Quantitative habitat models for the conservation of the endangered European crayfish Austropotamobius pallipes complex (Astacoidea: Astacidae)

Crayfish are the largest mobile freshwater invertebrates and are keystone species in European aquatic ecosystems particularly in small streams and rivers. The white-clawed crayfish Austropotamobius pallipes (a species complex) is currently classified by the IUCN Red List as an endangered species (EN), because its populations have decreased significantly over the last decades in a number of European countries including Italy, due mainly to habitat modifications and the introduction and spread of alien species. Data on the ecological requirements of A. pallipes are needed to quantify the effects of habitat alteration, to simulate restoration scenarios, and to implement effective conservation measures for this species. We describe here a new methodology for modelling the habitat requirements for this endangered crayfish using the mesohabitat scale approach based on data from crayfish living in small streams draining the Italian foothills of the Alps (Lombardy region) and in streams in the mountainous areas of the Gran Sasso and Monti della Laga National Park (Abruzzo region). Data from seven morphologically different streams were used to train and validate the habitat models. The Random Forests algorithm was used to identify the best and most parsimonious habitat model, and to define the lowest number of variables to be surveyed in the future. The best habitat models were applied to each stream and used to classify each mesohabitat into suitability categories. Habitat flow-rating curves were developed to analyze spatio-temporal variation of habitat availability, and habitat time series analysis were used to define detailed management schemes for environmental river management. Flow releases and water temperature regimes were assessed for individual water diversions in order (1) to represent how physical habitat changes through time, and (2) to identify stress conditions for A. pallipes created by the persistent limitation of habitat availability. Results indicated that the kind of substrate in the stream bed (such as the proportion of fine-grained substrates), the water depth (whether shallow or deep), and the available cover (such as the presence of boulders, woody debris, and undercut banks) were all significant factors governing the occurrence of crayfish. The habitat models performed well in both calibration and validation phases (with accuracy ranging from 71% to 79% in training and from 69% to 73% in validation) and can be considered to be a valuable tool to predict the distribution of A. pallipes over a wide range of stream types. An example of how to establish environmental standards for small streams is presented. The proposed habitat model provides a useful tool that can be applied even when other commonly used methodologies are unsuitable. As such, this habitat model can be used to develop regional rules for the conservation of the endangered crayfish A. pallipes complex and for defining more site-specific management criteria

A Novel Molecular Complex Expressed on Immature B Cells: A Possible Role in T Cell-Independent B Cell Development

To identify surface molecules that may play a role in regulating ileal Peyer's patch (PP) B cell growth, we generated monoclonal antibodies (mAbs) and then selected them for a unique reactivity with ileal PP B cells. Flow cytometric analysis identified a mAb (SIC4.8R) that labeled 97% of ileal and 50–60% of jejunal PP sIgM+B cells. SIC4.8R also labeled a subpopulation of cortical thymocytes but few B or T cells in other lymphoid tissues, including bone marrow. Immunohistochemistry revealed intense SIC4.8R staining of B cells in the cortex of ileal PP follicles. SIC4.8R also labeled bovine PP B cells, a murine pro-B cell line, and pre-B cells in human bone marrow. Protein chemistry revealed that a structurally similar molecular complex was expressed on sheep ileal PP B cells and thymocytes and murine pro-B cells. Addition of soluble SIC4.8R to cultured ileal PP B cells reduced apoptotic cell death, elevated proliferative responses, partially inhibited anti-Ig-induced cell death, and induced IL-4 responsiveness. In contrast, soluble SIC4.8R had an antiproliferative effect on a mouse pro-B cell line. Finally, SIC4.8R labeling declined following the stimulation of ileal PP B cells with CD40 ligand. In conclusion, the present investigation determined that SIC4.8R identified a novel molecular complex that is expressed at several stages of T cell-independent B cell development in a variety of mammalian species. This observation confirmed that PP B cells are developmentally distinct from other B cell populations in sheep and suggested that the bone marrow may not be a site of B lymphopoiesis in young lambs

Ordering of human bone marrow B lymphocyte precursors by single-cell polymerase chain reaction analyses of the rearrangement status of the immunoglobulin H and L chain gene loci

