Amplified Fragment Length Polymorphism Fingerprinting of Pseudomonas Strains from a Poultry Processing Plant

Molecular typing has been used previously to identify and trace dissemination of pathogenic and spoilage bacteria associated with food processing. Amplified fragment length polymorphism (AFLP) is a novel DNA fingerprinting technique which is considered highly reproducible and has high discriminatory power. This technique was used to fingerprint 88 Pseudomonas fluorescens and Pseudomonas putida strains that were previously isolated from plate counts of carcasses at six processing stages and various equipment surfaces and environmental sources of a poultry abattoir. Clustering of the AFLP patterns revealed a high level of diversity among the strains. Six clusters (clusters I through VI) were delineated at an arbitrary Dice coefficient level of 0.65; clusters III (31 strains) and IV (28 strains) were the largest clusters. More than one-half (52.3%) of the strains obtained from carcass samples, which may have represented the resident carcass population, grouped together in cluster III. By contrast, 43.2% of the strains from most of the equipment surfaces and environmental sources grouped together in cluster IV. In most cases, the clusters in which carcass strains from processing stages grouped corresponded to the clusters in which strains from the associated equipment surfaces and/or environmental sources were found. This provided evidence that there was cross-contamination between carcasses and the abattoir environment at the DNA level. The AFLP data also showed that strains were being disseminated from the beginning to the end of the poultry processing operation, since many strains associated with carcasses at the packaging stage were members of the same clusters as strains obtained from carcasses after the defeathering stage.     

Fluorination of 6-methyluracil and its nlcleosides.

6-Methyluracil and its perbenzoylated 1-(beta-D-ribofuranosyl) and 1-(2-deoxy-beta-D-ribofuranosyl) derivatives afford on treatment with elemental fluorine in acetic acid solutions, the corresponding derivatives of 5-fluoro-6-methyluracil and 5-fluoro-6-fluoromethyluracil. The free nucleosides have been obtained from the protected derivatives by methanolysis. The CH2F linkage in 5-fluoro-6-fluoromethyluracil derivatives is stable towards hydrolysis and nucleophilic agents.     

CD studies on the conformation of oligonucleotides complexed with divalent metal ions: interaction of Zn2+ with guanine favours syn conformation.

The interaction of the divalent metal ions Mg2+, Mn2+, Zn2+ and Cu2+ with GpG and several other dinucleoside monophosphates were investigated by means of circular dichroism. The spectra of the complexes of GpG, GpU analogues and ApGpG caused in the presence of Zn2+ and other transition metals show a close similarity in the spectral CD shape to that previously reported in the literature for GpG and GpU at low pH and for m7GpG. From the results it may be concluded that transition metal ions-particularly considered for Zn2+/- tends to favour the degree of stacking with Guo in syn conformation in GpG or GpU due to the coordination of the metal ion at N-7 of the 3'-bound position while shielding of the phosphate site by Mg2+ does not influence the sugar-base torsional angle under comparable conditions. Stereochemical aspects and selectivity of the Zn2+ mediated conformation of the dinucleoside phosphates are discussed.     

Spindle pole centrosomes of sea urchin embryos are partially composed of material recruited from maternal stores

The spindle poles of fertilized sea urchin eggs have commonly been modeled as being derived from the centrosomes of the fertilizing spermatozoon. Boveri's theory of fertilization, proposed at the turn of the century, states that the maternal centrosome is suppressed or inactivated during oogenesis and that the sperm centrosome is functionally dominant. In support of this proposal, more recent studies have shown that the sperm imports a determinant that is involved in centrosomal replication. Examination of sea urchin zygotes immunofluorescently labeled with a new anti-centrosomal antibody by quantitative confocal laser-scanning microscopy shows, however, that spindle pole centrosomes are not exclusively paternal structures, but additionally contain material derived from maternal pools. Furthermore, this maternal centrosomal material is divided among daughter blastomeres during cleavage. It therefore appears that although the sperm centrosome plays a dominant role in organizing the spindle poles, much of the centrosomal material within the spindle poles of the zygote is actually recruited from preexisting egg cytoplasmic stores. These data indicate that centrosomes of sea urchin embryos are biparentally derived, composite organelles.     

Disruption of nucleocytoplasmic trafficking of cyclin D1 and topoisomerase II by sanguinarine

Background  

The quaternary isoquinoline alkaloid sanguinarine is receiving increasing attention as a potential chemotherapeutic agent in the treatment of cancer. Previous studies have shown that this DNA-binding phytochemical can arrest a number of different types of transformed cells in G0/G1, and upregulate the CKIs p21 and p27 while downregulating multiple cyclins and CDKs. To more closely examine the responses of some of these cell cycle regulatory molecules to sanguinarine, we used immunocytochemical methods to visualize cyclin D1 and topoisomerase II behavior in MCF-7 breast cancer cells.     

