Plague: Current, comprehensive information on pathogenesis, microbiology, epidemiology, diagnosis, and treatment

Last updated March 24, 2009

Agent and Pathogenesis
Epidemiology
Plague as a Biological Weapon
Clinical Syndromes and Differential Diagnosis
Clinical Laboratory Testing
Environmental Testing
Treatment, Postexposure Prophylaxis, and Vaccines
Hospital Infection Control (Including Autopsies and Burial)
Public Health Reporting and Case Definitions
Images
References

Agent and Pathogenesis

Agent

Key microbiologic characteristics of Yersinia pestis include the following (see References: CDC/ASM/APHL 2002; Sneath 1986):

Pleomorphic gram-negative bacillus (1.0 to 2.0 mcm x 0.5 mcm); single cells or short chains in direct smears
Bipolar ("closed safety pin") staining with Giemsa, Wright's, or Wayson stains (may not be visible on Gram stain)
Facultative anaerobe
Nonmotile, nonsporulating
Non–lactose fermenter
Slow-growing in culture (colonies are pinpoint after 24 hours on sheep blood agar [SBA] and much smaller than other Enterobacteriaceae growing for 24 hours on SBA; colonies may not be visible on MacConkey or eosin methylene blue agar at 24 hours)
Catalase-positive, oxidase- and urease-negative (rarely, strains may be urease-positive)
Optimal growth at 28°C
"Stalactite pattern" in broth culture with clumps of cells from the side of the tube settling to the bottom if disturbed
At 48 to 72 hours of incubation on solid media, colonies have a raised, irregular, "fried egg" appearance under 4x enlargement, which becomes more pronounced as the culture ages; colonies also have been described as having a "hammered copper" shiny surface
Alkaline slant/acid butt (K/A) on triple sugar iron agar (TSI) without gas or H₂S
Generally susceptible to tetracyclines, chloramphenicol, aminoglycosides, sulfonamides (with or without trimethoprim), and fluoroquinolone antibiotics

Y pestis is divided into three classic biovars (see References: Dennis 1997).

Biovar antiqua (Africa, southeastern Russia, central Asia; thought to be the cause of the first pandemic)
Biovar medievalis (Caspian Sea; thought to be the cause of the second pandemic)
Biovar orientalis (Asia, Western Hemisphere; cause of the third pandemic)

Other classification and diversity information:

A nonvirulent strain, microtus, has been proposed as a fourth biovar (see References: Zhou 2004).
Y pestis is thought to have evolved from Yersinia pseudotuberculosis 1,500 to 20,000 years ago, and the two species remain closely related (see References: Achtman 2000). Whole-genome sequence comparisons have identified 32 chromosomal genes and 2 plasmids in Y pestis but not Y pseudotuberculosis (see References: Chain 2004).
The complete genomes of several strains have been sequenced and are available online (see References: National Center for Biotechnology Information, Parkhill 2001, Song 2004, Zhou 2002).

Pathogenesis

Virulence Factors

Virulence factors for Y pestis are primarily encoded on the chromosome and on three plasmids (the Pst plasmid, the Lcr plasmid, and the pFra plasmid) (see References: Dennis 1997).

The major virulence factors for Y pestis are responsible for the following activities (see References: Dennis 1997, McGovern 1997, Perry 1997, Titball 2003):

The ability of Y pestis organisms to adhere to cell surfaces is a key step in pathogenesis. Irreversible binding to host cell receptors via adhesins allows the organisms to then penetrate the cell surfaces (see References: Zhou 2006).
The F1 antigen is antiphagocytic, elicits a humoral response, and is a target for immunologic-based diagnostic tests. Most pathogenic Y pestis strains isolated from humans contain the F1 antigen.
Plasminogen activator (Pla) is a protease that appears to degrade fibrin and other extracellular proteins and to facilitate systemic spread from the inoculation site. Expression of Pla allows Y pestis to replicate rapidly in the airways. Pla is essential for Y pestis to cause primary pneumonic plague but is less important for dissemination during pneumonic than bubonic plague (see References: Lathem 2007).
The V and W antigens (produced at 37°C) cause the organisms to be resistant to phagocytosis; the V antigen is important for survival of Y pestis in macrophages.
Yersinia outer proteins (Yops) have a variety of activities, including inhibiting phagocytosis, inhibiting platelet aggregation, and preventing an effective inflammatory response.
Lipopolysaccharide (LPS) endotoxin (encoded on the chromosome) causes the classic features of endotoxic shock. LPS consists of three domains: the hydrophobic membrane anchor (lipid A), the surface-exposed O-antigen polysaccharide, and the core sugar region connecting the other two. Most of the effects of LPS are caused by lipid A (LPS-lipid A) (see References: Zhou 2006).
Phospholipase D (PLD) allows the bacilli to survive in the flea midgut.
Yersinia murine toxin (Ymt) is one of the factors required for maintaining Y pestis in fleas. Ymt is highly toxic for mice and rats but less active in other animals (see References: Zhou 2006).

Bubonic Plague

After a flea initially ingests Y pestis, the organisms elaborate a coagulase that clots ingested blood in the proventriculus (an organ between the esophagus and stomach) of the flea, thus blocking passage of the next blood meal into the flea's stomach. Fleas with this blockage regurgitate Y pestis into the bite wound while attempting to feed (see References: Perry 1997).

In a recent study, unblocked fleas given a single infectious blood meal transmitted Y pestis for up to 7 days following the meal and fleas given a "booster" infectious blood meal 5 days after the first meal transmitted Y pestis for the full 9-day duration of the study (see References: Eisen 2007: Temporal dynamics of early-phase transmission of Yersinia pestis by unblocked fleas).
Such research may explain mathematical models that predict rapidly spreading epizootics and epidemics (see References: Eisen 2006: Early-phase transmission), as well as discrepancies observed in plague epizootics among prairie dogs (see References: Webb 2006).

Between 25,000 and 100,000 Y pestis organisms are inoculated into the skin via the bite of an infected flea (see References: Reed 1970).
A papule, vesicle, pustule, or furuncle may occur at the site of the fleabite but is noted in less than 10% of patients (see References: Dennis 1997).
The organisms migrate through the cutaneous lymphatics to regional lymph nodes. Comparative studies in mice reveal that Y pestis virulence is associated with a distinct ability to massively infiltrate the draining lymph node without inducing an organized polymorphonuclear cell reaction (see References: Guinet 2008).
Once in the lymph nodes, they are phagocytized by polymorphonuclear leukocytes (PMNs) and mononuclear phagocytes. Organisms that are phagocytized by PMNs generally are destroyed, whereas those phagocytized by mononuclear cells proliferate intracellularly and develop resistance to further phagocytosis (see References: Perry 1997). These organisms are released when cell lysis occurs.
Initially, a thick, proteinaceous exudate that includes plague bacilli, PMNs, lymphocytes, and fewer macrophages can be found in affected nodes (see References: Dennis 1997).
Subsequently, the exudative pattern gives way to lakes of hemorrhagic necrosis, which obliterate the underlying lymph node architecture. A ground-glass amphophilic material that represents masses of bacilli may be present (see References: CDC 2004).
The inflammatory process creates swollen painful buboes and surrounding edematous tissues that are characteristic of bubonic plague.
The organisms often enter the bloodstream, causing hemorrhagic lesions in other lymph nodes and in organs throughout the body (initially the liver and spleen). Findings from a recent study using a mouse model suggest that the organisms replicate in splenic macrophages during the later stages of infection (see References: Lukaszewski 2005).
Septicemia, disseminated intravascular coagulation (DIC), and shock can ensue.
Unless treated promptly with appropriate antibiotic therapy, death usually results from overwhelming sepsis.

Pneumonic Plague

Y pestis can enter the lungs either through direct inhalation (primary pneumonic plague) or through hematogenous spread as a complication of bubonic or septicemic plague (secondary pneumonic plague).
Primary pneumonic plague is acquired naturally by inhaling respiratory droplets from infected humans or animals (such as cats).
The infectious dose by inhalation is estimated to be 100 to 500 organisms (see References: Franz 1997).
Marked intra-alveolar edema and congestion of the lungs are common (see References: CDC 2004). Pulmonary lesions include areas of central exudate with peripheral congestion. This pattern initially is lobular, but usually progresses to lobar consolidation (see References: Dennis 1997).
Distinguishing primary pneumonic plague from secondary hematogenous spread to the lungs can be difficult. Features that occur more commonly with primary pneumonic plague include the following (see References: Dennis 1997):

Tracheal and bronchial mucosal hemorrhages
Fibrinous pleuritis and subpleural hemorrhages overlying areas of exudative pneumonia
Less inflammation and necrosis and more exudation in lobular foci of the parenchyma
Foci of pneumonia along medium and large bronchi
More involvement of hilar lymph nodes
Less severe evidence of disease in organs other than the lungs, if such evidence is present

In primary pneumonic plague, as with bubonic plague, organisms often enter the bloodstream and cause multiorgan involvement, DIC, and shock.
In the absence of early antibiotic therapy (ie, within the first 24 hours), death occurs from overwhelming sepsis (usually within several days after illness onset). Without therapy, mortality approaches 100%.

Septicemic Plague

Primary septicemic plague is defined as systemic toxicity caused by Y pestis infection but without apparent preceding lymph node involvement. Secondary septicemic plague occurs commonly with either bubonic or primary pneumonic plague.
In primary septicemic plague, Y pestis organisms can disseminate from a fleabitesite through thelymphatic system (but without clinically apparent involvement of the lymph nodes), directly through the circulatory system, orboth (see References: Sebbane 2006).
Septicemic plague causes sepsis syndrome with multiorgan involvement, DIC, and shock. In the late stages of infection, high-density bacteremia often occurs, leading to identification of organisms on peripheral blood smears (see References: Butler 1991).
Spleen, liver, kidneys, skin, and brain are the most commonly affected organs. Meningitis can occur and is characterized by a thick, yellow, fibrinous-purulent exudate. Foci of necrosis with hemorrhage are common, as are characteristic lesions of DIC (such as fibrin thrombi in glomerular capillaries or purpuric skin lesions) (see References: Dennis 1997).

Y pestis may persist in necrotic tissues after antibiotic treatment despite negative blood cultures. Presumably, Y pestis becomes trapped in hypoperfused tissues and is able to persist because of: (1) inadequate delivery of antibiotics to affected areas and (2) the ability of the organisms to overcome local host defenses (see References: Guarner 2005).

Epidemiology

Reservoirs/Vectors/Modes of Transmission

Reservoirs

Animals (predominantly wild rodents) are considered to be the primary natural reservoirs for Y pestis. More than 200 mammalian species have been shown to be infected with Y pestis.

In addition, soil may be an important reservoir. In one model, soil serves as the ultimate reservoir, with burrowing animals as the first link in the chain of transmission, followed by spread to other animals and humans through ectoparasites (via fleaborne transmission) (see References: Drancourt 2006). Experimental studies document survival of Y pestis in soil from 24 days (see References: Eisen 2008: Persistence of Yersinia pestis) to 40 weeks (see References: Ayyadurai 2008); however, further study is needed to asses the role of soil in maintaining Y pestis in the natural environment (see References: Drancourt 2006).

