Monday, January 5, 2009

NYT: Portrait Emerges of Anthrax Suspect’s Troubled Life

Scott Shane's newest, detailed exploration of Bruce Ivins can be summarized in Shane's statement, "unless new evidence were to surface, the enormous public investment in the case would appear to have yielded nothing more persuasive than a strong hunch, based on a pattern of damning circumstances, that Dr. Ivins was the perpetrator."

Let me make a few comments on this very detailed story.
...anthrax specialists who have not spoken out previously said that, contrary to some skeptics’ claims, Dr. Ivins had the equipment and expertise to make the powder in his laboratory.
He may well have--but this begs the bigger question of how long it would have taken and whether he could have done so without being detected.

And most importantly:
The science alone could not close the case. “We could get to a lab, to a refrigerator, to a flask,” said Dwight E. Adams, the F.B.I. laboratory director until 2006. “But that didn’t put the letters in anyone’s hand.”

As the bureau’s undercover informant, Dr. Haigwood struck up a breezy e-mail correspondence about scientific grants, pets and travel. Dr. Ivins complained about psychological screening and other “rather obnoxious and invasive measures” imposed at Fort Detrick since the anthrax attacks.
Dr. Nancy Haigwood believed Ivins was the culprit and had additionally, "damaged my property, he impersonated me and he stalked me.” She was afraid he would send her an anthrax letter. Her goal in renewing their friendship was to get him to incriminate himself. So any evidence resulting from their correspondence would need to be viewed from this perspective and in its entirety.

Dr. Ivins still carried resentment from four decades earlier at Lebanon High School in Ohio, where he had been a nerdy, awkward teenager devoted to photography and, even then, to the study of bacteria.
What percent of Ph.D. scientists did not feel nerdy, awkward and excluded as teens, I wonder? (I do not mean to target scientists. The teen years are awkward for most people.)

Though a public debate had raged for years over whether the mailed anthrax had been “weaponized” with sophisticated chemical additives, the F.B.I. had concluded early on that it was not.
The jury is still out on this issue, but many believe otherwise.

By 2004, secret scientific testing established that the mailed anthrax had been grown somewhere near Fort Detrick.
Presumably this comment refers to properties of the water used to grow anthrax; whether such analysis can reliably pinpoint the Frederick, Maryland area remains open to question.


Anonymous said...

I'd like to further comment on Meryl's last statement "Presumably this comment refers to properties of the water used to grow anthrax; whether such analysis can reliably pinpoint the Frederick, Maryland area remains open to question."

Scott Shane confusingly calls this a chemical signature. He is likely referring to isotopic analysis - which is more properly called a nuclear signature (the only chemical signature in the mailed spores is that of silicon - and that points AWAY from Detrick, since none of the original Detrick RMR-1029 spores contained silicon, but that's another story).

Back to the isotope analysis. It's true that their are regional (and seasonal) differences in local water - due to differences in the oxygen isotope concentration. But these differences can only locate very broad regions (such as, for example, the North East US versus the South West US). Shane writes as if the town of Frederick has it's own unique O isotope - not so. This was likely leaked from an anonymous FBI source and reminds me of the initial leaks that the FBI made about the printing error on the Federal Eagle stamp on the envelope only being sold at the same Post Office where Dr Ivins had a PO Box. That was incorrect, of course. These envelopes were sold all over Virginia and Maryland and probably other states. Still, the initial leak served the FBI's purpose of creating the framework they desired to create. I'm surprised Shane wrote about the water being specific to Detrick without bothering to check out the science first.

But the second and bigger problem with using O isotopes to trace spores is that the water used in their preparation only provides a fraction of the final oxygen atoms in the spores. The rest of the oxygen atoms come from dozens of other laboratory reagents, chemicals and nutrients - all of which are unknowns. For example, amino acids added to the spore broth could come from dozens of different sources - the oxygen isotope ratios in these would be all over the map. Same holds for other necessary chemicals such as NH4SO4, MgSO4, MnSO4, ZnSO4, CuSO4, FeSO4, CaCl2.2H2O, K2HPO4. All of these salts contain oxygen and all are needed to make anthrax spores. All of these oxygen atoms would end up being incorporated in the final spores, destroying any signature of the local water.
Labs that have been sponsored by the FBI to do work like this have themselves admitted the problem.
see paper here:

Conclusion: The anonymously leaked claim that isotopic analysis of the mailed spores can be traced to Detrick water sounds highly dubious. One cannot solve a 15 variable equation with 14 unknowns.

