Frank likes bees too.

When I was an undergraduate, I spent about a year or so working as a beekeeper. It was a fun job, and I learned all sorts of fun facts about bees. By this time I had been interested in parasitoids for nearly a decade and a half, having raised parasitic wasps out of caterpillars since I was five. Naturally, I attempted to see if there were any parasitoids which attacked Apis mellifera but I always ended up empty handed and disappointed. This always confused me because there were parasitoids which attacked ants, termites and caterpillars living in ant nests. I never understood why parasitoids had never been documented attacking honeybees.

This changed earlier this week, when a description of a parasitoid fly which attacks bees was published in PLOS ONE: A New Threat to Honey Bees, the Parasitic Phorid Fly Apocephalus borealis by Core et al. Unfortunately, the authors tried way too hard to connect the fly to Colony Collapse Disorder, but I’ll discuss that later. First…

What are Phorid flies? What are parasitoids? Why do we care?

Figure 1 from Core et. al 2012. I labeled the identifying characteristic of the family Phoridae with a black arrow and the ovipositor which is convergent with a wasp stinger with a red arrow.

Parasitoids are insects which are parasitic as larvae, free living as adults and which kill their hosts after development is complete. Most parasitoids are either flies or wasps, with wasps being the best studied. They’re important to agriculture because they’re good at regulating the populations of their hosts by killing them in large numbers. My studies revolve around how these insects evade the immune system, which gives us a springboard to learn more about how insect immunity works on a biochemical level.

Parasitoids in the family Phoridae are particularly interesting. Most species, such as the very common Megaselia scalaris, are actually scavengers but some species have made the leap to parasitism. The paper lists a particularly great example of parasitoid phorids, the decapitating flies which are used as fire ant biocontrol. These flies can be identified by a bunch of scrunched up veins on their wings, labeled by a black arrow in the first picture.

Figure 1 B from Core et. al 2012. The fly is visible laying it's eggs into the abdomen of the bee to the left of the picture.

Parasitoids go through standard holometabolous development of egg, larvae, pupa and adult. The adults lay eggs either on or inside their hosts. The larvae develop within the hosts, pupate, and then hatch into adults. Each of these stages requires particular adaptations, and parasitoid wasps and flies use completely different strategies. Larvae, for example, must evade the immune system. Wasps tend to suppress the immune system through venom or polydnaviruses. Flies, on the other hand, tend to hide in tissues which aren’t easily accessible to the immune system. They also have a tendency to hijack the immune system in some rather impressive ways like using melanization machinery to build snorkels to keep the parasitoid larvae supplied with air.

Sometimes, however, parasitoid flies and wasps solve similar problems with similar solutions. This parasitoid oviposits inside the host like the wasp, but uses a hard spike on it’s fleshy ovipositor to help it insert it’s eggs into the bee. I labeled this with a red arrow in picture A.

In the picture above, you can see the fly laying it’s eggs into the abdomen of the bee. From here, the eggs hatch and hatch into larvae which feed on the bee’s tissues. After this the larvae emerge in a particularly gruesome and characteristic fashion, between the head and thorax. The larvae emerge from under the ‘chin’ of the bee after the bee leaves the colony at night.

Fly larvae emerging from honeybee host, highlighted with red arrows.

This is a particlularly interesting example of a host shift. Apocephalus borealis is a specialist on bees and wasps, so a shift to Apis mellifera makes sense because it has a similar immune system. Honeybees hail from Africa, whereas this parasitoid is uniquely American so this is a definite example of a native parasitoid infecting and adapting to a new host.

What about Colony Collapse Disorder?

Despite the good job the authors did documenting the development of this parasitoid inside the honeybee, they lose my enthusiasm when they get to the colony collapse stuff. I really think they did some good work on natural history in this paper, that is they did some good work on looking at how the flies develop in the bees in labs. However, there are some things which weren’t very well fleshed out in this paper which mostly pertain to CCD.

I really think they tried too hard to connect this fly to CCD and this part of their work wasn’t very well performed. In their defense, I think the main purpose of this paper was the natural history work on this parasitoid. The CCD work appears to be done as almost an afterthought, but given the high profile of the paper I thought this warranted it’s own dissection.

To investigate internal hive behavior and possible infections within a hive, we kept an observation hive in a laboratory near our primary study hive. Samples taken from the observation hive in June 2010 confirmed infection with A. borealis. Rates of infection varied between June 2010 and December 2010 (Mean = 25% Range = 12%–38%) peaking over the sample period in November at 38%. In September, the number of bees in the hive declined and we observed phorid pupae and empty pupal casings among dead bees at the bottom of the hive, indicating emergence of adult phorids within the hive and the potential for A. borealis to multiply within a hive and infect a queen.

Let’s overlook the fact this was observed in a single hive kept differently than how most bees are housed. There’s a bigger issue here.

At my university, I am the curator of the insect zoo. We used to have some pretty big problems with Megaselia scalaris in our roach colonies. M. scalaris is a very common phorid fly that develops on dead and dying insects. Weakened hives aren’t able to fight off scavengers, so it’s possible in my view that Megaselia could have been in the hive after the populations declined because the large number of fresh dead bees would be the perfect environment where this scavenger could develop. The authors didn’t explicitly explain how they differentiated Apocephalus puparia from Megaselia puparia, and I think this is a fatal flaw in their work. These are two very similar looking Phorids with ecological habits that couldn’t really be more different. Megaselia does not develop as a parasitoid, and would thus pose no threat to the bees.  I think this is a rather important oversight and I wouldn’t trust their conclusions without further explanation of how they differentiated between the two. Put bluntly, I don’t think this piece of data should have been in this paper without that information.

Secondly, they did a bunch of tests looking for bee pathogens in the Phorids. They looked for genetic material, correctly noting this didn’t necessarily indicate that the pathogens were growing inside the flies. Quite frankly, I think it would be very strange if a parasitoid which fed on tissues of a bee didn’t consume any of it’s pathogens even if those pathogens didn’t infect the fly. Many science writers have been confused by this result, and many articles give the impression that the authors thought the flies vectored the pathogens. I find that doubtful but I won’t completely rule it out. Either way, I need to point out that it doesn’t come anywhere near proving it because there is no data indicating the pathogens were growing inside the flies. They also pointed out a correlation between Phorid emergence and the point of the year when colonies collapse, but demonstrating causation needs more data. The authors explicitly stated all of this, but some writers didn’t realize this.

Third, the majority of the data dealing with how the bees act when infected with the flies appears to have been conducted on a very specific building of the campus of San Francisco State University. That’s well and good, but urban beekeepers are a very specific subset of beekeepers and this data might not be relevant to most beekeepers, and thus irrelevant to the hives most important to agriculture. I think they need to do a lot more work on behavior before we can make any solid conclusions on how parasitized bees act in the wild before emergence of the parasitoids.

I don’t blame them for trying to connect this parasitoid to CCD, because that’s the hot topic right now in bee biology. However, I don’t think this fly really has much to do with the collapse of honeybee colonies. Despite this, this is still a really important find because this fly could shed more light on CCD, but not in the direct way the researchers imply in this paper. This is an insect which invades the bee and evades it’s defenses. Figuring out how the fly evades the bee’s defenses could shed light on how the bee’s immune system works. Figuring out how the bee’s immune system works might help us figure out how whatever pathogen actually does cause CCD is also able to evade the defense mechanisms of the bees. Once we fully understand the bee’s defense mechanisms, we can then think about potential interventions based on this data.

And now… some gratuitous parasitoid videos. These are particularly impressive examples of physical grace and biochemical warfare from some of the most incredible critters in the world.

A special thanks to Bug Girl for pointing out the Biodiversity In Focus article.

Core, A., Runckel, C., Ivers, J., Quock, C., Siapno, T., DeNault, S., Brown, B., DeRisi, J., Smith, C., & Hafernik, J. (2012). A New Threat to Honey Bees, the Parasitic Phorid Fly Apocephalus borealis. PLoS ONE, 7 (1) DOI: 10.1371/journal.pone.0029639

I like to think of myself as a skeptical blogger. I like to engage in critical thinking about scientific issues because this is an important aspect of my job as a graduate assistant. When I move into the workforce, I’ll still need some basic skills to parse evidence because this is my job as a scientist. Mythbusting is a great opportunity to do this, and I enjoy discussing things which may help people who read my posts whenever I can. Being an entomologist gives me some rather interesting opportunities to do this, which is leading me to discuss head lice of all things.

Pediculus humanus capitis by Gilles San Martin via Flickr.

In my last post, Do OTC Head Louse Treatments Work? Part 1: Mechanisms, I explained how the most commonly used FDA-approved treatments worked. In addition to those science-based products, there are many products that have no evidence of efficacy behind their claims, and that rely on fear to make a sale. What I’ve seen deeply concerns me not only as a scientist trying to make the world a better place, but as a parent trying to raise my daughter the best that I can. In this post, I’ve taken a few commonly sold products and listed some ways in which I think they play fast and loose with their claims.

A very brief review of how the nervous system works and how pesticides work in general can be found in the video below:

How do we know if treatments work?

Before getting down to the business of mythbusting, I think it’s appropriate to discuss how we know various products work. Treatments are assessed through clinical trails where infested volunteers subject themselves to putative treatments which are known as in vivo trials. In some cases, the lice are removed from the volunteers and exposed to the treatments in petri dishes which are known as in vitro treatments. In vitro treatments must be performed with the louse’s biology in mind because removing the louse from the host means that the louse is no longer in its natural environment. If the louse is not in it’s natural environment, the results gained from such a test may not be applicable to a real infestation. In vitro tests can disprove that a product works under ideal conditions, but proof of efficacy ultimately requires that the product be tested in real world conditions.

Clinical trials must have large numbers of people (and large numbers of lice) and untreated control groups. After all, insects are surprisingly fragile critters and even water or non-insecticidal shampoos may result in a small amount of mortality which is insignificant to treatment. Water or noninsecticidal shampoos can also temporarily clog the insect’s spiracles, resulting in immobile lice which could be interpreted as dead by a careless counter. Removal can physically injure the lice, which could cloud trial results if results are drawn from collected lice.

