egg count

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Light, long-lasting Necator infection in a volunteer.

School of Public Health, Tulane University, New Orleans, Louisiana 70112.

“A Necator infection produced by percutaneous exposure of a volunteer to three larvae was followed with periodic egg counts for 4 months beyond 18 years when passage of eggs in the feces ceased permanently. During the 2nd year of infection, there was unintentional exposure to two additional larvae. Egg counts per gram of feces (EPG) were approximately 1,000 during the 1st year, 1,500 over the next 5 years, and less than or equal to 200 over the final 3 years, during which time there were 6 periods of approximately 1 month each when Kato thick-smear examinations revealed no eggs. During 6 of the 1st 14 years, when egg counts were done by the standard direct smear method, up to 37% of the eggs were infertile. Based on the assumption that two female worms were present during the 1st year, three during the next 3 years or more, and only one during the final 3 years, the estimated output per female in this light infection was 500 EPG in the worm’s prime of life, and less than 200 EPG in the final year of the 17 or 18 year life span.”

From: http://www.ncbi.nlm.nih.gov/pubmed/3189697

This study is quite amazing in that such a small infection could last so long. (I’d like to know how he was accidentally exposed to 2 larvae, and how did he know they were two? Was he fooling around under the microscope and his hand slipped and they splashed onto his arm? I wish they provided details!) Also, that anyone would do fecal egg counts for 18 years, and monthly! If such a high EPG is found from only a few females, then why are we messing around with doses of 25-50? I’m still probably going to dose with 10, I just want to do my egg counts to see what happens. And if they all die, I’ll think of this man, who hosted 5 for two decades. Clearly, the modern lifestyle didn’t affect his hookworm status. Amazing creatures!

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Dosing

I was thinking about worm dose and efficacy. (Brace yourself; this is
long and convoluted.) If one were to be dosed once with 10 larvae,
then accounting for attrition, there would be perhaps 7 left in the
gut. (And this number may be a high estimate.) Assuming a 50% gender
split, there would be 3 or 4 males. (And there could be 2, 1.) This
Cornell transcript claims the males die off after copulation.
(http://www.cumc.columbia.edu/dept/ps/2007/para/old/transcript_02_pd02.pdf)
Even if they don’t, this paper claims the lifespan, though averaging
3-5 years, is really much shorter than that.
(http://www.jimmunol.org/cgi/content/full/173/4/2699) Would 3 or 4
worms be enough to maintain a steady immunological state? How many
months/years would they live and would there be an average drop off
around the same time? (meaning; do they all die at once?) As
Nottingham is only studying their Crohn’s patients for 6 months, they
will not collect this data, which is unfortunate. (length of
efficacy, worm lifespan, wormcount after x number of months/years,
timing of necessary reinfection, etc.) So we’re on our own…

This study says that adding a population of 50 in two healthy people
who hosted existing hookworms resulted in some old worms dying and new
ones replacing them, with no netted increase in number.
(http://www.ncbi.nlm.nih.gov/pubmed/17035088) So for these two
people, adding worms at a high, one time dose, didn’t result in a
greater population. What the paper doesn’t address is age of the
worm. How many new worms replaced the old? Would one end up with a
mixed age population? That should extend efficacy, assuming the new
ones live longer than the first cohorts. (Some worms would be old
crones…surrounded by fresh whippersnappers.)

It seems like the only choice to maintain a healthy, thriving
population is to redose periodically, with a small enough number not
to illicit enough of an allergic response to expel the new or old
population. What is the number before adding becomes displacing? If
one started with 10, was left with 5, added 15, they merely displaced,
then one is left with 5 again. Would adding 2 be more effective?

And then I wonder how much of the acquisition of new larvae is part of
the immunological puzzle. If there is an additional, temporary
benefit from each subsequent infection, ( the bounce, or the “hookworm
high”) then perhaps maintaining both the immunological and
psychological health of the human and an effective worm burden is
based on the constant periodic addition of small numbers of worms.

