Both B. mallei and B. pseudomallei can be cultured in a laboratory; nutrient agar can be used to grow the bacteria. When grown in culture, B. mallei grows in smooth, grey, translucent colonies. In a period of 18 hours at 37 °C, a B. mallei colony can grow to about 0.5–1.0 mm in diameter. B. mallei culture growth on MacConkey agar is variable.
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B. mallei is responsible for causing glanders disease, which historically mostly affected animals, such as horses, mules, and donkeys, and rarely humans. Horses are considered the natural host for B. mallei infection and are highly susceptible to it. B. mallei infects and gains access to the cell of its host through lysis of the entry vacuole. B. mallei has bacterial protein-dependent, actin-based motility once inside the cell.
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B. mallei was first called "Bacillus mallei" and was in the genus Pseudomonas until the early 1990s. It has also been referred to as "farcy". It is now part of the genus Burkholderia.
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In regards to a vaccine against B. mallei, the closeness of B. mallei to B. pseudomallei may make it possible that a vaccine developed for either type would be effective against the other.
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In the first days of B. mallei infection, neutrophils, macrophages, and T cells go to the spleen in great quantities. The early cellular response to B. mallei infection involves Gr-1+ (antigen) cells, and implies their importance to immunity against this bacterial infection. T cells (nitric oxide) are actually more involved in combating B. mallei in the later stages of its infection of a host. Lipopolysaccharide isolated from B. mallei demonstrated significantly lower biological activity as compared to the LPS from Escherichia coli, in agreement with the lower degree of acylation of its lipid A: the major forms of B. mallei lipid A were penta- and tetraacylated, whereas classical lipid A from E. coli was hexaacylated.
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Most organisms within the Burkholderiaceae live in soil; however, B. mallei does not. Because B. mallei is an obligate mammalian pathogen, it must infect a host mammal to live and to be transmitted from one host to another.
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It includes some pathogenic species, such as Burkholderia mallei (glanders) and Burkholderia pseudomallei (melioidosis).
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This suggestion has found support from studies that compare strains of B. mallei to B. pseudomallei and indicate that their two respective genomes are very similar. The genes that allowed the bacterium to survive in a soil environment, like genes that gave B. mallei the capacity to protect against bactericidals, antibiotics, and antifungals, were likely deleted. Thus, the reason that B. mallei is not found outside of a host is because it lacks the genes necessary for survival in the soil. Genome comparisons also seem to indicate that the B. mallei is still evolving and adapting to an intracellular lifestyle.
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Multilocus sequence typing has revealed that B. mallei most likely evolved from a B. pseudomallei clone reduction. About 1000 B. pseudomellei genes are absent or varying in the B. mallei genome. B. mallei’s genome also has a large amount of insertion sequences.
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B. mallei and B. pseudomallei have a history of being on a list of potential biological warfare agents. The Centers for Disease Control and Prevention classifies B. mallei as a category B critical biological agent. As a result, research regarding B. mallei may only be done in biosafety level 3 facilities in the US and internationally. Though it is so highly infective and a potential biological weapon, little research has been conducted on this bacterium.
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However, the Japanese did not end up creating a biological weapon out of B. mallei. They did actually use B. mallei to test its effectiveness in contaminating water supplies, and the results of these tests were successful. The Russians' biological weapons program also took an interest in B. mallei and conducted field tests with it. Some of the researchers from the program were actually infected and killed by it during the course of their research.
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Interest in melioidosis has been expressed because it has the potential to be developed as a biological weapon. Another similar bacterium, B. mallei, was used by the Germans in World War I to infect livestock shipped to Allied countries. Deliberate infection of human prisoners of war and animals using B. mallei were carried out in China's Pingfang District by the Japanese during World War II. The Soviet Union reportedly used B. mallei during the Soviet–Afghan War in 1982 and 1984. B. pseudomallei, like B. mallei, was studied by both the US and Soviet Union as a potential biological warfare agent, but never weaponized.
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If the bacteria enter through the skin, a local skin infection can result, while inhaling B. mallei can cause septicemic or pulmonary, muscular, hepatic, or splenous infections. B. mallei infection has a fatality rate of 95% if left untreated, and a 50% fatality rate in individuals treated with antibiotics.
