Novel SPG20 mutation in an extended family with Troyer syndrome.

Troyer Syndrome (TRS) is a rare autosomal recessive complicated spastic paraplegia disorder characterized by various neurological and musculoskeletal manifestations. Pathogenicity stems from mutations in SPG20 which encodes Spartin, a multifunctional protein that is thought to be essential for neuron viability. Here we report on the clinical and molecular characterization of TRS in five patients from an extended consanguineous family in the United Arab Emirates. Molecular analysis involved Whole Exome Sequencing and Sanger sequencing for identification and confirmation of the causative variant respectively. In silico tools including CADD and Polyphen-2 were used to assess pathogenicity of the variant. The clinical description of these patients included spastic paraparesis, motor and cognitive delay, gait abnormalities, musculoskeletal features, as well as white matter abnormalities and emotional liability. Molecular analysis revealed a novel homozygous missense mutation in SPG20 (c.1324G > C; p.Ala442Pro) occurring at an evolutionarily conserved residue in the Plant-Related Senescence domain of Spartin. The mutation segregated with the clinical phenotype in all patients. In silico algorithms predict the mutation to be disease causing, and the variant had not been previously reported in public or ethnic specific variant repositories.

Association of HLA-DQA1 and -DQB1 alleles with type I diabetes in Arabs: a meta-analyses.

This study aimed at assessing the nature and significance of associations between various alleles of HLA-DQA1, HLA-DQB1, and type I diabetes (T1D) in Arab populations. Evidence from literature (published before 20 April 2015) was amassed and analysed through multiple meta-analyses, which yielded effect summary odds ratios and 95% confidence intervals for 24 alleles and 4 haplotypes. A total of 1273 cases and 1747 controls from 16 studies were analysed. High levels of significance were obtained to support higher T1D risk when harbouring DQA1*03:01. The alleles DQB1*02:01 and *03:02 and the haplotypes DR3 and DR4 were significant risk factors, albeit with high publication heterogeneity. The protective effects of DQA1*01:01, DQB1*05:03, *06:02, *06:03, and *06:04 were robustly suggested by all indicators of meta-analyses. The haplotypes DR7 and DR11 were strongly suggested to be protective in Arabs. A relatively small number of studies have emerged from Arab countries, mostly with inadequate power on an individual basis. This study fills the gap by providing significant size effect of human leukocyte antigen (HLA) alleles and completes the continuum of global ethnic differences in this context.

Root canal debridement efficacy of different final irrigation protocols.

AIM: To compare the smear layer and debris removal effectiveness of four root canal irrigation protocols as well as their effectiveness in removing remaining soft tissues in curved root canals. METHODOLOGY: The mesiobuccal and mesial root canals of 107 extracted human maxillary and mandibular molars were instrumented using Mtwo rotary NiTi instruments then randomly divided into four groups according to a final rinse protocol: Group 1 (n = 28) - manual agitation of 1% NaOCl and 15% EDTA; Group 2 (n = 26) - CanalBrush agitation of 1% NaOCl and 15% EDTA; Group 3 (n = 26) - 3% H(2) O(2) alternated with 1% NaOCl; Group 4 (n = 27) - passive ultrasonic agitation of 1% NaOCl and 15% EDTA. All irrigation protocols were performed in a closed system. Eleven roots per group were prepared and histologically stained (H&E) to assess percentage of remaining pulpal tissues in the apical thirds. The remaining specimens were split longitudinally and examined under scanning electron microscope at ×2000 magnification to assess smear layer and debris removal. Image Pro Plus 6.0 software was used to analyse smear layer and remaining pulp tissue. Debris presence was scored by two blinded investigators using a five-point scale. Data were analysed using Univariate analysis of variance (GenStat 13, α = 0.05). RESULTS: CanalBrush and passive ultrasonic irrigation were equally effective with significantly less smear layer and debris than manual agitation and H(2) O(2) alternated with NaOCl (P < 0.05). The H(2) O(2) alternated with NaOCl protocol was significantly more effective in removing pulp tissue remnants in the apical level than manual agitation (P = 0.009) and passive ultrasonic irrigation (P = 0.01). CONCLUSIONS: CanalBrush was as effective as passive ultrasonic irrigation in smear layer and debris removal. Alternating H(2) O(2) with NaOCl was effective in removing soft tissues from root canal complexities. Further studies are required to evaluate effectiveness of this regimen taking into account irrigant volume differences and effect of root canal system configuration.

