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Search Results: 1 - 10 of 229839 matches for " Alex R. Hoffmaster "
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Molecular approaches to identify and differentiate Bacillus anthracis from phenotypically similar Bacillus species isolates
Chung K Marston, Jay E Gee, Tanja Popovic, Alex R Hoffmaster
BMC Microbiology , 2006, DOI: 10.1186/1471-2180-6-22
Abstract: Six isolates were LRN real-time PCR-positive but resistant to gamma phage; MLVA data supported the identification of these isolates as gamma phage-resistant B. anthracis. Seventeen isolates were LRN real-time PCR-negative but susceptible to gamma phage lysis; these isolates appear to be a group of unusual gamma phage-susceptible B. cereus isolates that are closely related to each other and to B. anthracis. All six B. anthracis MLVA chromosomal loci were amplified from one unusual gamma phage-susceptible, motile, B. cereus isolate (although the amplicons were atypical sizes), and when analyzed phylogenetically, clustered with B. anthracis by MLST.MLVA and MLST aided in the identification of these isolates when standard microbiological methods and PCR could not definitely identify or rule out B. anthracis. This study emphasized the need to perform multiple tests when attempting to identify B. anthracis since relying on a single assay remains problematic due to the diverse nature of bacteria.Bacillus anthracis and B. cereus are two closely related gram-positive species belonging to the B. cereus group (B. anthracis, B. cereus, B. thuringiensis, B. weihenstephanensis, B. pseudomycoides and B. mycoides). Despite their close genetic relatedness, in general B. anthracis has several phenotypic characteristics making it easily differentiated from B. cereus and other members of the B. cereus group. B. anthracis is non-hemolytic on sheep blood agar, susceptible to penicillin, lysed by the gamma phage, and non-motile. Conversely, B. cereus is hemolytic on sheep blood agar, resistant to penicillin, resistant to lysis by the gamma phage, and motile. In addition, B. anthracis harbors two virulence plasmids, pXO1 and pXO2, which B. cereus lacks, although a recent reports have showed that rare B. cereus isolates harbor genes and similar plasmids closely related to pXO1 and pXO2 [1-6]. However, standard microbiologic identification can be complicated when isolates of B. anthracis are
Genetic diversity of clinical isolates of Bacillus cereus using multilocus sequence typing
Alex R Hoffmaster, Ryan T Novak, Chung K Marston, Jay E Gee, Leta Helsel, James M Pruckler, Patricia P Wilkins
BMC Microbiology , 2008, DOI: 10.1186/1471-2180-8-191
Abstract: A retrospective analysis of 55 clinical B. cereus isolates submitted to the Centers for Disease Control and Prevention between 1954 and 2004 was conducted. Clinical isolates from severe infections (n = 27), gastrointestinal (GI) illness (n = 18), and associated isolates from food (n = 10) were selected for analysis using MLST. The 55 isolates were diverse and comprised 38 sequence types (ST) in two distinct clades. Of the 27 isolates associated with serious illness, 13 clustered in clade 1 while 14 were in clade 2. Isolates associated with GI illness were also found throughout clades 1 and 2, while no isolates in this study belonged to clade 3. All the isolates from this study belonging to the clade 1/cereus III lineage were associated with severe disease while isolates belonging to clade1/cereus II contained isolates primarily associated with severe disease and emetic illness. Only three STs were observed more than once for epidemiologically distinct isolates.STs of clinical B. cereus isolates were phylogenetically diverse and distributed among two of three previously described clades. Greater numbers of strains will need to be analyzed to confirm if specific lineages or clonal complexes are more likely to contain clinical isolates or be associated