Comparison of antibacterial activity in the hemocytes of different crustacean species

Antibacterial activity in hemocytes of the squat lobster, Galathea strigosa, the Norway lobster, Nephrops norvegicus, the common shrimp, Crangon crangon, and the giant Antarctic isopod, Glyptonotus antarcticus, was investigated in vitro. For all species, the marine bacterium, Psychrobacter immobilis, was used as the test organism, although with G. antarcticus, the Gram positive bacteria, Planococcus citreus and BS 68 (an isolate from Antarctic waters), were also used. Hemocyte lysate supernatants (HLS) from all four species reduced the viable count of test bacteria over a period of 4 hr showing that their hemocytes contain factors able to neutralize bacteria in vitro. However, comparison of responses produced by serially diluted samples of HLS from G. strigosa, N. norvegicus and C. crangon, revealed that activity (per unit protein) is weaker than for Carcinus maenas. Using G. antarcticus, positive activity was also observed against P. citreus and BS 68; with the response effective against all of the bacteria at both 0°C and 20°C. These results show that: (1) the hemocytes from a range of crustacean species contain factor(s) able to neutralize bacteria in vitro; (2) antibacterial potency varies from species to species; and (3) antibacterial immunity in at least one polar invertebrate functions at low temperature.     

Transformation of various species of gram-negative bacteria belonging to 11 different genera by electroporation

Summary We have undertaken a systematic study to test the transformation of various species of gram-negative bacteria using the electroporation method. The data obtained show very clearly that a great variety of gram-negative bacteria — 15 different species belonging to 11 different genera — including freshly isolated wild-type strains can be transformed efficiently by use of the electric-field mediated transformation technique. These include species of the families Enterobacteriaceae, Pseudomonadaceae, Rhizobiaceae, photosynthetic bacteria and strains for which transformation could not be achieved, up to now, by other methods.     

Evaluation of in vitro antibacterial activity of some disinfectants on Escherichia coli serotypes

Three disinfectants commonly used in poultry farms (formalin, TH4+, and Virkon-S) were chosen for the present study. The effect of disinfectant concentration and the duration of exposure to these disinfectants on the survival of Escherichia coli serotypes (O114:K-, O86, O55:K39, and O86:K60) were investigated. Formalin (0.6%), TH4+ (0.06%), and Virkon (0.5%) all killed the four serotypes within 5 min of exposure. As the disinfectant concentration decreases, the length of exposure time to kill serotype increases. At 0.03%, 0.007%, and 0.03% of formalin, TH4+ and Virkon-S concentrations failed to kill the four E. coli serotypes within 360 min, respectively. An improvement of the inhibitory effect of these disinfectants occurred when added together with the inoculum instead of an established population. The influence of formalin, TH4+, and Virkon-S on the cell morphology of E. coli O55:K39 was investigated by using transmission electron microscopy. Formalin-treated cells exhibited normal cell morphology, with the exception that the treated cell was less fimbriated, and more destruction of pili increased when formalin concentrations were doubled. Cells treated with TH4+ (0.03%) showed destruction of the cell wall and cell surface membrane after 5 min. Cell filamentation occurred at 0.015% and increased with the increase of exposure time to this drug. Spheroplasts were observed only when cells were treated with 0.125% Virkon-S for 60 min, and cell lysis started to occur when 0.25% Virkon-S was applied for 15 min. Scanning electron microscope study revealed that Virkon-S at 0.03% and TH4+ at 0.007% completely prevented the adherence of E. coli O55:K39 serotype to chicken tracheal organ, whereas formalin (0.03%) disinfection minimized the adherence of E. coli cells to tracheal explants after 360 min of incubation.     

