or any skin lesion (3). The evolution of the disease is characterized by two phases: acute, which may last 2 months and can be asymptomatic or with symptoms appearing shortly after the infection, they include fever, headache, enlarged lymph nodes, pallor, muscle pain, labored breathing, swelling and abdominal or chest pain (3); and a chronic phase which may last for the entire life with symptoms appearing after a silent period which may take several years. During the chronic phase, up to 30% of infected people develop lesions that compromise the heart, and up to 10% develop digestive, neurological or mixed alterations (3). Two medications are commonly used for the treatment of the acute phase of trypanosomiasis, including nifurtimox and benznidazole, with 75–100% healing with prompt administration, particularly in cases of congenital infection (11). Both medications are effective during the acute phase but not in the chronic phase and this is one of the reasons why many strategies are developed to avoid vector's transmission. Despite these pharmacological alternatives, therapeutic management of the disease is complex as adverse effects may develop during the treatment. Many alternatives have been implemented for the interruption of disease spreading including early detection of seropositive patients and pharmacological treatment during the acute phase to avoid irreversible lesions in target organs, health campaigns and vectors surveillance by means of synthetic pesticides (e.g., pyrethroids) with residual properties (12). Pesticides are extensively used in many countries of Latin America with strong impact on non-target insects, animals and human health as well. Besides these issues, resistance to pesticide develops fast in some species of insects (13, 14) including T. infestans (15). Moreover, the exposition of pyrethroids to sunlight and water determines a substantial reduction of residual power (16), a common situation in rural areas. To minimize these factors new technologies and management strategies are necessary to obtain less hazardous and more resistant chemical or biological compounds. Regarding the latter, the use of plant extracts arises as a promising alternative nowadays, something that it is not necessarily new because botanical insecticides have been used for at least 2000 years in Asia and Middle East (17). Interest in these compounds relies on their low cost, efficacy, degradability and pharmacological activity on insects (18, 19). Melia azedarach L. (MA) also known as "chinaberry tree" is an ornamental species of the Meliaceae family considered to be native from Asia which grows from North to South America as well as Northern Australia, Africa and Southern Europe (20). It is a deciduous and evergreen 3–10 m tree with sweet-scented lilac flowers during autumn and spring, dark green leaves and a round-shaped fruit initially green and yellowish when mature (21). Trees are cultivated in countries with template to warm climates and in Argentina they are easily found in houses and parks as ornamental trees for protection against sunlight and winds. Melia azedarach has demonstrated to have both pharmacological and toxicological properties. Fruits have been studied for their phytochemical composition which includes melianoninol, melianol, melianone, meliandiol, vanillin, and vanillic acid (22). Toxic compounds are tetranortriterpens, known as meliatoxins, present in all the parts of the plants but specially concentrated in the ripe fruits (21). Aqueous and alcohol extracts prepared from different parts of MA have antibacterial (23), antiparasitic (24), antifungal (25), antiviral (26), and antioxidant properties (27) while the ingestion of foliage or fruit by cattle (28), pigs (29), dogs (30), and other species has caused intoxication with fatal outcomes in some cases. MA has also been tested for insecticide activity. Vergara et al. (31) and Carpinella et al. (25) state that this capability is due to the anti-feeding effect of tri-terpenoids that inhibit food intake capacity, which has been demonstrated in phytophagous insects, thus leading to death and malformations in next generations. Plant extracts prepared from leaves or fruits have been tested on bean weevil (32), mosquitoes (33, 34) and moths (35). Some information exists regarding repellent and insecticidal properties on T. infestans (36) but no up-to-date data are available. Purpose of this work is to present the results of tests performed with acetonic and methanolic extracts prepared from fruits of MA considering repellent and insecticidal capabilities on different evolutionary stages of T. infestans for a possible economic and easy to implement complement of traditional pesticides used for the control of the vector of Chagas disease. Materials and Methods Plant Material Ripe fruits of MA (1 kg) were collected in August 2017 from trees located close to the School of Veterinary Science, National University of La Plata (UNLP). Voucher specimens were deposited after botanical identification at the Laboratory of Pharmacological and Toxicological Studies ("LEFyT," from Spanish), School of Veterinary Sciences, UNLP. Plant material was put inside an Erlenmeyer (2 L) and was shaken, after this 10 g of fruits were separated for the assays. Fruits were washed with distilled water and excess of moisture was removed on adsorbent paper. Covers were separated from the seeds for a better extraction and all the plant material was placed in a Soxhlet cartridge. Melia Azedarach Extracts For the extraction, acetone or methanol (200 mL) was used as solvent for the preparation of the acetonic extract and methanolic extract, respectively. Ten grams of fruits were used to obtain each extract. The extraction temperature of the Soxhlet equipment ranged from 60 to 70°C and the process was carried out during 10 h under dim light to avoid possible inactivation of photosensitive compounds. The acetonic (AC) or methanolic (ME) extract was separated into two parts, 50 mL for phytochemical assay and 150 mL for biological assay in triatomines. The solvent of the 150 mL fraction (AC or ME extracts) was rotaevaporated at 60°C (Senko Ltd.) and a dark red residue was obtained which was resuspended in the same solvent (50 mL). This process was carried out as three consecutive extractions using 10 g of M. azedarach fruits each in order to standardize the amount of residue in the rotaevaporator flask and for each in vivo assay, obtaining in average 342 ± 50 mg/50 mL AC extract and 2,524 ± 150 mg/50 mL ME extract. These stock solutions were used to prepare a series of dilutions (1:1, 1:5, and 1:10). Qualitative chemical determinations were performed with the 50 mL of AC and ME extracts to determine the presence of compounds with potential repellent and insecticide activities. AC and ME total extracts were fractioned in three parts and chemical reactions were used for each fraction as follows: Fraction A: Shinoda for flavonoids, FeCl3 for tannins and phenolic OH, Iodine for lipids, Phenol 5% + concentrated H2SO4 for carbohydrates; fraction B: Liebermann-Burchard for steroids, Bornträger for antraquinones; fraction C: Dragendorff for alkaloids, Kedde for cardenolides and Rosenheim for leucoanthocyanins (37, 38). Only qualitative analysis of the extracts was performed to guarantee the presence of compounds with repellence and lethal activities (considering previous works by other researchers). Quantitative determinations considering chromatographic techniques will be part of future assays in order to determine exact ingredients of AC extract of Melia azedarach in our laboratory. Experimental Insects Nymphs of 1st and 5th stages of T. infestans susceptible to pyrethroids were from an insect colony grown at LEFyT-UNLP insectary. This colony was originated from triatomines provided by the Centro de Referencia de Vectores (CeReVe), Santa María de Punilla, Córdoba, Argentina. All the insects were free of T. cruzi infection. Colonies were fed with chicken blood once a week, kept at 26 ± 2°C, 60–70% relative humidity and a light cycle of 12:12. Repellent Activity of AC and ME Extracts on 1st and 5th T. infestans Stages For the evaluation of the repellent activity of AC and ME extracts the preference area method was used. Fifth stage nymphs were placed on a 9 cm diameter filter paper divided into two areas and then into Petri dishes (Figure 1). For 1st stage nymphs 3.5 cm filter papers were located in multiple-well plates (6 wells, Figure 2), purpose of this was to offer the insect the half of the contact surface treated with different concentrations of extract and the remaining without treatment. In case of positive repellent activity, the insect moves to the area of the paper free of extract. For controls, one half of the paper was treated with acetone o methanol. Treated halves were left to evaporate during 24 h before placement of insects and joined to the correspondent non-treated half using adhesive tape. Both nymph stages were tested using three dilutions of AC or ME extracts (1:1, 1:5, and 1:10), each concentration tested on ten insects. Volume for each dilution was 500 μL for 5th stage nymphs and 77 μL for 1st stage nymphs. Each insect was placed in the center of the paper and observation was performed after 1, 12, 