Structural variants contribute to phenotypic variation in maize.

Comprehensively identifying the loci shaping trait variation has been challenging, in part because standard approaches often miss many types of genetic variants. Structural variants, especially transposable elements are likely to affect phenotypic variation but we need better methods in maize for detecting polymorphic structural variants and TEs using short-read sequencing data. Here, we used a whole genome alignment between two maize genotypes to identify polymorphic structural variants and then genotyped a large maize diversity panel for these variants using short-read sequencing data. We characterized variation of SVs within the panel and identified SV polymorphisms that are associated with life history traits and genotype-by-environment interactions. While most of the SVs associated with traits contained TEs, only one of the SV's boundaries clearly matched TE breakpoints indicative of a TE insertion, whereas the other polymorphisms were likely caused by deletions. All of the SVs associated with traits were in linkage disequilibrium with nearby single nucleotide polymorphisms (SNPs), suggesting that this method did not identify variants that would have been missed in a SNP association study.

Genotype-by-environment interactions and local adaptation shape selection in the US National Chip Processing Trial.

KEY MESSAGE: We find evidence of selection for local adaptation and extensive genotype-by-environment interaction in the potato National Chip Processing Trial (NCPT). We present a novel method for dissecting the interplay between selection, local adaptation and environmental response in plant breeding schemes. Balancing local adaptation and the desire for widely adapted cultivars is challenging for plant breeders and makes genotype-by-environment interactions (GxE) an important target of selection. Selecting for GxE requires plant breeders to evaluate plants across multiple environments. One way breeders have accomplished this is to test advanced materials across many locations. Public potato breeders test advanced breeding material in the National Chip Processing Trial (NCPT), a public-private partnership where breeders from ten institutions submit advanced chip lines to be evaluated in up to ten locations across the country. These clones are genotyped and phenotyped for important agronomic traits. We used these data to interrogate the NCPT for GxE. Further, because breeders submitting clones to the NCPT select in a relatively small geographic range for the first 3 years of selection, we examined these data for evidence of incidental selection for local adaptation, and the alleles underlying it, using an environmental genome-wide association study (envGWAS). We found genomic regions associated with continuous environmental variables and discrete breeding programs, as well as regions of the genome potentially underlying GxE for yield.

Unloading potatoes: Potato breeding moves forward with only half the genome.

Potato is one of the most important food crops in the world and, in contrast to other staples, has not seen large improvements in yield. Agha, Shannon, and Morrell preview an article recently published in Cell, "Phylogenomic discovery of deleterious mutations facilitates hybrid potato breeding," which advances potato breeding strategies via a genetic approach.

A Comparative Study of Single-Layered Versus Double-Layered Intestinal Anastomosis.

BACKGROUND: Intestinal perforations requiring resection anastomosis of the gastrointestinal tract (GIT) or the formation and closure of temporary intestinal stoma are prevalent worldwide. This prospective comparative study was done to assess the efficacy and safety of single-layered anastomosis compared to a double-layered anastomosis. METHODS: Patients undergoing intestinal anastomosis with either of these two techniques were observed prospectively for various outcome parameters like time taken for anastomosis, and that for entire surgery, postoperative complications, etc. Data obtained were analyzed for statistical significance by applying the chi-square test and student's "t-test." RESULTS: Duration for fashioning the anastomosis was significantly lesser for a single layer anastomosis than double (mean [±SD] for single layer was 19.57 ± 2.25 minutes and for double layer group was 30 ± 2.59 minutes, p=0.002). There was no statistical difference in the postoperative complications between the two groups. The postoperative incomplete intestinal obstruction was reported in three cases of the double layer group. CONCLUSION: Single-layered gastrointestinal anastomosis (GIA) resulted in a significant reduction in time, without any difference in complications. Additionally, it is easier to train surgical residents in the single-layered technique which is particularly important in the setting of a teaching institute and can be recommended for intestinal anastomosis.

Rosuvastatin ameliorates cognitive impairment in rats fed with high-salt and cholesterol diet via inhibiting acetylcholinesterase activity and amyloid beta peptide aggregation.

