Effects of biocidal treatments to inhibit the growth of legionellae and other microorganisms in cooling towers.

The effects of biocidal treatments for cooling towers were examined through the use of chemicals and ultraviolet irradiation to inhibit the growth of legionellae and other microorganisms. In the water of cooling towers without continuous biocidal treatments, heterotrophic bacteria and bacterivorous protozoan first appeared, and then legionellae increased up to 10(4) CFU/100 ml. When a UV sterilizer was connected to the cooling tower, the legionellae count was 1/10 or 1/100 of that in the nontreated tower water. In the water of towers supplemented continuously with the biocidal chemicals, legionellae were not found during a 4-month period. The biocidal treatments tested were proved to suppress the increase of legionellae in cooling-tower water, and thus are useful in preventing the outbreak of legionellosis due to inhalation of contaminated aerosol from the cooling tower system.     

Legionella spp. in Puerto Rico cooling towers

Water samples from air conditioning cooling towers receiving different treatment protocols on five large municipal buildings in San Juan, P.R., were assayed for various Legionella spp. and serogroups by using direct immunofluorescence. Several water quality parameters were also measured for each sample. Guinea pigs were inoculated with water samples to confirm pathogenicity and recover viable organisms. Legionella pneumophila serogroups 1 to 6, L. bozemanii, L. micdadei, L. dumoffii, and L. gormanii were observed in at least one of the cooling towers. L. pneumophila was the most abundant species; its density reached 10(5) cells per ml, which is within the range that is considered potentially pathogenic to humans. A significantly higher density of L. pneumophila was observed in the cooling tower water that was not being treated with biocides. Percent respiration (INT) and total cell activity (acridine orange direct count) were inversely correlated with bacterial density. This study demonstrates that Legionella spp. are present in tropical air-conditioning cooling systems and that, without continuous biocide treatment, they may reach densities that present a health risk.     

Legionella spp. in Puerto Rico cooling towers.

Water samples from air conditioning cooling towers receiving different treatment protocols on five large municipal buildings in San Juan, P.R., were assayed for various Legionella spp. and serogroups by using direct immunofluorescence. Several water quality parameters were also measured for each sample. Guinea pigs were inoculated with water samples to confirm pathogenicity and recover viable organisms. Legionella pneumophila serogroups 1 to 6, L. bozemanii, L. micdadei, L. dumoffii, and L. gormanii were observed in at least one of the cooling towers. L. pneumophila was the most abundant species; its density reached 10(5) cells per ml, which is within the range that is considered potentially pathogenic to humans. A significantly higher density of L. pneumophila was observed in the cooling tower water that was not being treated with biocides. Percent respiration (INT) and total cell activity (acridine orange direct count) were inversely correlated with bacterial density. This study demonstrates that Legionella spp. are present in tropical air-conditioning cooling systems and that, without continuous biocide treatment, they may reach densities that present a health risk.     

Legionella in industrial cooling towers: monitoring and control strategies

Aims: Legionella contamination of industrial cooling towers has been identified as the cause of sporadic cases and outbreaks of legionellosis among people living nearby. To evaluate and control Legionella contamination in industrial cooling tower water, microbiological monitoring was carried out to determine the effectiveness of the following different disinfection treatments: (i) continuous chlorine concentration of 0·01 ppm and monthly chlorine shock dosing (5 ppm) on a single cooling tower; (ii) continuous chlorine concentration of 0·4 ppm and monthly shock of biocide P3 FERROCID 8580 (BKG Water Solution) on seven towers. Methods and Results: Legionella spp. and total bacterial count (TBC) were determined 3 days before and after each shock dose. Both strategies demonstrated that when chlorine was maintained at low levels, the Legionella count grew to levels above 10⁴ CFU l^?1 while TBC still remained above 10⁸ CFU l^?1. Chlorine shock dosing was able to eliminate bacterial contamination, but only for 10–15 days. Biocide shock dosing was also insufficient to control the problem when the disinfectant concentration was administered at only one point in the plant and at the concentration of 30 ppm. On the other hand, when at a biocide concentration of 30 or 50 ppm was distributed throughout a number of points, depending on the plant hydrodynamics, Legionella counts decreased significantly and often remained below the warning limit. Moreover, the contamination of water entering the plant and the presence of sediment were also important factors for Legionella growth. Conclusions: For effective decontamination of outdoor industrial cooling towers, disinfectants should be distributed in a targeted way, taking into account the possible sources of contamination. Significance and Impact of the Study: The data of the research permitted to modify the procedure of disinfection for better reduce the water and aerosol contamination and consequently the exposure risk.     