24, and 48 h. Repellent activity (RA) was calculated using the following equation: RA=Nc −NtNc +Nt x 100 Nc: number of insects in the control area; Nt: number of insects in the treated area. FIGURE 1 www.frontiersin.org Figure 1. Fifth stage nymphs located into Petri dishes. FIGURE 2 www.frontiersin.org Figure 2. First stage nymphs located in multiple-well plates (6 wells). Values of RA may be negative or positive. In the case that most of the insects stay in the untreated area (Nc > Nt) RA value will be positive and the assayed substance is considered to have repellent activity. Negative RA values (Nt > Nc) means that most of the insects stay in the treated area, thus considering the assayed extract to have attracting activity (37). Insecticidal Capabilities Assays were performed only with AC extract as it showed rapid onset of repellent activity, a characteristic that was absent in ME extract. Insecticidal capability of AC extract was determined on 5th stage nymphs by means of the calculation of the LD50, comparing fed to repletion vs. fasted triatomines. From the stock solution of the AC extract (342 ± 50 mg/50 mL), different work solutions to be assayed were prepared by means of dilution/concentration procedures in order to obtain a concentrations range between 1.37 and 30.52 μg/μL. Briefly, the stock solution was diluted with acetone (1:5) to obtain a secondary solution (1.37 μg/μL). Afterwards aliquots were taken from the stock or secondary solution, which were concentrated to dryness (SpeedDry Vacuum Concentrators Christ CD Plus, Germany) and then dissolved in different volumes of acetone to reach the final concentrations/μL to be tested on the triatomines. Calculation of LD50 in 5th Stage T. infestans Fasted Nymphs For the determination of the LD50 of AC extract, 5th stage T. infestans susceptible to pyrethroids was used according to the WHO protocol (38). Nymphs were fasted during 12–15 days after ecdysis and prior to their use. The assay considered a binary response—dead or alive—with an independent variable (dose). AC extract was applied topically on the dorsal of nymphs' abdomen (1 μL). For the dose-response curves, increasing doses of the extract were evaluated (1.37–30.52 μg/μL or 1.37–30.52 μg/insect). The same procedure was used for controls with the application of 1 μL acetone. Assay was replicated during three different days under similar conditions and all the extracts were recently prepared for each repetition. Ten nymphs were used for each dose and repetition (30 insects for each dose) and only 10 nymphs as controls. After topication insects were located in labeled flasks and observed after 24, 48, 72, and 168 h. An insect was considered dead when it was not able to move from the center of the filter paper by its own or by means of stimulation with tweezers. Dead insects were removed and placed in labeled containers and observed again after 24 h to corroborate death. Calculation of LD50 in 5th Stage T. infestans Fed to Repletion Nymphs For this experiment, 5th stage T. infestans nymphs fed to repletion with chicken blood were used in groups of 10 insects. Prior to this, nymphs were fasted for a period of 12–15 days after ecdysis. Each triatomine was individualized with different acrylic paint color marks (AD acrílico, Argentina) and weight before and after feeding with a precision scale (Denver Instruments, USA). After a feeding period of 30 min the quotient between weight after and before was calculated and only those insects with a 4-fold relation or more were used. For the determination of the LD50 the same procedure for fasted nymphs was used. Statistical Analysis To verify the relation among different doses of AC extract regarding mortality of nymphs the Probit method was used, which allows to associate mortality with a dose necessary to cause it. Mortality data for the treated nymphs were corrected taking into account the mortality of control nymphs by means of Abbott's Equation (39): Corrected % mortality=(% NT mortality−% NC mortality)(100%−% NC mortality) x 100 Where: NT: nymphs treated with different doses of AC extract diluted in acetone NC: control nymphs, only received acetone. From corrected mortality data of AC extract different dose-response curves were obtained using the POLO-PLUS software (LeOra Software Company, Petaluma, CA 2005). LD50 with its respective confidence interval 95% (CI 95%) was calculated as well. To determine differences in susceptibility between T. infestans nymph populations the ratio calculation between LD50 (RLD50) was calculated with confidence interval (CI) of 95%. LD50's are considered statistically different when 95% CI of RLD50 does not include number one (p < 0.05) (40). Results Phytochemistry of AC and ME Extracts Table 1 shows the results of the phytochemical analysis of the AC extract while Table 2 those for ME extract. TABLE 1 www.frontiersin.org Table 1. Qualitative analysis of AC extract. TABLE 2 www.frontiersin.org Table 2. Qualitative analysis of ME extract. Repellent Activities of AC and ME Extracts In Tables 3, 4 it can be observed that only AC extract showed repellent activity for both stages of T. infestans. Repellent activity was directly proportional to the concentration of the extract. Dilution 1:10 did not cause repellence or it was negligible in any stage. In the case of 1st nymph stage, 1:1 dilution showed weak repellent activity as early as 1 h of the initiation of the assay (Table 3), while 1:5 dilution demonstrated this activity at 24 h. Highest repellent activity (100%) was observed after 24 h and remained constant until 48 h. TABLE 3 www.frontiersin.org Table 3. Repellent activity of AC extract on 1st stage T. infestans nymphs. TABLE 4 www.frontiersin.org Table 4. Repellent activity of AC extract on 5th stage T. infestans nymphs. For the 5th stage repellent activity was lower compared to 1st stage. Again, only 1:1 dilution was effective and its activity started at 12 h after initiation of the experiment. This dilution reached 100% of repellence at 24 h and remained constant until 48 h. LD50 of AC Extract Table 5 shows the influence of nutritional state of nymphs on the lethal activity of AC extract. Fed to repletion nymphs were more sensitive to the lethal activity of the extract when compared to fasted nymphs. Both RLD50 were close to 2, this means that twice the dose was needed for fasted nymphs compared with fed to repletion nymphs. TABLE 5 www.frontiersin.org Table 5. Relation between nymph nutritional state and lethal activity of AC extract. Discussion Under our working conditions we found that AC extract showed efficacy in susceptible to pyrethroid nymphs regarding repellent and lethal activities, such situation was influenced by the feeding state of insects. A possible explanation of the higher lethal efficacy of AC extract in T. infestans nymphs fed to repletion could be associated to the ability of some insects to modify the mechanical properties of cuticle. This process, known as plasticization (41), involves modifications in the aqueous phase of cuticle due to changes in pH, leading to rupture of soft bonds among proteins and chitin microfibers. These reversible modifications allow procuticle to have a better elongation capacity. Plasticization has been largely studied in Rhodnius proxilus (42, 43), researchers found this process to be important during ecdysis and feeding, particularly in 5th stage nymphs when plasticization allows triatomines to considerably increase their original size. Moreover, in 5th stage nymphs of T. infestans it has been demonstrated that a feeding time as short as 1 min using an artificial feeding system is enough to start plasticization. During this process epicuticle folds are expanded and procuticle is more flexible, thus enhancing the penetration of different molecules from outside, such as pesticides (44). From our results it can be stated that in nymphs fed to repletion plasticization allowed an increased penetration of the active compounds present in AC extract resulting in a higher toxic response (LD50 fed to repletion insects < LD50 fasted insects). In 2nd stage T. infestans submitted to 14C-DDT, Fontán and Zerba (45) reported an increased penetration rate of the organochlorine in fed to repletion vs. fasted insects; this result is similar to the topication assay we performed with AC extract. Regarding repellent activity our results were similar to those of Valladares et al. (36) although our extract was acetonic, compared to the ethanolic extract of the researchers. They also determined that the ethanolic extract did not affect the survival of triatomines when they were submitted to papers impregnated with extracts. The latter could be a possible explanation to the differences with our results, because in the case of topication each insect receives a determined volume (dose) while with the contact method the amount of extract on insects depends on the locomotive activity of each individual. Besides, there are some similarities and differences in the chemical composition of extracts. The ethanolic extract from Valladares et al. (36) had limonoids, a group of insecticidal triterpenes. In AC extract, although we did not determine the chemical identity of triterpenes, they were abundant. As a difference, the ethanolic