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Bacillus cereus is typically considered a blood culture contaminant; however, its presence in blood cultures can indicate true bacteremia. We report 4 episodes of B. cereus bacteremia in 3 persons who inject drugs. Multilocus sequence typing showed that the temporally associated infections were caused by unrelated clones.

Background. Melioidosis results from infection with Burkholderia pseudomallei and is associated with casefatality rates up to 40%. Early diagnosis and treatment with appropriate antimicrobials can improve survival rates. Fatal and nonfatal melioidosis cases were identified in Puerto Rico in 2010 and 2012, respectively, which prompted contact investigations to identify risk factors for infection and evaluate endemicity. Methods. Questionnaires were administered and serum specimens were collected from coworkers, neighborhood contacts within 250 m of both patients’ residences, and injection drug user (IDU) contacts of the 2012 patient. Serum specimens were tested for evidence of prior exposure to B. pseudomallei by indirect hemagglutination assay. Neighborhood seropositivity results guided soil sampling to isolate B. pseudomallei. Results. Serum specimens were collected from contacts of the 2010 (n = 51) and 2012 (n = 60) patients, respectively. No coworkers had detectable anti–B. pseudomallei antibody, whereas seropositive results among neighborhood contacts was 5% (n = 2) for the 2010 patient and 23% (n = 12) for the 2012 patient, as well as 2 of 3 IDU contacts for the 2012 case. Factors significantly associated with seropositivity were having skin wounds, sores, or ulcers (odds ratio [OR], 4.6; 95% confidence interval [CI], 1.2–17.8) and IDU (OR, 18.0; 95% CI, 1.6–194.0). Burkholderia pseudomallei was isolated from soil collected in the neighborhood of the 2012 patient. Conclusions. Taken together, isolation of B. pseudomallei from a soil sample and high seropositivity among patient contacts suggest at least regional endemicityof melioidosis in Puerto Rico. Increased awareness of melioidosis is needed to enable early case identification and early initiation of appropriate antimicrobial therapy.

Cutaneous anthrax outbreaks occurred in Bangladesh from August to October 2009. As part of the epidemiological response and to confirm anthrax diagnoses, serum samples were collected from suspected case patients with observed cutaneous lesions. Anthrax lethal factor (LF), anti-protective antigen (anti-PA) immunoglobulin G (IgG), and anthrax lethal toxin neutralization activity (TNA) levels were determined in acute and convalescent serum of 26 case patients with suspected cutaneous anthrax from the first and largest of these outbreaks. LF (0.005-1.264 ng/mL) was detected in acute serum from 18 of 26 individuals. Anti-PA IgG and TNA were detected in sera from the same 18 individuals and ranged from 10.0 to 679.5 μg/mL and 27 to 593 units, respectively. Seroconversion to serum anti-PA and TNA was found only in case patients with measurable toxemia. This is the first report of quantitative analysis of serum LF in cutaneous anthrax and the first to associate acute stage toxemia with subsequent antitoxin antibody responses.

