In tuberculosis, infecting mycobacteria are phagocytosed by macrophages, which then migrate into deeper tissue and recruit additional cells to form the granulomas that eventually contain infection. Mycobacteria are exquisitely adapted macrophage pathogens, and observations in the mouse model of tuberculosis have suggested that mycobacterial growth is not inhibited in macrophages until adaptive immunity is induced. Using the optically transparent and genetically tractable zebrafish embryo-Mycobacterium marinum model of tuberculosis, we have directly examined early infection in the presence and absence of macrophages. The absence of macrophages led rapidly to higher bacterial burdens, suggesting that macrophages control infection early and are not an optimal growth niche. However, we show that macrophages play a critical role in tissue dissemination of mycobacteria. We propose that residence within macrophages represents an evolutionary trade-off for pathogenic mycobacteria that slows their early growth but provides a mechanism for tissue dissemination.

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Vlad tepes (vlt m651 ) is one of only five ''bloodless'' zebrafish mutants isolated through large-scale chemical mutagenesis screening. It is characterized by a severe reduction in blood cell progenitors and few or no blood cells at the onset of circulation. We now report characterization of the mutant phenotype and the identification of the gene mutated in vlt m651 . Embryos homozygous for the vlt m651 mutation had normal expression of hematopoietic stem cell markers through 24 h postfertilization, as well as normal expression of myeloid and lymphoid markers. Analysis of erythroid development revealed variable expression of erythroid markers. Through positional and candidate gene cloning approaches we identified a nonsense mutation in the gata1 gene, 1015C 3 T (Arg-339 3 Stop), in vlt m651 . The nonsense mutation was located C-terminal to the two zinc fingers and resulted in a truncated protein that was unable to bind DNA or mediate GATA-specific transactivation. A BAC clone containing the zebrafish gata1 gene was able to rescue the bloodless phenotype in vlt m651 . These results show that the vlt m651 mutation is a previously uncharacterized gata1 allele in the zebrafish. The vlt m651 mutation sheds new light on Gata1 structure and function in vivo, demonstrates that Gata1 plays an essential role in zebrafish hematopoiesis with significant conservation of function between mammals and zebrafish, and offers a powerful tool for future studies of the hematopoietic pathway.

Patients with arteriovenous malformations of the brain, who are subject to disabling or fatal recurrent hemorrhage, seizures, severe headache, and progressive neurologic deficits, may be considered unsuitable for conventional therapies (craniotomy with excision or embolization), usually because of the location, size, or operative risk of the lesion. We have treated such patients with stereotactic Bragg-peak proton-beam therapy and report the follow-up of 74 of the first 75, 2 to 16 years after treatment. Proton-beam therapy is intended to induce subendothelial deposition of collagen and hyaline substance, which narrows the lumens of small vessels and thickens the walls of the malformation during the first 12 to 24 months after the procedure. Two deaths from hemorrhage occurred in the first 12 months after treatment, but no lethal or disabling hemorrhages occurred after this interval. Seizures, headaches, and progressive neurologic deficits were in most cases arrested or improved. Bragg-peak proton-beam therapy appears to be a useful technique for treatment of intracranial arteriovenous malformations, especially those that are unsuitable for treatment by other methods.