July 2012

Editor:
Deborah Sesok-Pizzini, MD, MBA

• Using a clinical rule and platelet count to predict esophageal varices in children
• Using melting curve analysis on a PCR instrument to diagnose fragile X syndrome
• Diagnosis of invasive fungal disease using serum (1↓3)-β-D-glucan

Using a clinical rule and platelet count to predict esophageal varices in children

Children with chronic liver disease or portal vein obstruction may be at risk of developing esophageal varices and bleeding, which may lead to morbidity and mortality. Unlike for adults, esophagogastroduodenoscopy (EGD) is more selectively performed for children at risk for variceal bleeding because it is considered too invasive, time consuming, and expensive and has potential procedural risks. In addition, prophylactic therapy, treatment with nonselective β-blockade, and endoscopic variceal ligation have not been proven to benefit children. However, some clinicians will provide prophylactic therapy, so selecting which children should undergo EGD screening is of utmost importance. One approach to this is to develop noninvasive methods for identifying and monitoring esophageal varices in children. In a previous retrospective study, Gana and colleagues described a noninvasive test with good diagnostic accuracy for identifying children with esophageal varices. The authors used a ratio of the platelet count and spleen size z-score (P/SSAZ) and a clinical prediction rule (calculated from platelet count, spleen size z-score, and albumin concentration) to predict esophageal varices. The study discussed herein validated those measurements for predicting esophageal varices in children. The prospective study spanned eight centers that treated children younger than 18 years who had chronic liver disease or portal vein thrombosis between 2007 and 2010. Of 108 patients enrolled in the study, 69 percent had esophageal varices. In receiver operating characteristic curve analysis, the best predictors of esophageal varices of any size were P/SSAZ, clinical prediction rule (CPR), and platelet count. The authors showed that the best predictors for larger esophageal varices were platelet count followed by CPR. In summary, Gana and colleagues showed that when compared with EGD, CPR and platelet count are a good noninvasive test for esophageal varices and may be used to select children to undergo EGD to confirm and grade varices.

Gana JC, Turner D, Mieli-Vergani G, et al. A clinical prediction rule and platelet count predict esophageal varices in children. Gastroenterol. 2011;141(6):2009–2016.

Correspondence: Dr. Juan Cristobal Gana at jcgana@gmail.com
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Using melting curve analysis on a PCR instrument to diagnose fragile X syndrome

Fragile X syndrome is the most common inherited form of mental retardation and the second most frequent cause of mental retardation after Down syndrome. The incidence is approximately one in 4,000 for males and one in 6,000 for females. Cytosine-guanine-guanine (CGG) repeat expansions in the FMR1 gene are associated with fragile X syndrome (FXS), fragile X-associated tremor/ataxia syndrome, and fragile X-associated primary ovarian insufficiency. The mutation usually affects more males than females since the syndrome is linked to the X chromosome. Individuals with FSX may experience intellectual dysfunction, language delay, behavioral and social problems, and other physical malformations. In some patients, however, the presentation of FXS may be more subtle and the diagnosis more difficult. The clinical diagnosis of FXS is dependent on the molecular detection of FMR1 CGG repeat expansion mutations, which are most often full-mutation CGG repeat expansions in the 5´ untranslated region of the FMR1 gene. Pregnant women at risk for transmitting FXS to their offspring are screened by conventional polymerase chain reaction–based assays and Southern blot analysis. The authors conducted a study to describe and evaluate the feasibility of using melting curve analysis (MCA) on a real-time PCR instrument to diagnose FXS. Teo and colleagues described the current gold standard screening method for FMR1 expansion as labor intensive, time consuming, and requiring large amounts of DNA. The advantages of MCA are that it is a cost-effective, automated, and high-throughput screening assay that may be used to detect FMR1 expansions in males and females. The authors optimized the assay, validated it with known FMR1 reference samples, and performed a blinded validation study. Their results showed that 5’ dTP-PCR assay-generated MCA peak profiles that were different for normals compared with expanded alleles. In addition, the normal samples from males and females showed distinctive melt profiles compared to samples with permutation (55 to 200 repeats) and full mutation (more than 200 repeats). Despite improvements in FMR1 testing that have reduced the need for Southern blot analysis, a simpler strategy is required for high-throughput screening. Combining 5’ and 3’ dTP-PCR assays with MCA fulfills the requirements for a single-step, closed-tube, homogenous assay for rapid, large-scale, high-throughput, and high diagnostic-sensitivity screening for males and females. The authors suggested that in a population-based screening program, this assay should be followed by a more precise diagnostic test for samples testing positive for allele expansion.

Teo CRL, Law HY, Lee CG, et al. Fragile X mental retardation 1 gene CGG repeat expansion screening by melting curve analysis of combined 5´ and 3´ direct triplet-primed PCRs. Clin Chem. 2012;58:3:1–12.

Correspondence: Samuel S. Chong at paecs@nus.edu.sg
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Diagnosis of invasive fungal disease using serum (1↓3)-β-D-glucan

Early detection of invasive fungal disease is critical to patient care, especially in the immunosuppressed patient. The most recently reported invasive fungal diseases (IFDs) are invasive aspergillosis, invasive candidiasis, cryptococcosis, endemic mycoses, and zygomycosis. The high morbidity and mortality rates associated with IFDs are often due to delays in diagnosis. Because clinical signs and symptoms may be nonspecific, and radiological findings delayed and cultures slow to grow, a timely noninvasive test may greatly improve the early diagnosis of IFDs. To this end, the authors studied the (1↑3)-β-D-glucan (BG) assay. BG is a common cell wall constituent of most pathogenic fungi, and the BG assay is approved for diagnosing invasive fungal disease. However, the assay’s performance results are variable. The authors conducted a study to determine the overall accuracy of the BG assay for diagnosing IFDs. They analyzed the sensitivity, specificity, and positive and negative likelihood ratios (PLRs and NLRs) of BG for diagnosing IFDs using a bivariate meta-analysis. They also performed a subgroup analysis. The authors reviewed 12 reports, including 15 studies, with proven and probable IFD versus possible or no IFD. The sensitivity, specificity, PLR, and NLR were 0.76, 0.85, 5.05, and 0.28, respectively. The BG assay had higher specificities for patients with hematological disorders and a positive BG result with two consecutive samples. In addition, the combination of galactomannan and BG increased the specificity value to 0.98 for invasive aspergillosis. The authors concluded that the BG assay was clinically useful for diagnosing IFD in at-risk patients, especially patients with hematological disorders. However, the diagnostic performance of the BG assay was markedly decreased for solid-organ transplant recipients. The authors suggested that variations in some studies may occur due to differences in cutoff values as well as serum values measured with different kits. In summary, although useful for diagnosing IFDs, BG assay results should be interpreted in the context of clinical findings.

Lu Y, Chen YQ, Guo YL, et al. Diagnosis of invasive fungal disease using serum (1↑3)-β-D-glucan: a bivariate meta-analysis. Intern Med. 2011;50:2783–2791.

Correspondence: Dr. Ke Wang at walk7632@yahoo.com.cn
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