Anticipating Knowledge to Inform Species Management: Predicting Spatially Explicit Habitat Suitability of a Colonial Vulture Spreading Its Range

Background

The knowledge of both potential distribution and habitat suitability is fundamental in spreading species to inform in advance management and conservation planning. After a severe decline in the past decades, the griffon vulture (Gyps fulvus) is now spreading its breeding range towards the northwest in Spain and Europe. Because of its key ecological function, anticipated spatial knowledge is required to inform appropriately both vulture and ecosystem management.

Methodology/Findings

Here we used maximum entropy (Maxent) models to determine the habitat suitability of potential and current breeding distribution of the griffon vulture using presence-only data (N = 124 colonies) in north-western Spain. The most relevant ecological factors shaping this habitat suitability were also identified. The resulting model had a high predictive performance and was able to predict species'' historical distribution. 7.5% (∼1,850 km²) of the study area resulted to be suitable breeding habitat, most of which (∼70%) is already occupied by the species. Cliff availability and livestock density, especially of sheep and goats, around 10 km of the colonies were the fundamental factors determining breeding habitat suitability for this species.

Conclusions/Significance

Griffon vultures could still spread 50–60 km towards the west, increasing their breeding range in 1,782 km². According to our results, 7.22% of the area suitable for griffon vulture will be affected by wind farms, so our results could help to better plan wind farm locations. The approach here developed could be useful to inform management of reintroductions and recovery programmes currently being implemented for both the griffon vulture and other threatened vulture species.     

Anthrax Toxin Uptake by Primary Immune Cells as Determined with a Lethal Factor-β-Lactamase Fusion Protein

Background

To initiate infection, Bacillus anthracis needs to overcome the host innate immune system. Anthrax toxin, a major virulence factor of B. anthracis, impairs both the innate and adaptive immune systems and is important in the establishment of anthrax infections.

Methodology/Principal Findings

To measure the ability of anthrax toxin to target immune cells, studies were performed using a fusion of the anthrax toxin lethal factor (LF) N-terminal domain (LFn, aa 1–254) with β-lactamase (LFnBLA). This protein reports on the ability of the anthrax toxin protective antigen (PA) to mediate LF delivery into cells. Primary immune cells prepared from mouse spleens were used in conjunction with flow cytometry to assess cleavage and resulting FRET disruption of a fluorescent β-lactamase substrate, CCF2/AM. In spleen cell suspensions, the macrophages, dendritic cells, and B cells showed about 75% FRET disruption of CCF2/AM due to cleavage by the PA–delivered LFnBLA. LFnBLA delivery into CD4+ and CD8+ T cells was lower, with 40% FRET disruption. When the analyses were done on purified samples of individual cell types, similar results were obtained, with T cells again having lower LFnBLA delivery than macrophages, dendritic cells, and B cells. Relative expression levels of the toxin receptors CMG2 and TEM8 on these cells were determined by real-time PCR. Expression of CMG2 was about 1.5-fold higher in CD8+ cells than in CD4+ and B cells, and 2.5-fold higher than in macrophages.

Conclusions/Significance

Anthrax toxin entry and activity differs among immune cells. Macrophages, dendritic cells, and B cells displayed higher LFnBLA activity than CD4+ and CD8+ T cells in both spleen cell suspension and the purified samples of individual cell types. Expression of anthrax toxin receptor CMG2 is higher in CD4+ and CD8+ T cells, which is not correlated to the intracellular LFnBLA activity.     

Inactivation of Bacillus anthracis Spores by Liquid Biocides in the Presence of Food Residue

Biocide inactivation of Bacillus anthracis spores in the presence of food residues after a 10-min treatment time was investigated. Spores of nonvirulent Bacillus anthracis strains 7702, ANR-1, and 9131 were mixed with water, flour paste, whole milk, or egg yolk emulsion and dried onto stainless-steel carriers. The carriers were exposed to various concentrations of peroxyacetic acid, sodium hypochlorite (NaOCl), or hydrogen peroxide (H₂O₂) for 10 min at 10, 20, or 30°C, after which time the survivors were quantified. The relationship between peroxyacetic acid concentration, H₂O₂ concentration, and spore inactivation followed a sigmoid curve that was accurately described using a four-parameter logistic model. At 20°C, the minimum concentrations of peroxyacetic acid, H₂O₂, and NaOCl (as total available chlorine) predicted to inactivate 6 log₁₀ CFU of B. anthracis spores with no food residue present were 1.05, 23.0, and 0.78%, respectively. At 10°C, sodium hypochlorite at 5% total available chlorine did not inactivate more than 4 log₁₀ CFU. The presence of the food residues had only a minimal effect on peroxyacetic acid and H₂O₂ sporicidal efficacy, but the efficacy of sodium hypochlorite was markedly inhibited by whole-milk and egg yolk residues. Sodium hypochlorite at 5% total available chlorine provided no greater than a 2-log₁₀ CFU reduction when spores were in the presence of egg yolk residue. This research provides new information regarding the usefulness of peroxygen biocides for B. anthracis spore inactivation when food residue is present. This work also provides guidance for adjusting decontamination procedures for food-soiled and cold surfaces.     