In vitro activity of cefepime against ESBL-positive clinical strains of gram-negative rods

The aim of this study was to determine an in vitro activity of cefepime against ESBL-positive clinical strains of Gram-negative rods isolated from hospitalized patients. Experiments were performed with 100 ESBL-positive strains of Gram-negative rods isolated from clinical samples in 2004. Strains were identified with the use of automatic ATB Expression system and biochemical ID 32 GN tests (bioMdrieux sa). Extended-spectrum beta-lactamases (ESBLs) were detected by means of disc diffusion methods: the double-disc synergy test (DDST) and the diagnostic disc test (DD, Oxoid Ltd, UK). Susceptibility in vitro of ESBL producers to 4th generation--cefepime was determined with gradient diffusion method Etest (AB Biodisk, Solna, Sweden). MIC value of cefepime was assessed for each strain. Among 100 ESBL-producing strains, 94--belonged to enteric rods and 6--to nonfermentative rods. The greatest number of strains belonged to the species Serratia marcescens (27% of all strains) and next--to the species Enterobacter cloacae (21%). Fourteen strains were susceptible (S) in vitro to cefepime, 12--intermediately susceptible (I) and 74--resistant (R). Application of cefepime in a therapy of infections caused by ESBL-positive strains of Gram-negative rods highly susceptible in vitro to this antibiotic, should be considered.     

Evaluation of the specificity of Salmonella PCR primers using various intestinal bacterial species

AIMS: Nine sets of PCR primers targeting Salmonella were evaluated for their specificity with pure cultures of intestinal-associated bacteria prior to their application to Salmonella detection in faecal samples. METHODS AND RESULTS: Gene targets of PCR primers included: 16S rDNA, a Salmonella pathogenicity island I virulence gene, Salmonella enterotoxin gene (stn), invA gene, Fur-regulated gene, histidine transport operon, junction between SipB and SipC virulence genes, Salmonella-specific repetitive DNA fragment, and multiplex targeting invA gene and spvC gene of the virulence plasmid. Fifty-two Salmonella strains were used to determine sensitivity; five strains from related genera and 45 intestinal bacteria were used to evaluate specificity. All primers amplified DNA from Salmonella strains, although two primer sets failed to amplify Salmonella DNA from either Salmonella bongori (hilA) or subgroups VI or VII (16S rDNA). There was no detected amplification of DNA from related bacterial genera with any of nine PCR assays. Six of the PCR assays amplified DNA for some intestinal bacteria. CONCLUSIONS: Only three primer pairs were determined to be suitable for application of PCR amplification of Salmonella in faecal samples - 16S rDNA, stn and histidine transport operon. We are currently evaluating their sensitivity of detection of Salmonella in faecal samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the importance of internal lab validation of PCR primers prior to application to the type of samples of interest. Information from this evaluation can be applied in other labs to facilitate choosing Salmonella PCR primers.     

Chlorhexidine-silver sulfadiazine-impregnated central venous catheters: in vitro antibacterial activity and impact on bacterial adhesion

The aim of this study was to compare the in vitro activity and the impact on bacterial adhesion of two different catheters, one impregnated with chlorhexidine-silver sulfadiazine (C-SS) and the other not impregnated with antibacterial agents. The antimicrobial coating prevented the bacterial colonization by slime positive Staphylococcus epidermidis in the first two days. The antibacterial activity of the effluents from catheters impregnated with C-SS dissipated by day seven. Our results demonstrated that the surface treatment modified the composition of impregnated catheters and determined different contact angle values of the two catheters (impregnated and not impregnated). Examination of coated and uncoated catheter segments by scanning electron microscopy showed a good correlation with the results of adherence experiments. In conclusion, the findings suggest that C-SS coated catheters prevent in vitro bacterial adhesion.     

