Rapid screening of pyogenic Staphylococcus aureus for confirmation of genus and species, methicillin resistance and virulence factors by using two novel multiplex PCR

Objectives: Emergence of methicillin resistant Staphylococcus aureus [MRSA] is a major medical problem of current era. These bacteria are resistant to most drugs and rapid diagnosis can provide a clear guideline to clinicians. They possess specific virulence factors and relevant information can be very useful. We designed this study to develop multiplex PCRs to provide rapid information

Methods: We studied 60 Staphylococcus aureus isolates and detected methicillin resistance by cefoxitin sensitivity and targeting of mecA gene. After initial studies with uniplex PCRs we optimized two multiplex PCRs with highly reproducible results. The first multiplex PCR was developed to confirm genus, species and methicillin resistance simultaneously, and the second multiplex PCR was for screening of virulence factors

Results: We found 38.33% isolates as methicillin resistant, a-toxin, the major cytotoxic factor, was detected in 40% whereas 6-hemolysin was found in 25% cases. Panton Valentine leucocidin was detected in 8.33% and toxic shock syndrome toxin in5% cases. The results of uniplex and multiplex PCRs were highly compatible

Conclusions: These two multiplex PCRs when run simultaneously can provide vital information about methicillin resistance and virulence status of the isolate within a few hours as compared to several days needed by routine procedures

assessment of postoperative levels of pain with reference to validated protocols at a hospital in Karachi Pakistan

Objective: The aim of study is to assess the postoperative levels of pain in accordance with established protocols

Method: The type of study is a hospital wide survey conducted from February 2014 to April 2014, at department of surgery and associated wards, at a tertiary care hospital in Karachi Pakistan. World Health Organization pain scoring system was used as a base to develop a questionnaire to assess the level of pain. The results were correlation with various other variables involved

Results: The study was conducted on 200 patients out of whom 38% [n=76] were males having mean age of 44.16 +/- 20 and 62% [n=124] were females having mean age of 36 +/- 14.5 years. Generally patients experiences mild pain [40%], moderate [39%] severe [16%] and no pain [5%] when recorded after 24 hours postoperatively. While after 48 hours the scores recorded for the same patients were as, mild [56%], moderate to severe [34%] and in the no pain group [10%] of patients were recorded

Conclusion: It is noted that absolute pain free surgeries are not possible at the moment, but with the use of latest advancements in the field of health sciences, post-operative pain can be minimized and should be the one of the prime goals for a good surgical outcome

Pancytopenia; study for clinical features and etiological pattern at tertiary care settings in Abbottabad

Pancytopenia is an important hematological problem encountered in our day-to-day clinical practice. The aim of our study was to evaluate clinical features and etiological pattern of pancytopenia at tertiary care settings in Abbottabad. This prospective study was conducted at Northern Institue of Medical Sciences [NIMS] and Ayub Teaching Hospital Abbottabad from 25th August 2009 to 31st July 2010. A total of 85 patients fulfilling the criteria of pancytopenia were randomly selected by time-based sampling. Pancytopenia was diagnosed by anemia [hemoglobin

Rapid emergence of ESBL producers in E. coli causing urinary and wound infections in Pakistan

Objectives: Production of extended spectrum beta -lactamases [ESBLs] by clinical isolates of pathogenic E. coli is a very serious therapeutic threat. This study was aimed to investigate the prevalence of ESBLs and associated drug resistance in E. coli isolates from urine and pus, and to report the drift from 2005 to 2009-10

Methodology: Among 173 E. coli isolates, 82 were phenotypically detected as ESBL producers by standard cefotaxime / clavulanic acid and ceftazidime / clavulanic acid disc diffusion tests. Antimicrobial resistance of all ESBL producers was assessed by disc diffusion method. Presence of CTX-M, TEM, SHV and OXA groups was investigated by PCR

