Insights into the prevalence and underlying causes of clonal variation through transcriptomic analysis in Pichia pastoris

Clonal variation, wherein a range of specific productivities of secreted proteins are observed from supposedly identical transformants, is an accepted aspect of working with Pichia pastoris. It means that a significant number of transformants need to be tested to obtain a representative sample, and in commercial protein production, companies regularly screen thousands of transformants to select for the highest secretor. Here, we have undertaken a detailed investigation of this phenomenon by characterising clones transformed with the human serum albumin gene. The titers of nine clones, each containing a single copy of the human serum albumin gene (identified by qPCR), were measured and the clones grouped into three categories, namely, high-, mid- and low-level secretors. Transcriptomic analysis, using microarrays, showed that no regulatory patterns consistently correlated with titer, suggesting that the causes of clonal variation are varied. However, a number of physiological changes appeared to underlie the differences in titer, suggesting there is more than one biochemical signature for a high-secreting strain. An anomalous low-secreting strain displaying high transcript levels that appeared to be nutritionally starved further emphasises the complicated nature of clonal variation

A Landscape and Climate Data Logistic Model of Tsetse Distribution in Kenya

, biologically transmitted by the tsetse fly in Africa, are a major cause of illness resulting in both high morbidity and mortality among humans, cattle, wild ungulates, and other species. However, tsetse fly distributions change rapidly due to environmental changes, and fine-scale distribution maps are few. Due to data scarcity, most presence/absence estimates in Kenya prior to 2000 are a combination of local reports, entomological knowledge, and topographic information. The availability of tsetse fly abundance data are limited, or at least have not been collected into aggregate, publicly available national datasets. Despite this limitation, other avenues exist for estimating tsetse distributions including remotely sensed data, climate information, and statistical tools.Here we present a logistic regression model of tsetse abundance. The goal of this model is to estimate the distribution of tsetse fly in Kenya in the year 2000, and to provide a method by which to anticipate their future distribution. Multiple predictor variables were tested for significance and for predictive power; ultimately, a parsimonious subset of variables was identified and used to construct the regression model with the 1973 tsetse map. These data were validated against year 2000 Food and Agriculture Organization (FAO) estimates. Mapcurves Goodness-Of-Fit scores were used to evaluate the modeled fly distribution against FAO estimates and against 1973 presence/absence data, each driven by appropriate climate data.Logistic regression can be effectively used to produce a model that projects fly abundance under elevated greenhouse gas scenarios. This model identifies potential areas for tsetse abandonment and expansion

Methanobactin and the Link Between Copper and Bacterial Methane Oxidation

Methanobactins (mbs) are low-molecular-mass (<1,200 Da) copper-binding peptides, or chalkophores, produced by many methane-oxidizing bacteria (methanotrophs). These molecules exhibit similarities to certain iron-binding siderophores but are expressed and secreted in response to copper limitation. Structurally, mbs are characterized by a pair of heterocyclic rings with associated thioamide groups that form the copper coordination site. One of the rings is always an oxazolone and the second ring an oxazolone, an imidazolone, or a pyrazinedione moiety. The mb molecule originates from a peptide precursor that undergoes a series of posttranslational modifications, including (i) ring formation, (ii) cleavage of a leader peptide sequence, and (iii) in some cases, addition of a sulfate group. Functionally, mbs represent the extracellular component of a copper acquisition system. Consistent with this role in copper acquisition, mbs have a high affinity for copper ions. Following binding, mbs rapidly reduce Cu2+ to Cu1+. In addition to binding copper, mbs will bind most transition metals and near-transition metals and protect the host methanotroph as well as other bacteria from toxic metals. Several other physiological functions have been assigned to mbs, based primarily on their redox and metal-binding properties. In this review, we examine the current state of knowledge of this novel type of metal-binding peptide. We also explore its potential applications, how mbs may alter the bioavailability of multiple metals, and the many roles mbs may play in the physiology of methanotrophs