CD19+CD10+ human B lineage bone marrow cells were separated into cycling or resting cells, which differ in their expression of CD34, V(preB), recombination activating gene (RAG-1), and terminal deoxynucleotidyl transferase (TdT). Polymerase chain reaction analyses developed for D(H)J(H) and V(K)J(K) V(K)J(K)K((de)) and V(K)K((de)) rearrangements with DNA of single cells and a comparison with B lineage cell development in mouse bone marrow, allow to delineate the human B lymphocyte pathway of development as follows: CD34+V(preB)+RAG-1+TdT+, D(H)J(H)-rearranged, KL germline cycling pre-B I cells → CD34- V(preB)+μH chain+ (pre-B receptor+) RAG- 1+ TdT, V(H)D(H)J(H)-rearranged, KL germline, cycling pre-B II cells → CD34- V(preB)-, intracytoplasmic μH chain+ (pre-B receptor) RAG-1+ TdT, V(H)D(H)J(H)-rearranged, mainly KL germline cycling pre-B II cells → CD34+ V(preB)- intracytoplasmic μH chain+, RAG-1+ TdT, V(H)D(H)J(H)-rearranged, V(K)J(K)-rearranged, IgM-, resting pre-B II cells → CD34+ V(preB)-, sIgM+, RAG-1+ TdT-, V(H)D(H)J(H)- and V(K)J(K)-rearranged IgM+ immature B cells → CD34+, CD10- sIgM+/sIgD+ mature B cells. This order, for the first time established for human B lineage cells, shows striking similarities with that established for mouse B lineage cells in bone marrow.We thank Drs. Rod Ceredig and Thomas Winkler for critical reading of this manuscript. We are grateful to Marcus Dessing for his outstanding skill at the FACSÒ sorter and his extraordinary help during long, unusual hours. We thank Prof. A. Gratwohl, Dr. E. Signer, and Dr. U. Ramenghi for providing the bone marrow samples and Prof. F. Caligaris Cappio for continuous encouragement and discussions. We gratefully acknowledge Ms. Nadia Straube’s technical experience in DNA sequencing. The Basel Institute for Immunology was founded and is supported by F. Hoffmann-La Roche Ltd., Basel, Switzerland. E. Sanz was supported by contracts from the CSIC and grant CAM92/126, and A. de la Hera was supported by grants SAF-93-0925 and SAF-96-0201 from the CICY.Peer reviewedPeer Reviewe

Invariant NKT cells contribute to chronic lymphocytic leukemia surveillance and prognosis

Chronic lymphocytic leukemia (CLL) is characterized by the expansion of malignant CD5(+) B lymphocytes in blood, bone marrow and lymphoid organs. CD1d-restricted invariant Natural Killer T (iNKT) cells are innate-like T lymphocytes strongly implicated in tumor surveillance. We investigated the impact of iNKT cells in the natural history of the disease both in Eμ;-Tcl1 (Tcl1) CLL mouse model and 68 CLL patients. We found that Tcl1-CLL cells express CD1d and iNKT cells critically delay the disease onset, but become functionally impaired upon disease progression. In patients, disease progression correlates also with high CD1d expression on CLL cells and impaired iNKT cells. Conversely, disease stability correlates with negative/low CD1d expression on CLL cells and normal iNKT cells, suggesting an indirect leukemia control. iNKT cells indeed hinder CLL survival in vitro by restraining CD1d-expressing Nurse Like Cells, a relevant pro-leukemia macrophage population. Finally, multivariate analysis identifies iNKT cell frequency as independent predictor of disease progression. Together, these results support iNKT cell contribution to CLL immune-surveillance and highlight iNKT cell frequency as prognostic marker for disease progression

TP53 aberrations in chronic lymphocytic leukemia: an overview of the clinical implications of improved diagnostics

Chronic lymphocytic leukemia is associated with a highly heterogeneous disease course in terms of clinical outcomes and responses to chemoimmunotherapy. This heterogeneity is partly due to genetic aberrations identified in chronic lymphocytic leukemia cells such as mutations of TP53 and/or deletions in chromosome 17p [del(17p)], resulting in loss of one TP53 allele. These aberrations are associated with markedly decreased survival and predict impaired response to chemoimmunotherapy thus being among the strongest predictive markers guiding treatment decisions in chronic lymphocytic leukemia. Clinical trials demonstrate the importance of accurately testing for TP53 aberrations [both del(17p) and TP53 mutations] before each line of treatment to allow for appropriate treatment decisions that can optimize patient outcomes. The current report reviews the diagnostic methods to better detect TP53 disruption, the role of TP53 aberrations in treatment decisions and current therapies available for patients with chronic lymphocytic leukemia carrying these abnormalities. The standardization in sequencing technologies for accurate identification of TP53 mutations and the importance of continued evaluation of TP53 aberrations throughout initial and subsequent lines of therapy remain unmet clinical needs as new therapeutic alternatives become availabl