Microbiological quality of bottled water sold in retail outlets in Nigeria

M.T. OGAN. 1992. The microbiological quality of four brands of bottled water sold in retail outlets in Nigeria were assessed by routine methods in 90 samples. Samples of two brands were acidic in the pH range 3.5–5.9. Faecal coliforms and streptococci were not recovered from any sample. Heterotrophic plate counts (HPC) numbered 50–800 cfu/ml in two brands, A and B, and 100–87000 cfu/ml in C and D. Component colony types among the HPC bacteria in brands C and D produced water-soluble, fluorescent pigments on colony count and other agar media, and occurred in 11 of 16 batches: their numbers varied from 60 to 82000 cfu/ml. Presumptive antibiotic-resistant proportions of the HPC bacteria were also recovered from brands C and D on agar amended with ampicillin, penicillin or streptomycin. Five isolates from the green pigmented colonies, representing five batches of brands C and D were satisfactorily identified as strains of Pseudomonas aeruginosa. These strains resisted between four and nine of 14 antibiotics tested. Resistance to tetracycline was eliminated in one isolate when it was cured by incubation at 42C for 100 h, but gel electrophoretic resolution of the DNA did not reveal presence of a plasmid. Strains of Bacillus were the second most commonly isolated bacteria; they were the only colony types in most samples with low HPC counts.     

Microbiological quality of bottled water sold in retail outlets in Nigeria.

The microbiological quality of four brands of bottled water sold in retail outlets in Nigeria were assessed by routine methods in 90 samples. Samples of two brands were acidic in the pH range 3.5-5.9. Faecal coliforms and streptococci were not recovered from any sample. Heterotrophic plate counts (HPC) numbered 50-800 cfu/ml in two brands, A and B, and 100-87,000 cfu/ml in C and D. Component colony types among the HPC bacteria in brands C and D produced water-soluble, fluorescent pigments on colony count and other agar media, and occurred in 11 of 16 batches: their numbers varied from 60 to 82,000 cfu/ml. Presumptive antibiotic-resistant proportions of the HPC bacteria were also recovered from brands C and D on agar amended with ampicillin, penicillin or streptomycin. Five isolates from the green pigmented colonies, representing five batches of brands C and D were satisfactorily identified as strains of Pseudomonas aeruginosa. These strains resisted between four and nine of 14 antibiotics tested. Resistance to tetracycline was eliminated in one isolate when it was cured by incubation at 42 degrees C for 100 h, but gel electrophoretic resolution of the DNA did not reveal presence of a plasmid. Strains of Bacillus were the second most commonly isolated bacteria; they were the only colony types in most samples with low HPC counts.     

Long-term survival of Legionella pneumophila serogroup 1 under low-nutrient conditions and associated morphological changes

Abstract Extended survival of Legionella pneumophila , using both a clinical and an environmental isolate, was studied in drinking water, creek water, and estuarine water microcosms. Legionella populations were monitored by acridine orange direct counts (AODC) and viable count on buffered charcoal yeast extract agar amended with alpha-ketoglutarate (BCYEα). Initial colony counts of the clinical isolate in drinking and creek water microcosms were 2 × 10⁸ cfu/ml and, after incubation for 1.5 years, the plate counts decreased to 3 × 10⁶ cfu/ml. The AODC counts, however, did not change significantly. The clinical isolate in estuarine water decreased in plate counts to 10² (cfu/ml) over the same period. After incubation for 1.5 years at 15°C in the microcosms, Legionella plate counts of creek and drinking water decreased by two logs. Direct microscopic examination of aliquots removed from all microcosms revealed the presence of small bacilli, large bacilli and rare filamentous cells. The environmental isolate demonstrated only one colony morphology upon culture on BCYEα. Interestingly, after four months incubation in the microcosm, upon plating the clinical isolate on BCYEα, two distinct colony types were evident. Examination by immunofluorescent staining employing a monoclonal antibody against L. pneumophila revealed both bacillus and filamentous forms. The total cellular proteins of both morphotypes were examined by sodium dodecyl sulfate polyacrylyamide gel electrophoresis (SDS-PAGE), demonstrating identical protein patterns. Those Legionella cells remaining culturable during 1.5 years of incubation grew rapidly when transferred to BCYEα. Incubation was continued and it was found that some strains of L. pneumophila serogroup 1 can remain viable for longer than 2.4 years under low-nutrient conditions.     

