A cross sectional survey on the effect of COVID-19 related restrictions on undergraduate and postgraduate medical education in Qatar.

BACKGROUND: COVID-19 pandemic has affected all dimensions of day to day life across the world and medical education was no exception. With this study, we aimed to understand the effect of nationwide restrictions on medical education in Qatar, the models of education adopted during this period and perceptions of participants to the same. METHODS: We conducted a cross-sectional study utilizing an online questionnaire distributed via convenience sampling between April-October 2020. Study participants were faculty and trainees in governmental undergraduate and postgraduate medical education institutes. Two sets of questionnaires were designed for each group. They were asked a series of questions to assess pre- and post-COVID pandemic educational practices, their preferred teaching methods, and their familiarity with electronic teaching platforms. Faculty respondents were asked about their perceived barriers to delivery of medical education during the pandemic and their agreement on a 5-point Likert scale on specific elements. Trainees were asked a series of multiple-choice questions to characterize their pre- and post-COVID pandemic educational experiences. Both groups were asked open-ended questions to provide qualitative insights into their answers. Data were analysed using STATA software version 12.0. RESULTS: Majority of trainees (58.5%) responded that the pandemic has adversely affected medical education at both the undergraduate and postgraduate levels. Trainees (58.5%) and faculty (35.7%) reported an increased reliance on e-learning. Trainees preferred face-to-face education, while faculty preferred a combination of models of education delivery (33.5% versus 37.1%, p = 0.38). Although 52.5% of the faculty had no previous experience of delivering education through e-learning modalities, 58.9% however felt confident in using e-learning software. CONCLUSIONS: Faculty and trainees agree that the COVID-19 pandemic has had a significant impact on the provision of medical education and training in Qatar, with an increased dependence on e-learning. As trainee's prefer face-to-face models of education, we may have to consider restructuring of medical curricula in order to ensure that optimum learning is achieved via e-learning, while at the same time enhancing our use, knowledge and understanding of the e -learning methods. Further research is warranted to assess if these changes have influenced objective educational outcomes like graduation rates or board scores.

Uptake of hematite nanoparticles in maize and their role in cell cycle dynamics, PCNA expression and mitigation of cadmium stress.

Cadmium toxicity is considered a major threat to several crops worldwide. Hematite nanoparticles (NPs), due to their small size and large specific surface area, could be applied as an adsorbent for toxic heavy metals in soil. Also, they serve as an efficient nano-fertilizer, promoting Fe availability and biomass production in plants, thus enabling Cd2+ -induced stress tolerance. The phytotoxicity of five different concentrations of hematite NPs, ranging from 500 to 8,000 mg·kg-1 , and Cd2+ concentrations (110 or 130 mg·kg-1 Cd2+ ) alone or combined with 500 mg·kg-1 NPs was evaluated in maize. The changes in fresh weight, element analysis, cell cycle regulation, DNA banding patterns and proliferating cell nuclear antigen (PCNA) expression were used as biomarkers. The results revealed that increased fresh weight and fewest polymorphic DNA bands were detectable after treatment with 500 mg·kg-1 NPs. However, at 8,000 mg·kg-1 NPs, PCNA expression increased significantly, which resulted in cell cycle arrest at the G1/S checkpoint in roots. Significant reductions in fresh weight, altered nutrient profiles and cell cycle perturbations are considered symptoms of Cd2+ toxicity in maize. Conversely, amending 500 mg·kg-1 NPs with 130 mg·kg-1 Cd2+ increased fresh weight, Fe concentration and genomic template stability, while reducing Cd2+ uptake and PCNA1 expression. Overall, 8,000 mg·kg-1 hematite NPs interfered with the cellular homeostatic balance of maize, resulting in a cascade of genotoxic events, leading to growth inhibition. Although 500 mg·kg-1 hematite NPs alleviated Cd2+ -induced DNA damage to a certain extent, their impact on cell cycle progression requires further verification.

Relation between sex hormones and hepatocellular carcinoma.

