The emerging multidrug-resistant pathogenic yeast represents a serious threat to global health. Unlike most other species, this organism appears to be commonly transmitted within health care facilities and causes health care-associated outbreaks. To better understand the epidemiology of this emerging pathogen, we investigated the ability of to persist on plastic surfaces common in health care settings compared with that of, a species known to colonize the skin and plastics. Specifically, we compiled comparative and quantitative data essential to understanding the vehicles of spread and the ability of both species to survive and persist on plastic surfaces under controlled conditions (25°C and 57% relative humidity), such as those found in health care settings. When a test suspension of 10 cells was applied and dried on plastic surfaces, remained viable for at least 14 days and for at least 28 days, as measured by CFU. However, survival measured by esterase activity was higher for than throughout the 28-day study. Given the notable length of time species survive and persist outside their host, we developed methods to more effectively culture from patients and their environment. Using our enrichment protocol, public health laboratories and researchers can now readily isolate from complex microbial communities (such as patient skin, nasopharynx, and stool) as well as environmental biofilms, in order to better understand and prevent colonization and transmission.

Four swab materials were evaluated for their efficiency in recovery of Bacillus anthracis spores from steel coupons. Cotton, macrofoam, polyester, and rayon swabs were used to sample coupons inoculated with a spore suspension of known concentration. Three methods of processing for the removal of spores from the swabs (vortexing, sonication, or minimal agitation) and two swab preparations (premoistened and dry) were evaluated. Results indicated that premoistened swabs were more efficient at recovering spores than dry swabs (14.3% vs. 4.4%). Vortexing swabs for 2 min during processing resulted in superior extraction of spores when compared to sonicating them for 12 min or subjecting them to minimal agitation. Premoistened macrofoam and cotton swabs that were vortexed during processing recovered the greatest proportions of spores with a mean recovery of 43.6% (standard deviation [SD] 11.1%) and 41.7% (SD 14.6%), respectively. Premoistened and vortexed polyester and rayon swabs were less efficient, at 9.9% (SD 3.8%) and 11.5% (SD 7.9%), respectively.

Chlamydia trachomatis is an important human pathogen that replicates inside the infected host cell in a unique vacuole, the inclusion. The formation of this intracellular bacterial niche is essential for productive Chlamydia infections. Despite its importance for Chlamydia biology, a holistic view on the protein composition of the inclusion, including its membrane, is currently missing. Here we describe the host cell-derived proteome of isolated C. trachomatis inclusions by quantitative proteomics. Computational analysis indicated that the inclusion is a complex intracellular trafficking platform that interacts with host cells’ antero- and retrograde trafficking pathways. Furthermore, the inclusion is highly enriched for sorting nexins of the SNX-BAR retromer, a complex essential for retrograde trafficking. Functional studies showed that in particular, SNX5 controls the C. trachomatis infection and that retrograde trafficking is essential for infectious progeny formation. In summary, these findings suggest that C. trachomatis hijacks retrograde pathways for effective infection.

A protocol to recover Bacillus anthracis spores from a steel surface using macrofoam swabs was evaluated for its accuracy, precision, reproducibility, and limit of detection. Macrofoam swabs recovered 31.7 to 49.1% of spores from 10-cm 2 steel surfaces with a <32.7% coefficient of variation in sampling precision and reproducibility for inocula of >38 spores.Thousands of swab samples of Bacillus anthracis spores were taken during investigations of the 2001 bioterrorist events (3,7,10,11,12,13), but no validated protocol was available at the time (8). The swab protocol used during the incident was modified from swab methods used by the food industry (1), National and Aeronautics and Space Administration (5), and hospital epidemiologists (4), but these methods did not involve sampling for spores. A recent study found that macrofoam swabs recover Ն30% more spores than rayon and polyester swabs, premoistening the swab increases recovery by Ն30%, and vortexing the swab recovers Ͼ25% more spores than extraction by sonication (9).We evaluated a protocol that recovers spores from steel surfaces with premoistened macrofoam swabs and vortex processing. Steel coupons with 10-cm 2 surface area, as recommended for environmental and medical-device sampling (6), were chosen for their smooth surfaces. Ethanol was used as the inoculation medium in order to spread the spores evenly over the coupon. These conditions may increase the difficulty of removing spores with a swab compared to the removal of weapons grade powder spores. Five analysts from two laboratories performed the protocol, and the data were evaluated to determine accuracy, sampling precision, reproducibility, and limit of detection (LOD).B. anthracis Sterne 34F2 spores (Colorado Serum, Denver, CO), previously stored in 50% ethanol to prevent germination, were added to 95% ethanol to attain ϳ10 6 spores ml Ϫ1 . This suspension was serially diluted 1:10 in 95% ethanol to attain inocula (I) ranging from 4.0 ϫ 10 0 to 6.0 ϫ 10 4 spores ml Ϫ1 (Table 1). For each experiment, 10 to 15 sterile 10-cm 2 steel coupons were placed in glass petri dishes and inoculated with 0.5 ml of the appropriate inoculum of spores, as described previously (9). Petri dish lids remained partially open as the coupons dried in a biological safety cabinet for 2 h. A macrofoam swab (Critical Swab; VWR, Suwanee, GA; catalog no. 10812-046) was dipped in phosphate-buffered saline plus 0.04% Tween-80 (PBST; pH 7.2), and excess fluid was expressed from the head. The swab was swept in horizontal, vertical, and diagonal sweeps to cover the coupon during each orientation. The swab was vortexed in 5 ml PBST for 2 min at 10-s intervals (V1), placed into a second tube of PBST, and vortexed again (V2).Ten sampled and 10 unsampled coupons were processed to determine the number of spores left after swabbing (L) and recovered from inoculated controls (C), respectively. Control coupon tests were performed prior to swab experiments to ensure that spores remained on the coupons during drying. The coupons were sonicat...

In the UK, between 14% and 23% of infarct patients, between 33% and 56% of CABG patients, and between 6% and 10% of PTCA patients are enrolled into CR programs.

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This work was initiated to address the gaps identified by Congress regarding validated biothreat environmental sampling and processing methods. Nine Laboratory Response Network-affiliated laboratories participated in a validation study of a cellulose sponge wipe-processing protocol for the recovery, detection, and quantification of viable Bacillus anthracis Sterne spores from steel surfaces. Steel coupons (645.16 cm 2 ) were inoculated with 1 to 4 log 10 spores and then sampled with cellulose sponges (Sponge-Stick; 3M, St. Paul, MN). Surrogate dust and background organisms were added to the sponges to mimic environmental conditions. Labs processed the sponges according to the provided protocol. Sensitivity, specificity, and mean percent recovery (%R), between-lab variability, within-lab variability, and total percent coefficient of variation were calculated. The mean %R (standard error) of spores from the surface was 32.4 (4.4), 24.4 (2.8), and 30.1 (2.3) for the 1-, 2-, and 4-log 10 inoculum levels, respectively. Sensitivities for colony counts were 84.1%, 100%, and 100% for the 1-, 2-, and 4-log 10 inocula, respectively. These data help to characterize the variability of the processing method and thereby enhance confidence in the interpretation of the results of environmental sampling conducted during a B. anthracis contamination investigation.

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 cm 2 ). Steel and carpet coupons seeded in the chamber were sampled with swab (103 cm 2 ) or wipe or vacuum (929 cm 2 ) surface sampling methods and analyzed at three laboratories. Agar settle plates (60 cm 2 ) 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 cm 2 for sampling steel surfaces and 40, 9.2, and 28 CFU/100 cm 2 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.