Bacterial endotoxins and microorganisms in the oral cavities of patients on cancer therapy.

OBJECTIVES: This study investigated the presence of Streptococci, Staphylococci, aerobic gram negative bacteria (AGNB), Candida and bacterial endotoxins in the oral cavities of patients receiving chemo- and/or radiotherapy for cancer. METHODS: Samples of oral cavity rinse were collected from 100 patients on cancer treatment and 70 healthy individuals. Demographic and clinical data were recorded. Samples were cultured onto various agar plates for qualitative and quantitative analysis and tested for the presence of endotoxin. Results were analysed using the Mann-Whitney and chi-square tests. RESULTS: In cancer patients, S. aureus counts were high and 66.7% of patients on chemo- and radiotherapy carried these bacteria (p=<0.05). The Candida carrier rate was significantly (p < 0.01) high in cancer patients (54%). No significant difference was found in the carrier rate of Streptococci and AGNB between the healthy and cancer group as well as between the cancer patients with chemo and radio- and chemotherapy alone. No significant difference was found in the level of endotoxin between the cancer patients and healthy individuals, and cancer patients with and without AGNB. CONCLUSIONS: No differences in the prevalence of bacteria and bacterial endotoxins were found between the cancer patients and healthy individuals. Oral cavity endotoxins did not correlate with the carriage of AGNB. However, due to the high prevalence in cancer patients, the role of Candida species and S. aureus in the pathology may not be excluded.

THE EFFECT OF COMBINED ACTION OF ANTIBACTERIAL DRUGS WITH LOW-INTENSIVE LASER RADIATION ON CLINICAL STRAINS S. AUREUS AND S. SALIVARIUS IN THE ORAL CAVITY.

The oral mucosa is constantly contaminated by a large number of microorganisms that may cause diseases such as periodontitis and caries. The present paper aims to study the effectiveness of the antimicrobial effect of combined use of antibacterial drugs (AD) and low-intensity laser radiation (LLR) on S. aureus S. salivarius isolated from the oral cavity. The study included 20 individuals with dental caries, 20 individuals with periodontitis and 10 without any signs of dental disease. The material for the microbacterial study was collected from surfaces of the teeth, oral cavity with dental caries and periodontal pockets. The intensity of bacterial isolation was estimated by two factors: the frequency of isolation and percentage of other aerobic microorganisms. The obtained data demonstrated that the use of several antibacterial drugs had a different impact on the strains of S. salivarius and S. aureus, depending on the source of their collection. The collected isolates were used to determine the effect of a 5 minute laser radiation combined with antibacterial drugs. The simultaneous use of antibacterial therapy and laser radiation showed an increase in the therapeutic effect of all investigated antibiotics followed by the inhibition of the growth presentations in S. aureus and S. salivarius. The application of photodynamic therapy, e.g. LLR, combined with antibacterial drugs allowed to achieve a complete inhibition of the microbial growth.

Inactivation of Salmonella spp., pathogenic Escherichia coli, Staphylococcus spp., or Listeria monocytogenes in chicken purge or skin using a 405-nm LED array.

Raw poultry are sometimes contaminated with foodborne pathogens, which can lead to illness in humans. In recent years research has focused on a variety of light technologies to decontaminate food and food contact surfaces during meat and poultry processing. In this study we evaluated the ability of 405-nm light generated from an LED array to inactivate multi-isolate cocktails of either Salmonella spp., pathogenic Escherichia coli, Staphylococcus spp., or Listeria monocytogenes suspended in chicken purge or skin. When exposed to 180 J/cm2 405-nm light at two separate light intensities (300 mW/cm2/s or 150 mW/cm2/s) the maximum pathogen reduction on chicken skin was ca. 0.4 log. When the pathogens were suspended in chicken purge the maximum log reductions ranged from 0.23 to 0.68 log (180 J/cm2; 150 mW/cm2/s) versus 0.69 to 1.01 log (180 J/cm2; 300 mW/cm2/s). Log reductions of each pathogen, when they were subjected to heat shock prior to 405-nm light treatment, were reduced, indicating that thermal effects accounted for much of the bacterial inactivation.

