Bacteriocidal Effect Research Articles

Acetoin production as an indicator of growth and metabolic inhibition of Listeria monocytogenes.

It has been shown that Listeria monocytogenes produces acetoin from glucose under aerobic conditions. A defined medium with glucose as the sole carbon source was used in an aerobic shake flask culture to reliably produce acetoin. Acetoin, the reactive compound in the Voges-Proskauer test, was assayable in the medium and was used to quantify the metabolic response when inhibitors were added to the medium. Inhibitors such as lactic, acetic, propionic and benzoic acids were used to demonstrate the utility of acetoin production as an indicator of metabolic disruption. With increasing levels of inhibitor, the metabolic and growth responses were measured by acetoin production and optical density change, respectively. Both measurements decreased in a similar manner with increasing inhibitor concentrations. The data also showed the apparent mode of action of the inhibitors. A bacteriostatic effect was observed for the protonated organic acids, acetic (4 mmol l(-1)) and propionic (4 mmol l(-1)), whereas protonated lactic (4 mmol l(-1)) and benzoic (0.16 mmol l(-1)) acids gave an irreversible (apparent bacteriocidal) effect. Lactic, acetic, and propionic acids showed stimulation of metabolic activity at low concentrations, but benzoic did not. Acetoin production is a novel method for quantifying and assessing the mode of action of inhibitors against L. monocytogenes. This system can be used to screen inhibitors for applications in food safety.

Inhibition of Listeria monocytogenes on cold-smoked salmon by nisin and carbon dioxide atmosphere

The bacteriostatic and bacteriocidal effect of nisin in combination with carbon dioxide, NaCl and low temperature on the survival of Listeria monocytogenes was investigated in in vitro model studies and in trials with cold-smoked salmon. Addition of nisin caused various degrees of inhibition and sometimes death of L. monocytogenes in model experiments performed at 10°C. The antilisterial effect of nisin was improved in the presence of 100% CO2 and increasing NaCl concentrations (0.5 to 5.0% w/v). Minimal bactericidal concentrations (MBC) of nisin varied from 30 to more than 500 IU/ml. The most pronounced effect of nisin was found when 102 cfu/ml was grown in media with 5.0% NaCl and incubated in CO2 atmosphere (MBC=30 IU/ml). The bactericidal effect of nisin was reduced in air and vacuum, and did not increase systematically with increasing NaCl concentrations. In general, nisin concentration ≤50 IU/ml resulted in the survival and growth of L. monocytogenes in all combinations with other preservatives (NaCl, CO2). Addition of nisin (500 or 1000 IU/g) to cold-smoked salmon inoculated with L. monocytogenes and stored at 5°C delayed, but did not prevent growth of L. monocytogenes in vacuum-packs. Numbers of L. monocytogenes increased to 108 cfu/g in vacuum packed cold-smoked salmon in 8 days, whereas CO2 packing of cold-smoked salmon resulted in an 8-day lag phase of L. monocytogenes, with numbers eventually reaching 106 cfu/g in 27 days. Addition of nisin to CO2 packed cold-smoked salmon resulted in a 1 to 2 log reduction of L. monocytogenes followed by a lag phase of 8 and 20 days in salmon with 500 and 1000 IU nisin/g, respectively. The levels of L. monocytogenes remained below 103 cfu/g during 27 days of storage at both concentrations of nisin.

Growth inhibition of Prevotella ruminicola by protamine

Growth of Prevotella ruminicola strains B 14 (subsp. brevis) and D31d (subsp. ruminicola), was inhibited by protamine, a polycationic, low molecular mass protein. Results showed that protamine has a bacteriocidal effect when present in concentrations exceeding 30 μg ml −1. Protamine exerted its toxic effects by disrupting the outer membrane, which was demonstrated by: (i) an increased sensitivity to hydrophobic antibiotics (novobiocin and monensin) and (ii) release of the periplasmic enzyme alkaline phosphatase following short-term exposure to protamine. Although the concentrations of protamine inhibitory to P. ruminicola are relatively low, the effects of such a compound are probably too broad to permit its successful use in terms of manipulating ruminal proteolysis.