Another important aspect of clinical trials is blinding and randomization which make sure the person who is counting the lice isn’t aware of the treatment the person received. The human mind is a surprisingly bad tool for science because we tend to see patterns where none exist, and we may unintentionally superimpose patterns that don’t exist. Since everything in nature has some amount of variability (Anastasia is about six inches shorter than I am, Bug Girl is about a foot shorter than I am, and my boss is about a foot taller than I am for a quick example) we use statistics to tell us what the probability is that our results are due to random chance, eventually ending up with something known as a P-value. Followup observations are also required to show that the patient remained louse free, that is that there weren’t any hidden adults or unhatched eggs because the unhatched eggs can restart infestations.

Last week, I discussed some common OTC head louse treatments. While effective, there are some problems with resistance for some OTC treatments which results in failure of some treatments. This is a product of evolution where some lice are able to survive treatment because they have some random mutations which just so happen to be beneficial in a pesticide filled environment. The mechanisms of this resistance are actually similar to agricultural pests which have been treated with the same product.

One thing astute readers may have noticed is that I didn’t shy away from the use of the word ‘pesticide’ when discussing these treatments. One of my very first posts on Biofortified revolved around the definition of the word ‘pest’ which is completely anthropocentric. A pest is any critter which annoys us in the slightest, and a pesticide is a compound which kills a pest. Insecticides are used to kill insect pests and head louse treatments are referred to as ‘pediculicides’ because they kill lice. All pediculicides are insecticides (because they kill lice, which are insects), and many of the less toxic insecticides used in agriculture have been repurposed as pediculicides. Often times with head louse treatments, you hear companies claim with great pride that their products are pesticide free, are great at killing lice and that no resistance has evolved to their treatment.

Well… how do these claims stack up?

Uncomfortable Truths

Current packaging of products discussed in this article. Product images taken from the websites of their respective companies, used in accordance with the Fair Use Clause in US Copyright Law.

The use of agricultural insecticides to treat head lice is somewhat of an uncomfortable truth, and many companies have taken advantage of this to market head louse treatments. Despite what any label you read may say, any product which claims to kill lice is an insecticide by definition. It doesn’t matter if these are plant extracts, because pyrethrum falls straight into this category and it is classified as an insecticide. In fact, I would even go so far as to argue that a product is engaging in false advertising if it claims to kill headlice while being pesticide free. This, of course, doesn’t mean that all products must directly interfere with the inner workings of lice to be potential treatments.

Some compounds like mineral oil are used as insecticides in agriculture to kill aphids by suffocating them. The product marketed as ‘Lice MD‘ in the picture above claims to kill lice through a similar mechanism. The fact that these chemicals do not interfere with the neurological systems of insects does not mean that the product isn’t an insecticide. If the product claims to kill lice, as Lice MD does, it is claiming to be an insecticide.

A couple of examples of products that play fast and loose with advertising, in my opinion. Remember: natural products can be just as bad as synthetic products. Also, any product that claims to kill something is certainly not pesticide free. Both images taken from the websites of their respective companies, and used in accordance with the Fair Use Clause in US copyright law.

Uneasiness about insecticides has also given rise to many products which are derived from natural sources; these are popular because of a general assumption that natural products are safer than synthetic insecticides. The advantage of this from a company’s point of view is that these products don’t have to go through safety or efficacy testing, depending on how they’re marketed. The Dietary Supplement Health and Education Act of 1994 allows many products to go straight to market without testing under the guise of ‘supplements’ which allows them to make sometimes outlandish health-related claims. Homeopathic products are similarly exempt from safety and efficacy testing, which gives companies a great loophole to sell products which make medicinal claims.

This is a successful tactic because it plays on the unease parents have about treating their children with insecticides to kill lice. Unfortunately for these uneasy parents, the assumption that natural products are less harmful than synthetic products doesn’t always hold true. The LD50s for many synthetic pyrethroids are higher than their natural counterparts. Ricin and amantin are both incredibly powerful poisons derived from plants and fungi respectively. Eucalyptus oil, if used improperly as a head louse treatment, can have dire consequences including seizures and death. Many natural components can have chronic effects, too. Cyclopamine, derived from Vetratum californicum, causes some rather disturbing birth defects by inhibiting developmental pathways. Rotenone, a pesticide once used widely in organic agriculture, has been linked to Parkinson’s disease in workers exposed to sublethal doses of the toxin over the course of a very long time. Aflatoxins are powerful carcinogens produced by fungi which threaten food supplies all over the world. Even something as seemingly innocuous as a cockroach sex pheromone can be carcinogenic.

To be safe, it doesn’t matter if a chemical is derived from natural sources. Instead, safety depends on how the chemical interacts with the molecular machinery that keeps us alive. The safest way to make a new product is to construct it with chemicals where we know what everything does, as opposed to treating with soups of unknown composition. Unfortunately, this isn’t always possible because purifying and testing a compound for effects is extremely expensive and can take years of effort.

One of the components in the Quit Nits formula is a plant called Delphinium, a plant genus which is famed for its toxic alkaloids that poison cattle and make this plant genus a pest of cattle pastures. I’ll discuss this claim further in another paragraph, but the plant is in the formula at a concentration that is most likely too low to harm either lice or people. Other claims on this product are technically honest, but misleading. Some components of this product have been used in agriculture as insecticides. Lice consume a blood diet, and would probably have to eat these pesticides to see any effect. Components of the shampoo are toxic, but the concentrations these components are used in are harmless to people and are probably harmless to lice as well. As far as I can tell, most of these components haven’t been tested against lice in literature available to researchers. The statements made in the Quit Nits product comparison chart are misleading on multiple levels.

Misleading Statistics

Study methodology and results from the Lice Sheild website. This usage of the information from the company's website is in accordance with the Fair Use Clause of US copyright law.

Many products give misleading statistics that aim to trick parents into believing the product works. For example, let’s take a look at a product called Lice Sheild. They give a description of an experiment on their website that seems like a good test on the surface but is missing any information that actually allows you to draw any conclusions. For instance, they give P-values in their experimental setup, but do not give any information needed to verify their results. The P-values given imply statistical significance, but without any information on repetitions, sample sizes, means, or deviations the information is basically useless. There’s no way to check their math to see if the statistics were correctly performed. There are also no details on how many lice they used for the experiment. If they used two repetitions of five lice (the minimum required for 80% repellency with four lice moving between hair strands and away from the treatment), this would be a negligible result. If they used ten repetitions of 500 lice, the results would be a bit stronger. There simply isn’t enough information here to determine if the statistics were correctly performed.

Also lacking is a description of the experimental arena which is important because there are many ways in which you can test repellency that would potentially interfere with louse movement. If they placed the louse in a container in a patch of hair, one wouldn’t expect lice to move away from the hair at an appreciable rate. Many products are tested by placing the lice on a piece of filter paper and looking at the percentage of lice which move away from the product. While a test like this may appear to show some repellent activity, it’s not actually a very good measure of how good your product repels lice in real world conditions. Using in vitro tests means you’re trapping the lice in a place with the repellent and giving them essentially unlimited time to make their choice. This is a big problem because they’re using a timeframe that’s irrelevant to head louse transmission. Lice are mainly transmitted through hair to hair contact, and it’s rare for two people to be in hair to hair contact for this amount of time.

In short, showing 80% repellency is very different than saying that you have a 80% reduction in your chances of getting head lice. The company which makes Lice Shield uses questionable statistics to claim their product repels 80% of lice under conditions that don’t reflect the conditions where lice are transmitted, then turns around and claims in their FAQ this means that there is an 80% reduction in head louse transmission without any evidence for this claim. These two claims are quite different, because repellency doesn’t necessarily translate to a reduction of infestation.

Homeopathic Medicines are Marketed Differently than FDA Approved Drugs

Homeopathy is a system of beliefs which claim that serially diluting a ‘medicinal’ substance makes it stronger. These dilutions are pretty specific, for example the letter X denotes a 1:10 dilution. If a product is diluted 6X, it’s diluted to 10^-6 which is about one millionth of the original concentration. The idea that diluting a potential louse treatment makes it stronger is ridiculous because if the chemicals in any treatment interfere with insect biochemisty, they must be within a certain range to have an effect. Too much, the person gets poisoned (but the lice still die). Too little, and the lice survive and resistance can build up after the more susceptible individuals are culled from the population.

Homeopathy defies essentially every principle in science from biology to physics. Homeopaths claim that water has memory but, to paraphrase Tim Minchin, this ‘memory’ of water seems infinite when paired with some substances but water seems to have a case for amnesia when it comes to more harmful substances. Homeopathy has no a plausible mode of action.

The next paragraph of my post may get me into hot water with some of my skeptic friends. Many skeptical bloggers have taken on homeopathy. Let me restate: homeopathy has no plausible mode of action. I want to put this in writing to avoid the inevitable criticism from other skeptical bloggers. I also want to avoid the quote-mining from naturopaths who may want to say that I support homeopathy. I am not claiming the efficacy of homeopathy because there is no evidence that it works, and there is no plausible mechanism by which this practice could possibly work.

I am considering some homeopathic products as potentially effective. Why? Homeopathic formulas are exempt from safety and efficacy testing by the FDA which gives many products a free pass when it comes to clinical trials. Many products aren’t actually homeopathic because they contain ingredients in concentrations that could potentially have an effect. These products are often classified as homeopathic so they can make medicinal claims. A cold remedy product marketed under the name Zicam is a good example of this. Zicam was a solution of zinc which was marketed to treat the common cold after it was shown that zinc ions could interfere with viral replication in in vitro tests. The product was eventually recalled by the FDA because it was found to destroy the sense of smell. Another example of a product sold as a homeopathic remedy with potentially active components is sold under the name of Quit Nits.

Unlikely Modes of Introduction

Quit Nits bills itself as a homeopathic remedy and contains a bunch of plant extracts from several different species. As a result of intense selection by insect herbivory, all plants have some sort of anti-herbivore defense. Many plants have toxic components as a result of being under selective pressure to develop such components over the course of millions of years. Plants represent a wonderful treasure trove of different types of novel pesticide chemistries. After all, this is how we got pyrethrum. Despite the fact plant extracts are potentially plausible pesticides in and of themselves, we shouldn’t assume that any plant can kill any insect.

