Fructooligosaccharides and mannose affect Clostridium difficile adhesion and biofilm formation in a concentration-dependent manner.

The aim of this study was to investigate the effects that prebiotic and candidates for prebiotics on Clostridium difficile strains to adhere to various human epithelial cell lines and to compare the adhesive properties of specific C. difficile strains. We also sought to examine the effect of different concentrations of fructooligosaccharides and mannose on the formation of biofilms by C. difficile strains. The influence of cellobiose, fructooligosaccharides, inulin, mannose, and raffinose on the adherence properties of various C. difficile strains, including motile 630, non-motile M120, and 10 clinical motile ribotype 027 strains, to non-mucous secreting HT-29, mucous secreting HT-29 MXT, and CCD 841 CoN cells lines. The most effective prebiotics were used in biofilm formation assays. We demonstrated that all C. difficile strains adhered to all cell lines. However, the C. difficile M120 non-motile strain was statistically more likely to adhere to all three cell lines (CFU median, 40) compared to the motile strains (CFU median, 3; p < 0.001). Furthermore, among the carbohydrates examined, only fructooligosaccharides and mannose were found to significantly decrease adhesion (p < 0.001) of C. difficile strains. Alternatively, using a biofilm assay, we observed, via confocal laser scanning microscopy, that sub-inhibitory concentrations (1%) of fructooligosaccharides and mannose functioned to increase biofilm formation by C. difficile. We demonstrated that specific prebiotics and candidate prebiotics exhibit varying anti-adhesive properties towards C. difficile in vitro and that treatment with sub-inhibitory concentrations of prebiotics can cause an increase in biofilm formation by C. difficile.

The level of fecal calprotectin significantly correlates with Clostridium difficile infection severity.

INTRODUCTION: Fecal calprotectin (FC) rises significantly in intestinal inflammation accompanied by neutrophil activation - such as Clostridium difficile infection (CDI). The aim of the study was to evaluate the benefit of FC testing in assessing the severity of CDI. MATERIALS AND METHODS: The study group included 76 patients with CDI hospitalized in the Jagiellonian University Hospital in Krakow from July 2017 till January 2018. FC levels were measured using an EIA (Enzyme Immunoassay). Demographic, clinical information and blood tests were recorded using standardized data collection forms. The selection of patients into non-severe and severe groups was carried out in accordance with the ESCMID criteria (European Society of Clinical Microbiology and Infectious Diseases) and some modi cations to those criteria were proposed. RESULTS: the studied population included 76 patients (39 men and 37 women) with CDI aged from 24 to 98 years (mean: 72). Median calprotectin level was 739 (Q25-Q75: 612-799 µg/g), characteristic of patients with colitis. A statistically significant difference in FC concentration in patients with severe vs non-severe CDI was observed (severe - 770 vs non-severe - 659 µg/g, p = 0.009). FC directly correlated with platelets level; however, no correlation between FC level and the blood parameters prognostic for CDI (leukocyte, neutrophil count, albumin, creatinine levels) was found. CONCLUSION: FC level is an indication of ongoing intestinal inflammation in CDI patients. FC level significantly correlated with CDI severity, which demonstrates that FC could serve as a predictive marker for assessing CDI severity.

Hospital-based Clostridium difficile infection surveillance reveals high proportions of PCR ribotypes 027 and 176 in different areas of Poland, 2011 to 2013.

