Investigation of a pathogenic mtDNA microdeletion reveals a translation-dependent deadenylation decay pathway in human mitochondria

Human mtDNA is transcribed from both strands, producing polycistronic RNA species that are immediately processed. Discrete RNA units are matured by the addition of nucleotides at their 3' termini: -CCA trinucleotide is added to mt-tRNAs, whilst mt-rRNAs and mt-mRNAs are oligo- or polyadenylated, respectively. The cis-acting elements, enzymes and indeed the mechanisms involved in these processes are still largely uncharacterized. Further, the function of polyadenylation in promoting stability, translation or decay of human mt-mRNA is unclear. A microdeletion has been identified in a patient presenting with mtDNA disease. Loss of these two residues removes the termination codon for MTATP6 and sets MTCO3 immediately in frame. Accurate processing at this site still occurs, but there is a markedly decreased steady-state level of RNA14, the ATPase 8- and 6-encoding bi-cistronic mRNA unit, establishing that an mtDNA mutation can cause dysregulation of mRNA stability. Analysis of the polyadenylation profile of the processed RNA14 at steady state revealed substantial abnormalities. The majority of mutated RNA14 terminated with short poly (A) extensions and a second, partially truncated population, was also present. Initial maturation of mutated RNA14 was unaffected, but deadenylation occurred rapidly. Inhibition of mitochondrial protein synthesis showed that the deadenylation was dependent on translation. Finally, deadenylation was shown to enhance mRNA decay, explaining the decrease in steady-state RNA14. An hypothesis is presented to describe how an mtDNA mutation that results in the loss of a termination codon causes enhanced mt-mRNA decay by translation-dependent deadenylation.     

Evaluation of use of recombinant Em18 and affinity-purified Em18 for serological differentiation of alveolar echinococcosis from cystic echinococcosis and other parasitic infections

To further evaluate recombinant Em18 antigen (rEm18) for immunodiagnosis of human alveolar echinococcosis, 208 serum samples were examined by enzyme-linked immunosorbent assay (ELISA). To comparatively assess the results of rEm18-ELISA, ELISA and immunoblot analysis with two affinity-purified native antigens were also performed with 45 selected serum samples. The results indicate that rEm18 is highly useful for serodiagnosis.     

Targeting large molecules to mitochondria

Mitochondrial function is central to a range of cell processes and mitochondrial dysfunction contributes to a number of human diseases. Consequently there is growing interest in delivering large molecules such as nucleic acids, proteins, enzyme mimetics, drugs and probes to mitochondria within cells. The reasons for doing this are to understand how mitochondria function in the cell and to develop therapies for diseases involving mitochondrial damage. Here we review the methods that have been used to target large molecules to mitochondria and discuss some approaches under development.     

Cestode infections in animals: immunological diagnosis and vaccination

Cestode infections in animals are important because several species are zoonotic, causing cysticercosis and hydatidosis in man, and because of the economic losses incurred due to infections in livestock. Information on immunological diagnosis of and vaccination against cestode infection is restricted almost exclusively to the taeniid cestodes in which two groups of mammalian hosts are concerned: the intermediate host infected with the larval parasite and the definitive host infected with the adult tapeworm parasite. Research towards developing serological tests for the diagnosis of larval cestode infection in animals has been largely unsuccessful. Substantial problems remain, due to the frequent existence of multiple infections with different taeniid species and antigenic crossreactivity between these related parasites, and the low level of specific antibody response to infection. Problems with poor specificity and sensitivity of traditional serological tests for cysticercosis and hydatidosis have prevented the development of any practical test for ante-mortem diagnosis of infection. A recent new approach to the diagnosis of Taenia saginata infection by detecting circulating parasite antigen offers some prospect for the development of a practical diagnostic test for cysticercosis in cattle. The effectiveness of the arecoline purge for detection of Echinococcus granulosus in dogs has been reduced by the widespread availability of praziquantel. A serological method for diagnosis of E. granulosus in dogs has been developed which offers equivalent or superior diagnostic sensitivity compared with arecoline purge. This test should provide a valuable tool in hydatid control campaigns for the diagnosis of existing or recent past infections in dogs. Substantial progress has been made towards developing a practical vaccine for the prevention of T. ovis infection in sheep. An antigen derived from the parasite egg has been identified and produced in Escherichia coli using recombinant DNA techniques. The vaccine, which protects sheep against challenge infection with T. ovis, is the first highly effective defined antigen vaccine against any parasite infection of man or animals. Commercial development of this vaccine is in progress. The success achieved with the T. ovis vaccine augurs well for the rapid development of other recombinant vaccines against cysticercosis caused by other taeniid species and against hydatidosis in animals.     

