Maggot debridement therapy to treat hard-to-heal diabetic foot ulcers: a single-centre study.

OBJECTIVE: Maggot debridement therapy (MDT) has seen a resurgence in recent years in the treatment of hard-to-heal wounds, as a result of rising antibiotic resistance. The sterilised larvae of Lucilia cuprina have been used in MDT in Malaysia since 2003, with encouraging results for the treatment of hard-to-heal diabetic wounds. We report a case series of 30 patients selected from our clinic by convenient sampling with diabetic lower limb ulcers treated with MDT. The average age of patients receiving MDT was >50 years. Of the 30 patients in the study, nine were female and 21 were male. All patients had underlying diabetes, two patients had leg ulcers and 28 patients had diabetic foot ulcers. Sterilised Lucilia cuprina larvae were applied via a standard method of 10 maggots per square centimetre and dressed with sterile gauze. The study endpoint was defined as ≤5% coverage with slough or necrotic tissue following three successive applications of MDT. In this study, maximum debridement of wounds was achieved in 96.6% (29 patients) of our patients, with ≤5% coverage with slough or necrotic tissue, in addition to a reduction in wound-related pain, as assessed by a visual analogue scale. No adverse events were reported. The findings of this study support the use of MDT as a safe, efficacious, and cost-effective method of managing diabetic wounds.

Group A Streptococcus puerperal sepsis with invasive neonatal infection: A fatal case.

Neonatal invasive Group A Streptococcus (GAS) infection is a rare occurrence nowadays. Prior maternal vaginal colonization is an important factor in early neonatal disease. We report a case of invasive and fatal infection in a neonate. At Day 1 of life, a term baby was found to be lethargic, with poor feeding, and later became unresponsive. Consequently, the baby was immediately brought to the Emergency Department of Hospital Universiti Sains Malaysia, Kubang Kerian, Kelantan via ambulance. Despite the active resuscitation efforts in the hospital, the baby died. An autopsy was carried out to determine the cause of death. The mother was noted to have puerperal pyrexia secondary to vaginal discharge. Her high vaginal swab culture was positive for GAS. GAS was also isolated from the intracardiac blood, pleural fluid, peritoneal fluid, and umbilical swab of the baby, giving evidence to the aetiology of the mortality. Vaginal colonization of GAS is an important factor for high morbidity and mortality for both mother and infant due to its invasiveness and virulence.

First report and molecular characterization of Cryptosporidium spp. in humans and animals in Khartoum state, Sudan.

BACKGROUND AND AIM: Cryptosporidium is recognized to infect several mammalian species as well as humans, causing substantial economic losses and serious public health concern. Infected animals can be a source of environmental contamination and human infections. In general, the occurrence of Cryptosporidium species in animals and human in Sudan and zoonotic importance is not well documented. This study aimed to identify Cryptosporidium spp. infecting different animal species and humans and to compare between different isolates obtained. MATERIALS AND METHODS: To provide molecular information about Cryptosporidium in animals and humans, both modified Ziehl-Neelsen (MZN) specific stain and molecular assay were used. Concentration techniques followed by three protocols of DNA extraction were carried out. After microscopic screening of 263 fecal samples (goats [n=197], cattle [n=12], sheep [n=12], and human [n=42]), 61 positive and 30 negative, randomly selected samples were used in nested polymerase chain reaction (PCR) targeting part of the 18S RNA. RESULTS: Nested PCR amplification confirmed 91.8% (56/61) of microscopic-positive samples. 8.2% (5/61) of negative samples by PCR (positive by microscopy) were considered false negatives. Sequencing followed by alignment of the 14 isolates indicated that all samples were identical (100%) and belonged to Cryptosporidium parvum. CONCLUSION: MZN staining procedure is reliable for the routine diagnosis of Cryptosporidium; cetyltrimethylammonium bromide extraction buffer and nested PCR targeting 18S rRNA gene are reliable and useful in epidemiological studies of this parasite.

Zika virus modulates blood-brain barrier of brain microvascular endothelial cells.

