Helicobacter hepaticus HHGI1 is a pathogenicity island associated with typhlocolitis in B6.129-IL10 tm1Cgn mice

Helicobacter hepaticus strain 3B1 (H. hepaticus) contains a genomic island of approximately 71 kb, HHGI1, with some of the common features shared among known bacterial pathogenicity islands. In this study, we characterized the pathogenic potential of HHGI1 by infecting B6.129-IL10 tm1Cgn (IL10-/-) mice with an isogenic mutant (namely HhPAId1) lacking 19 predicted genes within HHGI1. In contrast to H. hepaticus (P<0.001), HhPAId1 did not cause typhlocolitis and hyperplasia in IL10-/- mice. Colonization levels of HhPAId1 were significantly higher in the cecum (P<0.007) and similar in the colon (P=0.27) when compared to H. hepaticus by 13 or 16 weeks post inoculation (WPI). The magnitude of the Th1-associated IgG2c response against HhPAId1 was less than that against H. hepaticus (P<0.004). There was no significant difference in Th2-associated IgG1 responses against these two strains. Cecal and colonic mRNA levels of proinflammatory cytokines IFN-gamma, TNF-alpha and IL-17a in the HhPAId1-infected mice were significantly lower than those in the H. hepaticus-infected mice (P<0.05) at 13 WPI. These results demonstrate that genes in the HHGI1 contribute to the pathogenicity of H. hepaticus, at least in part via up-regulation of proinflammatory mediators IFN-gamma, TNF-alpha and IL-17a.     

Is Helicobacter pylori a True Microaerophile?

BACKGROUND: There is no general consensus about the specific oxygen and carbon dioxide requirements of the human pathogen Helicobacter pylori. This bacterium is considered a microaerophile and consequently, it is grown under atmospheres at oxygen tensions 5-19% and carbon dioxide tensions 5-10%, both for clinical and basic and applied research purposes. The current study compared the growth of H. pylori in vitro, under various gas atmospheres, and determined some specific changes in the physiology of bacteria grown under different oxygen partial pressures. METHODS: Measurements of bacterial growth under various conditions were carried out employing classical solid and liquid culture techniques. Enzymatic activities were measured using spectrophotometric assays. RESULTS: H. pylori and all the other Helicobacter spp. tested had an absolute requirement for elevated carbon dioxide concentrations in the growth atmosphere. In contrast with other Helicobacter spp., H. pylori can tolerate elevated oxygen tensions when grown at high bacterial concentrations. Under 5% CO(2), the bacterium showed similar growth in liquid cultures under oxygen tensions from microaerobic (< 5%) to fully aerobic (21%) at cell densities higher than 5 x 10(5) cfu/ml for media supplemented with horse serum and 5 x 10(7) cfu/ml for media supplemented with beta-cyclodextrin. Evidence that changes occurred in the physiology of H. pylori was obtained by comparing the activities of ferredoxin:NADH (nicotinamide adenine dinucleotide) oxidoreductases of bacteria grown under microaerobic and aerobic atmospheres. CONCLUSIONS: H. pylori is a capnophile able to grow equally well in vitro under microaerobic or aerobic conditions at high bacterial concentrations, and behaved like oxygen-sensitive microaerophiles at low cell densities. Some characteristics of H. pylori cells grown in vitro under microaerobic conditions appeared to mimic better the physiology of organisms grown in their natural niche in the human stomach.     

The spectrum of Escherichia coli--Bacteroides fragilis pathogenic synergy in an intraabdominal infection model

Pathogenic synergy between Escherichia coli and Bacteroides fragilis was investigated in an intraabdominal infection model. Defined inocula of E. coli and B. fragilis, alone or in combination, were enmeshed within a fibrin clot and surgically implanted into the peritoneal cavity of rats. A spectrum of bacterial synergy ranging from synergistic abscess formation to synergistic lethality was demonstrated using this model. The type of synergy exhibited was dependent upon the initial E. coli inoculum. When combined with B. fragilis, high inocula of E. coli (greater than 10(8) cfu/clot) produced synergistic lethality while low inocula (2 x 10(2) to 2 x 10(7) cfu/clot) resulted in synergistic abscess formation. With respect to abscess formation, there was reciprocal synergy between E. coli and B. fragilis. Abscesses resulting from mixed inocula were larger and had significantly higher numbers of E. coli and B. fragilis than abscesses initiated by monomicrobial inocula. These studies define a clinically relevant model of bacterial interactions in the setting of intraabdominal infection and suggest that conclusions drawn from experimental models of bacterial synergy should consider the type of model examined, the strains of bacteria studied, and the number of bacteria inoculated.     

