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ABSTRACT: BACKGROUND: This study investigated a possible role of Escherichia coli in propagation and perpetuation of the chronic inflammation in ulcerative colitis (UC). The lesions of UC are located superficially on the rectal and/or colonic mucosa. It is suggested that the commensal bacteria of the digestive tract may play a role in the pathogenesis of UC. Several studies have demonstrated proliferation of E. coli in the gut of UC patients. An increase in the number of E. coli in the inflamed tissue is most probably related to the abundance of iron ions produced by the bacteria. METHODS: Colon mucosal biopsies were collected from 30 patients with acute-phase UC, both from tissues with inflammatory changes (n = 30) and unchanged tissue with no inflammatory changes (n = 30) from the same patient. Biopsies were also taken from 16 patients with irritable bowel syndrome diarrhea who comprised the control group. Quantitative and qualitative analysis of the biopsy specimens was performed using culture methods and real-time polymerase chain reaction (PCR). Genotyping of the E. coli isolates was done using pulsed-field gel electrophoresis. Multiplex PCR was used to compare the E. coli strains for the presence of genes responsible for synthesis of iron acquisition proteins: iroN, iutA, iha, ireA, chuA, and hlyA. RESULTS: We demonstrated that there was a significant increase in the number of E. coli at the sites of inflammation in patients with UC compared to the control group (P = 0.031). Comparative analysis of the restriction patterns of E. coli isolated from inflammatory and unchanged tissues showed that the local inflammatory changes did not promote specific E. coli strains. There was a significant difference in the frequency of the iroN gene in E. coli isolated from patients with UC as compared to the control group. CONCLUSIONS: The increase in the numbers of E. coli in the inflammatory tissues is related to the presence of chuA and iutA genes, which facilitate iron acquisition during chronic intestinal inflammatory processes.
BMC Gastroenterology 04/2013; 13(1):61. · 2.42 Impact Factor
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ABSTRACT: Biofilms are consortia of microorganisms (sessile cells) that form on various surfaces including mucosal membranes or teeth. Bacterial biofilms cause many human infections such as chronic sinusitis, acne vulgaris, periodontal diseases, and chronic wounds. These infections are persistent as they show increased resistance to antibiotics and host defense system. Taurine chloramine (TauCl) and taurine bromamine (TauBr) are the physiological products of activated neutrophils, resulting from the reaction between taurine with hypochlorous acid (HOCl) and hypobromous acid (HOBr), respectively. It has been shown in vitro that taurine haloamines exert antimicrobial properties against various pathogenic bacteria. Moreover, clinical studies have shown that both haloamines are effective in the local treatment of skin and mucose infections, including biofilm-related infections. Nevertheless, it has been not tested yet whether they can kill bacteria hidden in biofilm or disrupt biofilm structure. In this study we have investigated the capacity of TauCl and TauBr to inhibit in vitro the formation of P. aeruginosa biofilm. We have also tested their ability to destroy the mature biofilm. Our results suggest that TauBr is able to inhibit in vitro the formation of P. aeruginosa biofilm but cannot destroy the mature biofilm and effectively killed hidden bacteria. In further studies, the combined effect of TauBr and DNase, one of suggested biofilm inhibitors, was tested. Together, we conclude that TauBr is a better than TauCl candidate for local therapy of biofilm-related infections. However, a combined therapy, an application of TauBr together with other anti-biofilm agents (e.g., DNase), seems to be more promising.
Advances in experimental medicine and biology 01/2013; 775:269-83. · 1.09 Impact Factor
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ABSTRACT: In recent decades, the interest in probiotics as diet supplements or drugs has increased. In order to determine a specific bacterial isolate to be probiotic, it is necessary to describe precisely its probiotic characteristics and taxonomic properties, including the strain level. Most of the well-known genotyping methods were designed for the commonly-found pathogenic bacteria. The objective of this study is to undertake an attempt at standardization of FISH, RAPD and PFGE methods to genotype and identify the bacteria belonging to Lactobacillus fermentum, L. gasseri and L. plantarum species. The FISH probes have been designed and tested for Lactobacillus fermentum, L. gasseri and L. plantarum species and an endeavor has been made at standardization of RAPD and PFGE methods for these bacterial species. Moreover, the MLST method was applied to differentiate Lactobacillus plantarum strains. L. plantarum isolated from humans could not be genetically diversified with the use of RAPD, PFGE or MLST methods; only the strains originating from plants have displayed diversification among themselves and have been different from the strains of human origin.
