Mammalian adaptation of influenza A(H7N9) virus is limited by a narrow genetic bottleneck

Human infection with avian influenza A(H7N9) virus is associated mainly with the exposure to infected poultry. The factors that allow interspecies transmission but limit human-to-human transmission are unknown. Here we show that A/Anhui/1/2013(H7N9) influenza virus infection of chickens (natural hosts) is asymptomatic and that it generates a high genetic diversity. In contrast, diversity is tightly restricted in infected ferrets, limiting further adaptation to a fully transmissible form. Airborne transmission in ferrets is accompanied by the mutations in PB1, NP and NA genes that reduce viral polymerase and neuraminidase activity. Therefore, while A(H7N9) virus can infect mammals, further adaptation appears to incur a fitness cost. Our results reveal that a tight genetic bottleneck during avian-to-mammalian transmission is a limiting factor in A(H7N9) influenza virus adaptation to mammals. This previously unrecognized biological mechanism limiting species jumps provides a measure of adaptive potential and may serve as a risk assessment tool for pandemic preparedness.published_or_final_versio

Terrestrial behavior in titi monkeys (Callicebus, Cheracebus, and Plecturocebus) : potential correlates, patterns, and differences between genera

For arboreal primates, ground use may increase dispersal opportunities, tolerance to habitat change, access to ground-based resources, and resilience to human disturbances, and so has conservation implications. We collated published and unpublished data from 86 studies across 65 localities to assess titi monkey (Callicebinae) terrestriality. We examined whether the frequency of terrestrial activity correlated with study duration (a proxy for sampling effort), rainfall level (a proxy for food availability seasonality), and forest height (a proxy for vertical niche dimension). Terrestrial activity was recorded frequently for Callicebus and Plecturocebus spp., but rarely for Cheracebus spp. Terrestrial resting, anti-predator behavior, geophagy, and playing frequencies in Callicebus and Plecturocebus spp., but feeding and moving differed. Callicebus spp. often ate or searched for new leaves terrestrially. Plecturocebus spp. descended primarily to ingest terrestrial invertebrates and soil. Study duration correlated positively and rainfall level negatively with terrestrial activity. Though differences in sampling effort and methods limited comparisons and interpretation, overall, titi monkeys commonly engaged in a variety of terrestrial activities. Terrestrial behavior in Callicebus and Plecturocebus capacities may bolster resistance to habitat fragmentation. However, it is uncertain if the low frequency of terrestriality recorded for Cheracebus spp. is a genus-specific trait associated with a more basal phylogenetic position, or because studies of this genus occurred in pristine habitats. Observations of terrestrial behavior increased with increasing sampling effort and decreasing food availability. Overall, we found a high frequency of terrestrial behavior in titi monkeys, unlike that observed in other pitheciids

Ectopic expression of rhombotin-2 causes selective expansion of CD4-CD8- lymphocytes in the thymus and T-cell tumors in transgenic mice

Abstract Although the proto-oncogene rhombotin-2 (RBTN-2) is widely expressed in most tissues, it is not expressed in T cells. We investigated the potential for overexpression of RBTN-2 to cause tumors in T cells and other tissues by constructing transgenic mice that expressed RBTN-2 under control of the metallothionein-1 promoter. Despite overexpression of RBTN-2 in all tissues, transgenic mice developed T-cell tumors only, thus indicating that tumorigenesis caused by RBTN-2 is T-cell-specific. Thymic tumors were found between 37 and 71 weeks and were invariably associated with metastasis to nonlymphoid organs. Thymuses from apparently healthy transgenic mice were also examined. In some mice there was an 10-fold increase in the CD4-CD8- thymocyte subset, yet the total number of thymocytes was the same as that in wild-type mice. Thymic homeostasis was maintained by a compensatory reduction in the CD4+CD8+ subset. The expansion of CD4-CD8- thymocytes was associated with increased expression of RBTN-2 and with increased cell proliferation. No differences were found in the proportion of thymocytes undergoing apoptosis in transgenic mice. Furthermore, RBTN-2-induced expansion of CD4-CD8- cells did not block differentiation of these cells. Thymuses with 30% CD4-CD8- cells were essentially monoclonal, indicating that all thymic immunophenotypes were derived from a single clone. Overall, our data are consistent with the following scenario: (1) RBTN-2 expression in T cells causes selective and polyclonal proliferation of CD4-CD8-thymocytes accompanied by a compensatory decrease in other thymocyte subsets; (2) a clone with growth advantage and differentiation potential is selected and populates the thymus; and (3) this clone eventually breaches homeostasis of the thymus, accompanied or followed by metastasis to other organs.</jats:p

Ectopic expression of rhombotin-2 causes selective expansion of CD4-CD8- lymphocytes in the thymus and T-cell tumors in transgenic mice

