Uninvited Guests: a Chronology of Petri Dish Contaminations.

Petri dish contaminations are commonplace and personally witnessed by every microbiologist. The vast majority of such contaminations result in nothing more than annoyance following which the Petri dishes are discarded. However, a handful of incidents of contaminations have led to momentous outcomes, the most renowned of which being that perceived by Alexander Fleming on the basis of the immense number of lives saved by penicillin. Petri dish contaminations as reported upon in the literature fall broadly into two categories; those in which the contaminant caused antagonism toward the species being cultured, and those in which the contaminant was established to be a species novum. Accounts of both of these categories of contaminations are set out here.

The "Petri" Dish: A Case of Simultaneous Invention in Bacteriology.

Richard Julius Petri's status as inventor of the culture dish that bears his name has been subject to a number of challenges over the years. Both those bacteriologists who claimed self-recognition for the invention, and those to whom it was attributed by their various advocates were all contemporaries of Petri. The evidence assembled here indicates that no single individual-including Petri-ought to be accorded credit for the inception of that shallow, circular, covered culture dish which, it transpires, is a simultaneous invention made by half a dozen bacteriologists active in the mid-1880s and ultimately owes its emergence to the prevailing bacteriological zeitgeist.

Hans Enoch and vivicillin.

Hans Emmanuel Enoch (1896-1991) was born in Hamburg, the son of a manufacturer of sera and vaccines. Upon his father's death, he took charge of the Hamburg Serum Werke. Following the rise of Hitler, he came to be pilloried in the Nazi press for allegedly having poisoned the population of Hamburg and was imprisoned for a time. He immigrated to England in 1935 where he had secured a position with the International Serum Company in Norwich. Following the outbreak of war, he was interned as an enemy alien, eventually ending up in Canada. In 1941, he was permitted to return to England, but wartime conditions prevented him from continuing to manufacture sera. At about this time, penicillin was making the headlines, and coupled with accounts of its miraculous properties, was the news that all production was reserved exclusively for the armed forces. Enoch decided to meet the public clamour for penicillin by producing a crude version which he termed 'vivicillin.' News of this spread globally, and he came to incur the disdain of Howard Florey for the attendant publicity. Notwithstanding this, vivicillin was to prove itself effective and its use led to the saving of lives.

Miracle near 34th street: Wartime Penicillin Research at St John's University, NY.

In the spring of 1944 Sister Marie Immaculate was awarded a Master of Science degree for research on penicillin which she had conducted at St John's University in Brooklyn, New York. She gave her motivation for undertaking research in this topic as wishing to fulfil her patriotic duty by participating in the quest towards making penicillin more readily available to all who needed it. It is possible that contemporary media reports suggesting that the power of penicillin was comparable to a miracle cure contributed to her interest in the subject at the time. In practical terms, her work was to have no bearing in increasing the availability of penicillin, but simply by becoming engaged in this endeavour, it could be argued that she was enacting the beliefs underlying her religious calling. This article explores those beliefs, and proposes an ideological synergy between science and religion in respect to Sister Marie Immaculate's faith in penicillin's potential to cure the world's ills.

La Moisissure et la Bactérie: Deconstructing the fable of the discovery of penicillin by Ernest Duchesne.

Ernest Duchesne (1874-1912) completed his thesis on microbial antagonism in 1897 in Lyon. His work lay unknown for fifty years, but on being brought to light led to his being credited with having discovered penicillin prior to Alexander Fleming. The claims surrounding Duchesne are examined here both from the strictly microbiological perspective, and also for what they reveal about how the process of discovery is frequently misconstrued. The combined weight of evidence presented here militates strongly against the possibility that the species of Penicillium that Duchesne worked with produced penicillin.

Inactivation of Acanthamoeba spp. and Other Ocular Pathogens by Application of Cold Atmospheric Gas Plasma.

