Use of superabsorbent polymer gels for surface decontamination of Bacillus anthracis spores

Aims:  This study evaluated the inactivation of Bacillus anthracis Vollum spores dried on a nonporous surface using a superabsorbent polymer (SAP) gel containing commercially available liquid decontaminants.
Methods and Results:  The first phase determining the availability of the liquid decontaminant within the SAP showed that the SAP gel containing pH-adjusted sodium hypochlorite (NaOCl) inhibited B. anthracis growth while the water control SAP gel had no affect on growth. For testing surface decontamination, B. anthracis spores were dried onto steel coupons painted with chemical agent resistant coating and exposed to SAP containing either pH-adjusted NaOCl, chlorine dioxide (ClO₂) or hydrogen peroxide/peracetic acid (H₂O₂/PA) for 5 and 30 min. At contact times of both 5 and 30 min, all of the SAP gels containing pH-adjusted NaOCl, ClO₂ or H₂O₂/PA inactivated B. anthracis spores at levels ranging from 2·2 to ≥7·6 log reductions.
Conclusions:  Incorporation of three commercially available decontaminant technologies into a SAP gel promotes inactivation of B. anthracis spores without observable physical damage to the test surface.
Significance and Impact of the Study:  This work provides preliminary data for the feasibility of using SAP in inactivating B. anthracis spores on a nonporous surface, supporting the potential use of SAP in surface decontamination.     

A combined immunomagnetic separation and lateral flow method for a sensitive on-site detection of Bacillus anthracis spores – assessment in water and dairy products

Aim:  Combination of immunomagnetic separation (IMS) and lateral flow device (LFD) assays for the development of a sensitive, rapid, on-site methodology that enables concentration and detection of Bacillus anthracis spores in complex samples.
Methods and Results:  The data presents the development of an optimized, 30 min, IMS assay, with about 95% capture of B. anthracis spores from different dairy products ( n  = 38). No cross reactivity was detected with typical milk flora and some closely related Bacilli. To enable direct application of the IMS captured spores on the LFD, spores were eluted from the bead–spore complex utilizing 95% (v/v) formamide-10 mmol l⁻¹ EDTA for 30 s in a microwave oven. Detached spores were analysed on LFD enabling detection within 10 min. The combined IMS–LFD methodology (40 min) demonstrates a 60-fold improvement in sensitivity, relative to samples that were applied directly on the LFD without the IMS concentrating step.
Conclusions:  The IMS–LFD method is a powerful platform, combining rapidity, specificity and efficiency for concentrating and detecting B. anthracis from water and milk contaminated samples.
Significant and Impact of the Study:  The combination of IMS and LFD enhances the sensitivity and flexibility of B. anthracis spore detection from complex samples. This method can potentially be extended to other toxins and micro-organisms in a variety of matrices.     

Evaluation of different methods to discriminate Bacillus anthracis from other bacteria of the Bacillus cereus group

Aims:  To evaluate different methods that are useful for rapid and definitive discrimination of Bacillus anthracis from other bacteria of the Bacillus cereus group in environmental samples like letters claimed to contain anthrax spores.
Methods and Results:  Characterized strains and bacteria from environmental samples were analysed by microbiological and molecular methods (PCR and restriction analysis). Environmental isolates often shared several microbiological features with B. anthracis , e.g. lack of β -haemolysis and phospholipase C activity, and only the gamma phage assay was specific for B. anthracis . PCR assays targeting markers from the virulence plasmids exclusively detected B. anthracis , but other PCR targets were also detected in nonanthrax isolates. Additionally, the restriction pattern in an Alu I restriction analysis of the SG-749 fragment is not 100% specific. The loci used for multiple-locus variable-number tandem repeat analysis of B. anthracis are also present in other members of the B. cereus group, but amplicon sizes are usually different.
Conclusions:  Environmental samples often contain borderline isolates closely related to B. anthracis both on microbiological and genetic levels. Real-time PCR targeting plasmidal and chromosomal markers should be used for rapid and definitive exclusion of a virulent strain of B. anthracis in such samples.
Significance and Impact of the Study:  This study gives an overview of the current microbiological and molecular methods used for identification of B. anthracis and shows that most assays have limits when borderline isolates present in environmental samples are analysed.     

