Molecular characterization of the Yp11.2 region deletion in the Chinese Han population

The Y chromosome is male-specific and is important for spermatogenesis and male fertility. However, the Y chromosome is poorly characterized due to massive palindromes and inverted repeats, which increase the likelihood of genomic rearrangements, resulting in short tandem repeats on the Y chromosome or long fragment deletions. The present study reports a large-scale (2.573~2.648 Mb) deletion in the Yp11.2 region in a Chinese population based on the analysis of 34 selected Y-specific sequence-tagged sites and subsequent sequencing of the breakpoint junctions on the Y chromosome from 5,068,482–5,142,391 bp to 7,715,462–7,716,695 bp. The results of sequence analysis indicated that the deleted region included part or all of the following five genes: PCDH11Y, TSPY, AMELY, TBL1Y, and RKY. These genes are associated with spermatogenesis or amelogenesis and various other processes; however, specific physiological functions and molecular mechanisms of these genes remain unclear. Notably, individuals with this deletion pattern did not have an obvious pathological phenotype but manifested some degree of amelogenesis imperfecta.

Isolation and Identification of Subgroup J Avian Leukosis Virus Inducing Multiple Systemic Tumors in Parental Meat-Type Chickens

Avian leukosis virus (ALV) continues evolving to obtain new genomic characters to enhance its pathogenicity. In the present study, an ALV-J strain LH20180301 was isolated from broiler breeder chickens that reached the speak of paralyzation before 20-week-old. The necropsy chickens showed subcutaneous and muscular hemorrhage, and developed tumors in multiple organs including bone, liver, spleen, and kidney. The complete provirus was then cloned and sequenced to investigate the molecular characteristics and oncogenicity etiology of this virus associated with the outbreak of disease. The genomic structure of the reported ALV-J strain LH20180301 was highly conservative with other ALVs. Recombination events between the virus with endogenous virus were identified in the viral genome. Compared with the ALV-J original HPRS-103 strain, the major recombination sites of the viral genome with ev-1 were located in 5′ UTR-gag and 3′ UTR regions. Phylogenetic analysis of group specific antigen gp85 encoding protein showed that the LH20180301 branched with ALV-J prevalent in “yellow chickens” of local breeds in South China. Nine amino acids (N58, D60, K70, A71, K108, N112, N113, N121, R272) in the gp85 were highly conserved among ALV-J isolates before 2012, but various mutations were found in the late isolates including LH20180301. In addition, the LH20180301 strain also had the same deletion pattern of 3′ UTR with them. Therefore, LH20180301 might derive from the same ancestor with those viruses and may be the trend of ALV-J evolution in China. The defined new genomic characters in the gp85 and 3′ UTR region of ALV-J might provide the molecular basis for its enhanced oncogenicity.

Full genome sequence analysis of a 1-7-4-like PRRSV strain in Fujian Province, China

PRRS virus (PRRSV) has undergone rapid evolution and resulted in immense economic losses worldwide. In the present study, a PRRSV strain named FJ0908 causing high abortion rate (25%) and mortality (40%) was detected in a swine herd in China. To determine if a new PRRSV genotype had emerged, we characterized the genetic characteristics of FJ0908. Phylogenetic analysis indicated that FJ0908 was related to 1-7-4-like strains circulating in the United States since 2014. Furthermore, the ORF5 sequence restriction fragment length polymorphism (RFLP) pattern of FJ0908 was 1-7-4. Additionally, FJ0908 had a 100 aa deletion (aa329–428) within nsp2, as compared to VR-2332, and the deletion pattern was consistent with most of 1-7-4 PRRSVs. Collectively, the data of this study contribute to the understanding of 1-7-4-like PRRSV molecular epidemiology in China.

Development of an Evidence-Based Algorithm to Guide IGH-Based Reflex Testing for Plasma Cell Neoplasms

