nosema ceranae

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Nosema ceranae:

Kiss of Death or Much Ado about Nothing?

Randy Oliver

Once again Im interrupting a series of articles to update beekeepers on breaking napplications in bee management. This time the information is aboutNosema ceranreaders of the Journal might benefit from an update on recent research results.

Dr. Mariano Higes team (from the Centro Apicola Regionalin Spain) electrified the beekeeping world two years agoparasite, Nosema ceranae, appeared to be causing massive colony depopulations (Martn-Hernndez 2007). Sud

the cause of CCD! I followed up on this announcement with a series of articles on The Nosema Twinsall now pScientificBeekeeping.com. There has been a recent flurry of papers published onN. ceranae (several by the Centroproduced the largest body of research on the parasite), so I felt it appropriate to update my readers.

My tongue-in-cheek title forthis article refers to the diametrically opposed evaluations of the effects of the new noSpain. Papers published by Dr. Higes prolific team suggest that without antibiotic treatment, any colony that is infecdoomed. On the other hand, Antonio Pajuelo (Consultores Apcolas) and Jose Orantes-Bermejo (DirectorTcnico Ladamant that little if any correlation exists between N. ceranae infection and colony collapse!

This is more than a mere academic debate, as beekeepers worldwide are forced to make expensive management dantibiotic treatment and the sterilization of contaminated combs.

Here in the U.S., some beekeepers blame N. ceranae for taking down their colonies, and depend upon regular use

whereas others have yards of mildly infested colonies that thrive and produce great yields of honey (this is the case Beekeeperswhere we vigorously pursue the single- minded quest of exploring every conceivable way to lose mobeekeeper to think? Unfortunately, I cant yet answer that question, but I will try to digest and sort out the current s

The Nosema Twins are not twinsThey are actually more like cousins. Despite the titles of my previous series onN. ceranae, we have come to find threlated to N. vespula (from yellowjacket wasps) than it is to the old N. apis (Chen 2009). This finding has major co

we cant simply apply what we know about N. apis to N. ceranaeits a different critter.

In addition, there are different strains (haplotypes) ofN. ceranae, which may exhibit different degrees of virulence parasite, and can infect various hosts). And ifceranae is killing off colonies, then natural selection is certainly exertiarea towards the development of resistant bees.

Dr. Denis Anderson explained to me that Nosema [apis] follows a very much "temperature driven" pathology. Thainfected, they spread spores to neighboring bees in the winter cluster (forming 'pockets of infection' within the clustertoward the end of winter until they are completely eliminated in spring when the infected bees fly out and die. Gene

over summer, until autumn when there is a small peak, and again this is mostly temperature driven.

Nosema ceranae follows a different seasonality. Instead of spiking only in November and March like its cousin, it isthrives in summer. Colonies may struggle or collapse even during a spring or summer bloom.

The two nosemas both infect epithelial cells that line the bee gut, but the similarity ends there. Higes found thatN. c


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basal cells, and Dr. Chen found that it then apparently travels throughout the entire alimentary tract, including the hyglands. But only 20% of the fat bodies were infected, and no muscle tissue.

Raquel Martn-Hernndez (2009) found that N. ceranae infects bees held at 25C (77F) more quickly than does N. apisgrowth appears to be limited to a narrow temperature range. It grew at 25C (77F), which is a bit cool for a below side of brood nest temperature), but died off at 37C (98.6Ftypical of warmed bee flight muscles, or perhaps a

2000). N. ceranae, on the other hand, is able to survive at 37C.

The take home message is that N. ceranae is a somewhat more virulent parasite, and apparently more heat-adaptelonger count on the warmth of summer (or induced fever in the bees) to make nosema disappear.

Nosema apis is evidenced by the symptom of dysentery (above). An infection byN. ceranae is generally without symptomdisappear. Photos by the author.

Infection of QueensNosema apis was a nasty problem for queen bees, often causing early supersedure. From my reading of numerouparasite is most effectively transmitted from the attendants to the queen during the chilling and stress of shipment or(Lehnert 1973, Webster 2004). At that point, the queen can then become a Typhoid Maryinfecting the workers

are licked up by the attendant bees (Loskotova 1980).

