Tungsten Silicide Contacts to Polycrystalline Silicon and Silicon-Germanium Alloys

Novel CVD WSi2 technology with low series and contact resistance in SiGe HBTs was achieved. Specific contact resistance to Si1-xGex with 0

Specific growth rate plays a critical role in hydrogen peroxide resistance of the marine oligotrophic ultramicrobacterium Sphingomonas alaskensis strain RB2256

The marine oligotrophic ultramicrobacterium Sphingomonas alaskensis RB2256 has a physiology that is distinctly different from that of typical copiotrophic marine bacteria, such as Vibrio angustum S14. This includes a high level of inherent stress resistance and the absence of starvation-induced stress resistance to hydrogen peroxide. In addition to periods of starvation in the ocean, slow nutrient-limited growth is likely to be encountered by oligotrophic bacteria for substantial periods of time. In this study we examined the effects of growth rate on the resistance of S. alaskensis RB2256 to hydrogen peroxide under carbon or nitrogen limitation conditions in nutrient-limited chemostats. Glucose-limited cultures of S. alaskensis RB2256 at a specific growth rate of 0.02 to 0.13 h(-1) exhibited 10,000-fold-greater viability following 60 min of exposure to 25 mM hydrogen peroxide than tells growing at a rate of 0.14 h(-1) or higher. Growth rate control of stress resistance was found to be specific to carbon and energy limitation in this organism. In contrast, V. angustum S14 did not exhibit growth rate-dependent stress resistance. The dramatic switch in stress resistance that was observed under carbon and energy limitation conditions has not been described previously in bacteria and thus may be a characteristic of the oligotrophic ultramicrobacterium, Catalase activity varied marginally and did not correlate with the growth rate, indicating that hydrogen peroxide breakdown was not the primary mechanism of resistance. More than 1,000 spots were resolved on silver-stained protein gels for cultures growing at rates of 0.026, 0.076, and 0.18 h(-1). Twelve protein spots had intensities that varied by more than twofold between growth rates and hence are likely to be important for growth rate-dependent stress resistance. These studies demonstrated the crucial role that nutrient limitation plays in the physiology of S. alaskensis RB2256, especially under oxidative stress conditions.

Fasciola hepatica: Comparative effects of host resistance and parasite intra-specific interactions on size and reproductive histology in flukes from rats infected with isolates differing in triclabendazole sensitivity

The efficacies of putative fasciolicides and vaccines against Fasciola hepatica are frequently monitored in clinical and field trials by determination of fluke egg output in host faeces and by worm counts in the host liver at autopsy. Less often used are parameters based on fluke size and histology, yet these can provide important indications of specific effects on the development of particular germ-line or somatic tissues, especially in relation to the timing and profligacy of egg production. In this study. F. hepatica metacercariae of two distinct isolates, the triclabendazole (TCBZ)-sensitive Cullompton isolate and the TCBZ-resistant Oberon isolate, were administered to rats as single-isolate or mixed-isolate infections. At autopsy 16 weeks later individual adult flukes were counted, measured and the reproductive organs were examined histologically. The degree of development of the testis tubules in each fluke was represented by a numerical score, based on the proportion of the histological section profiles occupied by testis tissue. The level of anti-F. hepatica antibody in the serum of each rat was determined by ELISA. It was found that Cullompton flukes were significantly larger than Oberon flukes, and that significantly more Cullompton metacercariae developed to adults than Oberon metacercariae. The Cullompton flukes showed histological evidence of aspermy and spermatogenic arrest, which was reflected in quantitatively reduced testicular development, as compared with the Oberon isolate. In Cullompton flukes, parthenogenetic egg development is implied. The size of Cullompton and Oberon flukes was significantly related to the number of adult flukes recovered, to the number of metacercariae administered, and to the percentage success of infection. The testis development score in both isolates was significantly related to the number of adult flukes recovered but not to the number of metacercariae administered, or to the percentage success of infection. Fluke size was positively related to testis score for both isolates, and a significant negative relationship was found between percentage success of infection and metacercarial dose. The results are interpreted in terms of differing interactions between various numbers of young flukes and host immunity during invasion of and migration in the hepatic parenchyma, and of fluke intra-specific (possibly pheromonal) stimulatory effects in the final stages of development, within the host bile ducts. No significant relationships were found between host antibody levels and fluke size or testis score. False positive serological reactions were found in some rats that had been infected, but found to harbour no flukes at autopsy. Clearly the act of eliminating the flukes involved generation of an immune response. (C) 2011 Elsevier B.V. All rights reserved.

Chemical ablation of gastric inhibitory polypeptide receptor action by daily (Pro3) GIP administration improves glucose tolerance and ameliorates insulin resistance and abnormalities of islet structure in obesity-diabetes.

