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Editor-in-Chief: Diana Crow

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Contribu tors: Jen nifer Gil len

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Bard Science Journal April 2013 Volume 2, No.4


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How do I justify my liberal arts degree to my parents when they are both STEM folks? Lots of Bard students are faced with this issue. Sometimes your interests diverge from those of your parents, and that’s okay! Remind them that you’re passionate about what you’re studying, and that a liberal arts education makes you a well-rounded individual. Just remember Montaigne and Galileo – these guys dabbled in every area one can possibly dabble in. Or Goethe: while most know him as the famous 19th century poet, he was actually one of the first people to discuss refraction and chromatic aberration in his Theory of Colors (1840). Science and the liberal arts intertwine in a multitude of ways, and your parents are aware of that. After all, how would STEM be this popular and applicable without anyone to contextualize it? How do I get guys to notice me? As simple as this is going to sound, just smile at them. Or smile more in general! Happiness is by far the most attractive emotion, and smiles are contagious. If a guy sees you smiling, chances are he’ll smile back! Adjusting your body language can help as well. When we sit with our bodies folded into each other (arms crossed, hunched over, chest caved in, etc.) we give off a negative vibe because those positions make us seem uninviting. We look like we don’t want to be approached, or even noticed, even if that’s the opposite of what we’re thinking! Uncross those arms and lift up your chin. You’ll instantly look (and feel!) more attractive. And here’s a tiny bit of science: wearing red increases how sexually attractive you look. Color transmits powerful messages about how interested in you a guy might be – take advantage of that and throw on some red! And finally, you can just tell guys that you’re interested in them. Simply asking someone to coffee can do wonders. Best of luck!

moderation’s just around the corner for a lot of Bardians right now, so here are some general tips: First and foremost, don’t panic. Eat a banana. They contain tryptophan, which is an amino acid that is converted to serotonin (a neurotransmitter typically associated with happiness and overall well-being). Sleep a good eight hours the night before, and come at least 10 minutes early. Read over what you wrote before you go in. Some of the hardest questions people are faced with generally have to do with your future plans (“What do you want to get out of your last two years in college?” “What do you want to do with your degree?”). Tell them what you’re really interested in with regards to your major, how you want to show kids the fun in graph theory or how determined you are to publish your own translation of the entirety of Dostoevsky’s White Nights. Remember that these people know you have infinite potential, and that they’re interested in what you can offer to your field of interest and to Bard.

Introducing a new segment in which a neotenic salamander answers science related questions from Bard students! Axylotls, also called “water monsters,” are adorable salamanders that remain aquatic and gilled. Send in your questions to bardsciencejournal@gmail.com, or to bardsciencejournal.tumblr.com.

What should I do with my CitSci binder? It’s a lot of paper and we never used it. Recycle it! Or make a collage. Even if you personally don’t find the information interesting, the titles of articles alone could look really cool on the walls of your room. And if you’re feeling super creative, you can even make it into a sculpture. Call it “Post-Scientific Immersion.” How do I get people to stop stealing my lab stuff? I autoclave my pipettes myself, and people keep using them up. Tell Maureen if the lab is running out of pipettes! But otherwise you might have to claim some lab or closet space as your own to keep all of your supplies in check.

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ne of the most active and scientifically oriented clubs in Bard College is the Math Circle. Its goal is to expose kids to the cool ideas associated with math, showing them that math isn’t the sort of tedious, pedantic science that some schools make it out to be. The Math Circle brings in logic games and puzzles – all of which are tied to some sort of mathematical property – to fuel the attendees’ curiosity about whatever math topic is being showcased. With regards to how the club got its name, current head of the club and senior math major Jeffrey Pereira explains that “if we take a regular polygon and have each side represent a specific amount of fun, and we keep increasing the number of sides so that n number of sides approaches infinity, the amount of fun approaches infinity as well.” Just like in The Greedy Triangle, a children’s book geared towards mathematics. Founded five years ago by a couple of students and one professor, the program evolved into one of the most successful math enrichment programs in the area, hosting events in Tivoli, Kingston, and Red Hook. The Math Circle prides itself on bringing a new side of mathematics to local students. “It’s less on the tutoring side of things and more on the, ‘I’m really curious about math, so where can I go to get exposed to different aspects in this field’ side,” says Pereira. The Bard Math Circle was quick to disprove all negative associations that local students had with regards to the academic subject. What makes being part of the Math Circle worthwhile is the very contagious enthusiasm for mathematics as well as working with the people who attend its events. This is Pereira’s favorite part: interacting with the kids and their parents. “When you go and you bring a really cool mathematical idea to the table, you’re not only showing it to the student, but to the parent as well. For example, if

n=5

n = 10

you bring in a game that involves graph theory, you can show the kid how the game works, and you can connect the ideas of graph theory to the real world so that the parent sees how important it is in day-to-day life: like how graph theory connects to the internet.” However, there are some challenges to being part of the Math Circle. Pereira emphasizes that one really has to work with the community, which would be the towns of Tivoli and Kingston. Because of the somewhat abrasive relationship between Bard and Tivoli, promoting the club’s events has become difficult. Bard College is sending mixed messages to the surrounding community. While enrichment programs such as the Math Circle try to incorporate education in a fun and engaging manner, other members of our community “act the way they do on weekends,” explains Pereira. “Tivoli and Red Hook are part of the same school district, and unless we find a lot of math majors from Bard, it’s difficult to promote the idea of specifically mathematical enrichment since Bard is not a math-oriented school. There is a definite bias against math, and the Math Circle is trying to combat that and give people good experiences in math.” The Math Circle plays a big part in Pereira’s life, not only within the program itself but also in his senior project. His senior project revolves around a game often featured at Math Circle events: Set. If we take the standard game of Set, we notice that each of the shapes on any given card has three different states for each of the three properties – color, shape, and shading. Pereira invented a variation of the card game where there is a fourth state. He explains, “in the regular Set, every two cards imply a unique third card. To keep that property in the new Set game, I would add a new color, a new shape, a new type of shading, and

n=∞

N represents the number of sides in a regular polygon and the amount of fun one has in any given activity. If we increase the number of sides so that n approahes infinity, we eventually get a circle. Thus, the Math Circle represents an infinite amount of fun.

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a new number of shapes per card.” The rest of his project centers on the probability of finding a set and what changes in terms of how many cards one has to deal out before finding a set. The kids are most excited about the games and toys that math circle brings. This is because “they’re developing logic skills via a medium they’re not used to getting something educational out of. They’re developing really useful skills without it seeming like a chore – kind of like building up muscle by carrying an extra ten pounds in your backpack but without the ten pounds,” explains Pereira. When we volunteered at the Kingston Math Circle a few months ago, we distinctly remember the enthusiasm that Katie, a regular at Math Circle events, had with regards to polyhedral legos. She put the pieces together into a huge blue and yellow hat made up of two spheres. These she could dissemble and put back together within seconds. When it was time to pack up and head back to Bard, she was reluctant to part with her creation and wanted to experiment with the lego pieces for a little longer. It was incredible to see how much Katie enjoyed the mathematical concepts behind the legos. There’s also a wide diversity parents at certain events. Artists and business owners attend, as well as “teachers, engineers, architects and the like, all of whom are trying to push their kids to develop a love of math in a “this is what math has done for me” fashion, but also because math always leads to good things,” Pereira says. “It’s like a tool for the parents as well as the kids now!” Over the past five years, the biggest event that the Math Circle hosted was the American Math Competition 8 (AMC8). The AMC8 is an optional 25-question multiple choice exam designed to further the development of problem solving skills in middle school students and to enhance positive feelings towards mathematics. The club decided to host the exam because the nearest testing center was in Albany, which is inconvenient for many of the participating students. The Math Department invited families within a 50-mile radius of Bard and helped make the event free for the students. It was a spectacular success: Jim and Maria Belk gave a talk on mathematics, food was catered, and the test takers enjoyed a friendly, stress-free environment. “All that work and months of planning,” Pereira tells us, “yielded to about 60 students participating. There were two perfect scores in entirety of New York State, and one of those was taken at our event. We had high scores, 23/25, and people who regularly took part in the Math Circle’s events were on the higher end of the spectrum.” Imagine this: it’s a Friday afternoon in the Tivoli public library. We’re greeted by ten or so lively school kids who are excited to participate in the activities that the Math Circle has planned. We present them with a red wicker basket brimming with math-based puzzles and games. They are instantly entranced and, once their attention is hooked, we sit down with them one on one and subtly introduce math concepts through analogies relating to their interests. Seeing their excitement as they pick up a new skill like the addition of fractions or mul-

“They’re developing logic skills via a medium that they’re not used to getting something educational out of.” tiplication makes math circle an incredibly rewarding experience. Students immediately get to work playing, unaware that they are learning about geometry through tan grams and basic game theory through Set. Despite the general stereotype that kids dislike math in school, the personal attention and game-based learning makes them feel as if they aren’t working and let them leave with a new trove of knowledge. If the Math Cirlce sounds like something you would enjoy, email Jeffrey Pereira at jp3275@bard.edu.

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or the first time, Bard College’s Center for Civic Engagement has partnered up with Chancellor Livingston Elementary School to connect eager Bard students with teaching opportunities in the Elementary School. Since February, around twelve Bard undergraduates have been travelling to Rhinebeck Elementary School to mentor 4th and 5th grade students who are participating in the annual Rhinebeck Science fair. In this science fair, students are not only expected to develop and conduct independent experiments, they will also be presenting them on March 20th to their peers and the community. The students participating in this fair are walked through the process of conducting an experiment by their Bard Mentors. Using his or her question of interest, each participant forms a hypothesis and a null hypothesis, designs their experimental methods, and analyzes and compiles their data. For the 2013 Science Fair, the participants are conducting experiments on a wide range of scientific topics. Abdullah Nassim,

’16, a mentor for the science fair reports that his students plan on investigating not only the water content of fruit but also how much voltage different fruits possess. Another mentor, Rebecca Lansbury ’16 states that her students plan on measuring the effect of plant growth in different environments. When asked of their interest in mentoring the young scientists, the majority of the Bard mentors cited their interest in working with kids in the future. One of the mentors, Noah Richmond ’16 replied, “ I want to be a teacher and I also enjoy spending time with kids”. Bard senior Tiago Moura cites, “It is fun, enjoyable; It is interesting to see little kids to science!” Not only do the students get support form the Bard mentors, but also, some of their teachers provide them with the materials and resources needed to succeed. These materials include, a worksheet detailing the scientific method and materials for their experiments if available. However, the most important ingredient, the participant’s enthusiasm and motivation to conduct their experiments outside of school comes purely from the participants. All participants have to conduct their experiments and record all their data at home, without the supervision of the Bard mentors. However, help with analysis and compilation of their data is always available from the Bard mentors. After a brief period of inactivity, the science fair was relaunched in 2010 with the sponsorship of the Rhinebeck Science Foundation. This foundation funds many activities of scientific outreach in Rhinebeck including, a math fair at the Chancellor Livingston Elementary and also the established a LEGO engineering pilot program in the elementary school.

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ard is talking small steps to make big change. Three years ago, a few students from Bard decided to form the American Chemical Society Chapter. They brought their idea to the attention of their professor Christopher LaFratta, who agreed to be the chapter’s adviser. The ACS wanted Bard students to start volunteering at the Red Hook middle school. Dr. LaFratta sat down with the middle school faculty and asked them “How can we help you?” The Bard College ACS chapter and the middle school faculty came up with a three-phase plan to get young minds excited for science. Let’s be honest, by the time kids are in eighth grade for many there is a stigma associated with chemistry, biology and/or physics. Those subjects are often deemed purely for certain ”types” of students who are more or less mathematical thinkers. The writers, artists and those who haven’t even begun to figure out their passions are essentially left behind. Bard’s science society believes that there is a scientist in everyone; it is all about finding the right way to turn on the switch. In the fall of 2011, the three-phase plan began with the first phase known as “The Bard Science Day”. During this day, eighth graders from the Red Hook Middle School came to Bard and conducted their own science experiments using the Bard Science laboratory equipment. Bard students volunteered and helped the eighth graders in their experiments. The eighth graders extracted DNA from strawberries, played with robots, measured the concentration of salt in Gatorade using a flame photometer test, and tested the motion of pendulum.