Luckily, with N. ceranae we get a break! It doesnt appear to transmit readily to the queens, at least until the final spers comm). This is good news for queen producers, who may experience more trouble at eliminatingceranae than

Immune SuppressionNosema is tough on individual bees, not least of which because the infection inhibits their ability to digest food. Modigestive proteolytic activity in young bees infected with N. apis. Dr. Mariano Higes explains that bees infected by N


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in the midst of plenty due to lack of digestive function.

As if induced nutritional stress were not enough, N. ceranae also appears to suppress the bees immune functions. bees ramp up their immune systems in response to N. apis, but that system is apparently suppressed by N. ceranaeceranae depresses the level of the bee fountain of youth, vitellogenin, suggesting that infection may decrease thei

So the poor bees become both nutritionally stressed and immune suppressedlikely candidates for latent viruses to

add mite damage and miticide/pesticide stress. Since a nosema also broaches a bees main barrier to virus infectican suspect that well find N. ceranae/virus associations.

Just to make things more exciting, Santiago Plischuk (2009) has documented that N. ceranae has infected at least t

Argentina. Im not sure of the implications of this, but when a parasite hops between hosts, more virulent forms ca

Changes in BehaviorPerhaps the most noticeable effect ofN. ceranae infection is the lack of population buildup of infected colonies, due foragers. Of interest is that nosema-infected bees tend to forage at cooler temperatures. Woyciechowski (1998)engage in more risky foraging behavior, perhaps sensing that they do not have long to live. Additionally, infected bexhibiting an altruistic suicide to help prevent the infection of nestmates (Kralj 2006).

Another symptom, reported by several, and described by Bob Harrison on Bee-L, is that of bees not taking syrup, andivision board feeders. Bob feels that the symptom of going off feed is a good indicator forN. ceranae infection, wdrench of fumagillin syrup.

The drowning behavior may have an explanation in a recent paper by Chris Mayak (2009), in which he found that Nstress on infected bees, revealed in their elevated appetite and hunger level. infected bees attempt to compensatby feeding more Mayak suggests that such hungry, nutritionally stressed bees, exhibiting risky foraging behavidepopulation of infected colonies.

Development of Vegetative Stage/SporesMost nosema species in other insects infect the larvae. Neither of our nosema cousins do so. This is unusual, has a relatively short lifespan of only a few weeks during summer. That means that the parasite would be expecte

adult life, so that it can go through several generations before the bee dies, in order to multiply and produce an abun

The mode of transmission forN. apis is mainly by house bees ingesting spores when they clean soiled combs, andspores in stored pollen (Higes 2007). Both activities are performed mainly by very young house bees, so you wou

infected.

Curiously, although nosema can complete a reproductive cycle in just a few days, massive sporulation in young beehappen. Yefimenko (1994) found that inoculated caged bees did not develop substantial sporulation in the gut until 20 days old), despite the age that they were inoculated. Matthew Smart (working with Dr. Steve Sheppard, pers comarked cohorts of emerging bees in N. ceranae infected colonies.

This late development of spores should be apparent to all if one compares the average spore load of house bees tocolony. The mean spore counts for the older field bees will typically be 10-20 times higher than that of the house bfunction of behavior, than of chronology. Bees begin to age once they transition from house functions to foraging (swebsite).

So the question in my mind is whetherN. ceranae could multiply in the vegetative form in bees without producing spoverlooking actual infection by only looking for spores in the gut contents. This suspicion was bolstered when I spotechnician who works for Ken Olley in Australia. Zita previously studiedNosema bombycis, which infects silkworm l

a vegetative infection without producing spores.


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A partial answer is given by Raquel Martn-Hernndez (2009). She found that N. ceranae, in the early stages of infecells relative to spores than does N. apis. So a lack of spores in the gut does not necessarily mean that the bee is n

of an infection.

And how about Dr. Chens finding ofceranae DNA in the hypopharyngeal glands? Could the vegetative stage be pbetween workers? There are still a lot of unanswered questions about this parasite!