Results: The prevalence of ESBL producing E. coli increased significantly from 33.7% in 2005 to 60.0% in 2009-10 [urine: 31.8% to 62.9%; pus: 41.1% to 55.5%]. Resistance to cefotaxime, ceftazidime, ciprofloxacin, gentamicin, nalidixic acid, ticarcillin-clavulanic acid, and trimethoprim-sulfamethoxazole was above 85% in both sets of isolates. Imipenem and Fosfomycin resistance was non-existent in 2005 but ranged from 3-15% in 2009-10. Remarkable increase from 9.5% to 64.7% in urinary tract isolates and from 0 to 55% in pus isolates was observed in colistin sulphate resistance. The dissemination of genes encoding ESBLs was: CTX-M 3.5%; TEM 10.7%; both CTX-M and TEM 3.5% in 2005, and CTX-M 42.5%; TEM 48.1%; both CTX-M and TEM 29.6% in 2009-10

Conclusions: Our results showed very rapid emergence of multidrug resistant ESBL producing E. coli in Pakistan posing a very serious threat in the treatment of nosocomial and community acquired infections

novel multiplex PCR for detection of pseudomonas aeruginosa: a major cause of wound infections

Wound infections are often difficult to treat due to various bacterial pathogens. Pseudomonas aeruginosa is one of the common invaders of open wounds. Precise diagnosis of this etiological agent in wound infections is of critical importance particularly in treatment of problematic cases. The existing diagnostic methods have certain limitations particularly related to specificity. Our objective was to establish a comprehensive and reliable multiplex PCR to confirm diagnosis of P. aeruginosa. A multiplex PCR test was developed for rapid and comprehensive identification of P. aeruginosa. Four highly specific genes were targeted simultaneously for detection of genus, species and exotoxin production [16S rDNA, gyrB, oprL and ETA] in P. aeruginosa; additionally one internal control gene invA of Salmonella was used. The specificity of the multiplex PCR was confirmed using internal and negative controls. Amplified fragments were confirmed by restriction analysis and DNA sequencing. The developed method was applied on 40 morphologically suspected P. aeruginosa isolates [from 200 pus samples] and 18 isolates were confirmed as P. aeruginosa. In comparison, only 12 could be identified biochemically. Combination of the four reported genes in multiplex PCR provided more confident and comprehensive detection of P. aeruginosa which is applicable for screening of wound infections and assisting treatment strategy

Clinical evaluation of safoof-e-darchini [Cinnamomum zeylanicum blum.] in primary hyperlipidaemia

A single blind standard controlled study was designed to evaluate the efficacy of Safoof-e-Darchini in the management of Primary Hyperlipidaemia. The study was conducted at ZVM College Pune. Before starting study ethical clearance was obtained from Ethical Committee. Sixty diagnosed patients were selected on the basis of inclusion criterion. Patients were randomly divided into two groups 'A' and 'B' comprising 30 patients in each group. In group 'A' test drug was given in a dose of 3 gram twice a day whereas in group 'B' standard drug Atorvastatin 20 mg was given once a day for the period of 60 days. Assessment of efficacy was done on the basis of objective parameters. In this study both test and control group exhibited significant hypolipidaemic activity [P<0.001]

Frequency and pattern of retinopathy in newly diagnosed type 2 diabetic patients at tertiary care settings in Abbottabad

Retinopathy is one of the most frequent and serious complication of diabetes mellitus and leading cause of blindness worldwide. The objective of this study was to determine frequency and pattern of retinopathy in newly diagnosed type 2 diabetic patients at tertiary care settings in Abbottabad. This was a descriptive cross-sectional comparative study being conducted at Ayub Teaching Hospital and Northern Institute of Medical Sciences Abbottabad. It included 100 newly diagnosed type 2 diabetic patients who were attending diabetic clinics of these tertiary care hospitals. Already diagnosed type 2 patients taking anti-diabetic medication, type 1 diabetes mellitus, hypertension, retinal vasculitis, retinal vessel occlusion, and sickle cell retinopathy were excluded. After pupillary dilatation, detailed fundoscopic examination was carried out via direct ophthalmoscopy and further confirmed by an ophthalmologist. According to fundoscopic findings, retinopathy was graded into background, pre-proliferative and proliferative types. Total 100 patients were included, with mean age 45.1 +/- 3.2 years, 60% of them were females. Overall, 17% of type 2 diabetic patients had retinopathy within one month of diagnosis. Background retinopathy was predominant [12%] followed by pre-proliferative [4%] and proliferative [1%] lesions. Frequency of retinopathy in newly diagnosed type 2 diabetic patients seems to be higher than previous reports and background lesions were predominant. Detailed fundoscopic examination of all newly type 2 diabetic patients at the time of diagnosis is of paramount importance.