Comparative Killing Kinetics of Methicillin-Resistant Staphylococcus aureus by Bacitracin or Mupirocin

AbstractThe in vitro activities of bacitracin and mupirocin were compared for seven different strains of methicillin-resistant Staphylococcus aureus. Six of seven strains showed bacitracin minimum inhibitory concentrations (MICs) of 0.5 to 1.0 units/mL, and all seven had mupirocin MICs of 0.5 to 2 μg/mL. Time-kill studies revealed 2.6- to 4.5-log reduction in 24 hours with strains susceptible to bacitracin (4 units/mL) and 0 to 2.2 reduction with mupirocin (16 μg/mL). Bacitracin should be considered further for in vivo studies because of enhanced bacteriocidal effect and lower cost.

Comparative Killing Kinetics of Methicillin-Resistant Staphylococcus aureus by Bacitracin or Mupirocin

The in vitro activities of bacitracin and mupirocin were compared for seven different strains of methicillin-resistant Staphylococcus aureus. Six of seven strains showed bacitracin minimum inhibitory concentrations (MICs) of 0.5 to 1.0 units/mL, and all seven had mupirocin MICs of 0.5 to 2 μg/mL. Time-kill studies revealed 2.6- to 4.5-log reduction in 24 hours with strains susceptible to bacitracin (4 units/mL) and 0 to 2.2 reduction with mupirocin (16 μg/mL). Bacitracin should be considered further for in vivo studies because of enhanced bacteriocidal effect and lower cost.

Mechanism of lantibiotic-induced pore-formation

Nisin and other lantibiotics have a bacteriocidal effect against Gram-positive bacteria, and also inhibit the outgrowth of bacterial spores. The bacteriocidal effect appears to be due to the formation of pores in the bacterial membrane. In the absence of anionic membrane phospholipids, the lantibiotic nisin acts as an anion selective carrier. In the presence of anionic phospholipids, nisin forms nonselective, transient, multi-state pores in cells, proteoliposomes, liposomes and black lipid membranes. Pore formation involves distinct steps. First, nisin associates tightly with the anionic membrane surface leading to a high local concentration. This results in a disturbance of the lipid dynamics near the phospholipid polar head group-water interface, and an immobilization of lipids. In the presence of a transmembrane electrical potential above the threshold level, the molecules reorient, presumably as an aggregate, from a surface-bound into a membrane-inserted configuration. Co-insertion of bound, anionic phospholipids results in bending of the lipid surface giving rise to a wedge-like, nonspecific, water-filled pore.

Multicopy plasmid suppression of stationary phase chaperone toxicity in Escherichia coli by phosphogluconate dehydratase and the N-terminus of DnaK.

Overproduction of DnaK in Escherichia coli results in a bacteriocidal effect. This effect is most acute in stationary phase cells. A selection scheme was developed to isolate multicopy suppressors from an E. coli plasmid expression library, which overcome the stationary phase toxicity of excess DnaK. Two suppressor plasmids were recovered which contained inserts of 1.85 kb and 2.69 kb, respectively. Rearranged and deleted plasmid derivatives were constructed and used to further localize the suppressors. DNA sequence analysis demonstrated that one suppressor encoded phosphogluconate dehydratase (Edd) while the other suppressor encoded the N-terminal 237 amino acids of DnaK itself (DnaK'). Strains bearing the suppressor plasmids constitutively overproduced proteins with apparent masses of 66 kDa (Edd) and 37 kDa (DnaK') as determined by gel electrophoresis. Western blot analysis using polyclonal antisera specific for either Edd or DnaK confirmed the identity of these overproduced proteins. Suppression of DnaK toxicity was eliminated by the introduction of a + 1 frameshift mutation early in the respective coding regions of either of the two suppressors. These results suggest that suppressor gene translation plays a role in the mechanism of DnaK suppression.