Some animal populations are relatively resistant to the effects of Y pestis infection and serve as enzootic reservoirs (see References: Dennis 1997). Plague-resistant mammalian carnivores may aid the spread of plague by feeding on and removing infected prairie dog carcasses to other areas (see References: Salkeld 2007, Boone 2008).

Other animal species, particularly rodents, are more susceptible to disease caused by Y pestis and serve as epizootic hosts (see References: Butler 1991, Gage 1998). Examples of susceptible rodents include the following (see References: Dennis 1997, Gabastou 2000, Reed 1970):

Urban and domestic rats
Ground squirrels
Rock squirrels
Prairie dogs
Deer mice
Field mice
Gerbils
Voles
Chipmunks
Marmots
Guinea pigs
Kangaroo rats

Like humans, mammalian species other than rodents generally are incidental hosts for Y pestis. However, such animals also can serve as sources of human exposure (either through direct contact or through flea vectors). Examples of animals other than rodents that are susceptible to plague include the following (see References: Christie 1980, Dennis 1997, Gage 2000, Palmer 1971, Reed 1970, von Reyn 1976, Wild 2006):

Domestic and feral cats
Dogs
Lagomorphs (rabbits and hares)
Coyotes
Camels
Goats
Deer
Antelope
Lynx

Humans are incidental hosts for Y pestis and are not part of the natural life cycle of the organisms. Disease occurrence in humans is dependent on the frequency of infection in local rodent populations and the degree of contact between rodents and humans. Most human exposures to plague occur in the peridomestic environment, and free-roaming pets that bring infected rodent fleas into the home are considered to be a potential source of infection as well (see References: CDC 2006).

Human outbreaks usually are preceded by epizootics with increased deaths in susceptible animal hosts (see References: Butler 1991, Perry 1997).
Exposure to dogs was found to be a significant risk factor for plague among infected patients in New Mexico. Patients with plague were more likely to report having a sick dog or having slept in the same bed with a pet dog than were controls (see References: Gould 2008). Infected cats were the source of 7.7% of the 297 cases of plague in the United States from 1977 through 1998 (see References: Gage 2000).
In Africa, a study of fleas that have humans as their host (the "human flea," Pulex irritans) revealed that they may be an indicator of plague potential in rural areas and may play a role in plague epidemiology (see References: Laudisoit 2007). Cat fleas may also be secondary vectors for plague in Africa (see References Eisen 2008: Early-phase transmission of Yersinia pestis by cat fleas).
Analysis of serum samples from domestic dogs and cats in an area of China that had recent human plague cases suggests that these animals may serve as sentinel animals for surveillance (see References: Li 2008).

In another recent study, logistic regression models were used to identify landscape features associated with areas where humans have acquired plague from 1957 to 2004 in the four-corners region of the United States (Arizona, Colorado, New Mexico, and Utah). The overall accuracy of the model was >82% and the most conservative model predicted that 14.4% of the four-corners region represented a high risk area of peridomestic exposure to Y pestis (see References: Eisen 2007: Human plague in the southwestern United States, 1957-2004). Such information can be used to identify target areas for surveillance and control (see References: Eisen 2007: Residence-linked human plague in New Mexico).

Plague dynamics appear to be driven by climate variation and seasonal influences.

Studies of plague movement eastward from California show that factors such as climatic and environmental variables can influence spread (see References: Adjemian 2007).
Models of climate change suggest that over the next 50 years geographic shifts of zoonotic diseases such as tularemia and plague will occur. Plague disease ranges in the United States have the potential to expand from a current central focus in New Mexico north into Wyoming and Idaho (see References: Nakazawa 2007).
A recent study found that environmental conditions present during the emergence of recent plague epidemics (wetter springs and warmer summers) may be more common in the future and make outbreaks more likely in both endemic and new areas (see References: Stenseth 2008).
In a 3-year longitudinal study, serologic testing of gerbils in Kazakhstan suggested that there was a mid-summer peak in the abundance of infectious hosts and possible transmission from the reservoir to humans (see References: Begon 2006).
A recent study used logistic regression and geographic information system (GIS)–based modeling to identify environmental predictors of elevated risk for plague in the southwestern United States. Results showed that two factors (distance to pinon-juniper ecotones and amount of precipitation) accurately identified case locations as suitable for plague (producer accuracy, 93%) (see References: Eisen 2007: A spatial model of shared risk).

Vectors

The organisms most commonly are transmitted between animal reservoirs and to humans via bites of infected fleas. In order to survive in the flea midgut, Y pestis organisms require phospholipase D (PLD; formerly referred to as Yersinia murine toxin), which allows the organisms to be resistant to a cytotoxic digestion product of blood plasma in the flea gut. A recent study demonstrated that Y pestis acquired the PLD gene at some point in the past, which allowed transformation from a rather benign species of gut bacteria to a major global pathogen (see References: Hinnebusch 2002).

Of the more than 1,500 flea species, about 30 are known to be vectors for Y pestis. Examples of major flea vectors include the following (see References: Perry 1997):

Xenopsylla cheopis (the oriental rat flea; nearly worldwide in moderate climates)
Oropsylla montanus (United States)
Nosopsyllus fasciatus (nearly worldwide in temperate climates)
Xenopsylla brasiliensis (Africa, India, South America)
Xenopsylla astia (Indonesia and Southeast Asia)
Xenopsylla vexabilis (Pacific Islands)

Because of the poor vector competence of fleas, plague epizootics require a high flea burden per host, even when the susceptible host population density is high (see References: Lorange 2005).

Experimental studies of human body lice have demonstrated that lice can also serve as vectors of Y pestis. Infected lice were able to transmit two virulent strains of plague to uninfected rabbits that subsequently became septicemic and died of plague. Infections were transmitted to naïve lice that fed on infected rabbits (see References: Houhamdi 2006).

Modes of Transmission

Bubonic plague is transmitted from animal reservoirs via:

Bites from flea vectors
Bites or scratches from infected animals, such as cats (see References: Gage 2000)
Direct contact with infected animal carcasses, such as rodents (especially marmots), rabbits, hares, carnivores (eg, wild cats, coyotes), and goats (see References: Christie 1980, Reed 1970, von Reyn 1976)

Pneumonic plague is transmitted via:

Inhalation of respiratory droplets (ie, large droplets [>5 microns]) from infected animals such as cats (see References: Gage 2000)
Inhalation of respiratory droplets from a person with primary or secondary pneumonic plague
Handling Y pestis cultures in the laboratory setting (see References: Burmeister 1962)

Pharyngeal plague can be transmitted via ingestion of Y pestis organisms. A recent report identified five patients with plague who acquired infection after eating raw camel liver; four developed severe pharyngitis and one developed submandibular lymphadenitis (see References: Bin Saeed 2005). In another report from Jordan, pharyngeal plague developed in 12 people after they consumed contaminated camel meat; 11 ate the meat raw and one ate cooked meat (see References: Arbaji 2005)

The risk of infection among contacts of cases of pneumonic plague has not been well quantified and likely varies with intensity of exposure and possibly with environmental factors.

During one recent outbreak of pneumonic plague in Madagascar, investigators measured serum F1 antibodies among contacts and estimated that the secondary infection rate was 8.4% (see References: Ratsitorahina 2000).
Another recent report reviewed eight documented outbreaks of pneumonic plague and found that the average number of secondary transmissions per primary transmission ranged from 0.8 to 3.0 (mean, 1.5; median, 1.3) (see References: Gani 2004).
A third study involved one definite and three probable plague cases (two concurrent index patient-caregiver pairs) (see References: Begier 2006). Each index case transmitted infection to one caregiver, although there were 23 additional close contacts for the two index cases (yielding a secondary infection rate of 8%). Transmission was consistent with large-droplet spread.
Several experimental studies have been conducted to determine how far Y pestis organisms will spread from patients with pneumonic plague. These studies involved placing agar plates at various distances from the mouths of infected patients. The farthest distance of spread was 3.7 feet, indicating that close contact is necessary for transmission (see References: Kool 2005).

Historical Perspective

Three plague pandemics have been recorded throughout history (see References: WHO: Report on global surveillance 2000), with an estimated 200 million deaths (see References: Perry 1997). Brief descriptions of the three pandemics follow.

The first pandemic started in Egypt in 542 AD and continued for more than a century. Outbreaks in Europe, Central and Southern Asia, and Africa killed an estimated 100 million people.
The second pandemic began in Italy in 1347 and rapidly spread throughout Europe over the next several years, killing an estimated one third of the European population. Paleodemographic studies suggest that mortality was partially determined by relative health of those infected (see References: DeWitte 2008). During that time, plague became known as the Black Death. Outbreaks of plague continued to occur sporadically in Europe over the next several centuries.
The third pandemic began in 1894 in China and spread around the world over a 10-year period, predominantly by infected rats and their fleas aboard steamships. An estimated 12 million deaths occurred, mostly in India.

Although bubonic plague historically has been the most common form of disease, large outbreaks of pneumonic plague (with person-to-person transmission as the primary mode of spread) also have been reported (see References: Meyer 1961, Kool 2005).

Two large outbreaks of pneumonic plague occurred in Manchuria in the early 20th century (1910-1911 and 1920-1921). An estimated 60,000 deaths occurred in the former and an estimated 9,300 in the latter.
Two pneumonic plague outbreaks occurred in the United States in the early 1900s (see References: Anderson 1978, Kool 2005). The first occurred in 1919 in Oakland, California. The index case was a hunter who contracted bubonic plague from an infected squirrel. He subsequently developed plague pneumonia and transmitted the disease to 12 or 13 other persons. A second outbreak occurred in Los Angeles in 1924 and involved 39 cases of pneumonic plague.

Naturally Occurring Plague in the United States

Plague was first introduced into the United States in 1900 from China; the first plague epidemic occurred in San Francisco from 1900 to 1904 (see References: Caten 1966).
Up through 1926, plague occurred most commonly in urban settings (particularly in California) and was associated with infections in urban rat populations (see References: Kaufmann 1980).
After 1926, plague gradually became endemic in wild animal populations in the Western United States (generally in wild rodents), and human cases continued to occur in persons with exposure to such populations.
Cases also have occurred following exposure to infected cats (see References: Eidson 1988, Gage 2000). A recent report described 23 cases of cat-associated plague in the western United States from 1977 through 1998; five were cases of primary pneumonic plague (see References: Gage 2000).
Since the 1920s, most human plague cases in the United States have occurred in California, New Mexico, Arizona, and Colorado (see References: CDC 1994).
Plague in the United States generally is seasonal, with a higher incidence in the summer months (see References: Caten 1966, Kaufmann 1980).
From 1947 through 1996, 390 cases of plague were reported to the Centers for Disease Control and Prevention (CDC) (see References: CDC 1996):

The overall case-fatality rate was 15.4%.
Bubonic plague accounted for 327 (83.9%) cases.
Primary septicemic plague accounted for 49 (12.6%) cases.
Primary pneumonic plague accounted for seven (1.8%) cases.
Seven cases were unclassified.

