CASE CLOSED … what really happened in the 2001 anthrax attacks?

* FBI now admits the silicon concentration in the attack anthrax is too high … higher than can be explained by natural occurrence.

Posted by DXer on December 7, 2010

Microbial Forensics, Second Edition

Bruce Budowle (Editor), Steven E. Schutzer (Editor), Roger G. Breeze (Editor), Paul S. Keim (Editor), Stephen A. Morse (Editor)

The second edition of the treatise has just been released

and contains some startling admissions.

The silicon concentration of the Amerithrax spores, long downplayed by the FBI as a “natural occurrence” has finally been discussed in detail.

The FBI scientists finally admit that the concentration of the element silicon found in the attack spores is too high – higher than can be explained by natural occurrence.

Interestingly, the high silicon concentration was discussed in a Wall Street Journal aricle by Ed Epstein:

https://caseclosedbylewweinstein.wordpress.com/2010/01/25/epstein-wsj-1-24-10-the-fbi-says-ivins-was-the-sole-perpetrator-but-it-has-presented-no-evidence-to-support-that-conclusion-and-the-largest-case-in-fbi-history-is-still-open/

Ed Epstein’s article was disputed days later by FBI lab director Chris Hassell (although he gave no real explanation ahy he was disputing it):

https://caseclosedbylewweinstein.wordpress.com/2010/02/04/fbi-disputes-epsteins-wsj-opinion-piece/

Now things seem to have taken a complete 180 and the new book Microbial Forensics, basically agrees with Ed Epstein’s original article – which stated that the silicon concentration was too high and was not a “natural occurence”.

The book even goes as far as to state that

the high silicon content would have provided a serious problem

had any case against Dr Ivins have gone to court –

it would have affected the “prosecution narrative”.

Quote (from page 513):

“Thus if the estimates silicon concentrations in the Amerithrax spores are correct, they are not consistent with our current understanding of silica deposition or those materials must have indeed been produced under an unusual set of conditions. If the latter were true, the silica evidence might provide a significant bound on the credible growth and production scenarios that would be consistent with the prosecution narrative in this case.

  • Although this is a good step forward by the FBI, they appear to be still downplaying the NYP silicon concentration.
  • The AFIP lab report is still not mentioned nor is the >30% silicon content that is found in that sample.

******

LMW COMMENT …

The FBI’s case against Dr. Ivins is clearly bogus: no evidence, no witnesses, an impossible timeline, science that proves innocence instead of guilt. So what really happened? And why doesn’t the FBI offer America a credible story?

I can imagine only 3 possible “actual” scenarios …

  1. The FBI has more evidence against Dr. Ivins but is, for some undisclosed reason, withholding that evidence … POSSIBLE BUT NOT SO LIKELY
  2. The FBI, despite the most expensive and extensive investigation in its history, has not solved the case and has no idea who prepared and mailed the anthrax letters that killed 5 Americans in 2001 … EVEN LESS LIKELY
  3. The FBI knows who did it (not Dr. Ivins) but is covering up the actual perpetrators, for undisclosed reasons … THE MOST LIKELY SCENARIO

The “fictional” scenario in my novel CASE CLOSED has been judged by many readers, including a highly respected official in the U.S. Intelligence Community, as perhaps more plausible than the FBI’s unproven assertions regarding Dr. Ivins.

* buy CASE CLOSED at amazon *

******

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39 Responses to “* FBI now admits the silicon concentration in the attack anthrax is too high … higher than can be explained by natural occurrence.”

  1. DXer said

    Flask 1030 had a Silicon Signature. In 1997, a sample from Flask 1030 was shipped to Dugway. It was to serve as a standard in aerosol studies.