Comparison of two selective media for the detection and enumeration of Lactobacilli in human faeces

The enumeration of faecal bacteria is an important requirement for many studies of bowel health. One approach is the use of selective culture media for the culture and identification of genera or species from faeces. This study compares the culture of Lactobacilli from dilution series of faecal samples from six healthy human volunteers on two commonly used media, LAMVAB and Rogosa agar. Colonies were counted after a 72-h anaerobic incubation at 37 degrees C, and colony morphology recorded by a single observer. DNA was isolated from a representative number of colonies and genus-specific PCR, single-stranded conformation polymorphism (SSCP) and DNA sequencing performed. Total colony counts ranged from <3.00 to 7.48 log(10) cfu/g of faeces for LAMVAB and 5.09 to 7.66 log(10) cfu/g for Rogosa. For each subject, the total colony count was higher on Rogosa than that obtained with LAMVAB agar. SSCP analysis and DNA sequencing indicated that colony morphology was not an accurate predictor of genus identity. Growth of two species, Lactobacillus acidophilus and Lactobacillus gasseri, was not supported on LAMVAB medium. Rogosa agar was more likely to support growth of non-Lactobacillus species. Therefore, neither medium gave a fully accurate representation of the Lactobacilli species present in human faecal samples.     

Improvement of human melanoma colony formation in soft agar using boiled instead of autoclaved agar

The effect of agar sterilized by either boiling or autoclaving on human melanoma colony formation in soft agar was compared using cells from 17 biopsies of metastatic malignant melanoma. The frequency of colony formation was significantly increased for cells grown in boiled agar in 8 samples (47%), unchanged in 8 samples (47%), and decreased in only one sample (6%). There were increases in both cluster and colony formation for the melanomas which had augmented colony formation when grown in boiled agar. There was also qualitative morphological improvement, including rounder, smoother cells and less extracellular debris surrounding the colonies. These data suggest that melanoma colony formation is enhanced when cells are grown in agar which has been sterilized by boiling rather than autoclaving.     

A comparison of two air samplers for recovery of indoor bioaerosols

Summary A preliminary study was performed using two sampling instruments for airborne bacteria and fungi collection. A Reuter Centrifugal Sampler (RCS) and the open-faced type membrane filter sampler (Sartorius MD8) were compared for evaluating their capability of viable particles recovery. 61 series of parallel samples were collected in the air of a microbiological laboratory. Bacteria and fungi per cubic metre of air were enumerated using appropriate culture media and reported in terms of colony forming units (CFU). Performances of the two instruments for fungi were comparable and significantly correlated, particularly when the Rose Bengal Agar (RBA) medium was used (geometric mean: 237 CFU/m³ for RCS and 247 CFU/m³ for MD8; correlation coefficient: 0.78). Bacterial counts from MD8 resulted consistently lower than those obtained from RCS. The observed high variability suggests the existence of selective collection efficiencies which tend to underestimate the actual occurrence of airborne microrganisms.     

Quality control of membrane filters for bacteriological examination of water

A standard protocol for quality control of membrane filters for bacteriological examination of water is presented. It is based on the screening test proposed by Brenner and Rankin (1990) making use of Enterobacter aerogenes ATCC 49701 as a test strain and tryptone soya agar as medium for incubation. The protocol is standardized to achieve adequate statistical power. A diluted culture of the test strain is filtered through samples of 20 filters of a test batch and a reference batch. Filters and colony morphology are screened for visual defects and criteria for acceptance are given. Variation in the colony counts within the batches is checked for randomness, because filters of a batch must be of equal quality and must not cause overdispersion. The protocol requires that the mean count of the reference batch should lie between 50 and 80 cfu. Homogeneity of the counts (i.e. variation no greater than random) between the batches is tested at a 99% confidence level. In this way an actual difference in recovery between filter batches of 13% to 16% can be detected with a probability of 90%. Results of several tests within this laboratory show good reproducibility.     