アルゴンレーザーによる殺菌のメカニズム

黄色ブドウ球菌をはじめとするブドウ球菌, 大腸菌, 緑膿菌や他の細菌についてのレーザーによる殺菌効果に関する報告はいくらかみられるが, レーザーによる殺菌効果のメカニズムに関しては明らかではない。本実験において著者は, アルゴンレーザーによる殺菌効果のメカニズムについて次のような方法を用いて調べた。a. アルゴンレーザーによる殺菌効果について増殖阻止域の形成の有無を観察することにより調べた。b. MRSAをはじあとする5種類の細菌に対してレーザーを照射し, 殺菌に有効な照射出力と照射時間との関係を調べた。c. レーザー照射後のMRSAの形態学的変化について透過型電子顕微鏡を用いて観察した。その対照として, レーザー非照射菌, 紫外線を照射した細菌, 熱処理した細菌もあわせて観察した。d. アクリジンオレンジにより細菌を染色し, レーザー照射群と非照射群との間の蛍光性の違いを蛍光顕微鏡を用いて観察した。e. レーザー照射後のMRSAのDNA変化をアガロースゲル電気泳動を用いて観察した。以上の実験よりa. アルゴンレーザーによりすべての細菌について増殖阻止域の形成を認めた。b. 照射出力と照射時間とを変化させたところ殺菌効果の出現と照射出力との間には深い関係があり, また緑膿菌では他の細菌と比べてアルゴンレーザーにより殺菌され易かった。c. アルゴンレーザー照射後, 細菌の細胞質および核様体付近では電子密度が上昇していた。その変化は細菌を熱処理したものと非常に類似していた。d. レーザー照射前後で細菌のアクリジンオレンジの蛍光性に変化はなかった。e. レーザー照射前後で電気泳動上, 細菌のDNAのバンドパターンに変化はなかった。以上よりアルゴンレーザーによるMRSAに対する殺菌効果は, 光化学的作用よりも熱作用が主体であると考えられ. またアルゴンレーザーは細菌のDNAのバンドパターンには直接影響を及ぼさないことがわかった。

Control of Listeria monocytogenes on the surface of frankfurters by acid treatments

Small numbers of Listeria monocytogenes can be isolated from packaged frankfurters. Based on previous work from this laboratory, these L. monocytogenes cells undoubtedly represent surface recontamination during peeling and packaging. We investigated organic acid dips just prior to packaging as a secondary lethal step to destroy L. monocytogenes and observed both bacteriocidal and bacteriostatic effects depending on the concentration of acid used. Acetic acid when combined with citric acid (at 2·5% each) restricted the growth and development of L. monocytogenes on frankfurters stored vacuum-packaged at 5°C for up to 90 days. Other acids individually, such as lactic acid (5%) and acetic acid (5%), also suppressed the organism. A dose response effect was observed for all acids tested. A simple 2 min surface rinse in saline was adequate to remove L. monocytogenes from frankfurter surfaces and gave recoveries similar to a stomacher method. The acid treatments appeared to be lethal and did not injure the organism. Overall, organic acid treatments appear to provide a secondary lethal step in frankfurter processing and could be an additional factor in limiting the presence of L. monocytogenes in frankfurters.

Association of bacteria with Phaeocystis sp. In McMurdo Sound, Antarctica

We present a microscopic study of bacteria and Phaeocystis sp. during the annual bloom in McMurdo Sound, Antarctica. During the initial phases of the bloom when Phaeocystls sp. was actively growing, bacterial abundance and cell size both ~ncreased, suggesting that the bacterial community was also actively growing. At 5 and 25 m, the median density of bacteria associated with Phaeocystissp. colonies ranged from about 2to over 1 l-fold greater than over an equivalent area of a nearby region of the slide. On 7 out of 8 sampling dates, the length of bacteria associated with Phaeocystis sp. colonies was similar to or larger than pelagic bacteria from the same sample, suggesting that bacterial growth rates were not reduced by their proximity to the alga. In short, rather than bacteriocidal effects sometimes ascribed to Phaeocystjs sp., we observed a close association between both pelagic and epiphytic bacteria and Phaeocystis sp. throughout the bloom in McMurdo Sound. lnitial results with a strain of Phaeocystis pouchetii (CCMP 628) originally collected in Surinam and grown in culture suggest that differences between the results of this and previous studies might, at least in part, reflect strain differences. As the season progressed, disappearance of Phaeocystis sp. colonies in the upper water column corresponded to the appearance of a large bacterial bloom at and below 100 m depth. We speculate that in McMurdo Sound, the close alga-bacterial association might enhance remineralization of Phaeocystis sp. thus reducing the amount of organic material originatmy from the bloom which ultimately reaches the sediments.