Amyloid beta (Aß) peptide aggregation and cholinergic neurodegeneration are involved in the development of cognitive impairment. Therefore, in this article, we examined rosuvastatin (RSV), an oral hypolipidemic drug, to determine its potential as a dual inhibitor of acetylcholinesterase (AChE) and Aß peptide aggregation for the treatment of cognitive impairment. Molecular docking study was done to examine the affinity of RSV with Aß1-42 and AChE in silico. We also employed neurobehavioral activity tests, biochemical estimation, and histopathology to study the anti-Aß1-42 aggregation capability of RSV in vivo. Molecular docking study provided evidence that RSV has the best binding conformer at its receptor site or active site of an enzyme. The cognitive impairment in female Wistar rats was induced by high-salt and cholesterol diet (HSCD) ad libitum for 8 weeks. RSV ameliorated serum cholesterol level, AChE activity, and Aß1-42 peptide aggregations in HSCD induced cognitive impairment. In addition, RSV-treated rats showed greater scores in the open field (locomotor activity) test. Moreover, the histopathological studies in the hippocampus and cortex of rat brain also supported that RSV markedly reduced the cognitive impairment and preserved the normal histoarchitectural pattern of the hippocampus and cortex. Taken together, these data indicate that RSV may act as a dual inhibitor of AChE and Aß1-42 peptide aggregation, therefore suggesting a therapeutic strategy for cognitive impairment treatment.

The Significance of ERG and Androgen Receptor Expression in Adenocarcinoma Prostate.

Background Fusions of transmembrane protease, serine 2 (TMPRSS2) with erythroblast transformation specific transcription factors have been found in prostate cancer. The v-etserythroblastosis virus E26 oncogene homologue (ERG) is a proto-oncogene of the erythroblast transformation specific transcription factor family. TMPRSS2-ERG fusion is the most common molecular alteration present in about 50% of prostatic adenocarcinomas. Androgen receptor (AR) plays a key role in prostate development and is involved in the progression of prostate cancer. Objective To evaluate the significance of combined ERG and AR expression in cases of prostatic adenocarcinoma. Method The study was conducted at Dr. Ram Manohar Lohia Institute of Medical Sciences, Lucknow, Uttar Pradesh, India. Formalin fixed-paraffin embedded archival prostatic tissue specimens were obtained. A total of 10 cases of prostatic adenocarcinoma were included in the study. Immunohistochemistry for Androgen receptor was done by the standard protocol. Multiplex immunohistochemical staining was done for ERG+CK5 using a primary antibody cocktail of mouse and rabbit antibodies. Result Specific AR immunostaining was exclusively nuclear and was present in all 10 cases in varying intensity. Specific ERG immunostaining was nuclear and was present in seven cases (70%) and absent in three cases (30%). The three cases that were negative for ERG had a Gleason score of ≤ 6 and the AR staining was strong and present in about 90% of the cells. Gleason score was directly related to the ERG staining while AR staining was inversely related to the ERG staining. Conclusion The prognostic value of combined ERG and AR over-expression, its associated genes should be further investigated as potential therapeutic targets in prostate cancer progression. Preliminary data is being presented. Larger prospective studies with survival analysis are essential for prognostic significance.

Genomic characterization of two new enterovirus types, EV-A114 and EV-A121.

Enteroviruses cause a variety of illnesses of the gastrointestinal tract, central nervous system and cardiovascular system. Phylogenetic analysis of VP1 sequences has identified 106 different human enteroviruses classified into four enterovirus species within the genus Enterovirus of the family Picornaviridae. It is likely that not all enterovirus types have been discovered. Between September 2013 and October 2014, stool samples of 6274 apparently healthy children of up to 5 years of age residing in Gorakhpur district, Uttar Pradesh, India were screened for enteroviruses. Virus isolates obtained in RD and Hep-2c cells were identified by complete VP1 sequencing. Enteroviruses were isolated from 3042 samples. A total of 87 different enterovirus types were identified. Two isolates with 71 and 74 % nucleotide sequence similarity to all other known enteroviruses were recognized as novel types. In this paper we report identification and complete genome sequence analysis of these two isolates classified as EV-A114 and EV-A121.

How to identify, assess and utilise mobile medical applications in clinical practice.