Glucose-dependent insulinotropic polypeptide (gastric inhibitory polypeptide [GIP]) is an important incretin hormone secreted by endocrine K-cells in response to nutrient ingestion. In this study, we investigated the effects of chemical ablation of GIP receptor (GIP-R) action on aspects of obesity-related diabetes using a stable and specific GIP-R antagonist, (Pro3)GIP. Young adult ob/ob mice received once-daily intraperitoneal injections of saline vehicle or (Pro3)GIP over an 11-day period. Nonfasting plasma glucose levels and the overall glycemic excursion (area under the curve) to a glucose load were significantly reduced (1.6-fold; P <0.05) in (Pro3)GIP-treated mice compared with controls. GIP-R ablation also significantly lowered overall plasma glucose (1.4-fold; P <0.05) and insulin (1.5-fold; P <0.05) responses to feeding. These changes were associated with significantly enhanced (1.6-fold; P <0.05) insulin sensitivity in the (Pro3)GIP-treated group. Daily injection of (Pro3)GIP reduced pancreatic insulin content (1.3-fold; P <0.05) and partially corrected the obesity-related islet hypertrophy and ß-cell hyperplasia of ob/ob mice. These comprehensive beneficial effects of (Pro3)GIP were reversed 9 days after cessation of treatment and were independent of food intake and body weight, which were unchanged. These studies highlight a role for GIP in obesity-related glucose intolerance and emphasize the potential of specific GIP-R antagonists as a new class of drugs for the alleviation of insulin resistance and treatment of type 2 diabetes.

Mutations affecting the Rossman fold of isoleucyl-tRNA synthetase are correlated with low-level mupirocin resistance in Staphylococcus aureus.

The isoleucyl-tRNA synthetase (ileS) gene was sequenced in toto from 9 and in part from 31 Staphylococcus aureus strains with various degrees of susceptibility to mupirocin. All strains for which the mupirocin MIC was greater than 8 µg/ml contained point mutations affecting the Rossman fold via Val-to-Phe changes at either residue 588 (V588F) or residue 631 (V631F). The importance of the V588F mutation was confirmed by an allele-specific PCR survey of 32 additional strains. Additional mutations of uncertain significance were found in residues clustered on the surface of the IleS protein.

Chromosomal rearrangements affecting biofilm production and antibiotic resistance in a Staphylococcus epidermidis strain causing shunt-associated ventriculitis.

Ziebuhr W, Dietrich K, Trautmann M, Wilhelm M. Institut für Molekulare Infektionsbiologie, Würzburg, Germany. w.ziebuhr@mail.uni-wuerzburg.de During two clinical courses of shunt-associated meningitis in a 3-month-old child, five multiresistant S. epidermidis isolates were obtained and analyzed with regard to biofilm production and antibiotic susceptibility. Three S. epidermidis strains, which were initially isolated from the cerebrospinal fluid, produced biofilms on polystyrene tissue culture plates. Following antibiotic treatment and subsequent exchange of the shunt system, sterilization of the CSF was achieved. However, after three weeks a relapse of the infection occurred. The two S. epidermidis isolates obtained now were biofilm negative, but showed an identical resistance pattern as those from the previous infection, except that resistance to rifampicin and increased mininal inhibitory concentrations of aminoglycoside antibiotics had emerged. DNA fingerprinting by PFGE indicated the clonal origin of all isolates. However, some DNA rearrangements and differences in the IS256-specific hybridization patterns could be identified in the isolates from the second infection period that led to altered biofilm formation and increased expression of aminoglycoside resistance traits. The data evidence that variation of biofilm expression occurs in vivo during an infection and highlight the extraordinary genome flexibility of pathogenic S. epidermidis.

A new monoclonal antibody, P2A8(6), that specifically recognizes a novel epitope on the multidrug resistance-associated protein 1 (MRP1), but not on MRP2 nor MRP3

