Mixture Of Spores Research Articles

Growth responses of maritime sand dune plant species to arbuscular mycorrhizal fungi

In a pot experiment conducted in a greenhouse, the response of 6 plant species dominating in the succession of vegetation of a deflation hollow of the Łeba Bar to inoculation with arbuscular mycorrhizal fungi (AMF) was investigated. The inoculum was a mixture of soil, roots and spores of 5 species of AMF with the dominant species <i>Glomus aggregatum</i>. Except for <i>Corynephorus canescens</i> and <i>Festuca rubra</i> subsp. <i>arenaria</i>, both the growth and the dry matter of above-ground parts of plants of <i>Agrostis stolonifera, Ammophila arenaria, Corynephorus canescens, Juncus articulatus</i> and <i>J. balticus</i> inoculated with AMF were higher than those growing in soils lacking infection propagules of these fungi. Inoculation with AMF decreased the dry matter of root: shoot ratios in 5 plant species. This property was not determined in <i>Festuca rubra</i> subsp. <i>arenaria</i> due to the death of all control plants. The level of mycorrhizal infection was low and did not correlate with the growth responses found. The high growth reaction of <i>Juncus</i> spp. to AMF found in this study suggests that the opinion of non-mycotrophy or low dependence of plants of <i>Juncaceae</i> on AMF was based on results of investigations of plants growing in wet sites known to inhibit the formation of mycorrhizae.

Development of an eco-friendly approach for biogenesis of silver nanoparticles using spores of Bacillus athrophaeus

The biological synthesis methods have been emerging as a promising new approach for production of nanoparticles due to their simplicity and non-toxicity. In the present study, spores of Bacillus athrophaeus were used to achieve the objective of developing a green synthesis method of silver nanoparticles. Enzyme assay revealed that the spores and their heat inactivated forms (microcapsules) were highly active and their enzymatic contents differed from the vegetative cells. Laccase, glucose oxidase, and alkaline phosphatase activities were detected in the dormant forms, but not in the vegetative cells. Although no nanoparticle was produced by active cells of B. athrophaeus, both spores and microcapsules were efficiently capable of reducing the silver ions (Ag⁺) to elemental silver (Ag⁰) leading to the formation of nanoparticles from silver nitrate (AgNO₃). The presence of biologically synthesized silver nanoparticles was determined by obtaining broad spectra with maximum absorbance at 400 nm in UV-visible spectroscopy. The X-ray diffraction analysis pattern revealed that the nanoscale particles have crystalline nature with various topologies, as confirmed by transmission electron microscopy (TEM). The TEM micrograph showed the nanocrystal structures with dimensions ranging from 5 to 30 nm. Accordingly, the spore mixture could be employed as a factory for detoxification of heavy metals and subsequent production of nanoparticles. This research introduces an environmental friendly and cost effective biotechnological process for the extracellular synthesis of silver nanoparticles using the bacterial spores.

A NEW INTEGRATED STRATEGY FOR CONTROLLING THE DESERT LOCUST SCHISTOCERCA GREGARIA (FORSKAL)

Nowadays, there is an upsurge in the locust and grasshopper populations in many parts of the world. Any way despite of the increase use of agrochemicals, that reduced the attack by insects. These chemicals are still represents a high risk to field workers and consumers. Accordingly, the use of biological control in these days as a particle solution is too desirable as the only alternative solution to the problem of chemical pesticides usage for long time. Our study revealed that the mortality percentage of the first nymphal instar of the desert locust was 100% after 7, 10 And 19 days when treated with mixture of conc. 5 x 1010 added to 100ppm μl of 10% consult (inhibitor compound), and Metarhizium anisopliae fungal spores at concentrations 5 ×1010, 5 ×109 Spores/ml., respectively. On the other hand, mortality percentage reached 98 and 80% after 6 days by treatment with 100ppm and 0.1ppm of the inhibitor compound respectively. Also, the mortality percentage of the second nymphal instar attained 100% after 20 days by treatment with Conc. 5 x 109 Spores/ml. of fungal spores. Amazingly, the mortality percentage was 100% after 17 days for the third nymphal instar, this by mixing the fungal spores at higher Conc. 5 x 1010 Spores/ml. With the insecticide at Conc. of 100ppm interestingly, the third and fourth nymphal instar, were less sensitive toward lower concentration of insecticide than that of fungal spores usage. Any way, the biochemical pattern revealed that the carbohydrate concentration in haemolymph of infected adult locust was higher than that in the adult non infected control. On the other hand lipid contents and total cholesterol, of the growth inhibitor treated adult insect was sharply increased after 3 days of treatment. Moreover, results showed that, the treatment with fungal spores reduced both the lipid and total cholesterol contents in the insect nymph, while total protein in the nymph of adult locust insect was increased with treatment by both fungal spores and growth inhibitor as a mixture. Also results revealed that the mortality percentage was 100% in both cases of adult and immature insect after 35 days and, 72% after 24 days. Moreover the treatment with mixture of both fungal spores and growth inhibitor reduced the hatchability percentage up to 47.2% in case of treated immature insects compared with 95.5 to the control. However, results also showed that the percentage was 16.9% in case of adult when compared with control. Also sterility percentage was 93.12% in case of immature insect to control, and 97.82% for mature ones compared with the control.