Analysis of dye binding by and membrane potential in spores of Bacillus species

Aims:  To determine roles of coats in staining Bacillus subtilis spores, and whether spores have membrane potential.
Methods and Results:  Staining by four dyes and autofluorescence of B. subtilis spores that lack some ( cotE , gerE ) or most ( cotE gerE) coat protein was measured. Wild-type, cotE and gerE spores autofluorescenced and bound dyes, but cotE gerE spores did not autofluorescence and were stained only by two dyes. A membrane potential-sensitive dye DiOC₆(3) bound to dormant Bacillus megaterium and B. subtilis spores. While this binding was abolished by the protonophore FCCP, DiOC₆(3) bound to heat-killed spores, but not to dormant B. subtilis cotE gerE spores. However, DiOC₆(3) bound well to all germinated spores.
Conclusions:  The autofluorescence of dormant B. subtilis spores and the binding of some dyes are due to the coat. There is no membrane potential in dormant Bacillus spores, although membrane potential is generated when spores germinate.
Significance and Impact of the Study:  The elimination of the autofluorescence of B. subtilis spores may allow assessment of the location of low abundance spore proteins using fluorescent reporter technology. The dormant spore's lack of membrane potential may allow tests of spore viability by assessing membrane potential in germinating spores.     

Immuno-detection of anthrose containing tetrasaccharide in the exosporium of Bacillus anthracis and Bacillus cereus strains

Aims:  Bacillus anthracis strains of various origins were analysed with the view to describe intrinsic and persistent structural components of the Bacillus collagen-like protein of anthracis glycoprotein associated anthrose containing tetrasaccharide in the exosporium.
Methods and Results:  The tetrasaccharide consists of three rhamnose residues and an unique monosaccharide – anthrose. As anthrose was not found in spores of related strains of bacteria, we envisioned the detection of B. anthracis spores based on antibodies against anthrose-containing polysaccharides. Carbohydrate–protein conjugates containing the synthetic tetrasaccharide, an anthrose–rhamnose disaccharide or anthrose alone were employed to immunize mice. All three formulations were immunogenic and elicited IgG responses with different fine specificities. All sera and monoclonal antibodies derived from tetrasaccharide immunized mice cross-reacted not only with spore lysates of a panel of virulent B. anthracis strains, but also with some of the B. cereus strains tested.
Conclusions:  Our results demonstrate that antibodies to synthetic carbohydrates are useful tools for epitope analyses of complex carbohydrate antigens and for the detection of particular target structures in biological specimens.
Significance and Impact of the Study:  Although not strictly specific for B. anthracis spores, antibodies against the tetrasaccharide may have potential as immuno-capturing components for a highly sensitive spore detection system.     

The combined effects of high pressure and nisin on germination and inactivation of Bacillus spores in milk

Aims:  The aim of this work was to investigate the germination and inactivation of spores of Bacillus species in buffer and milk subjected to high pressure (HP) and nisin.
Methods and Results:  Spores of Bacillus subtilis and Bacillus cereus suspended in milk or buffer were treated at 100 or 500 MPa at 40°C with or without 500 IU ml⁻¹ of nisin. Treatment at 500 MPa resulted in high levels of germination (4 log units) of B. subtilis spores in both milk and buffer; this increased to >6 logs by applying a second cycle of pressure. Viability of B. subtilis spores in milk and buffer was reduced by 2·5 logs by cycled HP, while the addition of nisin (500 IU ml⁻¹) prior to HP treatment resulted in log reductions of 5·7 and 5·9 in phosphate buffered saline and milk, respectively. Physical damage of spores of B. subtilis following HP was apparent using scanning electron microscopy. Treating four strains of B. cereus at 500 MPa for 5 min twice at 40°C in the presence of 500 IU ml⁻¹ nisin proved less effective at inactivating the spores of these isolates compared with B. subtilis and some strain-to-strain variability was observed.
Conclusions:  Although high levels of germination of Bacillus spores could be achieved by combining HP and nisin, complete inactivation was not achieved using the aforementioned treatments.
Significance and Impact of the Study:  Combinations of HP treatment and nisin may be an appealing alternative to heat pasteurization of milk.     