Continuous glucose monitoring, oral glucose tolerance, and insulin - glucose parameters in adolescents with simple obesity.

In obese adolescents pancreatic beta-cells may not be able to cope with insulin resistance leading to hyperglycemia and type2 diabetes (T2DM To assess oral glucose tolerance, 72-h continuous blood glucose concentrations (CGM) and calculate homeostatic model assessment (HOMA), and the quantitative insulin sensitivity check index (QUICKI) in 13 adolescents with simple obesity (BMI SDS=4 ± 1.06). OGTT performed in 13 obese adolescents (13.47 ± 3 years) revealed 3 cases (23%) with impaired fasting glucose (IFG: fasting glucose >5.6 mmol/L), 4 cases (30%) with impaired glucose tolerance (IGT: 2h blood glucose >7.8 <11.1 mmol/L), and none with diabetes. Using the continuous glucose monitoring system ( CGMS), IFG was detected in 4 cases, the maximum serum blood glucose (BG : 2h or more after meal) was >7.8 and <11.1 mmol/L (IGT) in 9 children (69%) and >11.1 mmol/L (diabetes) in one case (7.6%). Five cases had a minimum BG recorded of <2.7 mmol/L (hypoglycemia). No glycemic abnormality was detected using HbA1C (5.7 ± 0.3%). 11/13 patients had HOMA values >2.6 and QUICKI values <0.35 denoting insulin resistance. Beta cell mass percent (B %) = 200 ± 94.8% and insulin sensitivity values (IS)=50.4 ± 45.5% denoted insulin resistance with hyper-insulinaemia and preserved beta cell mass. In obese adolescents, CGMS is superior to OGTT and HbA1C in detecting glycemic abnormalities, which appears to be secondary to insulin resistance.

Effect of Long-Term Treatment with Dexamethasone on the Liver and Kidney Histopathology, as well as Blood Biochemistry in Male Rabbits (Lepus Cuniculus).

Glucocorticoids have a wide range of pharmacological activities. Generally speaking, the steroid drugs, such as dexamethasone (DEX) can have severe side effects on the histology of different organs. In fact, glucocorticoids have been known as powerful medicines which can cure inflammation and work with the immune system to treat a wide range of health problems. Therefore, this study aimed to investigate the effects of DEX on the histological changes of the liver and kidney, as well as blood biochemical parameters. In total, 13 specific pathogen-free male Lepus Cuniculus rabbits aged 8-10 months old, with a mean weight of 1.12±0.13 kg were randomly divided into three groups. Group I (n=3) did not receive DEX, and they only received saline solution as a placebo (control). In Group II (n=5), the animals received 0.25 mg DEX/kg body weight/day for a period of 56 days, and the animals in Group III (n=5) received 0.5 mg DEX/kg body weight/day for 56 days. Blood was aspirated from the rabbit's marginal ear vein. All blood samples were centrifuged at 3000×g for 10 min to separate serum samples. Blood lipids and trace elements (zinc, copper, calcium, and iron) were measured. The microscopical analyses of the liver and kidney tissues were performed through the observation of the histological changes in the tissues. The results showed a significant (P≤0.05) decrease in the body and organ weight, as well as serum concentrations for the trace elements. On the other hand, lipid profile showed a significant increase (P≤0.05) in cholesterol, triglycerides, and low-density lipoprotein. However, a significant decrease was recorded in high-density lipoprotein in both treated groups with DEX, compared to the control group. The results of the histological evaluation showed some degrees of degeneration, necrosis, cell vacuolation, and lymphocyte infiltration in the kidney and liver tissues in the treatment groups.

Investigation of some trace elements and hematological and biochemical parameters in the blood of emaciated Omani goats.

BACKGROUND AND AIM: The analysis of hematological and biochemical parameters is widely used in assessing animal health status. Limited information is available on trace element levels and their association with hematological and biochemical parameters in Omani goats suffering from emaciation. Therefore, the current study aimed to determine the levels of some trace elements in emaciated Omani goats and their relationship with hematological and biochemical parameters. MATERIALS AND METHODS: Goats suffering from emaciation and muscular dystrophy (n=18) were compared with healthy goats (n=12). Venous blood samples for the hematological, biochemical, and trace element analysis were collected from the jugular vein. RESULTS: Emaciated goats showed significantly lower white blood cell, lymphocyte, and red blood cell counts than the healthy goats. In contrast, the percentages of monocytes and eosinophils were higher in emaciated goats than in healthy ones. In addition, emaciated goats showed higher levels of biochemical parameters alkaline phosphatase, alanine aminotransferase, gamma-glutamyl transferase, aspartate aminotransferase, creatine kinase, and total bilirubin but lower levels of albumin than the healthy goats. The results of trace element analysis revealed lower concentrations of zinc, iron, and selenium in serum from emaciated goats than in serum from healthy goats. CONCLUSION: This study identified significant differences in the serum levels of some trace elements and hematological and biochemical parameters between healthy and emaciated Omani goats. The identified differences represent valuable diagnostic biomarkers for the evaluation of the health status of Omani goats.