From 1990 through 2005, 107 cases of plague were reported to the CDC, with a median of 7 cases per year (see References: CDC 1999; CDC 2006).
Most human cases in the United States occur in two regions: (1) northern New Mexico, northern Arizona, and southern Colorado and (2) California, southern Oregon, and far western Nevada (see References: CDC: Plague: epidemiology). In April 2006, Los Angeles County health officials confirmed a case of bubonic plague in a local resident; this is the first human case of plague in a Los Angeles County resident since 1984 (see References: County of Los Angeles Department of Health Services).
Between February and July 2006, 13 human cases of plague were reported among residents of four states: New Mexico, 7; Colorado, 3; California, 2; and Texas, 1. This total is the largest number of cases reported in a single year in the United States since 1994.

The increased number of cases during this time frame is consistent with the predicted relationship between climate and frequency of human plague in the southwestern United States.
Two consecutive February-March periods with high precipitation and an intervening cool summer favor an increased number of cases of plague the following summer. The net effect was increased reproduction and survival rates among rodents and fleas (see References: CDC 2006; Enscore 2002).

Naturally Occurring Plague Worldwide

Natural enzootic foci of plague (and, therefore, areas with the highest incidence of human disease) include the following countries or regions (see References: CDC: Information on plague; WHO: Report on global surveillance 2000):

Madagascar
Eastern and southern Africa (eg, Uganda, Kenya, Tanzania, Mozambique, Botswana)
Southeast Asia (particularly Vietnam and Myanmar)
Pockets in South America (including areas in the Andean mountain regions of Peru, Bolivia, and Ecuador and in northeastern Brazil)
The western United States
Mongolia and northern China
Russia (the area of the Caucasus Mountains) and central Asia into the Middle East

Between 1954 and 1997, 38 countries reported cases of human plague to the World Health Organization (WHO) (see References: WHO: Report on global surveillance 2000). No plague cases have been reported from Europe since shortly after World War II and no cases have been reported from Australia. Overall, 80,613 cases were reported, with a mean of 1,832 cases per year (range, 200 to 6,004 cases).

The overall average case-fatality rate among reported cases was 11.8%.
Seven countries reported cases of plague for each of the 44 years: Brazil, Democratic Republic of the Congo, Madagascar, Myanmar, Peru, the United States, and Vietnam.
Large outbreaks occurred in Vietnam (from 1966 through 1972), India (1954, 1963, and 1994), Tanzania (1990 through 1992), and Madagascar (1994 through 1996).
The 1994 outbreak in India included cases of pneumonic plague and raised concerns about the spread of pneumonic plague to other areas of the world through airline travel (See References: Campbell 1995, Titball 1998).
In 1997, a localized outbreak of pneumonic plague involving 18 patients occurred in Madagascar; most of the patients had exposure to a traditional healer who died of the disease after treating the index patient (see References: Ratsitorahina 2000).

An outbreak of pneumonic plague was reported in the Democratic Republic of the Congo (DRC) in early 2005. Cases occurred between December 2004 and March 2005 in workers of a diamond mine where about 7,000 people worked under crowded conditions with poor sanitation. WHO reported 130 cases with 57 deaths in their last bulletin on the subject (see References: WHO: Plague in the Democratic Republic of the Congo). It is likely that the index case acquired Y pestis infection through a fleabite and then developed the pneumonic form of the disease and subsequently transmitted infection to other workers through the respiratory route. Additional outbreaks of pneumonic plague have occurred in 2006 in the Oriental province of the DRC (see References: WHO: Plague in the Democratic Republic of the Congo). More than 620 cases, including 42 deaths, were reported from the end of July to mid-October; however, investigators suspect that the total may be overestimated because of the low fatality rate observed among cases.

Plague as a Biological Weapon

Experience with plague as a biological weapon is limited; however, the following information supports the perspective that plague deserves serious consideration as a bioterrorist agent:

Plague was used as a biological weapon in the Middle Ages when armies catapulted dead plague victims into cities under siege in order to spread the disease (see References: Osterholm 2000).
Japan used plague as a biological weapon against the Chinese during World War II by dropping plague-infected fleas over populated areas and causing outbreaks of the disease (see References: McGovern 1997, Osterholm 2000).
In the years following World War II, biological weapons programs in the United States and the Soviet Union developed techniques for aerosolizing Y pestis, thus enhancing the effectiveness of this agent as a potential biological weapon (see References: Inglesby 2000).
A 1970 WHO report estimated that an aerosol release of 50 kg of dried powder containing 6 x 10¹⁵ Y pestis spores over a city of 5 million people in an economically developed country (such as the United States) would produce 150,000 incapacitating illnesses and up to 36,000 deaths (see References: WHO: Health aspects of chemical and biological weapons). These estimates did not take into consideration secondary cases that would occur through subsequent person-to-person contact.
Plague is a suitable pathogen for use as a biological weapon because:

The organisms can be delivered in an aerosol form.
Pneumonic plague causes a serious illness with a high case-fatality rate.
Pneumonic plague is communicable, and large outbreaks have occurred in the past.
A bioterrorist attack involving pneumonic plague would cause widespread fear and panic that would be difficult to contain, partly because of the communicable nature of the disease (see References: Campbell 1995; Inglesby 2001).
Y pestis could potentially be genetically altered to enhance virulence or create antibiotic-resistant strains (see References: Gilsdorf 2005).

Plague used as a bioterrorist weapon would be expected to have the following features:

Previously healthy patients would present with a severe and rapidly progressive pneumonia.
An acute multilobar pneumonia accompanied by hemoptysis, associated gastrointestinal symptoms, and a fulminant clinical course would be very suspicious for pneumonic plague.
Many similar cases would present over several days.
Illness onsets would generally occur 2 to 4 days after release, but could occur as soon as 1 day and up to 6 days later.
Buboes characteristic of bubonic plague would not be present.
Illness would likely occur in an urban area and patient would not have a history of recent travel to a plague-endemic region (ie, southwestern United States).
Patients would not necessarily have risk factors for plague exposure (eg, outdoor field work, veterinary work, recent outdoor recreational activity).
There would be no indication of a prior recent plague epizootic with rodent deaths in the affected community.
Antibiotic resistance may be present.

Investigators recently used univariate and multivariate modeling to assess key parameters for controlling a pneumonic plague outbreak (see References: Massin 2007). Using a hypothetical reference scenario of 1000 index cases of plague pneumonia in Paris, if interventions were taken 10 days after an attack, an estimated 2,500 deaths would occur. Rapidity of implementing interventions offered the greatest effect on final epidemic size. Other measures, in order, were wearing masks, treating contacts preventively, and quarantine. Limiting inter-regional mixing confined casualties to the region but did not reduce casualties in the model.

Clinical Syndromes and Differential Diagnosis

Overview

Yersinia pestis infection can cause the following clinical syndromes:

Bubonic plague
Primary septicemic plague
Primary pneumonic plague
Plague meningitis
Plague pharyngitis
Pestis minor
Subclinical infection

The classic forms of plague are bubonic plague, septicemic plague, and pneumonic plague; these are outlined in the tables below. Septicemic plague can be either primary or secondary to bubonic plague. Similarly, pneumonic plague can be either primary or secondary to septicemic plague or bubonic plague (ie, following hematogenous spread).

Brief information about other syndromes caused by Y pestis infection follows:

Plague meningitis occurs as a complication of bacteremia and may be the presenting clinical syndrome for some cases. Symptoms are typical of meningitis from other etiologies and include fever, headache, meningismus, and mental status changes. If meningitis occurs as a complication of bubonic plague, some data suggest that a bubo in the axillary region is a predisposing factor (see References: Butler 1976). The cerebrospinal fluid demonstrates PMNs; characteristic gram-negative organisms usually can be seen on Gram stain (see References: Butler 1991).
Plague pharyngitis occurs as a result of inhaling or ingesting Y pestis organisms. The clinical illness is similar to severe pharyngitis or acute tonsillitis of other causes (eg, streptococcal infection); inflamed cervical nodes usually are present (and usually have the features of a characteristic bubo or buboes). As with bubonic plague, septicemia can occur.
Pestis minor is a milder form of bubonic plague. Patients usually have a febrile illness with localized lymphadenopathy. The nodes drain and patients recover without therapy. Patients with this form of plague are more likely to have some preexisting immunity to Y pestis (see References: Legters 1970).
Subclinical infections can occur as evidenced by cross-sectional surveys of serum antibody titers in populations living in endemic areas (see References: Ratsitorahina 2000).

Clinical Features of Bubonic Plague
Feature	Characteristics
Incubation period	1-7 days
Presenting features	—Sudden onset of fever, chills, weakness —Usually within 1 day, painful swollen lymph node or group of nodes (bubo) occurs in groin, axilla, or cervical region: ~1-10 cm, smooth, uniform, unfixed, egg-shaped mass or irregular cluster of several nodes ~Extremely tender ~Region may be erythematous, with surrounding edema ~Buboes usually occur in only one location, but multiple buboes may be seen ~Rarely, buboes may suppurate and rupture —Skin lesions may occur at site of flea bite (ie, papules, vesicles, pustules) but are present in <10% of cases —Associated lymphangitis uncommonly occurs —Presenting symptoms for case series of 40 Vietnamese patients with bubonic plague^a: ~Fever (100%) (mean temperature for 32 patients: 102.9°F [39.4°C]) ~Chills (40%) ~Bubo (100%—groin, 88%; axilla, 15%; cervical, 5%; epitrochlear, 3%) ~Headache (85%) ~Prostration (75%) ~Altered mental status (38%) (lethargy, confusion, delirium, seizures) ~Anorexia (33%) ~Vomiting (25%) ~Abdominal pain (18%) ~Cough (25%) ~Chest pain (13%) ~Skin rash (23%) (petechiae, purpura, papular eruptions)
Laboratory features	—Laboratory features for case series of 40 Vietnamese patients^a: ~Mean WBC count: 21,500/mm³ (range, 6,000/mm³-100,000/mm³) (most patients had left shifts and 3 had leukemoid reactions) ~PMNs showed cytoplasmic vacuolation in 24 patients, Dohle bodies in 20 patients, toxic granules in 8 patients ~Mean platelet count: 210,000/mm³ (range, 72,000/mm³-496,000/mm³) (18 patients had platelet counts <150,000/mm³) ~SGOT elevated in 13 patients (20-92 M-IU) ~LDH elevated in 7 patients (308-900 units) ~Alkaline phosphatase elevated in 9 patients (33-116 units) ~PTT >10 seconds over control In 6 patients
Complications	—Secondary septicemia (can lead to DIC, shock, multisystem involvement) —Secondary pneumonic plague (5%-15% of patients)^b —Meningitis (may occur in patients with bubonic plague that was not adequately treated)^b —Buboes may become infected with other bacterial pathogens^a
Case-fatality rate	—Over 50% without antibiotic therapy^c —With appropriate antibiotic therapy, <5%^d
Abbreviations: DIC, disseminated intravascular coagulation; LDH, lactic dehydrogenase; PMNs, polymorphonuclear neutrophils; PTT, partial thromboplastin time; SGOT, serum glutamic oxaloacetic transaminase; WBC, white blood cell. ^aSee References: Butler 1972. ^bSee References: McGovern 1997. ^cSee References: Dennis 1997. ^dSee References: Butler 1991.

