Population Immunity and Polio Eradication.

The Global Polio Eradication Initiative made immense progress after its establishment in 1988 as a consequence of high coverage with various poliovirus vaccines in all populations of the world. Problems have arisen in recent years, however, related to security issues in some countries, to the circulation of vaccine-derived polioviruses, and to the recognition that individuals with certain immune deficiencies can remain infected and infectious for many months or years. As natural infection and different vaccines have different effects on the immune system, the patterns of humoral and mucosal immunity to polioviruses in the world today are complex but are crucial to the ultimate success of the eradication initiative. This paper describes the background of the current situation and current immunological patterns and discusses their implications for managing population immunity to polioviruses in the years ahead.

Contribution of remote M.tuberculosis infection to tuberculosis disease: A 30-year population study.

BACKGROUND: The importance of remote infection with M.tuberculosis as a cause of tuberculosis disease (TB) is unclear, with limited evidence of impact on TB rates beyond 10 years. Our objective was to assess rates of tuberculosis over 30 years by M.tuberculosis infection status at baseline in Karonga District, Northern Malawi. MATERIALS AND METHODS: Population-based surveys of tuberculin skin testing (TST) from the 1980s were linked with follow-up and TB surveillance in Karonga district. We compared rates of microbiologically-confirmed TB by baseline TST induration <5mm (no evidence of M.tuberculosis infection) and those with baseline TST >17mm (evidence of M.tuberculosis infection), using hazard ratios by time since baseline and attributable risk percent. The attributable risk percent was calculated to estimate the proportion of TB in those infected that can be attributed to that prior infection. We analysed whole genome sequences of M.tuberculosis strains to identify recent transmission. RESULTS: Over 412,959 person-years, 208 incident TB episodes were recorded. Compared to the small induration group, rates of TB were much higher in the first two years in the large induration group, and remained higher to 20 years: age, sex and area-adjusted hazard ratios (HR) 2-9 years post-TST 4.27 (95%CI 2.56-7.11); 10-19 years after TST 2.15 (1.10-4.21); ≥20 years post-TST 1.88 (0.76-4.65). The attributable risk percent of remote infection was 76.6% (60.9-85.9) 2-9 years post-TST, and 53.5% (9.1-76.2) 10-19 years post-TST. Individuals with large TST indurations had higher rates of unique-strain TB (HR adjusted for age, sex and area = HR 6.56 (95% CI 1.96-22.99)), suggesting disease following remote infection, but not of linked-strain TB (recent transmission). CONCLUSIONS: M.tuberculosis infection can increase the risk of TB far beyond 10 years, accounting for a substantial proportion of TB occurring among those remotely infected.

Genome-wide association study of leprosy in Malawi and Mali.

Leprosy is a chronic infection of the skin and peripheral nerves caused by Mycobacterium leprae. Despite recent improvements in disease control, leprosy remains an important cause of infectious disability globally. Large-scale genetic association studies in Chinese, Vietnamese and Indian populations have identified over 30 susceptibility loci for leprosy. There is a significant burden of leprosy in Africa, however it is uncertain whether the findings of published genetic association studies are generalizable to African populations. To address this, we conducted a genome-wide association study (GWAS) of leprosy in Malawian (327 cases, 436 controls) and Malian (247 cases, 368 controls) individuals. In that analysis, we replicated four risk loci previously reported in China, Vietnam and India; MHC Class I and II, LACC1 and SLC29A3. We further identified a novel leprosy susceptibility locus at 10q24 (rs2015583; combined p = 8.81 × 10-9; OR = 0.51 [95% CI 0.40 - 0.64]). Using publicly-available data we characterise regulatory activity at this locus, identifying ACTR1A as a candidate mediator of leprosy risk. This locus shows evidence of recent positive selection and demonstrates pleiotropy with established risk loci for inflammatory bowel disease and childhood-onset asthma. A shared genetic architecture for leprosy and inflammatory bowel disease has been previously described. We expand on this, strengthening the hypothesis that selection pressure driven by leprosy has shaped the evolution of autoimmune and atopic disease in modern populations. More broadly, our data highlights the importance of defining the genetic architecture of disease across genetically diverse populations, and that disease insights derived from GWAS in one population may not translate to all affected populations.

BCG re-vaccination in Malawi: 30-year follow-up of a large, randomised, double-blind, placebo-controlled trial.