In vitro bactericidal activity of blue light (465 nm) phototherapy on meticillin-susceptible and meticillin-resistant Staphylococcus pseudintermedius.

BACKGROUND: Staphylococcus pseudintermedius is the most common cause of bacterial skin infections in dogs. Meticillin-resistant infections have become more common and are challenging to treat. Blue light phototherapy may be an option for treating these infections. HYPOTHESIS/OBJECTIVES: The objective of this study was to measure the in vitro bactericidal activity of 465 nm blue light on meticillin-susceptible Staphylococcus pseudintermedius (MSSP) and meticillin-resistant Staphylococcus pseudintermedius (MRSP). We hypothesized that irradiation with blue light would kill MSSP and MRSP in a dose-dependent fashion in vitro as previously reported for meticillin-resistant Staphylococcus aureus (MRSA). METHODS: In six replicate experiments, each strain [MSSP, n = 1; MRSP ST-71 (KM1381) n = 1; and MRSA (BAA-1680) n = 1] were cultivated on semisolid media, irradiated using a 465 nm blue light phototherapeutic device at the cumulative doses of 56.25, 112.5 and 225 J/cm2 and incubated overnight at 35°C. Controls were not irradiated. Colony counts (CC) were performed manually. Descriptive statistics were performed and treatment effects assessed using the Wilcoxon-Mann-Whitney rank-sum test. Bonferroni-corrected rank-sum tests were performed for post hoc analysis when significant differences were identified. RESULTS: There was a significant decrease in CC with blue light irradiation at all doses for MRSA (P = 0.0006) but not for MSSP (P = 0.131) or MRSP (P = 0.589). CONCLUSIONS: Blue light phototherapy significantly reduced CC of MRSA, but not of MSSP or MRSP. The mechanism for the relative photosensitivity of the MRSA isolate is unknown, but is hypothesized to be due to an increased concentration of porphyrin in S. aureus relative to S. pseudintermedius, which would modulate blue light absorption.

Reduction of bacteria and human immunodeficiency virus Type 1 infectivity of platelet suspension in plasma using xenon flash-pulse light in a bench-scale trial.

BACKGROUND: Current pathogen reduction systems for platelet concentrates (PCs) require addition of chemical compounds and/or reduction of plasma content in PCs. We have investigated a new method using xenon (Xe) flash-pulse light without additional compounds or plasma replacement. STUDY DESIGN AND METHODS: An aliquot of apheresis platelets (PLTs) in plasma inoculated with bacteria or human immunodeficiency virus Type 1 (HIV-1) was irradiated with Xe flash-pulse light (Xe flash phototreatment). Bacterial growth was monitored up to 6 days of storage, whereas HIV-1 infectivity was assayed just after treatment. Pairs of Xe flash-phototreated and untreated PCs were examined for PLT lesion during the storage period. RESULTS: Under the current conditions, a low titer (1.8 colony-forming units [CFUs]/mL) of Staphylococcus aureus did not proliferate during the 6-day storage period, but grew in some cases at high-titer (24.0 CFUs/mL) inoculation. HIV-1 infectivity was reduced by 1.8 log. PLT recovery of the treated PCs was lower than untreated ones. An increase of mean PLT volume and glucose consumption, together with a decrease of hypotonic shock response and pH, were enhanced by the treatment. CD62P- and PAC-1-positive PLTs increased after the treatment, indicating the induction of PLT activation. Among biologic response modifiers, soluble CD40 ligand was significantly increased in the treated PCs on Day 6. CONCLUSIONS: Xe flash phototreatment could prevent bacterial proliferation and reduce HIV-1 infectivity in 100% plasma PCs without any additional compounds, but enhanced PLT storage lesions. Further improvement is required to increase the potency of pathogen inactivation with reducing PLT damage.

Photodynamic oxidation of Staphylococcus warneri membrane phospholipids: new insights based on lipidomics.