Anomalies in the enumeration of starved bacteria on culture media containing nalidic acid and tetracycline

Culturable counts of antibiotic resistant, genetically engineeredPseudomonas fluorescens were determined on antibiotic-containing plate count agar during starvation in water. Prior to starvation, colony counts obtained on all media separated into two groups. The mean of the colony counts on plate count agar with or without tetracycline (4.9 × 10⁶ ml⁻¹) was significantly higher than the mean colony counts on plate count agar containing either nalidixic acid or nalidixic acid plus tetraclycline (2.5×10⁶ ml⁻¹). After 20 days of starvation the highest mean colony counts continued to be obtained on plate count agar (7.2 × 10⁶ ml⁻¹) with slightly, but significantly, lower counts obtained on plate count agar containing either nalidixic acid (5.6 × 10⁶ ml⁻¹) or tetraclycline (1.5×10⁶ ml⁻¹). A combination of nalidixic acid and tetracycline in plate count agar, however, dramatically reduced colony counts (8.3 × 10² ml⁻¹) after this starvation period. The addition of catalase to plate count agar containing nalidixic acid and tetracycline negated the effect caused by this combination of antibiotics. When colony counts obtained over the entire 20 day incubation were considered, the addition of MgSO₄ to plate count agar containing nalidixic acid and tetracycline resulted in a significant increase in colony counts. Other combinations of antibiotics, nalidixic acid+carbenicillin, nalidixic acid+kanamycin, streptomycin+tetracycline, streptomycin+carbenicillin, rifampicin+tetracycline, rifampicin+carbenicillin, and rifampicin+kanamycin, did not inhibit colony formation of starved cells. Antibiotic resistant strains ofP. putida andEscherichia coli also displayed sensitivity to the combination of nalidixic acid and tetracycline in plate count agar after starvation.     

广州市空气微生物含量及其变化规律研究

用JWL-IIB新型固体撞击式空气微生物采样仪对广州市18个监测点的空气微生物含量及其月和季节变化势态进行了观察和研究。结果表明：广州市区室外空气微生物年平均含量为2,298cfu／m^3,室内空气微生物年平均含量为1,792cfu／m^3。各功能区的室外空气微生物平均含量（cfu／m^3）大小顺序为：垃圾压缩站（4,573）〉商业步行街（3,835）〉交通枢纽（1,580）〉居民住宅小区（1,413）〉工业厂区（1,197）〉中心公园（1,187）;室内空气微生物含量（cfu／m^3）大小顺序为：交通枢纽（2,511）〉旅游三星酒店（1,699）〉地铁站（1,167）。一年中,市区各监测点室外空气微生物月平均含量变化范围在1,073～4,096cfu／m^3,其中3月份的空气微生物平均含量最高,可达4,096cfu／m^3,而10月份的空气微生物平均含量最低仅为1,073cfu／m^3。四季中,各监测点室外空气细菌和真菌平均含量（cfu／m^3）变化均表现为春季（3,716）〉夏季（2,300）〉冬季（1,816）〉秋季（1,553）。可为广州市区公共场所的卫生防疫和环境治理提供科学依据。     

Effect of incubation temperature, ageing, and bisulfite content of unsupplemented Brucella agar on aerotolerance of Campylobacter jejuni

A mutant strain of Campylobacter jejuni ATCC 29428 was isolated that grows on unsupplemented Brucella agar at O2 levels as high as 21% at 37 degrees C. While measuring the degree of aerotolerance of this mutant on unsupplemented Brucella medium and comparing it with that of the wild type, we found considerable variation among our estimates. As measured by colony counts on unsupplemented Brucella agar incubated at various oxygen levels, the degree of aerotolerance was affected by incubation temperature and the age of the medium. Aerotolerance was consistently higher on plates incubated at 42 degrees C than at 37 degrees C. When the commercial dehydrated Brucella medium that was used to prepare the Brucella agar was stored in a beaker for 2.5 months, the degree of aerotolerance of C. jejuni was decreased. Addition of 0.01% sodium bisulfite reversed this inhibition. Storage of bottles of hydrated Brucella agar for 1.5 months also resulted in a decreased aerotolerance; again, in addition of 0.01% bisulfite reversed the effect. Aerotolerance was greatly decreased when Brucella agar was prepared from all its individual components except 0.01% bisulfite. The results indicate that the bisulfite component of Brucella agar deteriorates during storage of the dehydrated and hydrated media, and that this deterioration can affect measurements of aerotolerance.     

A versatile most probable number system to quantify lacZY-marked pseudomonads present at low cell numbers in the rhizosphere

Quantitation of introduced genetically modified micro-organisms (GMMs) in the rhizosphere is a key issue when their release in the environment is planned. An improved most probable number (MPN) system, using a titration plate for incubation of rhizosphere extracts and two microcomputer programmes made recently available, MPNES (Woomer et al. 1990) and MPN 2.80 (Klee 1993) to generate the MPNs, is described. The MPN system was compared with colony counts to assess colonization of sugarbeet roots by an introduced lac ZY-modified rhizosphere pseudomonad. The MPNs displayed wider confidence intervals compared with drop-plate counts but allowed the quantitation limit to be lowered to 2.30 log₁₀ cfu per root system. The MPN system proved useful to quantify GMMs present at low cell numbers in the rhizosphere of sugarbeet.     

