Identification of two new intimin types in atypical enteropathogenic Escherichia coli.

Stool specimens of patients with diarrhea or other gastrointestinal alterations who were admitted to Xeral-Calde Hospital (Lugo, Spain) were analyzed for the prevalence of typical and atypical enteropathogenic Escherichia coli (EPEC). Atypical EPEC strains (eae+ bfp-) were detected in 105 (5.2%) of 2015 patients, whereas typical EPEC strains (eae+ bfp+) were identified in only five (0.2%) patients. Atypical EPEC strains were (after Salmonella) the second most frequently recovered enteropathogenic bacteria. In this study, 110 EPEC strains were characterized. The strains belonged to 43 O serogroups and 69 O:H serotypes, including 44 new serotypes not previously reported among human EPEC. However, 29% were of one of three serogroups (O26, O51, and O145) and 33% belonged to eight serotypes (O10:H-, O26:H11, O26:H-, O51:H49, O123:H19, O128:H2, O145:H28, and O145:H-). Only 14 (13%) could be assigned to classical EPEC serotypes. Fifteen intimin types, namely, alpha1 (6 strains), alpha2 (4 strains), beta1 (34 strains), xiR/b2 (6 strains), gamma1 (13 strains), gamma2/q (16 strains), delta/k (5 strains), epsilon1 (9 strains), nuR/e2 (5 strains), zeta (6 strains), iota1 (1 strain), muR/iota2 (1 strain), nuB (1 strain), xiB (1 strain), and o (2 strains), were detected among the 110 EPEC strains, but none of the strains was positive for intimin types mu1, mu2, lambda, or muB. In addition, in atypical EPEC strains of serotypes O10:H-, O84:H-, and O129:H-, two new intimin genes (eae-nuB and eae-o) were identified. These genes showed less than 95% nucleotide sequence identity with existing intimin types. Phylogenetic analysis revealed six groups of closely related intimin genes: (i) alpha1, alpha2, zeta, nuB, and o; (ii) iota1 and muR/iota2; (iii) beta1, xiR/beta2B, delta/beta2O, and kappa; (iv) epsilon1, xiB, eta1,eta2, and nuR/epsilon2; (v) gamma1, muB, gamma2, and theta; and (vi) lambda. These results indicate that atypical EPEC strains belonging to large number of serotypes and with different intimin types might be frequently isolated from human clinical stool samples in Spain.     

Electrolyte effects on the intervalence transition within discrete binuclear cyano-bridged complexes. An estimation of activation free energy from static, optical and electrochemical data

A study of the metal-to-metal charge-transfer (MMCT) transition within the binuclear cyano-bridged complexes cis-[L¹³Co^III(μ-NC)Fe^II(CN)₅]⁻ (L¹³ = 12-methyl-1,4,7,10-tetraazacyclotridecan-12-amine), trans-[L¹⁴Co^III(μ-NC)Fe^II(CN)₅]⁻ (L¹⁴ = 6-methyl-1,4,8,11-tetraazacyclotetradecan-6-amine) and trans-[L¹⁵Co^III(μ-NC)Fe^II(CN)₅]⁻ (L¹⁵ = 10-methyl-1,4,8,12-tetraazacyclopentadecan-10-amine) has been carried out in electrolyte solutions at varying concentrations. Using these data, as well as the reaction free energies obtained from electrochemical measurements, the reorganisation and activation free energies for the forward and reverse thermal electron-transfer processes have been estimated. The changes of these parameters with the electrolyte concentration, as well as those of the energy of the maximum MMCT band and the reaction free energy, are mainly due to ion-pairing effects.     

Daunomycin-polypeptide conjugates with antitumor activity

We have developed a group of water-soluble drug conjugates in which daunomycin (Dau) is coupled to cationic, amphoteric or anionic branched polypeptides and a new conjugate containing a cationic polypeptide carrier modified with a cell penetrating octaarginine. We investigated in vitro physiological activity of these conjugates in several aspects: in vitro cytotoxicity and cytostatic effect, adhesion and cellular uptake were examined on murine (J774 and L1210) and human (MonoMac6 and HL-60) leukemia cell lines and on murine bone marrow derived macrophages. We found that these processes are dependent on the properties of the carrier, on experimental conditions like concentration and incubation time. We found that attachment of polypeptide and cell penetrating peptide to the bioactive agent, depending on the cell line, could significantly improve the antitumor activity of the drug.     

