Diagnostic Validity of Cica Beta Test 1 for the Detection of Extended Spectrum Beta-Lactamase (ESBL) Producing Gram Negative Bacteria by Comparing with Phenotypic Method

Background: Detection of extended spectrum beta lactamase producing bacteria is an important issue in the clinical settings. Objective: The purpose of the present study was to validate the Cica Beta Test 1 for detection of extended spectrum beta-lactamase (ESBL) producing bacteria. Method: This analytical type of cross-sectional study was carried out in the Department of Microbiology and Immunology at Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka from January 2006 to December 2006 for a period of one (01) year. All the patients presented with the clinical features of urinary tract infection and surgical as well as burn wound infection at any age with both sexes were selected as study population. All bacteria were isolated and identified by their colony morphology, staining characters, pigment production, motility and other relevant biochemical tests. Phenotypic confirmation of ESBLs producing isolates were done by inhibitor potentiated disc diffusion test according to CLSI recommendation. The Cica Beta Test 1 was performed according to the manufacturer’s instructions. Result: A total number of 288 Gram negative bacteria were isolated. Among these isolates Cica Beta test 1 was positive in 97 strains and phenotypic confirmatory test was positive in 89 strains. The test sensitivity of Cica Beta Test 1 was 100% (95% CI 95.9% to 100.0%). Specificity of the test was 96.0% (95% CI 92.2% to 98.2%). The positive predictive value (PPV) and negative predictive value (NPV) were 92.7% (95% CI 84.5% to 95.7%) and 100.0% (95% CI 98.0% to 100.0%) respectively. The accuracy of the test was 97.2% (95% CI 95.1% to 99.1%). Area under ROC curve = 0.980 (95% CI 0.964 to 0.996);p value 0.0001. Conclusion: In conclusion, Cica Beta Test 1 is very high sensitivity and specificity for the detection of ESBL from Gram negative bacteria.

Multiple Antibiotic Resistant Index of Gram-Negative Bacteria from Bird Droppings in Two Commercial Poultries in Enugu, Nigeria

Antimicrobial resistance refers to the ability of microorganisms to grow in the presence of an antimicrobial agent at a concentration that will normally kill or inhibit their growth. Antimicrobial resistance has become a major global threat making treatment of infections tougher especially with high cost of treatment in humans and animals. This study was done to determine the Multiple Antibiotic Resistant Index (MARI) of Gram-negative bacteria from bird droppings in two commercial poultries in Enugu. Forty (40) samples were collected from each of the poultries. Isolates were identified by standard microbiological methods. The isolates identified were Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Citrobacter spp, Proteus spp. and, Enterobacter spp. Antibiotic susceptibility testing was carried out using disc diffusion technique. The organisms were tested against pefloxacin, augmentin (amoxicillin and clavulanic acid), ceftazidime, streptomycin, ciprofloxacin, gentamycin, cephalothin, neomycin and ofloxacin. The result of the susceptibility test showed that Proteus spp had the highest resistance and MARI value of 0.5 and 1.0 respectively. The other MARI values were Escherichia coli (0.9), Klebsiella pneumonia (0.9), Pseudomonas aeruginosa (0.8), Citrobacter spp (0.8) and Enterobacter spp (0.7). These results suggest that bacterial organisms from poultry source can contribute significantly to the spread of multi-antibiotic resistant organisms. This could arise from the indiscriminate use of antibiotics in bird feeds in poultries.

Occurrence of tetracycline residues and antimicrobial resistance in gram negative bacteria isolates from cattle farms in Spain

The usage of antibiotics in animal husbandry has dramatically increased the concentration of antibiotic residues and has promoted the development and abundance of antibiotic resistance in manure. When it is spread onto agricultural land, both residues of antibiotics and bacteria carrying antibiotic resistance genes may be introduced into the environment. In this research, we isolated resistant gram negative bacteria from manure produced in two dairy and two beef cattle farms, located in Madrid (Spain), to determine their resistance to seventeen representative antibiotics commonly used in veterinary therapy. A total of 63 isolates were used to assess the overall bacterial antimicrobial resistance on cattle manure samples. Predominant species were Escherichia coli and Comamonas testosteroni accounting for 25% and 19.6% of the total, respectively. The most found antimicrobial resistances in gram-negative bacteria were to tetracycline (66.7%), sulphamethoxazole (55.6%), ampicillin (52.4%), cephalothin (46.0%), chloramphenicol (44.4%), nalidixic acid (39.7%) and trimethroprim- sulphamethoxazole (33.3%). The mean of resistance and the percentage of multi-resistant bacteria in beef farms were higher and statistically significant when compared to dairy farms which is opposite from the findings of the previous studies. The presence of three tetracyclines in all manure samples was also examined with stable recoveries (76% - 82%) and high sensitivity (limit of quantification 0.015 - 0.03 μg/kg). The concentrations of tetracyclines detected (<0.015 - 10 mg/kg) were consistent to the theoretical tetracycline levels in manure in Spain according to the ex- cretion rate of these antibiotics and the values re-ported in scientific literature in other European coun- tries.