with specific illness, similar to B. anthracis and emetic B. cereus isolates.The phylogenetically related species of the Bacillus cereus group include: B. cereus, B. anthracis, B. thuringiensis, B. mycoides and two recently described species, B. pseudomycoides and B. weihenstephanensis [1-4]. B. cereus, B. thuringiensis, and B. anthracis have been the most characterized due to their pathogenic nature. B. anthracis is the etiologic agent of anthrax, B. thuringiensis is an insect pathogen, and B. cereus can be associated with a variety of human infections. B. cereus is most frequently associated with food poisoning, characterized by strains producing emetic or diarrheal toxins. It is also an opportunistic pathogen resultin
Molecular Epidemiology of Anthrax Cases Associated with Recreational Use of Animal Hides and Yarn in the United States
Chung K. Marston,Christina A. Allen,Jodi Beaudry,Erin P. Price,Spenser R. Wolken,Talima Pearson,Paul Keim,Alex R. Hoffmaster
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0028274
Abstract: To determine potential links between the clinical isolate to animal products and their geographic origin, we genotyped (MLVA-8, MVLA-15, and canSNP analysis) 80 environmental and 12 clinical isolates and 2 clinical specimens from five cases of anthrax (California in 1976 [n = 1], New York in 2006 [n = 1], Connecticut in 2007 [n = 2], and New Hampshire in 2009[n = 1]) resulting from recreational handling of animal products. For the California case, four clinical isolates were identified as MLVA-8 genotype (GT) 76 and in the canSNP A.Br.Vollum lineage, which is consistent with the Pakistani origin of the yarn. Twenty eight of the California isolates were in the A.Br.Vollum canSNP lineage and one isolate was in the A.Br. 003/004 canSNP sub-group. All 52 isolates and both clinical specimens related to the New York and Connecticut cases were MLVA-8 GT 1. The animal products associated with the NY and CT cases were believed to originate from West Africa, but no isolates from this region are available to be genotyped for comparison. All isolates associated with the New Hampshire case were identical and had a new genotype (GT 149). Isolates from the NY, CT and NH cases diverge from the established canSNP phylogeny near the base of the A.Br.011/009. This report illustrates the power of the current genotyping methods and the dramatically different epidemiological conditions that can lead to infections (i.e., contamination by a single genotype versus widespread contamination of numerous genotypes). These cases illustrate the need to acquire and genotype global isolates so that accurate assignments can be made about isolate origins.
Identification of an unusual Brucella strain (BO2) from a lung biopsy in a 52 year-old patient with chronic destructive pneumonia
Rebekah V Tiller, Jay E Gee, David R Lonsway, Sonali Gribble, Scott C Bell, Amy V Jennison, John Bates, Chris Coulter, Alex R Hoffmaster, Barun K De
BMC Microbiology , 2010, DOI: 10.1186/1471-2180-10-23
Abstract: Standard biochemical profiles confirmed that the unusual strain was a member of the Brucella genus and the full-length 16S rRNA gene sequence was 100% identical to the recently identified B. inopinata sp. nov. (type strain BO1T). Additional sequence analysis of the recA, omp2a and 2b genes; and multiple locus sequence analysis (MLSA) demonstrated that strain BO2 exhibited significant similarity to the B. inopinata sp. compared to any of the other Brucella or Ochrobactrum species. Genotyping based on multiple-locus variable-number tandem repeat analysis (MLVA) established that the BO2 and BO1Tstrains form a distinct phylogenetic cluster separate from the other Brucella spp.Based on these molecular and microbiological characterizations, we propose that the BO2 strain is a novel lineage of the newly described B. inopinata species.Brucellosis is primarily a zoonotic disease, caused by members of the genus