- The Royal Colege of Physicians. Preparation for pandemic influenza.

www.rcplondon.ac.uk/pubs/brochure.aspx?e=276

- La gestione della sindrome influenzale

www.apss.tn.it/allegati/doc_625308_0.pdf

- World Health Organisation. Guidance for hospital medical specialities on adaptations needed for a pandemic influenza out break, WHO, 6/09.

www.who.int/csr/disease/influenza/pandemic/en

Javeed Kakroo

Culture is a technique where we grow microorganisms in a nutritional liquid or on a solid medium in order to increase numbers and simplify identification. It is also important as a first step in several laboratory techniques and for testing of antimicrobial sensitivity.

There are many types of medium available for culturing microorganisms, and so it is important to know which is best to use – especially for organisms that are difficult to grow.

Many of these media are available commercially, but for some it can be more cost effective to prepare the media yourself. For completeness, recipes for all media above are listed in alphabetical order. Adjustment of these recipes may be necessary for some cultures, but these will provide a good starting point. Most of the recipes include ‘agar’, which can be left out to make liquid cultures, but some rely on the use of nutrient agars which will require replacing with another nutrient source to make a liquid medium for culture.

Contents:

Bacteroides bile-esculin (with gentamicin and bile) agar
Bordet-Gengou agar with methicillin or cephalexin
Buffered Charcoal Yeast Extract agar
Campylobacter-selective agar
Cefsulodin-Irgasan-Novobiocin agar
Cystine Heart agar
Cystine-Tellurite Blood agar
Fletcher’s medium with rabbit serum
Gram-Negative Enrichment Broth
Hektoen agar
Horse Blood-Charcoal agar
Kanamycin-Vancomycin Laked Blood agar
Leptospira Medium with Bovine Serum Albumin (BSA)-Tween 80
Löffler’s Coagulated Serum Medium
MacConkey agar
MacConkey-Sorbitol agar
Modified Thayer-Martin agar
New York City agar
Pseudomonas cepacia agar
Regan-Lowe Cephalixin agar
Salmonella–Shigella agar
Selenite Enrichment Broth
Stuart Medium with rabbit serum
Thiosulfate-Citrate-Bile Salts-Sucrose agar
Tinsdale agar
Xylose-Lysine-Deoxycholate agar

Bacteroides bile-esculin with gentamycin and bile

Bacteroides bile-esculin (with gentamicin and bile) agar:

45g/l Tryptic Soy Agar
20g/l Ox bile
1 g/l Esculin
5g/l Ferric Ammonium Citrate
12mg/l Hemin
100mg/l Gentamicin

Bordet-Gengou agar with methicillin or cephalexin:

15g/l Agar
5g/l Potato Infusion
5g/l Sodium Chloride
3g/l Meat Peptone
2g/l Casein Peptone
10ml/l Glycerol
15% Sheep’s blood
Methicillin or Cephalexin as appropriate to your sample

Buffered Charcoal Yeast Extract Agar

Buffered Charcoal Yeast Extract agar:

17g/l Agar
5g/l Yeast Extract
10g/l ACES Buffer
3g/l Activated Charcoal
5g/l Potassium Hydroxide
1g/l alpha-Ketoglutarate
4g/l L-Cysteine
25g/l Ferric Pyrophosphate

Campylobacter-selective agar (for example):

5g/l Agar
12g/l Pancreatic Digest of Casein
5g/l Peptic Digest of Animal Tissue
5g/l Sodium Chloride
4g/l Activated Charcoal
3g/ Yeast Extract
3g/l Beef Extract
1 g/l Corn Starch
1g/l Cycloheximide
32mg/l Hemin
32mg/l Cefoperazone
20mg/l Vancomycin

Cefsulodin-Irgasan-Novobiocin agar

Cefsulodin-Irgasan-Novobiocin agar:

