Anthrax has been described as a veterinary disease of minor importance to clinical medicine, causing occasional occupational infections in single cases or clusters. Its potential for rapid and widespread epidemic transmission under natural circumstances has not been widely appreciated. A little-known 1770 epidemic that killed 15,000 people in Saint-Domingue (modern Haiti) was probably intestinal anthrax. The epidemic spread rapidly throughout the colony in association with consumption of uncooked beef. Large-scale, highly fatal epidemics of anthrax may occur under unusual but natural circumstances. Historical information may not only provide important clues about epidemic development but may also raise awareness about bioterrorism potential.

We used unpublished reports, published manuscripts, and communication with investigators to identify and summarize 49 anthrax-related epidemiologic field investigations conducted by the Centers for Disease Control and Prevention from 1950 to August 2001. Of 41 investigations in which Bacillus anthracis caused human or animal disease, 24 were in agricultural settings, 11 in textile mills, and 6 in other settings. Among the other investigations, two focused on building decontamination, one was a response to bioterrorism threats, and five involved other causes. Knowledge gained in these investigations helped guide the public health response to the October 2001 intentional release of B. anthracis, especially by addressing the management of anthrax threats, prevention of occupational anthrax, use of antibiotic prophylaxis in exposed persons, use of vaccination, spread of B. anthracis spores in aerosols, clinical diagnostic and laboratory confirmation methods, techniques for environmental sampling of exposed surfaces, and methods for decontaminating buildings.

In October 2001, the first inhalational anthrax case in the United States since 1976 was identified in a media company worker in Florida. A national investigation was initiated to identify additional cases and determine possible exposures to Bacillus anthracis. Surveillance was enhanced through health-care facilities, laboratories, and other means to identify cases, which were defined as clinically compatible illness with laboratory-confirmed B. anthracis infection. From October 4 to November 20, 2001, 22 cases of anthrax (11 inhalational, 11 cutaneous) were identified; 5 of the inhalational cases were fatal. Twenty (91%) case-patients were either mail handlers or were exposed to worksites where contaminated mail was processed or received. B. anthracis isolates from four powder-containing envelopes, 17 specimens from patients, and 106 environmental samples were indistinguishable by molecular subtyping. Illness and death occurred not only at targeted worksites, but also along the path of mail and in other settings. Continued vigilance for cases is needed among health-care providers and members of the public health and law enforcement communities.

On October 4, 2001, we confirmed the first bioterrorism-related anthrax case identified in the United States in a resident of Palm Beach County, Florida. Epidemiologic investigation indicated that exposure occurred at the workplace through intentionally contaminated mail. One additional case of inhalational anthrax was identified from the index patient’s workplace. Among 1,076 nasal cultures performed to assess exposure, Bacillus anthracis was isolated from a co-worker later confirmed as being infected, as well as from an asymptomatic mail-handler in the same workplace. Environmental cultures for B. anthracis showed contamination at the workplace and six county postal facilities. Environmental and nasal swab cultures were useful epidemiologic tools that helped direct the investigation towards the infection source and transmission vehicle. We identified 1,114 persons at risk and offered antimicrobial prophylaxis.

The index case of inhalational anthrax in October 2001 was in a man who lived and worked in Florida. However, during the 3 days before illness onset, the patient had traveled through North Carolina, raising the possibility that exposure to Bacillus anthracis spores could have occurred there. The rapid response in North Carolina included surveillance among hospital intensive-care units, microbiology laboratories, medical examiners, and veterinarians, and site investigations at locations visited by the index patient to identify the naturally occurring or bioterrorism-related source of his exposure.

On October 15, 2001, a U.S. Senate staff member opened an envelope containing Bacillus anthracis spores. Chemoprophylaxis was promptly initiated and nasal swabs obtained for all persons in the immediate area. An epidemiologic investigation was conducted to define exposure areas and identify persons who should receive prolonged chemoprophylaxis, based on their exposure risk. Persons immediately exposed to B. anthracis spores were interviewed; records were reviewed to identify additional persons in this area. Persons with positive nasal swabs had repeat swabs and serial serologic evaluation to measure antibodies to B. anthracis protective antigen (anti-PA). A total of 625 persons were identified as requiring prolonged chemoprophylaxis; 28 had positive nasal swabs. Repeat nasal swabs were negative at 7 days; none had developed anti-PA antibodies by 42 days after exposure. Early nasal swab testing is a useful epidemiologic tool to assess risk of exposure to aerosolized B. anthracis. Early, wide chemoprophylaxis may have averted an outbreak of anthrax in this population.