A longitudinal survey of African animal trypanosomiasis in domestic cattle on the Jos Plateau, Nigeria:prevalence, distribution and risk factors

BACKGROUND: Trypanosomiasis is a widespread disease of livestock in Nigeria and a major constraint to the rural economy. The Jos Plateau, Nigeria was free from tsetse flies and the trypanosomes they transmit due to its high altitude and the absence of animal trypanosomiasis attracted large numbers of cattle-keeping pastoralists to inhabit the plateau. The Jos Plateau now plays a significant role in the national cattle industry, accommodating approximately 7% of the national herd and supporting 300,000 pastoralists and over one million cattle. However, during the past two decades tsetse flies have invaded the Jos Plateau and animal trypanosomiasis has become a significant problem for livestock keepers. METHODS: In 2008 a longitudinal two-stage cluster survey on the Jos Plateau. Cattle were sampled in the dry, early wet and late wet seasons. Parasite identification was undertaken using species-specific polymerase chain reactions to determine the prevalence and distribution bovine trypanosomiasis. Logistic regression was performed to determine risk factors for disease. RESULTS: The prevalence of bovine trypanosomiasis (Trypanosoma brucei brucei, Trypanosoma congolense savannah, Trypanosoma vivax) across the Jos Plateau was found to be high at 46.8% (39.0 – 54.5%) and significant, seasonal variation was observed between the dry season and the end of the wet season. T. b. brucei was observed at a prevalence of 3.2% (1% – 5.5%); T. congolense at 27.7% (21.8% - 33.6%) and T. vivax at 26.7% (18.2% - 35.3%). High individual variation was observed in trypanosomiasis prevalence between individual villages on the Plateau, ranging from 8.8% to 95.6%. Altitude was found to be a significant risk factor for trypanosomiasis whilst migration also influenced risk for animal trypanosomiasis. CONCLUSIONS: Trypanosomiasis is now endemic on the Jos Plateau showing high prevalence in cattle and is influenced by seasonality, altitude and migration practices. Attempts to successfully control animal trypanosomiasis on the Plateau will need to take into account the large variability in trypanosomiasis infection rates between villages, the influence of land use, and husbandry and management practices of the pastoralists, all of which affect the epidemiology of the disease

Sleeping sickness and its relationship with development and biodiversity conservation in the Luangwa valley, Zambia

The Luangwa Valley has a long historical association with Human African trypanosomiasis (HAT) and is a recognised geographical focus of this disease. It is also internationally acclaimed for its high biodiversity and contains many valuable habitats. Local inhabitants of the valley have developed sustainable land use systems in co-existence with wildlife over centuries, based on non-livestock keeping practices largely due to the threat from African Animal Trypanosomiasis. Historical epidemics of human sleeping sickness have influenced how and where communities have settled and have had a profound impact on development in the Valley. Historical attempts to control trypanosomiasis have also had a negative impact on conservation of biodiversity. Centralised control over wildlife utilisation has marginalised local communities from managing the wildlife resource. To some extent this has been reversed by the implementation of community based natural resource management programmes in the latter half of the 20th century and the Luangwa Valley provides some of the earliest examples of such programmes. More recently, there has been significant uncontrolled migration of people into the mid-Luangwa Valley driven by pressure on resources in the eastern plateau region, encouragement from local chiefs and economic development in the tourist centre of Mfuwe. This has brought changing land-use patterns, most notably agricultural development through livestock keeping and cotton production. These changes threaten to alter the endemically stable patterns of HAT transmission and could have significant impacts on ecosystem health and ecosystem services. In this paper we review the history of HAT in the context of conservation and development and consider the impacts current changes may have on this complex social-ecological system. We conclude that improved understanding is required to identify specific circumstances where win-win trade-offs can be achieved between the conservation of biodiversity and the reduction of disease in the human population.Ecosystem Services for Poverty Alleviation (ESPA

Civil conflict and sleeping sickness in Africa in general and Uganda in particular