Males have higher incidence of hepatocellular carcinoma (HCC) than females. Sex hormones may be a risk factor. The aim was to determine the levels of sex hormones in male and female patients with HCC and cirrhosis versus controls and its possible relationship with HCC. This study was conducted on 90 subjects divided into 40 patients with HCC, 30 patients with liver cirrhosis and 20 apparently healthy subjects complete blood picture, liver function tests. Determination of AFP levels and hormonal assay of oestrogen, progesterone, total testosterone, prolactin, FSH and LH were performed on all subjects. Total testosterone levels were significantly decreased in the two patients groups compared with controls. While oestrogen levels were significantly decreased in the HCC group in comparison with other two groups, prolactin levels were significantly decreased in the HCC group compared with the liver cirrhosis group and increased in the liver cirrhosis group when compared to controls. FSH and LH levels were significantly increased in the HCC group when compared to controls. There is no significant correlation between sex hormones assay and both the size of HCC and degree of cirrhosis in both patient groups. It is concluded that there is no strong relation between sex hormones and HCC when the study was carried out on the levels of sex hormones in patients with HCC.

Finite element analysis of the biomechanical effects of PEEK dental implants on the peri-implant bone.

Dental implants are mostly fabricated of titanium. Potential problems associated with these implants are discussed in the literature, for example, overloading of the jawbone during mastication due to the significant difference in the elastic moduli of titanium (110 GPa) and bone (≈1-30 GPa). Therefore poly-ether-ether-ketone (PEEK) could represent an alternative biomaterial (elastic modulus 3-4 GPa). Endolign(®) represents an implantable carbon fiber reinforced (CFR)-PEEK including parallel oriented endless carbon fibers. According to the manufacturer it has an elastic modulus of 150 GPa. PEEK compounds filled with powders show an elastic modulus around 4 GPa. The aim of the present finite element analysis was to point out the differences in the biomechanical behavior of a dental implant of Endolign(®) and a commercial powder-filled PEEK. Titanium served as control. These three materials were used for a platform-switched dental implant-abutment assembly, whereas Type 1 completely consisted of titanium, Type 2 of a powder-filled PEEK and Type 3 of Endolign(®). A force of 100 N was applied vertically and of 30° to the implant axis. All types showed a minimum safety factor regarding the yield strength of cortical bone. However, within the limits of this study the Type 2 implant showed higher stresses within the adjacent cortical bone than Type 1 and Type 3. These implant assemblies showed similar stress distributions. Endless carbon fibers give PEEK a high stability. Further investigations are necessary to evaluate whether there is a distinct amount of endless carbon fibers causing an optimal stress distribution behavior of CFR-PEEK.

Impact of New Chemotherapeutic agents on the outcome of Egyptian patients with Advanced Malignant Pleural Mesothelioma.

UNLABELLED: Patients with malignant pleural mesothelioma (MPM) are well known to have poor response to chemotherapy. Aim of this work was to evaluate the efficacy and safety of new chemotherapeutic agents for the treatment of Egyptian MPM patients. PATIENTS AND METHODS: The first study was a non-randomized, open-label trial. It included 34 eligible patients who were assigned to receive either cisplatin/ pemetrexed or pemetrexed alone if cisplatin was contraindicated for a maximum of 8 cycles. In the second trial, 21 chemo-naïve patients with histologically proven advanced MPM were included. They received cisplatin and raltitrexed for a maximum of 6 cycles. RESULTS: In the first trial, the median age was 43.5 years (range 25-69), partial response (PR) was achieved in 37.5%, stable disease (SD) in 50%. Median time to progression (TTP) and overall survival (OS) were 7 and 14 months respectively. Survival at 1 year was 64.7%. No toxicity was observed in 17.6% of patients, grade 3-4 toxicity was evident in 11.8% (neutropenia), 8.8% (anemia), and 2.9% (vomiting and diarrhea). In the second trial, median age was 46 years (range 19- 71), PR was achieved in 23.2%, one complete remission (CR) was reported. SD was noticed in 61.9%. The median TTP and OS were 6 and 12 months respectively. Survival at 1 year was 51.6%. CONCLUSION: Both cisplatin/pemetrexed and cisplatin/ raltitrexed are effective and safe regimens in the treatment of MPM.

Comparison between colloid preload and crystalloid co-load in cesarean section under spinal anesthesia: a randomized controlled trial.

