American Foulbrood (afb)

[rrussell6870]

This thread is to help people to understand more about afb... what it is, where it comes from, how it spreads, how to identify it, and how to resolve it....

As my time is limited, I will make posts on the matter as I have time to do so, instead of all at once... I encourage anyone with experience on the matter to lend a hand and their input... also, I encourage anyone with questions to please feel free to ask them... there will be no stupid questions in this thread as it is intended to inform everyone whether they are considering getting their first hive or are running tens of thousands of hives...

[SPRUCE BEE]

Just how prevelant is afb these days? I've only encountered it in the late 60's when it seemed to be pretty common. My old beekeeper mentor at that time said it was more prevelant in the Italian bees at that time. Which breed is most resistant? I'm assuming genetics has played an important part in current resistance? It sounds like maintaining healthy un-stressed hives is the best prevention.

[apis maximus]

It sounds like maintaining healthy un-stressed hives is the best prevention.

If the AFB spores are present in all the hives...and only some get to express the disease clinically...the above statement makes a lot of sense.

[rrussell6870]

Actually, that is a great starting point and one of the greatest issues faced by people not knowing what to look for... In so many cases afb is found to be consistant in strong early buildup colonies as opposed to weak dwindling hives as most would expect... thus the italians would have been to more common targets with their early and heavy brood rearing...

to fully understand the complexities of just how the bacteria works, I will first explain the bacteria Paenibacillus larvae ssp. larvae which was once classified as Bacillus larvae, but still commonly referred to as Bacillus larvae by most in the bee community...

Paenibacillus spp. is used in many agricultural environments since many were shown to be important for agriculture and horticulture (e.g. P. polymyxa), industrial (e.g. P. amylolyticus), and medical applications (e.g. P. peoriate). These bacteria produce various extracellular enzymes such as polysaccharide-degrading enzymes and proteases, which can catalyze a wide variety of synthetic reactions in fields ranging from cosmetics to biofuel production. Various Paenibacillus spp. also produce antimicrobial substances that affect a wide spectrum of micro-organisms such as fungi, soil bacteria, plant pathogenic bacteria and even important anaerobic pathogens as Clostridium botulinium.

Several Paenibacillus species serve as efficient plant growth promoting rhizobacteria (PGPR). PGPR competitively colonize plant roots and can simultaneously act as biofertilizers and as antagonists (biopesticides) of recognized root pathogens, such as bacteria, fungi and nematodes. They enhance plant growth by several direct and indirect mechanisms. Direct mechanisms include phosphate solubilization, nitrogen fixation, degradation of environmental pollutants and hormone production. Indirect mechanisms include controlling phytopathogens by competing for resources such as iron, amino acids and sugars, as well as by producing antibiotics or lytic enzymes.

Competition for iron also serves as a strong selective force determining the microbial population in the rhizosphere. Several studies show that PGPR exert their plant growth-promoting activity by depriving native microflora of iron. Although iron is abundant in nature, the extremely low solubility of Fe3+ at pH 7 means that most organisms face the problem of obtaining enough iron from their environment. To fulfill their requirements for iron, bacteria have developed several strategies, including
1. the reduction of ferric to ferrous ions,
2. the secretion of high-affinity iron-chelating compounds, called siderophores, and
3. the uptake of heterologous siderophores. P. vortex's genome for example, harbors many genes which are employed in these strategies, in particular it has the potential to produce siderophores under iron limiting conditions.

3.Endospores do not form normally during active growth and cell division. Rather, their differentiation begins when a population of vegetative cells passes out of the exponential phase of growth, usually as a result of nutrient depletion. Typically one endospore is formed per vegetative cell. The mature spore is liberated by lysis of the mother cell (sporangium) in which it was formed.

