Nosema ceranae:

Kiss of Death or Much Ado about Nothing?

Randy Oliver

 
Once again I’m interrupting a series of articles to update beekeepers on breaking new
applications in bee management.  This time the information is about Nosema ceranae
readers of the Journal might benefit from an update on recent research results.

 
Dr. Mariano Higes’ team (from the Centro Apicola Regional in Spain) electrified the beekeeping world two years ago wit
parasite, Nosema ceranae, appeared to be causing massive colony depopulations (Martín-Hernández 2007).  Suddenly
the cause of CCD!  I followed up on this announcement with a series of articles on “The Nosema Twins”—all now poste
ScientificBeekeeping.com.  There has been a recent flurry of papers published on N. ceranae (several by the Centro Ap
produced the largest body of research on the parasite), so I felt it appropriate to update my readers.

 My tongue-in-cheek title for this article refers to the diametrically opposed evaluations of the effects of the “new” nosem
Spain. Papers published by Dr. Higes’ prolific team suggest that without antibiotic treatment, any colony that is infected
doomed. On the other hand, Antonio Pajuelo (Consultores Apícolas) and Jose Orantes-Bermejo (Director Técnico Labo
adamant 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 decis
antibiotic 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 of fu
whereas others have yards of mildly infested colonies that thrive and produce great yields of honey (this is the case in m
Beekeepers”—where we vigorously pursue the single- minded quest of exploring every conceivable way to lose money
beekeeper to think?  Unfortunately, I can’t yet answer that question, but I will try to digest and sort out the current state

The “Nosema Twins” are not twins

They are actually more like cousins.  Despite the titles of my previous series on N. ceranae,  we have come to find that 
related to N. vespula (from yellowjacket wasps) than it is to the “old” N. apis (Chen 2009).   This finding has major cons
we can’t simply apply what we know about N. apis to N. ceranae—it’s a different critter.

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

Dr. Denis Anderson explained to me that “Nosema [apis] follows a very much "temperature driven" pathology.  That is,
infected, they spread spores to neighboring bees in the winter cluster (forming 'pockets of infection' within the cluster).  
toward the end of winter until they are completely eliminated in spring when the infected bees fly out and die.  Generally
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 is pre
thrives 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 that N. cera
cells, and Dr. Chen found that it then apparently travels throughout the entire alimentary tract, including the hypopharyn
only 20% of the fat bodies were infected, and no muscle tissue.

Raquel Martín-Hernández (2009) found that N. ceranae infects bees held at 25°C (77°F) more quickly than does N. api
apisgrowth appears to be limited to a narrow temperature range.  It grew at 25°C (77°F), which is a bit cool for a bee, th
low side of brood nest temperature), but died off at 37°C (98.6F—typical of warmed bee flight muscles, or perhaps a bro
2000).  N. ceranae, on the other hand, is able to survive at 37°C.

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

 Nosema apis is a c
evidenced by the symptom of dysentery (above).  An infection by N. ceranae is generally without symptoms—
“disappear.”  Photos by the author.

Infection of Queens
Nosema apis  was a nasty problem for queen bees, often causing early supersedure.  From my reading of numerous stu
parasite is most effectively transmitted from the attendants to the queen during the chilling and stress of shipment or ho
(Lehnert 1973, Webster 2004).  At that point, the queen can then become a “Typhoid Mary”—infecting the workers via t
are licked up by the attendant bees (Loskotova 1980).

Luckily, with N. ceranae we get a break!  It doesn’t appear to transmit readily to the queens, at least until the final stage
pers comm).  This is good news for queen producers, who may experience more trouble at eliminating ceranae than th

Immune Suppression
Nosema is tough on individual bees, not least of which because the infection inhibits their ability to digest food.  Molone
digestive proteolytic activity in young bees infected with N. apis.   Dr. Mariano Higes explains that bees infected by N. ce
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.  Kar
bees ramp up their immune systems in response to N. apis, but that system is apparently suppressed by N. ceranae.  In
ceranae depresses the level of the bee “fountain of youth,” vitellogenin, suggesting that infection may decrease their life

So the poor bees become both nutritionally stressed and immune suppressed—likely candidates for latent viruses to ex
add mite damage and miticide/pesticide stress.  Since a nosema also broaches a bee’s main barrier to virus infection—
can suspect that we’ll find N. ceranae/virus associations.