Annals of Microbiology 12/2012; 62(4):1437-1445. · 0.69 Impact Factor
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ABSTRACT: The vaginal microflora is composed of many bacterial species and plays a major role in maintaining the balance of this complex environment. This study was conducted in order to assess the degree and persistence of the colonization of vaginal epithelium by strains from an orally administered mixture of lactobacilli, containing Lactobacillus fermentum 57A, Lactobacillus plantarum 57B and Lactobacillus gasseri 57C. We also monitored its effects on parameters of vaginal health, especially total lactobacilli counts, vaginal pH and Nugent score.
The patient group in this open study consisted of clinically healthy women with intermediate vaginal flora. Altogether 37 women were included in the study; 25 finished the full cycle consisting of 8 visits during 70 days. Lactobacillus mixture was administered as 1×10(8) c.f.u. once a day for 60 days. Lactobacillus isolates collected from vaginal and rectal samples from studied women during all visits were typed using molecular methods (PFGE for L. fermentum and L. gasseri and MLST for L. plantarum). Total lactobacilli counts, vaginal pH and Nugent score were also determined during the visits.
We confirmed that the ingested strains were able to reach and colonize both sites within the third and eighth visits, i.e. between the 20th and 70th days of the study. Maximal colonization was recorded between the fifth and seventh visits (31st-60th days). Moreover, ingestion of the Lactobacillus mixture was related to normalization of vaginal parameters (within 28-60 days after the initiation of the treatment). This was demonstrated by a decrease of vaginal pH and Nugent score together with an increase of total numbers of lactobacilli in the vagina and rectum. No adverse events were noted during the course of the study.
Oral application of the combination of the three probiotic strains derived from vaginal microbiota of healthy woman with high adherence abilities to both vaginal and colonic epithelium in vitro shows that both individual strains and their mixture can colonize vagina for some weeks, the effect of which is correlated with significant improvement of such parameters like pH and Nugent score values and total numbers of vaginal lactobacilli. This indicates that the mixture may be a good candidate for the planned double-blind, placebo-controlled randomized studies involving larger numbers of women.
European journal of obstetrics, gynecology, and reproductive biology 06/2012; 163(2):210-5. · 1.97 Impact Factor
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Annals of Microbiology 01/2012; · 0.69 Impact Factor
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ABSTRACT: A novel structure of exopolysaccharide from the lactic acid bacteria (LAB) Lactobacillus rhamnosus KL37B, from the human intestinal flora, is described. During the structural investigation of the exopolysaccharide it was found that the repeating unit is a nonasaccharide, which is the largest repeating unit found in LAB exopolysaccharides to date. The polysaccharide material was prepared by TCA extraction of a bacterial cell mass, purified by anion-exchange and gel permeation chromatography and characterized using chemical and enzymatic methods. On the basis of monosaccharide and methylation analysis and also 1D and 2D (1)H and (13)C NMR spectroscopy the exopolysaccharide was shown to be composed of the following nonasaccharide repeating unit: The physicochemical cell surface study and adhesive properties indicated distinct surface properties of Lactobacillus rhamnosus strain KL37B with high adhesive abilities to Caco-2 cells, hydrophobicity and slime production, in comparison to other Lactobacillus strains used as controls.
Carbohydrate research 10/2011; 346(18):2926-32. · 2.03 Impact Factor
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ABSTRACT: The commensal microbiota of the gastrointestinal tract plays an important role in the pathogenesis of inflammatory bowel disease. We examined the horizontal structure of the fecal microbiota in the colon in adolescents with Crohn disease or ulcerative colitis and a control group.
Fecal samples were collected in 3 fractions from patients with Crohn disease (n = 22), ulcerative colitis (n = 12), and controls (n = 24) during preparation for colonoscopy. Additionally, biopsies from colon tissue were taken. Samples were examined using a culture technique and a fluorescent in situ hybridization method. The mucin degradation assay was carried out.
