Metabolic modelling of polyhydroxyalkanoate copolymers production by mixed microbial cultures

Background: This paper presents a metabolic model describing the production of polyhydroxyalkanoate (PHA) copolymers in mixed microbial cultures, using mixtures of acetic and propionic acid as carbon source material. Material and energetic balances were established on the basis of previously elucidated metabolic pathways. Equations were derived for the theoretical yields for cell growth and PHA production on mixtures of acetic and propionic acid as functions of the oxidative phosphorylation efficiency, P/O ratio. The oxidative phosphorylation efficiency was estimated from rate measurements, which in turn allowed the estimation of the theoretical yield coefficients. Results: The model was validated with experimental data collected in a sequencing batch reactor (SBR) operated under varying feeding conditions: feeding of acetic and propionic acid separately (control experiments), and the feeding of acetic and propionic acid simultaneously. Two different feast and famine culture enrichment strategies were studied: (i) either with acetate or (ii) with propionate as carbon source material. Metabolic flux analysis (MFA) was performed for the different feeding conditions and culture enrichment strategies. Flux balance analysis (FBA) was used to calculate optimal feeding scenarios for high quality PHA polymers production, where it was found that a suitable polymer would be obtained when acetate is fed in excess and the feeding rate of propionate is limited to ∼0.17 C-mol/ (C-mol.h). The results were compared with published pure culture metabolic studies. Conclusion: Acetate was more conducive toward the enrichment of a microbial culture with higher PHA storage fluxes and yields as compared to propionate. The P/O ratio was not only influenced by the selected microbial culture, but also by the carbon substrate fed to each culture, where higher P/O ratio values were consistently observed for acetate than propionate. MFA studies suggest that when mixtures of acetate and propionate are fed to the cultures, the catabolic activity is primarily guaranteed through acetate uptake, and the characteristic P/O ratio of acetate prevails over that of propionate. This study suggests that the PHA production process by mixed microbial cultures has the potential to be comparable or even more favourable than pure cultures.publishersversionpublishe

Considering the Case for Biodiversity Cycles: Reexamining the Evidence for Periodicity in the Fossil Record

Medvedev and Melott (2007) have suggested that periodicity in fossil biodiversity may be induced by cosmic rays which vary as the Solar System oscillates normal to the galactic disk. We re-examine the evidence for a 62 million year (Myr) periodicity in biodiversity throughout the Phanerozoic history of animal life reported by Rohde & Mueller (2005), as well as related questions of periodicity in origination and extinction. We find that the signal is robust against variations in methods of analysis, and is based on fluctuations in the Paleozoic and a substantial part of the Mesozoic. Examination of origination and extinction is somewhat ambiguous, with results depending upon procedure. Origination and extinction intensity as defined by RM may be affected by an artifact at 27 Myr in the duration of stratigraphic intervals. Nevertheless, when a procedure free of this artifact is implemented, the 27 Myr periodicity appears in origination, suggesting that the artifact may ultimately be based on a signal in the data. A 62 Myr feature appears in extinction, when this same procedure is used. We conclude that evidence for a periodicity at 62 Myr is robust, and evidence for periodicity at approximately 27 Myr is also present, albeit more ambiguous.Comment: Minor modifications to reflect final published versio

Evaluation of the current knowledge limitations in breast cancer research: a gap analysis

BACKGROUND A gap analysis was conducted to determine which areas of breast cancer research, if targeted by researchers and funding bodies, could produce the greatest impact on patients. METHODS Fifty-six Breast Cancer Campaign grant holders and prominent UK breast cancer researchers participated in a gap analysis of current breast cancer research. Before, during and following the meeting, groups in seven key research areas participated in cycles of presentation, literature review and discussion. Summary papers were prepared by each group and collated into this position paper highlighting the research gaps, with recommendations for action. RESULTS Gaps were identified in all seven themes. General barriers to progress were lack of financial and practical resources, and poor collaboration between disciplines. Critical gaps in each theme included: (1) genetics (knowledge of genetic changes, their effects and interactions); (2) initiation of breast cancer (how developmental signalling pathways cause ductal elongation and branching at the cellular level and influence stem cell dynamics, and how their disruption initiates tumour formation); (3) progression of breast cancer (deciphering the intracellular and extracellular regulators of early progression, tumour growth, angiogenesis and metastasis); (4) therapies and targets (understanding who develops advanced disease); (5) disease markers (incorporating intelligent trial design into all studies to ensure new treatments are tested in patient groups stratified using biomarkers); (6) prevention (strategies to prevent oestrogen-receptor negative tumours and the long-term effects of chemoprevention for oestrogen-receptor positive tumours); (7) psychosocial aspects of cancer (the use of appropriate psychosocial interventions, and the personal impact of all stages of the disease among patients from a range of ethnic and demographic backgrounds). CONCLUSION Through recommendations to address these gaps with future research, the long-term benefits to patients will include: better estimation of risk in families with breast cancer and strategies to reduce risk; better prediction of drug response and patient prognosis; improved tailoring of treatments to patient subgroups and development of new therapeutic approaches; earlier initiation of treatment; more effective use of resources for screening populations; and an enhanced experience for people with or at risk of breast cancer and their families. The challenge to funding bodies and researchers in all disciplines is to focus on these gaps and to drive advances in knowledge into improvements in patient care

Reliability of Rapid Diagnostic Tests in Diagnosing Pregnancy-Associated Malaria in North-Eastern Tanzania.