On October 12, 2001, two envelopes containing Bacillus anthracis spores passed through a sorting machine in a postal facility in Washington, D.C. When anthrax infection was identified in postal workers 9 days later, the facility was closed. To determine if exposure to airborne B. anthracis spores continued to occur, we performed air sampling around the contaminated sorter. One CFU of B. anthracis was isolated from 990 L of air sampled before the machine was activated. Six CFUs were isolated during machine activation and processing of clean dummy mail. These data indicate that an employee working near this machine might inhale approximately 30 B. anthracis-containing particles during an 8-h work shift. What risk this may have represented to postal workers is not known, but the risk is approximately 20-fold less than estimates of sub-5 micron B. anthracis-containing particles routinely inhaled by asymptomatic, unvaccinated workers in a goat-hair mill.

At least four Bacillus anthracis–containing envelopes destined for New York City and Washington, D.C., were processed at the Trenton Processing and Distribution Center (PDC) on September 18 and October 9, 2001. When cutaneous anthrax was confirmed in a Trenton postal worker, the PDC was closed. Four cutaneous and two inhalational anthrax cases were identified. Five patients were hospitalized; none died. Four were PDC employees; the others handled or received mail processed there. Onset dates occurred in two clusters following envelope processing at the PDC. The attack rate among the 170 employees present when the B. anthracis–containing letters were sorted on October 9 was 1.2%. Of 137 PDC environmental samples, 57 (42%) were positive. Five (10%) of 50 local post offices each yielded one positive sample. Cutaneous or inhalational anthrax developed in four postal employees at a facility where B. anthracis–containing letters were processed. Cross-contaminated mail or equipment was the likely source of infection in two other case-patients with cutaneous anthrax.

After reports of the intentional release of Bacillus anthracis in the United States, epidemiologists, laboratorians, and clinicians around the world were called upon to respond to widespread political and public concerns. To respond to inquiries from other countries regarding anthrax and bioterrorism, the Centers for Disease Control and Prevention established an international team in its Emergency Operations Center. From October 12, 2001, to January 2, 2002, this team received 130 requests from 70 countries and 2 territories. Requests originated from ministries of health, international organizations, and physicians and included subjects ranging from laboratory procedures and clinical evaluations to assessments of environmental and occupational health risks. The information and technical support provided by the international team helped allay fears, prevent unnecessary antibiotic treatment, and enhance laboratory-based surveillance for bioterrorism events worldwide.

A two-component direct fluorescent-antibody (DFA) assay, using fluorescein-labeled monoclonal antibodies specific to the Bacillus anthracis cell wall (CW-DFA) and capsule (CAP-DFA) antigens, was evaluated and validated for rapid identification of B. anthracis. We analyzed 230 B. anthracis isolates; 228 and 229 were positive by CW-DFA and CAP-DFA assays, respectively. We also tested 56 non–B. anthracis strains; 10 B. cereus and 2 B. thuringiensis were positive by the CW-DFA assay, and 1 B. megaterium strain was positive by CAP-DFA. Analysis of the combined DFA results identified 227 of 230 B. anthracis isolates; all 56 strains of the other Bacillus spp. were negative. Both DFA assays tested positive on 14 of 26 clinical specimens from the 2001 anthrax outbreak investigation. The two-component DFA assay is a sensitive, specific, and rapid confirmatory test for B. anthracis in cultures and may be useful directly on clinical specimens.

