Challenge Organisms Research Articles

Fibrous Catalyst-Enhanced Acanthamoeba Disinfection by Hydrogen Peroxide.

SIGNIFICANCEHydrogen peroxide (H2O2) disinfection systems are contact-lens-patient problem solvers. The current one-step, criterion-standard version has been widely used since the mid-1980s, without any significant improvement. This work identifies a potential next-generation, one-step H2O2, not based on the solution formulation but rather on a case-based peroxide catalyst.PURPOSEOne-step H2O2 systems are widely used for contact lens disinfection. However, antimicrobial efficacy can be limited because of the rapid neutralization of the peroxide from the catalytic component of the systems. We studied whether the addition of an iron-containing catalyst bound to a nonfunctional propylene:polyacryonitrile fabric matrix could enhance the antimicrobial efficacy of these one-step H2O2 systems.METHODSBausch + Lomb PeroxiClear and AOSept Plus (both based on 3% H2O2 with a platinum-neutralizing disc) were the test systems. These were tested with and without the presence of the catalyst fabric using Acanthamoeba cysts as the challenge organism. After 6 hours' disinfection, the number of viable cysts was determined. In other studies, the experiments were also conducted with biofilm formed by Stenotrophomonas maltophilia and Elizabethkingia meningoseptica bacteria.RESULTSBoth control systems gave approximately 1-log10 kill of Acanthamoeba cysts compared with 3.0-log10 kill in the presence of the catalyst (P < .001). In the biofilm studies, no viable bacteria were recovered following disinfection in the presence of the catalyst compared with ≥3.0-log10 kill when it was omitted. In 30 rounds' recurrent usage, the experiments, in which the AOSept Plus system was subjected to 30 rounds of H2O2 neutralization with or without the presence of catalytic fabric, showed no loss in enhanced biocidal efficacy of the material. The catalytic fabric was also shown to not retard or increase the rate of H2O2 neutralization.CONCLUSIONSWe have demonstrated the catalyst significantly increases the efficacy of one-step H2O2 disinfection systems using highly resistant Acanthamoeba cysts and bacterial biofilm. Incorporating the catalyst into the design of these one-step H2O2 disinfection systems could improve the antimicrobial efficacy and provide a greater margin of safety for contact lens users.

Previous stress exposures influence subsequent UV-C resistance of Salmonella enterica in coconut liquid endosperm

This study determined the effects of prior exposures to stresses commonly encountered by cells in food production and processing environments on the subsequent UV-C inactivation rates of Salmonella enterica in coconut liquid endosperm. Seven different strains of S. enterica were separately exposed to individual stresses such as gradual acidification (final pH 4.5), abrupt desiccation (aw 0.96), or heat stress at 40 °C for 24 h, after which the test strains were cocktailed and subjected to UV-C challenge. Cells were also exposed to all possible combinations of the individual stresses and thereafter subjected to UV-C challenge. Cells exposed to all individual and combinations of stresses exhibited 1st order, log-linear inactivation behavior (R2 0.903 to 0.998). Cells previously exposed to singular heat stress had the highest UV-C resistance (DUV-C 43.8 mJ/cm2), while cells exposed to all simultaneous pH, aw, and heat stresses had the least (DUV-C 22.5 mJ/cm2). Heterologous adaptive mechanisms were observed after S. enterica cells were exposed to acid, acid + desiccation, heat, and acid + heat, with individual heat stress exposure resulting in the significantly most UV-C resistant cells. Results obtained in this study provide baseline information on the selection of appropriate challenge organism for establishment of UV-C process schedule for coconut liquid endosperm and similar commodities.