Molecular Variation of Sporisorium scitamineum in Mainland China Revealed by RAPD and SRAP Markers.

Sugarcane smut caused by Sporisorium scitamineum occurs worldwide, causing serious losses in sugar yield and quality. To study the molecular variation of S. scitamineum, 23 S. scitamineum isolates collected from the six primary sugarcane production areas in mainland China (Guangxi, Yunnan, Guangdong, Hainan, Fujian, and Jiangxi provinces) were assessed by random amplified polymorphic DNA (RAPD) and sequence-related amplified polymorphism (SRAP) markers. The results of RAPD, SRAP, and RAPD-SRAP combined analysis showed that, whereas the molecular variation of S. scitamineum was associated with geographic origin, there was no evidence of co-evolution between sugarcane and the pathogen. The results of RAPD, SRAP, or RAPD-SRAP combined analysis also did not provide any information about race differentiation of S. scitamineum. This suggests that the mixture of spores from sori collected from different areas should be used in artificial inoculations for resistance breeding and selection.

Botulism challenge studies of a modified atmosphere package for fresh mussels: inoculated pack studies.

A series of botulism challenge studies were performed to determine the possibility of production of botulinum toxin in mussels (Mytilus edulis) held under a commercial high-oxygen (60 to 65% O(2)), modified atmosphere packaging (MAP) condition. Spore mixtures of six strains of nonproteolytic Clostridium botulinum were introduced into mussel MAP packages receiving different packaging buffers with or without the addition of lactic acid bacteria. Dye studies and package flipping trials were conducted to ensure internalization of spores by packed mussels. Inoculated mussel packages were stored at normal (4°C) and abusive (12°C) temperatures for 21 and 13 days, respectively, which were beyond the packaged mussels' intended shelf life. Microbiological and chemical analyses were conducted at predetermined intervals (a total of five sampling times at each temperature), including total aerobic plate counts, C. botulinum counts, lactic acid bacterial counts, package headspace gas composition, pH of packaging buffer and mussel meat, and botulinum toxin assays of packaging buffer and mussel meat. Results revealed that C. botulinum inoculated in fresh mussels packed under MAP packaging did not produce toxin, even at an abusive storage temperature and when held beyond their shelf life. No evidence was found that packaging buffers or gas composition influenced the lack of botulinum toxin production in packed mussels.

Mortality and repellent effects of microbial pathogens on Coptotermes formosanus (Isoptera: Rhinotermitidae)

BackgroundTwo entomopathogenic fungi, Isaria fumosorosea and Metarhizium anisopliae, and one bacterium, Bacillus thuringiensis, were tested for their ability to cause mortality of Formosan subterranean termites (FST), Coptotermes formosanus (Shiraki), after liquid exposure, and for their lack of propensity to repel FST.ResultsThe fungus Isaria fumosorosea at 108 spores/ml caused 72.5% mortality on day 7, significantly higher than the control and 106 spores/ml treatment. On day 14, the 106 and 108 concentrations caused 38.8% and 92.5% mortality, respectively, significantly higher than the control. On day 21, 82.5% and 100% of the termites were killed by the 106 and 108 treatments, respectively. I. fumosorosea did not repel termites at 106 nor 108 spores/g in sand, soil or sawdust. The fungus Metarhizium anisopliae at 108 spores/ml caused 57.5% mortality on day 7, 77.5% mortality on day 14 and 100% mortality on day 21.ConclusionsOn all three days the rate of mortality was significantly higher than that of the control and 106 spores/ml treatment with I. fumosorosea. Neither I. fumosorosea nor M. anisopliae caused repellency of FST in sand, soil or sawdust. The bacterium Bacillus thuringiensis did not cause significant mortality on days 7, 14 or 21. When termites were exposed to cells of B. thuringiensis in sawdust and when termites were exposed to a mixture of spores and cells in sand, a significantly higher number remained in the control tubes. Repellency was not seen with B. thuringiensis spores alone, nor with the above treatments in the other substrates.

