Over the past few months, a large team of undergraduate, graduate, and postdoctoral researchers, and I, have been processing hundreds of samples from our scallop hatchery microbiome project. As 2022 winds down, so does the first phase of our lab work, and we are taking a well-deserved break over the holidays before we launch additional lab work, data analysis, and manuscript writing in 2023.
In 2021, the Ishaq Lab, collaborators at UMaine, and collaborators at the Downeast Institute ran a pilot project to investigate the bacteria that associate with sea scallop larvae in hatcheries, and how this is develops in relation to bacteria in hatchery tanks over time. For that project, we collected hundreds of culture plates with a specialized media that selects for certain species of bacteria.
When tanks are drained and cleaned every two days, cotton swabs are rolled across part of the bottom or side of the tank and used to inoculate bacteria onto these culture plates. This is part of a routine screening for pathogens, and don’t worry, we aren’t finding bacteria that causes disease in humans. But, these screening plates creates a useful starting point for our research on bacterial community dynamics.
Tank swab samples are used to inoculate TCBS plates to screen for Vibrio and similar bacteria
We received over 200 of these TCBS culture plates, and from them we isolated 140 bacteria in 2021 and early 2022 which we archived at -80 degrees Celsius. This was part of Sarah Hosler’s master’s of science thesis in August of 2022, and has since been passed to Ayodeji Olaniyi for part of his master’s of science thesis.
This fall, we were able to recover 115 of these isolates from the deep freeze, and tested them on 12 different media in duplicate, which created >1800 cultures plates and tubes, and 230 microscope slides!
This massive undertaking would not have been possible without a large team helping with the lab work, including rockstars Ayodeji Olaniyi, Sydney Shair, Keagan Rice, and Lacy Mayo who put in hours and hours leading the efforts on this. We are also grateful to Alaa Rabee, Aaron Williams, Lily Robbins, Ash VanNorwick, and Rebecca Kreeger who provided assistance with media making, inoculating, and the large amount of cleanup (we used glass or autoclavable plastic where possible, and sterilized some single-use plastics to be used as training tools for student education). We were also assisted by Bryanna Dube, who is working on creating outreach/education materials based on our results.
Now, our team will focus on analyzing the results of all these microbiology tests and look for trends. Some will also be heading to the Perry Lab to learn how to perform quantitative polymerase chain reactions (qPCR), in which we use a modified version of DNA replication to count the copies of specific genes. We will use this to look for genes which confirm the identity of our bacteria.
Beginning in summer 2022, the Ishaq Lab has also been part of a state-wide research and commercial collaboration to understand and improve sea scallop production in hatcheries and farms. As part of that project, we received 1500 DNA samples from different hatchery tanks or larvae over the summer/fall rearing season.
Gloria Adjapong is a Postdoctoral Fellow at the UMaine Cooperative Extension Veterinary Diagnostics Lab, and she has been graciously extracting these samples as part of her cross-training in the Ishaq Lab. We will use the extracted DNA to sequence the bacterial communities to identify which bacteria are present, and when, to understand microbial community dynamics over time and in relation to scallop health.
Last week, undergraduate Alexis Kirkendall concluded her 10-week Research Experience for Undergraduates (REU) program at the University of Maine. Over that time, she has been an integral member of the Ishaq lab, and had assisted with lab work for multiple projects, comparing microscopy staining protocols, training students, assisting with laboratory management and safety regulation checks, and more. She picked up skills in animal sample collection, microscopy and staining, parasitology, culture media preparation, DNA extraction, and data visualization in R. Alexis also helped create some of the marketing materials for the Microbes and Social Equity Symposium in July, and facilitated group discussions as a note taker.
At the end of the program, REU students create posters and short presentations of their efforts over the summer. The presentations were last week, but you can check out the poster below.
Alexis is heading back to Ohio for her next year at Heidelberg College where, in addition to studying science, she is leading initiatives to make the campus more accessible and inclusive. But, we hope to see her back in Maine sometime in the future!
The Research Experience for Undergraduates (REU) 2022 summer program has begun again, and the Ishaq Lab is pleased to welcome Alexis!
Alexis Kirkendall
Undergraduate Researcher, Biology, Heidelberg University
Alexis is from Ohio and is majoring in Biology at Heidelberg University. Her research interests are in genetics and she has a love for the fascinating world of microbes. She joined the lab through the Summer 2022 REU. She is researching Cryptosporidium in cows, helping in the MSE Symposium, and aiding in the Camel Rumen Microbiome Project this summer.