Burnout among clinical dental students at Jordanian universities.

Dentistry is a profession demanding physical and mental efforts as well as people contact, which can result in burnout. The level of burnout among 307 clinical dental students in 2 Jordanian universities was evaluated using the Maslach Burnout Inventory survey. Scores for the inventory's 3 subscales were calculated and the mean values for the students' groups were computed separately. Dental students in both universities suffered high levels of emotional exhaustion and depersonalization. The dental students at the University of Jordan demonstrated a significantly higher level of emotional exhaustion than their counterparts at the Jordan University of Science and Technology.

Recurrent acute liver failure and mitochondriopathy in a case of Wolcott-Rallison syndrome.

A 10-year-old Arabic boy of consanguineous parents has suffered eight episodes of acute liver failure with haemolysis triggered by intercurrent febrile illnesses. The first crisis occurred at 9 months of age, after which diabetes mellitus developed. By the age of 6 years, short stature, mild myopathy and later skeletal epiphyseal dysplasia also became evident. His psychosocial development and educational achievements have remained within normal limits. While there were no clear biochemical indicators of a mitochondrial disorder, an almost complete deficiency of complex I of the respiratory chain was demonstrated in liver but not in fibroblast or muscle samples. Molecular analysis of the eukaryotic translation initiation factor 2alpha kinase gene (EIF2AK3) demonstrated a homozygous mutation, compatible with a diagnosis of Wolcott-Rallison syndrome (WRS). This patient's course adds a new perspective to the presentation of WRS caused by mutations in the EIF2AK3 gene linking it to mitochondrial disorders: recoverable and recurrent acute liver failure. The findings also illustrate the diagnostic difficulty of mitochondrial disease as it cannot be excluded by muscle or skin biopsy in patients presenting with liver disease. The case also further complicates the decision-making process for liver transplantation in cases of acute liver failure in the context of a possible mitochondrial disorder. Such patients may be more likely to recover spontaneously if a mitochondrial disorder underlies the liver failure, yet without neurological features liver transplantation remains an option.

Emergency obstetric hysterectomy in a university hospital: A 25-year review.

Over the past 25 years, 43 peripartum hysterectomies were performed at the authors' institution, an incidence of 0.64/1,000 deliveries; 31 procedures followed caesarean section and 12 were performed for haemorrhage following vaginal delivery. The common indications for hysterectomy were abnormal placentation (39.5%), uterine atony (23.3%), uterine rupture (23.3%), and haemorrhage during caesarean section (11.6%). The risk factors for hysterectomy included advancing maternal age and parity, previous caesarean section scars and abnormal placentation. Subtotal hysterectomy was performed in 72.1% cases which appeared a quicker and safer procedure than total hysterectomy in desperately ill patients. Five (11.6%) maternal deaths occurred in the series. Mortality was associated with massive haemorrhage. With rising caesarean section rates worldwide, MRI and colour Doppler sonography is useful to diagnose antepartum placenta accreta/bladder involvement in order to plan elective surgery that is associated with reduced maternal morbidity and mortality. Early decision to perform an emergency hysterectomy is essential before the patient's condition deteriorates, besides availability of an experienced obstetrician to undertake a technically demanding operation.

Detection of a novel familial deletion of four genes between BP1 and BP2 of the Prader-Willi/Angelman syndrome critical region by oligo-array CGH in a child with neurological disorder and speech impairment.