20g/l Mannitol
17g/l Peptone
5g/l Agar
3g/l Proteose Peptone
2g/l Yeast Extract
2g/l Sodium Pyruvate
1g/l Sodium Chloride
5g/l Sodium Deoxycholate
5g/l Sodium Cholate
30mg/l Neutral Red
10mg/l Magnesium Sulfate
4mg/l Irgasan®
4mg/l Cefsulodin
5mg/l Novobiocin
1mg/l Crystal Violet

Cystine Heart agar:

15g/l Agar
10g/l Beef Heart (Infusion from 500g)
10g/l Proteose Peptone
10g/l Dextrose
5g/l Sodium Chloride
1g/l L-Cystine

Cystine-Tellurite Blood agar

Cystine-Tellurite Blood agar:

15g/l Agar
13g/l Casein Peptone
5g/l Sodium Chloride
5g/l Yeast Extract
2g/l Beef Heart (Infusion from 500g)
44mg/l L-Cystine
50ml/l 1% Potassium Tellurite
5% Sheep’s Blood

Fletcher’s medium with rabbit serum:

5g/l Agar
5g/l Sodium Chloride
3g/l Peptone
2g/l Beef Extract
80ml/l Rabbit Serum

Gram-Negative Enrichment Broth:

10g/l Pancreatic Digest of Casein
10g/l Peptic Digest of Animal Tissue
5g/l Sodium Chloride
5g/l Sodium Citrate
4g/l Dipotassium Phosphate
2g/l Mannitol
5g/l Monopotassium Phosphate
1g/l Dextrose
5g/l Sodium Deoxycholate

Hektoen agar

Hektoen agar:

5g/l Agar
12g/l Peptic Digest of Animal Tissue
12g/l Lactose
12g/l Sucrose
9g/l Bile Salts
5g/l Sodium Chloride
5g/l Sodium Thiosulfate
3g/l Yeast Extract
2g/l Salicin
5g/l Ferric Ammonium Citrate
1g/l Acid Fuchsin
64mg/l Bromothymol Blue

Horse Blood-Charcoal agar:

12g/l Agar
10g/l Beef Extract
10g/l Casein/Meat polypeptone
10g/l Starch
5g/l Sodium Chloride
4g/l Charcoal
1mg/l Nicotinic acid
10% Horse Blood

Kanamycin-Vancomycin Laked Blood agar:

20g/l Peptamin
17g/l Agar
5g/l Sodium Chloride
2g/l Yeast Extract
1g/l Dextrose
1g/l Sodium Bisulfite
5mg/l Hemin
1mg/l Vitamin K
40ml/l Vancomycin
40ml/l Kanamycin
50ml Laked Sheep’s Blood

Leptospira Medium with Bovine Serum Albumin (BSA)-Tween 80:

Base medium (900ml)11g/l Agar

1g/l Disodium Phosphate
1g/l Sodium Chloride
3g/l Monopotassium Phosphate
1ml/l 10% Glycerol
1ml/l 25% Ammonium Chloride solution
1ml/l 10% Sodium Pyruvate solution
1ml/l 0.5% Thiamine solution

Albumin Supplement (100ml)Magnesium Chloride-Calcium Chloride solution:100g/l BSA

125ml/l 10% Tween-80 solution
100ml/l 0.5% Iron Sulfate solution
10ml/l Magnesium Chloride-Calcium Chloride solution (see below)
10ml/l 0.02% Cyanocobalamin solution
10ml/l 0.4% Zinc Sulfate solution

Magnesium Chloride-Calcium Chloride solution:

15g/l Calcium Chloride Dihydrate
15g/l Magnesium Chloride Hexahydrate

Loeffler’s Coagulated Serum Medium

Löffler’s (also spelled ‘Loeffler’) Coagulated Serum Medium:

5g/l Dextrose
5g/l Gelatin Peptone
5g/l Beef Extract
25g/l Sodium Chloride
750ml/l Bovine Serum

MacConkey agar

MacConkey agar:

17g/l Peptone
5g/l Agar
10g/l Lactose
5g/l Sodium Chloride
5g/l Bile Salts
30mg/l Neutral Red
1mg/l Crystal Violet

