Azetidines Kill Multidrug-Resistant Mycobacterium tuberculosis without Detectable Resistance by Blocking Mycolate Assembly.

Tuberculosis (TB) is the leading cause of global morbidity and mortality resulting from infectious disease, with over 10.6 million new cases and 1.4 million deaths in 2021. This global emergency is exacerbated by the emergence of multidrug-resistant MDR-TB and extensively drug-resistant XDR-TB; therefore, new drugs and new drug targets are urgently required. From a whole cell phenotypic screen, a series of azetidines derivatives termed BGAz, which elicit potent bactericidal activity with MIC99 values <10 µM against drug-sensitive Mycobacterium tuberculosis and MDR-TB, were identified. These compounds demonstrate no detectable drug resistance. The mode of action and target deconvolution studies suggest that these compounds inhibit mycobacterial growth by interfering with cell envelope biogenesis, specifically late-stage mycolic acid biosynthesis. Transcriptomic analysis demonstrates that the BGAz compounds tested display a mode of action distinct from the existing mycobacterial cell wall inhibitors. In addition, the compounds tested exhibit toxicological and PK/PD profiles that pave the way for their development as antitubercular chemotherapies.

The prevalence of tuberculosis infection among foreign-born Canadians: a modelling study.

BACKGROUND: The prevalence of tuberculosis infection is critical to the design of tuberculosis prevention strategies, yet is unknown in Canada. We estimated the prevalence of tuberculosis infection among Canadian residents born abroad. METHODS: We estimated the prevalence of tuberculosis infection by age and year of migration to Canada for people from each of 168 countries by constructing country-specific and calendar year-specific trends for annual risk of infection using a previously developed model. We combined country-specific prevalence estimates with Canadian Census data from 2001, 2006, 2011, 2016 and 2021 to estimate the overall prevalence of tuberculosis infection among foreign-born Canadian residents. RESULTS: The estimated overall prevalence of tuberculosis infection among foreign-born people in Canada was 25% (95% uncertainty interval [UI] 20%-35%) for census year 2001, 24% (95% UI 20%-33%) for 2006, 23% (95% UI 19%-30%) for 2011, 22% (95% UI 19%-28%) for 2016 and 22% (95% UI 19%-27%) for 2021. The prevalence increased with age at migration and incidence of tuberculosis in the country of origin. In 2021, the estimated prevalence of infection among foreign-born residents was lowest in Quebec (19%, 95% UI 16%-24%) and highest in Alberta (24%, 95% UI 21%-28%) and British Columbia (24%, 95% UI 20%-30%). Among all foreign-born Canadian residents with tuberculosis infection in 2021, we estimated that only 1 in 488 (95% UI 185-1039) had become infected within the 2 preceding years. INTERPRETATION: About 1 in 4 foreign-born Canadian residents has tuberculosis infection, but very few were infected within the 2 preceding years (the highest risk period for progression to tuberculosis disease). These data may inform future tuberculosis infection screening policies.

Impact of Reversion of Mycobacterium tuberculosis Immunoreactivity Tests on the Estimated Annual Risk of Tuberculosis Infection.

A key metric in tuberculosis epidemiology is the annual risk of infection (ARI), which is usually derived from tuberculin skin test (TST) and interferon-γ release assay (IGRA) prevalence surveys carried out in children. Derivation of the ARI assumes that immunoreactivity is persistent over time; however, reversion of immunoreactivity has long been documented. We used a deterministic, compartmental model of Mycobacterium tuberculosis (Mtb) infection to explore the impact of reversion on ARI estimation using age-specific reversion probabilities for the TST and IGRA. Using empirical data on TST reversion (22.2%/year for persons aged ≤19 years), the true ARI was 2-5 times higher than that estimated from immunoreactivity studies in children aged 8-12 years. Applying empirical reversion probabilities for the IGRA (9.9%/year for youths aged 12-18 years) showed a 1.5- to 2-fold underestimation. ARIs are increasingly underestimated in older populations, due to the cumulative impact of reversion on population reactivity over time. Declines in annual risk did not largely affect the results. Ignoring reversion leads to a stark underestimation of the true ARI in populations and our interpretation of Mtb transmission intensity. In future surveys, researchers should adjust for the reversion probability and its cumulative effect with increasing age to obtain a more accurate reflection of the burden and dynamics of Mtb infection.

Modest increase of KIF11 expression exposes fragilities in the mitotic spindle, causing chromosomal instability.

