Genetic characterization of Toxoplasma gondii isolates from chickens in India by GRA6 gene sequence analysis.
Acta Parasitol. 2014 Oct;59(4):666-74
Authors: Biradar SS, Saravanan BC, Tewari AK, Sreekumar C, Sankar M, Sudhakar NR
PCR-RFLP and nucleotide sequencing based genotyping of Toxoplasma gondii Indian isolates (Izatnagar and Chennai isolates and Chennai clone) vis-a vis RH-IVRI strain was conducted by targeting GRA6 as genetic marker. The 791 bp GRA6 product was PCR amplified from the genomic DNA of different T. gondii Indian isolates, including the RH-IVRI strain. Tru1I restriction endonuclease based PCR-RFLP of GRA6 sequence produced polymorphic digestion pattern that discriminated the virulent RH-IVRI strain (as type I) from the moderately virulent local isolates as type III. The PCR amplicon of T. gondii GRA6 from RH-IVRI strain as well as from the local isolates were cloned in cloning vector and custom sequenced. The nucleotide and deduced amino acid sequences of T. gondii isolates were aligned with that of the type I, II and III strains (RH, BEVERLEY, ME49, C56, TONT and NED) available in public domain and analyzed in silico using MEGA version 4.0 software. Nucleotide sequencing and phylogenetic analysis of GRA6 marker from the Indian isolates revealed a close genetic relationship with type III strains of T. gondii. Further, detection of a single nucleotide polymorphism (SNP) at positions 162 and 171 of the GRA6 marker, established the lineage of Indian isolates as type III. This is the first report on characterization of T. gondii lineage as type III in selective chicken population of India based on PCR-RFLP and sequence analysis of GRA6 gene.
PMID: 25236278 [PubMed - in process]
Use of single-nucleotide polymorphisms (SNPs) to distinguish gene expression subtypes of chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME).
J Clin Pathol. 2014 Sep 19;
Authors: Shimosako N, Kerr JR
AIMS: We have reported gene expression changes in patients with chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) and the fact that such gene expression data can be used to identify subtypes of CFS/ME with distinct clinical phenotypes. Due to the difficulties in using a comparative gene expression method as an aid to CFS/ME disease and subtype-specific diagnosis, we have attempted to develop such a method based on single-nucleotide polymorphism (SNP) analysis.
METHODS: To identify SNP allele associations with CFS/ME and CFS/ME subtypes, we tested genomic DNA of patients with CFS/ME (n=108), patients with endogenous depression (n=17) and normal blood donors (n=68) for 504 human SNP alleles located within 88 CFS-associated human genes using the SNP Genotyping GoldenGate Assay (Illumina, San Diego, California, USA). 360 ancestry informative markers (AIM) were also examined.
RESULTS: 21 SNPs were significantly associated with CFS/ME compared with depression and normal groups. 148 SNP alleles had a significant association with one or more CFS/ME subtypes. For each subtype, associated SNPs tended to be grouped together within particular genes. AIM SNPs indicated that 4 subjects were of Asian origin while the remainder were Caucasian. Hierarchical clustering of AIM data revealed the relatedness between 2 couples of patients with CFS only and confirmed the overall heterogeneity of all subjects.
CONCLUSIONS: This study provides evidence that human SNPs located within CFS/ME associated genes are associated with particular genomic subtypes of CFS/ME. Further work is required to develop this into a clinically useful subtype-specific diagnostic test.
PMID: 25240059 [PubMed - as supplied by publisher]
An overview of SNP interactions in genome-wide association studies.
Brief Funct Genomics. 2014 Sep 19;
Authors: Li P, Guo M, Wang C, Liu X, Zou Q
With the recent explosion in high-throughput genotyping technology, the amount and quality of single-nucleotide polymorphism (SNP) data has increased exponentially. Therefore, the identification of SNP interactions that are associated with common diseases is playing an increasing and important role in interpreting the genetic basis of disease susceptibility and in devising new diagnostic tests and treatments. However, because these data sets are large, although they typically have small sample sizes and low signal-to-noise ratios, there has been no major breakthrough despite many efforts, making this a major focus in the field of bioinformatics. In this article, we review the two main aspects of SNP interaction studies in recent years-the simulation and identification of SNP interactions-and then discuss the principles, efficiency and differences between these methods.
PMID: 25241224 [PubMed - as supplied by publisher]
Pyrosequencing for Classification of Human FcγRIIIA Allotypes: A Comparison with PCR-Based Techniques.
Mol Diagn Ther. 2014 Sep 18;
Authors: Matlawska-Wasowska K, Gale JM, Nickl CK, Khalili P, Shirley B, Wilson BS, Vasef MA, Winter SS
BACKGROUND: Surface-specific antigens expressed by hematopoietic cells are attractive targets for antibody-mediated immunotherapy. Monoclonal antibodies (mAbs) involve various mechanisms to eliminate target cells, including antibody-dependent cellular cytotoxicity (ADCC)- and phagocytosis (ADCP)-mediated killing through natural killer (NK) and macrophage effector cells bearing FcγRIIIA (CD16). The clinical efficacy of ADCC is particularly impacted by a single nucleotide polymorphism (SNP) found in the gene encoding FcγRIIIA (FCGR3A), which generates a variable distribution of the 158 V/V, F/V or F/F CD16 allotypes (F = phenylalanine, V = valine) in the normal human population. Currently, most patients are not screened for CD16 allotypes, creating the potential to include in their treatment a mAb-based therapy that may have limited benefit. Therefore, it is important to identify CD16 allotypes when considering mAb therapies that require ADCC/ADCP.
OBJECTIVE: The objective of this study was to develop a reliable PCR-based assay for classification of human FcγRIIIA allotypes.
METHODS: We studied 42 normal human subjects for the incidence of FcγRIIIA-158 polymorphisms using comparative molecular approaches.
RESULTS: The results of our study showed 100 % accuracy in genotyping by pyrosequencing. In contrast, nested PCR-based allele-specific restriction assay and quantitative PCR techniques proved to be relatively less sensitive and less specific in distinguishing variant genotypes.
CONCLUSION: Since the efficacy of the mAb-based targeted immunotherapy may be highly dependent upon the CD16 polymorphism in a given individual, we recommend pyrosequencing for CD16 allotype testing.
PMID: 25230857 [PubMed - as supplied by publisher]