BACKGROUND: A large, double-blind, randomised, placebo-controlled trial of repeat BCG found 49% efficacy against leprosy but no protection against tuberculosis after 6-9 years' follow-up in 1995. We report here additional follow-up, which resulted in greatly increased tuberculosis case numbers, and allowed subgroup analysis. METHODS: Nearly 47 000 individuals of all ages living in northern Malawi with a BCG vaccine scar were randomly assigned (1:1) between 1986 and 1989 to receive a second BCG or placebo. The investigators and project staff remained masked to all interventions. Enhanced passive surveillance ensured ascertainment of tuberculosis and leprosy to the end of 2018. Tuberculosis case definitions included rigorous microbiological or histological confirmation. Prespecified subgroup analyses were by tuberculosis type, age at vaccination, time since vaccination, previous tuberculin reactivity, HIV status and Mycobacterium tuberculosis lineage. The original trial is registered with ISRCTN registry, ISRCTN11311670. FINDINGS: In follow-up until Dec 31, 2018, 824 participants had developed tuberculosis, including 786 with pulmonary disease, of whom 383 (63%) of 607 with known HIV status were HIV positive. There was no effect of a second BCG overall (odds ratio [OR] 0·92; 95% CI 0·80-1·05), or for pulmonary (0·93; 0·81-1·07), or lymph node tuberculosis (0·60; 0·31-1·17). The OR was lower for those with known HIV-negative tuberculosis (0·77; 0·59-1·00), for those vaccinated as children (aged <5 years, 0·74; 0·41-1·35; aged 5-14 years, 0·77; 0·60-0·99), and for cases arising at least 20 years after vaccination (0·79; 0·63-1·01). There were no differences by tuberculin status at vaccination, or lineage. There was no evidence of protection against leprosy beyond 10 years after vaccination (although there have been only nine diagnostically certain cases since 1995). INTERPRETATION: There was no evidence that repeat BCG vaccination provides appreciable protection against overall tuberculosis in this rural African population with a high prevalence of HIV. Subgroup effects should not be overinterpreted given the multiple analyses done. However, the evidence for modest protection against HIV-negative tuberculosis, and for a delayed benefit in those vaccinated as children, is consistent with other observations in the literature. FUNDING: LEPRA, Wellcome Trust, Bill & Melinda Gates Foundation.

The effect of BCG revaccination on all-cause mortality beyond infancy: 30-year follow-up of a population-based, double-blind, randomised placebo-controlled trial in Malawi.

BACKGROUND: Trials of BCG vaccination to prevent or reduce severity of COVID-19 are taking place in adults, some of whom have been previously vaccinated, but evidence of the beneficial, non-specific effects of BCG come largely from data on mortality in infants and young children, and from in-vitro and animal studies, after a first BCG vaccination. We assess all-cause mortality following a large BCG revaccination trial in Malawi. METHODS: The Karonga Prevention trial was a population-based, double-blind, randomised controlled in Karonga District, northern Malawi, that enrolled participants between January, 1986, and November, 1989. The trial compared BCG (Glaxo-strain) revaccination versus placebo to prevent tuberculosis and leprosy. 46 889 individuals aged 3 months to 75 years were randomly assigned to receive BCG revaccination (n=23 528) or placebo (n=23 361). Here we report mortality since vaccination as recorded during active follow-up in northern areas of the district in 1991-94, and in a demographic surveillance follow-up in the southern area in 2002-18. 7389 individuals who received BCG (n=3746) or placebo (n=3643) lived in the northern follow-up areas, and 5616 individuals who received BCG (n=2798) or placebo (n=2818) lived in the southern area. Year of death or leaving the area were recorded for those not found. We used survival analysis to estimate all-cause mortality. FINDINGS: Follow-up information was available for 3709 (99·0%) BCG recipients and 3612 (99·1%) placebo recipients in the northern areas, and 2449 (87·5%) BCG recipients and 2413 (85·6%) placebo recipients in the southern area. There was no difference in mortality between the BCG and placebo groups in either area, overall or by age group or sex. In the northern area, there were 129 deaths per 19 694 person-years at risk in the BCG group (6·6 deaths per 1000 person-years at risk [95% CI 5·5-7·8]) versus 133 deaths per 19 111 person-years at risk in the placebo group (7·0 deaths per 1000 person-years at risk [95% CI 5·9-8·2]; HR 0·94 [95% CI 0·74-1·20]; p=0·62). In the southern area, there were 241 deaths per 38 399 person-years at risk in the BCG group (6·3 deaths per 1000 person-years at risk [95% CI 5·5-7·1]) versus 230 deaths per 38 676 person-years at risk in the placebo group (5·9 deaths per 1000 person-years at risk [95% CI 5·2-6·8]; HR 1·06 [95% CI 0·88-1·27]; p=0·54). INTERPRETATION: We found little evidence of any beneficial effect of BCG revaccination on all-cause mortality. The high proportion of deaths attributable to non-infectious causes beyond infancy, and the long time interval since BCG for most deaths, might obscure any benefits. FUNDING: British Leprosy Relief Association (LEPRA); Wellcome Trust.