BACKGROUND: Hypotension is a common problem during spinal anesthesia for cesarean delivery. Intravenous fluid loading is used to correct preoperative dehydration and reduce the incidence and severity of hypotension. Different fluid regimens have been studied but colloid preload and crystalloid co-load have not been compared. METHODS: In this randomized double-blind study, 210 patients scheduled for elective cesarean section under spinal anesthesia were randomly allocated to receive either 6% hydroxyethyl starch 130/0.4 500 mL before spinal anesthesia (colloid preload) or Ringer's acetate solution 1000 mL administered rapidly starting with intrathecal injection (crystalloid co-load). Maternal hypotension (systolic blood pressure <80% of baseline or <90 mmHg) and severe hypotension (systolic blood pressure <80 mmHg) were treated with 5 and 10mg ephedrine boluses, respectively. The primary outcome was the incidence of hypotension. Secondary outcomes included the incidence of severe hypotension, total ephedrine dose, nausea and vomiting and neonatal outcome assessed by Apgar scores and umbilical artery blood gas analysis. RESULTS: Data analysis was performed on 205 patients; 103 in the colloid preload group and 102 in the crystalloid co-load group. There were no significant differences in the incidence of hypotension (52.4% vs. 42.2%; P=0.18) or severe hypotension (15.5% vs. 9.8%; P=0.31) between colloid preload and crystalloid co-load groups, respectively. The median [range] ephedrine dose was 5 [0-45]mg in the colloid preload group and 0 [0-35]mg in the crystalloid co-load group (P=0.065). There were no significant differences in maternal nausea or vomiting or neonatal outcomes between groups. CONCLUSION: The use of 1000 mL crystalloid co-load has similar effect to 500 mL colloid preload in reducing the incidence of hypotension after spinal anesthesia for elective cesarean delivery. Neither technique can totally prevent hypotension and should be combined with vasopressor use.

Silica coating and photocatalytic activities of ZnO nanoparticles: effect of operational parameters and kinetic study.

Silica-coating ZnO nanoparticles were prepared using the hydrothermal method. The prepared nanoparticles were characterized by X-ray diffraction (XRD), transmission electron microscopy (TEM), scanning electron microscopy (SEM) and energy dispersive X-ray Spectroscopy (EDX). It was found that ultrafine core/shell structured silica-coating ZnO nanoparticles were successfully obtained. TEM analysis revealed a continuous and uniform silica coating layer of about 8nm in thickness on the surface of ZnO nanoparticles. The photocatalytic performance of silica-coating ZnO core/shell nanoparticles in methylene blue aqueous solution was investigated. The effects of some operational parameters such as pH value, nanocatalyst loading and initial MB concentration on the degradation efficiency were discussed. Kinetic parameters were experimentally determined and a pseudo-first-order kinetic was observed. Thus, the main advantage of the coating is the stability of the photocatalysts and the better performance in acidic or alkaline solutions. Compared to ZnO the maximum apparent rate constant is obtained at pH 8.5 (pH 11.5 in case of bare ZnO). Moreover, the Langmuir adsorption model was applied to describe the equilibrium isotherm at different MB concentration. The applicability of the Langmuir isotherm suggests monolayer coverage of the MB onto surface of silica-coating ZnO nanoparticles. The kinetics of the adsorption with respect to the initial dye concentration, were also investigated. The pseudo-first-order and second-order kinetic models were used and the rate constants were evaluated. The kinetic studies revealed that the pseudo-second-order kinetic model better represented the adsorption kinetics, suggesting that the adsorption process may be chemisorption.

Evaluation and characterisation of A and B fragments of Corynebacterium diphtheriae toxin towards recombinant diphtheria vaccine.

BACKGROUND: Diphtheria is a highly communicable disease caused by toxin-producing strains of Corynebacterium diphtheriae. OBJECTIVES: To evaluate the efficacy of A and B subunits of diphtheria toxin (DT-A, DT-B) as potential vaccines against C. diphtheriae. A culture of C. diphtheriae (strain PW 8) was grown on Loeffler plates while Lingood medium was used for production of diphtheria toxin (DT). MATERIALS AND METHODS: DT was purified and digested to obtain pure DT-A and DT-B and detoxified to obtain diphtheria toxin. Four groups of mice were immunised with different antigens (Ag) of C. diphtheriae. RESULTS: The antibody (Ab) titres were significantly increased with immunised groups subsequent to three injections. On the other hand, Ab titres were estimated after the three immunisations and the levels of different Ab isotypes were comparatively measured. The levels of various isotypes immune responses showed variation between immunised groups where the IgG subclasses were significantly increased mainly with DPT immunised group. The IgM and IgA were significantly increased with DT-A more than others. Additionally, the evaluation of the cellular immune responses demonstrated that spleen cells from DPT and DT-A groups gave highly significant proliferative response with production of high levels of IL-2 and IFN-γ (Th1/Th2). Separation and purification of DT gene were performed using polymerase chain reaction (PCR) and sub-cloned in pGEM-T vector, for further studying of recombinant vaccine. CONCLUSION: Our results showed the possibility to prepare a potent recombinant vaccine containing whole DT gene or DT-A against C. diphtheriae or could be used in treatment of cancer as it give high levels of IL-2 and IFN-γ.