MacConkey-Sorbitol agar:

As above, but including 10g/l sorbitol instead of lactose

Modified Thayer-Martin agar:

23g/l Peptic Digest of Animal Tissue
20g/l Agar
5g/l Sodium Chloride
5g/l Dextrose
1g/l Starch
200ml/l Lysed Blood
5mg/l Colistin Methane Sulphate
3mg/l Vancomycin

New York City agar:

20g/l Agar
15g/l Proteose Peptone
5g/l Glucose
5g/l Sodium Chloride
4g/l Dipotassium Hydrogen Phosphate
1g/l Corn Starch
1g/l Potassium Dihydrogen Phosphate
100ml/l Horse Blood Cells
60ml/l Horse Plasma
Yeast Autolysate Supplement (Available commercially, recipe below)

10g/l Yeast Autolysate fractions
1g/l Glucose
15g/l Sodium Bicarbonate

NYC Supplement, also known as C.L.A.T. antibiotics (available commercially, recipe below)

5mg/l Colustin Methane Sulphonate

4mg/l Linomycin

1mg/l Amphotericin B

5mg/l Trimethoprim Lactate

Pseudomonas cepacia agar:

15g/l Agar
5g/l Sodium Pyruvate
3g/l Dipotassium Phosphate
1g/l Monopotassium Phosphate
5g/l Bile Extract
1g/l Enzymatic Digest of Animal Tissue
1g/l Ammonium Sulfate
2g/l Magnesium Sulfate
1g/l Ticarcillin
20mg/l Phenol Red
10mg/l Ferrous Ammonium Sulphate
1mg/l Crystal Violet
300 000Units Polymyxin B

Regan-Lowe Cephalixin agar:

As Horse Blood-Charcoal Agar with the inclusion of 40mg/l Cephalexin

Salmonella–Shigella agar

Salmonella–Shigella agar:

5g/l Agar
10g/l Lactose
5g/l Bile Salts
5g/l Sodium Citrate
5g/l Sodium Thiosulfate
5g/l Beef Extract
5g/l Enzymatic Digest of Casein
5g/l Enzymatic Digest of Animal Tissue
1g/l Ferric Citrate
25mg/l Neutral Red
33mg/l Brilliant Green

Selenite Enrichment Broth:

10g/l Sodium Phosphate
5g/l Pancreatic Digest of Casein
4g/l Lactose
4g/l Sodium Selenite

Stuart Medium with rabbit serum:

10g/l Sodium Glycerophosphate
3g/l Agar
1g/l Sodium Thioglycollate
1g/l Calcium Chloride
2mg/l Methylene Blue
80ml/l Rabbit Serum

Thiosulfate-Citrate-Bile salts-Sucrose agar

Thiosulfate-Citrate-Bile Salts-Sucrose agar:

20g/l Sucrose
15g/l Agar
10g/l Dipeptone
10g/l Sodium Citrate
10g/l Sodium Thiosulfate
10g/l Sodium Chloride
5g/l Yeast Extract
5g/l Ox Bile
3g/l Sodium Cholate
1g/l Ferric Citrate
40mg/l Bromothymol Blue
40mg/l Thymol Blue

Tinsdale agar:

20g/l Protease Peptone
15g/l Agar
5g/l Yeast Extract
5g/l Sodium Chloride
425g/l Sodium Thiosulfate
3445g/l Potassium Tellurite
24g/l L-Cystine
100ml Serum

Xylose-Lysine-Deoxycholate agar

Xylose-Lysine-Deoxycholate agar:

5g/l Agar
5g/l Lactose
5g/l Sucrose
8g/l Sodium Thiosulfate
5g/l L-Lysine HCl
5g/l Sodium Chloride
3g/l Yeast Extract
1g/l Sodium Deoxycholate
8g/l Ferric Ammonium Citrate
80mg/l Phenol Red

Robert Millar (University of Warwick)

Society for Applied Microbiology

He was a Scottish biologist and pharmacologist. Fleming published many articles on bacteriology, immunology and chemotherapy. His best-known achievements are the discovery of the enzyme lysozyme in 1923 and the antibiotic substance penicillin from the fungus Penicillium notatum in 1928, for which he shared the Nobel Prize in Physiology or Medicine in 1945 with Howard Walter Florey and Ernst Boris Chain.