'Bioluminescent' reporter phage for the detection of Category A bacterial pathogens

Yersinia pestis and Bacillus anthracis are Category A bacterial pathogens that are the causative agents of the plague and anthrax, respectively. Although the natural occurrence of both diseases' is now relatively rare, the possibility of terrorist groups using these pathogens as a bioweapon is real. Because of the disease's inherent communicability, rapid clinical course, and high mortality rate, it is critical that an outbreak be detected quickly. Therefore methodologies that provide rapid detection and diagnosis are essential to ensure immediate implementation of public health measures and activation of crisis management. Recombinant reporter phage may provide a rapid and specific approach for the detection of Y. pestis and B. anthracis. The Centers for Disease Control and Prevention currently use the classical phage lysis assays for the confirmed identification of these bacterial pathogens. These assays take advantage of naturally occurring phage which are specific and lytic for their bacterial hosts. After overnight growth of the cultivated bacterium in the presence of the specific phage, the formation of plaques (bacterial lysis) provides a positive identification of the bacterial target. Although these assays are robust, they suffer from three shortcomings: 1) they are laboratory based; 2) they require bacterial isolation and cultivation from the suspected sample, and 3) they take 24-36 h to complete. To address these issues, recombinant "light-tagged" reporter phage were genetically engineered by integrating the Vibrio harveyi luxAB genes into the genome of Y. pestis and B. anthracis specific phage. The resulting luxAB reporter phage were able to detect their specific target by rapidly (within minutes) and sensitively conferring a bioluminescent phenotype to recipient cells. Importantly, detection was obtained either with cultivated recipient cells or with mock-infected clinical specimens. For demonstration purposes, here we describe the method for the phage-mediated detection of a known Y. pestis isolate using a luxAB reporter phage constructed from the CDC plague diagnostic phage ΦA1122 (Figure 1). A similar method, with minor modifications (e.g. change in growth temperature and media), may be used for the detection of B. anthracis isolates using the B. anthracis reporter phage Wβ::luxAB. The method describes the phage-mediated transduction of a biolumescent phenotype to cultivated Y. pestis cells which are subsequently measured using a microplate luminometer. The major advantages of this method over the traditional phage lysis assays is the ease of use, the rapid results, and the ability to test multiple samples simultaneously in a 96-well microtiter plate format. Figure 1. Detection schematic. The phage are mixed with the sample, the phage infects the cell, luxAB are expressed, and the cell bioluminesces. Sample processing is not necessary; the phage and cells are mixed and subsequently measured for light.     

Modelling the photosensitization-based inactivation of Bacillus cereus

Aims:  To study and to develop a model for the photo-destruction of the foodborne pathogen Bacillus cereus , initially treated with a precursor of endogenous photosensitizers (5-aminolevulinic acid, ALA).
Materials and methods:  The cells were incubated in the presence of ALA (3 or 7·5 mmol l⁻¹) for incubation times ranging from 2 to 60 min, inoculated onto the surface of LB Agar plates and submitted to light irradiation. The Weibull model was used to describe the survival curves of B. cereus . Quadratic equations were used to describe the effects of ALA concentration and incubation time on the Weibull model parameters.
Results:  ALA-based photosensitization proved to be an effective tool for inactivation of B. cereus . The decrease in viable counts observed after 20 min of irradiation, ranged from 4 to 6 log CFU g⁻¹.
Conclusions:  The developed model proved to be a parsimonious and robust solution to describe the observed data.
Significance and Impact of the Study:  The study demonstrates the effectiveness of photosensitization on B. cereus on agar plates. The model developed may be useful to optimize inactivation treatments by photosensitization.     

Decontamination assessment of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surfaces using a hydrogen peroxide gas generator

AIMS: To evaluate the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials using hydrogen peroxide gas. METHODS AND RESULTS: Bacillus anthracis, B. subtilis, and G. stearothermophilus spores were dried on seven types of indoor surfaces and exposed to > or =1000 ppm hydrogen peroxide gas for 20 min. Hydrogen peroxide exposure significantly decreased viable B. anthracis, B. subtilis, and G. stearothermophilus spores on all test materials except G. stearothermophilus on industrial carpet. Significant differences were observed when comparing the reduction in viable spores of B. anthracis with both surrogates. The effectiveness of gaseous hydrogen peroxide on the growth of biological indicators and spore strips was evaluated in parallel as a qualitative assessment of decontamination. At 1 and 7 days postexposure, decontaminated biological indicators and spore strips exhibited no growth, while the nondecontaminated samples displayed growth. CONCLUSIONS: Significant differences in decontamination efficacy of hydrogen peroxide gas on porous and nonporous surfaces were observed when comparing the mean log reduction in B. anthracis spores with B. subtilis and G. stearothermophilus spores. SIGNIFICANCE AND IMPACT OF THE STUDY: These results provide comparative information for the decontamination of B. anthracis spores with surrogates on indoor surfaces using hydrogen peroxide gas.     