Across the U.S., REU programs are intended to engage undergraduates in research, including those who have no or very little research experience and no opportunity to do so at their home institution. The program I volunteer for is through the UMaine Initiative for One Health and the Environment, which brings about 10 students to campus for 10 weeks to engage in research and professional development. In addition to participating in research, the students attend workshops of coming up with research plans and questions, professional development plans, their resumes, posters, presentations, and other skills.
This year, the UMaine Student Research Symposium was held in person and virtually, and undergrads and grads from the Ishaq Lab shared their research with the Maine community. You can check out the recorded presentations in the links below.
Pelletier*, E., Taylor, T., Ishaq, S. Abstract 830. Assessing the Veterinary Needs of Rural Maine and Implementing an Effective Management Plan. UMaine Student Symposium (poster presentation). April 15, 2022.
Undergraduate Animal and Veterinary Science (AVS) students Natalie Sullivan (4th year), Maria Messersmith (3rd year), and Elaina Cobb (3rd year) were awarded a spring 2022 research award from the J. Franklin Witter Undergraduate Research Endowment Fund! Natalie, Maria, and Elaina will be part of a large student researcher team working on different aspects of this project over the next year, but the three of them composed the funding proposal as they have been working on this project the longest.
Background: Cryptosporidiosis, an infection caused by protozoa from the Cryptosporidium genus, often C. parvum, has emerged as a frequently reported intestinal disease of animals and humans since the 1980s when its zoonotic potential was recognized. C. parvum causes a brief diarrheal disease, which can lead to dehydration, weight loss, and, in extreme cases, death, particularly in children or neonatal animals. On farms, the disease results in intense diarrhea of neonatal calves and is associated with substantial economic losses in dairy farming worldwide as it can have long-term effects on weight gain and production efficiency. Cryptosporidiosis is transmissible via the fecal-oral route, either indirectly or directly, and C. parvum is almost always present in water and soil/bedding. There is no treatment, so disease management and prevention tend to be more focused on animal management practices involving nutrition, biosecurity, and cleanliness. This project centers around the tracking of cryptosporidium parvum in different aged dairy cattle populations at the University of Maine’s J.F. Witter Farm.
Soil samples from the farm were checked for the presence of parasites using microscopy.The calf stalls have bedding instead of soil floors, so we need to wash bedding samples to collect any protozoa that might have been left there.Cryptosporidium is most commonly spread through shared water sources so we are monitoring those.Once protozoa have been washed from the samples, the wash is put on glass slides, where dyes are used to stain the cells so we can see them using a microscope.Cows are cute but make sure you wash your hands after touching them to avoid infections.Photos from the Cryptosporidium project.
Importance/Impact: This project will track the trends and prevalence in C. parvum carriage in the Witter herd, and identify if there are certain conditions in which C. parvum is more prone to grow and multiply, especially in dairy calves. We anticipate that any findings will be used to implement new management practices for the control of Cryptosporidium parvum, specifically in bovine management, and reduce zoonosis at this farm. This project will potentially lead to making management recommendations for other farms in Maine.
Research Question, Objectives, and Hypotheses: What are the difference in disease prevalence and contamination between calves, yearlings, and adults in relation to Cryptosporidium parvum? Is infection rate due to risk of age range or animals or housing conditions?
Sarah Hosler
Master’s student, has been coordinating the project but will be defending thesis this summer and is transitioning project leadership over to Ayo
Ayodeji Olaniyi
New Master’s student Jan 2022, will be taking over project management, overseeing lab work
Natalie S.
AVS Capstone senior, graduating this May, has been collecting samples and doing microscopy since fall 2021
Maria M.
AVS Capstone junior, has been collecting samples and doing microscopy since fall 2021
Elaina C.
AVS Capstone junior, started in Jan 2022, will focus on sample collection at the farm
Jada M.
AVS Capstone junior, started in Jan 2022, will focus on sample processing in the lab
Lani P.
AVS Capstone junior, started in Jan 2022, will focus on sample processing in the lab
Sydney R.
AVS Capstone junior, started in Jan 2022, will focus on literature review
Josh F.
AVS Capstone junior, started in Jan 2022, will focus on literature review
Amatullah A.