In October 2001, four cases of inhalational anthrax occurred in workers in a Washington, D.C., mail facility that processed envelopes containing Bacillus anthracis spores. We reviewed the envelopes’ paths and obtained exposure histories and nasal swab cultures from postal workers. Environmental sampling was performed. A sample of employees was assessed for antibody concentrations to B. anthracis protective antigen. Case-patients worked on nonoverlapping shifts throughout the facility. Environmental sampling showed diffuse contamination of the facility, suggesting multiple aerosolization events. Potential workplace exposures were similar for the case-patients and the sample of workers. All nasal swab cultures and serum antibody tests were negative. Available tools could not identify subgroups of employees at higher risk for exposure or disease. Prophylaxis was necessary for all employees. To protect postal workers against bioterrorism, measures to reduce the risk of occupational exposure are necessary.

In October 2001, two inhalational anthrax and four cutaneous anthrax cases, resulting from the processing of Bacillus anthracis–containing envelopes at a New Jersey mail facility, were identified. Subsequently, we initiated stimulated passive hospital-based and enhanced passive surveillance for anthrax-compatible syndromes. From October 24 to December 17, 2001, hospitals reported 240,160 visits and 7,109 intensive-care unit admissions in the surveillance area (population 6.7 million persons). Following a change to reporting criteria on November 8, the average of possible inhalational anthrax reports decreased 83% from 18 to 3 per day; the proportion of reports requiring follow-up increased from 37% (105/286) to 41% (47/116). Clinical follow-up was conducted on 214 of 464 possible inhalational anthrax patients and 98 possible cutaneous anthrax patients; 49 had additional laboratory testing. No additional cases were identified. To verify the limited scope of the outbreak, surveillance was essential, though labor-intensive. The flexibility of the system allowed interim evaluation, thus improving surveillance efficiency.

On November 19, 2001, a case of inhalational anthrax was identified in a 94-year-old Connecticut woman, who later died. We conducted intensive surveillance for additional anthrax cases, which included collecting data from hospitals, emergency departments, private practitioners, death certificates, postal facilities, veterinarians, and the state medical examiner. No additional cases of anthrax were identified. The absence of additional anthrax cases argued against an intentional environmental release of Bacillus anthracis in Connecticut and suggested that, if the source of anthrax had been cross-contaminated mail, the risk for anthrax in this setting was very low. This surveillance system provides a model that can be adapted for use in similar emergency settings.

On November 11, 2001, following the bioterrorism-related anthrax attacks, the U.S. Postal Service collected samples at the Southern Connecticut Processing and Distribution Center; all samples were negative for Bacillus anthracis. After a patient in Connecticut died from inhalational anthrax on November 19, the center was sampled again on November 21 and 25 by using dry and wet swabs. All samples were again negative for B. anthracis. On November 28, guided by information from epidemiologic investigation, we sampled the site extensively with wet wipes and surface vacuum sock samples (using HEPA vacuum). Of 212 samples, 6 (3%) were positive, including one from a highly contaminated sorter. Subsequently B. anthracis was also detected in mail-sorting bins used for the patient’s carrier route. These results suggest cross-contaminated mail as a possible source of anthrax for the inhalational anthrax patient in Connecticut. In future such investigations, extensive sampling guided by epidemiologic data is imperative.

After public notification of confirmed cases of bioterrorism-related anthrax, the Centers for Disease Control and Prevention’s Emergency Operations Center responded to 11,063 bioterrorism-related telephone calls from October 8 to November 11, 2001. Most calls were inquiries from the public about anthrax vaccines (58.4%), requests for general information on bioterrorism prevention (14.8%), and use of personal protective equipment (12.0%); 882 telephone calls (8.0%) were referred to the state liaison team for follow-up investigation. Of these, 226 (25.6%) included reports of either illness clinically confirmed to be compatible with anthrax or direct exposure to an environment known to be contaminated with Bacillus anthracis. The remaining 656 (74.4%) included no confirmed illness but reported exposures to “suspicious” packages or substances or the receipt of mail through a contaminated facility. Emergency response staff must handle high call volumes following suspected or actual bioterrorist attacks. Standardized health communication protocols that address contact with unknown substances, handling of suspicious mail, and clinical evaluation of suspected cases would allow more efficient follow-up investigations of clinically compatible cases in high-risk groups.

