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Xenopus is a genus of African clawed frogs including two species, X. tropicalis and X. laevis that are extensively used in experimental biology, immunology, and biomedical studies. The availability of fully sequenced and annotated Xenopus genomes is strengthening genome-wide analyses of gene families and transgenesis to model human diseases. However, inaccuracies in genome annotation for genes involved in the immune system (i.e., immunome) hamper immunogenetic studies. Furthermore, advanced genome technologies (e.g., single-cell and RNA-Seq) rely on well-annotated genomes. The annotation problems of Xenopus immunome include a lack of established orthology across taxa, merged gene models, poor representation in gene pages on Xenbase, misannotated genes and missing gene IDs. The Xenopus Research Resource for Immunobiology in collaboration with Xenbase and a group of investigators are working to resolve these issues in the latest versions of genome browsers. In this review, we summarize the current problems of previously misannotated gene families that we have recently resolved. We also highlight the expansion, contraction, and diversification of previously misannotated gene families. 

Abstract The fms-related tyrosine kinase 3 (Flt3) and its ligand (Flt3lg) are important regulators of hematopoiesis and dendritic cell (DC) homeostasis with unsettled coevolution. Gene synteny and deduced amino acid sequence analyses identified conserved flt3 gene orthologs across all jawed vertebrates. In contrast, flt3lg orthologs were not retrieved in ray-finned fish, and the gene locus exhibited more variability among species. Interestingly, duplicated flt3/flt3lg genes were maintained in the allotetraploid Xenopus laevis. Comparison of modeled structures of X. laevis Flt3 and Flt3lg homoeologs with the related diploid Xenopus tropicalis and with humans indicated a higher conformational divergence between the homoeologous pairs than their respective counterparts. The distinctive developmental and tissue expression patterns of Flt3 and Flt3lg homoeologs in tadpoles and adult frogs suggest a subfunctionalization of these homoeologs. To characterize Flt3 cell surface expression, X. laevis–tagged rFlt3lg.S and rFlt3lg.L were produced. Both rFlt3lg.S and rFlt3lg.L bind in vitro Flt3.S and Flt3.L and can trigger Erk1/2 signaling, which is consistent with a partial overlapping function between homoeologs. In spleen, Flt3.S/L cell surface expression was detected on a fraction of B cells and a population of MHC class IIhigh/CD8+ leukocytes phenotypically similar to the recently described dual follicular/conventional DC-like XL cells. Our result suggests that 1) Flt3lg.S and Flt3lg.L are both involved in XL cell homeostasis and that 2) XL cells have hematopoietic origin. Furthermore, we detected surface expression of the macrophage/monocyte marker Csf1r.S on XL cells as in mammalian and chicken DCs, which points to a common evolutionary origin in vertebrate DCs. 

ABSTRACT Ranaviruses such as frog virus 3 (FV3) are large double-stranded DNA (dsDNA) viruses causing emerging infectious diseases leading to extensive morbidity and mortality of amphibians and other ectothermic vertebrates worldwide. Among the hosts of FV3, some are highly susceptible, whereas others are resistant and asymptomatic carriers that can take part in disseminating the infectious virus. To date, the mechanisms involved in the processes of FV3 viral persistence associated with subclinical infection transitioning to lethal outbreaks remain unknown. Investigation in Xenopus laevis has revealed that in asymptomatic FV3 carrier animals, inflammation induced by heat-killed (HK) Escherichia coli stimulation can provoke the relapse of active infection. Since Toll-like receptors (TLRs) are critical for recognizing microbial molecular patterns, we investigated their possible involvement in inflammation-induced FV3 reactivation. Among the 10 different TLRs screened for changes in expression levels following FV3 infection and HK E. coli stimulation, only TLR5 and TLR22, both of which recognize bacterial products, showed differential expression, and only the TLR5 ligand flagellin was able to induce FV3 reactivation similarly to HK E. coli . Furthermore, only the TLR5 ligand flagellin induced FV3 reactivation in peritoneal macrophages both in vitro and in vivo . These data indicate that the TLR5 signaling pathway can trigger FV3 reactivation and suggest a role of secondary bacterial infections or microbiome alterations (stress or pollution) in initiating sudden deadly disease outbreaks in amphibian populations with detectable persistent asymptomatic ranavirus. IMPORTANCE This study in the amphibian Xenopus laevis provides new evidence of the critical role of macrophages in the persistence of ranaviruses in a quiescent state as well as in the reactivation of these pathogens into a virulent infection. Among the multiple microbial sensors expressed by macrophages, our data underscore the preponderant involvement of TLR5 stimulation in triggering the reactivation of quiescent FV3 in resident peritoneal macrophages, unveiling a mechanistic connection between the reactivation of persisting ranavirus infection and bacterial coinfection. This suggests a role for secondary bacterial infections or microbiome alterations (stress or pollution) in initiating sudden deadly disease outbreaks in amphibian populations with detectable persistent asymptomatic ranavirus.