Occurrence, characterization and insecticidal activity of Bacillus thuringiensis strains isolated from argan fields in Morocco

Soils collected from five locations in the argan forest (an endemic plant) in Morocco were used to form the first collection of Bacillus thuringiensis (Bt) strains from this area (58 strains). Here we found that the argan forest is a major source of Bt, as 90.62% of the samples contained Bt strains. These strains produced mainly spherical or irregular crystals that in some cases remained adhered to the spore after cell lysis. There was no strain producing bipyramidal crystals, suggesting the absence of strains bearing cry1 genes. This was confirmed by PCR analysis using eight primer pairs that can potentially detect 13 different groups of cry and cyt genes. Strains containing cry7/8 were the most abundant (25.53%), followed by strains harbouring cry9A (14.89%), cry11 (8.51%) and cry4 (4.25%). The mixtures of spores and crystals as well as culture supernatants were assayed for toxicity towards Ceratitis capitata (Medfly), showing up to 30% mortality. Our findings suggest that the argan region is a suitable target for future and wider screening programmes looking for strains bearing toxins or combinations of them to develop more efficient Bt‐based formulates.

Dicty Tiger Nixes Cheaters

Polymorphic receptors prevent cheaters from avoiding altruistic cell death in Dictyostelium .

Interactions between Fusarium verticillioides and Fusarium graminearum in maize ears and consequences for fungal development and mycotoxin accumulation

Fungal interactions of Fusarium verticillioides and F. graminearum in maize ears and the impact on fungal development and toxin accumulation were investigated in a 2‐year field study at two locations in France. Maize ears were inoculated either with a spore mixture of F. graminearum and F. verticillioides or using a sequential inoculation procedure consisting of a first inoculation with F. graminearum followed by a second with F. verticillioides 1 week later. Toxin and fungal biomass were assessed on mature kernels, using HPLC and quantitative PCR. Correlation between the levels of DNA and toxin was high concerning F. graminearum DNA and deoxynivalenol (R² = 0·73) and moderate for F. verticillioides DNA and fumonisin (R² = 0·44). Fusarium graminearum DNA either decreased in mixed inoculations or was not influenced by subsequent inoculations with F. verticillioides, compared to single inoculations. In contrast, F. verticillioides DNA either significantly increased or was not affected in mixed and sequential inoculations. In two of the replicates, it can be assumed that natural contamination by F. verticillioides was favoured by previous contamination with F. graminearum. Overall, the results suggest that F. verticillioides has competitive advantages over the F. graminearum strains. Additionally, the data provide, for the first time, key evidence that previous contamination by F. graminearum in maize ears can facilitate subsequent infections by F. verticillioides.

Toxicity of several δ-endotoxins of Bacillus thuringiensis against the cotton pest Earias insulana (Lepidoptera: Noctuidae)

The toxicity of nine Bacillus thuringiensis Cry proteins against neonate Earias insulana larvae was tested using a mixture of crystals and spores. The mean lethal concentration (LC 50) of Cry1Ac was 1.99 μg/ml. Cry1Fa, Cry1Ca, Cry1Ja and Cry2Aa were more active than Cry1Ac, with LC 50 values of 0.22, 0.24, 0.29, 0.43 μg/ml, respectively. Cry1Da and Cry1Aa were considerably less active than Cry1Ac. The remaining proteins, Cry1Ba and Cry1Ab, displayed no activity. Relative potencies were also calculated. Cry1Ja and Cry1Fa were significantly more active (7.72 and 5.71 times, respectively) than Cry1Ac, while Cry1Ca was significantly (1.95 times) more active than Cry2Aa.