Studies of microbiological contamination of ultrasonic apparatus used in pediatric aerosol therapy

It was found that inflating tube is most rarely contaminated with microorganisms during the use of ultrasonic inhalator TUR USI 70. Glass cylinder is contaminated more frequently whereas a diaphragm, aerosol preparation, inhaling mask and a pipe joining it with the device are contaminated most frequently. Sporadic contamination of the inflating tube indicate an efficient work of air filters while frequent contamination of the diaphragm, aerosol preparation and glass cylinder prove that the contamination is caused by a coupling fluid. It was also found that ultrasound exerts a destructive effect on microorganisms in the aerosol preparation. The investigations have shown that the inhaling mask and tubes joining it with the device should be changed before each use while the other parts of an inhalator and aerosol preparation may be changed once per 15 inhalations. It was also noted that disinfection of different parts of the device by a 2% aqueous glutaric aldehyde (30 minutes at room temperature) is efficient in about 95%.     

Searching <Emphasis Type="Italic">Simkania negevensis</Emphasis> in environmental waters

Simkania negevensis is an obligate intracellular bacterium grouped into the order Chlamydiales. This new amoeba-resistant bacterium represents a novel aetiologic agent of bronchiolitis and community-acquired pneumonia in both adults and children. It has been suggested that Simkania could be an ubiquitous microorganism presented in water environments. In the natural history of infections with amoeba-related bacteria encountered in aquatic habitats, the transmissions by environmental aerosols or contaminated water/air systems have been extensively recognized. Therefore, understanding the feasibility of Simkania infection by these or similar routes is relevant. In the present work, we investigated the prevalence of this novel disease-associated microorganism in water samples from different sources by real-time PCR (qPCR). Our results show Simkania detection in 5 of 185 water analyzed samples (2.7%: 2 of 88 cooling towers and 3 of 8 waste water samples). However, no Simkania was detected in a drinking water.     

A survey of aflatoxin B<Subscript>1</Subscript> and total aflatoxin contamination in baby food,peanut and corn products sold at retail in Indonesia analysed by ELISA and HPLC

Aflatoxin contamination has been well known as a world-wide health-threatening problem in tropical countries including Indonesia. This research was undertaken to determine the degree of aflatoxin contamination in different Indonesian foodstuffs. A preliminary survey was carried out to evaluate the level of total aflatoxin (AfT) and aflatoxin B₁ (AfB₁) contamination of baby foods, peanut products, and corn products, which were purchased from traditional markets and supermarkets in Indonesia during the year 2001-2002. Eighty two peanut products, 12 baby foods products, and 11 corn products from different brands were analysed for AfT and AfB₁ using the Enzyme-Linked Immunosorbent Assay (ELISA) method. The results indicate that, of the brands analysed, 35% of the peanut products were contaminated with aflatoxins at various levels (range 5 to 870 μg/kg). Peanut-chilli sauces had the highest percentage of AfT contamination 9/12 (75%), which was followed by traditional snacks 5/11 (45%), peanut butter 4/11 (40%), flour egg coated peanut 6/16 (37%), and peanut cake 3/10 (30%). Fried peanuts and roasted peanut were found to contain aflatoxin at relatively lower percentages of 9% and 8%, respectively. From the 12 analysed baby food samples, on the other hand, no sample was found to be contaminated with aflatoxins. Two of 11 samples (18%) of corn based products were contaminated with AfT, ranging between 5.8 and 12.4 μg/kg. Additionally, 30 selected samples in different concentration ranges were further analysed to verify the correlation between ELISA and HPLC techniques and results were compared.     