Multidrug resistance (NIDR) is a major problem in the chemotherapeutic treatment of cancer. Overexpression of the multidrug resistance-associated protein 1 (MRP1), is associated with NIDR in certain tumors. A number of MRP1-specific MAbs, which facilitate both clinical and experimental investigations of this protein, are available. To add to this panel of existing antibodies, we have now generated an additional MRP1-specific monoclonal antibody (MAb), P2A8(6), which detects a unique heat stable epitope on the MRP1 molecule. Female Wistar rats were immunized via footpad injections with a combination of two short synthetic peptides corresponding to amino acids 235-246 (peptide A) and 246-260 (peptide B) of the MRP1 protein. Immune reactive B cells were then isolated from the popliteal lymph nodes for fusion with SP2/O-Ag14 myeloma cells. Resultant hybridoma supernatants were screened for MRP1-specific antibody production. Antibody P2A8(6) was characterized by Western blotting and immunocytochemistry on paired multidrug resistant (MRP1 overexpressing) and sensitive parental cell lines. The antibody detects a protein of 190 kDa in MRP1-expressing cell lines but not in MRP2- or MRP3-transfected cell lines. P2A8(6) stains drug-selected and MRP1-transfected cell lines homogeneously by immunocytochemistry and recognizes MRP1 by immunohistochemistry on formalin-fixed paraffin wax-embedded tissue sections. Peptide inhibition studies confirm that P2AS(6) reacts with peptide B (amino acids 246-260), therefore recognizing a different epitope from that of all currently available MRP1 MAbs. This new MAb, chosen for its specificity to the MRP1 protein, may be a useful addition to the currently available range of MRP1-specific MAbs.

Neural adaptations to resistance training - Implications for movement control

It has long been believed that resistance training is accompanied by changes within the nervous system that play an important role in the development of strength. Many elements of the nervous system exhibit the potential for adaptation in response to resistance training, including supraspinal centres, descending neural tracts, spinal circuitry and the motor end plate connections between motoneurons and muscle fibres. Yet the specific sites of adaptation along the neuraxis have seldom been identified experimentally, and much of the evidence for neural adaptations following resistance training remains indirect. As a consequence of this current lack of knowledge, there exists uncertainty regarding the manner in which resistance training impacts upon the control and execution of functional movements. We aim to demonstrate that resistance training is likely to cause adaptations to many neural elements that are involved in the control of movement, and is therefore likely to affect movement execution during a wide range of tasks.

Expression of multidrug resistance markers ABCB1 (MDR-1/P-gp) and ABCC1 (MRP-1) in renal cell carcinoma.

Background Renal cell carcinoma patients respond poorly to conventional chemotherapy, this unresponsiveness may be attributable to multidrug resistance (MDR). The mechanisms of MDR in renal cancer are not fully understood and the specific contribution of ABC transporter proteins which have been implicated in the chemoresistance of various cancers has not been fully defined in this disease. Methods In this retrospective study the expression of two of these transporter efflux pumps, namely MDR-1 P-gp (ABCB1) and MRP-1 (ABCC1) were studied by immunohistochemistry in archival material from 95 renal cell carcinoma patients. Results In the first study investigating MDR-1 P-gp and MRP-1 protein expression patterns in renal cell carcinoma patients, high levels of expression of both efflux pumps are observed with 100% of tumours studied showing MDR-1 P-gp and MRP-1 positivity. Conclusion Although these findings do not prove a causal role, the high frequency of tumours expressing these efflux pumps suggests that they may be important contributors to the chemoresistance of this tumour type.

A Systems Biology Approach Identifies SART1 as a Novel Determinant of Both 5-Fluorouracil and SN38 Drug Resistance in Colorectal Cancer

Chemotherapy response rates for advanced colorectal cancer remain disappointingly low, primarily because of drug resistance, so there is an urgent need to improve current treatment strategies. To identify novel determinants of resistance to the clinically relevant drugs 5-fluorouracil (5-FU) and SN38 (the active metabolite of irinotecan), transcriptional profiling experiments were carried out on pretreatment metastatic colorectal cancer biopsies and HCT116 parental and chemotherapy-resistant cell line models using a disease-specific DNA microarray. To enrich for potential chemoresistance-determining genes, an unsupervised bioinformatics approach was used, and 50 genes were selected and then functionally assessed using custom-designed short interfering RNA(siRNA) screens. In the primary siRNA screen, silencing of 21 genes sensitized HCT116 cells to either 5-FU or SN38 treatment. Three genes (RAPGEF2, PTRF, and SART1) were selected for further analysis in a panel of 5 colorectal cancer cell lines. Silencing SART1 sensitized all 5 cell lines to 5-FU treatment and 4/5 cell lines to SN38 treatment. However, silencing of RAPGEF2 or PTRF had no significant effect on 5-FU or SN38 sensitivity in the wider cell line panel. Further functional analysis of SART1 showed that its silencing induced apoptosis that was caspase-8 dependent. Furthermore, silencing of SART1 led to a downregulation of the caspase-8 inhibitor, c-FLIP, which we have previously shown is a key determinant of drug resistance in colorectal cancer. This study shows the power of systems biology approaches for identifying novel genes that regulate drug resistance and identifies SART1 as a previously unidentified regulator of c-FLIP and drug-induced activation of caspase-8. Mol Cancer Ther; 11(1); 119-31. (C) 2011 AACR.