Although the proto-oncogene rhombotin-2 (RBTN-2) is widely expressed in most tissues, it is not expressed in T cells. We investigated the potential for overexpression of RBTN-2 to cause tumors in T cells and other tissues by constructing transgenic mice that expressed RBTN-2 under control of the metallothionein-1 promoter. Despite overexpression of RBTN-2 in all tissues, transgenic mice developed T-cell tumors only, thus indicating that tumorigenesis caused by RBTN-2 is T-cell-specific. Thymic tumors were found between 37 and 71 weeks and were invariably associated with metastasis to nonlymphoid organs. Thymuses from apparently healthy transgenic mice were also examined. In some mice there was an 10-fold increase in the CD4-CD8- thymocyte subset, yet the total number of thymocytes was the same as that in wild-type mice. Thymic homeostasis was maintained by a compensatory reduction in the CD4+CD8+ subset. The expansion of CD4-CD8- thymocytes was associated with increased expression of RBTN-2 and with increased cell proliferation. No differences were found in the proportion of thymocytes undergoing apoptosis in transgenic mice. Furthermore, RBTN-2-induced expansion of CD4-CD8- cells did not block differentiation of these cells. Thymuses with 30% CD4-CD8- cells were essentially monoclonal, indicating that all thymic immunophenotypes were derived from a single clone. Overall, our data are consistent with the following scenario: (1) RBTN-2 expression in T cells causes selective and polyclonal proliferation of CD4-CD8-thymocytes accompanied by a compensatory decrease in other thymocyte subsets; (2) a clone with growth advantage and differentiation potential is selected and populates the thymus; and (3) this clone eventually breaches homeostasis of the thymus, accompanied or followed by metastasis to other organs.</jats:p

A gene transfer strategy for making bone marrow cells resistant to trimetrexate

Trimetrexate (TMTX) is an anticancer drug with potential advantages over the more commonly used antifolate, methotrexate (MTX); however, its use has been limited by severe myelosuppression. Retroviral vectors containing mutant dihydrofolate reductase (DHFR) genes have been used to protect bone marrow cells from MTX, suggesting a similar approach could be used for TMTX. We first screened six variants of human DHFR to determine which allowed maximal TMTX resistance in fibroblasts. A variant enzyme containing a Leu-to-Tyr mutation in the 22nd codon (L22Y) was best, allowing a 100-fold increase in resistance over controls. Murine hematopoietic progenitor cells transduced with an L22Y- containing retroviral vector also showed high-level TMTX resistance in vitro. Mice reconstituted with L22Y-transduced bone marrow cells were challenged with a 5-day course of TMTX to determine whether hematopoiesis could be protected in vivo. Transfer of the L22Y vector resulted in consistent protection from TMTX-induced neutropenia and reticulocytopenia at levels that correlated with the proviral copy number in circulating leukocytes. We conclude that the L22Y vector is highly effective in protecting hematopoiesis from TMTX toxicity and may provide a means for increasing the therapeutic utility of TMTX in certain cancers.</jats:p

The Wistar Furth rat: an animal model of hereditary macrothrombocytopenia

Abstract The mechanisms that determine and regulate platelet size are unknown. By phase microscopy, we observed that Wistar Furth (WF) rats had macrothrombocytopenia. In this study, we have characterized and compared platelets and megakaryocytes of WF rats with those of Wistar, Long-Evans hooded (LE), and Sprague-Dawley rats. In addition, we have examined the mode of inheritance of this WF rat platelet abnormality. The average platelet count of WF rats was only one-third that of the other three rat strains. In contrast, the mean platelet volume (MPV) of adult WF rats was twice that of the other rat strains; however, the average megakaryocyte diameter and DNA content distribution of WF rats were not significantly different from those of LE rats. The average megakaryocyte concentration was 30% lower in the WF strain compared with that of LE rats. Mazelike membrane formations were observed in WF platelets and megakaryocytes by electron microscopy. Reciprocal crosses of WF and LE rats resulted in offspring with MPVs and platelet counts like those of LE rats, indicating that the macrothrombocytopenic trait is recessive in its inheritance. Reciprocal marrow transplants between the WF and LE strains resulted in MPVs like those of the donor strain, demonstrating that the macrothrombocytopenia is an intrinsic marrow abnormality of the WF strain. Splenectomy did not alter the MPV of WF rats. The response of WF megakaryocytes and platelets to severe, acute thrombocytopenia was similar to that of LE rats except that the shift to higher megakaryocyte DNA contents was muted and platelet recovery was slower in the WF rats. In summary, the WF rat has a hereditary macrothrombocytopenia that is recessive in nature and not due to differences in megakaryocyte size or DNA content. These results suggest that the macrothrombocytopenia of WF rats results from the formation of fewer platelets per megakaryocyte, possibly resulting from a qualitative or quantitative defect in some component necessary for proper subdivision of megakaryocyte cytoplasm into platelets.</jats:p