Autochthonous leprosy in Spain: Has the transmission of Mycobacterium leprae stopped?

BACKGROUND: The aim of this study is to explore whether transmission of M. leprae has ceased in Spain, based upon the patterns and trends of notified cases. METHODOLOGY: Data on new cases reported to the National Leprosy Registry between the years 2003-2018 were extracted. In absence of detailed travel history, cases were considered "autochthonous" or "imported" based on whether they were born within or outside of Spain. These data were analyzed by age, sex, clinical type, country of origin, and location of residence at time of notification. PRINCIPAL FINDINGS: Data were available on 61 autochthonous and 199 imported cases since 2003. There were clear declines in incidence in both groups, and more imported than autochthonous cases every year since 2006. Autochthonous cases were more frequently multibacillary and had older age at diagnosis compared to imported cases. All the autochthonous cases had been born before 1985 and were more than 25 years old at diagnosis. Male-to-female ratio increased with time for autochthonous cases (except for the last time period). The imported cases originated from 25 countries, half of them from Brasil and Paraguay. Autochthonous cases were mainly distributed in the traditionally endemic regions, especially Andalucía and the eastern Mediterranean coast. CONCLUSIONS: Autochthonous and imported cases have different epidemiologic patterns in Spain. There was a clear decline in incidence rates of autochthonous disease, and patterns consistent with those reported from other regions where transmission has ceased. Autochthonous transmission of M. leprae is likely to have now effectively stopped in Spain.

The duration of protection of school-aged BCG vaccination in England: a population-based case-control study.

Background: Evidence of protection from childhood Bacillus Calmette-Guerin (BCG) against tuberculosis (TB) in adulthood, when most transmission occurs, is important for TB control and resource allocation. Methods: We conducted a population-based case-control study of protection by BCG given to children aged 12-13 years against tuberculosis occurring 10-29 years later. We recruited UK-born White subjects with tuberculosis and randomly sampled White community controls. Hazard ratios and 95% confidence intervals (CIs) were estimated using case-cohort Cox regression, adjusting for potential confounding factors, including socio-economic status, smoking, drug use, prison and homelessness. Vaccine effectiveness (VE = 1 - hazard ratio) was assessed at successive intervals more than 10 years following vaccination. Results: We obtained 677 cases and 1170 controls after a 65% response rate in both groups. Confounding by deprivation, education and lifestyle factors was slight 10-20 years after vaccination, and more evident after 20 years. VE 10-15 years after vaccination was 51% (95% CI 21, 69%) and 57% (CI 33, 72%) at 15-20 years. Subsequently, BCG protection appeared to wane; 20-25 years VE = 25% (CI -14%, 51%) and 25-29 years VE = 1% (CI -84%, 47%). Based on multiple imputation of missing data (in 17% subjects), VE estimated in the same intervals after vaccination were similar [56% (CI 33, 72%), 57% (CI 36, 71%), 25% (-10, 48%), 21% (-39, 55%)]. Conclusions: School-aged BCG vaccination offered moderate protection against tuberculosis for at least 20 years, which is longer than previously thought. This has implications for assessing the cost-effectiveness of BCG vaccination and when evaluating new TB vaccines.

Impact of foot-and-mouth disease on milk production on a large-scale dairy farm in Kenya.