4. The formation of endospores is a complex and highly-regulated form of development in a relatively simple (procaryotic) cell. In all Bacillus species, the process of spore formation is similar, and can be divided into seven defined stages.
The vegetative cell
1. begins spore development when the DNA coils along the central axis of the cell as an "axial filament"
2. The DNA then separates and one chromosome becomes enclosed in plasma membrane to form a protoplast
3. The protoplast is then engulfed by the mother cell membrane to form a intermediate structure called a forespore
4. Between the two membranes, The core (cell) wall, cortex and spore coats are synthesized
5. As water is removed from the spore and as it matures, it becomes increasingly heat resistant and more refractile
6. The mature spore is eventually liberated by lysis of the mother cell.

The entire process takes place over a period of 6-7 hours and requires the temporal regulation of more than 50 unique genes.

Mature spores have no detectable metabolism, a state that is described as cryptobiotic. They are highly resistant to environmental stresses such as high temperature (some endospores can be boiled for several hours and retain their viability), irradiation, strong acids, disinfectants, etc. Although cryptobiotic, they retain viability indefinitely so that under optimal environmental conditions, they germinate into vegetative cells.

Endospores are formed by vegetative cells in response to environmental signals that indicate a limiting factor for vegetative growth, such as exhaustion of an essential nutrient. They germinate and become vegetative cells when the environmental stress is relieved. Hence, endospore formation is a mechanism of survival rather than a mechanism of reproduction.

Differences between endospores and vegetative cells that form them. (Note- Collumns are provind to be quite difficult on beesource, the first column is the property/ next is the vegetative cells/ and next is the endospores. Each is seperated by /.)

Property / Vegetative cells / Endospores

Microscopic appearance / Nonrefractile / Refractile

Calcium dipicolinic acid / Absent / Present in core

Cytoplasmic water activity / High / Very low

Enzymatic activity / Present / Absent

Macromolecular synthesis / Present / Absent

Heat resistance / Low / High

Resistance to chemicals and acids / Low / High

Radiation resistance / Low / High

Sensitivity to lysozyme / Some sensitive; some resistant / Resistant

Sensitivity to dyes and staining / Sensitive / Resistant

Due to the resistance of their endospores to environmental stress, as well as their long-term survival under adverse conditions, most aerobic sporeformers are ubiquitous and can be isolated from a wide variety of sources. Hence, the occurrence of sporeforming bacteria in a certain environment is not necessarily an indication of habitat. However, it is generally accepted that the primary habitat of the aerobic endospore-forming bacilli is the soil. The Russian microbiologist, Winogradsky, considered them as "normal flora" of the soil.

In the soil environment the bacteria become metabolically-active when suitable substrates for their growth are available, and presumably they form spores when their nutrients become exhausted. This is a strategy used by other microbes in the soil habitat, including the filamentous fungi and the actinomycetes, which also predominate in the aerobic soil habitat. It is probably not a coincidence, rather an example of convergent evolution, that these three dissimilar groups of microbes live in the soil, form resting structures (spores), and produce antibiotics in association with their sporulation processes.

Since many endospore forming species can effectively degrade a series of biopolymers (proteins, starch, pectin, etc.), they are assumed to play a significant role in the biological cycles of carbon and nitrogen.

From soil, by direct contact or air-borne dust, endospores can contaminate just about anything that is not maintained in a sterile environment. They may play a biodegradative role in whatever they contaminate, and thereby they may be agents of unwanted decomposition and decay. Several Bacillus species are especially important as food spoilage organisms.

[rrussell6870]

Ok, so if you can wrap your head around all of the information in the above post, you will have discovered that...

1. The genus bacillus has many purposes in agriculture and is found almost anywhere...

2. The P. Bacillus that is in every bee colony is not the spores, but rather the vegitative cells...

3. P. Bacillus reproduces (produces endospores from vegetative cells) as it dies from lack of nutrition rather than while it thrives... thus it produces endospores as a survival mechanism rather than a reproductive mechanism... but this only takes place when the vegetative cell find an optimum temporary environment...

4. As vegetative cells, P. Bacillus can more easily be destroyed than the endospores... However, once destroyed from a surface, the chances of that surface coming in contact with more vegetative cells are almost immenant...

5. You need a stout cup of coffee before you sit down to read any of my more descriptive posts.

[rrussell6870]

Now that we know what bacteria we are dealing with and how it lives and multiplies, lets move on to how it works within a bee colony and how the social behavior of bees effect its developement...