Real-time quantification of Pseudomonas fluorescens cell removal from glass surfaces due to bacteriophage varphiS1 application

Aims:  To study the efficacy of the lytic phage φS1 in eliminating Pseudomonas fluorescens in the early stage of biofilm formation, using an in situ and real time methodology for cell quantification.
Methods and Results:  Cell adhesion and phage infection studies were carried out in a parallel plate flow chamber under laminar conditions. Cells were allowed to adhere until reaching 1·7–1·8 × 10⁶ cells cm⁻² and phage infection was performed with two different phage concentrations (2 × 10⁹ PFU ml⁻¹ and 1 × 10¹⁰ PFU ml⁻¹). Phage concentration clearly affects the speed of infection. The less concentrated phage solution promoted a three times slower rate of cell removal but did not affect the overall percentage of cell removal. In fact, after a longer infection period the less concentrated phage solution reached the same 93% cell removal value.
Conclusions:  Phages are efficient in the eradication of bacterial cells at the early stage of biofilm formation and their presence at the surface did not allow bacterial recolonization of the surface.
Significance and Impact of the Study:  To date, no published studies have been made concerning in situ and real time quantification of cell removal from surfaces due to phage action.     

Bactericidal and sporicidal performance of a polymer-encapsulated chlorine dioxide-coated surface

Aims:  To investigate the physical characteristics and the bactericidal and sporicidal potential of a polymer-encapsulated ClO₂ coating.
Methods and Results:  An antimicrobial coating based on polymer-encapsulated ClO₂ was developed. A low viscosity, water/oil/water double emulsion coating was formulated for easy on-site application. Escherichia coli , Pseudomonas aeruginosa , Bacillus subtilis and Staphylococcus aureus were applied onto the coating to study the bactericidal capabilities of the coating. The bactericidal performance of the coating increased when the contact time with the tested bacteria increased. Over 99% of the E. coli , Ps. aeruginosa , B. subtilis were killed with a contact time of 30 min. Although endospores of B. subtilis are more resistant, about 75% of the spores were killed after 72 h on the coating. Moreover, a sustained release of gaseous ClO₂ was achieved to maintain about 90% removal of B. subtilis with a 10-min contact time during a 28-day study period. The coating also exhibits antiadhesive properties against bacteria.
Conclusions:  A polymer-encapsulated ClO₂ coating with sustained release of ClO₂ and promising bactericidal and sporicidal features was tested for 28 days.
Significance and Impact of the Study:  This study provides a new direction for developing polymer-encapsulated ClO₂ coatings that possess persistent bactericidal and sporicidal properties.     

Development of a PCR assay for identification of the Bacillus cereus group species

Aims:  A PCR technique was developed as a reliable and rapid identification method for the Bacillus cereus group species, based on a unique conserved sequence of the motB gene (encoding flagellar motor protein) from B. cereus , Bacillus thuringiensis and Bacillus anthracis .
Methods and Results:  Primer locations were identified against eight strains of the B. cereus group spp. from nucleotide sequences available in the National Centre for Biotechnology Information database. The PCR assay was applied for the identification of 117 strains of the B. cereus group spp. and 19 strains from other microbial species, with special emphasis on foodborne pathogens.
Conclusion:  The designed cross-species primers are group specific and did not react with DNA from other Bacillus and non- Bacillus species either motile or not. The primers system enabled us to detect 10³ CFU of B. cereus cells per millilitre of sample.
Significance and Impact of the Study:  Bacillus cereus group spp. belongs to one of the most prevalent foodborne pathogens. Bacterial growth results in production of different toxins; therefore, consumption of food containing >10⁶ bacteria per gram may result in emetic and diarrhoeal syndromes. A rapid and sensitive bacterial detection method is significant for food safety.     

Nonisothermal heat resistance determinations with the thermoresistometer Mastia

Aims:  To design and build a thermoresistometer, named Mastia, which could perform isothermal and nonisothermal experiments.
Methods and Results:  In order to evaluate the thermoresistometer, the heat resistance of Escherichia coli vegetative cells and Alicyclobacillus acidoterrestris spores was explored. Isothermal heat resistance of E. coli was characterized by D _60°C = 0·38 min and z =  4·7°C in pH 7 buffer. When the vegetative cells were exposed to nonisothermal conditions, their heat resistance was largely increased at slow heating and fast cooling rates. Isothermal heat resistance of A. acidoterrestris was characterized by D _95°C = 7·4 min and z =  9·5°C in orange juice. Under nonisothermal conditions, inactivation was reasonably well predicted from isothermal data.
Conclusions:  The thermoresistometer Mastia is a very suitable instrument to get heat resistance data of micro-organisms under isothermal and nonisothermal treatments.
Significance and Impact of the Study:  The thermoresistometer Mastia can be a helpful tool for food processors in order to estimate the level of safety of the treatments they apply.     