Anonymous said...

The Times article is troubling for a number of different reasons.

One problem is what is not said in the Times article. In particular, the Times article never mentions the alledgedly comprehensive investigation by FBI agents that eliminated everyone who had access to RMR-1029 but Dr. Ivins as the person who prepared the attack spores. Interviews, lab notebook investigations, background investigations, etc., of what the FBI claimed were over 100 different individuals (Ft. Dietrick personnel say the number is closer to 300). If the FBI had actually put in the man hours required for such a massive investigation of over 100 different individuals, how could that not be a part of the story of the Ivins' investigation? Yet there's not a single sentence, not one, reflecting this massive investigation. I suspect it never happened.

Instead, we are told that the FBI relied on "secret scientific testing" to eliminate "military and intelligence research programs in Utah or Ohio", (The Times review found that the F.B.I. had disproved the assertion, widespread among scientists who believe Dr. Ivins was innocent, that the anthrax might have come from military and intelligence research programs in Utah or Ohio. By 2004, secret scientific testing established that the mailed anthrax had been grown somewhere near Fort Detrick.). The Times article further states, "By early 2004, F.B.I. scientists had discovered that out of 60 domestic and foreign water samples, only water from Frederick, Md., had the same chemical signature as the water used to grow the mailed anthrax".

Now about those water samples, as the previous Anonymous poster pointed out the term "chemical signature" is confusing and worrisome. It's confusing in that the "chemistry" of water doesen't vary; except at one point the "F.B.I. scientists" claimed that it did vary in silicon content -- but then the "F.B.I. scientists" reversed course and said the silicon came from an "additive". So was the elimination of 59 of 60 domestic and foreign water sources based on silicon content, or on hydrogen and oxygen isotope ratios as the previous Anonymous poster suspects?

A careful reading of Stable Isotope Ratios and Forensic Analysis of Microorganisms, cited by the prior anonymous poster does indeed reveal a number of critical limitations in the applicability of that analysis. For example, the studies in that paper were limited to spores grown in liquid media, as opposed to agar plates. (We present data only for spores produced in liquid media because most of the 32 different media used here were liquid media, and the presence of agar is often associated with evaporative enrichment of the water in the medium (evaporation increases the relative amounts of 2H and 18O in the residual water [40]) (45), thereby affecting the O and H isotope ratios of spores.). More specifically, the studies were limited to spores grown in local tap water. (Approximately 70% of the oxygen atoms in B. subtilis spores originate in culture water, while only 30% of the hydrogen atoms do). Moreover, if a scientist uses purchased or partially distilled water to prepare spores, the analysis is meaningless. (Additionally, a perpetrator could have purchased water for culture production that was not isotopically linked to his or her geographic location, a possibility that must be considered even when a suspected production location has been identified. Finally, although deionization does not affect the hydrogen and oxygen isotope ratios of water, incomplete distillation does. If a perpetrator used incompletely distilled water, its isotope ratios could also be different from those of the local water from which it was derived.)

How could the "F.B.I. scientists" have possibly assumed that the anthrax attack spores were grown in local tap water, and nothing else?

But there's even a larger issue hidden in the all of the "water analysis" assumptions of the "F.B.I. scientists". From the various comments of anthrax experts, and the scientists who worked with Dr. Ivins, it appears that Ivins grew anthrax samples on agar plates, and not in liquid media. His lab apparently wasn't equipped to grow anthrax in liquid media.

Thus, an article published by The Baltimore Examiner, quotes W. Russell Byrne, a former director of the bacteriology division at Fort Detrick, who was familiar with Ivins' work and his lab, as saying, he "never believed Ivins’ could have produced the preparations used in the anthrax letters working in the bacteriology division area of Building 1425.”... "Nearly 1 gram per contaminated letter would have required months of intensive labor and hundreds of agar “plates,” on which the spores are grown, Byrne says.".