Biology Honors, started in fall 2021, will focus on sample collection at the farm
Team Crypto in the Ishaq Lab
This project is a continuation of the pilot research that undergrad Emily Pierce, master’s student Alex Fahey, and master’s student Sarah Hosler worked on over the past two years.
Funded by the University of Maine Rural Health and Wellbeing Grand Challenge Grant Program, this project assesses pathogen carriage by mice and flying squirrels on or near farms in several locations in Maine. We live-capture mice and flying squirrels in traps, collect the poop they’ve left in the trap, and conduct a few other health screening tests in the field before releasing them. To maximize the information we collect while minimizing stress and interference to the animals, information is being collected for other projects in the Levesque Lab at the same time. We will be collecting samples for another few weeks, and then working on the samples we collected in the lab over the fall and winter.
One of the major goals of the funding program, and this project, is to engage students in research. After a few months on the project, some of our students describe their role and their experiences so far…
Marissa Edwards
Undergraduate in Biology
Levesque Lab
Hi! My name is Marissa Edwards and I am an undergraduate research assistant with Danielle Levesque. This summer, my role has been to set traps, handle small mammals, and collect fecal and tissue samples from deer mice.
One of the skills I’ve learned this summer is how to properly ear tag a mouse. To catch mice, we set traps across UMaine’s campus as well as other parts of Maine, including Moosehead Lake, Flagstaff Lake, and Presque Isle.
During our trip to Moosehead Lake, I saw a marten for the first time (it was in one of our traps). I did not know martens existed and initially thought it was a fisher cat. It was both a cool and terrifying experience!
Elise Gudde
Master’s Student of Ecology and Environmental Sciences
Levesque Lab
Hello, my name is Elise Gudde, and I am currently a master’s student at the University of Maine in the Ecology and Environmental Sciences program. I work in Dr. Danielle Levesque’s lab studying small mammal physiology in Maine.
This summer, as a part of the squirrel project, I work to trap small mammal species in Maine, such as white footed mice, deer mice, and flying squirrels in order to determine which species have shifted their range distributions as a result of climate change. Being a part of the research team, this summer has brought me all over Maine! I have been able to travel to Orono, Greenville, New Portland, and Aroostook County to study many interesting mammals. I even got to handle an Eastern chipmunk for the first time! As a member of the animal-handling side of the research team, I also collect fecal and tissue samples from the animals. These samples are then handed off for other members of the team to research in the lab!
Rebecca French
Undergraduate in Animal and Veterinary Sciences
Ishaq Lab
In the beginning of this project, I had no idea what I was getting myself into when I began researching flying squirrels and mice. I came into it with almost no in-person lab experience, so I had a lot to learn.
So far, I have been focusing on making media on petri dishes for culturing bacterial growth and after plating fecal bacteria on said plates; discerning what that growth can be identified as.
We are using media with specific nutrients, and colored dyes, and certain bacteria we are interested in will be able to survive or produce a color change. I have also been performing fecal flotations and viewing possible eggs and parasites under a microscope. What I’ve found most fun about this project is putting into practice what I have learned only in a classroom setting thus far. It is also very satisfying to be a part of every step of the project; from catching mice, to making media, to using that media to yield results and then to be able to have a large cache of information to turn it all into a full fledged project.
Some of the media used to culture bacteria from the feces of mice and squirrels.
Joe Beale
Undergraduate in Animal and Veterinary Sciences
Kamath Lab
Hello! My name is Joseph Beale, and I am an undergraduate at the University of Maine working on the squirrel project as a part of my capstone requirement for graduation. My primary responsibility in this project is the molecular testing of samples obtained from the field. Primarily I will be working with ear punch samples taken from flying squirrels and field mice. DNA extracts from these field samples will be run via qPCR. The results of this qPCR will tell us if these squirrels are carrying any pathogens.