Potato Protoplast-derived Callus Tissue Challenged with Erwinia carotovora subsp. carotovora: Survival,Growth and Identification of Resistant Callus Lines*

Abstract Potato (cv. Crystal) protoplast-derived callus tissue was evaluated for survival and growth when exposed to Erwinia carotovora subsp. carotovora (strain Ecc71). Calli were either directly exposed to the pathogen by inoculation or to metabolites produced by the pathogen via a bilayer medium. Individual calli were inoculated with 0.5 μl of bacterial suspensions at 10⁴, 10⁵, 10⁶, 10⁷, 10⁸ and 10⁹ cfu/ml. The bilayer consistedof 10 ml of callus proliferation medium supplemented with pectin (2 g/l) and contained bacteria at 10², 10³, 10⁴, 10⁵ and 10⁶ cfu/ml. This medium was overlaid with 10 ml of bacteria free callus induction medium. Mean callus diameter of the inoculated treatments increased for 24 h, then declined. Over 90% of the inoculated calli were killed within 5 days but some survived as long as 14 days. Calli grown on the bilayer medium containing 10⁶, 10⁵ and 10⁴ cfu/ml also decreased in size. Most were killed within 9 days but some survived 20 days. Calli exposed to 10³ and 10² cfu/ml experienced limited growth with 20% and 7%, respectively, surviving after 27 days. Reactions to the pathogen varied considerably within the callus populations and individual calli with extended survival were identified in both experiments.     

Adhering ability of <Emphasis Type="Italic">Stenotrophomonas maltophilia</Emphasis> is dependent on growth conditions

The growth conditions are known to influence the bacterial adhesion to different kinds of surfaces. In the present study the adhering ability of S. maltophilia, on growth in nutrient rich media (Tryptic Soy Broth (TSB)) and minimal media (Luria Bertani (LB)) was checked by viable cell count and spectrophotometric method. TSB grown S. maltophilia showed higher adhesion compared to bacteria grown in LB broth, to both biotic and abiotic surfaces. However, when bacteria were grown in LB broth supplemented with different concentrations of glucose, under aerobic conditions, the bacteria grown at lower glucose concentration (2 gm/l) showed maximumadhesion to abiotic surfaces (polystyrene microliter plate) compared to biotic surfaces (mouse trachea, mouse tracheal mucus and HEp-2 cells line). Maximum adhesion to biotic surfaces was seen with cells grown at 4 gm/l of glucose concentration. On the contrary if the cell was grown under microaerophilic conditions maximum adhesion to abiotic and biotic surfaces was achieved with bacteria grown at 1 gm/l and 2 gm/l of glucose concentration respectively. A negative correlation was observed between glucose concentrations and pH of media, the latter declined faster under microaerophilic conditions as compared to aerobic condition.     

Production and storage of Rhizobium leguminosarum cell concentrates for use as inoculants

Recovery of Rhizobium leguminosarum cells by centrifugation after growth in an industrial fermenter was 100-fold higher when cells were grown on yeast extract (5 g/1) as sole source of carbon and nitrogen when compared with the yields recovered when cells were grown in standard mannitol-yeast extract medium. Methods of storing concentrated suspensions of R. leguminosarum were investigated. Freeze-drying caused a marked decrease in viable cell numbers. Viable cell numbers of bacterial concentrates stored in peat decreased steadily from 10₁₁-10₁₂ cfu/g to 10₉ cfu/g or less during 26 weeks storage at room temperature or at 4°C. Cell concentrates stored in 40% glycerol at — 20°C maintained viable numbers higher than 10₁₁ cfu/ml during a 76 week storage period.     

Effects of ozone treatment on cell growth and ultrastructural changes in bacteria

Ozone appeared to inhibit growth and caused the death of gram negative and gram positive tested bacteria: Escherichia coli, Salmonella sp., Staphylococcus aureus and Bacillus subtilis. Bacterial cultures at 10(3), 10(4), 10(5), 10(6), and 10(7) cfu/ml dilution were exposed to 0.167/mg/min/L of ozone at different time intervals (0, 5, 10, 15, 30, 60, 90, 120, and 150 min). Cell viability was observed in all types of tested bacteria at 10(3), 10(4), 10(3) cfu/ml within 30 min after ozone exposure. However, cell inactivation was not significantly observed at concentrations of 10(6), 10(7) cfu/ml even after an exposure of 150 min. Ultrastructural changes of treated bacteria showed deformation, rough damage and surface destruction revealed by scanning electron microscopy. Some bacterial cells showed collapsed and shrunken patterns within 60 min and severe rupture and cellular lysis after 90 min of ozone treatment. This study supports the proposed mechanism of the bacteria inactivation by ozone that caused cell membrane destruction and finally lysis reaction. Thus, the precaution of using ozone as a biocide should be used to address appropriate concentrations of bacterial contamination in water.     