George Mason University professor and former Soviet bioweapons researcher Sergei Popov (who has also commented here), also distinguishes between the laboratory set up required for liquid fermentation spore growth verses agar plate spore growth. Thus the Baltimore Examiner article reports Dr. Popov's comments. Popov said that the only way the FBI scenario works is if someone else provided the spores to Ivins. “What if somebody fermented the spores for him?” he asks. “What’s in favor of this hypothesis is the presence of silica in the spores. This is a signature of a large-scale fermentation process.”

It thus appears quite possible that even aside from the numerous limitations of the "water analysis" issue, (which apparently played a large role in the F.B.I.'s investigation of Ivins), that "water analysis" was never applicable to Ivins' work at all, as his work didn't involve growing anthrax spores in water and his lab wasn't set up for liquid fermentation.

Anonymous said...

It's Anonymous poster 1 again. I'd like to follow up with some observations about the isotope issue based on what was posted by Anonymous 2.
As anonymous 2 pointed it, the paper I cited states that up to 80% of the oxygen atoms in B. subtilis spores may come from the water used in liquid preparations. The paper that determined that, by the same authors, is here:

That sounds impressive at first glance - but remember, this work used ideal conditions. The authors used all the same nutrients and salts from the same sources - there are at least a dozen of these and probably more (I listed some of the needed salts in my previous post). It's not surprising that they were able to trace the water in some cases, since they had already FIXED more than a dozen variables. When these many variables are NOT fixed, a completely different picture would emerge. Even one salt containing an unusually high or low delta 18^O would skew the results. So applying this to a sample of unkown nutrients and salts is a completely different ball game.

More interesting than that, the paper I link to in this post showed that they could not determine the water source in spores made in North Carolina and Ohio - the delta 18^O's were identical for the water.

Just for a laugh I used the publicly available water isotope database (here ) to determine the annual precipitation isotope ratios for 2 different grid locations (you can enter grid locations in a calculator).

Here are my results:

(1) Frederick, Maryland, 39N, 77W, altitude 90m
Delta 18^O = -7.6
Delta 2^H = -48

(2) Columbus, Ohio, 39N, 82W, altitude 300m
Delta 18^O = -7,6
Delta 2^H = -49

Not much difference here - identical oxygen and almost identical hydrogen.

That seems to say you can forget about isotope differences between water at Battelle and Frederick.

Anonymous said...

Anonymous 1 again. To follow up on the points I've been making, it's useful to look at a world map of oxygen 18 water levels. That can be seen here:

Notice that the North East US is all one color. That is consistent with the fact that the authors of this paper:

could not distinguish between spores grown in North Carolina versus spores grown in Ohio. They were, however, able to distinguish spores grown between Baton Rouge, LA; Los Alamos, NM; and Salt Lake City, UT. That is reflected in the regional variations seen in the map. Again, I stress that their study was ideal - they used common reagents for all the other oxygen 18 sources. Varying any of these would have killed this ideal study and created a confusing mess of measured 18^O in spores.

I'm cynical enough to imagine that when the FBI claim to have studied 60 different water sources that they cherry picked areas that were OUTSIDE of Detrick's region (this region is large and includes Battelle labs in Ohio). And then they used the same batches of starting materials (nutrients and salts) to ensure that only the water contributed to to the final delta 18^O's. This would be a meaningless exercise, of course, since it does not reflect reality.

Anonymous said...

Most of the material in RMR 1029 was fermentor grown by Dugway, very little material (in germs of volume of purified spores) was grown by Ivins in aqueous media, i.e. shaker flask system. Therefore, the stable oxygen, hydrogen isotopes in spores directly from RMR 1029 would reflect the isotope ratio of a region that includes Utah and not Maryland. The isotope region that includes Maryland also includes wide swaths of from Maine to Maryland and includes Ohio, West Virginia. So, where is the data that says stable isotope ratios can pinpoint Frederick Maryland. Sounds like over reaching to me.

Anonymous said...

“And anthrax specialists who have not spoken out previously said that, contrary to some skeptics’ claims to the contrary, Dr. Ivins had the equipment and expertise to make the powder in his laboratory.”

One of the most crucial aspects of the case is the equipment used for dry spore production. For starters, any lyophylizer used for this purpose would have been significantly contaminated with anthrax spores. It would have been identified early in the investigation by the FBI as the one and only smoking gun. But no evidence from any lyophylizer has been presented, a silence that speaks volumes.