Effects of immunostimulatory CpG ODN on experimental allergic conjunctivitis caused by aspergillus fumigatus

Background Researches demonstrated that CpG ODN,a immunostimulatory sequences,has preventing and treating effect on allergic conjunctivitis caused by protein allergen.However,its effect on allergic conjunctivitis caused by fungal allergen is unclear. Objective This study aimed to investigate into whether the Th1-Th2 switching immunostimulatory CpG ODN could reverse the response in the murine allergic conjunctivitis model caused by aspergillus fumigatus. Methods A mixture of spores and hyphae of aspergillus fumigatus strain was used to induce allergic conjunctivitis in male BALB/C mice aged 6-8 weeks.This experiment was designed into preventive or therapeutical treatment program.Under both settings,allergic conjunctivitis of the animals were treated with CpG ODN,nonstimulatory GpC ODN or PBS.After the last challenge with the allergen,the clinical symptoms of the animals were scored based on the criteria of Magone.The animals were sacrificed and the histopathological examination of conjunctiva was performed.Expression of TLR4 mRNA in conjunctiva was analyzed by real-time PCR assay.The responsiveness and populations of lymphocytes in spleen and draining lymph nodes were analyzed by flow cytometry.The use complied with the Standard of Association for Research in Vision and Ophthalmology. Results In the prevention mode.CpG ODN decreased subconjunctival infiltration compared with GpC ODN and PBS groups with the average neutrophil count index(21.25 ±11.59/section,30.75 ±11.44 section and 69.00±9.90/section,respectively).Expression of TLR4 mRNA was up-regulated significantly by CpG ODN.The clinical scores for CpG ODN group were insignificantly lower than those in GpC ODN group and PBS group(P＞0.05).In the therapeutic mode,compared with GpC ODN and PBS groups,the allergic symptom score in CpG ODN group manifested significantly lower(t=4.000.t=2.750,P＜0.01)and showed fewer cellular infiltration(t=4.870,t=3.829,P＜0.01)and higher expression of TLR4 mRNA(P＜0.01).In cultured splenic and draining lymph node cells,increased percentages of CD4+ CD25+ and CD4+ CD25+ CD69+ in CpG ODN group were observed compared with control groups(|P＜0.05). Conclusion CpG ODN can relieve aspergillus fumigatus-induced allergic conjunctivitis via either subconjunctival injection or topical application by upregulating expression of TLR4 and activating Treg lymphocytes. Key words: Aspergillus fumigatus; Allergic conjunctivitis; CpG ODN; T-Lymphoeytes; Regulatory

Biological Control of Hemp sesbania (Sesbania exaltata) and Sicklepod (Senna obtusifolia) in Soybean with Anthracnose Pathogen Mixtures

In greenhouse and field experiments conducted over 3 yr, tank mixtures of spores of the fungiColletotrichum truncatumandColletotrichum gloeosporioides, formulated in unrefined corn oil and Silwet L-77 surfactant, were evaluated as a mycoherbicide mixture for simultaneous control of hemp sesbania and sicklepod, respectively. In greenhouse tests, 100% mortality and dry weight reduction of hemp sesbania occurred 6 d after treatment (DAT), whereas 15 d were required to achieve 100% mortality and dry weight reduction of sicklepod. In field experiments conducted in narrow-row (51-cm) soybean test plots, a single application of the fungal mixture formulated as described controlled hemp sesbania and sicklepod 94% and 88%, respectively, 28 DAT. Neither fungus, applied as aqueous conidial suspensions, provided control of their respective weed hosts. These results indicate that tank mixtures of these anthracnose-forming pathogens can effectively control hemp sesbania and sicklepod with a single application.

Bacterial mixture identification using Raman and surface‐enhanced Raman chemical imaging

AbstractThe ability of normal Raman and surface‐enhanced Raman scattering (SERS) to identify and detect bacteria has shown great success in recent studies. The addition of silver nanoparticles to bacterial samples not only results in an enhanced Raman signal, but it also suppresses the native fluorescence associated with biological material. In this report, Raman chemical imaging (RCI) was used to analyze individual bacteria and complex mixtures of spores and vegetative cells. RCI uses every pixel or a binned pixel group (BPG) of the Raman camera as an independent Raman spectrograph, allowing collection of spatially resolved Raman spectra. The advantage of this technique resides primarily in the analysis of samples in complex backgrounds without the need for physically isolating or purifying the sample. Using a chemical imaging Raman microscope, we compare normal RCI to SERS‐assisted chemical imaging of mixtures of bacteria. In both cases, we are able to differentiate single bacterium in the Raman microscope's field of view, with a 60‐fold reduction in image acquisition time and a factor of 10 increase in the signal‐to‐noise ratio for SERS chemical imaging over normal RCI. Copyright © 2010 John Wiley & Sons, Ltd.