During the second phase, Bard students Olja Simoska and Madison Fletcher gave lectures in the 8th grader’s classroom. They taught eighth graders about the history of the atom and about the discoveries of the proton, neutron and electron. The third phase consisted of volunteers mentoring and judging the students’ annual science fair projects. The science fair allowed students to tap into their creativity and combine it with their new knowledge of science. Chemistry major and an active volunteer, Olja Simoska, says, “Science is best learned through the experience of experimentation and doing research. Reading a science textbook cannot teach me what a lab experiment can. It’s just not as fun.” The notion that science can be fun seems to be a relatively foreign concept but extremely essential to inspire new generations of scientists. This project has also received honorable mention for the year 2012 by the American Chemical Society. The students who volunteer for these activities do not have to be science majors; they are trained in performing all aspects of the program’s experiments. This project started out with the hope to introduce a little bit of science into the middle school community, but has spread into an enterprise dedicated to encouraging new and unlikely minds into scientific fields. This works perfectly with Bard’s philosophy to integrate all subjects into the curriculum to inspire those to attempt something out of their comfort zone of one’s intended major.

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s an atheist, I like to think that my way of looking the world is a step away from bigotry. To me, the world isn’t divided into good and bad; it’s an enormous mishmash of interconnected people, organisms, and molecules, where every action has an equal and opposite reaction. Contrary to what religious leaders tell their followers, this “godlessness” isn’t a detriment to my moral development. Because I know that what I do has an effect on other people and organisms, I know that my actions matter. I don’t need to know that a supreme being is tallying my rights and wrongs, because I can see the effects of my own actions in my immediate surroundings, even if the differences are small and subtle. That’s not to say that all atheists think this way, or that those of us who do are all perfectly attentive to the effects of our actions all the time. Neither of those is true by a longshot. But I like to think that people who subscribe to an empirically-grounded form of atheism are more able to recognize their own hypocrisy. Unfortunately, they are not. I recently read an essay by an academic named Jackson Lears critiquing a neuroscientist and author named Sam Harris. Harris, a vehement “New Atheist” had written a book in the wake of 9/11 called The End of Faith: Religion, Terror, and the Future of Reason, attacking religion and Islam in particular, as the cause of all social problems. Harris argues that without religion people would not commit most of the evil acts they do and that religious beliefs should no longer be tolerated since they impede the otherwise inevitable development into a moral and scientific society. Lears rips into these assumptions, pointing out Harris’s relentless bigotry against believers and the illogic of his blind faith in the curative powers of science and technology. It’s a blistering essay; Lears thoroughly destroys Harris’ credibility as an authority on the subject. One point in particular felt like a blow in the stomach: Harris was not a lone crackpot but rather one of several “intellectuals the media chose to anoint” as champions of the scientific cause and one of the self-proclaimed New Atheists, who argue that religion should no longer be tolerated due to its long track record of doing harm. In other words, Harris’ ideas weren’t coming from the fringe; these ideas are well established in atheist discourse, and many leading atheists advocate these ideas. Among them, Richard Dawkins. It would be hard to overstate the impact Richard Dawkins has had on the way I think. I first got into his writing in high school, reading excerpts from his book The Selfish Gene for a philosophy club discussion. In that book, Dawkins introduces the concept of memes (not to be confused with internet memes), small units of culture or thought that behave like organisms in an ecosystem of ideology. By characterizing units of thought in terms of biological principles, it’s possible to formulate a natural-selection-driven model for explaining and predicting how different ideas interact with each other. For example, fiscal conservative

memes like “Raising taxes on corporations kills jobs” have developed a symbiotic relationship with socially conservative memes like “Marriage is the union between one man and one woman”, because the partnership between these groups of ideas allows both to colonize new territories. People who are inclined to accept socially conservative memes into their intellectual ecosystems will be more likely to accept fiscally conservative memes if the two seem to be a package deal. The more I thought about it the more sense it made: Just like organisms in the physical world, ideas must be able to spread and reproduce themselves (through speech, writing, art, etc.) in order to last more than one generation. And just like organisms, small, simple, energetically efficient memes are easy to recreate and remember spread faster than larger, more complex ones; replicating these big ideas in other people’s minds requires more effort (and inevitably, there will be some mutations), so these big ideas be powerful enough to displace other competing memes that fulfill its niche. Even though attempts to reproduce and spread memes are often consciously initiated by human beings (e.g. publicity campaigns, sermons, works of art), they are still subject to the same laws and limitations that govern natural selection. This idea changed me. Up until that point in my life, I had viewed science and humanities as separate spheres; I was a writer, and science, when I had to learn it, was an imposition. A well-intentioned and sometimes interesting imposition, but science wasn’t something I was going to use very much myself. Dawkins blew my high school mind wide open and set me off on the path toward becoming a science writer. So it was a shock to me to read his name on a list of overzealous anti-theistic bigots. Of course, I had always known that Dawkins was a vociferous atheist, but I had always seen his snarky and dismissive remarks on the subject of Creationists as the natural attitude of someone who’s spent their entire adult life studying evolution and its implications. Dawkins is a really sharp thinker, I thought, Surely, he’s not as delusional as this Sam Harris guy. He is. Even a cursory glance at his work on the subject of religion and atheism is pretty conclusive. Dawkins is a vehement anti-theist. His main book on the subject, The God Delusion is everything its title suggests: a vicious attack on everything related to Western religion. Dawkins insists that he is an equal opportunity anti-theist, writing, “I decry supernaturalism in all its forms, and the most effective way to proceed will be to concentrate on the form that impinges most threateningly on our societies”(Dawkins 36), but this is really a paper-thin excuse for spewing insults at the three Abrahamic religions. The book is saturated with diatribes against Christianity, Judaism, and Islam. For instance, in a chapter called “The God Hypothesis”, where Dawkins sets himself up as a scientist evaluating evidence about whether or not there is a God Bard Science Journal April 2013 Volume 2, No.4

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(in the general sense of “Is there a supernatural being with intent and power to work its will upon the world?”), he still takes the time to throw in barbs, such as, “It is unfair to attack such an easy target. The God Hypothesis should not stand or fall with its most unlovely instantiation, Yahweh, nor his insipidly opposite Christian face, ‘Gentle Jesus, meek and mild’”(Dawkins 31). For Dawkins, finding new and creative insults for the monotheistic establishment seems to be at least as important as the logical argument against the existence of God that his book supposedly hinges on The more I read, the more stupid I felt for not having noticed the level of bigotry inherent in Dawkins’ ideas about religion. They had always been there, even in the more biologically focused books like The Selfish Gene and The Ancestor’s Tale (albeit as sidenotes rather than central theses), but I had always blithely read past them. They completely blended in with my ideological preconceptions; as a lifelong atheist with a penchant for writing self-assigned reports about the Permian-Triassic extinction, I felt like annoyance and even rage at Creatonist attempts to obviate evolution from school curricula were completely justified reactions. But, in reading The God Delusion, I had to face the hypocrisy and inconsistencies inherent in the anti-theist position square-on. Hinduism, Buddhism, and Confucianism are only mentioned three times each in the entire book. “Indeed, there is something to be said for treating these not as religions at all but as ethical systems or philiosophies of life” (Dawkins 38). In addition to highlighting his hypocrisy in terms of which religions he chooses to attack, this statement begs the question of “What exactly does Richard Dawkins think a religion is?” He seems to view it as a system of beliefs that affirms the existence of a God, which he defines as a “supernatural intelligence who deliberately designed and created the universe and everything in it, including us” (Dawkins 31). In other words, the only religions that Dawkins attacks are the ones whose narratives conflict with the theory of evolution. And, evolution, is, of course, at the center of all of Dawkins’ writing. In other words, this isn’t an attack on religion in general. It’s a demagogue from one ideology trying to displace the older, established ideology that has long oppressed the non-believers. All I can think of when I read these passages is Paolo Freire’s Pedagogy of the Oppressed, specifically one passage where Freire wrote, “Almost always, during the initial stage of the struggle, the oppressed, instead of striving for liberation, tend themselves to become oppressors or ‘sub-oppressors’. The very structure of their thought has been shaped by the contradictions of the existential situation by which they were shaped. Their ideal is to be men; but for them, to be men, is to be oppressors.” (Freire 45). In Dawkins’ case, as a well-educated white guy and a devout adherent to Darwinian thought in a world dominated by other well-educated white guys who use their education and access to the altern to deride and discredit competing ideologies, is fighting back by using the exact same strategies to discredit religion and glorify atheism. The bullied kid has become a bully himself. This type of atheist bullying is everywhere. Just yesterday, I logged onto Reddit, and one of the first things I saw was a picture of Dawkins standing at a podium with the caption: “The only difference between the Christian God and a mass murderer is that the mass murderer actually exists”- Richard Dawkins. The post had over 5,000 upvotes. Granted, there were slightly under 4,000

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=0A62:/%?0%>.<2 downvotes, but there’s no denying that the atheism subreddit has become synonymous not with discussions about the non-existence of God but rather a dumping ground for opposition to the deeply held beliefs of the religious right. In contrast, religious and conservative bloggers complain that liberals don’t take them seriously, and honestly, I can’t say that they’re wrong. There are still aspects of this debate that I’m ambivalent about. I still think the fact that Creationists try to keep evolution from being taught in schools is unconscionable; biology cannot be taught without it, and without a basic background in biology, there’s no way people can make informed decisions about environmental health issues or even their own healthcare. I feel like I have a responsibility to demand that evolution is given its rightful place, as a central and indisputable force in our world. But at the same time, who am I to demand that others recognize the exceptionalism of my own belief system? If atheism leads to bigotry of the Dawkins variety and a blind faith in natural selection, just as religion can lead to bigotry and blind faith in “God”, then maybe atheism doesn’t merit exceptional treatment in the church-and-state debate... I do know this much: ignorance cannot be fought with ignorance. An atheist who believes in evolution simply because it’s the party line that good atheists and agnostics are supposed to subscribe to is no better than someone who believes in Creationism. An atheist who argues that religion should not be tolerated simply because it conflicts with his or her personal beliefs is no better than a believer who does the same to atheists. The antidote to bigotry is not the removal of religion but rather thought and investigation. I like to use empiricism and memetics to develop my understanding of the world, and as such, I cannot completely dissociate myself from Dawkins. But I can continue to question and interrogate what others, even the authorities who’ve shaped my thinking, tell me.


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A translation of Hydraulic Fracturing Threats to Species with Restricted Geographic Ranges in the Eastern United States.

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igh-volume horizontal hydraulic fracturing (fracking) is a process used to extract natural gas, and it has recently gained a lot of attention for its potential harmful effects on drinking water and human health. The process consists of drilling vertically and horizontally through natural gas shale beds, and then pumping water, sand, and a mixture of chemicals at high pressures into the shales, which releases the natural gas. Some of the sand and chemical water mix returns to the surface as “frack water,” which includes benzene, a known carcinogen, many other toxins, and salt from the gas shales. It’s understandable why most of the focus has been on drinking water and public health issues, but impacts on biodiversity and species’ habitats also need to be considered. Native plants and animals will be affected not only by the toxic chemicals, but also by the salt, truck traffic, habitat fragmentation, air pollution, and noise that will result from fracking. We live near the largest occurrence of commercially exploitable gas shales: the Marcellus-Utica shale region. This region, located mostly in New York, Pennsylvania, Ohio, and West Virginia, supports high species diversity and many endemic species with narrow habitat affinities and small geographic ranges. The paper I wrote with Erik Kiviat, who is a wetland scientist and the executive director of Hudsonia, reviews the potential impacts of fracking on fifteen species—all of which have geographic ranges that overlap with 35% or more of the Marcellus-Utica shale region. We studied each species’ habitat needs, natural histories, and legal statuses to determine how they would be affected by the introduction of fracking to their habitats. The species reviewed include eight salamanders, two fish, three plants, one mammal, one butterfly. The eight salamander species selected for this study are lungless plethodontids (family Plethodontidae) that respire solely through their skin and thus require continuous contact with moisture. This family of salamanders is particularly sensitive to water quality, and the frack water that comes back up to the surface may be harmful to them, especially for those with aquatic larvae. One of the species, the West Virginia spring salamander, is found in only one West Virginian cave. They are dependent on the stream that flows through the cave, and with only about 250 mature individuals, this species could be at risk for extinction if the stream becomes polluted with chemicals and salt from the frack water.