The economic impact of foot-and-mouth disease (FMD) has been poorly characterised particularly in endemic settings where such knowledge is important for decision-making on disease control with limited resources. In order to address this, a study was designed using individual animal data from a large-scale dairy farm in Kenya to estimate the impact of an FMD outbreak due to serotype SAT2 virus on milk yield. Daily milk yields from 218 mainly European-breed cattle that were lactating during the 29-day outbreak period were considered in the analysis. At the herd level, the average daily yields decreased from around 20 to 13kg per cow, recovering approximately 2 months after the commencement of the outbreak. Generalised estimating equations (GEE) and an autoregressive correlation matrix were used to compare yields of reported clinical FMD cases and non-cases. No difference was found between reported clinical and non-clinical cases suggesting inaccurate case recording, poor sensitivity of the case definition and subclinical infections being present. To further investigate the impact of FMD, yields were predicted for each individual animal based on historic data from the same herd using a similar GEE approach. For cattle lactating during the outbreak, comparisons were made between actual and predicted yields from the commencement of the outbreak to 305 days lactation using a linear regression model. Animals produced significantly less than predicted if in parity 2 or greater and between 0 and 50 days in milk (DIM) at the start of the outbreak period. The maximum effect was seen among animals in parity ≥4 and between 0 and 50 DIM at the start of the outbreak, producing on average 688.7kg (95%CI 395.5, 981.8) less milk than predicted for their remaining lactation, representing an average 15% reduction in the 305 day production for these animals. Generalisation of the results requires caution as the majority of Kenyan milk is produced in smallholder farms. However, such farms use similar genetics and feeding practices to the study farm, and such systems are increasingly important in the supply of milk globally. These results make an important and unique contribution to the evidence base on FMD impact among dairy cattle in an endemic setting.

Impact of foot-and-mouth disease on mastitis and culling on a large-scale dairy farm in Kenya.

Foot and mouth disease (FMD) is a highly transmissible viral infection of cloven hooved animals associated with severe economic losses when introduced into FMD-free countries. Information on the impact of the disease in FMDV-endemic countries is poorly characterised yet essential for the prioritisation of scarce resources for disease control programmes. A FMD (virus serotype SAT2) outbreak on a large-scale dairy farm in Nakuru County, Kenya provided an opportunity to evaluate the impact of FMD on clinical mastitis and culling rate. A cohort approach followed animals over a 12-month period after the commencement of the outbreak. For culling, all animals were included; for mastitis, those over 18 months of age. FMD was recorded in 400/644 cattle over a 29-day period. During the follow-up period 76 animals were culled or died whilst in the over 18 month old cohort 63 developed clinical mastitis. Hazard ratios (HR) were generated using Cox regression accounting for non-proportional hazards by inclusion of time-varying effects. Univariable analysis showed FMD cases were culled sooner but there was no effect on clinical mastitis. After adjusting for possible confounders and inclusion of time-varying effects there was weak evidence to support an effect of FMD on culling (HR = 1.7, 95% confidence intervals [CI] 0.88-3.1, P = 0.12). For mastitis, there was stronger evidence of an increased rate in the first month after the onset of the outbreak (HR = 2.9, 95%CI 0.97-8.9, P = 0.057).

The decline of leprosy in the Republic of Korea; patterns and trends 1977-2013.

BACKGROUND: Though the World Health Organization declared the 'elimination of leprosy as public health problem' in 2000, the disease remains endemic in many countries. Current trends in incidence of infection and disease are unclear. METHODS: Data on leprosy prevalence between 1977-2013 and data on new leprosy cases detected in the Republic of Korea between 1989-2013 were analysed by age, sex, clinical types, mode of detection, family history, disability grading and geographical distribution. RESULTS: Both prevalence and incidence have declined greatly. There has been a shift to an increased proportion of multibacillary disease, and older age groups, consistent with a dramatic decrease in infection transmission in recent decades. An increase in proportion of cases with family history of disease is consistent with these declines. There is evidence that declines in infection and disease have been greater in the north of the country, as revealed in patterns by place of birth over time. Cases in immigrants now form a substantial proportion of leprosy disease in the Republic of Korea. CONCLUSIONS: Leprosy has declined dramatically in the Republic of Korea in recent decades, and transmission of M. leprae may have effectively stopped. There remains a burden of care for individuals whose disease developed in the past, and there may be some additional newly detected cases among immigrants and among older individuals who acquired autochthonous infections decades ago.

Protection by BCG vaccine against tuberculosis: a systematic review of randomized controlled trials.