Indirect detection of Bacillus anthracis using real-time PCR to detect amplified gamma phage DNA

Typical real-time PCR methods used to identify Bacillus anthracis do not distinguish between viable and non-viable spores, which would be critical in any first response and remediation scenarios. This study combined both real-time PCR, using primers specifically designed for gamma phage, with the highly specific gamma phage amplification into one simple assay to indirectly detect Bacillus anthracis. Since the amplification of gamma phage only occurs in the presence of a suitable host, the detection of increasing concentrations of progeny gamma phage DNA using real-time PCR implies the presence of viable Bacillus anthracis cells. This method detected a starting Bacillus anthracis concentration of 207 cfu/mL, equivalent to less than one cell in 20 microL, in less than 5 h.     

A DNA microarray facilitates the diagnosis of Bacillus anthracis in environmental samples

Aims:  In order to improve the diagnosis of Bacillus anthracis in environmental samples, we established a DNA microarray based on the ArrayTube technology of Clondiag.
Methods and Results:  Total DNA of a bacterial colony is randomly biotinylated and hybridized to the array. The probes on the array target the virulence genes, the genomic marker gene rpoB , as well as the selective 16S rDNA sequence regions of B. anthracis , of the Bacillus cereus group and of Bacillus subtilis . Eight B. anthracis reference strains were tested and correctly identified. Among the analysed environmental Bacillus isolates, no virulent B. anthracis strain was detected.
Conclusions:  This array clearly differentiates B. anthracis from members of the B. cereus group and other Bacillus species in environmental samples by chromosomal ( rpoB ) and plasmid markers. Additionally, recognition of B. cereus strains harbouring the toxin genes or atypical B. anthracis strains that have lost the virulence plasmids is feasible.
Significance and Impact of the Study:  The array is applicable to the complex diagnostics for B. anthracis detection in environmental samples. Because of low costs, high security and easy handling, the microarray is applicable to routine diagnostics.     

Inactivation of Mycobacterium avium ssp. paratuberculosis in milk by UV treatment

Aims:  To determine the effect of UV radiation on the viability of two strains of Mycobacterium avium ssp. paratuberculosis (Map) inoculated into milk.
Methods and Results:  Mycobacterium avium ssp. paratuberculosis in a ultra heat treated milk matrix was subjected to increasing doses of UV-C radiation from 0 to 1836 mJ ml⁻¹ using a pilot-scale UV reactor (20 l capacity). Survival of Map was monitored by culture on Herrold's egg yolk medium, Middlebrook 7H10 medium and the FASTPlaque TB™ phage assay. Differences in sensitivity to UV treatment were observed between strains, however, at 1000 mJ ml⁻¹ a Map kill rate of 0·1–0·6 log₁₀ was achieved regardless of strain used or method employed to enumerate Map. Although the inactivation trend was similar on the culture and phage assay, the former gave a consistently higher viable count.
Conclusions:  The use of UV radiation alone does not represent an alternative to current pasteurization regimes for a large reduction in viable Map in milk.
Significance and Impact of the Study:  To the authors' knowledge the work here represents the first pilot-scale UV treatment process used to assess UV efficacy to inactivate Map in milk. The results are similar to those obtained with a laboratory-scale process indicating the difficulties associated with UV treatment of an opaque liquid and the recalcitrance of Map towards inimical treatments.     

Bovicin HC5 reduces thermal resistance of Alicyclobacillus acidoterrestris in acidic mango pulp

Aims:  To test the effect of bovicin HC5 against vegetative cells and endospores of Alicyclobacillus acidoterrestris DSMZ 2498 in synthetic media and in acidic mango pulp.
Methods and Results:  Alicyclobacillus acidoterrestris was grown in synthetic medium at 40°C and pH 4·0. The effect on vegetative cells was assayed by adding bovicin HC5 to synthetic medium (40–160 AU ml⁻¹) or to mango pulp (100 AU ml⁻¹) at various pH values and determining the effect on growth (OD_600nm) and viable cell number, respectively. The effect of bovicin HC5 on spore germination and thermal sensitivity of A. acidoterrestris was tested in mango pulp (pH 4·0) containing 80 AU ml⁻¹ of bovicin HC5. Bovicin HC5 was bactericidal against vegetative cells of A. acidoterrestris at different pH values and showed sporicidal activity against endospores of this bacterium. When spores of A. acidoterrestris were heat treated in the presence of bovicin HC5, D -values decreased 77% to 95% compared to untreated controls at temperatures ranging from 80 to 95°C.
Conclusion:  Bovicin HC5 was bactericidal and sporicidal against A. acidoterrestrsi DSMZ 2498.
Significance and Impact of the Study:  These results indicated that bovicin HC5 has potential to prevent spoilage of acidic fruit juices by thermocidophilic spore-forming bacteria.     

