Better Know a Scientist: Interview with Entomologist Erfan Vafaie

A few people have asked me to review papers about bugs. I started reading papers about butterflies and I kept falling asleep. So I decided to do the next best thing: interview an entomologist. I’m going to be interviewing my friend, Erfan Vafaie (@sixleggedaggie), who also has a blog up on sixleggedaggie.com  We’ve sent each other papers that we’ve found interesting over the past few years, and I’ve asked him about bugs and GMOs in the past to get his insights and perspectives. Like me, he is also an Iranian-Canadian, but lives in Texas. This will hopefully be the first in many interviews with people whose research I know nothing about in a series entitled “Better Know a Scientist”. Here we go!

Q: Most people spend a lifetime trying to avoid bugs yet you chose to study them. Why?

Monarch Butterfly.
Image from Wikimedia Commons. Kenneth Dwain Harrelson.

Oh boy! Because insects are super cool? Whether looking at how bees communicate with their colony-mates where a good nectar/pollen source is via a bee dance, or ants building intricate networks under the ground including graveyards, or the ability of a woolly bear caterpillar to tolerate freezing at temperatures as low as -70ºC, insects are incredibly fascinating in their behaviors, lifecycles, and physiology. One of my favorites are parasitic wasps, which kill their host by laying an egg inside it (endoparasitoids). That egg becomes a larva that slowly feeds on the insides of the host insect, while the host continues to feed and live normally. After a while, the larva will eat everything on the inside, metamorphose inside (similar to a caterpillar to a butterfly), and become a new adult parasitoid - That’s basically the movie Alien happening EVERYWHERE in your garden!

But it’s the combination of my fascination with insects and their importance in sustainable and fruitful agriculture (pun intended) that really motivates me. Farmers are some of the hardest working and under-appreciated individuals in our society. So what better job than to be able to help them whilst studying these mini-aliens we call insects?

Q: What bugs are you currently studying?

At this moment, the crape myrtle bark scale, which is an invasive insect of crape myrtles in Southern US. We are currently studying their population dynamics throughout the summer, as well as best management practices in the landscape and nursery. I also just finished up some work on aphids and will be doing some work on thrips and mosquitoes soon. Being in my position, you tend to work on a wide variety of insects on different commodities, depending on the needs of the industry in your area at the time.

Q: How does it make you feel that you’re not the only Iranian-Canadian-Baha’i living in the US with a blog about science? Does it make you want to take out my kneecaps so that you can claim the keys to the blog-dom?

Kneecaps would be unnecessary, seeing as how I’m the tallest Iranian-Canadian-Baha’i living in the US with a blog about science. I actually really admire your blog, writing style, and ability to relay information to a wide audience!

[Biochica’s note: I think that Erfan is trying to make me lower my defenses with this response. Well played, sir, well played.]

Q: A common criticism I’ve read is that studies on transgenic crops only examine health impacts in mammals, and not the impact on non-target bugs and critters. Do you think this is fair criticism?

I’m not so sure that’s fair. There are many studies on the subject and the research continues to investigate the impact of transgenic crops on non-target insects. Wolfenbarger and friends assimilated and analyzed results from several other research articles (meta-analysis) and found hundreds of studies with specific criteria related to the impact of transgenic crops (Bt specifically) on non-target organisms of cotton, maize, and potato. So there’s a substantial amount there and research continues being done in that area.

[Biochica note: the Bt trait will be mentioned throughout this interview. It is a bacterial protein in some transgenic crops that targets specific insects. It is also a common pesticide used in organic farming. For more information on the trait, please see this post.]

Q: Why do you think that the topic of neonicotinoid pesticides has gotten confounded with the issue of GMOs? As I understand it, they’re two separate issues.

I agree, they are two very separate issues. Neonicotinoids are a class of insecticides that were first born in the commercial market in 1985 in response to older chemistries (i.e. organophosphates and carbamates) that had higher mammalian, avian, and environmental toxicity compared to neonics (short for neonicotinoids). Neonics have since become one of the most popular insecticide classes used. They do not need to be used in conjunction with GMOs, nor do I know of any GMO seeds that depend on the use of neonics; thus making the two issues unrelated. There are activists that would name neonics as the primary factor responsible for bee colony collapse disorder, whereas other activists accuse GMOs for causing a completely different set of consequences.

People may be confounding the two issues because neonics have often been used as a ‘seed treatment’; the seeds are coated in neonics, and as a result, are taken up by the plant and protect it for the first 3 - 4 weeks of growth, which can be thought of as being similar to introducing a gene, such as the transgene in Bt-corn that protects the plant from within. The EPA has since published a document (‘Benefits of Neonicotinoid Seed Treatments to Soybean Production’) demonstrating that neonic coated seeds are ubiquitous in nature and not very advantageous in most situations. That’s the only reason why I can think the two may have been confounded in the past.