Salamanders also have difficulty crossing roads—in some studies, roads have reduced salamander movement by up to 51%. Roads are necessary for fracking installations, as trucks have to transport so much water and other materials back and forth, and these roads could be detrimental to species of salamanders that have small habitat ranges. Salamanders are already affected by human activities like logging, pollution, and habitat destruction, and the habitat fragmentation, truck traffic, and water pollution associated with fracking will intensify the threats these creatures already face. The two fish selected for the study, the tonguetied minnow and the bluebreast darter, both require good water quality, and the darter is critically imperiled in New York, and imperiled in Ohio and Virginia. The probability of water pollution from fracking wastewater is high, and thus, both of these species are particularly vulnerable to fracking. Plants will be affected by fracking through habitat fragmentation and increased soil salinity levels. The northern blue monkshood is listed as federally threatened, and because it is sensitive to disturbance and grows slowly, fracking could cause it to become endangered or extinct. Developmental practices like fracking, that cause forest fragmentation, often aid in the spread of invasive plants, which could also compete with and threaten these native plants. The mammal selected for this study, the Appalachian cottontail, was only recently separated from the New England cottontail, and because we do not yet know what its habitat needs are, it cannot be properly managed. Local populations of the species are declining due to a number of factors, including habitat destruction and forest fragmentation. A main source population for the species is located in West Virginia, a state where fracking is common, and if this large population were disturbed, those smaller populations dependent on the source population for gene flow would also be negatively affected. The Appalachian azure, a native butterfly, lives in deciduous forests and has difficulty moving between forest fragments. Their larval food plant, black cohosh, is threatened by both non-native plants and white-tailed deer, both of which are likely to benefit from fracking. In conclusion, fracking poses a large number of threats to a diverse group of species, ranging from salamanders and fish to butterflies and plants. Some of the species reviewed are not well known enough to successfully manage or understand how fracking would affect them. An average of 30 acres of forest is impacted with each fracking site (including edge effects), and this impact alone

An average of 30 acres of forest is impacted by each fracking site.

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will be harmful to many populations. Fracking is not yet allowed in New York, so there is still an opportunity to study and protect some of these species. And there are countless other species that were not reviewed in the paper that could be severely affected by fracking, perhaps some that have not been discovered yet. These last paragraphs represent my opinion; I am not speaking for Erik Kiviat. Fracking, especially if allowed in New York, needs more regulation—we can’t pump a mixture of unknown chemicals into the ground, with much of that returning to the surface, and not ;:110B:>%C0//:1

expect it to harm the species that live there. And we shouldn’t allow companies to have the freedom to drill wherever they want to when the process they use has not been properly studied. It will take years to study how fracking affects biodiversity (and drinking water), and many people do not want to wait, but is the natural gas gained now worth the long-term problems it may cause? We also do not know how long the chemicals will remain in the environment—we don’t know if an ecosystem can recover after fracking stops, or if those chemicals will continue to shape and alter the environment even after fracking installations are removed. I understand that each company’s goal is to make a profit, but is it still considered profitable if the costs, in this case environmental costs, outweigh the benefits? I don’t think a monetary value can be placed on biodiversity, and its importance cannot be emphasized enough. Biodiversity is important to the health of ecosystems, and the elimination of certain species could have serious detrimental effects on ecosystem functions. We do not know what every species’ function is, or how species’ interactions affect their ecosystem. But our lack of knowledge does not mean biodiversity is not important—in fact, it means the opposite. We might not know how important a species is until it is eliminated, but we don’t want to reach that point. Let’s learn from the past—think of how DDT affected wildlife through biomagnification, how PCBs still persist in the Hudson—and thoroughly study how a novel practice will affect the environment before we use it throughout the state (and the country). And keep in mind that biodiversity isn’t threatened by fracking alone; habitat destruction, climate change, pollution, invasive species, and urbanization are other threats, among many more, that species have to endure. Consequently, we cannot study one of these threats without recognizing that their cumulative effect is what endangers so many species, and what may, ultimately, drive them to extinction.

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baby is born, and within an instant, she both colonizes and becomes colonized. She enters this world suddenly. Sights and sounds all around her. Her world becomes filled with vibrancy and texture—bright lights, doctor’s hands, and loud noises. We’ll welcome her, the latest passenger on a turbulent ship. Her body, once immaculate in the womb of her mother, becomes host to a vast assortment of microbes the minute she passes from womb to world. To the microbes, the neonate’s body is a pure frontier, a vast land to be inhabited. The baby will grow, and eventually, she’ll come to be composed of more bacterial cells than human cells. But these bacterial cells are not unforgiving invaders of her body—they are part of her. They contribute to the metabolism and homeostasis of her body like little worker bees. Undoubtedly, some microbes the child may encounter will cause her harm. However, these select few are outliers that give the rest of the microbial community a bad name. In other words, many microbes are not only helpful, but also required for proper homeostasis in our bodies. The array of microbial cells found within a specific place of the body is known as a microbiome. The baby will develop different microbiomes in different parts of her body, depending on where certain microbes thrive best. She’ll have a skin microbiome, a gut microbiome, a vaginal microbiome, and so on. Each community is unique in their composition and effects.

For instance, it has been found that the composition of Lactobacillus bacteria in the vagina, directly correlate with the likelihood of females contracting the disease bacterial vaginosis, or BV. It was found that if females lack the normal amount of Lactobacilli in the vagina, they have higher odds of contracting BV. Microbes are an integral part of a body’s makeup; if the balance of bacteria in a microbiome is disrupted, then disease can occur. For instance, if a woman uses douching agents that kill off helpful Lactobacilli, then she may indeed be putting herself at risk for BV (White et al. 2011). Maybe eliminating our body’s natural microbial agents can impact our health, but how did we acquire all these bacteria if they aren’t innate? Let’s go back to birth. When a baby is conceived, she’ll get a set of genes from her parents. And maybe acquisition of the microbiome is not much different. In 2010, a team of scientists headed by Dr. Maria Dominguez-Bello, studied the effects of vaginal birth versus Cesarean section birth on the development of microbiomes in the neonate. They found a pattern of initial microbial colonization: If the baby is born vaginally, the microbes of her mother’s birth canal colonize her. However, if the baby is born through Caesarian section, the microbes of her mother’s skin microbiome colonize the child instead. It is logical—if the baby touches skin fist, she gets skin bacteria, and if the baby goes through the vagina, she gets vaginal bacteria. Vaginal-born infants are dominated with Lactobacillus, Prevotella, and Sneathia species, whereas C-section delivered infants are dominated by Staphlococcus, Corynebacterium, and Propionibacterium species upon birth (Dominguez et al. 2010). According to Dominguez-Bello’s research, mode of delivery is vital in establishing initial communities of bacteria in a baby. However, it is not as clear how this initial colonization may affect the baby as he or she grows into adulthood. To determine the long-term health differences between neonates born through c-section versus and those born vaginally, there is a need for greater longitudinal studies. Since the study of microbiomes is a relatively new and rapidly expanding field, this logical next step in research will certainly be in the works for years to come. Colonization is changing. Years ago, before the perfection of Cesarean sections, microbiomes, and in turn, life-long immune processes, could have looked very different than they do now, with the advent of modern medical practices. When a baby is born, her own life is not only established, but also the lives of millions of little bacteria. Maybe it’s time to pay our respects to these colonizers of our body, as they thrive, work, and integrate with our human cells and bones. Works Cited Dominguez-Bello, M. et al. (2010). Delivery mode shapes the acquisition and structure of the initial microbiota across multiple body habitats in newborns. PNAS 107. 11971-11975. White, B, et al. (2011). The vaginal microbiome in health and disease. Trends in Endocrinology and Metabolism. 22. 389-393.

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This was the first time Harry had done the boogie. The Mayflower Compact had definitively integrated childlike wonder into all of their sparkling eyes. It was a site to behold, the two of them getting it on. Like a pair of frogs in a quickly evaporating South-Saharan pool. Her fathers hadn’t seen her in days, but she hadn’t written him. The purple pumkin-tree skins were smoldering over the open flame and all were gathered round. __________________________ Behavioral critics had pointed to democracy as the major cause of the lunar eclipse. But as the Thirst Echelon vied for status, the Slambovian army hauled ass to make sure the chimps got their way. The chimps got away. Earth was left barren with dust-dusted cupcakes on top of muffin-top courtship errors. Enemies beheld flotillas upon flotillas of frozen bananas and fecal matter, on top which dung-beatles and cockroaches danced a merry dance. Kangaroo brisket was all they could eat after the 3rd apocalypse, and there was nothing left in the sky but Saturn. Pico and his 3-legged dog Martin took the steamboat across the lagoon to loot for some metal, as they did on most lunar eclipse days. Today they happened upon an ancient prosthetics factory, a graveyard of silicone and valleys of uncanny arms and legs. After a ruffle through the rubble, they splashed around in the beer fountain outside the remnants of the great fallen University. The liquid was stagnant and tepid, just right for a summer day on the southernmost tip of the Continent. Melinda came up to the outer lip of the pool and swung her purple mane to the side, displaying her perfectly rotund memory glands. She must have been spending all of her time learning these days, and it was obvious. Pico imagined fitting his hard third eye into her external memory reception socket, and the subsequent eloquent mist of delicious precious knowledge and subjectivity that might flood his thirsty dendrites. He had never shared Knowledge with a female before. His third eye was embarrassingly engorged with neurotransmitter as he pondered Melinder, which is what he liked to call her when he spoke to her in his mind, and he had to dash to the ancient University library to find something, anything, to quench his unbearable desire. He felt he would burst at any moment, spewing immaturity and ignorance everywhere and feel horrendously sophomoric in front of the purple and divine Melinda. As for Melinda, she did love Pico, thought she would never said so. She loved him in a way that made her face numb whenever she went near a cactus. __________________________ Elvis felt this way often. Laxatives weren’t his favorite dessert, but they did the job, especially in this desert. Survival was priority, and that meant appeasing the gods with daily offerings of hastily excreted jumbalayah. Sundae enemas were also tolerated under the scriptures of the Temple, depending on which hemisphere’s interpretation one ascribed to, but they lacked the spiritual exorcism of intestinal heaving. The gods were most gleeful when they saw a bead of sweat on the brow, but they accepted any form of worship. Worship was worship. Enemas were a quick and dirty form of ritualized appeasement, and could be purchased as easily as a gasmask or mantid hallucination kit at any gasmask station.

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Speaking of mantid hallucination kits, or MHKs, children and adults were grappling with each other to obtain this apparently limited resource now that Limes had been obliterated. Bootleg copies of the mantid hallucination kits were widely distributed, with claims that these were the very last by way of authenticity, and that they even contained the blood of the last mantids seen on earth. Teenagers bragged to their friends about their latest MHK experiences, though really all they had experienced was a brief and intense niacin flush, followed by a pancreatic jolt that sent a few repressed memories into temporary frontal lobe con sciousness. Still, the vendors of such purported MHKs made dollops of profit, sending the dough to their families back at the starvation centers in the plasma-harvesting communities. Today was the Lime Parade, being that it was the third lunar eclipse since the coming of the 3rd apocalypse. Bootleg MHKs were being disseminated in droves, and little girls from the Temple, adorned in pearls and the customary lime-green gauze, gnashed their canines and serpent tongues at the lascivious livestock owners as they passed by in ceremonial display. It was a day of slimy, inauthentic appreciation for the temporary embargo on xeno-manufactured goods that had been achieved by the newest ambassador. For some, that meant a boost to Earth-based businesses. For others, it meant waiting for the next dust storm before being able to harvest, produce offspring, and make their pilgrimage to the Hub. The ambassadorâ&#x20AC;&#x2122;s job, after all, was not to represent the dwellers of the Hub domain, but to assert dominance over the threat of the rapidly multiplying and over-crowding settlements of the plasma-harvesting communities. On this facade of an auspicious day, the trum pets were festooned with articles and pronouns of the weekly bias-fest. Fractals danced across the sunrise and the buvalo loved it. They swung their gigantic juggalos into a frenzy of song. Strife was purple today and reached forward to grab the nearest exit.