Zika virus (ZIKV) is a mosquito-borne Flaviviruses. ZIKV is known to cause birth defect in pregnant women, especially microcephaly in the fetus. Hence, more study is required to understand the infection of Zika virus towards human brain microvascular endothelial cells (MECs). In this study, brain MECs were infected with ZIKV at MOI of 1 and 5 in vitro. The changes in barrier function and membrane permeability of ZIKV-infected brain MECs were determined using electric cell-substrate impedance sensing (ECIS) system followed by gene expression of ZIKV-infected brain MECs at 24 hours post infection using one-color gene expression microarray. The ECIS results demonstrated that ZIKV infection enhances vascular leakage by increasing cell membrane permeability via alteration of brain MECs barrier function. This was further supported by high expression of proinflammatory cytokine genes (lnc-IL6-2, TNFAIP1 and TNFAIP6), adhesion molecules (CERCAM and ESAM) and growth factor (FIGF). Overall, findings of this study revealed that ZIKV infection could alter the barrier function of brain MECs by altering adhesion molecules and inflammatory response.

Zika virus modulates blood-brain barrier of brain microvascular endothelial cells

@#Zika virus (ZIKV) is a mosquito-borne Flaviviruses. ZIKV is known to cause birth defect in pregnant women, especially microcephaly in the fetus. Hence, more study is required to understand the infection of Zika virus towards human brain microvascular endothelial cells (MECs). In this study, brain MECs were infected with ZIKV at MOI of 1 and 5 in vitro. The changes in barrier function and membrane permeability of ZIKV-infected brain MECs were determined using electric cell-substrate impedance sensing (ECIS) system followed by gene expression of ZIKV-infected brain MECs at 24 hours post infection using one-color gene expression microarray. The ECIS results demonstrated that ZIKV infection enhances vascular leakage by increasing cell membrane permeability via alteration of brain MECs barrier function. This was further supported by high expression of proinflammatory cytokine genes (lnc-IL6-2, TNFAIP1 and TNFAIP6), adhesion molecules (CERCAM and ESAM) and growth factor (FIGF). Overall, findings of this study revealed that ZIKV infection could alter the barrier function of brain MECs by altering adhesion molecules and inflammatory response.

Chronic magnesium deficiency and human disease; time for reappraisal?

Numerous epidemiological, experimental and clinical studies over the last 30 years have consistently shown that chronic magnesium deficiency is associated with and/or exacerbates a number of major disorders (Table 1). Yet chronic magnesium deficiency is not widely recognized and a major reason for this failure is that serum magnesium levels do not accurately reflect body magnesium stores. Specifically, in chronic magnesium deficiency, serum magnesium levels are often within the normal reference range (usually lowest quartile) and may not progress to overt hypomagnesaemia. This raises serious questions namely (i) should chronic magnesium deficiency be considered in high-risk patients irrespective of serum magnesium, even when 'normal'? and (ii) if recognized, should oral magnesium supplement be given to restore body stores? Appreciating the vital role of magnesium for normal cellular function and bone health may help in formulating a well-considered and justifiable approach to these questions. Pragmatic tests for assessing magnesium status in the adult are suggested and discussed.

Injection safety among primary health care workers in Jazan Region, Saudi Arabia.

BACKGROUND: Occupational exposure to percutaneous injuries is a substantial source of infections with blood-borne pathogens among health-care workers. Few studies evaluated injection safety practices in Saudi Arabia. OBJECTIVE: To examine the structure and process of injection safety at primary health care level in Jazan health district, to evaluate knowledge, attitudes, and practices of primary health care physicians and nurses towards injection safety, and to determine the incidence of needle stick injuries among health care workers in Jazan region, Saudi Arabia. METHODS: A cross-sectional study was conducted in Jazan primary health care centers (PHCCs), Saudi Arabia from September 2011 to March 2012. Data were collected using an observational checklist and data collection sheet. Jazan city health district was chosen at random from the 14 health sectors in Jazan region. All the 33 (10 urban, and 23 rural) PHCCs of Jazan city were included in this study to get the predetermined sample size of health care workers. 200 health care workers (HCWs) were recruited (29% physicians, and 71% nurses). RESULTS: Syringes in the PHCCs were disposable (100%), individually packed (92%), and available at all volumes (98%). Methods of safe disposal of needles and sharps were also operated through contracting with professional companies in 84.8% of instances. Urban PHCCs had more posts for injection safety promotion than rural centers (p=0.02). Continuous Medical Education (CME) programs on infection control were present in only 60% of PHCCs. At least 95% of HCWs in Jazan believed that sharp objects should be kept in a puncture-proof container, kept in a closed container, or disposed by a professional company. More than 80% of HCWs washed their hands by soap and water and cleaned them by alcohol before giving injection, and also got the three doses of hepatitis B vaccine.The rate of needle stick injury in the past year was 14%, without a significant difference between nurses and physicians (p=0.8). CONCLUSION: Jazan PHCCs have reasonable facilities that prevent needle-stick injuries. We need to design and implement more educational programs on safety injection, and increase promotion of safety injection posters, especially in rural PHCCs.