BACKGROUND: There are thousands of medical applications for mobile devices targeting use by healthcare professionals. However, several factors related to the structure of the existing market for medical applications create significant barriers preventing practitioners from effectively identifying mobile medical applications for individual professional use. AIMS: To define existing market factors relevant to selection of medical applications and describe a framework to empower clinicians to identify, assess and utilise mobile medical applications in their own practice. MATERIALS AND METHODS: Resources available on the Internet regarding mobile medical applications, guidelines and published research on mobile medical applications. RESULTS: Mobile application stores (e.g. iTunes, Google Play) are not effective means of identifying mobile medical applications. Users of mobile devices that desire to implement mobile medical applications into practice need to carefully assess individual applications prior to utilisation. DISCUSSION: Searching and identifying mobile medical applications requires clinicians to utilise multiple references to determine what application is best for their individual practice methods. This can be done with a cursory exploration of mobile application stores and then moving onto other available resources published in the literature or through Internet resources (e.g. blogs, medical websites, social media). Clinicians must also take steps to ensure that an identified mobile application can be integrated into practice after carefully reviewing it themselves. CONCLUSION: Clinicians seeking to identify mobile medical application for use in their individual practice should use a combination of app stores, published literature, web-based resources, and personal review to ensure safe and appropriate use.

Assessment of fluoride level in groundwater and prevalence of dental fluorosis in Didwana block of Nagaur district, Central Rajasthan, India.

BACKGROUND: In India, for the high concentration of fluoride in groundwater, people are at risk of dental fluorosis. The problem is common in various states of India. The condition in Rajasthan is worse where all districts have such a problem. OBJECTIVE: To study the fluoride concentration in groundwater and prevalence of dental fluorosis in Didwana block of Nagaur district, Central Rajasthan, India. METHODS: The fluoride concentration in water of 54 villages was measured electrochemically, using fluoride ion selective electrode. Dental fluorosis was assessed in 1136 people residing in study area by Dean's classification for dental fluorosis. RESULTS: The fluoride concentration in groundwater in studied sites ranged from 0.5 to 8.5 mg/L. The concentration of fluoride was more than the maximum permissible limit set by WHO and Bureau of Indian Standards (1 mg/L) in 48 groundwater sources. Of 1136 people studied, 788 (69.4%; 95% CI: 66.7%-72.1%) had dental fluoros---252 had mild and 74 had severe dental fluorosis. CONCLUSION: High level of fluoride in drinking water of Didwana block of Nagaur district, Central Rajasthan, India, causes dental fluorosis in most people in the region and is an important health problem that needs prompt attention.

Diabetes mellitus and bladder cancer--an epidemiological relationship?

An epidemiological association between diabetes mellitus and transitional cell carcinoma of the bladder has been proposed. This study looked retrospectively at 125 patients with transitional cell carcinoma of the bladder as a study group and 80 other hospital patients with conditions not specifically associated with diabetes mellitus as a control group. Diabetic patients had an increased, significant odds ratio for bladder cancer compared with non diabetics even after adjustment for smoking and age [OR: 2.69 p=0.049 (95% CI 1.006-7.194)] A history of smoking OR 2.16 p=0.013 (95% C.I. 1.175-3.964) is a significant independent association with transitional cell carcinoma of the bladder as is age: p=0.001 OR 1.07. We propose potential pathogenic pathways for transitional cell carcinoma of the bladder in diabetic patients based on altered integrin and cadherin distribution in urothelial cells in diabetic patients. A larger study is planned to confirm an association between diabetes mellitus and transitional cell carcinoma of the bladder.

Plk3 functionally links DNA damage to cell cycle arrest and apoptosis at least in part via the p53 pathway.