extract did not have alkaloids, compounds that were present in AC extract and are probably responsible for the lethal activity, as these secondary metabolites are used as defense mechanism against insects and herbivores (46). Such compounds, acting as protective agents for plants, are known as allelochemics (47). Acetone is the recommended solvent for experimental use in T. infestans according to the World Health Organization (38). We found that repellent activity of AC extract was slightly greater in 1st stage compared to 5th stage nymphs, this could be related to anatomical and physiological differences between both stages, such as penetration rate of substances through cuticle, presence of sensory organs specific to each stage and augmentation of metabolic activity (48). Repellent activity of ME extract was considerably low when compared to AC extract, this could be due to the higher concentration of some compounds in AC extract such as triterpenes, as previously demonstrated for similar compounds by other researchers (49, 50). Although health campaigns have been implemented in developing countries the infected human population in Latin America is still high and there is concern about international immigration in countries where the disease is non-endemic. Synthetic insecticides are useful tools for the control of pests, but their excessive use has led to negative consequences such as toxicity against farmers, consumers and both wild and domestic animals as well as interruption of natural control and pollination, water pollution and development of resistance (50–53). Moreover, some populations of T. infestans have developed resistance to these pesticides (52, 54) Melia azedarach is present in many countries of Latin America where it is usually used in folk medicine by means of maceration of fruits and leaves to prepare extracts due to their repellent and insecticide properties against many crop pests and human disease vectors. Such conditions together with our findings could justify the use of plant preparations as an accessible complement together with the traditional use of pesticides for the control of T. infestans with probable synergism or potentiation of actions between molecules in susceptible nymphs. Next step in our research will be the assay of MA extracts on T. infestans that are resistant to pyrethroids for a possible synergism between AC extract and the pesticides. Such situation, if successful, may allow to use less concentrations of synthetic insecticides during aspersion, which in turn may cause less impact on environment as well as human population. Part of this work has already started. Author Contributions MD, PZ, and NM conceived and designed the experiments. PZ performed the Melia azedarach Extracts. FB Managed and maintained the triatomines. All authors contributed to the redaction, revision and approved the final manuscript. Conflict of Interest Statement The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. 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In vitro antitoxin activity of aqueous extracts of selective medicinal herbs against Naja naja venom
Vol. 7, Issue 1 (2018) AUTHOR(S): Mani M, Revathi P, Prabhusaran N and Pramila MABSTRACT: There is increasing global health problems related to snake bites and its complications in India. In the medical practice, the only therapy for treating snake bite victims is anti-snake venom serum, but it has huge acute and chronic including late adverse reactions in humans. Thus this study was aimed to analyze the potential of selective medicinal herbs against Naja naja toxin by in vitro methods. The aqueous extracts of fruit, root, seed, bark and leaf of Emblica officinalis, Hemidesmus indicus, Tamarindus indicus, Mangifera indica and Vitex negundo were prepared respectively. Various concentrations of the extracts were used to evaluate in vitro anti toxin activity by acetylcholinesterase, protease, direct hemolysis, phospholipase and procoagulant activities. The results indicate that the root and bark extracts of H. indicus and M. indica possesses significant bioactive compounds that can neutralize the toxins of N. Naja.PAGES: 585-589 | 117 VIEWS 8 DOWNLOADSDOWNLOAD (564KB)
Trans Mountain pipeline halted after Canadian court overturns approval
https://www.theguardian.com/world/2018/aug/30/trans-mountain-pipeline-latest-canada-court-overturns In a unanimous decision, the federal court of appeal said the government failed to consider the concerns of some First Nations Ashifa Kassam in Toronto @ashifa_k Thu 30 Aug 2018 22.06 BST Last modified on Thu 30 Aug 2018 23.11 BST members of the Anishinaabe tribe march with others against the expansion of the Trans Mountain project in Burnaby, British Columbia on 10 March. members of the Anishinaabe tribe march with others against the expansion of the Trans Mountain project in Burnaby, British Columbia on 10 March. Photograph: Jason Redmond/AFP/Getty Images A Canadian court has overturned Ottawa's approval of a hotly-contested pipeline project – throwing plans to nearly triple the flow of Alberta's landlocked bitumen to the west coast into limbo – in a ruling hailed by environmentalists and Indigenous groups. In a unanimous decision released on Thursday, the federal court of appeal said the Liberal government – led by Justin Trudeau – failed to adequately consider the concerns of some First Nations regarding the Trans Mountain expansion project. The ruling also found that the country's energy regulator, the National Energy Board, did not properly consider the impact of increased tanker traffic due to the project. 'Our land is our home': Canadians build tiny homes in bid to thwart pipeline Read more The scheme had been a crucial test for Trudeau and his government, who swept into office in 2015 on promises of striking a balance between economic growth, environmental concerns and repairing the country's fraught relationship with Indigenous peoples. While the project could allow Alberta to get its bitumen to markets in Asia and reduce its reliance on the US market, there has been opposition over the potential for oil spills and the impact that a dramatic rise in tanker traffic could have on the region's southern resident killer whales, a population already on the knife edge of extinction. The court ruling will halt construction of the 1,150km project, spearheaded by Texas-based Kinder Morgan, until the energy regulator and government can show they have complied with the court's demands – a process that could take years. Several Indigenous groups and environmentalists applauded the ruling, which emerged from a legal challenge backed by more than a dozen groups, including the city of Vancouver, several First Nations and environmental organisations. "Thankfully, the court has stepped in where Canada has failed to protect and respect our rights and our water," Coldwater Indian Band chief Lee Spahan said in a statement. Recent months have seen increased Indigenous-led protests against the project, with thousands of people demonstrating in the streets. Hundreds – including two federal MPs – have been arrested for blocking the entrance of a facility belonging to Trans Mountain. The federal government, who earlier this year announced it would spend $4.5bn to purchase the pipeline in a bid to push the expansion forward, said it was reviewing the decision. "We are absolutely committed to moving forward with this project," Bill Morneau, Canada's finance minister, told reporters on Thursday, saying that the government had yet to decide whether it would appeal. The court's decision was announced minutes before Kinder Morgan shareholders voted to approve the sale of the pipeline to the Canadian government. Morneau defended the purchase, describing it as a "sound investment", given that any private sector company would have found it difficult to bear the risks involved with the project. "We believe this project is in the national interest, we believe that it's critically important for our economy, critically important to allow us … to get to international markets," said Morneau. Thursday's ruling puts the government in a difficult position, said Stewart Phillip, grand chief of the Union of British Columbia Indian Chiefs. "They actually bought the pipeline that nobody wanted," he said. "And they paid too much for it." The ruling also exposes the federal government's shortcomings when it comes to meaningful dialogue with Indigenous peoples – despite Trudeau's oft-repeated promises to do better. "Clearly the eloquent words of Prime Minister Trudeau do not reconcile with his actions on the ground," said Phillip. Describing the decision as a "massive victory", Greenpeace Canada noted that the ruling offers the federal government a second chance of sorts. "Now it's time for Prime Minister Trudeau to read the writing on the wall, dump this pipeline and shift the billions of public dollars slated for this problem-plagued project into Canada's renewable energy economy," Mike Hudema of the organisation said in a statement. He added: "This summer's fires, floods and choking smoke make it impossible to ignore the rising costs of climate inaction, so the prime minister should welcome the opportunity created by today's ruling to get on the right side of history."