The Bacillus anthracis Protein MprF Is Required for Synthesis of Lysylphosphatidylglycerols and for Resistance to Cationic Antimicrobial Peptides

During inhalational anthrax, Bacillus anthracis survives and replicates in alveolar macrophages, followed by rapid invasion into the host's bloodstream, where it multiplies to cause heavy bacteremia. B. anthracis must therefore defend itself from host immune functions encountered during both the intracellular and the extracellular stages of anthrax infection. In both of these niches, cationic antimicrobial peptides are an essential component of the host's innate immune response that targets B. anthracis. However, the genetic determinants of B. anthracis contributing to resistance to these peptides are largely unknown. Here we generated Tn917 transposon mutants in the ΔANR strain (pXO1⁻ pXO2⁻) of B. anthracis and screened them for altered protamine susceptibility. A protamine-sensitive mutant identified carried the transposon inserted in the BA1486 gene encoding a putative membrane protein homologous to MprF known in several gram-positive pathogens. A mutant strain with the BAS1375 gene (the orthologue of BA1486) deleted in the Sterne 34F2 strain (pXO1⁺ pXO2⁻) of B. anthracis exhibited hypersusceptibility not only to protamine but also to α-helical cathelicidin LL-37 and β-sheet defensin human neutrophil peptide 1 compared to the wild-type Sterne strain. Analysis of membrane lipids using isotopic labeling demonstrated that the BAS1375 deletion mutant is unable to synthesize lysinylated phosphatidylglycerols, and this defect is rescued by genetic complementation. Further, we determined the structures of these lysylphosphatidylglycerols by using various mass spectrometric analyses. These results demonstrate that in B. anthracis a functional MprF is required for the biosynthesis of lysylphosphatidylglycerols, which is critical for resistance to cationic antimicrobial peptides.     

Complete Genome Sequence of Halalkalicoccus jeotgali B3T,an Extremely Halophilic Archaeon

Halalkalicoccus jeotgali B3^T, isolated from salt-fermented seafood from South Korea, is an extremely halophilic archaeon belonging to the family Halobacteriaceae. Here, we present the complete genome sequence of the type strain H. jeotgali B3^T (3,698,650 bp, with a G+C content of 62.5%), which consists of one chromosome and six plasmids. This is the first complete genome sequence of the Halalkalicoccus species.Extremely halophilic archaea (haloarchaea) are adapted to hypersaline environments and grow optimally in NaCl solutions of 2.6 M or higher (12). These haloarchaea are classified within the family Halobacteriaceae in the order Halobacteriales; currently, this family comprises 28 genera (3), and only 11 complete genome sequences in Halobacteriaceae have been reported. In a study of archaeal diversity in salt-fermented small shrimp or shellfish from South Korea, our laboratory isolated and characterized 5 novel, extremely halophilic archaeal strains of Halobacteriaceae. These strains included Natronococcus jeotgali (9), Halalkalicoccus jeotgali (11), Halorubrum cibi (7), Haloterrigena jeotgali (10) and Haladaptatus cibarius (8). We have now sequenced the genome of Halalkalicoccus jeotgali B3^T; genome sequencing had not been completed or initiated for any strain in this genus when our sequencing project was begun. The genus Halalkalicoccus currently contains only two species, Halalkalicoccus tibetensis (13) and H. jeotgali, and these species exhibit 98.6% gene sequence similarity in their 16S rRNA. The genome of H. jeotgali B3^T is the first of this genus to be sequenced.The complete genome sequence of H. jeotgali B3^T was determined by a whole-genome shotgun strategy using Roche 454 GS (FLX Titanium) pyrosequencing (898,168 reads totaling ∼348 Mb; ∼94-fold coverage of the genome) and a fosmid library (514 reads totaling ∼680 kb) at the Genome Resource Center, KRIBB (Korea Research Institute of Bioscience and Biotechnology). Genome sequences from pyrosequencing were processed by Roche''s software according to the manufacturer''s instructions, and sequences from the fosmid library were processed by PESTAS (6). A total of 898,196 reads were assembled using Newbler Assembler 2.3 (454 Life Science), which generated 54 large contigs (>100 bp in size) with bases having quality scores of 40 and above. The gaps between contigs were closed by primer walking and sequencing of PCR products across the gaps. The annotation was done by merging results obtained from the RAST (Rapid Annotation using Subsystem Technology) pipeline (1), Glimmer 3.02 (2), tRNAscan-SE 1.21 (5), and RNAmmer 1.2 (4).The H. jeotgali B3^T genome is 3,698,650 bases long with a 62.5% G+C content. The chromosome consists of a single circular chromosome (2,809,118 bp, with a G+C content of 65.0%) and six plasmids (406,285 bp, 55.3%; 363,534 bp, 54.2%; 44,576 bp, 58.9%; 44,459 bp, 54.9%; 23,727 bp, 47.6%; 6,951 bp, 60.6%). The genome contains 3,860 predicted coding sequences and 52 RNA genes (determined using RAST). The chromosome is predicted to contain 3,101 coding sequences with a coding intensity of 90.0%, including 47 tRNA genes, 1 5S rRNA gene, 1 16S rRNA gene, and 1 23S rRNA gene. The largest plasmid contains 466 coding sequences with a coding intensity of 81.2% and 2 tRNA genes, while the other five plasmids contain 425, 44, 48, 29, and 5 coding sequences with coding intensities of 80.2%, 84.2%, 83.0%, 69.6%, and 22.8%, respectively (determined using Glimmer3). More detailed analysis of this genome and comparative analysis with other haloarchaea will provide further insight into the genomic differences and metabolism of the extremely halophilic archaea.     

