The importance of homologous recombination in the generation of large deletions in hybrid plasmids in Amycolatopsis mediterranei

The whole nucleotide sequence of pT3.2I, the smallest plasmid of the acidophilic bacterium Thiobacillus T3.2, has been determined. pT3.2I is 15,390 bp long with a 53.7% GC content. Different regions can be defined in it: one 2569-bp putative insertion sequence similar to other insertion sequences of some Agrobacterium Ti plasmids; and a longer sequence, which occurs in two almost identical copies, differing only in a 1-bp deletion (6406 and 6405 bp). Several open reading frames and some smaller sequences were found in this duplicated region: ORFA and ORFG, encoding a putative polyol dehydrogenase and a putative RepA replication protein, respectively, an 83-bp sequence which could code for an antisense RNA, and a 36-bp region highly homologous to ori sequences of ColE2- and ColE3-related plasmids. Another putative gene, ORFH, is only present in the longer copy of this region (it is deleted in the short copy) and might encode a 90-amino-acid polypeptide which could act as a second replication protein, RepB. Based on sequence comparisons, pT3. 2I can be related to plasmids in the pColE2-CA42 incB incompatibility group.     

Clonal propagation of Acacia catechu Willd. by shoot tip culture

A method is described for in vitromicropropagation through shoot apices of Acaciacatechu Willd., a semi-arid tree valued for Katha (atanin-like substance obtained from red heart wood of10–20 year old trees) and timber. Explants wereexcised from 15-days-old in vitro grownseedlings raised from superior seed stocks. Shoot budinduction from shoot apex explants was observed onMurashige and Skoog's (MS) [12] medium containingvarious growth regulators. A maximum of 12 shoots wasobtained on MS medium supplemented with 1.5 mg/l 6-benzylaminopurine (BAP) and 1.5 mg/l kinetin.Well-developed shoots (3–4 cm long) were rooted on strength MS medium with 3.0 mg/l indole-3-acetic acid (IAA) and sucrose 1.5%. In vitro regenerated plantlets of A. catechu were transferred to field conditions.     

A critical evaluation of copper metabolism in indian wilson’s disease children with special reference to their phenotypes and relatives

Wilson’s disease is an autosomal recessive disorder of copper accumulation in various organs, with most common clinical manifestations such as hepatic, neurological, and renal dysfunctions. Serum copper and ceruloplasmin in Wilson’s disease were significantly lower as compared to normals, controls, and relatives of Wilson’s disease patients, whereas marked hypercupriuria (145 ± 7 μg/24 h) was observed in Wilson’s children only. A good correlation (r = 0.92) was found between non-ceruloplasmin-bound copper and 24-h urinary copper excretion in Wilson’s disease patients. Further, copper studies among the different phenotypes of Wilson’s disease revealed substantially low serum ceruloplasmin and a marked hypercupriuria in Wilson’s disease children associated with renal tubular acidosis as compared to the patients with either hepatological or neurological manifestations. Serum ceruloplasmin levels in 14 patients of Wilson’s disease were between 14 and 20 mg/dL. These patients of Wilson’s disease were confirmed by measuring liver biopsy copper, which was about nine times higher than normal hepatic copper content. During the family screening by copper studies, four asymptomatic siblings were diagnosed for Wilson’s disease. These subjects were then started on D-penicillamine therapy because presymptomatic treatment prevents progression of the disease complications.     

Gastrointestinal parasites of the chimpanzee population introduced onto Rubondo Island National Park,Tanzania