RATIONALE: The photodynamic process involves the combined use of light and a photosensitizer, which, in the presence of oxygen, originates cytotoxic species capable of oxidizing biological molecules, such as lipids. However, the effect of the photodynamic process in the bacterial phospholipid profile by a photosensitizer has never been reported. A lipidomic approach was used to study the photodynamic oxidation of membrane phospholipids of Staphylococcus warneri by a tricationic porphyrin [5,10,15-tris(1-methylpyridinium-4-yl)-20-(pentafluorophenyl)porphyrin triiodide, Tri-Py(+)-Me-PF]. METHODS: S. warneri (10(8) colony forming units mL(-1)) was irradiated with white light (4 mW cm(-2), 21.6 J cm(-2)) in the presence of Tri-Py(+)-Me-PF (5.0 µM). Non-photosensitized bacteria were used as control (irradiated without porphyrin). After irradiation, total lipids were extracted and separated by thin-layer chromatography (TLC). Isolated fractions of lipid classes were quantified by phosphorus assay and analyzed by mass spectrometry (MS): off-line TLC/ESI-MS, hydrophilic interaction (HILIC)-LC/MS and MS/MS. RESULTS: The most representative classes of S. warneri phospholipids were identified as phosphatidylglycerols (PGs) and cardiolipins (CLs). Lysyl-phosphatidylglycerols (LPGs), phosphatidylethanolamines (PEs), phosphatidylcholines (PCs) and phosphatidic acids (PAs) were also identified. After photodynamic treatment, an overall increase in the relative abundance of PGs was observed as well as the appearance of new oxidized species from CLs, including hydroxy and hydroperoxy derivatives. Formation of high amounts of lipid hydroperoxides was confirmed by FOX2 assay. Photodynamic oxidation of phospholipid standards revealed the formation of hydroperoxy and dihydroperoxy derivatives, confirming the observed CL oxidized species in S. warneri. CONCLUSIONS: Membrane phospholipids of S. warneri are molecular targets of the photoinactivation process induced by Tri-Py(+) -Me-PF. The overall modification in the relative amount of phospholipids and the formation of lipid hydroxides and hydroperoxides indicate the lethal damage caused to photosensitized bacterial cells.

Effects of ultraviolet light and ultrasound on microbial quality and aroma-active components of milk.

BACKGROUND: Heat treatment is the most common way to extend the shelf life of milk. However, alternative technologies such as ultraviolet (UV) light and ultrasound (US), which are non-thermal methods for processing milk, have been developed to replace heat treatment. These technologies do not have any adverse effects on the quality of milk. The major purpose of this study was to investigate the effects of UV and US on different micro-organism groups and aroma compounds in milk. RESULTS: Heat pasteurisation at 65 °C for 30 min was used as thermal control treatment. The growth of total coliform group bacteria, Escherichia coli and Staphylococcus spp. was completely reduced by UV treatment. Application of US was not sufficient to reduce the numbers of yeasts and moulds. In neutral/basic fractions, 3-methylthiophene (plastic), hexanal (grass) and 1-hexen-3-one (floral) were major volatiles in milk samples. CONCLUSION: UV had a major effect on total coliforms, E. coli and Staphylococcus spp., but US was not as effective as UV and heat treatment in reducing certain groups of micro-organisms. No major differences were observed in terms of aroma-active compounds and flavour of milk following the different treatments. However, some new volatiles and change in rheological properties were generated by UV and US treatments.

UVc-irradiation sublethal stress does not alter antibiotic susceptibility of staphylococci (MRSA, MSSA, and coagulase-negative staphylococci) to ß-lactam, macrolide, and fluoroquinolone antibiotic agents.