Ecological modelling for the conservation of Gluta travancorica Bedd. - An endemic tree species of southern Western Ghats,India

Endemic species are highly adapted to grow exclusively in a specific geographical area. The goal of the current study is to determine the probable habitat distribution range of the narrowly endemic species Gluta travancorica. An ecological niche modelling is carried out, using four different models viz., BioClim, MaxEnt, Random Forest and Deep Neural Networks (DNN). A total of 506 G. travancorica cluster locations were surveyed and used for this study with thirty different ecogeographic, edaphic and bioclimatic environmental parameters. After a preliminary investigation using multi-collinearity analysis, soil parameter variables like pH, cation exchange capacity (CEC), silt and clay content are used for final modelling. Factor analysis of ecological niche revealed the soil parameters like pH, CEC, silt and clay content as the key predictors. The result is validated using true skill statistics, sensitivity, specificity, kappa statistic and AUC-ROC. Results of the present study show that DNN have exceptional prediction performance, demonstrated by its AUC score of 0.959. DNN model projected 32.37% (938.18 km²) of the study region to have a ‘highly suitable habitat’, whereas 67.63% (1960.82 km²) was classified as having ‘low habitat suitability’. Besides, back-to-field assessments have also proven DNN's potential in predicting the habitat suitability of G. travancorica. The study results will facilitate the prioritization of conservation and seedling restoration strategies. The forest range explored in this work is a component of one of the most important global biodiversity hotspots, and it has significant implications for regional biodiversity conservation.     

Novel Data on the Ecology of Cochranella mache (Anura: Centrolenidae) and the Importance of Protected Areas for This Critically Endangered Glassfrog in the Neotropics

We studied a population of the endangered glassfrog, Cochranella mache, at Bilsa Biological Station, northwestern Ecuador, from 2008 and 2009. We present information on annual abundance patterns, behavioral ecology, habitat use and a species distribution model performed with MaxEnt. We evaluate the importance of the National System of Protected Areas (SNAP) in Colombia and Ecuador, under scenarios of climate change and habitat loss. We predicted a restricted environmental suitability area from 48,509 Km² to 65,147 Km² along western Ecuador and adjacent Colombia; ∼8% of the potential distribution occurs within SNAP. We examined four aspects of C. mache ecology: (1) ecological data suggests a strong correlation between relative abundance and rainfall, with a high probability to observe frogs through rainy months (February–May); (2) habitat use and the species distribution model suggest that this canopy dweller is restricted to small streams and rivulets in primary and old secondary forest in evergreen lowland and piedmont forest of western Ecuador, with predictions of suitability areas in adjacent southern Colombia; (3) the SNAP of Colombia and Ecuador harbor a minimum portion of the predicted model of distribution (<10%); and (4) synergetic effects of habitat loss and climate change reduces in about 95% the suitability areas for this endangered frog along its distributional range in Protected Areas. The resulting model allows the recognition of areas to undertake conservation efforts and plan future field surveys, as well as forecasting regions with high probability of C. mache occurrence in western Ecuador and southern Colombia. Further research is required to assess population tendencies, habitat fragmentation and target survey zones to accelerate the discovery of unknown populations in unexplored areas with high probability of suitability. We recommend that Cochranella mache must be re-categorized as “Critically Endangered” species in national and global status, according with criteria and sub-criteria A4, B1ab(i,ii,iii,iv),E.     

Ecological Niche Modeling of Bacillus anthracis on Three Continents: Evidence for Genetic-Ecological Divergence?

We modeled the ecological niche of a globally successful Bacillus anthracis sublineage in the United States, Italy and Kazakhstan to better understand the geographic distribution of anthrax and potential associations between regional populations and ecology. Country-specific ecological-niche models were developed and reciprocally transferred to the other countries to determine if pathogen presence could be accurately predicted on novel landscapes. Native models accurately predicted endemic areas within each country, but transferred models failed to predict known occurrences in the outside countries. While the effects of variable selection and limitations of the genetic data should be considered, results suggest differing ecological associations for the B. anthracis populations within each country and may reflect niche specialization within the sublineage. Our findings provide guidance for developing accurate ecological niche models for this pathogen; models should be developed regionally, on the native landscape, and with consideration to population genetics. Further genomic analysis will improve our understanding of the genetic-ecological dynamics of B. anthracis across these countries and may lead to more refined predictive models for surveillance and proactive vaccination programs. Further studies should evaluate the impact of variable selection of native and transferred models.     