Wednesday, 29 August 2018
Effect of Evening Primrose Oil on Korean Patients With Mild Atopic Dermatitis: A Randomized, Double-Blinded, Placebo-Controlled Clinical Study.
Ann Dermatol. 2018 Aug;30(4):409-416. doi: 10.5021/ad.2018.30.4.409. Epub 2018 Jun 27. Chung BY1, Park SY1, Jung MJ1, Kim HO1, Park CW1. Author information 1 Department of Dermatology, Kangnam Sacred Heart Hospital, Hallym University College of Medicine, Seoul, Korea. Abstract BACKGROUND: Atopic dermatitis (AD) is related to a deficiency of delta-6-desaturase, an enzyme responsible for converting linoleic acid to gamma-linolenic acid (GLA). Evening primrose oil (EPO) as a source of GLA has been of interest in the management of AD. OBJECTIVE: The aim of this randomized, double-blinded, placebo-controlled clinical study is to evaluate the efficacy and safety of EPO in Korean patients with AD. METHODS: Fifty mild AD patients with an Eczema Area Severity Index (EASI) score of 10 or less were enrolled and randomly divided into two groups. The first group received an oval unmarked capsule containing 450 mg of EPO (40 mg of GLA) per capsule, while placebo capsules identical in appearance and containing 450 mg of soybean oil were given to the other group. Treatment continued for a period of four months. EASI scores, transepidermal water loss (TEWL), and skin hydration were evaluated in all the AD patients at the baseline, and in months 1, 2, 3, and 4 of the study. RESULTS: At the end of month 4, the patients of the EPO group showed a significant improvement in the EASI score (p=0.040), whereas the patients of the placebo group did not. There was a significant difference in the EASI score between the EPO and placebo groups (p=0.010). Although not statistically significant, the TEWL and skin hydration also slightly improved in the EPO patients group. CONCLUSION: We suggest that EPO is a safe and effective medicine for Korean patients with mild AD. KEYWORDS: Atopic dermatitis; Gamma-linolenic acid; Linoleic acid
The Distribution of Bovine Tuberculosis in Cattle Farms Is Linked to Cattle Trade and Badger-Mediated Contact Networks in South-Western France, 2007–2015
ORIGINAL RESEARCH ARTICLE Front. Vet. Sci., 26 July 2018 | https://doi.org/10.3389/fvets.2018.00173 Malika Bouchez-Zacria1, Aurélie Courcoul2 and Benoit Durand2* 1Epidemiology Unit, Paris-Sud University, Laboratory for Animal Health, French Agency for Food, Environment and Occupational Health and Safety (ANSES), Maisons-Alfort, France 2Epidemiology Unit, Paris-Est University, Laboratory for Animal Health, French Agency for Food, Environment and Occupational Health and Safety (ANSES), Maisons-Alfort, France Bovine tuberculosis (bTB), mainly caused by Mycobacterium bovis, can affect domestic and wild animals as well as humans. Identifying the major transmission mechanisms in an area is necessary for disease control and management. In this study, we aimed to evaluate the involvement of different types of contact in M. bovis transmission between cattle farms of south-western France between 2007 and 2015. We analyzed an empirical contact network of cattle farms as nodes, with known infection status and molecular types (16 circulated during the study period of which 14 affected only cattle and two both badgers and cattle). Edges were based on cattle trade data (T-edges) and on spatial neighborhood relationships between farms, either direct (P-edges) or badger-mediated, when two farms neighbored the same badger home range (B-edges), or two distinct but neighboring badger home ranges (D-edges). Edge types were aggregated so that the contact network contained only unique edges labeled by one or several edge types. The association between the contact network structure and bTB infection status was assessed using a non-parametric test, each molecular type being considered a marker of an independent epidemic. Using a logistic regression model, we estimated the contribution of each edge type to the probability for an edge originating from an infected farm to end at another infected farm. A total number of 1946 cattle farms were included in the study and were linked by 54,243 edges. Within this contact network, infected farms (whatever the molecular type) always belonged to the same component, suggesting the contact network may have supported bTB spread among those farms. A significant association between the pattern of bTB-infected farms and the structure of the contact network was observed when all the molecular types were simultaneously considered. The logistic regression model showed a significant association between M. bovis infection in direct neighbors of infected farms and the connection by T-, B- and D-edges, with odds-ratios of 7.4, 1.9, and 10.4, respectively. These results indicate a multifactorial M. bovis transmission between cattle farms of the studied area, with varying implication levels of the trade, pasture and badger networks according to the molecular type. Introduction Since its discovery by Theobald Smith in the late 1800's (1) Mycobacterium bovis, the main agent of bovine tuberculosis (bTB) has been found in a wide variety of domestic and wild animal hosts, as well as in humans (2, 3). In Europe, the main host of M. bovis is cattle (4–6), but sheep (7), pigs (8) and goats (9) can be affected too. Wildlife species found infected on this continent include red deer (Cervus elaphus) (10, 11), roe deer (Capreolus capreolus) (12), red fox (Vulpes vulpes) (13–16), wild boar (Sus scrofa) (17, 18) and badger (Meles meles) (19–21). Different routes may allow M. bovis transmission between wild and domestic hosts. The largest part of M. bovis shedding seems to occur through aerosols (respiratory tract secretions) and to a lesser extent through saliva, urine, feces (20, 22, 23), milk in cattle (24) and even wound exudates in badgers (20). Therefore close contacts (e.g., nose to nose) between infected individuals and susceptible ones can allow the transmission of M. bovis. However, several studies have shown that M. bovis may survive outside a host in a favorable environment for several months (24–26), allowing transmission through indirect contacts. M. bovis transmission between cattle can also involve different susceptible species either wild (27) or domestic [although the implication of other domestic species than cattle remains unclear regarding cattle transmission (24)]. At the herd level, several risk factors of bTB have been identified such as larger herd sizes, neighborhood with other herds, cattle movements, farm management practices such as grazing, dispersion of slurry on pastures or the share of water points (24, 28–31). Environmental risk factors have also been studied, with certain environmental conditions favoring the survival and persistence of M. bovis (such as shade, moisture or even some soil types) that foster M. bovis transmission (24–26). A third category of risk factors involves wildlife interactions, especially with badgers, wild boars and deer. For the latter two species, the sharing of feed or water on pastures appears to be a risk factor of M. bovis indirect transmission (23, 32, 33). The transmission between badgers and cattle seems a bit more complex, with uncertain direct contacts on pastures (34–36) and/or inside farm buildings (37). This interspecies transmission could occur on pastures through the shedding of the mycobacteria in urine and feces of infected badgers (24), and in respiratory tract secretions and feces of infected cattle (6, 29). BTB molecular types are stable (38, 39) and can be used to trace independent epidemics (4). In France, while the officially bTB-free status was obtained in 2000, M. bovis infection has persisted in several regions. In 2014, 46% of incident outbreaks were detected in south-western France, with a national number of 105 cattle herds newly detected infected (40). Molecular typing methods spoligotyping (39) combined to MLVA (Multiple Loci Variable Number of Tandem Repeats, VNTR Analysis) based on MIRU-VNTR [Mycobacterial Interspersed Repetitive Unit–VNTR; (4, 38)] have allowed identifying 16 molecular types in this area between 2007 and 2015 from cattle isolates, two of which were shared between cattle and wildlife (4). Because spoligotype and MIRU-VNTR are considered stable markers (at least at a time horizon of several years), these 16 molecular types allow identifying 16 independent epidemics spreading in the same area during