Ok, so understanding that this bacteria is virtually impossible to stay away from, How do we avoid issues with it?

1. The bacteria is no threat in a vegetative cell state...

2. Heavy amounts of vegetative cells must enter the correct age larvae via the larvae's feed... the optimal age is within the 24 hrs after the larvae has emerged from the egg and this window of opportunity is closed forever when the larvae reaches 3 days of age... only the larvae that injest heavy amounts of vegetative cells within that first 24 hrs will be suitable hosts for the bacteria to produce spores that can infect the colony, as the older larvae will develop to a harder state so the bees can remove them without releasing the P. Bacillus spores...

3. Once the vegetative cells recognise that the lavae is no longer providing the nutrient resources that they require (ie, the larvae has died from lack of nutrients due to the bacteria stealing it all), the vegetative cells will begin to produce endospores which are all racing to get to the vegetative cell state and is also how the bacteria is spread throughout the hive by the bees trying to remove the dead larvae that has turned in to a pool of goo in the cell...

This is where the "roping" effect comes from... the dead larvae is a host to millions of endospores at this point and it has become a high elasticity substance... as the bees pull and tug trying to remove it from the cell, it simply streches out, then snaps back in, releasing endospores on to the bees that are tugging and the edge of the cells around it... these endospores are then passed along all throughout the colony...

Keep in mind, that although endospores are now all over the colony, they still have to find their way into the feed for the perfect aged larvae in high numbers, just not quite as high as the vegetative cells... and since they are being produced directly within the colony, the possiblity of them finding a suitable host is greater than before... and since they are now travelling by nurse be, the possibilty of them being fed to a larvae is greater than before...

Once 4-8 larvae have been infected and become endospore production sites, the possiblities of spores finding their way into the guts of the right aged larvae have been multiplied several times over...

So what are the WARNING signs that you could run into AFB problems?
1. You keep honey bees...
2. Your honey bees buildup heavily...
3. Your honey bees have some level of hygenic behavior...

Wait a second!!! Those are not bad things in bee keeping!! What gives??

Thats exactly right, these are not bad things for the bees... the reality is that the chances of an infection are Extremely rare... In the natural state, honey bees have lived and thrived with P. larvae and AFB for millinium... some colonies die from it, sometimes it manages to spread from one colony to another... but the chances are litterally 1 in 1,000,000, in the natural state... But as bee keepers, we do not keep bees in a natural state... how could we? Could you imagine spacing your hives throughout the forests one hive every mile or so?? Not me...

So just simply needing to keep multiple hives within one space we have raised our chances of seeing P. bacillus create AFB by a very large amount...

Still the for an infection to start, a heavy stress is usually required, ie, heavy comb manipulation in cold and poor weather, unmanagable combs forcing the seperation of the enternal workings of the colony, etc...

Panic time? No..

Now its time to learn how to identify infection of different levels and test questionable findings both during your inspections and in the lab (if necessary)...

So now lets move on to identifying AFB infections... Starting with what it actually looks like, then explaining the different testing methods...

[jrbbees]

Thanks Russell,
You are blessing us. Would we have some one do this for every problem we are facing.
Moderators: Can this become a sticky or something always at the top?

[Oldtimer]

Just thought I'd add a little snipet to that (if I may, Robert).

It can be hard to understand why hives can have bacillus larvae bacteria in them but not get sick. One of the main reasons is that a bee larva has to be killed by the disease and turn into the sticky stuff Robert described, for the disease to take hold in the hive. But the way the bacteria kill the larva, is when the larva pupates, the bacteria penetrate it's gut wall, and kill it. If there are only a few bacteria, they cannot cause enough damage at pupation to kill the larva. The larva survives, and the infection is not spread. If the bee larva only gets infected with one bacteria, the bacteria cannot multiply into enough bacteria by the time the larva pupates, to kill the bee larva. The bee larva has to ingest quite a few bacteria while it is still quite young, for the bacteria to multiply into enough to kill the larva.