Quantitative composition of the microbiota was different in the consecutive 3 fecal fractions and in the colon tissue of the study groups, but in patients from the control group, the composition of microbiota in the consecutive fractions was similar. Statistical analyses showed that the total distribution of the studied bacterial taxons in the contents in all 3 fecal fractions and in the colon tissue in the given disease group, and in the control group was characteristic for the studied patient group. Differences in species distribution among the cohorts studied were highly significant (P < 0.0001). Moreover, it was shown that in the fecal fraction I and in the colon tissue samples, there is no significant difference for any of the analyzed bacterial groups, using the culture methods or fluorescent in situ hybridization, but significant results were demonstrated in the II and III fractions for specific bacterial groups. The bacterial flora attached to the mucus layer in the UC group had significantly more degraded mucus in comparison with the control group (P = 0.045).
Distribution of the microbiota in the colon is layered, which can be called horizontal distribution of the fecal flora. Only in the ulcerative colitis group, the bacterial flora attached to the mucous layer exerts action on the mucin.
Journal of pediatric gastroenterology and nutrition 07/2011; 54(1):20-7. · 2.18 Impact Factor
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ABSTRACT: Macrolide resistance in 169 Streptococcus agalactiae [group B streptococcus (GBS)] isolates originating from pregnant carriers was investigated. Using multiplex PCR the presence of genes encoding erythromycin resistance and capsular polysaccharides, as well as surface proteins, was determined. Random amplification of polymorphic DNA (RAPD) and PFGE were used to characterize specific clones among the isolates. In the examined population of women, erythromycin-resistant strains were found in 4.5 % of patients, whereas clindamycin-resistant strains were found in 3 % of patients, which was 16 % of strains resistant to erythromycin and 10 % of strains resistant to clindamycin among GBS isolates, respectively. Among the isolates, the largest percentage was represented by the constitutive macrolide-lincosamide-streptogramin B (cMLS(B)) phenotype (63 %), then the inductive macrolide-lincosamide-streptogramin B (iMLS(B)) phenotype (26 %) and the macrolide resistance (M) phenotype (11 %). The ermB gene was indicated in all isolates with the cMLS(B) phenotype and V serotype, whereas mefA/mefE genes were found in isolates with the M phenotype and Ia serotype. Among resistance isolates, serotype V was predominant (67 %), followed by serotypes II (15 %), Ia (11 %) and III (7 %). The most common surface protein encoding genes were alp3 (70 %), then rib (11 %), epsilon (7.5 %), bca (7.5 %) and alp2 (4 %). A statistically significant relationship between macrolide resistance, serotype V and the alp3 gene was demonstrated. PFGE, in comparison to the RAPD method, gave better genetic discrimination of GBS isolates. A relatively high genetic diversity among investigated strains was shown. In addition, the largest genetic homogeneity was found in serotype V.
Journal of Medical Microbiology 03/2010; 59(Pt 7):780-6. · 2.50 Impact Factor
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ABSTRACT: Streptococcus agalactiae (Group B streptococci--GBS) is one of the coexistent components of vaginal microflora. Lactic Acid Bacteria (LAB) are the predominant bacteria in the physiological flora of vagina. The production of antagonistic substances like short chain fatty acids, hydrogen peroxide, bacteriocin-like substances protects against multiplication of pathogenic microorganisms. However, during pregnancy there is a disturbance of vaginal microflora and there are optimal conditions for bacterial or candidal vaginitis. Despite sufficient protection of LAB during pregnancy, Streptococcus agalactiae is one of the microorganisms, which can multiply and it is dangerous for newborns. The aim of the study was the assessment of antagonistic activity of chosen species of LAB to GBS strains. Antagonism between LAB and GBS was tested in a mixture of fluid 24 hrs cultures and the results were determined quantitatively by serial dilutions. Susceptibility of GBS strains to LAB activity was analyzed according to serotypes of GBS strains (Ia, Ib-V) and to place of isolation of GBS strains (there are three groups: GBS strains isolated from vagina of carriers; vaginal GBS strains isolated from women, who had clinical symptoms of vaginitis and GBS strains isolated from blood of children, who were ill with sepsis). The obtained results indicated strong inhibitory activity of Lactobacillus species to GBS strains after 2 hrs duration. Statistically significant dependence was found in the susceptibility of GBS strains to Lactobacillus. L. plantarum has shown the strongest activity against GBS strains. There were no statistically significant relationship found in the susceptibility of GBS to LAB activity between serotypes of GBS and between investigated groups of GBS.