Which means the patients who get one dose only are incomparable to the
ones who get 3 doses, who are incomparable to Jasper, for example,
who’s had multiple tiny doses. Perhaps his efficacy would have
waned if a small, steady population wasn’t supplied?

If there is a psychological component to small reinfections, along
with a temporary immunological benefit, perhaps man evolved to have
this periodically. Sort of a natural antidepressant to go with the
immune regulation. This study finds antidepressant qualities in the
soil bacteria itself
(http://www.medicalnewstoday.com/articles/66840.php), showing even
greater depth to the whole hygiene hypothesis. Depression as a
consequence of our sterility. We may need not only the worms, but the
soil they came from.

Which means trying to recreate the natural state by supplying one
aspect of it; the worms, in an unnatural dose (one time, a great
number) and removing everything else may not work. What is the
natural number and frequency of infection in the wild for a healthy
adult anyway?

This really argues against a one time inoculation, at least based on
the theory that the worms’ benefit will last as long as they are
alive. Yet it also argues against multiple inoculation therapy, as
that might cause one to end up with the same amount as one began with,
or less. It seems the only way to recreate the natural state is to
add 1 or 2 worms at a time.

There are no answers to any of these questions yet…Will we die
before we learn how to properly live?

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So I took a standard stool test through Quest lab, and all 3 samples came back negative for ova. I did use one stool for all three samples, rather than 3 different days. One has to wonder, are light infections producing enough eggs for the standard test to find them? In the Nottingham dose ranging study, there were a few weeks where no eggs were found:

“Egg counts for individual subjects were variable and two subjects had one or two weeks in which eggs were not seen, having previously been detected.”

Those that got 50 worms had higher counts:

“The highest egg counts occurred in the people who received 50 larvae; median egg counts were similar in participants allocated to the two lower doses.”

I got a total of 37 larvae; but only 10 were given in the first dose. Then, by 2′s and 3′s, and one has to wonder if any of those attached. I’m having AIT incubate a sample for me to see if anything grows. I should know tomorrow.

IF I still have worms, which by my symptoms I would guess I have few, then the question becomes how to raise the population. If I have adequate worms, then I suppose I wait and hope I’m like the video of the guy with psoriasis, who had 60 worms, got better, relapsed, then waited, and got better after 11 months or so. I don’t know though, regression happened when I added worms. It really points to a lower population.

I guess I’ll know soon enough. Then I’ll have to figure out what to do…

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So I’ve had a very eventful week calling labs across California to see if they’d do a quantitative egg count for me. The answer from all? No.

“Why do you want to do an egg count?”
“I’m trying to determine my aproximate worm burden for an experimental
hookworm infection.”
“How do you do an egg count anyway?”
“I was hoping you knew. From what I’ve read online, one measures eggs
per gram of feces, using a grid slide.”
“But I take a small amount of that stool and look under a microscope.
We can tell you light infection, heavy infection, but cannot count.”

Or will not?

And so on.

My favorite was calling UCDavis and being transferred first to the
rodent division, who were very nice, but said I needed the large
animal division. At this point I was becoming desperate and asked if
I could just be a large animal, though I think the dog vets would be
more familiar with hookworm egg counts. No one would help me.

Finally, I found this:

http://www.docstoc.com/docs/599020/Macmaster-counting-Technique-pp

http://cal.vet.upenn.edu/projects/parasit06/website/lab1.htm#techniques

Nottingham’s dose-ranging study used the Macmaster counting technique. It seems rather simple. You mix a small amount of measured feces with salt water and a tongue dispenser, then pass it through a sieve. Then suck up a measured quantitity, and put it on a grid slide. Then, going slowly up and down the grid, you count the visible eggs, and multiply it by the correct number to get your eggs per gram of feces. If we all learned to do this accurately, we could monitor our egg output post infection, and watch it through the months/years to track a decline in adult hookworm population.

What I’d like to know is how one cleans the lab equipment without the lab. Somehow I’m not that thrilled. I’d rather bake a cake.

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