Predictors of ischaemic heart disease in a Malaysian population with the metabolic syndrome.

AIMS: Cardiovascular disease is the foremost cause of mortality in Malaysia but little is known about the prevalence of the metabolic syndrome and its associations with other known cardiovascular risk markers. We undertook a population-based study to examine these. METHODS: For the study, 4341 subjects were selected using a multistage stratified sampling method. Subjects were interviewed for personal and past medical history. Biomedical markers and anthropometric indices were measured. The metabolic syndrome was defined using the harmonized criteria. The associations between the metabolic syndrome and cardiovascular risk markers, including high-sensitivity C-reactive protein, microalbuminuria and HbA(1c) were examined. RESULTS: The prevalence of the metabolic syndrome was 42.5%. Subjects with the metabolic syndrome are significantly more likely to have higher BMI (> 25 kg/m(2)), HbA(1c) [≥ 42 mmol/mol (6.0%)], LDL (≥ 2.6 mmol/l), elevated albumin:creatinine ratio (> 2.5 µg/mmol creatinine for men, 3.5 µg/mmol creatinine for women) and high-sensitivity C-reactive protein (> 3 mg/l); odds ratio 5.48, 6.14, 1.44, 3.68 and 1.84, respectively, P < 0.001. The presence of an elevated albumin:creatinine ratio and high-sensitivity C-reactive protein are strong predictors for the presence of a higher number of positive criteria of the metabolic syndrome. HbA(1c) > 48 mmol/mol (6.5%) is associated with increased relative risk of elevated albumin:creatinine ratio, high-sensitivity C-reactive protein and LDL (relative risk 3.10, 2.46 and 1.65 respectively, P < 0.001). CONCLUSIONS: We confirmed the high prevalence of the metabolic syndrome in Malaysia. Our study revealed a strong relationship between risk markers of elevated BMI, HbA(1c), LDL, albumin:creatinine ratio and high-sensitivity C-reactive protein with the presence of the metabolic syndrome, putting them at a statistically high risk for cardiovascular mortality.

Effects of high-mobility group a protein application on canine adipose-derived mesenchymal stem cells in vitro.

Multipotency and self-renewal are considered as most important features of stem cells to persist throughout life in tissues. In this context, the role of HMGA proteins to influence proliferation of adipose-derived mesenchymal stem cell (ASCs) while maintaining their multipotent and self-renewal capacities has not yet been investigated. Therefore, extracellular HMGA1 and HMGA2 application alone (10-200 ng/mL) and in combination with each other (100, 200 ng/mL each) was investigated with regard to proliferative effects on canine ASCs (cASCs) after 48 hours of cultivation. Furthermore, mRNA expression of multipotency marker genes in unstimulated and HMGA2-stimulated cASCs (50, 100 ng/mL) was analyzed by RT-qPCR. HMGA1 significantly reduced cASCs proliferation in concentrations of 10-200 ng/mL culture medium. A combination of HMGA1 and HMGA2 protein (100 and 200 ng/mL each) caused the same effects, whereas no significant effect on cASCs proliferation was shown after HMGA2 protein application alone. RT-qPCR results showed that expression levels of marker genes including KLF4, SOX2, OCT4, HMGA2, and cMYC mRNAs were on the same level in both HMGA2-protein-stimulated and -unstimulated cASCs. Extracellular HMGA protein application might be valuable to control proliferation of cASCs in context with their employment in regenerative approaches without affecting their self-renewal and multipotency abilities.

Neurobehavioral performance among agricultural workers and pesticide applicators: a meta-analytic study.