__________________________

Crab-boy and his puppet Jay remained in the picture frame, hiding from Pope Luci fer. Jim Lucifer resented his twin for stealing his wife and money, but oddly enough his largest pitfall was the constant crippling shame he felt over his last pair of boating shoes. The only rule--the number one rule, he always said--was Donâ&#x20AC;&#x2122;t leave your hunter behind. Well, that, and Donâ&#x20AC;&#x2122;t leave your boating shoes out or the scorpions and rattlesnakes will get them. They use them as bait. __________________________

Female dogfish sharks can sense the fear emanating from the desperately vibrating sperm inside each canine. They used this fear for echolocation and for triangulating the field of pheromonal hemorrhage of the predominant species in their zone. This adaptation became particularly useful when dogfish became able to navigate on land, due to the change in both atmospheric humidity concentration and gill apparatus evolution.

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If you add enough oxygen and energy you can get anything to burn I’ve got big lungs and a strong heart and I leave a trail of sparks Incineration is kind of my thing. But being a child of fire leads to a strange kind of desire you find yourself in emotional badlands striped down dry and everything cracks if you let it. I guess long story short or short story shorter I’m better at lighting matches than I am at quenching flames I prefer letters to text messages because a paper trail at least, you can burn. And don’t worry I know what flames can do The woman who would have raised me if I let her nearly lost her house to an inferno from the west. And some nights I wake up shaking thinking ‘what would I take from the fire?’ What would it take from me? But there’s a part of me that wants flint shoes and a hole in a backpack full of gasoline. Tinderbox heart. Tymbal machine. 600 years ago I would have been hunted and burned for having hair the color of burning. Witchery comes easily to redheads, I’m told. I’ve heard Adam’s first wife was ginger. Yes, Lilith came before Eve. She was banished and God decreed paradise was no place for a wildling craving autonomy and with an anomaly in her pigment anatomy. She had a little too much spark. And yet descendent that I am of history’s catalysts, derivative of these flawed disciples of flame, I’ve been carrying a bucket of H2O lately, And I flicker every time I hear your name. See! You didn’t expect a love poem. I love the unexpected. But it’s harder to burn as of late. The oxygen’s gone. You take it every time you do that wicked smile that just reminds me that everybody falls for you, Plummets for you, And I’m just pilling on. I leave people smoking. You leave them in ruins. And no pun intended but at least fire sparks new life.

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They used to set fire to the badlands. But earthquakes are good for nothing except heartache And cheep demolition. It’s like a knife against a sea of rubble. It is a book of matches in the pouring rain. It’s the fact that you think I’m ordinary. Just another lass falling for you again and again. Hate to break it to you, sweet heart. My first rodeo, this is not. People say that you’re an out of sight, out of mind, kind of guy. What’s that make me. Blind and beyond belief? I’ve been in a stalemate with the horizon for years. And who needs lock and load when I’m a walking fuse. I’m not I’m not scared of leaving a harbor I’m scared of getting back to land and finding I can’t run Everybody knows I’m clumsy to begin with Everybody knows the mirror must have been unlucky in love because i’m a little more singed around the edges every time I catch my reflection I want to wear heals too tall and a smile too wide because everyone knows the girl with the sexiest walk has the most to loose and the one thing that I refuse loosing is the one, if I had a choice, I wouldn’t choose. This is the part where I say I hate you I don’t hate you. This is the part where you remember it’s a script. This is the part where I say I hate you I can’t hate you. It’s a script but I mean every word of it. Has it ever occurred to you that fireflies are worse than butterflies in your stomach? Torch metal and the particles realign. Which is why it sucks that I’m combustible. A little heat and metal’s matrix is fine. Everything cracks it you let it. Truthfully, I hope it’s me. That you’ll come spooling out of my stomach like thread I was saving for winter, for all the little repairs. But if it’s you that cracks I make gorgeous kindling. My hands are always cold but I would keep you warm. And wouldn’t a Russian nesting doll life be nice? You inside my heart inside your heart inside mine There’s days I wish the fire would tuck its know-it-all self into some forgotten chimney. I’d cut off all my hair and be sensible and good. Would you recognize if I didn’t laugh like a hyena? I’d rather not take the chance.

-./012%34/256

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!"#$%&'$()*+$,-../+#0$)/$%/123 “Don’t eat me, bro.”

Macrophages (immune cells that eat microbes and cancer cells) and other blood cellls have a secret handshake that lets them recognize each other. Blood cells have a surface marker called CD47, and macrophages have a surface marker called SIRP-α. They fit together like a lock and key, and when they bind to each other, it sets off a chemical reaction that stops the macrophage from eatng the blood cell.

%"#4$5&46#1$789&62+$)"#$,8:4&;3 “Don’t eat me, bro.”

Some cancer cells also have CD47 surface markers. When they encounter macrophages, CD47 and SIRP-α bind, producing the “don’t eat me” signal, and the cancer cell escapes.

%"#4$,68#4)8+)+$<00$<4)8=5>?@$<4)8A/08#+3 “Don’t eat me,”

“Eat me. Eat me now!” The anti-CD47 antibody fits into the CD47 receptor slot and stops the cancer cell’s CD47 from binding to binding to the macrophage’s SIRP-α. Shutting down the CD47-SIRP-α interaction allows macrophages to bind to other cancer cell surface markers and recognize the cancer cells as enemies. Then the macrophage eats the cancer cell. (-%2%'63;

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8LI4SXIRXMEPSJÂą(SR´X)EX1I²7MKREP'(EWE8EVKIXJSV%RXMFSH]8LIVET] 78%?0212%A>.@ The immune system is best known for its role in suppressing infectious disease, but it also plays a crucial role in protecting the body against tumors. Studies have shown that selectively immunodeficient individuals are more likely to develop spontaneous tumors than their immunocompetent peers, even without exposure to any carcinogenic materials (Jaiswal et al 2010). In the past, most investigations into the role of the immune system in tumor suppression have focused on the learned immune system on the premise that cancer cells produce â&#x20AC;&#x153;not self â&#x20AC;? antigens that can be recognized by B-cells and Natural Killer-cells (Jaiswal et al 2010). However, more recent studies have shown that the innate immune systemâ&#x20AC;&#x2122;s macrophages, which recognize mutant â&#x20AC;&#x153;self â&#x20AC;? cells by their abnormal surface markers, also play an important role in killing off tumor cells (Jaiswal et al 2009). Researchers have found that it is possible to give the innate immune system a boost by adding monoclonal antibodies (mAbs) that to bind to surface markers expressed by cancer cells. These antibody-macrophage interactions can either increase antibody dependent cellular cytoxicity (ADCC) or inhibit anti-phagocytic signals, depending on which surface marker they bind to (Cheson and Leonard 2008). Although it is a relatively new development in the field of cancer therapy, antibody-based therapy is a promising new strategy. In contrast to chemotherapy, which usually attacks cells across tissue types indiscriminately, antibody therapy can be used to target cells expressing certain surface makers. Increased tissue specificity in targeting would result in fewer severe side effects and hopefully allow researchers to develop faster and more effective cancer therapies (Jaiswal et al 2010). However, this lack of general cytotoxicity (the extent to which a chemical or biological agent is lethal to cells) is a double-edged sword. Most existing antibody therapies are not deadly enough to kill off all of the cancer cells on their own; they must be combined with more cytotoxic chemotherapy strategies to be fully effective (Cheson and Leonard 2008). However, it may be possible to develop an alternative to general cytotoxic chemicals by developing therapies that use different combinations of mAbs to increase efficacy and protect against a greater range of tumors. One chemical receptor that isof particular interest in the development of a general antibody-based therapy is CD47 (Chao et al 2010a). Although cancer cells are an extremely heterogeneous group, there are certain chemical receptors that are more likely to be expressed in cancerous cells than in normal cells of the same tissue type. One of the most commonly expressed surface receptors in cancer cells is an immunoglobulin-like protein called CD47, which binds to signal regulatory protein- Îą (SIRP-Îą), a protein which is expressed on the surface of macrophages (Jaiswal et al 2009). Once these two proteins are bound, the SIRP-Îą is phosphorylated, resulting in a signaling cascade that stops the macrophage from eating the cell expressing CD47 (Jaiswal et al 2009). Any cell that expresses CD47 on its surface is much less likely to be phagocytized (killed by a macrophage)

=0A62:/%?0%>.<2 than a cell that does not. However, researchers have demonstrated that the addition of a monoclonal antibody which binds to CD47 and blocks the receptor increases the rate of phagocytic signals. In one study, Jaiswal and colleagues added green fluorescent protein (GFP) to a line of identical cancer cells which had been altered to express varying levels of CD47 and found that there was a correlation between the amount of CD47 expression and the probability of the cell being phagocytized (Jaiswal et al 2009). The researchers then added a CD47 antibody and found that with the antibody, all other things being equal, cells had an equal probability of being phagocytized, regardless of CD47 expression level (Jaiswal et al 2009). This result has been corroborated by an independent study by Zhao and colleagues. They investigated whether or not the anti-phagocytic signal would persist in mice where the long cytoplasmic tail of SIRP-Îą had been partially knocked out. They found out that the presence of functional SIRP-Îą binding sites did not affect the rate of tumor formation, but that macrophages SIRP-Îą deficient mice were more likely to attack tumor cells than macrophages from fully immune-competent mice (Zhao et al 2011). The team also confirmed that the addition of CD 47 antibodies slowed the growth of breast cancer cells, particularly when used in conjunction with an antibody called Trastuzumab, which binds to breast cancer cells and produces a prophagocytic signal. They found that they were able to induce a phagocytic response in breast cancer cells (Zhao et al 2011). Studies have overwhelmingly indicated that the CD47- SIRP-Îą interaction is potentially a very useful target for developing anti-cancer therapies in general and for myeloid cancers (cancers that occur in immune cells, such as leukemia) in particular (Jaiswal et al 2009). CD47 is heavily expressed in hematopoietic stem cells (stem cells that give rise to all types of blood cells, including immune cells) and their descendants (Jaiswal et al 2009). CD47 is important for maintaining a homeostatic Bard Science Journal April 2013 Volume 2, No.4