Anthrax is endemic throughout Africa, causing considerable livestock and wildlife losses and severe, sometimes fatal, infection in humans. Predicting the risk of infection is therefore important for public health, wildlife conservation and livestock economies. However, because of the intermittent and variable nature of anthrax outbreaks, associated environmental and climatic conditions, and diversity of species affected, the ecology of this multihost pathogen is poorly understood. We explored records of anthrax from the Serengeti ecosystem in north-west Tanzania where the disease has been documented in humans, domestic animals and a range of wildlife. Using spatial and temporal case-detection and seroprevalence data from wild and domestic animals, we investigated spatial, environmental, climatic and species-specific associations in exposure and disease. Anthrax was detected annually in numerous species, but large outbreaks were spatially localized, mostly affecting a few focal herbivores. Soil alkalinity and cumulative weather extremes were identified as useful spatial and temporal predictors of exposure and infection risk, and for triggering the onset of large outbreaks. Interacting ecological and behavioural factors, specifically functional groups and spatiotemporal overlap, helped to explain the variable patterns of infection and exposure among species. Synthesis and applications. Our results shed light on ecological drivers of anthrax infection and suggest that soil alkalinity and prolonged droughts or rains are useful predictors of disease occurrence that could guide risk-based surveillance. These insights should inform strategies for managing anthrax including prophylactic livestock vaccination, timing of public health warnings and antibiotic provision in high-risk areas. However, this research highlights the need for greater surveillance (environmental, serological and case-detection-orientated) to determine the mechanisms underlying anthrax dynamics. Keywords: Bacillus anthracis, disease ecology, exposure, infectious disease, multihost, serology, surveillance, susceptibility, zoonosis Introduction Bacillus anthracis (Cohn), the causative agent of anthrax, is a multihost pathogen affecting human, livestock and wildlife populations. The disease has a world-wide distribution, but has declined in many developed countries because of the implementation of livestock vaccination programmes and sanitary measures. However, anthrax is still endemic in Africa, with severe outbreaks causing significant losses in domestic and wild animal populations (Prins & Weyerhaeuser 1987; Shiferaw et al. 2002; Siamudaala 2006; Clegg et al. 2007; Wafula, Patrick & Charles 2008). Non-fatal cutaneous anthrax acquired through contact with infected carcasses or animal products accounts for >95% of all reported human cases globally (World Health Organization 2008). More severe forms, particularly gastrointestinal anthrax because of handling and consumption of inadequately cooked products from infected animals, may also exert a substantial burden that is both underreported and under-diagnosed (Sirisanthana & Brown 2002). Predicting the risk of infection is therefore important from the perspectives of public health, wildlife conservation and livestock economies. Despite the infamous reputation of anthrax, the ecology and transmission of the disease under natural conditions are not well understood (Hugh-Jones & de Vos 2002). The difficulty lies in the intermittent and variable nature of outbreaks, with considerable variation in the species affected, and associated environmental and climatic conditions (Table 1). Table 1 Anthrax outbreaks in Eastern and Southern Africa for which details were available. Predominant species are emboldened, followed by species in which > 5 cases were documented The life cycle of B. anthracis comprises a multiplication phase in the mammalian host and a persistence phase of spores in the soil. Transmission to herbivores largely occurs indirectly, so risk factors are usually associated with exposure to spores, rather than direct animal-to-animal transmission. Survival mechanisms of the pathogen outside the host remain unclear. Bacillus anthracis has been shown to be associated with plant roots (Saile & Koehler 2006), and this is suggested to be an adaptation that increases the likelihood of infecting ungulate hosts (Raymond et al. 2010). Certain environmental factors affect long-term spore survival and thus increase the risk of anthrax (Blackburn et al. 2007). For example, the endemicity of B. anthracis in some areas has been associated with calcium-rich and neutral-to-alkaline soils (van Ness 1971; Dragon et al. 2005; Hugh-Jones & Blackburn 2009), although strain differences exist in soil chemistry preferences (Smith et al. 2000). Strain differences, in fact, may also govern spread in anthrax epidemics (Blackburn et al. 2007; Garofolo et al. 2010). Seasonal incidence patterns suggest climatic factors (precipitation and ambient temperature) play an important role in triggering outbreaks, although these are not consistent between locations (Table 1), and therefore underlying mechanisms are debated. In some African ecosystems, outbreaks have been reported late in the dry season (Prins & Weyerhaeuser 1987; Turnbull et al. 1991; Lindeque & Turnbull 1994; de Vos & Bryden 1996; Shiferaw et al. 2002; Clegg et al. 2007; Muoria et al. 2007), leading to suggestions that transmission may be facilitated by close grazing, nutritional stress and congregation at water holes (Hugh-Jones & de Vos 2002). Elsewhere, outbreaks have been associated with heavy rains (Lindeque & Turnbull 1994; Mlengeya et al. 1998; Wafula, Patrick & Charles 2008; Bellan 2010), which are hypothesized to unearth spores and amplify vector populations (Durrheim et al. 2009; Hugh-Jones & Blackburn 2009; Lewerin et al. 2010). Experimental spore germination and vegetative growth in the rhizosphere provide a mechanism by which bursts of rainfall could spark epidemics (Saile & Koehler 2006). This would parsimoniously explain anthrax occurrences during dry summers in North America and Australia that have been preceded by rains and when animals are often in good body condition (Turner et al. 1999; Hugh-Jones & de Vos 2002; Parkinson, Rajic & Jenson 2003; Hugh-Jones & Blackburn 2009; Epp, Waldner & Argue 2010). Although anthrax infects a wide range of species, outbreaks are typically associated with just a few (Table 1), for example impala Aepyceros melampus, kudu Tragelaphus strepsiceros, buffalo Syncerus caffer and zebra Equus spp. (Prins & Weyerhaeuser 1987; Mlengeya et al. 1998; Clegg et al. 2007; Muoria et al. 2007; Wafula, Patrick & Charles 2008). Even within a single ecosystem, different species may be affected at different times (Lindeque & Turnbull 1994). Reasons for this remain unclear, but could be attributed to host-specific differences in susceptibility and exposure as a result of behavioural and ecological traits as well as differences in pathogen strains. The overall ecological and genetic factors that contribute to environmental persistence of anthrax, and the underlying conditions that initiate outbreaks and underlie patterns of circulation are poorly known. In the Serengeti ecosystem, anthrax has been reported in a variety of species, with sporadic outbreaks occurring in relatively localized endemic foci and mostly affecting a few focal species (Lembo et al. 2011). The ecosystem covers a large area (over 20 000 km2, Sinclair et al. 2008) encompassing a variety of habitats and environmental gradients, which may influence anthrax occurrence. Here, we draw on data gathered opportunistically including serology indicative of exposure and case reports from human, domestic and wild animal populations in the greater Serengeti ecosystem, to explore ecological factors affecting infection patterns in a range of species. Combined, the investigation into environmental and climatic predictors of anthrax and their associations with species-specific patterns of exposure and mortality provides a holistic picture of ecological drivers of anthrax dynamics and identifies priorities for further research in anthrax endemic ecosystems.

