A quantitative method to assess the role of indoor air decontamination to simultaneously reduce contamination of environmental surfaces: testing with vegetative and spore-forming bacteria.

Indoor air can spread pathogens, which can be removed/inactivated by a variety of means in healthcare and other settings. We quantitatively assessed if air decontamination could also simultaneously reduce environmental surface contamination in the same setting. Two types of vegetative bacteria (Staphylococcus aureus and Acinetobacter baumannii), and a bacterial spore-former (Geobacillus stearothermophilus) were tested as representative airborne bacteria. They were separately aerosolized with a Collison nebulizer into a 24-m3 aerobiology chamber and air samples collected with a programmable slit-to-agar sampler. Settling airborne particles were collected on culture plates placed at, and collected from, five different locations on the floor of the chamber with a custom-built remote plate-placement and -retriever system. Experimentally contaminated air in the chamber was decontaminated for 45 min with a device based on HEPA filtration and UV light. The plates were incubated and CFU counted. The device reduced the viability levels of all tested bacteria in the air by >3 log10 (>99·9%) in 45 min. Based on two separate tests, the average reductions in surface contamination for S. aureus, A. baumannii and G. stearothermophilus were respectively, 97, 87 and 97%. We thus showed that air decontamination could substantially and simultaneously reduce the levels of surface contamination in the same setting irrespective of the type of pathogen present. SIGNIFICANCE AND IMPACT OF THE STUDY: The innovative and generic test protocol described can quantitatively assess the reduction in environmental surface contamination from microbial decontamination of indoor air in the same setting. This added advantage from air decontamination has implications for infection prevention and control in healthcare and other settings without the need for additional expense or effort. Continuous operation of an air decontamination device, such as the one tested here, can lead to ongoing reductions in pathogens in air and on environmental surfaces.

Assessing the activity of microbicides against bacterial spores: knowledge and pitfalls.

Bacterial endospores (spores) have a higher intrinsic resistance to microbicides as compared to other microbial forms, most likely due to their impermeable outer layers and low water content. Though structural differences between the spores of various bacterial species may account for observed variations in their resistance to microbicides, flaws in methods for testing the sporicidal activity of microbicides often exaggerate the differences. This has major implications when considering the selection of one or more surrogates to assess microbicides against clinically relevant spore-formers such as Clostridium difficile. The mounting significance of Cl. difficile as a pathogen is leading to a corresponding increase in the number of commercially available microbicidal formulations claiming activity against its spores without proper differentiation between the product's sporistatic and sporicidal actions. In this review we critically assess the situation and the implications of product claims on the field use of microbicidal products.

Biological control of Phlebotomus papatasi larvae by using entomopathogenic nematodes and its symbiotic bacterial toxins.

The sand fly Phlebotomus papatasi is an important disease-bearing vector. Five entomopathogenic nematodes (EPNs) - Steinernema carpocapsae DD136, Steinernema sp. (SII), S. carpocapsae all, S. abbasi, and Heterorhabditis bacteriophora HP88 - were applied as biocontrol agents against the late third instar larvae of P. papatasi. In addition, the effect of toxin complexes (TCs) of Xenorhabdus nematophila and Photorhabdus luminescens laumondii bacteria was evaluated. Results revealed that S. carpocapsae DD136 was the most virulent species followed by Steinernema sp. (SII) and S. carpocapsae all where LC50 were 472, 565, 962 IJs/ml, respectively. Also, the crude TCs were slightly more active and toxic than their fractionated protein. Histopathological examination of infected larvae with H. bacteriophora HP88 showed negative effect on their midgut cells. In conclusion, EPNs with their symbiotic bacteria are more effective as biocontrol agents than the crude or fractionated TCs against sand fly larvae.

Chloraminated drinking water does not generate bacterial resistance to antibiotics in Pseudomonas aeruginosa biofilms.