Forensic Application of Microbiological Culture Analysis To Identify Mail Intentionally Contaminated with Bacillus anthracis Spores

The discovery of a letter intentionally filled with dried Bacillus anthracis spores in the office of a United States senator prompted the collection and quarantine of all mail in congressional buildings. This mail was subsequently searched for additional intentionally contaminated letters. A microbiological sampling strategy was used to locate heavy contamination within the 642 separate plastic bags containing the mail. Swab sampling identified 20 bags for manual and visual examination. Air sampling within the 20 bags indicated that one bag was orders of magnitude more contaminated than all the others. This bag contained a letter addressed to Senator Patrick Leahy that had been loaded with dried B. anthracis spores. Microbiological sampling of compartmentalized batches of mail proved to be efficient and relatively safe. Efficiency was increased by inoculating culture media in the hot zone rather than transferring swab samples to a laboratory for inoculation. All mail sampling was complete within 4 days with minimal contamination of the sampling environment or personnel. However, physically handling the intentionally contaminated letter proved to be exceptionally hazardous, as did sorting of cross-contaminated mail, which resulted in generation of hazardous aerosol and extensive contamination of protective clothing. Nearly 8 × 10⁶ CFU was removed from the most highly cross-contaminated piece of mail found. Tracking data indicated that this and other heavily contaminated envelopes had been processed through the same mail sorting equipment as, and within 1 s of, two intentionally contaminated letters.     

Uncertainty Quantification in Life Cycle Assessments: Interindividual Variability and Sensitivity Analysis in LCA of Air‐Conditioning Systems

The life cycle environmental profile of energy‐consuming products, such as air conditioning, is dominated by the products’ use phase. Different user behavior patterns can therefore yield large differences in the results of a cradle‐to‐grave assessment. Although this variation and uncertainty is increasingly recognized, it remains often poorly characterized in life cycle assessment (LCA) studies. Today, pervasive sensing presents the opportunity to collect rich data sets and improve profiling of use‐phase parameters, in turn facilitating quantification and reduction of this uncertainty in LCA. This study examined the case of energy use in building cooling systems, focusing on global warming potential (GWP) as the impact category. In Singapore, building cooling systems or air conditioning consumes up to 37% of national electricity demand. Lack of consideration of variation in use‐phase interaction leads to the oversized designs, wasted energy, and therefore reducible GWP. Using a high‐resolution data set derived from sensor observations, energy use and behavior patterns of single‐office occupants were characterized by probabilistic distributions. The interindividual variability and use‐phase variables were propagated in a stochastic model for the life cycle of air‐conditioning systems and simulated by way of Monte Carlo analysis. Analysis of the generated uncertainties identified plausible reductions in global warming impact through modifying user interaction. Designers concerned about the environmental profile of their products or systems need better representation of the underlying variability in use‐phase data to evaluate the impact. This study suggests that data can be reliably provided and incorporated into the life cycle by proliferation of pervasive sensing, which can only continue to benefit future LCA.     

Air conditioning in a tropical climate: Impacts upon European residents in Darwin,Australia

The efficacy of current practices in air conditioning is investigated in the two monsoonal seasons in Darwin. Assessment is made of atmospheric parameters, clothing, metabolic rate. Some 1000 questionnaires are applied dealing with adaptations, health perceptions and preferences as related to air cooling and ventilation. The findings are discussed with reference to energy balance calculations and current models of psychological control in thermoregulation. The results indicate that Darwin's population is considerably overcooled, and contrary to assumptions and practice, air conditioning is not desired in office buildings during the Dry. In the home, air conditioning is not regarded as essential. The indications are that a rationalization of air cooling to comply with natural variability in warmth would lead to a significant reduction in energy consumption, and an overall enhancement to the health and comfort of the population through the greater ventilation rates that would be economically feasible were design temperatures lifted.     