Bacillus anthracis, the bacterium that causes anthrax, is responsible for varying death rates among animal species. Difficulties in case detection, hazardous or inaccessible carcasses, and misdiagnosis hinder surveillance. Using case reports and a new serologic assay that enables multispecies comparisons, we examined exposure to and illness caused by B. anthracis in different species in the Serengeti ecosystem in Tanzania during 1996-2009 and the utility of serosurveillance. High seroprevalence among carnivores suggested regular nonfatal exposure. Seropositive wildebeest and buffalo showed that infection was not invariably fatal among herbivores, whereas absence of seropositivity in zebras and frequent detection of fatal cases indicated high susceptibility. Exposure patterns in dogs reflected known patterns of endemicity and provided new information about anthrax in the ecosystem, which indicated the potential of dogs as indicator species. Serosurveillance is a valuable tool for monitoring and detecting anthrax and may shed light on mechanisms responsible for species-specific variability in exposure, susceptibility, and mortality rates.

Bacillus anthracis, the etiological agent of anthrax, is a gram-positive, spore-forming rod, with colonies exhibiting a unique ground-glass appearance, and lacking hemolysis and motility. In addition to these phenotypes, several others traits are characteristic of B. anthracis such as susceptibility to gamma phage, the presence of two virulence plasmids (pX01 and pX02), and specific cell wall and capsular antigens that are commonly detected by direct fluorescent-antibody assays. We report on the identification and characterization of 14 Bacillus megaterium and four Bacillus sp. clinical isolates that are nonhemolytic, nonmotile, and produce a capsule antigenically similar to B. anthracis. This work furthers our understanding of Bacillus diversity and the limitations of the assays and phenotypes that are used to differentiate species in this genus. Further work is necessary to understand whether these strains are opportunistic pathogens or just contaminates.

Currently there are no commercially available selective media indicated for the isolation of Burkholderia mallei and Burkholderia pseudomallei. Ashdown's agar, a custom selective medium for isolation of B. pseudomallei, is well described in the literature but unavailable commercially. Three commercially available media, Burkholderia cepacia selective agar (BCSA), oxidative-fermentative-polymyxin B-bacitracin-lactose (OFPBL) agar, and Pseudomonas cepacia (PC) agar are recommended for isolation of B. cepacia from respiratory secretions of cystic fibrosis patients. We evaluated the sensitivity and selectivity of these four media using 20 B. mallei, 20 B. pseudomallei, 20 Burkholderia spp., and 15 diagnostically challenging organisms. Ashdown's agar was the most sensitive medium for the isolation of B. pseudomallei, but it was unable to support growth of B. mallei. Pseudomonas cepacia agar was highly sensitive and selective for both organisms. In non-endemic areas, we suggest the use of the commercially available PC agar for the isolation of B. mallei and B. pseudomallei.

The cheetah (Acinonyx jubatus) is a vulnerable species, with estimates of only 6700 animals left in the wild. Namibia, an anthrax-endemic country, is home to the world's largest and most viable free-ranging population (~3000 animals), which predominantly resides on unprotected private farmlands (Durant, 2015). For over 170 years, anthrax has been reported in African wildlife species with sporadic outbreaks across Namibia (Beyer et al., 2012). Anthrax is regularly reported from zebra (Equus quagga), hartebeest (Alcelaphus buselaphus), springbok (Antidorcas marsupialis) and kudu (Tragelaphus strepsiceros) (Turner et al., 2014; Wafula, Patrick & Charles, 2007); all cheetah prey species. Anthrax epidemics occur annually in Namibia's Etosha National Park (ENP), whereas the establishment of a government mandated livestock vaccination programme in 1973 reduced the occurrence of anthrax on the surrounding farmlands (Bellanet al., 2012; Schneider, 1994; Turner et al., 2013). However, sporadic epidemics still occur on private farmlands throughout Namibia (Shaanika, 2013).

Aims: To evaluate two selective media, polymyxin, lysozyme, ethylenediaminetetraacetic acid, thallium acetate (PLET) agar and R&F Anthracis chromogenic agar (ChrA), for the isolation and selection of Bacillus anthracis. Methods and Results: Sixteen genotypically diverse B. anthracis strains were sub-cultured onto PLET and ChrA to test the sensitivity (ability of B. anthracis to grow and produce expected colony morphology) of both media. Fourteen of the 16 B. anthracis strains produced the expected morphology on PLET (88% sensitive) while 13/16 produced the expected morphology on the ChrA medium (81% sensitive). Seventeen other Bacillus strains and 18 nonBacillus spp. strains were used to evaluate the media’s selectivity (ability to inhibit non-B. anthracis growth). PLET inhibited growth of 14/35 strains (40% selective), including six Bacillus strains, while ChrA inhibited 3/35 (9% selective). In addition, we did not observe any differences between the recovered CFU on PLET or ChrA when plating extractions of spiked soil. Conclusions: Polymyxin, lysozyme, ethylenediaminetetraacetic acid, thallium acetate agar was more selective and sensitive than ChrA. Significance and Impact of the Study: Although both media are more expensive than sheep blood agar, for samples with high numbers of bacteria, they can be used to isolate B. anthracis with proper training and experience and with the knowledge that there are limitations to each media.