Chromosomal instability (CIN), the process of increased chromosomal alterations, compromises genomic integrity and has profound consequences on human health. Yet, our understanding of the molecular and mechanistic basis of CIN initiation remains limited. We developed a high-throughput, single-cell, image-based pipeline employing deep-learning and spot-counting models to detect CIN by automatically counting chromosomes and micronuclei. To identify CIN-initiating conditions, we used CRISPR activation in human diploid cells to upregulate, at physiologically relevant levels, 14 genes that are functionally important in cancer. We found that upregulation of CCND1, FOXA1 and NEK2 resulted in pronounced changes in chromosome counts, and KIF11 upregulation resulted in micronuclei formation. We identified KIF11-dependent fragilities within the mitotic spindle; increased levels of KIF11 caused centrosome fragmentation, higher microtubule stability, lagging chromosomes or mitotic catastrophe. Our findings demonstrate that even modest changes in the average expression of single genes in a karyotypically stable background are sufficient for initiating CIN by exposing fragilities of the mitotic spindle, which can lead to a genomically diverse cell population.

Whole genome sequencing for tuberculosis in Victoria, Australia: A genomic implementation study from 2017 to 2020.

Background: Whole genome sequencing (WGS) is increasingly used by tuberculosis (TB) programs to monitor Mycobacterium tuberculosis (Mtb) transmission. We aimed to characterise the molecular epidemiology of TB and Mtb transmission in the low-incidence setting of Victoria, Australia, and assess the utility of WGS. Methods: WGS was performed on all first Mtb isolates from TB cases from 2017 to 2020. Potential clusters (≤12 single nucleotide polymorphisms [SNPs]) were investigated for epidemiological links. Transmission events in highly-related (≤5 SNPs) clusters were classified as likely or possible, based on the presence or absence of an epidemiological link, respectively. Case characteristics and transmission settings (as defined by case relationship) were summarised. Poisson regression was used to examine associations with secondary case number. Findings: Of 1844 TB cases, 1276 (69.2%) had sequenced isolates, with 182 (14.2%) in 54 highly-related clusters, 2-40 cases in size. Following investigation, 140 cases (11.0% of sequenced) were classified as resulting from likely/possible local-transmission, including 82 (6.4%) for which transmission was likely. Common identified transmission settings were social/religious (26.4%), household (22.9%) and family living in different households (7.1%), but many were uncertain (41.4%). While household transmission featured in many clusters (n = 24), clusters were generally smaller (median = 3 cases) than the fewer that included transmission in social/religious settings (n = 12, median = 7.5 cases). Sputum-smear-positivity was associated with higher secondary case numbers. Interpretation: WGS results suggest Mtb transmission commonly occurs outside the household in our low-incidence setting. Further work is required to optimise the use of WGS in public health management of TB. Funding: The Victorian Tuberculosis Program receives block funding for activities including case management and contact tracing from the Victorian Department of Health. No specific funding for this report was received by manuscript authors or the Victorian Tuberculosis Program, and the funders had no role in the study design, data collection, data analysis, interpretation or report writing.

Modeling the Cost-Effectiveness of Latent Tuberculosis Screening and Treatment Strategies in Recent Migrants to a Low-Incidence Setting.

Many tuberculosis (TB) cases in low-incidence settings are attributed to reactivation of latent TB infection (LTBI) acquired overseas. We assessed the cost-effectiveness of community-based LTBI screening and treatment strategies in recent migrants to a low-incidence setting (Australia). A decision-analytical Markov model was developed that cycled 1 migrant cohort (≥11-year-olds) annually over a lifetime from 2020. Postmigration/onshore and offshore (screening during visa application) strategies were compared with existing policy (chest x-ray during visa application). Outcomes included TB cases averted and discounted cost per quality-adjusted life-year (QALY) gained from a health-sector perspective. Most recent migrants are young adults and cost-effectiveness is limited by their relatively low LTBI prevalence, low TB mortality risks, and high emigration probability. Onshore strategies cost at least $203,188 (Australian) per QALY gained, preventing approximately 2.3%-7.0% of TB cases in the cohort. Offshore strategies (screening costs incurred by migrants) cost at least $13,907 per QALY gained, preventing 5.5%-16.9% of cases. Findings were most sensitive to the LTBI treatment quality-of-life decrement (further to severe adverse events); with a minimal decrement, all strategies caused more ill health than they prevented. Additional LTBI strategies in recent migrants could only marginally contribute to TB elimination and are unlikely to be cost-effective unless screening costs are borne by migrants and potential LTBI treatment quality-of-life decrements are ignored.