Carbon : nitrogen : phosphorus ratios influence biofilm formation by Enterobacter cloacae and Citrobacter freundii

AIMS: To test the effects of C : N : P ratio modification of a well-known nutrient medium formulation, the Endo formulation on biofilm formation by Enterobacter cloacae Ecl and Citrobacter freundii Cf1 in both single-species and binary species biofilms. METHODS AND RESULTS: The C : N : P atom : atom ratio of a well-known nutrient medium formulation, the Endo formulation, that has been applied in fermentative biohydrogen studies, was modified to include two different C concentrations, one containing 17.65 g l(-1) and the other 8.84 g l(-1) sucrose, each containing four different C : N : P ratios, two at higher C : N : P ratios (334 : 84 : 16.8 and 334 : 84 : 3) and two at lower C : N : P ratios (334 : 28 : 5.6 and 334 : 28 : 1). Attached cells were enumerated after dislodging the biofilms that had formed on granular activated carbon (GAC). The modified medium containing 17.65 g l(-1) sucrose and having a C : N : P ratio of 334 : 28 : 5.6 resulted in significantly (P < 0.05) higher counts of attached cells for both single-species biofilms at 7.73 log(10) CFU g(-1) GAC and 9.3 log(10)CFU g(-1) GAC for Ent. cloacae Ecl and Cit. freundii Cf1, respectively, and binary species biofilms at 8.2 log(10) CFU g(-1) GAC and 6.34 log(10) CFU g(-1) GAC for Ent. cloacae Ecl and Cit. freundii Cf1, respectively. Scanning electron micrographs showed qualitative evidence that the 334 : 28 : 5.6 ratio encouraged more complex and extensive biofilm growth for both single-species and binary species biofilms. CONCLUSIONS: The differences in the attachment numbers between the different ratios were found not to be a result of the individual actions of the bacterial isolates involved but rather because of the effects of the various C : N : P ratios. The 334 : 28 : 5.6 ratio showed significantly (P < 0.05) higher counts of attached cells for both single-species and binary species biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: This study indicates that C : N : P ratios should be a key consideration with regard to maximizing biofilm formation in shake flask and fluidized bed bioreactor studies as well as understanding fundamental factors affecting biofilm growth in natural environments.     

Serum neurotrophin profile in systemic sclerosis

Background

Neurotrophins (NTs) are able to activate lymphocytes and fibroblasts; they can modulate angiogenesis and sympathic vascular function. Thus, they can be implicated in the three pathogenic processes of systemic sclerosis (SSc). The aims of this study are to determine blood levels of Nerve Growth Factor (NGF), Brain-Derived Neurotrophic Factor (BDNF) and Neurotrophin-3 (NT-3) in SSc and to correlate them with clinical and biological data.

Methods

Serum samples were obtained from 55 SSc patients and 32 control subjects to measure NTs levels by ELISA and to determine their relationships with SSc profiles.

Findings

Serum NGF levels were higher in SSc patients (288.26±170.34 pg/mL) than in control subjects (170.34±50.8 pg/mL, p<0.001) and correlated with gammaglobulins levels and the presence of both anti-cardiolipin and anti-Scl-70 antibodies (p<0.05). In contrast, BDNF levels were lower in SSc patients than in controls (1121.9±158.1 vs 1372.9±190.9 pg/mL, p<0.0001), especially in pulmonary arterial hypertension and diffuse SSc as compared to limited forms (all p<0.05). NT-3 levels were similar in SSc and in the control group (2657.2±2296 vs 2959.3±2555 pg/mL, NS). BDNF levels correlated negatively with increased NGF levels in the SSc group (and not in controls).

Conclusion

Low BDNF serum levels were not previously documented in SSc, particularly in the diffuse SSc subset and in patients with pulmonary hypertension or anti-Scl-70 antibodies. The negative correlation between NGF and BDNF levels observed in SSc and not in healthy controls could be implicated in sympathic vascular dysfunction in SSc.     

Kinetics of Binding of Multisubstrate Analogue Inhibitor (2-Amino-9-[2-(Phosphonomethoxy)Ethyl]-6-Sulfanylpurine) with Trimeric Purine Nucleoside Phosphorylase

Complex formation of multisubstrate analogue inhibitor—2-amino-9-[2-(phosphonomethoxy)ethyl]-6-sulfanylpurine (PME-6-thio-Gua) with trimeric purine nucleoside phosphorylase from Cellulomonas sp. was investigated using a stopped-flow spectrofluorimetric approach. Results obtained indicate that, in contrast to binding of guanine, i.e., the transition-state conformation trapping ligand, for which binding at each active site is followed by the enzyme conformational change, association of the ground-state analogue PME-6-thio-Gua is a one-step process.