Polo-like kinase 3 (Plk3, previously termed Prk) contributes to regulation of M phase of the cell cycle (Ouyang, B., Pan, H., Lu, L., Li, J., Stambrook, P., Li, B., and Dai, W. (1997) J. Biol. Chem. 272, 28646-28651). Plk3 physically interacts with Cdc25C and phosphorylates this protein phosphatase predominantly on serine 216 (Ouyang, B., Li, W., Pan, H., Meadows, J., Hoffmann, I., and Dai, W. (1999) Oncogene 18, 6029-6036), suggesting that the role of Plk3 in mitosis is mediated, at least in part, through direct regulation of Cdc25C. Here we show that ectopic expression of a kinase-active Plk3 (Plk3-A) induced apoptosis. In response to DNA damage, the kinase activity of Plk3 was rapidly increased in an ATM-dependent manner, whereas that of Plk1 was markedly inhibited. Recombinant Plk3 phosphorylated in vitro a glutathione S-transferase fusion protein containing p53, but not glutathione S-transferase alone. Recombinant Plk1 also phosphorylated p53 but on residues that differed from those targeted by Plk3. Co-immunoprecipitation and pull-down assays demonstrated that Plk3 physically interacted with p53 and that this interaction was enhanced upon DNA damage. In vitro kinase assays followed by immunoblotting showed that serine 20 of p53 was a target of Plk3. Furthermore, expression of a kinase-defective Plk3 mutant (Plk3(K52R)) resulted in significant reduction of p53 phosphorylation on serine 20, which was correlated with a decrease in the expression of p21 and with a concomitant increase in cell proliferation. These results strongly suggest that Plk3 functionally links DNA damage to cell cycle arrest and apoptosis via the p53 pathway.

DNA polymerase beta expression differences in selected human tumors and cell lines.

A long-standing question in cancer biology has been the extent to which DNA repair may be altered during the process of carcinogenesis. We have shown recently that DNA polymerase beta (beta-pol) provides a rate-determining function during in vitro repair of abasic sites by one of the mammalian DNA base excision repair pathways. Therefore, altered expression of beta-pol during carcinogenesis could alter base excision repair and, consequently, be critical to the integrity of the mammalian genome. We examined the expression of beta-pol in several cell lines and human adenocarcinomas using a quantitative immunoblotting method. In cell lines from normal breast or colon, the level of beta-pol was approximately 1 ng/mg cell extract, whereas in all of the breast and colon adenocarcinoma cell lines tested, a higher level of beta-pol was observed. In tissue samples, colon adenocarcinomas had a higher level of beta-pol than adjacent normal mucosa. Breast adenocarcinomas exhibited a wide range of beta-pol expression: one tumor had a much higher level of beta-pol (286 ng/mg cell extract) than adjacent normal breast tissue, whereas another tumor had the same level of beta-pol as adjacent normal tissue. Differences in beta-pol expression level, from normal to elevated, were also observed with prostate adenocarcinomas. All kidney adenocarcinomas tested had a slightly lower beta-pol level than adjacent normal tissue. This study reveals that the base excision repair enzyme DNA polymerase beta is up-regulated in some types of adenocarcinomas and cell lines, but not in others.

An epidemiologic study of cystitis among Muslim women attending an out-patients clinic in Karachi.

Cystitis is considered to be one of the most common infections afflicting women of all age groups. It has been suggested that personal hygiene customs may influence the prevalence of cystitis. In this study, 130 women of child bearing age attending the out-patients clinic were interviewed in order to determine the frequency of cystitis and its associated risk factors including personal hygiene practices. Twenty seven percent of the women reported suffering from cystitis atleast once in the past, comparable to reported studies from the United States but higher than reported from other Muslim countries. Sixty-three percent of women reported their first episode during pregnancy and a significant relationship was observed between parity and cystitis (p value < 0.000). However, personal hygiene practices following micturition or sexual intercourse bore no significant association with cystitis. Cystitis, commonly reported among Pakistani married women, occurs primarily during pregnancy. Preventive measures during pregnancy are suggested.

Extracorporeal shock wave lithotripsy for the rescue of post-endoscopic or surgically retained biliary duct stones: does saline irrigation affect the outcome?

Extracorporeal shock wave lithotripsy (ESWL) has been found valuable in situations where obstructing stones in the common bile or intrahepatic ducts are retained following surgery or attempted endoscopic removal. However, success rates are dependent on the type of ESWL system employed and upon a high frequency rate of repeated treatment sessions. We outline our experience with 23 cases of retained, obstructing bile duct stones, ranging in size from 10 to 40 mm diameter, treated with Dornier HM3 ESWL. In the initial 12 patients in the series, successful stone fragmentation occurred in 83% of cases with a median 1.6 treatment sessions. In the latter 11 cases in the series, patients were treated prone and the stone-bearing biliary duct was irrigated with saline solution during ESWL delivery. With this technique, successful stone break up was achieved in all patients (100%) with a single treatment session. Endoscopic sphincterotomy is, however, a prerequisite for extracorporeal lithotripsy and, despite the high success rates now available with new techniques, we believe the ESWL should continue to be employed in support of primary endoscopic methods of management for obstructing bile duct stones.