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immune system, because without it, macrophages tend to attack other blood cells, and if this phagocytosis continues unchecked, it can wipe out much of the immune system (Chao et al 2011). The CD47- SIRP-α interaction is one of the immune system’s key defenses against itself. It is also important for its role in preventing macrophages from consuming essential tissues elsewhere in the body(Jaiswal et al 2010). Because of its prevalence in blood cells, CD47 appears to be a highly effective method for treating cancers in immune cells. These cancers are often particularly problematic, because they are so dispersed throughout the body and because they are cancers of one of the key mechanisms for suppressing tumors (Edris et al 2011). However, several studies have indicated that anti-CD47 treatments enable the immune system to eliminate cancerous blood cells (Chao et al 2011a; Chao et al 2010b; Edris et al 2011). For example, one study by Chao and colleagues found that the addition of monoclonal CD47 antibodies eliminated virtually all traces of engrafted Acute Lymphoblastic Leukemia (ALL) in humanized mice (mice where the murine hematopoietic stem cells have been removed and replaced with human hematopoietic stem cells. The human HSCs divide and develop into a functional human immune system housed in a mouse’s body. This technique allows researchers to test how human immune systems will interact with cancer therapies). In this study, they used flow cytometry to investigate whether or not CD47 was overexpressed in ALL cells compared to normal BM cells, a stem cell type that is a precursor to myeloid cells (Chao et al 2011a). They found that with the addition of the antibody, macrophages were able to phagocytize all of the ALL cells in vitro and slowed down tumor growth significantly in vivo (Chao et al 2011a). Later that year another study by Chao and colleagues found that CD47 antibody therapy was also capable of stopping the spread of non-Hodgkin lymphoma (Chao et al 2011b). In this study, non-Hodgkin’s lymphoma (NHL) cells were injected into mice. They found that NHL cells in which CD47 had been knocked out were unable to disseminate themselves and had a much lower tumor engraftment rate. Most strikingly, in trials where CD47 had been knocked out, there were hardly any secondary lesions whatsoever, suggesting that CD47 is a crucial defense mechanism which permits cancer cells to migrate through the bloodstream unharmed, meaning knocking out CD47 is an incredibly effective means of preventing metastasis (Chao et al 2011b). This result is consistent with the previous studies by Chao, Jaiswal, and colleagues. Shortly thereafter, an independent team conducted a study investigating whether or not CD47 could be used to treat leiomyosarcoma (LMS), a cancer of smooth muscle cells. LMScan arise in any region of the body where soft tissue is present and is particularly problematic because it is very adept at recruiting tumor associated macrophages (TAMs) (Edris et al 2012). TAMs are macrophages that have essentially “changed alignment”. Instead of attacking the tumor cells, TAMs help the tumor acquire adaptive immunity mechanisms that help it evade other macrophages and help mediate tumor angiogenesis. In short, TAMs do everything within their power to produce a favorable microenvironment for the growing tumor and can have a huge impact on the progress of the disease (Edris et al

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2012). In the Edris study, researchers first confirmed that CD47 expression was higher in LMS cells than in cells from benign tumors and normal smooth muscle. This increased CD47 expression level corresponded to increased levels of TAM markers CD68 and CD163. Then they added LMS cells to mice and found that the addition of anti-CD47 mAbs resulted in much higher levels of phagocytosis than in mice that were treated with a control antibody. They concluded that since, in untreated LMS, the majority of macrophages found near tumors were TAMs, the antibody must be turning off the “don’t eat me” signal from the CD47 and allowing the would-be TAMs to recognize the cancer cells as cells that needed to be destroyed (Edris et al 2012). This result is not isolated or unprecedented: Upregulation of CD47 is vital for the migration of hematopoetic stem cells (HSCs) to inflamed areas. Previous studies have shown that mice with CD47 knocked out are unable to recruit HSCs to swollen regions, resulting in the absence of tumor-associated macrophages (TAMs) (Jaiswal et al 2010). Additionally, in Willingham and colleagues’ 2012 study they found that not only did the presence of CD47 antibody prevent tumors from recruiting TAMs, it also allowed would-be TAMs to reverse the “alignment change” and act as tumor-suppressing phagocytes. (Willingham et al 2012). The ability to prevent macrophages from becoming TAMs is unique to antibody therapies, and since TAMs are often such a key factor in the cancer prognosis, this is a significant advantage that should be investigated further (Willingham et a 2012; Edris et al 2012). The main disadvantage to using CD47 as a therapeutic target is the fact that many healthy cells in the body also express it. Simply turning off the CD47 “don’t eat me” signal is not specific enough. For instance, in the study where Willingham and colleagues used CD47 antibodies to test whether or not these antibodies could prevent metastasis of breast cancer in mice, a few of the mice given the antibody treatment were not “viable” after three weeks, meaning that although the tumors had been eliminated, the test subjects were dead or dying due to side-effects of CD47- SIRP-α interference(Willingham et al 2012). The underlying problem is that CD47 antibody therapy by itself it is often not sufficiently specific. In the absence of prophagocytic signals, macrophages will attack cells that express CD47 at random, meaning that there will be no particular preference given to cancer cells, unless they express a prophagocytic signal that attracts cytocidal macrophages to the cancer cell (Chao et al 2010a). However, several studies have suggested that antibody therapies work well when used in combination with each other (Chao et al 2010a; Zhao et al 2011). In yet another study by Chao and colleagues, the researchers decided to test whether or not an anti-CD47 antibody could be used to boost the efficacy of Rituximab, an antibody therapy used to treat non-Hodgkin lymphoma. Rituximab attaches to a surface marker called CD20, which is frequently expressed on the surface of NHL B-cell lymphomas. The other end of the Rituximab binds to an immune cell receptor called FcR, which occurs on the surfaces of macrophages and natural killer cells and produces a strong prophagocytic (“Come eat me!”) signal. However, some forms of NHL are somewhat resistant to Rituximab therapy, and even in cases where it is effective, the antibody often does not fully eradicate the cancer (Chao et


al 2010a). To test this hypothesis, they injected NHL cells into humanized mice and measured tumor engraftment. They found that a combination of Rituximab and anti-CD47 was by far the most effective treatment. Most mice that were treated with just one of the two antibodies died within a month, but 60% of those that were treated with the combination therapy survived for over 182 days (Chao et al 2010a). It appears that the CD47- SIRP-α interaction was to some extent cancelling out the effects of Rituximab, but with the CD47 knocked out via antibody, the prophagocytic signal was able to come through loud and clear (Chao et al 2010a). In a corroborating study, Zhao and colleagues found that CD47 was most effective at eliminating when used in conjunction with Trastuzumab, which produces a prophagocytic signal (Zhao et al 2011). However, further investigations into which types of tumors tend to express which surface markers and into which antibody treatments are most effective are still needed. Unless clinicians know which prophagocytic antibodies should be used in conjunction with CD47, its effectiveness will be limited (Chao et al 2010b). Fortunately, many cancer cells express prophagocytic signals naturally. For instance, one common cancer cell marker that Chao and colleagues have identified is calreticulin. It is found in abnormally high abundances in a diverse range of cancers including non-Hodgkin’s lymphoma, neuroblastoma, and bladder cancer.Although calreticulin releases a strong prophagocytic signal, most cancer cells that express calreticulin also express CD47, which effectively cancels out the calreticulin signal. However, when CD47 antibodies are added, knocking out the “don’t eat me” signal, macrophages attack the calreticulin-expressing cells. Chao and colleagues have postulated that markers like calreticulin are the reason macrophages tend to attack tumor cells more frequently than they attack normal, healthy cells (Chao et al 2010b). The CD47- SIRP-α interaction will continue to be a key topic in future research into antibody-mediated therapy, partly because of its ubiquity in cancer cells and partly because it has a synergistic effect when used in conjunction with other antibodies. Although the CD47 antibodies do appear to cause some severe side effects, it is essential to block the “don’t eat me” signal so that the prophagocytic signals induced other tumor-specific antibodies can be more effective (Chao et al 2010a). Traditional anti-cancer chemotherapies also have severe side effects and are typically far less tissue specific than even widespread cell surface markers like CD47. Additionally, the production of monoclonal antibodies also offers the possibility of customization. Therapeutic antibodies are typically produced through a technique called hybridoma, which involves injecting the target cancer cells into an immune competent mouse. The mouse’s immune system recognizes the cancer cells as “not self ’ and produces an antigen for that specific cancer, which scientists can replicate through hybridoma technology. This technique could be carried out with cancers from individual patients. (Willingham et al 2012). This type of approach could be particularly useful in cancers where there are novel variants in the cancer phenotype and in cases where cancer is spread throughout the body. Because antibodies travel through the bloodstream and cause minimal collateral damage, they may be a more effective and more survivable means of treating cancer than traditional ther-

apies. There is a lot of work that remains to be done in terms of determining which antibodies and combinations of antibodies will eliminate which types of cancer and what the possible side effects and complications may be, but it is a field that clearly merits further investigation. B#C#1#46#+

Chao MP, Alizadeh AA, Tang C, Myklebust J, Varghese B, Gill S, Jan M, Cha AC, Chan CK, Tan BT, Park CY, Zhao F, Kohrt HE, Malumbres R, Briones J, Gascoyne RD, Lossos IS, Levy R, Weissman IL, and Majeti R. (2010). Anti-CD47 Antibody synergizes with Rituximab to promote phagocytosis and eradicate Non-Hodgkin Lymphoma. Cell, 142 (5): 699-713. Chao MP, Jaiswal S, Weissman-Tsukamoto R, Alizadeh AA, Gentles AJ, Volkmer J, Weiskopf K, Willingham SB, Raveh T, Park CY, Majeti R, and Weissman IL. (2010). Calreticulin is the dominant pro-phagocytic signal on multiple human cancers and is counterbalanced by CD47. Science: Translational Medicine, Vol. 2, Issue 63, p. 63ra94. Chao MP, Alizadeh AA, Tang C, Jan M, Weissman-Tsukamoto R, Zhao F, Park CY, Weissman IL, and Majeti M. (2011) Therapeutic Antibody Targeting of CD47 Eliminates Human Acute Lymphoblastic Leukemia. Cancer Res. 71:1374-1380. Chao MP, Tang C, Pachynski RK, Chin R, Majeti R, and Weissman IL. (2011). Extradnodal dissemination of non-Hodgkin lymphoma requires CD47 and is inhibited by anti-CD47 antibody therapy. Blood, 118: 4890-4901. Cheson BD and Leonard JP. (2008). Monoclonal antibody therapy for B-cell non-Hodgkin’s lymphoma. N. Engl. J. Med., 359: 613–626. Edris B, Weiskopf K, Volkmer AK, Volkmer JP, Willingham SB, Conteras-Trujilo H, Liu J, Majeti R, West RB, Fletcher JA, Beck AH, Weissman IL, and van de Rijn. (2012). Antibody therapy targeting the CD47 protein is effective in a model of aggressive metastatic leiomyosarcoma. PNAS 110: 3501-3506. Jaiswal S, Jamieson CHM, Pang WW, Park CY, Chao MP, Majeti R, Traver D, van Roojen N, and Weissman IL. (2009) CD47 is Upregulated on Circulating Hematopoietic Stem Cells and Leukemia Cells to Avoid Phagocytosis. Cell, 138: 271-285. Jaiswal, S, Chao MP, Majeti R, and Weissman IL. (2010). Macrophages as mediators of tumor surveillance. Trends in Immunology, 31: 212219. Willingham SB, Volkmer JP, Gentles AJ, Sahoo D, Dalerba P, Mitra SS, Wang J, Contreras-Trujillo H, Martin R, Cohen JD, Lovelace P, Scheeren FA, Chao MP, Weiskopf K, Tang C, Volkmer AK, Naik TJ, Storm TA, Mosley AR, Edris B, Schmid SM, Sun CK, Chua MS, Murillo O, Rajendran P, Cha AC, Chin RK, Kim D, Adorno M, Raveh T, Tseng D, Jaiswal S, Enger PØ, Steinberg GK, Li G, So SK, Majeti R, Harsh GR, van de Rijn M, Teng NN, Sunwoo JB, Alizadeh AA, Clarke MF, Weissman IL. The CD47-signal regulatory protein alpha (SIRPa) interaction is a therapeutic target for human solid tumors. ,109(17):6662-7 Zhao XW, van Beek EM, Schornagel K, Van der Maaden H, Van Houdt M, Otten MA, Finetti P, Van Egmond M, Matozaki T, Kraal G, Birnbaum D, van Elsas A, Kuijpers TW, Vertucci F, and van den Berg TK. CD47-signal regulatory protein-alpha interactions form barrier for antibody-mediated tumor cell destruction. PNAS, 108 (45) 18342-18347.

