A total of 172,411 patients were seen by participating practitioners during the study period, of whom 678 (0.4%) had ILI. After a nadir ILI rate of 0.2% in week 21, the rate gradually increased to 0.4% in week 31 before increasing more sharply to a peak of 0.9% in week 36. Swabs were collected from 478 (71%) ILI patients, among whom 170 (36%) had positive influenza test results and the remainder were negative. Influenza-positive patients were detected during weeks 26–40, which was defined as the influenza season (Figure). A total of 142 patients were excluded from further analysis because vaccination status was unknown (n = 11), symptom onset date was unknown (n = 33), time between symptom onset and specimen collection was >4 days (n = 43), or the specimen was collected outside the influenza season (n = 82). A significantly higher proportion of influenza-negative patients (13%) than influenza-positive patients (4%) were excluded because >4 days had elapsed between symptom onset and specimen collection (p = 0.001). No significant difference was found by age group for whether study participants had a specimen collected within 4 days after symptom onset (p = 0.10).

Of the remaining 336 patients, 156 (46%) had positive influenza test results. Most (89%) influenza case-patients had pandemic (H1N1) 2009, 6% had unspecified type A influenza, 4% had influenza A (H3N2), and 1% had influenza type B (Figure). After exclusion of the other influenza patients, 139 pandemic (H1N1) 2009 case-patients and 180 controls were included in the study analysis. Most (57%) participants were 20–49 years of age, and case-patients were significantly younger than controls (p = 0.001); no case-patient was >65 years of age (Table 1). No statistically significant difference was found between male and female study participants by case or control status (p = 0.60) or by vaccination status (p = 0.09). The high proportion of case-patients detected in August resulted in a significant difference between case-patients and controls by month of swab collection (p<0.001).

Overall, 59 (18%) study participants were reported as vaccinated with any vaccine, but the proportion was higher among controls (26%) than among case-patients (9%; p<0.001). The proportion of controls, who were mostly older, who had received the trivalent seasonal vaccine was higher than the proportion of controls who had received the monovalent vaccine (Table 1). Similarly, controls who had received both vaccines were all >20 years of age. Only case-patients who were 5–19 and 20–49 years of age were reported as vaccinated. Influenza vaccine type was not specified for 1 case-patient and 1 control, each of whom was reported as vaccinated.

Reflecting the availability of each vaccine, the median period between vaccination and visit to a general practitioner was significantly shorter for those who received seasonal vaccine (114 days) than for those who received monovalent vaccine (223 days; p<0.0001). No significant difference in the time from vaccination to practitioner visit was found between case-patients and controls for seasonal (p = 0.70) or monovalent vaccine (p = 0.95).

In general, point estimates of vaccine effectiveness adjusted for patient age and month of specimen collection differed little from crude estimates (Table 2). A significant protective effect was observed for seasonal vaccine only (adjusted vaccine effectiveness 79%; 95% CI 33%–93%) and seasonal and monovalent vaccines (adjusted vaccine effectiveness 81%; 95% CI 7%–96%). The adjusted vaccine effectiveness for receipt of any (either or both the seasonal and monovalent) vaccine was lower at 67% because of the 47% vaccine effectiveness for monovalent vaccine. The absence of vaccinated case-patients and controls meant vaccine effectiveness could not be estimated for several of the 5 age groups (Table 1); therefore, age was collapsed into 3 variables: children (0–19 years), working-age adults (20–64 years), and elderly persons (>65 years). Estimates of vaccine effectiveness for working adults were 0%–14% higher than the overall adjusted estimates; estimates for children were either undefined because no controls were vaccinated or were without a significant protective effect. Vaccine effectiveness could not be calculated for elderly persons because there were no case-patients in this age group.

Sensitivity analyses to determine the effects of certain assumptions resulted in variations in the adjusted vaccine effectiveness point estimates of 0%–3% and no changes to their relative significance. The effects considered were as follows: assumption that those patients with unspecified influenza type A had pandemic (H1N1) 2009, no exclusion of patients if >4 days had elapsed between symptom onset and specimen collection, and no exclusion of patients if they were identified outside the defined influenza season.

Our results indicate that the 2010 seasonal trivalent influenza vaccine is >80% effective against pandemic (H1N1) 2009 virus, regardless whether given by itself or in addition to monovalent vaccine. Groups in Europe and Canada have estimated the effectiveness of monovalent seasonal influenza vaccine against pandemic (H1N1) 2009 virus to be 72%–100% (13–17). However, the effectiveness of any vaccine (monovalent, seasonal, or both) against pandemic (H1N1) 2009 virus was lower (67%, 95% CI 33%–84%) because effectiveness for monovalent vaccine only was 47% (95% CI –62% to 82%). The lower effectiveness of monovalent influenza vaccine against pandemic (H1N1) 2009 virus compared with seasonal trivalent influenza vaccine is difficult to explain. Both vaccines contain the same quantities (15 µg) of hemagglutinin; and although the monovalent vaccine does not contain adjuvant and was available ≈6 months before the seasonal vaccine, it has been shown to be strongly immunogenic (3,9,10). Immunogenicity does not necessarily correlate directly with vaccine effectiveness, and we cannot exclude waning immunity as an explanation for the lower effectiveness of monovalent vaccine in our study. Waning immunity after receipt of monovalent vaccine has been suggested after an interim study from the United Kingdom for the 2010–11 influenza season (26). The finding could also be a product of the relatively small number of case-patients and controls who received only the monovalent vaccine, given that vaccine effectiveness estimates can change considerably by the inclusion or exclusion of 1–2 vaccinated study participants.