Conflict and war have long been recognized as determinants of infectious disease risk. Re-emergence of epidemic sleeping sickness in sub-Saharan Africa since the 1970s has coincided with extensive civil conflict in affected regions. Sleeping sickness incidence has placed increasing pressure on the health resources of countries already burdened by malaria, HIV/AIDS, and tuberculosis. In areas of Sudan, the Democratic Republic of the Congo, and Angola, sleeping sickness occurs in epidemic proportions, and is the first or second greatest cause of mortality in some areas, ahead of HIV/AIDS. In Uganda, there is evidence of increasing spread and establishment of new foci in central districts. Conflict is an important determinant of sleeping sickness outbreaks, and has contributed to disease resurgence. This paper presents a review and characterization of the processes by which conflict has contributed to the occurrence of sleeping sickness in Africa. Conflict contributes to disease risk by affecting the transmission potential of sleeping sickness via economic impacts, degradation of health systems and services, internal displacement of populations, regional insecurity, and reduced access for humanitarian support. Particular focus is given to the case of sleeping sickness in south-eastern Uganda, where incidence increase is expected to continue. Disease intervention is constrained in regions with high insecurity; in these areas, political stabilization, localized deployment of health resources, increased administrative integration and national capacity are required to mitigate incidence. Conflict-related variables should be explicitly integrated into risk mapping and prioritization of targeted sleeping sickness research and mitigation initiatives

Low Concentrations of Methamphetamine Can Protect Dopaminergic Cells against a Larger Oxidative Stress Injury: Mechanistic Study

Mild stress can protect against a larger insult, a phenomenon termed preconditioning or tolerance. To determine if a low intensity stressor could also protect cells against intense oxidative stress in a model of dopamine deficiency associated with Parkinson disease, we used methamphetamine to provide a mild, preconditioning stress, 6-hydroxydopamine (6-OHDA) as a source of potentially toxic oxidative stress, and MN9D cells as a model of dopamine neurons. We observed that prior exposure to subtoxic concentrations of methamphetamine protected these cells against 6-OHDA toxicity, whereas higher concentrations of methamphetamine exacerbated it. The protection by methamphetamine was accompanied by decreased uptake of both [3H] dopamine and 6-OHDA into the cells, which may have accounted for some of the apparent protection. However, a number of other effects of methamphetamine exposure suggest that the drug also affected basic cellular survival mechanisms. First, although methamphetamine preconditioning decreased basal pERK1/2 and pAkt levels, it enhanced the 6-OHDA-induced increase in these phosphokinases. Second, the apparent increase in pERK1/2 activity was accompanied by increased pMEK1/2 levels and decreased activity of protein phosphatase 2. Third, methamphetamine upregulated the pro-survival protein Bcl-2. Our results suggest that exposure to low concentrations of methamphetamine cause a number of changes in dopamine cells, some of which result in a decrease in their vulnerability to subsequent oxidative stress. These observations may provide insights into the development of new therapies for prevention or treatment of PD

A Multi-Host Agent-Based Model for a Zoonotic, Vector-Borne Disease. A Case Study on Trypanosomiasis in Eastern Province, Zambia

Background: This paper presents a new agent-based model (ABM) for investigating T. b. rhodesiense human African trypanosomiasis (rHAT) disease dynamics, produced to aid a greater understanding of disease transmission, and essential for development of appropriate mitigation strategies. Methods: The ABM was developed to model rHAT incidence at a fine spatial scale along a 75 km transect in the Luangwa Valley, Zambia. The method offers a complementary approach to traditional compartmentalised modelling techniques, permitting incorporation of fine scale demographic data such as ethnicity, age and gender into the simulation. Results: Through identification of possible spatial, demographic and behavioural characteristics which may have differing implications for rHAT risk in the region, the ABM produced output that could not be readily generated by other techniques. On average there were 1.99 (S.E. 0.245) human infections and 1.83 (S.E. 0.183) cattle infections per 6 month period. The model output identified that the approximate incidence rate (per 1000 person-years) was lower amongst cattle owning households (0.079, S.E. 0.017), than those without cattle (0.134, S.E. 0.017). Immigrant tribes (e.g. Bemba I.R. = 0.353, S.E.0.155) and school-age children (e.g. 5–10 year old I.R. = 0.239, S.E. 0.041) were the most at-risk for acquiring infection. These findings have the potential to aid the targeting of future mitigation strategies. Conclusion: ABMs provide an alternative way of thinking about HAT and NTDs more generally, offering a solution to the investigation of local-scale questions, and which generate results that can be easily disseminated to those affected. The ABM can be used as a tool for scenario testing at an appropriate spatial scale to allow the design of logistically feasible mitigation strategies suggested by model output. This is of particular importance where resources are limited and management strategies are often pushed to the local scale. © 2016 Alderton et al