Caffeic acid derivatives: in vitro and in vivo anti-inflammatory properties

Caffeic acid and some of its derivatives such as caffeic acid phenetyl ester (CAPE) and octyl caffeate are potent antioxidants which present important anti-inflammatory actions. The present study assessed the in vitro and in vivo effects of five caffeic acid derivatives (caffeic acid methyl, ethyl, butyl, octyl and benzyl esters) and compared their actions to those of CAPE. In the model of LPS-induced nitric oxide (NO) production in RAW 264.7 macrophages, the pre-incubation of all derivatives inhibited nitrite accumulation on the supernatant of stimulated cells, with mean IC₅₀ (μM) values of 21.0, 12.0, 8.4, 2.4, 10.7 and 4.80 for methyl, ethyl, butyl, octyl, benzyl and CAPE, respectively. The effects of caffeic acid derivatives seem to be related to the scavenging of NO, as the compounds prevented SNAP-derived nitrite accumulation and decreased iNOS expression. In addition, butyl, octyl and CAPE derivatives significantly inhibited LPS-induced iNOS expression in RAW 264.7 macrophages. Extending the in vitro results, we showed that the pre-treatment of mice with butyl, octyl and CAPE derivatives inhibited carrageenan-induced paw edema and prevented the increase in IL-1β levels in the mouse paw by 30, 24 and 36%, respectively. Butyl, octyl and CAPE derivatives also prevented carrageenan-induced neutrophil influx in the mouse paw by 28, 49 and 31%, respectively. Present results confirm and extend literature data, showing that caffeic acid derivatives exert in vitro and in vivo anti-inflammatory actions, being their actions mediated, at least in part by the scavenging of NO and their ability to modulate iNOS expression and probably that of other inflammatory mediators.     

Reproductive ecology of distylous Palicourea padifolia (Rubiaceae) in a tropical montane cloud forest. II. Attracting and rewarding mutualistic and antagonistic visitors

By definition, the floral morphs of distylous plants differ in floral architecture. Yet, because cross-pollination is necessary for reproductive success in both morphs, they should not differ in attributes that contribute to attracting and rewarding floral visitors. Floral and vegetative attributes that function in distylous polymorphism in hummingbird-pollinated Palicourea padifolia (Rubiaceae) and the responses of pollinators and insect herbivores to the resources offered by both morphs were investigated. The performance of each morph along multiple stages of the reproductive cycle, from inflorescence and nectar production to fruit production, was surveyed, and pollinator behavior and nectar standing crops were then observed. Costs associated with such attractiveness were also evaluated in terms of herbivore attack and of plant reproductive fitness (female function) as a function of leaf herbivory. The number of inflorescences, floral buds, open flowers, and ripe fruits offered by either floral morph were similar, but short-styled plants almost doubled the number of developing fruits of long-styled plants. Long-styled flowers produced higher nectar volumes and accumulated more nectar over time than short-styled flowers. Measures of nectar standing crop and data on pollinator behavior suggest that hummingbirds respond to this morph-specific scheduling of nectar production. Lastly, long-styled plants suffered a higher herbivore attack and lost more leaf area over time than those with short-styled flowers. Herbivory was negatively correlated with fruit number and fruit mass, and long-styled plants set significantly less fruit mass than short-styled plants. The results suggest that pollinators and herbivores may exert selective pressures on floral and vegetative traits that could also influence gender function.     

A contrastive view of Irish language dynamics

This paper presents an analysis of the linguistic anthropology which underpins the language dynamics of two Gaeltacht (Irish-speaking) areas, Ros Muc in Conamara, Co. Galway and Ráth Cairn in Co. Meath. This research highlights what could be considered a socio-linguistic paradox: the community (Ráth Cairn) which engages more vigorously in language maintenance endeavors, and exhibits a greater awareness of language policy and of linguistic ideology among members of the community, fares less favorably in socio-linguistic terms to the contrasting community (Ros Muc) which has to endure a more challenging socio-economic climate than that of Ráth Cairn. The relative socio-economic success of the Ráth Cairn community appears to be masking a greater malaise of socio-cultural fragility and language endangerment. In contrast, the language obsolescence issues faced by the Ros Muc community, though superficially not as severe, are enmeshed in what would be considered more pressing issues of socio-economic marginalization.     

