R plasmids in environmental Vibrio cholerae non-O1 strains

The occurrence of drug resistance and its plasmid-mediated transferability was investigated in 140 environmental strains of Vibrio cholerae non-O1 and 6 strains of Vibrio cholerae, both O1 and non-O1, of clinical origin. Of the 146 strains tested, 93% were resistant to at least one drug and 74% were resistant to two or more antibiotics. The O1 strains were susceptible to all antibiotics used. A total of 26 of 28 selected resistant wild strains carried R plasmids that were transferable by intraspecific and intergeneric matings. The most common transmissible R factor determined resistance to ampicillin, amoxicillin, and sulfanilamide (30%), followed by resistance to ampicillin and amoxicillin (13%) and resistance to ampicillin, amoxicillin, phosphomycin, and sulfanilamide (9%). Comparison of the three methods of plasmid analysis showed that the method of Birnboim and Doly (Nucleic Acids Res. 7:1513-1523, 1979) without EDTA and lysozyme was optimal for isolation of both large and small plasmids in environmental V. cholerae strains. Most strains harbored more than one plasmid, and the molecular sizes ranged from 1.1 to 74.8 megadaltons. The plasmids of high molecular size (around 74 megadaltons) were responsible for the resistance pattern transferred and were maintained with high stability in the hosts.     

Surface and virulence properties of environmental Vibrio cholerae non-O1 from Albufera Lake (Valencia, Spain).

A total of 140 environmental Vibrio cholerae non-O1 isolates, together with several culture collection strains from both environmental and clinical sources, were studied in relation to hemagglutination, surface hydrophobicity, and the enzymatic, hemolytic, cytotoxic, and enterotoxic activities of their extracellular products. A total of 78 and 62% of the strains produced hemagglutinins and exohemagglutinins, respectively. Four different hemagglutinating and two exohemagglutinating activities were found by using eight sugars in the inhibition assays. Cell-bound mannose-sensitive hemagglutination was detected mainly in chicken blood, whereas fucose-sensitive hemagglutination was recorded only in human blood. Cell-bound hemagglutinin resistant to all sugars tested was the only one related to surface hydrophobicity. The surface properties varied along the growth curves. The non-O1 strains displayed strong enzymatic and hemolytic activities, except for esculin hydrolysis. Of 26 non-O1 isolates selected for cytotoxin and enterotoxin production, 23 showed a wide spectrum of cytotoxic effects on cell lines of poikilothermic and homoiothermic species, but they were weakly enterotoxigenic in the infant mouse test. All extracellular products of cytotoxic strains were proteolytic, lipolytic, and hemolytic, and a high percentage produced hemagglutination of chicken blood. The cytotoxic factors in the non-O1 strains analyzed were not R plasmid mediated.     

Iron-binding compounds and related outer membrane proteins in Vibrio cholerae non-O1 strains from aquatic environments

A total of 156 strains of Vibrio cholerae non-O1 from aquatic origins were examined for the presence of iron uptake mechanisms and compared with O1 strains and other Vibrio species. All non-O1 strains were able to grow in iron-limiting conditions, with MICs of ethylenediaminedi (O-hydroxyphenylacetic acid) ranging from 20 microM to 2 mM. The production of siderophores was demonstrated by growth in chrome azurol S agar and cross-feeding assays. All strains produced phenolate-type compounds, as assessed by the chemical tests and by bioassays with Salmonella typhimurium enb-7. Some of the strains also promoted the growth of S. typhimurium enb-1 (which can use only enterobactin as a siderophore) as well as some strains of Vibrio anguillarum deficient in the anguibactin-mediated system. The chromatographic analyses and absorption spectra of siderophores extracted from culture supernatants suggest that vibriobactin may be produced by the strains examined. Interestingly, some strains also produced hydroxamate-type compounds, as determined by chemical tests, and were able to promote the growth of an aerobactin-deficient strain of Escherichia coli. These results were confirmed by the absorption spectra and chromatographic analyses of the culture extracts. The synthesis of iron-regulated outer membrane proteins in representative strains was also examined. The molecular sizes of the main induced proteins ranged from 70 to 78 kilodaltons. These results indicate that several iron uptake mechanisms which could be involved in environmental survival and pathogenicity are present in environmental V. cholerae non-O1 strains.     

Siderophore-mediated iron acquisition mechanisms in Vibrio vulnificus biotype 2.

Vibrio vulnificus biotype 2 is a primary pathogen for eels and, as has recently been suggested, an opportunistic pathogen for humans. In this study we have investigated the ability of V. vulnificus biotype 2 to obtain iron by siderophore-mediated mechanisms and evaluated the importance of free iron in vibriosis. The virulence degree for eels was dependent on iron availability from host fluids, as was revealed by a reduction in the 50% lethal dose for iron-overloaded eels. This biotype produced both phenolate- and hydroxamate-type siderophores of an unknown nature and two new outer membrane proteins of around 84 and 72 kDa in response to iron starvation. No alterations in lipopolysaccharide patterns were detected in response to iron stress. Finally, our data suggest that V. vulnificus biotype 2 uses the hydroxamate-type siderophore for removal of iron from transferrin rather than relying on a receptor for this iron-binding protein.     

An Immunological Assay for Detection and Enumeration of Thermophilic Biomining Microorganisms

A specific, fast, and sensitive nonradioactive immunobinding assay for the detection and enumeration of the moderate thermophile Thiobacillus caldus and the thermophilic archaeon Sulfolobus acidocaldarius was developed. It employs enhanced chemiluminescence or peroxidase-conjugated immunoglobulins in a dot or slot blotting system and is very convenient for monitoring thermophilic bioleaching microorganisms in effluents from industrial bioleaching processes.     