Recovery Efficiency and Limit of Detection of Aerosolized Bacillus anthracis Sterne from Environmental Surface Samples

After the 2001 anthrax incidents, surface sampling techniques for biological agents were found to be inadequately validated, especially at low surface loadings. We aerosolized Bacillus anthracis Sterne spores within a chamber to achieve very low surface loading (ca. 3, 30, and 200 CFU per 100 cm²). Steel and carpet coupons seeded in the chamber were sampled with swab (103 cm²) or wipe or vacuum (929 cm²) surface sampling methods and analyzed at three laboratories. Agar settle plates (60 cm²) were the reference for determining recovery efficiency (RE). The minimum estimated surface concentrations to achieve a 95% response rate based on probit regression were 190, 15, and 44 CFU/100 cm² for sampling steel surfaces and 40, 9.2, and 28 CFU/100 cm² for sampling carpet surfaces with swab, wipe, and vacuum methods, respectively; however, these results should be cautiously interpreted because of high observed variability. Mean REs at the highest surface loading were 5.0%, 18%, and 3.7% on steel and 12%, 23%, and 4.7% on carpet for the swab, wipe, and vacuum methods, respectively. Precision (coefficient of variation) was poor at the lower surface concentrations but improved with increasing surface concentration. The best precision was obtained with wipe samples on carpet, achieving 38% at the highest surface concentration. The wipe sampling method detected B. anthracis at lower estimated surface concentrations and had higher RE and better precision than the other methods. These results may guide investigators to more meaningfully conduct environmental sampling, quantify contamination levels, and conduct risk assessment for humans.Anthrax, the spectrum of diseases caused by infection with Bacillus anthracis, is not considered a communicable disease but is generally acquired via environmental exposures. Many anthrax cases through history have been the result of agricultural or industrial exposure to B. anthracis spores (33). The disease most often presents itself as a cutaneous infection; however, there are both gastrointestinal and inhalational forms of the disease. Inhalational anthrax is typically rapidly fatal, even with treatment. In general, inhalation exposures require specific conditions, such as poor ventilation and activities that disturb dust containing B. anthracis spores (13).Because diagnosing anthrax in its early stages in human and animal hosts is difficult and B. anthracis spores are extremely stable in the environment, this microorganism has been investigated, developed, and deployed as a biological weapon throughout the 20th century. Use of this microorganism has seen varied success during World War I (9) and subsequently. It is generally accepted that there was an accidental release of B. anthracis spores from a weapons manufacturing or development facility in 1979 in Sverdlovsk, USSR (now Yekaterinaburg, Russia) (10, 26). In 1993, an attempt by a civilian group, Aum Shinrikyo, to use this microorganism to attack a civilian population in a Tokyo suburb did not result in any casualties (22, 28).In 2001, envelopes containing a powder formulation of B. anthracis were mailed in the United States to several individuals. These letters were the presumed cause of 22 cases of clinical anthrax, 11 inhalational and 11 cutaneous, with 5 fatalities, all of whom suffered from inhalational disease (34). According to congressional testimony, the powdered spore suspension was “easily dispersed into the air” (29). Of the 11 individuals with inhalational disease, 2 had no history of handling mail or having any other direct contact with these threat letters (11, 21). Of the remaining nine individuals, eight were thought to have been exposed through handling or processing mail (20) but may never have picked up or directly handled the actual threat letters. Thus, some individuals who contracted inhalational disease may have been exposed to aerosols that were generated from residual spore material deposited on contaminated surfaces. This conclusion was borne out by a study conducted on the scene of one contamination incident, which demonstrated that spores could be reaerosolized from surfaces during simulated office activities—e.g., paper handling, foot traffic, moving containers—after a period of no entry and no ventilation for several days (38). McCleery et al. (25) found that reaerosolization of spores is possible in postal facilities.In the mail-related instance of 2001, aerosol exposures occurred. Since spore-contaminated surfaces can become sources for aerosol generation, nonporous surfaces (walls, desks, lockers, etc.) were decontaminated to reduce risk while porous surfaces (draperies and sofas) were removed. To determine the efficacy of decontamination, contaminated buildings were first sampled for the presence of B. anthracis spores followed by treatment by a variety of techniques. Postdecontamination sampling was used to determine efficacy (37) and to assess the safety for reoccupancy.The Government Accountability Office (GAO) reported that additional methodological validation of sampling collection and analytical methods should be conducted to enhance the interpretation of negative sampling results because initial samples from two postal facilities were negative, but later samples were positive (17). The GAO (17) report defined validation as “… a formal and independently administered empirical process. For validation, the overall performance characteristics of a given method must be certified as meeting the specified requirements for intended use and as conforming with applicable standards.” Currently, there is no preexisting standard for a presumable safe level of surface contamination with B. anthracis spores that may be assessed through sampling and analysis.Development of independent standards for assessing the requirements for surface sampling methods requires an understanding of the rate at which spores leave surfaces to become entrained in aerosols, the potential for aerosol exposure by humans, and the infectivity of inhaled spores. Inhalation infectivity has been researched, but estimates of a lethal dose vary (14, 15). Bartrand et al. (5) conducted a risk analysis on the mortality of guinea pigs and rhesus monkeys exposed to B. anthracis spores and found a 50% lethal dose (LD₅₀; i.e., the dose at which 50% of subjects die) of about 100,000 spores inhaled for 1-μm particles. Limitations of relating exposure to inhalation infectivity include quantification of the ability of spores to move from stasis on a surface to entrainment as an aerosol, quantification of exposures to the resultant aerosol, uptake by humans, room size and ventilation characteristics, and exposure time. Despite these limitations, it is necessary to standardize the performance of surface sampling methods.Brown et al. evaluated wipe (6), swab (7), and vacuum (8) spore collection methods with B. atrophaeus. These studies have added significant information to the understanding of recovery efficiencies for these three sampling methods; however, sampling performance was not evaluated at very low spore surface loading concentrations. Sampling performance measures at very low surface loading of B. anthracis are needed to aid in the decision making for decontamination and other interventions (31, 38).The goal of this study was to evaluate the current CDC environmental surface sampling methods for B. anthracis (12) as slightly modified based on subsequent CDC research (19, 30). We estimated B. anthracis Sterne sampling limit of detection (LOD), recovery efficiency (RE), and measurement precision for three sampling methods (swab, wipe, and vacuum) and two surfaces (steel and carpet) by allowing spores to settle from an aerosol in a controlled environment. In addition, we compared sample analyses performed at three laboratories to determine the level of interlaboratory variability.     