In 2001, the intentional release of anthrax spores in the eastern United States increased concern about exposure to anthrax nationwide, and residents of Idaho sought assistance. Response from state and local agencies was required, increasing the strain on epidemiologists, laboratorians, and communications personnel. In late 2001, Idaho’s public health communications system handled 133 calls about suspicious powders. For each call, a multiagency bridge call was established, and participants (public health officials, epidemiologists, police, Federal Bureau of Investigation personnel, hazardous materials officials, and others) determined which samples would be tested by the state public health laboratory. A triage system for calls helped relieve the burden on public safety and health systems.

In October 2001, the greater New York City Metropolitan Area was the scene of a bioterrorism attack. The scale of the public response to this attack was not foreseen and threatened to overwhelm the Bioterrorism Response Laboratory’s (BTRL) ability to process and test environmental samples. In a joint effort with the Centers for Disease Control and Prevention and the cooperation of the Department of Defense, a massive effort was launched to maintain and sustain the laboratory response and return test results in a timely fashion. This effort was largely successful. The development and expansion of the facility are described, as are the special needs of a BTRL. The establishment of a Laboratory Bioterrorism Command Center and protocols for sample intake, processing, reporting, security, testing, staffing, and quality assurance and quality control are also described.

The bioterrorism-associated human anthrax epidemic in the fall of 2001 highlighted the need for a sensitive, reproducible, and specific laboratory test for the confirmatory diagnosis of human anthrax. The Centers for Disease Control and Prevention developed, optimized, and rapidly qualified an enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G (IgG) antibodies to Bacillus anthracis protective antigen (PA) in human serum. The qualified ELISA had a minimum detection limit of 0.06 µg/mL, a reliable lower limit of detection of 0.09 µg/mL, and a lower limit of quantification in undiluted serum specimens of 3.0 µg/mL anti-PA IgG. The diagnostic sensitivity of the assay was 97.8%, and the diagnostic specificity was 94.2%. A competitive inhibition anti-PA IgG ELISA was also developed to enhance diagnostic specificity to 100%. The anti-PA ELISAs proved valuable for the confirmation of cases of cutaneous and inhalational anthrax and evaluation of patients in whom the diagnosis of anthrax was being considered.

Molecular subtyping of Bacillus anthracis played an important role in differentiating and identifying anthrax strains during the 2001 bioterrorism-associated outbreak. Because B. anthracis has a low level of genetic variability, only a few subtyping methods, with varying reliability, exist. We initially used multiple-locus variable-number tandem repeat analysis (MLVA) to subtype 135 B. anthracis isolates associated with the outbreak. All isolates were determined to be of genotype 62, the same as the Ames strain used in laboratories. We sequenced the protective antigen gene (pagA) from 42 representative outbreak isolates and determined they all had a pagA sequence indistinguishable from the Ames strain (PA genotype I). MLVA and pagA sequencing were also used on DNA from clinical specimens, making subtyping B. anthracis possible without an isolate. Use of high-resolution molecular subtyping determined that all outbreak isolates were indistinguishable by the methods used and probably originated from a single source. In addition, subtyping rapidly identified laboratory contaminants and non-outbreak–related isolates.

In a bioterrorism event, a tool is needed to rapidly differentiate Bacillus anthracis from other closely related spore-forming Bacillus species. During the recent outbreak of bioterrorism-associated anthrax, we sequenced the 16S rRNA generom these species to evaluate the potential of 16S rRNA gene sequencing as a diagnostic tool. We found eight distinct 16S types among all 107 16S rRNA gene seqs fuences that differed from each other at 1 to 8 positions (0.06% to 0.5%). All 86 B. anthracis had an identical 16S gene sequence, designated type 6; 16S type 10 was seen in all B. thuringiensis strains; six other 16S types were found among the 10 B. cereus strains. This report describes the first demonstration of an exclusive association of a distinct 16S sequence with B. anthracis. Consequently, we were able to rapidly identify suspected isolates and to detect the B. anthracis 16S rRNA gene directly from culture-negative clinical specimens from seven patients with laboratory-confirmed anthrax.