Current status of fluoroquinolone and cephalosporin resistance in Salmonella enterica serovar Typhi isolates from Faisalabad, Pakistan

Currently fluoroquinolones and cephalosporins are at the forefront of typhoid treatment. The objective of this study was to assess the current drug resistance status of S. Typhi isolates from Faisalabad region by conventional and molecular methods. Drug resistance pattern of 30 clinical isolates [2011] against 8 drugs [nalidixic acid, ciprofloxacin, ofloxacin, gatifloxacin, cephradine, cefixime, ceftriaxone and cefpodoxime] was determined. MICs were noted by E-test. ESBL production was also tested. Relevant drug resistance genes bla[TEM], bla[OXA], gyrA, gyrB, parC, parE, qnrS and qnrA were targeted and QRDR regions of gyrA, gyrB, parC, and parE were sequenced for mutations. Nalidixic acid and ciprofloxacin resistance were seen in 30.0% and 10.0% of isolates respectively. No resistance was detected towards ofloxacin and gatifloxacin. Resistance to cephradine, cefixime, cefpodoxime and ceftriaxone was found in 46.7%, 13.3%, 16.7%, and 3.3% of isolates respectively. In ciprofloxacin resistant isolates a single mutation at codon Ser83 in gyrA gene was detected. A slow increase in ciprofloxacin resistance was indicated. However, newer fluoroquinolones ofloxacin and gatifloxacin are still very effective. Among cephalosporins, ceftriaxone showed promising results but emerging resistance was evident. Fortunately no ESBL producing isolate was detected. No correlation between two groups was detected in emergence of drug resistance

Knowledge, attitudes and practices of general practitioners towards diabetes mellitus in Hyderabad, Sindh

Objectives: To ascertain the knowledge, attitude and practice of general practitioners towards diabetes mellitus

Study Design: Cross sectional study

Duration of Study: From July 2009 to October 2009

Place of the study: The cases were collected from various areas of Hyderabad like Latifabad, Qasimabad, Hussainabad, Pretabad and Tando Jam, Hosrri

Material and Methods: One hundred general practitioners were selected in this study were excluded. A written questionnaire was filled by each doctor that included the 4 questions of each domain of knowledge, practice and attitudes. The knowledge was assessed by levels of blood glucose, random and fasting, practices assessed by treatment options and attitudes by advising to their patients for obesity, diet and pregnancy. The frequencies were taken of known or unknown

Results: out of 100 participants 35 belonged to periphery and 65 to major areas of Hyderabad. The mean +/- SD age was 41+/- 3.5 years and means +/- SD duration of practice was 10.4+/- 5.7 years. Each doctor had seen 10 diabetic patients per day. Out of 100 doctors, 55 answered correctly and 45 were poor to all domain

Conclusion: This study explored the knowledge, attitudes and practices regarding diabetes in Hyderabad among general practitioners and identified the need for improvement in managing diabetes. Awareness and education regarding diabetes is recommended to improve the health-care services provided by general practitioners

acute effects of Dokha smoking on the cardiovascular and respiratory systems among male university students

To assess the acute effects of dokha smoking on the cardiovascular and respiratory systems among male university students in the UAE. A quasi-experimental study was conducted during the months of November-December, 2009 among 97 male volunteers aged more than 17 years. Blood pressure, heart rate and respiratory rate of each participant, were measured before and immediately after smoking. A self administered questionnaire was used to collect personal details and data about smoking pattern. Mean increases in systolic bloc pressures [12 +/- mmHg], heart rates [20 +/- bpm] and respiratory rates [4 +/- 1 breaths/min] were observed [p< 0.001]. A mean decrease in diastolic blood pressures [1 +/- 1 mmHg] was observed [p = 0.483] smoking dokha has a significant acute effect on systolic blood pressure, heart rate and respiratory rate. Anti smoking campaigns must address the ill effects of this form of smoking. Results from the study warrant further research in this method of smoking which is becoming popular

Frequency and clinical presentation of dengue fever at tertiary care hospital of Hyderabad/Jamshoro