Inhibition of Listeria monocytogenes by Lactobacillus sake strains of meat origin

The ability of two Lactobacillus sake strains of meat origin to inhibit the growth of Listeria monocytogenes at 4, 8, 15, 24 and 32°C in a conventional liquid media was investigated. Growth of L. monocytogenes was affected by Lac. sake strains at all temperatures. The inhibition was higher at 15, 24 and 32°C than at refrigeration temperatures. The inhibitory activity of both lactobacilli was similar perhaps due to the fact that Lac. sake 148 produces a bacteriocin inhibitory to L. monocytogenes, while Lac. sake 23 is a strong lactic acid producer. The antagonism exhibited by the lactobacilli on the L. monocytogenes strains seems to display a bacteriostatic rather than a bacteriocidal effect.

The Effect of pH on the Growth of MRSA

The aim of this experiment was to determine the influence of pH on MRSA growth. MRSA growth wau suppressed in acidic medium, and the degree of suppression paralleled the degree of acidity. The results showed that MRSA hardly grew in the media with a pH below 4, few cultures were identified at pH 2 and 3. A bacteriocidal effect was noted in the media with a pH below 3. At pH 5, cultures were markedly inhibited, and even at pH 6, culture failed to yield certain strains of MRSA.The usefulness of antibiotics can not be denied, but proper implementation of infection control should be excuted to avoid the outbreaks of resistant pathogens. Although treatment with dilute hydrochloric acid (pH 2) may only be applicable in superficial infections such as MRSA colitis, it seems to be promising as an attempt of antimicrobial therapy without any induction of resistant strains.

A bacteriocidal effect of 2-deoxy-D-galactose on Escherichia coli due to inhibition of amino sugar metabolism.

A bacteriocidal effect of 2-deoxy-D-galactose (dGal) in addition to the well known static effect was newly demonstrated on the basis of various physiological and biochemical events in Escherichia coli. Unlike D- galactose-induced lysis of a E. coli galE mutant via the process of uridine 5′-diphosphate galactose accumulation, dGal-induced growth inhibition was characterized by cell burst without swelling and elongation of the original rod shaped cells. 2-Deoxy-D-galactose 1-phosphate (dGal1P) was the most probable effector for a bacteriocidal effect of dGal owing to the relationship between dGal-sensitive phenotype and plasmid-dependent galK gene expression. The synthesis of cell wall polysaccharides was functional in dGal-grown cells with the added D-glucosamine and N-acetyl-D-glucosamine, which were protective against dGal-induced growth inhibition. The present study proposes inhibition of fructose 1, 6- bisphosphatase related to de novo synthesis of amino sugar with accumulation of dGal1P as a cause of overall growth inhibition by dGal.

Effects of Organic Solvents on Growth ofEscherichia coliK-12

Growth of Escherichia coli K-12 JA300 on agar and in medium in the presence of 10 different organic solvents has been investigated to obtain information on solvent tolerance. Cells grew readily in nutrient medium containing n-octane or isooctane, and some tolerant cells grew in the presence of n-hexane. Cyclohexane and p-xylene were toxic. Bacteriocidal effects of organic solvents agreed with their bacteriostatic effects observed previously. The toxicity correlated with the common logarithm of the partition coefficient of the solvents between n-octanol and water. When 13 cations were assessed for their effects on solvent tolerance, it was found that the organism grown in the presence of Mg2+, Ca2+, Ba2 +, or Sr2+ had a higher tolerance level towards n-hexane or cyclohexane. The addition of EDTA to medium decreased the solvent tolerance level of the organism.