The first thing that raises a red flag for me in the Quit Nits formula is the mode of introduction of this pesticide. Some pesticides (see this RNAi post, for example) must be eaten to be toxic, and these are referred to as stomach poisons. Others can be absorbed, and are referred to as contact poisons. While this product does have toxic components, the fact these plants have natural toxins doesn’t automatically mean that they’ll be absorbed by the lice. Because the lice feed by inserting their mouthparts into the host, it seems very unlikely to me that they’d actually be able to pick up any pesticide by eating it unless the pesticide was in the blood of the host in appreciable amounts. Thus, any active ingredient would have to be absorbed through the exoskeleton.

A second thing that I am concerned about is the formulation. Spraying plant extracts on crops and lathering the same stuff into hair and then washing it off are very different modes of introduction. Pesticidal activity may not be preserved by the shampoo, even if the substance is downright toxic to bugs when dissolved in water. This stuff needs to be tested on lice in the formulation offered for sale before it can be said to have insecticidal activity. The mode of introduction and dosage play vital roles in the insecticidal activity. It’s possible the active ingredients wouldn’t retain their insecticidal activity in shampoo or that they wouldn’t be in contact with the lice long enough to be toxic. No pesticide kills every insect with equal efficacy in every situation. This is why the ultimate test of any pesticide is to test it on the pest in the situation you’re going to use it in, in the formulation in which it will be used.

Third, the mode of action of the two active ingredients means that they are unlikely to affect lice. Extract of the plant Delphinium and extract of a plant called Sabadilla (Schoenocaulon sp.) are listed as active ingredients at one part per million. There appears to be little work evaluating Delphinium for insecticidal activity, but Sabadilla and Delphinium both contain veratridine which acts as a stomach poison in insects. Because lice feed by inserting their mouthparts into the skin of the host and sucking blood from capillaries under the skin, I have a tough time believing they’d actually pick the insecticide up in appreciable amounts unless the toxins were absorbed directly into the bloodstream. Since the active ingredient is toxic to humans, there would probably be some major issues with the product if it made it’s way into the bloodstream. The mode of action here renders me skeptical that the lice would pick up a toxic dose of the pesticide in the first place.

Pesticides Used in Doses Unlikely to be Effective

The biggest problem with Quit Nits is the concentrations of the active ingredients. It’s the dose that makes the poison and if you look at the label of the product in question, there are two ingredients that are listed as parts per million and one component that’s in there at a 1:100 dilution.

Purified components of Sabadilla have been used as pesticides for high value orchards like oranges and mangoes. The lowest concentration for semi-purified Sabadilla alkaloids is about .1 g/l, or about one part in 10,000 if we’re going by weight. The extract of the plant seeds, of which the alkaloids are only a small part, is about a hundred times lower than this in the quit nits shampoo. The seeds of Schoenocaulon contain 2-4% insecticidal alkaloids by weight, which means the alkaloids from Sabadilla are present at one part in 25,000,000 in the shampoo.

Delphinium contains veratridine in appreciable amounts as well and has a large amount of other toxic alkaloids in addition to veratridine. It’s difficult to know what concentrations the insecticidal alkaloids are present in Delphinium because there are simply many potentially insecticidal alkaloids in these plants. However, we can make some educated guesses because researchers have purified alkaloids from Delphinium. The individual components are present in milligram amounts with all the alkaloids being present at about 6 grams per kilogram of plant tissue. If we assume the insecticidal alkaloids are present at a concentration of five grams per kilogram of plant material to make our math easy, this means that the insecticidal components comprise about one part in two hundred per unit weight. The concentration of plant in the shampoo is about two parts per million, which means the alkaloids are present at one two hundredth (1/200) this concentration. Given generous assumptions of grams per kilogram amounts, the active ingredients would be present part per hundred million concentrations if we assumed all of the alkaloids in the plants had insecticidal activity.

These plants combined are in about one part in 500,000 in the shampoo. This means that the concentrations of the insecticidal alkaloids is about one part in 4-6*10^-8 parts depending on the alkaloid concentrations of the plants used. Because the lowest concentration of this pesticide used in agriculture is one part in ten thousand, this comes out to a ballpark figure of somewhere around 1,000 times lower than the lowest dose used in agriculture. The dose the lice will be exposed to in the shampoo won’t be great, as there will only be a couple grams of the shampoo used on the entire scalp. To give you an idea of what the pesticidal concentrations are in other louse products, pyrethrum is generally in antilouse shampoos at one part per hundred (one percent). Malathion is generally present at one part in two hundred parts, or one-half percent. This means that the crude alkaloids from the plant extracts would be present at one one millionth the concentration of the active ingredients that have known insecticidal activity. The improbable mode of action combined with the low amounts of active ingredients in the plant means that I would assume these ingredients are essentially inert without proof that they kill lice at these concentrations.

These ingredients aren’t the main stuff in the Quit Nits treatment, though. The plant extracts listed above are in parts per million, but Quassia amara extract is present at a 1:100 dilution…about 10,000 times higher than Sabadilla and Delphinium. Furthermore, it’s in the ballpark of the Pyrethrum extract. So what about Quassia?

Ingredients with No Proof of Efficacy

Quassia amara is an interesting plant because it contains one of the most bitter substances in the world. These substances are called quassinoids, and have been examined for insecticidal and antifeedant activities against a wide range of pests. In many cases extracts and purified components from Quassia have been shown to have insecticidal and antifeedant activity, but it wasn’t always clear to me whether the antifeedant activity was so strong that it led to mortality. In other words, it was difficult to tell if the substance made the food taste so bad to the bug that they’d rather starve than eat. Either way, we’re interested in it’s activity against lice in particular.

There have only been two papers which have examined Quassia extracts against lice. One appears in a Dutch journal in 1978 and another in a Spanish journal in 1991. Since these papers are in rather obscure low impact journals, I was not able to access them directly through my library and instead had to rely on their descriptions in review articles. The review articles weren’t exactly favorable towards Quassia as a louse treatment. The Dutch paper claimed high efficacy, but the experiment was apparently ran as an un-controlled, un-randomized, un-blinded experiment and counts as nothing as far as proof goes. The Spanish paper claimed high efficacy, but the review states that the Spanish paper concluded that Quassia would only have repellent effects but didn’t mention whether the extract had a clinically relevant success rate. There have been no well performed tests of Quassia as a head louse treatment, and the few tests that have been performed have yielded conflicting results. There’s simply no proof that the “active” ingredients in Quit Nits work.

Quit Nits also sells a repellent spray that has undergone independent testing. One paper compared it’s repellent activity using a filter paper repellency test, incubating the lice with filter paper treated with repellent on one side and water on the other. The objective was to measure what percentage of the lice moved away from the treatment. At the earliest time point measured (two hours), Quit Nits performed about as well as water. At later time points, there was some non-significant repellent activity. Another paper looked at the repellency of Quit Nits under real world (or close to real world) conditions and looked at whether lice would transfer to hair under approximated hair-hair contact conditions, if the lice would move on the hair treated with Quit Nits repellent spray, or if the lice would feed on the forearm of one of the authors who performed the study. For the hair tests, KY jelly was used to simulate greasy hair and Quit Nits fared no better than this. For the skin tests, bare skin not receiving any treatment was used and the lice exposed to Quit Nits treated skin fed just as well as those on bare skin. Quit Nits repellent spray simply doesn’t repel lice, as far as the current experiments show.

Some Products Have no Plausible Mode of Action

The first example of a product with no mode of action is a product called X-pel. In fairness, I’ve only seen this at a few small grocery stores in Iowa, but the fact something like this is sold at all really worries me. The product is a shampoo which consists of ground up honeybees, phenol, and an uncommon species of Rhododendron at femptogram concentrations (15X), or one part in one quadrillion. On their website, they give a couple of vague descriptions of various tests. The tests contain very little methodology and give no statistical information about their results. They claim a few ‘major universities’ were involved in the testing of the product, but neglect to give any sort of contact information or any publications generated as I described in the Quit Nits treatment. They have a video on their website, below, where they show an in-vitro test that consists of them drowning a louse in the shampoo. Because lice can be inactive for a long time following immersion in water, there is no evidence given that the lice in this video were actually killed. They also show an uncontrolled, un-randomized, un-blinded test of a single subject without any apparent followup as proof that their product works. Phenol here is the most likely ingredient for insecticidal activity, as the concentration of the Rhododendron is far too low to do anything at all. Quite frankly, I’m not sure how honeybees are supposed to kill head lice unless we assume the venom glands were somehow involved, but I find this unlikely. The ingredients used in this product are only used in vanishingly small concentrations, and there’s really no way to justify using ground up honeybees to treat head lice.

Another product marketed under the name Licefreee! is little more than a concentrated sodium chloride solution. While it’s plausible the product could suffocate the lice, the data for suffocants in head lice treatment isn’t exactly convincing. Because lice are coated in a waxy layer that prevents dehydration, I find the claim that a 10% salt solution will kill lice suspect. As far as I can tell, there’s no evidence of this product works either because I’ve only seen this mentioned in passing under ‘folk treatment’ sections of review articles. I’ve seen no primary literature articles dealing with concentrated salt solutions as lice-killers.

Conclusion

Many of these companies use a variety of tactics to sell their products that have nothing to do with efficacy. Many use highly questionable advertising methods, like capitalizing on patient fears of synthetic medicines and pretending to identify with their customers to sell them products of uncertain effectiveness. Some of these products even go as far as to claim to be pesticide free while still claiming to kill lice. Many of these products claim to have been invented by parents, but as a parent myself I cannot imagine marketing a questionable head louse treatment and this is a big part of why I’ve written this post.

The science-based products currently on the market that I mentioned in Part 1 have been thoroughly studied and activity proven with the obvious exception for strains of lice that are resistant to some treatments. Even though there is a risk to any product you’re bound to use, the risks of these products have been investigated and have been taken into consideration when formulating treatment regimens. I can certainly understand anxiety about exposing kids to pesticides, but when looking at alternative treatments one needs to ask whether they’re safe and effective. Extraordinary claims require extraordinary evidence. If a product you spend money on makes any sort of claim, you should consider the claim extraordinary and ask for evidence behind the claim.