Feedback regulation of an Agrobacterium catalase gene katA involved in Agrobacterium–plant interaction

Catalases are known to detoxify H2O2, a major component of oxidative stress imposed on a cell. An Agrobacterium tumefaciens catalase encoded by a chromosomal gene katA has been implicated as an important virulence factor as it is involved in detoxification of H2O2 released during Agrobacterium-plant interaction. In this paper, we report a feedback regulation pathway that controls the expression of katA in A. tumefaciens cells. We observed that katA could be induced by plant tissue sections and by acidic pH on a minimal medium, which resembles the plant environment that the bacteria encounter during the course of infection. This represents a new regulatory factor for catalase induction in bacteria. More importantly, a feedback regulation was observed when the katA-gfp expression was studied in different genetic backgrounds. We found that introduction of a wild-type katA gene encoding a functional catalase into A. tumefaciens cells could repress the katA-gfp expression over 60-fold. The katA gene could be induced by H2O2 and the encoded catalase could detoxify H2O2. In addition, the katA-gfp expression of one bacterial cell could be repressed by other surrounding catalase-proficient bacterial cells. Furthermore, mutation at katA caused a 10-fold increase of the intracellular H2O2 concentration in the bacteria grown on an acidic pH medium. These results suggest that the endogenous H2O2 generated during A. tumefaciens cell growth could serve as the intracellular and intercellular inducer for the katA gene expression and that the acidic pH could pose an oxidative stress on the bacteria. Surprisingly, one mutated KatA protein, exhibiting no significant catalase activity as a result of the alteration of two important residues at the putative active site, could partially repress the katA-gfp expression. The feedback regulation of the katA gene by both catalase activity and KatA protein could presumably maintain an appropriated level of catalase activity and H2O2 inside A. tumefaciens cells.     

Dependence of Staphylococcus epidermidis on non-transferrin-bound iron for growth

The ability of Staphylococcus epidermidis strains to grow in the presence of human transferrin and varying amounts of ferric iron was studied. At initial bacterial densities up to 10(4) cfu ml(-1), none of the three strains grew when transferrin iron saturation was below the full saturation point, whereas the bacteria grew consistently when transferrin was fully iron-saturated and there was non-transferrin-bound iron in the medium. Precultivation of the bacteria under iron-restricted conditions to induce siderophore production did not abolish the growth dependence on non-transferrin-bound iron. At initial bacterial densities of 10(6) cfu ml(-1), the bacteria proliferated consistently also in the presence of partially saturated transferrin. The results indicate that at low bacterial densities, S. epidermidis cannot utilise transferrin-bound iron for growth and that its proliferation is dependent on non-transferrin-bound iron.     

去乙酰化酶抑制剂TSA通过激活ERK和p38抑制间充质干细胞成脂分化

本文研究了丝裂原活化蛋白激酶(mitogen activated protein kinases, MAPKs)信 号通路在组蛋白去乙酰化酶抑制剂(histone deacetylase inhibitors, HDACis)曲古抑 菌素A(trichostatin A, TSA)抑制间充质干细胞(mesenchymal stem cells, MSCs) C3H10T1/2成脂分化中的调节机制.首先利用 MTT法检测TSA对其增殖活性的影响;Western印迹法首先检测MAPKs信号通路中pERK和p-p38蛋白在间充质干细胞C3H10T1/2成 脂分化过程中的表达情况,以及不同浓度、不同时间TSA处理对pERK和p-p38蛋白差异变 化情况;其次再用Western印迹检测TSA对成脂分化过程中间充质干细胞pERK和p-p38蛋 白表达的影响.MTT结果显示,TSA浓度在1 nmol/L~100 nmol/L范围内抑制C3H10T1/2细胞 的增殖活性,且TSA浓度约为60 nmol/L时即抑制一半以上的C3H10T1/2细胞增殖活 性.Western印迹结果显示,TSA处理5 min~80 min,及浓度在1 nmol/L~100 nmol/L范围内 激活MAPK信号通路中pERK和p-p38蛋白的表达;C3H10T1/2细胞成脂分化过程中,胞内pERK和p-p38蛋白的表达呈现下调趋势;而TSA抑制了成脂分化过程中C3H10T1/2细胞内pERK和p-p38蛋白的表达变化.本研究结果提示,在C3H10T1/2细胞成脂分化过程中,MAPK信号途径分子pERK和p-p38表达下调;TSA可能是通过活化pERK和p-p38进而抑制间充质干细胞C3H10T1/2成脂分化.     