Clinical Features of Primary Septicemic Plague
Feature	Characteristics
Incubation period	1-4 days
Presenting features^a,b	—10%-25% of US plague cases present with primary septicemic plague^b —Presenting symptoms for 18 cases of primary septicemic plague in New Mexico: ~Fever (100%) ~Chills (61%) ~Nausea (44%) ~Headache (44%) ~Vomiting (50%) ~Diarrhea (39%) ~Abdominal pain (39%) ~Any gastrointestinal symptom (72%) —Presenting signs for 18 cases of primary septicemic plague in New Mexico^a: ~Mean temperature: 38.5°C (range, 35.4°C-40.4°C) ~Mean pulse: 109 (range, 72-160) ~Mean respiratory rate: 31 (range, 16-60) ~Mean systolic BP: 104 (range, 80-130) ~Mean diastolic BP: 66 (range, 36-80) —Mental status changes commonly occur (delirium, obtundation, coma)
Laboratory features	—Laboratory features consistent with severe bacterial infection and sepsis syndrome (as often seen with bubonic plague and secondary septicemia) —Leukocytosis, leukopenia, or normal WBC count may be seen —If plague pneumonia present, CXR shows patchy alveolar infiltrates (usually bilateral), often with consolidation —Findings noted for 18 patients with septicemic plague: ~Mean WBC count: 18,950/mm³ (range, 3,000/mm³-68,700/mm³); all had marked left shifts ~Bacteria seen on peripheral blood smear (17.6%)
Complications^a,c,d	—Illness rapidly progresses to sepsis syndrome often with DIC, shock, and multisystem involvement —Skin lesions reflect DIC (may be similar to meningococcemia)^c: ~Purpura ~Petechiae ~Ecchymoses ~Gangrene of acral regions (caused by small artery thromboses) ~Ecthyma gangrenosum (rare) —Meningitis^d —Secondary plague pneumonia (about 25% of patients)*
Case-fatality rate^d	—Overall 30%-50%^b —High CFR related to delay in appropriate diagnosis and antibiotic therapy —Without antibiotic therapy, CFR approaches 100%
Abbreviations: BP, blood pressure; WBC, white blood cell; CXR, chest x-ray. ^aSee References: Hull 1987. ^bSee References: Perry 1997. ^cSee References: McGovern 1999. ^dSee References: Dennis 1997.

Clinical Features of Primary Pneumonic Plague^a
Feature	Characteristics
Incubation period	1-4 days
Presenting features	—Symptoms of primary plague pneumonia^b: ~Fever ~Chest pain ~Dyspnea ~Productive cough (sputum may be purulent or watery, frothy, blood-tinged) ~Hemoptysis ~Tachypnea (particularly in young children) ~Cyanosis ~Bubo not present (rarely, cervical bubo may be noted) —Gastrointestinal symptoms (nausea, vomiting, abdominal pain, diarrhea) common
Laboratory features	—Findings consistent with severe bacterial infection and sepsis syndrome (as often seen with bubonic and primary septicemic plague) —CXR findings in series of 9 cases of secondary pneumonic plague^c: ~Alveolar infiltrates (100%) ~Pleural effusion (55%) ~One patient developed cavitary lesion 3 weeks after illness onset —Consolidation common on CXR; massive mediastinal adenopathy occurs rarely —Organisms usually seen on sputum Gram stain
Complications	—Septicemia with sepsis syndrome —Meningitis
Case-fatality rate	—Close to 100% without appropriate antibiotic therapy (generally, fatality rates are high if antibiotic therapy is not instituted soon after symptom onset [ie, within 24 hr]; however, patients may survive even if appropriate therapy is instituted beyond 24 hr)^d —Death often occurs 2-5 days after illness onset^e
Abbreviations: CXR, chest x-ray. ^aNote: Few detailed clinical descriptions of primary plague pneumonia are readily available since the condition is relatively rare. ^bSee References: Dennis 1997. ^cSee References: Alsofrom 1981. ^dSee References: Begier 2006, Gage 2000, Butler 1991. ^eSee References: Inglesby 2000.

Pediatric Considerations

The clinical presentation of plague in children is similar to that in adults. Several studies have made the following observations about pediatric plague:

Children with bubonic plague may have a slightly increased risk for development of secondary pneumonic plague or meningitis. In one study of 38 pediatric patients with plague, 16% developed pneumonia and 11% developed meningitis (see References: Mann 1982). In two other case series, most of the meningitis cases occurred among children (see References: Crook 1992, Reed 1970).
Vomiting may be more common in children at the time of presentation of illness than in adults (ie, about 50% and 30%, respectively) (see References: Burkle 1973, Mann 1982).
In cases of bubonic plague, node pain is more common in children than lymph node swelling or a bubo; this may manifest as limb immobility (such as from painful axillary nodes).
Retroperitoneal adenopathy may be responsible for vomiting and/or abdominal pain.
Children may be more likely to have seizures as part of the presenting symptom complex (see References: Butler 1991). Most often these are febrile seizures, although they may be caused by plague meningitis, which may be more common in children.
The diagnosis often is not considered at time of initial presentation for pediatric cases, even in a plague-endemic area (see References: Mann 1982).

Presenting symptoms for 38 pediatric patients with bubonic or septicemic plague diagnosed in New Mexico between 1970 and 1980 are shown in the table below. The overall case-fatality rate was 15.8%.

Presenting Symptoms for 38 Pediatric Patients With Bubonic or Septicemic Plague
Symptom	Patients With Bubonic Plague (N=31)	Patients With Septicemic Plague (N=7)
Fever Chills Vomiting Headache Abdominal distress or nausea Diarrhea Lethargy, malaise, anorexia	30 (97%) 11 (35%) 16 (52%) 11 (35%) 8 (26%) 3 (10%) 12 (39%)	36 (95%) 0 3 (43%) 0 2 (29%) 0 3 (43%)
Adapted from Mann 1982 (see References).

In the pre-antibiotic era, pregnant women with plague usually either died or had an adverse outcome of pregnancy (eg, spontaneous abortion, stillbirth). However, more recent reports have demonstrated successful outcomes with antibiotic therapy, including normal gestational periods and delivery of healthy infants (see References: Mann 1977, Welty 1985). Potential adverse effects to the fetus are governed by the time of antibiotic therapy; however, during outbreaks and bioterrorism emergencies, treatment benefits for the mother outweigh fetal risk (see References: Cono 2006). Antibiotics should be administered to infants born to infected mothers (see References: Welty 1985).

Breast-feeding women and their infants should be treated with the same antibiotic. The medication that is safest for the infant generally should be considered the first choice (ie, gentamicin in the contained casualty setting and doxycycline in the mass casualty setting). Fluoroquinolone antibiotics would be the recommended alternative in both settings (see References: Inglesby 2000).

Differential Diagnosis

Differential Diagnosis for Pneumonic, Bubonic, and Septicemic Plague
Condition	Distinguishing Features
PNEUMONIC PLAGUE^a
Inhalational anthrax (Bacillus anthracis)	—Widened mediastinum and pleural effusions seen on CXR or chest CT —Not true pneumonia; minimal sputum production —Hemoptysis uncommon (if present, suggests diagnosis of plague)
Tularemia (Francisella tularensis)	Clinical course not as fulminant as pneumonic plague
Community-acquired bacterial pneumonia —Mycoplasmal pneumonia (Mycoplasma pneumoniae) —Pneumonia caused by Chlamydia pneumoniae —Legionnaires' disease (Legionella pneumophila or other Legionella species) —Psittacosis (Chlamydia psittaci) —Other bacterial agents (eg, Staphyloccocus aureus, Streptococcus pneumoniae, Haemophilus influenzae, Klebsiella pneumoniae, Moraxella catarrhalis)	—Rarely as fulminant as pneumonic plague —Legionellosis and many other bacterial agents (S aureus, S pneumoniae, H influenzae, K pneumoniae, M catarrhalis) usually occur in persons with underlying pulmonary or other disease or in the elderly —Bird exposure with psittacosis —Gram stain of sputum may be useful —Community outbreaks caused by other etiologic agents not likely to be as explosive as pneumonic plague outbreak —Outbreaks of S pneumoniae usually institutional —Community outbreaks of Legionnaires' disease often involve exposure to cooling towers
Viral pneumonia —Influenza —Hantavirus —RSV —CMV	—Influenza generally seasonal (October-March in United States) or involves history of recent cruise ship travel or travel to tropics —Exposure to excrement (urine or feces) of mice with Hantavirus —RSV usually occurs in children (although may be cause of pneumonia in elderly); tends to be seasonal (winter/spring) —CMV usually occurs in immunocompromised patients
Q fever (Coxiella burnetii)	—Exposure to infected parturient cats, cattle, sheep, goats —Severe pneumonia not prominent feature
BUBONIC PLAGUE^b
Streptococcal or staphylococcal adenitis (S aureus, Streptococcus pyogenes)	—Purulent or inflamed lesion often noted distal to involved nodes (ie, pustule, infected traumatic lesion) —Involved nodes more likely to be fluctuant —Associated ascending lymphangitis or cellulitis may be present (generally not seen with plague)
Tularemia (F tularensis)^c	—Ulcer or pustule often present distal to involved nodes —Clinical course rarely as fulminant as in plague —Systemic toxicity uncommon
Cat scratch disease (Bartonella henselae)	—History of contact with cats; usually history of cat scratch —Indolent clinical course; progresses over weeks —Primary lesion at site of scratch often present (small papule, vesicle) —Systemic toxicity not present
Mycobacterial infection, including scrofula (Mycobacterium tuberculosis and other Mycobacterium species)	—With scrofula, adenitis occurs in cervical region —Usually painless —Indolent clinical course —Infections with species other than M tuberculosis more likely to occur in immunocompromised patients
Lymphogranuloma venereum (Chlamydia trachomatis)	—Adenitis occurs in the inguinal region —History of sexual exposure 10-30 days previously —Suppuration, fistula tracts common —Although LGV buboes may be somewhat tender, exquisite tenderness usually absent —Although patients may appear ill (headache, fever, myalgias), systemic toxicity not present
Chancroid (Hemophilus ducreyi)	—Adenitis occurs in the inguinal region —Ulcerative lesion present —Systemic symptoms uncommon; toxicity does not occur
Primary genital herpes	—Herpes lesions present in genital area —Adenitis occurs in the inguinal region —Although patients may be ill (fever, headache), severe systemic toxicity not present
Primary or secondary syphilis (Treponema pallidum)	—Enlarged lymph nodes in the inguinal region —Lymph nodes generally painless —Chancre may be noted with primary syphilis
Strangulated inguinal hernias	Evidence of bowel involvement
SEPTICEMIC PLAGUE
Meningococcemia	More likely to have evidence of meningitis (but not always present)
Septicemia caused by other gram-negative bacteria	Underlying illness usually present
Abbreviations: CMV, cytomegalovirus; CT, computed tomography; CXR, chest x-ray; LGV, lymphogranuloma venereum; RSV, respiratory syncytial virus. ^aOther causes of pneumonia also may be considered, depending on the clinical presentation and setting (eg, tuberculosis, fungal infections). ^bInfectious causes of generalized lymphadenopathy (eg, CMV infection, toxoplasmosis, mononucleosis) also may be considered, depending on the clinical presentation. ^cSee References: Butler 1979; Cleri 1997.

