Retargeting of adenovirus vectors through genetic fusion of a single-chain or single-domain antibody to capsid protein IX.

Adenovirus (Ad) vectors are the most commonly used system for gene therapy applications, due in part to their ability to infect a wide array of cell types and tissues. However, many therapies would benefit from the ability to target the Ad vector only to specific cells, such as tumor cells for cancer gene therapy. In this study, we investigated the utility of capsid protein IX (pIX) as a platform for the presentation of single-chain variable-fragment antibodies (scFv) and single-domain antibodies (sdAb) for virus retargeting. We show that scFv can be displayed on the capsid through genetic fusion to native pIX but that these molecules fail to retarget the virus, due to improper folding of the scFv. Redirecting expression of the fusion protein to the endoplasmic reticulum (ER) results in correct folding of the scFv and allows it to recognize its epitope; however, ER-targeted pIX-scFv was incorporated into the Ad capsid at a very low level which was not sufficient to retarget virus infection. In contrast, a pIX-sdAb construct was efficiently incorporated into the Ad capsid and enhanced virus infection of cells expressing the targeted receptor. Taken together, our data indicate that pIX is an effective platform for presentation of large targeting polypeptides on the surface of the virus capsid, but the nature of the ligand can significantly affect its association with virions.

Microbial contamination of two urban sandstone aquifers in the UK.

Development of urban groundwater has historically been constrained by concerns about its quality. Rising urban water tables and overabstraction from rural aquifers in the UK have led to a renewed interest in urban groundwater, particularly the possibility of finding water of acceptable quality at depth. This study assessed the microbial quality of groundwater collected from depth-specific intervals over a 15-month period within the Permo-Triassic Sherwood Sandstone aquifers underlying the cities of Nottingham and Birmingham. Sewage-derived bacteria (thermotolerant coliforms, faecal streptococci and sulphite-reducing clostridia) and viruses (enteroviruses, Norwalk-like viruses, coliphage) were regularly detected to depths of 60 m in the unconfined sandstone and to a depth of 91 m in the confined sandstone. Microbial concentrations varied temporally and spatially but increased frequency of contamination with depth coincided with geological heterogeneities such as fissures and mudstone bands. Significantly, detection of Norwalk-like viruses and Coxsackievirus B4 in groundwater corresponded with seasonal variations in virus discharge to the sewer system. The observation of low levels of sewage-derived microbial contaminants at depth in the Triassic Sandstone aquifer is explained by the movement of infinitesimal proportions of bulk (macroscopic) groundwater flow along preferential pathways (e.g., fissures, bedding planes). The existence of very high microbial populations at source (raw sewage) and their extremely low detection limits at the receptor (multilevel piezometer) enable these statistically extreme (microscopic) flows to be traced. Rapid penetration of microbial contaminants into sandstone aquifers, not previously reported, highlights the vulnerability of sandstone aquifers to microbial contamination.

Dietary proteins as environmental modifiers of type 1 diabetes mellitus.

Type 1 diabetes is an autoimmune disease in which the patient's immune system destroys the insulin-secreting beta-cells in the pancreatic islets of Langerhans. A majority of cases is thought to occur as a result of gene-environment interactions. The identity of the environmental factors remains unknown mainly because of the difficulty in linking past exposures with later disease development. Overall, the data suggest a model in which individuals develop diabetes by several different pathways, each influenced by numerous genetic and environmental variables. The most investigated environmental factors are diet and viruses. In this review, we examine the evidence that the source of dietary proteins can modify diabetes outcome, describe new approaches to identify candidate diabetes-related dietary agents, examine possible links with gut dysfunction, discuss some of the limitations, and propose a multifactorial model for dietary modification of diabetes. The key to diabetes pathogenesis, its prevention, and the ultimate success of beta-cell replacement therapies lies in understanding how the environment controls disease expression. Dietary proteins could be one of these keys.