Currently there are estimated to be approximately 3.7 million contact lens wearers in the United Kingdom and 39.2 million in North America. Contact lens wear is a major risk factor for developing an infection of the cornea known as keratitis due to poor lens hygiene practices. While there is an international standard for testing disinfection methods against bacteria and fungi (ISO 14729), no such guidelines exist for the protozoan Acanthamoeba, which causes a potentially blinding keratitis most commonly seen in contact lens wearers, and as a result, many commercially available disinfecting solutions show incomplete disinfection after 6 and 24 h of exposure. Challenge test assays based on international standard ISO 14729 were used to determine the antimicrobial activity of cold atmospheric gas plasma (CAP) against Pseudomonas aeruginosa, Candida albicans, and trophozoites and cysts of Acanthamoeba polyphaga and Acanthamoeba castellanii P. aeruginosa and C. albicans were completely inactivated in 0.5 min and 2 min, respectively, and trophozoites of A. polyphaga and A. castellanii were completely inactivated in 1 min and 2 min, respectively. Furthermore, for the highly resistant cyst stage of both species, complete inactivation was achieved after 4 min of exposure to CAP. This study demonstrates that the CAP technology is highly effective against bacterial, fungal, and protozoan pathogens. The further development of this technology has enormous potential, as this approach is able to deliver the complete inactivation of ocular pathogens in minutes, in contrast to commercial multipurpose disinfecting solutions that require a minimum of 6 h.

The role of the media in influencing public attitudes to penicillin during World War II.

Penicillin's trajectory towards becoming an effective antibacterial chemotherapeutic agent took place during World War II. Its strategic military value was immediately recognised by the Allies, and mass production was undertaken with the prime objective of meeting the needs of the armed forces. News of its development came to be widely reported on in the media and is examined here. These reports frequently combined accounts of penicillin's prodigious clinical effectiveness with the fact that it was to remain unavailable to the civilian population essentially until the war had ended. More penicillin was to be made available to the civilian population in the United States than in Britain, but the sense that it was severely rationed remained as high. It was in response to this that the idea of "homemade penicillin" was hatched. News of this was also widely promulgated by both the British and American media. Although the numbers treated with penicillin produced in this way was never to be significant, knowledge of the existence of such endeavours may have served to assuage in some measure the feelings of frustration felt by the civilian population at penicillin's non-availability.

Emerging applications of low temperature gas plasmas in the food industry.

The global burden of foodborne disease due to the presence of contaminating micro-organisms remains high, despite some notable examples of their successful reduction in some instances. Globally, the number of species of micro-organisms responsible for foodborne diseases has increased over the past decades and as a result of the continued centralization of the food processing industry, outbreaks now have far reaching consequences. Gas plasmas offer a broad range of microbicidal capabilities that could be exploited in the food industry and against which microbial resistance would be unlikely to occur. In addition to reducing the incidence of disease by acting on the micro-organisms responsible for food spoilage, gas plasmas could also play a role in increasing the shelf-life of perishable foods and thereby reduce food wastage with positive financial and environmental implications. Treatment need not be confined to the food itself but could include food processing equipment and also the environment in which commercial food processing occurs. Moreover, gas plasmas could also be used to bring about the degradation of undesirable chemical compounds, such as allergens, toxins, and pesticide residues, often encountered on foods and food-processing equipment. The literature on the application of gas plasmas to food treatment is beginning to reveal an appreciation that attention needs also to be paid to ensuring that the key quality attributes of foods are not significantly impaired as a result of treatment. A greater understanding of both the mechanisms by which micro-organisms and chemical compounds are inactivated, and of the plasma species responsible for this is forming. This is significant, as this knowledge can then be used to design plasma systems with tailored compositions that will achieve maximum efficacy. Better understanding of the underlying interactions will also enable the design and implementation of control strategies capable of minimizing variations in plasma treatment efficacy despite perturbations in environmental and operational conditions.

Cold atmospheric gas plasma disinfection of chicken meat and chicken skin contaminated with Listeria innocua.

