醋酸钙复盐对微生物生长的影响

应用四醋酸钙对黑曲霉、黄曲霉、米曲霉、链格孢霉等霉菌和大肠杆菌、枯草杆菌、绿脓杆菌、金黄色葡萄球菌等细菌生长影响作了研究,并将四醋酸钙与醋酸钙、苯甲酸钠对上述微生物的生长抑制作用加以比较。0.4％四醋酸钙对上述细菌的生长有明显的抑制作用。对上述霉菌生长亦有一定的抑制作用。当其与苯甲酸钠协同使用时,能更明显地抑制微生物生长。实验结果表明四醋酸钙很可能是一种安全性高、成本低的新型食品防腐剂。     

Conversion of native lignin to a highly phenolic functional polymer and its separation from lignocellulosics

An original reaction system (the phase separative reaction system) has been designed for derivatizing native lignins to highly phenolic, functional polymers. This system is composed of a phenol derivative and concentrated acid, which are not miscible at room temperature. The key point of the lignin functionalization process, including the phase separative system, is that lignin and carbohdrates, which are totally different in structures and reactivitie, are modified individually in the different phases: lignin is present in the organic phase and carbohydrates in the aqueous phase. Through the process, lignin was modified selectively at Calpha-positions of side chains, the most reactive sites, to give highly phenolic, light-colored, diphenylmethane-type materials which still retained original interunit linkages formed by the dehydrogenative polymerization during the biosynthesis. The carbohydrates were swollen, followed by partial hydrolysis and dissolution in the acid solution, resulting in the perfect decomposition of interpenetrating polymer network structures in the cell wall. Therefore, the functionalization of lignin and the separation of resulting lignin from carbohydrates were quickly achieved at room temperature, independent of wood species. This process would be a powerful tool for estimating strutures and reactivities of lignins as well as the functionalization of lignins, because of the selective structural modifications. (c) 1995 John Wiley & Sons, Inc.     

Synthesis of an artificial glycoconjugate polymer carrying Pk-antigenic trisaccharide and its potent neutralization activity against Shiga-like toxin.

Fluorescence-labeled glycoconjugate polymers carrying carbohydrate segments of a globotriaosyl ceramide (Gb3) were synthesized and subjected to biological assays using Escherichia coli O-157 strains and Shiga-like toxins (Stx-I and Stx-II). For the fluorescence labeling, a new polymerizable fluorescent monomer with a TBMB carbonyl chromophore (Ex. 325 nm, Em. 410 nm) was designed. A glycosyl monomer of the trisaccharide segment of Gb3 was prepared from p-nitrophenyl beta-lactoside and copolymerized with acrylamide and the fluorescent monomer to prepare a fluorescence-labeled glycoconjugate copolymer carrying [alpha-D-galactopyranosyl-(1-->4)-beta-D-galactopyranosyl]-(1-->4)-beta- D-glucopyranoside. The polymer showed potent neutralization activity against Stx-I and also binding activity onto E. coli O-157 strains.     

Superquenching as a detector for microsphere-based flow cytometric assays.

BACKGROUND: Fluorescent conjugated polymers display high fluorescence quantum yields and enhanced sensitivity to quenching (superquenching) by oppositely charged quenchers through energy or electron transfer. Fluorescent polymers and their quenchers are used in bead-based biosensor applications where the polymers are coated on particles. In this work, we investigate a detection method that utilizes superquenching on microspheres, which can be used for flow cytometric assays. METHODS: Microspheres were coated with the fluorescent cationic polyelectrolyte poly(p-phenylene-ethynylene) (PPE), and its superquenching by 9,10-anthraquinone-2,6-disulfonic acid (AQS) was examined by fluorometric methods in presence and in absence of a barrier to superquenching in the form of an anionic lipid bilayer. RESULTS: Flow cytometry detected superquenching of PPE on microspheres (MS-PPE) by AQS where high levels of reduction in fluorescence were observed. Adding different concentrations of AQS to MS-PPE yielded a Stern-Volmer quenching constant of 0.8x10(6) M-1. While forming an anionic lipid bilayer around the MS-PPE acted as a barrier to superquenching by AQS, disrupting the lipid bilayer allowed superquenching to take place. CONCLUSIONS: The sensitivity of flow cytometry in detecting fluorescence of microspheres and the amplified quenching sensitivity of fluorescent conjugated polymers both offer advantages over other fluorometric methods and conventional quenching detection. This study used superquenching of fluorescent polymers as a new tool in flow cytometry, thus combining the advantages offered by both method and detector. In addition, we employed the formation and the disruption of a supported lipid bilayer in mediating superquenching to offer new biosensing applications.     