Transmission of SporesWhen a colony dies with N. apis, the combs are generally soiled with brown spotting and streaks from the spore-ladbees. This contamination has been clearly demonstrated to infect fresh bees placed back on the combs.

When a bee ingests nosema spores, its digestive fluids cause the spores to release the harpoon-like polar body. Ia bee, the harpoon must get past the protective peritrophic membrane, and penetrate an epithelial cell. Even thenovercome the innate and induced immune responses of the cell in order to reproduce.

Most spores are not successful. Bees (and humans) are constantly bombarded by spores (you breathe in plenty esick every day! Scientists use the term ID50the infectious dose necessary to infect 50% of the host population. has found that the ID50 for either nosema to be close to 100 spores. Fewer than that, and half the bees wont get

Once a spore does take hold, the infection can multiply rapidly. The infected cell can release new spores into the gspores may autoinfect the bee, rather than passing out of the gut. The infection can then spread throughout the (in the lab) dying on day 7 (from N. ceranae).

But not all spores are created equal. As with hardened plant seeds, or the cysts of brine shrimp, some organismseggssome which hatch quickly, and some that stay dormant for a time. Youssef (1971) found thatN. apis can psummer and winter. We have no idea as to whetherN. ceranae does the same.

With N. apis, it is pretty clear that transmission is via feceseither due to dysentery of infected bees confined during(Czekoska 2000). WithN. ceranae it was easy to assume that the same would apply. However, in beekeeping, conclusions.

There is no dysentery associated with a N. ceranae infection. So how do the spores get transmitted? Higes foun

hive, and demonstrated that it could be infective. But no one has clearly demonstrated the normal means of trans

This has important practical implications, since many beekeepers are assumingthat they should sterilize combs of other means) before putting them back to use. At the national conventions this year I asked around to see if any recombs to confirm that they were contaminated with N. ceranae spores. None had.

It occurred to me that it might be foolish to be going to all the expense and effort of comb sterilization based only upcombs were loaded with spores. So I invited Florida apiary inspector Dave Westerveld to visit myN. ceranae test yadetail our findings in the photos below.

A Quick and Dirty Field Search for Spores


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Dave Westerveld using the Suck-a-Bee to confirm the infection level ofN. ceranae. We tested combs frombeen infected with ceranae for at least two years at about 5 million spores in the foragers, and from a deadbees tested at 10M spores.


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Nancy Gentry (promoter of the Florida Honey Standard of Identity) helping out at our makeshift field la bor


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We found that a swab with hot water from a thermos quickly dissolved and lifted residues from the face of ththe cells.


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We swabbed, ground, crushed, and squeezed numerous samples in our search for spores.


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This is a typical view of comb surface residue at 400x magnification, which includes pollen coats, dirt, etc. contain any nosema spores whatsoever!


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The next day, Dr. Jerry Bromenshenk (sans white lab coat) stopped by to share i n the fun. We looked fohoney in the infected colonies. Again, they were few and far between.


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A typical view of diluted fresh beebread a mixture of pollen grains, but again, it was hard to find a single s

I reported our findings to Dr. Robert Cramer. He checked a comb that had been sent to him from anN. ceranae deaspores. So I asked him to send the comb to me so that I could confirm my ability to detect spores by swabbing. I dtechnique works), but not many. He didnt know the history of this comb, so Im not sure why there were more spotested.

Bottom line: Im not convinced that there are enough spores on combs of colonies infected with a moderate level ospores/bee) that a young bee would be likely to ingest the ID50 during the normal processes of comb cleaning and this hypothesis once I free up my incubator.

Sterilization ofCombs/Viability of SporesOK, so lets say that youre the cautious type, and still want to kill any N. ceranae spores on the combs of your deadradiation are options, and Dr. Cramer has found that bleach or lye also work.