Gas plasmas generated at atmospheric pressure and ambient temperatures offer a possible decontamination method for poultry products. The efficacy of cold atmospheric gas plasmas for decontaminating chicken skin and muscle inoculated with Listeria innocua was examined. Optimization of operating conditions for maximal bacterial inactivation was first achieved using membrane filters on which L. innocua had been deposited. Higher values of AC voltage, excitation frequency and the presence of oxygen in the carrier gas resulted in the greatest inactivation efficiency, and this was confirmed with further studies on chicken muscle and skin. Under optimal conditions, a 10 s treatment gave > 3 log reductions of L. innocua on membrane filters, an 8 min treatment gave 1 log reduction on skin, and a 4 min treatment gave > 3 log reductions on muscle. These results show that the efficacy of gas plasma treatment is greatly affected by surface topography. Scanning electron microscopy (SEM) images of chicken muscle and skin revealed surface features wherein bacteria could effectively be protected from the chemical species generated within the gas plasma. The developments in gas plasma technology necessary for its commercial application to foods are discussed.

A novel bioreactor for immobilized phototrophs.

A novel configuration of photobioreactor is described in which filaments of alginate containing immobilized cells of a leaky mutant of Dunaliella parva are wound round a central light well which is located within a glass outer chamber so that a liquid medium is caused to flow in the annular space between the outside chamber and the alginate filaments. Glycerol production by D. parva was maintained for 700 h and the highest concentration of glycerol attained was approx. 12 mg l(-1).

Chapter 5: Zones of inhibition? The transfer of information relating to penicillin in Europe during World War II.

Alexander Fleming published his first description of penicillin in 1929, but the journal articles that were to propel penicillin from its relative obscurity were those of Howard Florey and his co-workers at Oxford University. These were published in The Lancet in the early years of World War II and although wartime conditions restricted the flow of information on penicillin throughout Europe, they never succeeded in shutting it off altogether. In Germany an information-gathering initiative was established in the early phases of the war to systematically copy and distribute British and American scientific articles. A similar, though less well-resourced, operation was permitted to function in Occupied France. Both these operations were to yield up information on penicillin to their respective scientists. However, workers in other countries of occupied Europe fared less well; there was a dearth of information on penicillin in Holland but despite this, activity to produce the antibiotic still took place. Central to the production of penicillin at this time was access to a strain of Fleming's strain of Penicillium notatum, and an attempt to explain how this particular strain found its way to various European laboratories is given here.

Cold atmospheric plasma disinfection of cut fruit surfaces contaminated with migrating microorganisms.

The efficacy of cold atmospheric gas plasmas against Escherichia coli type 1, Saccharomyces cerevisiae, Gluconobacter liquefaciens, and Listeria monocytogenes Scott A was examined on inoculated membrane filters and inoculated fruit surfaces. Inoculated samples were exposed to a cold atmospheric plasma plume generated by an AC voltage of 8 kV at 30 kHz. The cold atmospheric plasma used in this study was very efficient in reducing the microbial load on the surfaces of filter membranes. However, its efficacy was markedly reduced for microorganisms on the cut surfaces. This lack of effect was not the result of quenching of reactive plasma species responsible for microbial inactivation but principally the result of the migration of microorganisms from the exterior of the fruit tissue to its interior. The velocity of migration through melon tissues was estimated to be around 300 microm min(-1) for E. coli and S. cerevisiae and through mango tissues to be 75 to 150 microm min(-1). These data can serve as operational targets for optimizing the performance of gas plasma inactivation processes. The current capabilities of cold atmospheric plasmas are reviewed and ways to improve their bactericidal efficacy are identified and discussed. Considerable scope exists to enhance significantly the efficacy of cold atmospheric plasmas for decontaminating fresh cut fruits.

Listeria monocytogenes biofilm-associated protein (BapL) may contribute to surface attachment of L. monocytogenes but is absent from many field isolates.

Listeria monocytogenes is a food-borne pathogen capable of adhering to a range of surfaces utilized within the food industry, including stainless steel. The factors required for the attachment of this ubiquitous organism to abiotic surfaces are still relatively unknown. In silico analysis of the L. monocytogenes EGD genome identified a putative cell wall-anchored protein (Lmo0435 [BapL]), which had similarity to proteins involved in biofilm formation by staphylococci. An insertion mutation was constructed in L. monocytogenes to determine the influence of this protein on attachment to abiotic surfaces. The results show that the protein may contribute to the surface adherence of strains that possess BapL, but it is not an essential requirement for all L. monocytogenes strains. Several BapL-negative field isolates demonstrated an ability to adhere to abiotic surfaces equivalent to that of BapL-positive strains. BapL is not required for the virulence of L. monocytogenes in mice.

