Physical and genetic mapping of cloned ribosomal DNA from Toxoplasma gondii: primary and secondary structure of the 5S gene.

The ribosomal DNA (rDNA encoding rRNA) of the obligately intracellular protozoan parasite, Toxoplasma gondii, was identified, cloned, physically mapped, its copy number determined, and the 5S gene sequenced. Using total RNA as a probe, a collection of recombinant lambda phages containing copies of rDNA were isolated from a lambda 2001 tachyzoite genomic library. Northern gel hybridization confirmed specific homology of the 7.5-kb rDNA unit, subcloned into pTZ18R, to T. gondii rRNA. The mapped rDNA found in pTOX1 contained small ribosomal subunit (SS; 18S)- and large ribosomal subunit (LS; 26S)-encoding genes localized using intragenic heterologous probes from the conserved sequences of the SS (18S) and LS (28S) Xenopus laevis genes. the physical mapping data, together with partial digestion experiments and Southern gel hybridization, confirmed a 7.5-kb rDNA unit arranged in a simple head-to-tail fashion that is tandemly repeated. We estimated the rDNA repeat copy number in T. gondii to be 110 copies per haploid tachyzoite genome. Parts of the SS gene and the complete 5S gene were sequenced. The 5S gene was found to be within the rDNA locus, a rare occurrence found only in some fungi and protozoa. Secondary-structure analysis revealed an organization remarkably similar to the 5S RNA of eukaryotes.     

Disentangling thermal from alternative drivers of reflectance in jewel beetles: A macroecological study

Aim

To predict future colour–climate relationships, it is important to distinguish thermal drivers of reflectance from other evolutionary drivers. We aimed to achieve this by comparing relationships between climate and coloration in ultraviolet–visible (UV–Vis) and near-infrared (NIR) light, separately.

Location

Samples were distributed primarily across Australia and North America, with some from Africa and Asia.

Major taxa studied

Coleoptera: Buprestidae.

Methods

We used jewel beetles as models to identify climatic drivers of reflectance, because jewel beetles have highly diverse coloration and a wide distribution and are often active in hot conditions. Specifically, we tested the association between climate, body size and reflectance using a phylogenetic comparative analysis for three wavebands (UV–Vis, NIR and total).

Results

Reflectance of jewel beetles was more strongly predicted by body size than by climate. NIR reflectance and total reflectance were not associated with climate, but larger beetles had higher NIR reflectance. For UV–Vis reflectance, small beetles were darker in warmer and more humid environments, whereas there was no association with climate for large beetles.

Main conclusions

Our study suggests that variation in reflectance of jewel beetles is not driven by thermal requirements and highlights the importance of considering NIR reflectance when evaluating explanations of the effects of colour on thermoregulation.     

Stress relaxation of swine growth plate in semi-confined compression: depth dependent tissue deformational behavior versus extracellular matrix composition and collagen fiber organization

Mechanical environment is one of the regulating factors involved in the process of longitudinal bone growth. Non-physiological compressive loading can lead to infantile and juvenile musculoskeletal deformities particularly during growth spurt. We hypothesized that tissue mechanical behavior in sub-regions (reserve, proliferative and hypertrophic zones) of the growth plate is related to its collagen and proteoglycan content as well as its collagen fiber orientation. To characterize the strain distribution through growth plate thickness and to evaluate biochemical content and collagen fiber organization of the three histological zones of growth plate tissue. Distal ulnar growth plate samples (N = 29) from 4-week old pigs were analyzed histologically for collagen fiber organization (N = 7) or average zonal thickness (N = 8), or trimmed into the three average zones, based on the estimated thickness of each histological zone, for biochemical analysis of water, collagen and glycosaminoglycan content (N = 7). Other samples (N = 7) were tested in semi-confined compression under 10 % compressive strain. Digital images of the fluorescently labeled nuclei were concomitantly acquired by confocal microscopy before loading and after tissue relaxation. Strain fields were subsequently calculated using a custom-designed 2D digital image correlation algorithm. Depth-dependent compressive strain patterns and collagen content were observed. The proliferative and hypertrophic zone developed the highest axial and transverse strains, respectively, under compression compared to the reserve zone, in which the lowest axial and transverse strains arose. The collagen content per wet mass was significantly lower in the proliferative and hypertrophic zones compared to the reserve zone, and all three zones had similar glycosaminoglycan and water content.Polarized light microscopy showed that collagen fibers were mainly organized horizontally in the reserve zone and vertically aligned with the growth direction in the proliferative and hypertrophic zones. Higher strains were developed in growth plate areas (proliferative and hypertrophic) composed of lower collagen content and of vertical collagen fiber organization. The stiffer reserve zone, with its higher collagen content and collagen fibers oriented to restrain lateral expansion under compression, could play a greater role of mechanical support compared to the proliferative and hypertrophic zones, which could be more susceptible to be involved in an abnormal growth process.     

Metabolic vulnerability of cisplatin‐resistant cancers

Cisplatin is the most widely used chemotherapeutic agent, and resistance of neoplastic cells against this cytoxicant poses a major problem in clinical oncology. Here, we explored potential metabolic vulnerabilities of cisplatin‐resistant non‐small human cell lung cancer and ovarian cancer cell lines. Cisplatin‐resistant clones were more sensitive to killing by nutrient deprivation in vitro and in vivo than their parental cisplatin‐sensitive controls. The susceptibility of cisplatin‐resistant cells to starvation could be explained by a particularly strong dependence on glutamine. Glutamine depletion was sufficient to restore cisplatin responses of initially cisplatin‐resistant clones, and glutamine supplementation rescued cisplatin‐resistant clones from starvation‐induced death. Mass spectrometric metabolomics and specific interventions on glutamine metabolism revealed that, in cisplatin‐resistant cells, glutamine is mostly required for nucleotide biosynthesis rather than for anaplerotic, bioenergetic or redox reactions. As a result, cisplatin‐resistant cancers became exquisitely sensitive to treatment with antimetabolites that target nucleoside metabolism.     

Gill tissue recovery after copper exposure in Carassius gibelio (Pisces: Cyprinidae)

We investigated the influence of copper in a long-time treatment with concentrations of 0.05 mg L^?1 and 0.1 mg L^?1 to track the histopathological changes in gills of Carassius gibelio, and to find at what extent they will recover after the effect of the copper concentrations stops. Treatment with copper lasted 21 days and the recovery time was of the same duration. The results of histological examination showed degenerative changes (resulting in thinner secondary lamellae and filamentary epithelium), and hyperplastic and hypertrophic changes (proliferation, vasodilatation, aneurysms, epithelial interstitial edema, and fusion) in gills under the influence of two concentrations. The degenerative changes have higher prevalence at low concentrations, while hyperplastic and hypertrophic ones — at high concentrations. After the period of recovery they remained the same, but the extent of expression on the surface of gill filaments changed. The long-time copper intoxication in low concentrations of copper affects gill structure, causing severe changes whose recovery is a slow process that requires a longer period of time.     

Phenomic and genomic approaches to studying the inhibition of multiresistant Salmonella enterica by microcin J25

In livestock production, antibiotics are used to promote animal growth, control infections and thereby increase profitability. This practice has led to the emergence of multiresistant bacteria such as Salmonella, of which some serovars are disseminated in the environment. The objective of this study is to evaluate microcin J25 as an inhibitor of Salmonella enterica serovars of various origins including human, livestock and food. Among the 116 isolates tested, 37 (31.8%) were found resistant to at least one antibiotic, and 28 were multiresistant with 19 expressing the penta-resistant phenotype ACSSuT. Microcin J25 inhibited all isolates, with minimal inhibitory concentration values ranging from 0.06 μg/ml (28.4 nM) to 400 μg/ml (189 μM). Interestingly, no cross-resistance was found between microcin J25 and antibiotics. Multiple sequence alignments of genes encoding for the different proteins involved in the recognition and transport of microcin J25 showed that only ferric-hydroxamate uptake is an essential determinant for susceptibility of S. enterica to microcin J25. Examination of Salmonella strains exposed to microcin J25 by transmission electronic microscopy showed for the first-time involvement of a pore formation mechanism. Microcin J25 was a strong inhibitor of several multiresistant isolates of Salmonella and may have a great potential as an alternative to antibiotics.