Current Progress and Future Prospect of Wheat Genetics Research towards an Enhanced Nitrogen Use Efficiency.

To improve the yield and quality of wheat is of great importance for food security worldwide. One of the most effective and significant approaches to achieve this goal is to enhance the nitrogen use efficiency (NUE) in wheat. In this review, a comprehensive understanding of the factors involved in the process of the wheat nitrogen uptake, assimilation and remobilization of nitrogen in wheat were introduced. An appropriate definition of NUE is vital prior to its precise evaluation for the following gene identification and breeding process. Apart from grain yield (GY) and grain protein content (GPC), the commonly recognized major indicators of NUE, grain protein deviation (GPD) could also be considered as a potential trait for NUE evaluation. As a complex quantitative trait, NUE is affected by transporter proteins, kinases, transcription factors (TFs) and micro RNAs (miRNAs), which participate in the nitrogen uptake process, as well as key enzymes, circadian regulators, cross-talks between carbon metabolism, which are associated with nitrogen assimilation and remobilization. A series of quantitative genetic loci (QTLs) and linking markers were compiled in the hope to help discover more efficient and useful genetic resources for breeding program. For future NUE improvement, an exploration for other criteria during selection process that incorporates morphological, physiological and biochemical traits is needed. Applying new technologies from phenomics will allow high-throughput NUE phenotyping and accelerate the breeding process. A combination of multi-omics techniques and the previously verified QTLs and molecular markers will facilitate the NUE QTL-mapping and novel gene identification.

Hepatitis C prevalence and elimination planning in Pakistan, a bottom-up approach accounting for provincial variation.

In Pakistan, substantial changes to hepatitis C virus (HCV) programming and treatment have occurred since the 2008 nationwide serosurvey estimated a 4.8% anti-HCV prevalence. In the absence of an updated national study, this analysis uses provincial data to estimate a national prevalence and the interventions needed to achieve elimination. Using a Delphi process, epidemiologic HCV data for the four provinces of Pakistan (accounting for 97% of the population) were reviewed with 21 subject-matter experts in Pakistan. Province-level estimates were inputted into a mathematical model to estimate the national HCV disease burden in the absence of intervention (Base), and if the World Health Organization (WHO) elimination targets are achieved by 2030 (80% reduction in new infections, 90% diagnosis coverage, 80% treatment coverage, and 65% reduction in mortality: WHO Elimination). An estimated 9,746,000 (7,573,000-10,006,000) Pakistanis were living with viraemic HCV as of January 1, 2021; a viraemic prevalence of 4.3% (3.3-4.4). WHO Elimination would require an annual average of 18.8 million screens, 1.1 million treatments, and 46,700 new infections prevented anually between 2022 and 2030. Elimination would reduce total infections by 7,045,000, save 152,000 lives and prevent 104,000 incident cases of hepatocellular carcinoma from 2015 to 2030. Blood surveys, programmatic data, and expert panel input uncovered more HCV infections and lower treatment numbers in the provinces than estimated using national extrapolations, demonstrating the benefits of a bottom-up approach. Screening and treatment must increase 20 times and 5 times, respectively, to curb the HCV epidemic in Pakistan and achieve elimination by 2030.

Yield-Related QTL Clusters and the Potential Candidate Genes in Two Wheat DH Populations.

In the present study, four large-scale field trials using two doubled haploid wheat populations were conducted in different environments for two years. Grain protein content (GPC) and 21 other yield-related traits were investigated. A total of 227 QTL were mapped on 18 chromosomes, which formed 35 QTL clusters. The potential candidate genes underlying the QTL clusters were suggested. Furthermore, adding to the significant correlations between yield and its related traits, correlation variations were clearly shown within the QTL clusters. The QTL clusters with consistently positive correlations were suggested to be directly utilized in wheat breeding, including 1B.2, 2A.2, 2B (4.9-16.5 Mb), 2B.3, 3B (68.9-214.5 Mb), 4A.2, 4B.2, 4D, 5A.1, 5A.2, 5B.1, and 5D. The QTL clusters with negative alignments between traits may also have potential value for yield or GPC improvement in specific environments, including 1A.1, 2B.1, 1B.3, 5A.3, 5B.2 (612.1-613.6 Mb), 7A.1, 7A.2, 7B.1, and 7B.2. One GPC QTL (5B.2: 671.3-672.9 Mb) contributed by cultivar Spitfire was positively associated with nitrogen use efficiency or grain protein yield and is highly recommended for breeding use. Another GPC QTL without negatively pleiotropic effects on 2A (50.0-56.3 Mb), 2D, 4D, and 6B is suggested for quality wheat breeding.