The pathogens we will be testing for are those found in Ixodes ticks. The qPCR panel which we will be running the extracted DNA from the ear punches on tests for Borrelia burgdorferi, the causative agent of Lyme disease, Anaplasma phagocytophilum, the causative agent of anaplasmosis, and Babesia microti, the causative agent of Babesiosis. These pathogens and respective diseases discussed are all transmitted through Ixodes ticks. Deer ticks are the most common and famous of the Ixodes genus. The Ixodes genus encapsulates hard-bodied ticks. Along with deer ticks, Ixodes ticks found in Maine include: woodchuck ticks, squirrel ticks, mouse ticks, seabird ticks, and more. Mice and squirrel are ideal hosts for these Ixodes ticks, therefore becoming prime reservoirs for these diseases. In our research, we are interested in determining the prevalence of these diseases in squirrels and mice as these hosts can spread these diseases to humans and other animals in high tick areas.
qPCR, quantitative polymerase chain reaction, allows for the quantification of amplified DNA in samples. This will help tell us if these pathogens are present in samples and in what capacity. In qPCR provided DNA strands are added to the reaction. These strands match with the genome of the intended pathogens. If the pathogens are present in our samples, the provided DNA strands will bind to the present pathogen DNA. PCR will then work to manufacture billions of copies of this present pathogen DNA.
When not working on this project, I also work in the University of Maine Cooperative Extension Diagnostic Research Laboratory as a part of the Tick Lab. In this position I have honed the molecular biology skills that I will in turn use for the squirrel project.
Hello everyone! My name is Yvonne Booker and I am a rising senior, animal and poultry science major at Tuskegee University in Tuskegee, Alabama. I am interested in animal health research, with a particular focus in veterinary medicine. I’ve always wanted to be a veterinarian, but as I progressed throughout college, I became interested in learning more about animal health and how I might help animals on a much larger and impactful scale–which led me to the REU ANEW program. Currently climate change is causing an increase in global temperatures, putting pressure on animals’ ability to interact and survive within their environment. Consequently, scientists are now attempting to understand not just how to prevent climate change, but how these creatures are adapting to this emerging challenge.
My research experience this summer is geared toward addressing this global issue. I am currently working in Dr. Danielle Levesque’s Lab, which aims to study the evolutionary and ecological physiology of mammals in relation to climate. My project involves conducting a literature review of the microbiome of mammals, to learn more about how their microbial community plays a role in how they adapt in a heat-stressed environment.
Our knowledge of vertebrate-microbe interactions derives partly from research on ectotherms. While this research paves the path for a better understanding of how organisms react to temperature changes, fewer studies have focused on how mammals deal with these extreme temperature shifts—specifically, the abrupt surge in climate change. The ability of endotherms to thermoregulate alters our knowledge of (1) how mammals create heat tolerance against these environmental challenges and (2) how this internal process alters mammals’ adaptability and physiology over time. We suggest that the microbiome plays an essential part in understanding mammals’ heat tolerance and that this microbial community can help researchers further understand the various processes that allow mammals to survive extreme temperatures.
As a student of the REU ANEW program my goal was to go out of my comfort zone and study animals in an applied fashion that would impact animal health on an environmental and ecological scale; and this program was just that! My mentor, Dr. Levesque was wonderful in guiding me through conducting this research, while giving me the independence to create my own voice. The program directors, Dr. Anne Lichtenwalner and Dr. Kristina Cammen, have also been extremely supportive throughout this entire program equipping students with the tools they need to succeed as researchers. Although research was my primary focus this summer, some of my favorite memories involved building community with the students and the staff. From weekly check-ins on zoom to virtual game nights of complete smiles and laughter, this program has been one for the books! The One Health and the Environment approach to this Research Experience for Undergraduate students has encouraged me to build on my curiosity within the field of science, and I’m looking forward to applying what I’ve learned to my career in the future.
Did you know that camels have three stomach chambers or that they have to throw up their own food in order to digest their food properly? Have you felt excluded from science spaces before? Then this blog post is for you!
Allow me to introduce myself.
My name is Myra, and I am a rising senior at SUNY Stony Brook University, where my major is Ecosystems and Human Impact, with a biology minor. In a nutshell, my major is interdisciplinary with a focus on conservation and ecology within human societies.
If I were to describe my college experience in one word I’d pick “surprises”. I never actually saw myself being a scientist in my middle and high school years. I found it hard to care about abstract concepts or theories that felt so far removed from humanity, particularly minority communities. But, during college I found myself falling in love with environmental studies, and along with it, the beautiful complexities that come with being human in our increasingly anthropogenic world.