FISH testing on bone marrow aspirates is routinely used in the genetic workup for patients with plasma cell neoplasms. However, characteristic low cellularity and low in vitro proliferation rates may limit the number of plasma cells available to comprehensively test recurrent and prognostically significant genetic markers. Plasma cell selection via CD138+ cell isolation or cytoplasmic Ig staining is often required to improve sensitivity, and still may yield insufficient or poor quality cells, and possibly false negative results. Our current Multiple Myeloma panel (MMP) is designed as a reflex test, where positive IGH break-apart (IGH-BAP) and negative t(11;14)/IGH-CCND1 fusion patterns direct subsequent testing for t(4;14)/IGH-FGFR3 and t(14;16)/IGH-MAF fusions. Included in the first tier panel are probes for common trisomies 9 and 15, 1q/CKS1B gain, and 17p/TP53 deletion. This strategy is cost-effective by prioritizing testing for the most common IGH translocation (11;14) first, but can be problematic with low cellularity and/or low percentage patterns. To improve this process, we retrospectively reviewed data from 294-reflexed MMPs encountered in our clinical laboratory in order to develop an evidence-based algorithm for reflex testing. IGH-BAP-positive (IGH-BAP+) signal patterns included RGF (rearrangement), RF (5' deletion), and GF (3' deletion). Positive results for t(4;14) or t(14;16) dual fusion (DF) probes were observed in 75 (25.5%) and 40 (13.9%) samples, respectively. The median differential positivity rate for BAP versus DF probes was 8.5% (range: 0-39.5%), demonstrating similar detection rates between these probe sets (r=0.94, Pearson correlation). Reflex testing was performed on 263 (89.5%) CD138+ enriched cell pellets. IGH-BAP positivity (IGH-BAP+) rates ranged from 3.5-98.0% in this cohort, where higher rates correlated with positive t(4;14) and t(14;16) results (median DF+ =78.5%; median DF- =50.8%; p=2.97E-9, t-Test). Of 68 CD138+ samples with low percentage IGH-BAP+ (≤30%), only seven (10.1%) cases were DF+, six of which had additional abnormal MMP signal patterns at a similar percentage to IGH-BAP, suggestive of a low-level plasma cell population in these samples. The single positive case showed the typical GRF pattern for IGH-BAP (12.0%; DF=15.0%). Of 61 DF- cases, IGH-BAP+ was the sole finding in 23 (37.7%) cases, 16 (26.2%) cases had additional patterns at a similar percentage to IGH-BAP, and 22 (36.0%) cases showed additional signal patterns at much higher percentages, >2.5 times that of IGH-BAP. These findings suggest reflex testing has limited value for CD138+ samples with low IGH-BAP+ rates, especially with IGH-BAP as the sole abnormality, as well as for reflex cases with additional patterns at a much higher percentage, as t(4;14) and t(14;16) should represent primary alterations in these samples. Across all cases in this dataset, the IGH-BAP RF or GF (partial deletion) pattern was observed in 10/110 (9.0%) DF+ cases compared to 75/178 (42.1%) DF- cases, indicating these patterns are less likely to represent true IGH rearrangements. The 5' IGH probe may be deleted due to normal VDJ recombination. Of the four 5' IGH deletion cases that were DF+, all cases showed a complex partial deletion pattern (e.g. 2R2F, 2RF, etc) whereas 44 (81.5%, n=54) DF- cases showed a simple RF pattern, suggesting reflex may have limited utility for the simple RF pattern. Of 31 unsorted samples, 19 (61.3%) were DF+, 14 of which (73.7%) had IGH-BAP+ ≤30%. There was no difference between median IGH-BAP+ percentages in this cohort (DF+=14.0%, DF-=17.5%; range=4.5-77.0%; p=0.236, t-Test). Most DF+ cases (89.5%, n=17) had other abnormal MMP signal patterns in addition to IGH-BAP; two cases showed IGH-BAP+ as a sole finding, both with GRF patterns. These findings indicate reflex testing should be performed for all unsorted samples with IGH-BAP pattern above established cut-offs unless additional MMP signal patterns are present at much higher percentage. We present an algorithm for PCN sample processing that incorporates sample type, signal patterns, and relative detection rates for IGH and other MMP probes in order to guide appropriate IGH reflex testing in the clinical laboratory setting. This method predicts a reduction of greater than 10-15 percent of reflexed cases based on retrospective analysis. Prospective results from implementation will also be presented. Disclosures No relevant conflicts of interest to declare.