In 1999, Time Magazine named Fleming one of the 100 Most Important People of the 20th Century for his discovery of penicillin, and stated; “It was a discovery that would change the course of history. The active ingredient in that mold, which Fleming named penicillin, turned out to be an infection-fighting agent of enormous potency. When it was finally recognized for what it was—the most efficacious life-saving drug in the world—penicillin would alter forever the treatment of bacterial infections. By the middle of the century, Fleming’s discovery had spawned a huge pharmaceutical industry, churning out synthetic penicillins that would conquer some of mankind’s most ancient scourges, including syphilis, gangrene and tuberculosis”.

Fleming served throughout World War I as a captain in the Army Medical Corps, and was mentioned in dispatches. He and many of his colleagues worked in battlefield hospitals at the Western Front in France. In 1918 he returned to St. Mary’s Hospital, which was a teaching hospital. He was elected Professor of Bacteriology in 1928.

After the war Fleming actively searched for anti-bacterial agents, having witnessed the death of many soldiers from septicemia resulting from infected wounds. Unfortunately antiseptics killed the patients’ immunological defences more effectively than they killed the invading bacteria. In an article he submitted for the medical journal The Lancet during World War I, Fleming described an ingenious experiment, which he was able to conduct as a result of his own glass blowing skills, in which he explained why antiseptics were actually killing more soldiers than infection itself during World War I. Antiseptics worked well on the surface, but deep wounds tended to shelter anaerobic bacteria from the antiseptic agent, and antiseptics seemed to remove beneficial agents produced that actually protected the patients in these cases at least as well as they removed bacteria, and did nothing to remove the bacteria that were out of reach. Sir Almroth Wright strongly supported Fleming’s findings, but despite this, most army physicians over the course of WWI continued to use antiseptics even in cases where this worsened the condition of the patients.

“When I woke up just after dawn on September 28, 1928, I certainly didn’t plan to revolutionize all medicine by discovering the world’s first antibiotic, or bacteria killer,” Fleming would later say, “But I guess that was exactly what I did”.

By 1928, Fleming was investigating the properties of staphylococci. He was already well-known from his earlier work, and had developed a reputation as a brilliant researcher, but his laboratory was often untidy. On 3 September 1928, Fleming returned to his laboratory having spent August on vacation with his family. Before leaving he had stacked all his cultures of staphylococci on a bench in a corner of his laboratory. On returning, Fleming noticed that one culture was contaminated with a fungus, and that the colonies of staphylococci that had immediately surrounded it had been destroyed, whereas other colonies further away were normal. Fleming showed the contaminated culture to his former assistant Merlin Price who said “that’s how you discovered lysozyme.” Fleming identified the mould that had contaminated his culture plates as being from the Penicillium genus, and—after some months’ of calling it “mould juice”— named the substance it released penicillin on 7 March 1929.

He investigated its positive anti-bacterial effect on many organisms, and noticed that it affected bacteria such as staphylococci, and many other Gram-positive pathogens that cause scarlet fever, pneumonia, meningitis and diphtheria, but not typhoid fever or paratyphoid fever—which are caused by Gram-negative bacteria—for which he was seeking a cure at the time. It also affected Neisseria gonorrhoeae, which causes gonorrhoea although this bacterium is Gram-negative.

Fleming published his discovery in 1929, in the British Journal of Experimental Pathology, but little attention was paid to his article. Fleming continued his investigations, but found that cultivating penicillium was quite difficult, and that after having grown the mould, it was even more difficult to isolate the antibiotic agent. Fleming’s impression was that because of the problem of producing it in quantity, and because its action appeared to be rather slow, penicillin would not be important in treating infection. Fleming also became convinced that penicillin would not last long enough in the human body (in vivo) to kill bacteria effectively. Many clinical tests were inconclusive, probably because it had been used as a surface antiseptic. In the 1930s, Fleming’s trials occasionally showed more promise, and he continued, until 1940, to try to interest a chemist skilled enough to further refine usable penicillin.

