Phylogenetic analysis of members of the genus Aeromonas based on gyrB gene sequences.

The phylogenetic relationships of all known species of the genus Aeromonas were investigated by using the sequence of gyrB, a gene that encodes the B-subunit of DNA gyrase. Nucleotide sequences of gyrB were determined from 53 Aeromonas strains, including some new isolates, which were also characterized by analysis of the 16S rDNA variable regions. The results support the recognition of the family Aeromonadaceae, as distinct from Plesiomonas shigelloides and other enteric bacteria. This phylogenetic marker revealed strain groupings that are consistent with the taxonomic organization of all Aeromonas species described to date. In particular, gyrB results agreed with 16S rDNA analysis; moreover, the former showed a higher capacity to differentiate between species. The present analysis was useful for the elucidation of reported discrepancies between different DNA-DNA hybridization sets. Additionally, due to the sequence diversity found at the intraspecies level, gyrB is proposed as a useful target for simultaneous identification of species and strains. In conclusion, the gyrB gene has proved to be an excellent molecular chronometer for phylogenetic studies of the genus Aeromonas.

Detection of Legionella pneumophila in bioaerosols by polymerase chain reaction.

Most studies focusing on detecting microorganisms in air by polymerase chain reaction (PCR) have used a liquid impinger to sample bioaerosols, mainly because a liquid sample is easy to be processed by PCR analysis. Nevertheless, the use of multiple-hole impactors for the analysis of bioaerosols by PCR has not been reported despite its great utility in culture analysis. In this study we have modified the impaction onto an agar surface sampling method to impaction onto a liquid medium using the MAS-100 air sampler (Merck) (single-stage multiple-hole impactor). To evaluate the recovery of airborne microorganisms of both sampling methods, a suspension containing Escherichia coli was artificially aerosolized and bioaerosols were collected onto Tergitol-7 agar and phosphate-buffered saline (PBS) with the MAS-100. A linear regression analysis of the results showed a strong positive correlation between both sampling methods (r = 0.99, slope 0.99, and y intercept 0.07). Afterwards, the method of impingement into a liquid medium was used to study airborne Legionella pneumophila by PCR. A total of 64 samples were taken at a wastewater treatment plant, a chemical plant, and an office building and analyzed by culture and PCR. Results showed that three samples were positive both by PCR and plate culture, and that nine samples negative by plate culture were positive by PCR, proving that L. pneumophila was present in bioaerosols from these three different environments. The results demonstrate the utility of this single-stage multiple-hole impactor for sampling bioaerosols, both by culture and by PCR.

Rapidness and reliability: important factors in the actual and future development of the microbiological control of water

Se presentan metodos para la deteccion de microorganismos indicadores de contaminacion fecal en el agua para consumo. Las metodologias presentadas se basan en analisis moleculares, como la amplificacion del ADN o PCR

Rapidness and reliability: important factors in the actual and future development of the microbiological control of water

Se presentan metodos para la deteccion de nicroorganismos indicadores de contaminacion fecal en el agua para consumo. Las metodologias presentadas se basan en analisis moleculares, como la amplificacion del ADN o PCR

Detection of Legionella pneumophila in wastewater by nested polymerase chain reaction.

The study of Legionella in treated wastewater acquires special importance when this water is used in irrigation by spray, as Legionella is transmitted via the inhalation of aerosols and may consequently represent a health risk. In this study, we applied polymerase chain reaction (PCR) amplification as an alternative method to plate culture for detecting L. pneumophila in twelve heavily biocontaminated samples from a wastewater treatment plant. Moreover, we studied the efficiency of rapid gel filtration methods and filtration through chelating ion exchange resin in the elimination of PCR inhibitors from wastewater samples. When Legionella was investigated by PCR without any previous treatment, no amplification occurred, and when we used chromatographic methods to eliminate PCR inhibitors, nine out of twelve samples became positive. These results indicate the abundant presence of Legionella in wastewater, and although the methods used to eliminate PCR inhibitors are effective in the preparation of clean samples, the possible presence of different metal-organic matter compounds, which are not eliminated, may produce false-negative results.

A simple method for the eradication of Legionella pneumophila from potable water systems.

In this paper we describe a simple method, noncorrosive to pipes, for the eradication of Legionella pneumophila from potable water systems. This method is based on the systematic purging of the pipe networks with cold water containing 1-1.5 mg residual chlorine/L. In the hot water system, a new pipe bypassing the water heater was installed, whereas in the air conditioning system, the circuit is purged with water from the tap water system. The feasibility of this method was studied in two hotels in which the presence of Legionella was detected despite treatment of the water by the hyperchlorination method. The evolution of the presence of Legionella was studied by culture and polymerase chain reaction. Eighty samples from hotel A and sixty-seven samples from hotel B were analyzed during the time that the eradication method was applied. Our results showed that this method permitted the effective elimination of L. pneumophila after 5 months in hotel A and 7 months in hotel B.

Nested polymerase chain reaction for detection of Legionella pneumophila in water.

A highly sensitive nested polymerase chain reaction (PCR) method was evaluated for detection of Legionella pneumophila in water. Two sets of primers homologous to the coding region of the L. pneumophila macrophage infectivity potentiator (mip) gene were used. Even when starting from minute amounts of L. pneumophila DNA, the double PCR products were readily detected by direct visualization in ethidium-bromide-stained agarose gels. The method was tested on 34 potable water samples from a hospital building and compared with standard culture isolation. L. pneumophila was isolated in only twelve samples, whereas, by nested PCR, 19 samples were positive, 12 of them coincidental with culturing. These results indicate that nested PCR permits detection of L. pneumophila in samples where culturing fails, with the advantage of a rapid turnaround time, simplicity and the ability to detect non-culturable cells, fulfilling the requirements of sensitivity and specificity for routine use in an environmental laboratory.

[Comparative study of the quality of water from the supply system and after decalcification: health aspects]. / Estudio comparativo de la calidad del agua de la red y tras descalcificación: aspectos sanitarios.

A study is carried out on drinking water after decalcification. What is notable is that only 51.8% of the decalcification systems studied operated under adequate conditions. The hardness levels found after the softening process are much lower than the limits set by the present Health-Technical Regulations, while sodium concentrations were five times greater than the network's water, exceeding in the majority of cases the limits set by the Council of European Communities Directive. Water obtained after an uncontrolled decalcification may be a risk factor in cardiovascular diseases.