Cold atmospheric plasma decontamination of the pericarps of fruit.

This investigation describes the inactivation by cold atmospheric plasmas of one pathogenic and three spoilage organisms on the pericarps of mangoes and melons. The operating voltage necessary for efficient microbial decontamination of fruit pericarps was first established using Escherichia coli at a concentration of 10(7) CFU/cm2 on the surface of mango. It was found that, when the plasma was sustained slightly above its breakdown voltage of 12 kV (peak to peak), no inactivation was detected when cells were plated onto tryptone soya extract agar (TSA). However, when plated onto eosin methylene blue agar, sublethal injury corresponding to approximately 1 log reduction was achieved, whereas on TSA supplemented with 4% NaCl a greater reduction of 1.5 log was revealed. When the voltage was increased by 33% to 16 kV, a reduction in cell counts of 3 log was achieved on all three plating media. Further investigations at these new operating conditions were conducted using a range of spoilage microorganisms (Saccharomyces cerevisae, Pantoea agglomerans, and Gluconacetobacter liquefaciens) all at a surface concentration of 106 CFU/cm2 on the pericarps of mango and melon. P. agglomerans and G. liquefaciens were reduced below the detection limit (corresponding to 3 log) after only 2.5 s on both fruits, whereas E. coli required 5 s to reach the same level of inactivation. S. cerevisae was the most resistant organism studied and was reduced in numbers below the detection limit after 10 s on mango and 30 s on melon. The optical emission spectra generated by the cold atmospheric plasma at both high and low operating voltages were compared in order to identify putative lethal species. It was shown that an increase in the applied voltage led to more efficient production of reactive plasma species, particularly oxygen atoms, and the production of oxygen atoms was related to the level of bacterial inactivation achieved. Production of atomic oxygen could be used as an indicator of inactivation efficiency for scaling up cold plasma systems for whole fruit.

Production of streptomycin from chitin using Streptomyces griseus in bioreactors of different configuration.

Streptomyces griseus was cultured in three different bioreactors in a medium containing chitin flakes. When a conventional bioreactor stirred by two sets of Rushton impellers and operated at high speed was used, the yield of streptomycin (3.1mg/l) was the highest observed and occurred at approximately 500 h. Cultivation of S. griseus in a bioreactor stirred at low speed by a U-shaped paddle resulted in a lower yield of streptomycin (1.8 mg/l) but this was achieved in a shorter period of time (400 h). Increasing the concentration of chitin from 5% to 10% w/v had no significant effect on either of these two parameters. The use of a novel vertical basket bioreactor in which the chitin flakes were contained within a wire mesh basket and were gently fluidised by air, enabled comparatively high yields of streptomycin (2.8 mg/l) in the relatively short time of 300 h.

The resistance to detachment of dairy strains of Listeria monocytogenes from stainless steel by shear stress is related to the fluid dynamic characteristics of the location of isolation.

Strains of Listeria monocytogenes isolated from artisanal Portuguese cheese-making dairies were divided into two categories on the basis of the locations from which they were isolated: strains from dynamic locations were those that were habitually exposed to flowing liquids during the process of cheese-making, whereas those from static locations were rarely, if ever, exposed to the shear stresses generated by liquid flows. The strength of attachment to stainless steel discs of all of these strains was obtained using a radial flow chamber. Initial attachment strengths to stainless steel (after a 0.5 h contact time) of L. monocytogenes strains were greater for the 5 isolates from surfaces exposed to flow (dynamic isolates) than for most (3 out of 4) of those that were not (static isolates). After a 24 h contact time, attachment strength of all isolates reached similar levels. These results suggest that strains having high initial attachment strength are more likely to persist on surfaces exposed to flow than strains having low initial attachment strength. The numerical values of shear forces obtained could prove useful in the rational design of cleaning and decontamination procedures in food processing facilities.