Biocidal efficacy of multipurpose solutions against Gram‐negative organisms associated with corneal infiltrative events

BackgroundBecause corneal infiltrative events () may result from bacterial components on contact lenses, which can come from contaminated lens cases, we evaluated the biocidal efficacy of five multipurpose solutions against Gram‐negative commonly isolated and ‐associated organisms.MethodsOf the multipurpose solutions tested, one contained polyhexamethylene biguanide (PHMB)/polyquaternium‐1 (PQ‐1; Bausch & Lomb Incorporated: Biotrue), one contained alexidine dihydrochloride (alexidine)/PQ‐1 (AMO: RevitaLens OcuTec) and three contained PQ‐1/myristamidopropyl dimethylamine (MAPD; Alcon: Opti‐Free PureMoist, PQ‐1/‐1; Opti‐Free RepleniSH, PQ‐1/MAPD‐2; Opti‐Free Express, PQ‐1/MAPD‐3). Challenge organisms were CIE‐associated Achromobacter xylosoxidans, Delftia acidovorans and Stenotrophomonas maltophilia at manufacturer‐recommended durations (stand‐alone), in lens cases without lenses (up to seven days) and in lens cases with etafilcon A lenses (up to 30 days).ResultsIn stand‐alone testing against CIE‐associated organisms, PHMB/PQ‐1 and alexidine/PQ‐1 were significantly superior versus MAPD‐based multipurpose solutions against A. xylosoxidans (all p ≤ 0.01), D. acidovorans (all p ≤ 0.001) and S. maltophilia (all p ≤ 0.05). In lens cases, PHMB/PQ‐1 and alexidine/PQ‐1 achieved greater than 3‐log reductions against all challenge organisms at all times evaluated. PQ‐1/MAPD‐1 achieved a greater than 3‐log reduction against D. acidovorans at 24 hours; PQ‐1/MAPD‐1 and PQ‐1/MAPD‐3 achieved greater than 3‐log reductions at seven days against all organisms. In lens cases with lenses, PHMB/‐1 and alexidine/PQ‐1 achieved greater than 3‐log reductions against all organisms at all times. PQ‐1/MAPD‐1 and PQ‐1/MAPD‐3 achieved greater than 3‐log reductions at seven or more days against all organisms. PQ‐1/MAPD‐2 did not achieve a greater than 3‐log reduction at any time; some regrowth was observed.ConclusionsPHMB‐ and alexidine‐based multipurpose solutions demonstrated significantly greater biocidal activity compared with PQ‐1/MAPD‐based agents against Gram‐negative organisms commonly isolated and CIE‐associated pathogens.

Tetraselmis as a challenge organism for validation of ballast water UV systems

Transport and release of waterborne organisms as a result of ballasting and de-ballasting operations is widely acknowledged to represent an important mechanism for invasions by non-indigenous species. Regulatory requirements have been implemented globally to require treatment of ballast water before its release to the environment as a means of minimizing risks of invasion. UV-based processes represent an option for ballast water treatment; however, their use will require development of appropriate methods for reactor validation. To address this need, Tetraselmis was examined as challenge organism using a most probable number (MPN) assay for quantification of the concentration of viable (reproductively active) cells in suspension. A low pressure collimated-beam reactor was used to investigate UV254 dose-response behavior of Tetraselmis. Based on the experimental conditions applied, Tetraselmis indicated 4.5–5 log10 units of inactivation for UV254 doses of approximately 120 mJ/cm2, with no apparent change of resistance resulting from repeated exposure. A medium pressure UV collimated-beam reactor equipped with a series of narrow bandpass optical filters was used to investigate the action spectrum of Tetraselmis for wavelengths ranging from 228 nm–297 nm. Radiation with wavelengths in the range 254–280 nm was observed to be most efficient for inactivation of Tetraselmis. Additionally, DNA was extracted from Tetraselmis to allow measurement of its absorption spectrum. These results indicated strong absorbance from 254 nm to 280 nm, thereby suggesting that damage to DNA plays an important role in the inactivation of Tetraselmis sp. However, deviations of the action spectrum shape from the shape of the DNA absorption spectrum suggest that UV-induced damage to biomolecules other than DNA may contribute to Tetraselmis inactivation at some wavelengths in the UVC range.