Burgess, I. (2009). Current treatments for pediculosis capitis Current Opinion in Infectious Diseases, 22 (2), 131-136 DOI: 10.1097/QCO.0b013e328322a019

Leone, P. (2007). Scabies and Pediculosis Pubis: An Update of Treatment Regimens and General Review Clinical Infectious Diseases, 44 (Supplement 3) DOI: 10.1086/511428

Canyon, D., & Speare, R. (2007). Do head lice spread in swimming pools? International Journal of Dermatology, 46 (11), 1211-1213 DOI: 10.1111/j.1365-4632.2007.03011.x

Heukelbach, J., Canyon, D., Olivera, F., Muller, R., & Speare, R. (2008). Efficacy of over-the-counter botanical pediculicides against the head louse based on a stringent standard for mortality assessment. Medical and Veterinary Entomology, 22 (3), 264-272 DOI: 10.1111/j.1365-2915.2008.00738.x

Lebwohl, M., Clark, L., & Levitt, J. (2007). Therapy for Head Lice Based on Life Cycle, Resistance, and Safety Considerations PEDIATRICS, 119 (5), 965-974 DOI: 10.1542/peds.2006-3087

Leone, P. (2007). Scabies and Pediculosis Pubis: An Update of Treatment Regimens and General Review Clinical Infectious Diseases, 44 (Supplement 3) DOI: 10.1086/511428

Greive KA, & Barnes TM (2011). In vitro comparison of four treatments which discourage infestation by head lice. Parasitology research PMID: 22030833

Canyon, D., & Speare, R. (2007). A comparison of botanical and synthetic substances commonly used to prevent head lice (Pediculus humanus var. capitis) infestation International Journal of Dermatology, 46 (4), 422-426 DOI: 10.1111/j.1365-4632.2007.03132.x

Rossini, C., Castillo, L., & González, A. (2007). Plant extracts and their components as potential control agents against human head lice Phytochemistry Reviews, 7 (1), 51-63 DOI: 10.1007/s11101-006-9026-0

When lice strike

"We all have lice" by Antonia Hayes via Flickr.

Head lice are something almost everyone has to deal with, and head lice treatments are something someone buys every once and awhile. These are big business in and of themselves. Because they’re big business, many firms have started popping up offering louse treatments with varying degrees of effectiveness.

A while back, I went through my own head louse ordeal with my daughter. Treatment was complicated by a family member who didn’t realize they were infested. We originally thought the lice were resistant to treatment, so I had to get a second treatment. Since then, I’ve become curious about what is for sale in stores for Over The Counter (OTC) head louse treatments and generally take a look at whatever treatments I can when I get the chance. Over the years, I’ve become surprised at how many dubious treatments are offered for sale (although perhaps I shouldn’t be) and how many of these use questionable advertising techniques mostly built upon fear rather than science. Many treatments offered for sale over the counter are either unproven, or have been proven not to work.

First, let’s discuss some headlouse biology. Then, let’s discuss how the treatments currently FDA approved work. In Do OTC Head Louse Treatments Work? Part 2: Questionable treatments, I’ll discuss the dubious treatments.

Disclaimer: I have worked in entry level positions at companies which sell these products. This, has not influenced my position. I should also mention that I’m not a medical professional and will not be discussing side-effects or risk-benefit analysis. I am an entomology graduate student who studies insect physiology and although I’ll mention the side effects of these products in passing I will not discuss them in detail. This post will discuss the science behind head louse therapies, how they work, and why some aren’t thought to work. I’ll also discuss the evidence that would be required to show that they work. This post deals with insect physiology and should not be mistaken for medical advice. Always consult a doctor if you think you may have health issues because blogs are notoriously bad places for medical advice*.

Lice biology

Male head louse, via Wikipedia.

Head lice are small hemimetabolous insects - basically booklice that have evolved to be parasitic. They start life as an egg or nit attached to hair, then go through a series of nymphal stages before maturing to an adult.  The adults and nymphs both feed, injecting saliva that causes small localized immune reactions which is why you itch. They’re very well adapted to hair and grasp it with clawlike legs. They can only grasp onto some kinds of hair which is why you don’t get pubic lice and head lice occurring on the same body parts. They spend all their lives on hair, even staying on the hair while they feed. Actually, off hair (or similar

fibrous material) human lice are nearly useless and have trouble getting around. They must feed every few hours, otherwise they quickly starve to death or die of dehydration off the host. Lice are transmitted mainly through direct hair to hair contact, with objects like combs and hats playing a potential minor role in transmission.

The nervous system of head lice is surprisingly similar to ours, with differences that are minor as far as we’re concerned. The nervous system is composed of several thousand cooperating neurons and is involved with every aspect of a louse’s life, movement, feeding and reproduction and many products target this system. When a nerve fires, sodium and potassium channels open which causes potassium to flow out of the cell and sodium to flow back in. Often, this process is touched off by the binding of another messenger such as acetylcholine which causes these channels to open. The charge changes from a negative charge to a positive charge, known as depolarization. The positive charge is very localized and moves down the nerve cell as a result of the sodium/potassium channels opening and closing in a very tightly regulated sequence that is essential to function. Other channels can prevent the nerves from firing such as GABA which binds to a receptor and causes the opening of chloride channels which prevent the nerve cell from firing by causing it to attain a very negative electrical charge. The pesticides used in headlouse treatments target all these systems, all of which can lead to a dysregulation of the nervous system and a collapse of nervous system function.

How do louse treatments work?

The safest products are sold over the counter and are used as commonly available first line treatments. Pyrethroids are generally considered to be the least toxic product, and are the most widely available. Lindane is a bit more toxic than either pyrethrum or malathion but most adverse reactions are still due to misuse. All three of these pesticides target different systems in the louse. Resistance has been documented in lindane and pyrethroid insecticides, but not malathion in the US. Pyrethroid based insecticides are used as a first line of attack with lindane and malathion being listed as a second and third route of attack respectively due to resistance of lindane and lack of resistance to malathion.

Pyrethroids are compounds similar to pyrethrum which is derived from the chrysanthemum plant.

Chemical structure of pyrethrum, the most commonly used pyrethroid derived from chrysanthemum plants.

Pyrethrum is a botanical product, while pyrethrins are artificial versions of this compound which have varying degrees of effectiveness on insects. In general, the artificial versions are more toxic to insects and less toxic to mammals based on LD50 values. Pyrethrum is the compound used in head lice treatments. Pyrethrum acts by propping open the sodium channels, allowing a sodium influx into the nerve cells. The nerve cells then become depolarized in unison, which results in the discoordination of the nervous system. The nervous system eventually shuts down, followed by the louse’s vital systems.

Resistance to this pesticide exists in two forms, knockdown resistance and cytochrome p450 degredation. Cytochrome p450s are enzymes which detoxify various compounds and catalyze a wide variety of breakdown reactions. These enzymes are present in humans as well, and also serve to detoxify the small amount of pyrethroids which are absorbed during treatment. In many resistant strains, the cytochrome p450s are upregulated, or overproduced. The overproduction of specific cytochrome p450 enzymes results in the increased breakdown of the pesticide. To combat this, a common additive called piperonyl butoxide is added as a cytp450 inhibitor. Knockdown resistance occurswith a change in the sodium channel that decreases the sensitivity to the pyrethrum, which is more difficult to combat. This is a wonderful example of evolution in action because it’s something which has evolved in direct response to usage of pyrethroids in headlouse treatment.

Chemical structure of lindane, courtesy of wikipedia commons. The molecule consists of a 6 membered ring decorated with chlorine atoms.

Lindane acts by binding to the GABA receptor and permanently inhibiting it. This results in the influx of chloride ions. With the GABA receptor stuck to the ‘on’ position, the nerves are unable to transmit any signals. This  mechanism is semi-complex mechanism but briefly the chloride ions reduce the charge in the cells, so much so that when the potassium flows out the nerve cell is still negatively charged and never fires. The nerves are unable to fire, and are in effect turned off. With the nervous system turned off, the louse becomes permanently paralyzed and dies as a result of not being able to feed. Resistance has been documented to lindane, but I’m not sure what the mechanism is. This product is one of the more toxic substances on the market for head louse treatment, and generally isn’t prescribed for children.

A third mechanism revolves around acetylcholinesterase, an enzyme not directly involved in the transmission of nerve signals. Acetylcholine Is used as a neurotransmitter, being sent between nerve cells to cause them to fire. When an action potential reaches the end of a nerve cell, the nerve cell releases acetylcholine which results in the nerve cell firing. Acetylcholine is degraded by an enzyme called acetylcholinesterase. Without acetylcholinesterase, the nerve remains permanently depolarized and the ion gradients collapse.

Chemical structure of malathion. The active portion of the molecule is the phosphate-like group on the far left which modifies the place in the enzyme responsible for catalyzing the reaction which shuts off nerve cells temporarily.

Even though acetylcholinesterase isn’t directly involved in the transmission of the signal, the enzyme is still important in ensuring the proper working of the nervous system. Organophosphates such as malathion knock the enzyme out, killing the insects. Malathion is an interesting molecule in and of itself. Toxicity requires degredation to another product, which happens better in insects than in mammals. Malathion is sold in a solution that contains isopropyl alcohol and tea tree oil which both synergize the effects of malathion by mechanisms which aren’t well understood. They work either by denaturing protiens in the lice as in isopropyl alcohol or by acting as a supplementary antiacetylcholinesterase as in tea tree oil.

Another method which has been used to cure head lice is what I refer to as the ‘nuclear option’ (or, to use a rare euphemism… landscaping for crab lice), and that’s simply removing the child’s hair. Without hair, the lice cannot hold onto their host and simply fall off. While side effects of the above treatments are relatively rare when the pesticide is used properly, this is by far the safest and most effective method of louse control. Unfortunately, this may not be acceptable for many people. When my daughter had head lice, she did not want to have her head shaved and this is the case for many little girls.

Are there treatment risks?

Although I’m keeping this post focused mainly on the mechanisms of these pesticides, remember that it’s the dose which makes the poison and any substance can be toxic when given in a high enough dose. Exposing yourself to a small amount of pesticide is OK so long as you allow it to break down and leave your system. Repeated exposure over a very long period isn’t a good thing because these products do inhibit neuronal function, leaving the door open for possible neurodevelopmental effects. Because of this, these products are not reccomended for long term use and treatment regimens are designed to last as short as possible. Head lice generally take about ten days to two weeks to mature into adults which is why retreatment is recommended within a week. Many products (except lindane) do not kill eggs, so any leftover eggs will hatch and eventually grow to reproductive adults if a followup treatment isn’t performed. The active ingredients have proven useful in a variety of contexts, including agriculture, but in this case the trick is to treat the patient with a dose high enough to kill most of the lice but low enough to not cause symptoms in the human.