Forensic application of microbiological culture analysis to identify mail intentionally contaminated with Bacillus anthracis spores

The discovery of a letter intentionally filled with dried Bacillus anthracis spores in the office of a United States senator prompted the collection and quarantine of all mail in congressional buildings. This mail was subsequently searched for additional intentionally contaminated letters. A microbiological sampling strategy was used to locate heavy contamination within the 642 separate plastic bags containing the mail. Swab sampling identified 20 bags for manual and visual examination. Air sampling within the 20 bags indicated that one bag was orders of magnitude more contaminated than all the others. This bag contained a letter addressed to Senator Patrick Leahy that had been loaded with dried B. anthracis spores. Microbiological sampling of compartmentalized batches of mail proved to be efficient and relatively safe. Efficiency was increased by inoculating culture media in the hot zone rather than transferring swab samples to a laboratory for inoculation. All mail sampling was complete within 4 days with minimal contamination of the sampling environment or personnel. However, physically handling the intentionally contaminated letter proved to be exceptionally hazardous, as did sorting of cross-contaminated mail, which resulted in generation of hazardous aerosol and extensive contamination of protective clothing. Nearly 8 x 10(6) CFU was removed from the most highly cross-contaminated piece of mail found. Tracking data indicated that this and other heavily contaminated envelopes had been processed through the same mail sorting equipment as, and within 1 s of, two intentionally contaminated letters.     

Green bio-dispersant removal efficacy estimation for controlling biofilms in cooling towers

In this study the effect of green bio-dispersants on cooling water biofilm attachment has been investigated. As monitoring biofilm growth in industrial cooling systems is extremely complex and difficult, we developed a simple method to additively evaluate the effectiveness of green bio-dispersants on biofilms grown in two different lab-scale systems using, as inoculum, make-up water from an industrial cooling tower. The proposed method uses spectrophotometry to estimate the detached biomass after bio-dispersant action. The biofilm detachment rate and the removal percentage of biofilm were calculated after the use of six green bio-dispersants, two non-green dispersants and three controls at two different concentrations. Under all tested conditions, green bio-dispersants showed higher biofilm removal percentage and detachment rate than non-green additives. These results suggest that highly performing green additives could be recommended as a reasonable replacement for non-green additives for controlling biofilms in cooling towers, in accordance to the European Union’s REACH regulation.     

Genotypic variability and persistence of Legionella pneumophila PFGE patterns in 34 cooling towers from two different areas

Genotypic variability and clonal persistence are important concepts in molecular epidemiology as they facilitate the search for the source of sporadic cases or outbreaks of legionellosis. We studied the genotypic variability and persistence of Legionella pulsed-field gel electrophoresis (PFGE) patterns over time (period > 6 months) in 34 positive cooling towers from two different areas. In area A, radius of 70 km, 52 indistinguishable PFGE patterns were differentiated among the 27 cooling towers. In 13 cooling towers we observed ≥ 2 PFGE patterns. Each cooling tower had its own indistinguishable Legionella PFGE pattern which was not shared with any other cooling tower. In area B, radius of 1 km, 10 indistinguishable PFGE patterns were obtained from the seven cooling towers. In four, we observed ≥ 2 PFGE patterns. Three of these 10 indistinguishable PFGE patterns were shared by more than one cooling tower. In 27 of 34 cooling towers the same PFGE pattern was recovered after 6 months to up to 5 years of follow-up. The large genotypic diversity of Legionella observed in the cooling towers aids in the investigation of community outbreaks of Legionnaires' disease. However, shared patterns in small areas may confound the epidemiological investigation. The persistence of some PFGE patterns in cooling towers makes the recovery of the Legionella isolate causing the outbreak possible over time.     