As part of the European Clostridium difficile infections (CDI) surveillance Network (ECDIS-Net), which aims to build capacity for CDI surveillance in Europe, we constructed a new network of hospital-based laboratories in Poland. We performed a survey in 13 randomly selected hospital-laboratories in different sites of the country to determine their annual CDI incidence rates from 2011 to 2013. Information on C. difficile laboratory diagnostic testing and indications for testing was also collected. Moreover, for 2012 and 2013 respectively, participating hospital-laboratories sent all consecutive isolates from CDI patients between February and March to the Anaerobe Laboratory in Warsaw for further molecular characterisation, including the detection of toxin-encoding genes and polymerase chain reaction (PCR)-ribotyping. Within the network, the mean annual hospital CDI incidence rates were 6.1, 8.6 and 9.6 CDI per 10,000 patient-days in 2011, 2012, and 2013 respectively. Six of the 13 laboratories tested specimens only on the request of a physician, five tested samples of antibiotic-associated diarrhoea or samples from patients who developed diarrhoea more than two days after admission (nosocomial diarrhoea), while two tested all submitted diarrhoeal faecal samples. Most laboratories (9/13) used tests to detect glutamate dehydrogenase and toxin A/B either separately or in combination. In the two periods of molecular surveillance, a total of 166 strains were characterised. Of these, 159 were toxigenic and the majority belonged to two PCR-ribotypes: 027 (n=99; 62%) and the closely related ribotype 176 (n=22; 14%). The annual frequency of PCR-ribotype 027 was not significantly different during the surveillance periods (62.9% in 2012; 61.8% in 2013). Our results indicate that CDIs caused by PCR-ribotype 027 predominate in Polish hospitals participating in the surveillance, with the closely related 176 ribotype being the second most common agent of infection.

Antimicrobial susceptibility patterns of Clostridium difficile strains belonging to different polymerase chain reaction ribotypes isolated in Poland in 2012.

In the beginning of 2012, a study was conducted to obtain an overview of Clostridium difficile infections (CDIs) in Polish hospitals. The collection of 83 toxigenic C. difficile isolates obtained from this hospital-based survey was used to identify antimicrobial susceptibility patterns. Among the C. difficile isolates analyzed, 48 (57.8%) belonged to PCR ribotype 027, 21 (25.3%) to its closely related PCR ribotype 176, and 14 (16.9%) to different PCR ribotypes. Seventy one (85.5%) isolates were resistant to erythromycin, whereas 23 (27.7%) had high-level clindamycin resistance, having minimum inhibitory concentrations (MICs) greater than 256 mg/L. All strains were ciprofloxacin resistant and 69 (83.1%) were moxifloxacin resistant. Seventy-three (87.9%) strains were imipenem resistant, but only 2 (2.4%) strains were resistant to tetracycline. All strains were sensitive to tigecycline. Metronidazole and vancomycin were generally effective against the C. difficile isolates, both having an MIC90 value of 0.75 mg/L. Isolates belonging to PCR ribotype 027 and the closely related PCR ribotype 176, showed higher resistance. All ribotype 027 and 176 C. difficile isolates demonstrated high-level resistance to erythromycin (MIC ≥ 256 mg/L), and 95,2% of ribotype 176 isolates were co-resistant to erythromycin and clindamycin. The MIC of moxifloxacin for this epidemic strain was very high (≥32 mg/L). Resistance to erythromycin, moxifloxacin, and rifampicin was observed in 15 (18%) of the isolates, all of which belonged to PCR ribotype 027. Multidrug resistance (MDR), defined as resistance at least to three classes of antimicrobial agents was observed in 85.5% (n = 71) of toxigenic C. difficile strains.

[Evaluation of growth of clinical Clostridium difficile strains belonging to different PCR-ribotypes on chromID C. difficile Agar]. / Ocena wzrostu na podlozu chromID C. difficile Agar klinicznych szczepów Clostridium difficile nalezacych do róznych PCR-rybotypów.

