Biology question for FRAMEshift

[BlueBee]

Hey Frameshift, since you made the mistake of telling us you're a biochemist, now you're going to get all our Bio questions :-D

Have you read this article about a technique to mess up the machinery that runs the Varroa cell life?  They hope to muddle up the cell machinery enough that the cells die and mites along with them. 

http://www.apinews.com/es/noticias/item/12833-reino-unido-destruccion-de-un-gen-en-la-plaga-varroa-destructor-en-las-abejas-con-un-enfoque-no-invasivo-estudios-del-glutation-s-transferasa

I'm wondering if you might be able to summarize in layman's terms how this might work?  A lot of us probably have a descent science background, and know the basics of cell operation.  However this idea of throwing a wrench in the DNA/RNA transcription process is just a shade beyond what I know.

This is probably a dumb question, but how would you get RNA into a mite cell (or any cell) without injecting it through a vector (virus) or through mechanical means (a needle)?  The paper is suggesting you could basically plop your mites (and bees?) into a water solution with some RNA and the RNA would be able to get into the mite cells and clog up their transcription resulting in cell and mite mortality. 

[FRAMEshift]

As I read it, they are targeting GST which is an enzyme in mites.  GST gives the mites resistance against pesticides, so if you could knock it out, the pesticides would be more effective.  I doubt that  the RNA technique would actually be used against mites.  They are just setting up a model for studying which genes are important in resistance.  So the next step will be to take these mites and treat them with pesticides to see if they are more susceptible with this gene knocked out.  Once they verify a target gene, they will find some other way to knock it out or block it's action.

These guys are just working with whole live mites, not cells.  And apparently the cells inside the mite take up the RNA unaltered, whether you inject the mites with it or just soak them in a water solution of the RNA.  If they develop some other way to stop GST they might introduce it though a virus vector as you suggest.

[Finski]

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Beekeepers have just undersigned notes: No gene manipulation!

[FRAMEshift]

Finski, this has nothing to do with gene modification.  They are not trying to introduce an improved GM mite.   :-D   They are using genetic techniques to study mites to see why they are resistant to pesticides.  

I'm not interested in their research goals because I do not and will never use pesticides on a bee hive.   I also would not be interested in a gene modified bee, it anyone ever comes up with one.  But that has nothing to do with this research.

[hardwood]

That's whats wrong with teens these days! They modified their jeans and now they won't stay up around their waists!

Scott

[FRAMEshift]

Exactly!   :-D

[Course Bee]

I wondered what was wrong the jeans these days. I saw a kid(I must be getting old) the other day at a gas station and he was having trouble walking his pants were hanging down so far. I Wonder if my bees are having the same problem since they've been hold up in the hive for two months.

Tim

[Burl]

  Just for discussions sake ---  When I was a teenager ( many , many years ago ) I remember seeing a filmstrip in science class about how a parasite that troubled sheep ( screw worm , if I remember correctly ) was found to be kept in check by sorting out adult males of that species and then exposing them to radiation until they were sexually sterile .  Then they were released into sheep grazing areas .  The film stated that the sterile males then mated , causing the females to lay sterile eggs.  Is this an angle that has been considered for V-mites ?   If not, well then would it be ?              ---Burl---

[FRAMEshift]

The film stated that the sterile males then mated , causing the females to lay sterile eggs.

I would believe you if you told me the female sheep did not get pregnant.  I'm having more trouble with the idea that sheep lay eggs.  :-D

No wonder the teenage pregnancy rate is so high.  Mix tight jeans with crazy science class filmstrips and you have the recipe for disaster.  

[Burl]

  Gosh  , did I word it that badly ?   Sorry for confusing you , really !  Maybe if we understood that the subject was to do with parasites , in this case what I remember to be the screw worm fly ( funny name , ha ha ) .  And what was being explained as a successful way of lessening their population .  Now let's suppose we are on a forum discussing a honeybee parasite and how it's population might be lessened .  Seems to be a predominately important topic to beekeepers .  Any serious answers as to whether or not anyone is aware of the thought of somehow sexually sterilizing male v-mites and introducing them into a colony of honeybees that has a v-mite infestation to see if it has a noticible effect on the v-mite poulation ?  Hope this is clear and understandable .                                          ---Burl---

[FRAMEshift]

I'm sure that works for some species.  The issue with mites is that the males hatch from eggs laid inside a  comb cell after it is capped.  They mate with females and then die, all inside the cell.  All the mites that you see attached to adult bees are female.  So the issue would be, how would you get a large population of male mites to irradiate.  And then how would you introduce those to the already capped cells where the females are mating? 