Just to make things more exciting, Santiago Plischuk (2009) has documented that N. ceranae has infected at least thre
Argentina.  I’m not sure of the implications of this, but when a parasite hops between hosts, more virulent forms can em

Changes in Behavior
Perhaps the most noticeable effect of N. ceranae infection is the lack of population buildup of infected colonies, due to t
foragers.  Of interest is that nosema-infected bees tend to forage at cooler temperatures.  Woyciechowski (1998) sugge
engage in more risky foraging behavior, perhaps sensing that they do not have long to live.  Additionally, infected bees
exhibiting 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, and th
division board feeders.  Bob feels that the symptom of going “off feed” is a good indicator for N. ceranae infection, whic
drench of fumagillin syrup. 

The drowning behavior may have an explanation in a recent paper by Chris Mayak (2009), in which he found that “N. ce
stress on infected bees, revealed in their elevated appetite and hunger level…. infected bees attempt to compensate fo
by feeding more…”  Mayak suggests that such hungry, nutritionally stressed bees, exhibiting risky foraging behavior, m
depopulation of infected colonies.

Development of Vegetative Stage/Spores

Most nosema species in other insects infect the larvae.  Neither of our nosema “cousins” do so.  This is unusual, espec
has a relatively short lifespan of only a few weeks during summer.  That means that the parasite would be expected to t
adult life, so that it can go through several generations before the bee dies, in order to multiply and produce an abundan

The mode of transmission for N. apis  is mainly by house bees ingesting spores when they clean “soiled” combs, and pe
spores in stored pollen (Higes 2007).  Both activities are performed mainly by very young house bees, so you would exp
infected.

Curiously, although nosema can complete a reproductive cycle in just a few days, massive sporulation in young bees do
happen. Yefimenko (1994) found that inoculated caged bees did not develop substantial sporulation in the gut until they
days old), despite the age that they were inoculated.  Matthew Smart (working with Dr. Steve Sheppard, pers comm) ob
marked 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 to the
colony.  The mean spore counts for the older field bees will typically be 10-20 times higher than that of the house bees.
function of behavior, than of chronology.  Bees begin to age once they transition from house functions to foraging (see “
website).

So the question in my mind is whether N. ceranae could multiply in the vegetative form in bees without producing spore
overlooking actual infection by only looking for spores in the gut contents.  This suspicion was bolstered when I spoke w
technician who works for Ken Olley in Australia.  Zita previously studied Nosema bombycis, which infects silkworm larva
a vegetative infection without producing spores.

A partial answer is given by Raquel Martín-Hernández (2009). She found that N. ceranae, in the early stages of infectio
cells relative to spores than does N. apis.  So a lack of spores in the gut does not necessarily mean that the bee is not s
of an infection.

And how about Dr. Chen’s finding of ceranae DNA in the hypopharyngeal glands?  Could the vegetative stage be pass
between workers?  There are still a lot of unanswered questions about this parasite!

Transmission of Spores
When a colony dies with N. apis, the combs are generally soiled with brown spotting and streaks from the spore-laden d
bees. 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.  In ord
a bee, the harpoon must get past the protective peritrophic membrane, and penetrate an epithelial cell.  Even then, the
overcome 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 each d
sick every day! Scientists use the term “ID50”—the infectious dose necessary to infect 50% of the host population.  Dr.
found that the ID50 for either nosema to be close to 100 spores.  Fewer than that, and half the bees won’t get infected.

Once a spore does take hold, the infection can multiply rapidly.  The infected cell can release new spores into the gut in
spores may “autoinfect” the bee, rather than passing out of the gut.  The infection can then spread throughout the epithe
(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 organisms can
“eggs”—some which hatch quickly, and some that stay dormant for a time.  Youssef (1971) found that N. apis  can prod
summer and winter.  We have no idea as to whether N. ceranae does the same.