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In addition, lipid A from B. mallei contains 4-amino-4-deoxyarabinose residue in almost half of the molecules, which would partially neutralize the negative charge of the phosphate groups necessary for the interaction with the positively charged amino acids of TLR4. At the same time, lipid A acyl chains in B. mallei were on the average longer (14–16 carbon atoms) than those in E. coli (14 carbon atoms), yet LPS from B. mallei appeared to be a weaker activator. B. mallei may employ LPS with low biological activity to evade proper recognition by the TLR4/MD-2 complex of innate immune system, dampening the host immune response and increasing the risk of bacterial dissemination.
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It has been suggested that the Russians eventually used B. mallei during their war in Afghanistan against the mujahideen.
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Glanders is caused by infection with the Burkholderia mallei, usually by ingestion of contaminated feed or water.B. mallei is able to infect humans, so is classed as a zoonotic agent. Transmission occurs by direct contact with infected animal's body fluid and tissues and entry is through skin abrasions, nasal and oral mucosal surfaces, or inhalation.
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The genome for B. mallei is made up of two circular chromosomes. Chromosome 1 is where genes relating to metabolism, capsule formation, and lipopolysaccharide biosynthesis are located. B. mallei has a polysaccharide capsule which indicates its potential as a pathogen. Chromosome 2 is where most of the information regarding secretion systems and virulence-associated genes are located.
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Burkholderia mallei is a Gram-negative, bipolar, aerobic bacterium, a human and animal pathogen of genus Burkholderia causing glanders; the Latin name of this disease (malleus) gave its name to the species causing it. It is closely related to B. pseudomallei, and by multilocus sequence typing it is a subspecies of B. pseudomallei. B. mallei evolved from B. pseudomallei by selective reduction and deletions from the B. pseudomallei genome. Unlike B. pseudomallei and other genus members, B. mallei is nonmotile; its shape is coccobacillary measuring some 1.5-3.0 μm in length and 0.5-1.0 μm in diameter with rounded ends.
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B. mallei has been eradicated in the United States and most Western countries, but still affects animals in Africa, Asia, the Middle East, Central America, and South America. Many Western countries were able to eliminate the disease through glanders control programs and laws requiring notification of cases of infection to health departments and the destruction of any animal affected with B. mallei.
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B. mallei is a clone of B. pseudomallei that has lost substantial portions of its genome as it adapted to live exclusively in mammals.
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B. mallei is very closely related to B. pseudomallei, being 99% identical in conserved genes when compared to B. pseudomallei. B. malllei has about 1.4 Mb less DNA than B. pseudomallei. B. mallei may have actually evolved from a strain of B. pseudomallei after the latter had infected an animal. The bacterium would have lost the genes that were not necessary for living in an animal host.
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No vaccine is currently available for humans or animals to protect against B. mallei infection. An animal model that will predict immune responses necessary to create immunity to the bacterium is needed before a vaccine can be developed. Mice are fairly close to humans in their susceptibility to B. mallei and would be the ideal choice of animal for creating a model for the vaccine.
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Many microbiologists are unfamiliar with B. mallei and as a result it has frequently been misidentified as a Pseudomonas species or as a contaminant in a culture.
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No standardised system exists for differentiating between B. mallei and B. pseudomallei. The methods that have been used to differentiate and identify one strain from the other include ribotyping, pulsed-field gel electrophoresis, multilocus enzyme electrophoresis, random amplified polymorphic DNA analysis, and multilocus sequence typing. Comparing the DNA of B. mallei and B. pseudomallei must be done at the 23S rDNA level, however, since no identifiable difference is found between the two species at the 16S rDNA level.
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B. mallei and B. pseudomallei under the policy of Institutional Oversight of Life Sciences Dual Use Research of Concern would be subject to oversight to ensure the responsible investigation of these agents.
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Horses chronically infected with B. mallei with glanders disease typically experience mucus-containing nasal discharge, lung lesions, and nodules around the liver or spleen. Acute infection in horses results in a high fever, loss of fat or muscle, erosion of the surface of the nasal septum, hemorrhaging or mucus discharge. The bacterium mostly affects the lungs and airways. Human infection with B. mallei is rare, although it occasionally occurs among laboratory workers dealing with the bacteria or those who are frequently near infected animals.
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Although Mallein is the most commonly used form of testing for glanders, cross reactions were reported between Burkholderia mallei and Streptococcus equi, which is a bacteria-caused contagious upper respiratory tract infection of equines. This resulted in false-positive reactions.
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The mallein test is a sensitive and specific clinical test for glanders. Mallein (ATCvet code: ), a protein fraction of the glanders organism (B. mallei), is injected intradermopalpebrally or given by eye drop. In infected animals, the eyelid swells markedly in 1 to 2 days.