The release of any species into a novel environment can evoke transmission of parasites that do not normally parasitize the host as well as potentially introducing new parasites into the environment. Species introductions potentially incur such risks, yet little is currently known about the parasite fauna of introduced primate species over the long term. We describe the results of long‐term monitoring of the intestinal parasite fauna of an unprovisioned, reproducing population of chimpanzees introduced 40 years earlier (1966–1969) onto Rubondo Island in Lake Victoria, Tanzania, a non‐native habitat for chimpanzees. Two parasitological surveys (March 1997–October 1998 and October 2002–December 2005) identified Entamoeba spp. including E. coli, Iodamoeba buetschlii, Troglodytella abrassarti, Chilomastix mesnili, Trichuris sp., Anatrichosoma sp., Strongyloides spp., Strongylida fam. gen. sp., Enterobius anthropopitheci, Subulura sp., Ascarididae gen. sp., and Protospirura muricola. The parasite fauna of the Rubondo chimpanzees is similar to wild chimpanzees living in their natural habitats, but Rubondo chimpanzees have a lower prevalence of strongylids (9%, 3.8%) and a higher prevalence of E. anthropopitheci (8.6%, 17.9%) than reported elsewhere. Species prevalence was similar between our two surveys, with the exception of Strongyloides spp. being higher in the first survey. None of these species are considered to pose a serious health risk to chimpanzees, but continued monitoring of the population and surveys of the parasitic fauna of the two coinhabitant primate species and other animals, natural reservoir hosts of some of the same parasites, is important to better understand the dynamics of host–parasite ecology and potential long‐term implications for chimpanzees introduced into a new habitat. Am. J. Primatol. 72:307–316, 2010. © 2009 Wiley‐Liss, Inc.     

Role of root-mediated interactions in phytotoxic interference of Ageratum conyzoides with rice (Oryza sativa)

Ageratum conyzoides L. (billy goat weed; Asteraceae) is an annual invasive weed native of tropical America and has now naturalized worldwide, particularly in Southeast Asia. The present study investigated the nature and potential of root-mediated allelopathic interference of A. conyzoides against rice (Oryza sativa). Root and shoot length and biomass accumulation of rice were significantly reduced (by 18–30%) when grown in the rhizosphere soil of the weed indicating the release of putative allelochemicals from the weed into the soil. The growth of rice was also progressively reduced in the soil amended with increasing amounts of root residues (5, 10 and 20 g kg⁻¹ soil) of A. conyzoides. The addition of activated charcoal, an inert material with high affinity for organic biomolecules, partly ameliorated the negative effects of root residues amended in the soil. Further, there was no negative effect on the availability of soil nutrients in the root-amended soils. These were rather nutrient rich with greater electrical conductivity, and higher amount of organic matter, thus indicating no role in observed growth reduction. The reduction in allelopathic effects of root residue upon charcoal addition further indicated that putative phytotoxins released from the weed roots are water-soluble phenolic compounds. A significant amount of water-soluble phenolics were present in rhizosphere (∼6-times higher) and root-amended soils (∼5–10-fold higher) and their content was reduced (to ∼3.6–7.0-fold higher) when charcoal was added. The observed growth reduction in Ageratum rhizospheric or root-amended soils was concomitant with the amount of phenolic compounds. Upon HPLC analyses, these were identified as p-coumaric acid, gallic acid, ferulic acid, p-hydroxybenzoic acid and anisic acid. Under laboratory conditions, these phenolic acids reduced the root length and seedling weight of rice individually as well as in equimolar mixture, though no synergistic effect was noticed. The study concludes that root exudates and residues of A. conyzoides suppress the growth of rice by releasing phenolic allelochemicals into the soil rhizosphere and not through alteration of soil nutrients, and allelopathy plays a significant role in root-mediated negative interference of A. conyzoides.     

Design,synthesis and evaluations of acridone derivatives using Candida albicans—Search for MDR modulators led to the identification of an anti-candidiasis agent

In order to search for MDR modulators, rationally designed acridone derivatives were investigated for their effect on influx or efflux of Rhodamine6G (R6G) in CAI4 cells. Results of these investigations indicate that in presence of compound 12, inhibition of growth of CAI4 cells and also an increased influx/efflux of R6G in CAI4 cells have been observed. This seems to be occurring due to the cell wall rupturing of Candida albicans. Compound 12 may be a suitable candidate for candidiasis therapy.     