Evaluation of antibacterial activity of crude protein extracts from seeds of six different medical plants against standard bacterial strains

A huge group of natural antimicrobial compounds are active against a large spectrum of bacterial strains causing infectious threat. The present study was conducted to investigate the crude extracts of antimicrobial protein and peptide efficacy from six medicinal plant seeds. Extraction was carried out in Sodium phosphate citrate buffer, and Sodium acetate buffer using different pH. Antimicrobial activities of these plants were determined by the microbiological technique using Agar well diffusion Assay. Extremely strong activity was observed in the seed extracts of Allium ascolinicum extracted in sodium phosphate citrate buffer at pH (5.8) against Proteus vulgaris, Escherichia coli and Staphylococcus aureus with zone of inhibition 17 mm, 17 mm and 15 mm and Rumex vesicarius at pH (7.6), Ammi majus at pH (6.8), Cichorium intybus at pH (7.4) and Cucumis sativus at pH (7.8) also showed better sensitivity against the bacterial strains with zone of inhibition ranges 16–10 mm and some of the strains were found to be resistant. Antibacterial activity pattern of different plant extracts prepared in sodium acetate buffer pH (6.5), among all the plant seed extracts used Foeniculum vulgare had shown good inhibition in all the bacterial strains used, with zone of inhibition ranges 11–12.5 mm, The extracts of C. intybus and C. sativus were found to be effective with zone of inhibition 11–6 mm and some of the strains were found to be resistant. Most of the strains found to have shown better sensitivity compared with the standard antibiotic Chloramphenicol (25 mcg). Our results showed that the plants used for our study are the richest source for antimicrobial proteins and peptides and they may be used for industrial extraction and isolation of antimicrobial compounds which may find a place in medicine industry as constituents of antibiotics.     

Effect of titanium-ion on the growth of various bacterial species

There are a number of studies that explain the metabolism and roles of metallic titanium and titaniumion. One of the most intriguing results from these studies is the finding of metallic titanium having no bacteriostatic effects on oral bacterial species. In this research, the effects of titanium-ion on the growth of twenty-two bacterial species, some of which are commonly found in foods such as yoghurt, kimchi, and soy fermented products, were investigated. All but two bacteria, Escherichia coli and Pseudomonas aeruginosa appeared to be sensitive to titanium-ion. These two species were grown on 360 microg/ml of titanium-ions, and they were found to be resistant to the titanium-ion. Both the wild-type and plasmid-cured E. coli showed good growth in a medium with 200 microg/ml of titanium-ions. These results suggest that titanium-resistance was independent from the effects of the plasmid in E. coli.     

New derivatives of salicylamides: Preparation and antimicrobial activity against various bacterial species

Three series of salicylanilides, esters of N-phenylsalicylamides and 2-hydroxy-N-[1-(2-hydroxyphenylamino)-1-oxoalkan-2-yl]benzamides, in total thirty target compounds were synthesized and characterized. The compounds were evaluated against seven bacterial and three mycobacterial strains. The antimicrobial activities of some compounds were comparable or higher than the standards ampicillin, ciprofloxacin or isoniazid. Derivatives 3f demonstrated high biological activity against Staphylococcus aureus (?0.03 μmol/L), Mycobacterium marinum (?0.40 μmol/L) and Mycobacterium kansasii (1.58 μmol/L), 3g shows activity against Clostridium perfringens (?0.03 μmol/L) and Bacillus cereus (0.09 μmol/L), 3h against Pasteurella multocida (?0.03 μmol/L) and M. kansasii (?0.43 μmol/L), 3i against methicillin-resistant S. aureus and B. cereus (?0.03 μmol/L). The structure–activity relationships are discussed for all the compounds.     

Comparative in vitro activity of cefepime and other antibiotics against clinical strains of gram-negative bacteria]

In vitro activity of cefepime against etiologically significant strains of gramnegative microflora of patients treated in the Reanimation and Intensive Care Unit after cardiosurgical operations was evaluated. Sixty four strains of gramnegative aerobic bacteria isolated within the period from October 2001 to April 2002 were tested. The isolates susceptibility was determined by the disk diffusion method. Cefepime had an obvious advantage over the 3rd generation cephalosporins. Low incidence of cefepime resistant strains of the problem organisms in the Reanimation and Intensive Care Unit should be taken into account in empirical therapy of infections due to such pathogens.     

