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.
Listeria monocytogenes is an important foodborne pathogen with high mortality. L. monocytogenes and five other Listeria species can frequently be found in the same sample. To identify Listeria isolates found in foods to the species level, two multiplex PCRs were designed. The PCR and conventional biochemical methods were compared for the identification of 456 Listeria isolates collected from routine food quality monitoring schemes between June 2004 and February 2006 and for 62 L. monocytogenes isolates from patients between 1999 and 2005. The results showed that the PCR and biochemical methods had 100% agreement in Listeria identification. The distribution of Listeria species from foods was as follows: L. monocytogenes, 50.4%; L. innocua, 33.8%; L. welshimeri, 14.9%; L. seeligeri, 0.7%; L. grayi, 0.2%; and L. ivanovii, 0.0%. Additional analyses were performed to identify the major serotypes (1/2a, 1/2b, 1/2c, and 4b) and the three lineages of L. monocytogenes isolates from foods and patients, with 1/2a (69.6%) and 1/2b (21.7%) dominating the food isolates and 1/2b (54.8%) and 4b (30.7%) dominating the patient isolates. The lineage results showed that isolates of 1/2a and 1/2c belonged to lineage II and that isolates of 1/2b and 4b belonged to lineage I. The multiplex PCRs for Listeria identification that have been established provide an accurate and rapid method for food quality control. This study has provided the basic knowledge of distribution of Listeria species and L. monocytogenes serotypes in Queensland, Australia, which is useful for epidemiological investigations of listeriosis.
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...
Despite having a low occurrence rate, Listeria monocytogenes is one of the most prominent foodborne pathogens in Australia. The organism is responsible for severe outbreaks with high case fatality and substantial economic losses due to food recalls. In this study, we analyze the incidence trends of listeriosis in Australia during 2001-2010, discuss the relevance of food recalls, and investigate the pathogen's role in foodborne outbreaks. A significant epidemiological finding was a consistently high national age-specific rate recorded for individuals aged 60 years and over. Analysis of Australian Listeria outbreak and food recall data suggests deficiencies in food safety programs of food manufacturing businesses implicated in Listeria outbreaks and revealed that ready-to-eat foods are high-risk vehicles for transmitting listeriosis. Highlighted is Australia's highly efficient Listeria management and surveillance systems bolstered by the introduction of Listeria molecular subtyping in 2010 coupled with a nationally standardized questionnaire by the "Australian foodborne disease surveillance network (OzFoodNet)." The detection of clusters and therefore outbreaks was now possible, allowing cases to be linked across multiple jurisdictions and enabling timely public health action. Considering current changes in food production and consumption patterns, continuous monitoring and improvement of surveillance systems will provide ongoing public health benefits and be crucial to future development of food safety policy for Australia.
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