Skin tanning, either by exposure to natural sunlight or through use of UV sunbeds, has become a popular practice in the US, where it is estimated that approximately 1 million times per day someone in the US uses UV radiation for skin tanning, equating to 30 million Americans (circa 10% of the US population) who use a tanning bed. As well as exposing the host to periods of UV radiation, such practices also expose commensal skin bacteria, including Staphylococcus aureus, to such UV radiation. Previous work has indicated that environmental stresses on bacteria may lead to an upregulation of stress responses, in an attempt for the organism to combat the applied stress and remain viable. UV light may act as an environmental stress on bacteria, and so it was the aim of this study to examine the effect of UVc light on the antibiotic susceptibility of commensal skin bacteria, to determine if UV radiation would increase the antibiotic resistance of such skin flora and thus lead to a potential skin flora with increased antibiotic resistance. Previously, it has been shown that UVc light has a greater mutational effect on bacteria compared to lower-energy UV forms, including UVa and UVb light. Therefore, we decided to employ UVc light in our study to amplify the potential for mutational events occurring in skin staphylococci organisms (n=8) including methicillin-sensitive Staphylococcus aureus (n=2), methicillin-resistant Staphylococcus aureus (n=4), and coagulase-negative staphylococci (Staphylococcus haemolyticus) (n=2) were exposed to varying degrees of sublethal radiation via UVc light, and their minimum inhibitory concentration (MIC) susceptibility was determined by broth dilution assay against three classes of commonly used antibiotics, namely ß-lactams (penicillin), macrolides (erythromycin), and fluoroquinolones (ciprofloxacin). There was no significant difference between antibiotic susceptibility before UVc exposure and until maximum sublethal stress, prior to cell death due to fatal UVc exposure with the cells. These results indicate that UV environmental stress/exposure does not upregulate antibiotic resistance, and therefore these data indicate that UVc radiation does not lead to a more antibiotic-resistant population in the staphylococci organisms post-exposure.

Effects of combined treatment of sodium hypochlorite/ionizing radiation and addition of vitamin B(1) on microbial flora of oyster and short-necked clam.

To determine the synergistic disinfection effect of the combined treatments of sodium hypochlorite (NaClO), irradiation, and vitamin B(1), the bactericidal effects of the treatments on natural microflora of oyster and short-necked clam were investigated. Then, bacteria isolated from the samples were identified by 16S rDNA sequencing. Oyster and short-necked clam were mainly contaminated with Vibrio spp. and Bacillus spp. Total number of aerobic bacteria ranged from 10(2) to 10(4) colony forming units (CFU)/g initially. More than 100 mg/L of NaClO with 1000 mg/L vitamin B(1) and 2 kGy irradiation treatment for oyster and short-necked clam can reduce the total aerobic bacteria to the level of lower than a detection limit (10 CFU/g). Synergistic effects were observed for all combined treatment against natural microflora. The results suggest that a significant synergistic benefit can be achieved by a combination of NaClO-ionizing radiation treatment with the addition of vitamin B(1) to reduce the microbial population contaminated in oyster and short-necked clam.

In vitro antibacterial effects of non-thermal atmospheric plasma irradiation on Staphylococcus pseudintermedius and Pseudomonas aeruginosa.

In the last decade, atmospheric plasma has been used to treating bacterial infections in humans due to its bactericidal effects; however, its efficacy in dogs is unclear. This study evaluated the in vitro bactericidal efficacy of atmospheric plasma on Staphylococcus pseudinter- medius and Pseudomonas aeruginosa, two of the most important bacterial agents isolated from canine pyodermas. Three isolates each of S. pseudintermedius and P. aeruginosa obtained from dogs with pyoderma were subjected to atmospheric plasma. The isolates from the control group were not exposed to plasma, while those from the treatment groups were exposed to plasma for 15 (7.5 J/cm2), 30 (15 J/cm2), 60 (30 J/cm2), or 90 (45 J/cm2) seconds. After each treatment, a reduction in colony formation was observed. Bacterial viability was evaluated using the LIVE/ DEAD® BacLight™ Bacterial Viability Kit. The antibacterial effects were evaluated with Image J software and significance was assessed statistically in comparison to the control group. The bactericidal effect of atmospheric plasma against both bacteria increased significantly in a time-dependent manner. These results demonstrate the bactericidal capacity of atmospheric plasma, and suggest that it could serve as an alternative treatment method for canine pyoderma. Further studies are needed to evaluate the safety and efficacy of atmospheric plasma in dogs.