[Jason]

Hello Everyone,
Here is a re-post of some information I wrote in the "article" section for wd.  The original article was about RNAi.
I think the original question was about getting the RNA into the critter.  The RNAi technique uses very small RNA molecules.  If stabilized properly or in some kind of transport solution the RNA will enter the cells, expresion vectors can also be used, but then this may lead to a "permanent" incorporation into the mite or host.  One of the good aspects I think about RNAi is its transient nature...

Here is my original post:
The researchers that originally observed the process of RNAi (RNA interference) were recently given the Nobel prize.
RNAi has been used in multiple molecular biology techniques, mostly as diagnostic and research tools.  Recently some researchers are focusing on therapeutic uses for the technology (antivirals, and vaccines).  It looks like researchers are looking at RNAi for mite control.

RNAi are small single stranded oligos of RNA (ribo-nucleic acid, very similar to DNA) that range between 15-30 bases.  RNAi works by specifically attaching to targeted mRNA molecules (mRNA contains the information to make protein).  The RNAi oligo sequence can be made specific to only hybridize (bind to) to a specific mRNA.  When an RNAi molecule binds to an mRNA molecule it sets in motion a chain of enzymatic events in the cell.  The end result is the destruction of the mRNA molecule and therefore the protein the the mRNA codes for.  This mRNA destruction pathway is just one of the ways the cell normally controls protein expression.  Researchers can put the pathway into overdrive by providing additional RNAi to the cells (organism).

I figure the researchers would comb the apis and mite genomes (I think they are both finished now) looking for mRNA sequences that are divergent from each other in important genes needed for mite life.  The trick is to find RNAi's that only recognize mite sequence and not bee sequence.  Many important cellular genes are similar from organism to organism, but with the entire genome to compare I am positive differences will be found.  Also another thing to consider is multiple targets.  It would be feasible with RNAi to target multiple genes for protein expression knockdown.  With multiple targets it is less likely that resistance will be generated in the mite population.

The down side is that RNA is easily degraded.  So the half life would be low, but I guess for some that is a Plus.

If you have any other questions please ask.  I am a molecular biologist, not that that means much, but I don't have my bees yet ( I hope to start two hives in April) so this is the only question I can answer on a bee forum site right now :)

[WPG]

Hello Everyone,
....
If you have any other questions please ask.  I am a molecular biologist, not that that means much, but I don't have my bees yet ( I hope to start two hives in April) so this is the only question I can answer on a bee forum site right now :)

Welcome, glad to have ya here.

You may become one of the heavy hitters in the bee world.
This does become a lifelong addiction.

Goodluck with your bees.

[BlueBee]

Fantastic answer Jason, thanks so much! 

You did an excellent job summarizing the process in layman's terms, and I think I understand it better know.   

I'm guessing the RNAi molecules don't get copied and multiplied in the host cells like a virus, but instead each one locks onto a mRNA and prevents that mRNA/cell from making some life sustaining protein. 

This is not my area of expertise, but I wonder if you can really get enough RNAi into the mites to shut down enough cells to kill the mite.  If the RNAi got copied and multiplied like a virus, then the numbers would clearly on the side of the RNAi vs the mite.  However without enough RNAi, does the mite still win?

This technology does sound exciting, but it also sounds very spooky! 

[Jason]

Hey Bluebee,

Yeah I don't think the RNAi molecules will be "copied" or be made by the cell like a virus.  There is some speculation that as the mRNA is degraded that some of the sequence by products of the degrading mRNA will act as RNAi and further the process, however I think that will be very mRNA specific if it happens.

to the question of amounts that get into the mite: I think like any type of therapeutic the dose would need to be determined...and I think that may depend on the route of administration (like spraying the mites etc).  It may also take a much lower concentration of RNAi in the mites hemolymph if multiple mite genes are targeted.  If multiple needed protiens are downregulated at the same time then the effect could be much quicker....but this is just me making some guesses :)

I think that this technology could be very exciting, but great care would have to go into looking at the chosen mite target genes and making sure that Bee sequence, and beneficial microbe sequence etc do not match.  Cause you are right it is kinda spooky :)