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

* The DOJ and FBI Should Produce To GAO All Forensic Analyses and Documents Relating to Use of Fatty Acid Methyl Ester Profiles For Determination Of The Culture Media Used To Grow The Anthrax Used In The Fall 2001 Anthrax Mailings

Posted by Lew Weinstein on April 17, 2012

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10 Responses to “* The DOJ and FBI Should Produce To GAO All Forensic Analyses and Documents Relating to Use of Fatty Acid Methyl Ester Profiles For Determination Of The Culture Media Used To Grow The Anthrax Used In The Fall 2001 Anthrax Mailings”

  1. DXer said

    Chemical and Physical Signatures for Microbial Forensics
    Infectious Disease, 2012, 35-52, DOI: 10.1007/978-1-60327-219-3_3

    Fatty Acids and Lipids

    James M. Robertson, Christopher J. Ehrhardt and Jason Bannan

    • Abstract

    Lipids are an integral component of the bacterial membrane and show great structural diversity within the cell. While lipid composition has been an important signature of bacterial phylogeny for many decades, it also has the potential to provide information on the resources and procedures used to culture an organism. Chemical factors like nutritional substrates, temperature, and physical dynamics during growth all can influence the types of lipids and their relative ratios inside the cell and potentially leave diagnostic biosignatures that are unique to a specific production process.

    In this chapter, we examine the structural diversity of lipids in the cell and the factors that affect lipid composition during laboratory culturing with particular emphasis on Bacillus organisms. Methods used to extract, separate, and detect fatty acids are reviewed. In addition, the potential utility of lipid profiles in forensic investigations is discussed in the context of specific examples from the literature. Lastly, validation considerations and quality assurance are highlighted as important aspects in the implementation of lipid analysis for microbial forensics.

  2. DXer said

    Infectious Disease 2012, Chemical and Physical Signatures for Microbial Forensics

    Extracellular Signatures as Indicators of Process Methods

    Karen L. Wahl
    Pages 107-115

    Abstract
    Much of the focus within microbial forensics has been on characteristics of the specific agent involved, principally the genetic makeup. The phenotypic changes in the microbial cell have also been characterized by analyzing changes in the structure, elemental, chemical, or biochemical content of the cell produced in response to the growth environment. The same analytical methods used to detect cellular constituents can be applied to detecting residual indicators of materials used for production and preservation. Specific examples, such as residual agar or blood components from solidified growth medium, have been targeted for assay development. However, the same methods can be applied to detection of other carbohydrate and protein components derived from growth medium or used for dry stabilization. Taken together, these residual components comprise a potentially rich signature of the method used to produce a biological agent.

  3. DXer said

    Wasn’t the already published data regarding to tin was exculpatory of Dr. Ivins?

    Given that the FBI’s lead scientist’s hypothesis described above was that the tin came from the water used, isn’t what was already known by the USGS and in the public domain exculpatory of Dr. Ivins?

    As an evidentiary matter, a federal district court judge could have taken judicial notice of such data.

    Tin

    http://mrdata.usgs.gov/catalog/science.php?thcode=2&term=1596

    • DXer said

      http://www.lenntech.com/periodic/water/tin/tin-and-water.htm

      The naturally occurring amount of tin in soils and water is relatively small. The release of tin from anthropogenic processes exceeds tin release from geological processes 110 times.

      Tin is applied as a protective layer on iron cans to prevent corrosion. This property is mainly applied in conserve can production, and relatively large amounts of tin may enter the food chain through weathering from unvarnished cans. Conserve cans are partially recycled. Tin is present in various alloys, such as soldering tin, bronze and amalgam for dentistry. However, most tin is applied in cans.

      Read more: http://www.lenntech.com/periodic/water/tin/tin-and-water.htm#ixzz1slszTqC4

      Let’s consider manmade as opposed to geological sources.