Evaluation of an Automatic Electronic Device for Counting Bacterial Colonies

An automatic colony counter was tested extensively with colonies of two bacterial species, Serratia marcescens and Bacillus subtilis var. niger, grown on agar media. A stable relationship was established between machine counts and counts done visually by technicians. The calibration curve and estimates of the efficiency of the machine are presented and discussed. It is estimated that a 40% reduction in colony counting time is feasible through use of the machine.     

Impaction onto a Glass Slide or Agar versus Impingement into a Liquid for the Collection and Recovery of Airborne Microorganisms

To study impaction versus impingement for the collection and recovery of viable airborne microorganisms, three new bioaerosol samplers have been designed and built. They differ from each other by the medium onto which the bioaerosol particles are collected (glass, agar, and liquid) but have the same inlet and collection geometries and the same sampling flow rate. The bioaerosol concentrations recorded by three different collection techniques have been compared with each other: impaction onto a glass slide, impaction onto an agar medium, and impingement into a liquid. It was found that the particle collection efficiency of agar slide impaction depends on the concentration of agar in the collection medium and on the sampling time, when samples are collected on a nonmoving agar slide. Impingement into a liquid showed anomalous behavior with respect to the sampling flow rate. Optimal sampling conditions in which all three new samplers exhibit the same overall sampling efficiency for nonbiological particles have been established. Inlet and collection efficiencies of about 100% have been achieved for all three devices at a sampling flow rate of 10 liters/min. The new agar slide impactor and the new impinger were then used to study the biological factors affecting the overall sampling efficiency. Laboratory experiments on the total recovery of a typical environmental microorganism, Pseudomonas fluorescens ATCC 13525, showed that both sampling methods, impaction and impingement, provided essentially the same total recovery when relatively nonstressed microorganisms were sampled under optimal sampling conditions. Comparison tests of the newly developed bioaerosol samplers with those commercially available showed that the incorporation of our research findings into the design of the new samplers yields better performance data than data from currently available samplers.     

Inhibitory effects of titanium (III) citrate on enumeration of bacteria from rumen contents.

Titanium citrate (TC) or L-cysteine-sodium sulfide was added as a reducing agent to buffers and agar media used for enumeration of bacteria from rumen contents of high-forage-fed steers. Approximately equal colony counts were found on TC and L-cysteine-sodium sulfide-reduced media with rumen contents taken 8 h postfeeding, when active bacterial growth was occurring. The colony counts on TC medium were only 56% of those with L-cysteine-sodium sulfide medium with rumen contents taken 1 h prefeeding when bacterial growth was minimal. When colonies from L-cysteine-sodium sulfide medium were transferred to TC medium and vice versa, almost all colonies grew. The data indicate that TC can be inhibitory to bacteria upon their initial isolation from natural habitats, particularly when growth rates are low in these habitats.     

Optimal growth temperature for the isolation of Plesiomonas shigelloides, using various selective and differential agars.

The growth characteristics of known strains of Plesiomonas shigelloides were compared with those of Aeromonas species (the major competing species in environmental waters) on plesiomonas differential agar, inositol brilliant green bile salt, and modified salmonella-shigella agar at incubation temperatures of 37, 42, and 44 degrees C. Using local isolates from clinical and environmental sources, optimal growth conditions, as determined by colony counts and the colony characteristics, plesiomonas differential agar proved to be ideal when incubated at 44 degrees C. Contrary to earlier recommendations for 48 h incubation, the colonies could be recognized readily after an incubation of 24 h.     

Collection of airborne micro-organisms on Nuclepore filters, estimation and analysis--CAMNEA method

The total number of airborne micro-organisms collected on Nuclepore filters was determined by acridine orange staining and epifluorescence microscopy. The viable fraction of the total numbers varied significantly when actinomycete and fungal spores from different environments were stored on the filter surface for 1 week, although the microflora composition was not altered. A high correlation between viable and total counts was noted in environments where the airborne flora was dominated by fungal spores, while a low correlation was found for airborne bacteria. Peak values of the total counts registered in some work environments varied between 10(7) and 10(11) micro-organisms/m3. Size analysis showed a dominating fraction of respirable micro-organisms (aerodynamical diameter less than 5 micron). The investigation shows that it is of the utmost importance to combine viable counts with total count enumeration in the study of exposure to micro-organisms in work-related situations.     