Clinical Laboratory Testing

Specimen Collection and Transport

Collection and Transport of Clinical Laboratory Specimens for the Diagnosis of Plague
Condition	Collection and Transport^a,b,c
Pneumonia	Sputum: —May be collected for culture/direct examination, but yield on culture may be low owing to likely overgrowth of normal flora^d —Transport at room temperature (22°C-28°C) if transport <2 hr —If transport expected to be 2-24 hr, refrigerate (2C°C-8°C) —Order culture, Gram stain, and Giemsa, Wright’s, or Wayson stain —If suspicion of plague high, contact local health department and LRN laboratory for instructions on ordering DFA or other tests Bronchial wash (>1.0 mL): —Bronchoscopy may be indicated in certain situations where sputum specimens are negative [Note: In such situations, the bronchoscopy team should follow appropriate barrier precautions, including use of masks and other personal protective equipment; if a bronchoscopy is performed on a patient who later is found to have pneumonic plague and the team did not wear respiratory protection, then post-exposure prophylaxis is indicated for members of the team] —Use same specimen handling conditions and test orders as described for sputum specimens Blood: —Collect volume and number of sets per institution’s standard protocol —Transport to laboratory and hold at ambient temperature until placed into incubator or blood culture instrument —DO NOT REFRIGERATE —Follow established laboratory protocol for processing blood cultures —If high suspicion of plague, order additional blood or broth culture (general nutrient broth) for incubation at room temperature (22°C-28°C), the optimal growth temperature range for Y pestis. Cultures should be incubated without shaking [Note: An additional culture set is needed because holding cultures at room temperature will delay or negate growth of other common bacterial pathogens] Serum: —An acute-phase serum sample may be collected and stored at 4°C until plague can be ruled out —If plague cannot be ruled out, contact public health officials and LRN system for further instructions
Septicemia	Blood: —Collect volume and number of sets per institution’s standard protocol —Transport to laboratory and hold at ambient temperature until placed into incubator or blood culture instrument —DO NOT REFRIGERATE —Follow established laboratory protocol for processing blood cultures —If high suspicion of plague, order additional blood or broth culture (general nutrient broth) for incubation at room temperature (22°C-28°C), the optimal growth temperature range for Y pestis. Cultures should be incubated without shaking [Note: An additional culture set is needed because holding cultures at room temperature will delay or negate growth of other common bacterial pathogens.] —Polychromatic staining of direct peripheral smear may demonstrate organisms Serum: —An acute-phase serum sample may be collected and stored at 4°C until plague can be ruled out —If plague cannot be ruled out, contact public health officials and LRN system for further instructions
Bubonic plague	Bubo aspirate: —Sterile saline flush (1.0 mL saline in 10 mL syringe with 20-guage needle) may be needed to obtain adequate material for culture —Transport capped, taped syringe without needle (or contents of syringe in sterile container) at room temperature (22°C-28°C) for immediate processing. —Refrigerate (2°C-8°C) if processing will be delayed —Order culture, Gram stain, and Giemsa, Wright’s, or Wayson stain —If high suspicion of plague, contact local health department and LRN laboratory for instructions on ordering direct fluorescent antibody (DFA) or other tests Tissues: —Place in sterile containers with 1-2 drops sterile nonbacteriostatic normal saline to keep moist —Transport at room temperature to laboratory for immediate processing —Use same specimen handling conditions and test orders as described for bubo aspirates —Swabs of tissue are not recommended Blood: —Collect volume and number of sets per institution’s standard protocol —Transport to laboratory and hold at ambient temperature until placed into incubator or blood culture instrument —DO NOT REFRIGERATE —Follow established laboratory protocol for processing blood cultures —If high suspicion of plague, order additional blood or broth culture (general nutrient broth) for incubation at room temperature (22°C-28°C), the optimal growth temperature range for Y pestis. Cultures should be incubated without shaking [Note: An additional culture set is needed because holding cultures at room temperature will delay or negate growth of other common bacterial pathogens] Serum: —Acute-phase serum sample may be collected and stored at 4°C until plague can be excluded —If plague cannot be ruled out, contact public health officials and LRN system for further instructions
Meningitis	CSF: —Use standard collection and transport protocols for Gram stain and culture Blood: —Collect volume and number of sets per institution’s standard protocol —Transport to laboratory and hold at ambient temperature until placed into incubator or blood culture instrument —DO NOT REFRIGERATE —Follow established laboratory protocol for processing blood cultures —If high suspicion of plague, order additional blood or broth culture (general nutrient broth) for incubation at room temperature (22°C-28°C), the optimal growth temperature range for Y pestis. Cultures should be incubated without shaking [Note: An additional culture set is needed because holding cultures at room temperature will delay or negate growth of other common bacterial pathogens]
Autopsy	Blood, tissue from buboes, liver, spleen, lungs, and bone marrow —Transport fresh or frozen on dry ice —Use holding medium such as Cary-Blair
Abbreviations: DFA, direct fluorescent antibody; LRN, Laboratory Response Network. ^aThese collection criteria are primarily designed for ruling out a diagnosis of plague at level A laboratories in the Laboratory Response Network (LRN). Higher-level testing for confirmation of plague occurs at levels B, C, and D LRN laboratories. If medical suspicion of plague is high, or if there has been an announced or recognized attack, local public health authorities should be contacted for further specimen collection information and access to testing through the LRN system. ^bSpecimens for culture should be taken before administration of antimicrobial therapy. ^cSee References: Aleksic 1999; CDC: Public health emergency response and preparedness/specimen selection; CDC/ASM/APHL 2002; McGovern 1997; WHO: Plague manual. ^dSee References: CDC: Information on plague/Diagnosis.

Guidelines have been published for the packing and shipping of infectious substances, diagnostic specimens, and biological agents from suspected bioterrorism (see References: ASM: Sentinel laboratory guidelines for suspected agents of bioterrorism: packing and shipping infectious substances, diagnostic specimens, and biological agents). Y pestis is classified under World Health Organization (WHO) risk group 4. Specimens and isolates that are reasonably suspected to contain Y pestis must be transported as "infectious substances." International Air Transport Association (IATA) rules require training of all individuals involved in the transport of dangerous goods, including infectious substances. Once Y pestis is identified, isolates and specimens are regulated as select agents and are subject to additional transport requirements (see below). Chain of custody should be documented for material that may constitute evidence of criminal activity.

Laboratory Biosafety and Biosecurity Information

Laboratory-acquired Y pestis infections have been reported, although rarely. Hazards to laboratory personnel include:

Direct contact with cultures and infectious materials from humans or rodents
Infectious aerosols or droplets generated during manipulation of cultures and infected tissues
Accidental autoinoculation, ingestion, or bites from infected fleas during necropsy of rodents

Biosafety level 2 (BSL-2) conditions are recommended in the following situations (see References: CDC 1999):

Culture of specimens
Handling of cultures that may contain Y pestis
Necropsy of potentially infected animals

Once the organism has been identified, it is prudent to work in a higher-level containment environment (see References: Aleksic 1999).
Biosafety level 3 (BSL-3) practices are required for the following activities (see References: CDC 1999):

Those with high potential for droplet or aerosol production
Those involving work with antibiotic-resistant strains
Those involving production quantities or concentrations of infectious materials

Vaccination is not recommended for clinical laboratory personnel (see References: CDC: Laboratory Information>Biosafety>Vaccines).
Laboratory safety practices associated with Y pestis and other potential agents of bioterrorism have been reviewed elsewhere (see References: Sewell 2003).
Yersinia pestis is classified as a select agent and therefore is regulated under 42 CFR part 73 (Possession, Use, and Transfer of Select Agents and Toxins), which was published in final form in the Federal Register in March 2005 (see References: HHS). As specified in the Public Health Security and Bioterrorism Preparedness and Response Act of 2002, 42 CFR part 73 provides requirements for laboratories that handle select agents (including registration, security risk assessments, safety plans, security plans, emergency response plans, training, transfers, record keeping, inspections, and notifications). These new requirements went into effect on February 7, 2003, and override earlier government requirements regarding possession and transfer of select agents. For more information about CDC's Select Agent Program, see References: CDC: Select Agent Program. In addition, CDC published additional guidelines for enhancing laboratory security for laboratories working with select agents (see References: CDC 2002).

The Laboratory Response Network

The Laboratory Response Network (LRN) is a national network of approximately 150 laboratories. The network includes the following types of labs: federal, state and local public health, military, food testing, environmental, veterinary, and international (located in Canada, the United Kingdom, and Australia) (see References: CDC: Facts about the Laboratory Response Network).

The LRN structure for bioterrorism designates laboratories as either national, reference, or sentinel. Designation depends on the types of tests a laboratory can perform and how it handles infectious agents to protect workers and the public.

National laboratories have unique resources to handle highly infectious agents and the ability to identify specific agent strains.
Reference laboratories, sometimes referred to as “confirmatory reference,” can perform tests to detect and confirm the presence of a threat agent. These laboratories ensure a timely local response in the event of a terrorist incident. Rather than having to rely on confirmation from laboratories at the CDC, reference laboratories are capable of producing conclusive results; this allows local authorities to respond quickly to emergencies. These are mostly state or local public health laboratories with BSL-3 containment facilities that have been given access to nonpublic testing protocols and reagents.
Sentinel laboratories represent the thousands of hospital-based labs that are on the front lines. Sentinel laboratories have direct contact with patients. In an unannounced or covert terrorist attack, patients provide specimens during routine patient care. Sentinel laboratories could be the first facility to spot a suspicious specimen. A sentinel laboratory’s responsibility is to refer a suspicious sample to the right reference laboratory. These laboratories generally have at least BSL-2 containment.

Standard Tests for Detection of Y pestis

Direct stains for bacterial micromorphology

Approved for LRN sentinel laboratories.
Gram stain:

Yersinia pestis organisms in direct specimens or in specimen cultures appear as single cells or short chains of plump, gram-negative rods (1.0 to 2.0 mcm x 0.5 mcm).
Specimen types in order of likely positive smear results are: bubo aspirates, tissue, blood, sputum.
In direct smears, bacterial cells may be inside or outside of leukocytes.
The Gram smear morphology is suggestive but not specific for Y pestis.
Streptococcus pneumoniae may be visualized on the Gram stain (gram-positive lancet-shaped diplococci) in pneumonic plague cases with secondary infection (see References: CDC/ASM/APHL 2002).

Polychromic stain:

Yersinia pestis stains as a bipolar "closed safety pin" with Giemsa, Wright's, or Wayson stains. Bipolar morphology may not be evident on Gram stain.
Bipolar staining is not exclusive to Y pestis and therefore is considered only suggestive of the diagnosis (see References: CDC/ASM/APHL 2002).

Limited information is available on the sensitivity of direct staining for detecting Y pestis organisms in clinical specimens. The following findings have been reported:

Investigation of an outbreak of pneumonic plague in Madagascar demonstrated that a sputum Gram stain was positive in only 1 (14%) of 7 patients tested; of these, all except 1 (not the patient with the positive stain) had been receiving streptomycin for 48 hours prior to specimen collection (see References: Ratsitorahina 2000).
A study in New Mexico found that gram-negative bacteria were seen on direct examination of peripheral smears from 3 (17.6%) of 17 patients with septicemic plague and 4 (9.5%) of 42 patients with bubonic plague (see References: Hull 1987).
In 22 Vietnamese patients with bubonic plague, Wayson staining of bubo aspirates demonstrated characteristic organisms in 13 (59%) of the patients (see References: Butler 1974).
Another Vietnamese study found that plague bacilli were visible in blood smears from plague patients with blood concentrations of more than 10⁶ colony-forming units per milliliter. The authors concluded that patients with high concentrations of bacilli in the blood, whose specimens are most likely to be positive upon direct examination, have a poorer prognosis than patients with lower concentrations (see References: Butler 1976).
Although aspiration of buboes is more difficult early in the course of disease, early samples are more likely to yield characteristic gram-negative bacilli that to exhibit bipolar staining with polychromic stains (see References: Goldenberg 1968).

