Adenosine kinase of Trypanosoma brucei and its role in susceptibility to adenosine antimetabolites

Trypanosoma brucei cannot synthesize purines de novo and relies on purine salvage from its hosts to build nucleic acids. With adenosine being a preferred purine source of bloodstream-form trypanosomes, adenosine kinase (AK; EC 2.7.1.20) is likely to be a key player in purine salvage. Adenosine kinase is also of high pharmacological interest, since for many adenosine antimetabolites, phosphorylation is a prerequisite for activity. Here, we cloned and functionally characterized adenosine kinase from T. brucei (TbAK). TbAK is a tandem gene, expressed in both procyclic- and bloodstream-form trypanosomes, whose product localized to the cytosol of the parasites. The RNA interference-mediated silencing of TbAK suggested that the gene is nonessential under standard growth conditions. Inhibition or downregulation of TbAK rendered the trypanosomes resistant to cordycepin (3'-deoxyadenosine), demonstrating a role for TbAK in the activation of adenosine antimetabolites. The expression of TbAK in Saccharomyces cerevisiae complemented a null mutation in the adenosine kinase gene ado1. The concomitant expression of TbAK with the T. brucei adenosine transporter gene TbAT1 allowed S. cerevisiae ado1 ade2 double mutants to grow on adenosine as the sole purine source and, at the same time, sensitized them to adenosine antimetabolites. The coexpression of TbAK and TbAT1 in S. cerevisiae ado1 ade2 double mutants proved to be a convenient tool for testing nucleoside analogues for uptake and activation by T. brucei adenosine salvage enzymes.     

RHINOCEREBRAL MUCORMYCOSIS DIABETES MELLITUS AND ADRENOGENITAL SYNDROME

A case of rhinocerebral mucormycosis in a diabetic is described. The case is unusual as the mucormycosis developed in the absence of ketoacidosis and because the patient had concomitant adrenogenital syndrome.     

Specific antibody responses to vaccination with bivalent CM235/SF2 gp120: detection of homologous and heterologous neutralizing antibody to subtype E (CRF01.AE) HIV type 1

HIV-1 CRF.AE-01 (formerly subtype E) infection is highly prevalent in Southeast Asia. Despite success with public health measures, the development of an effective CRF01.AE vaccine is critical to the control of this epidemic. Sera from the open-label arms of the first clinical trial of a bivalent HIV gp120 SF2/CM235 (subtypes B and CRF.AE-01, respectively) vaccine were evaluated for the presence of gp120-specific binding (BAb) and neutralizing antibody (NAb). Twelve pre- and postvaccination sera pairs were tested for CM235 BAb; anti-gp120 CM235 BAb was found in all postvaccination samples. The 12 pre- and postvaccination (1 month after third vaccination) serum pairs were evaluated in several neutralization formats: heterologous T cell line adapted (TCLA) NP03/H9, homologous CM235/PBMC, CM235/dendritic cell, and CM235M4-C4.6/A3R5. A3R5 is a CCR5+ T cell line, and CM235M4-C4.6 is the homologous CM235 virus adapted to growth in A3R5 cells. All volunteers developed BAb, but meaningful NAb was not demonstrable against primary isolate CM235. Using the TCLA CRF01.AE virus NP03 in H9 cells, 9 of 12 persons had NAb with a geometric mean titer (GMT) of 46. The CM235M4-C4.6 virus in A3R5 cells also detected NAb in 9 of 12 persons, with a GMT of 41. CM235M4-C4.6/A3R5 detected NAb in two persons with negligible NAb to NP03/H9 and vice versa. Whether the NAb detected by the CM235M4-C4.6/A3R5 system is qualitatively different from those in more traditional NP03/H9 assays will require further study.     

Pharyngeal pump and esophageal transit

In deglutition the pharynx appears to act as a pump to inject boluses into the esophagus. A new method for measuring the velocity profile of the leading edge of a radionuclide bolus has been developed and applied to boluses of different viscosity—water and treacle—in nine normal volunteers. The results show that the more viscous bolus (treacle) acquires a slower initial injection velocity (152 mm/sec vs 236 mm/sec) that only propels it over the proximal half of the esophagus. Peristaltic action must drive the bolus over the distal half. With water boluses, however, the higher initial velocity is sufficient to propel a part of the bolus at least to the gastroesophageal junction leaving minimal work to be performed by esophageal peristalsis. This confirms the important role of the pharyngeal pump in deglutition. The pump may be the major mechanism for ingestion of nonviscous liquids (water), peristalsis merely being required to sweep up what remains in the esophagus.