Prevalence and genetic diversity of bartonella species detected in different tissues of small mammals in Nepal

Bartonellae were detected in a total of 152 (23.7%) of 642 tissues from 108 (48.4%) of 223 small mammals trapped in several urban areas of Nepal. Based on rpoB and gltA sequence analyses, genotypes belonging to seven known Bartonella species and five genotypes not belonging to previously known species were identified in these animals.     

Tissue engineering for clinical applications

Tissue engineering is increasingly being recognized as a beneficial means for lessening the global disease burden. One strategy of tissue engineering is to replace lost tissues or organs with polymeric scaffolds that contain specialized populations of living cells, with the goal of regenerating tissues to restore normal function. Typical constructs for tissue engineering employ biocompatible and degradable polymers, along with organ-specific and tissue-specific cells. Once implanted, the construct guides the growth and development of new tissues; the polymer scaffold degrades away to be replaced by healthy functioning tissue. The ideal biomaterial for tissue engineering not only defends against disease and supports weakened tissues or organs, it also provides the elements required for healing and repair, stimulates the body's intrinsic immunological and regenerative capacities, and seamlessly interacts with the living body. Tissue engineering has been investigated for virtually every organ system in the human body. This review describes the potential of tissue engineering to alleviate disease, as well as the latest advances in tissue regeneration. The discussion focuses on three specific clinical applications of tissue engineering: cardiac tissue regeneration for treatment of heart failure; nerve regeneration for treatment of stroke; and lung regeneration for treatment of chronic obstructive pulmonary disease.     

Generation of mice with a conditional null allele for Wntless

The Wnt‐signaling pathway is necessary in a variety of developmental processes and has been implicated in numerous pathologies. Wntless (Wls) binds to Wnt proteins and facilitates Wnt sorting and secretion. Conventional deletion of Wls results in early fetal lethality due to defects in body axis establishment. To gain insight into the function of Wls in later stages of development, we have generated a conditional null allele. Homozygous germline deletion of Wls confirmed prenatal lethality and failure of embryonic axis formation. Deletion of Wls using Wnt1‐cre phenocopied Wnt1 null abnormalities in the midbrain and hindbrain. In addition, conditional deletion of Wls in pancreatic precursor cells resulted in pancreatic hypoplasia similar to that previously observed after conditional β‐catenin deletion. This Wls conditional null allele will be valuable in detecting novel Wnt functions in development and disease. genesis 48:554–558, 2010. © 2010 Wiley‐Liss, Inc.     

Procathepsin D expression correlates with invasive and metastatic phenotype of MDA-MB-231 derived cell lines

Procathepsin D (pCD) is a glycoprotein secreted abundantly by cancerous cells with a documented role in tumor development. The levels of pCD in primary tumors are highly correlated with an increased incidence of metastasis. Our earlier studies have shown that pCD exerts its effect on cancer cells through its activation peptide (AP) and involves both autocrine and paracrine modes of action. In this study, we analyzed the expression and role of pCD in MDA-MB-231 and its derived cell lines 1833 and 4175 possessing discrete metastatic abilities. Our results demonstrated a direct relationship between expression and secretion of pCD to the differential invasive potential of these cells. Also, the cell lines responded to AP treatment by enhancing their invasive potential, proliferation and induction of secretion of various cytokines, suggesting that pCD plays a role in metastasis through its AP region.     

Phenotypes and fed-batch fermentation of ubiquinone-overproducing fission yeast using ppt1 gene

Ubiquinone (UQ), a component of the electron transfer system in many organisms, has been widely used for pharmaceuticals and cosmetics. In this study, we cloned and overexpressed the full-length ppt1 (MTppt1) gene, which encodes p-hydroxybenzoate:polyprenyltransferase and ERppt1 gene, which was modified to be localized on endoplasmic reticulum in fission yeast. The yeast MTppt1 and ERppt1 transgenic lines showed about 3.7 and 5.1 times increment in UQ content and the recombinant yeasts with a higher UQ level are more resistant to H(2)O(2), Cu(2+) and NaCl, and interestingly their growth was also faster than the wild type at lower temperature. For large-scale cultivation, the direct feedback control of glucose using an on-line ethanol concentration monitor for ubiquinone production of yeast ERppt1 by high-cell-density fermentation was investigated and the fermentation parameters (e.g., dissolved oxygen, pH, ethanol concentration, oxygen uptake rate, carbon dioxide evolution rate and respiration quotient) were also discussed. After 90 h cultures, the yeast dry cell weight reached 57 gl(-1) and the ubiquinone yield reached 23 mgl(-1). In addition, plasmid stability was maintained at high level throughout the fermentation.     

An entomopathogenic bacterium, Xenorhabdus nematophila, inhibits hemocyte phagocytosis of Spodoptera exigua by inhibiting phospholipase A(2)

Phagocytosis is a hemocytic behavior against bacterial infection. An entomopathogenic bacterium, Xenorhabdus nematophila, inhibits immune responses of target insects and causes hemolymph septicemia. This study analyzed how X. nematophila could inhibit phagocytosis to increase its pathogenicity. Granular cells and plasmatocytes were the main phagocytic hemocytes of Spodoptera exigua determined by observing fluorescence-labeled bacteria in the cytosol. X. nematophila significantly inhibited phagocytosis of both hemocytes, while heat-killed X. nematophila lost its inhibitory potency. However, co-injection of X. nematophila with arachidonic acid did not show any significant inhibition of hemocyte phagocytosis. In fact, hemocytes of S. exigua infected with X. nematophila showed significant reduction in phospholipase A(2) (PLA(2)) activity. Dexamethasone, a specific PLA(2) inhibitor, significantly inhibited phagocytosis of both cell types. However, the inhibitory effect of dexamethasone was recovered by addition of arachidonic acid. Incubation of hemocytes with benzylideneacetone, a metabolite of X. nematophila, inhibited phagocytosis in a dose-dependent manner. These results suggest that X. nematophila produces and secretes PLA(2) inhibitor(s), which in turn inhibit the phagocytic response of hemocytes.     