Effects of a soybean protein diet on ovariectomised female albino rats subjected to myocardial infarction.

INTRODUCTION: Cardiovascular disease is the leading cause of death among menopausal women in developed countries, mostly due to the loss of endogenous oestrogen protection. Soybean protein (SP) is rich in isoflavone phytoestrogens. This study aimed to determine the effect of SP on ovariectomised rats subjected to myocardial infarction and its possible cardio-protection. METHODS: The study was conducted on 30 adult female albino rats, which were divided into three groups: Group I comprised the sham-operated rats; Group II, the ovariectomised (OVX) rats fed a standard diet; and Group III, OVX rats fed a standard diet supplemented with SP (OVX plus SP). The rats were anaesthetised, and electrocardiograms were conducted. The rats were then sacrificed, after which their hearts and livers were removed, weighed and subjected to histopathological examination. Blood was collected to determine the lipid profile, and the levels of total triiodothyronine, tetraiodothyronine (T4), thyroid-stimulating hormone (TSH), creatinine phosphokinase (CPK), lactate dehydrogenase, superoxide dismutase (SOD) and malonedialdehyde (MDA). RESULTS: The biochemical studies showed a significant increase in plasma CPK (Group II), MDA and triacylglycerol (Groups II and III) levels compared to Group I. The plasma SOD showed a significant decrease in Group II compared to Group I. Total cholesterol, low and very low density lipoprotein cholesterol levels showed a significant increase in Group II, and a significant decrease compared to Group I. Significant increases in T4 and TSH were found in Group III compared to Group II. CONCLUSION: SP intake can be valuable in protecting the heart against an attack of acute myocardial infarction.

The use of a quantitative real-time polymerase chain reaction assay for identification and enumeration of Lactobacillus buchneri in silage.

AIMS: To detect and quantify Lactobacillus buchneri in plant samples with the aid of polymerase chain reaction (PCR) methods. METHODS AND RESULTS: DNA from silage samples spiked with different amounts of L. buchneri cells was isolated using a lysozyme/sodium dodecyl sulfate lysis and phenol/chloroform extraction method. The DNA served as a template for PCR amplification with primers specific for the bacterium. The primers were developed by comparison of 16S rDNA sequences from different lactic acid bacteria (LAB) and testing for specificity with 11 different strains of LAB. As few as 100 L. buchneri colony-forming units per gram of silage could be detected. Additionally, the technique was successfully applied to quantify the population of L. buchneri in two cultivars of corn with or without inoculation. CONCLUSIONS: The PCR assay provided a specific and rapid tool for identifying and enumerating L. buchneri in silage samples. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of microbial inoculants for silage production is a safe and environment friendly practice, but the full potential of such additives can only be achieved with a better understanding of the fate and activity of the microbes involved. The current study describes a methodology to detect and enumerate L. buchneri, a micro-organism used as an inoculant.

Prediabetes: a must to recognise disease state.

UNLABELLED: Prediabetes mellitus (PDM) is defined as a state of abnormal glucose homeostasis in which deficiency or resistance to insulin is the hallmark. PDM precedes the development of overt type 2 DM. It is associated with increased mortality and morbidity and thus fits well with the criteria of a disease condition. Framing PDM as a disease and not a risk or a 'pre' stage for diabetes is needed to facilitate early management. AIM: This review aims therefore to increase awareness of PDM as a disease state. METHODS: To do so, we shall preview guidelines for its diagnosis. Its prevalence and hazards will be then discussed. Finally, we shall elaborate on the current treatment guidelines. RESULT: Enough evidence support the notion that PDM is a curable disease state. CONCLUSIONS: The current recommendations for the treatment of PDM should be adhered. In addition, there is a room for the use of other pharmacological agents.

An immunoglobulin E-reactive chimeric human immunoglobulin G1 anti-idiotype inhibits basophil degranulation through cross-linking of FcepsilonRI with FcgammaRIIb.