      By way of background, in Fall 2001, the Armed Forces Institute of Pathology (”AFIP”) had detected silicon dioxide (silica) in the attack anthrax — with a characteristic big spike for the silicon. No silica was observable on the SEMs images that Dr. Alibek and Dr. Matthew Meselson saw. The Daschle product was “pure spores.” Was silicon dioxide used as part of a “Microdroplet Cell Culture” process used prior to drying to permit greater concentration? Joe Michaels’ research, as interpreted by Dr. Majidi, the top FBI WMD scientist, indicates that the silica could have been in the culture medium. As explained in a later related patent, the silica could be removed from the surface of the spore through repeated centrifugaton or an air chamber.

      Is there also a source for in in the water at GMU? Dr. Bannan used to work as a collection scientist in the Bacteriology Division at ATCC which shared space at GMU — and so he may know. GAO should ask him.
      Dr. Alibek and Dr. Bailey had filed a patent application in mid-March 2001 involving a microdroplet cell culture technique that used silicon dioxide in a method for concentrating growth of cells. The patent was granted and the application first publicly disclosed in the Spring of 2002. Weren’t the SEMS images and AFIP EDX finding both consistent with use of this process in growing the culture? It’s been suggested informally to me that perhaps the silicon analytical peak was more likely due to silanol from hydrolysis of a silane, used in siliconizing glassware. But didn’t the AFIP in fact also detect oxygen in ratios characteristic of silicon dioxide? Wasn’t the scientist, now deceased, who performed the EDX highly experienced and expert in detecting silica? Hasn’t the AFIP always stood by its report? In its report, AFIP explained: “AFIP experts utilized an energy dispersive X-ray spectrometer (an instrument used to detect the presence of otherwise-unseen chemicals through characteristic wavelengths of X-ray light) to confirm the previously unidentifiable substance as silica.”

      Perhaps the nuance that was lost — or just never publicly explained for very sound reasons — was that silica was used in the cell culture process and then removed from the spores through a process such as centrifugation. Or perhaps sol gel was used as a drying agent such as under an old USAMRIID technique (this is the hypothesis favored by Dr. Alibek when I last consulted). Or perhaps it is due to rice hull contamination. Or use of a silicone sealant on the inside of the envelope which the AQ manual instructs the mailer of a poisonous letter to use to avoid killing the mailman. Beats me. That’s why we have the wonderful experts hopefully available to be consulted by GAO addressing the issue. The applicants in March 2001 for an international patent relating to vaccines were a leading aerosol expert, Herman R. Shepherd, and a longstanding anthrax biodefense expert, Philip Russell.

      Dr. Morozov is co-inventor along with Dr. Bailey for a patent “Microdroplet Cell Culture” that explains how the silicon dioxide can be removed from the surface. Perhaps it is precisely this AFIP finding of silicon dioxide (without silica on the SEMs) that is why the FBI came to suspect Al-Timimi in 2003 (rightly or wrongly, we don’t know). The FBI would have kept these scientific findings secret to protect the integrity of the confidential criminal/national security investigation. There was still a processor and mailer to catch — still a case to prove. That is, until DOJ and FBI screwed the pooch and liked it.

      An example from October 2006 of equipment that went missing from GMU’s Discovery Hall was a rotissery hybridization oven belonging to the Center for Biomedical Genomics. “This equipment can be used to manufacture biological agents and genetically modified agents, which could potentially be used as biological weapons,” Corinne Verzoni explained in her PhD 2007 thesis. “Upon hearing about instances or missing equipment in Discovery Hall, the author contacted campus security who was unaware of instances of missing equipment. Missing equipment should be reported to the equipment liaison. Missing equipment may not be reported to campus security because labs tend to share equipment. Equipment also goes missing because it is not inventoried if it is under $2,000.”

      One of her other examples was equally dramatic:

      “A DI system is a de-ionized water system, which removes the ions that are found in normal tap water. The assistant director for operations noticed the DI system in Discovery Hall was using the entire 100 gallons in two days, which is an enormous amount of water for the four DI taps in the whole building. According to the assistant director for operations, it is difficult to calculate the reason for that much water since no leak was found. A large amount of water used over a short period of time for unknown reasons could indicate that the research is being conducted covertly.”