the same time period. An effective way of representing the structure of contacts between hosts of an infectious disease consists in building networks (41), with epidemiological units as nodes, to which an infection status is associated. Edges linking nodes represent the contacts between epidemiological units that may allow the transmission of the disease agent. Regarding M. bovis transmission between cattle in France and in light of the above, nodes can represent cattle farms and edges may represent direct or indirect contacts between them. Two types of direct contacts may be featured by edges between farms: (i) contacts due to the trade of live cattle (42, 43) and (ii) contacts due to pasture neighborhood between cattle belonging to different farms but with nose to nose contacts over the fence (31, 44, 45). Besides, indirect contacts between cattle farms due the presence of wildlife may also be represented by edges. Concerning the badger, a known susceptible species to M. bovis infection (21, 40), the spatial organization of social groups with stable home ranges around setts (46, 47) allows us to represent indirect contacts with cattle based on the spatial intersection between farm pastures and home ranges (48). The aim of our study was to analyze M. bovis transmission between cattle farms in a south-western area of France using contact networks and molecular types as infection status information. We built different networks featuring possible direct and indirect contacts between cattle farms and analyzed the association between their structure and the observed pattern of infected farms. Materials and Methods Cattle Data The study population was made up of the 1946 farms having reported cattle between January 2007 and March 2016 (end of the 2015 herd skin-testing period) and owning at least one pasture included in a 2,735 km2 study area, an area straddling the border of Pyrénées-Atlantiques and Landes French departments (Figure 1). Pastures were defined as land parcels used by cattle for grazing according to the "Relevé Parcellaire Graphique" (RPG) of 2013 provided by the French Ministry of Agriculture. Two pastures were considered neighbors if the minimal distance between their borders was less than 3 m. Farm sizes (number of bovine females over two years old) and types (dairy, beef, fattening, mixed, small and other herds) were obtained from the French cattle tracing system ("Base de Données Nationale d'Identification" denoted below BDNI) (Table 1). FIGURE 1 www.frontiersin.org Figure 1. Location of the study area (A), at the border between Pyrénées-Atlantiques (south) and Landes (north) French departments (B). TABLE 1 www.frontiersin.org Table 1. Description of cattle farms included in the study population. BTB surveillance data were provided by the French Ministry of Agriculture. Herd skin-testing was performed each year in the study area in communes (the smallest French administrative subdivision) where infected farms had been detected the previous year, as well as in the neighboring ones, using either single intradermal comparative tuberculin tests (SICTT) (in all dairy farms or in farms located in the communes with confirmed infected farms) or single intradermal tuberculin tests (SITT) (in all the other situations), both performed in the cervical region. In the other communes of the study area, herd testing was biennial in Landes department, and triennial in Pyrénées-Atlantiques department. M. bovis infection was confirmed by polymerase chain reaction (PCR) and/or bacterial culture (either following a positive skin test or the detection of a suspect lesion during routine meat inspection at a slaughterhouse) (40) in 69 cattle farms of the study area during the study period; all the cattle of these farms were subsequently slaughtered and molecular typing was performed on each bovid found infected (with a mean of four cattle per farm detected infected during the study period). Molecular typing results were provided by the National Reference Laboratory (NRL) (Anses, Maisons-Alfort). The combination of spoligotyping and MLVA based on MIRU-VNTR allowed identifying 16 distinct molecular types (Table 2). A unique molecular type was identified in all of the 69 detected infected farms, except two where several molecular types were identified. TABLE 2 www.frontiersin.org Table 2. Number of cattle farms detected infected per molecular type during the study period and within the study area. A farm was classified infected by a given molecular type if this type had been detected at least once in the farm during the study period. Because of the geographic differences in the frequency of skin testing, having detected M. bovis earlier in a given farm than in another one does not imply that the former had been infected earlier than the latter. For this reason, the detection dates could not be taken into account. Badger Data Two thousand four hundred and 25 badger setts were identified and geolocalised by hunters in the study area, between 2013 and 2015. Around those setts, considered as main setts (i.e., hosting a social group), we defined badger home ranges using a two-step procedure: (i) a Dirichlet tessellation was first built around all setts [in which the perpendicular bisectors of each segment between two adjacent setts delineate the home range around one given sett, thus assuming that boundaries were located halfway between neighboring main setts (47)] and (ii) to avoid unrealistically home range large sizes, a home range was defined as the intersection of a tile with a 1,000 m-radius buffer area drawn around the setts (48). Two setts were considered neighbors if the corresponding home ranges were adjacent. A sett and a farm were considered neighbors if one of the farm pastures intersected with the badger home range. BTB surveillance data were provided by the French Ministry of Agriculture. In the study area, bTB surveillance in badgers was performed according to the "Sylvatub" surveillance network, which started in 2012 in the study area (49). Surveillance protocol included badger trapping (i) within a 1.5 km-radius around confirmed infected farms, (ii) within a 2 km radius around setts with confirmed infected badgers and (iii) in communes at less than 5 km of communes where confirmed infected farms were located (one badger per sett). Trapping was performed using stopped restraints (https://www.plateforme-esa.fr/filedepot_download/35377/100) and snares were checked the morning after the day they were set up within the 2 h following sunrise, in order to limit the stress of trapped badgers. Trapped badgers were culled by head shot except in a minority of cases where they were found already dead (due to trap related injuries that sometimes occurred when snares were placed on sloping terrain, with no possible alternative). Road-killed badgers were also considered. Stopped restraints used for trapping were placed near sett entrances, those setts being considered as the sett of the trapped animals. Where badgers were found dead along roads, hunters reported the most probable sett according to their knowledge of the area (48). All the trapped and road-killed badgers were tested for M. bovis infection. Among 401 analyzed badgers (4.5% were road-killed badgers), 11.2% were detected infected (45 animals, one was a road-killed badger), of which 39 harbored the SB0821 molecular type and 6 the SB0832 molecular type, both molecular types having also been found in cattle (Table 2). All the badgers trapped could be attributed to 113 distinct setts, of which 33.6% hosted at least one infected badger (32 setts with at least one badger detected infected by SB0821 and 6 by SB0832). Road-killed badgers were attributed to five distinct setts. For four of these setts, the analysis of road-killed badgers did not provide additional information as they had also been subjected to trapping measures. For the fifth sett, the analysis of one road-killed badger allowed the detection of infection (SB0821 molecular type), not revealed by trapping. Setts with at least two badgers tested negative were considered as uninfected (n = 75). All the remaining setts, either with only one badger tested negative or without analyzed badger were considered of unknown status. Contact Network A contact network was built using farms of the study population as nodes, and four types of edges (Figure 2): - A trade edge (denoted T-edge below) from farms i to