The SARS-CoV-2 Nsp3 macrodomain reverses PARP9/DTX3L-dependent ADP-ribosylation induced by interferon signaling.

SARS-CoV-2 nonstructural protein 3 (Nsp3) contains a macrodomain that is essential for coronavirus pathogenesis and is thus an attractive target for drug development. This macrodomain is thought to counteract the host interferon (IFN) response, an important antiviral signalling cascade, via the reversal of protein ADP-ribosylation, a posttranslational modification catalyzed by host poly(ADP-ribose) polymerases (PARPs). However, the main cellular targets of the coronavirus macrodomain that mediate this effect are currently unknown. Here, we use a robust immunofluorescence-based assay to show that activation of the IFN response induces ADP-ribosylation of host proteins and that ectopic expression of the SARS-CoV-2 Nsp3 macrodomain reverses this modification in human cells. We further demonstrate that this assay can be used to screen for on-target and cell-active macrodomain inhibitors. This IFN-induced ADP-ribosylation is dependent on PARP9 and its binding partner DTX3L, but surprisingly the expression of the Nsp3 macrodomain or the deletion of either PARP9 or DTX3L does not impair IFN signaling or the induction of IFN-responsive genes. Our results suggest that PARP9/DTX3L-dependent ADP-ribosylation is a downstream effector of the host IFN response and that the cellular function of the SARS-CoV-2 Nsp3 macrodomain is to hydrolyze this end product of IFN signaling, rather than to suppress the IFN response itself.

Quantifying the rates of late reactivation tuberculosis: a systematic review.

The risk of tuberculosis is greatest soon after infection, but Mycobacterium tuberculosis can remain in the body latently, and individuals can develop disease in the future, sometimes years later. However, there is uncertainty about how often reactivation of latent tuberculosis infection (LTBI) occurs. We searched eight databases (inception to June 25, 2019) to identify studies that quantified tuberculosis reactivation rates occurring more than 2 years after infection (late reactivation), with a focus on identifying untreated study cohorts with defined timing of LTBI acquisition (PROSPERO registered: CRD42017070594). We included 110 studies, divided into four methodological groups. Group 1 included studies that documented late reactivation rates from conversion (n=14) and group 2 documented late reactivation rates in LTBI cohorts from exposure (n=11). Group 3 included 86 studies in LTBI cohorts with an unknown exposure history, and group 4 included seven ecological studies. Since antibiotics have been used to treat tuberculosis, only 11 studies have documented late reactivation rates in infected, untreated cohorts from either conversion (group 1) or exposure (group 2); six of these studies lasted at least 4 years and none lasted longer than 10 years. These studies found that tuberculosis rates declined over time, reaching approximately 200 cases per 100 000 person-years or less by the fifth year, and possibly declining further after 5 years but interpretation was limited by decreasing or unspecified cohort sizes. In cohorts with latent tuberculosis and an unknown exposure history (group 3), tuberculosis rates were generally lower than those seen in groups 1 and 2, and beyond 10 years after screening, rates had declined to less than 100 per 100 000 person-years. Reinfection risks limit interpretation in all studies and the effect of age is unclear. Late reactivation rates are commonly estimated or modelled to prioritise tuberculosis control strategies towards tubuculosis elimination, but significant gaps remain in our understanding that must be acknowledged; the relative importance of late reactivation versus early progression to the global burden of tuberculosis remains unknown.

Self-clearance of Mycobacterium tuberculosis infection: implications for lifetime risk and population at-risk of tuberculosis disease.

Background: it is widely assumed that individuals with Mycobacterium tuberculosis (Mtb) infection remain at lifelong risk of tuberculosis (TB) disease. However, there is substantial evidence that self-clearance of Mtb infection can occur. We infer a curve of self-clearance by time since infection and explore its implications for TB epidemiology. Methods and findings: data for self-clearance were inferred using post-mortem and tuberculin-skin-test reversion studies. A cohort model allowing for self-clearance was fitted in a Bayesian framework before estimating the lifetime risk of TB disease and the population infected with Mtb in India, China and Japan in 2019. We estimated that 24.4% (17.8-32.6%, 95% uncertainty interval (UI)) of individuals self-clear within 10 years of infection, and 73.1% (64.6-81.7%) over a lifetime. The lifetime risk of TB disease was 17.0% (10.9-22.5%), compared to 12.6% (10.1-15.0%) assuming lifelong infection. The population at risk of TB disease in India, China and Japan was 35-80% (95% UI) smaller in the self-clearance scenario. Conclusions: the population with a viable Mtb infection may be markedly smaller than generally assumed, with such individuals at greater risk of TB disease. The ability to identify these individuals could dramatically improve the targeting of preventive programmes and inform TB vaccine development, bringing TB elimination within reach of feasibility.