Affibody-attached hyperbranched conjugated polyelectrolyte for targeted fluorescence imaging of HER2-positive cancer cell

A hyperbranched conjugated polyelectrolyte (HCPE) with a core-shell structure is designed and synthesized via alkyne polycyclotrimerization and click chemistry. The HCPE has an emission maximum at 565 nm with a quantum yield of 12% and a large Stokes shift of 143 nm in water. By virtue of its poly(ethylene glycol) shell, this polymer naturally forms spherical nanoparticles that minimize nonspecific interaction with biomolecules in aqueous solution, consequently allowing for efficient bioconjugation with anti-HER2 affibody via carbodiimide-activated coupling reaction. The resulting affibody-attached HCPE can be utilized as a reliable fluorescent probe for targeted cellular imaging of HER2-overexpressed cancer cells such as SKBR-3. Considering its low cytotoxicity and good photostability, the HCPE nanoprobe holds great promise in practical imaging tasks. This study also provides a molecular engineering strategy to overcome the intrinsic limitations of traditional fluorescent polymers (e.g., chromophore-tethered polymers and linear conjugated polyelectrolytes) for bioconjugation and applications.     

Characterization and conservation of the inner E2 core domain structure of branched-chain alpha-keto acid dehydrogenase complex from bovine liver. Construction of a cDNA encoding the entire transacylase (E2b) precursor

A cDNA clone encoding the entire transacylase (E2b) precursor of the bovine branched-chain alpha-keto acid dehydrogenase complex has been constructed from two overlapping incomplete cDNA clones which were isolated from a lambda ZAP library prepared from bovine liver poly(A)+ RNA. Nucleotide sequencing indicates that this bovine E2b cDNA insert (bE2-11) is 2701 base pairs in length with an open reading frame of 1446 base pairs. The bE2-11 cDNA insert encodes a leader peptide of 61 residues and a mature E2b polypeptide of 421 amino acid residues with a calculated monomeric molecular mass of 46,518 daltons. The molecular mass of the native E2b component isolated from bovine liver is 1,110,000 daltons as determined by sedimentation equilibrium. This value establishes the 24-subunit octahedral model for the quaternary structure of bovine E2b. The amino-terminal sequences of two tryptic fragments (A and B) of the E2b protein have been determined. Fragment A comprises residues 175 to 421 of the E2b protein and is the inner E2 core domain which contains the transacylase active site. Fragment B, produced by further tryptic cleavage of fragment, comprises residues 205 to 421, but does not have transacylase activity. Both fragments A and B confer the highly assembled 24-mer structure. The primary structure of the inner E2 core domain of bovine E2b (fragment A) is very similar to those of three other E2 proteins (human E2p, Escherichia coli E2p, and E. coli E2k). These similarities suggest that these E2 proteins are structurally and evolutionarily related.     

Designing 1,5‐Naphthyridine‐2,6‐dione‐Based Conjugated Polymers for Higher Crystallinity and Enhanced Light Absorption to Achieve 9.63% Efficiency Polymer Solar Cells

Highly crystalline conjugated polymers represent a key material for producing high‐performance thick‐active‐layer polymer solar cells (PSCs). However, despite their potential, a limited number of crystalline polymers are used in PSCs because of the lack of highly coplanar acceptor building blocks and insufficient light absorptivity (α < 10⁵) of most donor (D)–acceptor (A)‐type polymers. This study reports a series of novel 3,7‐di(thiophen‐2‐yl)‐1,5‐naphthyridine‐2,6‐dione (NTDT) acceptor‐based conjugated polymers, PNTDT‐2T, PNTDT‐TT, and PNTDT‐2F2T, synthesized with 2,2′‐bithiophene (2T), thieno[3,2‐b]thiophene (TT), and 3,3′‐difluoro‐2,2′‐bithiophene (2F2T) donor units, respectively. PNTDT‐2F2T exhibits superior polymer crystallinity and a much higher absorption coefficient than those of PNTDT‐2T or PNTDT‐TT because of adequate matching between highly coplanar A (NTDT) and D (2F2T) building blocks. A bulk heterojunction solar cell based on PNTDT‐2F2T exhibits a power conversion efficiency of up to 9.63%, with a high short circuit current of 18.80 mA cm^?2 and fill factor of 0.70, when a thick active layer (>200 nm) is used, without postfabrication hot processing. The findings demonstrate that the polymer crystallinity and absorption coefficient can be effectively controlled by selecting appropriate D and A building blocks, and that NTDT is a novel and versatile A building block for highly efficient thick‐active‐layer PSCs.     