Chronic low level exposure of agricultural workers and applicators to pesticides has been found to be associated with different degrees of decrement in cognitive and psychomotor functions. The goal of this study was to use meta-analysis to (1) identify and quantify neurobehavioral deficits among agricultural workers and pesticide applicators, and (2) analyse the potential confounders or moderators of these neurobehavioral deficits. Seventeen studies, reporting on 21 independent cohort groups, were included in the meta-analysis. These studies involved 16 neuropsychological tests providing 23 different performance measures that constitute the neurobehavioral constructs. All tests and measures of the neurobehavioral functions of attention, visuomotor integration, verbal abstraction and perception constructs showed significant decrements for exposed participants. One out of three tests of memory, two of five tests of sustained attention, and four of eight tests of motor speed constructs also showed significant decrements. Nine out of these 15 effect size distributions demonstrated significant heterogeneity across cohorts. A search for cohort-level variables (eg, agricultural workers vs applicators, duration of exposure, age and percentage of male participants) to explain this heterogeneity was largely unsuccessful. However, for one test, Block Design, the duration of exposure was positively associated with performance decrements. Furthermore, it was also found that performance decrements on this test were smaller for older participants. Increasing the number of studies and using more consistent methodologies in field studies are needed.

Anaerobic capacity of physically active eumenorrheic females at mid-luteal and mid-follicular phases of ovarian cycle.

AIM: The present study was undertaken to evaluate the anaerobic capacity in repeated sprint cycling bouts during mid-luteal (ML) and mid-follicular (MF) phases of ovarian cycle. METHODS: Twelve physically active females aged 22.41±1.68 years, with normal regular menstrual cycle and VO2max of 34.92±4.85 mL·kg-1·min-1 volunteered as subjects in this study. The menstrual phases were verified through daily basal body temperature recording and serum progesterone analysis. Anaerobic capacity was quantified by measuring maximal accumulated oxygen deficit (MAOD). In this study, MAOD was measured following well established method. Initially, the VO2max of the subjects was measured following a graded exercise protocol on a cycle ergometer. On separate days, the subjects performed sub-maximal cycling exercise at 50%, 60%, 70% and 80% of VO2max for 10 minutes. The linear regression determined from the sub-maximal VO2-power relationship was used to estimate the supra-maximal power output at 120% VO2max. The subjects performed repeated sprint cycling for 3 times at 120% of VO2max with 20 minutes rest between consecutive sprints during MF and ML phases. RESULTS: Results indicated there was no significant difference in maximal accumulated oxygen deficit (MAOD) and sprint performance between MF and ML phases in repeated sprint cycling. Serum progesterone was significantly lower in luteal phase after repeated sprints. CONCLUSION: Hence, it is concluded that the anaerobic capacity is unaffected by ovarian phases in women with regular menstrual cycle. Lower serum progesterone after repeated sprints might be due to the repeated anaerobic activity.

Reproducibility of laser Doppler fluximetry and the process of iontophoresis in assessing microvascular endothelial function using low current strength.

Iontophoresis of acetylcholine (ACh) and sodium nitroprusside (SNP) combined with laser Doppler fluximetry (LDF) is a tool used to determine microvascular endothelial function. Our aim was to study the reproducibility of different parameters of this technique using iontophoresis with low current strength on the forearm skin of healthy subjects. Baseline skin perfusion was done before application of five current pulses with 1 min of current-free interval. Current strength of 0.007 mA, current density of 0.01 mA/cm(2) and charge density of 6 mC/cm(2) were used, along with 1% ACh and 1% SNP. The absolute maximum change in perfusion (max), percent change in perfusion (% change), peak change in perfusion (peak) and area under the curve during iontophoresis (AUC) at the anodal and cathodal leads were recorded. Measurements were performed in three sessions for 2 days. The coefficient of variation (CV) was calculated for each parameter. Among the parameters studied, maximum change in perfusion and peak flux were the most reproducible parameters.

Type 2 diabetes mellitus patients with poor glycaemic control have lower quality of life scores as measured by the Short Form-36.

INTRODUCTION: This study aimed to compare the quality of life based on the Short Form-36 (SF-36) between two different groups of type 2 diabetes mellitus patients with glycaemic control: those with a glycosylated haemoglobin (HbA1c) level at or below 7.5 percent and those above 7.5 percent. METHODS: In this cross-sectional study, a generic SF-36 questionnaire was self-administered to patients with type 2 diabetes mellitus. Based on the HbA1c level, the mean SF-36 scale scores were compared. The analysis of covariance was used to obtain the adjusted mean scores of the SF-36 scales while controlling for age and duration of type 2 diabetes mellitus. RESULTS: 150 patients with type 2 diabetes mellitus were analysed. There were 63 (42 percent) women and 87 (58 percent) men, and their mean HbA1c level was 8.9 percent (SD 2.4 percent). When comparing the two groups of patients with different HbA1c levels, the adjusted means of four scales: physical health functioning, general health, social functioning and mental health, differed significantly between the two. The SF-36 scale scores in type 2 diabetes mellitus patients were also lower than those of the SF-36 norms for the Malaysian population. CONCLUSION: Type 2 diabetes mellitus patients with poor glycaemic control had lower mean SF-36 scores in physical functioning, general health, social functioning and mental health, and the SF-36 scores in these patients were also lower than the SF-36 norms of the Malaysian population.