The effect of bacterial contamination on the growth and gas evolution ofIn vitro cultured apricot shoots

Summary Shoots of “San Castrese” and “Portici” apricots (Prunus armeniaca L.) free of cultivable bacteria, shoots of the same origin exhibiting bacterial contamination after repeated subcultures, and contaminated shoots treated with cefotaxime were compared for gas exchange, proliferation rate, and fresh and dry weight. Cultures of San Castrese contaminated byBacillus circulans andSphingomonas paucimobilis, and of Portici contaminated withStaphylococcus hominis andMicrococcus kristinae, including those treated with cefotaxime, showed comparable shoot weights and lower proliferation rates than healthy cultures. Bacteria, even if not visible until the end of subculture, markedly influenced the gaseous composition of the jar headspace. Healthy cultures clearly showed photosynthetic activity at 60 μM·m⁻²·s⁻¹ photosynthetically active radiation; in contrast, oxygen quickly decreased and carbon dioxide increased in contaminated cultures, including those treated with cefotaxime, in which bacteria became visible in the culture medium only after repeated subcultures.     

Study on the aerosol transmission of Hantaan virus

We studied some important aspects constituting aerosol transmission of Hantaan virus, including the possibility of viral aerosol generated by rodents, airborne stability, rodent’s susceptibility to aerosol challenge, and field air sampling for the virus. Our results showed that Hantaan virus aerosol could be generated through the activities of infected mice, and cause specific infection among the exposed animals. Several kinds of rodents such asApodemus agrarius, weaning mice and suckling mice were found to be rather sensitive to the aerosol challenge of Hantaan virus. The 50% of inhaled lethal dose (LD₅₀) of suckling mice is 0.73 (1.4–0.37) plaque-forming unit (pfu). Hantaan virus aerosol was relatively stable in the air at 18–20°C and 70–90% relative humidity. The biological decay rate of the viral aerosol was 4.1% per min during 90 min. We also successfully sampled and isolated Hantaan virus from the working field atmosphere. The data obtained in the study provided more solid evidence for Hantaan virus aerosol transmission among rodents and from rodents to human-beings.     

Feasibility of using real-time optical methods for detecting the presence of viable bacteria aerosols at low concentrations in clean room environments

An experimental investigation was carried out to determine the agreement between two methods of viable bacteria aerosol detection. Various amounts of Bacillus globigii (BG) spores were aerosolized in 1-s bursts into a HEPA-filtered air stream and sampled simultaneously with a fluorescence aerosol particle sensor (FLAPS) and a slit to agar biological air sampler. The slit sampler incorporated 150-mm malt extract culture plates, which were incubated at 37°C for at least 12 h before culturable BG particles were counted in terms of colony-forming units (CFU). A relationship between CFU and optically detected viable bacteria particles was determined as culturable particle concentrations decreased. Through further analytical procedures, the FLAPS showed a limit of detection (LOD) of 4.2 bacterial particle/2.5 l of sampled air or 1.7 × 10³ m⁻³. This real-time bacteria aerosol monitor could be used to detect burst contamination events during a surgical procedure. The technology may be used for developing a dose–response relationship between bacterial particle exposure and infection, a tool potentially helpful in determining patient risk.     

Protocol for sampling environmental sites for legionellae

A protocol for sampling environmental sites was developed and used to identify possible sources of Legionella species in support of epidemiologic investigations at two hospitals. In hospital A, legionellae were isolated from 43 of 106 (40%) different sites. Three separate Legionella pneumophila serotypes and a previously unrecognized species were present in different combinations in the positive samples. Two of five cooling towers contained the same L. pneumophila serogroup 1 monoclonal type (1,2,4,5) as was isolated from patients. The same monoclonal type was also isolated from make-up water for the two cooling towers, a hot water tank, water separators in four main air compressor systems for respiratory therapy, and cold and hot water faucets. In hospital B, 13 of 37 (38%) sample sites contained legionellae, all of which were L. pneumophila serogroup 1. The monoclonal type matching isolates from patients (1,2,4,5) was found at the highest concentration in a hot water tank, but it was also present at four other sample sites. Since legionellae not related to disease may be found in many of the sites sampled, an epidemiologic association with the probable source should be established before intervention methods, such as disinfection, are undertaken.     