AIM: To determine if exposure of Pseudomonas aeruginosa biofilms to chloraminated drinking water can lead to individual bacteria with resistance to antibiotics. METHODS AND RESULTS: Biofilms of P. aeruginosa PA14 were grown in drinking water in a Kadouri drip-fed reactor; the biofilms were treated with either 0.5 mg l(-1) or 1.0 mg l(-1) of chloramine for 15 or 21 days; control biofilms were grown in water without chloramine. Fewer isolates with antibiotic resistance were obtained from the chloramine-treated biofilms as compared to the control. Minimum inhibitory concentrations (MIC) for selected antibiotic-resistant isolates were determined using ciprofloxacin, tobramycin, gentamicin, rifampicin and chloramphenicol. All of the isolates tested had increased resistance over the wildtype to ciprofloxacin, rifampicin and chloramphenicol, but were not resistant to tobramycin or gentamicin. CONCLUSIONS: Under these test conditions, there was no detectable increase in antibiotic resistance in P. aeruginosa exposed as biofilms to disinfectant residues in chloraminated drinking water. SIGNIFICANCE AND IMPACT OF THE STUDY: Chloramine in drinking water, while unable to kill biofilm bacteria, does not increase the potential of P. aeruginosa to become resistant to antibiotics.

Survival of calicivirus in foods and on surfaces: experiments with feline calicivirus as a surrogate for norovirus.

Although there is a large body of evidence incriminating foods as vehicles in the transmission of norovirus, little is known about virus survival in foods and on surfaces. Feline calicivirus was used as a surrogate for norovirus to investigate its survival in representative foods of plant and animal origin and on metal surfaces. Known concentrations of feline calicivirus in a natural fecal suspension were deposited onto lettuce, strawberries, ham, or stainless steel and incubated for 7 days at refrigeration or room temperatures. Virus was recovered at 1-day intervals, and the titers of the virus were determined by plaque assay. Infectious virus was recoverable until day 7 from lettuce, ham, and stainless steel. Statistically higher titers of feline calicivirus (P < 0.05) were recovered from ham under all conditions than from lettuce, strawberries, or stainless steel. These data provide valuable information for epidemiological and monitoring purposes as well as for the development of food processing practices and appropriate strategies to inactivate norovirus and control its transmission via foods and surfaces.

Virucidal activity of a new hand disinfectant with reduced ethanol content: comparison with other alcohol-based formulations.

A new formula with reduced ethanol content (55%) in combination with 10% propan-1-ol, 5.9% propan-1.2-diol, 5.7% butan-1.3-diol and 0.7% phosphoric acid exhibited a broad spectrum of virucidal activity. In quantitative suspension tests, with and without protein load, this formulation reduced the infectivity titre of seven enveloped (influenza A and B, herpes simplex 1 and 2, bovine corona, respiratory syncytial, vaccinia, hepatitis B, bovine viral diarrhoea) and four non-enveloped (hepatitis A, polio, rota, feline calici) viruses >10(3)-fold within 30s. In comparative testing, only 95% ethanol showed similar levels of activity. In fingerpad tests, the formulation produced a log10 reduction factor of the titre of poliovirus type 1 (Sabin) of 3.04 in 30s compared with 1.32 by 60% propan-2-ol. Testing against feline calicivirus produced a log10 reduction factor of 2.38 by the test formulation; in contrast, the log10 reduction factors with 70% ethanol and 70% propan-1-ol were 0.68 and 0.70, respectively.

Predicting serum gastrin levels among men during Ramadan fasting.

Increased gastric acidity is common when fasting during Ramadan. Our study aimed to develop a regression equation to predict fasting serum gastrin levels using parameters commonly analysed in clinical laboratories. Fasting blood samples from six men were taken on days 1, 10, 19, 26 and 28 of Ramadan. Serum gastrin, total cholesterol, urea and uric acid were analysed. All 5 samples from each man were included in multiple regression analysis and the prediction equation obtained was: serum gastrin, pg/mL = 198.27-0.199 total cholesterol (mg/dL) + 2.525 urea (mg/dL)--103.238 uric acid (mg/dL) + 10.923 uric acid (mg/dL)2 + 3.683 body mass index, r2 = 0.75, P < 0.001. This equation might be used to estimate gastrin levels and plan dietary and medicinal measures to avoid high gastric acidity during Ramadan.