When stratified by age, estimates of vaccine effectiveness for working-age adults were higher and more precise than those for children. We previously demonstrated that the sentinel practitioner surveillance program in Victoria is well suited for estimating vaccine effectiveness among working-age adults, who account for most of the surveillance population (18), and the 2010 results were consistent with this observation. The relatively few participants in the young (childhood) age groups meant the study had insufficient power to produce defined or significant estimates of vaccine effectiveness. At the other end of the age spectrum, 2% of study participants (5 controls and 0 case-patients) in 2010 were >65 years of age compared with an average of 7% in this age group during 2003–07 (18). Although the absence of pandemic (H1N1) 2009 case-patients >65 years of age is not surprising, given that older adults have been shown to have relatively higher levels of cross-reactive antibodies to pandemic (H1N1) 2009 virus (27–29), the reason for the low proportion of controls in this age group remains unclear. Among the several explanations are a true lower rate of ILI in older persons during 2010, a lower rate of visits to practitioners for ILI by persons in this age group (or treatment at other health services such as hospitals), or preferential sampling of younger persons by practitioners (and perhaps awareness that pandemic [H1N1] 2009 was the predominant circulating influenza virus subtype).

In addition to having a sample size large enough to provide vaccine effectiveness estimates by age group and influenza type, several other considerations with regard to design of case–control studies of influenza vaccine effectiveness have been proposed: 1) whether the control group best represents the vaccination coverage of the source population and 2) whether collection and confounding variables have been adjusted for, particularly underlying chronic conditions for which vaccine is recommended and previous influenza vaccination history (30). A 2010 survey of pandemic vaccination suggests that monovalent vaccine coverage in the control group was generally consistent with that in the general population and that use of monovalent vaccine was ≈17% among those from Victoria, compared with 13% among controls (31). No equivalent survey of 2010 seasonal vaccine usage was available for comparison.

Data about concurrent conditions of study participants that would indicate need for influenza vaccination were not collected during the 2010 influenza season; thus, adjustment of the vaccine effectiveness estimates for this potentially confounding variable could not be conducted. Such confounding by indication (or negative confounding), in which persons at higher risk for influenza are more likely to be vaccinated, underestimates effectiveness of influenza vaccine but may be counteracted by healthy vaccinee bias (or positive confounding), which overestimates effectiveness (30,32). The extent to which these biases occur is likely to vary and may explain the positive and negative variation of crude influenza vaccine effectiveness estimates after adjustment for chronic conditions in several similar test-negative case–control studies (33–35). Speculation about the relative effects of these biases on how many received monovalent vaccine is also difficult; vaccination was funded for the entire population of Australia, but at the end of February 2010, only 18% had been vaccinated (31).

Similar methods using test-negative controls to assess seasonal and pandemic vaccine effectiveness against both seasonal and pandemic influenza viruses have been applied in North America and Europe (13,16,17,33–39). Observational studies provide a convenient and timely way to assess influenza vaccine effectiveness without the ethical, practical, and financial stringencies associated with clinical trials for vaccine efficacy, but they also have limitations. Modeling suggests that the test-negative case–control design generally underestimates true vaccine effectiveness under most conditions of test sensitivity, specificity, and the ratio of influenza to noninfluenza attack rates (25), although quantifying the extent of this effect in this study is difficult because the precise sensitivity and specificity of the test are not known. We attempted to limit ascertainment bias by censoring records that indicated specimen collection >4 days after symptom onset and restricting the analysis to case-patients and controls tested within the influenza season only, although sensitivity analyses indicated little effect if these restrictions were relaxed. Of note, these findings apply predominantly to working-age adults receiving medical care in the general practice setting; the study did not include those who did not seek medical care for ILI. Thus, the study measured effectiveness of vaccine against illness severe enough to require a visit to a practitioner; the results cannot necessarily be generalized to other parts of the population, in particular young children and elderly persons. We were also unable to determine whether participants had previously been infected with pandemic (H1N1) 2009 virus, which may result in overestimation of vaccine effectiveness.

In conclusion, we applied a test-negative case–control study design to an established sentinel surveillance system to estimate effectiveness of a trivalent seasonal influenza vaccine, which included an A/California/7/2009 (H1N1)–like virus, the pandemic (H1N1) 2009 influenza virus strain. This strain is also a component of the trivalent influenza vaccine for the 2010–11 Northern Hemisphere influenza season (40). The trivalent vaccine provided significant protection against laboratory-confirmed pandemic (H1N1) 2009 virus infection.