Protocol for sampling environmental sites for legionellae.

A protocol for sampling environmental sites was developed and used to identify possible sources of Legionella species in support of epidemiologic investigations at two hospitals. In hospital A, legionellae were isolated from 43 of 106 (40%) different sites. Three separate Legionella pneumophila serotypes and a previously unrecognized species were present in different combinations in the positive samples. Two of five cooling towers contained the same L. pneumophila serogroup 1 monoclonal type (1,2,4,5) as was isolated from patients. The same monoclonal type was also isolated from make-up water for the two cooling towers, a hot water tank, water separators in four main air compressor systems for respiratory therapy, and cold and hot water faucets. In hospital B, 13 of 37 (38%) sample sites contained legionellae, all of which were L. pneumophila serogroup 1. The monoclonal type matching isolates from patients (1,2,4,5) was found at the highest concentration in a hot water tank, but it was also present at four other sample sites. Since legionellae not related to disease may be found in many of the sites sampled, an epidemiologic association with the probable source should be established before intervention methods, such as disinfection, are undertaken.     

Development of a new CARD-FISH protocol for quantification of Legionella pneumophila and its application in two hospital cooling towers

Aims: Open cooling towers are frequent sources of infections with Legionella pneumophila. The gold standard for the detection of Leg. pneumophila is based on cultivation lasting up to 10 days and detecting only culturable cells. Alternative fluorescence in situ hybridization (FISH) protocols have been proposed, but they result in faint fluorescence signals and lack specificity because of cross‐hybridization with other Legionella species. Our aim was thus to develop a new FISH protocol for rapid and specific detection of Leg. pneumophila in water samples. Methods and Results: A novel catalysed reporter deposition FISH (CARD‐FISH) protocol for the detection of Leg. pneumophila was developed, which significantly enhanced signal intensity as well as specificity of the probe through the use of a novel competitor probe. The developed protocol was compared with the culture method for monitoring the seasonal development of culturable and nonculturable Leg. pneumophila in two hospital cooling tower systems. Seasonal fluctuations of Leg. pneumophila concentrations detected via CARD‐FISH were related to the development of the total bacterial community in both cooling towers, with temperature and biocide as the main factors controlling this development. Conclusions: Our results clearly showed that the majority of the Leg. pneumophila cells were in a nonculturable state. Thus, detection of Leg. pneumophila with culture methods may underestimate the total numbers of Leg. pneumophila present. Significance and Impact of the Study: Rapid, sensitive and specific detection and quantification of Leg. pneumophila in water systems is prerequisite for reliable risk estimation. The new protocol significantly improves current methodology and can be used to monitor and screen for Leg. pneumophila concentrations in cooling towers or other water systems.     

Combined effects of experimental heavy-metal contamination (Cu, Zn, and CH3Hg) and starvation on quail’s body condition

Combined effects of heavy-metal contamination (Cu, Zn, and CH₃Hg) and starvation were tested on common quails (Coturnix coturnix japonica) and used as a model for comparison with a wild common guillemot (Uria aalge) population found stranded at the Belgian coast. Appropriate heavy-metal levels were given to the quails to obtain concentrations similar to those found in the seabirds’s tissues. The contaminated animals were then starved for 4 d to simulate the evident malnutrition symptoms observed at the guillemot’s level. In such conditions, food intake and total-body weight are shown to decrease in contaminated individuals with simultaneous significant hepatic and renal increase of the heavy-metal concentrations. Like guillemots, higher heavy-metal levels were observed in those contaminated quails that had also developed a cachectic status characterized by a general atrophy of their pectoral muscle and complete absence of subcutaneous and/or abdominal fat depots. Although likely the result of a general protein catabolism during starvation, it is suggested that these higher metal levels could as well enhance a general muscle wasting process (cachectic status).