Medycyna doświadczalna i mikrobiologia 01/2010; 62(2):153-61.
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Krzysztof Fyderek, Magdalena Strus,
Kinga Kowalska-Duplaga,
Tomasz Gosiewski,
Andrzej Wedrychowicz,
Urszula Jedynak-Wasowicz,
Małgorzata Sładek,
Stanisław Pieczarkowski,
Paweł Adamski,
Piotr Kochan,
Piotr B Heczko
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ABSTRACT: To assess the mucosa-associated bacterial microflora and mucus layer in adolescents with inflammatory bowel disease (IBD).
Sixty-one adolescents (mean age 15 years, SD+/-4.13) were included in the study. Intestinal biopsies from inflamed and non-inflamed mucosa of IBD patients and from controls with functional abdominal pain were cultured under aerobic and anaerobic conditions. The number of microbes belonging to the same group was calculated per weight of collected tissue. The mucus thickness in frozen samples was measured under a fluorescent microscope.
The ratios of different bacterial groups in inflamed and non-inflamed mucosa of IBD patients and controls were specific for particular diseases. Streptococcus spp. were predominant in the inflamed mucosa of Crohn's disease (CD) patients (80% of all bacteria), and Lactobacillus spp. were predominant in ulcerative colitis patients (90%). The differences were statistically significant (P=0.01-0.001). Lower number of bifidobacteria was observed in the whole IBD group. A relation was also found between clinical and endoscopic severity and decreased numbers of Lactobacillus and, to a lesser extent, of Streptococcus in biopsies from CD patients. The mucus layer in the inflamed sites was significantly thinner as compared to controls (P=0.0033) and to non-inflamed areas in IBD patients (P=0.031).
The significantly thinner mucosa of IBD patients showed a predominance of some aerobes specific for particular diseases, their numbers decreased in relation to higher clinical and endoscopic activity of the disease.
World Journal of Gastroenterology 11/2009; 15(42):5287-94. · 2.47 Impact Factor
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ABSTRACT: This paper describes the structure of neutral exopolysaccharide (EPS) produced by Lactobacillus johnsonii 142, strain of the lactic acid bacteria isolated from the intestine of mice with experimentally induced inflammatory bowel disease (IBD). Sugar and methylation analyses along with (1)H and (13)C NMR spectroscopy, including two-dimensional (1)H,(1)H COSY, TOCSY, NOESY, and (1)H,(13)C HSQC experiments revealed that the repeating unit of the EPS is a pentasaccharide: -->3)-alpha-D-Galp-(1-->3)-beta-d-Glcp-(1-->5)-beta-D-Galf-(1-->3)-alpha-D-Galp-(1-->3)-alpha-D-Galp-(1--> The rabbit antiserum raised against whole cells of L. johnsonii 142 reacted with homologous EPS, and cross-reacted with exopolysaccharide from Lactobacillus animalis/murinus 148 isolated also from mice with IBD, but not reacted with EPS of L. johnsonii 151 from healthy mice.
Carbohydrate research 09/2009; 345(1):108-14. · 2.03 Impact Factor
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ABSTRACT: The study was arranged to assess the actual rates of colonization of pregnant women and their children with group B streptococcus (GBS) in a Polish university hospital. Resistance of these cocci to macrolides and clindamycin was also tested and routes of transmission of GBS were followed in some cases using molecular typing. Colonization with GBS was checked in 340 pregnant women living in the south-eastern region of Poland (Małopolska) in the years 2004-2006. Women with a complicated pregnancy were more often colonized than those with a normal pregnancy (20.0 % versus 17.2 %). Moreover, women with a complicated pregnancy were twice as often colonized with GBS strains with the MLS(B) phenotype indicating resistance to macrolides and clindamycin. Regarding neonatal colonization by GBS, we found that neonates born from the colonized mothers with a complicated pregnancy were more often colonized with GBS than those from the mothers with a normal pregnancy (35 % versus 26.7 %). By molecular typing of the GBS strains isolated from mothers and their newborns we have been able to suggest the possibility of horizontal transmission of the strains from the hospital environment to newborns. Our results clearly indicate that rates of GBS colonization among pregnant women and neonates in a Polish university hospital have reached levels comparable to those reported in other European clinical centres.