With N. apis, it is pretty clear that transmission is via feces—either due to dysentery of infected bees confined during wi
(Czekońska 2000).  With N. ceranae it was easy to assume that the same would apply.  However, in beekeeping, assu
conclusions.

There is no dysentery associated with a N. ceranae infection.  So how do the spores get transmitted?  Higes found spo
and demonstrated that it could be infective.  But no one has clearly demonstrated the normal means of transmission.

This has important practical implications, since many beekeepers are assuming that they should sterilize combs of dea
other means) before putting them back to use.  At the national conventions this year I asked around to see if any resear
combs 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 upon
combs were loaded with spores.  So I invited Florida apiary inspector Dave Westerveld to visit my N. ceranae test yard
our findings in the photos below.

A Quick and Dirty Field Search for Spores

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

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

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

 
We swabbed, ground, crushed, and squeezed numerous samples in our search for spores.
This is a typical view of comb surface residue at 400x magnification, which includes pollen coats, dirt, etc.  B
contain any nosema spores whatsoever!
  

The next day, Dr. Jerry Bromenshenk (sans white lab coat) stopped by to share in the fun.  We looked for sp
in the infected colonies.  Again, they were few and far between.
A typical view of diluted fresh beebread—a mixture of pollen grains, but again, it was hard to find a single sp

I reported our findings to Dr. Robert Cramer.  He checked a comb that had been sent to him from an N. ceranae  deado
spores. So I asked him to send the comb to me so that I could confirm my ability to detect spores by swabbing.  I did ind
technique works), but not many.  He didn’t know the history of this comb, so I’m not sure why there were more spores th
tested.

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

Sterilization of Combs/Viability of Spores

OK, so let’s say that you’re the cautious type, and still want to kill any N. ceranae spores on the combs of your deadout
radiation 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 yard ba
that when he put the spores into the ‘fridge over the weekend, that a lot of them appeared to die (become nonviable) ov
great surprise, since N. apis spores are routinely frozen for storage, and spores in general survive better when chilled.

Dr. Cramer (unpublished data) analyzed groups of 10,000 spores.  They started at about 87% viability.  After only an ho
dropped to 70%.  After an hour in the freezer, viability dropped to 10-50%.  At -80ºC (-112ºF) for an hour, generally only
survived.  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 bees with
recently (2009) published some stunning similar results (by the way, Google translate does a great job going from Swed
superimposed English labels onto his graph below.
 

Effect of chilling on the infectiveness of nosema spores.  Unchilled spores of both species (bottom bars) infe
1000-spore dose, and 100% with a 10,000 spore dose.  The infectivity of N ceranae spores dropped substan
refrigeration, and dramatically after a week of freezing.  Modified from Fries (Bitidningen 107, 2009) by permis

Note:  The above study has important implications to beekeepers.  What is tells you is that in most of the U.S.,
enough to kill most of the spores of N. ceranae!   This supports the observation of a number of commercial beekeep
equipment “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 diseas
when stored at cold temperatures (there is normally more degradation at higher temperatures).  None of us would have
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 infectiv
freezing!  Note in the graph above how the 1000-spore inoculum of N. apis infected 100% percent of the bees after hav
percentage than with unchilled or refrigerated spores!  Dr. Fries is planning further investigation into this surprising phen

The susceptibility of N. ceranae spores to freezing may help to explain why it appears to be more of a problem in warm
be lost on those Midwestern beekeepers who take colonies to California for almond pollination—any deadout e
experience the “sterilizing” effect of cold winter temperatures.

So what if you live in a warm climate?  You’re still in luck, since ceranae spores appear to be rather fragile. Dr. Cramer
quickly to elevated temperatures, to wit, 120°F for 90 minutes.  Cramer hopes to develop a time/temperature curve of s
beekeeper can determine how much time at what temperature it takes to kill most of the spores.