Molecular hydrogen from water radiolysis as an energy source for bacterial growth in a basin containing irradiating waste

Although being deionized, filtered and therefore normally deeply oligotrophic, the water from a basin containing irradiating waste presented relatively high bacterial concentrations (ca 10(5) cfu ml(-1)) and biofilm development at its surface and on the walls. This water was characterized by a high concentration of molecular H2 due to water radiolysis, while its electrochemical potential was around +400 mV due the presence of dissolved O2 and active oxygen compounds. This combination of H2 availability and of an oxidant environment is completely original and not described in nature. From surface and wall biofilms, we enumerated the autotrophic populations ( approximately 10(5) bacteria ml(-1)) able to grow in presence of H2 as energy source and CO2 as carbon source, and we isolated the most abundant ones among cultivable bacteria. They efficiently grew on a mineral medium, in the presence of H2, O2 and CO2, the presence of the three gases being indispensable. Two strains were selected and identified using their rrs gene sequence as Ralstonia sp. GGLH002 and Burkholderia sp. GGLH005. In pure culture and using isotope exchange between hydrogen and deuterium, we demonstrated that these strains are able to oxidize hydrogen as energy source, using oxygen as an electron acceptor, and to use carbon dioxide as carbon source. These chemoautotroph hydrogen-oxidizing bacteria probably represent the pioneer bacterial populations in this basin and could be primary producers in the bacterial community.     

Survival of Escherichia coli O157 : H7 in yoghurt during preparation and storage at 4°C

Cow's milk was inoculated with ca 10³ and 10⁷ cfu ml⁻¹ Escherichia coli O157 : H7. After fermentation at 42°C for 0–5 h, the yoghurt was stored at 4°C. Two kinds of yoghurt were used : traditional yoghurt (TY), made with Streptococcus thermophilus and Lactobacillus bulgaricus starter cultures, and 'bifido' yoghurt (BY), made with the two starter cultures plus Bifidobacterium bifidum . After 7 d E. coli O157 : H7 decreased from 3·52 to 2·72 log₁₀ cfu ml⁻¹ and from 7·08 to 5·32 log₁₀ cfu ml⁻¹ in TY, and from 3·49 to 2·73 log₁₀ cfu ml⁻¹ and from 7·38 to 5·41 log₁₀ cfu ml⁻¹ in BY. The pH values of yoghurt dropped from 6·6 to 4·5 and 4·4 in TY (for low and high pathogen inocula, respectively), and from 6·6 to 4·6 and 4·5 in BY (for low and high pathogen inocula, respectively).     

The Uptake and Utilization of Bacteria, Amino Acids and Carbohydrate by the Rumen Ciliate Entodinium longinucleatum in Relation to the Sources of Amino Acids for Protein Synthesis

Entodinium longinucleatum grown in vitro in the presence of bacteria engulfed a wide range of bacterial species at rates of 130–3400 bacteria/h/protozoon (from suspensions of 10 bacteria/ml), but showed a preference for Klebsiella aerogenes and Proteus mirabilis which occurred in the growth medium. Some of the bacteria were digested with release of soluble material into the medium. Free amino acids were incorporated by the protozoa in the presence of chloramphenicol at rates of 5·4–15·1 nmol/h/10⁶ protozoa and approximately 40% of the amino acid-carbon was incorporated into protein. There was no appreciable synthesis of protozoal protein from carbohydrate. Evidence was obtained that the protozoa obtained the amino acids required for growth largely from engulfed bacteria.     

Sub-lethal damage of Listeria monocytogenes after long-term chilled storage at 4 degrees C

The possibility that long term in vitro chilled storage may result in sub-lethal damage to Listeria monocytogenes cells was investigated by comparing growth of chill-stored (starvation at 4 degrees C) and fresh cultures on selective and non-selective media. Growth of freshly grown cells was minimally (3-8%) affected by selective LSAMM agar compared with non-selective Brain Heart Infusion agar. In contrast, numbers of chill-stored strains were reduced by greater than 99% after direct plating on the same selective and non-selective media. Furthermore, chill-stored strains were able to grow in standard selective broth (Listeria Selective broth and Fraser broth) only if undiluted inocula (approximately 10(5)-10(6) cfu ml-1) were used, whereas they were capable of growth in Brain Heart Infusion broth even when the lowest dilutions were used (approximately 10(1) cfu ml-1). The potential public health consequences of this finding for the isolation of Listeria monocytogenes from foods is considered.     