Inactivation behaviors of selected bacteria in ultraviolet-C-treated human breast milk

Abstract This study determined the inactivation behaviors, which include inactivation pattern, sublethal injury development, and inactivation kinetic parameters of selected bacteria in UV-C treated human breast milk. Multi strain mixtures of Salmonella enterica, Escherichia coli O157:H7, non-pathogenic E. coli , Pseudomonas aeruginosa , Listeria monocytogenes, and a lone strain of Staphylococcus aureus were inoculated into donated breast milk prior to UV-C inactivation studies. Results showed that the test organisms exhibited two types of inactivation patterns, making it difficult to directly compare UV-C resistance. S. enterica and non-pathogenic E. coli exhibited a monophasic log-linear inactivation pattern within the 60-min UV-C treatment period. All other challenge organisms exhibited a biphasic inactivation pattern characterized by a faster initial log linear inactivation, followed by a slower inactivation tailing. Sublethal injury rates were small in all test organisms except for S. aureus , which developed total sublethal injury throughout the UV-C exposure. Despite the injury rates, S. aureus had the least total population reduction (2.63 log CFU) after the 60-min treatment, while S. enterica had the greatest reduction (3.88 log CFU). The inactivation behaviors established in this work provide baseline information in the development of a UV-C pasteurization equipment and process for breast milk.

Evolution of evolvability and phenotypic plasticity in virtual cells

BackgroundChanging environmental conditions pose a challenge for the survival of species. To meet this challenge organisms adapt their phenotype by physiological regulation (phenotypic plasticity) or by evolving. Regulatory mechanisms that ensure a constant internal environment in the face of continuous external fluctuations (homeostasis) are ubiquitous and essential for survival. However, more drastic and enduring environmental change, often requires lineages to adapt by mutating. In vitro evolutionary experiments with microbes show that adaptive, large phenotypic changes occur remarkably quickly, requiring only a few mutations. It has been proposed that the high evolvability demonstrated by these microbes, is an evolved property. If both regulation (phenotypic plasticity) and evolvability can evolve as strategies to adapt to change, what are the conditions that favour the emergence of either of these strategy? Does evolution of one strategy hinder or facilitate evolution of the other strategy?ResultsHere we investigate this with computational evolutionary modelling in populations of Virtual Cells. During a preparatory evolutionary phase, Virtual Cells evolved homeostasis regulation for internal metabolite concentrations in a fluctuating environment. The resulting wild-type Virtual Cell strains (WT-VCS) were then exposed to periodic, drastic environmental changes, while maintaining selection on homeostasis regulation. In different sets of simulations the nature and frequencies of environmental change were varied. Pre-evolved WT-VCS were highly evolvable, showing rapid evolutionary adaptation after novel environmental change. Moreover, continued low frequency changes resulted in evolutionary restructuring of the genome that enables even faster adaptation with very few mutations. In contrast, when change frequency is high, lineages evolve phenotypic plasticity that allows them to be fit in different environments without mutations. Yet, evolving phenotypic plasticity is a comparatively slow process. Under intermediate change frequencies, both strategies occur.ConclusionsWe conclude that evolving a homeostasis mechanisms predisposes lineage to be evolvable to novel environmental conditions. Moreover, after continued evolution, evolvability can be a viable alternative with comparable fitness to regulated phenotypic plasticity in all but the most rapidly changing environments.

A combined low‐pressure hydrogen peroxide evaporation plus hydrogen plasma treatment method for sterilization – Part 1: Characterization of the condensation process and proof‐of‐concept

A combined hydrogen peroxide evaporation and hydrogen low‐pressure plasma treatment process for sterilization is introduced and investigated. The combination of hydrogen peroxide evaporation followed by hydrogen plasma treatment offers an advantage regarding sterilization in complex metal geometries or in sealed sterile bags, where plasma treatment alone faces challenges. Within this contribution, the droplet size and film homogeneity after condensation is investigated by optical diagnostics. Sterilization tests with common challenge organisms show the sterilization capabilities of the combined process in a process challenge device, mimicking the worst‐case‐scenario for plasma treatment in a small metal box. Furthermore, sterilization in sealed sterile bags clearly demonstrates the advantage of the combined process, showing full spore inactivation solely for the combined process.

The visual assessment of broth cultures for tissue bank samples.

The bioburden screening process of allograft musculoskeletal tissue samples received at the South Eastern Area Laboratory Services includes the routine use of solid agar and cooked meat (CM) broth media. CM has been routinely sub-cultured onto solid agar plates after aerobic incubation at 35°C. This study will evaluate whether a visual assessment of CM can replace sub-culture by an in vitro inoculation and a prospective study. Eight challenge organisms were serially diluted and inoculated into CM. The average inoculum of 0.5-5.5CFU produced visible turbidity of CM after 24-h incubation for 7 of the challenge organisms with one organism producing turbidity after 48-h incubation. The prospective study evaluated 222 CM of which 213 were visually clear and no-growth on sub-culture and 9 turbid CM which were culture positive. Broth cultures are an integral part of the bioburden screening process of allograft musculoskeletal tissue and swab samples and visual assessment of CM can replace sub-culture.