At UMaine, we focus on the One Health Initiative, which views the health of humans, animals, and the environment as interconnected. When COVID-19 caused everyone to go into lockdown, I was fortunate to find this farm was looking for crew members, with a focus on food security. While certainly not how I planned to spend the summer of 2020, farming for underserved communities is where I saw how impactful One Health was. Organic farmers commonly use plastic mulch as a popular alternative to pesticides for weed suppression. At my home institution, I lead a project on the impacts of microplastics on earthworm health, an Ecotoxicology lab (students of the lab affectionately gave it the nickname “the Worm lab”). We use earthworm health as an indicator of soil health, which in turn is crucial for crop flourishment. The Worm Lab and farming emboldened me to pursue science and, ergo, look for this REU!
At UMaine, I am a member of the Ishaq Lab where I work on the camel metagenome project. Basically, scientists in Egypt raised camels on different diets, then used samples from their feces to sequence their microbial genome. These microbes live in the camel rumen (part of the camel stomach), and help the camel digest their food. What I do with Dr. Ishaq’s lab is, I perform data analysis on these sequences to see how the microbial gene profile changes with different diets. Camels are essential for transportation and food for the communities that rely on them, so finding the most efficient feed for them is important. Camels also release methane depending on their diet so it’s possible humans could control methane production of camels through their diet.
Being a part of the REU ANEW program for 2021 definitely has been an interesting experience, since it is the first time this program has been conducted virtually. Even though I would have loved to have seen everyone in person and spent time in lovely Orono, Maine, I’m glad for the research opportunity as it has further solidified my love of research and the One Health initiative.
Myra’s poster for the REU Research Symposium, virtual, Aug 13, 2021.
Nick Hershbine, undergraduate student majoring in Ecology and Environmental Sciences in the Ishaq Lab has been awarded a 2020-2021 University of Maine Undergraduate Student Employee of the Year!!!!
Nick began working with me in summer 2020, on a collaborative project looking at microbes in the soil around low bush blueberry plants. At the time, he was early in his undergraduate program and having no prior laboratory skills. In addition, because of the pandemic, we have minimized interactions between students, such that Nick has primarily been working independently with occasional meetings with me to discuss his progress. Despite all of that, he has exceeded my expectations for his capacity to handle this tricky project, and more than that, to have the independence to make it his own and contribute to protocol development. Nick has had a positive impact on the lab, within the confines of the limited interactions between students allowed at this time. He is kind and friendly, and makes the lab a welcoming place to work. Congrats on the award!!
Rebecca French, an undergraduate researcher in Animal and Veterinary Science, is beginning her time in the Ishaq Lab with an auspicious start: she has been awarded a 2021 research award from the J. Franklin Witter Undergraduate Research Endowment Fund! The fund supports AVS undergraduate student involvement in faculty supervised research which involves the J. Franklin Witter Teaching & Research Center.
Rebecca’s project will involve zoonotic disease tracking in rodent populations that live near farms/human development versus those which live in more natural areas, and will take place at the Witter farm and a paired natural ecosystem. Her project is part of a larger collaboration between myself and a team of researchers, which was recently funded by the University of Maine, but which has not yet been announced (details soon).
Rebecca formally joined my research lab in February of this year, but I have had the pleasure of teaching her in my data analysis class since January, which will be a handy skillset later in the project. She also learn and perform microbial culturing, qPCR, Sanger sequencing, and even some animal trapping, handling, and identification; mammal physiology data collection and analysis.
Rebecca French
Undergraduate Researcher, Animal and Veterinary Sciences
Rebecca is an animal and veterinary science student with a concentration in pre-veterinary medicine. She joined the Ishaq lab team in 2021 as a part of her capstone project, which is focused on flying squirrels and mice that are carrying zoonotic pathogens into Maine.
Emily joined the lab in early 2020 to work on a project investigating calf health and gut microbes, but very soon after joining the lab, the SARS-CoV-2 pandemic emerged and changed the way we were able to interact on campus. Without missing a beat, Emily shifted her efforts from helping me wrangle the lab renovations and sorting out our inventory, to helping me improve my teaching materials, to diving deep into previous literature to dig up protocols for her experiment in 2021: “Ideal Conditions for Cryptosporidium Attachment and Infection.“
We’ll be performing the experiment itself over the winter break, and then using the spring to analyze the data and write them up. As part of the CUGR award, Emily will be presenting her work at the 2021 Student Symposium in April, which will be held virtually this year. You’ll have to wait till then to get more details!