Non-genetic mechanisms communicating antibiotic resistance:rethinking strategies for antimicrobial drug design

Introduction: Infections by multidrug-resistant bacteria are of great concern worldwide. In many cases, resistance is not due to the presence of specific antibiotic-modifying enzymes, but rather associated with a general impermeability of the bacterial cell envelope. The molecular bases of this intrinsic resistance are not completely understood. Moreover, horizontal gene transfers cannot solely explain the spread of intrinsic resistance among bacterial strains. Areas covered: This review focuses on the increased intrinsic antibiotic resistance mediated by small molecules. These small molecules can either be secreted from bacterial cells of the same or different species (e.g., indole, polyamines, ammonia, and the Pseudomonas quinolone signal) or be present in the bacterial cell milieu, whether in the environment, such as indole acetic acid and other plant hormones, or in human tissues and body fluids, such as polyamines. These molecules are metabolic byproducts that act as infochemicals and modulate bacterial responses toward antibiotics leading to increasing or decreasing resistance levels. Expert opinion: The non-genetic mechanisms of antibiotic response modulation and communication discussed in this review should reorient our thinking of the mechanisms of intrinsic resistance to antibiotics and its spread across bacterial cell populations. The identification of chemical signals mediating increased intrinsic antibiotic resistance will expose novel critical targets for the development of new antimicrobial strategies.

Reconstitution of O-specific lipopolysaccharide expression in Burkholderia cenocepacia strain J2315, which is associated with transmissible infections in patients with cystic fibrosis

Burkholderia cenocepacia is an opportunistic bacterium that infects patients with cystic fibrosis. B. cenocepacia strains J2315, K56-2, C5424, and BC7 belong to the ET12 epidemic clone, which is transmissible among patients. We have previously shown that transposon mutants with insertions within the O antigen cluster of strain K56-2 are attenuated for survival in a rat model of lung infection. From the genomic DNA sequence of the O antigen-deficient strain J2315, we have identified an O antigen lipopolysaccharide (LPS) biosynthesis gene cluster that has an IS402 interrupting a predicted glycosyltransferase gene. A comparison with the other clonal isolates revealed that only strain K56-2, which produced O antigen and displayed serum resistance, lacked the insertion element inserted within the putative glycosyltransferase gene. We cloned the uninterrupted gene and additional flanking sequences from K56-2 and conjugated this plasmid into strains J2315, C5424, and BC7. All the exconjugants recovered the ability to form LPS O antigen. We also determined that the structure of the strain K56-2 O antigen repeat, which was absent from the LPS of strain J2315, consisted of a trisaccharide unit made of rhamnose and two N-acetylgalactosamine residues. The complexity of the gene organization of the K56-2 O antigen cluster was also investigated by reverse transcription-PCR, revealing several transcriptional units, one of which also contains genes involved in lipid A-core oligosaccharide biosynthesis.

A PCR-based method to characterise and identify benzimidazole resistance in Helminthosporium solani

Control of Helminthosporium solani, the cause of silver scurf in potato tubers, has been impaired by selection of benzimidazole-resistant strains as a result of repeated use of the fungicide thiabendazole. Identification of thiabendazole-resistant strains of H. solani by conventional techniques takes several weeks. Primers designed from conserved regions of the fungal beta-tubulin gene were used to PCR amplify and sequence a portion of the gene. A point mutation was detected at codon 198 in thiabendazole-resistant isolates causing a change in the amino acid sequence from glutamic acid to alanine or glutamine. Species-specific PCR primers designed to amplify this region were used in conjunction with a restriction endonuclease to cause cleavage in sensitive isolates only and thus provide a rapid diagnostic test to differentiate field isolates.

Identification of benzimidazole resistance in Cladobotryum dendroides using a PCR-based method

Cladobotryutn dendroides, causal agent of cobweb disease of Agaricus bisporus, has become increasingly resistant to methylbenzimidazole carbamate (MBC) fungicides following the extensive use of MBC in cultivated mushroom production in Ireland. Of 38 isolates of C. dendroides obtained from Irish mushroom units, 34 were resistant to carbendazim. Primers based on conserved regions of the -tubulin gene were used to amplify and sequence a portion of the -tubulin gene in C. dendroides. A point mutation was detected at codon 50 in isolates resistant to benzimidazole fungicides, causing an amino acid substitution from tyrosine to cysteine. Species-specific PCR primers were designed to amplify the region of the -tubulin gene containing this substitution. The point mutation removed an Ace I restriction site in the -tubulin gene sequence of resistant isolates. Digestion of the PCR product with Ace I thus provides a rapid diagnostic test to differentiate sensitive and resistant isolates of this fungus. EMBL accession number: YI2256.