Genetic Ancestry-Smoking Interactions and Lung Function in African Americans: A Cohort Study

Background: Smoking tobacco reduces lung function. African Americans have both lower lung function and decreased metabolism of tobacco smoke compared to European Americans. African ancestry is also associated with lower pulmonary function in African Americans. We aimed to determine whether African ancestry modifies the association between smoking and lung function and its rate of decline in African Americans. Methodology/Principal Findings: We evaluated a prospective ongoing cohort of 1,281 African Americans participating in the Health, Aging, and Body Composition (Health ABC) Study initiated in 1997. We also examined an ongoing prospective cohort initiated in 1985 of 1,223 African Americans in the Coronary Artery Disease in Young Adults (CARDIA) Study. Pulmonary function and tobacco smoking exposure were measured at baseline and repeatedly over the follow-up period. Individual genetic ancestry proportions were estimated using ancestry informative markers selected to distinguish European and West African ancestry. African Americans with a high proportion of African ancestry had lower baseline forced expiratory volume in one second (FEV1) per pack-year of smoking (-5.7 ml FEV1/ smoking pack-year) compared with smokers with lower African ancestry (-4.6 ml in FEV1/ smoking pack-year) (interaction P value = 0.17). Longitudinal analyses revealed a suggestive interaction between smoking, and African ancestry on the rate of FEV1 decline in Health ABC and independently replicated in CARDIA. Conclusions/Significance: African American individuals with a high proportion of African ancestry are at greater risk for losing lung function while smoking. © 2012 Aldrich et al

Infections with Immunogenic Trypanosomes Reduce Tsetse Reproductive Fitness: Potential Impact of Different Parasite Strains on Vector Population Structure

The parasite Trypanosoma brucei rhodesiense and its insect vector Glossina morsitans morsitans were used to evaluate the effect of parasite clearance (resistance) as well as the cost of midgut infections on tsetse host fitness. Tsetse flies are viviparous and have a low reproductive capacity, giving birth to only 6–8 progeny during their lifetime. Thus, small perturbations to their reproductive fitness can have a major impact on population densities. We measured the fecundity (number of larval progeny deposited) and mortality in parasite-resistant tsetse females and untreated controls and found no differences. There was, however, a typanosome-specific impact on midgut infections. Infections with an immunogenic parasite line that resulted in prolonged activation of the tsetse immune system delayed intrauterine larval development resulting in the production of fewer progeny over the fly's lifetime. In contrast, parasitism with a second line that failed to activate the immune system did not impose a fecundity cost. Coinfections favored the establishment of the immunogenic parasites in the midgut. We show that a decrease in the synthesis of Glossina Milk gland protein (GmmMgp), a major female accessory gland protein associated with larvagenesis, likely contributed to the reproductive lag observed in infected flies. Mathematical analysis of our empirical results indicated that infection with the immunogenic trypanosomes reduced tsetse fecundity by 30% relative to infections with the non-immunogenic strain. We estimate that a moderate infection prevalence of about 26% with immunogenic parasites has the potential to reduce tsetse populations. Potential repercussions for vector population growth, parasite–host coevolution, and disease prevalence are discussed