    “On February 2, 2005, BRUCE IVINS left a voice mail message for Supervisory Special Agent (SSA) _______… as a follow-up to a matter discussed in an interview of IVINS on January 13, 2005. IVINS provided the following information:

    After reviewing the shipping form for the Bacillus anthracis Ames strains samples he sent to Dugway Proving Ground in 1997, IVINS believes the tubes he sent were samples of RMR 1030, a mixed batch of spores prepared by IVINS and ______________________. The reason he believes this to be true is that the shipping form indicates the samples were spores in liquid form, unfrozen. The only Ames spores IVINS had unfrozen in liquid form in his laboratory at that time were the spores known as RMR 1030. Also, he compared the concentration of spores in the shipment to the concentration of samples he had in his laboratory at the time of the shipment. The documented concentration for RMR 1030 closely matched that on the shipping form for the spores sent as seed stock to Dugway for the mass production of spores to be used in aerosol challenge studies.”

    • DXer said

      So Flask 1030, with the Silicon Signature, was sent to Dugway as seed stock to Dugway for the mass production of spores to be used in aerosol challenge studies, before the former Zawahiri associate went there for aerosol studies.

      • DXer said

        2/22/2005

        BRUCE IVINS was interviewed at his place of employment. …

        IVINS advised the printout of the electronic USAMRIID foreign visiting scientist which listed ___________ as a Point of Contact (POC) was errant. IVINS further advised he was the USAMRIID POC for an Egyptian male, [Tarek Hamouda] from the University of Michigan circa ________________. IVINS advised the electronic USAMRIID foreign visiting scientist record had mispelled ________ as ________________________.

        IVINS advised prior to May 1998 he was contacted by ______________________ [James Baker] from the University of Michigan Medical Center who was ______________________ of a ______________________________________________. IVINS advised _________ wished to collaborate with IVINS in order to test the effectiveness of a new anti-sporicidal material against anthrax spores. IVINS further advised ____________ [Baker] had numerous visits to USAMRIID, however “never accessed the B3 suite.”

        IVINS advised [Baker] SENT [Tarek Hamouda and Michael Hayes] , both from the University Michigan, to conduct experiments. IVINS advised [HAMOUDA and HAYES] underwent safety traning and provided documentation of their shot records. IVINS advised when HAMOUDA showed up at USAMRIID, USAMRIID personnel realized HAMOUDA was not a U.S. citizen. IVINS advised, during the May 1999 time period, a email request for approval was all that was required for “green card holders” to visit USAMRIID. IVINS further interjected and advised “the request for _______ to visit USAMRIID did not come from command it came from [IVINS].” __________ further advised “this is different from which I had previously told [the interviewing Postal Inspector and SA].”

        IVINS advised _________ had contacted IVINS and ______________ were come to USAMRIID and conduct the research on the collaboration project.

        IVINS advised [HAMOUDA and HAYES] ______________ worked with the Ames strain of BL-3 laboratory” for three or four days in May 1998. IVINS further advised HAMOUDS and HAYES were never left alone in the BL-3 laboratory and either IVINS or __________________ were with [them].

        IVINS advised he does not recall whether or not ___________ and _________ had their own access into the B3 suite or whether or not someone had to let them in. IVINS further advised the interviewing Postal Inspector and SA to check USAMRIID key card entries.

        [NOTE: Key card entries only go back to mid-1998 and do not extend to May 1998, the date of this visit].

        iVINS further advised ______________________ were “at eat others throats.” IVINS advised _________ was “a jerk” and had problems with everyone to include ____________.”

        Comment: I called Michael Hayes to ask the details of this visit but he said “You don’t want to know.” I emailed Tarek long before posting on these issues but he did not respond.

    • DXer said

      2/03/2005

      IVINS stated in the voice message that the concentration on the 1997 shipping form made him believe that RMR 1030 was the Ames material he sent to Dugway for use in the mass production of spores for the making of RMR 1029. He advised that the information to support his belief could be found in his USAMRIID laboratory notebook #3655, page 86.

  2. DXer said

    5/07/04 IVINS Interview Statement –

    IVINS has never had to add antifoam or any other chemical to his production method for producing Ames spores. The shaking of the shaker flasks in the broth production is not enough to create a need for antifoam or any similar product. He is not aware of any protocol used at USAMRIID which calls for the use of antifoam with Ames. The protocol for the aerosol challenges does not call for antifoam.

  3. Anonymous said

    It is now 45 days since the NAS anthrax science report was supposed to be completed and published.