Introgression of an expressed HMW 1Ay glutenin subunit allele into bread wheat cv. Lincoln increases grain protein content and breadmaking quality without yield penalty.

KEY MESSAGE: An expressed HMW glutenin subunit Glu-Ay showed positive impacts on a range of wheat processing quality and yield traits. The grain protein compositions are significantly optimised for baking, resulting in a better breadmaking quality. The unique breadmaking properties of wheat flour are related to the quality and quantity of high-molecular weight glutenin subunits (HMW-GSs) present in the grain. In the current study, the silent 1Ay HMW-GS allele, present in most bread wheat cultivars, was replaced by the expressed 1Ay21* allele, which was introgressed into Australian bread wheat cultivar Lincoln by a backcrossing and selfing scheme. Stability of gene expression and the effect of the introgressed 1Ay21* subunit on protein composition, agronomic traits, flour functionality, and breadmaking quality were studied using BC4F5 grain grown in glasshouse and field. Field phenotyping and grain quality testing showed that the 1Ay21* gene conferred significant improvements to a range of traits, including an increase in grain protein content by up to 9%, UPP% by up to 24%, bread volume by up to 28%. The glasshouse experiment and one of the field trials showed positive 1Ay21* effects on yield, while one field trial showed one significant effects. This indicates that expression of the 1Ay21* gene has the potential of simultaneously increasing protein content and grain yield under certain environment. The qualitative improvements of the grain also led to a reduction of the energy required during the baking process in addition to the significant positive effects on bread quality.

NAM gene allelic composition and its relation to grain-filling duration and nitrogen utilisation efficiency of Australian wheat.

Optimising nitrogen fertiliser management in combination with using high nitrogen efficient wheat cultivars is the most effective strategy to maximise productivity in a cost-efficient manner. The present study was designed to investigate the associations between nitrogen utilisation efficiency (NUtE) and the allelic composition of the NAM genes in Australian wheat cultivars. As results, the non-functional NAM-B1 allele was more responsive to the nitrogen levels and increased NUtE significantly, leading to a higher grain yield but reduced grain protein content. Nitrogen application at different developmental stages (mid-tillering, booting, and flowering) did not show significant differences in grain yield and protein content. The NAM-A1 allelic variation is significantly associated with the length of the grain-filling period. While the NAM-A1 allele a was associated with a short to moderate grain-filling phase, the alleles c and d were related to moderate to long grain-filling phase. Thus, selection of appropriate combinations of NAM gene alleles can fine-tune the duration of growth phases affecting sink-source relationships which offers an opportunity to develop high NUtE cultivars for target environments.

Health informatics to optimize complex laboratory developed test configurations.

BACKGROUND: Configuration of complex Laboratory Developed Tests (LDTs) is a time-consuming and complicated task, potentially leading to inconsistent LDTs in which features constraints remain unresolved and important features could remain unselected. OBJECTIVE: Our objective is to address these issues by presenting an automated, health informatics solution which autonomously optimizes feature selection in complex LDTs through Particle Swarm Optimization (PSO). The optimization goal is to minimize inconsistencies and configuration time, and maximize the number of selected features. METHODS: We implemented our technology in a local, secondary-care hospital in Pakistan which configures LDT for a local epidemic disease. First, a list of inconsistent LDT configurations is generated. This is used to initially estimate optimal PSO parameters, which are then used for optimization process. RESULTS: Results show that PSO is able to minimize 91% inconsistencies between 9 and 11 seconds. The number of selected critical features also increases by 100% in the optimized LDT configuration. CONCLUSIONS: We present a novel and the first application of computational optimization to solve LDT configuration issues.