In vitro induction of IgM synthesis and secretion in rabbit spleen cells by soluble Concanavalin A

Rabbit spleen cells are not activated by Concanavalin A (Con A) conjugated to Sepharose 4B but are stimulated by soluble Con A which induces DNA and protein synthesis. At optimal concentration (5 μg/ml) one notes an increased intracellular protein and IgM synthesis and then secretion. This increase in protein synthesis is seen at all phases of the culture. At the intracellular level, IgM is found in the form of 7S molecules and a significant proportion of polymers with a centrifugation constant smaller than 19S. Fully assembled 19S polymers are found in the fluid phase. These results are compatible with a model of cellular cooperation, the basis of which is not the presentation of the inducer (mitogen or antigen) by a cell type to another, but rather the secretion of mediators by one of the cell populations making other cells responsive to the inducer.     

Effects of trehalose polycation end-group functionalization on plasmid DNA uptake and transfection

In this study, we have synthesized six analogs of a trehalose-pentaethylenehexamine glycopolymer (Tr4) that contain (1A) adamantane, (1B) carboxy, (1C) alkynyl-oligoethyleneamine, (1D) azido trehalose, (1E) octyl, or (1F) oligoethyleneamine end groups and evaluated the effects of polymer end group chemistry on the ability of these systems to bind, compact, and deliver pDNA to cultured HeLa cells. The polymers were synthesized in one-pot azide-alkyne cycloaddition reactions with an adaptation of the Carothers equation for step-growth polymerization to produce a series of polymers with similar degrees of polymerization. An excess of end-capping monomer was added at the end of the polymerizations to maximize functionalization efficiency, which was evaluated with GPC, NMR, and MALDI-TOF. The polymers were all found to bind and compact pDNA at similarly low N/P ratios and form polyplexes with plasmid DNA. The effects of the different end group structures were most evident in the polyplex internalization and transfection assays in the presence of serum as determined by flow cytometry and luciferase gene expression, respectively. The Tr4 polymers end-capped with carboxyl groups (1B) (N/P = 7), octyne (1E) (N/P = 7), and oligoethyleneamine (1F) (N/P = 7), were taken into cells as polyplex and exhibited the highest levels of fluorescence, resulting from labeled plasmid. Similarly, the polymers end-functionalized with carboxyl groups (1E at N/P = 7), octyl groups (1E at N/P = 15), and in particular oligoethyleneamine groups (1F at N/P = 15) yielded dramatically higher reporter gene expression in the presence of serum. This study yields insight into how very subtle structural changes in polymer chemistry, such as end groups can yield very significant differences in the biological delivery efficiency and transgene expression of polymers used for pDNA delivery.     

Fluorescence labeling method for estimation of soluble polymers in the living material

The highly fluorescent derivatives of fluorescein, bearing the aliphatic primary amino groups, N-(2-aminoethylcarbonyl)-5(6)-aminofluorescein and 5-[N′-(2-aminoethyl)thioureido]fluorescein, were prepared for labeling of soluble polymers. The absorption and emission properties of these labels and polymers labeled with them were compared with properties of fluorescein and fluorescein isothiocyanate (FITC)-bovine serum albumin conjugate. Effects of the chemical structure of the polymer on the relative fluorescence quantum yield of a covalently attached label were evaluated using ionogenic, olefinic, or phenolic groups in side chains. The fluorescence of labeled polymers was adequate for their tracing in all the cases studied. The most pronounced quenching of fluorescence in the presence of phenolic groups is comparable with the quenching of fluorescence of FITC observed in FITC-protein conjugates. The long-term stability of the polymer-fluorochrome bond was checked in solutions of pH 2.10, 7.46, and 11.84; a higher stability of simple amide over amide plus the thiourea bond was found. The quantitative method of measurement of the concentration of labeled polymers in the biological material in a range of about 10 ng was developed; factors affecting the reproducibility are discussed.     

Mechanism of enhancement of microbial cell hydrophobicity by cationic polymers.