Effect of sublethal temperatures on pulsed light inactivation of bacteria

Abstract This study investigated the effect of sublethal temperatures on the efficacy of pulsed light (PL) treatment for the inactivation of Listeria innocua, Escherichia coli ATCC 25922, and Pseudomonas fluorescens. A thin layer of clear, liquid phosphate buffer inoculated with one of the challenge organisms, at a concentration of about 108 CFU/mL, was equilibrated to a temperature ranging from 5 °C to 50 °C and then treated with PL, at doses between 1.02 and 12.29 J/cm2. All treatments were performed in triplicate. In the temperature range of 5 °C to 40 °C, the average maximum reductions for L. innocua, E. coli, P. fluorescens were 6.27 ± 0.23 log CFU, 6.66 ± 0.36 log CFU, and 6.15 ± 0.19 log CFU, respectively. Temperature did not affect PL inactivation of E. coli or P. fluorescens, but a modest synergistic effect between PL and temperature was observed for L. innocua treated above 40 °C. Industrial relevance This study suggests that PL inactivation of bacteria is independent of temperature in the sublethal range 5 °C to 40 °C. The practical benefit of this finding is that PL treatments can be conducted in a wide range of environmental temperatures, without any change in the outcome of the treatment.

Cefmetazole for bacteremia caused by ESBL-producing enterobacteriaceae comparing with carbapenems.

BackgroundESBL (Extended spectrum beta-lactamase) producing enterobacteriaceae are challenging organisms with little treatment options. Carbapenems are frequently used, but the emergence of carbapenem resistant enterobacteriaceae is a concerning issue, which may hinder the use of carbapenems. Although cephamycins such as cefoxitin, cefmetazole or cefotetan are effective against ESBL-producers in vitro, there are few clinical data demonstrating effects against bacteremia caused by these organisms.MethodsWe performed a retrospective observational study on cases of bacteremia caused by ESBL-producers to investigate the efficacy of cefmetazole compared with carbapenems. We also evaluated whether the trend of antibiotic choice changed over years.ResultsSixty-nine patients (male 34, age 69.2 ± 14.4), including two relapse cases, were reviewed for this analysis. The most common causative organisms were Escherichia coli (64, 93 %), followed by Klebsiella pneumoniae and K. oxytoca (2 each, 4 %). The group that received carbapenem therapy (43, 62 %) had increased severity in the Pittsburgh Bacteremic score than the group that received cefmetazole therapy, (1.5 ± 1.5 vs 2.5 ± 2.1, p = 0.048), while analysis of other factors didn’t reveal any statistical differences. Five patients in the carbapenem group and one patient in the cefmetazole group died during the observation period (p = 0.24). CTX-M-9 were predominant in this series (59 %). Infectious disease physicians initially recommended carbapenems at the beginning of the current research period, which gradually changed over time favoring the use of cefmetazole instead (p = 0.002).ConclusionCefmetazole may be safely given to patients with bacteremia caused by ESBL-producers as a definitive therapy, if one can select out relatively stable patients.

Mycobacterium goodii: An Emerging Nosocomial Pathogen: A Case Report and Review of the Literature.

Mycobacterium goodii, a rapidly growing nontuberculous mycobacterium, is an emerging pathogen in nosocomial infections. Its inherent resistance patterns make it a challenging organism to treat, and delays in identification can lead to poor outcomes. We present a case of cardiac device pocket infection with M. goodii, complicated by both antibiotic resistance and drug reactions that highlight the challenges faced by clinicians trying to eradicate these infections. We also present a brief review of the English literature surrounding this disease, including a table of all reported cases of M. goodii infections and their outcomes to act as guide for clinicians formulating treatment plans for these infections. A clear understanding of diagnostic methods and treatment caveats is essential to curing infections caused by these organisms.