Perspectives of Vietnamese, Sudanese and South Sudanese immigrants on targeting migrant communities for latent tuberculosis screening and treatment in low-incidence settings: A report on two Victorian community panels.

BACKGROUND: Tuberculosis (TB) elimination strategies in Australia require a focus on groups who are at highest risk of TB infection, such as immigrants from high-burden settings. Understanding attitudes to different strategies for latent TB infection (LTBI) screening and treatment is an important element of justifiable elimination strategies. METHOD: Two community panels were conducted in Melbourne with members of the Vietnamese (n = 11), Sudanese and South Sudanese communities (n = 9). Panellists were provided with expert information about LTBI and different screening and health communication strategies, then deliberated on how best to pursue TB elimination in Australia. FINDINGS: Both panels unanimously preferred LTBI screening to occur pre-migration rather than in Australia. Participants were concerned that post-migration screening would reach fewer migrants, noted that conducting LTBI screening in Australia could stigmatize participants and that poor awareness of LTBI would hamper participation. If targeted screening was to occur in Australia, the Vietnamese panel preferred 'place-based' communication strategies, whereas the Sudanese and South Sudanese panel emphasized that community leaders should lead communication strategies to minimize stigma. Both groups emphasized the importance of maintaining community trust in Australian health service providers, and the need to ensure targeting did not undermine this trust. CONCLUSION: Pre-migration screening was preferred. If post-migration screening is necessary, the potential for stigma should be reduced, benefit and risk profile clearly explained and culturally appropriate communication strategies employed. Cultural attitudes to health providers, personal health management and broader social vulnerabilities of targeted groups need to be considered in the design of screening programs.

Estimating Long-term Tuberculosis Reactivation Rates in Australian Migrants.

BACKGROUND: The risk of progression to tuberculosis (TB) disease is greatest soon after infection, yet disease may occur many years or decades later. However, rates of TB reactivation long after infection remain poorly quantified. Australia has a low incidence of TB and most cases occur among migrants. We explored how TB rates in Australian migrants varied with time from migration, age, and gender. METHODS: We combined TB notifications in census years 2006, 2011, and 2016 with time- and country-specific estimates of latent TB prevalences in migrant cohorts to quantify postmigration reactivation rates. RESULTS: During the census years, 3246 TB cases occurred among an estimated 2 084 000 migrants with latent TB. There were consistent trends in postmigration reactivation rates, which appeared to be dependent on both time from migration and age. Rates were lower in cohorts with increasing time, until at least 20 years from migration, and on this background there also appeared to be increasing rates during youth (15-24 years of age) and in those aged 70 years and above. Within 5 years of migration, annual reactivation rates were approximately 400 per 100 000 (uncertainty interval [UI] 320-480), dropping to 170 (UI 130-220) from 5 to 10 years and 110 (UI 70-160) from 10 to 20 years, then sustaining at 60-70 per 100 000 up to 60 years from migration. Rates varied depending on age at migration. CONCLUSIONS: Postmigration reactivation rates appeared to show dependency on both time from migration and age. This approach to quantifying reactivation risks will enable evaluations of the potential impacts of TB control and elimination strategies.

Perspective: Potential Impact and Therapeutic Implications of Oncogenic PI3K Activation on Chromosomal Instability.

Genetic activation of the class I PI3K pathway is very common in cancer. This mostly results from oncogenic mutations in PIK3CA, the gene encoding the ubiquitously expressed PI3Kα catalytic subunit, or from inactivation of the PTEN tumour suppressor, a lipid phosphatase that opposes class I PI3K signalling. The clinical impact of PI3K inhibitors in solid tumours, aimed at dampening cancer-cell-intrinsic PI3K activity, has thus far been limited. Challenges include poor drug tolerance, incomplete pathway inhibition and pre-existing or inhibitor-induced resistance. The principle of pharmacologically targeting cancer-cell-intrinsic PI3K activity also assumes that all cancer-promoting effects of PI3K activation are reversible, which might not be the case. Emerging evidence suggests that genetic PI3K pathway activation can induce and/or allow cells to tolerate chromosomal instability, which-even if occurring in a low fraction of the cell population-might help to facilitate and/or drive tumour evolution. While it is clear that such genomic events cannot be reverted pharmacologically, a role for PI3K in the regulation of chromosomal instability could be exploited by using PI3K pathway inhibitors to prevent those genomic events from happening and/or reduce the pace at which they are occurring, thereby dampening cancer development or progression. Such an impact might be most effective in tumours with clonal PI3K activation and achievable at lower drug doses than the maximum-tolerated doses of PI3K inhibitors currently used in the clinic.