Bacteriologic culture

Approved for LRN sentinel laboratories.
Standard procedures for culture are as follows (see References: CDC/ASM/APHL 2002):

Culture setup procedure: Use standard protocol for specimen type.
Tape plates shut if plague is suspected.
Add media for 28^oC incubation (optimal growth temperature of Y pestis) if plague is suspected. Media incubated at standard 35^o to 37^oC incubation is necessary for development of the F1 antigen of Y pestis and for recovery of other pathogens that may be present.
Atmosphere: ambient or 5% CO₂
Incubation time: 5 days, 7 days if the patient has been treated with a bacteriostatic antibiotic
Colony morphology, 24 hours (SBA): gray-white, pinpoint
Colony morphology, 48 hours (SBA): 1- to 2-mm gray-white to pale yellow nonhemolytic, described as having a "fried-egg" or "hammered-copper" appearance
Broth culture colonial morphology, 24 hours: "flocculant" or "stalactite" clumps
Broth culture colonial morphology, 48 hours: "white fluff" on bottom of tube

Preliminary identification

Approved for LRN sentinel laboratories (see References: CDC/ASM/APHL 2002).
Isolates identified as Y pestis (or isolates that cannot be ruled out as Y pestis) should be forwarded to LRN reference or higher laboratories for confirmatory testing.
Biochemical screening tests:

TSI K/A (no gas or H₂S)
Oxidase-negative
Catalase-positive
Urea-negative
Indole-negative

Commercial identification tests:

As Y pestis is relatively nonreactive, commercial tests may identify it as Shigella species, H₂S-negative Salmonella, Acinetobacter species, or Yersinia pseudotuberculosis [Note: THIS IS EXTREMELY IMPORTANT TO CONSIDER].
Many automated commercial identification systems do not include Y pestis in the data bank.

Additional Tests for Detection, Confirmation, and Characterization of Y pestis

Some of the tests outlined below are available through LRN reference and national laboratories.

Direct fluorescent antibody stain (DFA):

Based on the F1 antigen.
Offers presumptive identification of Y pestis in patient samples and for cultures.
Specimens can be falsely negative if refrigerated for more than 30 hours.
Since the antigen is only expressed at temperatures above 35^oC, DFA tests performed on cultures incubated at lower temperatures may be falsely negative (see References: WHO: Plague manual).
The test may not work directly on fleas (see References: Perry 1997).

Bacteriophage lysis:

Used for culture confirmation.
Unlike F1 antigen expression, Y pestis is susceptible to phage lysis at both 25^oC and 37^oC (see References: CDC Plague Home Page/Diagnosis).

Antimicrobial susceptibility tests
Standard biochemical identification
Serology:

Paired sera (collected 4 to 6 weeks apart) can be used for retrospective diagnosis (see References: WHO: Plague manual).
Single titer of more than 1:10 is considered presumptively positive for plague if the patient has not been vaccinated previously or has a history of infection.
Single titer of more than 1:128 or a fourfold increase in paired sera is considered confirmatory (see References: CDC: Plague Home Page/Diagnosis).

An F1 antigen-capture enzyme-linked immunsorbent assay (ELISA) (see References: Chanteau 2000, Splettstoesser 2004, Seramun Diagnostica)
Polymerase chain reaction (PCR)-based assays (see References: Loiez 2003, McAvin 2003, Norkina 1994, Tomaso 2003, Zhou 2004: Identification of signature genes for rapid and specific characterization of Yersinia pestis)

A simultaneous real-time PCR assay can detect Bacillus anthracis (anthrax), Francisella tularenesis (tularemia), and Y pestis (see References: Skottman 2007).
A new multiplex real-time PCR assay proved sensitive and specific, with a lower detection limit of 10 to 100 fg of extracted Y pestis DNA (see References: Matero 2009).

Mouse inoculation is used to increase recovery of Y pestis with contaminated specimens (see References: CDC: Information on plague/Diagnosis).
Molecular-based subtyping tests (see References: Achtman 2004, Drancourt 2004, Huang 2002, Klevytska 2001, Lowell 2005, Pourcel 2004, Ciammaruconi 2008, Torrea 2006):

Multilocus variable-number tandem-repeat analysis (MLVA/VNTR)
Restriction length polymorphism (RFLP)
Pulsed-field gel electrophoresis (PFGE)
Multiple spacer typing (MST)
IS100 typing
Genome-wide synonymous single nucleotide polymorphisms (SNPs)

Immunohistochemistry (IHC):

IHC can be performed on formalin-fixed tissues.
One assay utilizing anti-F1 antibody has been described (see References: Guarner 2002).

Antimicrobial Susceptibility Studies

Four studies (see References) have examined antimicrobial susceptibilities of Y pestis strains to various antibiotics. Data from these studies are shown in the table below.

The major conclusions from each of the studies are as follows:

Frean 2003: Cefditoren and new fluoroquinolones were the most active, and macrolides the least active, in vitro.
Wong 2000: Low minimal inhibitory concentrations (MICs) were found for ampicillin, cefixime, ceftazidime, ceftriaxone, chloramphenicol, doxycycline, gentamicin, streptomycin, trimethoprim-sulfamethoxazole, and trovafloxacin. Significant resistance to imipenem and rifampin was noted.
Frean 1996: The most active agents in vitro were cefotaxime, levofloxacin, and ofloxacin. All isolates were susceptible to chloramphenicol, trimethoprim-sulfamethoxazole, tetracycline, doxycycline, streptomycin, and amoxicillin.
Smith 1995: Antibiotics used as traditional treatment options (eg, streptomycin, tetracycline, chloramphenicol) were less active than ceftriaxone, ciprofloxacin, ofloxacin, and ampicillin. Azithromycin showed poor activity against all strains.

MICs of Various Antibiotics for Yersinia pestis Isolates as Identified in Four Detailed Studies
	Study
	Frean 2003^a		Wong 2000^b		Frean 1996^c		Smith 1995^d
*Antibiotic*	*MIC range^e*	MIC₉₀^e	MIC range^e	MIC₉₀^e	MIC range^e	MIC₉₀^e	MIC range^e	MIC₉₀^e
ABT	0.008-0.016	0.016	—	—	—	—	—	—
Amoxicillin	—	—	—	—	<0.03-0.25	0.12	—	—
Ampicillin	—	—	0.094-0.38	0.38	—	—	0.125-0.5	0.5
Azithromycin	—	—	—	—	—	—	4.0-32.0	32.0
Cefdinir	0.063-0.125	0.125	—	—	—	—	—	—
Cefditoren	0.031-0.063	0.063	—	—	—	—	—	—
Cefotaxime	—	—	—	—	<0.03	<0.03	—	—
Cefixime	—	—	0.006-0.032	0.023	—	—	—	—
Ceftazidime	—	—	0.016-0.19	0.125	—	—	—	—
Ceftriaxone	—	—	0.006-0.032	0.023	—	—	0.008-0.031	0.031
Cethromycin	2-4	4	—	—	—	—	—	—
Chloramphenicol	—	—	0.25-4.0	2.0	0.06-2.0	1.0	0.5-4.0	4.0
Ciprofloxacin	0.016-0.031	0.031	—	—	—	—	0.008-0.062	0.062
Clarithromycin	4->32	>32	—	—	—	—	—	—
Doxycycline	0.25-0.5	0.5	0.125-2.0	1.5	<0.03-4.0	1.0	0.25-1.0	1.0
Erythromycin	16-32	32	—	—	<0.03->16.0	16.0	—	—
Gentamicin	—	—	0.19-1.0	0.75	—	—	0.25-1.0	1.0
Imipenem	—	—	0.094->32	>32	—	—	—	—
Levofloxacin	—	—	—	—	<0.03-0.06	<0.03	—	—
Ofloxacin	—	—	—	—	<0.03-0.12	<0.03	0.031-0.25	0.25
Olamufloxacin	0.008	0.016	—	—	—	—	—	—
Penicillin	—	—	—	—	—	—	0.25-2.0	2.0
Rifampin	—	—	2.0-32	16	<0.03-8.0	8.0	2.0-8.0	8.0
Streptomycin	—	—	1.5-4.0	3.0	<0.03-2.0	0.5	4.0-8.0	4.0
Temafloxacin	0.016-0.031	0.031	—	—	—	—	—	—
Tetracycline	—	—	—	—	<0.03-2.0	2.0	0.5-4.0	4.0
TMP-SMX	—	—	0.012-0.047	0.032	<0.03/ 0.59- 0.06/1.18	0.06/1.18	0.5/2.0-1.0/32.0	1.0/16.0
Tosufloxacin	0.004-0.008	0.008	—	—	—	—	—	—
Trovafloxacin	—	—	0.006-0.047	0.032	—	—	—	—
Abbreviations: MIC, minimal inhibitory concentration; MIC₉₀, minimal inhibitory concentration for 90% of isolates tested; TMP-SMX, trimethoprim-sulfamethoxazole. ^aHuman strains from Southern Africa (n=28) 1982-1991 (see References: Frean 2003). ^bHuman and animal strains from California (n=92) 1977 to 1998 (see References: Wong 2000). ^cHuman strains from Southern Africa (n=100) 1982 to 1991 (see References: Frean 1996). ^dHuman and animal strains from the Central Highlands of Vietnam (n=78) 1985 to 1993 (see References: Smith 1995). ^eValues in mcg/mL.

Additional antimicrobial susceptibility studies include the following:

Studies in the Russian literature have found that alternative aminoglycosides (eg, tobramycin and amikacin) are effective against Y pestis using in vitro and animal models (see References: Romanov 2001, Romanov 2001, Ryzhko 1998, Shcherbaniuk 1992).
A strain of Y pestis isolated from a patient in Madagascar was found to have plasmid-mediated multidrug resistance. The isolate was resistant to chloramphenicol, streptomycin, tetracycline, sulfonamides, tetracycline, ampicillin, kanamycin, spectinomycin, and minocycline. It was susceptible to cephalosporins, other aminoglycosides, quinolones, and trimethoprim (see References: Galimand 1997). Investigators have recently shown that this strain contains a self-transmissible plasmid (pIP1202) that confers resistance to many of the antimicrobials recommended for plague treatment and prophylaxis (see References: Welch 2007).
Gatifloxicin and moxifloxacin appeared efficacious against experimental plague in a mouse model (see References: Steward 2004).