Ectomycorrhizal fungi as an alternative to the use of chemical fertilisers in nursery production of Pinus pinaster

Addition of fertilisers is a common practice in nursery production of conifer seedlings. The aim of this study was to evaluate whether ectomycorrhizal (ECM) fungi can be an alternative to the use of chemical fertilisers in the nursery production of Pinus pinaster. A greenhouse nursery experiment was conducted by inoculating seedlings obtained from seeds of P. pinaster plus trees with a range of compatible ECM fungi: (1) Thelephora terrestris, (2) Rhizopogon vulgaris, (3) a mixture of Pisolithus tinctorius and Scleroderma citrinum, and (4) a mixture of Suillus bovinus, Laccaria laccata and Lactarius deterrimus, using forest soil as substrate. Plant development was assessed at two levels of N-P-K fertiliser (0 or 600 mg/seedling). Inoculation with a mixture of mycelium from S. bovinus, L. laccata and L. deterrimus and with a mixture of spores of P. tinctorius and S. citrinum improved plant growth and nutrition, without the need of fertiliser. Results indicate that selected ECM fungi can be a beneficial biotechnological tool in nursery production of P. pinaster.

Incorporation of Bentazone with <i>Exserohilum rostratum</i> for Controlling <i>Cyperus iria</i>

Problem statement: Various methods are used to counter the population of Cyperus iria in rice fields and among the most common method are chemical approaches. Chemical control methods are preferred by rice farmers due to their efficacy, speed, ease of storage and affordable cost. The concern of chemical herbicides is on its effects on the environment. Less attention had been given to biological control methods. Limitations such as host specificity, formulations and storage requirements are some of the obstacles that have to be overcome before implementation. Although it is environmentally friendly, a total shift from chemical control methods to biological seems unreasonable. Therefore incorporation of biological control with reduced chemical herbicides is expected to produce a better and sustainable control of weeds. Approach: A study was conducted to determine the potential of using Exserohilum rostratum as a biocontrol agent (incorporated with chemical herbicide, bentazone) to control Cyperus iria. The biocontrol agent (E. rostratum) was confirmed to be pathogenic to C. iria by applying Koch’s postulates. Preliminary laboratory experiments were carried out to ensure the interactions within the mixture was not inhibitory. Results: Bentazone was confirmed to be compatible with E. rostratum with an ED50 value of 22064570.0 µgai mL-1. In order to determine the efficacy of the incorporations, several mixtures were tested in a greenhouse environment. It was found that the mixtures of E. rostratum (106 spores mL-1) with bentazone (0.4 kgai ha-1) was significantly a better control of Cyperus iria compared to the usage of bentazone alone (0.4 kgai ha-1) and spores alone (106 spores mL-1). It was significantly more effective than the mixture of RBD palm oil (10%), Tween 80 (5%) and spores (106 spores mL-1) which acted as the conventional bioherbicide approach. Synergistic effects were exhibited with 50% less herbicide concentrations from the recommended rates. The mixture exhibited 50% disease severity in 3.5 days compare to bentazone alone which requires 4.9 days and the conventional bioherbicide mixture was relatively less effective requiring 4.7 days. The mixture of spores alone required 7.6 days to achieve 50% disease severity. This suggests that the mixture can save 50% of the cost on bentazone while providing a faster control. Conclusion: The incorporations were confirmed to be successful in providing a better and environmentally friendlier method than the conventional methods of weed control.

Generation characteristics of fungal spore and fragment bioaerosols by airflow control over fungal cultures

We investigated the generation characteristics of airborne fungal fragments and spores by means of a newly developed fungal bioaerosol generation system that directs airflow to the fungal cultures horizontally. By controlling the airflow over the fungal cultures inside the generation system, we can produce three aerosolization modes of fungal bioaerosols: (i) fungal spores only, (ii) fungal fragments only, and (iii) a mixture of fragments and spores. As concerns over fungal fragments and spores have grown due to the adverse health effects of fungal bioaerosols, this generation study can be applied to develop techniques for controlling airborne fungal particles through means such as antifungal material tests, fungal bioaerosol filtering or sampling tests, and fungal bioaerosol inactivation tests. In addition, the generation system presented here simulates the environmental dispersal of fungal bioaerosols. Therefore, the results of this study can be interpreted as equivalent to the natural dispersal characteristics of fungal bioaerosols.