Mammalian DNA ligase III: molecular cloning, chromosomal localization, and expression in spermatocytes undergoing meiotic recombination.

Three biochemically distinct DNA ligase activities have been identified in mammalian cell extracts. We have recently purified DNA ligase II and DNA ligase III to near homogeneity from bovine liver and testis tissue, respectively. Amino acid sequencing studies indicated that these enzymes are encoded by the same gene. In the present study, human and murine cDNA clones encoding DNA ligase III were isolated with probes based on the peptide sequences. The human DNA ligase III cDNA encodes a polypeptide of 862 amino acids, whose sequence is more closely related to those of the DNA ligases encoded by poxviruses than to replicative DNA ligases, such as human DNA ligase I. In vitro transcription and translation of the cDNA produced a catalytically active DNA ligase similar in size and substrate specificity to the purified bovine enzyme. The DNA ligase III gene was localized to human chromosome 17, which eliminated this gene as a candidate for the cancer-prone disease Bloom syndrome that is associated with DNA joining abnormalities. DNA ligase III is ubiquitously expressed at low levels, except in the testes, in which the steady-state levels of DNA ligase III mRNA are at least 10-fold higher than those detected in other tissues and cells. Since DNA ligase I mRNA is also present at high levels in the testes, we examined the expression of the DNA ligase genes during spermatogenesis. DNA ligase I mRNA expression correlated with the contribution of proliferating spermatogonia cells to the testes, in agreement with the previously defined role of this enzyme in DNA replication. In contrast, elevated levels of DNA ligase III mRNA were observed in primary spermatocytes undergoing recombination prior to the first meiotic division. Therefore, we suggest that DNA ligase III seals DNA strand breaks that arise during the process of meiotic recombination in germ cells and as a consequence of DNA damage in somatic cells.

Specific inhibition of DNA polymerase beta by its 14 kDa domain: role of single- and double-stranded DNA binding and 5'-phosphate recognition.

DNA polymerase beta (beta-polymerase) has been implicated in short-patch DNA synthesis in the DNA repair pathway known as base excision repair. The native 39 kDa enzyme is organized into four structurally and functionally distinct domains. In an effort to examine this enzyme as a potential therapeutic target, we analyzed the effect of various beta-polymerase domains on the activity of the enzyme in vitro. We show that the 14 kDa N-terminal segment of beta-polymerase, which binds to both single- and double-stranded DNA, but lacks DNA polymerase activity, inhibits beta-polymerase activity in vitro. Most importantly, the 8, 27 and 31 kDa domains of beta-polymerase do not inhibit beta-polymerase activity, demonstrating that the inhibition by the 14 kDa domain is specific. The inhibition of beta-polymerase activity in vitro is abolished by increasing the concentrations of both of the substrates (template-primer and deoxynucleoside triphosphate). In contrast, an in vitro base excision repair assay is inhibited in a domain specific manner by the 14 kDa domain even in the presence of saturating substrates. The inhibition of beta-polymerase activity by the 14 kDa domain appears specific to beta-polymerase as this domain does not inhibit either mammalian DNA polymerase alpha or Escherichia coli polymerase I (Klenow fragment). These data suggest that the 14 kDa domain could be used as a potential inhibitor of intracellular beta-polymerase and that it may provide a means for sensitizing cells to therapeutically relevant DNA damaging agents.

Purification and characterization of DNA ligase III from bovine testes. Homology with DNA ligase II and vaccinia DNA ligase.