In addition, it is certain that spores would have escaped from the lyophylizer during or after use for making the anthrax spore powder. The consequences of that are all too obvious: someone at USAMRIID gets sick with inhalational anthrax.

But that did not happen, in spite of the fact that the lyophylizer available to Bruce in the Bacteriology Division area is not in a BSL-3 containment area, and has not been in the containment area at any time in the last 10 years. The lyophylizer is situated next to a wide, well traveled hallway and in very close proximity to employee work spaces and non-containment labs. There is nothing to keep any aerosol escaping from that lyophylizer from disseminating throughout Building 1425, just as occurred in the Senate Office Building.

And that means, if you believe the powder was made by Dr. Ivins at the USAMRIID Bacteriology Division, that the same spores that killed people miles distant from the letters themselves did not infect one – not one - of the approximately 500 USAMRIID employees who worked in Building 1425, most of whom have never been immunized against anthrax.

In addition, if anyone had been so reckless as to use the lyophylizer in the Bacteriology Division area to produce dry anthrax spores, and lucky enough to infect no one who walked by, worked near, or worked with the lyophylizer, there would have been the task of cleaning up after the crime. The lyophylizer would have been dirty with anthrax spores after the job was done, and that would have to be fixed to avoid detection of the process.

But complete topical decontamination of the lyophylizer with, for instance, bleach, would have been impossible. The vacuum that draws water molecules out of the frozen material being lyophilized would have drawn the spores into the machine where they could not be reached by hand. The person making the powder would have known that.

It might have been possible to completely decontaminate the lyophylizer by paraformaldehyde aerosol in an airlock, but that could not have been done without attracting a lot of attention and suspicion.

Why do I say that? Stick with me for a minute, this is going to be a little tedious. First, the decontamination would have involved moving the lyophylizer into the airlock, an activity that would require several people and would have been noticed by a number of individuals, not the least of whom would have been the hand receipt holder, who would have wanted to know where the hell it was going. All government equipment that has purchase value above a nominal minimum is hand receipted to an individual. If that equipment is missing, the hand receipt holder is responsible, so employees pay close attention to equipment that they are hand receipted for. If a piece of equipment can’t be located, and it’s on your hand receipt, you pay for it. Not only that, you pay the price the government paid for it, not matter how old, beat up, or obsolete it is. Once a year, hand receipts have to be reconciled. If a piece of equipment on your hand receipt is missing, it comes out of your paycheck. Believe me, USAMRIID employees pay attention to equipment they are hand receipted for. If that lyophylizer had been moved, it could not have gone unnoticed.

But back to the decon required to cover up the crime: the airlocks are rooms about 10 feet square, with ceilings about 8 feet high and large steel doors on two sides, facing each other. The doors have small windows. One door leads into the BSL-3 containment suite, the other leads to the non-containment area.

The next step for the person who made the spore powder would be to request an airlock decontamination. Then the decon would be scheduled and subsequently performed by the Safety Office. The actual performance of the decon is usually about a 24 hour process. But when it's done, you don't just open the door and wheel your equipment out of the airlock when no one is looking. The decontamination has to be demonstrated to be effective enough to kill anthrax spores - no one touches anything until the Safety Office certifies the decon and OKs removal of the items in the airlock. All decons are done so that contaminated articles from the BSL-3 area can be safely transferred to non-containment areas - anywhere else, because the equipment now poses no risk. The whole process of scheduling and completing an airlock decon usually takes at least a week or two. It is serious business.

Then the lyophylizer would have been moved back into the non-containment area by the hallway from which it came. This would have made no sense and would certainly have attracted a lot of questions, about why it had been deconned and then moved right back where it came from.

Why do I say this? Because using the lyophylizer is not like making a cup of coffee - the runs take hours, or longer, frequently overnight. When it’s being used it is obvious. And people working in a lab with bugs that can kill you notice when experiments are being run or something different is happening, like using or deconning a piece of equipment for no apparent reason. The people who work in those labs have to know, for their own safety and everyone else's, what's happening, who’s doing it, and why. Someone else’s mistake can really hurt you, so you have to pay attention to what’s going on around you all the time.

The use of the lyophylizer would have been noticed, questioned, and remembered. There is absolutely no doubt about that.