farm j represented the sale of one or several cattle by farm i to farm j during the study period, at one or several occasions; - A pasture neighborhood edge (denoted P-edge below) between farms i and j represented the fact that a pasture owned by i and another one owned by j were neighbors; - A simple badger-mediated edge (denoted B-edge below) between farms i and j represented the fact that both farms were neighbors of a given sett; - A second level badger-mediated edge (denoted D-edge below) between farms i and j represented the fact that (i) farm i was neighbor of a sett k1, (ii) farm j was neighbor of a sett k2, and (iii) the setts k1 and k2 were themselves neighbors. FIGURE 2 www.frontiersin.org Figure 2. Schematic representation of the four types of edges between cattle farms in the contact network (A), trade edge (noted T); (B), pasture neighborhood edge (noted P); (C), simple badger-mediated edge (noted B); (D), second level badger-mediated edge (noted D). To avoid duplicated edges, the types of edges (T, P, B and D) were aggregated at the edge level. The full contact network thus contained only unique edges labeled by one or several edge types (Table 3). Because the T-edges are directed, each undirected P-, B- and D-edge was transformed into two symmetric directed edges. The full contact network was thus a directed network. TABLE 3 www.frontiersin.org Table 3. Label of edges in the different networks of contacts between cattle farms in the study area. Subnetworks were extracted from the full contact network by restricting the edges to those of specific types (Table 3). These subnetworks are termed below T-network, P-network, B-network and D-network. Similarly, we used edge types to split the full contact network in three non-overlapping subnetworks: - the cattle-specific network incorporated edges labeled T, P or T-P, thus representing only contacts induced by cattle breeding practices; - the badger-specific network incorporated edges labeled B, D or B-D, thus representing only badger-mediated contacts; - the mixed network incorporated all the remaining edges, thus representing the co-occurrence of cattle-specific and badger-mediated contacts. Statistical Analysis Each of the 16 molecular types of M. bovis identified in the study area was considered as a marker of an independent epidemic. For a given molecular type, the contact network may be considered as supporting M. bovis transmission between two farms only if a path exists in the network between these farms. The transmission tree rooted on a detected infected farm should then be entirely located in a single component of the network. The contact network may then be considered as supporting the spread of a given molecular type if most of the farms infected by this molecular type are located in the same component of the contact network. We thus first computed, for each molecular type identified in more than one farm, the number of components in which these infected farms were located (50). For the same subset of molecular types, we also computed, for each infected farm, the length of the shortest path to another farm where the same molecular type was detected. To evaluate whether the observed pattern of bTB infected farms may have resulted from transmission processes in the contact network, we used the k-test proposed by VanderWaal et al. (51). This permutation-based test is based upon the calculation of the k-statistic: the mean number of infected cases among the neighbors of an infected node (the approach is easily extended to neighborhoods of order >1). The observed value of this statistic is then compared to the distribution of the same statistic obtained by randomly reallocating the location of cases, thus simulating a possible pattern of cases under the null hypothesis of an absence of association between bTB case location and network structure. The empirical p-value of the k-test is then the proportion of permutations for which the k-statistic is greater than the observed one. We adapted this test to a multi-type epidemic by redefining the k-statistic as the mean number of cases among the neighbors of a node, which were infected by the same molecular type as that node. The k-test was first performed on the full contact network. It was then applied on the cattle-specific, badger-specific and mixed subnetworks; and this, for two groups of molecular types: those observed in cattle only and those observed in cattle and in badgers. Seven tests were thus performed and the Bonferroni correction was applied. Ten thousand permutations were used to compute the empirical p-value. To further analyse the association between edge types and bTB occurrence, we focused on edges originating from infected farms. A binary status was assigned to each of these edges, with a value of 1 when the destination node was infected by the same molecular type as the originating node, and 0 otherwise. The association between this status and the edge type was then assessed using a case-control design: cases were edges having a status of 1, and controls the edges having the status 0. Four binary explicative variables were defined, based on the types labeling the edge: T, P, B, and D. In addition, we took into account the size (number of bovine females over the age of 2 years) of the edge originating and destination farms, herd size being a well-known risk factor for bTB detection in cattle farms (24). We thus modeled the probability for an edge starting from a detected infected farm to end at a farm detected infected by the same molecular type, using a logistic regression model including six independent variables: four binary variables (presence/absence of the T, P, B and D edge type) and two quantitative variables (sizes of the originating and destination farms). We checked the absence of multicollinearity using variance inflation factors (VIF) with a threshold of 10 (52). Odds ratios (OR) and their associated 95% confidence intervals were computed. Finally, attributable risk fractions (AF) were computed for each edge type. The definition of badger-mediated edges was based upon the neighborhood between pastures and one (B-edges) or two (D-edges) badger home ranges. For some of the corresponding setts, the trapping results allowed defining an infection status: setts were considered as (i) infected when at least one trapped badger had been found infected with an identified molecular type and (ii) uninfected when at least two trapped badgers had been tested negative and no occupant badger had been found infected [for more details, see (48)]. Based on these data, we finally used a Fisher exact test to analyze the association between the status of B- or/and D-edges and the infection status of the corresponding setts. Dirichlet tessellations were computed using the deldir package (53) and buffers using the sp package (54). Network analyses were carried out using the igraph package (55) and variance inflation factors were computed using the car package (56). Attributable risk fractions were finally computed using the AF package (57). All those cited packages were used in R 3.3.2 (58). Results Within the full contact network, the most frequent edge type was the combination of B- and D-edges, followed by single D-, T-, and B-edges. The P-edge type was less frequent alone than in combination with the other types (Figure 3). FIGURE 3 www.frontiersin.org Figure 3. Distribution of the different types and combinations of types for the edges of the full contact network between cattle farms in the study area between 2007 and 2015 (T, trade edge; P, pasture neighborhood edge; B, simple badger-mediated edge; D, second level badger-mediated edge; edges having only one type are in light gray and combinations of several types are in gray). The largest weak component of the full contact network incorporated 99.8% of the study population. Regarding the four edge-type-specific networks, the proportion of nodes included in the largest component was higher in trade and badger related networks (94.4% for the T-network, 94.7% for the B-network and 93.6% for the D-network) than in the pasture network (50.4%) (Table 4) (a more detailed analysis of networks topology is given in Supplementary Tables 2–4 and Supplementary Figures 1, 2). TABLE 4 www.frontiersin.org Table 4. Description of the full contact network and of the four edge-type specific networks. For each of the 16 molecular types, the farms