An improved MALDI-TOF mass spectrometry procedure and a novel DNA marker for identifying over-expressed Bx7 glutenin protein subunit in wheat.

Wheat bread-making quality is mainly determined by glutenin proteins in the grain, which exist in a wide range of variable alleles with differential influence on processing attributes. A recently identified allele, Bx7 over-expression (Bx7(oe) ), has been showing highly significant positive effects on wheat dough strength over the normally expressed Bx7 allele. SDS-PAGE and normal RP-HPLC procedures failed to separate the two alleles. In the current study, an extensively optimised MALDI-TOF based procedure and a refined DNA based marker for efficiently differentiating Bx7(oe) from normal Bx7 allele were established. Results indicated that the MALDI-TOF procedure is cost effective, high throughput, and proven reliable, while the refined PCR marker only amplifies Bx7(oe) allele, a clear advantage over the previously developed codominant marker.

Identification of technical item flaws leads to improvement of the quality of single best Multiple Choice Questions.

OBJECTIVE: The purpose of the study was to identify technical item flaws in the multiple choice questions submitted for the final exams for the years 2009, 2010 and 2011. METHODS: This descriptive analytical study was carried out in Islamic International Medical College (IIMC). The Data was collected from the MCQ's submitted by the faculty for the final exams for the year 2009, 2010 and 2011. The data was compiled and evaluated by a three member assessment committee. The data was analyzed for frequency and percentages the categorical data was analyzed by chi-square test. RESULTS: Overall percentage of flawed item was 67% for the year 2009 of which 21% were for testwiseness and 40% were for irrelevant difficulty. In year 2010 the total item flaws were 36% and 11% testwiseness and 22% were for irrelevant difficulty. The year 2011 data showed decreased overall flaws of 21%. The flaws of testwisness were 7%, irrelevant difficulty were 11%. CONCLUSION: Technical item flaws are frequently encountered during MCQ construction, and the identification of flaws leads to improved quality of the single best MCQ's.

Outcome of allogeneic haematopoietic stem cell transplantation in aplastic anaemia.

OBJECTIVE: To analyze factors associated with survival, rejection and graft versus host disease in aplastic anaemia patients undergoing allogeneic haematopoietic stem cell transplantation (SCT) from HLA matched sibling donors. STUDY DESIGN: Analytical study. PLACE AND DURATION OF STUDY: Armed Forces Bone Marrow Transplant Centre, Rawalpindi, Pakistan from July 2001 to June 2010. METHODOLOGY: Consecutive aplastic anaemia (AA) patients undergoing haematopoietic stem cell transplantation from HLA-matched sibling donors at this centre were included in this study. Potential factors affecting overall survival, rejection, disease-free survival and graft versus host disease were analyzed. Survival analysis was done by Kaplan-Meier method. Cox regression model was applied for multivariate analysis. RESULTS: Ninety male and thirty-five female patients with AA were included in the study. Median age was 18 years. Conditioning regimens used were cyclophosphamide (Cy) plus antilymphocyte globulin (ALG) or antithymocyte globulin (ATG), fludarabine (FLU) +Cy+ATG, Campath 1-H +Cy in 89, 30 and 6 cases respectively. GVHD prophylaxis used was ciclosporin (CSA) plus prednisolone and short methotrexate in 81 while 44 received CSA plus prednisolone. At a median follow-up of 1185 days OS and DFS were 84% and 78% respectively. Factors associated with better OS were male sex, Flu/Cy/ATG conditioning and use of bone marrow as stem cell source. CONCLUSION: Flu/Cy/ATG conditioning regimen, bone marrow as stem cell source and CSA, prednisolone and short methotrexate regimen were associated with better survival in AA.

Does entry test make any difference on the future performance of medical students?