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U.S. interest in glanders (agent LA) continued through the 1950s, except it had an inexplicable tendency to lose virulence in the lab, making it difficult to weaponize. Between 1982 and 1984, the Soviet Union allegedly used weaponized B. mallei during the Soviet–Afghan War.
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B. mallei was intentionally used to infect animals and humans during World War I. The Germans used B. mallei to infect animals that were being sent from neutral countries to the Allies with glanders. The Germans' plans for biological warfare started in 1915 on the East Coast of the United States; they intended to infect and kill the livestock that were being sent to the Allies and facilitate the transfer of the disease to humans. The East Coast was where many animals were being assembled for shipment to the Allies fighting in Europe. The Germans also targeted Romania, Norway, and Spain's animal supplies with cultures of glanders.
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Mallein, a protein fraction of B. mallei, is usually injected by an eye-drop. If an animal is infected, the animal will show swelling in the eye from around 48 hours of injection and may be accompanied by secretion and conjunctivitis. Mallein is non toxic to normal animals.
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Glanders is a contagious zoonotic infectious disease that occurs primarily in horses, mules, and donkeys. It can be contracted by other animals, such as dogs, cats, goats, and humans. It is caused by infection with the bacterium Burkholderia mallei. Glanders is endemic in Africa, Asia, the Middle East, and Central and South America.
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This is before the land connections were developed with Europe in the early Cenozoic era. The middle ear of nine families of golden moles (family Chrysochloridae) were examined to see the ossicular apparatus. The Amblysomus species have ossicles typical of mammals. The Chrysospalax, Chrysochloris, Cryptochloris and Eremitalpa species do not. They ”have enormously hypertrophied mallei.
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Burkholderia is a genus of Proteobacteria whose pathogenic members include the Burkholderia cepacia complex, which attacks humans and Burkholderia mallei, responsible for glanders, a disease that occurs mostly in horses and related animals; Burkholderia pseudomallei, causative agent of melioidosis; and Burkholderia cepacia, an important pathogen of pulmonary infections in people with cystic fibrosis (CF). The Burkholderia (previously part of Pseudomonas) genus name refers to a group of virtually ubiquitous Gram-negative, obligately aerobic, rod-shaped bacteria that are motile by means of single or multiple polar flagella, with the exception of Burkholderia mallei, which is nonmotile. Members belonging to the genus do not produce sheaths or prosthecae and are able to use poly-beta-hydroxybutyrate (PHB) for growth. The genus includes both animal and plant pathogens, as well as some environmentally important species.
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In animals, another similar organism named Burkholderia mallei is the causative agent of the disease glanders. B. pseudomallei can be differentiated from another closely related, but less pathogenic species, B. thailandensis, by its ability to assimilate arabinose. B. pseudomallei is highly adaptable to various host environments ranging from inside mycorrhizal fungi spores to amoebae. Its adaptability may give it a survival advantage in the human body.
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Mallein test being performed in 1918 The mallein test is a sensitive and specific clinical test for glanders, a common bacterial disease of equids (horses, donkeys, mules). This test is an allergic hypersensitivity test used as a diagnosis for glanders. It is caused by a bacterium called Burkholderia mallei, which is contagious for humans and other species. The occurrence of glanders must be reported to the World Organisation for Animal Health.
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Wilhelm Schütz and Friedrich Löffler first isolated B. mallei in 1882. It was isolated from an infected liver and spleen of a horse. This bacterium is also one of the first to be identified containing a type VI secretion system which is important for its pathogenicity. In 1885, the German Botanist and Bacteriologist, Wilhelm Zopf (1846-1909) gave the pathogen its binomial name, after analyzing samples of the bacterium.
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Major Alfred Whitmore (1876-1946) was an English pathologist who, together with C.S. Krishnaswami, identified Burkholderia pseudomallei, the causative agent of melioidosis (also known as "Whitmore's disease") in opium addicts in Rangoon in 1911. He differentiated it from Burkholderia mallei, the causative agent of glanders, by clinical and microbiological features. He was initially a Captain, and later a Major, in the Indian Medical Service. Later, he also became director of the Rangoon Medical School.
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B. pseudomallei is not fastidious and grows on a large variety of culture media (blood agar, MacConkey agar, EMB, etc.). Ashdown's medium (or Burkholderia cepacia medium) may be used for selective isolation. Cultures typically become positive in 24 to 48 hours (this rapid growth rate differentiates the organism from B. mallei, which typically takes a minimum of 72 hours to grow). Colonies are wrinkled, have a metallic appearance, and possess an earthy odour.