Review: Gram Negative Bacteria in Brewing

Gram negative aerobic bacteria such as Acetic Acid Bacteria, which include Acetobacter and Gluconobacter, have historically caused significant problems to brewers. Although incidences of spoilage have recently reduced as a result of improvements in beer packaging, these bacteria are still a concern in dispense systems in pub breweries, public houses and cask conditioned beers. Gram negative facultative bacteria of the genus Zymomonas can spoil primed cask conditioned beer and cider. There is a wide range of Enterobacteraeceace which are found within brewery environments and they serve as indicator microorganisms for hygiene and sanitation. Gram negative strictly anaerobic bacteria such as Pectinatus and Megasphaera have recently emerged as a significant threat due to the improvement in reduction of oxygen levels in beer and an increase in production of unpasteurised beer. Pectinatus and Megasphaera are sensitive to routine cleaning agents used in breweries, but they can survive and proliferate in biofilms eventually causing spoilage of beer. This review focuses on Gram negative aerobic, facultative anaerobic and strictly anaerobic brewery related spoilage bacteria.

Meso-substituted cationic 3-and 4-N-Pyridylporphyrins and their Zn(II)derivatives for antibacterial photodynamic therapy

Photodynamic inactivation of microorganisms known as antibacterial photodynamic therapy(APDT)is one of the most promising and innovative approaches for the destruction of pathogenic microorganisms.Among the photosensitizers(PSs),compounds based on cationic porphyrins/metalloporphyrins are most successfully used to inactivate microorganisms.Series of meso-substituted cationic pyridylporphyrins and metalloporphyrins with various peripheral groups in the third and fourth positions of the pyrrole ring have been synthesized in Armenia.The aim of this work was to determine and test the most e®ective cationic porphyrins and metalloporphyrins with high photoactivity against Gram negative and Gram positive microorganisms.It was shown that the synthesized cationic pyridylporphyrins/metalloporphyrins exhibit a high degree of phototoxicity towards both types of bacteria,including the methicillinresistant S.aureus strain.Zinc complexes of porphyrins are more phototoxic than metal-free porphyrin analogs.The e®ectiveness of these Zn–metalloporphyrins on bacteria is consistent with the level of singlet oxygen generation.It was found that the high antibacterial activity of the studied cationic porphyrins/metalloporphyrins depends on four factors:the presence in the porphyrin macrocycle of a positive charge(+4),a central metal atom(Zn2þÞand hydrophobic peripheral functional groups as well as high values of quantum yields of singlet oxygen.The results indicate that meso-substituted cationic pyridylporphyrins/metalloporphyrins cannd wider application in photoinactivation of bacteria than anionic or neutral PSs usually used in APDT.

Isolation and Identification of Multi-Drug Resistant Strains of Non-Lactose Fermenting Bacteria from Clinical Refuses in Major Hospitals of Khyber Pakhtunkhwa, Pakistan

Purpose: The present studied was performed in order to investigate the drug resistance of different non-lactose fermenting gram negative bacteria from clinical isolates. The bacteria were morphologically characterized through culturing and gram staining techniques were used for the identification of different bacterial strains. Methods: A total of 324 samples were collected from patients, after they were diagnosed by physicians at different hospitals at district Peshawar. Samples were morphologically identified by blood agar, MacConkey agar and Eosine Methylene Blue, identified by gram staining techniques. Modified Kirby-Bauer Disc diffusion method was used to test the in-vitro susceptibility of the identified isolates to different antibiotics. Results: The non-lactose fermenting gram negative bacteria were isolated from samples of blood (33.30%), pus/ wound (33.30%), urine (23.30%) and from ascetic/pleural fluids (10.20%). The study revealed that Pseudomonas aeroginosa showed high resistance against Gentamicin (74%) and Aztreonam (74%), followed by Ciprofloxacin (59.20%) and Amikacin (33.30). Tazocin was active as low resistance (18.50%) is shown. More resistance was seen in Morganella morganii against Aztreonam (77.7%) followed by Gentamicin (62.90%), Ciprofloxacin (40.70%). Tazocin show low resistance (3.70%). Multidrug resistant Proteus mirabillis was highly resistance to Gentamicin (66.60%), followed by Aztreonam (62.90%), Amikacin (55.50%), Ciprofloxacin (40.20%) and low resistance to Tazocin was (22.20%). Salmonella typhi demonstrated high resistance against Amikacin (62.90%), followed by Aztreonam (48.10%), Tazocin (40.70%). Gentamicin showed low resistance (29.60%), and hence it is more active against S. typhi. Conclusions: It can be concluded from the present study that different species of non-lactose fermenting gram negative bacteria have shown a different resistivity pattern. This study is a gate way for better and suitable management strategy for the infections caused by non-Lactose fermenting bacteria in the sampling region.