THERMODURIC ORGANISMS IN MILK. PART V. A SIMPLIFIED TESTING TECHNIQUE

SUMMARY: Thermoduric colony counts at 30° of laboratory pasteurized milk determined by the roll tube or agar strip methods were lower than those obtained by the standard Petri plate method. The differences in colony count were not of such magnitude, however, as to be likely to result in many errors in grading if a thermoduric bacterial content of greater than 10⁴/ml is accepted as an index of unsatisfactory cleansing of dairy equipment.
The three methods examined were simpler and more economical than the Petri plate technique, but the agar strip method, as described, using the standard loop, was the simplest and gave a sufficiently reliable estimate of the thermoduric colony count for advisory purposes.     

ADVISORY MICROBIOLOGICAL STANDARDS AND TOLERANCES FOR RAW MILK

SUMMARY: To study the value of recent modifications of microbiological tests used for advisory purposes, samples of tuberculin tested milk, taken at weekly intervals over a 5 year period from the Trawscoed Experimental Husbandry Farm and selected at random during a 12 months period from farms in Wales, were examined by a temperature compensated keeping quality test at 22°, colony count on Yeastrel milk agar in 3 days at 30° and coli-aerogenes colony count on violet red bile agar in 20–24 hr at 30°.
The results show that milk produced and handled under hygienic conditions can be expected to have colony counts of less than 2 × 10⁴/ml and coli-aerogenes colony counts less than 10/ml when examined within 18 hr of milking.     

Colonisation of soft lining materials by micro‐organisms

doi:10.1111/j.1741‐2358.2009.00300.x
Colonisation of soft lining materials by micro‐organisms Objective: This study evaluated the in vitro adherence of pathogenic micro‐organisms, Candida albicans, Staphylococcus aureus and Pseudomonas aeruginosa, to soft lining materials and their inhibitory effect on these micro‐organisms. Materials and Methods: To measure adherence, specimens of Molloplast B and Ufi Gel P were inoculated [10⁷ colony‐forming units per millimetre (cfu/ml)] with TSB media containing the micro‐organisms. To determine the number of micro‐organisms in the 10^?2–10^?5 dilutions, 25 μl of the suspension were transferred to plates of selective media. Colony counts of each specimen were quantified (cfu/ml). The surface roughness was measured with a perfilometer to assess the relationship between the adherence of micro‐organisms and surface roughness of each material. For the inhibition test, specimens of materials were placed in agar plates inoculated individually with the micro‐organisms. After 48 h, the inhibition zones around the specimens were measured. Results: None of the materials exhibited inhibition zones. The number of cfu/ml of S. aureus and P. aeruginosa were significantly greater than C. albicans for both materials. The Ufi Gel P exhibited greater adherence of C. albicans than Molloplast B. No correlation was observed between the adherence of micro‐organisms and surface roughness. Conclusion: The surface roughness of the materials is not the only factor governing micro‐organism adherence.     

Efficiency of Various Growth Media in Recovering Oral Bacterial Flora from Human Dental Plaque

MM10 sucrose blood agar (MM10 SB agar), N(2)C agar, Schaedler agar (SH agar), and mitis salivarius agar (MS agar) were tested for their ability to recover human dental plaque flora by a continuous anaerobic procedure and by a conventional anaerobic method. MM10 SB agar yielded higher recovery of bacteria from plaque samples as determined by the enumeration of colony-forming units (CFU). The CFU on N(2)C agar, SH agar, and MS agar were lower than MM10 SB agar when the continuous anaerobic procedure was used. The superior performance of MM10 SB agar was much more apparent when used for the cultivation of dental plaque by the conventional anaerobic method. Under these conditions the counts were consistently higher on MM10 SB agar as compared to the other media tested. However, the differential counts of Streptococcus sanguis and S. mutans from carious plaque samples were in general comparable on all culture media. Deletion of blood from MM10 SB agar did not lower counts. The elimination of dithiothreitol from this medium resulted in a significantly lower recovery of bacteria from the plaque samples when cultured by the conventional anaerobic method. The storage of MM10 SB agar for varying periods of time aerobic conditions did not seem to affect its performance. These findings suggest that MM10 SB agar is an ideal culture medium for the isolation, nonselective enumeration, and differential counts of bacteria present in normal and disease-associated plaques.