BACKGROUND: Randomized trials assessing BCG vaccine protection against tuberculosis have widely varying results, for reasons that are not well understood. METHODS: We examined associations of trial setting and design with BCG efficacy against pulmonary and miliary or meningeal tuberculosis by conducting a systematic review, meta-analyses, and meta-regression. RESULTS: We identified 18 trials reporting pulmonary tuberculosis and 6 reporting miliary or meningeal tuberculosis. Univariable meta-regression indicated efficacy against pulmonary tuberculosis varied according to 3 characteristics. Protection appeared greatest in children stringently tuberculin tested, to try to exclude prior infection with Mycobacterium tuberculosis or sensitization to environmental mycobacteria (rate ratio [RR], 0.26; 95% confidence interval [CI], .18-.37), or infants (RR, 0.41; 95% CI, .29-.58). Protection was weaker in children not stringently tested (RR, 0.59; 95% CI, .35-1.01) and older individuals stringently or not stringently tested (RR, 0.88; 95% CI, .59-1.31 and RR, 0.81; 95% CI, .55-1.22, respectively). Protection was higher in trials further from the equator where environmental mycobacteria are less and with lower risk of diagnostic detection bias. These associations were attenuated in a multivariable model, but each had an independent effect. There was no evidence that efficacy was associated with BCG strain. Protection against meningeal and miliary tuberculosis was also high in infants (RR, 0.1; 95% CI, .01-.77) and children stringently tuberculin tested (RR, 0.08; 95% CI, .03-.25). CONCLUSIONS: Absence of prior M. tuberculosis infection or sensitization with environmental mycobacteria is associated with higher efficacy of BCG against pulmonary tuberculosis and possibly against miliary and meningeal tuberculosis. Evaluations of new tuberculosis vaccines should account for the possibility that prior infection may mask or block their effects.

Control of (multi)drug resistance and tuberculosis incidence over 23 years in the context of a well-supported tuberculosis programme in rural Malawi.

BACKGROUND: The rise in tuberculosis (TB) incidence following generalized HIV epidemics can overwhelm TB control programmes in resource-limited settings, sometimes accompanied by rising rates of drug resistance. This has led to claims that DOTS-based TB control has failed in such settings. However, few studies have described the effect of a sustained and well-supported DOTS programme on TB incidence and drug resistance over a long period. We present long-term trends in incidence and drug resistance in rural Malawi. METHODS: Karonga District in northern Malawi has an adult HIV prevalence of ≈ 10%. A research group, the Karonga Prevention Study, collaborates with the National Tuberculosis Programme to support core TB control activities. Bacteriological, demographic and clinical (including HIV status) information from all patients starting TB treatment in the District have been recorded since 1988. During that period isolates from each culture-positive TB patient were exported for drug sensitivity testing. Antiretroviral therapy (ART) has been widely available since 2005. RESULTS: Incidence of new smear-positive adult TB peaked at 124/100,000/year in the mid-90 s, but has since fallen to 87/100,000/year. Drug sensitivity information was available for 95% (3132/3307) of all culture-positive cases. Initial resistance to isoniazid was around 6% with no evidence of an increase. Fewer than 1% of episodes involved a multi-drug resistant strain. DISCUSSION: In this setting with a generalised HIV epidemic and medium TB burden, a well-supported DOTS programme enhanced by routine culture and drug sensitivity testing may well have reduced TB incidence and maintained drug resistance at low levels.

Patterns and trends of leprosy in Mexico: 1989-2009.

Data from the Mexican national leprosy control programme 1989-2009 are described and analysed. After initial increases associated with the introduction of MDT and the start of the global elimination initiative in the early 1990 s, both prevalence and incidence declined dramatically throughout most of the country. Reported prevalence fell below 1 per 10000 in 1994 and has remained below that level ever since. There is considerable geographic heterogeneity, with highest case detection rates in western states bordering the Pacific and lowest in the south east. Reasons for these geographic differences are unclear. There is evidence of increases in average age of cases, and in proportions male and MB, as in several other populations with declining leprosy. There is some evidence of increasing leprosy in states bordering on Texas, USA, where M. leprae is known to be harboured in armadillos. The relevance of armadillos for leprosy in Mexico is unclear but a priority question.

Profile: the Karonga Health and Demographic Surveillance System.