Photoinactivation Sensitivity of Staphylococcus carnosus to Visible-light Irradiation as a Function of Wavelength.

Inactivation properties of visible light are of increasing interest due to multiple possible fields of application concerning antibacterial treatment. For violet wavelengths, the generation of reactive oxygen species by porphyrins is accepted as underlying mechanism. However, there is still little knowledge about photosensitizers at blue wavelengths. While flavins were named as possible candidates, there is still no experimental evidence. This study investigates the photoinactivation sensitivity of Staphylococcus carnosus to selected wavelengths between 390 and 500 nm in 10- to 25-nm intervals. Absorption and fluorescence measurements in bacterial lysates confirmed inactivation findings. By means of a mathematical calculation in MATLAB® , a fit of different photosensitizer absorption spectra to the measured action spectrum was determined to gain knowledge about the extent to which specific photosensitizers are involved. The most effective wavelength for S. carnosus at 415 nm could be explained by the involvement of zinc protoporphyrin IX. Between 450 and 470 nm, inactivation results indicated a broad plateau, statistically distinguishable from 440 and 480 nm. This observation points to flavins as responsible photosensitizers, which furthermore seem to be involved at violet wavelengths. A spectral scan of sensitivities might generally be an advantageous approach for examining irradiation impact.

Isolation of UV-B resistant bacteria from two high altitude Andean lakes (4,400 m) with saline and non saline conditions.

Laguna (L.) Negra and L. Verde are high altitude Andean lakes located at the 4,400 m altitude in the Andean desert (Puna) in the Argentine northwest. Both lakes are exposed to extreme weather conditions but differ in salinity contents (salinity 6.7% for L. Negra and 0.27% for L. Verde). The aim of this work was to isolate ultraviolet B fraction (UV-B) resistant bacteria under UV-stress in order to determine, a possible connection, between resistance to UV-B and tolerance to salinity. DNA damage was determined by measuring CPDs accumulation. Connection among pigmentation production and UV resistance was also studied. Water samples were exposed to artificial UV-B radiation for 24 h. Water aliquots were plated along the exposition on different media, with different salinity and carbon source content (Lake medium (LM) done with the lake water plus agar and LB). CFU were counted and DNA damage accumulation was determined. Isolated bacteria were identified by 16S rDNA sequence. Their salinity tolerance, were measured at 1, 5 and 10% NaCl and their pigment production in both media was determined. In general it was found that UV resistance and pigment production were the optimum in Lake Medium done with lake water which maintained similar salinity. The most resistant bacteria in L. Negra were different strains of Exiguobacterium sp. and, in L. Verde, Staphylococcus sp. and Stenotrophomonas maltophilia. These bacteria showed the production and increase of UV-Vis absorbing compounds under UV stress and in LM. Bacterial communities from both lakes were well adapted to high UV-B exposure under the experimental conditions, and in many cases UV-B even stimulated growth. The idea that resistance to UV-B could be related to adaptation to high salinity is still an open question that has to be answered with future experiments.

Leu169Trp substitution in MnSOD from Staphylococcus equorum created an active new form of similar resistance to UVC irradiation.

Manganese superoxide dismutase from Staphylococcus equorum (MnSODSeq) maintains its activity after up to 45 minutes of UVC radiation. The enzyme occurs in a dimeric form that likely contributes to its activity and stability. Therefore, maintaining the dimeric form could be a way to improve the enzyme's stability. One of the main interactions for dimer formation occurs between Tyr168 and His31, of which the latter is also involved in the enzymatic reaction. UVC radiation may cause alterations in the electronic structure of the phenolic ring in the Tyr168 side chain: this may disrupt the Tyr168-His31 pairing and lead to enzyme instability and/or activity loss. In this report, a Leu169Trp substitution was carried out to protect the Tyr168 residue by introducing an amino acid with an aromatic side chain for better photon absorption of the UV light. Interestingly, although the substitution appeared to have a minor effect on enzyme stability and activity upon UVC irradiation, the melting temperature (TM) of the Leu169Trp mutant was different. Unlike the native protein, the TM of the mutant had not changed after UV irradiation. Thus, our effort to extend the resistance to UVC radiation was not successful, but we have discovered a biologically active new form. The present finding provides evidence that MnSODSeq maintains most of its activity and resistance to UVC irradiation as long as the dimer and its glutamate-bridge are intact, despite an alteration that destabilizes its monomeric structure. The present finding further unravels the relationship between the structure of the enzyme and its activity. Furthermore, the results may provide further insight in how to modify the enzyme to improve its characteristics for application in medicine or cosmetics.