Disinfectant wipes are appropriate to control microbial bioburden from surfaces: use of a new ASTM standard test protocol to demonstrate efficacy.

BACKGROUND: The use of disinfectant pre-soaked wipes (DPW) to decontaminate high-touch environmental surfaces (HTES) by wiping is becoming increasingly widespread in the healthcare environment. However, DPW are rarely tested using conditions simulating their field use, and the label claims of environmental surface disinfectants seldom include wiping action. AIM: To evaluate the new E2967-15 standard test specific to wipes, particularly their ability to decontaminate surfaces and to transfer acquired contamination to clean surfaces. METHODS: ASTM Standard E2967-15 was used by three independent laboratories to test the efficacy of five types of commercially available wipe products. All data generated were pulled together, and reproducibility and repeatability of the standard were measured. FINDINGS: All the commercial DPW tested achieved a >4log10 (>99.99%) reduction in colony-forming units (CFU) of Staphylococcus aureus and Acinetobacter baumanii with 10s of wiping, but only one DPW containing 0.5% accelerated H2O2 prevented the transfer of bacteria to another surface. CONCLUSION: This newly introduced standard method represents a significant advance in assessing DPW for microbial decontamination of HTES, and should greatly assist research and development, and in making more relevant and reliable claims on marketed DPW.

Biological control of Phlebotomus papatasi larvae by using entomopathogenic nematodes and its symbiotic bacterial toxins

@# The sand fly Phlebotomus papatasi is an important disease-bearing vector. Five entomopathogenic nematodes (EPNs) – Steinernema carpocapsae DD136, Steinernema sp. (SII), S. carpocapsae all, S. abbasi, and Heterorhabditis bacteriophora HP88 – were applied as biocontrol agents against the late third instar larvae of P. papatasi. In addition, the effect of toxin complexes (TCs) of Xenorhabdus nematophila and Photorhabdus luminescens laumondii bacteria was evaluated. Results revealed that S. carpocapsae DD136 was the most virulent species followed by Steinernema sp. (SII) and S. carpocapsae all where LC50 were 472, 565, 962 IJs/ml, respectively. Also, the crude TCs were slightly more active and toxic than their fractionated protein. Histopathological examination of infected larvae with H. bacteriophora HP88 showed negative effect on their midgut cells. In conclusion, EPNs with their symbiotic bacteria are more effective as biocontrol agents than the crude or fractionated TCs against sand fly larvae.

Neutralizing monoclonal antibody against enterovirus-70 reacts with viral proteins 1C and 1D.

A library of murine monoclonal antibodies against the prototype Enterovirus-70 (EV-70) strain, J670/71, was made for the purpose of studying the immunologically reactive determinants of the virus. Each of the monoclonal antibodies reacted with several other strains of Enterovirus-70 when tested by immunofluorescence. However, none of these monoclonal antibodies reacted with any other picornavirus tested. It was found that all of the monoclonal antibodies precipitated EV-70 viral proteins 1C and 1D in radio-immunoprecipitation assays. However, only one of these monoclonal antibodies, an IgG3 kappa, was capable of neutralizing the virus.

Activity of an alcohol-based hand gel against human adeno-, rhino-, and rotaviruses using the fingerpad method.

OBJECTIVE: To assess the activity against three non-enveloped viruses (an adeno-, a rhino- and a rotavirus) of a gel containing 60% ethanol, using experimentally contaminated thumb- and fingerpads of 12 panelists, as per standard procedure E-1838-96 of the American Society of Testing and Materials. DESIGN: Each digit received 10 microL of the test virus suspension. The inoculum from the thumbs was eluted immediately with 990 microL of Earle's balanced salt solution (EBSS) to assess the amount of virus on each digit (0-minute control). The inoculum on the fingers was allowed to dry (20-25 minutes), and virus was eluted from two fingerpads to determine the loss in virus infectivity upon drying (baseline titer). Then the dried inoculum on randomly selected fingers was exposed to 1 mL of the test product or standard hard water (200-ppm calcium carbonate) for 20 seconds. The virus remaining was eluted with 1 mL of EBSS, titrated to determine the amounts eliminated, and compared to the baseline titer. RESULTS: Each digit received at least 10(4) plaque-forming units of virus in 10 microL. The amounts of adeno-, rhino-, and rotaviruses surviving the drying were 30%, 75%, and 42%, respectively. The product reduced the infectivity titers of the three viruses by 3 to >4 log10 when compared to a reduction of < or =1 log10 for the hard-water rinse. CONCLUSION: The level of virus reduction by gel was statistically significantly higher than that seen with the water control. Evidence for such activity against non-enveloped viruses supports further investigation of the benefits of this product.