    When will this $1M report be released?

    http://www8.nationalacademies.org/cp/projectview.aspx?key=49105

    The start date for the project is 4/24/2009.

    A report will be issued at the end of the project.

    Project Duration: 18 months

  4. Old Atlantic said

    ““Thus if the estimates silicon concentrations in the Amerithrax spores are correct, they are not consistent with our current understanding of silica deposition or those materials must have indeed been produced under an unusual set of conditions. If the latter were true, the silica evidence might provide a significant bound on the credible growth and production scenarios that would be consistent with the prosecution narrative in this case.””

    “We are all agreed that your theory is crazy. The question that divides us is whether it is crazy enough to have a chance of being correct.”

    Niels Bohr

    “under an unusual set of conditions”

    Like al Qaeda guys making it up as they go along under constrained circumstances.

  5. Old Atlantic said

    Another paper on permeability and also discussion of flotation of spores.

    http://www.ncbi.nlm.nih.gov/pmc/articles/PMC279244/pdf/jbacter00474-0143.pdf

    “PERMEABILITY OF BACTERIAL SPORES
    1. CHARACTERIZATION OF GLUCOSE UPTAKE
    S. H. BLACK AND PHILIPP GERHARDT
    Department of Bacteriology, The University of Michigan Medical School, Ann Arbor, Michigan
    Received for publication May 15, 1961”

  6. Old Atlantic said

    Also available”

    http://aem.asm.org/cgi/reprint/6/5/327.pdf

    Separation and Concentration of Bacterial Spores and Vegetative
    Cells by Foam Flotation
    W. A. BOYLES1 AND R. E. LINCOLN
    U. S. Army Chemical Corps, Fort Detrick, Frederick, Maryland
    Received for publication February 12, 1958

    “Selective methods for removing and/or concentrating
    bacterial spores and vegetative cells from the growth
    medium are essential in studies on spore physiology
    and antigenicity, phagocytosis, and similar problems
    in which cell preparations free of cellular debris are desired.
    Development of a method to obtain “clean”
    preparations followed the observation that masses of
    material collected above the liquid level in the head of
    foam when Bacillus anthracis was grown in aerated deep
    cultures. When this material was smeared and stained,
    microscopic observation indicated that the material was
    composed of essentially clean spores. Frequently, on
    those occasions when masses of the material collected
    or when foam was incompletely controlled and lost
    through the air vent, the spore count of the culture was
    low. These observations led to the conclusion that a
    collection process could be developed that would
    separate B. anthracis spores from vegetative cells and
    cellular debris contained in the culture medium. Such a
    method of purification by flotation was developed and
    is reported in this paper. While this manuscript was
    being edited, it was learned that similar independent
    observations had been made and the same conclusions
    I Deceased.
    drawn by Black et al. (1958) who observed the loss of
    Bacillus cereus spores from deep culture fermentations
    after uncontrolled foaming.
    In the present study, simple glass cylinders equipped
    with spargers or diffusers were used as foaming chambers.
    Serratia marcescens cells were first successfully
    separated and collected and, with the development of
    equipment and a working procedure, cells of Brucella
    suis and Pasteurella (Bacterium) tularensis and spores of
    B. anthracis and Bacillus subtilis var. niger (B. globigii)
    were processed. Although major emphasis has been
    placed on the results obtained with B. anthracis, data
    on several other species are given to supply additional
    information on the flotation of cells or spores

  7. Old Atlantic said

    “Indices for evaluation. In flotation, as in any physical
    separation process, evaluation of the effectiveness of
    the operation requires the quantitative measurement
    (or assay) of either the preferred or unwanted constituent
    in a product, or, better still, of both. These
    assays are translated into ratios or indices which make
    clear the degree of success in separation. Two indices
    of effectiveness commonly used in flotation are “grade”
    (or purity) and “recovery.” The grade expresses the
    number of parts of a constituent per hundred parts of
    dry product, for example, a concentrate containing 2 g
    of spores and 3 g of other solids would have a grade of
    40 per cent. The recovery is the percentage of the total
    amount of a constituent from a feed which is found in
    a product or
    100 Cc (3)
    Ff
    A flotation test cannot be evaluated unless both
    recovery and grade are measured. A feed containing
    10 g of spores and 1 g of vegetative cells might yield a
    concentrate containing 9 g of the spores, and the test
    would be judged a success from the standpoint of recovery.
    But if the concentrate also contained 0.9 g of
    the vegetative cells, the grade has not been altered.
    Since the object of flotation is the separation of solids
    from each other, the analysis of a flotation test should
    be made on the basis of both grade and recovery.