Environmental Testing

Environmental testing generally is not considered necessary, since Y pestis does not sporulate and the organisms generally do not persist in the environment for prolonged periods of time. In a recent study, Y pestis cells were viable for up to 6 hours after drying on steel, 7 hours on glass, 24 hours on polyethylene, and up to 5 days on paper under controlled conditions (see References: Rose 2003).
If environmental testing is considered, samples should be tested at LRN reference or higher laboratories (see References: CDC/ASM/APHL 2002).
An autonomous pathogen detection system (APDS) has been described for Y pestis. It consists of an aerosol collector, an immunoassay subsystem, a flow-through PCR subsystem with sequential injection analysis, and a multianalyte flow-cytometry subsystem for PCR product detection (see References: McBride 2003, Hindson 2004, Hindson 2005, Langlois).
The handheld advanced nucleic acid analyzer (HANAA) is a real-time PCR analyzer utilizing a miniaturized thermal cycling process. It has been developed for anthrax and plague (see References: LLNL 2002).
A variety of other systems are available for detection/identification of Y pestis in the environment, but the practical applications of these systems are not always clear (see References: Alexeter Technologies, New Horizons Diagnostics, Defense Advanced Research Projects Agency, Research International, Rider 2003, Kenny 2008).

Treatment, Postexposure Prophylaxis, and Vaccines

General Considerations

Traditionally, streptomycin, tetracycline, and doxycycline have been used for the treatment of plague and are approved by the FDA for this indication. Gentamicin also has been shown to be efficacious in the treatment of plague (although it is not currently approved by the FDA) (see References: Boulanger 2004, Crook 1992, Welty 1985).

Doxycycline and gentamicin regimens have been compared in mice receiving aerosolized Y pestis. Survival was similar with both drugs, although because doxycylcine behaved in vivo as a bacteriostatic drug, it required an intact immune system for clearance of infection after challenge (see References: Heine 2007).

Clinical experience with the fluoroquinolones in treating plague is limited; however, animal studies have suggested efficacy in this setting (see References: Russell 1996). An in vitro pharmacodynamic infection model showed that a regimen of levofloxacin was superior to a regimen of streptomycin (see References: Louie 2007). Streptomycin therapy caused a reductionin the number of bacteria over 24 hours, followed by regrowth withstreptomycin-resistant mutants. Levofloxacin resulted in a greater reduction in the number of bacteria within 12 hours, and ultimatelysterilized the culture without resistance selection.

A recent randomized, comparative, open-label clinical trial involving monotherapy with gentamicin or doxycycline found that both antibiotics were effective in treatment of plague (see References: Mwengee 2006). The patients studied had bubonic, septicemic, or pneumonic plague; 35 patients were randomized to receive gentamicin and 30 received doxycycline. Three patients died (two were treated with gentamicin and one with doxycycline); all had advanced disease or complications at the start of therapy. The overall effectiveness of treatment was 94% for gentamicin and 97% for doxycycline.

Rifampin, aztreonam, ceftazidime, cefotetan, and cefazolin have been shown to not be efficacious and should not be used to treat plague.

Some investigators have suggested alternative therapies for treatment of plague, such as immunotherapy, non-pathogen-specific immunomodulatory therapy, bacteriocin therapy, treatment with inhibitors of virulence factors, and phage therapy (see References: Anisimov 2006).

Treatment for Pneumonic Plague

The Working Group on Civilian Biodefense has developed consensus-based recommendations for treatment of pneumonic plague during a bioterrorist attack (see References: Inglesby 2000). The working group made the following recommendations:

In a contained casualty setting where the medical care delivery system can effectively manage the number of patients, parenteral antibiotics should be administered to all patients whenever possible, according to the table below.
In a mass casualty setting where the medical care delivery system is not able to meet the demands for patient care, use of oral antibiotics may be necessary. In such a situation, the medications listed above under the table on postexposure antibiotic prophylaxis should be used, except that treatment should be given for 10 days instead of 7 days.
Supportive care of patients also is critical, including fluid management and hemodynamic monitoring. Many patients would require intensive care with respiratory support owing to complications of gram-negative sepsis (eg, shock, adult respiratory distress syndrome, multisystem failure, DIC).
Bioterrorist use of a Y pestis strain resistant to conventional antibiotic therapy is of concern and should be considered, particularly if patients deteriorate despite early initiation of antibiotic therapy. Recently, a clinical isolate of Y pestis with plasmid-mediated multidrug resistance was identified in Madagascar (see References: Galimand 1997).

Recommendations From the Working Group on Civilian Biodefense for Treatment of Pneumonic Plague During a Bioterrorism Event
Choices by Patient Category	Therapy Recommendations^a
Adults: Preferred choices	Streptomycin, 1 gm IM twice daily for 10 days^b or Gentamicin, 5 mg/kg IM or IV once daily or 2 mg/kg loading dose followed by 1.7 mg/kg IM or IV 3 times daily for 10 days^c,d
Adults: Alternative choices^e	Doxycycline, 100 mg IV twice daily or 200 mg IV once daily for 10 days^d or Ciprofloxacin, 400 mg IV twice daily for 10 days^d,f or Chloramphenicol, 25 mg/kg IV 4 times daily for 10 days^g
Children: Preferred choices	Streptomycin, 15 mg/kg IM twice daily (maximum daily dose, 2 gm) or Gentamicin 2.5 mg/kg IM or IV 3 times daily for 10 days^c
Children: Alternative choices^e	Doxycycline: >45 kg, give adult dosage <45 kg, give 2.2 mg/kg IV twice daily for 10 days (maximum, 200 mg/day) or Ciprofloxacin, 15 mg/kg IV twice daily for 10 days (maximum daily dose, 1 gm)^f or Chloramphenicol, 25 mg/kg IV 4 times daily for 10 days (maximum daily dose, 4 gm)^g,h
Abbreviations: IM, intramuscularly; IV, intravenously. ^aThese recommendations are most appropriate for the contained casualty setting where resources are adequate to treat all patients with intravenous antibiotics. In the mass casualty setting where the medical care delivery system is not able to meet the demands for patient care, oral antibiotics may need to be substituted for intravenous antibiotics for treatment of patients with plague. In such a situation, the recommendations in the table on postexposure prophylaxis should be followed for treatment, except that treatment should be continued for 10 days instead of 7 days. ^bStreptomycin is not acceptable for use in pregnant women because irreversible deafness in children exposed in utero has been reported. ^cAminoglycosides must be adjusted according to renal function. ^dAcceptable for pregnant women. Although fetal toxicity may occur with doxycycline use, the Working Group recommended doxycycline or ciprofloxacin if gentamicin is not available or if oral antibiotics must be used. ^eTrimethoprim-sulfamethoxazole has been used successfully to treat plague; however, the Working Group considers this agent as a second-tier choice. ^fOther fluoroquinolones may be substituted at dosages appropriate for age. ^gConcentration should be maintained between 5 and 20 mcg/mL; concentrations >25 mcg/mL can cause reversible bone marrow suppression. The oral formulation is available only outside the United States. Some experts have recommended that chloramphenicol be used to treat patients with plague meningitis, since chloramphenicol penetrates the blood-brain barrier (see References: Dennis 1997, Butler 1991, AAP 2006); however, controlled trials to verify improvement in outcome have not been performed. ^hAccording to the Working Group, children younger than 2 years of age should not receive chloramphenicol. However, the American Academy of Pediatrics (AAP) has recommended chloramphenicol as the drug of choice for treating plague meningitis in children. The AAP Red Book does not indicate that chloramphenicol should not be given to children with serious infections who are beyond the newborn period but younger than 2 years of age (see References: AAP 2006). Adapted from Inglesby 2000 (see References).

Treatment for Bubonic Plague

The initial cases in a bioterrorist attack would be expected to be pneumonic plague from exposure to the initial aerosol release and the occurrence of secondary cases with respiratory droplet transmission. It is also conceivable that, as the epidemic progressed, some cases of bubonic plague might occur from contact with infected animals in the area (ie, bites, scratches) or even from subsequent flea bites (see References: McGovern 1999).

Antibiotic treatment of bubonic plague is the same as therapy for pneumonic plague. Usually buboes will recede without intervention, but if they become fluctuant or secondarily infected, they made need incision and drainage.

Postexposure Prophylaxis for Pneumonic Plague

Antibiotic prophylaxis (with tetracycline, doxycycline, or trimethoprim-sulfamethoxazole) following exposure to a person with primary or secondary pneumonic plague has been recommended as a public health control measure (see References: ACIP/CDC 1996). Tetracycline and doxycycline are approved by the Food and Drug Administration (FDA) for this indication. Clinical experience with the fluoroquinolones for prophylaxis against pneumonic plague is limited; however, animal studies have suggested efficacy in this setting (see References: Russell 1996).

Few data are available on the efficacy of postexposure prophylaxis in this setting; however, according to a CDC report published in 1984, over 2,000 persons had been placed on prophylactic antibiotics and no cases of person-to-person transmission had been reported (see References: CDC 1984). In fact, the CDC has not received any reports of person-to-person Y pestis transmission in the United States since the last US outbreak of pneumonic plague in Los Angeles in the 1920s.

The Working Group on Civilian Biodefense developed consensus-based recommendations in 2000 for treatment and postexposure prophylaxis of pneumonic plague during a bioterrorist attack (see References: Inglesby 2000). The working group made the following recommendations:

Any potentially exposed persons in the affected community in whom a temperature of 38.5°C or higher or a new cough develops should be evaluated and placed on appropriate parenteral therapy (if available) for presumptive pneumonic plague. (Note: the Working Group did not recommend mass use of antibiotic prophylaxis for the entire population in the affected community in response to a release of Y pestis organisms.)
Persons who have close contact with a patient with pneumonic plague who has not received at least 48 hours of appropriate antibiotic therapy should receive antibiotic postexposure prophylaxis, as outlined in the table below. Since transmission occurs through respiratory droplets, close contact is defined as contact at less than 2 meters.
Persons who develop fever or cough while receiving antibiotic prophylaxis should be evaluated immediately for pneumonic plague and treated appropriately if plague is suspected.

Recommendations from the Working Group on Civilian Biodefense for Antib iotic Postexposure Prophylaxis During an Outbreak of Pneumonic Plague Following a Bioterrorism Event
Choices by Patient Category	Therapy Recommendations^a
Adults: Preferred choices	Doxycycline, 100 mg PO twice daily for 7 days^b,c or Ciprofloxacin, 500 mg PO twice daily for 7 days^c,d
Adults: Alternative choice^e	Chloramphenicol, 25 mg/kg PO 4 times daily for 7 days^f
Children: Preferred choices	Doxycycline: if >45 kg, give adult dosage; if <45 kg, give 2.2 mg/kg PO twice daily for 7 days^b o Ciprofloxacin, 20 mg/kg PO twice daily for 7 days (maximum daily dose, 1 gm)^d
Children: Alternative choice^e	Chloramphenicol, 25 mg/kg PO 4 times daily for 7 days (maximum daily dose, 4 gm)^f,g
Abbreviation: PO, orally. ^aRecommendations were reached by consensus of the Working Group on Civilian Biodefense and may not necessarily be approved by the Food and Drug Administration. Although these recommendations are intended for postexposure prophylaxis, they also can be used for treatment of plague cases in the mass casualty setting where the number of patients is too great for all patients to receive intravenous antibiotics and oral antibiotics must be substituted (except that treatment should be continued for 10 days instead of 7 days as for prophylaxis). ^bTetracycline can be substituted for doxycycline at a dose of 10-25 mg/kg/day divided into 2-4 doses. ^cAcceptable for pregnant women. Although fetal toxicity may occur with doxycycline use and toxic effects on the liver in pregnancy have been noted with the tetracycline class, the Working Group recommended doxycycline or ciprofloxacin for postexposure prophylaxis of pregnant women or for treatment of infection in the mass casualty setting. ^dOther fluoroquinolones may be substituted at dosages appropriate for age. ^eTrimethoprim-sulfamethoxazole (40 mg sulfa/kg/day administered orally in 2 divided doses for 7 days) has been recommended for postexposure prophylaxis in children younger than 8 years old and pregnant women (see References: AAP 2006, McGovern 1997). ^fConcentration should be maintained between 5 and 20 mcg/mL; concentrations >25 mcg/mL can cause reversible bone marrow suppression. The oral formulation is available only outside the United States ^gAccording to the Working Group, children younger than 2 years of age should not receive chloramphenicol. Adapted from Inglesby 2000 (see References).