The crystal structure of a high affinity RNA stem-loop complexed with the bacteriophage MS2 capsid: further challenges in the modeling of ligand-RNA interactions

We have determined the structure to 2.8 A of an RNA aptamer (F5), containing 2'-deoxy-2-aminopurine (2AP) at the -10 position, complexed with MS2 coat protein by soaking the RNA into precrystallised MS2 capsids. The -10 position of the RNA is an important determinant of binding affinity for coat protein. Adenine at this position in other RNA stem-loops makes three hydrogen bonds to protein functional groups. Substituting 2AP for the -10 adenine in the F5 aptamer yields an RNA with the highest yet reported affinity for coat protein. The refined X-ray structure shows that the 2AP base makes an additional hydrogen bond to the protein compared to adenine that is presumably the principal origin of the increased affinity. There are also slight changes in phosphate backbone positions compared to unmodified F5 that probably also contribute to affinity. Such phosphate movements are common in structures of RNAs bound to the MS2 T = 3 protein shell and highlight problems for de novo design of RNA binding ligands.     

The A-204C polymorphism in the cholesterol 7alpha-hydroxylase (CYP7A1) gene determines the cholesterolemia responsiveness to a high-fat diet

The aim of the study was to ascertain whether the A-204C polymorphism in the cholesterol 7 -hydroxylase (CYP7A1) gene plays any role in determining LDL-cholesterol (LDL-C) concentration responsiveness to a high-fat diet. The concentrations of total cholesterol and LDL-cholesterol were measured in eleven healthy men (age: 30.9+/-3.2 years; BMI: 24.9+/-2.7 kg/m(2);;) who were homozygous for either the -204A or -204C allele, after 3 weeks on a low-fat (LF) diet and 3 weeks on a high-fat (HF) diet. During both dietary regimens, the isocaloric amount of food was provided to volunteers; LF diet contained 22 % of energy as a fat and 2.2 mg of cholesterol/kg of body weight a day, HF diet 40 % of fat and 9.7 mg of cholesterol/kg of body weight a day. In six subjects homozygous for the -204C allele, the concentrations of cholesterol and LDL-cholesterol were significantly higher on HF than on LF diet (cholesterol: 4.62 vs. 4.00 mmol/l, p<0.05; LDL-C: 2.15 vs. 1.63 mmol/l, p<0.01, respectively); no significant change was observed in five subjects homozygous for the -204A allele. There were no other differences in lipid and lipoprotein-lipid concentrations. Therefore, the polymorphism in the cholesterol 7alpha-hydroxylase promotor region seems to be involved in the determination of cholesterol and LDL-C responsiveness to a dietary fat challenge.     

Primate models for human immunodeficiency virus infection. Evolution of receptor use during pathogenesis

Animal models greatly facilitate understanding of transmission, pathogenesis and immune responses in HIV and SIV infection and provide models for studies on the effect of candidate drugs or vaccines. However, there are several aspects that one should consider when drawing conclusions from results obtained from animal models. First, the genetic relationship of primate lentiviruses cannot be disregarded because it is known that HIV-1 is more closely related to SIV of chimpanzee origin (SIVcpz) than to SIV from sooty mangabey (SIVsm) origin. Nevertheless, SIVsm and SIVmac are the ones most often used as model systems. Second, there are differences in the biological properties, like CXCR4 use and CD4-independent coreceptor use, of HIV and SIV. These differences might be relevant in virus transmission, pathogenesis and in evoking immune responses. Third, in vivo and in vitro selection may influence the results. Neutralizing antibodies may play a role in selection of variant viruses since neutralization sensitive, CD4-independent SIVsm variants seemed to be suppressed in animals that mounted a neutralizing antibody response. It is tempting to speculate that neutralizing antibodies shape the SIV/HIV infection by selecting variants with a more "closed" envelope conformation with consequences for both receptor binding and neutralization sensitivity. The SIV/monkey model, although it has important advantages, may not answer all questions asked about HIV-1 infection in human.