Evaluation of selected alcoholic antiseptics activity against clinical bacterial strains

The aim of this study was to evaluate antimicrobial activity of selected alcoholic antiseptics against clinical strains, which possessed in majority a high level of drug resistance: MRSA (7), MSSA (3), E. coli: (9): strains producing ESBL (4), P. aeruginosa: (4), E. cloacae: (3), K. pneumoniae: (3). These strains were defined by MIC value, using antibiotic agar dilution method according to NCCLS. Fourteen alcoholic antiseptics were used in this study. Beside alcohol, they contained other active substances like iodine, hydrogen peroxide, chlorhexidine. Some additional agents were included for easier application, such as: gelling, moisturizing, aromatic or coloring substances. The objective of this study was also to determine the dependence of bactericidal activity on preparations (concentration). Product undiluted and diluted two and four times in water was analyzed according to prEN 12054 standard; 30 seconds and 1 minute contact time was used. The obtained data indicate that all tested undiluted antiseptics possessed bactericidal activity described by producers. However antiseptics (dilution leads to decrease and even loss of bactericidal activity. Two-times dilution of gel almost completely inactivated the product. Antimicrobial activity after 30 seconds of contact time was not affected by presence of additional agents in the tested antiseptics.     

Occurrence of protein phosphorylation in various bacterial species.

1. The occurrence of protein phosphorylation in Escherichia coli B, Bacillus megaterium, Bacillus sphaericus, Pseudomonas fluorescens and Arthrobacter S1-55, was investigated by means of both in vivo and in vitro experiments. 2. In each bacterial species the presence of several phosphorylated proteins was evidenced by gel electrophoresis and autoradiography after either labelling of growing cells with [32P]orthophosphate or incubating cellular extracts with radioactive ATP. 3. The analysis of the radioactive moiety of proteins showed that they contained phosphoserine, phosphothreonine and phosphotyrosine. These three phosphoamino acids were found in varying proportions depending both on the bacterial species and, within the same species, on the conditions used for labelling proteins, either in vivo or in vitro. 4. By measuring the effect of cyclic nucleotides on the extent of protein phosphorylation in cellular extracts, it was observed that, in all five bacterial species analyzed, neither cyclic AMP nor cyclic GMP was able to stimulate the activity of protein kinases. 5. All together these results bring evidence that protein phosphorylation catalyzed by protein kinases is a post-translational modification widespread among prokaryotes.     

In vitro reactivation of sarin-inhibited brain acetylcholinesterase from different species by various oximes

In vitro as well as in vivo evaluation of the reactivating efficacy of various oximes against nerve agent-inhibited acetylcholinesterase has been usually done with the help of animal experiments. Nevertheless, previously published data indicate that the reactivation potency of oximes may be different in human and animal species, which may hamper the extrapolation of animal data to human data. Therefore, to better evaluate the efficacy of various oximes (pralidoxime, obidoxime, HI-6, K033) to reactivate brain acetylcholinesterase inhibited by sarin by in vitro methods, human, rat and pig brain acetylcholinesterase were used to calculate kinetic parameters for the reactivation. Our results show differences among the species, depending on the type of oxime, and indicate that data from animal experiments needs to be carefully evaluated before extrapolation to humans.     

In vitro reactivation of sarin-inhibited brain acetylcholinesterase from different species by various oximes

In vitro as well as in vivo evaluation of the reactivating efficacy of various oximes against nerve agent-inhibited acetylcholinesterase has been usually done with the help of animal experiments. Nevertheless, previously published data indicate that the reactivation potency of oximes may be different in human and animal species, which may hamper the extrapolation of animal data to human data. Therefore, to better evaluate the efficacy of various oximes (pralidoxime, obidoxime, HI-6, K033) to reactivate brain acetylcholinesterase inhibited by sarin by in vitro methods, human, rat and pig brain acetylcholinesterase were used to calculate kinetic parameters for the reactivation. Our results show differences among the species, depending on the type of oxime, and indicate that data from animal experiments needs to be carefully evaluated before extrapolation to humans.