BACKGROUND: IgE binds to mast cells and basophils via its high-affinity receptor, FcepsilonRI, and cross-linking of FcepsilonRI-bound IgE molecules by allergen leads to the release of allergic mediators characteristic of type I hypersensitivity reactions. Previous work has shown that cross-linking of FcepsilonRI with FcgammaRIIb, an ITIM-containing IgG receptor, leads to inhibition of basophil triggering. 2G10, a chimeric human IgG1 anti-idiotype, has broad reactivity with human IgE and as such has the potential to bind simultaneously to FcepsilonRI-bound IgE, via its Fab regions, and the negative regulatory receptor, FcgammaRIIb, via its Fc region. OBJECTIVE: To assess the ability of human 2G10 to inhibit anti-IgE and allergen-driven basophil degranulation through cross-linking of FcepsilonRI-bound IgE with FcgammaRIIb. METHODS: 2G10 was assessed for its ability to bind to FcgammaRIIb on transfected cells and on purified basophils. In the basophil degranulation assay, basophils were purified from peripheral blood of atopic individuals and activated with either anti-IgE or the house dust mite allergen Der p 1, in the presence or absence of human 2G10. Basophil activation was quantified by analysis of CD63 and CD203c expression on the cell surface, and IL-4 expression intracellularly, using flow cytometery. RESULTS: Human 2G10 was able to bind to FcgammaRIIb on transfected cells and on purified basophils, and induce a dose-dependent inhibition of both anti-IgE and Der p 1-driven degranulation of basophils. CONCLUSION: The inhibition of basophil degranulation by the human IgG1 anti-idiotype 2G10 highlights the therapeutic potential of IgE-reactive IgG antibodies in restoring basophil integrity through recruitment of the inhibitory receptor FcgammaRIIb.

Plasmid-mediated high-level ceftriaxone resistance in a Salmonella enterica serotype typhimurium isolate.

OBJECTIVE: To present the first documented case of acute infectious gastroenteritis caused by high-level ceftriaxone-resistant Salmonella enterica serotype typhimurium in Kuwait. SUBJECT AND METHODS: Isolation from stool specimen and species identification of current enteric pathogen was carried out according to standard methods. Susceptibility to antibiotics was determined by the disc diffusion method on Mueller-Hinton agar. Minimal inhibitory concentrations (MICs) were measured with E-test strips. The production of extended spectrum beta-lactamase (ESBL) was studied by the double disc synergy method and E-test ESBL strips. Plasmid DNA isolation was performed by the rapid alkaline lysis method. Plasmid DNA was transferred by conjugation to a recipient strain of Escherichia coli. RESULTS: The isolate of S. enterica serotype typhimurium was resistant to ceftriaxone (MIC >256 mg/l), cefotaxime and ceftazidime, and produced ESBL. Ceftriaxone and cefotaxime resistance were co-transferred on a 3.2-kb plasmid to the E. coli recipient strain. Loss of the 3.2-kb plasmid from the transconjugant resulted in the co-loss of ceftriaxone and cefotaxime resistance confirming the carriage of ceftriazone resistance on the 3.2-kb plasmid. CONCLUSION: Plasmid-mediated high-level resistance to ceftriaxone and ESBL production in Salmonella serotype typhimurium is an emerging problem among Salmonella that requires closer monitoring of antimicrobial resistance among these bacterial species.

Incidence of bloodstream infections in a speciality hospital in Kuwait: 8-year experience.

OBJECTIVES: To determine the frequency of isolation and antibiotic-susceptibility patterns of clinically significant bacterial pathogens isolated from blood. MATERIALS AND METHODS: The study was conducted over a period of 8 years (1995-2002) at Infectious Diseases Hospital (IDH), Kuwait. Demographic and clinical data were obtained from medical records. 18,535 blood cultures were analyzed. Disk diffusion method was used to perform antibiotic-susceptibility testing. Minimum inhibitory concentrations of 9 antimicrobials were determined using E-test. Double disk (potentiation) test and E-test ESBL strips were used to detect the production of extended-spectrum beta-lactamases (ESBLs). RESULTS: Salmonella spp. and Brucella spp. were predominant blood isolates, and represented 60.6 and 30.0% of all clinically significant episodes of bloodstream infections, respectively. Among the Salmonella, Salmonella enterica serotypes typhi and paratyphi A were most frequently isolated. The percentage of multidrug resistance (MDR) among them varied from 22 to 51%. A high percentage (40%) of MDR S. enterica serotypes typhi and paratyphi A also showed reduced susceptibility to ceftriaxone and ciprofloxacin. CONCLUSION: During the study period, Salmonella spp. and Brucella spp. were predominant blood isolates. MDR S. enterica serotypes typhi and paratyphi A, with reduced susceptibility to ceftriaxone and ciprofloxacin, are among the most frequent causes of bloodstream infections in IDH, suggesting the need to monitor their susceptibility.