Plague Vaccines

Killed Whole-Cell Vaccine

A licensed vaccine against plague was available in the United States until 1999 (see References: ACIP/CDC 1996), when the sole manufacturer discontinued production.

The vaccine was a formalin-inactivated whole-cell vaccine.
The vaccine primarily induced antibodies to the F1 capsular antigen.
The vaccine protected against bubonic plague but did not protect against pneumonic plague.
Data on efficacy are limited; however, US military servicemen were vaccinated during the Vietnam War and even though plague was prevalent in the local animal and human populations at the time, only eight cases of plague occurred among immunized US servicemen (see References: Titball 2001). The estimated plague incidence in Vietnamese civilians was 333 cases per 10⁶ population versus 1 case per 10⁶ population among US servicemen.
Administration of a heat-killed vaccine against plague provided long-lasting T-cell responses, comparable to those of conventional vaccines such as pertussis or tetanus-diphtheria, among Gulf War participants. The study was conducted among British soldiers vaccinated at the time of the first Gulf War (1990-1991) (see References: Allen 2006).

The EV76 Live Attenuated Vaccine

The EV76 vaccine was initially developed in the early 1900s and has been used since that time in some parts of the world (particularly in the former Soviet Union). However, the vaccine strain is not avirulent and its safety in humans has been questioned. This vaccine preparation is not available in the United States.

New Plague Vaccines

Research is ongoing to develop new and improved plague vaccines, particularly in light of the current bioterrorist threat and concerns about intentional dissemination of aerosolized plague organisms. Approaches for new vaccine development include the following:

One approach is to create vaccines using live attenuated mutant strains of Y pestis or related organisms (such as Salmonella or attenuated Y pseudotuberculosis) (see References: Bubeck 2007, Calhoun 2006, Liu 2007, Yang 2007, Blisnick 2008).

Another approach is to create subunit vaccines that selectively target antigens on the surface of the plague bacillus. Most candidate vaccines involve the F1 capsular antigen and the V antigen (also referred to as LcrV [low-calcium-response] antigen) (see References: Alvarez 2006, Anderson 1998, Chiuchiolo 2006, Elvin 2006, Jones 2003, Palin 2007, Rocke 2008, Titball 2001, Tripathi 2006, Wang 2007, Williamson 2007, Thomas 2009).
Combinatorial vaccines involving antigens against more than one bacterial agent may represent the next generation in biodefense vaccines (DuBois 2007).

Hospital Infection Control (Including Autopsies and Burial)

Isolation Precautions

Droplet Precautions plus eye protection, in addition to Standard Precautions, should be implemented for patients with pneumonic plague. Although Droplet Precautions are generally accepted as adequate for protection against pneumonic plague, some experts have questioned whether or not Droplet Precautions are sufficient to protect healthcare workers.

Patients are considered infectious for 48 to 72 hours after initiation of appropriate antibiotic therapy with evidence of clinical improvement (see References: APIC/CDC 1999; Inglesby 2000; Weber 2001).

Standard Precautions include the following practices related to direct patient care (see References: CDC/HICPAC 1996):

Handwashing:

Wash hands after touching blood, body fluids, secretions, excretions, or contaminated items, whether or not gloves are worn.
Wash hands immediately after gloves are removed, between patient contacts, and when otherwise indicated to avoid transfer of microorganisms to other patients or environments.

Gloves:

Wear gloves when touching blood, body fluids, secretions, excretions, or contaminated items; put on clean gloves just before touching mucous membranes and nonintact skin.
Change gloves between tasks and procedures on the same patient after contact with material that may contain a high concentration of microorganisms.
Remove gloves promptly after use, before touching noncontaminated items or environmental surfaces, and before going to another patient, and wash hands immediately to avoid transfer of microorganisms to other patients or environments.

Masks, eye protection, face shields:

Wear a mask and eye protection or a face shield to protect mucous membranes of the eyes, nose, and mouth during procedures and patient-care activities that are likely to generate splashes or sprays of blood, body fluids, secretions, or excretions.

Gowns:

Wear a gown to protect skin and prevent soiling of clothing during procedures and patient-care activities that are likely to generate splashes or sprays of blood, body fluids, secretions, or excretions.
Select a gown that is appropriate for the activity and amount of fluid likely to be encountered.
Remove a soiled gown as promptly as possible and wash hands.

Patient-care equipment:

Handle used equipment soiled with blood, body fluids, secretions, or excretions in a manner that prevents skin and mucous membrane exposures, contamination of clothing, and transfer of microorganisms to other patients and environments.
Ensure that reusable equipment is not used for the care of another patient until it has been appropriately cleaned and reprocessed; single-use items should be appropriately discarded.

Droplet Precautions include the following (see References: CDC/HICPAC 1996):

Place the patient in a private room or in a room with other patients who have the same infection (ie, cohort).
When a private room is not available and cohorting is not possible, a spatial separation of at least 3 ft should be maintained between the patient and other patients or visitors. (Note: Other sources suggest that contact within 2 m [6.5 ft] can spread the disease.)
Healthcare workers should wear a mask when working within 3 ft of the patient; a standard surgical mask is considered adequate.
Patient transportation should be limited to essential purposes only. If transport or movement is necessary, minimize dispersal of droplets from the patient by masking the patient (using a standard surgical mask).
Negative-pressure rooms or other special air-handling measures are not necessary for routine care and doors may remain open.

Contact and Droplet Precautions should be implemented when buboes are being aspirated or irrigated, owing to the propensity for aerosolization of infectious material. Contact Precautions include the following (see References: CDC/HICPAC 1996):

Place the patient in a private room, or if a private room is not available, place the patient in a room with a patient who has an active infection with the same pathogen (ie, cohort). When a private room is not available and cohorting is not possible, a spatial separation of at least 3 ft should be maintained between the infected patient and other patients and visitors. (Note: Other sources suggest that contact within 2 m [6.5 ft] can spread the disease.)
Gloves should be worn when entering the room and removed before leaving the room; hands should be washed with an antimicrobial agent or a waterless handwashing agent immediately after removing gloves, and clean hands should not touch potentially contaminated items or environmental surfaces. Gloves should be changed during the course of patient care following contact with infective material that may contain high concentrations of microorganisms.
Gowns should be worn when entering the room if it is anticipated that clothing will have substantial contact with the patient, environmental surfaces, or items in the room; the gown should be removed before leaving the patient's environment.
Patient transport should be limited to essential purposes only; if the patient is transported out of the room, precautions should be maintained.
Noncritical patient-care equipment should be dedicated whenever possible. If equipment cannot be dedicated, then it should be adequately cleaned and disinfected between patients.

Decontamination

In general, environmental decontamination following an aerosol event has not been recommended, since experts have estimated that an aerosol of Y pestis organisms would be infectious for only about 1 hour (see References: Inglesby 2000).
A recent study demonstrated that Y pestis can survive on selected environmental surfaces for at least several days, with survival being the longest on paper (cells were still viable after 120 hours) (see References: Rose 2003), although the potential for re-aerosolization of these organisms was not addressed.
Commercially available bleach or 0.5% hypochlorite solution (1:10 dilution of household bleach) is considered adequate for cleaning. Experiments show that vapor-phase hydrogen peroxide may be used for decontamination in circumstances in which liquid or heat decontamination may not be suitable (eg, to meet time-sensitive schedules and activities) (see References: Rogers 2008).

Issues Related to Autopsies and Burial

Autopsy Practices

Recent guidelines from the CDC indicate that Standard Precautions should be used for postmortem care. These include using a surgical scrub suit, surgical cap, impervious gown or apron with full sleeve coverage, a form of eye protection (eg, goggles or face shield), shoe covers, and double surgical gloves with an interposed layer of cut-proof synthetic mesh (see References: CDC 2004).
In addition, autopsy personnel should wear N-95 respirators during all autopsies, regardless of suspected or known pathogens. Powered air-purifying respirators (PAPRs) equipped with N-95 or high-efficiency particulate air (HEPA) filters should be considered.
Other experts have recommended that aerosol-generating procedures (such as bone sawing) should be avoided during autopsies if possible. If such procedures are necessary, then high-efficiency particulate air–filtered masks and negative-pressure rooms should be used (see References: Inglesby 2000; Weber 2001).

Burial

Contact with corpses should be limited to trained personnel and routine precautions should be implemented when transporting corpses.
Bodies infected with biological terrorism agents should not be embalmed (see References: CDC 2004). Bodies infected with Y pestis can be directly buried without embalming.

Public Health Reporting and Case Definitions

Plague is a reportable disease. Suspect or confirmed cases must be reported to public health authorities as soon as possible, according to disease-reporting rules within each state or local jurisdiction.

In 1997, the Council of State and Territorial Epidemiologists (CSTE) and the CDC revised the public health case definitions for conditions under public health surveillance (see References: CDC 1997). The current public health case definitions for plague include the following.

Clinical Description

Plague is transmitted to humans by fleas or by direct exposure to infected tissue or respiratory droplets; the disease is characterized by fever, chills, headache, malaise, prostration, and leukocytosis that manifests in one or more of the following principal clinical forms:

Regional lymphadenitis (bubonic plague)
Septicemia without an evident bubo (septicemic plague)
Plague pneumonia, resulting from hematogenous spread in bubonic or septicemic cases (secondary pneumonic plague) or inhalation of infectious droplets (primary pneumonic plague)
Pharyngitis and cervical lymphadenitis resulting from exposure to larger infectious droplets or ingestion of infected tissue (pharyngeal plague)

Laboratory Criteria for Diagnosis

Presumptive:

Serum antibody titer(s) to Y pestis F1 antigen of 1:10 or more (without documented fourfold or greater change) in a patient with no history of plague vaccination or
Detection of F1 antigen in a clinical specimen by fluorescent assay

Confirmatory:

Isolation of Y pestis from a clinical specimen or
Fourfold or greater change in serum antibody titer to Y pestis F1 antigen

Case Classifications

Suspected: A clinically compatible case without presumptive or confirmatory laboratory results

Probable: A clinically compatible case with presumptive laboratory results

Confirmed: A clinically compatible case with confirmatory laboratory results

Images

Images of Y pestis organisms, characteristic buboes, skin lesions, and chest films can be found at the following CDC Web sites (see References):

CDC: Plague images
CDC: Public Health Image Library

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