But, of course, none of this happened – not the questionable/suspicious use of the lyophylizer, not the inexplicable movement of a large piece of equipment, not the decon. The general statements such as the one quoted above ignore vital specifics and details, to the point that such comments are meaningless at best, misleading at worst.

If the FBI had any case against Bruce, the lyophylizer used to produce the powder would have been quickly identified, after which the DNA evidence confirming the lyophylizer’s use for refinement of anthrax spores would have followed in short order. Keep in mind that even if you decontaminate a piece of equipment well enough to kill all the anthrax spores – and this is not easy to do - there will still be plenty of DNA left behind. Figuring out which equipment was used to make the anthrax spore powder would have been a very simple, straightforward task.

This aspect of the case is so fundamental that I am shocked that it has not already been thoroughly reviewed and discussed. There is no way around it: the smoking gun for the anthrax letters case is not at USAMRIID. That means it has to be somewhere else, and that it was done by someone else.

Anonymous said...

Here is a Fall 2008 presentation by the FBI's consulting scientists.

Integration of Stable Isotope and other Mass Spectral Data for Microbial Forensics
Kreuzer-Martin, H. W.; Jarman, K. H.

American Geophysical Union, Fall Meeting 2008, abstract #H51C-0812

The nascent field of microbial forensics requires the development of diverse signatures as indicators of various aspects of the production environment of microorganisms. We have characterized isotopic relationships between Bacillus subtilis ATCC 6051 spores and their growth environment, using as a database the carbon, nitrogen, oxygen and hydrogen stableisotope ratios of a total of 247 separate cultures of spores produced on a total of 32 different culture media. We have analyzed variation within individual samples, between cultures produced in tandem, and between cultures produced in the same medium but at different times in the context of using stable isotope ratios as a signature for sample matching. We have correlated the stable isotope ratios of carbon, nitrogen, oxygen, and hydrogen of growth medium nutrients or water and spores and show examples of how these relationships can be used to exclude nutrient or water samples as possible growth substrates for specific cultures. The power of stable isotope ratio data can be greatly enhanced by combining it with orthogonal datasets that speak to different aspects of an organism's production environment. We developed a Bayesian network that follows the causal relationship from culture medium recipe to spore elemental content as measured by secondary ion mass spectrometry (SIMS), carbon and nitrogen stable isotope ratios, and to the presence of residual agar by electrospray ionization MS (ESI-MS). The network was developed and tested on data from three replicate cultures of B. subtilis ATCC 49760 in broth and agar-containing versions of four different nutrient media. To test the network, data from SIMS analyses of B. subtilis 49760 produced in a different medium, from approximately 200 ESI MS analyses of B. thuringensis ATCC 58890 and B. anthracis Sterne grown in five additional media, and the stable isotope data from the 247 cultures of B. subtilis 6051 spores were used. This network was able to characterize Bacillus spores grown under multiple culture conditions with an error rate of less than 0.07 in characterizing carbon and nitrogen source, addition of metals, and presence of agar, and an error rate of 0.19 in characterizing the culture medium recipe. The integration of multiple analytical techniques allowed us to maximize the amount of information obtained from unknown source microorganisms. The Bayesian network approach allowed us to combine scientific understanding with well established statistical methodologies to characterize a microbe's growth environment without the need for reference signatures. Similar approaches could be applied to data from other scientific disciplines, as well as to other problems of attribution.

Anonymous said...

Thank you to the previous poster.

The isotope analysis techniques could certainly provide valuable information in appropriately controlled circumstances and environments. Nevertheless, its important to understand that this technique, as applied to the case of the anthrax attack spores, presumes that whoever prepared those spores used local tap water to prepare the growth medium.

I personally question whether there is any reasonable basis whatsoever for making such an assumption. Tap water varies from place to place, particularly in its pH and in the inclusion of different amounts of trace minerals. In "layman's languages", these differences are reflected in characterizations of tap water as "hard" or "soft". Because of the unknowns associated with tap water, in chemical and analytical labs, laboratory work is virtually always conducted using distilled or demineralized water purchased from an outside vendor. One would certainly expect the same to be true in the field of microbiology since tap water can contain bacteria and viral microbes in addition.