However, Dr. Cramer reported some very interesting observations when I first supplied him with spores from my yarthat when he put the spores into the fridge over the weekend, that a lot of them appeared to die (become nonviable)great surprise, since N. apis spores are routinely frozen for storage, and spores in general survive better when chille

Dr. Cramer (unpublished data) analyzed groups of 10,000 spores. They started at about 87% viability. After onlydropped to 70%. After an hour in the freezer, viability dropped to 10-50%. At -80C (-112F) for an hour, generalsurvived. These are preliminary figures, and he is continuing this line of research.

At about the same time, Dr. Ingemar Fries and Eva Forsgren in Sweden also were having troubles at infecting beesrecently (2009) published some stunning similar results (by the way, Google translate does a great job going from S

superimposed English labels onto his graph below.


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Effect of chilling on the infectiveness of nosema spores. Unchilled spores of both species (bottom bars

1000-spore dose, and 100% with a 10,000 spore dose. The infectivity ofN ceranae spores dropped subsrefrigeration, and dramatically after a week of freezing. Modified from Fries (Bitidningen 107, 2009) by perm

Note: The above study has important implications to beekeepers. What is tells you is that in most of the Uenough to kill most of the spores ofN. ceranae!This supports the observation of a number of commercial beekeequipment rest for a month at cold temperatures resulted in better colonies when restocked in the spring.

When I and other researchers first heard of this observation, we scratched our heads, since in general, spores of diswhen stored at cold temperatures (there is normally more degradation at higher temperatures). None of us would

ceranae spores would be so intolerant of cold storage!

A very interesting additional observation by Fries was that N. apis spores actually appear to become even more infefreezing! Note in the graph above how the 1000-spore inoculum ofN. apis infected 100% percent of the bees after

percentage than with unchilled or refrigerated spores! Dr. Fries is planning further investigation into this surprising

The susceptibility ofN. ceranae spores to freezing may help to explain why it appears to be more of a problem in wabe lost on those Midwestern beekeepers who take colonies to California for almond pollination any deadou

experience the sterilizing effect of cold winter temperatures.

So what if you live in a warm climate? Youre still in luck, sinceceranae spores appear to be rather fragile. Dr. Cramquickly to elevated temperatures, to wit, 120F for 90 minutes. Cramer hopes to develop a time/temperature curve

beekeeper can determine how much time at what temperature it takes to kill most of the spores.


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Thresholds forTreatmentA recent paper by Martn-Hernndez (2009) found that both N. ceranae and N. apis can complete their life cycles (sgut cells, and that N. ceranae initially produced a larger proportion of vegetative cells relative to spores than did N. a

implications as to the reliability of using spore counts alone as a means of determining how badly a bee (or a colonysuggests, with scientific terseness, that spore counts are unreliable as a tool to evaluate the health status of inthere is no easy or cheap alternative available to the beekeeper. Again, I caution the beekeeper about putting too

especially when taken from either inside the colony, or from samples of only a few bees.

The main point to clarify is that there is a difference between being infected by a parasite, and suffering from diseasea mean spore count (of foragers) of 5M, there are only a very small proportion of the total population of bees in thedegree. The true concern is when the infection level grows to include a substantial proportion of house bees, at whiccollapse. Sampling of field bees will indicate if the colony is infected. Sampling of house bees may be theinfection is serious.

Perhaps the simplest way to evaluate the effect ofN. ceranae on your bees is to monitor how well your colonies arediscounting the lack of interest in syrup). Ifceranae is indeed killing off a noticeable proportion of field bees, you willbuild its population (although Higes describes a false recovery by which the colony compensates by massive broo

In my own test yard, the only colonies that I find that appear to suffer when infected with N. ceranae are those that athe brood. There appears to be a (new?) brood disease that has appeared in the past three years, in which th(cells filled with brood of mixed ages, rather than a nice pattern), the dying larvae turn yellowish, and die wispoke yesterday with Dr. Jeff Pettis--he and Dennis vanEnglesdorp are also seeing it on the East Coast. Theyve bof snotty brood. In my own test yards, the disease does not appear to spread easily from colony to colony, and I antibiotic mayhelp, and that bees can be successfully restocked on to deadouts without treatment. Im eager for D

Wick to figure out how to get brood samples to run through the IVDS machine, to see if they can identify the pathog

Most researchers in various countries, other than the Higes team in Spain, do not see a strong correlation betweenwinter losses. Nosema infection of any sort is clearly a stressor to the colony, and an invitation for viral infections. diseases, colonies do their best in conditions of good forage and weather. If your yards are crowded or the naturalthere are mite or other disease issues, then it would be prudent to be more concerned aboutN. ceranae.