  8. Old Atlantic said

    “Media and preparation of cultures. The organism
    used throughout this study was Bacillus subtilis var.
    niger. Spore concentrates were obtained from the U. S.
    Army Chemical Corps, Fort Detrick, Frederick,
    Maryland, or were produced at Massachusetts Institute
    of Technology (M.I.T.). Spores from Fort Detrick
    were grown in a casein hydrolyzate medium (Boyles and
    Lincoln, 1958) in metal fermentors and, after autolysis,
    were harvested by centrifugation at 50,000 X g. The
    spore concentrate, or paste, was shipped both frozen
    and unfrozen, with and without an additional washing
    operation in the centrifuge. Freezing was accomplished
    by extruding the paste into an acetone-Dry Ice bath.
    Frozen preparations were stored at -20 C, whereas
    unfrozen preparations were kept at 4 C. Frozen preparations
    contained about 50 X 1010 viable spores per g
    of dry wt.”

    From part I.

  9. Old Atlantic said

    The Gaudin paper has a part I also available.

    http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1057576/pdf/applmicro00328-0028.pdf


    The separation of bacterial cells from the growth
    menstruum is a routine operation which continues to
    demand more than its fair share of effort from workers
    in such fields as physiology, immunology and cytology.
    Preparation of “clean” cell suspensions by centrifugation
    is arduous and yields are often low.
    The observation that masses of cells collected in the
    foam above the liquid level in fermentors led Boyles
    and Lincoln (1958) to use vigorous aeration of the
    culture medium after growth as a means for removing
    and concentrating spores of Bacillus anthracis. Quantitative
    estimates of the degree of separation and/or
    concentration were based on viable counts. Success in
    separating spores of Bacillus cereus T (formerly called
    Bacillus cereus var. terminalis) from autolyzed culture
    by frothing was reported by Black et al., (1958).
    The concept of the separation of particles in liquid
    by foaming has long been known in the field of mineral
    engineering, and is the basis for “flotation” processes.

    “Definitions. 1’lotation may be defined as a process
    for separatinig finely divided solids from each other
    (Gaudin, 1957). The solids are suspended in water
    through which gas bubbles are caused to flow. Separation
    takes place when particles of one type adhere
    to gas bubbles and are carried to the top of the liquid
    as a froth, whereas particles of other types adhere to
    the liquid and remain in suspension.”

    This part of the paper has some basic explanations that even I can understand.

  10. Old Atlantic said

    Ezzell and the mail anthrax preparer both were working with daughters of RMR-1029. Ezzell did not end up with the same silicon content.

    Ezzell got white spores v tan for the Daschle Leahy. The NY Post letter had very high silicon and more debris and vegetative cells.

    Thus the numerical data of the letters and Ezzell’s lab is different. The numerical outcome is different on these dimensions.

    Thus the mathematical operations that take you from starting with a sample from RMR-1029 to the numerical outcomes are different. Thus the experimental steps were different.

    Its finding the experimental steps for the letters that is our task. The data we have is the numerical outcomes. We can then infer different mathematical operations for the processing steps of Ezzell v the letters. We can then try to fit an experimental procedure to the letter anthrax processing steps.

    We are thus looking for different processing steps and likely ones with less efficiency than Ezzell had and also an attempt to use all of the original spores and not discard those clinging to debris or vegetative cells. The flotation separation of vegetative cells, debris and spores was likely less for the letter preparer than for Ezzell.