Mammalian cell nuclei contain three biochemically distinct DNA ligases. In the present study we have found high levels of DNA ligase I and DNA ligase III activity in bovine testes and have purified DNA ligase III to near homogeneity. The high level of DNA ligase III suggests a role for this enzyme in meiotic recombination. In assays measuring the fidelity of DNA joining, we detected no significant differences between DNA ligases II and III, whereas DNA ligase I was clearly a more faithful enzyme and was particularly sensitive to 3' mismatches. Amino acid sequences of peptides derived from DNA ligase III demonstrated that this enzyme, like DNA ligase II, is highly homologous with vaccinia DNA ligase. The absence of unambiguous differences between homologous peptides from DNA ligases II and III (10 pairs of peptides, 136 identical amino acids) indicates that these enzymes are either derived from a common precursor polypeptide or are encoded from the same gene by alternative splicing. Based on similarities in amino acid sequence and biochemical properties, we suggest that DNA ligases II and III, Drosophila DNA ligase II, and the DNA ligases encoded by the pox viruses constitute a distinct family of DNA ligases that perform specific roles in DNA repair and genetic recombination.

The aetiological and clinical pattern of childhood urolithiasis in Saudi Arabia.

The clinical and aetiological pattern in 85 stone-forming children presenting to an integrated nephrourological service in Riyadh is reviewed. All patients were below the age of 15 years, the male to female ratio being 2:1. Only 2 children presented with bladder calculi. The remaining all had upper tract stones and, in 12 cases, these were bilateral. Of 34 calculi recovered for analysis, one-third was predominantly calcium oxalate and a further third was composed of uric acid or urate. Four patients had cysteine stones and the remaining 7 presented mixed calcium stones, 6 (17.6%) being struvite and infection-related. Of the 85 patients 55 were treated successfully with extracorporeal shock wave lithotripsy, 16 underwent surgery and 7 had their stones removed by endourological procedures. In the remaining 7 children, stones dissolved or were passed spontaneously during medical therapy. Nine children (10.6%) showed a primary metabolic defect leading to their stone formation, 10 (11.8%) had a predisposing anatomical anomaly and 15 (17.6%) presented with urinary tract infection. Of the remaining 51 patients (60%) with idiopathic disease, 6 showed hypercalciuria on investigation and 2 children may have formed their stones due to prolonged recumbency.

Mammalian DNA ligase II is highly homologous with vaccinia DNA ligase. Identification of the DNA ligase II active site for enzyme-adenylate formation.

Mammalian cells contain three biochemically distinct DNA ligases. In this report we describe the purification of DNA ligase II to homogeneity from bovine liver nuclei. This enzyme interacts with ATP to form an enzyme-AMP complex, in which the AMP moiety is covalently linked to a lysine residue. An adenylylated peptide from DNA ligase II contains the sequence, Lys-Tyr-Asp-Gly-Glu-Arg, which is homologous to the active site motif conserved in ATP-dependent DNA ligases. The sequences adjacent to this motif in DNA ligase II are different from the comparable sequences in DNA ligase I, demonstrating that these enzymes are encoded by separate genes. The amino acid sequences of 15 DNA ligase II peptides exhibit striking homology (65% overall identity) with vaccinia DNA ligase. These peptides are also homologous (31% overall identity) with the catalytic domain of mammalian DNA ligase I, indicating that the genes encoding DNA ligases I and II probably evolved from a common ancestral gene. Since vaccinia DNA ligase is not required for DNA replication but influences the ability of the virus to survive DNA damage, the homology between this enzyme and DNA ligase II suggests that DNA ligase II may be involved in DNA repair.

Multisite point-of-care potassium testing for patient-focused care.

In the past two decades, a dominant paradigm has been the main laboratory, which is often located far from the patient and characterized by slow response times. The invention of whole blood biosensors and the innovation of point-of-care testing have initiated a paradigm shift in diagnostic medicine that supports the trend toward patient-focused care. The objective of this study was to compare point-of-care potassium testing performed with a handheld potassium analyzer (STAT K, PDx Technologies Inc, Westlake Village, Calif) in the cardiac and intensive care units with potassium measurements obtained similarly in the main laboratory. Two critical care nurses performed point-of-care testing for critically ill patients. In a series of 56 specimens, the mean +/- SD potassium levels were 3.91 +/- 0.53 and 3.94 +/- 0.57 mmol/L when testing was performed at the bedside and in the main laboratory, respectively. The mean paired difference, -0.03 mmol/L, between point-of-care and main laboratory results was not statistically or clinically significant. Point-of-care potassium testing is accurate and precise, as well as clinically efficient for use in patient-focused care settings.