Antimicrobial efficacy of irradiation with visible light on oral bacteria in vitro: a systematic review.

AIM: Resistances to antibiotics employed for treatment of infectious diseases have increased to alarming numbers making it more and more difficult to treat diseases caused by microorganisms resistant to common antibiotics. Consequently, novel methods for successful inactivation of pathogens are required. In this instance, one alternative could be application of light for treatment of topical infections. Antimicrobial properties of UV light are well documented, but due to its DNA-damaging properties use for medical purposes is limited. In contrast, irradiation with visible light may be more promising. METHODS: Literature was systematically screened for research concerning inactivation of main oral bacterial species by means of visible light. RESULTS: Inactivation of bacterial species, especially pigmented ones, in planktonic state showed promising results. There is a lack of research examining the situation when organized as biofilms. CONCLUSION: More research concerning situation in a biofilm state is required.

Improved contact lens disinfection by exposure to violet radiation.

BACKGROUND: Conventional procedures for contact lens disinfection, based on solutions with aggressive chemical ingredients, not only affect microorganisms but operate likewise damaging towards the epithelial eye surface. OBJECTIVE: The aim of this study was to evaluate the applicability of an alternative or complementary disinfection procedure for contact lenses based on irradiation within the visible wavelength range. METHODS: Suspensions of S. auricularis, B. subtilis and E. coli were exposed to 405 nm irradiation, for determining the disinfection efficacy. Surviving rates were analyzed by membrane filtration as well as a semi-quantitative analysis using DipSlides. RESULTS: A significant antibacterial effect of the 405 nm irradiation is verifiable for all probed bacteria. Using S. auricularis, there has been no colony forming after an irradiation exposure of 2 hours. CONCLUSION: The hitherto existing results give reason for the assumption that violet LEDs integrated in contact lens cases will provide a subsidiary disinfection activity and maybe even offer the reduction of chemical ingredients in lens cleaning solutions to become gentler to the eye. In addition the danger of a rerise of the germ concentration after the completion of the disinfection procedure will be reduced.

Overall biochemical changes in bacteria photosensitized with cationic porphyrins monitored by infrared spectroscopy.

BACKGROUND: Photodynamic inactivation of micro-organisms is a promising nonantibiotic multitarget approach to treat localized and superficial infections through oxidative stress. Herein, the changes occurring on major cellular components of Escherichia coli and Staphylococcus warneri, induced by photosensitization with cationic porphyrins (Tri-Py(+)-Me-PF and Tetra-Py(+)-Me) and white light, were monitored by infrared spectroscopy. RESULTS: In E. coli, most of the changes occurred on proteins and lipids, suggesting a key effect on lipopolysaccharides in the first irradiation times. In S. warneri, proteins were the major molecular targets of oxidative damage but phospholipids and polysaccharides were also affected. CONCLUSION: Infrared spectroscopy is a very interesting tool to monitor biochemical changes induced by photosensitization in bacteria and also to infer on its mechanism of action.

Genetic control of chromosomal and plasmid recombination in Staphylococcus aureus.

Recombination-deficient mutants of Staphylococcus aureus have been isolated and found to have properties similar to those of recombination-deficient Escherichia coli. In addition, one Rec(-) mutant was found to be defective in the restriction and modification of DNA. There is a marked reduction ( approximately 10(4)-fold) in recombination between penicillinase plasmids in the Rec(-) mutants suggesting that these elements do not encode an efficient recombination system. There is, however, a demonstrable residuum of interplasmid recombination; evidence is lacking on whether this residuum is a plasmid or host function. In the absence of the generalized host recombination system it has been possible to demonstrate that interplasmid recombination occurs during vegetative bacteriophage growth and is presumably mediated by a phage-determined recombination system.