Polycationic polymers have been noted for their effects in promoting cell adhesion to various surfaces, but previous studies have failed to describe a mechanism dealing with this type of adhesion. In the present study, three polycationic polymers (chitosan, poly-L-lysine, and lysozyme) were tested for their effects on microbial hydrophobicity, as determined by adhesion to hydrocarbon and polystyrene. Test strains (Escherichia coli, Candida albicans, and a nonhydrophobic mutant, MR-481, derived from Acinetobacter calcoaceticus RAG-1) were vortexed with hexadecane in the presence of the various polycations, and the extent of adhesion was measured turbidimetrically. Adhesion of all three test strains rose from near zero values to over 90% in the presence of low concentrations of chitosan (125 to 250 micrograms/ml). Adhesion occurred by adsorption of chitosan directly to the cell surface, since E. coli cells preincubated in the presence of the polymer were highly adherent, whereas hexadecane droplets pretreated with chitosan were subsequently unable to bind untreated cells. Inorganic cations (Na+, Mg2+) inhibited the chitosan-mediated adhesion of E. coli to hexadecane, presumably by interfering with the electrostatic interactions responsible for adsorption of the polymer to the bacterial surface. Chitosan similarly promoted E. coli adhesion to polystyrene at concentrations slightly higher than those which mediated adhesion to hexadecane. Poly-L-lysine also promoted microbial adhesion to hexadecane, although at concentrations somewhat higher than those observed for chitosan. In order to study the effect of the cationic protein lysozyme, adhesion was studied at 0 degree C (to prevent enzymatic activity), using n-octane as the test hydrocarbon. Adhesion of E. coli increased by 70% in the presence of 80 micrograms of lysozyme per ml. When the negatively charged carboxylate residues on the E. coli cell surface were substituted for positively charged ammonium groups, the resulting cells became highly hydrophobic, even in the absence of polycations. The observed "hydrophobicity" of the microbial cells in the presence of polycations is thus probably due to a loss of surface electronegativity. The data suggest that enhancement of hydrophobicity by polycationic polymers is a general phenomenon.     

The selective detection of cell surface determinants by means of antibodies and acetylated avidin attached to highly fluorescent polymer microspheres

Procedures are described for the synthesis of 500 A-diameter polymer microspheres containing a novel fluorescent cross-linking agent. These microspheres have very high fluorophore concentration without quenching of the fluorescence and show very low nonspecific interaction with cells. When monoclonal anti-Thy-1.2 is attached to the fluorescent microspheres, specific binding results in 10(4) spheres being attached per thymocyte while non-specific binding is less than 1%. Similar values are obtained for an indirect staining procedure. The high non-specific binding of cationic avidin to negative cell surfaces is shown to be decreased to negligible levels by acetylation of the amine groups of the protein without decreasing its high-affinity binding to biotin. The use of acetyl-avidin (pI = 6.7) directly, or when attached to fluorescent microspheres, resulted in a highly selective detection of biotinyl groups on the erythrocyte or lymphocyte cell surface. Attachment of biotinyl groups to the hinge carbohydrates of antibodies did not affect their specificity. It allowed their detection by means of microspheres-acetyl-avidin conjugates.     

FACTORS AFFECTING THE REDISTRIBUTION OF SURFACE-BOUND CONCANAVALIN A ON HUMAN POLYMORPHONUCLEAR LEUKOCYTES

Human neutrophil polymorphonuclear leukocytes (PMN) were studied to determine the influence of cellular locomotion upon the redistribution and capping of concanavalin A (Con A). Con A was detected by fluorescence (using Con A conjugated to fluorescein isothiocyanate [Con A-FITC]), or on shadow-cast replicas (using Busycon canaliculatum hemocyanin as a marker for Con A). After labeling with Con A 100 µg/ml at 4°C and warming to 37°C, locomotion occurred, and the Con A quickly aggregated into a cap at the trailing end of the cell. When locomotion was inhibited (with cytochalasin B, or by incubation in serum-free medium at 18°C) Con A rapidly formed a cap over the central region of the cell. Iodoacetamide inhibited capping. PMN labeled with FITC, a monovalent ligand, developed caps at the tail only on motile cells; FITC remained dispersed on immobilized cells. PMN exposed to Con A 100 µg/ml at 37°C bound more lectin than at 4°C, became immobilized, and showed slow central capping. The Con A soon became internalized to form a perinuclear ring. Such treatment in the presence of cytochalasin B resulted in the quick formation of persistent central caps. Colchicine (or prior cooling) protected PMN from the immobilizing effect of Con A, and tail caps were found on 30–40% of cells. Immobilization of colchicine-treated cells caused Con A to remain in dispersed clusters. Thus, capping on PMN is a temperature- and energy-dependent process that proceeds independently of cellular locomotion, provided a colchicine-sensitive system is intact and the ligand is capable of cross linking receptors. On the other hand, if the cell does move, it appears that ligands may be swept into a cap at the tail whether cross-linking occurs or not.     