OBJECTIVE: To observe the influence/efficiency of entry test in the selection of students for MBBS. To see the effect of FSC and Entry test marks on the future performance of students in the coming years. METHODS: This was an Analytical Prospective Longitudinal study carried out in a private medical college from December 2009 to April 2011. Sampling was universal as all the students of year 2009 who succeeded in getting admission were included. The subjects were selected at the time of admission and the whole class (admission intake) comprising of 100 students during the admission year 2009 were studied. The same cohort was observed over a period of three years. The base line data was collected from the record, documents submitted at the time of admission and verified from the original record. RESULTS: The results of FSC and entry test are consistent in approximately all the percentage groups who got above 70% marks in FSC. Consistency was only disturbed in students who acquired 61-70% marks in FSC and entry test. The students who got 81-90% in FSC and Entry test performed well in first and second year MBBS. The males who had 66-70% marks in FSC were the poor performers (100% failed 5 out of 5) while among the females (33% failed) who got 71-75% in FSC. Best performance was shown by those who got 81-85% in FSC i.e. 91% males and 90% females passed in annual examinations. The results of FSC and Entry test are highly significant in statistical application and hence there was no association between the entry test marks and future performance. CONCLUSION: FSC marks are of prime importance, and are the best marker for merit admission in medical college. Entry test is the tool for equalization in between different institutions, boards and provinces. It controls the errors and omissions done deliberately or unintentionally by students, board mal-practitioners and other confounders at institution and board level in FSC.

Deferiprone challenge test for monitoring iron overload in hepatitis positive thalassemic major patients.

Viral hepatitis is common among beta-thalassemia major (BTM) children in Pakistan. Transfusional iron overload in BTM is usually monitored by serum ferritin. But its levels are falsely raised in viral hepatitis and do not reflect the true iron body burden in thalassemic patients. The objective of the study was to develop a test for monitoring iron overload in 'Hepatitis B&C' positive BTM patients by urinary iron excretion (UIE) after oral deferiprone chelation as compared to serum ferritin. We recruited 130 BTM patients from the registry of Thalassaemia centre at Rawalpindi, Pakistan. The patients were grouped into Hepatitis positive and Hepatitis negative based on ELISAtest. Serum ferritin levels were analyzed by kit on Access II. Each patient was given 75mg/kg of deferiprone at morning. Baseline UIE before deferiprone, and 4, 8 12 hours (hrs) UIE after deferiprone were analyzed on Selectra E. One hundred and thirty BTM patients aged 3 to 23 years comprising of Hepatitis positive (n=69) and Hepatitis negative (n=61) participated in the study. Hepatitis positive thalassemic patients had significantly high serum ferritin median (IQ) 4349 (2782-5927) mug/Lthan 3338 (2189-5506) mug/Lin the Hepatitis negative (p=0.001). We did not find any significant change in UIE at 4, 8, and 12 hours between two groups after Deferiprone intake (p=NS). We observed significant positive correlation between serum ferritin and 4 hours UIE in Hepatitis negative patients (r=0.57; p=0.01) but not in the Hepatitis positive patients (r=0.16; p=NS). Deferiprone challenge with measurement of 4 hours UIE is cost effective and non-invasive test rather than serum ferritin for monitoring iron overload in Hepatitis' positive BTM patients.

Formulation and evaluation of antisebum secretion effects of sea buckthorn w/o emulsion.

PURPOSE: This study was designed to formulate and evaluate the anti-sebum secretion effects of a topical skin-care cream (w/o emulsion) of sea buckthorn versus its vehicle (Base) as control. MATERIALS AND METHODS: Concentrated sea buckthorn (H.rhamnoides) fruit extract was entrapped in the inner aqueous phase of w/o emulsion. Base containing no extract and a Formulation containing 1% concentrated extract of H.rhamnoides was formulated. Lemon oil was incorporated to the odor. Both the Base and the Formulation were stored at different storage conditions for a period of 4 weeks to predict their stability. Different stability parameters i.e.; physical stability, centrifugation, and pH were monitored at different time intervals. Both the Base and the Formulation were applied to the cheeks of 10 healthy human volunteers (n=10) for a period of 8 weeks. RESULT: The expected organoleptic stability of creams was achieved from 4 weeks in-vitro study period. Odor disappeared with the passage of time due to volatilization of lemon oil. The pH of the Formulation showed significant (P = 0.0002) decline due to high concentration of organic acids present in sea buckthorn. Similarly the Formulation showed statistically significant (P < 0.05) effects on skin sebum secretion. CONCLUSION: The in vitro results showed a good stability over 4 weeks of observation period of both the Base and Formulation and the Formulation has anti sebum secretion effects over 8 weeks of observation period.