Classifying these chemicals as pesticides sounds scary to many, and many companies have figured out how to take advantage of the unease many parents feel about treating their kids to sell products which have no evidence of efficacy. Next week, I’m going to expose many of these products and further explain the science behind clinical trials for these products.

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* One of my favorite fellow entomobloggers, Bug Girl, even has a page titled ‘I will not diagnose you’ and this applies to me as well. Do not contact me asking for any diagnosis because any E-mails of this character will be sent directly to my junk E-mail folder as it is outside of my duties as an entomologist to perform this sort of work and would be completely irresponsible.

However, the internet gives me some hope. Online science based communities such as this one, scienceblogs, freethought blogs and others are extremely popular and don’t show any sign of declining. Another site’s been a favorite of mine, a site on the immensely popular internet website Memebase called Dropping the Science. It’s chock-full of nerdy inside jokes and descriptions like the one below of how science *really* works. I even like the name, Dropping the Science. It’s defiant and confident… something you’d say when you’re about to unapologetically present a winning argument. Go check it out.

see more Dropping The Science

Tip ‘o the hat to ERV

The topic of the debate was “can organic agriculture solve food scarcity problems?”. The subjects were randomly chosen and don’t necessarily support the views of those engaged in the debate, so I will not speak for anybody but myself. I was on the con team, and we were charged with arguing that organic agriculture is an inferior method of food production. We were up against a very good team and all day folks were coming up to us and telling us how much they enjoyed our debate. Ultimately, we won the best overall debate team and took home an engraved trophy and left the meeting $125 richer after splitting a $500 prize between the four of us.

My role on the team was to look into the pesticides used in organic agriculture and their treatment regimes. To my surprise, I found that organic operations actually increase the amount of inputs put into the environment by requiring higher concentrations and more frequent applications of pesticides. The insecticides used in organic ag are often less effective, less selective, and can have greater non-target effects than synthetic insecticides. Some organic pesticides, like the biopesticide Beauveria bassiana, are assumed to have a very low environmental impact quotient (EIQ), but haven’t been tested for potential ecological side effects. My position (and position on the debate team) is that GMOs like Bt corn are better for the environment because they decrease the amount of pesticides that we must put on crops and that organic pesticides are worse for the environment because they must be constantly reapplied in very high concentrations.

This, however, wasn’t the idea that earned me my stripes during the debate. During the Q&A session, somebody asked us to clarify why we thought organic ag was able to innovate to a lesser extent than sustainable or conventional agriculture. My response was that we can modify pesticides to become less toxic, more easily degradeable and more difficult for insects to detoxify by producing insecticides synthetically and making it more or less difficult for the insecticides to degrade. While organic ag could certainly benefit from new chemistries, they reject them as soon as modifications such as these take place because the new pesticide is synthetic. In short, organic producers are unable to take advantage of novel chemistries. I used the example of adding carbon atoms or benzene rings in a specific place to keep beta-lactam antibiotics medically relevant during the debate, but there was a much better example I could have used but unfortunately neglected to discuss. But, hey… that’s what the blogosphere’s for isn’t it?

Very recently, the lab of Reddy Palli has figured out a way to genetically modify an organism to become a spray-on pesticide. To fully understand and appreciate what’s going on, there are a lot of things I need to explain. Fortunately, I’ve got about 12 hours of travel time ahead of me. Awesome, right?

A Colorado potato beetle. USDA photograph by Scott Bauer via Wikipedia.

First, let’s talk about the animal discussed in the paper. The Colorado Potato Beetle is what’s referred to as a ‘superpest’. It’s highly prolific, and essentially bulletproof. This insect specializes on solanaceous crops like potatoes and tomatoes, the crops most closely related to nightshade plants. These plants are famous for defending themselves by producing deadly secondary metabolites. By specializing on these plants, the Colorado Potato Beetle has evolved with some incredible detoxification mechanisms which shields it from our pesticides. As an unfortunate (for us) side effect, it manages to become resistant to every pesticide we throw at it very quickly. It can defoliate entire potato fields, and we can’t stop it very easily. We’re desperate for new chemistry to counteract this pest.

Next, let’s talk about a very basic part of insect physiology. Insects, like humans, are made from proteins encoded by DNA. When a protein needs to be made, an RNA polymerase translates DNA to RNA, and a ribosome transcribes the RNA molecule to protein. This is pretty constant throughout the kingdom of life plants, humans and insects all use a similar system and there is RNA in everything you consume. It can get a bit more complicated than this (see below), but there’s one thing I need to point out – mRNA is always single stranded in eukaryotic organisms. Some viruses use a double-stranded RNA (dsRNA) molecule. This is kind of like DNA, but it’s made out of slightly different stuff. Insect immune systems are good at picking up stuff that looks like it shouldn’t be there and dsRNA sticks out like a sore thumb.

The beetle has an immune system just like us. Ever get sick? Did you get better? That’s your immune system working. Beetles are exposed to pathogens just like we are every day. A good example of this is a cypovirus, which is kind of like an insect rotavirus. When the beetle gets a cypovirus, a series of enzymes pick the dsRNA it makes from the crowd of mRNA and selectively degrades it by using that dsRNA as a template to scan all the RNA in the insect and then degrade it. This is called RNA interference, or RNAi.

How can we use this to our advantage?

The experimental setup Palli's team used. Everything's labeled pretty well, and very self explanatory. The larvae eat the leaf, eat the dsRNA which causes their own body to shut down vital systems.

Unlike our antibody production system the RNAi system is kind of stupid and won’t distinguish self from nonself mRNA. The reason for this is that RNAi is also used to make sure the beetle doesn’t produce too much of a particular protein. If it wants to shut down certain specific proteins, it can make small interfering RNA (siRNA) and allow the RNAi system to destroy the RNA. It’s physiologically important for the beetle to be able to do this, but there’s no doublecheck system. The beetle can’t tell if it produced the RNA or if the dsRNA came from another source.

Reddy Palli’s lab did something ingenious with bacteria. They inserted several sequences into a bacterium that made double stranded RNA to a variety of important proteins. These included the muscle protein actin, sec23 which is a protein involved in the transport of newly produced proteins, and a couple ATPases which are responsible for producing the ATP energy currency of the cell. After killing the bacteria but preserving the RNA, they sprayed the bacteria onto potato plants which contained Colorado Potato Beetles. They also did this with just straight dsRNA. The beetles eat the plants, they eat the bacteria and a whole load of dsRNA.

What happened?

Here’s the cool part: it actually worked. To me this is mind blowing because RNA is incredibly unstable, thanks to an oxygen attached in a rather unfortunate place which allows it to break the backbone of the molecule. There are also nucleases which degrade RNA so the bacteria had to be modified so they wouldn’t produce these enzymes. Keeping the molecule double stranded helps by making it more difficult for either of these reactions to occur, so dsRNA is more stable than regular mRNA. But it’s still an incredible thing to me that this even worked.

If you want to show this works, you need to first show that the mRNA levels drop in response to the treatment. Turns out that they do for all the genes involved. Actin is the muscle protein, the ATPases produce energy and sec23 and CopB are involved in protein transport. The control was something which has relatively constant transcription that wasn't target by RNAi.

The beetles ate the killed bacteria, digested the outer wall and released the dsRNA. The cells take up the RNA, and the RNAi process occurs just as described above. The RNA coding for actin gets degraded, so that the beetles don’t make new actin or repair their existing actin polymers. In short, their muscles fall apart, their cells don’t divide. Even their sperm wouldn’t move…all these processes are dependent on actin.  As a direct result, the beetles stop eating, stop moving and die. Similar things happened with the other genes. When sec23 and COPB are silenced, their proteins don’t properly get transported and modified, resulting in a buildup of nonfunctional machinery. When the ATPases are silenced, ATP is no longer produced and the beetle can’t produce enough energy to maintain vital life functions. From this research, it would appear there are a great diversity of genes we could target which opens up a lot more avenues of attack when making pesticides.

Now, there are some neat implications to this research. This was a ‘proof of concept’ paper, which means that this works on a particular organism with a particular set of proteins under ideal lab conditions but doesn’t directly deal with the economics, field conditions or range of pests that could be targeted. It’s exciting and this technique has a lot of potential, but a lot more research needs to be done before we could use this in the field. That doesn’t mean there aren’t good reasons to be excited to see this further developed, though. Even though this may be a somewhat limited technique (see below), I could still see this being used to create very highly specific insecticides that quickly degrade in the environment.

In general, this would be the ideal pesticide for an environmentalist because RNA is all around you, as are nucleases. The Colorado Potato Beetle produces RNA and siRNA. We produce RNA and siRNA. Bacteria produce RNA, but I’m not sure if they produce siRNA. This is essentially all-natural, with the only difference being that we’re telling the beetle to degrade proteins at the wrong time and at a much higher rate than it normally would. RNA degrades by itself pretty easily and RNA degrading nucleases can be found almost anywhere you look. The bacteria can degrade in the environment and have no components which aren’t found in soil bacteria except foreign RNA sequences. There’s no reason to think there would be any issues with the bacteria staying around in the soil for an extended period such as we’d see with DDT.

Despite my enthusiasm for this clever technique, I also don’t want to give anybody the impression this is a ‘magic bullet’ for pest control. Some critters take up RNA better than others. RNAi was discovered in nematodes using this technique, so we could potentially use this on nematodes as well as beetles. Honeybees are able to ingest RNA and acheive silencing, so we might even be able to target sawflies. We could not use this on moth pests because lepidopterans are notoriously difficult to perform RNAi in, which has led to caterpillars being more of a biochemistry rather than genetic model organism. Since a lot of pests like aphids pierce the plant and suck the juices out, this would be useless against them because they’re not actually ingesting anything on the outside of the plant. There also may be better ways to introduce the dsRNA and for all we know using viral machinery may be a better way to introduce and replicate the dsRNA. There’s a lot more basic research which needs to be done on this before I’d be willing to say ‘we could use this’. With this paper, there are good reasons to think this would work.

The results from the experiment. This is a survival curve, with the percentage of the larvae surviving plotted on the Y axis and the time of survival plotted on the X axis. As you can see, the survival curves dropped far below the controls. A is the bacteria encapsulated dsRNA, while B is the dsRNA without bacteria. Both work, despite the limitations I explained earlier.