Thresholds for Treatment

A recent paper by Martín-Hernández (2009) found that both N. ceranae and  N. apis can complete their life cycles (spor
gut cells, and that N. ceranae  initially produced a larger proportion of vegetative cells relative to spores than did N. apis
implications as to the reliability of using spore counts alone as a means of determining how badly a bee (or a colony) is
suggests, with scientific terseness, that spore counts are “unreliable as a tool to evaluate the health status of infec
there is no easy or cheap alternative available to the beekeeper.  Again, I caution the beekeeper about putting too much
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 disease ca
a mean spore count (of foragers) of 5M, there are only a very small proportion of the total population of  bees in the colo
degree. The true concern is when the infection level grows to include a substantial proportion of house bees, at which p
collapse. Sampling of field bees will indicate if the colony is infected.  Sampling of house bees may be the best
infection is serious.

Perhaps the simplest way to evaluate the effect of N. ceranae on your bees is to monitor how well your colonies are bui
discounting the lack of interest in syrup).  If ceranae  is indeed killing off a noticeable proportion of field bees, you will no
build its population (although Higes describes a “false recovery” by which the colony compensates by massive broodrea

In my own test yard, the only colonies that I find that appear to suffer when infected with N. ceranae are those that also
the brood.  There appears to be a (new?) brood disease that has appeared in the past three years, in which the b
(cells filled with brood of mixed ages, rather than a nice pattern),  the dying larvae turn yellowish, and die with “
spoke yesterday with Dr. Jeff Pettis--he and Dennis vanEnglesdorp are also seeing it on the East Coast.  They’ve besto
of “snotty brood.”  In my own test yards, the disease does not appear to spread easily from colony to colony, and I find t
antibiotic may help, and that bees can be successfully restocked on to deadouts without treatment.  I’m eager for Dr. Je
Wick to figure out how to get brood samples to run through the IVDS machine, to see if they can identify the pathogen(s

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

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

To monitor N. ceranae levels, be sure to collect bees from the entrance.  A number of beekeepers have built Suck-a-Be
website). The feedback is that the 16V model is better, since it holds much more charge, and you don’t need a separate
vacuum, you can brush bees off the landing boards into a widemouth jar containing rubbing alcohol (a small piece of sc
make it easier—screen works better than a block of wood).  Be sure to collect at least 50 bees in every sample—we suc
several colonies in each yard all into the same jar as “yard samples,” and take them home in labeled jars.
If you don’t have a bee vacuum, you can simply sweep bees off the landing board into a wide mouthed jar of
judge the mite level of a yard by sampling a single colony, in order to determine the yard average for nosema
bees from as many colonies as possible.  Be aware that sweeping the entrance really sets the bees off—glo

Regarding microscopes, over the past year, I’ve had the opportunity to view nosema spores through a number of micro
expensive! I’ve yet to see a scope that gives a clearer image of spores than the Omano, which I still feel is the best buy

Treatments
We’re still struggling to determine the most effective and appropriate treatments for N. ceranae.  Large-scale field trials
Pernal have been bedeviled by the spontaneous disappearance of N. ceranae from the control groups. This inconvenie
one to call into question any beekeepers’ reports that any certain treatment was effective, if they didn’t run untreated co
treated colonies.

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

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

HoneyBHealthy appears to be somewhat beneficial.  I don’t have enough data on the other herbal products, such as Vit
ApiHerb. And I’m still curious to see if I can get thymol to have an effect.

We don’t know why N. ceranae  appears to displace N. apis.   If a bee is inoculated with both species, both grow (Fries 2
to be an inhibitory competition.  However, treatment with fumagillin may favor N. ceranae.

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

Bottom Line
 

1. Nosema ceranae is different than Nosema apis, and exhibits different modes of infection, virulence, epi
2. N. ceranae appears to be more of a problem in some areas and operations than others.  It may be more
areas. 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 haven’t 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 stre
feed.” In the later stages, house bees become infected, bees may drown en mass in division board feed
suddenly collapse.
6. We’re not really clear as to how N. ceranae is transmitted from bee to bee.
7. We’re not really clear about the best treatments, or application thereof, or if treatment in general is even
8. Luckily, N. ceranae spores are more delicate than those of N. apis—they are readily killed by exposure t
temperatures.
9. This last point is of major practical application—it may be unnecessary to attempt to “sterilize” combs i
cold (or warm) temperatures.

Acknowledgements

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

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