      “A student with legitimate access to Discovery Hall,” she explained, “has easy accessibility to equipment. A student with access to the loading dock could steal equipment on the weekend when campus security is not present in Discovery Hall. A student could also walk out of the entrance with equipment on the weekend without security present.” She concluded: “The events at GMU demonstrate opportunity to create a clandestine lab, the ability to sell items illegally, or the ability to exploit school equipment.” In a late September 2001 interview on NPR on the anthrax threat, Dr. Alibek said: “When we talk and deal with, for example, nuclear weapons, it’s not really difficult to count how much of one or another substance we’ve got in the hands. When you talk about biological agents, in this case it’s absolutely impossible to say whether or not something has been stolen.”

      Dr. Bannan, can you tell the GAO whether Ali Al-TImimi had unrestricted access to the specimens you kept at your bacteriology collection? Did you participate in the determination that virulent Ames was not kept in ATCC’s confidential patent repository? A whisteblower (who has a JD and PhD) tells me that she was let go by ATCC as soon as she complained about lax security there; she had BL-3 and BL-4 experience and so was quite experienced and knew lax security or a total lack of security when she saw it.)

      That’s what I do. I represent whistleblowers. Everyone’s good faith will be presumed. But everyone needs to go back to their files and give the GAO the documents that they have not produced yet. In this day and age, there are no secrets that can be assumed will be kept. There are just people who are ahead of the curve — like the former Walmart person who came forward … and then there are people who fall behind the curve and get run over.

      • DXer said

        Dr. Alibek, for his part, told me in 2003 that their research with virulent Ames was not done on campus and was done instead by Southern Research Institute. (SRI was in Frederick, MD). See also his company’s press release. Dr. Ivins’ chief accuser, PF, came to head the BL-3 lab there. Dr. Bannan’s former employer, ATCC (at GMU), had BL-3 labs but GMU’s BL-3 lab at the time was used for storage.

      • DXer said

        DARPA funded Dr. Alibek and Dr. Bailey’s program — DARPA had given the Center $12 million prior to 9/11 as I recall. Dr. Bruce Ivins supported DARPA research by supplying virulent Ames from Flask 1029, such as the Ames given to the FBI’s lead scientist, JE, to make a dried powder.

        Amerithrax represents the greatest failure in intelligence analysis in United States history.

        Dr. Zawahiri may have needed Dr. Fadl to guide him in clinical practice with treating people at the clinic in Saudi Arabia. But Dr. Zawahiri now has had quite a lot of experience and become quite accomplished in killing people. Thus, this is not a matter in which AUSA Lieber and Agent Montooth’s good faith mistakes can go uncorrected.

        From the start, dating to 2001, FBI and CIA analysts should have zeroed in on who Dr. Ayman knew to recruit in accomplishing his stated plan to kill people using anthrax.

  4. DXer said

    This issue had previously been the subject of study by FBI scientists in connection with the mailed anthrax and was addressed again this year by Dr. Bannan and colleagues.

    Was heme detected in the testing of the Fall 2001 anthrax mailings? Based on iron content? (Although not a scientist, I venture yes based on earlier articles on the subject).

    The authors in Appl Environ Microbiol. 2008 June; 74(11): 3573–3582, “Bayesian-Integrated Microbial Forensics” explain:

    “The utilities of various analytical techniques and approaches for characterizing an organism’s growth environment have been explored. Valentine et al. (31) demonstrated reproducible differences in matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) signatures of Bacillus subtilis spores grown in different culture media. Kreuzer-Martin and Jarman (20) have discussed the usefulness of 13C/12C and 15N/14N isotope ratios for characterizing culture media. Cliff et al. (8) suggested secondary-ion MS (SIMS) as a means for identifying a culture medium based on the metal content. Edberg et al. (H. C. Edberg, C. E. Petersen, N. B. Valentine, D. S. Wunschel, and K. L. Wahl, presented at the 54th ASMS Conference on Mass Spectrometry and Allied Topics, Seattle, WA, 2006; H. C. Edberg, C. E. Petersen, N. B. Valentine, and K. L. Wahl, unpublished data) demonstrated a method for detecting the presence of agar in microbial samples based on electrospray ionization (ESI) MS and derivatization gas chromatography MS. Whiteaker et al. (34) developed a MALDI MS-based method for detecting heme on Bacillus spores.”

