Genotype-specific interactions and the trade-off between host and parasite fitness

Background  

Evolution of parasite traits is inextricably linked to their hosts. For instance one common definition of parasite virulence is the reduction in host fitness due to infection. Thus, traits of infection must be viewed in both protagonists and may be under shared genetic and physiological control. We investigated these questions on the oomycete Hyaloperonospora arabidopsis (= parasitica), a natural pathogen of the Brassicaceae Arabidopsis thaliana.     

Long-term,large scale cryopreservation of insect cells at −80 °C

Standard tissue culture methods advise freezing cells in small aliquots (≤1 × 10⁷ cells in 1 mL), and storing in liquid nitrogen. This is inconvenient for laboratories culturing large quantities of insect cells for recombinant baculovirus expression, owing to the length of time taken to produce large scale cultures from small aliquots of cells. Liquid nitrogen storage requires use of specialized cryovials, personal protective equipment and oxygen monitoring systems. This paper describes the long-term, large scale cryopreservation of 8 × 10⁸ insect cells at −80 °C, using standard 50 mL conical tubes to contain a 40 mL cell suspension. Sf9, Sf21 and High 5 cells were recovered with a viability > 90 % after storage for one year under these conditions, which compared favorably with the viability of cells stored in liquid nitrogen for the same length of time. Addition of green fluorescent protein encoding baculovirus demonstrated that cells were “expression ready” immediately post thaw. Our method enables large scale cultures to be recovered rapidly from stocks cryopreserved at −80 °C, thus avoiding the inconvenience, hazards and expense associated with liquid nitrogen.

Electronic supplementary material

The online version of this article (doi:10.1007/s10616-014-9781-5) contains supplementary material, which is available to authorized users.     

Does P-Cresylglucuronide Have the Same Impact on Mortality as Other Protein-Bound Uremic Toxins?

Background

Uremic toxins are emerging as important, non-traditional cardiovascular risk factors in chronic kidney disease (CKD). P-cresol has been defined as a prototype protein-bound uremic toxin. Conjugation of p-cresol creates p-cresylsulfate (PCS) as the main metabolite and p-cresylglucuronide (PCG), at a markedly lower concentration. The objective of the present study was to evaluate serum PCG levels, determine the latter’s association with mortality and establish whether the various protein-bound uremic toxins (i.e. PCS, PCG and indoxylsulfate (IS)) differed in their ability to predict mortality.

Methodology/Principal Findings

We studied 139 patients (mean ± SD age: 67±12; males: 60%) at different CKD stages (34.5% at CKD stages 2–3, 33.5% at stage 4–5 and 32% at stage 5D). A recently developed high-performance liquid chromatography method was used to assay PCG concentrations. Total and free PCG levels increased with the severity of CKD. During the study period (mean duration: 779±185 days), 38 patients died. High free and total PCG levels were correlated with overall and cardiovascular mortality independently of well-known predictors of survival, such as age, vascular calcification, anemia, inflammation and (in predialysis patients) the estimated glomerular filtration rate. In the same cohort, free PCS levels and free IS levels were both correlated with mortality. Furthermore, the respective predictive powers of three Cox multivariate models (free PCS+other risk factors, free IS+other risk factors and free PCS+other risk factors) were quite similar - suggesting that an elevated PCG concentration has much the same impact on mortality as other uremic toxins (such as PCS or IS) do.

Conclusions

Although PCG is the minor metabolite of p-cresol, our study is the first to reveal its association with mortality. Furthermore, the free fraction of PCG appears to have much the same predictive power for mortality as PCS and IS do.     

Production of active pediocin PA-1 in Escherichia coli using a thioredoxin gene fusion expression approach: cloning, expression, purification, and characterization

Antimicrobial peptides possess cationic and amphipathic properties that allow for interactions with the membrane of living cells. Bacteriocins from lactic acid bacteria, in particular, are currently being studied for their potential use as food preservatives and for applications in health care. However, bacteriocin exploitation is often limited owing to low production yields. Gene cloning and heterologous protein or peptide production is one way to possibly achieve overexpression of bacteriocins to support biochemical studies. In this work, production of recombinant active pediocin PA-1 (PedA) was accomplished in Escherichia coli using a thioredoxin (trx) gene fusion (trx-pedA) expression approach. Trx-PedA itself did not show any biological activity, but upon cleavage by an enterokinase, biologically active pediocin PA-1 was obtained. Recombinant pediocin PA-1 characteristics (molecular mass, biological activity, physicochemical properties) were very similar to those of native pediocin PA-1. In addition, a 4- to 5-fold increase in production yield was obtained, by comparison with the PA-1 produced naturally by Pediococcus acidilactici PAC 1.0. The new production method, although not optimized, offers great potential for supporting further investigations on pediocin PA-1 and as a first-generation process for the production of pediocin PA-1 for high-value applications.     

Physico-chemical approach to adhesion of Alicyclobacillus cells and spores to model solid materials

Extremophiles - Acidothermophilic bacteria of the genus Alicyclobacillus are frequent contaminants of fruit-based products. This study is the first attempt to characterize the physico-chemical...