Even assuming for the sake of argument that some microbiological labs do use tap water, it would be quite a jump to assume that microbiology preparations are always made using tap water. But if the Times report is true, the analysis of anthrax attack spores would have been made based on precisely this assumption, and apparently numerous labs and individuals were eliminated as suspects on the basis of such questionable assumptions.

Anonymous said...

I do not think Battelle was responsible or complicit. But isn't any scientist at a Battelle-managed facility disqualified given that, for example, there is a $50 million dollar lawsuit against Battelle for negligence in allowing access to the pathogen?

Bruce Ivins reportedly gave Ames from the flask to Battelle, which had it at more than one location. See mid-August 2008 transcript referring to quasi-governmental entity that had it at more than location. (Battelle managed the life sciences lab at Dugway).

Question: If Battelle was a suspect in the anthrax mailings, am I correct that Bruce Ivins would not be allowed to manage the microbial forensic investigation? (or factual investigation)

Question: If Bruce Ivins was a suspect in the anthrax mailings, am I correct that Battelle would not be allowed to manage the microbial forensic investigation? (or factual investigation)

Relatedly, would a scientific study and associated expert testimony by a scientist at a Battelle-managed facility be subject to a motion to strike on the grounds of a conflict of interest?

The Journal of Philosophy, Science & Law
Volume 7, April 16, 2007

Conflicts of Interest in Scientific Research
Related to Regulation or Litigation

David B. Resnik, JD, PhD*

* Bioethicist at the National Institute of Environmental Health Sciences (NIEHS)
"To have a better understanding of our subject matter, it will be useful to set forth some key definitions. To frame the discussion, let’s consider some cases that most people agree would constitute a conflict of interest:

• A father is asked to referee his daughter’s soccer game;
• A judge presides over a trial involving a close friend;
• A researcher owns $50,000 worth of stock in a medical device company that makes a product that he is testing;
• A pharmaceutical company invites a physician to an all-expense-paid seminar in the Aspen, Colorado to learn about its products;
• A county commissioner votes on a zoning law that will increase the value of his land by $10,000;
• A woman, who is adamantly opposed to capital punishment, sits on a jury in a murder case in which the defendant could receive the death penalty;
• A scientist is asked to review an article that reports results that could refute a hypothesis that she has defended for over twenty years.

In each of these cases, a person has interests that may interfere with the performance of his or her ethical or legal duties. In case 1, the father has a duty to referee impartially, but he may have a difficult time doing this because his daughter is on one of the teams. In case 4, the physician has a duty to prescribe medications to promote the health of his patients, but he may be tempted to prescribe medications to promote the interests of the drug company. The interests in several of the cases (cases 3-5) are financial interests, but the interests also include personal interests (case 1 and 2), political interests (case 6), and professional interests (case 7). "

Anonymous said...

Bayesian-Integrated Microbial Forensics
Appl Environ Microbiol. 2008 June; 74(11): 3573–3582.
[Battelle-managed] Pacific Northwest National Laboratory, P.O. Box 999, Richland, WA 99352

*Corresponding author. Mailing address: Pacific Northwest National Laboratory, [Battelle Boulevard], Richland, WA 99352.

In the aftermath of the 2001 anthrax letters, researchers have been exploring ways to predict the production environment of unknown-source microorganisms. Culture medium, presence of agar, culturing temperature, and drying method are just some of the broad spectrum of characteristics an investigator might like to infer. The effects of many of these factors on microorganisms are not well understood, but the complex way in which microbes interact with their environments suggests that numerous analytical techniques measuring different properties will eventually be needed for complete characterization. In this work, we present a Bayesian statistical framework for integrating disparate analytical measurements. We illustrate its application to the problem of characterizing the culture medium of Bacillus spores using three different mass spectral techniques. The results of our study suggest that integrating data in this way significantly improves the accuracy and robustness of the analyses.

Would the FBI and US DOJ have been relying on the work of scientists at the Battelle-run PNNL to convict Bruce Ivins in a court of law? Do they plan on relying on the work of scientists at the Battelle-run PNNL to convict Bruce Ivins in the court of public opinion? Then I hereby move in the court of public opinion to strike from the record all evidence implicating Bruce Ivins done by scientists at labs managed by Battelle Memorial Institute. (I assume the good faith of all the scientists and Battelle -- a disqualifying conflict of interest exists even assuming good faith as I do).