The Karonga Health and Demographic Surveillance System (Karonga HDSS) in northern Malawi currently has a population of more than 35 000 individuals under continuous demographic surveillance since completion of a baseline census (2002-2004). The surveillance system collects data on vital events and migration for individuals and for households. It also provides data on cause-specific mortality obtained by verbal autopsy for all age groups, and estimates rates of disease for specific presentations via linkage to clinical facility data. The Karonga HDSS provides a structure for surveys of socio-economic status, HIV sero-prevalence and incidence, sexual behaviour, fertility intentions and a sampling frame for other studies, as well as evaluating the impact of interventions, such as antiretroviral therapy and vaccination programmes. Uniquely, it relies on a network of village informants to report vital events and household moves, and furthermore is linked to an archive of biological samples and data from population surveys and other studies dating back three decades.

What happens to ART-eligible patients who do not start ART? Dropout between screening and ART initiation: a cohort study in Karonga, Malawi.

BACKGROUND: Routine ART programme statistics generally only provide information about individuals who start treatment. We aimed to investigate the outcome of those who are eligible but do not start ART in the Malawi programme, factors associated with this dropout, and reasons for not starting treatment, in a prospective cohort study. METHODS: Individuals having a first screening visit at the ART clinic at Karonga District Hospital, northern Malawi, between September 2005 and July 2006 were interviewed. Study follow-up to identify treatment outcomes was conducted at the clinic and in the community. Logistic regression models were used to identify factors associated with dropout before ART initiation among participants identified as clinically eligible for ART. RESULTS: 88 participants eligible for ART at their first screening visit (out of 633, 13.9%) defaulted before starting ART. Participants with less education, difficulties in dressing, a more delayed ART initiation appointment, and mid-upper arm circumference (MUAC) < 22 cm were significantly less likely to have visited the clinic subsequently. Thirty-five (58%) of the 60 participants who defaulted and were tracked at home had died, 21 before their ART initiation appointment. CONCLUSIONS: MUAC and reported difficulties in dressing may provide useful screening indicators to identify sicker ART-eligible individuals at high risk of dropping out of the programme who might benefit from being brought back quickly or admitted to hospital for observation. Individuals with less education may need adapted health information at screening. Deaths of ART-eligible individuals occurring prior to ART initiation are not included in routine programme statistics. Considering all those who are eligible for ART as a denominator for programme indicators would help to highlight this vulnerable group, in order to identify new opportunities for further improving ART programmes.

Changes in Mycobacterium tuberculosis genotype families over 20 years in a population-based study in Northern Malawi.

BACKGROUND: Despite increasing interest in possible differences in virulence and transmissibility between different genotypes of M. tuberculosis, very little is known about how genotypes within a population change over decades, or about relationships to HIV infection. METHODS AND PRINCIPAL FINDINGS: In a population-based study in rural Malawi we have examined smears and cultures from tuberculosis patients over a 20-year period using spoligotyping. Isolates were grouped into spoligotype families and lineages following previously published criteria. Time trends, HIV status, drug resistance and outcome were examined by spoligotype family and lineage. In addition, transmissibility was examined among pairs of cases with known epidemiological contact by assessing the proportion of transmissions confirmed for each lineage, on the basis of IS6110 RFLP similarity of the M tuberculosis strains. 760 spoligotypes were obtained from smears from 518 patients from 1986-2002, and 377 spoligotypes from cultures from 347 patients from 2005-2008. There was good consistency in patients with multiple specimens. Among 781 patients with first episode tuberculosis, the majority (76%) had Lineage 4 ("European/American") strains; 9% had Lineage 3 ("East-African/Indian"); 8% Lineage 1 ("Indo-Oceanic"); and 2% Lineage 2 ("East-Asian"); others unclassifiable. Over time the proportion of Lineage 4 decreased from >90% to 60%, with an increase in the other 3 lineages (p<0.001). Lineage 1 strains were more common in those with HIV infection, even after adjusting for age, sex and year. There were no associations with drug resistance or outcome, and no differences by lineage in the proportion of pairs in which transmission was confirmed. CONCLUSIONS: This is the first study to describe long term trends in the four M. tuberculosis lineages in a population. Lineage 4 has probably been longstanding in this population, with relatively recent introductions and spread of Lineages1-3, perhaps influenced by the HIV epidemic.