Accurate diagnosis and prompt treatment of pregnancy-associated malaria (PAM) are key aspects in averting adverse pregnancy outcomes. Microscopy is the gold standard in malaria diagnosis, but it has limited detection and availability. When used appropriately, rapid diagnostic tests (RDTs) could be an ideal diagnostic complement to microscopy, due to their ease of use and adequate sensitivity in detecting even sub-microscopic infections. Polymerase chain reaction (PCR) is even more sensitive, but it is mainly used for research purposes. The accuracy and reliability of RDTs in diagnosing PAM was evaluated using microscopy and PCR. A cohort of pregnant women in north-eastern Tanzania was followed throughout pregnancy for detection of plasmodial infection using venous and placental blood samples evaluated by histidine rich protein 2 (HRP-2) and parasite lactate dehydrogenase (pLDH) based RDTs (Parascreen™) or HRP-2 only (Paracheck Pf® and ParaHIT®f), microscopy and nested Plasmodium species diagnostic PCR. From a cohort of 924 pregnant women who completed the follow up, complete RDT and microscopy data was available for 5,555 blood samples and of these 442 samples were analysed by PCR. Of the 5,555 blood samples, 49 ((proportion and 95% confidence interval) 0.9% [0.7 -1.1]) samples were positive by microscopy and 91 (1.6% [1.3-2.0]) by RDT. Forty-six (50.5% [40.5 - 60.6]) and 45 (49.5% [39.4 - 59.5]) of the RDT positive samples were positive and negative by microscopy, respectively, whereas nineteen (42.2% [29.0 - 56.7]) of the microscopy negative, but RDT positive, samples were positive by PCR. Three (0.05% [0.02 - 0.2]) samples were positive by microscopy but negative by RDT. 351 of the 5,461 samples negative by both RDT and microscopy were tested by PCR and found negative. There was no statistically significant difference between the performances of the different RDTs. Microscopy underestimated the real burden of malaria during pregnancy and RDTs performed better than microscopy in diagnosing PAM. In areas where intermittent preventive treatment during pregnancy may be abandoned due to low and decreasing malaria risk and instead replaced with active case management, screening with RDT is likely to identify most infections in pregnant women and out-performs microscopy as a diagnostic tool

Alpha-1-antitrypsin phenotypes in adult liver disease patients

Alpha-1-antitrypsin (AAT) is an important serine protease inhibitor in humans. Hereditary alpha-1-antitrypsin deficiency (AATD) affects lungs and liver. Liver disease caused by AATD in paediatric patients has been previously well documented. However, the association of liver disease with alpha-1-antitrypsin gene polymorphisms in adults is less clear. Therefore, we aimed to study AAT polymorphisms in adults with liver disease. We performed a case-control study. AAT polymorphisms were investigated by isoelectric focusing in 61 patients with liver cirrhosis and 9 patients with hepatocellular carcinoma. The control group consisted of 218 healthy blood donors. A significant deviation of observed and expected frequency of AAT phenotypes from Hardy-Weinberg equilibrium (chi-square = 34.77, df 11, P = 0.000) in the patient group was caused by a higher than expected frequency of Pi ZZ homozygotes (f = 0.0143 and f = 0.0005, respectively, P = 0.000). In addition, Pi M homozygotes were more frequent in patients than in controls (63% and 46%, respectively, P = 0.025). Our study results show that Pi ZZ homozygosity in adults could be associated with severe liver disease. Presence of Pi M homozygosity could be associated with liver disease via some mechanism different from Z allele-induced liver damage through accumulation of AAT polymers

C9orf72-ALS human iPSC microglia are pro-inflammatory and toxic to co-cultured motor neurons via MMP9

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive motor neuron loss, with additional pathophysiological involvement of non-neuronal cells such as microglia. The commonest ALS-associated genetic variant is a hexanucleotide repeat expansion (HRE) mutation in C9orf72. Here, we study its consequences for microglial function using human iPSC-derived microglia. By RNA-sequencing, we identify enrichment of pathways associated with immune cell activation and cyto-/chemokines in C9orf72 HRE mutant microglia versus healthy controls, most prominently after LPS priming. Specifically, LPS-primed C9orf72 HRE mutant microglia show consistently increased expression and release of matrix metalloproteinase-9 (MMP9). LPS-primed C9orf72 HRE mutant microglia are toxic to co-cultured healthy motor neurons, which is ameliorated by concomitant application of an MMP9 inhibitor. Finally, we identify release of dipeptidyl peptidase-4 (DPP4) as a marker for MMP9-dependent microglial dysregulation in co-culture. These results demonstrate cellular dysfunction of C9orf72 HRE mutant microglia, and a non-cell-autonomous role in driving C9orf72-ALS pathophysiology in motor neurons through MMP9 signaling

Safety and immunogenicity of an FP9-vectored candidate tuberculosis vaccine (FP85A), alone and with candidate vaccine MVA85A in BCG-vaccinated healthy adults: a phase I clinical trial.