Two common classes of deletions are described in the literature in individuals with Prader-Willi/Angelman syndrome (PWS/AS): one between breakpoint 1 (BP1) to BP3 and the other between BP2 to BP3 of the PWS/AS critical region on chromosome 15q11-->q13. We present here a novel observation of an approximately 253-kb deletion between BP1 and BP2 on 15q11.2, in a 3(1/2)-year-old boy, who was referred to us with a clinical suspicion of having Angelman syndrome and presenting with mental retardation, neurological disorder, developmental delay and speech impairment. Karyotype and FISH results were found to be normal. The microdeletion between BP1 and BP2 includes four genes - NIPA1, NIPA2, CYFIP1 and TUBGCP5 which was detected by a high-resolution oligonucleotide array-CGH that was further validated by a Multiplex Ligation-dependent Probe Amplification (MLPA) assay. The same deletion was observed in the father who presented with similar but relatively milder clinical features as compared to the affected son. Methylation studies by methylation-specific MLPA (MS-MLPA) of the SNRPN imprinting center (IC) showed a normal imprinting pattern, both in the patient and the father. To our knowledge a microdeletion limited only to the BP1-BP2 region has not yet been reported. The familial genetic alteration together with the striking clinical presentation in this study are interesting, but from our single case study it is difficult to suggest if the deletion is causative of some of the abnormal features or if it is a normal variant. The study however further strengthens the fact that genome-wide analysis by array CGH in individuals with developmental delay and mental retardation is very useful in detecting such hidden interstitial chromosomal rearrangements.

A novel missense mutation in ATRX uncovered in a Yemeni family leads to alpha-thalassemia/mental retardation syndrome without alpha-thalassemia.

BACKGROUND: Intellectual disability (ID) features in numerous heritable medical conditions that result from ATRX mutations. Alpha-thalassemia mental retardation syndrome (ATR-X syndrome) is the most notable manifestation of ATRX dysfunction. In addition to ID, genitourinary and craniofacial abnormalities are regularly observed with or without alpha-thalassemia. AIMS: The study sought to characterize two cases of ATR-X in a Yemeni family clinically and molecularly. METHODS: PCR amplification and Sanger sequencing were used to study the ATRX gene in a Yemeni family. Also, methylation-sensitive PCR was used to perform X-inactivation studies. CADD, SNAP2 and PolyPhen-2 helped to predict the functional consequences of the variant. RESULTS: Molecular testing revealed a novel hemizygous missense mutation (c.5666T>G) in the ATRX gene in the two Yemeni brothers. This mutation was found in a heterozygous state in the mother, with the chromosome harboring the mutated allele being under strongly skewed X-inactivation. CONCLUSIONS: The mutated gene is predicted to have a disrupted SNF-2 domain at a conserved residue; p.Leu1889Trp, which is deemed functionally damaging. This report offers, for the first time, full clinical and molecular characterization of a novel ATRX variant in an Arab family.

High prevalence and level of resistance to metronidazole, but lack of resistance to other antimicrobials in Helicobacter pylori, isolated from a multiracial population in Kuwait.

BACKGROUND: The primary treatment regimen for Helicobacter pylori infection for Kuwaitis does not contain metronidazole, but that for expatriates does. There is also increasing failure of antimicrobial therapy. AIM: To determine the susceptibility of H. pylori from upper gastrointestinal biopsies of Kuwaitis and non-Kuwaitis to find out if differences existed in the susceptibilities of the isolates from the two different populations. METHODS: The susceptibilities of 96 H. pylori isolates were tested against metronidazole, amoxicillin, clarithromycin and tetracycline by the E test. The rdxA gene was analysed from selected metronidazole-susceptible and metronidazole-resistant strains to find out polymorphism and the basis of metronidazole resistance. RESULTS: Approximately, 70% of isolates from both populations were metronidazole resistant with 65% isolates showing high minimum inhibitory concentration values of >256 mug/mL. No resistance to the other three antimicrobials was found. There were novel nonsense and missense mutations with no deletion in the rdxA gene by insertion of mini-IS605. CONCLUSIONS: The prevalence and level of metronidazole resistance in H. pylori in the two populations was high with no difference, in spite of different treatment regimens. Metronidazole resistance in this transitional country appeared to be independent of prior metronidazole use for treatment of H. pylori infection.

The profile of HLA-DRB1 alleles in Arabs with type 1 diabetes; meta-analyses.

Genes from the HLA complex have a major contribution in type 1 diabetes (T1D), which results from an interplay between environmental and genetic factors. The latter can explain some of the geographic variability in T1D occurrence around the world. Of a particular importance in this regard are the HLA-DR, -DP and -DQ loci. Consequently, we aimed at elucidating the collective genetic profiles of various alleles relating to HLA-DRB1 and -DP in T1D patients throughout the Arab World using the tools of meta-analysis. As for HLA-DQA1 and DQB1 alleles; this analysis was completed and published previously (see Introduction). As a result of limited availability of relevant studies of the HLA-DP locus, only HLA-DRB1 alleles were tackled in this paper. Our study showed that significant increases in T1D risk resulted from harboring the alleles DRB1*03:01 and *04:05 (odds ratio 7.76 and 7.52, respectively). DRB1*04:01 and *04:02 were also predisposing for T1D in Arabs. Very strong evidence supported the protective effects of DRB1*10:01, *13:01, *15:02 and *16:01, with low heterogeneity and no publication bias. The results from the series of meta-analyses performed in this study help to complete the global genetic epidemiological map of T1D by providing statistically robust data from the Arab World.