Journal of Medical Microbiology 03/2009; 58(Pt 2):228-33. · 2.50 Impact Factor
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ABSTRACT: The most popular method of GBS identification in Poland currently is by culturing on enriched agar and verifying the Lancefield Group using special latex agglutination kits. However, the classical methods are time-consuming and their sensitivity is insufficient therefore it is becoming more common to try and apply molecular methods which are characterized by high sensitivity and rapid results. Moreover, molecular methods give us the possibility to carry out epidemiological investigations and gene detection, for instance for antibiotic resistance. It was confirmed that PCR and FISH procedures may be effective in rapid detection of GBS. Thanks to RAPD methods we showed that newborns born to colonized mothers were colonized by GBS strains which originated from the mother, irrespective of the way and the course of labour. Additionally, we detected GBS colonization in children who were born to mothers who were not colonized by GBS. These children were probably colonized with strains coming from hospital environment. More studies are needed to elucidate the route of transmission and the role of colonization of the medical staff. Using multiplex PCR we showed the presence of ermA, ermB and ermC genes in phenotypically confirmed MLS, GBS strains.
Medycyna doświadczalna i mikrobiologia 02/2008; 60(2):91-9.
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ABSTRACT: Streptococcus agalactiae, group B streptococci (GBS) are a constituent of normal vaginal bacterial microflora which often do not give any clinical symptoms. On the other hand, during pregnancy there are optimal conditions for GBS multiplication in the vagina, which may have very serious consequences for both the mother and her child. The women (n = 563) that participated in our study were in their 3rd trimester and they were divided into groups: normal pregnancy or high risk pregnancy. We also examined their newborns. GBS identification was done basing on traditional culture method and its modification recommended by the CDC. We showed a slightly improved (about 4%) effectiveness of GBS detection in pregnant women using the CDC method. In high risk pregnancy GBS colonization was 20% (among them 35% newborns were colonized) and in normal pregnancy it was found to be 17.2% (among them 26.7% newborns were colonized). Both in the high risk group and their newborns we confirmed a higher and statistically significant frequency of detection of GBS strains which had MLS(B) mechanism of antibiotic resistance. In newborns we confirmed two cases which were fatal. The results of our study show the need and necessity for implementing unified procedures recommended by the CDC in Poland.
Medycyna doświadczalna i mikrobiologia 01/2008; 60(1):5-12.
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ABSTRACT: Visible light induced photocatalytic inactivation of bacteria (Escherichia coli, Staphylococcus aureus, Enterococcus faecalis) and fungi (Candida albicans, Aspergillus niger) was tested. Carbon-doped titanium dioxide and TiO2 modified with platinum(IV) chloride complexes were used as suspension or immobilised at the surface of plastic plates. A biocidal effect was observed under visible light irradiation in the case of E. coli in the presence of both photocatalysts. The platinum(IV) modified titania exhibited a higher inactivation effect, also in the absence of light. The mechanism of visible light induced photoinactivation is briefly discussed. The observed detrimental effect of photocatalysts on various microorganism groups decreases in the order: E. coli > S. aureus approximately E. faecalis>C. albicans approximately A. niger. This sequence results most probably from differences in cell wall or cell membrane structures in these microorganisms and is not related to the ability of catalase production.
Photochemical and Photobiological Sciences 07/2007; 6(6):642-8. · 2.58 Impact Factor
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ABSTRACT: Lactobacillus rhamnosus (573L/1-3) strains are considered effective in the treatment of rotaviral diarrhoea in children. The colonisation of the gastrointestinal (G.I.) tract by the Lactobacillus rhamnosus strains and the determining factors are discussed reporting data of a prospective, double-blinded, placebo-controlled randomized study in children between the 2nd month and 6th year of life with acute diarrhoea lasting not longer than 5 days. The examined strains were detected in 37/46 (80.43%) patients after 5 days and in 19/46 (41.3%) patients after 14 days since the start of the treatment. L. rhamnosus 573L/1 strain colonised the G.I. tract more persistently. L. rhamnosus strains are effective in colonising the G.I. tract during acute diarrhoea. Persistence of colonisation is dependent on the properties of administered probiotic strains.
Digestive and Liver Disease 01/2007; 38 Suppl 2:S274-6. · 3.05 Impact Factor
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ABSTRACT: This study presents a series of experiments carried out in order to elucidate the role of H2O2 in antimicrobial activity of lactobacilli. Vaginal swabs were collected from 60 premenopausal women and checked for pH and Nugent score, and Lactobacillus species were cultured, phenotyped and genotyped. The main outcome measures involved: (1) species of vaginal lactobacilli most effective in liberating H2O2, (2) minimal microbicidal concentrations of added H2O2, (3) kinetics of H2O2 liberation in relation to oxygen tension, (4) antimicrobial activity of pure H2O2 versus one produced by selected vaginal lactobacilli and the total activity of their culture supernatants. Results showed that H2O2 was liberated especially by: Lactobacillus delbrueckii, Lactobacillus acidophilus, Lactobacillus crispatus, Lactobacillus johnsonii and L. gasseri. Hydrogen peroxide reached concentrations from 0.05 to 1.0 mM, which under intensive aeration increased even up to 1.8 mM. Microorganisms related to vaginal pathologies show varied resistance to the action of pure H2O2. Most potent inhibitory activity against bacteria and yeasts was presented by Lactobacillus culture supernate producing H2O2, followed by the nonproducing strain and pure H2O2. To conclude - the antimicrobial activity of lactobacilli is a summation of various inhibitory mechanisms in which H2O2 plays some but not a crucial role, in addition to other substances.
FEMS Immunology & Medical Microbiology 11/2006; 48(1):56-63. · 2.44 Impact Factor
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ABSTRACT: In this work we studied interactions between bacterial antigens and receptors on the surface of macrophages using atomic force microscopy (AFM). We used two bacterial cell wall components: lipopolysaccharide (LPS) derived from gram-negative Escherichia coli and exopolysaccharide (EPS) derived from gram-positive Lactobacillus rhamnosus. Interactions between these bacterial antigens and immune cell receptors were studied in peritoneal macrophages derived from two strains of mice, CBA and C3H/J, in which the Toll-like receptor 4 (TLR4) is genetically disabled. We collected 500 force-distance curves for LPS-activated cells using an EPS-covered AFM tip, and for EPS-activated cells using an LPS-covered AFM tip. Nonactivated cells were tested as reference cells. The results show that LPS-primed macrophages decrease their ability to bind EPS. Surprisingly, EPS-activated macrophages maintain or even increase their ability to bind LPS. This may suggest that in vivo commensal enteric bacteria, such as lactobacilli, will enhance the defense potential of local macrophages against pathogens expressing LPS.
Nanomedicine: nanotechnology, biology, and medicine 07/2006; 2(2):82-8. · 5.44 Impact Factor
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ABSTRACT: Lactobacilli, the predominant vaginal microorganisms in healthy premenopausal women, control other members of the vaginal microflora and thus protect against bacterial vaginosis and urinary tract infections. It has been claimed that some lactobacilli are also protective against Candida vaginitis. Little is known, however, about the mechanisms by which these lactobacilli can control vaginal populations of Candida and prevent vaginitis. To address this question, vaginal Lactobacillus strains with known antagonistic properties against bacteria were tested for their cell surface properties, adhesion to vaginal cell lines in vitro and antagonistic activities against Candida. A small proportion of the lactobacilli tested adhered strongly to cultured vaginal epithelial cells and inhibited growth of Candida albicans but not of C. pseudotropicalis. This anticandidal activity was in some Lactobacillus strains related to hydrogen peroxide (H2O2) production, but catalase treatment did not suppress this activity in other Lactobacillus strains, suggesting alternative mechanism(s). Moreover, tested vaginal Candida strains were resistant to relatively high concentrations of H2O2 that markedly exceeded those produced by even the most active Lactobacillus strains.
Infectious Diseases in Obstetrics and Gynecology 07/2005; 13(2):69-75.