Interruption of rotavirus spread through chemical disinfection.

INTRODUCTION: Rotaviruses, which are among the most important infectious causes of acute diarrhea, frequently cause outbreaks in hospitals, daycare centers, schools, and nursing homes. These viruses can remain viable on inanimate surfaces for many days and infectious rotavirus particles have been recovered from hands and a variety of surfaces and objects. Casual contact can lead to the transfer of these viruses from contaminated to clean surfaces. Therefore, animate and inanimate surfaces may play a complementary role in the spread of these viruses. OBJECTIVE: In this study, we compared the capacity of a disinfectant spray (0.1% o-phenylphenol and 79% ethanol), a domestic bleach (6% sodium hypochlorite diluted to give 800 ppm free chlorine), a quarternary ammonium (quat)-based product (7.05% quat diluted 1:128 in tap water), and a phenol-based product (14.7% phenol diluted 1:256 in tap water) to interrupt the transfer of a human rotavirus (DS-1) from stainless steel disks to fingerpads of volunteers with a 10-second contact at a pressure of 1 kg/cm2. DESIGN: Each disk received a 10 microL inoculum containing 1.0 x 10(4) to 7.0 x 10(4) plaque-forming units (PFU) of the virus suspended in 10% feces. The inoculum was dried for 1 hour and overlaid with 20 microL of either tap water or the test product. RESULTS: A 10-minute exposure to tap water reduced the virus titer by 52.3% +/- 11.7%. The disinfectant spray was able to reduce virus infectivity by > 99.99% after a contact of 3 to 10 minutes. The loss in virus infectivity after a 10-minute treatment with the quat was almost the same (54.7% +/- 17.8%) as seen with tap water. The activities of the bleach and the phenolic were very similar with losses in PFU of 97.9% +/- 0.4% and 95% +/- 5.36%, respectively. No detectable virus was transferred to fingerpads from disks treated with disinfectant spray, the bleach, and the phenolic. Contact of the fingerpads with tap water- or quat-treated disks resulted in the transfer of 5.6% +/- 1.1% and 7.6% +/- 2.5% of the remaining infectious virus, respectively. CONCLUSION: These findings emphasize the care needed in the selection of environmental surface disinfectants in preventing the spread of rotaviral infections.

Feasibility of a combined carrier test for disinfectants: studies with a mixture of five types of microorganisms.

BACKGROUND: There is mounting concern regarding the efficacy of many germicides on the market because officially recognized germicidal tests for various classes of microorganisms vary widely and often lack reproducibility and proper quantitation. We report here a carrier method for simultaneously and quantitatively assessing the efficacy of liquid chemical germicides against a mixture of microorganisms of varying degrees of resistance. METHODS: In the test, each small glass cup (10 mm wide x 14 mm long) was contaminated with 10 microliters of a standardized mixture of Staphylococcus aureus, Mycobacterium bovis bacille Calmette-Guérin, Trichophyton mentagrophytes spores, Sabin poliovirus type 1, and Bacillus stearothermophilus spores in 5% fetal bovine serum. The inoculum was dried for 60 minutes under ambient conditions and covered with 60 microliters of the disinfectant under test or a balanced salt solution control for the desired contact time. The carrier was then placed in 2940 microliters of an eluent and the eluates assayed separately for the five microorganisms. Tap water was used to dilute the test product as needed. RESULTS: Of the 11 products tested, 2% alkaline glutaraldehyde, 0.6% sodium hypochlorite (about 5000 ppm free chlorine), and a 0.4% quarternary ammonium compound containing 23% hydrochloric acid were effective against all five challenge organisms. A hard-surface spray containing 0.1% o-phenylphenol with 79% ethanol was effective against all but bacterial spores; 70% (volume/volume) ethanol alone and povidone-iodine (1% available iodine) were effective against S. aureus, the mycobacterium, and the fungus; a 3% solution of peroxygen compounds was effective only against S. aureus and the poliovirus; 1.5% chlorhexidine gluconate, 0.06% quaternary ammonia compound, and 0.03% o-phenylphenol + 0.03% p-tertiary amylphenol could inactivate nothing but S. aureus; and 3% hydrogen peroxide was ineffective in all tests. CONCLUSIONS: This method shows promise for use with various classes of microorganisms, individually or as mixtures. Its application should enable the classification of germicides according to spectrum of activity.