    Yazid Sufaat’s Green Lab was involved in blood testing. In Afghanistan, the cover for his laboratory was blood testing. Marwan Hadid worked at Omar Hospital assisting Yazid Sufaat. Hadid was a hematology technician.

    https://caseclosedbylewweinstein.wordpress.com/2011/05/09/marwan-hadid-worked-at-omar-hospital-assisting-yazid-sufaat-hadid-was-a-hematology-technician-in-may-2001-yazid-sufaat-reportedly-was-attempting-to-cultivate-anthrax-there-who-does-marwan-had/

    Does Dr. Bannan interpret the detection of heme as pointing to Dr. Ivins rather than Yazid? Why? Because of all the exsanguination of rabbits going on in B3 in the first week of October that the FBI neglected to mention? (Evidence that the prosecutors and FBI knew of Dr. Ivins’ work with rabbits would include internal documents, if GAO manages to force their production, showing it was their speculation that Dr. Ivins used the blood from the rabbits to grow the anthrax!)

    Or did Dr. Bannan interpret the presence of heme as pointing to Dr. Ivins just because like many scientists, he used sheep blood agar as well as Leighton-Doi?

    If the FBI had disclosed all the rabbits that were dying in B3 in the first week of October 2001, it could have been pointed out that it was INCONSISTENT with the narrative and claim that Dr. Ivins was using the space to grow and dry anthrax using hundreds of plates. Reporters still need to conduct interviews of those who assisted with the exanguination of rabbits that first week of October 2001.

    In Wunschel et al., “Detection of agar, by analysis of sugar markers, associated with Bacillus anthracis spores, after culture,” J Microbiol Methods. 2008 Aug;74(2-3):57-63. Epub 2008 Apr 12, we see that D. Fetterolf of the FBI provided irradiated Ames. The authors explained “determining information about growth characteristics remains a challenging task that has only recently begun to be addressed. It is vital to be able to assay trace contaminants. Furthermore, in chemical analysis of such a precious sample there is only a finite amount of material that can’t be replenished (e.g. by growth or polymerase chain amplification, PCR, as for molecular diagnostics). Markers for agar which is, of course, a constituent of all widely used solid media could be used to differentiate prior growth in liquid media. Other components of media may be indicative of a specific medium (e.g. one containing animal constituents, such as sheep blood agar …”

    More recently, Dr. Bannan notably is co-author of the chapter in 2012 on this subject. (Dr. Robertson also is from the FBI’s Laboratory Division, Counterterrorism and Forensic Unit. )

    Chemical and Physical Signatures for Microbial Forensics
    Infectious Disease, 2012, 35-52, DOI: 10.1007/978-1-60327-219-3_3
    Fatty Acids and Lipids

    James M. Robertson, Christopher J. Ehrhardt and Jason Bannan

    The Abstract states:

    “Lipids are an integral component of the bacterial membrane and show great structural diversity within the cell. While lipid composition has been an important signature of bacterial phylogeny for many decades, it also has the potential to provide information on the resources and procedures used to culture an organism. Chemical factors like nutritional substrates, temperature, and physical dynamics during growth all can influence the types of lipids and their relative ratios inside the cell and potentially leave diagnostic biosignatures that are unique to a specific production process.

    In this chapter, we examine the structural diversity of lipids in the cell and the factors that affect lipid composition during laboratory culturing with particular emphasis on Bacillus organisms. Methods used to extract, separate, and detect fatty acids are reviewed. In addition, the potential utility of lipid profiles in forensic investigations is discussed in the context of specific examples from the literature. Lastly, validation considerations and quality assurance are highlighted as important aspects in the implementation of lipid analysis for microbial forensics.”