where the type had been observed were always located in the same component of the full contact network. This was also the case for the B-network, but not for the T-, P-, and D- networks (Table 5). TABLE 5 www.frontiersin.org Table 5. Distribution of detected infected farms in the components of the full contact network and in the four edge-type-specific networks for the molecular types identified in more than one farm. Four molecular types were observed in at least two detected infected farms (Table 2). For 87% of these farms, the path to the closest farm detected infected by the same molecular type was made of a single edge. It included one intermediary cattle farm in 11% of cases (Figure 4 and Supplementary Table 2). This result suggests a prominence of M. bovis transmission between an infected farm and its direct neighbors in the full contact network. FIGURE 4 www.frontiersin.org Figure 4. Distribution of the shortest path lengths in the full contact network between pairs of farms detected infected by the same molecular type (only the four molecular types found in at least two farms are considered). We computed the proportion of shortest paths made of a single edge between farms infected (i) by molecular types found only in cattle and (ii) by molecular types found both in badgers and cattle. The difference between these two proportions was not significant (Fisher exact test: p = 0.13). Using k-tests, a significant association was observed between the pattern of bTB detected infected farms and the structure of the full contact network (observed k-statistic: 2.3; distribution obtained by randomly reallocating the location of cases: mean = 0.39, SD = 0.12; p < 7.14*10−3, threshold after Bonferroni correction) (Figure 5). No significant association was observed for the cattle-specific network, neither for the molecular types observed in cattle only, nor for those found both in cattle and badgers. Conversely, a significant association was observed between the pattern of farms detected infected by molecular types shared between badgers and cattle and the structure of the badger-specific network (p < 7.14*10−3). Finally, the structure of the mixed network was significantly associated with the pattern of bTB-infected farms for both groups of molecular types (p = 0.006 and p < 7.14*10−3 respectively) (Table 6). FIGURE 5 www.frontiersin.org Figure 5. Graphical representation of the k-test results for the full contact network [dot-dashed line: k-statistic computed in the observed network; gray density plot: distribution obtained by randomly reallocating the location of cases; this last distribution was clearly lower than the k-statistic observed (p < 7.14*10−3, threshold after Bonferroni correction)]. TABLE 6 www.frontiersin.org Table 6. Results of the k-tests for the cattle-specific, the badger-specific and for the mixed subnetworks of the full contact network, for the molecular types only found in cattle only and for those found both in badgers and cattle. The four edge types were included in the logistic regression model as no significant multicollinearity was detected. T-, B-, and D-edge types were significantly associated to the probability of being a case with an OR of 7.13 for the T-edge type (95% CI: [3.39–15.06]), 1.89 for the B-edge type (95% CI: [1.32–2.76]) and 10.44 for the D-edge type (95% IC: [4.38–26.66]). The size of the destination farm of the edge was also significantly associated to the probability of being a case. Regarding edge types, attributable risk fractions were 84% for the D edge type, 32% for the B edge type, and 12% for the T edge type (Table 7). TABLE 7 www.frontiersin.org Table 7. Logistic model of the probability of an edge starting from a detected infected cattle farm to join another detected infected cattle farm and with the same molecular type according to the type of edge. Among edges representing badger-mediated transmission (i.e., B- and D-edges), the infection status of badger setts involved (one sett regarding B-edges and at least one of the two setts regarding D-edges) was known for 264 edges (5%) originating from a farm infected by one of the two molecular types shared between badgers and cattle. Among them, 44 were case edges (i.e., the destination farm had also been found infected by the same molecular type) of which 38 (86%) were supported by positive badger setts; and 220 were control edges of which 102 were supported by positive badger setts (46%). These differences were significant (Fisher exact test: p < 0.0001) with an associated OR of 7.3 [95% CI: (2.9–21.9)]. Discussion The objective of this study was to provide a better understanding of M. bovis transmission mechanisms between cattle farms in south-western France using networks which represented the direct and indirect contacts that may allow M. bovis transmission among farms of this area between 2007 and 2015. Four types of edges were represented because of their potential involvement in M. bovis transmission between cattle farms and we assumed that they represented the main transmission mechanisms in the study area. Cattle movements due to trade are a known M. bovis transmission route in Great Britain (59, 60), but also in France (42). The neighborhood with an infected farm through adjoining pastures (allowing over the fence contacts between herds) has also been identified as a potential risk factor for the M. bovis transmission between French cattle farms (31). The intersection of badger home ranges with cattle pastures and between each other's was considered a proxy for badger-mediated transmission, considering the territoriality of badgers (36) and the ability of M. bovis to survive in the soil (25, 26). BTB surveillance measures in badgers were not homogeneous among setts of the study area, as they were dependent on bTB detection in the cattle farms in their vicinity. For this reason, although the location of setts was known, we did not model badger setts as nodes in the contact network (we would have been unable to attribute an infection status to each of them). Instead of that, sett location data were used to represent badger-mediated contacts between farms by specific edges, based on neighboring badger home ranges. Two types of badger-mediated contacts were thus modeled by edges. B-edges represented a situation in which two farms neighbored the same badger home range: farm to farm M. bovis transmission through such edges thus only assumed cattle to badger and badger to cattle transmission. Conversely, D-edges represented a situation in which two farms neighbored two distinct but neighboring badger home ranges: farm to farm transmission through such edges thus also assumed badger to badger transmission in animals from neighboring setts. Because the epidemiological unit of this study was the farm, P-, B-, and D-edges were built based on the aggregation of pastures of each cattle farm. In the study area, cattle are often moved from one pasture to another one belonging to the same farm, e.g., when rotational grazing is used, we thus assumed that this simplification was meaningful. The frequency of testing cattle was different in the different parts of the study area and this could have biased our results. However, testing was performed each year in communes where infected herds had been detected, and was also performed reactively in farms identified by contact tracing from these herds, based on cattle trade data and on pasture neighborhood. For these reasons, farms directly connected (in the full contact network) to a herd detected infected were considered having been submitted to similar testing regimens, both for B and D edge types (as in most cases the connected farms were located in the same commune), and for the T and P edge types (because of contact tracing). As only edges originating from herds detected infected were considered in the k-tests and in the logistic regression model, the corresponding results should not have been biased by geographic variations of the frequency of testing in the study area. Taking into account the molecular types of isolates allowed considering 16 independent epidemics, of which 12 appeared restricted to a single farm, and 14 to less than 10 farms. All of these 14 molecular types affected only cattle. This predominance of molecular types found in a single cattle farm (75%) was in line with a previous study carried out in France between 1979 and 2000 in which a large majority