In Slovakia, dairy products made from ewes' milk have a long tradition. These products include the lactic acid product called "žinčica" which is a by-product occurring during the preparation of ewes' lump cheese. There is no information in the literature regarding the special properties of the microbiota, especially lactic acid Firmicutes, which can survive in "žinčica." From the safety aspect, enterococci are a controversial group of bacteria, and those from "žinčica" have never been tested for their properties. The "žinčica" used in our study was supplied by several different agrofarms producing ewes' lump cheese in central Slovakia. The species Enterococcus faecium (strains EF30E1, EF32E1, EF34E1, EF34E5) and Enterococcus faecalis (strains EE30E4, EE35E1, E31E2, altogether 7) were detected in samples from "žinčica" identified using MALDI-TOF spectrometry with secure genus identification/probable species identification and then confirmed by means of PCR. Enterococci were hemolysis-negative and the genes of the typical enterococcal virulence factors were mostly absent; the gelE gene was found in two E. faecium strains (EF30E1 and EF32E1), the agg gene was detected in E. faecalis EE35E1, and the esp gene was found in two E. faecalis strains (EE30E4 and EE31E2). No strains harbored the cytolysin A gene. Biofilm formation was detected in four strains (EF30E1, EF32E1, EF34E1, and EF34E5), indicating highly positive and low-grade positive biofilm formation. Enterococci were mostly susceptible to antibiotics tested for their phenotype. This is the first study to analyze enterococci in "žinčica."

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální proteiny genetika   MeSH
• bezpečnost potravin * MeSH
• biofilmy růst a vývoj   MeSH
• Enterococcus faecalis účinky léků   genetika   MeSH
• Enterococcus faecium účinky léků   genetika   MeSH
• Enterococcus klasifikace   účinky léků   patogenita   MeSH
• faktory virulence genetika   MeSH
• mikrobiální testy citlivosti MeSH
• mikrobiota MeSH
• ovce MeSH
• potravinářská mikrobiologie * MeSH
• sýr mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Slovenská republika MeSH

Enterococcus faecalis is a Gram-positive facultative anaerobe involved in many fatal or refractory infections of humans. Silver, often used as silver ions (Ag+) or nanoparticles (AgNPs), is a strong and broad-spectrum antibacterial agent, but E. faecalis shows resistance against it. Despite this, the knowledge about the resistance of E. faecalis against silver is still lacking. In this study, the silver-resistant E. faecalis strains (AgR and ANR E. faecalis) were established through a serial selection method. Their biological and silver-resistant features as well as the Gene Ontology (GO) in comparison with the original E. faecalis were evaluated. The results showed that the silver-resistant E. faecalis could proliferate as original bacteria and had strong resistance against both Ag+ and AgNPs. The minimum bactericidal concentrations (MBCs) of AgNO3 on original, AgR, and ANR E. faecalis were 400 mg/L, 600 mg/L, and 500 mg/L, and the MBCs of AgNPs on these strains were 80 mg/L, 110 mg/L, and 130 mg/L, respectively. GO analysis revealed significant difference (P < 0.05) in gene expressions of biological process (BP), cellular component (CC), and molecular function (MF) among original, AgR, and ANR E. faecalis. These findings provided a significant basis for further understanding and managing the silver-resistance of E. faecalis in infection-control environments. The mechanism behind Ag+/AgNPs resistance of E. faecalis needs to be further investigated.

• MeSH
• antibakteriální látky chemie   farmakologie   MeSH
• bakteriální léková rezistence účinky léků   genetika   MeSH
• bakteriální proteiny genetika   MeSH
• dusičnan stříbrný metabolismus   farmakologie   MeSH
• Enterococcus faecalis účinky léků   genetika   růst a vývoj   fyziologie   MeSH
• kovové nanočástice chemie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• regulace genové exprese u bakterií MeSH
• stříbro chemie   farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Vancomycin-resistant enterococci (VRE) are nosocomial pathogens of increasing medical importance. This study involved 121 VRE selectively obtained from a representative set of 1464 samples collected from various sources in the north-eastern part of the Czech Republic. In total, 119 VRE belonged to Enterococcus faecium and two to Enterococcus faecalis. All isolates of E. faecium were resistant to at least three antibiotic classes. The resistance genes vanA, erm(B), tet(M), tet(L), aac(3)-IIIa and aac(6')-aph(2'') were detected. We assigned the E. faecium to sequence types ST5, ST18, ST38, ST64, ST92, ST273, ST549 and ST640. In E. faecium isolates, we identified the presence of replicases rep20pLG1 , rep2pRE25 , rep17pRUM , rep21pVEF1/2 and rep14pRI1 , as well as relaxases relpEF1 , relpLG1 , relpCIZ2 , relpRE25 and relpRUM . The presence of the toxin-antitoxin system axe-txe was detected mainly among isolates of hospital origin. The A and D types of transposon Tn1546 were those occurring most frequently. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first extensive study of vancomycin-resistant enterococci of diverse origin in a single well-defined area of the Czech Republic. The isolates were investigated for their antibiotic resistance, epidemiological characteristics and plasmid characteristics. Based on the results obtained, we can make assumptions as to the ways that vancomycin resistance is disseminated throughout the environment including humans and animals.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální proteiny genetika   MeSH
• Enterococcus faecalis účinky léků   genetika   izolace a purifikace   MeSH
• Enterococcus faecium účinky léků   genetika   izolace a purifikace   MeSH
• enterokoky rezistentní vůči vankomycinu klasifikace   genetika   izolace a purifikace   MeSH
• grampozitivní bakteriální infekce epidemiologie   mikrobiologie   MeSH
• lidé MeSH
• plazmidy genetika   MeSH
• rezistence na vankomycin genetika   MeSH
• systémy toxin-antitoxin genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH

• MeSH
• bakteriální léková rezistence genetika   imunologie   MeSH
• Enterococcus faecalis genetika   účinky léků   MeSH
• Enterococcus faecium genetika   účinky léků   MeSH
• enterokoky rezistentní vůči vankomycinu * genetika   patogenita   účinky léků   MeSH
• fenotyp MeSH
• genotyp MeSH
• glykopeptidy farmakologie   metabolismus   terapeutické užití   MeSH
• infekce spojené se zdravotní péčí mikrobiologie   prevence a kontrola   MeSH
• lidé MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH
• přehledy MeSH

The genes encoding aminoglycoside resistance in Enterococcus faecalis may promote collateral aminoglycoside resistance in polymicrobial wounds. We studied a total of 100 diabetic foot ulcer samples for infection and found 60 samples to be polymicrobial, 5 to be monomicrobial, and 35 samples to be culture negative. A total of 65 E. faecalis isolates were screened for six genes coding for aminoglycoside resistance, antibiotic resistance patterns, and biofilm production. Infectious Diseases Society of America/International Working Group on the Diabetic Foot system was used to classify the wound ulcers. Majority of the subjects with culture-positive wound were recommended conservative management, while 14 subjects underwent amputation. Enterococcal isolates showed higher resistance for erythromycin, tetracycline, and ciprofloxacin. Isolates from grade 3 ulcer showed higher frequency of aac(6')-Ie-aph(2″)-Ia, while all the isolates were negative for aph(2″)-Ib, aph(2″)-Ic, and aph(2″)-Id. The isolates from grade 3 ulcers showed higher resistance to aminoglycosides as well as teicoplanin and chloramphenicol. All the 39 biofilm producers were obtained from polymicrobial wound and showed higher resistance when compared to biofilm non-producers. Higher frequency of isolates carrying aac(6')-Ie-aph(2″)-Ia in polymicrobial community showing resistance to key antibiotics suggests widespread distribution of aminoglycoside-resistant E. faecalis and their role in worsening diabetic foot ulcers.

• MeSH
• bakteriální léková rezistence * MeSH
• diabetická noha mikrobiologie   MeSH
• dospělí MeSH
• Enterococcus faecalis klasifikace   účinky léků   genetika   izolace a purifikace   MeSH
• grampozitivní bakteriální infekce mikrobiologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

To evaluate the microbiological safety of tilmicosin on human intestinal microflora, four chemostat models of healthy human colonic ecosystems were exposed to tilmicosin (0, 0.436, 4.36, and 43.6 μg/mL) for 7 days. Prior to and during drug exposure, three microbiological endpoints were monitored daily including short-chain fatty acids, bacterial counts and macrolide susceptibility. Colonization resistance of each community was determined by 3 successive daily challenges of Salmonella typhimurium. Genes associated with virulence and macrolide resistance in Enterococcus faecalis were determined by PCR. Transcriptional expression of the virulence gene (gelE) in E. faecalis was determined by real-time RT-PCR. Our results showed that different concentrations of tilmicosin did not disrupt the colonization resistance in each chemostat. During exposure to 4.36 and 43.6 μg/mL tilmicosin, the Bacteroides fragilis population was significantly decreased while the proportion of resistant Enterococci increased. After long-term exposure to the highest concentration (43.6 μg/mL) of tilmicosin, the gelE gene was significantly up-regulated in the high-level macrolide resistant strains that also contained the ermB resistance gene. This study was the first of its kind to evaluate the microbiological toxicity of tilmicosin using a chemostat model. These findings also provide new insight into the co-occurrence of macrolide resistance and virulence in E. faecalis under tilmicosin selective pressure.