INTRODUCTION: Clostridium difficile is main reason of antibiotic-associated diarrhea in hospitalized patients. Diagnostic method for detection of Clostridium difficile infection (CDI) are limited to an enzyme immunoassays (EIAs), while the culture of toxigenic strains is still seen as the gold standard for the laboratory diagnosis. The aim of this study was to compare growth of C. difficile strains belonging to different polymerase chain reaction (PCR) ribotypes on new ChromID C. difficile Agar (CDIFF, bioMérieux, Marcy l'Etoile, France). MATERIALS AND METHODS: One hundred thirty one of clinical C. difficile strains stored. in Anaerobic Laboratory were cultured on ChromID C. difficile Agar. Ten faecal samples were cultured on the same chromogenic medium and incubated at 37°C for 24 h under anaerobic conditions. Isolates were confirmed as C. difficile on the basis of well-known criteria. PCR-ribotyping was performed by visually comparison of patterns of PCR products of the 16S-23S rRNA intergenic spacer region. We examined the occurrence of beta-glucosidase gene, responsible for the dark color of the colony C. difficile on ChromID C.difficile Agar using a pair of primers: gluF (5'-AAGGT GTAAATTTAGGAGGTTGGTT-3') i gluR (5'-AGGTCCCAACTATCCC ATCC-3'). RESULTS: Among ten C. dfficile isolates obtained from stool specimens one formed colorless colonies. We received 8 colorless isolates from 131 additional examined strains. All C. difficile isolates forming colorless colonies belonged to PCR ribotype 023. The prevalence of PCR-ribotype 023 was about 6%. We detected lack of beta-glucosidase gene in PCR-ribotype 023 isolates. CONCLUSIONS: There are some C. difficile strains forming colorless colonies on ChromID C.difficile Agar. This appearance is important in routine diagnostic use this chromogenic culture medium.

Occurrence of Clostridium difficile PCR-ribotype 027 and it's closely related PCR-ribotype 176 in hospitals in Poland in 2008-2010.

Since 2003, a rising incidence of Clostridium difficile infection (CDI) in North America and Europe has coincided with outbreaks of C. difficile PCR ribotype 027. This ribotype was not observed in Poland until 2008. In the period 2008-2010, outbreaks of antibiotic-associated diarrhoea occurred in three different hospitals in Poland. Of 30 C. difficile isolates available for microbiological characterisation, 17 (56%) were positive for binary toxin genes and belonged to PCR ribotype 027 (n = 7) and its closely related PCR ribotype 176 (n = 10). All 17 binary toxin-positive C. difficile strains demonstrated high-level resistance to fluoroquinolones (minimum inhibitory concentration (MIC) ≥ 32 mg/L), including ciprofloxacin, gatifloxacin, and moxifloxacin, as well as erythromycin and clindamycin (MIC ≥ 256 mg/L for both). Of 14 patients from whom clinical information was available, 50% had a severe form of CDI, defined by fever (>38.5 °C), decreased kidney function, and high leucocyte count. We conclude that outbreaks of CDI associated with hypervirulent strains belonging to PCR ribotypes 027 and 176 occurred in hospitals in Poland. Further studies evaluating the clinical impact of type 176 are urgently needed.

Three-step diagnostic algorithm in diagnosing patients suspected of Clostridium difficile-associated diarrhea.

UNLABELLED: Clostridium difficile is a predominant etiological agent of healthcare-associated infectious diarrhea. Immunoenzymatic tests for detecting toxins A/B from faecal samples are still used in routine diagnosis of Clostridium difficile-associated diseases in a number of healthcare centers in Poland. Recently, however, new diagnostic tests were introduced which allow for detecting toxigenic strains of C. difficile in a more effective and precise manner. It is of importance, especially in the light of hypervirulent strain occurrence. AIM: The aim of the present paper was to evaluate the efficacy of three-step algorithm in the diagnosis of Clostridium difficile-associated diseases (CDAD), considering the occurrence of false negative test results for toxins while using exclusively immunoenzymatic tests. MATERIALS AND METHODS: In the present study, faecal samples collected from patients presenting diarrhea were tested. Immunoenzymatic tests were used for detecting glutamate dehydrogenase (GDH) and toxins A/B. Culture and RT-PCR were also employed. RESULTS: Of 615 study participants, toxigenic strains GDH (+) TOX (+) were identified in 108 patients while for 67 patients, test results remained unspecified GDH (+) TOX (-). Further analysis of unspecified samples revealed 32 patients infected with toxigenic strains, i.e. 22.9% of all positive test results (n=140). CONCLUSION: Three-step diagnostic algorithm is an effective and reliable tool for diagnosing C.difficile- associated diseases.

[Assessment of antagonistic activity in vitro Lactobacillus spp. strains againts Clostridum difficile strains isolated from gastrointestinal tract of patients hospitalized in three hospitals in region Mazovia]. / Ocena antagonistycznego dzialania w warunkach in vitro szczepów z rodzaju Lactobacillus wobec toksynotwórczych szczepów Clostridium difficile wyhodowanych z przewodu pokarmowego pacjentów leczonych w szpitalach na terenie województwa mazowieckiego.

INTRODUCTION: The aim of this study was to evaluate the antagonistic activity of Lactobacillus strains against clinical C. difficile strains isolated from faecal samples of adults patients with diarrhea. A total 61 strains of C. difficile randomly selected isolated in the period 2007-2008 from the gastrointestinal tract of hospitalized patients in three hospitals province Mazovia, Poland. To determination of antagonistic activity ofprobiotic Lactobacillus spp. strains used four reference strains: Lactobacillus plantarum 2017405, Lactobacillus fermentum 353, Lactobacillus acidophilus DSM 21007 and Lactobacillus rhamnosus GG. METHODS: Isolation of C. difficile was performed on selective Columbia agar supplemented with cycloserine/cefoxitine and amphothericin B (CLO medium, bioMérieux, France). The plates were incubated in an anaerobic chamber for 48 h at 37 degrees C. Isolates were identified as C. difficile by the characteristic morphology of the colonies and horse-like odour, green yellow fluorescence under UV. Toxigenicity of of C. difficile strains was determined in PCR to detection of fragments of genes encoding toxin A (tcdA), toxin B (tcdB) and binary toxin (cdtA and cdtB). The study of antagonistic activity four Lactobacillus spp. strains against 61 clinical C. difficile strains was performed according to standard methods. Lactobacillus strains were inoculated on MRS medium and incubated in oxygen-free atmosphere and cut the bars of MRS agar and applied to the plate with cultures of C. difficile strains. RESULTS: Assessment of antagonist activity of Lactobacillus spp. strains was performed by measuring the zone of inhibition of grown of C. difficile strains. The study shows that of probiotic Lactobacillus spp. strains interacted antagonistically in vitro against all toxigenic (A+B+CDT- and A+B+CDT+) of C. difficile strains. CONCLUSIONS: The differences in the antagonistic activity of Lactobacillus spp. strains against different toxigenic clinical C. difficile strains were not observed.

[Assessment of susceptibility of strictly anaerobic bacteria originated from different sources to fluoroquinolones and other antimicrobial drugs]. / Ocena wrazliwosci szczepów bezwzglednych beztlenowców pochodzacych z róznych zródel na fluorochinolony oraz inne leki stosowane w lecznictwie szpitalnym.

INTRODUCTION: During the past 20 years, several studies at a national level in different countries followed resistance trends for Bacteroides sp. and Clostridium difficile. This study analysed antimicrobial susceptibility 73 anaerobic bacteria strains of Bacteroides fragilis group (BFG) and C. difficile to fluoroquinolones and other antimicrobial drugs. METHODS: The strictly anaerobes strains isolated in different hospitals were sent to the Department of Medical Microbiology, Medical Uniwersity of Warsaw, where species determination was carried out with the API20 ANA (bioMerieux SA, Marcy-l'Etoile, France) system. Susceptibility to antimicrobials was determined using E-test. RESULTS: The rates of high resistance to ciprofloxacin and moxifloxacin of BFG was respectively 84% and 31% and among of C. difficile strains respectively 92% and 36%). The percentage of BFG strains resistant to erythromycin and clindamycin were respectively 84% and 46%. The percentage of C. difficile strains resistant to erythromycin and clindamycin was 52%. Reduced level of susceptibility of BFG strains to amoxicillin/clavulanic acid (8%) was confirmed. Resistance to cefoxitin was 16% of BFG strains. All tested strains as well as BFG and C. difficile were susceptible to metronidazole. Was observed reduced leve (EUCAST) of susceptibility of C. difficile strains to vancomycin (13%). CONCLUSIONS. Increasing resistance to various antimicrobial agents is a significant problem in Poland. This demonstrate the need to continue with antibiotic resistance testing and surveys in anaerobic bacteria.

[Assessment of the usefulness of chromogenic medium for the isolation of Clostridium difficile from the gastrointestinal tract of children with diarrhoea]. / Ocena przydatnosci podloza chromogennego do izolacji Clostridium difficile z przewodu pokarmowego dzieci z biegunka.

INTRODUCTION: Clostridium difficile is well known as an important cause of nosocomial infection. Laboratory diagnostics have included bacterial culture or more commonly, direct detection of preformed toxin in stool samples using different assays. The aim of this study was to evaluate and compare two selecitve media to isolation of C. difficile from paediatric diarrhoeal stool samples. METHODS: Fifty nine stool samples, collected from 43 children with diarrhoea, were examined for routine laboratory diagnosis of C. difficile infection. Commercially available tests for detection of A/B toxins of C. difficile were performed. The same stool samples were cultured on two selective media for strain isolation: CLO and CDIFF (bioMerieux S.A., France) and incubated 48h and 24h respectively. RESULTS: Twenty two samples gave positive results for toxins A/B C. difficile. From 24 samples inoculated on selective media C. difficile strains were cultured: from 8 samples on CLO medium and from 16 samples on CDIFF medium. CONCLUSIONS: CDIFF medium is more effective for isolation of C. difficile strains from stool samples collected from children with diarrhoea.

[Prevalence of Clostridium difficile in the gastrointestinal tract of hospitalized children under two years of age]. / Wystepowanie Clostridium difficile w przewodzie pokarmowym hospitalizowanych dzieci do drugiego roku zycia.

The aim of this study was to determine prevalence of C. difficile in the gastrointestinal tract of hospitalized children under two years of age and a comparison of phenotypic and genotypic features. Hundred and seventy-eight samples collected from the faecal samples of children aged 2 months to 2 years, hospitalized in 2003-2006 were examined for the presence of toxin A/B of C. difficile. Toxigenicity of strains was confirmed using PCR. Susceptibility to antimicrobials was determined using E-test. The percentage of children infected with C. difficile was 68.6%. Toxigenic of C. difficile strains A+B+CDT- accounted for 35%, A-B+CDT- 10%, and 5% were strains of A+B+CDT+. 50% of the cultivated strains were non-toxigenic. The percentage of strains resistant to erythromycin and clindamycin was respectively 52% and 42%. Resistance to ciprofloxacin was widespread concern 98% of strains, and the moxifloxacin and gatifloxacin was 8%. The percentage of resistant strains to imipenem was 50%. All tested strains were susceptible to metronidazole and vancomycin.

First isolation of Clostridium difficile PCR-ribotype 027/toxinotype III in Poland.

Of 175 Clostridium difficile strains isolated from patient hospitalized in one academic hospital in Warsaw between 2005-2006, one isolate belonged to PCR-ribotype 027/toxinotype III. This isolate had tcdA, tcdB, binary toxin genes (cdtA and cdtB), a 18-bp deletion and a 1 bp deletion at 117 position in the tcdC gene. Antimicrobial susceptibility tests revealed high level resistance to erythromycin, moxifloxacin and gatifloxacin. This is a first report of the 027 strain of C. difficile in Poland.

Prevalence of Clostridium difficile infection in hospitalized patients with diarrhoea: Results of a Polish multicenter, prospective, biannual point-prevalence study.