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De-worming White-Footed Mice as a Strategy for Reducing Microparasite Transmission to Ticks D'$,)#."&48#$>-44E

<0H8+/13$I#;868&$J##+84:

Abstract. This study applies the concept that a host organism’s immune response to simultaneous infections involves a system of tradeoffs. Past research has shown that chronic intestinal worm parasitism has a negative impact on the host’s immune response to subsequent infections. The white-footed mouse (Peromyscus leucopus) is an excellent reservoir for the agents of Anaplasmosis, Lyme disease, and Babesiosis, passing these microparasites to ticks at high rates. Thus, pathogen dynamics within this host are of major interest in the effort to prevent human infection. Here, I investigate the potential impact of intestinal parasites on the transmission of microparasites to ticks. I hypothesized that treating wild P. leucopus for intestinal worms would reduce the rate at which they transmit blood-borne pathogens to ticks. I treated mice with a commercially available de-worming drug and measured their ability to transmit pathogens to ticks. I found that infected hosts treated with de-wormer pass Anaplasma phagocytophilium at a significantly lower rate than their untreated counterparts. However, de-wormer had no significant impact on a host’s reservoir competence for Borrelia burgdorferi or Babesia microti. (4)1/0-6)8/4 Past research in disease ecology has focused on the impact of a single parasite on a single host species. This simplification neglects a level of complexity that exists in nature. Realistically, a host organism may be faced with many pathogens simultaneously. Focusing on a subset of the potential interactions among affected species provides a limited understanding of infectious diseases. Recently, research has explored instances of multiple simultaneous infections and their consequences (Jolles et al. 2008, Ostfeld 2008). For instance, in populations that experience high rates of HIV, tuberculosis (TB) and malaria, there is also a high likelihood of simultaneous helminth infection (Borkow et al. 2000). Chronic helminth infection alters and impairs immune function (Bentwich et al. 2010, Elias et al. 2007). Studies have suggested that co-infection with helminth parasites may impede treatment efforts and vaccinations against difficult to treat diseases such as HIV and tuberculosis (TB) (Borkow et al. 2000). Patients suffering from these diseases may benefit from anti-helminth treatment in the form of “de-wormer” medication (Bentwich et al. 2008, Bentwich et al. 2010). There is also evidence that intestinal worms weaken immune response to additional pathogens and treatment (Elias et al. 2007, Bentwich et al. 2010). Thus, the removal of intestinal worms using a de-worming drug should aid in the fight against infections that are more challenging to treat. Most research into the effects of co-infection has been conducted in humans. However, co-infection affects a range of non-human animals. For example, Jolles et al. (2008) found that African buffalo are likely to be simultaneously infected with macroparasites such as intestinal helminths and microparasites that cause bovine tuberculosis. This study was one of the first to explore the importance of immunological trade-offs for pathogen dynamics in the wild. Some wildlife diseases affect wild species exclusively. Other wildlife diseases may affect a range of hosts including humans and these are called zoonoses. Zoonotic disease research calls for consideration of multiple factors such

as vectors, host interaction, immune reaction, and the interaction effects of simultaneous infections. Three zoonotic diseases of increasing concern in the United States are Anaplasmosis, Lyme disease, and Babesiosis. According to the Center for Disease Control (CDC), Lyme disease is the most commonly reported vector-borne disease in the United States. In humans, early symptoms for Lyme disease, Anaplasmosis, and Babesiosis include flu-like symptoms such as fatigue, fever, and joint pain (CDC 2006, Steere et al. 2004, Vennier and Krause 2012, Wormser et al. 2006). However, most symptoms of these diseases are non-specific. Between 20 and 50 percent of Lyme disease patients do not experience the rash (EM) that distinguishes Lyme disease from its common flu-like symptoms (Wormser et al. 2006). In some Babesiosis patients, symptoms may be absent (Vennier and Krause 2012, Wormser et al. 2006). Consequently, these diseases are often difficult to diagnose. Untreated Anaplasmosis and Babesiosis can result in severe symptoms and death (CDC 2006, Demma et al. 2005, Vannier and Krause 2012). These tick-borne pathogens can be difficult to treat, so control efforts are focused on prevention (Auwaerter et al. 2004). Co-infection has not been widely studied in the case of these emerging zoonoses. However, parallel situations in HIV and TB may offer insights. Patients infected with HIV and TB are often co-infected with intestinal worms (Bentwich et al. 2008). Helminth parasites are a significant burden on a host’s immune system and further hinder treatment efforts for diseases such as TB, malaria and HIV (Borkow et al. 2000, Bentwich et al. 2008, Bentwich et al. 2010, Elias at al. 2007). This phenomenon can be explained with a deeper understanding of the tradeoffs involved in the immune system. An important group of cells that play an integral role in regulating immune function are T helper cells. T-helper type 1 (Th1) cells activate cytotoxic cells that are specialized for killing infected host cells. T-helper type 2 (Th2) cells activate mostly B-cells, thus promoting the production of antibodies. The immune system operates with limited resources. As a result, Th1 and Th2 cells are cross-regulated. This means

E=$,#48/1$F1/9#6)G$D8/;/:'G$D&10$5/;;#:#$ $ <0H8+/13$I#;868&$J##+84: KK!"8+$+#48/1$.1/9#6)$"&+$A##4$&A180:#0L$I/1$C-;;$M#)"/0+$&40$0&)&G$6/4)&6)$)"#$&-)"/1LKK 5/4)&6)3$+)#."&48#40-44N:M&8;L6/MG$2##+84:NA&10L#0-

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that a heightened response by one T helper subgroup actually the individual level. Specifically, we asked whether the presence suppresses the activity of the other (Kinat et al. 2007, Long et al. of a helminth infection impacts the rate at which white-footed 2004, Pedersen and Fenton 2006). mice pass Anaplasma phagocytophilium, Borrelia burgdorferi and Anaplasmosis, Lyme disease, and Babesiosis are highly Babesia microti to previously uninfected ticks. concentrated in the Northeast and upper Midwestern United The potential for de-worming treatment of white-footStates (Demma et al. 2005, Wormser et al. 2006). Although ed mice to reduce the rate at which microparasites are passed to helminth parasites are not endemic to human populations in this ticks depends on the immune response of the mice to the simularea, these pathogens are passed to humans taneous infections. As shown in past from animals that experience a range of research of human co-infection, the simultaneous infections in the wild. In the burden of a chronic helminth infecNortheast U.S., the agents of Anaplasmosis, tion alters immune response, creating Lyme disease, and Babesiosis are spread a heightened Th2 background (Elias through the bite of a black-legged tick et al. 2007, Kinat et al. 2007). Since (Ixodes scapularis). The reservoir host is Th1 and Th2 cells are cross-regulated, an environment in which the microparait can be postulated that a heightened site may survive and multiply, later to be Th2 background diminishes the actransmitted to additional hosts by the bite tivity of Th1 cells (Kinat et al. 2007). of a vector. For Borrelia burgdorferi, these I aimed to illuminate the possible reservoir hosts include humans, whiteinteraction effects of helminth and tailed deer (Odocoileus virginiaus), veeries microparasite infection on Peromy(Catharus fuscescens), and white-footed scus leucopus. To do so, I treated a mice (Peromyscus leucopus) (Keesing et al. group of wild white-footed mice with 2009, LoGiudice et al. 2002). Hosts vary de-wormer and measured individual in the rate at which they transmit microhosts’ reservoir competence for three parasites to tick vectors. The proportion of tick-borne pathogens. ticks receiving infection from a reservoir I de-wormed mice on host describes the host’s “reservoir compeexperimental forest grids (Hudson et tence”. P. leucopus are the primary reservoir al. 1998, Pedersen and Greives 2007). hosts for B. burgdorferi, infecting 40 to 90 Following the de-worming period, percent of uninfected larval ticks that feed mice were captured and brought to a on them (LoGiudice et al. 2002, Ostfeld and I8:-1#$EL$!"#$%&'()'*+")($,)' holding facility where larval ticks fed Keesing 2001). P. leucopus are also highly competent reservoir to repletion and dropped off the hosts. Ticks were collected and hosts for the agents of Anaplasmosis and Babesiosis. used to measure reservoir competence for the agents of AnaHuman populations likely to contract Anaplasmosis, plasmosis, Lyme disease, and Babesiosis (Anaplasma phagocyLyme disease, and Babesiosis are not likely to be co-infected with tophilum, Borrelia burgdorferi and Babesia microti, respectively) intestinal worms. However, it may be possible to apply de-worm- (Courtney et al. 2004, Hersh et al. 2012, Keesing et al. 2012). ing strategies to the control of these tick-borne diseases. This study focuses on a potential prevention strategy that integrates O#)"/0+ knowledge of the ecology of blood-borne pathogens and the Field Work zoonotic reservoir hosts. I tested whether it is possible to reduce I. Trapping the rate at which microparasites are passed to uninfected ticks, Field studies were conducted at the Cary Institute of thereby reducing instances of human infection. In this study, we Ecosystem Studies (IES) in Dutchess County, New York (lat. target the primary reservoir host of Anaplamsa phagocytophil418509N, long. 738459W). Here, small mammals occupying ium, Borrelia burgdorferi and Babesia microti the white-footed three pairs of trapping grids in forest habitat are monitored using mouse (Peromyscus leucopus). capture-mark-recapture techniques. Within each pair, one grid P. leucopus are often infected with more than a sinis designated experimental and the other control. These trapping gle parasite at once. For instance, P. leucopus are susceptible grids consist of Sherman mouse traps arranged in pairs on an to intestinal worms. In general, helminth parasites (including 11x11 grid. nematodes) have been known to negatively impact host fitness For this study, P. leucopus were baited and trapped ac(Borkow et al. 2000, Elias et al. 2007, Jolles et al. 2008, Hudson cording to standard procedure on two experimental forest plots: et al. 1998, Pedersen and Greives 2007). Helminth parasites Henry experimental and Tea experimental (Ostfeld et al. 2006). impact mouse populations as a whole. The helminth PterygoAll procedures were conducted with approval from the Cary dermatites peromysci plays a role in the population dynamics of Institute of Ecosystem Studies Institutional Animal Care and Use the white-footed mouse (Vandegrift and Hudson 2008). When a Committee. We began trapping for this study in mid-June 2012 large portion of a host population is treated for intestinal worms and continued through July, trapping every three weeks. During using a commercial de-wormer, P. leucopus tend not to experithese sessions, traps were baited with oats, set at 1600 hours and ence characteristic population crashes (Hudson et al. 1998). We checked between 0800 hours and 1300 hours the next day. aimed to explore the potential impact of helminth parasites on Bard Science Journal April 2013 Volume 2, No.4