• MeSH
• antibakteriální látky škodlivé účinky   MeSH
• Bacteroides fragilis účinky léků   genetika   MeSH
• bakteriální geny genetika   MeSH
• Enterococcus faecalis účinky léků   genetika   MeSH
• feces mikrobiologie   MeSH
• kolon mikrobiologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti metody   MeSH
• Salmonella typhimurium účinky léků   genetika   MeSH
• střevní mikroflóra účinky léků   genetika   MeSH
• tylosin škodlivé účinky   analogy a deriváty   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

UNLABELLED: The bacterial stringent response (SR) is a conserved stress tolerance mechanism that orchestrates physiological alterations to enhance cell survival. This response is mediated by the intracellular accumulation of the alarmones pppGpp and ppGpp, collectively called (p)ppGpp. In Enterococcus faecalis, (p)ppGpp metabolism is carried out by the bifunctional synthetase/hydrolase E. faecalis Rel (RelEf) and the small alarmone synthetase (SAS) RelQEf. Although Rel is the main enzyme responsible for SR activation in Firmicutes, there is emerging evidence that SASs can make important contributions to bacterial homeostasis. Here, we showed that RelQEf synthesizes ppGpp more efficiently than pppGpp without the need for ribosomes, tRNA, or mRNA. In addition to (p)ppGpp synthesis from GDP and GTP, RelQEf also efficiently utilized GMP to form GMP 3'-diphosphate (pGpp). Based on this observation, we sought to determine if pGpp exerts regulatory effects on cellular processes affected by (p)ppGpp. We found that pGpp, like (p)ppGpp, strongly inhibits the activity of E. faecalis enzymes involved in GTP biosynthesis and, to a lesser extent, transcription of rrnB by Escherichia coli RNA polymerase. Activation of E. coli RelA synthetase activity was observed in the presence of both pGpp and ppGpp, while RelQEf was activated only by ppGpp. Furthermore, enzymatic activity of RelQEf is insensitive to relacin, a (p)ppGpp analog developed as an inhibitor of "long" RelA/SpoT homolog (RSH) enzymes. We conclude that pGpp can likely function as a bacterial alarmone with target-specific regulatory effects that are similar to what has been observed for (p)ppGpp. IMPORTANCE: Accumulation of the nucleotide second messengers (p)ppGpp in bacteria is an important signal regulating genetic and physiological networks contributing to stress tolerance, antibiotic persistence, and virulence. Understanding the function and regulation of the enzymes involved in (p)ppGpp turnover is therefore critical for designing strategies to eliminate the protective effects of this molecule. While characterizing the (p)ppGpp synthetase RelQ of Enterococcus faecalis (RelQEf), we found that, in addition to (p)ppGpp, RelQEf is an efficient producer of pGpp (GMP 3'-diphosphate). In vitro analysis revealed that pGpp exerts complex, target-specific effects on processes known to be modulated by (p)ppGpp. These findings provide a new regulatory feature of RelQEf and suggest that pGpp may represent a new member of the (pp)pGpp family of alarmones.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• deoxyguanosin analogy a deriváty   biosyntéza   chemie   MeSH
• dipeptidy biosyntéza   chemie   MeSH
• Enterococcus faecalis účinky léků   enzymologie   genetika   metabolismus   MeSH
• fyziologický stres MeSH
• guanosindifosfát metabolismus   MeSH
• guanosinpentafosfát metabolismus   MeSH
• guanosintetrafosfát biosyntéza   MeSH
• guanosintrifosfát metabolismus   MeSH
• hořčík MeSH
• ligasy genetika   metabolismus   MeSH
• molekulární struktura MeSH
• regulace genové exprese enzymů MeSH
• regulace genové exprese u bakterií MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

We studied the vanA-carrying vancomycin-resistant enterococci (VRE) isolated from American crows in the United States during the winter 2011/2012. Faecal samples from crows were cultured selectively for VRE and characterized. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were used to examine epidemiological relationships of vanA-containing VRE. Isolates were tested in vitro for their ability to horizontally transfer the vancomycin resistance trait. VRE with the vanA gene were found in 15 (2.5%) of 590 crows samples, from which we obtained 22 different isolates. Enterococcal species were Enterococcus faecium (14) and E. faecalis (8). One, two and 19 isolates originated from Kansas, New York State and Massachusetts, respectively. Based on MLST analysis, E. faecium isolates were grouped as ST18 (6 isolates), ST555 (2), and novel types ST749 (1), ST750 (3), ST751 (1), ST752 (1). Enterococcus faecalis isolates belonged to ST6 (1), ST16 (3) and ST179 (4). All isolates were able to transfer the vancomycin resistance trait via filter mating with very high transfer range. Clinically important enterococci with the vanA gene occur in faeces of wild American crows throughout the United States. These migrating birds may contribute to the dissemination of VRE in environment over large distances. [Correction added after first online publication on 06 August 2013: The number of E. faecium ST752 isolate is now amended to '1', consistent with that shown in the 'Results' section and Figure 2.].