PURPOSE: We aimed to measure the underdiagnosis of Clostridium difficile infection across Poland and the distribution of PCR-ribotypes of C. difficile. MATERIAL AND METHODS: Twenty seven Polish healthcare facilities (HCFs) participated in this prospective study. Each HCF systematically sent all diarrhoeal stools received from inpatients at their laboratories on two days (one in January 2013 and one in July 2013), independently of CDI test request, to the National Coordinating Laboratory (NCL) for standardized testing of CDI. Positive samples (using two-stage algorithm), had CDI, confirmed by qPCR and toxigenic culture. C. difficile isolates were characterized by PCR-ribotyping. Hospitals were questioned about their methods and testing policy for CDI during the study period: September 2011 to August 2013. RESULTS: During the study period, participating hospitals reported a mean of 33.2 tests for CDI per 10 000 patient-days and a mean of 8.4 cases of CDI per 10 000 patient-days. The overall prevalence of positive CDI patients at NCL was 16.5%. Due to absence of clinical suspicion, 19.1% of these patients were not diagnosed by the local diagnostic laboratory. We identified 23 different PCR-ribotypes among 87C. difficile strains isolated from patients. PCR-ribotype 027 (48%) was the most prevalent. CONCLUSIONS: The incidence of CDI in Poland in study period was very high. It should be noted however, that there is a lack of clinical suspicion and underestimation of the need to perform diagnostic tests for CDI in hospitalized patients. This will have an impact on the reported epidemiological status of CDI in Poland.

Laboratory diagnosis of antibiotic-associated diarrhea: a Polish pilot study into the clinical relevance of Clostridium difficile and Clostridium perfringens toxins.

In the present study, we investigated the prevalence of the Clostridium perfringens enterotoxin (CPEnt) in stool samples originally submitted for detection of Clostridium difficile toxins. Fifty-two fecal samples from inpatients were screened simultaneously for C. difficile and C. perfringens toxins: 75% of the specimens were positive for TcdA/TcdB toxins, 40% were positive for CPEnt, and 31% gave positive test results for both. It is interesting to note that only a relatively small number of C. perfringens isolates were positive for the cpe gene. All C. difficile strains were susceptible to metronidazole, but intermediate metronidazole resistance was documented for the C. perfringens isolates, which decreased upon in vitro passaging in the absence of metronidazole. We recommend that CPEnt detection should be included when diagnosing patients with presumed antibiotic-associated diarrhea.

[Survey of susceptibility of clinical Clostridium diffiicile strains isolated from patients hospitalised in different departments of paediatric hospital to antimicrobial agents]. / Przeglad wrazliwosci klinicznych szczepów Clostridium difficile izolowanych z przewodu pokarmowego dzieci hospitalizowanych w oddzialach pediatrycznych w warszawie na leki stosowane w lecznictwie szpitalnym.

This study was performed to determine the susceptibility of 50 C. difficile strains isolated from faecal samples of children suspected to antibiotic associated diarrhea (AAD) to antimicrobial agents: metronidazole, vancomycin, erythromycin, clindamycin, ciprofloxacin, moxifloksacin, gatifloksacin and imipenem. The all C. difficile strains were sensitived to metronidazole and vancomycin. Twenty six per cent of strains were resistant to erythromycin and clindamycin (MLS(B) type resistance). Resitance to ciprofloxacin, moxifloxacin, gatifloxacin and imipenem was detected in 98%, 8%, 8% and 30% of C. difficile strains, respectively.

Prevalence and association of PCR ribotypes of Clostridium difficile isolated from symptomatic patients from Warsaw with macrolide-lincosamide-streptogramin B (MLSB) type resistance.

