Babesia caballi is an intra-erythrocytic parasite causing equine piroplasmosis. Three B. caballi genotypes (A, B, and C) have been identified based on the 18 S rRNA and rhoptry-associated protein ...(rap-1) gene sequences. These variant parasite genotypes compromise the diagnostic utility of the WOAH-recommended serological assays in declaring horses free of equine piroplasmosis. Although a gene encoding a spherical body protein 4 (sbp4) has recently been identified as a potential antigen for the serological detection of B. caballi, the ability of this antigen to detect the different geographical strains has not been determined. The molecular distinction between variant B. caballi genotypes is limited and therefore we developed molecular typing assays for the rapid detection and quantification of distinct parasite genotypes. Field samples were screened for the presence of B. caballi using an established multiplex equine piroplasmosis qPCR assay. In this study, B. caballi genotype A was not detected in any field samples screened. However, phylogenetic analysis of the amplified sbp4 and 18 S rRNA genes confirmed the phylogenetic groupings of the South African isolates into either B. caballi genotypes B or C. A multiple sequence alignment of the sbp4 gene sequences obtained in this study together with the published sbp4 sequences representing B. caballi genotype A, were used to identify conserved regions within the gene to design three primer pairs and three genotype-specific TaqMan minor-groove binder (MGB™) probes. The qPCR assays were shown to be specific and efficient in the detection and differentiation between B. caballi genotypes A, B, and C and could be used as a diagnostic assay to prevent the unintentional spread of variant B. caballi genotypes globally.
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•Three B. caballi genotypes (A, B, C) occur that complicate diagnostics.•The sbp4 gene was targeted for molecular typing qPCR assays.•Phylogenetic analysis confirmed the genotype grouping.•Conserved regions of the sbp4 gene were identified for assay design.•qPCR assays detect and differentiate genotypes efficiently.
The guidelines for the implementation and reporting of HLA nomenclature for the World Marrow Donor Association have served as a reliable standard for communication of HLA data in the hematopoietic ...cell transplantation process. Wider use of next‐generation sequencing made a special provision of the guidelines increasingly pertinent: how to communicate novel HLA alleles. Novel alleles need to be recognized by the WHO Nomenclature Committee for Factors of the HLA system to obtain official allele designations. Until then they have to be handled according to the specific rules. Leaving the actual rules basically unchanged we give some advice on how to communicate novel alleles to best facilitate the search process for cases where novel alleles are identified on donor or patient side.
Although the taxonomy of
has been extensively scrutinized, significant uncertainty remains regarding the generic boundaries and composition of this large and heterogeneous taxon. Here we used the ...amino acid and nucleotide sequences of 106 conserved proteins from 92 species to infer robust maximum likelihood phylogenies with which to investigate the generic structure of
sensu lato. These data unambiguously supported five distinct lineages, of which four correspond to
sensu stricto and the newly introduced genera
,
, and
. The fifth lineage was represented by
. Based on these findings, we propose 13 new combinations for those species previously described as members of
but that form part of
. These findings also suggest revision of the taxonomic status of
as it is does not form part of any of the genera currently recognized in
sensu lato. From a phylogenetic point of view,
sensu stricto has a sister relationship with the
+
clade. Also, the lineages represented by
and
, respectively, emerged prior to the radiation of the
sensu stricto+
+
clade. Our findings therefore constitute a solid framework, not only for supporting current and future taxonomic decisions, but also for studying the evolution of this assemblage of medically, industrially and agriculturally important species.
Functional genomics studies in minimal mycoplasma cells enable unobstructed access to some of the most fundamental processes in biology. Conventional transposon bombardment and gene knockout ...approaches often fail to reveal functions of genes that are essential for viability, where lethality precludes phenotypic characterization. Conditional inactivation of genes is effective for characterizing functions central to cell growth and division, but tools are limited for this purpose in mycoplasmas. Here we demonstrate systems for inducible repression of gene expression based on clustered regularly interspaced short palindromic repeats-mediated interference (CRISPRi) in Mycoplasma pneumoniae and synthetic Mycoplasma mycoides, two organisms with reduced genomes actively used in systems biology studies. In the synthetic cell, we also demonstrate inducible gene expression for the first time. Time-course data suggest rapid kinetics and reversible engagement of CRISPRi. Targeting of six selected endogenous genes with this system results in lowered transcript levels or reduced growth rates that agree with lack or shortage of data in previous transposon bombardment studies, and now produces actual cells to analyze. The ksgA gene encodes a methylase that modifies 16S rRNA, rendering it vulnerable to inhibition by the antibiotic kasugamycin. Targeting the ksgA gene with CRISPRi removes the lethal effect of kasugamycin and enables cell growth, thereby establishing specific and effective gene modulation with our system. The facile methods for conditional gene activation and inactivation in mycoplasmas open the door to systematic dissection of genetic programs at the core of cellular life.
The rhesus macaque (Macaca mulatta) is an abundant primate species that diverged from the ancestors of Homo sapiens about 25 million years ago. Because they are genetically and physiologically ...similar to humans, rhesus monkeys are the most widely used nonhuman primate in basic and applied biomedical research. We determined the genome sequence of an Indian-origin Macaca mulatta female and compared the data with chimpanzees and humans to reveal the structure of ancestral primate genomes and to identify evidence for positive selection and lineage-specific expansions and contractions of gene families. A comparison of sequences from individual animals was used to investigate their underlying genetic diversity. The complete description of the macaque genome blueprint enhances the utility of this animal model for biomedical research and improves our understanding of the basic biology of the species.
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