I was recently speaking with a large commercial beekeeper from a warm climate who found that at spore counts of anot build. However, another very large operator from Southern California does not even worry about counts in theCalifornia foothills, counts of 5M do not appear to affect colony production or survivability.

To monitorN. ceranae levels, be sure to collect bees from the entrance. A number of beekeepers have built Suck-website). The feedback is that the 16V model is better, since it holds much more charge, and you dont need a sepavacuum, you can brush bees off the landing boards into a widemouth jar containing rubbing alcohol (a small piece omake it easierscreen works better than a block of wood). Be sure to collect at least 50 bees in every samplew

from several colonies in each yard all into the same jar as yard samples, and take them home in labeled jars.


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If you dont have a bee vacuum, you can simply sweep bees off the landing board into a wide mouthed jar oudge the mite level of a yard by sampling a single colony, in order to determine the yard average for nosem

bees from as many colonies as possible. Be aware that sweeping the entrance really sets the bees off

Regarding microscopes, over the past year, Ive had the opportunity to view nosema spores through a number of m

expensive! Ive yet to see a scope that gives a clearer image of spores than the Omano, which I still feel is the best b

TreatmentsWere still struggling to determine the most effective and appropriate treatments forN. ceranae. Large-scale field triPernal have been bedeviled by the spontaneous disappearance ofN. ceranae from the control groups. This inconveone to call into question any beekeepers reports that any certain treatment was effective, if they didnt run untreated

treated colonies.

Fumagillin often works, but not always. It appears to me that one may need to increase the recommended dose return a few months after treatment.

Researchers and beekeepers are still experimenting with alternative treatments. The strong cup o soup (8 oz ofin 7 gal of syrup) gets good feedback. Nosevit appears to work, if applied precisely by label instructions (1/4 tsp these treatments indicate that giving the dose of active ingredient in cups, rather than gallons, of syrup (other than fo

effective.

HoneyBHealthy appears to be somewhat beneficial. I dont have enough data on the other herbal products, such a

ApiHerb. And Im still curious to see if I can get thymol to have an effect.

We dont know why N. ceranae appears to displace N. apis. If a bee is inoculated with both species, both grow (Frie


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to be an inhibitory competition. However, treatment with fumagillin may favorN. ceranae.

I caution against treating your bees unnecessarily! Not only can treatments be expensive, but some, if not all, are

Bottom Line

1.Nosema ceranae is different than Nosema apis, and exhibits different modes of infection, virulence, e

2.N. ceranae appears to be more of a problem in some areas and operations than others. It may be moareas. Fortunately, in some areas, the spore counts never progress above the 4 -8M range.

3.It is likely a waste of money to treat if you havent confirmed that your bees are indeed infected.

4.N. ceranae appears to only infect queens shortly before the collapse of the colony.

5.Initial signs of infection may be lack of colony growth, due to early death of foragers and nutritional sfeed. In the later stages, house bees become infected, bees may drown en mass in division board fesuddenly collapse.

6.Were not really clear as to howN. ceranae is transmitted from bee to bee.

7.Were not really clear about the best treatments, or application the reof, or if treatment in general is eeffective!

8.Luckily, N. ceranae spores are more delicate than those ofN. apisthey are readily killed by exposutemperatures.

9.This last point is of major practical applicati onit may be unnecessary to attempt to sterilize comcold (orwarm) temperatures.

AcknowledgementsI greatly appreciate the time given to me by researchers Mariano Higes, Antonio Pajuelo, Jose Orantes-Bermejo, JefForsgren. Thanks also to commercial beekeeper Bob Harrison, and especially to Peter Borst for his invaluable as

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