Fleming soon abandoned penicillin, and not long after Florey and Chain took up researching and mass producing it with funds from the U.S and British governments. They started mass production after the bombing of Pearl Harbor. When D-day arrived they had made enough penicillin to treat all the wounded allied forces.

ISO-14644-1 Classification by Airborne Particles ISO-14644-2 Monitoring for Compliance

ISO-14644-3 Measurement & Testing ISO-14644-4 Design, Construction and Start-up

ISO-14644-5 Cleanroom Operations ISO-14644-6 Terms, Definitions & Units

ISO-14644-7 Separative Enclosures ISO-14644-8 Molecular Contamination 

The obligate anaerobe micro-organisms are not able to detoxifying these active forms of oxygen.

It is therefore imperative to adopt an anaerobic cabinet equipped with air locks and filled with inert gases to cultivate stringent anaerobes.

An antiviral pill proved 100 percent effective in preventing new cases of HIV, according to a new Kaiser Permanente study.

Truvada was administered to 657 sexually-active people over 32 months – and there were no new cases of HIV reported, according to the study, published in the journalClinical Infectious Diseases this month.

“Our study is the first to extend the understanding of the use of PrEP (pre-exposure prophylaxis) in a real-world setting and suggests that the treatment may prevent new HIV infections even in a high-risk setting,” said Jonathan Volk, physician and epidemiologist at Kaiser Permanente San Francisco Medical Center, and the lead author.

Ninety-nine percent of the subjects were men who have sex with men, according to the study. They were sexually active: within 12 months of the initiation of the Truvada regimen, 50 percent of the patients had contracted at least one kind of sexually-transmitted infection, like rectal infections, chlamydia, gonorrhea and syphilis.

But none contracted HIV.

A subset of the group reported not changing their number of sex partners over the time of treatment.

However, the researchers could not be sure if the antiviral treatments encouraged risky behaviors.

“Without a control group, we don’t know if these STI rates were higher than what we would have seen without PrEP,” said Julia Marcus, postdoctoral fellow at the Kaiser Permanente Division of Research, and a co-author. “Ongoing screening and treatments for STIs, including hepatitis C, are an essential component of a PrEP treatment program.”

Truvada was approved by the FDA in in July 2012 for daily oral use.

The results could even prove to be a breakthrough, according to an editorial that accompanied the study, written by staff from the University of California at San Francisco and the San Francisco AIDS Foundation.

“Given historical devastation wrought by HIV/AIDS within the city of San Francisco and elsewhere, this is tremendously good news,” they wrote. “The data published by Volk and colleagues demonstrates meaningful progress towards the goal of halting new infections.”

However, additional attention to growing rates of other STIs is necessary, they added.

“It is time for a vigorous conversation about sexually-transmitted infections, too long eclipsed by fear of HIV infection,” they added. “While HIV testing and trends are frequently in the news, notice of STI trends remain unnoticed in technical reports… Feeling safer from HIV infection while using PrEP creates spaces for a more robust discussion of STIs.”

Another potentially promising study was published last week in the journal Cell. Scientists from the California Institute of Technology have honed in on a particular antibody that naturally occurs in some patients who are naturally able to fend off HIV – and the researchers said it could prove to be a route to eradicate the infection.

Font >>> http://www.laboratoryequipment.com/

Scanning electron microscopy of particles of 4 families of giant viruses that have now been identified. The largest dimensions can reach between 0.6 microns (Mollivirus) and 1.5 microns (Pandoravirus). © IGS CNRS/AMU

Scientists have issued another climate change warning—be careful not to awaken prehistoric viruses that we’ve never seen the likes of before.