The Genome of a Bacillus Isolate Causing Anthrax in Chimpanzees Combines Chromosomal Properties of B. cereus with B. anthracis Virulence Plasmids

Anthrax is a fatal disease caused by strains of Bacillus anthracis. Members of this monophyletic species are non motile and are all characterized by the presence of four prophages and a nonsense mutation in the plcR regulator gene. Here we report the complete genome sequence of a Bacillus strain isolated from a chimpanzee that had died with clinical symptoms of anthrax. Unlike classic B. anthracis, this strain was motile and lacked the four prohages and the nonsense mutation. Four replicons were identified, a chromosome and three plasmids. Comparative genome analysis revealed that the chromosome resembles those of non-B. anthracis members of the Bacillus cereus group, whereas two plasmids were identical to the anthrax virulence plasmids pXO1 and pXO2. The function of the newly discovered third plasmid with a length of 14 kbp is unknown. A detailed comparison of genomic loci encoding key features confirmed a higher similarity to B. thuringiensis serovar konkukian strain 97-27 and B. cereus E33L than to B. anthracis strains. For the first time we describe the sequence of an anthrax causing bacterium possessing both anthrax plasmids that apparently does not belong to the monophyletic group of all so far known B. anthracis strains and that differs in important diagnostic features. The data suggest that this bacterium has evolved from a B. cereus strain independently from the classic B. anthracis strains and established a B. anthracis lifestyle. Therefore we suggest to designate this isolate as “B. cereus variety (var.) anthracis”.     

Predicting the Distribution Pattern of Small Carnivores in Response to Environmental Factors in the Western Ghats

Due to their secretive habits, predicting the pattern of spatial distribution of small carnivores has been typically challenging, yet for conservation management it is essential to understand the association between this group of animals and environmental factors. We applied maximum entropy modeling (MaxEnt) to build distribution models and identify environmental predictors including bioclimatic variables, forest and land cover type, topography, vegetation index and anthropogenic variables for six small carnivore species in Mudumalai Tiger Reserve. Species occurrence records were collated from camera-traps and vehicle transects during the years 2010 and 2011. We used the average training gain from forty model runs for each species to select the best set of predictors. The area under the curve (AUC) of the receiver operating characteristic plot (ROC) ranged from 0.81 to 0.93 for the training data and 0.72 to 0.87 for the test data. In habitat models for F. chaus, P. hermaphroditus, and H. smithii “distance to village” and precipitation of the warmest quarter emerged as some of the most important variables. “Distance to village” and aspect were important for V. indica while “distance to village” and precipitation of the coldest quarter were significant for H. vitticollis. “Distance to village”, precipitation of the warmest quarter and land cover were influential variables in the distribution of H. edwardsii. The map of predicted probabilities of occurrence showed potentially suitable habitats accounting for 46 km² of the reserve for F. chaus, 62 km² for V. indica, 30 km² for P. hermaphroditus, 63 km² for H. vitticollis, 45 km² for H. smithii and 28 km² for H. edwardsii. Habitat heterogeneity driven by the east-west climatic gradient was correlated with the spatial distribution of small carnivores. This study exemplifies the usefulness of modeling small carnivore distribution to prioritize and direct conservation planning for habitat specialists in southern India.     