Microbial fouling community analysis of the cooling water system of a nuclear test reactor with emphasis on sulphate reducing bacteria

Culture and molecular-based techniques were used to characterize bacterial diversity in the cooling water system of a fast breeder test reactor (FBTR). Techniques were selected for special emphasis on sulphate-reducing bacteria (SRB). Water samples from different locations of the FBTR cooling water system, in addition to biofilm scrapings from carbon steel coupons and a control SRB sample were characterized. Whole genome extraction of the water samples and SRB diversity by group specific primers were analysed using nested PCR and denaturing gradient gel electrophoresis (DGGE). The results of the bacterial assay in the cooling water showed that the total culturable bacteria (TCB) ranged from 10(3) to 10(5)?cfu?ml(-1); iron-reducing bacteria, 10(3) to 10(5)?cfu?ml(-1); iron oxidizing bacteria, 10(2) to 10(3)?cfu?ml(-1) and SRB, 2-29?cfu?ml(-1). However, the counts of the various bacterial types in the biofilm sample were 2-3 orders of magnitude higher. SRB diversity by the nested PCR-DGGE approach showed the presence of groups 1, 5 and 6 in the FBTR cooling water system; however, groups 2, 3 and 4 were not detected. The study demonstrated that the PCR protocol influenced the results of the diversity analysis. The paper further discusses the microbiota of the cooling water system and its relevance in biofouling.     

Cross-talk between endothelial and breast cancer cells regulates reciprocal expression of angiogenic factors in vitro

Reciprocal growth factor exchange between endothelial and malignant cells within the tumor microenvironment may directly stimulate neovascularization; however, the role of host vasculature in regulating tumor cell activity is not well understood. While previous studies have examined the angiogenic response of endothelial cells to tumor-secreted factors, few have explored tumor response to endothelial cells. Using an in vitro co-culture system, we investigated the influence of endothelial cells on the angiogenic phenotype of breast cancer cells. Specifically, VEGF, ANG1, and ANG2 gene and protein expression were assessed. When co-cultured with microvascular endothelial cells (HMEC-1), breast cancer cells (MDA-MB-231) significantly increased expression of ANG2 mRNA (20-fold relative to MDA-MB-231 monoculture). Moreover, MDA-MB-231/HMEC-1 co-cultures produced significantly increased levels of ANG2 (up to 580 pg/ml) and VEGF protein (up to 38,400 pg/ml) while ANG1 protein expression was decreased relative to MDA-MB-231 monocultures. Thus, the ratio of ANG1:ANG2 protein, a critical indicator of neovascularization, shifted in favor of ANG2, a phenomenon known to correlate with vessel destabilization and sprouting in vivo. This angiogenic response was not observed in nonmalignant breast epithelial cells (MCF-10A), where absolute protein levels of MCF-10A/HMEC-1 co-cultures were an order of magnitude less than that of the MDA-MB-231/HMEC-1 co-cultures. Results were further verified with a functional angiogenesis assay demonstrating well-defined microvascular endothelial cell (TIME) tube formation when cultured in media collected from MDA-MB-231/HMEC-1 co-cultures. This study demonstrates that the angiogenic activity of malignant mammary epithelial cells is significantly enhanced by the presence of endothelial cells.     

Expression of deuterium-isotope-labelled protein in the yeast Pichia pastoris for NMR studies

Deuterium isotope labelling is important for NMR studies of large proteins and complexes. Many eukaryotic proteins are difficult to express in bacteria, but can be efficiently produced in the methylotrophic yeast Pichia pastoris. In order to facilitate NMR studies of the malaria parasite merozoite surface protein-1 (MSP1) complex and its interactions with antibodies, we have investigated production of the MSP1-19 protein in P. pastoris grown in deuterated media. The resulting deuteration patterns were analyzed by NMR and mass spectrometry. We have compared growth characteristics and levels of heterologous protein expression in cells adapted to growth in deuterated media (95% D₂O), compared with expression in non-adapted cells. We have also compared the relative deuteration levels and the distribution pattern of residual protiation in protein from cells grown either in 95% D₂O medium with protiated methanol as carbon source, or in 95% D₂O medium containing deuterated methanol. A high level of uniform C deuteration was demonstrated, and the consequent reduction of backbone amide signal linewidths in [¹H/¹⁵N]-correlation experiments was measured. Residual protiation at different positions in various amino acid residues, including the distribution of methyl isotopomers, was also investigated. The deuteration procedures examined here should facilitate economical expression of ²H/¹³C/¹⁵N-labelled protein samples for NMR studies of the structure and interactions of large proteins and protein complexes.     