Brucella, which currently constitutes several species based on pathogenicity, host preferences and phenotypic characteristics: B. abortus (cattle), B. canis (dogs), B. melitensis (goats), B. suis (pigs), B. ovis (rams), B. neotomae (desert rats), B. ceti and B. pinnipedialis (marine mammals), and B. microti (common vole) [1-6]. Recently, a novel species, Brucella inopinata, associated with a human infection has been recognized as the newest member of the genus Brucella [7,8]. In early 1985, whole genome hybridization analysis studies revealed a high degree of genetic homology among the Brucella species, which led to the proposal that the genus Brucella was a mono-specific species with B. melitensis being the primary species and all others as sub-species and biovars [9-11]. However, due to the limited acceptability of the one-species concept, the traditional classification of Brucella spp. based on phenotypic characteristics has been re-instated by the Brucella Taxonomy Subcommittee in 2006 [3].Brucella are facultative intracellular pathogens that infect many organs and
Comparison of DNA Extraction Kits for Detection of Burkholderia pseudomallei in Spiked Human Whole Blood Using Real-Time PCR
Nicole L. Podnecky, Mindy G. Elrod, Bruce R. Newton, Leslie A. Dauphin, Jianrong Shi, Sutthinan Chawalchitiporn, Henry C. Baggett, Alex R. Hoffmaster, Jay E. Gee
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0058032
Abstract: Burkholderia pseudomallei, the etiologic agent of melioidosis, is endemic in northern Australia and Southeast Asia and can cause severe septicemia that may lead to death in 20% to 50% of cases. Rapid detection of B. pseudomallei infection is crucial for timely treatment of septic patients. This study evaluated seven commercially available DNA extraction kits to determine the relative recovery of B. pseudomallei DNA from spiked EDTA-containing human whole blood. The evaluation included three manual kits: the QIAamp DNA Mini kit, the QIAamp DNA Blood Mini kit, and the High Pure PCR Template Preparation kit; and four automated systems: the MagNAPure LC using the DNA Isolation Kit I, the MagNAPure Compact using the Nucleic Acid Isolation Kit I, and the QIAcube using the QIAamp DNA Mini kit and the QIAamp DNA Blood Mini kit. Detection of B. pseudomallei DNA extracted by each kit was performed using the B. pseudomallei specific type III secretion real-time PCR (TTS1) assay. Crossing threshold (CT) values were used to compare the limit of detection and reproducibility of each kit. This study also compared the DNA concentrations and DNA purity yielded for each kit. The following kits consistently yielded DNA that produced a detectable signal from blood spiked with 5.5×104 colony forming units per mL: the High Pure PCR Template Preparation, QIAamp DNA Mini, MagNA Pure Compact, and the QIAcube running the QIAamp DNA Mini and QIAamp DNA Blood Mini kits. The High Pure PCR Template Preparation kit yielded the lowest limit of detection with spiked blood, but when this kit was used with blood from patients with confirmed cases of melioidosis, the bacteria was not reliably detected indicating blood may not be an optimal specimen.
Diagnostic Accuracy of Real-Time PCR Assays Targeting 16S rRNA and lipl32 Genes for Human Leptospirosis in Thailand: A Case-Control Study
Janjira Thaipadunpanit,Wirongrong Chierakul,Vanaporn Wuthiekanun,Direk Limmathurotsakul,Premjit Amornchai,Siriphan Boonslip,Lee D. Smythe,Roongrueng Limpaiboon,Alex R. Hoffmaster,Nicholas P. J. Day,Sharon J. Peacock
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0016236
Abstract: Rapid PCR-based tests for the diagnosis of leptospirosis can provide information that contributes towards early patient management, but these have not been adopted in Thailand. Here, we compare the diagnostic sensitivity and specificity of two real-time PCR assays targeting rrs or lipL32 for the diagnosis of leptospirosis in northeast Thailand.