Identification of the high affinity Mn2+ binding site of bacteriophage lambda phosphoprotein phosphatase: effects of metal ligand mutations on electron paramagnetic resonance spectra and phosphatase activities

Bacteriophage lambda phosphoprotein phosphatase (lambdaPP) has structural similarity to the mammalian Ser/Thr phosphoprotein phosphatases (PPPs) including the immunosuppressant drug target calcineurin. PPPs possess a conserved active site containing a dinuclear metal cluster, with metal ligands provided by a phosphoesterase motif plus two additional histidine residues at the C-terminus. Multiple sequence alignment of lambdaPP with 28 eubacterial and archeal phosphoesterases identified active site residues from the phosphoesterase motif and in many cases 2 additional C-terminal His metal ligands. Most highly similar to lambdaPP are E. coli PrpA and PrpB. Using the crystal structure of lambdaPP [Voegtli, W. C., et al. (2000) Biochemistry 39, 15365-15374] as a structural and active site model for PPPs and related bacterial phosphoesterases, we have studied mutant forms of lambdaPP reconstituted with Mn(2+) by electron paramagnetic resonance (EPR) spectroscopy, Mn(2+) binding analysis, and phosphatase kinetics. Analysis of Mn(2+)-bound active site mutant lambdaPP proteins shows that H22N, N75H, and H186N mutations decrease phosphatase activity but still allow mononuclear Mn(2+) and [(Mn(2+))(2)] binding. The high affinity Mn(2+) binding site is shown to consist of M2 site ligands H186 and Asn75, but not H22 from the M1 site which is ascribed as the lower affinity site.     

Flow cytometric measurement of fluorescence resonance energy transfer on cell surfaces. Quantitative evaluation of the transfer efficiency on a cell-by-cell basis

A method has been developed for the determination of the efficiency (E) of the fluorescence resonance energy transfer between moieties on cell surfaces by use of a computer-controlled flow cytometer capable of dual wavelength excitation. The absolute value of E may be calculated on a single-cell basis. The analysis requires the measurement of samples stained with donor and acceptor conjugated ligands alone as well as together. In model experiments HK 22 murine lymphoma cells labeled with fluorescein-conjugated concanavalin A (Con A) and/or rhodamine conjugated Con A were used to determine energy transfer histograms. Using the analytic solution to energy transfer in two dimensions, a high surface density of Con A binding sites was found that suggests that the Con A receptor sites on the cell surface are to a degree preclustered . We call this technique flow cytometric energy transfer ( FCET ).     

APPLICATIONS OF TIME-RESOLVED FLUOROIMMUNOASSAY TO DETECT MAGNETIC BEAD CAPTURED ESCHERICHIA COLI O157:H7

A time-resolved fluorescence technique was developed to detect Escherichia coli O157:H7 in ground beef burger. After a 4.5 h enrichment period, streptavidin coated magnetic beads conjugated with biotin-labeled anti E. coli O157:H7 were used to capture the bacteria. The bacteria were, at the same time, also labeled by a nonfluorescent, europium (Eu)-tagged anti-E. coli O157:H7 antibody. The sandwiched bacterial complexes were then concentrated using a magnetic particle concentrator and washed to remove other solution components. Upon addition of an enhancement buffer, the Eu-labels were then released from the antibodies and chelated to nitrilo-triacetic acid (NTA) and trioctylphosphine oxide (TOPO) to form highly fluorescent Eu-(2-NTA)₃(TOPO)_2–3 micellar complexes. Delayed fluorescence associated with these complexes was measured and its intensity was used to estimate the original bacterial concentration spiked in hamburger. This approach was applied to detect E. coli O157:H7 spiked in hamburgers. The results indicated this method is able to detect 1 CFU/g of the bacteria after a brief enrichment for four and half hours at 37C. Specificity studies indicated that the approach exhibited no or limited cross reactivity to Salmonella typhimurium, E. coli K-12 or Shigella dysenteriae spiked in hamburgers. Thus, the developed approach may be used as a rapid screening procedure for E. coli O157 bacteria in foods.     