Q: I think it’s fair to say that you’re very tall. So tall, in fact, that it seems “unnatural”. How should my readers know that you’re not a genetically modified human being, who is advocating the safety of transgenic crops as the first step in some quest for eventual world domination?

Standing at 6 foot 7 on a good day (depending on atmospheric pressure, humidity, and sock thickness), the daily fight to prove that I’m 100% natural and organic has been a losing battle. Let us thus assume, for the sake of argument, that I am genetically modified or genetically enhanced in some manner. We can draw on the great anecdotal and scientifically accurate example of the mutants from the Marvel Universe and X-Men.

In that arena, there are genetically enhanced humans that continue to promote the welfare of humanity (i.e. Xavier and his school gifted youngsters) and those that are selfish and desire for the world to crumble (Magneto and his hard-headed goons). We can differentiate between the will to do good by viewing the actions, motivations, and thoughts of the person as a whole. Xavier continues to build strong and positive ties with common-folk, empowers the younger generation to embrace their inner powers, and shows integrity in goodwill towards mankind. Magneto, on the other hand, continues to harm humans and expresses that “Nature has made us superior. We are the living future of this mighty planet. This world is ours world now! Take it!” (X-Men: Graduation Day, S.5.E.4, 1997). I think those who know me personally would agree that my actions are more in line with the Xavier team than Magneto’s.

[Biochica’s note: Again, I think that Erfan is trying to portray himself as the “good guy” with this amazing response. That’s exactly what a genetically modified alien would try to do. Erfan 2, Biochica 0]

Q: If there’s one thing that you want people to know about bugs and GMOs, what would it be?

There’s a common misconception that the use of GMOs promotes more insecticide use and can hurt more non-target insects (i.e. beneficial insects like butterflies and honeybees) than conventional agriculture . Brookes and Barfoot estimated a drop in global herbicide and insecticide use as a result of GM adoption to be 172.5 million kg (between the years of 1996 - 2004), with cotton crops specifically decreasing just over 14% in total herbicide and insecticide applications.

Gianessi and Carpenter (2001) calculated that between 1995 (year before Bt varieties were introduced) to 1999, the amount of insecticides used decreased by 2.7 million kg of formulated product in just six states (AR, AZ, CA, LA, MS, and TX), which represents 14% of all insecticides used in those six states. In addition the number of spray applications/ha was reduced by 15 million which represented a 22% reduction in spray applications. The Arizona Cotton Research and Protection Council (2000) has stated that Bt cotton has helped to reduce insecticide use in Arizona cotton to the lowest levels in the past 20 years! That’s a win for reduction of pesticide use.

In addition, incorporating the insecticide into the crop is an intuitive way to keep the non-target organisms safe; by integrating the insect toxin into the plant, you’re effectively targeting the insects that are eating your crops! Marvier and her friends analyzed several studies (meta-analysis) and found non-target insects to be more abundant in fields where GM crops were used compared to non-GM crops with insecticide sprays (using pyrethroids). It should be noted, however, that fields without GM crops and were not sprayed had higher numbers of non-target organisms than fields with GM-crops, so the GM crop does have some impact on non-target organisms, whether it be directly or indirectly (i.e. killing the insect that carnivorous insects eat).

Q: The spouse is from Texas, so we’ve been to quite a few country concerts. Have you been to one yet? If so, who? (You cannot lie on my blog, so you have to admit if it was Taylor Swift). If not, whose show would you go to?

Unfortunately, I have not been to any concerts in Texas as of yet. I have been to some local gigs/musical performances in Tyler Texas. They have some good talent here! I have, however, been to an A&M football game, with just over 109K people there. Let me tell ya - that was insane!

[Biochica note: As a country music fan who has shaken hands with Tim McGraw and has had a cowboy hat signed by Alan Jackson, this response is completely unacceptable. Erfan 2, Biochica 10000]

[Erfan note to Biochica note: Can we review how the point system works here?]

[Biochica note to Erfan: No. My interview, my rules :P]

Q: Are you aware of any evidence indicating that transgenic crops are responsible for CCD (colony collapse disorder) in bees? What about the decline of butterflies?

Colony collapse disorder, a very specific set of symptoms that have characterized large losses of honeybee hives since 2006, may be caused by several different factors: environmental stress, transportation of the bees, malnutrition, varroa mites, insect pathogens, and pesticides. Insect-resistant GMO crops incorporate genes from the bacteria Bacillus thuringiensis, which produce toxins that have relatively specific binding sites in the gut of lepidopteran (i.e. moth and butterfly larva/caterpillars) and some coleopteran (i.e. beetle) insects . As such, insects like honeybees are left unharmed. A meta-analysis (i.e. research article that looks at many studies) by Duan and friends analyzed 25 studies and found there to be no negative impact of Bt on honeybees in a lab setting. A more recent study by Dai and friends looked at the impact of Bt corn on honeybees in the field and found there to be “no difference in immature stages, worker survival, bee body weight, hypopharyngeal gland weight, colony performance, foraging activity or olfactory learning abilities”. So it appears that the current consensus is that transgenic crops are not a player in the whole CCD dealio.

Butterflies are a slightly different story, because as I mentioned just above, the toxins from Bt are specific to moth and butterfly larva/caterpillars. The very intent of the incorporated gene is to control/kill the younger stages of butterflies… so of course it’s going to kill them! The question we should ask, however, is whether the Bt is killing non-target butterflies on neighboring plants (i.e. milkweed) that we don’t want killed - afterall, monarch  butterflies feed specifically on milkweed, which is often considered a weed in the context of agriculture. A study by Losey and friends found that Bt pollen can reduce the growth, survival, and leaf consumption of monarch butterfly caterpillars. This study was trying to investigate what is referred to as “pollen drift”, which in the transgenic world means - “is the pollen from the transgenic crop drifting onto neighboring plants and causing unforeseen consequences?”. However, this study was done in a lab (i.e. unrealistic environmental factors), the pollen was artificially introduced to the monarch’s host plant in an amount that ‘seemed’ equivalent to the field, and there was no comparison to alternatives (i.e. Bt spray), so the jury is still out on whether Bt crops are a major concern for non-target insects.  One thing to consider, however, is that an entire corn field may take only up to two weeks to release all of its pollen, which is a very short exposure time to Bt-pollen compared to a weekly spray of Bt (or some other insecticide), which was the norm pre-Bt transgenics.

[Biochica note to Erfan: nobody's used "dealio" since 2002. There aren't enough points to subtract from your score to make up for this offense.]

Q: Do you secretly wish that you were a human geneticist like me? Do you want to join the private industry and swear a secret oath in exchange for never-ending wealth? I can put in a good word with my overlords for you at our next sacrificial ritual.

No. Certainly not.

(But secretly, yes! I submitted an online query to the League of Villainous Scientists through their contact page, with a full cover letter of why I’d like to do evil things to people and our planet for never-ending wealth, but haven’t heard back. A ‘good word’ would be most appreciated!)

Erfan and I would like to close with a more serious note: had our parents not immigrated out of Iran, both of us would have been barred from receiving education due to our religious background, and the world would be two scientists shorter. We’d like to bring awareness to the fact that many Baha’is in Iran are being excluded from receiving higher education and dozens are imprisoned for their beliefs. Please join the #educationisnotacrime campaign on Twitter and Facebook. For more information, please visit www.educationisnotacrime.me

Gene editing and GMOs

This month, I'm tackling gene editing. It's finally time I read papers on the topic because I got an email advertisement announcing a new gene editing kit and I realized that I don't know the mechanistic details of the system (yes... There's science email spam...).

I'll be summarizing a 2014 review from the journal Cell. The title of the review is "Development and Applications of CRISPR-Cas9 for Genome Engineering" (unfortunately, the review is behind a paywall).

As you can imagine, gene editing is somewhat of a holy grail. To be able to erase undesired mutations in DNA would be a dream for many clinicians/doctors. But there are many different applications that don't necessarily have to do with erasing what we don't want, rather, we could introduce variations that we want: creating an animal model for a disease, developing crops with desired traits, etc.

The paper starts by providing a brief overview of how genome engineering has been done in the past (focusing primarily on mammalian cells), outlining the pros and cons of each method. The difficulties inherent in current methods has led to the development of programmable gene editing technologies, the most promising of which is the CRISPR-Cas9 system which is the focus of the paper.

Spouse: I need to clarify something before I continue. The system isn't quite as simple as I'll describe below. There are additional DNA/RNA sequences and enzymes that are part of the complex, and I'll be omitting them because I could hear your voice in my head complaining about all the acronyms and jargon. So consider this to be the uber-abridged Cliffs Notes on gene editing.

Cas9 is an endonuclease, meaning that it's an enzyme that can cut both strands of DNA's double helix. Endonucleases can be random, meaning that they can cut anywhere along the length of the DNA. In fact, one of the fears of scientists working with DNA is nuclease contamination, which can render your samples to DNA dust. Other endonucleases, such as restriction enzymes, search for a specific DNA sequence and cut at that site. However, the cut site cannot be specified and can be found frequently in a genome (i.e. restriction enzymes can cut thousands of times).

Unlike restriction enzymes, the bacterial Cas9 cuts at a specific site but the DNA sequence where it cuts can be specified. Cas9 is associated with the CRISPR system, which guides Cas9 to its target using a small piece of RNA. In nature, this small piece of RNA generally encodes for a viral (phage) sequence. Cas9 searches for the viral sequence and then hacks it up, which is why the CRISPR system is part of the bacteria's antiviral defense mechanism. However, the small piece of RNA that guides Cas9 can be replaced with a sequence of the researcher's choice. Part of the system's benefit is the fact that you can provide more than one guide molecule, meaning that you could direct the system to cut more than one place, if desired. The system can be specific in the DNA sequence that it cuts, however, as this paper highlights, off-target edits can occur and are an area of ongoing study/research.

The paper provides a history of the many discoveries surrounding CRISPR-Cas9. There are different mechanisms by which the CRISPR system gets activated, and after many years of research, it was decided that the CRISPR-Cas9 was the most promising in terms of trying to find a programmable system for gene editing. By 2013, researchers successfully engineered the CRISPR system from two types of bacteria, including the one used to make yogurt, to edit genes in mammalian cells.

So far, I've described how to get the system to cut where you want it to cut. But then what? If you think of editing as deleting something that's incorrect and typing in something else, how do you get "what's right" or "what you want" into DNA?

Once the DNA is cut, the cell's natural repair mechanism kicks in and one of two things can happen (see my bee-u-ti-ful graphic below):

• The two loose ends of the DNA get glued back together again. The system is error prone but easy to use, so if your goal is to create a protein that doesn't function or to delete it altogether, this may be the way to go. This process is known as non-homologous end joining.
• The break is detected by enzymes that look around for the proper template to use to fill in the gap. If that template is provided artificially, then it will copy in that sequence. The template that researchers provide can contain the desired sequence, additional sequence, etc. This process is known as homology directed repair.

CRISPR-Cas9 can also be modified so that the "search" function of the system remains intact, but the cutting function is disabled. As such, researchers can create a complex where they guide their enzyme of choice to a specific region. I think that the simplest example that the paper provides is one where Cas9 was fluorescently labelled/tagged, so researchers could visualize the location of the DNA sequence they were studying.

In reviewing this article, the spouse asked if we could write a movie script where the villian sprinkled Cas9 along with the DNA specific to his arch-nemesis' genome into the latter's cereal. Would it be the perfect crime? Would the CRISPR-Cas9 enter the arch-nemesis' body and hack up his DNA? Unfortunately, no. Keep in mind that our DNA is within the nucleus of our cells and isn't very accessible. Additionally, as non-bacterial species, we don't have CRISPR-Cas9 in our cells. Getting the CRISPR-Cas9 into the nucleus isn't all that simple and requires a bit of fancy lab work.

To date, there's no medical therapy on the market developed using gene editing. Likewise, there's no crop on the market that has been engineered using CRISPR-Cas9. However, studies have demonstrated that crops can be modified using the system (this paper provides an example of successful gene editing using CRISPR-Cas9 in rice). Consequently, many wonder whether crops generated through gene editing would be considered GMOs.

As I've previously described, what are currently known as a GMOs or genetically modified organisms are transgenic crops, meaning that a gene from a different species has been added to their genome (NOTE: in this post, I note that anti-GMO activists have a very different definition of a GMO). But in the case of crops modified using CRISPR-Cas9, what's edited was there to begin with. Technically, nothing has been added from a different species. So how will regulatory agencies categorize these crops?

This paper published just last month provides a great summary: it states that the USDA has concluded that if you cannot distinguish an edit from a naturally occurring mutation, then it's not a GMO. Additionally, if a gene is deleted using the cell's own repair mechanism (as is the case with non-homologous end joining), then it isn't a GMO either. Interestingly, the paper states that the USDA has waived regulations on two crops generated using gene editing, because they fell within these categories. The European Union has yet to determine how these crops will be classified, because they consider something to be genetically modified if "it is altered in a way that does not occur naturally by mating and/or natural recombination" (although crops generated through mutagenesis are not regulated in the EU. Please see my previous post on mutagenesis for more information on this technique). There are two additional points that the paper makes that I completely agree with: 1) if the EU's definition of a GMO does not end up aligning with the USDA's, the regulation of these crops for import will be very difficult since there will not be an easy way to detect if the crop is a product of gene editing. 2) If the EU's definition of a GMO does not end up aligning with the USDA's then the cost of getting a crop through regulatory hurdles will limit the development of these plants to large biotech companies, which will stifle innovation; i.e: if you want someone other than Monsanto, Syngenta, et al to make a biotech crop, these crops should not be considered GMOs.

To conclude, here's my first "infographic" on the different methods or ways used to develop new traits in crops and feedback would be appreciated. My perspective on gene editing is the same as it is on transgenesis and mutagenesis: crops should be regulated based on the trait introduced/modified, not on the way that the introduction/modification was generated.