Wednesday, May 30, 2012

Demonstration of Bacterial Biofilms in Chronic Otitis Media


Objective: To establish the presence of biofilms in surgical tissue specimens from patients with chronic otitis media.

Material and Methods: 22 patients with chronic otitis media scheduled for tympanomastoid surgery were enrolled in this study between September 2007 and January 2008. Biopsies of the middle ear mucosa and cultures were taken at the time of surgery. Tissues were cultured using conventional methods for Haemophilus influenzae, Pseudomonas
aeruginosa, Streptococcus pneumoniae and Staphylococcus aureus. Bacteria identification was performed using the Becton Dickinson automatic identification system. Slime forming ability was tested on congo red agar for culture positive bacteria. The presence of icaA and icaD DNA
were detected by polymerase chain reaction using forward and reverse primers for icaA and icaD for staphylococcus aureus.

Results: 6 of 22 patients’ tissue specimens were culture-positive(72.7%). 5 Staphylococcus aureus and 11 Pseudomonas aeruginosa were identified on 16 specimens. Bacterial biofilms were present on 9 of 16(56.2%) culture-positive specimens. 2 of 5(40%) Staphylococcus aureus and 7 of 11(63.6%) Pseudomonas aeruginosa produced bacterial biofilms.

Conclusion: Pseudomonas aeruginosa was the most commonly bacteria in chronic otitis media. Biofilm forming ability was higher in Pseudomonas aeruginosa compared with other bacteries. The presence of biofilms on the mucosa of patients with chronic otitis media offers a possible cause of antimicrobial therapy failure.

source:  Mediterr J Otol 2008; 4: 64-68

Friday, May 18, 2012

Polymerase Chain Reaction (PCR) a diagnostic tool.

What is PCR?

Sometimes called "molecular photocopying," the polymerase chain reaction (PCR) is a fast and inexpensive technique used to amplify, or make many copies of, small segments of DNA. This is necessary because methods used for analyzing DNA (determining the DNA base pair sequence) require more DNA than may be in a typical sample. A particularly useful feature of PCR is that it allows the amplification process to be limited to specifically targeted segments of the DNA mixture--such as the Y chromosome markers used in genealogical testing.
When your vials of cheek cells arrive at the lab for testing, they are first mixed with a detergent which causes the cells to burst open and release their DNA along with other cell contents. The mixture is then washed with a phosphate containing buffer (mild salt) solution to dilute cellular debris. With minimal preparation, the sample is ready and the DNA on the targeted area of a chromosome can be amplified.

How does PCR work?

PCR is a process based on the ability of a DNA polymerase enzyme that can synthesize a complementary strand to a targeted segment of DNA in a test tube mixture of the four DNA base. In addition, the mixture must also contain two DNA fragments, each about 20 bases long, called primers, that have sequences complementary to areas adjacent to each side of the target sequence. (To do PCR, you need to know the DNA sequence around the region you want to amplify.) These primers can be constructed in the lab, or purchased from commercial suppliers. If chosen well, the 20-25 base pair sequence will be unique in the entire human genome so will match only the place specifically chosen thus limiting and defining the area to be copied.
The mixture is first heated to denature (separate) the sides of the double- stranded DNA and then cooled to allow (1) the primers to find and bind to their complementary sequences on the separated strands and (2) the polymerase to extend the primers into new complementary strands. Repeated heating and cooling cycles multiply the target DNA exponentially, since each new double strand separates to become two templates for further synthesis. In about 1 hour, 20 PCR cycles can amplify the target by a millionfold. In 32 cycles at 100% efficiency, 1.07 billion copies of targeted DNA region are created.

From: The National Human Genome Research Institute Office of Science Education and Outreach

The entire cycling process of PCR is automated and can be completed in just a few hours. It is directed by a machine called a thermocycler, which is programmed to alter the temperature of the reaction every few minutes to allow DNA denaturing and synthesis. To avoid the destruction of needed enzymes in the mixtures by the high temperatures needed to denature the DNA, enzymes from bacteria that thrive in hot springs are used for the process.

Why is PCR useful?

Once amplified, PCR products can be used in many different laboratory procedures; for example, most mapping techniques in the Human Genome Project rely on PCR.
PCR is also valuable in a number of newly emerging laboratory and clinical techniques, including DNA fingerprinting, detection of bacteria or viruses (particularly AIDS), and diagnosis of genetic disorders and preparing samples for genealogical DNA testing.


Thursday, May 17, 2012

Tips for Effective PowerPoint Presentations

  1. Select sans-serif fonts such as Arial or Helvetica.  Avoid serif fonts such as Times New Roman or Palatino as they are sometimes more difficult to read.
  2. Use no font size smaller than 24 point.
  3. Clearly label each screen.  Use a larger font (35-45 points) or different color for the title.Use a single sans-serif font for most of the presentation.  
  4. Use different colors, sizes and styles (bold, underline) for impact.
  5. Avoid italicized fonts as they are difficult to read quickly.
  6. No more than 6-8 words per line.
  7. For bullet points, use the 6 x 6 Rule.  One thought per line with no more than 6 words per line and no more than 6 lines per slide.
  8. Use dark text on light background or light text on dark background.  However, dark backgrounds sometimes make it difficult for some people to read the text. 
  9. Do not use all caps except for titles.
To test the font, stand back six feet from the monitor and see if you can read the slide.

Graphics and Design
  1. Keep the background consistent and subtle.
  2. Use only enough text when using charts or graphs to explain clearly label the graphic.
  3. Keep the design clean and uncluttered.  Leave empty space around the text and graphics.
  4. Use quality clipart and use it sparingly.  The graphic should relate to and enhance the topic of the slide.
  5. Try to use the same style graphics throughout the presentation (e.g. cartoon, photographs).
  6. Limit the number of graphics on each slide.
  7. Check all graphics on a projection screen before the actual presentation.
  8. Avoid flashy graphics and noisy animation effects unless they relate directly to the slide.
  9. Limit the number of transitions used.  It is often better to use only one so the audience knows what to expect.
  1. Limit the number of colors on a single screen.
  2. Bright colors make small objects and thin lines stand out.  However, some vibrant colors are difficult to read when projected.
  3. Use no more than four colors on one chart.
  4. Check all colors on a projection screen before the actual presentation. They may project differently than what appears on the monitor.
General Presentation
  1. Check the spelling and grammar.
  2. Do not read the presentation.  Practice the presentation so you can speak from bullet points.  The text should be a cue for the presenter rather than a message for the viewer.
  3. Give a brief overview at the start.  Then present the information.  Finally review important points.
  4. It is often more effective to have bulleted points appear one at a time so the audience listens to the presenter rather than reading the screen.
  5. Use a wireless mouse or pick up the wired mouse so you can move around as you speak.
  6. If sound effects are used, wait until the sound has finished to speak.
  7. If the content is complex, print out the slides so the audience can take notes.
  8. Do not turn your back on the audience.  Try to position the monitor so you can speak from it.


Why study Microbiology?

Microbiology is the study of organisms of microscopic size, including bacteria, protozoa, viruses, and certain algae and fungi which affect every aspect of life on Earth. They have amazing diversity of form and can live in a wide range of habitats ranging from hot springs to the human body and the depths of the ocean. Although some microbes cause diseases, like measles, meningitis or AIDS, the majority are completely harmless. In fact these small life forms are essential to the cycling of nutrients in the ecosystems of the planet. If these processes did not occur, life on this planet would soon grind to a halt.
Activities of microbes can be harnessed in many ways to benefit humans, other animals, plants and the environment. Food, healthcare, chemical, and waste treatment industries rely heavily on the powers of microbes.
Microbiology is a vast subject which overlaps with other life sciences such as genetics, biochemistry, molecular biology and evenengineering. Microbiologists can be found at work in many different places, but they are normally based in a laboratory.
As there are many different types of microbes there are many different types of microbiologists: bacteriologists, mycologists (who study fungi) and virologists - all working within even smaller areas of specialisation; the variations are endless!

Obviously you need to be interested in science and biology. An enquiring mind, a methodical approach and an enthusiasm for solving problems are equally important. You should be a good communicator, as you will need to describe your findings clearly to other people, and be able to work well as a part of a team. Scientists today seldom work alone and most are members of multi-disciplinary groups. In industry you will also have to liaise with staff from non-scientific departments.



Food, pharmaceutical, agrochemical, biotechnological, biorefinery, environmental, pollution control and bioremediation, companies all need microbiologists to develop new products, monitor the production of existing ones and solve problems. 
In the Field
Agriculture - environmental and health specialists study the role of microbes in plant disease, pest control, nutrition and soil fertility, or monitor and control pollution and devise biological waste treatment approaches. The field of mariculture also relies on microbiologists to monitor production and solve problems.
Medicine & Health Care
Hospitals, public health laboratories, research institutes and pharmaceutical companies offer work in diagnosis, prevention and treatment of illnes.


Wednesday, May 16, 2012

Enteric fever pathogens and their antimicrobial susceptibility pattern in Bharatpur, Nepal.

Acharya A1, Nepal HP2, Gautam R3 and Shrestha S4

 1Associate Professor, 2,3,4Lecturer, Department of Microbiology, Chitwan Medical College Teaching Hospital, Bharatpur, Nepal.


Enteric fever is one of the common clinical conditions in patients presenting to the hospitals.  The study was carried out to assess the rate of isolation of common serotypes of enteric fever pathogens and their antimicrobial susceptibility pattern which is of utmost importance to institute effective therapy.

A prospective study was carried out in the Department of Microbiology, Chitwan Medical College Teaching Hospital from 15th June 2009 to 14th June 2010. A total of 4355 blood culture samples from both admitted patients and outpatients of the hospitals were processed by standard microbiological technique to identify the causative agents and their susceptibility pattern to commonly used antimicrobial agents in compliance with CLSI guidelines.

Isolation rate of Salmonella species was 0.96%. Among a total of 42 Salmonella isolates, 24 (57.1%) isolates were Salmonella Paratyphi A and 18 (42.9%) were Salmonella Typhi. Male preponderances were seen in infections caused by both the organisms. On performing antimicrobial susceptibility by Kirby Bauer disc diffusion method, Salmonella Paratyphi A demonstrated 100% susceptibility to Amikacin, Chloramphenicol and Ofloxacin. Similarly, Salmonella Typhi was highly susceptible to Ceftriaxone (94.1%) followed by Ofloxacin (90.9%) and Cephotaxime (90%) and both were least susceptible to Ampicillin (S.Paratyphi A 21.7% and S.Typhi 29.4%). Multidrug resistance was found to be 16.66% among the Salmonella Typhi isolates.

Isolation of Salmonella species is relatively low in Bharatpur. Salmonella Paratyphi A is the most common agent of enteric fever. Moreover, these pathogens have developed resistance to all commonly used antimicrobials.

Bacterial Involvement in Causing Lower Respiratory Tract Infection in Adults Visiting Tribhuvan University Teaching Hospital and their Antibiotic Susceptibility Pattern


Introduction: Respiratory conditions impose enormous burden on society. Reports indicated that the top five respiratory diseases accounted for 17.4 percent of all deaths and 13.3 percent of all Disability-Adjusted Life Years (DALYs). Also, out of total acute respiratory disease, 20-24 percent of deaths are accounted for by Lower Respiratory Tract Infection (LRTI). In developing countries like Nepal the need for timely diagnosis of the cases and the administration of appropriate therapy based on the antibiotic susceptibility test of the causative agents is critical. However, emergence of resistant strains may occur during antibiotic therapy, which is one of the contributing factors for the increase in the frequency of LRTI in recent years in the adult population of Nepal as well.

 Objectives: The study was undertaken to have a better understanding on the current trend of microbial involvement in causing LRTI in adults and to determine the efficacy of antimicrobial agents in-use in treating the infections.

Method: A hospital based cross-sectional study was carried out from March 2002 to February 2003. Total 181 adults presenting with LRTI defined by a new or increasing cough, productive sputum, chest pain, fever, anorexia, haemoptysis, headaches and throat ache were enrolled with their consent. This is a prospective study which included bacteriological culture, microscopic examination and sensitivity testing of bacterial
isolates in vitro in Health Research laboratory following Standard Operating Procedures (SOPs).

Results: Lower Respiratory Infection was established in 75 cases (41.4%). Males (61.3%) were found more at risk to LRTI than females (38.7%). LRTI was found most prevalent in 50-59 year age groups (21.3%). Altogether 15 different types of bacteria were identified majority of which were Gram-negative bacteria (72.4%). Haemophilus influenzae was the commonest isolate at 23.0 percent followed by Klebsiella pneumoniae (18.3%). Among Gram- positive isolates Streptococcs pneumoniae was predominant (12.7%) followed by
Staphylococcus aureus(9.3%).The in vitro antibiotic susceptibility test of the isolates showed that Chloramphenicol(100%) was the most effective antibiotic against Gram-negative bacteria followed by Amikacin (79.1%) and Ciprofloxaxin (66.7%), and the least effective was Co-trimoxazole (20.6%). Similarly, for the Gram-positive bacteria Ciprofloxacin (79.2%) was the most effective antibiotic and the least effective was Co-trimoxazole.

Conclusion: The study shows increasing number of respiratory pathogens resistant to antimicrobials in-use to treat the infection.

Key words: LRTI, Antibiotic susceptibility pattern.

Source: Gauchan Pa, Lekhak B b and Sherchand JBc