Les anticorps fluorescents dans le diagnostic bactériologique des Escherichia coli entéro-pathogènes: Comparaison avec l'isolement de ces germes par culture classique

Enteropathogenic E. coli were sought routinely by the fluorescent antibody technique, using monovalent and polyvalent conjugates (rhodamine sulfonyl· chloride, fluorescein and rhodamine isothiocyanate).In 2061 stool specimens examined with monovalent antibody to E. coli 0127:B8, there were 61 false positives, 14 of which were from previously known cases of E. coli 0127:B8 infection, and 33 specimens that were negative by fluorescence but positive on culture. In 457 stool specimens examined with polyvalent antiserum, there were 15 false positives, five of which came from cases previously infected by the corresponding serotypes, and there were 20 specimens negative by fluorescence but positive on culture. The disagreement amounted, therefore, to 4.6% in the former instance and 7.6% in the latter. This fluorescent technique permits rapid sufficiently precise detection of enteropathogenic E. coli in stools.     

Self-assembly of conjugated polymers and ds-oligonucleotides directed fractal-like aggregates

Highly ordered nanostructures between conjugated polymers and ds-oligonucleotides have been first fabricated by simply controlling the self-assembly processes, which shows a novel concept for fabricating fractal-like structures. The formation of polymer/DNA fractal-like aggregates is a diffusion-limited aggregation (DLA) process. The fractal dimension is independent of the polymer/DNA concentration but only related to the polymer/DNA charge ratio. More interestingly, the different fluorescent resonance energy transfer (FRET) behavior between the polymer and the DNA can be used to distinguish dsDNA from ssDNA.     

Study of the interaction of polyols with polymers containing n-substituted [(4-boronophenyl)methyl]-ammonio groups

Polymers, based on dextran and cellulose, having 2-{[(4-boronophenyl)-methyl]-ethylammonio}ethyl and -diethylammonio～ethyl groups were prepared. It was shown that these polymers could be employed for absorption of cis-diol compounds. The polymers were found to be highly specific towards polyols, carbohydrates, nucleosides, and nucleotides over a wide range of pH. The polymer based on DEAE-Sephadex A-25 was used in separating nucleosides, and in fractionating mononucleotides and carbohydrates. The chromatographic behavior of carbohydrates is defined by their structure and conformation, which are also responsible for different stabilities of the boronic complexes generated.     

Virulence potential and genomic mapping of the worldwide clone Escherichia coli ST131

Recently, the worldwide propagation of clonal CTX-M-15-producing Escherichia coli isolates, namely ST131 and O25b:H4, has been reported. Like the majority of extra-intestinal pathogenic E. coli isolates, the pandemic clone ST131 belongs to phylogenetic group B2, and has recently been shown to be highly virulent in a mouse model, even though it lacks several genes encoding key virulence factors (Pap, Cnf1 and HlyA). Using two animal models, Caenorhabditis elegans and zebrafish embryos, we assessed the virulence of three E. coli ST131 strains (2 CTX-M-15- producing urine and 1 non-ESBL-producing faecal isolate), comparing them with five non-ST131 B2 and a group A uropathogenic E. coli (UPEC). In C. elegans, the three ST131 strains showed intermediate virulence between the non virulent group A isolate and the virulent non-ST131 B2 strains. In zebrafish, the CTX-M-15-producing ST131 UPEC isolates were also less virulent than the non-ST131 B2 strains, suggesting that the production of CTX-M-15 is not correlated with enhanced virulence. Amongst the non-ST131 B2 group isolates, variation in pathogenic potential in zebrafish embryos was observed ranging from intermediate to highly virulent. Interestingly, the ST131 strains were equally persistent in surviving embryos as the non-ST131-group B2 strains, suggesting similar mechanisms may account for development of persistent infection. Optical maps of the genome of the ST131 strains were compared with those of 24 reference E. coli strains. Although small differences were seen within the ST131 strains, the tree built on the optical maps showed that these strains belonged to a specific cluster (86% similarity) with only 45% similarity with the other group B2 strains and 25% with strains of group A and D. Thus, the ST131 clone has a genetic composition that differs from other group B2 strains, and appears to be less virulent than previously suspected.