Studies on stage-specific immunity against Taenia Taeniaeformis metacestodes in mice

The possible existence of stage-specific immune responses to Taenia taeniaeformis infection was investigated in C3H/He mice vaccinated with antigens prepared from either the oncosphere or metacestode stages. Mice were immunized twice, 2 weeks apart, with antigen in Freund's complete adjuvant. Two weeks after the second immunization they were challenged with 250 T. taeniaeformis eggs and killed day 0, 5, 10, 15, 20, 25, 30, 45 and 60 after infection. Gross examination of the livers revealed marked differences between oncosphere (TtO) and metacestode (TtM) vaccinated mice. Very few metacestodes were found in the first group but most of those that evaded the initial host attack developed like the cysts found in the control group. In contrast, many degenerating metacestodes were found in the TtM vaccinated group. In a subsequent experiment groups of mice were vaccinated with varying doses of either TtO or TtM to determine whether the qualitative differences observed above were due to antigen dose effects. However, varying antigen doses gave the same results. These data show that vaccination with oncospheres generates an immune response capable of killing invading larvae soon after infection whereas vaccination with TtM results in larvae being killed at a later stage, suggesting that there are stage-specific, host-protective antigens.     

Intercity transferability of land use regression models for estimating ambient concentrations of nitrogen dioxide

Land use regression (LUR) is a method for predicting the spatial distribution of traffic-related air pollution. To facilitate risk and exposure assessment, and the design of future monitoring networks and sampling campaigns, we sought to determine the extent to which LUR can be used to predict spatial patterns in air pollution in the absence of dedicated measurements. We evaluate the transferability of one LUR model to two other geographically comparable areas with similar climates and pollution types. The source model, developed in 2003 to estimate ambient nitrogen dioxide (NO(2)) concentrations in Vancouver (BC, Canada) was applied to Victoria (BC, Canada) and Seattle (WA, USA). Model estimates were compared with measurements made with Ogawa passive samplers in both cities. As part of this study, 42 locations were sampled in Victoria for a 2-week period in June 2006. Data obtained for Seattle were collected for a different project at 26 locations in March 2005. We used simple linear regression to evaluate the fit of the source model under three scenarios: (1) using the same variables and coefficients as the source model; (2) using the same variables as the source model, but calculating new coefficients for local calibration; and (3) developing site-specific equations with new variables and coefficients. In Scenario 1, we found that the source model had a better fit in Victoria (R(2)=0.51) than in Seattle (R(2)=0.33). Scenario 2 produced improved R(2)-values in both cities (Victoria=0.58, Seattle=0.65), with further improvement achieved under Scenario 3 (Victoria=0.61, Seattle=0.72). Although it is possible to transfer LUR models between geographically similar cities, success may depend on the between-city consistency of the input data. Modest field sampling campaigns for location-specific model calibration can help to produce transfer models that are equally as predictive as their sources.     

Vaccines to combat the neglected tropical diseases

The neglected tropical diseases (NTDs) represent a group of parasitic and related infectious diseases such as amebiasis, Chagas disease, cysticercosis, echinococcosis, hookworm, leishmaniasis, and schistosomiasis. Together, these conditions are considered the most common infections in low- and middle-income countries, where they produce a level of global disability and human suffering equivalent to better known conditions such as human immunodeficiency virus/acquired immunodeficiency syndrome and malaria. Despite their global public health importance, progress on developing vaccines for NTD pathogens has lagged because of some key technical hurdles and the fact that these infections occur almost exclusively in the world's poorest people living below the World Bank poverty line. In the absence of financial incentives for new products, the multinational pharmaceutical companies have not embarked on substantive research and development programs for the neglected tropical disease vaccines. Here, we review the current status of scientific and technical progress in the development of new neglected tropical disease vaccines, highlighting the successes that have been achieved (cysticercosis and echinococcosis) and identifying the challenges and opportunities for development of new vaccines for NTDs. Also highlighted are the contributions being made by non-profit product development partnerships that are working to overcome some of the economic challenges in vaccine manufacture, clinical testing, and global access.     

Prevention and control of cystic echinococcosis

Human cystic echinococcosis (hydatid disease) continues to be a substantial cause of morbidity and mortality in many parts of the world. Elimination is difficult to obtain and it is estimated that, using current control options, achieving such a goal will take around 20 years of sustained efforts. Since the introduction of current (and past) hydatid control campaigns, there have been clear technological improvements made in the diagnosis and treatment of human and animal cystic echinococcosis, the diagnosis of canine echinococcosis, and the genetic characterisation of strains and vaccination against Echinococcus granulosus in animals. Incorporation of these new measures could increase the efficiency of hydatid control programmes, potentially reducing the time required to achieve effective prevention of disease transmission to as little as 5-10 years.