In addition to needing to pay attention to the pests this could work on, we need to pay attention to the kinds of beneficial insects and other animals this would potentially harm just as we would any other pesticide. Actin tends to be pretty similar in all organisms. The other genes are really important, and are probably very conserved in genetic sequence. I would think this could have some potential nontarget effects on other beetles, flies or wasps that I’d be pretty concerned about the potential for syrphid flies to eat aphids coated in dsRNA filled bacteria, for example. I think it’s unlikely that RNAi would be able to be done for humans in this manner because we’re coated in nucleases and to perform RNAi we must actually envelope dsRNA viral components in artificial cell walls to prevent degredation in the bloodstream if we inject RNA into the body as we would with medication used to treat ebola. I’ll go into more detail about this in the next paragraph but even if we found that we could potentially perform RNAi in humans by doing this I wouldn’t expect any big nontarget effects because we could choose the systems interfered with in the insects and avoid using systems humans and insects have in common. We aren’t able to do this with conventional insecticides as well as we could with dsRNA because they often target systems humans and insects have in common like sodium channels and acetylcholinesterase. We do OK by making pesticides less toxic to humans (synthetic pyrethroids have LD50s 10x less than natural pyrethrum for a quick example), but we could always do better.

I’m not sure how big of a problem resistance would be, but I can kind of sort of speculate on this. RNA is difficult for some organisms to take up, so I don’t think it’s impossible for the organism to change its ability to uptake RNA. As far as easily imaginable forms of resistance go, I think this would be the most problematic form of resistance. Increased nuclease activity in the digestive tract would be an issue from a resistance management standpoint, as well. The beauty of this technique is that we can put any sequence of RNA into the bacteria to perform this technique. If we were to target insect specific insulin-like peptides, we could kill the beetles by causing growth deformities or by putting the insect in a diabetic coma. If we found that we could silence some of the metabolic machinery in a species specific manner we could target this. We could target single genes, or groups of genes and thus custom-tailor our pesticides to the pest itself. If the sequence of the RNA changed in response to the management, we could just determine if a different RNA sequence would work. It’s very exciting stuff, and it uses chemistry that’s already existing all around (and even inside) you.

It’s a good example of how technology can be applied in novel ways. In this particular example, we are doing something very simple-genetically modifying bacteria-to accomplish the relatively simple goal of killing crop pests. If we were to develop this further and get it ready for field use, organic agriculture proponents would be sadly unable to take advantage of this technique because they ban both synthetic insecticides and genetically modified organisms. Organic agriculture rejects many tools which could help them further goals which are certainly admirable. Unfortunately organic agriculture proponents attempt to maintain a false dichotomy between synthetic insecticides, genetically modified organisms and environmental issues. A lot of this stems from simple chemophobia, the idea that synthetic things are inherently bad. This causes the field to reject many good tools like this based on little more than fear and misunderstanding. Unfortunately, as a result of this I reject organic agriculture and refuse to buy anything organically produced despite the fact I agree with their goals wholeheartedly. I sincerely hope the field moves in a direction which places an emphasis on environmentally friendly solutions rather than perceived naturalness of interventions. Unfortunately, from what I’ve seen I don’t expect that to happen anytime soon.

Zhu, F., Xu, J., Palli, R., Ferguson, J., & Palli, S. (2011). Ingested RNA interference for managing the populations of the Colorado potato beetle, Leptinotarsa decemlineata Pest Management Science, 67 (2), 175-182 DOI: 10.1002/ps.2048
Zehnder, G., Gurr, G., Kühne, S., Wade, M., Wratten, S., & Wyss, E. (2007). Arthropod Pest Management in Organic Crops Annual Review of Entomology, 52 (1), 57-80 DOI: 10.1146/annurev.ento.52.110405.091337

Bahlai, C., Xue, Y., McCreary, C., Schaafsma, A., & Hallett, R. (2010). Choosing Organic Pesticides over Synthetic Pesticides May Not Effectively Mitigate Environmental Risk in Soybeans PLoS ONE, 5 (6) DOI: 10.1371/journal.pone.0011250

Kovach, J., Petzoldt, C., Degni, J., & Tette, J. (1992). A Method to Measure the Environmental Impact of Pesticides New York’s Food and Life Sciences Bulletin

If you’re a graduate student like I am, PHD comics is an essential source of comfort. Emotionally and physically, graduate work is difficult. You’re working long hours with little pay and often (but not always), little gratitude. There are reasons for this, but to get the job in the first place you’ve got to be bright and have at least some modicum of passion to get through it.

There’s a lot to be said for obtaining an undergraduate degree. The course work can be really difficult and it’s often done at a time when you’ve got no real idea what you want to do with life. Aside from the *ahem* classical experimentation, you’re exposed to a lot of new ideas for the first time and for many, this is the first taste of independence. Personally, I spent my undergrad years living half an hour away from school while trying to figure out how to work two jobs that were a 20 minute commute from one another. I did this all while juggling classes, relationships and a kid.

I got accepted to a school halfway across the nation, and met up with a new set of challenges both personally and intellectually. I’d spent some time away from home as a teenager, so I was emotionally prepared to some degree. However, I’ve found it difficult to maintain a long-distance relationship with my child while having a really heavy workload. The lab I work in is also pretty heavy in biochemistry, but the last chemistry class I’d taken was in 2007 so I had to quickly re-learn some pretty basic skills.

When I say that being a graduate student is taxing, it may sound like I’m whining but I’m really not. Mentally, science can be very difficult. Many find the knowledge generated by their chosen field simply amazing, but the work required to generate that knowledge tiring and boring. I’ve met more than a handful of students who truly love their field, but absolutely hate their day to day work.

There are good reasons for this. My undergraduate work consisted of caring for bees, with the occasional PCR work. I had a good idea of how to generally read some data, but when it came down to reading some of the western blots I was generating there were a lot more subtleties than I realized.

These things are really quite normal as far as I can tell, and every student deals with them to some extent. There’s a lot of self doubt which comes along with the job because you’re learning to do things which are very precise for the first time. When you’re learning in undergraduate work, the experiments are pretty much pre-designed and almost always cookbook type labs which have a high probability of success. Mentally, it’s hard to prepare a student for a three month string of failed tests.

Undergraduate learning is mostly about learning whether you’re capable of understanding the generalities of the field. Proteins do this, this is how the organism makes them… that sort of thing. The answers are in books, and there’s a lot of wrote memorization but unless you land a research position there’s not a whole lot of real applied work. If you do land a research position in undergrad, it may or may not necessarily be similar to what you do in graduate school… and what I’m doing now can’t possibly be any more different than what I did while I was an undergrad.

Truth be told, I don’t know if I’m good at what I do. I think I am, or at least with a bit more experience I think I will be. I’m generating data and learning how to be more efficient at what I do. With every test, I’m learning more and more about how to pick out the subtleties of the tests despite the fact I still have a tendency to over-read my tests or occasionally overlook a relatively simple test which could answer a question I have about something or other. I’ve come a long way since I got here, but I’ve still got a hell of a way to go.

I’ve opined before that scientists are rarely treated as actual people. Science is hard work, and to make it in the field you’ve got to be good at what you do. Every one of us has their own personal backstory, some tragic, others not so much… but ultimately we’re all people who’ve got a passion for a certain subject and rack ourselves day in and day out to try to make the world a better place by making sense of the universe.

If this movie is anything like Cham’s comic (and the scenes in the trailer do correspond to Cham’s comics…I’ve read every single one) this movie is a must-see if you want to see what it’s like to be a scientist.

One of many B-movies based on a giant insect rampage.

I like sci-fi. I’m not your typical Star-Wars nerd, instead I like B-movies. You know… the low-budget creature feature movies that entail some giant creature killing everything in sight? They’re fun, campy, not at all meant to be taken seriously, yet can be useful in teaching about biology due to their reliance upon urban legends. Still, some things about them do get on my nerves.

Let’s take an episode of the television show Fringe: Immortality (13th episode of the 3rd season). Fringe is your typical X-Files wannabe show with writing that’s sub-par even for prime-time TV. The show centers around investigators who investigate apparent criminal abuses of science. And there’s a doomsday device in a parallel universe, somehow woven into the plotline, which feels like a very uncreative and poorly done rip-off of the parallel universe in the Doctor Who episode Rise of the Cybermen.

Anyway, the Immortality episode is about entomology, in which a mad scientist genetically modified a sheep parasite which somehow has a protein which cures a deadly flu. The episode made no sense to me for reasons I’m going to get into in a few moments, but there’s something more important I’d like to get to first because I think it’s an important part of how scientists are viewed in popular culture.

If you watch the episode, something jumps out at you rather quickly. These are people who are chasing a man who uses insects to commit murder. Yet, the people who actually know stuff about insects are given less than 10 minutes of airtime. Furthermore (at least in this show) scientists are generally written as shallow, boring or creepy people and this episode was the epitome of that. “Bug Girl”  (at right) in this episode was little more than eye candy and the main characters seemed really impressed by someone who was spewing statistics jargon with the grace of a freshman student who is struggling to get a C. It was sad, and just another reminder how commonly scientists are treated information spewing machines by television writers.

A friend of mine (the original Bug Girl) wrote a piece critical of the episode. In particular, she criticized the resident entomologist for looking a bit too stereotypical. Entomologists have gone through a transition in how folks view them. People used to see entomologists as a nerdy guy in a khaki vest, but more recently we get someone decked out in full goth regalia. Truth be told, I’m fine with this because it’s not as bad as how people view other closely related disciplines like geneticists. Plus, I’ve met entomologists who dress fairly similarly to this in real life.  Some of my fellow graduate students regularly don full biker regalia to work. Ultimately, I think it’s funny how you almost never see a scientist in their traditional gear: jeans and a cheap T-shirt.

Blaberus discoidales by DigbyRigby via PhotoBucket.

When doing a TV show that involves insects, you’re beset by some rather interesting limitations. There are only so many insects available on the market and the type of storyline you do revolves around what you can get and how easy it is to handle. In this particular case, they used some sort of Blaberus species roach. I’m thinking Blaberus discoidales, mostly because they look about right, are fairly cheap, and would be easy for a TV producer to get.