Therefore there can be low levels of bacillus larvae bacteria in a hive, but the likelihood of a larva getting enough to cause a fatal infection is very low, so the hive never shows symptoms of the disease.

Actually looking back I think that's kinda what Robert already said, but I'll leave it there, understanding this bit is part of the key to understanding the mechanics of this disease.

[Hawkster]

I am impressed both with the knowledge expressed and withDr Russell's response to an uniformed post implying he was quarantined. he turned it into a teaching event !

[jrbbees]

If I remember right the issue for Russells was EFB and not AFB and it only involved one small location and a small number of hives in that location. More important, he did the right thing. Worked with inspectors, pulled that yard completely out of production of bees or queens, and went through the treatment processes and has established a self imposed isolation of that yard that exceeds the inspectors recomendations. A man can't do better than that.

If you keep enough bees long enough you will take your turn with every major issue we face. You just wait your turn and have to do what is right when the hammer falls on you!

[preciousbees]

Now this is what I like to see, good solid information, a real educational opportunity here. All too often there is mis-information on problems we encounter with our hives. I want to thank the more experienced Beekeepers for sharing thier knowlege with us.

[arcowandbeegirl]

Wow, thanks for all the great information!!
How many people actually treat with tetracycline or tylan as a preventative?
I understand there are alot of people that dont treat unless they have to, but just wondered.

[AstroBee]

Dr. Russell,

Thank you for posting such a complete and well written source of information on AFB. This thread already is deserving of being promoted to a sticky.

Others, I respectfully request that we dedicate this thread to the opportunity to truly educate beekeepers on AFB. Please, lets not be tempted to dredge up issues being discussed elsewhere that do not pertain to the core mission of this wonderful thread. Thank you.

[megank]

Whoa..I think I just took mirco again. I will add that I agree with Dr. Russell. AFB is in every hive the same way anthrax is present on your kitchen floor, however, I disagree that hygienic bees are more susceptible as I believe he implied.

If the dead or dying larvae are removed before the ropey stage then that's 10 million innocuous spores that will be tossed out of the hive.

Also I'd like to add that for new infections to occur, three conditions must be present. A virulent pathogen (p. bacillus), a mode of transmission and a susceptible host. Break the link between any one and no new infections will occur.

Has any studies been done to determine the number of p. bacillus in either form required to cause a symptomatic infection of the larvae?

[megank]

I only treat with antibiotics when I receive comb of unknown origin. After which I quickly replace as winter approaches.

[Oldtimer]

Has any studies been done to determine the number of p. bacillus in either form required to cause a symptomatic infection of the larvae?

Yes that's been done.

I've actually read the info on it and had to pass an exam on it to qualify as a bee inspector, but now I cannot remember the exact figures.

However there's a certain number of bacillus larvae bacteria that a bee larva has to consume to create a fatal infection if the bee larva is less than 24 hours old. When the bee larva gets older, say, day two, it has to ingest a heckuva lot more before the infection would be fatal to it. The exact bacteria numbers needed have been established by testing under laboratory conditions.

Sorry I can't give exact numbers off my head, however the general idea is there.

[megank]

Thanks Oldtimer...If there's been studies, I can find it.

[Oldtimer]

I'll also see if I can get the info. I origionally borrowed someone elses literature to learn the "technical" stuff I had to know to pass the exam, and that was a few years ago. So I'll track it down but it might be a few days before I get back with it.

Pretty likely it will be on the web somewhere also, if anyone else gets it before me, I may not come back about it.

[Riskybizz]

Reading Robert Russell's latest posts regarding AFB properties in light of the recent allegations of his "quarantined" apiaries, PRICELESS

[WLC]

I've found the following paper to be interesting:

http://aem.asm.org/cgi/reprint/75/10/3344.pdf

It shows that there is more than one strain of AFB, and not only provides for a way to differentiate between strains, but also helps to bolster arguments against proscribed burning of hives.

Besides, if it's from Genersch, I would pay attention.

Yes Doc, it does help your cause and research objectives.