    By way of additional background, Dr. Ivins and co-authors used both sheep blood agar and Leighton-Doi.

    http://www.freepatentsonline.com/6387665.html

    “On sheep blood agar plates containing heat shocked culture material from both sheep blood agar cultures and Leighton-Doi medium cultures, there was confluent growth.” See also 1995 Vaccine article describing method used to prepare the Ames used in testing the Ann Arbor biocidal and decontamination agent that looks like skim milk.

    The study Bruce co-authored with Pat and Mara, revising it October 2001, involved both sheep blood agar and Leighton-Doi

    “B. anthracis isolates Ames, Vollum 1B, Zimbabwe and Namibia were each inoculated onto 5% sheep blood agar and incubated overnight at 37 °C. The following day, an inoculum was prepared from each plate by suspending a loopful of growth into 5 ml of phosphate-buffered saline (PBS). Two-liter Erlenmeyer flasks containing 250 ml of Leighton–Doi [10] broth were inoculated with 0.2 ml of the cell suspension and incubated at 37 °C for 3 days with moderate shaking of 100 reciprocating strokes per minute. The spores were harvested by pelleting at 10,000×g, washed twice in sterile water for injection (McGaw, Inc., Irvine, CA), suspended in 1% phenol and stored at 4 °C. Just before challenge, the spores were heat shocked at 60 °C for 45 min. The appropriate dilution for challenge was made in sterile water for injection and prepared in a 0.2 ml dose.”

    See also studies submitted in 2000 and 2001.

    In vitro correlate of immunity in a rabbit model of inhalational anthrax Vaccine, Volume 19, Issue 32, 14 September 2001, Pages 4768-4773
    M. L. M. Pitt, S. F. Little, B. E. Ivins, P. Fellows, J. Barth, J. Hewetson, P. Gibbs, M. Dertzbaugh, A. M. Friedlander

    Efficacy of a human anthrax vaccine in guinea pigs, rabbits, and rhesus macaques against challenge by Bacillus anthracis isolates of diverse geographical origin Vaccine, Volume 19, Issues 23-24, 30 April 2001, Pages 3241-3247

    P. F. Fellows, M. K. Linscott, B. E. Ivins, M. L. M. Pitt, C. A. Rossi, P. H. Gibbs, A. M. Friedlander

  5. DXer said

    The Washington Post journalist who wrote that importance piece today is interviewed on NPR.

    And according to an NPR blog,

    “And Tuesday night, forensic science comes under more scrutiny in a documentary by PBS Frontline, working with ProPublica and the Investigative Reporting Program at the University of California, Berkeley.

    Their reporting looks at how flaws in fingerprint analysis led to the false arrest of an Oregon attorney, who found himself on trial for participating in the 2004 terrorist bombings in Spain that killed 191 people. After a judge dismissed the case against Brandon Mayfield, the FBI offered a rare apology.

    The Frontline documentary also looks at other dubious uses of forensic evidence, from the testimony of a “smell” expert in the trial of Casey Anthony, who was acquitted of the first-degree murder of her 2-year-old daughter, to a story NPR reported last year on how flawed bite-mark identification was used to convict two innocent men in Mississippi.”

    • DXer said

      In Amerithrax, investigators allowed a report to be promoted in the press that extensively relied upon the statements of part-time counselor who wrote a 2009 book explaining that her actions in 2000 and 2001 were controlled by an alien who had implanted a microchip in her butt. She would receive her instructions each night and would have emergency exorcisms to rid herself of the murderous astral entities that attached themselves to her clients such as Dr. Ivins. The FBI relied on her statements in concluding that Dr. Ivins was murderous. Not disclosing these facts — not withdrawing the EBAP’s reliance on the witness or the Amerithrax Investigative Summary’s reliance — constitutes extremely serious violation of the standards that govern. … and that’s whether you consider them constitutional, statutory or moral.

    • DXer said

      Here is a link to the webpage for the Frontline documentary, “The Real CSI”.

      http://www.pbs.org/wgbh/pages/frontline/real-csi/

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