Enzymic detection of adhesion of enteropathogenic Escherichia coli to HEp-2 cells

We established a new method for detecting enteropathogenic Escherichia coli adhering to HEp-2 cells. An essential part of the method is an assay of beta-galactosidase activity of adhered bacterial cells. It consisted of the following steps: (1) culture of bacterial cells in a medium containing isopropyl-thio-beta-D-galactoside, an inducer of beta-galactosidase; (2) incubation of a bacterial culture with monolayered HEp-2 cells in a 96-well culture plate; (3) washing wells to remove bacterial cells which did not adhere to HEp-2 cells, and (4) enzymic reaction for beta-galactosidase activities. However, a calibration curve for the enzyme activity, obtained from each bacterial sample, showed that 10(5) bacteria per well permitted an accurate estimation. The enzyme activity of adhered bacteria to the monolayered cells showed that 10(7) bacteria were appropriate for the adherence assay. The number of adhered bacteria thus obtained was in good agreement with a viable cell count. The result indicates that the new method is more reliable than a widely used method, counting the number of bacteria under a microscope. The present method also makes it easy to detect adherent strains of E. coli in large numbers of specimens.     

Antibiofilm properties of chemically synthesized silver nanoparticles found against Pseudomonas aeruginosa

Nanomedicine is now being introduced as a recent trend in the field of medicine. It has been documented that metal nanoparticles have antimicrobial effects for bacteria, fungi and viruses. Recent advances in technology has revived the use of silver nanoparticles in the medical field; treatment, diagnosis, monitoring and control of disease. It has been used since ancient times for treating wide range of illnesses. Bacterial cells adheres to surfaces and develop structures known as biofilms. These structures are natural survival strategy of the bacteria to invade the host. They are more tolerant to commonly used antimicrobial agents, thus being more difficult to be controlled. This leads to increase in severity of infection. In this study, we have investigated the effect of silver nanoparticles in the formation of biofilm in multidrug resistant strains of Pseudomonas aeruginosa. Observation showed that biofilm formation occurred at bacterial concentration of 10⁶ cfu/ml for the sensitive strain of P. aeruginosa while in the resistant strain, the biofilm was evident at bacterial concentration of about 10³ cfu/ml. The biofilm were then tested against various concentrations of silver nanoparticles to determine the inhibitory effect of the silver nanoparticles. In the sensitive strain, 20 μg/ml of silver nanoparticles inhibited the growth optimally at bacterial concentration of 10⁴ cfu/ml with an inhibition rate of 67%. Similarly, silver nanoparticles inhibited the formation of biofilm in the resistant strain at an optimal bacterial concentration of 10⁵ cfu/ml with an inhibition rate of 56%. Thus, silver nanoparticles could be used as a potential alternative therapy to reduce severity of disease due to P. aeruginosa infections.     

Cytotoxin production in 100 strains of Haemophilus ducreyi from different geographic locations

Abstract One-hundred strains of Haemophilus ducreyi , representing isolates from different parts of the world, including the reference strains, were obtained from different collections and characterized with special reference to cytotoxin production in vitro. The cytotoxic activity on cultured epithelial cells (HEp-2) was examined with two methods. The activity in bacterial sonicates was tested on freshly trypsinated cells and strains manifesting little or no cytotoxic activity in sonicates were investigated using attached living bacteria on HEp-2 cell-monolayers. Sonicates from the majority of the H. ducreyi strains (89%) produced significant cytotoxic effects on HEp-2 cells. The reciprocal cytotoxic titers of the sonicates ranged from 2.4 × 10² to 5.3 × 10⁵. Sonicates of 11 strains had low cytotoxic titers ( 1:3 to 1:81), eight of those originating from Asia and three from Africa. These 11 strains caused no damage to the cell monolayer, indicating that the 11 strains produce little or no cytotoxic activity in vitro. In summary, the majority of H. ducreyi isolates produce cytotoxic activity, which support the hypothesis that the cytotoxin may be an important virulence factor of this species.