Impairment of nucleoid segregation and cell division at high osmolarity in a strain of Escherichia coli overproducing the chaperone DnaK.

Escherichia coli strain WM1390, which overproduces 20-fold the chaperone protein DnaK, was able to grow exponentially without apparent abnormality in 300 mosM medium. In contrast, it showed both aberrant nucleoid segregation and strong inhibition of septation when shifted to high osmolarity. These impairments could not be accounted for by a bacteriocidal effect of the high DnaK content. Rather, the DnaK content appeared to promote faster growth than that of the parent C600, at least at high osmolarity in the presence of the osmoprotectant glycine betaine.

Impairment of nucleoid segregation and cell division at high osmolarity in a strain of Escherichia coli overproducing the chaperone DnaK

Escherichia coli strain WM1390, which overproduces 20-fold the chaperone protein DnaK, was able to grow exponentially without apparent abnormality in 300 mosM medium. In contrast, it showed both aberrant nucleoid segregation and strong inhibition of septation when shifted to high osmolarity. These impairments could not be accounted for by a bacteriocidal effect of the high DnaK content. Rather, the DnaK content appeared to promote faster growth than that of the parent C600, at least at high osmolarity in the presence of the osmoprotectant glycine betaine.

Clinical evaluation of imipenem/cilastatin sodium and fosfomycin as second-line combination chemotherapy in severe infections associated with hematologic disorders

Imipenem/cilastatin sodium (IPM/CS) which is a broad-spectrum agent against both Gram-positive and -negative bacteria was used in combination with fosfomycin (FOM) as a second-line chemotherapy for severe infections associated with hematologic disorders. FOM was partnered with IPM because FOM may enhance the bacteriocidal effects of IPM when given as pretreatment to IPM/CS therapy. Fifty two patients were treated with IPM/CS plus FOM. Of them, 41 were evaluated for effectiveness. Eleven patients were not evaluated: 4 were treated with a combination of other regimens such as cefixime, gamma-globulin, G-CSF and a large dose of methyl prednisolone; 2 were given IPM/CS plus FOM as a first choice; 3 were observed to have gastrointestinal side effects such as nausea which led to the discontinuation of the combination therapy; and 2 were thought to be suffering from not infectious but tumor fever. An excellent response was observed in 15 (36.6%) patients and a good response in 10 (24.4%), for a overall efficacy rate of 61.0%. Efficacies was 71.4% (5/7) in patients with sepsis, and 60.0% (9/15) in patients whose peripheral granulocyte count was below 100/microliters before chemotherapy. The elimination rates of Gram-positive and -negative bacteria were 57.1% (4/7) and 75.0% (6/8), respectively. In particular, 75.0% (3/4) of Pseudomonas aeruginosa identified were eliminated. Two patients who suffered from tumor fever, 2 who did not receive chemotherapy before the combination chemotherapy and 3 who did not receive a full course of the combination chemotherapy because of side effects, were included in the final evaluation of safety. Side effects were observed in 18 of 48 patients (37.5%). In 1 patient, skin eruption occurred 3 days after the initiation of the combination chemotherapy. In 17 patients, gastrointestinal symptoms such as nausea and vomiting were identified after a few days of IPM/CS plus FOM administration. Degree's of the symptoms were mild, however. Therefore, the treatment was not withdrawn. No abnormal laboratory results such as eosinophilia, liver disfunction or renal disfunction were observed. These results show that IPM/CS plus FOM is effective as a second-line combination chemotherapy for the treatment of severe infections in patients with hematologic disorders.