Impact of changing societal trends on the spread of infections in American and Canadian homes.

Infectious diseases continue to exert a heavy toll on human health even in industrialized countries. Recent data from the World Health Organization suggests that infectious diseases are the leading cause of death in the world. Many changing trends in our society have a known or potential impact on infectious disease spread and may have an impact on the normal routine of home hygiene. Important amongst these societal trends are increasing population and life expectancy, changes in urbanization, grouping of susceptibles, increased ambulatory and home care, increased immunosuppression, increased and faster travel, changes in technology, increasing antibiotic resistance as a result of misuse of antibiotics, changes in food and water consumption, and changes in personal cleaning, washing, and laundry practices. This review will highlight these factors and their impact on home hygiene and steps that may be needed to reduce the risk from infections.

Preventing the spread of hepatitis B and C viruses: where are germicides relevant?

Hepatitis B virus (HBV) and hepatitis C virus (HCV) are the most prevalent bloodborne pathogens. Infections caused by these organisms can become chronic and may lead to liver cirrhosis and carcinoma. Limited chemotherapy is now available, but only HBV can be prevented through vaccination. Both viruses are enveloped and relatively sensitive to many physical and chemical agents; their ability to survive in the environment may not be as high as often believed. As a result, their spread occurs mainly through direct parenteral or percutaneous exposure to tainted body fluids and tissues. Careful screening of and avoiding contact with such materials remain the most effective means of protection. Nevertheless, the indirect spread of these viruses, although much less common, can occur when objects that are freshly contaminated with tainted blood enter the body or contact damaged skin. Germicidal chemicals are important in the prevention of HBV and HCV spread through shared injection devices, sharps used in personal services (such as tattooing and body piercing), and heat-sensitive medical/dental devices (such as flexible endoscopes) and in the cleanup of blood spills. Microbicides in vaginal gels may also interrupt their transmission. General-purpose environmental disinfection is unlikely to play a significant role in the prevention of the transmission of these viruses. Testing of low-level disinfectants and label claims for such products against HBV and HCV should be discouraged. Both viruses remain difficult to work with in the laboratory, but closely related animal viruses (such as the duck HBV) and the bovine viral diarrhea virus show considerable promise as surrogates for HBV and HCV, respectively. Although progress in the culturing of HBV and HCV is still underway, critical issues on virus survival and inactivation should be addressed with the use of these surrogates.

Comparison of cloth, paper, and warm air drying in eliminating viruses and bacteria from washed hands.

We compared the efficiency of paper, cloth, and electric warm air drying in eliminating rotaviruses and Escherichia coli remaining on finger pads washed with 70% isopropanol, a medicated liquid soap, an unmedicated liquid soap, or tap water alone. The contaminated area on the finger pads of a volunteer was exposed to the hand-washing agent for 10 seconds and then rinsed in 40 degrees C tap water. The washed areas were dried for 10 seconds by one of the three methods. Irrespective of the hand-washing agent used, electric air drying produced the highest and cloth drying the lowest reduction in the numbers of both test organisms. These findings indicate the importance of selecting the right means for drying washed hands, particularly when less effective hand-washing agents are used.