Proteome-wide antigen discovery of novel protective vaccine candidates against Staphylococcus aureus infection

Methicillin-resistant Staphylococcus aureus (MRSA) is a rapidly growing problem, especially in hospitals where MRSA cause increased morbidity and mortality and a significant rise in health expenditures. As many strains of MRSA are resistant to other antimicrobials in addition to methicillin, there is an urgent need to institute non-antimicrobial measures, such as vaccination, against the spread of MRSA. With the aim of finding new protective antigens for vaccine development, this study used a proteome-wide in silico antigen prediction platform to screen the proteome of S. aureus strain MRSA252. Thirty-five different S. aureus proteins were identified, recombinantly expressed, and tested for protection in a lethal sepsis mouse model using S. aureus strain MRSA252 as the challenge organism. We found that 13 of the 35 recombinant peptides yielded significant protection and that 12 of these antigens were highly conserved across 70 completely sequenced S. aureus strains.Thus, this in silico platform was capable of identifying novel candidates for inclusion in future vaccines against MRSA.

Adenine and benzimidazole-based mimics of REP-3123 as antibacterial agents against Clostridium difficile and Bacillus anthracis: Design, synthesis and biological evaluation

As part of our ongoing research efforts to develop new antibacterial agents acting on novel molecular targets, a series of twenty-two adenine and benzimidazole-based mimics of the lead compound REP-3123 was designed to target methionyl-tRNA synthetase of Clostridium difficile and Bacillus anthracis based on a homology model. Structures of the target compounds were elucidated by means of 1H and 13C NMR spectral data and their purity was confirmed by HRMS or microanalyses. The target compounds were tested for their in vitro antibacterial activity against those two challenging organisms by the microdilution method in brain heart infusion broth. Unfortunately, six of the target compounds were not biologically tested due to inadequate solubility in DMSO under the assay conditions. Only the fluoro-substituted adenine-based sulfamide (18) showed activity against C. difficile with an MIC of 85.33μg/mL. The adenine-based thiourea (32) and diamine (36) were the most promising antibacterial agents against B. anthracis with an MIC of 92.16μg/mL. The rest of the tested compounds either showed inferior activity (MIC=102.4μg/mL) or were totally inactive. Although the compounds were not very active, the biological data are employed as a basis for our currently underway investigation for structure optimization.

The efficacy of a commercial competitive exclusion product on Campylobacter colonization in broiler chickens in a 5-week pilot-scale study

The efficacy of the commercial competitive exclusion product Broilact against Campylobacter jejuni was evaluated in broiler chickens in a 5-week pilot-scale study. Newly-hatched broiler chicks were brought from a commercial hatchery. After arrival 50 seeder chicks were challenged orally with approximately 103 cfu of C. jejuni, wing marked, and placed back in a delivery box and moved to a separate room. The rest of the chicks (contact chicks) were placed in floor pens, 100 chicks per pen. Birds in two pens were treated orally on the day of hatch with the commercial competitive exclusion (CE) product Broilact, and three pens were left untreated. The following day 10 seeder chicks were introduced into the Broilact treated and untreated control pens. One pen was left both untreated and unchallenged (0-control). Each week the ceca of 10 contact chicks and one seeder chick were examined quantitatively for Campylobacter. The treatment prevented or significantly reduced the colonization of the challenge organism in the ceca during the two first weeks; the percentage of colonized birds being 0% after the first week and 30% after the second week in the Broilact treated groups but was 100% in the control groups the entire 5-week rearing period. During the third rearing week the proportion of Campylobacter positive birds started to increase in the treated pens, being 80% after the third week and 95 and 90% after the fourth and fifth rearing weeks, respectively. Similarly the average count of Campylobacter in the cecal contents of the Broilact treated chicks started to increase, the difference between the treated and control chicks being 1.4 logs at the end of the rearing period. Although the protective effect was temporary and occurred only during the first two weeks of the rearing period, the results of this study support the earlier observations that CE flora designed to protect chicks from Salmonella may also reduce Campylobacter colonization of broiler chickens.