Clinical and biochemical parameters of children and adolescents applying pesticides.

BACKGROUND: The primary agricultural product in Egypt is the cotton crop. Children and adolescents work seasonally in the cotton fields applying pesticides. OBJECTIVE: To examine the effect of pesticide exposure on clinical and biochemical parameters in children and adolescents applying pesticides. METHODS: Male children currently applying pesticides and aged between 9 and 19 years (n = 50) were recruited for this study. They were asked to complete work, health, and exposure questionnaires; examined for any medical and neurological problems with particular attention to sensory and motor functions including cranial nerves, sensory and motor system, and reflexes. From each participant, a blood sample was taken to measure acetylcholinesterase activity, and liver and kidney functions. Children who have never worked in agriculture (n = 50), matched on age, education, and socioeconomic status were also studied and served as controls. RESULTS: More neuromuscular disorders were identified in pesticide applicators than controls. A significant lower level of acetylcholinesterase was found in the applicator group compared to the controls. There was also a significant difference in hematological, renal and hepatic indices in the exposed children compared to the control children. Working more days in the current season and also working more years as a pesticide applicator were both associated with an increase in the prevalence of neuromuscular abnormalities and significant changes in the laboratory tests. CONCLUSION: Children and adolescent pesticide applicators working in farms of Egypt are at risk of developing serious health problems similar to those of adults.

Interference from endogenous antibodies in automated immunoassays: what laboratorians need to know.

Full automation of laboratory procedures confers numerous advantages over semi-automated/manual tests because equipment, reagents and the computation of results are offered as an integrated package. Automation has allowed millions of immunoassay tests to be performed with good sensitivity and excellent precision but inaccuracy caused by interference from endogenous immunoglobulins/antibodies remained a problem (irrespective of the immunoassay's format). Interference leading to a falsely high or low result affects a specific sample and may not be obvious despite the strictest laboratory control schemes. Reporting and interpreting such potentially erroneous data remained however the responsibility of the clinical laboratory despite the limited information supplied by their providers. The focus of this review is on highlighting the potential downside of current disjointed and blurred arrangement between the developers/providers of immunoassays, and the laboratorians responsible for providing these data to their clinical colleagues. These limitations can be addressed by drawing attention to the importance of the key fundamentals underpinning these immunologically based analyses which, if carefully considered, could help to formulate pragmatic strategies to reduce errors in immunoassays. In this review, the inherent fallibility of the binding reaction between an antigen and antibody will be reiterated. The difficulties in defining reaction rate kinetics in non-equilibrium automated assays, the potential clinical error rate and the need for minimising analytical error rate of these automated technologies will be highlighted.

The reliability and validity of the Malay version of the 18-item audit of Diabetes Dependent Quality of Life (the Malay ADDQOL) questionnaire.

The objective of this study was to examine the reliability and validity of the Malay version of the 18-item Audit of Diabetes Dependent Quality of Life (the Malay ADDQOL). Patients with type 2 diabetes mellitus who fulfilled the inclusion criteria were systematically selected. The Malay ADDQOL linguistically validated from the 18-item English version ADDQOL was self-administered twice at a 1-week interval. Two hundred eighty-eight respondents were included in the study. Analysis involved checking the feasibility, floor and ceiling effects, internal consistency, test-retest reliability and factor analysis. Item means and standard deviations fulfilled the Likert scale assumptions. The Cronbach's alpha was 0.943 (lower bound of the 95% CI of 0.935) and the intraclass correlation coefficient was 0.81 (95% CI from 0.72 to 0.87). Exploratory one factor analysis showed factor loadings above 0.5 for all the 18 items. The Malay ADDQOL has acceptable linguistic validity. It is feasible, has excellent reliability, content, construct validity, and is recommended to be used among Malay-speaking diabetic patients.

Effect of therapeutic lifestyle changes on insulin sensitivity of non-obese hyperlipidemic subjects: preliminary report.