The bacteriocidal effects of transition metal complexes containing the NO+group on the food-spoilage bacteriumClostridium sporogenes

Abstract The chemical and molecular mechanism of toxicity of nitrite towards food-spoilage bacteria such as Clostridium botulinum or Clostridium sporogenes is not well understood. In order to discover the active species and explore its chemistry, a number of compounds related to nitrite were synthesized. Their bacteriocidal effects on C. sporogenes were investigated in Oxoid nutrient broth No. 2 growth medium at pH 7.0. Inhibition of cell growth, expressed as the concentration which causes 50% cell inhibition, was observed with nitrite at 10 mM, whereas [Fe4S3(NO)7]− (the anion of Roussin's black salt) and (Fe2(SCH2CH2OH)2(NO)4] (a water-soluble Roussin's red salt ester) were found to be effective at 0.001 mM and 0.005 mM, respectively, confirming previous reports that iron-sulphur-nitrosyl complexes are much more toxic to these organisms than nitrite itself. The nitroprusside anion, [Fe(CN)5NO]2− was found to be toxic at 0.030 mM and the corresponding chromium species, [Cr(CN)5NO]3−, at 0.1 mM. Therefore, on the basis of the number of NO groups present, the nitrosylcyano complexes are comparable in activity with the iron-sulphur-nitrosyl compounds. These results show that neither iron nor sulphur are essential for the bacteriostatic effect of the Roussin's type compounds. The property that all these compounds have in common is that they contain NO+. It is proposed that this is the active species responsible for the preservative effect of nitrite, and that a relationship may exist between the NO stretching frequency, a measure of the NO+ character, and the toxicity of these NO+-containing complexes.

Physiological consequences of DnaK and DnaJ overproduction in Escherichia coli.

The physiological consequences of molecular chaperone overproduction in Escherichia coli are presented. Constitutive overproduction of DnaK from a multicopy plasmid containing large chromosomal fragments spanning the dnaK region resulted in plasmid instability. Co-overproduction of DnaJ with DnaK stabilized plasmid levels. To examine the effects of altered levels of DnaK and DnaJ in a more specific manner, an inducible expression system for dnaK and dnaJ was constructed and characterized. Differential rates of DnaK synthesis were determined by quantitative Western blot (immunoblot) analysis. Moderate levels of DnaK overproduction resulted in a defect in cell septation and formation of cell filaments, but co-overproduction of DnaJ overcame this effect. Further increases in the level of DnaK terminated culture growth despite increased levels of DnaJ. DnaK overproduction was found to be bacteriocidal, and this effect was also partially suppressed by DnaJ. The bacteriocidal effect was apparent only with cultures which were allowed to enter stationary phase, indicating that DnaK toxicity is growth phase dependent.

The bacteriocidal effects of transition metal complexes containing the NO+ group on the food-spoilage bacterium Clostridium sporogenes.

The chemical and molecular mechanism of toxicity of nitrite towards food-spoilage bacteria such as Clostridium botulinum or Clostridium sporogenes is not well understood. In order to discover the active species and explore its chemistry, a number of compounds related to nitrite were synthesized. Their bacteriocidal effects on C. sporogenes were investigated in Oxoid nutrient broth No. 2 growth medium at pH 7.0. Inhibition of cell growth, expressed as the concentration which causes 50% cell inhibition, was observed with nitrite at 10 mM, whereas [Fe4S3(NO)7]-(the anion of Roussin's black salt) and (Fe2(SCH2CH2OH)2(NO)4] (a water-soluble Roussin's red salt ester) were found to be effective at 0.001 mM and 0.005 mM, respectively, confirming previous reports that iron-sulphur-nitrosyl complexes are much more toxic to these organisms than nitrite itself. The nitroprusside anion, [Fe(CN)5NO]2- was found to be toxic at 0.030 mM and the corresponding chromium species, [Cr(CN)5NO]3-, at 0.1 mM. Therefore, on the basis of the number of NO groups present, the nitrosylcyano complexes are comparable in activity with the iron-sulphur-nitrosyl compounds. These results show that neither iron nor sulphur are essential for the bacteriostatic effect of the Roussin's type compounds. The property that all these compounds have in common is that they contain NO+. It is proposed that this is the active species responsible for the preservative effect of nitrite, and that a relationship may exist between the N-O stretching frequency, a measure of the NO+ character, and the toxicity of these NO(+)-containing complexes.