In the wild, they mostly live in caves eating bat poo, but in the Fringe ‘verse they’re apparrently known as Skelter beetles which were a parasite of sheep which became extinct in an unexplained manner which Abrams thought was somehow irrelevant to the plot. Now… if they’d have mentioned they were related to something like water pennies or said something about a ‘missing gonopore’, I’d have forgiven the crappy writing, but they didn’t.

I halfheartedly jotted down some notes while watching the episode. Here’s my summary of this train-wreck interspersed with some scientific facts.

The episode opens with two guys in an airport-type building. The older dude switches the younger dude’s drink and ominous music ensues. Fortunately for the younger guy, instead of getting hit with a roofie he instead drank some invisible beetle eggs. The eggs somehow managed to hatch and consume him from the inside out, gaining an incredible amount of biomass within a few minutes.

Parasitoid wasps grow from an egg the size of a comma to a grain of rice in a week. Percentagewise, this is a HUGE increase in biomass. Total biomass, not so much. These beetles take about 20 minutes to go from microscopic eggs to inch-and a half long insects.

Anyways, the old dude follows the now sick young dude into a bathroom stall and some screaming ensues after which you see the older guy getting the hell out of there while still managing to keep his shoes clean. At first, I was thinking roofie… but then you see tons of beetles and then you suddenly remember that you’re watching bad sci-fi.

After a rather poorly compiled intro sequence, you see some federal agents bantering and happily collecting bugs from a guy who just died. There are a few scenes with a bunch of stuff that’s probably relevant to some sub-plot I don’t care about but they eventually arrive at a relatively stereotypical entomology lab where there are people pinning a seemingly unrelated assortment of bugs for unexplained reasons with someone’s pet tarantula laying randomly about on a table.

If we assume it was a taxonomy lab, they generally work on one group of insects whether it’s a single family of beetles, flies or mantids. This was just a hodgepodge of random invertebrates.

Bug Girl incorrectly ID’s a cockroach as a beetle and hits on a fairly uncharasmatic agent who has no real role other than to sit around and look kind of menacing. During the course of her mis-ID, she announces that an aircraft station was an unusual habitat for beetles while seemingly oblivious to the fact that beetles are found almost everywhere on earth.

A few minutes after she goes away, one of the creepy statisticians informs the crew that they weren’t in the midst of an outbreak… which kind of makes me wonder why they weren’t wearing protective gear in the first scene as well as why they were traveling and thus exposing more potential targets to the terror attack. She also tells them that ‘statistics’ suggest they’ll get at least two calls if the agents tell the public there might be a biological terrorist attack going on. This impresses the agents, who apparrently don’t realize that putting out a phone number and asking for information is essentially telling every crank in the world that they’re willing to listen to any sort of insanity.

Of course, creepy stats girl was correct because they get inundated with waaay more than two useless calls as well as one which tells them about a beetle expert which Bug Girl should have been able to find with a quick google search. They eventually decide to track him down… and hey, guess what… he’s the bad guy! This is rather convenient for the team because they’ve got like half an hour to track down a fairly incompetent scientist.

It turns out the beetle expert is trying to rear the beetles in humans to create a flu vaccine from the adult beetle.Why he didn’t merely create a cDNA library before his beloved beetles went extinct (or from preserved specimens stored at -80) and order an expression kit will forever remain a mystery. Making copies of the beetle genes for the desired proteins and then using your colorimetric assay to find the protein of interest after producing them in bacteria seems a lot easier than killing multiple people and working in an underground lab. Either way, the crew tracks him down and one of them manages to get nabbed by the not-so-bright scientist.

There’s some confusion by the Fringe team over whether or not the cute girl is infected after she got kidnapped. To fill the rest of the time slot, the scientist gets yelled at, some guns are drawn, he pulls a beetle out of his neck and it turns out cute girl isn’t infected with Skelter beetles (which would have made the show marginally better), she’s just pregnant and her boyfriend isn’t the dad.

This show is in it’s third season. Firefly didn’t last one season. We’re in desperate need of good science fiction and good roles for scientists on TV… this is just more proof of that.

One of the things I’ve been talking about here on Biofortified is the concept of a ‘pest’, which is a completely anthropocentric term. Different insects can be pests at one part of their life cycle and be totally cool in another. It’s one of those weird science paradoxes which make the field of entomology so much fun.

In my last series of posts I discussed a new way to bring down pest populations by letting transgenic insects mate with wildtype insects and letting their offspring wither and die from a toxin which builds up inside of them. It’s really just a variation of a technique that’s been around since the ’50s that uses a gene that codes for a toxin (part 2) in place of radiation (part 1). It’s great for everyone involved (except for the mosquitoes – part 3), and it could even lead to the technique becoming more widespread by nixing the use of radiation all together.

It turns out that on top of all that we may be able to make mosquitoes work to our advantage.

Vaccines work by stimulating an immune response. In short, vaccines are made by either injecting a weakened version of the pathogen or a part of a pathogen like an important protein that’s part of the pathogen into a person. This coaxes the body into producing proteins called ‘antibodies’ which bind to the pathogen. From here, one of two things may happen… either a white blood cell finds the antibody-coated intruder and eats it (think pac-man) or a complex forms on the surface of the intruder and makes lots of holes all over the intruder which will eventually kill it. Injecting bits of pathogens or weakened pathogens into the body essentially gives your immune system a ‘cheat-sheet’ for something it may eventually encounter.

Your body can make antibodies to pretty much any non-self protein, which is incredibly useful. When mosquitoes bite you, they inject numerous proteins in their saliva which act as local anesthetics, anti-coagulants, vasodialators, so on and so forth. You actually make antibodies to these proteins and that itching after a mosquito bite is actually an immune reaction to the saliva. I work with antibodies in my lab. I essentially look for insect proteins which allow cells to communicate by using antibodies bound to enzymes to cause a chemical reaction that I can see. These antibodies are produced by injecting a protein into an animal and then harvesting them so we can work with them.

So in short you have an immune system which creates antibodies to foreign biomolecules and uses them mark targets for destruction. You’re constantly under assault from parasitic flies which inject you with proteins to which you create antibodies. We can use antibodies raised in other animals to look for proteins by using antibodies raised to antibodies that are attached to enzymes.

If you put these two bits of information together, you begin to see how this could be a useful tool because you may potentially be able to use mosquitoes to produce and distribute vaccines. A paper was recently published in the journal Insect Molecular Biology which I think is a good start towards this goal and makes me grumble a bit near the end…  but we’ll get to that when the time comes.

Let’s first walk through the image above to explain how everything works (click to see larger image). When a gene gets turned to RNA and then protein, the first step is binding of an enzyme called ‘RNA Polymerase’ at a promoter. Promoters essentially tell RNA polymerase how much RNA to make and where to make it. Above, Yoshida et. al took a sequence that’s almost always expressed and put it close to a promoter for a protein called ‘Anopheline antiplatelet protein (AAPP)’ that is expressed in mosquito saliva. Doing this allowed them to create a protein which is always expressed, but only in the saliva.

Now for the big question… what did they put into the mosquito’s saliva?

You know how I love paradoxes? Sand flies are an awesome example of paradoxes in action. Sand flies carry Leishmania which is a horrible parasite which causes your flesh to form a giant sore and pretty much melts your flesh off. Different Leishmania species cause sores in different areas. Some cause mucocutaneous leishmaniasis which is where the victim’s face slowly gets eaten away, and there’s another form called visceral leishmaniasis which is almost always fatal. It’s a gruesome, devastating disease.

Here’s the odd part: Sand fly saliva aids in parasite transmission, but can also protect those who have been bitten. When a sand fly bites someone, something in their saliva seems to help along the infection. However, those who have been bitten by a sand fly recently are better able to fight off Leishmania infection. It’s weird, I know. Those who are vaccinated with SP-15 from sand flies are protected against Leishmania infection.

The test below the picture of the gene’s set up is a Southern blot, which is essentially the scientists looking for the DNA of the genes they inserted to make sure they got there.

There’s some other stuff on there, as well. They fused SP-15 with a red fluorescent protein to be able to see if SP-15 was being produced because production of the desired proteins are just as important as making sure the DNA actually got into the mosquito. The results, shown below, are pretty neat looking (click to see larger image).

What you’re seeing is a fluorescent red protein fused with SP-15, which allows us to see that SP-15 is being produced (along with a western blot of SP-15 to make sure nothing funky’s going on).

To look to see if they could use the mosquitoes to vaccinate mice, they fed the mosquitoes on the mice repeatedly with the mice receiving 1500 bites over the course of four days. After feeding the mosquitoes on the mice, they looked for mouse antibodies by immobilizing recombinant or synthetically produced SP-15 and then washing it with filtered mouse blood, which would let any antibodies present bind to SP-15. The mouse antibodies which bound to the SP-15 then were detected with goat antibodies raised to mouse antibodies linked to an enzyme which allowed the researchers to visualize that mouse antibodies had bound to the SP-15.

So… I think they showed that mice could potentially be vaccinated against SP-15. Their titers were low (activity at 1:300 dilutions was the highest), and the authors correctly noted this. To give you an idea of what ‘normal’ activity would be, I use 1:5,000 and 1:10,000 dilutions of antibodies for my work and that’s a bit high. Not seeing activity at a 1:300 dilution means the antibody concentrations are pretty low in that plasma.

They also didn’t actually do tests which would allow them to see if the mice actually fought the Leishmania parasites off. I also don’t like the fact they didn’t do a western blot on the mosquito saliva, but the fact the saliva was glowing red from the red fluorescent protein fused to SP-15 means that chances were pretty good the mosquito was salivating the protein. I don’t like the fact they didn’t make an effort to quantify how much SP-15 was coming out in the saliva. The fact the mice produced antibodies to SP-15 is good enough for now because it shows that the SP-15 protein was actually leaving the mosquito, but it seems like a big step to go from SP-15 in the salivary glands straight to vaccination. Minor gripe… it shows the same thing but with a lot of extra added effort. It’s not a fatal flaw, but I would have done things a bit differently.

Moving this system to humans would be another story altogether. 1500 bites over four days is about par for the course when it comes to animals. Humans, on the other hand… I’m not sure 400 bites a day would happen on most Americans. Also, if you consider the mouse’s size and the low antibody titers they got, I’m not even sure this would be sufficient to replace traditional needle delivered vaccines. I’m just not convinced that this would be useful in it’s current form. As proof of the concept, however, I think it shows potential and warrants further development. Not much more than that at this point, though.