AIM: To determine the effects of lipid lowering by TLC on insulin sensitivity and secretory status of non-obese normoglycemic hyperlipidemic subjects. METHODS: An intervention study was undertaken on 16 non-obese normoglycemic hyperlipidemic subjects. They underwent 6 months of a TLC regimen. Their insulin sensitivity and lipid status were assessed at baseline and after six months. A control group containing 16 age, sex and body mass index (BMI) matched normolipidemic subjects was also enrolled to compare the change in lipid levels and insulin sensitivity in the hyperlipidemic subjects. RESULTS: The intervention showed significant reductions in insulin resistance (HOMA-IR reduced from 3.8 to 1.4, p<0.001) and improvement of insulin sensitivity (HOMA%S increased from 50.1% to 121.2%, p=0.004) in hyperlipidemic subjects with associated reductions in lipid levels. CONCLUSION: Lipid lowering in non-obese hyperlipidemic subjects may be associated with improvement of insulin sensitivity.

Rib fractures predict incident limb fractures: results from the European prospective osteoporosis study.

Population studies suggest that rib fractures are associated with a reduction in bone mass. While much is known about the predictive risk of hip, spine and distal forearm fracture on the risk of future fracture, little is known about the impact of rib fracture. The aim of this study was to determine whether a recalled history of rib fracture was associated with an increased risk of future limb fracture. Men and women aged 50 years and over were recruited from population registers in 31 European centres for participation in a screening survey of osteoporosis (European Prospective Osteoporosis Study). Subjects were invited to complete an interviewer-administered questionnaire that included questions about previous fractures including rib fracture, the age of their first fracture and also the level of trauma. Lateral spine radiographs were performed and the presence of vertebral deformity was determined morphometrically. Following the baseline survey, subjects were followed prospectively by annual postal questionnaire to determine the occurrence of clinical fractures. The subjects included 6,344 men, with a mean age of 64.2 years, and 6,788 women, with a mean age of 63.6 years, who were followed for a median of 3 years (range 0.4-5.9 years), of whom 135 men (2.3%) and 101 women (1.6%) reported a previous low trauma rib fracture. In total, 138 men and 391 women sustained a limb fracture during follow-up. In women, after age adjustment, those with a recalled history of low trauma rib fracture had an increased risk of sustaining 'any' limb fracture [relative hazard (RH)=2.3; 95% CI 1.3, 4.0]. When stratified by fracture type the predictive risk was more marked for hip (RH=7.7; 95% CI 2.3, 25.9) and humerus fracture (RH=4.5; 95% CI 1.4, 14.6) than other sites (RH=1.6; 95% CI 0.6, 4.3). Additional adjustment for prevalent vertebral deformity and previous (non-rib) low trauma fractures at other sites slightly reduced the strength of the association between rib fracture and subsequent limb fracture. In men, after age adjustment, there was a small though non-significant association between recalled history of rib fracture and future limb fracture. Our data highlight the importance of rib fracture as a marker of bone fragility in women.

A simple and rapid genotyping method for beta-2 receptor (beta2 AR) gene using allele specific multiplex PCR.

BACKGROUND: Polymorphism of the beta2-adrenergic receptor (beta2 AR) gene is an important determinant of the function of this receptor. It affects receptor down-regulation and beta2-agonist responses. It has also been a focus of interest in attempts to elucidate the genetic basis of asthma, hypertension, obesity and cystic fibrosis. Several different techniques have been established to determine beta2 AR genotypes but none of these methods are simple enough to detect simultaneously all the five alleles of our research interest (Arg16/Gly16, -20T/C, Gln27/Glu27, -47T/C and Thr164/Ile164). OBJECTIVES: To develop a simple and rapid PCR based method for the simultaneous detection of five beta2 AR alleles. METHOD: DNA was extracted from whole blood using standard alkali lysis method. Primers specific at the 3' end for the polymorphic sites were designed. The nested allele specific PCR was optimized for reproducibility and specificity. Parameters investigated included concentrations of MgCl2, Taq polymerase, primers and annealing temperature, to produce specific bands of interest. DNA samples were selected at random and submitted for direct PCR sequencing. RESULT: The first PCR produced a fragment of size 710 bp, which was used as template for the subsequent duplex and triplex PCR to detect the mutation sites of the five alleles. The method was found reproducible and specific when used to genotype patients with bronchial asthma. The sequencing results confirmed the specificity of the PCR method. CONCLUSION: The simple and rapid method for the simultaneous detection of the five beta2 AR alleles is suitable for the study of beta2 polymorphism and its clinical consequences.