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Leaving the cell early also keeps the bacteria from being destroyed by lysosomal defensins and other pathogen-killing agents. MNGCs may help protect the bacteria from immune responses. B. mallei’s ability to live within the host cell makes developing a vaccine against it difficult and complex. The vaccine would need to create a cell-mediated immune response, as well as a humoral response to the bacteria in to be effective in protecting against B. mallei.
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Mobile genetic insertion sequences can play a role in genome rearrangement activities. Pathogens that do not live in an isolated environment have been found to contain a large number of insertion sequence elements and various repetitive segments of DNA. The combination of these two genetic elements is thought help mediate homologous recombination. There are pathogens, such as Burkholderia mallei, and Burkholderia pseudomallei which have been shown to exhibit genome-wide rearrangements due to insertion sequences and repetitive DNA segments.
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On Gram staining, the organism is a Gram-negative rod with a characteristic "safety pin" appearance (bipolar staining). On sensitivity testing, the organism appears highly resistant (it is innately resistant to many antibiotics including colistin and gentamicin) and that again differentiates it from B. mallei, which is in contrast, exquisitely sensitive to many antibiotics. For environmental specimens only, differentiation from the nonpathogenic B. thailandensis using an arabinose test is necessary (B. thailandensis is never isolated from clinical specimens).
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The bacterium is susceptible to numerous disinfectants including benzalkonium chloride, iodine, mercuric chloride, potassium permanganate, 1% sodium hypochlorite, and ethanol. The micro-organism can also be destroyed by heating or ultraviolet light. Antibiotics such as streptomycin, amikacin, tetracycline, doxycycline, carbapenems, ceftazidime, amoxicillin/clavulanic acid, piperacillin, chloramphenicol, and sulfathiazole have been reported to be effective against the bacteria in vitro. B. mallei, like B. pseudomallei, is also resistant to a number of antibiotics including aminoglycosides, polymyxins, and beta-lactams.
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The next month, his symptoms seemed to disappear after treatment with clarithromycin, but after the medication was stopped, the symptoms reappeared. After conducting multiple tests on cultures from the researcher’s blood and a biopsied portion of a liver abscess, the bacterium was identified as B. mallei. Once it was established what infected the researcher, another course of antibiotics was given (imipenem and doxycycline) with 6 months of treatment. After a year, the researcher made a full recovery.
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The German biological sabotage eventually spread to Argentina, where agents would rely on bacterial cultures from Spain to infect the cattle, horses, and mules that Argentina was supplying to the Allies. The German use of microbes as weapons is one of the only documented attacks of intentionally using biological weapons against neutral countries. The Japanese used B. mallei in their biological warfare research units. The most notable and notorious unit, Unit 731, used the bacterium to conduct experiments on live human subjects.
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A pulmonary anthrax infection starts with ordinary influenza-like symptoms and progresses to a lethal hemorrhagic mediastinitis within 3–7 days, with a fatality rate that is 90% or higher in untreated patients. Finally, friendly personnel and civilians can be protected with suitable antibiotics. Agents considered for weaponization, or known to be weaponized, include bacteria such as Bacillus anthracis, Brucella spp., Burkholderia mallei, Burkholderia pseudomallei, Chlamydophila psittaci, Coxiella burnetii, Francisella tularensis, some of the Rickettsiaceae (especially Rickettsia prowazekii and Rickettsia rickettsii), Shigella spp.
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In particular, B. xenovorans (previously named Pseudomonas cepacia then B. cepacia and B. fungorum) is renowned for being catalase positive (affecting patients with chronic granulomatous disease) and its ability to degrade chlororganic pesticides and polychlorinated biphenyls. The conserved RNA structure anti-hemB RNA motif is found in all known bacteria in this genus. Due to their antibiotic resistance and the high mortality rate from their associated diseases, B. mallei and B. pseudomallei are considered to be potential biological warfare agents, targeting livestock and humans.
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Other agents attempted to introduce the disease in the United States and Argentina. This had an effect on troop and supply convoys, as well as on artillery movement, which were dependent on horses and mules. Human cases in Russia increased with the infections during and after WWI. The Japanese deliberately infected horses, civilians, and prisoners of war with B. mallei at the Unit 731 Pingfang (China) Institute and Unit 100 facilities during World War II. The U.S. studied this agent as a possible biological weapon in 1943–44, but did not weaponize it.