External Quality Assessment Scheme (IEQAS) in haematology at National Institute of Health, Islamabad, Pakistan.

OBJECTIVE: To document overall performance and improvement, if any, gained through participation in an International External Quality Assessment Scheme (IEQAS). STUDY DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: The Haematology Department, Public Health Laboratories Division, National Institute of Health, Islamabad, Pakistan, from January 1996 to December 2006. METHODOLOGY: Overall performance of blood parameters and parasite identification were analysed. Individual values were assessed against consensus value (mean + or - SD) and deviation index (DI) from the mean, whereas coefficients of variation (CV) were calculated for years 1996 to 2006. The results are expressed as percentage of accurate versus inaccurate results, deviation index (DI) and coefficient of variation (%CV). RESULTS: The laboratory achieved 87.74% of values within acceptable limits for haemoglobin, 72.03% for white blood count, 69.49% for platelet and 77.03% for reticulocyte estimation. Results were satisfactory, having DI values less than 3 for all four parameters. %CV values was found to be dependent on the type of test performed and varied among different parameters. Difficulty has been observed in identifying Plasmodium malariae and ovale. CONCLUSION: Participation in External Quality Assessment Schemes is extremely beneficial for the improvement of laboratory performance and quality of care. Evaluation of the survey results on a regular basis serves as a useful guide to assess overall performance of the laboratory. Standardization of analytical procedures, equipments and reagents, continuous monitoring of personnel competency and thorough investigation of discordant results significantly contributes to the delivery of quality diagnostic services.

Malignant tumors of the minor salivary glands in northern Pakistan: a clinicopathological study.

BACKGROUND: Malignant tumors of the minor salivary glands comprise a small but significant proportion of oral cancers. We analyzed this group of tumors in our population. METHODS: The records of all cases of malignant minor salivary gland tumors diagnosed at the Armed Forces Institute of Pathology (AFIP), Rawalpindi, Pakistan, during a period of 10 years (1994-2003) are described. The institute receives biopsy material from armed forces and public and private sector hospitals in northern Pakistan as well as referrals for second opinion. RESULTS: A total of 21168 tumors were recorded at the AFIP Tumor Registry during the study period. These included 70 malignant minor salivary gland tumors. Twenty-three of these (32.8%) arose in the palate while the remaining tumors originated at other sites in the oral cavity. The commonest histological type was adenoid cystic carcinoma, constituting 30 cases (42.8%). The next most common type was mucoepidermoid carcinoma comprising 26 (37.1%) cases. The mean age of cases was 43.4 years and the male-to-female ratio was 1:1.4. There were 2 cases each of polymorphous low-grade adenocarcinoma and clear cell adenocarcinoma. The remaining cases included undifferentiated carcinomas, adenocarcinomas (not otherwise specified) and a few other rare tumors. CONCLUSION: Adenoid cystic carcinoma was the commonest malignant neoplasm of the minor salivary glands followed by mucoepidermoid carcinoma. The palate was the commonest location of these tumors.

A crimean - congo haemorrhagic Fever outbreak in northern balochistan.

OBJECTIVE: To describe the clinical characteristics, epidemiology, predictors of fatal outcome (PFO), and management effects of Crimean-Congo haemorrhagic fever (CCHF) patients during an outbreak in Northern Balochistan. DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: Fatima Jinnah Hospital and Combined Military Hospital, Quetta, from June to October, 2001. PATIENTS AND METHODS: Patients presenting with a fever of less than 2 weeks duration and bleeding manifestations were included. Clinical history was recorded and patients were placed on oral ribavirin, and hematological support. Diagnosis was established by PCR for CCHF or detection of CCHF specific IgM and IgG. RESULTS: Eighty-four patients were received, 34 (40.5%) were suspected of suffering from classical CCHF. All 34 (100%) patients presented with a history of fever and bleeding (epistaxis, gum bleeding, melena or haematuria). Mean platelet count was 30 x 109/L and mean ALT (alanine transferase) was 288 U/L. Among fatal cases, the mean platelet count was 18.4 x 109/L and ALT was 781 units/L. PCR for CCHF viral RNA performed on 10 patients was positive in 3 (30%) patients. CCHF specific IgM and IgG was positive in 17.6% (6/34). Four patients were brought in moribund condition and expired before treatment could be started, 4 patients expired during treatment and 76.5% (26/34) were cured. The overall mortality was 23.5% (8/34). Main predictors of fatal outcome were ALT Z 150 units/L, activated partial thromboplastin time(aPT) Z 60 seconds, prothrombin time (PT) Z 34 seconds, aspartate transferase (AST) Z 200 units/L, platelets 20 x 109/L, and fibrinogen 110 mg/dL. CONCLUSION: In this series of CCHF occurring in Northern parts of Balochistan, gastrointestinal tract bleeding was the worst prognostic factor associated with fatal outcome. Providing education to healthcare workers and at risk populations, hematological support, anti-viral drugs, and barrier nursing may help reduce mortality.