The role of a heat shock protein from V. cholerae 0139 in the gut immune response

An immunodominant heat shock protein (Hsp 24) was purified from Vibrio cholerae O139 at 42 degrees C and used as an immunomodulator for studying the gut immune response. T cell clone and T cell line specific for the Hsp 24 were generated from the lymphocytes of lamina propria and intra-epithelial lymphocytes of mice orally infected with V. cholerae O139, respectively. The T cell clone was TCR alphabeta(+), CD4(+) and appeared to play an important role in the functioning of gut B-lymphocytes. The T cell line had heterogenous population of CD8+ and CD4+ cells, most of which were found to be TCR alphabeta(+) and a minor population was TCR gammadelta(+). The lymphokine profile of T cell line showed IFN-gamma to be the most abundant lymphokine followed by IL-2 and IL-4. The possible involvement of alternative pathway of activation for T cell clone was also addressed in this study. The splenocytes showed an up-regulation of their CD2 receptor expression on stimulation with the Hsp-24. The pattern of lymphokines released by splenocytes stimulated with the Hsp-24 showed no particular cell type to be responsible for mounting immune response. Thus, there is involvement of both, mucosal and peripheral arm of the immune system.     

Galactose-Specific Fimbrial Adhesin of Enteroaggregative <Emphasis Type="Italic">Escherichia coli</Emphasis>: A Possible Aggregative Factor

A galactose-specific adhesin was isolated from the fimbriae of an enteroaggregative Escherichia coli (EAEC) strain. The adhesin was found to be a high molecular weight aggregate of the 18-kDa monomer. The dimeric (36 kDa) and tetrameric (76 kDa) forms appeared in sodium dodecyl sulphate polyacrylamide gel electrophoresis when a higher concentration of the adhesin was used. The IgG_AD (IgG against adhesin) obtained from the immune sera raised in rabbits against purified adhesin could detect all three forms of the adhesin even from the crude fimbrial preparation. The IgG_AD failed to recognize the adhesin in the presence of galactose, thereby suggesting the antibody-binding site and the sugar-binding site on the adhesin might be same or overlapping. Furthermore, the IgG_AD could localize the adhesin exclusively on the fimbriae as observed in immunogold electron microscopy. The aggregative adherence of the bacteria to HEp-2 cells was reduced to 70% in the presence of the IgG_AD. A glycoprotein (34 kDa) present in the membrane fraction of HEp-2 cells interacted with the purified adhesin in a galactose-specific manner. The IgG_AD could recognize the adhesin from the crude fimbrial preparation of 9 out of 10 clinical isolates of EAEC strains but failed to identify any protein from the crude fimbrial preparation of Salmonella typhimurium (fim +ve as well as fim −ve strain), Vibrio cholerae (WO7) or Escherichia coli DH5α.     

PCR-based specific detection of Ralstonia solanacearum by amplification of cytochrome c1 signal peptide sequences

A polymerase chain reaction (PCR)-based method was developed to detect the DNA of Ralstonia solanacearum, the causal agent of bacterial wilt in various crop plants. One pair of primers (RALSF and RALSR), designed using cytochrome c1 signal peptide sequences specific to R. solanacearum, produced a PCR product of 932 bp from 13 isolates of R. solanacearum from several countries. The primer specificity was then tested using DNA from 21 isolates of Ralstonia, Pseudomonas, Burkholderia, Xanthomonas, and Fusarium oxysporum f. sp. dianthi. The specificity of the cytochrome c1 signal peptide sequences in R. solanacearum was further confirmed by a DNA-dot blot analysis. Moreover, the primer pair was able to detect the pathogen in artificially inoculated soil and tomato plants. Therefore, the present results indicate that the primer pair can be effectively used for the detection of R. solanacearum in soil and host plants.     

Identification of two entomopathogenic bacteria from a nematode pathogenic to the Oriental beetle, Blitopertha orientalis

A pathogenic nematode, Butlerius sp., was isolated from Oriental beetle, Blitopertha orientalis. The infective juveniles exhibited dose- as well as time-dependent entomopathogenicity on the larvae of B. orientalis. Two bacterial species, Providencia vermicola (KACC 91278) and Flavobacterium sp. (KACC 91279), were isolated from the infective juveniles and identified. P. vermicola outnumbered Flavobacterium sp. in the nematode host, in which the colony density of P. vermicola was found to be 21 times higher than that of Flavobacterium sp. However, when the two bacterial species were cocultured in culture media without the nematode host, they showed similar growth rates. Both bacteria induced significant entomopathogenicity against Spodoptera exigua larvae infesting economically important vegetable crops, where P. vermicola was more potent than Flavobacterium sp.