Isolation and Identification of Multi-Drug Resistant Strains of Non-Lactose Fermenting Bacteria from Clinical Isolates

Purpose: We studied the drug resistance of different microbes from clinical isolates. The morphological characteristics of bacteria were observed through culture characteristics and by carrying out gram staining techniques while the biochemical characteristics of bacteria were carried out by biochemical test. Methods: A total of 324 samples were collected from suspected patients visiting different hospitals at district Peshawar. For morphological identification, samples of clinical isolates were analyzed by blood agar, MacConkey agar and Eosine Methylene Blue, identified by gram staining and characterized by different biochemical tests. Antibiotic Sensitivity test by Modified Kirby-Bauer Disc diffusion method was used to test the in-vitro susceptibility of the identified isolates to different antibiotics such as Ceftazidime, Ceftazidime, Ceftriaxone, Cefepime and Imipenem. Results: These resistant non-lactose fermenting gram negative bacteria were isolated from samples of pus/wound (33.30%, n = 108/324), blood (33.30%, n = 108/324), urine (23.30%, n = 75/324) and from ascetic/pleural fluids (10.20%, n = 33/324). The study revealed that the percentage of non-fermenting bacterial infection was higher in females (53%) as compared to males (47%) along with higher infection observed in the age group of 11 - 30 years. Pseudomonas aeroginosa showed high resistance against Cefepime (88.80%), followed by Cefoperazone (55.50%), Ceftazidime (48.10%), Ceftriaxone (33.30%). Imipenem was active with low resistance (7.40%). More resistance was seen in Morganella morganii against Imipenem (66.70%) followed by Cefoperazone (55.50%), Ceftriaxone (55.50%). Cefepime showed low resistance (11%). Multi-drug resistant Proteus mirabillis was highly resistance to Ceftriaxone (74.07%), followed by Cefepime (59.20%), Cefoperazone (44.40%) and low resistance for Imipenem (25.90%). Salmonella typhi demonstrated high resistance against Imipenem (74.07%), followed by Ceftriaxone (40.70%), Ceftazidime (37.03%). Cefepime showed low resistance (3.70%), hence it is more active against S. typhi. Conclusions: The different species of non-lactose fermenting gram negative bacteria have shown a different resistivity pattern in the present study. Therefore identification of non-lactose fermenting gram negative bacteria and looking after their resistivity/susceptibility pattern are important for suitable management of the infections caused by them.

In vitro antibacterial effect of aqueous and ethanolic Moringa leaf extracts

Objective:To evaluate the antibacterial effect of aqueous and ethanolic moringa leaf extracts (Moringa oleifera) on the growth of gram-positive and negative bacteria.Methods:Paper disks were soaked with 100,200,300 and 400μL of extract at 20 g/180mL and 10 g/190 mL.All extracts were tested against Escherichia coli(ATCC25922),Staphylococcus aureus(ATCC25923),Vibrio parahaemolyticus,Enterococcus faecalis(ATCC29212),Pseudomonas aeruginosa(ATCC27853), Salmonella enteritidis(IH) and Aeromonas caviae.The susceptibility tests were performed using the modified disk diffusion method.Results:The strains E.coli,P.aeruginosa and S.enteritidis (IH) were resistant to all treatments.In general,disks with 400μL extract were the most efficient against S.aureus,V.parahaemolyticus,E.faecalis and A.caviae.Conclusions:The study indicates a promising potential for aqueous and ethanolic Moringa leaf extracts as alternative treatment of infections caused by the tested strains.

