Functional Characterization of AbeD,an RND-Type Membrane Transporter in Antimicrobial Resistance in Acinetobacter baumannii

Background

Acinetobacter baumannii is becoming an increasing menace in health care settings especially in the intensive care units due to its ability to withstand adverse environmental conditions and exhibit innate resistance to different classes of antibiotics. Here we describe the biological contributions of abeD, a novel membrane transporter in bacterial stress response and antimicrobial resistance in A. baumannii.

Results

The abeD mutant displayed ~ 3.37 fold decreased survival and >5-fold reduced growth in hostile osmotic (0.25 M; NaCl) and oxidative (2.631 μM–6.574 μM; H₂O₂) stress conditions respectively. The abeD inactivated cells displayed increased susceptibility to ceftriaxone, gentamicin, rifampicin and tobramycin (~ 4.0 fold). The mutant displayed increased sensitivity to the hospital-based disinfectant benzalkonium chloride (~3.18-fold). In Caenorhabditis elegans model, the abeD mutant exhibited (P<0.01) lower virulence capability. Binding of SoxR on the regulatory fragments of abeD provide strong evidence for the involvement of SoxR system in regulating the expression of abeD in A. baumannii.

Conclusion

This study demonstrates the contributions of membrane transporter AbeD in bacterial physiology, stress response and antimicrobial resistance in A. baumannii for the first time.     

Ear culture as a technique to overcome hybrid necrosis in wheat

Hybrid necrosis in wheat is a problem for gene transfer in wheat breeding. Hybrid necrosis occurs due to dominant complementary interaction of two genes, Ne1 and Ne2. A cross between wheat (Triticum aestivum L.) varieties C306 (drought tolerant, Ne1 carrier) and WL711(high yielding, Ne2 carrier) produced necrotic F1 hybrids, which died before ear emergence and produced no seeds. To overcome the problem of hybrid necrosis, ears enclosed in the leaf sheath were taken and cultured to maturity in liquid medium containing 5% sucrose and 0.04% glutamine. The necrotic hybrids produced only a few seeds per ear compared to parents, but individual grain weight was similar in the hybrid and the parents. The F1 ear culture study has been repeated for three years and F2 seeds obtained. In 1996–97, the cultured ears of F1 hybrids produced 62 seeds, of which only 52 showed germination and were grown under normal field conditions. Out of the 52 seeds, 50% were non-necrotic and showed segregation for various physiological traits. The results reveal that hybrids ears had the potential to form viable seeds. Culturing of wheat ears before ear emergence and production of viable F2 seeds from necrotic hybrids is a simple and efficient method for overcoming the problem of hybrid necrosis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Minimum fluidization velocity and friction factor in a liquid-solid inverse fluidized bed reactor

In the present study the hydrodynamic characteristics (bed expansion and pressure drop) of low-density polyethylene (LDPE) and polypropylene (PP) are studied in a liquid-solid inverse fluidized bed reactor as a function of particle diameter, liquid viscosity and density. The bed expansion and pressure drop data are used to determine the minimum fluidization velocity, U_mf and friction factor, P. It was found that the U_mf increased with an increase in the particle diameter and a decrease in solid density and was independent of initial bed height (solid loading). In addition, the U_mf decreased with an increase in the CMC concentration. The friction factor Reynolds number plot was found similar to that of classical fluidization. Dimensionless equations were proposed for the. prediction of the friction factor and the U_mf.     

树鼩的骨元和骨细胞穴微观结构超微影像的定性比较

近年来,由于树鼩与灵长类动物的亲缘关系,它们引起了人类发展和疾病研究的兴趣。在这项对树鼩,鼠,狗,狒狒和人类的骨骼超微形态的比较研究中,我们定性分析了骨骼的微观结构和形态,以评估树鼩对人类的亲近程度。在3只成年雄性树鼩 (滇西亚种) (Tupaia belangeri chinensis）的股骨中,使用荧光素异硫氰酸盐样品制备和染色共聚焦成像研究了皮质骨的骨元结构,并使用酸蚀刻SEM观察了骨细胞穴的形态。总体而言,树鼩中骨样形成物的密度和结构以及骨细胞穴的形态更像鼠, 与人类,狗和狒狒都明显不同。这些发现表明,尽管树鼩在系统发育上比鼠更接近人类,但它们的骨骼超微形态仍与鼠接近。这是除狗和狒狒之外,第一次对树鼩的骨元和骨细胞穴进行超微影像的研究。这个比较研究的结果丰富了我们对早期灵长类动物骨骼发育,适应性和进化的理解。未来有必要进行进一步的定量比较研究来表征树鼩骨骼的微观形态。     

Molecular cloning and expression of the IL-10 gene from guinea pigs

The Guinea pig (Cavia porcellus) is one of the most relevant small animals for modeling human tuberculosis (TB) in terms of susceptibility to low dose aerosol infection, the organization of granulomas, extrapulmonary dissemination and vaccine-induced protection. It is also considered to be a gold standard for a number of other infectious and non-infectious diseases; however, this animal model has a major disadvantage due to the lack of readily available immunological reagents. In the present study, we successfully cloned a cDNA for the critical Th2 cytokine, interleukin-10 (IL-10), from inbred Strain 2 guinea pigs using the DNA sequence information provided by the genome project. The complete open reading frame (ORF) consists of 537 base pairs which encodes a protein of 179 amino acids. This cDNA sequence exhibited 87% homology with human IL-10. Surprisingly, it showed only 84% homology with the previously published IL-10 sequence from the C4-deficient (C4D) guinea pig, leading us to clone IL-10 cDNA from the Hartley strain of guinea pig. The IL-10 gene from the Hartley strain showed 100% homology with the IL-10 sequence of Strain 2 guinea pigs. In order to validate the only published IL-10 sequence existing in Genbank reported from C4D guinea pigs, genomic DNA was isolated from tissues of C4D guinea pigs. Amplification with various sets of primers showed that the IL-10 sequence reported from C4D guinea pigs contained numerous errors. Hence the IL-10 sequence that is being reported by us replaces the earlier sequence making our IL-10 sequence to be the first one accurate from guinea pig. Recombinant guinea pig IL-10 proteins were subsequently expressed in both prokaryotic and eukaryotic cells, purified and were confirmed by N-terminal sequencing. Polyclonal anti-IL-10 antibodies were generated in rabbits using the recombinant IL-10 protein expressed in this study. Taken together, our results indicate that the DNA sequence information provided by the genome project is useful to directly clone much needed cDNAs necessary to study TB in the guinea pig. The newly cloned guinea pig IL-10 cDNA and recombinant proteins will serve as valuable resources for immunological studies in the guinea pig model of TB and other diseases.