Cardiac manifestations of acute carbamate and organophosphate poisoning.

OBJECTIVE: To study the frequency, extent, and pathogenesis of the cardiac complications accompanying organophosphate and carbamate poisoning. DESIGN: Retrospective study. SETTING: A medical intensive care unit (MICU) of a general hospital. SUBJECTS: 46 adult patients admitted over a five year period with a diagnosis of organophosphate or carbamate poisoning. RESULTS: Cardiac complications developed in 31 patients (67%). These were: non-cardiogenic pulmonary oedema, 20 (43%); cardiac arrhythmias, 11 (24%); electrocardiographic abnormalities including prolonged Q-Tc interval, 31 (67%); ST-T changes, 19 (41%); and conduction defects, 4 (9%). Sinus tachycardia occurred in 16 patients (35%) and sinus bradycardia in 13 (28%). Hypertension developed in 10 patients (22%) and hypotension in eight (17%). Eight patients (17%) needed respiratory support because of respiratory depression. Although more than two thirds of the patients (67%) had a prolonged Q-Tc interval, none had polymorphic ventricular tachycardia of the torsade de pointes type. Two patients died from ventricular fibrillation, an in hospital mortality of 4%. CONCLUSIONS: Cardiac complications often accompany poisoning with these compounds, particularly during the first few hours. Hypoxaemia, acidosis, and electrolyte derangements are major predisposing factors. Intensive supportive treatment in intensive or coronary care facilities with administration of atropine in adequate doses early in the course of the illness will reduce the mortality.

Exhaled nitric oxide; relationship to clinicophysiological markers of asthma severity.

Bronchial asthma is an airway disorder associated with bronchial hyperresponsiveness, variable airflow obstruction and elevated levels of nitric oxide (NO) in exhaled air. The variables all reflect, in part, the underlying airway inflammation in this disease. To understand their interrelationships we have investigated the relationship between exhaled NO levels and clinicophysiological markers of asthma severity. Twenty-six steroid naive atopic asthmatics participated in the analysis. All were given diary cards and were asked to record their peak expiratory flow (PEF) rates twice daily together with their asthma symptom scores and beta-agonist use. Diary cards were collected 2 weeks later and measurements of exhaled NO levels, FEV1 and histamine bronchial hyperreactivity (PC20 histamine) were undertaken. Exhaled NO levels were significantly higher in our study population than in normal control subjects and correlated negatively with PC20 histamine (r = -0.51; P = 0.008) and positively with PEF diurnal variability (r = 0.58; P = 0.002), but not with symptom scores, beta-agonist use of FEV1 (%). We conclude that a significant relationship exists between exhaled NO levels and the two characteristic features and markers of asthma severity, namely bronchial hyperreactivity and PEF diurnal variability. The lack of correlation between symptom score and beta-agonist use, of FEV1 (%) predicted and exhaled NO suggests that these measures are reflective of differing aspects of asthma.

Development of contralateral pleural effusion during chemotherapy for tuberculous pleurisy.

Paradoxical worsening of tuberculous lesions, despite effective chemotherapy, has been reported in intracranial tuberculomas, lymph nodes, pulmonary disease, and tuberculous pleurisy. However, development of contralateral pleural effusion during treatment of tuberculous pleurisy is very rare. We report the case of a 22 year old female patient who presented with right sided pleural effusion and was treated with antituberculous drugs. Four weeks later although her right sided pleural effusion was subsiding she developed a left sided pleural effusion. Closed pleural biopsy on the left side showed granulomatous inflammation with early caseation. Antituberculous drugs were continued and a short course of oral prednisolone was added. She recovered completely and her chest x-ray became normal after finishing her treatment.

Exhaled nitric oxide levels in exacerbations of asthma, chronic obstructive pulmonary disease and pneumonia.