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II. Field Data Collection Individuals that have been trapped are labeled with ear tags displaying unique identification numbers. Since 1995, data have been collected on individuals during two-day trapping periods. These consist of the individual’s identification number, weight, age, reproductive status, and ecto-parasite burden including a count of Ixodes scapularis on the head and neck. In order to assess helminth burden, fecal samples were collected from mice as available. To minimize contamination, the crew collected samples directly from the mouse and into Eppendorf tubes. Fecal samples were stored in a refrigerator for no longer than one week before processing. III. De-worming I administered the commercial de-wormer Levamisole Hydrochloride (Prohibit™) to half of the mice captured during two consecutive trapping sessions at each grid. De-wormer solution was prepared and stored according to procedures used in a similar project in 2011 (Vandegrift, pers. comm.). We are approved to administer a dose of 20 mg per kg of body weight. Treatment groups were randomly selected for the initial trapping session. During the second trapping session, mice that had not previously been trapped were assigned to the treatment group in order to increase the sample of treated and naturally helminth-infected mice. Mice assigned to the control group received water in doses equal to those given to the treatment group. IV. Rearing Facility Upon the third and final trapping session, I transported animals to the Rearing Facility at the Cary Institute. The third trapping session was held 15-17 days following the second de-wormer administration. During this final session, Sherman traps were baited and checked as described previously. However, this session lasted one day. I used a ranked list of previously captured mice to assemble groups of approximately thirty mice to be transported to the Rearing Facility. Priority was given to mice

that had been identified as “wormy” prior to treatment. This ensured that all groups (treatment/control, wormy/not wormy) were represented in the reservoir competence analysis. Mice were kept in the rearing facility for at least 60 h, in which time most larval ticks could feed to repletion. Ticks that dropped off were collected in glass tick vials. I then transported mice to the field site and released animals to the trapping station from which they were originally found. Ticks were stored in the Ostfeld lab at the Cary Institute for 6 to 8 weeks until they molted into nymphs. Laboratory Procedures I. Tick Preparation Once Ixodes scapularis ticks had successfully molted, I transferred them to cryogenic freezing vials. I froze ticks using liquid nitrogen according to standard protocol (Hersh et al. 2012). These were stored at -80ºC then transported to the Keesing Biology laboratory at Bard College (Annandale-on-Hudson, 12504) for qPCR analysis. Frozen I. scapularis tissues were tested for three tick-borne pathogens: Anaplasma phagocytophilium, Borrelia burgdorferi, and Babesia microti. II. Quantitative Real-Time Polymerase Chain Reaction To obtain DNA from ticks, we extracted total genomic DNA using the DNEasy Blood and Tissue Kit (QIAGEN, Hilden, Germany). To amplify DNA after extraction, we used the multiplex protocol described by Courtney et al. (2004) for Anaplasma phagocytophilium and Borrelia burgdorferi. To amplify extracted DNA for Babesia microti, we used a protocol developed by Hersh et al. (2012). We accounted for potential contamination during the extraction process using extracted DNA from unfed larval ticks as a negative control. Real-time PCR was performed using a CFX96 Real-Time PCR System (Bio-Rad, Hercules, CA, USA). We used ultrapure water as a negative control to account for potential contamination during the PCR process. For all pathogens, we used cloned fragments as positive controls (Keesing et al. 2012, Hersh et al. 2012). We conducted 3 replicate PCRs per tick. For

!&A;#$EL$>#+618.)8H#$+)&)8+)86+$C/1$1#+#1H/81$6/M.#)#46#$84$)1#&)M#4)$&40$6/4)1/;$:1/-.+$/C$&;;$.&)"/:#4+L Sample number, mean, standard deviation, and standard error are shown for treatment and control groups of all hosts. These same statistics are shown for exclusively infected individuals (excluding those with reservoir competence = 0).

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I8:-1#$PL$O#&4$1#+#1H/81$6/M.#= )#46#$Q.1/./1)8/4$/C$)862+$84C#6)#0R$ 6/M.&1#0$A#)S##4$)1#&)M#4)$&40$ 6/4)1/;$:1/-.+L$All hosts were included in this analysis of mean reservoir competence. Data for all three pathogens did not fit the normal distribution. Therefore, Wilxocon Signed-Rank tests were used to compare treatment and control groups. De-wormer had no significant impact on reservoir competence for the three pathogens: A. phagocytophilium (S = 336, Z = -0.37, P = 0.71), B. burgdorferi (S = 309, Z = -1.15, P = 0.25), and B. microti (S = 312, Z = -1.04, P = 0.29).

all pathogens, ticks were considered positive if at least 1 of 3 replicates amplified DNA for that pathogen compared to negative controls. Ticks with marginal results underwent confirmatory testing as described in Keesing et al. (2012) and Hersh et al. (2012). Ticks were considered negative if all 3 replicates showed marginal or negative results. Reservoir competence for each pathogen was calculated as the percentage of ticks infected per host. III. Fecal Egg Counts In order to determine the helminth infection status of individual mice, I used the fecal float method on a present/ absent basis (Sloss and Kemp, 1978). For this process, I prepared a float solution of de-ionized water saturated with solid magnesium sulfate. This solution allows helminth eggs to float while debris sinks. I combined fecal samples with this solution at a ratio of 10 uL per 0.1 g of fecal matter. Next, I homogenized, filtered, and injected the mixture into McMaster egg count chamber slides using a sterile micropipette and tip. Samples were viewed under the compound microscope at an overall magnification of 125x for detection and 250x for verification. Slides were searched exhaustively for parasite ovum. I took note of visible eggs, specifically identifying nematode species including Pterygodermatites peromysci, Rictularia, and Capillaria. Data Analysis/Statistics A similar de-worming project took place throughout June and November 2011. At the Cary Institute, Peromycus leucopus captured on 8 x 8 sub-plots on experimental grids were de-wormed. Individuals captured outside of this square received water as a control treatment. I used data from this study in order to assess the impact of de-worming drugs on the natural tick burden of wild white-footed mice. For the 2012 study, my general approach was to assess whether there was a significant difference in the mean reservoir competence depending on de-worming treatment For this purpose, I used a t-test when data were normally distrib-

uted. For data that did not fit the normal distribution, I used a non-parametric Wilcoxon Signed-Rank test (JMP 9.0; 2012). I tested whether there was a significant difference in the number of uninfected mice (reservoir competence = 0) using a Fisher’s Exact test. Equivalent statistical analyses were run for all three pathogens (Anaplasma phagocytophilium, Borrelia burgdorferi and Babesia microti). Tick burden data are used to confirm that the use of ProhibitTM as a de-wormer does not have a significant impact on ecto-parasite burden, thus verifying that any observed effect of de-wormer on reservoir competence is due to an internal mechanism. I also used a Wilcoxon Signed-Rank test in order to assess any difference in tick burden between treated and control mice. B#+-;)+

Using data from a similar de-worming project in 2011, I assessed the effect of de-wormer on the natural tick burden of wild Peromyscus leucopus. In 2011, the average mouse had 7.14 ticks on its head and neck. Mice that received de-wormer (n = 160) treatment had an average tick burden of 7.04 ± 3.20. Mice that did not receive treatment (n = 183) had an average tick burden of 7.25 ± 2.60. I found that de-wormer has no significant effect on tick burden of mice on any of the experimental grids at the Cary Institute (S = 26599, z = -1.012, P = 0.312). We determined reservoir competence (percentage of ticks infected per host) for three tick-borne pathogens: Anaplasma phagocytophilium, Borrelia burgdorferi, and Babesia microti. We compared mean reservoir competence between treatment and control groups. I first determined that there was no significant difference in mean reservoir competence between grids for any of the three pathogens: A. phagcytophilium (S = 416.5, Z = 0.811, P = 0.42), B. burgdorferi (S = 430.5, Z = 1.19, P = 0.24), or B. microti (S = 422, Z = 0.92, P = 0.36) . In subsequent analyses, I treated individual mice as the unit of replication regardless of the mouse’s grid of origin. I used a non-parametric Wilcoxon Signed-Rank Test to assess the potential difference in mean reservoir competence Bard Science Journal April 2013 Volume 2, No.4

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*

I used a non-parametric Wilcoxon Signed-Rank Test to assess the potential difference in mean reservoir competence between individuals receiving de-wormer treatment and those in the control group. When compared between all treated and control hosts, de-wormer was shown to have no significant effect on mean reservoir competence of A. phagocytophilium, B. burgdorferi, or B. microti (Fig. 3). When I analyzed reservoir competence of only infected hosts (reservoir competence > 0), I found that de-wormer significantly reduced reservoir competence of hosts infected with Anaplasma phagocytophilium. However, de-wormer treatment had no significant effect on the reservoir competence of hosts infected with Borrelia burgdorferi or Babesia microti (Fig. 4). I found that for all pathogens, hosts treated with de-wormer are no more likely to be uninfected than those not receiving treatment. Fisherâ&#x20AC;&#x2122;s Exact tests returned non-significant P-values for Anaplasma phagocytophilium (P = 1.0), Borrelia burgdorferi (P = 0.43), and Babesia microti (P = 0.34). >8+6-++8/4 When all hosts were included, de-wormer treatment had no significant impact on tick burden or the mean reservoir competence for Anaplasma phagocytophilium, Borrelia burgdorferi, or Babesia microti. Additionally, de-wormer treatment did not increase the likelihood of a mouse being uninfected (passing microbial infection to exactly zero ticks). When only infected mice were included, treatment had no significant impact on B. burgdorferi or B. microti transmission. However, hosts treated with de-wormer and infected with A. phagocytophilium passed infection to significantly fewer ticks than their untreated counterparts. The fecal float method I used has a high potential frequency of false negatives. Infected hosts are not consistently shedding detectable levels of parasite eggs. A host determined to have no visible nematode eggs in its fecal matter may indeed be infected with intestinal parasites. However, similar studies have confirmed that de-wormer treatment is effective for eliminating intestinal worms in Peromyscus leucopus and none of the de-

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I8:-1#$TL$O#&4$1#+#1H/81$6/M.#)#46#$Q.1/= ./1)8/4$/C$)862+$84C#6)#0R$/C$84C#6)#0$"/+)+$ 6/M.&1#0$A#)S##4$)1#&)M#4)$&40$6/4)1/;$ :1/-.+LL Only infected hosts were included in this analysis of the mean reservoir competence based on treatment status. Borrelia burgdorferi data did not fit the normal distribution, so a Wilcoxon Signed-Rank test was used. De-wormer treatment had no significant impact on reservoir competnce for Borrelia burgdorferi (S = 292, Z = 0.85, P = 0.39) or Babesia microti (t = 0.026, df = 18.13, P < t = 0.51). A one-tailed T-test shows that de-wormer had a significant negative impact on reservoir competence for Anaplasma phagocytophilium (t = -2.07, df = 12.57, P = 0.03). Significant differences in mean reservoir competence are marked by an asterisk (*). wormed mice in this study were found to have eggs post-treatment. Finally, fecal floats were performed as a confirmatory method to the ultimate hypothesis that de-wormer treatment affects the rate at which P. leucopus pass microbes to Ixodes scapularis. Oral administration of de-wormer can lead to inaccurate dosage in the field. It is more likely for an animal to receive an entire dose of de-wormer using an oral gavage or injection method. However, past studies have determined that orally administrating de-wormer using a micropipette is an effective method for the treatment of helminth infection in P. leucopus (Pedersen and Greives 2007, Vandegrift and Hudson 2008). Mice selected to be taken to the Rearing Facility and subsequently to be assessed for reservoir competence were given a priority if they were treated with de-wormer more than once, thus increasing the likelihood that the host received an effective dose. Additionally, I did not find any of the post-de-wormer treatment mice to be shedding helminth eggs. I found no significant effect of treatment on natural Ixodes scapularis burden of Peromyscus leucopus hosts. This indicates that the de-wormer Prohibit TM has no impact on the number of ticks feeding on individual mice. Other de-wormers were not tested for this effect, and it is possible that the use of an alternative anti-helminthic drug could reduce ecto-parasite burden. However, the aim of this study was not to kill ticks or defer them from murine hosts. These strategies are likely to have negative ecological effects. Rather, I investigated the potential to reduce the likelihood of human infection through reducing the transmission of microparasites from zoonotic hosts to vectors. Treated hosts were no more likely than control hosts to have a reservoir competence equal to zero for any of the pathogens. I found no evidence that de-wormer increased the chances that a host would be uninfected. Any significant difference in reservoir competence due to de-wormer treatment is therefore associated with a significant reduction as opposed to a complete elimination of pathogen transmission. In analyses including both infected and uninfected hosts, I found that de-wormer did not impact reservoir competence for any of three pathogens:


Anaplasma phagocytophilium, Borrelia burgdorferi, or Babesia microti. However, we were primarily concerned with the cases in which an uninfected tick feeds on an infected mouse, thus promoting disease transmission. For this reason, I conducted a similar analysis of data after excluding uninfected hosts. A host was deemed uninfected for a certain pathogen if its individual reservoir competence was exactly zero. When I removed uninfected hosts from subsequent analyses, I found that de-wormer had no significant effect on a host’s reservoir competence for B. burgdorferi and B. microti in either direction. Given this evidence, it appears that intestinal parasites neither hinder nor aid the response of a host’s immune system to B. burgdorferi or B. microti. This could indicate that intestinal parasites are not hindering a mouse from carrying out a defense against these microbial pathogens. It is also possible that the immune system requires more time to recover from chronic intestinal worms than this study allowed. The immune system could be permanently damaged by chronic helminth infection. Future research is required in order to draw a more pointed conclusion on the effects of helminth infection on the immune system of P. leucopus. A more experimental study manipulating the helminth and microbial infection would be necessary. However, this study’s focus is the feasibility of implementing a de-wormer treatment being to reduce reservoir competence of P. leucopus in nature. Essentially, this study concludes that elimination of helminth parasites with a de-wormer does not significantly impact the transmission of certain pathogens from P. leucopus to I. scapularis. Furthermore, the presence of helminth parasites is not explanatory of the high percentage of larval ticks that obtain B. burgdorferi or B. microti from P. leucopus. When I included only infected hosts, those infected with A. phagocytophilium and treated with de-wormer had a significantly reduced mean reservoir competence. This could be due to a mechanistic effect of the drug on the bacteria itself. More importantly, this result may indicate that helminth parasites hinder the host’s immune response to A. phagocytophilium. Relief from intestinal worms allows the immune system to carry out a more successful response to and removal of these bacteria. It is curious that de-wormer has a significant impact on reservoir competence for A. phagocytophilium and not another bacterium such as B. burgdorferi. What is it about these microbial parasites that makes de-wormer a potential strategy for disease prevention or not? Past research has indicated that although B. burgdorferi is essentially an extracellular pathogen, it may instigate an immune response similar to that of an intracellular pathogen (Craig-Mylius et al. 2006, Liang et al. 2012, Yssel et al. 1991). There is also evidence that Peromyscus leucopus produce antibodies to B. burgdorferi (Magnarelli et al. 1997) and yet P. leucopus hold their status as the most competent reservoirs for the bacteria. It is apparent that B. burgdorferi is well adapted to evade immune response. It is essential for public health efforts that studies continue to pursue an explanation for the high rates at which P. leucopus pass B. burgdorferi to I. scapularis. Ultimately, for B. burgdorferi and B. microti, administering de-wormer has no significant impact on transmission of these pathogens to tick vectors in the wild. We looked to reduce human infection by reducing transmission to ticks. This study helps us to determine that de-wormer is not a plausible

prevention strategy for B. burgdorferi and B. microti. However, de-wormed hosts infected with A. phagocytophilium transmit the bacteria to a significantly reduced proportion of previously uninfected larval ticks than those that remain untreated. Future efforts may be made to de-worm mice in particularly high-risk areas in order to reduce transmission to vectors and subsequent transmission to uninfected hosts. Although it would be difficult to ensure treatment is widespread and sufficient, it is possible to provide attractive food sources containing de-wormer treatment for mice in the wild. As de-wormer is commercially available and considered safe and effective, this prevention strategy may prove to be more feasible, more widely accessible, and overall more successful than expensive vaccinations. <624/S;#0:#M#4)+ Funding for this project was provided by NSF. Research facilities were provided by Bard College and the Cary Institute. I would like to thank the Ostfeld field crew and the Keesing laboratory staff for their assistance with field work and tick sample analysis, respectively. This project benefitted from the assistance and support of those at the Cary Institute, especially Dr. Richard Ostfeld, Kelly Oggenfuss, Patti Smith, Dr. Alan Berkowitz, and the REU students. I am grateful for the hospitality of The Pennsylvania State University and the direction of Dr. Kurt Vandegrift. I thank Drs. Brooke and Craig Jude, Dr. William Maple and Dr. Michael Tibbetts for helpful discussions. Finally, I owe my deepest gratitude to Dr. Felicia Keesing without whose mentorship this project would not have been possible. U8)#1&)-1#$58)#0 Barbour, A. G., and A. Jasinskas. 2008. A genome-wide proteome array reveals a limited set of Immunogens in natural infections of humans and white-footed mice with Borrelia burgdorferi. Infection and Immunity 76(8):3374-3389. Belperron, A. A., C. M. Dailey, C. J. Booth, and L. K. Bockenstedt. 2007. Marginal zone B-cell depletion impairs murine host defense against Borrelia burgdorferi infection. Infectious Immunology 75(7):3354-3360. Bentwich, Z., C. L. Teicher, and G. Borkow. 2008. The helminth HIV connection: time to act. AIDS 22:1611-1614. Bentwich, Z., R. Horner, and G. Borkow. 2010. De-worming in developing countries as a feasible and affordable means to fight co-endemic infectious diseases. The Open Biology Journal 3:97-103. Borkow, G., and Z. Bentwich. 2000. Eradication of helminthic infections may be essential for successful vaccination against HIV and tuberculosis. Bulletin of the World Health Organization 78(11):1368-1369. Courtney, J. W., L. M. Kostelnik, N. S. Zeidner, and R. F. Massung. 2004. Multiplex real-time PCR for detection of Anaplasma phagocytophilium and Borrelia burgdorferi. Journal of Clinical Microbiology 42(7):31643168. Craig-Mylius, K., G. F. Weber, J. Coburn, and L. Glickstein. 2006. Borrelia burgdorferi, an extracellular pathogen, circumvents osteopontin in inducing an inflammatory cytokine response. Journal of Leukocyte Biology 77:710-718. Center for Disease Control. 2006. Diagnosis and Management of Tickborne Rickettsial Diseases: Rocky Mountain spotted fever, Ehrlichiosis, and Anaplasmosis- United States. MMWP Recommended Reports 55(1) Bard Science Journal April 2013 Volume 2, No.4

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Dahlgren, F. S., E. J. Mandel, J. W. Krebs, R. F. Massung, and J. H. McQuiston. 2011. Increasing incidence of Ehrlichia chaffeensis and Anaplasma phagocytophilium in the United States, 2000-2007. American Journal of Tropical Medicine and Hygiene. 85(1):124-131.

Magnarelli, L. A., J. F. Anderson, K. C. Stafford 3rd. and J. S. Dumler. 1997. Antibodies to multiple tick-borne pathogens of babesiosis, ehrlichiosis, and Lyme borreliosis in white-footed mice. Journal of Wildlife Diseases 33(3):466-473.

Demma, L. J., R. C. Holman, J. H. McQuiston, J. W. Krebs, D. L. Swerdlow. 2005. Epidemiology of human ehrlichiosis and anaplasmosis in the United States, 2001-2002. American Journal of Tropical Medicine and Hygiene 73(2):400-409

Martin L. B., Z. M. Weil, and R. J. Nelson. 2007. Immune defense and reproductive pace of life in Peromyscus mice. Ecology 88(10): 25162528.

Elias, D., S. Britton, A. Kassu, and H. Akuffo. 2007. Chronic helminth infections may negatively influence immunity against tuberculosis and other diseases of public health importance. Expert Reviews of Anti-Infective Therapy 5(3):475-484. Embers, M. E., R., Ramamoorthy, and M. T. Philipp. 2004. Survival strategies of Borrelia burgdorferi, the etiologic agent of Lyme disease. Microbes and Infection 6:312-318. Ghayad Z., and C. Hou. 2012. Erythema migrans in early disseminated lyme disease. Journal of the American Osteopathic Association 112(11):748 Hersh, M. H., M. T. Tibbetts, M. Strauss, R. S. Ostfeld, and F. Keesing. 2012. Quantifying reservoir competence for Babesia microti of wildlife host species using real-time PCR. Emerging Infectious Diseases http:// dx.doi.org/10.3201/eid1812.111392 Herwaldt, B. L., J. V. Linden, E. Bosserman, C. Young, D. Olkowska, and M. Wilson. 2011. Transfusion-associated babesiosis in the United States: a description of cases. Annals of Internal Medicine 155:509-519. Hudson, P. J., A. P. Dobson, and D. Newborn. 1998. Prevention of population cycles by parasite removal. Science 282(5397):2256-2258 Jolles, A. E., V. O. Ezenwa, R. S. Etienne, W. C. Turner, and H. Olff. 2008. Interactions between macroparasites and microparasites drive infection patterns in free-ranging African Buffalo. Ecology 89(8):2239-2250. Keesing, F., M. H. Hersh, M. Tibbetts, D. J. McHenry, S. Duerr, J. Brunner, M. Killilea, K. LoGiudice, K. A. Schmidt, and R. S. Ostfeld. 2012. Reservoir competence of vertebrate hosts for Anaplasma phagocytophilium. Emerging Infectious Diseases http://dx.doi.org/10.3201/ eid1812.120919http://www.cdc.gov/Other/disclaimer.html Kinat, T. J., B. A. Osborne, and R. A. Goldsby. 2007. Kuby immunology, sixth edition. W. H. Freeman and Company, New York, New York, USA. Krupp, L. B., L. G. Hyman, R. Grimson, P. K. Coyle, P. Melville, S. Ahnn, R. Dattwyler, and B. Chandler. 2003. Study and treatment of post Lyme disease (STOP-LD): a randomized double masked clinical trial. Neurology 60(12):1923-1930. Liang, F. T., F. K. Nelson, and E. Fikrig. 2002. Molecular adaption of Borrelia burgdorferi in he murine host. Journal of Experimental Medicine 196(2):275-280. LoGiudice, K., R. S. Ostfeld, K. A. Schmidt, and F. Keesing. 2002. The ecology of infectious disease: effects of host diversity and community composition on Lyme disease risk. Long, K. Z., and N. Nanthakumar. 2004. Energetic and nutritional regulation of the adaptive immune response and trade-offs in ecological immunology. American Journal of Human Biology 16(5):499-507.

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Ostfeld, R. S., and F. Keesing. 2000. The function of biodiversity in the ecology of vector-borne zoonotic diseases. Canadian Journal of Zoology 78:2061-2078. Ostfeld, R. S. 2008. Parasites as weapons of mouse destruction. Journal of Animal Ecology 77(2):201-204. Pedersen, A. B., and A. Fenton. 2007. Emphasizing the ecology in parasite community ecology. Trends in Ecology and Evolution 22(3):133139. Pedersen, A. B., and T. J. Greives. 2007. The interaction of parasites and resources cause crashes in a wild mouse population. Journal of Animal Ecology 77(2):370-377. Pedersen, A. B., and S. A. Babayan. 2011. Wild immunology. Molecular Ecology 20(5):872-880. Sloss, M. W., and R. L. Kemp. 1978. Veterinary Clinical Parasitology. Iowa State University Press, Ames, Iowa, USA. Steere, A. C., J. Coburn, and L. Glickstein. 2004. The emergence of Lyme disease. Journal of Clinical Investigation 113(8):1093. Steere, A. C., R. T. Schoen, and E. Taylor. 1987. The clinical evolution of Lyme arthritis. Annals of Internal Medicine 107:725-731. Vandegrift, K. J., and P. J. Hudson. 2008. Could parasites destabalize mouse populations? The potential role of Pterygodermatites peromysci in the population dynamics of free-living mice, Peromyscus leucopus. International Journal for Parasitology 39:1253-1262. Vannier E., and P. J. Krause. 2012. Human babesiosis. New England Journal of Medicine 336:2397-2407. Wormser, G. P., R. J. Dattwyler, E. D. Shapiro, J. J. Halperin, A. C. Steere, M. S. Klempner, P. J. Krause, J. S. Bakken, F. Strie, G. Stanek, L. Bockenstedt, D. Fish, J. S. Dumler, R. B. Nadelman. 2006. The clinical assessment, treatment, and prevention of lyme disease, human granulocytic anaplasmosis, and babesiosis: clinical practice guidelines by the Infectious Diseases Society of America. Clinical Infectious Diseases 43(9):1089-1134. Yazdanbakhsh, M., P. G. Kremsner, and R. van Ree. 2002. Allergy, parasites, and the hygiene hypothesis. Science 296:490-494. Yssel, H., M. Shanafelt, C. Soderberg, P.V. Schneider, J. Anzola, and G. Peltz. 1991. Borrelia burgdorferi activates a T Helper Type 1-like T Cell subset in Lyme Arthritis. Journal of Experimental Medicine 174:593601.


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April 2013  

BSJ's most colorful issue yet, featuring articles on everything from science outreach to vaginal microbiomes, plus paper reviews, art and fi...

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