of molecular types (84%) were found at a low frequency (less than 10 farms). This result has been interpreted as the sign of a poor spread of these strains (61), which could be traces of older epidemics that would have spread prior to 2007, but without significant transmission afterwards. Indeed, in our study, the 14 molecular types found in less than 10 farms were all detected not later than 2012 (Table 2). Farms detected infected by a given molecular type were always located in the same large weak component of the full contact network that contained 99.8% of farms, whereas it was not the case for three of the edge-type-specific networks: the T-, P-, and D- networks. This indicated that, although the T-, P-, and D-edge-type-specific networks could not alone have supported the spread of bTB infection within the study area (contrary to the B-network), the strong connectivity resulting from the union of the four networks into the full contact network provided a structure that might enable the spread of the M. bovis infection in the study area. This result is in line with multifactorial mechanisms of bTB spread previously suggested by other studies (24, 29). As an example in Great Britain, dynamic modeling of cattle taking into account farm environment helped understanding M. bovis transmission routes (62). Prominent identified routes of M. bovis transmission were moving infected cattle between farms and reinfection from an environmental reservoir. The conclusion of this study was that control measures should simultaneously address several transmission routes to be effective. Using k-tests, a significant association was observed between the pattern of bTB-infected farms and the structure of the full contact network. Moreover, the structures of the badger-specific and mixed networks were significantly associated with the pattern of farms detected infected by molecular types shared between badgers and cattle. This result was expected and confirmed that badger-mediated edges could be viewed as paths for the interspecies M. bovis transmission. In addition, the structure of the mixed network was significantly associated with the pattern of bTB-infected farms for molecular types found only in cattle, whereas it was not the case for the cattle-specific network. We could assume that the spread of cattle molecular types would be more efficient when direct contact (trade and/or pasture neighborhood) are associated with indirect badger-mediated contacts. In addition, we should be cautious about the cattle specificity of these molecular types, as these molecular types may be (or have been) present in the badger population without being observed, because of the relatively low sensitivity of bTB surveillance in the badger population. Considering edges originating from detected infected farms, we used a case-control design and a logistic model to analyse the relationship between the types of an edge and the detection of the same molecular type at the originating and destination farm of the edge (case edges) or at the originating farm only (control edges). Because the detection dates could not be considered in the study to infer dates of infection, the co-occurrence of the same molecular type at both ends of case edges does not model the transmission of M. bovis through the edge, although the edges of the full contact network represent possible transmission paths for the bacteria and case edges thus represent possible transmission events. The largest odds-ratio was attributed to the D edge type, followed by the T edge type. This predominance of badger-mediated edges reflects the specific situation of the study area, where molecular types shared between badgers and cattle were predominant (84% of detected infected farms Table 2), and the predominant effect of the D edge type suggests a probable spread of M. bovis between badgers from neighboring setts, and not only between badgers and cattle. However, B and D edges were defined based on a geographic representation of home ranges, with a maximal distance of 1,000 m to the sett. This distance threshold, the Dirichlet tessellation used to model home ranges, and the fact that some setts may have been unoccupied, are three elements that may have led to an underestimation of home range size, and to an overestimation of the role of the D edge type. The T edge type was also associated with a putative transmission of M. bovis (AF = 12%). This result is in agreement with a previous French study conducted at the national scale, according to which the population attributable risk fraction of bTB infection had been estimated at 12% [5–18%] for cattle trade (42), often allowing long distance bTB spread. In a previous study conducted in France, pasture neighborhood was found significantly associated with the farm infection status (31). However, in the present study, the P edge type was not significantly associated with M. bovis transmission when using the case-control design. This may be first explained by the fact that some of farmers of the study area use rotational grazing, with some pastures left unoccupied for grass re-growth. Furthermore, P-edges were defined based on a direct neighborhood between pastures (<3 m). This short distance does not allow other opportunities of direct contacts between cattle, such as the wandering of livestock, to be represented. The badger-specific edges (B and D edge types) were defined based on sett locations, one or two setts being associated to each. For some of these setts, an infection status could be determined based on bTB surveillance data. We showed that this infection status was significantly associated with the fact that the sett as well as the originating and the destination farms had all been found infected by isolates of the same molecular type (OR = 7.3; 95% CI:[2.9-21.9]). This result supports an actual badger-mediated transmission through these types of edges. Nevertheless, wild boars have also been found infected with M. bovis within the study area. Indeed, among 548 analyzed wild boars between 2011 and 2015, 15 (2.7%) were found infected. The corresponding molecular types found in these wild boars were the two molecular types shared between badgers and cattle. Therefore we cannot exclude the role of this wild species that we could not consider in this study because of a lack of field data that would have allowed its spatial organization (captured through radio tracking, for example) to be represented. Not considering wild boars in our analyses could have led to an over-estimate of the role of B and D edge types in M. bovis transmission between cattle farms. Other indirect contacts through herd practices could also have contributed to the predominance of the D edge type. Indeed, this type of edge created links between farms without direct contacts at pasture but being in a kind of vicinity. As examples, the sharing of material or the loan of animals could create links between farms that may overlap the D edges. However, no data were available to investigate this assumption. Its confirmation or refutation would require supplementary investigation. In conclusion, this study supports the multifactorial nature of M. bovis transmission between cattle farms within the Pyrénées-Atlantiques–Landes area, France from 2007 to 2015. The largest part of bTB spread seemed to be due to badger-mediated contacts, however cattle trade played a significant role. Consequently, to be truly effective, control measures should not focus on a single type of contact but ought to act on the different mechanisms we raised. Author Contributions MB-Z, AC, and BD conceived and designed the study. MB-Z prepared the data for the analysis. MB-Z and BD performed the analysis. MB-Z wrote the manuscript. MB-Z, AC, and BD revised the manuscript. All the authors approved the submitted version of the manuscript. Conflict of Interest Statement The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. 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Edited by: Andrew William Byrne, Agri Food and Biosciences Institute, United Kingdom Reviewed by: Joseph Crispell, University College Dublin, Ireland Helen R. Fielding, University of Exeter, United Kingdom Copyright © 2018 Bouchez-Zacria, Courcoul and Durand. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. *Correspondence: Benoit Durand, firstname.lastname@example.com
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