Estimating the prevalence of latent tuberculosis in a low-incidence setting: Australia.

Migration is a key driver of tuberculosis (TB) in many low-incidence settings, with the majority of TB cases attributed to reactivation of latent TB (LTBI) acquired overseas. A greater understanding of LTBI risk in heterogeneous migrant populations would aid health planning. We aimed to estimate the LTBI prevalence and distribution among locally born and overseas-born Australians.Annual risks of TB infection estimates were applied to population cohorts (by country of birth, year of arrival and age) in Australian census data in 2006, 2011 and 2016.Both the absolute number and proportion of Australian residents with LTBI increased from 4.6% (interquartile range (IQR) 4.2-5.2%) in 2006 to 5.1% (IQR 4.7-5.5%) in 2016, due to the increasing proportion of the population born overseas (23.8% in 2006 to 28.3% in 2016). Of all residents estimated to have LTBI in 2016; 93.2% were overseas born, 21.6% were aged <35 years and 34.4% had migrated to Australia since 2007.The overall prevalence of LTBI in Australia is low. Some residents, particularly migrants from high-incidence settings, may have considerably higher risk of LTBI, and these findings allow for tailored public health interventions to reduce the risk and impact of future TB disease.

Comparing tuberculosis management under public and private healthcare providers: Victoria, Australia, 2002-2015.

BACKGROUND: Private healthcare providers are important to tuberculosis (TB) management globally, although internationally there are reports of suboptimal management and disparities in treatment commencement in the private sector. We compared the management of TB patients receiving private versus public healthcare in Victoria, an industrialised setting with low tuberculosis (TB) incidence. METHODS: Retrospective cohort study: 2002-2015. Private healthcare provision was included as an independent variable in several multivariate logistic and Cox proportional hazard regression models that assessed a range of outcome variables, encompassing treatment commencement delays, management and treatment outcomes. RESULTS: Of 5106 patients, 275 (5.4%) exclusively saw private providers, and 4714 (92.32%) public. Private care was associated with a shorter delay to presentation (HR 1.36, p = 0.065, 95% CI 1.02-2.00). Private patients were less likely to have genotypic testing (OR 0.66, p = 0.009, 95% CI 0.48-0.90), those with pulmonary involvement were less likely to have a sputum smear (OR 0.52, p = 0.011, 95% CI 0.31-0.86) and provided samples were less likely to be positive (OR 0.54, p = 0.070, 95% CI 0.27-1.05). Private patients with extrapulmonary TB were less likely to have a smear sample (OR 0.7, 95% CI 0.48-0.90, p = 0.009) and radiological abnormalities (OR 0.71, p = 0.070, 95% CI 0.27-1.05). Treatment commencement delays from presentation were comparable for cases with pulmonary involvement and extrapulmonary TB, although public extrapulmonary TB patients received radiological examinations slightly earlier than private patients (HR 0.79, p = 0.043, 95% CI 0.63-0.99) and public patients with pulmonary involvement from high burden settings commenced treatment following an abnormal CXR more promptly than their private counterparts (HR 0.41, p = 0.011, 95% CI 0.21-0.81). Private patients were more likely to receive <4 first-line medications (OR 2.17, p = 0.001, 95% CI 1.36-3.46), but treatment outcomes were comparable between sectors. CONCLUSIONS: The differences we identified are likely to reflect differing case-mix as well as clinician practice. Sputum smear status was an important covariable in our analysis; with its addition we found no significant disparity in the health-system delay to treatment commencement between sectors. Our study highlights the importance of TB programs engaging with private providers, enabling comprehensive data collection that is necessary for thorough and true comparison of TB management and optimisation of care.

Risk of Active Tuberculosis in the Five Years Following Infection . . . 15%?