Effect of low-level laser therapy on the healing of second-degree burns in rats: a histological and microbiological study.

This paper presents the results of a study on the effects of two different doses of low-level laser therapy on healing of deep second-degree burns. Sixty rats were randomly allocated to one of four groups. A deep second-degree burn was inflicted in each rat. In the control group burns remained untreated; in two laser treated groups the burns were irradiated daily with low-level helium-neon laser with energy densities of 1.2 and 2.4 J/cm2, respectively. In the fourth group the burns were treated topically with 0.2% nitrofurazone cream every day. The response to treatments was assessed histologically at 7, 16 and 30 days after burning, and microbiologically at Day 15. The number of macrophages at day 16, and the depth of new epidermis at day 30, was significantly less in the laser treated groups in comparison with control and nitrofurazone treated groups (P=0.000). Staphylococcus epidermidis was found in the 70% of rat wounds in the laser treated groups in comparison with 100% of rats in the control group. S. aureus was found in the 40% rat wounds in the nitrofurazone treated group, but there was not found in the wounds of laser treated, and control groups. It is concluded that low-level laser therapy of deep second-degree burn caused significant decrease in the number of macrophage and depth of new epidermis. In addition, it decreased incidence of S. epidermidis and S. aureus.

Enhancing DNA electro-transformation efficiency on a clinical Staphylococcus capitis isolate.

Clinical staphylococcus isolates possess a stronger restriction-modification (RM) barrier than laboratory strains. Clinical isolates are therefore more resistant to acceptance of foreign genetic material than laboratory strains, as their restriction systems more readily recognize and destroy foreign DNA. This stronger barrier consequently restricts genetic studies to a small number of domestic strains that are capable of accepting foreign DNA. In this study, an isolate of Staphylococcus capitis, obtained from the blood of a very low birth-weight baby, was transformed with a shuttle vector, pBT2. Optimal conditions for electro-transformation were as follows: cells were harvested at mid-log phase, electro-competent cells were prepared; cells were pre-treated at 55°C for 1min; 3µg of plasmid DNA was mixed with 70-80µL of competent cells (3-4×10(10)cells/mL) at 20°C in 0.5M sucrose, 10% glycerol; and electroporation was conducted using 2.1kV/cm field strength with a 0.1cm gap. Compared to the conventional method, which involves DNA electroporation of Staphylococcus aureus RN4220 as an intermediate strain to overcome the restriction barrier, our proposed approach exhibits a higher level (3 log10 units) of transformation efficiency. Heat treatment was used to temporarily inactivate the recipient RM barrier. Other important parameters contributing to improved electro-transformation efficiency were growth stage for cell harvesting, the quantity of DNA, the transformation temperature and field strength. The approach described here may facilitate genetic manipulations of this opportunistic pathogen.

Bacterial growth rates are influenced by cellular characteristics of individual species when immersed in electromagnetic fields.

Previous studies have shown that exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) have negative effects on the rate of growth of bacteria. In the present study, two Gram-positive and two Gram-negative species were exposed to six magnetic field conditions in broth cultures. Three variations of the 'Thomas' pulsed frequency-modulated pattern; a strong-static "puck" magnet upwards of 5000G in intensity; a pair of these magnets rotating opposite one another at ∼30rpm; and finally a strong dynamic magnetic field generator termed the 'Resonator' with an average intensity of 250µT were used. Growth rate was discerned by optical density (OD) measurements every hour at 600nm. ELF-EMF conditions significantly affected the rates of growth of the bacterial cultures, while the two static magnetic field conditions were not statistically significant. Most interestingly, the 'Resonator' dynamic magnetic field increased the rates of growth of three species (Staphylococcus epidermidis, Staphylococcus aureus, and Escherichia coli), while slowing the growth of one (Serratia marcescens). We suggest that these effects are due to individual biophysical characteristics of the bacterial species.