Integration and decontamination of Bacillus cereus in Pseudomonas fluorescens biofilms

Aims:  The hypothesis that surrogate planktonic pathogens ( Bacillus cereus and polystyrene microspheres) could be integrated in biofilms and protected from decontamination was tested.
Methods and Results:  Pseudomonas fluorescens biofilms were grown on polyvinyl chloride coupons in annular reactors under low nutrient conditions. After biofilm growth, B. cereus spores and polystyrene microspheres (an abiotic control) were introduced separately. Shear stress at the biofilm surface was varied between 0·15 and 1·5 N m⁻². The amount of surrogate pathogens introduced ranged from approximately 10⁵ CFU ml⁻¹ to 10¹⁰ spheres ml⁻¹. The quantity of surrogate pathogens integrated in the biofilm was proportional to the amount introduced. In 14 of the 16 cases, 0·4–3·0% of the spores or spheres introduced were measured in the biofilms. The other two cases had 10% and 21% of the spores detected. Data suggested that the spores germinated in the system. The amount of surrogate pathogens detected in the biofilms was higher in the mid-shear range. Chlorine treatment reduced the quantity of both surrogate pathogens and biofilm organisms. In one experiment, the biofilms and B. cereus recovered when the chlorine treatment was terminated.
Conclusions:  Planktonic surrogate pathogens can be integrated in biofilms and protected from chlorination decontamination.
Significance and Impact of the Study:  This knowledge assists in understanding the impact of biofilms on harbouring potential pathogens in drinking-water systems and protecting the pathogens from decontamination.     

Comparison of direct and indirect immunoradiometric assays (IRMA) for Bacillus anthracis spores immobilised on multispot microscope slides

A solid phase immunoradiometric assay (IRMA) is described in which Bacillus anthracis spores were heat fixed to the wells of glass multispot microscope slides. Assays for spores of B. anthracis Vollum and Sterne strains with ³H labels were evaluated in the direct and indirect versions. Neither signal nor signal-to-noise characteristics of indirect assays were greatly improved by the use of immunopurified antibody (IPAB) or IgG anti-bacterial reagents rather than antiserum. However, the specificity of the direct and indirect assays for B. anthracis strains and B. cereus NCTC 8035 was altered by immunopurification of the anti-bacterial reagent. Although the signal-to-noise ratio was sometimes higher in indirect than in direct assays, signal values were usually no better. Evidence was produced that the overall ratio of the indirect: direct antibody molecules bound by preparations of B. anthracis spores rarely exceeded two but the antibody-molecular ratio for antigens on extracellular material in spore preparations was much higher than the ratio for antigens on the spores themselves.     

Formaldehyde gas inactivation of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials

AIMS: To evaluate the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials using formaldehyde gas. METHODS AND RESULTS: B. anthracis, B. subtilis, and G. stearothermophilus spores were dried on seven types of indoor surfaces and exposed to approx. 1100 ppm formaldehyde gas for 10 h. Formaldehyde exposure significantly decreased viable B. anthracis, B. subtilis, and G. stearothermophilus spores on all test materials. Significant differences were observed when comparing the reduction in viable spores of B. anthracis with B. subtilis (galvanized metal and painted wallboard paper) and G. stearothermophilus (industrial carpet and painted wallboard paper). Formaldehyde gas inactivated>or=50% of the biological indicators and spore strips (approx. 1x10(6) CFU) when analyzed after 1 and 7 days. CONCLUSIONS: Formaldehyde gas significantly reduced the number of viable spores on both porous and nonporous materials in which the two surrogates exhibited similar log reductions to that of B. anthracis on most test materials. SIGNIFICANCE AND IMPACT OF THE STUDY: These results provide new comparative information for the decontamination of B. anthracis spores with surrogates on indoor surfaces using formaldehyde gas.