    The letter mailer came up with worse numerical outcomes than Ezzell. So the letter mailer was likely someone with less skill and know-how than Ezzell. That may have been a person from a semiconductor background who thought to use silicon possibly in an alkaline solution and who those got hung up on the hard road to get flotation separation of debris, vegetative cells and spores.

    So they got the different numerical outcomes in the letter anthrax from Ezzell’s numerical outcomes in his anthrax even though they both started from RMR-1029.

    This implies someone with a radically different background from Ezzell. That is not Ivins but some other scientist/engineer type completely.

    • BugMaster said

      “This implies someone with a radically different background from Ezzell. That is not Ivins but some other scientist/engineer type completely.”

      Very well put, Old Atlantic.

  11. Old Atlantic said

    We can think about solving the problem with this scheme.

    1. We start with the NY Post high debris and veg cells relatively and high silicon and Daschle better purity on spores but still high silicon relative to normal.

    2. We try to fit math to what we observe. We think of the math operations that would produce the observed numerical data.

    3. We then try to find the experimental procedure that has the math operations as its result or that are consistent with the math operations.

    The suggestions below may or may not be right, but this reverse step approach is one to try to understand what happened.

    Math operations that give us silicon accumulation are a possible solution. We then look for experimental steps that would result in such an accumulation mechanism. We also want to get out the result of more silicon in the less pure spore sample. We still want to have high silicon even in the Senate letters.

    • BugMaster said

      Some of the silicon could have been added in the form of a silicon-based antifoam (far in excess of what one would add to liquid fermentation media).

      Perhaps the perp felt this additional material would impart some “desirable” characteristics.

      Since this material is polymerized, it would be in addition to the trace amounts of material found in the spore coat. And it may not have contributed much to the Senate material’s “desired” characteristics, but rather, could have aided in the purification in some way.

      • anonymous said

        They tried using silicon antifoams and still couldn’t reproduce the silicon levels in the attack spores. This is mentioned by Weber in his presentation to NAS.

        The obvious inference is that the spores were treated post growth with a siloxane. It would penetrate the exosporium and polymerize in situ on the spore coat – giving exactly the observed silicon signature.

        Details on treating cellular material like this can be found here:
        Detection and distribution analysis of organosilicon compounds in
        wood by means of SEM-EDX and micro-CT (just Google it).

        Such spores treated like this become hydrophobic (repel water) – exactly what the attack spores were observed to do.

        • BugMaster said

          “The obvious inference is that the spores were treated post growth with a siloxane. It would penetrate the exosporium and polymerize in situ on the spore coat – giving exactly the observed silicon signature.”

          Sounds plausable. But could silicon from a concentrated sucrose solution (present in forms of silicic acid) have given the same signature? (also could explain the tin, if it was cane sugar).

  12. Old Atlantic said

    The search silicon alkaline solutions is interesting. The Gaudin paper indicates that alkaline variations don’t work.

    “The grade of spores in the float is not appreciably
    affected by increasing alkalinity, except perhaps at the
    highest level.”

    If they were using silicon in an alkaline solution they may have had a hard time in separation.

    The search

    silicon alkaline solutions semiconductors

    brings up hits.

    If for some reason their technology base was oriented to semiconductor thinking or experience and they used a silicon alkaline solution then they might have had a hard time to get the separation of debris, vegetative cells and spores based on the result of the Gaudlin paper. To repeat that

    “The grade of spores in the float is not appreciably
    affected by increasing alkalinity, except perhaps at the
    highest level.”

    See page 5 of 7 of pdf page 95 of the paper.

  13. Old Atlantic said

    Tuning problem. RMR-1029 was a mixture. If the separation method and additives used work for some parts of the mixture but not for others to separate the floating debris, vegetative cells, and spores, then you can’t tune both at the same time. Thus if you tune your additives and conditions to get good separation of floating spores for one part of the RMR-1029 you are not in tune for the others and so you end up with a mixture of debris, vegetative cells and spores at different heights.

    Thus RMR-1029 may have presented special separation problems for the additives being used. Or they may not have found the right tuning for it because it was so unusual given the method they were using.