Ectomycorrhizal fungi as an alternative to the use of chemical fertilisers in nursery production of Pinus pinaster

Addition of fertilisers is a common practice in nursery production of conifer seedlings. The aim of this study was to evaluate whether ectomycorrhizal (ECM) fungi can be an alternative to the use of chemical fertilisers in the nursery production of Pinus pinaster. A greenhouse nursery experiment was conducted by inoculating seedlings obtained from seeds of P. pinaster plus trees with a range of compatible ECM fungi: (1) Thelephora terrestris, (2) Rhizopogon vulgaris, (3) a mixture of Pisolithus tinctorius and Scleroderma citrinum, and (4) a mixture of Suillus bovinus, Laccaria laccata and Lactarius deterrimus, using forest soil as substrate. Plant development was assessed at two levels of N-P-K fertiliser (0 or 600 mg/seedling). Inoculation with a mixture of mycelium from S. bovinus, L. laccata and L. deterrimus and with a mixture of spores of P. tinctorius and S. citrinum improved plant growth and nutrition, without the need of fertiliser. Results indicate that selected ECM fungi can be a beneficial biotechnological tool in nursery production of P. pinaster.

Discrimination between Bacillus Species by Impedance Analysis of Individual Dielectrophoretically Positioned Spores

We combine the use of dielectrophoretic positioning with electrical impedance measurements to detect and discriminate between individual bacterial spores on the basis of their electrical response. Using lithographically defined microelectrodes, we use dielectrophoresis to manipulate individual bacterial spores between the electrodes. The introduction of a single spore between the microelectrodes produces a significant change in electrical response that is species-dependent. When positioned between two electrodes and an AC voltage was applied, single spores caused current increases averaging 6.8 (+/-2.4) pA for Bacillus mycoides to 1.18 (+/-0.37) pA for Bacillus licheniformis. Using a mixture of spores of two different species, we demonstrate the ability to distinguish the species of individual spores in real time. This work demonstrates the feasibility of using impedance measurements for real-time detection and discrimination between different types of spores.

Inactivation of food poisoning bacteria and Geobacillus stearothermophilus spores by high pressure carbon dioxide treatment

Inactivation of the vegetative foodborne pathogens by high pressure carbon dioxide treatment (HPCT) was investigated. Pseudomonas aeruginosa IFO3445, Aeromonas hydrophila IFO13286, Salmonella enteritidis JCM3313, Salmonella typhimurium IID1000, Yersinia enterocolitica JCM1677, Escherichia coli O157:H7 ATCC35150, E. coli O157:H7 ATCC43889, Staphylococcus aureus FDA209P and Listeria monocytogenes 1/2a were used in this study. In all cases, HPCT at 35 °C, 10 MPa for 1 min reduced initial numbers by approximately 6 logs. In addition HPCT inactivation of 9 pathogenic strains of Bacillus cereus (mixture of spores and vegetative cells) was investigated. HPCT at 75 °C, 10 MPa for 120 min resulted in significant inactivation in all the strains. The effect of HPCT (30 MPa at 95 °C for up to 120 min) on the inactivation of Geobacillus stearothermophilus spores in the presence of sodium chloride (3% and 6% w/v), glucose (6% and 12% w/v) and ethanol (10% and 20% v/v) was also studied. Both sodium chloride and glucose had a protective effect and the level of inactivation was reduced. The effect was in proportion to the solute concentration. However, 10% and 20% ethanol did not significantly affect the level of inactivation.

Waterborne Pathogen Detection Using Raman Spectroscopy

Raman spectroscopy is being evaluated as a candidate technology for waterborne pathogen detection. We have investigated the impact of key experimental and background interference parameters on the bacterial species level identification performance of Raman detection. These parameters include laser-induced photodamage threshold, composition of water matrix, and organism aging in water. The laser-induced photodamage may be minimized by operating a 532 nm continuous wave laser excitation at laser power densities below 2300 W/cm(2) for Grampositive Bacillus atrophaeus (formerly Bacillus globigii, BG) vegetative cells, 2800 W/cm(2) for BG spores, and 3500 W/cm(2) for Gram-negative E. coli (EC) organisms. In general, Bacillus spore microorganism preparations may be irradiated with higher laser power densities than the equivalent Bacillus vegetative preparations. In order to evaluate the impact of background interference and organism aging, we selected a biomaterials set comprising Gram-positive (anthrax simulants) organisms, Gram-negative (plague simulant) organisms, and proteins (toxin simulants) and constructed a Raman signature classifier that identifies at the species level. Subsequently, we evaluated the impact of tap water and storage time in water (aging) on the classifier performance when characterizing B. thuringiensis spores, BG spores, and EC cell preparations. In general, the measured Raman signatures of biological organisms exhibited minimal spectral variability with respect to the age of a resting suspension and water matrix composition. The observed signature variability did not substantially degrade discrimination performance at the genus and species levels. In addition, Raman chemical imaging spectroscopy was used to distinguish a mixture of BG spores and EC cells at the single cell level.