Pre-Columbian Origins for North American Anthrax
Leo J. Kenefic, Talima Pearson, Richard T. Okinaka, James M. Schupp, David M. Wagner, Jacques Ravel, Alex R. Hoffmaster, Carla P. Trim, Wai-Kwan Chung, Jodi A. Beaudry, Jeffrey T. Foster, James I. Mead, Paul Keim
PLOS ONE , 2009, DOI: 10.1371/journal.pone.0004813
Abstract: Disease introduction into the New World during colonial expansion is well documented and had a major impact on indigenous populations; however, few diseases have been associated with early human migrations into North America. During the late Pleistocene epoch, Asia and North America were joined by the Beringian Steppe ecosystem which allowed animals and humans to freely cross what would become a water barrier in the Holocene. Anthrax has clearly been shown to be dispersed by human commerce and trade in animal products contaminated with Bacillus anthracis spores. Humans appear to have brought B. anthracis to this area from Asia and then moved it further south as an ice-free corridor opened in central Canada ~13,000 ybp. In this study, we have defined the evolutionary history of Western North American (WNA) anthrax using 2,850 single nucleotide polymorphisms (SNPs) and 285 geographically diverse B. anthracis isolates. Phylogeography of the major WNA B. anthracis clone reveals ancestral populations in northern Canada with progressively derived populations to the south; the most recent ancestor of this clonal lineage is in Eurasia. Our phylogeographic patterns are consistent with B. anthracis arriving with humans via the Bering Land Bridge. This northern-origin hypothesis is highly consistent with our phylogeographic patterns and rates of SNP accumulation observed in current day B. anthracis isolates. Continent-wide dispersal of WNA B. anthracis likely required movement by later European colonizers, but the continent's first inhabitants may have seeded the initial North American populations.
Bacillus anthracis in China and its relationship to worldwide lineages
Tatum S Simonson, Richard T Okinaka, Bingxiang Wang, W Ryan Easterday, Lynn Huynh, Jana M U'Ren, Meghan Dukerich, Shaylan R Zanecki, Leo J Kenefic, Jodi Beaudry, James M Schupp, Talima Pearson, David M Wagner, Alex Hoffmaster, Jacques Ravel, Paul Keim
BMC Microbiology , 2009, DOI: 10.1186/1471-2180-9-71
Abstract: The canSNP study indicates that there are 5 different sub-lineages/sub-groups in China out of 12 previously described world-wide canSNP genotypes. Three of these canSNP genotypes were only found in the western-most province of China, Xinjiang. These genotypes were A.Br.008/009, a sub-group that is spread across most of Europe and Asia; A.Br.Aust 94, a sub-lineage that is present in Europe and India, and A.Br.Vollum, a lineage that is also present in Europe. The remaining two canSNP genotypes are spread across the whole of China and belong to sub-group A.Br.001/002 and the A.Br.Ames sub-lineage, two closely related genotypes. MLVA typing adds resolution to the isolates in each canSNP genotype and diversity indices for the A.Br.008/009 and A.Br.001/002 sub-groups suggest that these represent older and established clades in China.B. anthracis isolates were recovered from three canSNP sub-groups (A.Br.008/009, A.Br.Aust94, and A.Br.Vollum) in the western most portion of the large Chinese province of Xinjiang. The city of Kashi in this province appears to have served as a crossroads for not only trade but the movement of diseases such as anthrax along the ancient "silk road". Phylogenetic inference also suggests that the A.Br.Ames sub-lineage, first identified in the original Ames strain isolated from Jim Hogg County, TX, is descended from the A.Br.001/002 sub-group that has a major presence in most of China. These results suggest a genetic discontinuity between the younger Ames sub-lineage in Texas and the large Western North American sub-lineage spread across central Canada and the Dakotas.Ancient Chinese medical books suggest that an anthrax-like disease has been present in China for more than 5,000 years and that by 500–600 A.D. the epidemiology and symptoms of anthrax had been described [1]. A 1995 report from China described the results of an anthrax surveillance and control project in 10 provinces in China between 1990–1994 [2]. Stations in these 10 provinces (Sic
A Single Multilocus Sequence Typing (MLST) Scheme for Seven Pathogenic Leptospira Species