Disrupting Protein Expression with Peptide Nucleic Acids Reduces Infection by Obligate Intracellular Rickettsia

Peptide Nucleic Acids (PNAs) are single-stranded synthetic nucleic acids with a pseudopeptide backbone in lieu of the phosphodiester linked sugar and phosphate found in traditional oligos. PNA designed complementary to the bacterial Shine-Dalgarno or start codon regions of mRNA disrupts translation resulting in the transient reduction in protein expression. This study examines the use of PNA technology to interrupt protein expression in obligate intracellular Rickettsia sp. Their historically intractable genetic system limits characterization of protein function. We designed PNA targeting mRNA for rOmpB from Rickettsia typhi and rickA from Rickettsia montanensis, ubiquitous factors important for infection. Using an in vitro translation system and competitive binding assays, we determined that our PNAs bind target regions. Electroporation of R. typhi and R. montanensis with PNA specific to rOmpB and rickA, respectively, reduced the bacteria’s ability to infect host cells. These studies open the possibility of using PNA to suppress protein synthesis in obligate intracellular bacteria.     

Therapeutic targeting of the thrombospondin-1 receptor CD47 to treat liver cancer

CD47 is a signaling receptor for the matricellular protein thrombospondin-1 and a counter-receptor for signal regulatory protein-α (SIRPα) on macrophages. Following its initial discovery in 1992 as a cell surface protein that is over-expressed by ovarian carcinoma, elevated CD47 expression has emerged as a negative prognostic factor for a variety of cancers. CD47 is also a potential therapeutic target based on the ability of CD47 blockade to cause regression of tumors in mice, and a humanized CD47 antibody has recently entered phase I clinical trials. CD47 blockade may control tumor growth by inhibiting thrombospondin-1 signaling or by preventing inhibitory SIRPα signaling in tumor-associated macrophages. A recent publication by Lee et al. (Hepatology 60:179–191, 2014) provides evidence that blocking CD47 signaling specifically depletes tumor-initiating stem cells in hepatocellular carcinoma and implicates cathepsin-S/protease-activated receptor-2 signaling in mediating this therapeutic response.     

Quality of Artemisinin-Based Combination Formulations for Malaria Treatment: Prevalence and Risk Factors for Poor Quality Medicines in Public Facilities and Private Sector Drug Outlets in Enugu,Nigeria

BackgroundArtemisinin-based combination therapies are recommended by the World Health Organisation (WHO) as first-line treatment for Plasmodium falciparum malaria, yet medication must be of good quality for efficacious treatment. A recent meta-analysis reported 35% (796/2,296) of antimalarial drug samples from 21 Sub-Saharan African countries, purchased from outlets predominantly using convenience sampling, failed chemical content analysis. We used three sampling strategies to purchase artemisinin-containing antimalarials (ACAs) in Enugu metropolis, Nigeria, and compared the resulting quality estimates.MethodsACAs were purchased using three sampling approaches - convenience, mystery clients and overt, within a defined area and sampling frame in Enugu metropolis. The active pharmaceutical ingredients were assessed using high-performance liquid chromatography and confirmed by mass spectrometry at three independent laboratories. Results were expressed as percentage of APIs stated on the packaging and used to categorise each sample as acceptable quality, substandard, degraded, or falsified.ResultsContent analysis of 3024 samples purchased from 421 outlets using convenience (n=200), mystery (n=1,919) and overt (n=905) approaches, showed overall 90.8% ACAs to be of acceptable quality, 6.8% substandard, 1.3% degraded and 1.2% falsified. Convenience sampling yielded a significantly higher prevalence of poor quality ACAs, but was not evident by the mystery and overt sampling strategies both of which yielded results that were comparable between each other. Artesunate (n=135; 4 falsified) and dihydroartemisinin (n=14) monotherapy tablets, not recommended by WHO, were also identified.ConclusionRandomised sampling identified fewer falsified ACAs than previously reported by convenience approaches. Our findings emphasise the need for specific consideration to be given to sampling frame and sampling approach if representative information on drug quality is to be obtained.