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It is also able to initiate host cell fusion that results in multinucleated giant cells (MNGCs). The consequence of MNGCs has yet to be determined, but it may allow the bacteria to spread to different cells, evade responses by the infected host’s immune system, or allow the bacteria to remain in the host longer. B. mallei is able to survive inside host cells through its capabilities in disrupting the bacteria-killing functions of the cell. It leaves the vacuoles early, which allows for efficient replication of the bacteria inside the cell.
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In March 2000, one of the first cases since the 1940s of glanders in the United States occurred in a young microbiologist working for the U.S. Army Medical Research Institute for Infectious Diseases. The researcher had type 1 diabetes and had been working with B. mallei for about two years, but he did not always wear gloves while conducting his research. The researcher experienced enlargement of the lymph nodes and a fever which lasted for 10 days even with antibiotic treatment. In the following weeks, the researcher experienced fatigue, rigors, night sweats, and loss of weight.
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Over the course of its history, the Soviet program is known to have weaponized and stockpiled the following eleven bio-agentsCook, Michelle Stem and Amy F. Woolf (April 10, 2002), Preventing Proliferation of Biological Weapons: U.S. Assistance to the Former Soviet States, (Congressional Research Service Report for Congress), pg 3. (and to have pursued basic research on many more): Bacillus anthracis (anthrax), Yersinia pestis (plague), Francisella tularensis (tularemia), Burkholderia mallei (glanders), Brucella sp. (brucellosis), Coxiella burnetii (Q-fever), Venezuelan equine encephalitis virus, (VEE), Botulinum toxin, Staphylococcal enterotoxin B, Smallpox, and Marburg virus. These programs became immense and were conducted at 52 clandestine sites employing over 50,000 people.
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Glanders was a significant problem for civilian use of horses, as well. In the 18th- century veterinary hospital at the École Nationale Vétérinaire d'Alfort, glanders was the most common disease among their equine patients and the one most likely to cause death. Due to the high mortality rate in humans and the small number of organisms required to establish infection, B. mallei is regarded as a potential biological warfare or bioterrorism agent, as is the closely related organism, B. pseudomallei, the causative agent of melioidosis. During World War I, glanders was believed to have been spread deliberately by German agents to infect large numbers of Russian horses and mules on the Eastern Front.
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Stipple effects were used in conjunction with other engraving techniques by artists as early as Giulio Campagnola (c.1482 – c. 1515) and Ottavio Leoni (1578–1630), although some of Campagnola's small prints were almost entirely in stipple.Mark J. Zucker in Kristin L. Spangenberg (ed), Six Centuries of Master Prints: Treasures from the Herbert Greer French collection, Cincinnati Art Museum, 1993, nos 39 & 40, In Holland in the seventeenth century, the printmaker and goldsmith Jan Lutma developed an engraving technique, known as opus mallei, in which the dots are punched into the plate by an awl struck with a hammer, while in England the faces of portraits were engraved with stippled dots by William Rogers in the sixteenth century and Lucas Vorsterman in the seventeenth.
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When World War I began, Dilger was in Germany, but he returned to the United States in 1915 with cultures of anthrax and glanders with the intention of biological sabotage on behalf of the German government's biological sabotage officer Rudolf Nadolny. The U.S. was then neutral, but Germany wanted to prevent neutral countries from supplying Allied forces with livestock, and the fact that Dilger had a U.S. passport from 1908 onwards made it easy for him to travel to and from America. Along with his brother Carl, Dilger established a laboratory in the Chevy Chase district north of Washington, DC in which cultures of the causative agents of anthrax and glanders—Bacillus anthracis and Burkholderia mallei—were produced. A 1941 report reveals that the bacteria were to be painted onto the nostrils of horses.
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Pathema was one of the eight bioinformatics resource centers funded by the National Institute of Allergy and Infectious Diseases (NIAID), a component of the National Institute of Health (NIH), which is an agency of the United States Department of Health and Human Services. Pathema was funded for five years from 2004 through a contract to The J. Craig Venter Institute, and is currently led by PI Granger Sutton. Pathema is the web resource for JCVI's NIAID-funded Bioinformatics Resource Center, and was one of eight such centers designed to support bio-defense and infectious disease research. The overarching goal of Pathema is to provide a core resource that will accelerated scientific progress towards understanding, detection, diagnosis and treatment of diseases caused by six clades of Category A-C pathogens (Bacillus anthracis, Clostridium botulinum, Burkholderia mallei, Burkholderia pseudomallei, Clostridium perfringens, and Entamoeba histolytica) involved in new and re-emerging infectious diseases.
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