Cutaneous anthrax associated with handling carcasses of animals that died suddenly of unknown cause: Arua District,Uganda, January 2015–August 2017

BackgroundAnthrax is a zoonotic disease that can be transmitted to humans from infected animals. During May–June 2017, three persons with probable cutaneous anthrax were reported in Arua District, Uganda; one died. All had recently handled carcasses of livestock that died suddenly and a skin lesion from a deceased person tested positive by PCR for Bacillus anthracis. During July, a bull in the same community died suddenly and the blood sample tested positive by PCR for Bacillus anthracis. The aim of this investigation was to establish the scope of the problem, identify exposures associated with illness, and recommend evidence-based control measures.MethodsA probable case was defined as acute onset of a papulo-vesicular skin lesion subsequently forming an eschar in a resident of Arua District during January 2015–August 2017. A confirmed case was a probable case with a skin sample testing positive by polymerase chain reaction (PCR) for B. anthracis. Cases were identified by medical record review and active community search. In a case-control study, exposures between case-patients and frequency- and village-matched asymptomatic controls were compared. Key animal health staff were interviewed to learn about livestock deaths.ResultsThere were 68 case-patients (67 probable, 1 confirmed), and 2 deaths identified. Cases occurred throughout the three-year period, peaking during dry seasons. All cases occurred following sudden livestock deaths in the villages. Case-patients came from two neighboring sub-counties: Rigbo (attack rate (AR) = 21.9/10,000 population) and Rhino Camp (AR = 1.9/10,000). Males (AR = 24.9/10,000) were more affected than females (AR = 0.7/10,000). Persons aged 30–39 years (AR = 40.1/10,000 population) were most affected. Among all cases and 136 controls, skinning (OR_M-H = 5.0, 95%CI: 2.3–11), butchering (OR_M-H = 22, 95%CI: 5.5–89), and carrying the carcass of livestock that died suddenly (OR_M-H = 6.9, 95%CI: 3.0–16) were associated with illness.ConclusionsExposure to carcasses of animals that died suddenly was a likely risk factor for cutaneous anthrax in Arua District during 2015–2017. The recommendations are investigation of anthrax burden in livestock, prevention of animal infections through vaccinations, safe disposal of the carcasses, public education on risk factors for infection and prompt treatment of illness following exposure to animals that died suddenly.     

Potential impacts of climate change on geographical distribution of three primary vectors of African Trypanosomiasis in Tanzania’s Maasai Steppe: G. m. morsitans,G. pallidipes and G. swynnertoni

In the Maasai Steppe, public health and economy are threatened by African Trypanosomiasis, a debilitating and fatal disease to livestock (African Animal Trypanosomiasis -AAT) and humans (Human African Trypanosomiasis—HAT), if not treated. The tsetse fly is the primary vector for both HAT and AAT and climate is an important predictor of their occurrence and the parasites they carry. While understanding tsetse fly distribution is essential for informing vector and disease control strategies, existing distribution maps are old and were based on coarse spatial resolution data, consequently, inaccurately representing vector and disease dynamics necessary to design and implement fit-for-purpose mitigation strategies. Also, the assertion that climate change is altering tsetse fly distribution in Tanzania lacks empirical evidence. Despite tsetse flies posing public health risks and economic hardship, no study has modelled their distributions at a scale needed for local planning. This study used MaxEnt species distribution modelling (SDM) and ecological niche modeling tools to predict potential distribution of three tsetse fly species in Tanzania’s Maasai Steppe from current climate information, and project their distributions to midcentury climatic conditions under representative concentration pathways (RCP) 4.5 scenarios. Current climate results predicted that G. m. morsitans, G. pallidipes and G swynnertoni cover 19,225 km², 7,113 km² and 32,335 km² and future prediction indicated that by the year 2050, the habitable area may decrease by up to 23.13%, 12.9% and 22.8% of current habitable area, respectively. This information can serve as a useful predictor of potential HAT and AAT hotspots and inform surveillance strategies. Distribution maps generated by this study can be useful in guiding tsetse fly control managers, and health, livestock and wildlife officers when setting surveys and surveillance programs. The maps can also inform protected area managers of potential encroachment into the protected areas (PAs) due to shrinkage of tsetse fly habitats outside PAs.     

Decontamination Options for Bacillus anthracis-Contaminated Drinking Water Determined from Spore Surrogate Studies

