We evaluated the efficiency of five membrane filters for recovery of Cryptosporidium parvum oocysts and Giardia lamblia cysts. These filters included the Pall Life Sciences Envirochek (EC) standard filtration and Envirochek high-volume (EC-HV) membrane filters, the Millipore flatbed membrane filter, the Sartorius flatbed membrane filter (SMF), and the Filta-Max (FM) depth filter. Distilled and surface water samples were spiked with 10 oocysts and 10 cysts/liter. We also evaluated the recovery efficiency of the EC and EC-HV filters after a 5-s backwash postfiltration. The backwashing was not applied to the other filtration methods because of the design of the filters. Oocysts and cysts were visualized by using a fluorescent monoclonal antibody staining technique. For distilled water, the highest percent recovery for both the oocysts and cysts was obtained with the FM depth filter. However, when a 5-s backwash was applied, the EC-HV membrane filter (EC-HV-R) was superior to other filters for recovery of both oocysts (n ‫؍‬ 53 ؎ 15.4 per 10 liters) and cysts (n ‫؍‬ 59 ؎ 11.5 per 10 liters). This was followed by results of the FM depth filter (oocysts, 28.2 ؎ 8, P ‫؍‬ 0.015; cysts, 49.8 ؎ 12.2, P ‫؍‬ 0.4260), and SMF (oocysts, 16.2 ؎ 2.8, P ‫؍‬ 0.0079; cysts, 35.2 ؎ 3, P ‫؍‬ 0.0079). Similar results were obtained with surface water samples. Giardia cysts were recovered at higher rates than were Cryptosporidium oocysts with all five filters, regardless of backwashing. Although the time differences for completion of filtration process were not significantly different among the procedures, the EC-HV filtration with 5-s backwash was less labor demanding.

In epidemiological investigations of community legionellosis outbreaks, knowledge of the prevalence, distribution, and clinical significance (virulence) of environmental Legionella isolates is crucial for interpretation of the molecular subtyping results. To obtain such information for Legionella pneumophila serogroup 1 isolates, we used the standardized amplified fragment length polymorphism (AFLP) protocol of the European Working Group on Legionella Infection to subtype L. pneumophila SG1 isolates obtained from patients and water sources in Queensland, Australia. An AFLP genotype, termed AF1, was predominant in isolates from both patients (40.5%) and water (49.0%). The second most common AFLP genotype found in water isolates was AF16 (36.5%), but this genotype was not identified in the patient isolates. When virulence gene-based PCR assays for lvh and rtxA genes were applied to the isolates from patients and water, nearly all (65 of 66) AF1 strains had both virulence genes, lvh and rtxA. In contrast, neither the lvh nor the rtxA gene was found in the AF16 strains, except for one isolate with the rtxA gene. It appears that this may explain the failure to find this genotype in the isolates from patients even though it may be common in the environment. In view of the evidence that the AF1 genotype is the most common genotype among strains found in patients and water sources in this region, any suggested epidemiological link derived from comparing the AF1 genotype from patient isolates with the AF1 genotype from environmental isolates must be interpreted and acted on with caution. The use of virulence gene-based PCR assays applied to environmental samples may be helpful in determining the infection potential of the isolates involved.

The distribution of 19 major virulence genes and the presence of plasmids were surveyed in 141 Legionella pneumophila serogroup (SG) 1 isolates from patients and water in Queensland, Australia. The results showed that 16 of the virulence genes examined were present in all isolates, suggesting that they are life-essential genes for isolates in the environment and host cells. The 65 kb pathogenicity island identified originally in strain Philadelphia-1 T was detected more frequently in isolates from water (44?2 %) than in those from patients (2?7 %), indicating that the 65 kb DNA fragment may aid the survival of L. pneumophila in the sampled environment. However, the low frequency of the 65 kb fragment in isolates from patients suggests that the pathogenicity island may not be necessary for L. pneumophila to cause disease. Plasmids were not detected in the L. pneumophila SG1 isolates from patients or water studied. There was an association of both lvh and rtxA with the virulent and predominant genotype detected by amplified fragment length polymorphism, termed AF1, whereas the avirulent common isolate from water termed AF16 did not have lvh or rtxA genes, with the exception of one isolate with rtxA. It was found that a PCR detection test strategy with lvh and rtxA as pathogenesis markers would be useful for determining the infection potential of an isolate. INTRODUCTIONLegionellae, the aetiologic agents of legionellosis, are ubiquitous worldwide in rivers and lakes (Fliermans et al., 1981), as well as in man-made water systems such as cooling towers and spas (Fields et al., 2002). The vast majority of cases of legionellosis are caused by Legionella pneumophila, mostly serogroup (SG) 1 strains (Yu et al., 2002). To understand the pathogenicity of L. pneumophila, a number of virulence genes of L. pneumophila have been wellcharacterized and extensively reviewed (Cianciotto, 2001;Dowling et al., 1992). The virulence factors characterized include genes required for the whole infection process, such as bacterial cell attachment to host cells, survival and intracellular replication and cell-to-cell spread. The products of genes involved in the initial attachment to host cells and early stages of intracellular infection include type IV pili, the 60 kDa heat-shock protein Hsp60, the poreformation protein RtxA, the macrophage infectivity potentiator Mip and the macrophage-specific infectivity protein MilA (Cianciotto & Fields, 1992;Cirillo et al., 2001;Garduń o et al., 1998;Harb & Abu Kwaik, 2000). The genes required for bacterial survival and intracellular replication are a group of genes called icm (intracellular multiplication) or dot (defect in organelle trafficking) (Vogel et al., 1998). They form a type IV secretion system to deliver effectors to host cells to control organelle trafficking . A number of effectors have been characterized, including RalF (Nagai et al., 2002), LidA (Conover et al., 2003) and LepA/B (Chen et al., 2004) and other recently identified effectors (Shohdy et al., 2005).In addition, the typ...