OBJECTIVE: Nitric oxide is known to be present in the exhaled air of normal subjects and at higher concentrations in asthmatics. The aim of this study was to measure exhaled nitric oxide levels in patients admitted to hospital with acute exacerbations of asthma, or chronic obstructive pulmonary disease, or with pneumonia. METHODS: Within 24 hours of admission exhaled nitric oxide levels were measured by a chemiluminescent analyzer in 11 patients with acute sever asthma, 19 patients with acute exacerbation of chronic obstructive pulmonary disease, and in 12 patients with pneumonia. In asthmatics measurements were made on 3 occasions, at day 1, 4, and 28 and were related to changes in peak expiratory flow rate. RESULTS: On admission median exhaled nitric oxide levels (range) were significantly higher in asthmatics 22 (9.3-74) parts per billion in comparison to patients with chronic obstructive pulmonary disease 10.3 (2.7-34) parts per billion; p < 0.01, pneumonia 7 (4-17) parts per billion; p<0.001, and normal subjects 8.7 (5-13.3) parts per billion; p < 0.001. Following treatment the asthmatics had a significant reduction in their exhaled nitric oxide levels from 22 (9.3-74) parts per billion on day 1 to 9.7 (5.7-18.3) parts per billion on day 28; p = 0.005. Peak expiratory flow rate measurements increased from 200 (120-280) l/min on day 1 to 280 (150-475) l/min on day 4; p < 0.05 and to 390 (150-530) l/min on day 28; p < 0.01. A strong negative correlation existed between peak expiratory flow rate measurements and exhaled nitric oxide levels in asthmatics on day 28 (r = -0.70; p = 0.017). CONCLUSION: Acute exacerbations of asthma are associated with increased levels of exhaled nitric oxide in contrast to exacerbations of chronic obstructive pulmonary disease and acute pneumonia. Exhaled nitric oxide may be a useful indirect marker of asthmatic airway inflammation. The differing time course of response of nitric oxide to peak flow measures suggests that these two measures are reflecting differing airway events.

Risk of seizure recurrences after first unprovoked seizure during childhood.

OBJECTIVE: To find out incidence of seizure recurrence and its risk factor after the first unprovoked attack in children below 12 years of age. METHODS: A prospective study was carried out between 30th April 1996 - 1st May 1997 with the aim to find out the incidence of seizure recurrence and its risk factor after the first unprovoked attack in children below 12 years of age. All patients aged between 2 months - 12 years who presented with first unprovoked seizure at Hamad General Hospital, Doha, Qatar were studied. Seizures due to fever, metabolic causes, post-traumatic,and myoclonic seizures, infantile spasms, absence epilepsy and pseudo-seizures were excluded. All patients were followed for a minimum of one year. Possible risk factors such as sex, age, family history of epilepsy, delayed development, focal onset, clustered onset (two or more attacks within 24 hours), abnormal neurological findings and epileptogenic activity of electroencephalography were considered. RESULTS: There was a total of 33 patients (14 male and 19 female), 11 patients (33%) had recurrence. Significant risk factors for seizure recurrence were abnormal encephalograms (p=0.009), positive family history (p=0.03) and cluster onset (p=0.04). Presence of one or more risk factor (p=0.01) was significant. The other risk factors were not statistically significant. CONCLUSION: The incidence of seizure recurrence is higher in-patients who have one or more risk factors. The patient and the family should be aware about possibility seizure recurrence and should be involved in the decision of starting treatment after first attack.

Rapid non-enzymatic extraction method for isolating PCR-quality camelpox virus DNA from skin.

Molecular diagnostic investigations of orthopoxvirus (OPV) infections are performed using a variety of clinical samples including skin lesions, tissues from internal organs, blood and secretions. Skin samples are particularly convenient for rapid diagnosis and molecular epidemiological investigations of camelpox virus (CMLV). Classical extraction procedures and commercial spin-column-based kits are time consuming, relatively expensive, and require multiple extraction and purification steps in addition to proteinase K digestion. A rapid non-enzymatic procedure for extracting CMLV DNA from dried scabs or pox lesions was developed to overcome some of the limitations of the available DNA extraction techniques. The procedure requires as little as 10mg of tissue and produces highly purified DNA [OD(260)/OD(280) ratios between 1.47 and 1.79] with concentrations ranging from 6.5 to 16 microg/ml. The extracted CMLV DNA was proven suitable for virus-specific qualitative and, semi-quantitative PCR applications. Compared to spin-column and conventional viral DNA extraction techniques, the two-step extraction procedure saves money and time, and retains the potential for automation without compromising CMLV PCR sensitivity.