Synthesis, Structure Analysis and Antibacterial Activity of New Potent Sulfonamide Derivatives

Modification of sulfonamid drug using different principles of chemical reactions was investigated. These reactions involve the condensation of an amino group with triethyl orthoformate and dimethylformamide dimethyl acetal. Ability of sulfa to condense with active keto compounds, like ethyl pyruvate and piprazine carboxyaldehye was studied. Alkyation of sulfa with different chloro derivatives was also reported. The structure of the isolated compound was elucidated and confirmed using elemental analysis and spectral data. The bioactivity of the ob-tained compounds was investigated against different gram positive and gram negative bacteria. The study reveals that most of the modified drugs show high to moderate antibacterial activity.

Sterilization of Turmeric by Atmospheric Pressure Dielectric Barrier Discharge Plasma

In this study atmospheric pressure dielectric barrier discharge （DBD） plasma has been employed for sterilizing dry turmeric powders. A 6 kV, 6 kHz frequency generator was used to generate plasma with Ar, Ar/Ou, He, and He/O2 gases between the 5 mm gap of two quartz covered electrodes. The complete sterilization time of samples due to plasma treatment was measured. The most important contaminant of turmeric is bacillus subtilis. The results show that the shortest sterilization time of 15 rain is achieved by exposing the samples to Ar/O2 plasma. Survival curves of samples are exponential functions of time and the addition of oxygen to plasma leads to a significant increase of the absolute value of time constant of the curves. Magnitudes of protein and DNA in treated samples were increased to a similar value for all samples. Taste, color, and solubility of samples were not changed after the plasma treatment.

Antibacterial Action of Chemically Synthesized and Laser Generated Silver Nanoparticles against Human Pathogenic Bacteria

Silver nanoparticles in the range of 10-40 nm were synthesized chemically and by laser ablation, em- ployed for in vitro antibacterial action against human pathogenic bacterium. Their formation was evidenced by UV-visible spectrophotometer; particle size confirmed by atomic force microscopy, crystal structure determined by X-ray diffraction and chemical composition investigated by Fourier transform infrared spec- troscopy. The calculated MIC （minimum inhibitory concentration） of chemically synthesized nanoparticles with 30-40 nm in size are 2.8 μg/mL 4.37 μg/mL 13.5μg/mL and 2.81 p.g/mL for E. coil, S. aureus, B. subtillis and Salmonella, respectively. Whereas laser ablated nanoparticles exhibit MIC of 2.10 μg/mL 2.36 μg/ mL and 2.68 μg/mL for E. coli, S. aureus and Salmonella, respectively.

Regulating dual-use research:Lessons from Israel and the United States

Over the past 20 years,and with greater urgency since the H5N1 papers controversy(2012),dual-use research of concern(DURC)has been extensively debated by the scientific community,regulators,and the public at large.Many have raised the need to regulate DURC;however,despite the urgency of this issue,to date,only two countries have implemented specific regulations,United States and Israel(notably,China is in the final stages of creating a set of DURC-related regulations).In this paper,I have compared these policies as case studies that may benefit other countries.Therefore,the present study aimed to critically compare the DURC policies of the US and Israel and determine whether their different approaches provide a rigorous framework for the regulation of DURC.I have shown that in some aspects,they have adopted similar approaches;for example,allowing local committees to play a role in assessing the research.However,they differ in certain important aspects;for instance,the US policies have laid out a mechanism through which academic institutions and the government decide jointly on how to address risky studies,while the Israeli law places the authority solely in government hands.My aim in comparing the policies is twofold;first,to reveal the weaknesses and strengths of each policy,thereby aiding their improvement,and second,to provide those that are considering developing policies in this field an overview of what the current approaches look like,thereby enabling them to articulate their own approach while taking into account the experience gained by the US and Israel.

Development of designated laboratories under the United Nations Secretary-General’s Mechanism

The United Nations Secretary-General’s Mechanism(UNSGM)for investigating the alleged use of biological weapons was proposed in early 2013,with support from the World Organization for Animal Health(OIE)and World Health Organization(WHO).1 The UNSGM and related documents call for designated laboratories to support an investigation of the alleged use of biological weapons.Designed laboratories participating in the UNSGM investigations must not report false results,positive or negative.It is not affordable for whatever reason.

