Mutations in the DMD gene result in Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). Readily available clinical tests detect only deletions of one exon or greater, which are found in approximately 60% of cases. Mutational analysis of other types of DMD mutations, such as premature stop codons and small frameshifting insertions or deletions, has historically been hampered by the large size of the gene. We have recently reported a method that allows the rapid and economical sequencing of the entire coding region of the DMD gene, and that is more sensitive than methods based on single-strand conformational polymorphism (SSCP) screening or other preliminary screening steps. Here we use single condition amplification/internal primer (SCAIP) sequencing analysis, in combination with multiplex amplifiable probe hybridization (MAPH) analysis of duplications, to report the frequency of mutations in a large cohort of unselected dystrophinopathy patients from a single clinic. Our results indicate that 7% of dystrophinopathy patients do not have coding region mutations, suggesting that intronic mutations are not uncommon. The availability of rapid and thorough mutation analysis from peripheral blood samples, along with an improved estimate of the percentage of non-coding region mutations, will be of benefit for improved genetic counseling and in identification of cohorts for clinical trials.

Weight-bearing exercise is traditionally recommended for improving bone health in postmenopausal women. Effects of swim exercise were studied as an alternative to weight-bearing exercise in ovariectomized rats. Rats in a swim group (Sw, n = 8) swam for 12 wk, 5 days/wk for 60 min per session. A control group (Con, n = 9) engaged in no structured exercise. Femurs were analyzed for bone mineral density and for bone mineral content by dual energy X-ray absorptiometry, biomechanical properties by three-point bending (Instron), and bone structure and formation by histomorphometry. Food intake did not differ among groups. Final body weights were significantly lower in Sw compared with Con (P < 0.05). Swimmers had significantly greater femoral shaft bone mineral density and content (P < 0.05) compared with Con. Femurs of the Sw group had greater mechanical properties (P < 0.05) compared with Con. Histomorphometric data were significantly better in the Sw group compared with Con after the 12-wk intervention (P < 0.05). In conclusion, data from this study demonstrate some beneficial effects of swim exercise on bone structure, turnover, and strength.

Genetic tests are routinely ordered by health care providers (HCPs) within a wide range of medical specialties. Many providers have limited knowledge or experience with ordering and interpreting genetic tests; thus, test order errors are common. Rigorous review of genetic test orders by genetic counselors (GCs) can provide a direct financial benefit to medical institutions, patients and insurers. GCs at ARUP (Associated Regional University Pathologists) Laboratories routinely perform a preanalytic assessment of complex molecular genetic test orders that includes reviewing clinical and family history information and considering the clinical utility and cost-effectiveness of ordered tests. GCs contact the ordering institution and/or HCP as needed to collect additional clinical information and confirm the test order or suggest alternative testing based on the provided information. A retrospective review of the GC-facilitated test changes over a 21-month period at ARUP laboratories was performed. Approximately 26% of all requests for complex genetic tests assessing germ line mutations were changed following GC review. Testing fees associated with canceled tests were summed to estimate the cost-savings resulting from GC-facilitated test reviews. The test review process resulted in an average reduction in charges to the referring institutions of $48,000.00 per month. GC review of genetic test orders for appropriateness and clinical utility reduces healthcare costs to hospitals, insurers, and patients.

Lynch syndrome is characterized by mutations in the mismatch repair genes MLH1, MSH2, MSH6, and PMS2. In PMS2, detection of mutations is confounded by numerous pseudogenes. Detection of 3' deletions is particularly complicated by the pseudogene PMS2CL, which has strong similarity to PMS2 exons 9 and 11-15, due to extensive gene conversion. A newly designed multiplex ligation-dependent probe amplification (MLPA) kit incorporates probes for variants found in both PMS2 and PMS2CL. This provides detection of deletions, but does not allow localization of deletions to the gene or pseudogene. To address this, we have developed a methodology incorporating reference samples with known copy numbers of variants, and paired MLPA results with sequencing of PMS2 and PMS2CL. We tested a subset of clinically indicated samples for which mutations were either unidentified or not fully characterized using existing methods. We identified eight unrelated patients with deletions encompassing exons 9-15, 11-15, 13-15, 14-15, and 15. By incorporating specific, characterized reference samples and sequencing the gene and pseudogene it is possible to identify deletions in this region of PMS2 and provide clinically relevant results. This methodology represents a significant advance in the diagnosis of patients with Lynch syndrome caused by PMS2 mutations.

Objective Recent changes in the epidemiology of C. difficile infection include an increase in the incidence of C. difficile-associated disease (CDAD), and the identification of patients with inflammatory bowel disease (IBD) as a group-at-risk. In addition, effectiveness of antimicrobial therapies has been questioned. Our aim was to estimate the incidence of CDAD in a pediatric IBD population, and review treatment efficacy. Methods We identified patients aged ≤18 years from our center’s IBD database who tested positive for C. difficile toxin A and/or B between 8/1/07 and 12/31/08. Demographic information and treatment details were recorded. Chi-square and Fisher’s exact test were used to compare categorical variables and student’s t-test was used for continuous variables. Results From 372 pediatric IBD patients, we identified 29 patients who experienced a total of 40 cases of CDAD. The annualized incidence rate of CDAD was 7.2%. Initial treatment was successful in 17 cases (43%). Eventual success was documented with metronidazole in 15 cases (41%), with vancomycin in 16 cases (43%), and with other agents or a combination of agents in 6 cases (16%). Age, sex, and IBD type were not associated with initial treatment outcome or recurrence. The choice of initial antimicrobial treatment was not associated with treatment outcome. The type of IBD therapy medication was not associated with likelihood of CDAD recurrence although use of anti-inflammatory therapy was positively associated with initial antimicrobial treatment success. Conclusions CDAD occurred frequently in our cohort of pediatric IBD patients. Antimicrobial treatment success was achieved equally with either metronidazole or vancomycin. Initial treatment failed more than half the time, regardless of medication choice. Apparent lack of antimicrobial efficacy in resolving symptoms may reflect resistant C. difficile infection or increased IBD severity in a subset of patients who are C. difficile carriers. Awareness of the potential for a high incidence of CDAD and frequent failure rate of initial therapy is important in the management of children with IBD.