Study on Coinfection of Mycoplasma gallisepticum and Low Pathogenic Avian Influenza Virus H9 in Broilers

Avian influenza viruses (AIVs) belong to the family Orthomyxoviridae which consists of three genera known as Influenza virus -A, -B and -C. The influenza viruses, which can cause infections in avian species belong to genus influenza virus A which can produce two different types of diseases on the basis of their virulence and pathogenicity. Thus are classified as a highly pathogenic avian influenza virus (HPAIV) and a low pathogenic avian influenza virus (LPAIV). Recent outbreaks of LPAIV H9N2 in Asian countries such as in Pakistan and China revealed that this subtype has become endemic in poultry population of these countries. Meager literature is available about its pathogenicity. This study was made to ascertain the pathogenesis of H9 subtype AIVs in broilers along with most prevalent respiratory pathogen Mycoplasma gallisepticum under field conditions. In first part of study, challenge organisms (H9 virus and Mycoplasma gallisepticum) were isolated from field samples and experimental inoculum was prepared. In second part, pathogenesis of H9 virus in association with M. gallisepticum was carried out under controlled environmental conditions. The virus prevailing in Pakistan was found nonpathogenic as only slight diarrhoea and depression in few birds was seen while clinical signs and lesions produced grossly and microscopically were more severe in form of severe conjunctivitis and facial edema and respiratory involvement with hemorrhagic and pneumonic lungs and degenerative and lytic necrosis in kidneys in birds infected co-infected with H9 virus and Mycoplasma gallisepticum as compared to other groups which were infected with single pathogens. Immunohistochemical detection of viral antigen revealed that replication of virus was more severe in birds infected with H9 virus and Mycoplasma gallisepticum as compared to other groups. In conclusion, the virus subtype H9 circulating in Pakistan is low pathogenic in nature with affinity for respiratory system and urinary system while coinfection with Mycoplasma gallisepticum is the most important factor enhancing the pathogenicity of H9 virus under field conditions.

Antimicrobial Activity of a Novel Vascular Access Film Dressing Containing Chlorhexidine Gluconate

BackgroundCovering insertion sites with chlorhexidine impregnated dressings has been proven to be clinically effective in reducing catheter related blood stream infections (CR-BSI). Two chlorhexidine gluconate (CHG)-impregnated dressings are commercially available, a polyurethane foam disk and a film dressing containing a chlorhexidine gluconate-impregnated gel pad. While both have demonstrated efficacy in clinical settings, the major drawback of high cost and impaired IV insertion site visibility limits their usage. A new, simple film dressing containing CHG within its adhesive layer is now available. The objective of this study was to test the in vitro antimicrobial efficacy of the new dressing in comparison to the CHG-impregnated gel dressing.MethodsQuantitative aliquots of suspensions (concentration of 1.0x106 to 5.0x106 cfu/sample) of clinically relevant challenge organisms (Staphylococcus species, gram-negative bacilli, Candida albicans) were incubated in contact with the new CHG-containing film dressing, a placebo version of the same (negative control) and the commercially available CHG-impregnated gel dressing (positive control). Serial dilutions of the surviving organisms were quantified using the pour plate after 1, 3, 5, and 7 days of incubation in order to calculate an antimicrobial log10 reduction for each organism/dressing combination at each point in time.ResultsThe new CHG-containing film dressing delivered greater than 5.0 log10 reduction throughout the 7 days on all aerobic gram-negative bacilli and Staphylococcus species tested. As of day 1 the CHG-containing film dressing provided greater than 5.0 log10 reduction on Candida albicans. There were no statistically significant differences in the log10 reduction between the two dressings tested.ConclusionThe new CHG-containing film dressing was found to be as effective as the chlorhexidine gluconate-impregnated gel dressing on clinically relevant microbes.

Laboratory replication of filtration procedures associated with Serratia marcescens bloodstream infections in patients receiving compounded amino acid solutions.