Reduced longevity and fecundity in Leishmania-infected sand flies.

Phlebotomus papatasi and P. langeroni were infected with Leishmania major and L. infantum by membrane feeding. Each sand fly ingested approximately 200 parasites per blood meal. Higher mortality in both sand fly species was seen with mixed infections than with a single parasite species. There was no significant difference between infections with either L. major or L. infantum in their natural vectors or experimental hosts. Infection significantly depressed the mean number of eggs laid per female.

Rapid concentration and detection of hepatitis A virus from lettuce and strawberries.

Immunomagnetic beads-PCR (IM-PCR), positively-charged virosorb filters (F), or a combination of both methods (F-IM-PCR) were used to capture, concentrate and rapidly detect hepatitis A virus (HAV) in samples of lettuce and strawberries experimentally contaminated. Direct reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of the collected HAV-beads complex showed a detection limit of 0.5 plaque forming units (PFU) of the virus present in 1-ml of wash solution from the produce, which was several hundred-fold more sensitive than that demonstrated by RT-PCR. In separate trials, virus-containing wash solutions from the produce were passed through the filters and the captured virus was eluted with 10 ml volumes of 1% beef extract. Of the 62% filter-captured HAV, an average of 34.8% was eluted by the 1% beef extract. PCR amplification of 2 microl from this eluate failed to produce a clear positive band signal. As little as 10 PFU, present on each piece of the lettuce or strawberry, was detectable by the F-IM-PCR, which was almost 20 times less sensitive than the detection limit of 0.5 PFU by the IM-PCR. However, considering the large volumes (< or =50 ml) used in the F-IM-PCR, the sensitivity of detection could be much greater than that of the IM-PCR, which was restricted to < or =20 ml volumes. These data indicate that the F-IM-PCR method provides the potential for a greater sensitivity of detection than the IM-PCR, since low levels of virus could be detected from large volumes of sample than possible by the IM-PCR method. Although positively-charged filters captured a greater amount of virus than both the IM-PCR and F-IM-PCR methods, direct PCR amplification from beef extract eluates was not successful in detecting HAV from produce.

Rotavirus concentration from raw water using positively charged filters.

Positively charged (PC) Zeta Plus (30 S) filters and talc-Celite (TC) layers were tested for their ability to concentrate human rotavirus (strain Wa) from seeded samples of raw water (RW), collected from the Ottawa River. In the TC technique, the RW sample was preconditioned by adding Earle's balanced salt solution (1:100) and reducing the pH to 6.0 before passing it through the TC layer. Almost 6% of the input virus was lost in the filtrate. Using 1 X tryptose phosphate broth (TPB) at pH 9.0 as an eluent, 48% of the virus could be recovered from the layers. With the 30 S PC filters, no sample preconditioning was required and no virus was detected in the filtrates. In virus elution from these filters, 1 X TPB (pH 9.0), and a mixture of 6% glycine + 6% arginine (pH 9.0) gave virus recoveries of 44% and 46%, respectively. When these same two eluents were used in sequence to recover the rotavirus from 30 S PC filters, a virus recovery of about 70% was obtained. However, when 20 1 vols. of seeded raw water were concentrated using 30 S PC filters (142 mm diameter), and polyethylene glycol hydroextraction was used for second-step concentration of the eluates, only 16% of the input infectious virus could be recovered. The 30 S PC filters were then used to recover naturally occurring rotaviruses from the waters of the Ottawa River. After concentration, these viruses were detected and quantitated in MA-104 cells using an indirect immunoperoxidase staining technique.

Development of methods to study the survival of airborne viruses.

A number of viruses have been shown to be transmitted by the airborne route. It is the ability of these viruses to retain their infectivity for living hosts which play a key role in their aerial dissemination. Data generated by a number of workers on the airborne survival of viruses varies considerably because laboratory techniques have not been standardized. About 5 yr ago we started studies on the airborne survival of a number of animal and human viruses. This paper describes the methodology developed to study the aerobiology of these viruses. These methods should be useful in the aerobiological work of other viruses.