Molecular phylogenetics reveals the evolutionary history of marine fishes (Actinopterygii) endemic to the subtropical islands of the Southwest Pacific

Remote oceanic islands of the Pacific host elevated levels of actinopterygian (ray-finned fishes) endemism. Characterizing the evolutionary histories of these endemics has provided insight into the generation and maintenance of marine biodiversity in many regions. The subtropical islands of Lord Howe, Norfolk, and Rangitāhua (Kermadec) in the Southwest Pacific are yet to be comprehensively studied. Here, we characterize the spatio-temporal diversification of marine fishes endemic to these Southwest Pacific islands by combining molecular phylogenies and the geographic distribution of species. We built Bayesian ultrametric trees based on open-access and newly generated sequences for five mitochondrial and ten nuclear loci, and using fossil data for time calibration. We present the most comprehensive phylogenies to date for marine ray-finned fish genera, comprising 34 species endemic to the islands, including the first phylogenetic placements for 11 endemics. Overall, our topologies confirm the species status of all endemics, including three undescribed taxa. Our phylogenies highlight the predominant affinity of these endemics with the Australian fish fauna (53%), followed by the East Pacific (15%), and individual cases where the closest sister taxon of our endemic is found in the Northwest Pacific and wider Indo-Pacific. Nonetheless, for a quarter of our focal endemics, their geographic affinity remains unresolved due to sampling gaps within their genera. Our divergence time estimates reveal that the majority of endemic lineages (67.6%) diverged after the emergence of Lord Howe (6.92 Ma), the oldest subtropical island in the Southwest Pacific, suggesting that these islands have promoted diversification. However, divergence ages of some endemics pre-date the emergence of the islands, suggesting they may have originated outside of these islands, or, in some cases, ages may be overestimated due to unsampled taxa. To fully understand the role of the Southwest Pacific subtropical islands as a 'cradle' for diversification, our study advocates for further regional surveys focused on tissue collection for DNA analysis.fals

Identification of lymphoma-associated antigens using SEREX.

Serological analysis of recombinant complementary deoxyribonucleic acid (cDNA) expression libraries (SEREX) is a powerful approach to identify immunogenic cancer-associated proteins using antibodies that are naturally present in the serum of cancer patients. This technique involves the screening of a relevant cDNA expression library with patient serum that has been cleaned to remove any antibodies that may recognize bacterial and/or viral proteins. Once antigens have been identified and their reactivity has been confirmed with a second round of screening, the gene encoding the protein can be sequenced and identified. Antigens can then be screened with a panel of allogenic sera from other patients and control individuals. This identifies disease-specific antigens, which may be useful diagnostic markers or, alternatively, targets for immunotherapy. This chapter describes the SEREX methodology in full

Serologic detection of diffuse large B-cell lymphoma-associated antigens.

Diffuse large B-cell lymphoma (DLBCL) accounts for 30-40% of all adult non-Hodgkin's lymphomas, yet the understanding of its underlying genetic abnormalities remains poor. Our present study used the serological analysis of recombinant cDNA expression libraries (SEREX) technique to identify DLBCL-associated antigens. SEREX screening of testis libraries has previously identified cancer-testis antigens (CTAs) that may act as disease-specific targets for immunotherapy. Screening a testis cDNA expression library with serum from a DLBCL patient identified a total of 94 positive clones, representing 28 distinct antigens. Two of these antigens were novel, 8 were previously uncharacterised, and the remainder were proteins of known function. Screening of the antigens with sera from DLBCL (n = 10), acute myeloid leukaemia (AML, n = 10) and chronic myeloid leukaemia (CML, n = 10) patients, alongside normal healthy donor controls (n = 20), revealed that 7 of the antigens were recognised by DLBCL sera but not normal donor sera, whilst 2 of these antigens were also recognised by leukaemic sera. Some of the genes identified here were already known to be transcribed in DLBCL. The mRNA expression of the majority of the remaining antigens was confirmed in DLBCL cell lines using reverse-transcriptase PCR (RT-PCR). Our study identified a number of DLBCL associated antigens that may be suitable as prognostic/diagnostic markers and/or for the immunotherapy of haematologic malignancies

A panel of cancer-testis genes exhibiting broad-spectrum expression in haematological malignancies.

Cancer-testis (CT) antigens/genes show restricted expression in normal tissues but widespread expression in many tumour types. This, coupled with their ability to induce cytotoxic T-lymphocyte responses, makes them attractive vaccine candidates. Following our identification of PASD1, we have used RT-PCR to analyse the mRNA expression profile of a large panel of CT genes in cell lines derived from haematological malignancies, and have studied Sp17 protein expression in the same cell lines and diffuse large B-cell lymphoma (DLBCL) biopsies. Our extensive analysis revealed multiple CT transcripts exhibiting widespread expression across cell lines derived from 21 B- and 4 T-cell malignancies. The broadest mRNA expression profiles were observed for the following eight CT genes: Sp17 (25/25), PRAME (25/25), CSAGE (24/25), PASD1 (22/25), CAGE/DDX53 (19/25), CTAGE1 (19/25), HAGE/DDX43 (16/25) and PLU-1/JARID1B (15/25). Cell lines derived from more aggressive lymphoma subtypes generally expressed CT transcripts at higher frequency. Sp17 protein was detected in a number of cell lines and in six of eleven (54.5%) DLBCL biopsies. Analysis of Sp17 protein expression, by immunohistochemistry and Western blotting, broadens the scope of this CT antigen as a potentially relevant clinical target in haematological malignancies. Further studies of protein expression are now needed to validate these antigens as vaccine candidates