Five parameters were evaluated with surrogates of Bacillus anthracis spores to determine effective decontamination alternatives for use in a contaminated drinking water supply. The parameters were as follows: (i) type of Bacillus spore surrogate (B. thuringiensis or B. atrophaeus), (ii) spore concentration in suspension (10² and 10⁶ spores/ml), (iii) chemical characteristics of the decontaminant (sodium dichloro-S-triazinetrione dihydrate [Dichlor], hydrogen peroxide, potassium peroxymonosulfate [Oxone], sodium hypochlorite, and VirkonS), (iv) decontaminant concentration (0.01% to 5%), and (v) exposure time to decontaminant (10 min to 1 h). Results from 138 suspension tests with appropriate controls are reported. Hydrogen peroxide at a concentration of 5% and Dichlor or sodium hypochlorite at a concentration of 2% were highly effective at spore inactivation regardless of spore type tested, spore exposure time, or spore concentration evaluated. This is the first reported study of Dichlor as an effective decontaminant for B. anthracis spore surrogates. Dichlor''s desirable characteristics of high oxidation potential, high level of free chlorine, and a more neutral pH than that of other oxidizers evaluated appear to make it an excellent alternative. All three oxidizers were effective against B. atrophaeus spores in meeting the EPA biocide standard of greater than a 6-log kill after a 10-min exposure time and at lower concentrations than typically reported for biocide use. Solutions of 5% VirkonS and Oxone were less effective as decontaminants than other options evaluated in this study and did not meet the EPA''s efficacy standard for a biocide, although they were found to be as effective for concentrations of 10² spores/ml. Differences in methods and procedures reported by other investigators make quantitative comparisons among studies difficult.Developing a decontamination approach that can be safely and effectively applied to civilian water resources and facilities following a terrorist or catastrophic release of Bacillus anthracis spores poses many challenges. For example, if a municipal drinking water system were contaminated directly or indirectly during or after such an incident, it would be essential to assess the potential health risks posed by water consumption or other water uses (e.g., recreational and bathing) and then to apply one or more proven technologies, if deemed necessary, to decontaminate the water supply quickly and cost-effectively. Treatment of drinking water implies the use of a decontamination approach that would not pose adverse health risks to humans or result in unacceptable damage to the environment. A major obstacle in killing spores of Bacillus spp. on or in virtually any matrix is their high level of resistance to treatments such as harsh chemicals, heat, desiccation, and UV light (14, 20). Because of the substantial and widely reported resistance of Bacillus spores to inactivation, a decontaminant proven to be efficacious in killing such spores for site-specific applications is likely to be effective against all other biological warfare agents as well.Whereas nearly all biological warfare agents are intended for aerosol application, many have strong potential as waterborne threats and could inflict heavy casualties when ingested (2). B. anthracis in particular has been identified as a “probable” (12) or an actual (24) water threat. Even though the principal risk associated with the consumption of water containing B. anthracis spores would likely arise from an ingestion hazard, water used for bathing, showering, or recreational purposes might also pose cutaneous as well as aerosol exposure hazards. There is controversy regarding the long-term viability of B. anthracis in water, and experimental evidence is limited. However, according to a review of nonkinetic studies on survival of virulent strains in the environment (21), B. anthracis spores can survive from 2 to 18 years in pond water and 20 months in seawater or distilled water. B. anthracis spores have been reported by others to be stable in water for 2 years (24).Various decontamination approaches have been evaluated for efficacy against biological warfare agents, including Bacillus spores, on hard, nonporous surfaces. Recommendations by the U.S. Environmental Protection Agency (EPA) include the use of sodium hypochlorite (1:9 dilution of bleach to 5,250 to 6,000 ppm, corrected to pH 7, with a 60-min contact time at 20°C [6, 17]), and liquid chlorine dioxide with a 30-min wet contact time at 20°C (7). Liquid hydrogen peroxide/peroxyacetic acid (known as peroxy compounds and marketed as ready-to-use solutions), generally with a 15- to 20-min wet contact time and concentration as specified by the manufacturer, has also been recommended (13). Other products, such as hydrogen peroxide solution (3 to 25%) and potassium peroxymonosulfate, have been evaluated for efficacy against Bacillus spores as well (27). Although disinfectants at various concentrations have been tested previously against the spores of B. anthracis and their surrogates, wide variations in test protocols make meaningful comparisons among studies virtually impossible (9, 11, 17).In contrast to surface cleanup of spores, fewer assessments of efficacy utilizing suspension tests with the aforementioned chemicals or other methods have been reported for the decontamination of Bacillus species spores in water, and much of the published work has assessed only relatively high concentrations of spores in water. For example, one previous investigation commenced evaluations with 0.2-ml suspensions of approximately 10⁹ spores/ml of various Bacillus spp. to which 20 ml of aqueous ozone or 20 ml of hydrogen peroxide solution was added to assess sporicidal action (10), and others have reported mechanisms of deactivating B. subtilis spores prepared in concentrations of up to approximately 10⁸ spores/ml (26) and approximately 10⁹ spores/ml (17). Inactivation by chlorination of various Bacillus spp. with initial concentrations of approximately 1 × 10⁴ CFU/ml has also been tested (16). However, relatively low spore concentrations would be expected to result from dilutions following contamination of a large public water system. Therefore, it is reasonable to evaluate the effectiveness of decontaminants or other methods against even lower spore concentrations in water than have been previously assessed. In addition to assessing the parameter of Bacillus spore concentration in water, it is essential to identify the most effective commercially available chemical that will kill all the spores or minimize population growth, while considering the effects of the chemical on the environment and in humans.Several objectives served to focus our investigation. First, five potential candidate decontaminants were selected because of their relative safety and ultimate degradation in the environment without substantive adverse consequences. The five chemicals were also chosen as a way of comparing the effectiveness of available free chlorine content, pH, and oxidation potential on spore inactivation. From an evaluation of those chemical parameters, we sought to determine the most effective option for inactivating Bacillus spore surrogates suspended in water. As a second objective, we attempted to identify the lowest concentration of the selected chemicals necessary to achieve the EPA''s biocide standard of a >6-log kill. As a third objective, we wanted to assess the effect of reduced spore concentration on chemical biocide efficacy. As an important step in ascertaining an efficient, safe, and cost-effective water treatment method that could potentially provide safe water to the general population in the event of B. anthracis contamination—and limit the potential risk of contracting gastrointestinal or cutaneous anthrax as well—the following parameters were evaluated: chemical decontaminant type, chemical decontaminant concentration (0.01% to 5%), contact time of spores with chemical decontaminant (10 min to 1 h), spore type (Bacillus atrophaeus or Bacillus thuringiensis), and low versus relatively high spore concentrations (approximately 10² and 10⁶ spores/ml, respectively).Use of B. atrophaeus and B. thuringiensis spores as surrogates for B. anthracis is widely reported in the literature. For example, Szabo et al. (23) used B. atrophaeus subsp. globigii spores as a surrogate for B. anthracis to investigate the persistence and decontamination of those surrogates on corroded iron in a model drinking water system, and Rice et al. (16) used spores of B. thuringiensis as an “appropriate surrogate for spores of B. anthracis” for determining the sporicidal activity of chlorination as commonly used in drinking water treatment. Furthermore, the EPA (5) concluded that “B. globigii can serve as a conservative surrogate for B. anthracis during studies of inactivation by chlorination.”     