That is according to French scientists who have discovered and plan to reawaken a 30,000-year-old “giant” prehistoric virus in the permafrost of northern Siberia.

Called Mollivirius sibericum, this is the fourth prehistoric virus discovered since 2003, and it raises questions about what other dangerous microscopic pathogens might be locked in the frozen tundra—which is quickly becoming a non-frozen tundra.

With melting permafrost and climbing temperatures, this specific area of the Artic is increasingly interesting for its mining and oil resources.

“A few viral particles that are still infectious may be enough, in the presence of a vulnerable host, to revive potentially pathogenic viruses,” Jean-Michel Claverie, one of the lead authors of the study published in theProceedings of the National Academy of Sciences, told the AFP. “If we are not careful, and we industrialize these areas without putting safeguards in place, we run the risk of one day waking up viruses such as smallpox that we thought were eradicated.”

Giant viruses measure longer than 0.5 microns, placing Mollivirius sibericum in the category by just 0.1 micron. The scientists drew on microscopic, genomic, transcriptomic, proteomic and metagenomic technologies to draw a detailed portrait of the 0.6-micron ancient virus. According to CNRS, this is the first time all the analytical techniques applicable to living beings have been used simultaneously to characterize a virus.

Mollivirius sibericum takes the form of a roughly spherical particle containing a genome of approximately 650,000 base pairs coding for more than 500 proteins. What is surprising about this virus, as well as previously discovered ancient viruses, is how more complex genetically they are than modern-day viruses. For example, previously discovered ancient virus Pithovirus sibericum (located in the same area) boasts 2,500 genes. By contrast, today’s Influenza A virus has only eight genes.

In safe laboratory conditions, Claverie and colleagues will attempt to revive Mollivirius sibericum by placing it with single-cell amoeba, which will serve as its host. In 2013, Claverie and colleagues were successful in their attempts to revive Pithovirus sibericum in a petri dish. Before they wake the virus up, the researchers said they plan to verify that it is indeed harmless to animals and humans. 

Font >>> http://www.laboratoryequipment.com/

The Hantavirus is a very dangerous and very deceiving illness that mimics the symptoms of the common flu. This infographic from USC, Master of Public Health Online shows that within the first few days all symptoms appear as the flu would, but after the four day mark symptoms elevate, and become more strenuous. Carried by rodents, the Hantavirus is spread through their waste, which becomes airborne when their living space is disturbed. Any home or place of business should take premeditated actions against the virus by fixing holes around the building, as well as by dealing with the rodents currently inside. If anyone has been exposed to infested areas and starts to show flu-like symptoms, get help as soon as possible, because early treatment greatly increases your chance of survival.

Fonte: Genengnews

The spread of the Ebola virus in West Africa dominated headlines during the second half of the year. To date, more than 18,000 people have been infected with Ebola, and nearly 7,000 have died, according to the World Health Organization (WHO). While grim, these statistics are lower than the dire predictions released by the WHO and the US Centers for Disease Control and Prevention (CDC) earlier this year. And a recent epidemiological analysis suggested that the number of unreported cases is lower than previous estimates.

In September, the United Nations Security Council declared the Ebola epidemic a “threat to international peace and security,” spurring large-scale international aid efforts, which have helped to curb the spread of the disease, particularly in Liberia. “But the outbreak continues to surge in Sierra Leone, and there has been a troubling spread in Guinea’s capitol city,” CDC director Thomas Frieden said in a statement this week (December 22). “We’ve got a long way to go and this is no time to relax our grip on the response.”

Fonte: The Scientist

Living bacteria divide and proliferate; if not, they are considered dead. However, some bacteria can go into a strange, in-between state where they are biologically active – producing energy and making proteins – but do not divide. “It’s a kind of living-dead, zombie existence,” said John McKinney, whose postdoc, Giulia Manina, led the study on M. tuberculosis. “The bacteria are somewhat active, but they’re neither growing nor dividing. We refer to this state as ‘Non-Growing but Metabolically Active’ or ‘NGMA’”.

Fonte: Bioscience technology