Objectives: The aim of this study is to observe the clinical presentation and frequency of dengue as a cause of fever in our setup

Methods: This retrospective study comprising of clinically suspected dengue infection, admitted to Liaquat University Hospital Hyderabad, during an epidemic from August 2006 to August 2007. Only adults with acute febrile illness were evaluated for clinical features of dengue fever, dengue hemorrhagic fever and dengue shock syndrome

Results: Patients with acute febrile illness were evaluated during this study. Fifty [5%] patients presented with typical features of dengue fever. Age of the patients ranged between 13 and 70 years. All patients were males with mean age of 35 years. Only 20/50 [40%] were dengue proven while 30/50 [60%] were dengue suspected. Out of dengue proven, 18 patients had dengue fever and 2 had dengue hemorrhagic fever. Typical clinical features included chills and rigors in 16 [80%], myalgia in 14 [70%], vomiting in 12 [60%], headache in 10 [50%], rash in 5 [25%]. Unusual clinical features were pharygitis in 7 [35%] and bleeding manifestations in 5% of patients. Laboratory investigations showed leucopenia [<4.0x10[9]/L] in 80%, thrombocytopenia [<150x10[9]/L] in 90%, and serum ALT was elevated [>40 U/L] in 40% cases

Conclusions: Fever associated with chills and rigors, bodyaches, bone pain, headache, myalgia, rash, low platelet count, decreased total leukocyte count, raised serum ALT, and hemorrhagic manifestations are satisfactory and important parameters to screen the cases of suspected dengue virus infection; however the diagnosis cannot be confirmed unless supported by molecular studies or dengue specific IgM

Nested PCR based diagnosis of salmonella enterica serovar paratyphi a directly from blood samples

Development of a rapid, reliable PCR - based method for molecular identification of Salmonella enterica serovar Paratyphi A directly from blood samples. S. Paratyphi A isolates were used for regular PCR targeting specific region of fliC-a gene. New primers were designed and conditions were optimized for a nested PCR that could be directly applicable on blood samples. The procedure was tested on 70 blood samples from suspected cases of typhoidal infection and comparison made with blood culture. Blood culture was able to diagnose only four patients as infected with S. Paratyphi A. Regular PCR was unable to detect S. Paratyphi A directly from blood where as nested PCR detected S. Paratyphi A in blood of thirteen patients. S. Paratyphi A, which is emerging as a major pathogen can be detected with better sensitivity by nested PCR as compared with blood culture

Study of snake bite cases at Liaquat University Hospital Hyderabad/Jamshoro

Envenoming resulting from snakebites is an important public health hazard in many regions. It is common in rural areas not to delay access to life saving anti-venom. The objectives of this study were to know about common types of snakes in local areas, clinical features in snakebite victims, complications in snakebite case, and mortality rate in snakebite victims in rural Sindh. This descriptive study was conducted at 4 medical wards of Liaquat University Hospital Hyderabad/Jamshoro, Sindh from 1st January 2006 to December 2006. One hundred cases with history of snakebite were analysed. Both genders were included in study. Patients with history of trauma, insect bite or thorn prick were excluded from the study. Clotting time [CT] was the main bedside procedure, to assess the degree of envenomation. One hundred [100] cases from both genders, from 8 to 55 years age were reviewed. There were 57 [95%] viper bites [haemotoxic] having haemostatic abnormalities and 3 [5%] elapid [neurotoxic] bites presented with neuroparalytic symptoms. Most cases were from Tando Mohammad Khan and Hyderabad [rural] districts of Sindh. All victims had localized oedema at the site of bite. Fang/teeth marks were noted in [90%] cases. Majority [80%] were bitten on the legs below knee. Some 40% of the cases of snakebite occurred when the patient was asleep. Urban to rural ratio was 1:4.5 and male to female ratio was 4:1. Mean time to arrival at our hospital after the bite was 3 hours and mean duration of hospital stay was 4 days. One patient had acute renal failure [ARF] and disseminated intravascular coagulation [DIC], 3% cases of elapid bites were shifted to ICU for assisted ventilation, 4 patients [5.5%] had adverse effects after anti-venom administration and needed intravenous hydrocortisone, promethazine and subcutaneous adrenaline. The average dose of anti-venom was 60 vials for viper bites and 10 vials for elapibites. Overall mortality rate was 4%. Snakebites are common in the rural population of developing countries. There is need to educate the public about the hazards of snake bite, early hospital referral and treatment

