Abstract: The availability of genotyping platforms for comprehensive genetic analysis of complex traits has resulted in a plethora of studies reporting the association of specific single-nucleotide polymorphisms (SNPs) with common diseases or drug responses. However, detailed genetic analysis of these associated regions that would correlate particular polymorphisms to phenotypes has lagged. This is primarily due to the lack of technologies that provide additional sequence information about genomic regions surrounding specific SNPs, preferably in haploid form. Enrichment methods for resequencing should have the specificity to provide DNA linked to SNPs of interest with sufficient quality to be used in a cost-effective and high-throughput manner. We describe a simple, automated method of targeting specific sequences of genomic DNA that can directly be used in downstream applications. The method isolates haploid chromosomal regions flanking targeted SNPs by hybridizing and enzymatically elongating oligonucleotides with biotinylated nucleotides based on their selective binding to unique sequence elements that differentiate one allele from any other differing sequence. The targeted genomic region is captured by streptavidin-coated magnetic particles and analyzed by standard genotyping, sequencing or microarray analysis. We applied this technology to determine contiguous molecular haplotypes across a approximately 150 kb genomic region of the major histocompatibility complex.
Abstract: Necrotizing pneumonia and fatal septic shock were caused by Panton-Valentine leukocidin-positive, community-acquired, methicillin-resistant Staphylococcus aureus (CA-MRSA) in a previously healthy, 61-y-old female. This patient did not belong to any high-risk group (e.g. homosexuals, military recruits, sports team members, etc.). CA-MRSA infection should be suspected in any adult with severe pneumonia/sepsis.
Abstract: Substantial evidence has been accumulated suggesting that T cells in patients with epithelial ovarian carcinoma (EOC) exhibit an antigen-driven immune response directed against the tumor cells. In the context of human leukocyte antigen (HLA), this suggests its possible involvement in the disease. Therefore, we examined the distribution of the HLA-DRB1*, -DQA1*, and -DQB1* alleles in 47 patients with EOC and 67 healthy Caucasian women. The frequency of D(70) and E(71) polymorphic residues of the DRB1 alleles was significantly reduced in EOC patients versus controls (pD(70)E(71) = 0.009), suggesting a protective role against the disease. The DQalpha residues R(52) and Y(11)R(55) were increased in the patients (p = 0.008 and 0.012, respectively). Because residues 11 and 55 participate in the formation of pocket 1, they may be functionally important amino acid positions that influence disease susceptibility. The frequency of the DQalpha susceptibility epitope (R(52)Y(11)R(55)) among the DRbetaD(70)E(71)-positive EOC patients was increased when compared with DRbetaD(70)E(71)-positive controls (EOC, 100%; control, 52%; p = 0.028). Among individuals without the DQalpha susceptibility epitope, the distribution of DRbetaD(70)E(71)-positive cases was significantly different between EOC patients and controls (EOC, 0%; control, 60%; p = 0.039). Therefore, it appears that the presence of DQalpha susceptibility elements overrides the protective effect of the DRbetaD(70)E(71) epitope and suggests an interactive relationship between DRbeta and DQalpha epitopes that may be of importance for disease susceptibility. Because positions DRbeta 70,71 and DQalpha 52 have been implicated in immunologic diseases, it is likely that besides being critical for T-cell recognition, they may also play a role in T-cell development and acquisition of the T-cell repertoire.
Abstract: Whipple's disease is a rare chronic multi-systemic infectious disorder caused by the Gram-positive bacillus, Tropheryma whippelii. Infection may involve any organ in the body, and most commonly affects white men in the fourth to sixth decades of life. The most common presenting symptoms are gastrointestinal and include abdominal pain, diarrhoea, anorexia and associated weight loss. However, the variability in presentation is considerable and some patients may present with intermittent low-grade fever, neurological abnormalities (nystagmus, ophthalmoplegia, cranial nerve defects), migratory arthralgia, lymphadenopathy, or involvement of the cardiovascular system. In typical Whipple's disease, the most severe changes are seen in the proximal small intestine and biopsy reveals mucosal and lymph node infiltration with large, foamy histocytes, containing granules that stain positive with periodic acid-Schiff (PAS) reagent and represent intact or partially degraded bacteria. Extended antibiotic treatment (up to 1-year) is indicated. Life-long surveillance for recurrence is essential, once primary treatment has been completed. We report the case of a 58-year-old man who developed a rare infection with the actinobacterium, T. whippelii. The patient had suffered intermittent episodes of varying clinical symptoms associated with multiple hospital admissions and clinical diagnoses, spanning a period of 22 years. Historically, arthralgia was the primary manifestation in this patient and also was the chief complaint for which he was first hospitalized 22 years ago. At his most recent admission to our hospital department, his presenting symptoms were persistent fatigue, weight loss, arthralgia and diarrhoea. Thus, it is essential that clinicians retain a high index of suspicion for T. whippelii infection in patients who have a long-term history of arthritis, fever and diarrhoea.