The safety and immunogenicity of a new candidate tuberculosis (TB) vaccine, FP85A was evaluated alone and in heterologous prime-boost regimes with another candidate TB vaccine, MVA85A. This was an open label, non-controlled, non-randomized Phase I clinical trial. Healthy previously BCG-vaccinated adult subjects were enrolled sequentially into three groups and vaccinated with FP85A alone, or both FP85A and MVA85A, with a four week interval between vaccinations. Passive and active data on adverse events were collected. Immunogenicity was evaluated by Enzyme Linked Immunospot (ELISpot), flow cytometry and Enzyme Linked Immunosorbent assay (ELISA). Most adverse events were mild and there were no vaccine-related serious adverse events. FP85A vaccination did not enhance antigen 85A-specific cellular immunity. When MVA85A vaccination was preceded by FP85A vaccination, cellular immune responses were lower compared with when MVA85A vaccination was the first immunisation. MVA85A vaccination, but not FP85A vaccination, induced anti-MVA IgG antibodies. Both MVA85A and FP85A vaccinations induced anti-FP9 IgG antibodies. In conclusion, FP85A vaccination was well tolerated but did not induce antigen-specific cellular immune responses. We hypothesize that FP85A induced anti-FP9 IgG antibodies with cross-reactivity for MVA85A, which may have mediated inhibition of the immune response to subsequent MVA85A. ClinicalTrials.gov identification number: NCT00653770

Recruitment of regulatory T cells is correlated with hypoxia-induced CXCR4 expression, and is associated with poor prognosis in basal-like breast cancers

Introduction: Basal-like breast cancers behave more aggressively despite the presence of a dense lymphoid infiltrate. We hypothesised that immune suppression in this subtype may be due to T regulatory cells (Treg) recruitment driven by hypoxia-induced up-regulation of CXCR4 in Treg.Methods: Immunoperoxidase staining for FOXP3 and CXCL12 was performed on tissue microarrays from 491 breast cancers. The hypoxia-associated marker carbonic anhydrase IX (CA9) and double FOXP3/CXCR4 staining were performed on sections from a subset of these cancers including 10 basal-like and 11 luminal cancers matched for tumour grade.Results: High Treg infiltration correlated with tumour CXCL12 positivity (OR 1.89, 95% CI 1.22 to 2.94, P = 0.004) and basal phenotype (OR 3.14, 95% CI 1.08 to 9.17, P = 0.004) in univariate and multivariate analyses. CXCL12 positivity correlated with improved survival (P = 0.005), whereas high Treg correlated with shorter survival for all breast cancers (P = 0.001), luminal cancers (P < 0.001) and basal-like cancers (P = 0.040) that were confirmed in a multivariate analysis (OR 1.61, 95% CI 1.02 to 2.53, P = 0.042). In patients treated with hormone therapy, high Treg were associated with a shorter survival in a multivariate analysis (OR 1.78, 95% CI 1.01 to 3.15, P = 0.040). There was a tendency for luminal cancers to show CXCL12 expression (102/138, 74%) compared to basal-like cancers (16/27, 59%), which verged on statistical significance (P = 0.050). Up-regulation of CXCR4 in Treg correlated with the basal-like phenotype (P = 0.029) and tumour hypoxia, as indicated by CA9 expression (P = 0.049).Conclusions: Our data show that in the setting of hypoxia and CXCR4 up-regulation in Treg, CXCL12 expression may have the negative consequence of enhancing Treg recruitment and suppressing the anti-tumour immune response. © 2011 Yan et al.; licensee BioMed Central Ltd

The Elite-Plus stem migrates more than the flanged Charnley stem: A clinical, radiographic, and radiostereometric analysis of 114 patients with an average of 7 years follow-up

Background and purpose The Charnley Elite-Plus stem was introduced in 1993 as a presumed improvement of the flanged Charnley stem. We started this study in 1996 to investigate the migratory pattern of the Elite-Plus stem. Patients and methods We followed 114 patients with osteoarthritis and a primary total hip replacement with the Elite-Plus stem. Mean age at the time of operation was 64 (50-76) years. The mean follow-up time was 6.5 (2-7) years. Radiographs were evaluated with respect to cementing technique, migration, and wear measured by radiostereometry (RSA). Results The stem survival was 98% (CI: 96-100) at 7 years and 92% (CI: 86-97) at 10 years. Mean migration of the femoral head was 0.35 mm (SD 0.3) medially, 0.51 mm (SD 0.6) distally, and 1.1 mm (SD 1.8) in the dorsal direction. Mean total point motion was 1.7 mm (SD 1.7). The migration of the stems stabilized after 5 years in the medial and dorsal directions, but continued to subside slightly. Migration along any of the axes was higher if the cementing technique was inferior. Interpretation Patients with a Charnley Elite-Plus stem and defects in the cement mantle or other signs of inferior implantation technique should be carefully monitored