Projected impacts of climate change on snow leopard habitat in Qinghai Province,China

Assessing species’ vulnerability to climate change is a prerequisite for developing effective strategies to reduce emerging climate‐related threats. We used the maximum entropy algorithm (MaxEnt model) to assess potential changes in suitable snow leopard (Panthera uncia) habitat in Qinghai Province, China, under a mild climate change scenario. Our results showed that the area of suitable snow leopard habitat in Qinghai Province was 302,821 km² under current conditions and 228,997 km² under the 2050s climatic scenario, with a mean upward shift in elevation of 90 m. At present, nature reserves protect 38.78% of currently suitable habitat and will protect 42.56% of future suitable habitat. Current areas of climate refugia amounted to 212,341 km² and are mainly distributed in the Sanjiangyuan region, Qilian mountains, and surrounding areas. Our results provide valuable information for formulating strategies to meet future conservation challenges brought on by climate stress. We suggest that conservation efforts in Qinghai Province should focus on protecting areas of climate refugia and on maintaining or building corridors when planning for future species management.     

Habitat suitability assessment of Sichuan sika deer in Tiebu Nature Reserve during periods of green and dry grass

Habitat suitability assessment is an essential and dynamic research method for determining and evaluating the environmental pressures faced by wildlife. From March to November 2011, we investigated the quality of habitat available to Sichuan sika deer (Cervus nippon sichuanicus) in Tiebu Nature Reserve, Ruoergai County, Sichuan Province, China. A habitat evaluation model established by the fuzzy assignment quadrature method was used to assess habitat suitability for Sichuan sika deer within the reserve by using the GIS spatial analysis function. The results showed that the area of actual available habitat was 220.8 km² during the wet season and 213.2 km² during the dry season, accounting for 80.8% and 78.02% of the total nature reserve area, respectively. The area of suitable habitat for Sichuan sika deer was much lower, 128.01 km² during the wet season and 109.17 km² during the dry season, accounting for 46.84% and 39.95% of the total nature reserve area respectively. The difference between available and suitable habitat is likely due to potentially good habitat having been lost as a result of human disturbance. Lost habitat makes up 4.55% of the total area while grass is green and 5.52% while grass is dry. Human disturbance levels in the form of roads and residential areas were constant throughout the year, but grazing by domestic animals had a higher impact during the dry season. Habitat suitability during this time, already reduced by the withering of the grass, was thus further reduced by the grazing of livestock.     

Bacillus anthracis Diversity and Geographic Potential across Nigeria,Cameroon and Chad: Further Support of a Novel West African Lineage

Zoonoses, diseases affecting both humans and animals, can exert tremendous pressures on human and veterinary health systems, particularly in resource limited countries. Anthrax is one such zoonosis of concern and is a disease requiring greater public health attention in Nigeria. Here we describe the genetic diversity of Bacillus anthracis in Nigeria and compare it to Chad, Cameroon and a broader global dataset based on the multiple locus variable number tandem repeat (MLVA-25) genetic typing system. Nigerian B. anthracis isolates had identical MLVA genotypes and could only be resolved by measuring highly mutable single nucleotide repeats (SNRs). The Nigerian MLVA genotype was identical or highly genetically similar to those in the neighboring countries, confirming the strains belong to this unique West African lineage. Interestingly, sequence data from a Nigerian isolate shares the anthrose deficient genotypes previously described for strains in this region, which may be associated with vaccine evasion. Strains in this study were isolated over six decades, indicating a high level of temporal strain stability regionally. Ecological niche models were used to predict the geographic distribution of the pathogen for all three countries. We describe a west-east habitat corridor through northern Nigeria extending into Chad and Cameroon. Ecological niche models and genetic results show B. anthracis to be ecologically established in Nigeria. These findings expand our understanding of the global B. anthracis population structure and can guide regional anthrax surveillance and control planning.