Specific deviations from United States Pharmacopeia standards were analyzed to investigate the factors allowing an outbreak of Serratia marcescens bloodstream infections in patients receiving compounded amino acid solutions. Filter challenge experiments using the outbreak strain of S. marcescens were compared with those that used the filter challenge organism recommended by ASTM International (Brevundimonas diminuta ATCC 19162) to determine the frequency and degree of organism breakthrough. Disk and capsule filters (0.22- and 0.2-μm nominal pore size, respectively) were challenged with either the outbreak strain of S. marcescens or B. diminuta ATCC 19162. The following variables were compared: culture conditions in which organisms were grown overnight or cultured in sterile water (starved), solution type (15% amino acid solution or sterile water), and filtration with or without a 0.5-μm prefilter. Small-scale, syringe-driven, disk-filtration experiments of starved bacterial cultures indicated that approximately 1 in every 1,000 starved S. marcescens cells (0.12%) was able to pass through a 0.22-μm nominal pore-size filter, and about 1 in every 1,000,000 cells was able to pass through a 0.1-μm nominal pore-size filter. No passage of the B. diminuta ATCC 19162 cells was observed with either filter. In full-scale experiments, breakthrough was observed only when 0.2-μm capsule filters were challenged with starved S. marcescens in 15% amino acid solution without a 0.5-μm prefiltration step. Laboratory simulation testing revealed that under certain conditions, bacteria can pass through 0.22- and 0.2-μm filters intended for sterilization of an amino acid solution. Bacteria did not pass through 0.2-μm filters when a 0.5-μm prefilter was used.

Novel in situ liquefying antimicrobial wrap for preventing tissue expander infections following breast reconstructive surgeries.

Breast reconstruction surgeries using tissue expanders (TEs) have highly reported infection rates. To decrease this, we developed a method for disinfecting TEs and surgical pockets, where an antimicrobial solution was applied as a solid film at implantation that subsequently liquefied in situ to provide extended prophylaxis. Silicone discs cut from TEs were covered with gelatin-based films containing minocycline (M) and rifampin (R). Discs and films soaked in saline were subsequently challenged with pathogen at days 1, 3, 7, and 10 and quantified for potential biofilm formation. Discs that were not harvested at each specific time points were refreshed with sterile saline. The discs were challenged with clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant Staphylococcus epidermidis (MRSE), and multidrug-resistant Pseudomonas aeruginosa (MDR-PA). Recoveries of adherent organisms from uncovered silicone discs and gelatin-wrapped discs without added antimicrobial agents were >5 × 10(4) CFU/disc for each organism at each time point. Experimental 0.1%M/0.05%R gelatin films completely inhibited all challenge organisms from attaching to the silicone (p < 0.05) at each time point through day 10. Cytotoxicity was assessed by incubating films with HEK-293T human fibroblasts. There were no significant differences in HEK-293T cell survival between controls and any of the antimicrobial films. The in situ liquefying, bioabsorable, antimicrobial wrap prevented biofilm formation by microorganisms on silicone surfaces in vitro with minimal cytotoxicity.

The impact of dose, irradiance and growth conditions on Aspergillus niger (renamed A. brasiliensis) spores low-pressure (LP) UV inactivation

The use of Aspergillus niger (A. niger) fungal spores as challenge organism for UV reactor validation studies is attractive due to their high UV-resistance and non-pathogenic nature. However A. niger spores UV dose-response was dependent upon sporulation conditions and did not follow the Bunsen–Roscoe Principle of time-dose reciprocity. Exposure to 8 h of natural sunlight for 10 consecutive days increased UV resistance when compared to spores grown solely in dark conditions. Application of 250 mJ cm−2 at high irradiance (0.11 mW cm−2) resulted in a 2-log inactivation; however, at low irradiance (0.022 mW cm−2) a 1-log inactivation was achieved. In addition, surface electron microscopy (SEM) images revealed morphological changes between the control and UV exposed spores in contrast to other well accepted UV calibrated test organisms, which show no morphological difference with UV exposure.

RT-PCR testing of allograft musculoskeletal tissue: is it time for culturebased methods to move over?

Allograft musculoskeletal tissue samples are assessed for microbial bioburden to reduce the risk of post-transplant infection. Traditionally, solid agar and broth culture media have been used however, nucleic acid testing, such as real-time (RT) polymerase chain reaction (PCR), has been described as more sensitive. This study evaluated the recovery of low numbers of challenge organisms from inoculated swab and musculoskeletal biopsy samples using solid agar culture, cooked meat medium, blood culture bottles and a RT-PCR assay. It was found that broth culture methods were the most sensitive with RT-PCR unable to detect low numbers of bacteria from these samples. Investigation of other non-culture methods may be worthwhile.