    Thus they had to do multiple centrifugings. They may also have tried different additive mixtures each round to get better separation of spores from vegetative cells and debris.

    Another issue is whether Daschle and Leahy are the better separations from the same processing as the first letters that were held back for some reason or represent further processing or a new start. The missing letter mailed to Florida that had anthrax also is said by some to be better than the NY Post, so it might be better separation results from an earlier processing stage.

    • BugMaster said

      “The missing letter mailed to Florida that had anthrax also is said by some to be better than the NY Post, so it might be better separation results from an earlier processing stage.”

      Or the mailer just had more time to prepare it. And soon after he mailed the letter to Boca Raton, he had to hustle to “jump on the 9-11 bandwagon”.

      Thus the crude that was sent out on 9-18. In fact, he could have started with old inocula left over from the previous prep (a possible mechanism by which the contaminating b. subtilis establised itself in the culture).

  14. Old Atlantic said

    The Gaudin et al paper has the following implication:

    If the methodology used for separation of spores, debris and vegetative cells is off somewhat, you might get a hard time in separation. You might get vegetative cells and debris floating or spores clinging to the vegetative cells or debris if they don’t float.

    Thus depending on the method used to achieve separation of spores from debris and vegetative cells, the degree of separation of debris, veg cells and spores in the float may have been lower or higher.

    If the degree of separation was lower, then more rounds of centrifuging would be needed.

    If we assume the New York Post letter had high silicon and a lot of debris and vegetative cells compared to Daschle or to Ezzell’s work, then the method used to prepare New York Post may have hung up on a bad combination of additives and other conditions including possibly the strain or strains in this mixture.

  15. Old Atlantic said

    More from this paper at MIT

    “Tables 3 and 4 show that:
    1. The grade of spores in the float tends to increase
    with increasing acidity and then levels off.
    2. The grade of spores in the float is not appreciably
    affected by increasing alkalinity, except perhaps at the
    highest level.
    3. The recovery in the float of both spores and debris
    increases markedly with increasing acidity, and is but
    slightly affected by increased alkalinity.
    4. A significant improvement in selectivity index is
    not obtained by changes in pH with the particular acid
    and alkali used.
    Figure 6 shows the variation in the grade of spores in
    the float as a function of pH. The big change in grade
    near the neutral point is not due to pH variations but
    to the use of different lots of organisms for the two
    series of tests.
    The recovery of spores and of debris as a function of
    pH is shown in figure 7. It is seen that essentially complete
    recovery of spores and debris as a float product
    is obtainable by suitable pH adjustment, although the
    separation efficiency of spores from debris is poor. Just
    why pH has this effect is not known.”

    “DIscUSSION
    It is clear that there are differences in floatability
    between spores, vegetative cells, and debris. Relatively
    speaking, these differences are small but experiments
    show they can be increased. Thus, the addition of
    secondary amines seems to favor flotation of spores
    over debris, whereas the addition of carboxylic acids increases the floatability of vegetative cells and debris
    over that of spores. There can be little doubt that this
    is related to the polar groups available at the surfaces
    of spores and vegetative cells. The behavior of the two
    forms of the organism suggest that both cationic and
    anionic sites are available at the surface of both forms
    but that the relative abundance of the sites is different,
    the spores having an excess of carboxyl groups and the
    vegetative cells an excess of amino groups.
    Preliminary experiments with another strain of B.
    subtilis indicates that its behavior is not the same as
    that of the niger variety, hence that the outer coats of
    an organism differs from strain to strain.
    Pilot experiments with other organisms further indicate
    differences between one species and another, as
    might be expected. The prospect of relating flotation
    behavior to chemical composition, structure, and molecular
    orientation at the surface of spores and vegetative
    cells is an area to which attention should be
    directed.”

    • Old Atlantic said

      “SUMMARY
      Among the factors which influence the flotation behavior
      of Bacillus subtilis var. niger spore preparations
      are the age of the spore material at harvest, the solublesalt
      content of the flotation feed, prior heating of the
      pulp, pH of the pulp, and the addition of amine or fattyacid
      collectors.
      It is shown that judicious use of flotation “tools”
      can effect suitable separations, but much work remains
      to be done in correlating flotation behavior with surface
      properties.”