Peripheral blood-based polymerase chain reaction in diagnosis of pulmonary tuberculosis.

BACKGROUND: The rapid diagnosis of infectious diseases, particularly those that represent a public health problem, like tuberculosis, is a challenging problem. By using nucleic acid amplification techniques like PCR, one may be able to diagnose, the disease on the day of arrival of specimen in the laboratory. For diagnosis of tuberculosis by direct methods like PCR, specimens from site of infection are required. In certain cases it is difficult to get the specimens from site of infection and in such situations; some researchers have tried to detect the DNA of Mycobacterium tuberculosis complex from blood of these patients. The purposive of this study is to determine the diagnostic efficacy of peripheral blood-based polymerase chain reaction for diagnosis of pulmonary tuberculosis. METHODS: This was a simple descriptive study, carried out in Department of Microbiology, Armed Forces Institute of Pathology, Rawalpindi from Jan 2004 to Dec 2004. Sputum and blood samples were collected from 96 suspected patients of pulmonary tuberculosis. Sputum samples processed for ZN staining and AFB culture (gold standard) and blood samples processed for PCR. RESULTS: Out of 96 cases, 60 (62.5%) were culture positive. PCR was positive in 14 (14.5%). AFB smear positive were 34 (35.4%). The overall sensitivity and specificity of the PCR assay was 20% and 94.4% respectively and the positive and negative predictive values were 85.71% and 41.46% respectively. The overall efficiency of the test was 47.91%. CONCLUSION: Due to low sensitivity; a negative PCR assay does not rule the disease. However, this test may be helpful in cases where specimens from the site of infection are not available.

The pattern of malignant tumours: tumour registry data analysis, AFIP, Rawalpindi, Pakistan (1992-2001).

OBJECTIVE: To provide information regarding frequency of malignant tumours through data retrieved from pathology based tumour registry of AFIP, Rawalpindi, Pakistan. METHODS: All malignant tumours recorded with the AFIP tumour registry over a period of 10 years (1992-2001) were analysed in terms of age group, gender and type of tumour with relation to site. A comparison with the previously published material from same setting, national and international studies were also done. RESULTS: The total malignant tumours in the 10 years period were 21,168. Out of these, 12584 (59.5%) were seen in male patients while 8584 (40.5%) were in females. Total malignant tumours in pediatric age group were 927 (4.4%). The common malignant tumours in males in order of decreasing frequency were, those of prostate, skin, lymph node, leukaemia, urinary bladder, colorectum, bone, lung, stomach and liver. In females, breast carcinoma was on top followed by skin, leukaemia, ovary, coloretum, lymph node, bone, liver, cervix and gall bladder. In females, contrary to the Western studies and India, ovarian tumours were more frequent than cervical cancers. Comparison of this analysis with our previous analysis, national and international studies showed some interesting features. CONCLUSION: It was found that in males, tumours of the prostate were the most frequent as compared to the previous study, which showed lymphomas and leukemias to be the most common. On the other hand in females, tumours of the breast remained to be consistently most frequent.

Dengue fever in malaria endemic areas.