The authors conclude with a statement I disagree with:

The concept of a ‘flying vaccinator’ transgenic mosquito is not likely to be a practicable method of disease control, because a ‘flying vaccinator’ is an unacceptable way to deliver vaccine without issues of dosage and informed consent against current vaccine programmes. These difficulties are further complicated by the issues of public acceptance to release of transgenic mosquitoes. Therefore, we intend only that the present study makes available a model system using a salivary gland-specific promoter as a potential tool to elucidate the saliva–malaria sporozoite interactions

I think they’re right in that this couldn’t be used directly for humans, but I think this could still be very useful in the elimination of diseases. I think they’re right in that there are pretty serious ethical concerns as well as practical concerns with using these guys as vaccine vectors. Gene linkage, pathogen mutation (such as what we see the flu do yearly), and and a bunch of other things like how quickly we’re able to produce new vaxquitoes could potentially render this approach untenable. More research is needed before we can even consider this as a public health tool.

Despite the ethical concerns, I still think this might have some use because not all diseases are strictly human diseases. Many have animal counterparts in their lifecycle and we could potentially use these mosquitoes to vaccinate potential reservoirs.

Let’s ignore their crappy titers for a second and assume we can get this working to the point where we could get this working on a large scale. Introduce the mosquitoes to the wild, get them feeding on non-human hosts by… say, removing the equipment that allows them to find mammalian hosts. We could potentially use this system to vaccinate a massive amount of wildlife with the intention of eliminating disease reservoirs which are important in the transmission cycles of many different diseases.

Let’s take West Nile as an example. West Nile is mainly transmittesd between birds by bird-feeding mosquitoes. It only gets into humans when a bird-feeding mosquito feeds on humans in lieu of avian bloodmeals. If we were to take mosquitoes which feed exclusively on birds, transform them with this gene and then make it so their larvae die using the last set of genes I talked about (part 2) and then release them before their hosts migrate for the winter, this would create a system which would vaccinate birds of something like West Nile without the issues of informed consent raised in humans.

If we look at this as a tool to vaccinate disease reservoirs, we could achieve the same goal of reducing the prevalence of or eliminating the disease while avoiding the ethical concerns of informed consent raised by using these guys to vaccinate people. I’d say it’s a win all around.

Being able to use mosquitoes to deliver vaccines is a pretty neat tool, but let’s face it… I can see the antivaxxers needlessly freaking out over this.

Yamamoto, D., Nagumo, H., & Yoshida, S. (2010). Flying vaccinator; a transgenic mosquito delivers a Leishmania vaccine via blood feeding Insect Molecular Biology, 19 (3), 391-398 DOI: 10.1111/j.1365-2583.2010.01000.x

If you live in certain areas of the south, it’s really actually very similar. There are lots of rowcrops… peanuts and soybeans instead of corn and soybeans but still a similar concept. Lots of crops. Everything’s green and pretty without a whole lot of biodiversity. There’s one other major difference, though…lots of areas look like this:

Image courtesy of Wikipedia Commons

The green curtain draped over everything? Kudzu.

Kudzu was a vine originally planted to control erosion which grew out of control. It grows quickly, is hard to kill and covers everything with a green blanket and crowds everything out by keeping sunlight from reaching the plants. The trees under that green carpet are all dead.

So… how can things get worse?

Simple, really… just introduce something which lives on kudzu.

In 2009, a graduate student at the University of Georgia’s department of entomology found a very odd insect which resembled a beetle, but wasn’t. It had sucking mouthparts and a bunch of other features which landed it in a group of insects called the Heteroptera. The problem is that the insect wasn’t able to be identified with any of the keys available for the US. It was a new family which hadn’t been recorded in the New World before. The insect was eventually identified by another group of researchers as belonging to the family Plataspidae, and the species found was Megacopta cribraria, which lives on Kudzu but also on beans.

Megacopta species are known pests of various beans, including soybeans. From a recent review of Megacopta biology:

A number of authors report that Megacopta spp. are pests of soybeans. Soybean yield loss ranged from 1-50% depending on density of the bugs. The reported pest status ranges from minor to severe. As an introduced species, this bug appears to have potential to be a pest of legume crops in the United States.

They also invade houses during the fall while looking for a place to overwinter and can be smelled from some distance away, so they’re an urban pest as well.

The pest status of this species isn’t really certain; experimental crops infested with the species showed no apparent damage, I can’t find any information about thresholds for this species in particular and a lot of the information out there seems to be for the genus level and not this particular species so I can’t tell how well it’s been studied in it’s native range. There just simply isn’t enough information at this time to say if it’ll be apocalyptic, a flash-in-the-pan concern or something in between which is dependent on region, weather and/or biotype.

However, it’s considered a pest species in most of it’s range and this still presents a serious potential problem for soybean growers in the south because we now have a species that quickly reproduces, can grow to huge populations and which has a refuge which quite literally covers the entire south.

D. R. Suiter,1 J. E. Eger, Jr.,2 W. A. Gardner, R. C. Kemerait,3 J. N. All,4 P. M. Roberts,5 J. K. Greene,6 L. M. Ames,, & G. D. Buntin, T. M. Jenkins, and G. K. Douce5 (2010). Discovery and Distribution of Megacopta cribraria (Hemiptera: Heteroptera: Plataspidae) in Northeast Georgia Journal of Integrated Pest Management

So… part 3. What’s the next step? Put it to the test!

A system is only good if we can actually use it. We can test stuff in a laboratory and under cages all we want, but the tests only matter if they have an effect on real-world populations. This step has now been taken. Mosquitoes which have been modified by the Oxford-based company Oxitec to carry tetracycline repressible lethal genes (as described in part 2) are being released into the wild to combat outbreaks of Dengue fever in the Cayman Islands (see press release). This real-world test will help us to see if this technique will work in real-world situations.

It's kind of pretty in a 'they're gonna kill us all' kind of way. Image via Wikimedia.

Dengue is largely spread by Aedes aegypti mosquitoes which feed pretty much only on humans and are present in the highest populations around our cities…they’re mosquitoes which have adapted to an urban lifestyle. They bite during the day, so we can’t use bed nets. They’re notoriously resistant to many insect repellents… and they’re evolving resistance to insecticides used in vector control programs. Carbamates, pyrethroids… even DDT shows resistance in many populations of these guys. We simply need new tools.

Thanks to Medfly research, we now have new tools. One of these new tools, the RIDL genes explained in part two, is what is currently being tested in the Cayman Islands. Genetically modified Aedes aegypti have been released in an effort to bring the mosquito populations down past the number where they can spread disease. They’re only releasing male mosquitoes which don’t feed on blood, so they can’t transmit the disease to humans.

An Aedes aegypti larva. Image via Wikimedia.

There’s an even cooler ecological quirk to this technique, though. The lethal protein takes awhile to build up in the mosquito larvae, which means the mosquito larvae take a while to die. Before they die, they actually compete with wild type larvae for resources which should help to keep populations from quickly rebounding. The larvae of Aedes aegypti live in water where they feed on detritus. They don’t feed on blood and are unable to transmit disease.

In an AP article titled Mutant mosquitoes fight dengue in Cayman Islands, Oxitec is predicting an 80% reduction in Aedes mosquitoes from their test release of modified mosquitoes. I’m muting my enthusiasm until I see some data from this test as well as an decrease in Dengue transmission. It’s one thing to release the mosquitoes with a prediction the populations will fall, but ultimately I’d still like to see proof that this will work on this system. This is a good step in the right direction, though.

However, the program is meeting a lot of resistance from many including anti-GMO groups. From the AP article:

“If we remove an insect like the mosquito from the ecosystem, we don’t know what the impact will be,” said Pete Riley, campaign director of GM Freeze, a British non-profit group that opposes genetic modification.

He said mosquito larvae might be food for other species, which could starve if the larvae disappear. Or taking out adult mosquito predators might open up a slot for other insect species to slide in, potentially introducing new diseases.

Human ecology is a weird, wonderful thing. We bring in all sorts of animals wherever we go from dogs to birds to rats, and there are urban mosquitoes which specialize on all of these. RIDL relies on species-specific patterns of mating and reproduction, so we can target it pretty effectively. If the Aedes aegypti mosquitoes die out, something else will take its place and Riley is correct on this. It could be something like Aedes albopictus which also spreads Dengue or Culex quinquefasciatus which also spreads disease (not Dengue, though) but the mosquitoes could also be potentially be replaced by something which feeds on birds and rarely bites humans. We won’t know until this happens because nobody can predict the future.

Of course, because there are mosquito species other than Aedes aegypti those poor mossie predators will be OK because that will be the only species affected by the program. Either way, though… Riley’s arguing against mosquito control rather than the genetic modification of mosquitoes as a tool. I think this is weird because, vector borne diseases kill millions of people across the globe every year. I think it’s simply a good idea to try to eliminate the disease.

The next statement I found more than a bit odd because Riley seems to be using the concept of accidental species introduction or removal as an argument against pest control:

Humans have a patchy track record of interfering with natural ecosystems, Riley said. In the past, such interventions have led to the overpopulation of species including rabbits and deer. “Nature often does just fine controlling its problems until we come along and blunder into it.”

More than half the world’s population is at risk for dengue and over 50 million cases occur per year around the world. I’m not sure why he’s concluding nature’s doing just fine on this one because from the perspective of dengue sufferers… nature’s doing a crappy job managing this problem.

We have a patchy track record when it comes to ecology, and we should take it for exactly what it is. There are both successes and failures embedded in our history, even in mosquito control. The point is that we learn from both our failures and our successes. If something doesn’t work, we try again with a new technique. The Sterile Insect Technique has been a smashing success for all pests that it has been used for so far, and this is merely a re-invention of the technique which is an improvement over the original.

Consider this, though… the main reason the Panama Canal took two attempts to finish was because of yellow fever. We eventually figured out that if we eliminate mosquitoes from our living areas, yellow fever cases dropped. We were then able to finish the Panama Canal, and today it’s one of the main hubs for international trade routes.

This problem did not ‘work itself out’. Nature did not ‘do just fine controlling the problem’. We did this… we triumphed over the hostile forces of nature.