  16. Old Atlantic said

    “In this paper, it is shown experimentally that separations
    can be made, and that the flotation behavior
    of spores and debris of B. subtilis var. niger is dependent
    upon the age of the bacterial components at harvest,
    the content of soluble liquid components in the flotation
    feed, the pH of the pulp, the prior heat treatment
    of the flotation feed (if any), and the addition of flotation
    reagents.”

    http://aem.asm.org/cgi/reprint/8/2/91.pdf

    Separation of Microorganisms by Flotation
    IL. Flotation of Spores of Bacillus subtilis var. niger1
    A. M. GAUDIN,2 A. L. MULAR,3 AND R. F. O’CONNOR3
    Department of Metallurgy, Massachusetts Institute of Technology, Cambridge, Massachusetts
    Received for publication August 10, 1959

    “Bacillus subtilis. var. niger spore preparations were
    either shipped direct from the U. S. Chemical Corps,
    Fort Detrick, Frederick, Maryland, or were grown at
    the Massachusetts Institute of Technology (M.I.T.)
    laboratories using Fort Detrick inoculum.”

    Early experiments with Fort Detrick spore preparations
    proved that considerable variation in flotation
    behavior existed according to whether the preparation
    consisted of spent liquor as well as organisms, whether
    it had been washed, and whether the sediment was
    then frozen. To avoid working with spore material of a
    variable nature, spores were grown at M.I.T. This
    provided a flotation feed of reproducible behavior.
    Among the factors which appeared to influence the
    behavior of Fort Detrick spore preparations were, the
    amount of soluble components present, the length of
    time in storage, the heat-shocking of sediment, and the
    freezing of sediment in an acetone-Dry Ice bath. These
    observations were sufficient indication that age and
    soluble liquid components are variables which, along
    with the usual mineral-flotation variables such as
    temperature, pH, and reagents, merit more detailed
    study.

  17. Old Atlantic said

    http://invam.caf.wvu.edu/fungi/taxonomy/Paraglomaceae/occultum/occultum.htm

    “Spores often float on the surface of sucrose following density gradient centrifugation and also float in water with swirling. They also tend to cling to organic debris (including spores of other arbuscular fungi).”

    These are not anthrax spores that I could tell.

    • BugMaster said

      Never tryed anthrax spores (don’t have any and don’t want any), but I did try the technique on some plate-grown b. subtilis material (assumed it was subtilis, the goddamn thing had contaminated one of my cultures).

      Note that is was a very small scale “proof of concept” exercise.

      Worked real well, actually. In fact, it was kind of creepy!

      • Old Atlantic said

        I was just emphasizing these were funghi spores in the article I linked to. So your subtilis results are much more directly relevant than these funghi results.

  18. Old Atlantic said

    More Crazy Silicon Accumulation Mechanisms. The following are offered as a way to stimulate thought on how silicon might have accumulated. Maybe these have problems too. Bug Master or others please point those out.

    You start with 2 test tubes. One is empty, one is full of your prep. You centrifuge the prep with sugar cane added and some of your spores float to the top. You pour those off into the tube 2, The Good Accumulator.

    Now you centrifuge tube 1 again, adding more sugar cane.(You might pour out the water through a screen if such a screen exists. Then you pour in fresh water and add more sugar cane.)

    Again, what floats to top is poured into the Good Accumulator Tube. You keep repeating this process.

    If you have a third tube, then you can also centrifuge the Good Tube, pour the top off into the 3rd tube and then you have a couple choices. Call this stage, Pt A.

    You could centrifuge the remainder in tube 2 adding more sucrose, and pour its top into tube 3. You then pour its bottom into tube 1, so that you can centrifuge tube 3 and pour its top into tube 2.

    At point A, you could instead pour tube 2 into tube 1 and centrifuge tube 1, pour the top into tube 3 and then centrifuge tube 3 and pour its top into tube 2, etc.

    With more tubes you have more options.