OBJECTIVE: To determine the clinical presentation and frequency of dengue fever in patients presenting with acute febrile illness. DESIGN: Observational study. PLACE AND DURATION OF STUDY: Combined Military Hospital, Attock in collaboration with Armed Forces Institute of Pathology, Rawalpindi from November 2003 to October 2004. PATIENTS AND METHODS: Patients presenting with acute febrile illness were screened for clinical features of dengue fever (DF). Dengue specific IgM was performed in clinically suspected cases. In addition, peripheral smear for malarial parasites and complete blood counts were performed. RESULTS: Eight hundred patients with fever of less than 2 weeks duration were encountered. Twenty-two (2.75%) presented with the characteristic features of DF. Dengue specific IgM was detected in 11 patients (50%). Nine out of 11 (81.8%) dengue specific IgM positive patients were also positive for malarial parasites on peripheral smear. Out of 11 DF patients, 8 patients (78%) were cured and 3 (28%) died. CONCLUSION: Dengue fever should be suspected if patient presents with bleeding manifestations, retrobulbar headache, severe myalgias and/or thrombocytopenia. Malaria and dengue may co-exist; dengue should be excluded in clinically suspected cases by laboratory investigations. Furthermore, surveillance strategies, preventive measures and healthcare workers' education is critical for curtailing this problem.

Prevalence of prothrombin gene mutation (G-A 20210 A) in general population: a pilot study.

The objective of this study was to determine the prevalence of prothrombin gene mutation in a sample population from Pakistan. Two hundred apparently healthy unrelated adults (older than 18 years) were included in the study. The sample population comprised 100 Punjabis (male 50, female 50) and 100 Pathans (male 50, female 50). Patients with a history of previous thromboembolism were excluded from the study. Five milliliters (5 mL) of whole blood was drawn in an EDTA bottle. The DNA was extracted by the standard phenol-chloroform method. The DNA was amplified between exon number 14 and the 3'-untranslated region of the prothrombin gene by a polymerase chain reaction in a thermal cycler. Amplified products were digested overnight with HindIII at 37 degrees C. The digested products were electrophoresed on 6% polyacrylamide gel. The fragments were visualized by silver nitrate staining. A heterozygous wild type and an uncut amplified product were included in the electrophoresis strip for quality control. The wild type of DNA ran as a 350-bp fragment and internal control was cut as 550- and 150-bp fragments. The abnormal prothrombin gene was cut into 350-, 322-, and 28-bp fragments. Only two cases of heterozygous prothrombin gene mutation G-A 20210A were found in the sample studied, giving an overall carrier rate of 01% (95% CI 0.4-2.4%) in the target population. Prothrombin gene mutation is present in our population but at a lower frequency than in the white population.

Chicken pox associated thrombocytopenia in adults.

OBJECTIVE: To determine the frequency and magnitude of thrombocytopenia associated with chicken pox in adults. DESIGN: Observational descriptive study. PLACE AND DURATION OF STUDY: Combined Military Hospital, Attock, from July 2003 to June 2004. PATIENTS AND METHODS: All patients of age 15 years and above with history of fever, followed by appearance of the typical vesicular chicken pox rash, were inducted after informed consent. Two milliliters of whole blood was collected on day 1 of admission, and blood counts were performed. Patients were admitted and given 800 mg oral acyclovir, 5 times/day, for 7 days, in addition to symptomatic treatment. Patients were followed till 8 weeks. RESULTS: A total of 410 patients of chicken pox were received, out of which 270 were included. Age of patients ranged between 15 and 40 years with median age of 21 years. Platelet count on the day of admission ranged between 29 x 10(9)/L to 513 x 10(9)/L, mean platelet count 178 x 10(9)/L. Platelet count < 150 x 10(9)/L was detected in 80/270 (30%) patients. Platelet count in thrombocytopenia patients was from 29 x 10(9)/L to 149 x 10(9)/L with mean 121 x 10(9)/L. Thrombocytopenia recovered within 02 weeks in 78/80 (97%) patients. In 2 patients, thrombocytopenia recovered in 3 weeks. None of the patients developed purpuric spots, ecchymosis or bleeding manifestations. CONCLUSION: Thrombocytopenia in chicken pox is a common entity. Platelet count remains above 25 x 10(9) /L, which is usually not associated with bleeding manifestations. None of the patients in this series developed purpura. No specific pattern of total leukocyte counts was predictive of the progression or regression in platelet count.