Duchenne muscular dystrophy: A immunohistochemical profile and deletion pattern in dystrophin gene in North Indian population

Background: Duchenne muscular dystrophy (DMD), one of the most common X linked muscular disorder, affecting 1 in 3500 male births and is caused by mutation in dystrophin gene. 65% of DMD cases are caused by large deletion of dystrophin gene, followed by duplication (5-10%) or point mutation (25-30%). There is wide mutation spectrum of the mutations in dystrophin gene. Hence, population specific information is needed on mutation spectrum and frequency of common mutations occurring in that particular population for appropriate counseling, prenatal diagnosis and for developing genetic therapy in future. Aims and Objectives: To find out the frequency and distribution of deletion in dystrophin gene in DMD patients along with contribution of pathology and genetic testing in diagnosis of DMD and Becker muscular dystrophy (BMD) in North Indian population.Materials and Methods: Dystrophin gene was screened for deletion by multiplex polymerase chain reaction (PCR). Out of 41 patients, 09 patients underwent muscle biopsy, on which immunohistochemistry was performed for dystrophin, sarcoglycan, dysferlin and merosin.Results: Majority of the deletions were located in distal hotspot region (26/39 ~66.66%) which includes the exons 45-55 and 15.38% of deletions were located at the proximal hotspot region (2- 19 exons).Conclusion: In the present study, 34% patients only showed deletion. Hence complete work up of any muscular dystrophy requires immnohistochemical analysis to see the expression of muscle proteins along with multipleplex PCR test to detect any exon deletion, multiplex ligation-dependent probe amplification (MLPA) to detect point mutation and duplication and western blotting to quantify the dystrophin protein.Asian Journal of Medical Sciences Vol.8(6) 2017 13-18

DEVIASI FONOLOGIS TUTURAN BAHASA INDONESIA ANAK

This article explains the phonological deviations in Indonesian utterancesof children with the age range from 1.0 to 2.6, which represents a stage of languagedevelopment with the children of the age range from 1.0 to 1.6 producing their firstsingle-word utterances (SWU), those of the age range from 1.6 to 2.0 producingtheir first word combinations (FWC), and those of the age range from 2.0 to 2.6producing their first simple and complex sentences (SCS). The data show that 11phonological deviation patterns are found in SWU, 13 phonological deviationpatterns are found in FWC, and 7 phonological deviation patterns are found inSCS. On the basis of these available patterns, it could be stated that (1) the mostproductive patterns are stopping, gliding, nasalizing, and cluster reduction, (2) thedeviation patterns tend to decrease as age increases, (3) as children get older, thereduplication pattern is abandoned and the deletion pattern decreases, and (4) alateralization pattern occurs when children begin producing SCS. Most of thepatterns are universal since they also occur in the utterances of subjects of otherresearchers.Keywords : patterns of phonological deviations, children utterances, underlyingrepresentation, phonetic representation, SWU, FWC, SCS

DEVIASI FONOLOGIS TUTURAN BAHASA INDONESIA ANAK

This article explains the phonological deviations in Indonesian utterancesof children with the age range from 1.0 to 2.6, which represents a stage of languagedevelopment with the children of the age range from 1.0 to 1.6 producing their firstsingle-word utterances (SWU), those of the age range from 1.6 to 2.0 producingtheir first word combinations (FWC), and those of the age range from 2.0 to 2.6producing their first simple and complex sentences (SCS). The data show that 11phonological deviation patterns are found in SWU, 13 phonological deviationpatterns are found in FWC, and 7 phonological deviation patterns are found inSCS. On the basis of these available patterns, it could be stated that (1) the mostproductive patterns are stopping, gliding, nasalizing, and cluster reduction, (2) thedeviation patterns tend to decrease as age increases, (3) as children get older, thereduplication pattern is abandoned and the deletion pattern decreases, and (4) alateralization pattern occurs when children begin producing SCS. Most of thepatterns are universal since they also occur in the utterances of subjects of otherresearchers.Keywords : patterns of phonological deviations, children utterances, underlyingrepresentation, phonetic representation, SWU, FWC, SCS

Deletion pattern in the dystrophin gene in Duchenne muscular dystrophy patients in northeast India

ABSTRACTDuchenne muscular dystrophy (DMD) is an X‑linked recessive disorder that affects 1 in 3,500 males and is caused by mutations in the dystrophin gene. In this paper, we have reported DNA analysis of DMD patients by multiplex polymerase chain reaction (PCR) from various states of northeast India. Of the 69 clinically suspected patients of DMD, deletion was detected by multiplex PCR in 49 (71%) patients. Majority of the deletions (42/49, 85.7%) were located at distal hot spot region that encompasses exons 44-55 and 14.3% of the deletions were located at the proximal hot spot region (exons 2-19). In this study population, the deletion rate was 71% and was more frequent in the distal end exon.