Etiology and antibiotic susceptibility patterns of community-acquired urinary tract infections in a Kuwait hospital.

OBJECTIVE: The aim of this study was to determine the distribution and antibiotic susceptibility patterns of bacterial strains isolated from patients with community-acquired urinary tract infections (UTIs) at the Infectious Diseases Hospital, Kuwait. MATERIALS AND METHODS: The study was conducted over a 7-year period. Patient information was obtained from medical record files. Antibiotic-sensitivity testing was performed by disk diffusion. E test and double disk diffusion methods were used to study the production of extended spectrum beta-lactamases. RESULTS: Of the 14,042 urine samples processed, significant bacteriuria (>10(5) cfu/ml) was detected in 1,606 (11.4%). The majority (74.5%) of the isolates were from women while the remaining 25.5% were from men. The majority of infections (75%) were due to Enterobacteriaceae, coagulase-negative staphylococci (10.3%) and group B streptococci (8.7%). Among the gram-negative enteric bacilli high prevalence of resistance to ampicillin, amoxicillin/clavulanic acid, cephalothin, and trimethoprim/sulfamethoxazole was observed. Increasing resistance to ciprofloxacin and gentamicin was observed in E. coli isolates over the 7 years. Multiple resistance was detected in 53.8 and 41% of E. coli and Klebsiella spp. strains, respectively. No glycopeptide-resistant enterococci were isolated. CONCLUSION: This study revealed that Enterobacteriaceae were the predominant bacterial pathogen of community-acquired UTIs in Infectious Diseases Hospital, Kuwait. It also demonstrated an increasing resistance to ciprofloxacin, gentamicin and the production of extended spectrum beta-lactamase among UTI pathogens in the community.

Seroepidemiological and microbiological study of brucellosis in Kuwait.

OBJECTIVE: The primary objective of the study was to determine the prevalence of brucellosis and the antimicrobial susceptibility pattern of local Brucellae isolates in the Infectious Diseases Hospital, Kuwait. SUBJECTS AND METHODS: A single serum sample was collected from each of 1,836 patients of different nationalities from January 2000 to December 2001. Any patient with a provisional diagnosis of fever or brucellosis had a standard tube agglutination (STA) test for the quantitation of Brucella antibodies. Blood cultures were done in 166 of 455 patients with significant STA titers, using the Bactec system. Antimicrobial susceptibility testing of 123 isolates of Brucella spp. was done against 8 antimicrobial agents. RESULTS: A total of 455 serum samples (24.8%) having an STA titer of > or =1:160 were presumptively diagnosed as cases of brucellosis. The peak isolation was in April and May. Brucella spp. were isolated from 123 blood cultures (74.1%). The blood culture isolation rate was significantly higher in patients with an STA titer of >/=1:1,280 than in those with an STA titer of

A double-screening method to identify reliable candidate non-synonymous SNPs from chicken EST data.

Discovery of non-synonymous single nucleotide polymorphisms (nsSNP), which cause amino acid substitutions, is important because they are more likely to alter protein function than synonymous SNPs (sSNP) or those SNPs that do not result in amino acid changes. By changing the coding sequences, nsSNP may play a role in heritable differences between individual organisms. In the chicken and many other vertebrates, the main obstacle for identifying nsSNP is that there is insufficient protein and mRNA sequence information for self-species referencing and thus, determination of the correct reading frame for expressed sequence tags (ESTs) is difficult. Therefore, in order to estimate the correct reading frame at nsSNP in chicken ESTs, a double-screening approach was designed using self- or cross-species protein referencing, in addition to the ESTScan coding region estimation programme. Starting with 23 427 chicken ESTs, 1210 potential SNPs were discovered using a phred/phrap/polyphred/consed pipeline process and among these, 108 candidate nsSNP were identified with the double screening method. A searchable SNP database (chicksnps) for the candidate chicken SNPs, including both nsSNPs and sSNPs is available at http://chicksnps.afs.udel.edu. The chicken SNP data described in this paper have been submitted to the data base SNP under National Center for Biotechnology Information assay ID ss4387050-ss4388259.