Significant incidence of typhoid in hepatitis C patients

Only proven way of transmission of Hepatitis C is through blood. The origin is unknown in nearly half the cases. Pollution is suspected as a cause but it is impossible to prove this relationship directly. We thought that typhoid being a proven pollution related disease, determination of its confection in Hepatitis C patients representing same Socio-economic group would be of interest. A typhoid in Hepatitis C patients can easily be overlooked because symptoms like fever and abdominal discomfort are present in both diseases. Blood samples were collected from three groups of study as mentioned in materials and methods. These samples were processed for 4[th] generation HCV ELISA. PCR for HCV, PCR for typhoid. Blood culture for typhoid and widal test as required [details are given in methodology]. Finally the data thus obtained was analysed and conclusions were drawn. Health Biotechnology Division, National Institute for Biotechnology and Genetic Engineering [NIBGE] and Millat Laboratory Faisalabad. April 2004 to Oct 2004. The parameters included were PCR, blood culture and widal test. There were three groups of study, PCR and ELISA positive patients of Hepatitis C [105] - further subdivided into two groups, with history of exposure to known causes of spread of HCV in last one year [65] and those without such history [40]; clinically diagnosed cases of typhoid [30]; and healthy controls [50]. In the three groups, PCR was positive in 9.5[7.7 and 12.5], 63.3, and 2.0% cases respectively. Figures for blood culture were 4.7[3.1 and 7.5], 33.3, and 0% in the same order, and the respective figures for widal test were 34.2[33.8 and 35.0], 56.6, and 24.0%. The increase in PCR and blood culture positivity in Hepatitis C cases as compared with normal subjects is statistically significant [P< 0.05]. These results clearly suggest that the source of infection for the two diseases is same in many cases, and therefore, provides a strong indication of a relationship between pollution and Hepatitis C

Left ventricular ejection fraction [LVEF] determination in post myocardial infarction patients. A comparison of gated blood pool [GBP] ventriculography and contrast ventriculography

To compare the efficacy of gated blood pool versus contrast ventriculography [Gold standard] for determination of ejection fraction in post myocardial infarction patients. Patients and Gated blood pool imaging [BPI] was performed by gated blood pool radionuclide ventriculography or MUGA in 30 patients within 48 hours after single plane [RAO-30] contrast ventriculography. Statistical analysis was done by student "t-test" [unpaired] and by Pearson's correlation co-efficient. Result: There was no significance difference in their mean ejection fraction [44.04 +/- 11.93 Vs 48.46 ' 12.78%, P = NS] and has statistically significant correlation [r=0.86, P=0.171] GBP ventriculography can therefore be utilized accurately for determination of ejection fraction for prognostic and diagnostic evaluation of post myocardial infarction patients

Early detection of typhoid by polymerase chain reaction

Typhoid is a common problem in developing countries. Cultivation of bacteria and serology [especially Widal test] give unacceptable levels of false-negative and false-positive results, respectively. Patients and In this study, a recently introduced polymerase chain reaction-based technique [which has 100% specificity for Salmonella typhi] was compared with blood culture and Widal test during the first week of illness of 82 suspected cases of typhoid. The respective figures of positivity for PCR, blood culture and Widal test were 71.95%, 34.1%, and 36.5%. A control group of 20 healthy persons gave figures of 0%, 0%, and 33.3%, respectively. We conclude that this PCR-based technique is not only absolutely specific, but also very sensitive and, therefore, much superior to blood culture and Widal test for the early diagnosis of typhoid

Measuring national stereotypes

The study makes an attempt to review the various techniques of measuring national stereotypes in the hope of stimulating young social scientists, particularly of developing countries to conduct researches on the subject which has much interdisciplinary significance and value. The techniques that have been discussed are adjective checklist, free association method content analysis semantic differential scale, percentage attribution method and stereotype differential techniques. In addition to these commonly used techniques. The methods of cross- cultural comparison across two or more cultures and historical comparison within a culture have been discussed. The paper suggests the merits of classical adjective checklist technique, semantic differential scales and content analysis for the study of national and international images