    This is contrast to some other procedures. Only keep what you pour off the top off and get rid of all else. This procedure has lower yield of stuff, although it is pure.

    Unlimited tubes. You always separate into 2 or more tubes, and then divide those into 2 or more tubes. Eventually, you combine the ones that are of the same grade.

    If you have 10 tubes, you run against the ten barrier, but you can practice this type of grade judgment method so that you have accumulator tubes of various grades. These may vary in time so that your best tubes are higher in grade and your worst are worse. But you try to avoid discarding anything that contains spores, including those clinging to debris or vegetative cells.

    If you have 3 or more tubes and centrifuge the good stuff, and the bad stuff, adding sugar cane or something with silicon at each stage, then you get a silicon accumulator into both good stuff and the bad stuff.

    One problem with this is whether the spores will take up silicon in the good stuff and whether the debris and vegetative cells in the residue tubes will take up the silicon if the spores in those tubes won’t.

    Maybe there is some sort of X that will help spores take up silicon after they have already formed spores. Perhaps the tin or other elements? Something that was used up or didn’t persist in the final prep?

    The advantage of this method is that if your anthrax supply is fixed at the point you do this or costly to produce, then you save more of the spores including those that stubbornly cling to debris or vegetative cells.

    • Old Atlantic said

      In some of these schemes, the purer spores will either not become pure white or will do so more slowly. If you have 3 tubes, and you are alternating centrifuging the purer one and the least pure and using the extra tube as an intermediate, then you are limited in purifying the purer spores to white. You are giving up the speed of getting purer spores for the sake of not discarding spores from the 3 tube system even those clinging to vegetative cells or debris.

    • Old Atlantic said

      Suppose have one tube with prep and 2 empty tubes. You centrifuge first tube with prep and pour top into tube 3. Then centrifuge tube 3 and pour top into tube 2. Pour bottom into tube 1. Continue this pouring the bottom in tube 1. This purifies towards white.

      If the sequence involves centrifuging tube 1 again or the bottom in tube 2 or 3 after pouring the top into another tube, then the whiteness of the spores has to be less than in the first sequence above.

      • BugMaster said

        Depending on the density of the sucrose, the spores float to the top without centrifugation (let them set overnight, this could have also given them the time to absorb silicic acid and tin).

        And it seems a lot of the gunk and junk gets left behind in the viscous liquid.

        This could have been used as a preliminary prep, with more conventional centrifugation / wash steps like Ezzel described applied later.

  19. Anonymous said

    This seems to really vindicate Epstein’s article:

    —————————–

    Epstein wrote on 1/24/10:

    “The FBI’s answer was that the anthrax contained only traces of silicon, and those, it theorized, could have been accidently absorbed by the spores from the water and nutrient in which they were grown.
    No such nutrients were ever found in Ivins’s lab, nor, for that matter, did anyone ever see Ivins attempt to produce any unauthorized anthrax (a process which would have involved him using scores of flasks.)
    Natural contamination was an elegant theory that ran into problems after Congressman Jerry Nadler pressed FBI Director Robert Mueller in September 2008 to provide the House Judiciary Committee with a missing piece of data: the precise percentage of silicon contained in the anthrax used in the attacks.
    The answer came seven months later on April 17, 2009.
    According to the FBI lab, 1.4% of the powder in the Leahy letter was silicon.”

    ——————————————
    Then Hassell responded on 2/3/10:

    “Monday’s opinion piece, “The Anthrax Attacks Remain Unsolved,” was filled with inaccuracies and omitted several relevant facts that are necessary for a balanced discussion of the science applied in the anthrax investigation.

    The FBI is confident in the scientific findings that were reached in this investigation. We utilized established biological and chemical analysis techniques and applied them in an innovative manner to reach these findings.”

    ——————————————————–
    The FBI have now thus significantly changed their position. Everything they write in the new Microbial Forensics is consistent with what Ed Epatein wrote. They now admit that even by deliberately adding silicon to the broth in the form of silica the HIGHEST CONCENTRATION THAT COULD EVER BE ACHIEVED IS 0.3% !!!!

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