Surveillance of antibiotic sensitivity pattern in chronic suppurative otitis media of an Indian teaching hospital

Introduction:Chronic suppurative otitis media (CSOM) is a common problem in worldwide and untreated CSOM leads to fatal complications like facial nerve paralysis,lateral sinus thrombosis,labyrinthitis,meningitis and brain abscess in developing country like India.Objective:To isolate causative bacteria and antibiotic sensitivity pattern for CSOM and to know the prevalence of extended spectrum beta lactamases (ESBL) and Metallobetalactamases (MBL) in CSOM patients.Methods:A total of 500 ear swabs of clinical suspected CSOM patients were cultured on specific cultured medium and identified the bacteria with conventional methods.Then all the identified bacteria were subjected with specific antibiotics by the Kirby-Bauer's method to know the resistance pattern of antibiotics.ESBL and MBL strains were detected by double disc diffusion test.Results:A total of 384 bacteria were isolated from 500 CSOM patients,among them 86 P.aeruginosa (22.40%),112 Staphylococcus aureus (29.17%),53 A.baumannii (13.80%),32 E.aerogenes (18%),26 C.freundii (6.77%),24 K.oxytoca (6.25%),23 P.vulgaris (5.99%),18 K.pneumoniae (4.69%) and 10 P.mirabilis (2.60%) identified with conventional methods.From antibiotic disc diffusion methods 74.22% ESBL strains and 9.90%MBL strains were documented.Multidrug resistant strains of P.aeruginosa (86/384,22.40%) were more prevalent than those of S.aureus (112/384,29.17%) and other bacteria in ear discharges.Imipenem and vancomicin could control to gram negative bacteria and gram positive bacteria respectively.Conclusion:Continuous and periodic evaluation of microbiological profile and antimicrobial sensitivity pattern of bacterial is essential for optimum management of CSOM patients.

Assessment and impact of a biosecurity course in raising the awareness of students at the Jinnah post graduate Medical Center,Pakistan

Introduction:Pakistan Biological Safety Association(PBSA)is a not for profit,nongovernmental,professional organization established in 2008 and is striving to promote and implement Bio risk management in Pakistan.PBSA accelerated its program for strategic training of the national life scientists from 2013 and is still working on that mandate with the help of Fogarty International Center(FIC)&National Institute of Health USA and other international organizations working on this Global Health Security Initiative and subsequently Agenda(GHSA)Aims&Objectives:This biosafety training program was developed for capacity building of M.Phil.and PhD graduates the future laboratory users of Basic Medical Sciences Institute(BMSI),Jinnah Post Graduate Medical Center which is the largest tertiary care hospital in Karachi,Pakistan involved in Research&Development(R&D)in medical sciences.Materials&Methods:This Pre post study was carried at Basic Medical Sciences Institute(JPMC),during the period of October 2018 to January 2019.A total of 48(16 PhD.32 M.Phil.)Grads students participated in the following study.The training program is administered through,custom made 4-month course having 16 contact hours also 4 hours of practical training.Training was evaluated using written assessment.The pre and post-training test was conducted after each session.Each test comprised of 15 multiple choice questions(MCQs)pre and post session making it 30 MCQs in total for each session.A final assessment consisting 30 MCQs conducted at the end of course.Results:There was vast difference observed in the post assessment results as compared to the pre assessment results.At the end of the course final assessment was carried out which gave an overall impression of vast change(83.3%)in awareness regarding bio risk management among the graduate students,which reflected the efficacy of this training.To ensure the validity of results and the intention to improvise the quality of course in future,feedback regarding the course and trainer was obtain through Likert scale.Reliability of Likert scale carried through Cronbach’s alpha,showed all variables with a maximum score of 0.9,indicating high reliability of the generated feedback evaluation.Conclusion:This study highlighted that through concerted efforts of professional organizations,proper awareness of public and concerned personals,regarding Biosafety&Biosecurity can certainly reduce the chances of errors and will ensure potential safety and security to the laboratory workers,community and environment,potentially exposed to these pathogens&toxins which are regarded as biohazards.

A Survey on Biosafety Practices in Lab Personnel in 12 Selected Areas of Karachi,Pakistan

The objective of this study was to realize the degree of biosafety precautions and measures recorded among different Private Hospitals and Diagnostic labs personnel situated in different areas of Karachi,Pakistan.One thirty two private hospital based laboratory technicians were selected from all 12 selected areas in Karachi.The laboratory technicians were interviewed then a survey form was completed by the interviewer.The survey was conducted in 3 months duration from May 2017 to July 2017.The total number of members in this study was 132,of which 85 were males and 47 were females.Results presented that 65%of the laboratory technicians did not use any type of PPE and 35%of the respondents recapped used syringes frequently while 25%recapped occasionally.For avoiding the reuse of syringes,Pakistani governments recommend that they be cut before discarding;however,only 65%of the respondents followed the given procedure.Even though mouth pipetting is considered outdated,45%of the technicians continue to do so for numerous purposes.Results of this survey established an absence of awareness about good and proper lab practices and lab biosafety measures among lab technicians in Karachi,Pakistan.