Methylation of ribosomal proteins in Bacillus subtilis

We measured the methylation of ribosomal proteins from the 30S and 50S subunits of Bacillus subtilis after growing the cells in the presence of [1-14C]methionine and [methyl-3H]methionine. Two-dimensional polyacrylamide gel electrophoretic analysis revealed a preferential methylation of the 50S ribosomal proteins. Proteins L11 and L16, and possibly L9, L10, L18, and L20, were methylated. On the other hand, only two possibly methylated proteins were found on the 30S subunit. A comparison of these results with those for Escherichia coli suggests a common methylation pattern for the bacterial ribosomal proteins.     

Phosphate starvation affects the synthesis of outer membrane proteins in Thiobacillus ferrooxidans

The outer membrane protein (omp40) component from the chemolithoautotrophic acidophilic Thiobacillus ferrooxidans is apparently regulated by the external pH and the concentration of phosphorus. Its amino-terminal sequence showed little identity with the Escherichia coli OmpC, OmpF or PhoE porins, but was 38.5% identical to the outer membrane channel-forming protein NosA from Pseudomonas stutzeri, whose expression is also regulated environmentally. In addition, the partial amino acid sequence of T. ferrooxidans omp40 showed between 34 and 38% identity with the amino-terminal end of the small outer membrane proteins Rck and PagC from Salmonella typhimurium and OmpX from Enterobacter cloacae.     

Direct inhibition of hexokinase activity by metformin at least partially impairs glucose metabolism and tumor growth in experimental breast cancer

Emerging evidence suggests that metformin, a widely used anti-diabetic drug, may be useful in the prevention and treatment of different cancers. In the present study, we demonstrate that metformin directly inhibits the enzymatic function of hexokinase (HK) I and II in a cell line of triple-negative breast cancer (MDA-MB-231). The inhibition is selective for these isoforms, as documented by experiments with purified HK I and II as well as with cell lysates. Measurements of ¹⁸F-fluoro-deoxyglycose uptake document that it is dose- and time-dependent and powerful enough to virtually abolish glucose consumption despite unchanged availability of membrane glucose transporters. The profound energetic imbalance activates phosphorylation and is subsequently followed by cell death. More importantly, the “in vivo” relevance of this effect is confirmed by studies of orthotopic xenografts of MDA-MB-231 cells in athymic (nu/nu) mice. Administration of high drug doses after tumor development caused an evident tumor necrosis in a time as short as 48 h. On the other hand, 1 mo metformin treatment markedly reduced cancer glucose consumption and growth. Taken together, our results strongly suggest that HK inhibition contributes to metformin therapeutic and preventive potential in breast cancer.     

Risks and Benefits of Early Antithrombotic Therapy after Thrombolytic Treatment in Patients with Acute Stroke

Background

Current guidelines recommend withholding antithrombotic therapy (ATT) for at least 24 h in patients with acute ischemic stroke treated with thrombolytic therapy. Herein, we report a retrospective analysis of a single-centre experience on the safety and efficacy of antithrombotic therapy (ATT) started before or after 24 h of intravenous thrombolysis in a cohort of acute ischemic stroke patients.

Methods

A total of 139 patients (Rapid ATT group) received antithrombotic therapy before 24 h of thrombolysis, and 33 patients (Standard ATT group) after 24 h. The brain parenchyma and vessel status were assessed using simple CT scan on admission, multimodal CT scan at the end of thrombolysis, and angio-CT/MRI scan at day 3. Functional outcome was scored using the modified Rankin Scale (mRS) at day 90.

Results

The two ATT groups had similar demographics, stroke subtypes, baseline NIHSS, thrombolytic strategies, vessel-patency rates at the end of thrombolysis, and incidence of bleeding complications at follow up. At day 3, the Rapid ATT group had a non-significant improved vessel-patency rate than the Standard ATT group. At day 90, a greater proportion of patients in the rapid ATT group had shifted down the mRS, and had improved in the NIHSS score.

Conclusions

ATT initiated before 24 h of intravenous thrombolytic therapy in acute stroke patients disclosed no safety concerns compared with a conventional antithrombotic therapy delay of 24 h and showed better functional outcome at follow up. The value of early initiation of ATT after thrombolysis deserves further assessment in randomized controlled trials.     

Potential geographical distribution of the red palm mite in South America

Among pests that have recently been introduced into the Americas, the red palm mite, Raoiella indica Hirst (Prostigmata: Tenuipalpidae), is the most invasive. This mite has spread rapidly to several Caribbean countries, United States of America, Mexico, Venezuela, Colombia and Brazil. The potential dispersion of R. indica to other regions of South America could seriously impact the cultivation of coconuts, bananas, exotic and native palms and tropical flowers such as the Heliconiaceae. To facilitate the development of efficacious R. indica management techniques such as the adoption of phytosanitary measures to prevent or delay the dispersion of this pest, the objective of this paper was to estimate the potential geographical distribution of R. indica in South America using a maximum entropy model. The R. indica occurrence data used in this model were obtained from extant literature, online databases and field sampling data. The model predicted potential suitable areas for R. indica in northern Colombia, central and northern Venezuela, Guyana, Suriname, east French Guiana and many parts of Brazil, including Roraima, the eastern Amazonas, northern Pará, Amapá and the coastal zones, from Pará to north of Rio de Janeiro. These results indicate the potential for significant R. indica related economic and social impacts in all of these countries, particularly in Brazil, because the suitable habitat regions overlap with agricultural areas for R. indica host plants such as coconuts and bananas.