Isolates (79 in total) of Clostridium difficile obtained over a 2 year period from 785 patients suspected of having C. difficile-associated diarrhoea (CDAD) and being hospitalized in the University Hospital in Warsaw were characterized by toxigenicity profile and PCR ribotyping. Furthermore, their susceptibility to clindamycin and erythromycin was determined. Among the 79 C. difficile isolates, 35 were classified as (A+)B+, 1 as (A+)(B+)CDT+, 36 as (A-)B+ and 7 as (A-)B-. A total of 21 different PCR ribotypes was detected. Two main (A+)B+ strains circulated in our hospital: ribotype 014 and ribotype 046. Unexpectedly, the predominant PCR ribotype was type 017, a known (A-)B+ strain, and this accounted for about 45.5 % of all isolates cultured from patients with CDAD. Isolates belonging to PCR ribotype 017 were found in cases from epidemics of antibiotic-associated diarrhoea in the internal and surgery units. High-level resistance (MIC > or = 256 mg l(-1)) to clindamycin and erythromycin was found in 39 (49 %) of the C. difficile isolates. Interestingly, 34 (94 %) of macrolide-lincosamide-streptogramin B (MLSB) type resistance strains did not produce toxin A, but produced toxin B and were (A-)B+ ribotype 017. Thirty-seven of the high-level resistance strains harboured the erythromycin-resistance methylase gene (ermB). C. difficile isolates (2/29) that had high-level clindamycin and erythromycin resistance, and belonged to PCR ribotype 046, were ermB negative. These investigations revealed that the predominant C. difficile strain isolated from symptomatic patients hospitalized in University Hospital in Warsaw was MLSB-positive clindamycin/erythromycin-resistant PCR ribotype 017.

[Influence of selected Lactobacillus sp. on Clostridium difficile strains with different toxigenicity profile]. / Wplyw wybranych gatunków paleczek Lactobacillus sp. na szczepy Clostridium difficile o róznym profilu toksynotwórczosci.

This study was performed for determination of antagonistic activity of Lactobacillus spp. (L. plantarum 2017405, L. rhamnosus GG, L. acidophilus DSM 21007 and L. fernmentumn 353) on Clostridiunl difficile strains belonging to different toxigenicity profiles. Forty strains C. difficile isolated from patients suffering from antibiotic associated diarrhea (AAD) were used. Among C. difficile strains 13 produced toxin A and B (A+B+CDT-), 14 produced only toxin B (AB'CDT), 9 produced toxins A and B and possessing of binary toxin genes (A+B+CDT-) and 4 were non-toxigenic (A-B-CDT-). We did not observe relationship between degree of antagonistic activity Lactobacillus spp. and profile of toxigenicity of C. difficile strains.

Detection of binary-toxin genes (cdtA and cdtB) among Clostridium difficile strains isolated from patients with C. difficile-associated diarrhoea (CDAD) in Poland.

Clostridium difficile A+ B+ and A- B+ strains isolated from stool samples of patients with C. difficile-associated diarrhoea (CDAD) were selected from the University Hospital Warsaw collection. The binary-toxin genes cdtA and cdtB were detected by PCR in five of the 41 A+ B+ strains tested, but in none of the 17 A- B+ strains tested, giving 8.6 % prevalence (5/58) of binary-toxin-positive strains. All of the strains that were positive for binary-toxin genes were grouped into toxinotype IV, suggesting that in this institution toxinotype IV might dominate among the population of C. difficile with binary-toxin genes.

[Profile of toxigenicity of Clostridium difficile strains isolated from paediatric patients with clinical diagnosis of antibiotic associated diarrhea (AAD)]. / Profil toksynotwórczosci szczepów Clostridium difficile izolowanych z przewodu pokarmowego dzieci z klinicznym rozpoznaniem biegunki poantybiotykowej.

This study was performed to determine profile of toxigenicity of 18 Clostridium difficile strains isolated from paeditric patients suffering from antibiotic associated diarrhea (AAD). Toxigenicity of C. difficile strains was tested for detection toxin A and toxin B by phenotypic methods and for detection of the tcdA and tcdB genes using of PCR. Changes in the repeating regions of the tcdA genes were detected with the NK9/NKV011 primer pairs. For detection of binary toxin (CDT) cdtA and cdtB genes, cdtApos/cdtArev i cdtBpos/cdtBrev two pair primers in PCR was used. Among C. difficile strains was detected three profiles of toxigenicity: C. difficile strains possesing of tcdA and tcdB genes but not possesing cdtA and cdtB genes of binary toxin (A+B+CDT-), strains possesing tcdA and tcdB and cdtA and cdtB genes (A+B+CDT+), strains with deletion of toxin A gene (A-B+CDT-). This is the first report on the occurence of binary positive C. difficile strains isolated from paediatric patients.