We collected data during postexposure antimicrobial prophylaxis campaigns and from a prophylaxis program evaluation 60 days after start of antimicrobial prophylaxis involving persons from six U.S. sites where Bacillus anthracis exposures occurred. Adverse events associated with antimicrobial prophylaxis to prevent anthrax were commonly reported, but hospitalizations and serious adverse events as defined by Food and Drug Administration criteria were rare. Overall adherence during 60 days of antimicrobial prophylaxis was poor (44%), ranging from 21% of persons exposed in the Morgan postal facility in New York City to 64% of persons exposed at the Brentwood postal facility in Washington, D.C. Adherence was highest among participants in an investigational new drug protocol to receive additional antibiotics with or without anthrax vaccine—a likely surrogate for anthrax risk perception. Adherence of <60 days was not consistently associated with adverse events.

After inhalational anthrax was diagnosed in a Connecticut woman on November 20, 2001, postexposure prophylaxis was recommended for postal workers at the regional mail facility serving the patient’s area. Although environmental testing at the facility yielded negative results, subsequent testing confirmed the presence of Bacillus anthracis. We distributed questionnaires to 100 randomly selected postal workers within 20 days of initial prophylaxis. Ninety-four workers obtained antibiotics, 68 of whom started postexposure prophylaxis and 21 discontinued. Postal workers who stopped or never started taking prophylaxis cited as reasons disbelief regarding anthrax exposure, problems with adverse events, and initial reports of negative cultures. Postal workers with adverse events reported predominant symptoms of gastrointestinal distress and headache. The influence of these concerns on adherence suggests that communication about risks of acquiring anthrax, education about adverse events, and careful management of adverse events are essential elements in increasing adherence.

In October 2001, two envelopes containing Bacillus anthracis spores were processed at the Washington, D.C., Processing and Distribution Center of the U.S. Postal Service; inhalational anthrax developed in four workers at this facility. More than 2,000 workers were advised to complete 60 days of postexposure prophylaxis to prevent inhalational anthrax. Interventions to promote adherence were carried out to support workers, and qualitative information was collected to evaluate our interventions. A quantitative survey was administered to a convenience sample of workers to assess factors influencing adherence. No anthrax infections developed in any workers involved in the interventions or interviews. Of 245 workers, 98 (40%) reported full adherence to prophylaxis, and 45 (18%) had completely discontinued it. Experiencing adverse effects to prophylaxis, anxiety, and being <45 years old were risk factors for discontinuing prophylaxis. Interventions, especially frequent visits by public health staff, proved effective in supporting adherence.

During an investigation conducted December 17–20, 2001, we collected environmental samples from a U.S. postal facility in Washington, D.C., known to be extensively contaminated with Bacillus anthracis spores. Because methods for collecting and analyzing B. anthracis spores have not yet been validated, our objective was to compare the relative effectiveness of sampling methods used for collecting spores from contaminated surfaces. Comparison of wipe, wet and dry swab, and HEPA vacuum sock samples on nonporous surfaces indicated good agreement between results with HEPA vacuum and wipe samples. However, results from HEPA vacuum sock and wipe samples agreed poorly with the swab samples. Dry swabs failed to detect spores >75% of the time they were detected by wipe and HEPA vacuum samples. Wipe samples collected after HEPA vacuum samples and HEPA vacuum samples after wipe samples indicated that neither method completely removed spores from the sampled surfaces.

The biological attacks with powders containing Bacillus anthracis sent through the mail during September and October 2001 led to unprecedented public health and law enforcement investigations, which involved thousands of investigators from federal, state, and local agencies. Following recognition of the first cases of anthrax in Florida in early October 2001, investigators from Centers for Disease Control and Prevention (CDC) and the Federal Bureau of Investigation (FBI) were mobilized to assist investigators from state and local public health and law enforcement agencies. Although public health and criminal investigations have been conducted in concert in the past, the response to the anthrax attacks required close collaboration because of the immediate and ongoing threat to public safety. We describe the collaborations between CDC and FBI during the investigation of the 2001 anthrax attacks and highlight the challenges and successes of public health and law enforcement collaborations in general.