BACKGROUND: It is often stated that the lifetime risk of developing active TB after an index infection is 5% to 10%, one-half of which accrues in the 2 to 5 years following infection. The goal of this study was to determine whether such estimates are consistent with local programmatic data. METHODS: This study included close contacts of individuals with active pulmonary TB notified in the Australian state of Victoria from January 1, 2005, to December 31, 2013, who we deemed to have been infected as a result of their exposure. Survival analysis was first performed on the assumption of complete follow-up through to the end of the study period. The analysis was then repeated with imputation of censorship for migration, death, and preventive treatment, using local mortality and migration data combined with programmatic data on the administration of preventive therapy. RESULTS: Of 613 infected close contacts, 67 (10.9%) developed active TB during the study period. Assuming complete follow-up, the 1,650-day cumulative hazard was 11.5% (95% CI, 8.9-14.1). With imputation of censorship for death, migration, and preventive therapy, the median 1,650-day cumulative hazard over 10,000 simulations was 14.5% (95% CI, 11.1-17.9). Most risk accrued in the first 5 months after infection, and risk was greatest in the group aged < 5 years, reaching 56.0% with imputation, but it was also elevated in older children (27.6% in the group aged 5-14 years). CONCLUSIONS: The risk of active TB following infection is several-fold higher than traditionally accepted estimates, and it is particularly high immediately following infection and in children.

Health risk assessment for cyanobacterial toxins in seafood.

Cyanobacteria (blue-green algae) are abundant in fresh, brackish and marine waters worldwide. When toxins produced by cyanobacteria are present in the aquatic environment, seafood harvested from these waters may present a health hazard to consumers. Toxicity hazards from seafood have been internationally recognised when the source is from marine algae (dinoflagellates and diatoms), but to date few risk assessments for cyanobacterial toxins in seafood have been presented. This paper estimates risk from seafood contaminated by cyanobacterial toxins, and provides guidelines for safe human consumption.

Reported waterborne outbreaks of gastrointestinal disease in Australia are predominantly associated with recreational exposure.

OBJECTIVE: To examine the frequency and circumstances of reported waterborne outbreaks of gastroenteritis in Australia. METHOD: Examination of data reported to OzFoodNet between 2001 and 2007. RESULTS: During these seven years, 6,515 gastroenteritis outbreaks were reported to OzFoodNet, most of which were classified as being transmitted person-to-person or from an unknown source. Fifty-four (0.83%) outbreaks were classified as either 'waterborne' or 'suspected waterborne', of which 78% (42/54) were attributed to recreational water and 19% (10/54) to drinking water. Of the drinking water outbreaks, implicated pathogens were found on all but one occasion and included Salmonella sp. (five outbreaks), Campylobacter jejuni (three outbreaks) and Giardia (one outbreak). CONCLUSIONS: There have been few waterborne outbreaks detected in Australia, and most of those reported have been associated with recreational exposure. However, there are difficulties in identifying and categorising gastroenteritis outbreaks, as well as in obtaining microbiological and epidemiological evidence, which can result in misclassification or underestimation of water-associated events. IMPLICATIONS: Gastroenteritis surveillance data show that, among reported water-associated gastroenteritis outbreaks in Australia, recreational exposure is currently more common than a drinking water source. However, ongoing surveillance for waterborne outbreaks is important, especially as drought conditions may necessitate replacement of conventional drinking water supplies with alternative water sources, which could incur potential for new health risks.

Sporadic gastroenteritis and recreational swimming in a longitudinal community cohort study in Melbourne, Australia.

The relation between sporadic gastroenteritis and recreational swimming was examined in a cohort of 2,811 people in Melbourne, Australia, over a 15-month period (September 1997-February 1999). Data from a prospective community-based study of gastroenteritis were used for a Poisson analysis of temporality between reported swimming (in public or private pools/spas and in marine or freshwater settings) and a highly credible gastroenteritis (HCG) event. Overall, HCG events were more likely in participants who had swum in a public pool/spa (incidence rate ratio (IRR) = 1.25, 95% confidence interval (CI): 1.10, 1.42; P = 0.001) or river/lake/dam (IRR = 1.77, 95% CI: 1.13, 2.79; P = 0.014) during the previous week or had swum in a public pool/spa (IRR = 1.29, 95% CI: 1.13, 1.46; P < 0.001) during the previous 2 weeks. Subanalysis by age showed that HCG episodes were also more likely in adults who had swum in a private pool/spa (IRR = 1.56, 95% CI: 1.02, 2.39; P = 0.042) during the previous week or swum at an ocean/beach (IRR = 1.78, 95% CI: 1.12, 2.81; P = 0.014) during the previous 2 weeks, demonstrating significant associations between all swimming locations and gastrointestinal symptoms. This study showed that although the incremental risk of recreational swimming is significant, it is relatively small.