Genetic markers and their application in poultry breeding.

The current chicken genetic map contains at least 1,965 loci within 50 linkage groups, and it covers about 4,000 cM. About 235 of these loci have homology with known human or mammalian genes. The remaining loci are anonymous molecular DNA markers, including microsatellites, amplified fragment length polymorphism (AFLP), randomly amplified polymorphic DNA (RAPD), CR1 elements, and others. A third generation genetic map for human uses single nucleotide polymorphisms (SNP), which have allowed the mapping of complex traits by linkage disequilibrium. One advantage of SNP is that they are usually linked to the gene of interest, and association of the SNP with traits of economic importance can be analyzed using candidate gene approaches. With the tremendous advancements in characterizing chicken expressed sequence tags (EST), the identification of genetic polymorphisms such as SNP in chicken genes has become a reality. Our laboratory has undertaken an in silico analysis of the chicken EST at the University of Delaware by using a Phred/Phrap/Polyphred/Consed pipeline to identify candidate chicken SNP. Initial scanning of 23,427 chicken EST identified a total of 1,209 candidate SNP, with at least 182 non-synonymous SNP that result in an amino acid change observed. Validation of these candidate chicken SNP is ongoing. Placement of the SNP on the chicken genetic map will enhance marker density, thus allowing for mapping of complex traits through linkage analysis and linkage disequilibrium. Application of SNP to identify disease resistance genes in chickens is of special interest to our laboratory, especially in regards to Marek's disease and coccidiosis.

Mapping quantitative trait loci associated with resistance to coccidiosis and growth.

To map QTL associated with disease resistance to avian coccidiosis and growth, two commercial broiler lines with different degrees of resistance to the disease were crossed to generate an F1 generation that was intercrossed to produce 314 F2 generation offspring. The F2 offspring were inoculated with sporulated oocysts of Eimeria maxima. Five disease-associated phenotypes were measured after the infection. Intertrait comparisons revealed that oocyst shedding was a good parameter for evaluating disease resistance or susceptibility. One hundred nineteen microsatellite markers, covering 80% of the chicken genome with an average marker interval of 25 cM, were used for genotyping of F1 parents and F2 offspring. Statistical analysis based on the data of four families revealed a locus on chromosome 1 associated with oocyst shedding (logarithm of odds = 3.46). The genetic mechanism of this locus appeared additive. The genomic scan also identified three potential growth QTL on chromosomes 1, 6, and 8. These results provide the foundation for further investigation to validate the QTL.

Cell proteins TIA-1 and TIAR interact with the 3' stem-loop of the West Nile virus complementary minus-strand RNA and facilitate virus replication.

It was reported previously that four baby hamster kidney (BHK) proteins with molecular masses of 108, 60, 50, and 42 kDa bind specifically to the 3'-terminal stem-loop of the West Nile virus minus-stand RNA [WNV 3'(-) SL RNA] (P. Y. Shi, W. Li, and M. A. Brinton, J. Virol. 70:6278-6287, 1996). In this study, p42 was purified using an RNA affinity column and identified as TIAR by peptide sequencing. A 42-kDa UV-cross-linked viral RNA-cell protein complex formed in BHK cytoplasmic extracts incubated with the WNV 3'(-) SL RNA was immunoprecipitated by anti-TIAR antibody. Both TIAR and the closely related protein TIA-1 are members of the RNA recognition motif (RRM) family of RNA binding proteins. TIA-1 also binds to the WNV 3'(-) SL RNA. The specificity of these viral RNA-cell protein interactions was demonstrated using recombinant proteins in competition gel mobility shift assays. The binding site for the WNV 3'(-) SL RNA was mapped to RRM2 on both TIAR and TIA-1. However, the dissociation constant (K(d)) for the interaction between TIAR RRM2 and the WNV 3'(-) SL RNA was 1.5 x 10(-8), while that for TIA-1 RRM2 was 1.12 x 10(-7). WNV growth was less efficient in murine TIAR knockout cell lines than in control cells. This effect was not observed for two other types of RNA viruses or two types of DNA viruses. Reconstitution of the TIAR knockout cells with TIAR increased the efficiency of WNV growth, but neither the level of TIAR nor WNV replication was as high as in control cells. These data suggest a functional role for TIAR and possibly also for TIA-1 during WNV replication.