Siriphan Boonsilp equal contributor,Janjira Thaipadungpanit equal contributor ,Premjit Amornchai,Vanaporn Wuthiekanun,Mark S. Bailey,Matthew T. G. Holden,Cuicai Zhang,Xiugao Jiang,Nobuo Koizumi,Kyle Taylor,Renee Galloway,Alex R. Hoffmaster,Scott Craig,Lee D. Smythe,Rudy A. Hartskeerl,Nicholas P. Day,Narisara Chantratita,Edward J. Feil,David M. Aanensen,Brian G. Spratt,Sharon J. Peacock
PLOS Neglected Tropical Diseases , 2013, DOI: 10.1371/journal.pntd.0001954
Abstract: Background The available Leptospira multilocus sequence typing (MLST) scheme supported by a MLST website is limited to L. interrogans and L. kirschneri. Our aim was to broaden the utility of this scheme to incorporate a total of seven pathogenic species. Methodology and Findings We modified the existing scheme by replacing one of the seven MLST loci (fadD was changed to caiB), as the former gene did not appear to be present in some pathogenic species. Comparison of the original and modified schemes using data for L. interrogans and L. kirschneri demonstrated that the discriminatory power of the two schemes was not significantly different. The modified scheme was used to further characterize 325 isolates (L. alexanderi [n = 5], L. borgpetersenii [n = 34], L. interrogans [n = 222], L. kirschneri [n = 29], L. noguchii [n = 9], L. santarosai [n = 10], and L. weilii [n = 16]). Phylogenetic analysis using concatenated sequences of the 7 loci demonstrated that each species corresponded to a discrete clade, and that no strains were misclassified at the species level. Comparison between genotype and serovar was possible for 254 isolates. Of the 31 sequence types (STs) represented by at least two isolates, 18 STs included isolates assigned to two or three different serovars. Conversely, 14 serovars were identified that contained between 2 to 10 different STs. New observations were made on the global phylogeography of Leptospira spp., and the utility of MLST in making associations between human disease and specific maintenance hosts was demonstrated. Conclusion The new MLST scheme, supported by an updated MLST website, allows the characterization and species assignment of isolates of the seven major pathogenic species associated with leptospirosis.
Development of a Prototype Lateral Flow Immunoassay (LFI) for the Rapid Diagnosis of Melioidosis
Raymond L. Houghton,Dana E. Reed,Mark A. Hubbard,Michael J. Dillon,Hongjing Chen,Bart J. Currie,Mark Mayo,Derek S. Sarovich,Vanessa Theobald,Direk Limmathurotsakul,Gumphol Wongsuvan,Narisara Chantratita,Sharon J. Peacock,Alex R. Hoffmaster,Brea Duval,Paul J. Brett,Mary N. Burtnick,David P. AuCoin
PLOS Neglected Tropical Diseases , 2014, DOI: 10.1371/journal.pntd.0002727
Abstract: Burkholderia pseudomallei is a soil-dwelling bacterium and the causative agent of melioidosis. Isolation of B. pseudomallei from clinical samples is the “gold standard” for the diagnosis of melioidosis; results can take 3–7 days to produce. Alternatively, antibody-based tests have low specificity due to a high percentage of seropositive individuals in endemic areas. There is a clear need to develop a rapid point-of-care antigen detection assay for the diagnosis of melioidosis. Previously, we employed In vivo Microbial Antigen Discovery (InMAD) to identify potential B. pseudomallei diagnostic biomarkers. The B. pseudomallei capsular polysaccharide (CPS) and numerous protein antigens were identified as potential candidates. Here, we describe the development of a diagnostic immunoassay based on the detection of CPS. Following production of a CPS-specific monoclonal antibody (mAb), an antigen-capture immunoassay was developed to determine the concentration of CPS within a panel of melioidosis patient serum and urine samples. The same mAb was used to produce a prototype Active Melioidosis Detect Lateral Flow Immunoassay (AMD LFI); the limit of detection of the LFI for CPS is comparable to the antigen-capture immunoassay (~0.2 ng/ml). The analytical reactivity (inclusivity) of the AMD LFI was 98.7% (76/77) when tested against a large panel of B. pseudomallei isolates. Analytical specificity (cross-reactivity) testing determined that 97.2% of B. pseudomallei near neighbor species (35/36) were not reactive. The non-reactive B. pseudomallei strain and the reactive near neighbor strain can be explained through genetic sequence analysis. Importantly, we show the AMD LFI is capable of detecting CPS in a variety of patient samples. The LFI is currently being evaluated in Thailand and Australia; the focus is to optimize and validate testing procedures on melioidosis patient samples prior to initiation of a large, multisite pre-clinical evaluation.
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