• MeSH
• bakteriální proteiny genetika   MeSH
• Enterococcus faecalis genetika   izolace a purifikace   MeSH
• Enterococcus faecium genetika   izolace a purifikace   MeSH
• feces mikrobiologie   MeSH
• ligasy tvořící vazby C-O genetika   MeSH
• multilokusová sekvenční typizace MeSH
• přenos genů horizontální MeSH
• rezistence na vankomycin genetika   MeSH
• vrány mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Spojené státy americké MeSH

The influence of specific and non-specific antibiotic pressure on in vivo spread of macrolide-lincosamide-streptogramin B (MLSB) resistance was evaluated in this study. Chickens repeatedly inoculated with Enterococcus faecalis harbouring the plasmid pAMβ1 carrying the erm(B) gene were perorally treated for one week with tylosin, lincomycin (both specific antibiotic pressure) and chlortetracycline (non-specific antibiotic pressure). Antibiotic non-treated but E. faecalis inoculated chickens served as a control. To quantify the erm(B) gene and characterise intestinal microflora, faecal DNA was analysed by qPCR and 454-pyrosequencing. Under the pressure of antibiotics, a significant increase in erm(B) was observed by qPCR. However, at the final stage of the experiment, an increase in erm(B) was also observed in two out of five non-treated chickens. In chickens treated with tylosin and chlortetracycline, the increase in erm(B) was accompanied by an increase in enterococci. However, E. faecalis was at the limit of detection in all animals. This suggests that the erm(B) gene spread among the gut microbiota other than E. faecalis. Pyrosequencing results indicated that, depending on the particular antibiotic pressure, different bacteria could be responsible for the spread of MLSB resistance. Different species of MLSB-resistant enterococci and streptococci were isolated from cloacal swabs during and after the treatment. PFGE analysis of MLSB-resistant enterococci revealed four clones, all differing from the challenge strain. All of the MLSB-resistant isolates harboured a plasmid of the same size as pAMβ1. This study has shown that MLSB resistance may spread within the gut microbiota under specific and non-specific pressure and even in the absence of any antimicrobial pressure. Finally, depending on the particular antibiotic pressure, different bacterial species seems to be involved in the spread of MLSB resistance.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální léková rezistence genetika   MeSH
• chlortetracyklin farmakologie   MeSH
• DNA primery genetika   MeSH
• druhová specificita MeSH
• Enterococcus faecalis účinky léků   genetika   MeSH
• feces mikrobiologie   MeSH
• grampozitivní bakteriální infekce veterinární   MeSH
• kur domácí * MeSH
• linkosamidy farmakologie   MeSH
• makrolidy farmakologie   MeSH
• methyltransferasy genetika   MeSH
• mikrobiální testy citlivosti MeSH
• molekulární sekvence - údaje MeSH
• nemoci drůbeže mikrobiologie   MeSH
• neparametrická statistika MeSH
• plazmidy genetika   MeSH
• polymerázová řetězová reakce veterinární   MeSH
• přenos genů horizontální genetika   MeSH
• pulzní gelová elektroforéza veterinární   MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• streptogramin B farmakologie   MeSH
• tylosin farmakologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Identification of vaginal fluids is an important step in the process of sexual assaults confirmation. Advances in both microbiology and molecular biology defined technical approaches allowing the discrimination of body fluids. These protocols are based on the identification of specific bacterial communities by microfloraDNA (mfDNA) amplification. A multiplex real time-PCR assay (ForFLUID kit) has been developed for identifying biological fluids and for discrimination among vaginal, oral and fecal samples. In order to test its efficacy and reliability of the assay in the identification of vaginal fluids, an interlaboratory evaluation has been performed on homogeneous vaginal swabs. All the involved laboratories were able to correctly recognize all the vaginal swabs, and no false positives were identified when the assay was applied on non-vaginal samples. The assay represents an useful molecular tool that can be easily adopted by forensic geneticists involved in vaginal fluid identification.

• MeSH
• cervikální hlen mikrobiologie   MeSH
• DNA bakterií genetika   MeSH
• dospělí MeSH
• Enterococcus faecalis genetika   izolace a purifikace   MeSH
• kvantitativní polymerázová řetězová reakce * MeSH
• laboratoře normy   MeSH
• Lactobacillus genetika   izolace a purifikace   MeSH
• lidé MeSH
• multiplexová polymerázová řetězová reakce * MeSH
• soudní lékařství normy   MeSH
• Staphylococcus genetika   izolace a purifikace   MeSH
• Streptococcus genetika   izolace a purifikace   MeSH
• vagina mikrobiologie   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH