Human Immunodeficiency Virus Infection in Adolescents and Mode of Transmission in Southern Africa: A Multinational Analysis of Population-Based Survey Data.

BACKGROUND: Adolescents aged 10-19 years living with human immunodeficiency virus (HIV) (ALHIV), both perinatally infected adolescents (APHIV) and behaviorally infected adolescents (ABHIV), are a growing population with distinct care needs. We characterized the epidemiology of HIV in adolescents included in Population-based HIV Impact Assessments (2015-2017) in Zimbabwe, Malawi, Zambia, Eswatini, and Lesotho. METHODS: Adolescents were tested for HIV using national rapid testing algorithms. Viral load (VL) suppression (VLS) was defined as VL <1000 copies/mL, and undetectable VL (UVL) as VL <50 copies/mL. Recent infection (within 6 months) was measured using a limiting antigen avidity assay, excluding adolescents with VLS or with detectable antiretrovirals (ARVs) in blood. To determine the most likely mode of infection, we used a risk algorithm incorporating recency, maternal HIV and vital status, history of sexual activity, and age at diagnosis. RESULTS: HIV prevalence ranged from 1.6% in Zambia to 4.8% in Eswatini. Of 707 ALHIV, 60.9% (95% confidence interval, 55.3%-66.6%) had HIV previously diagnosed, and 47.1% (41.9%-52.3%) had VLS. Our algorithm estimated that 72.6% of ALHIV (485 of 707) were APHIV, with HIV diagnosed previously in 69.5% of APHIV and 39.4% of ABHIV, and with 65.3% of APHIV and 33.5% of ABHIV receiving ARV treatment. Only 67.2% of APHIV and 60.5% of ABHIV receiving ARVs had UVL. CONCLUSIONS: These findings suggest that two-thirds of ALHIV were perinatally infected, with many unaware of their status. The low prevalence of VLS and UVL in those receiving treatment raises concerns around treatment effectiveness. Expansion of opportunities for HIV diagnoses and the optimization of treatment are imperative.

Comparison of HIV prevalence, incidence, and viral load suppression in Zambia population-based HIV impact assessments from 2016 and 2021.

BACKGROUND: The Zambian government has implemented a public health response to control the HIV epidemic in the country. Zambia conducted a population-based HIV impact assessment (ZAMPHIA) survey in 2021 to assess the status of the HIV epidemic to guide its public health programs. METHODS: ZAMPHIA 2021 was a cross-sectional two-stage cluster sample household survey among persons aged ≥15 years conducted in Zambia across all 10 provinces. Consenting participants were administered a standardized questionnaire and whole blood was tested for HIV according to national guidelines. HIV-1 viral load (VL), recent HIV infection, and antiretroviral medications were tested for in HIV-seropositive samples. Viral load suppression (VLS) was defined as <1000 copies/ml. ZAMPHIA 2021 results were compared to ZAMPHIA 2016 for persons aged 15-59 years (i.e., the overlapping age ranges). All estimates were weighted to account for nonresponse and survey design. RESULTS: During ZAMPHIA 2021, of 25 483 eligible persons aged ≥15 years, 18 804 (73.8%) were interviewed and tested for HIV. HIV prevalence was 11.0% and VLS prevalence was 86.2% overall, but was <80% among people living with HIV aged 15-24 years and in certain provinces. Among persons aged 15-59 years, from 2016 to 2021, HIV incidence declined from 0.6% to 0.3% ( P -value: 0.07) and VLS prevalence increased from 59.2% to 85.7% ( P -value: <0.01). DISCUSSION: Zambia has made substantial progress toward controlling the HIV epidemic from 2016 to 2021. Continued implementation of a test-and-treat strategy, with attention to groups with lower VLS in the ZAMPHIA 2021, could support reductions in HIV incidence and improve overall VLS in Zambia.

People Who Self-Reported Testing HIV-Positive but Tested HIV-Negative: A Multi-Country Puzzle of Data, Serology, and Ethics, 2015-2021.

During population-based HIV impact assessments (PHIAs), some participants who self-reported testing HIV-positive (PSRP) tested negative in one or more subsequent survey HIV tests. These unexpected discrepancies between their self-reported results and the survey results draw into question the validity of either the self-reported status or the test results. We analyzed PSRP with negative test results aged 15-59 years old using data collected from 2015 to 2021 in 13 countries, assessing prevalence, self-report status, survey HIV status, viral load, rapid tests and confirmatory tests, and answers to follow-up questions (such as years on treatment). Across these surveys, 19,026 participants were PSRP, and 256 (1.3%) of these were concluded to be HIV-negative after additional survey-based testing and review. PSRP determined to be HIV-negative trended higher in countries with a higher HIV prevalence, but their number was small enough that accepting self-reported HIV-positive status without testing would not have significantly affected the prevalence estimates for HIV or viral load suppression. Additionally, using more detailed information for Uganda, we examined 107 PSRP with any negative test results and found no significant correlation with years on treatment or age. Using these details, we examined support for the possible reasons for these discrepancies beyond misdiagnosis and false reporting. These findings suggest that those conducting surveys would benefit from a nuanced understanding of HIV testing among PSRP to conduct surveys ethically and produce high-quality results.

Accuracy of point-of-care HIV and CD4 field testing by lay healthcare workers in the Botswana Combination Prevention Project.

Accurate HIV and CD4 testing are critical in program implementation, with HIV misdiagnosis having serious consequences at both the client and/or community level. We implemented a comprehensive training and Quality Assurance (QA) program to ensure accuracy of point-of-care HIV and CD4 count testing by lay counsellors during the Botswana Combination Prevention Project (BCPP). We compared the performance of field testing by lay counsellors to results from an accredited laboratory to ascertain accuracy of testing. All trained lay counsellors passed competency assessments and performed satisfactorily in proficiency testing panel evaluations in 2013, 2014, and 2015. There was excellent agreement (99.6 %) between field and laboratory-based HIV test results; of the 3002 samples tested, 960 and 2030 were concordantly positive and negative respectively, with 12 misclassifications (kappa score 0.99, p < 0.0001). Of the 149 HIV-positive samples enumerated for CD4 count in the field using PIMA at a threshold of ≤ 350 cells/µl; there was 86 % agreement with laboratory testing, with only 21 misclassified. The mean difference between field and lab CD4 testing was - 16.16 cells/µl (95 % CI -5.4 to 26.9). Overall, there was excellent agreement between field and laboratory results for both HIV rapid test and PIMA CD4 results. A standard training package to train lay counsellors to accurately perform HIV and CD4 point-of-care testing in field settings was feasible, with point-of-care results obtained by lay counsellors comparable to laboratory-based testing.

Point of Care CD4 Testing in National Household Surveys - Results and Quality Indicators from Eleven Population-Based HIV Impact Assessment (PHIA) Surveys.

Population-based HIV Impact Assessments (PHIAs) are national household (HH) surveys that provide HIV diagnosis and CD4 testing with an immediate return of results. Accurate CD4 results improve HIV-positive participants' clinical care and inform the effectiveness of HIV programs. Here, we present CD4 results from the PHIA surveys that were conducted in 11 countries in sub-Saharan Africa between 2015 and 2018. All of the HIV-positive participants and 2 to 5% of the HIV-negative participants were offered Pima CD4 (Abbott, IL, USA) point-of-care (POC) tests. The quality of the CD4 test was ensured by conducting instrument verification, comprehensive training, quality control, a review of testing errors and an analysis of unweighted CD4 data by HIV status, age, gender, and antiretroviral (ARV) treatment status. Overall, CD4 testing was completed for 23,085 (99.5%) of the 23,209 HIV-positive and 7,329 (2.7%) of the 270,741 negative participants in 11 surveys. The instrument error rate was 11.3% (range, 4.4% to 15.7%). The median CD4 values among HIV-positive and HIV-negative participants (aged 15+) were 468 cells/mm3 (interquartile range [IQR], 307 to 654) and 811 cells/mm3 (IQR, 647 to 1,013), respectively. Among the HIV-positive participants (aged 15+), those with detectable ARVs had higher CD4 values (508 cells/mm3) than those with undetectable ARVs (385.5 cells/mm3). Among the HIV-positive participants (aged 15+), 11.4% (2,528/22,253) had a CD4 value of less than 200 cells/mm3, and approximately half of them (1,225/2,528 = 48.5%) had detectable ARVs, whereas 51.5% (1,303/2,528) had no detectable ARVs (P < 0.0001). We successfully implemented high quality POC CD4 testing using Pima instruments. Our data come from nationally representative surveys in 11 countries and provide unique insights regarding the CD4 distribution among HIV-positive individuals as well as the baseline CD4 values among HIV-negative individuals. IMPORTANCE The manuscript describes CD4 levels among HIV-positive individuals and baseline CD4 levels among HIV-negative individuals from 11 sub-Saharan countries, thereby highlighting the importance of CD4 markers in the context of the HIV epidemic. Despite increased ARV access in each country, advanced HIV disease (CD4 < 200 cells/mm3) persists among approximately 11% of HIV-positive individuals. Therefore, it is important that our findings are shared with the scientific community to assist with similar implementations of point-of-care testing and to conduct a review of HIV programmatic gaps.

High HIV prevalence and associated factors in Lesotho: Results from a population-based survey.

Despite extensive global efforts, sub-Saharan Africa remains disproportionately affected by the HIV epidemic. This generalized epidemic can be seen in Lesotho which in 2014 the HIV prevalence rate of those aged 15-49 years was 24.6%, with and incidence of 1.9 new infections per 100-person-year exposures. To better understand the impact of Lesotho's national HIV response and significant predictors associated with HIV infection, the Lesotho Population-based HIV Impact Assessment was conducted. This survey provided a nationally representative sample of individuals aged 15-59 years old in which participants were tested for HIV and given an individual questionnaire that included socio-demographic and behavioral risk questions. The association of factors between survey questions and HIV incident was assessed using logistic regression. Multivariate logistic regression models for men and women were constructed for each outcome using variables known to be or plausibly associated with recent or chronic infection. Overall annualized incidence among people aged 15-49 was 1.19% (95% CI 0.73-1.65) per year. The overall prevalence of HIV was 25.6% with women having significantly higher prevalence. Multiple variables, including decreased wealth status, lower education levels, marital status, condom use at first sex, and circumcision (men only) were identified as being significantly associated with HIV infection for both men and women. In combination with improving the awareness of HIV status, an increased focus is needed on AGYW and men 35-49 years old to prevent new infections. HIV education and prevention programs should focus heavily on younger age groups prior to and soon after sexual debut to prevent HIV transmission. The findings of the survey showed significant room for improvement in increasing awareness of HIV status and reinforcing the need for continued HIV prevention and treatment efforts in Lesotho to prevent new infections.

Progress towards controlling the HIV epidemic in urban Ethiopia: Findings from the 2017-2018 Ethiopia population-based HIV impact assessment survey.

INTRODUCTION: In 2014, the Joint United Nations Programme on HIV/AIDS set an 'ambitious' 90-90-90 target for 2020. By 2016, there were disparities observed among countries in their progress towards the targets and some believed the targets were not achievable. In this report, we present the results of data from the Ethiopia Population-based HIV Impact Assessment survey analyzed to assess progress with the targets and associated factors. METHODS: We conducted a nationally representative survey in urban areas of Ethiopia. Socio-demographic and behavioural data were collected from consenting participants using a structured interview. HIV testing was done following the national HIV rapid testing algorithm and seropositivity confirmed using a supplemental laboratory assay. HIV viral suppression was considered if the viral load was <1,000 RNA copies/ml. Screening antiretroviral drugs was done for efavirenz, lopinavir, and tenofovir, which were in use during the survey period. In this analysis, we generated weighted descriptive statistics and used bivariate and logistic regression analysis to examine for associations. The 95% confidence interval was used to measure the precision of estimates and the significance level set at p<0.05. RESULTS: Of 19,136 eligible participants aged 15-64 years, 614 (3% [95% CI: 0.8-3.3]) were HIV-positive, of which 79.0% (95% CI: 4.7-82.7) were aware of their HIV status, and 97.1% (95% CI: 95.0-98.3 were on antiretroviral therapy, of which 87.6% (95% CI: 83.9-90.5) achieved viral load suppression. Awareness about HIV-positive status was significantly higher among females (aOR = 2.8 [95% CI: 1.38-5.51]), significantly increased with age, the odds being highest for those aged 55-64 years (aOR = 11.4 [95% CI: 2.52-51.79]) compared to those 15-24 years, and was significantly higher among those who used condom at last sex in the past 12 months (aOR = 5.1 [95% CI: 1.68-15.25]). Individuals with secondary education and above were more likely to have achieved viral suppression (aOR = 8.2 [95% CI: 1.82-37.07]) compared with those with no education. CONCLUSION: Ethiopia made encouraging progress towards the UNAIDS 90-90-90 targets. The country needs to intensify its efforts to achieve the targets. A particular focus is required to fill the gaps in knowledge of HIV-positive status to increase case identification among population groups such as males, the youth, and those with low education.

Prevalence of and factors associated with late diagnosis of HIV in Malawi, Zambia, and Zimbabwe: Results from population-based nationally representative surveys.

INTRODUCTION: Late diagnosis of HIV (LD) increases the risk of morbidity, mortality, and HIV transmission. We used nationally representative data from population-based HIV impact assessment (PHIA) surveys in Malawi, Zambia, and Zimbabwe (2015-2016) to characterize adults at risk of LD and to examine associations between LD and presumed HIV transmission to cohabiting sexual partners. METHODS: We estimated the prevalence of LD, defined as CD4 count <350 cells/µL, among adults newly diagnosed with HIV during the surveys and odds ratios for associated factors. We linked newly diagnosed adults (index cases) to their household sexual partners and calculated adjusted odds ratios for associations between LD of the index case, viral load of the index case, and duration of HIV exposure in the relationship, and the HIV status of the household sexual partner. RESULTS: Of 1,804 adults who were newly diagnosed with HIV in the surveys, 49% (882) were diagnosed late. LD was associated with male sex, older age, and almost five times the odds of having an HIV-positive household sexual partner (adjusted odds ratio [aOR], 4.65 [95% confidence interval: 2.56-8.45]). Longer duration of HIV exposure in a relationship and higher viral load of the index case were both independently associated with higher odds of having HIV-positive household sexual partners. Individuals with HIV exposure of more than 5 years had more than three times (aOR 3.42 [95% CI: 1.63-7.18]) higher odds of being HIV positive than those with less than 2 years HIV exposure. The odds of being HIV positive were increased in individuals who were in a relationship with an index case with a viral load of 400-3499 copies/mL (aOR 4.06 [95% CI 0.45-36.46]), 3,500-9,999 copies/mL (aOR 11.32 [95% CI: 4.08-31.39]), 10,000-49,999 copies/mL (aOR 17.07 [95% CI: 9.18-31.72]), and ≥50,000 copies/mL (aOR 28.41 [95% CI: 12.18-66.28]) compared to individuals who were in a relationship with an index case with a viral load of <400 copies/mL. CONCLUSIONS: LD remains a challenge in Southern Africa and is strongly associated with presumed HIV transmission to household sexual partners. Our study underscores the need for earlier HIV diagnosis, particularly among men and older adults, and the importance of index testing.

Performance of HIV rapid testing algorithm in Nigeria: Findings from a household-based Nigeria HIV/AIDS Indicator and Impact Survey (NAIIS).

BACKGROUND: The Nigeria AIDS Indicator and Impact Survey (NAIIS), a cross-sectional household survey, was conducted in 2018 with primary objectives to estimate HIV prevalence, HIV-1 incidence, and status of UNAIDS 90-90-90 cascade. We conducted retrospective analysis of the performance of HIV rapid tests and the national HIV testing algorithm used in Nigeria. METHODS: The national algorithm included Determine HIV-1/2 as test 1 (T1), Unigold HIV-1/2 as test 2 (T2), and StatPak HIV-1/2 as the tie-breaker test (T3). Individuals reactive with T1 and either T2 or T3 were considered HIV-positive. HIV-positive specimens from the algorithm were further confirmed for the survey using supplemental test Geenius HIV-1/2. If Geenius did not confirm HIV-positive status, HIV-1 Western blot was performed. We calculated the concordance between tests and positive predictive value (PPV) of the algorithm on unweighted data. RESULTS: Of 204,930 participants (ages ≥18 months) 5,103 (2.5%) were reactive on T1. Serial testing of T1 reactive specimens with T2 or if needed by tiebreaker T3 identified 2958 (1.44%) persons as HIV-positive. Supplemental testing confirmed 2,800 (95%) as HIV-positive (HIV-1 = 2,767 [98.8%]; HIV-2 = 5 [0.2%]; dual infections = 22 [0.8%]). Concordance between T1 and T2 was 56.6% while PPV of the national algorithm was 94.5%. CONCLUSIONS: Our results show high discordant rates and poor PPV of the national algorithm with a false-positive rate of about 5.5% in the NAIIS survey. Considering our findings have major implications for HIV diagnosis in routine HIV testing services, additional evaluation of testing algorithm is warranted in Nigeria.

Factors associated with unawareness of HIV-positive status in urban Ethiopia: Evidence from the Ethiopia population-based HIV impact assessment 2017-2018.

BACKGROUND: The HIV epidemic in Ethiopia is concentrated in urban areas. Ethiopia conducted a Population-based HIV Impact Assessment (EPHIA) in urban areas between October 2017 and April 2018 to measure the status of the country's response to the epidemic. METHODS: We conducted field data collection and HIV testing in randomly selected households using the national, rapid testing algorithm with laboratory confirmation of seropositive samples using a supplemental assay. In addition to self-report on HIV diagnosis and treatment, all HIV-positive participants were screened for a set of HIV antiretroviral (ARV) drugs indicative of the first- and second-line regimens. We calculated weighted frequencies and 95% confidence intervals to assess regional variation in participants' level of unawareness of their HIV-positive status (adjusted for ARV status). RESULTS: We interviewed 20,170 survey participants 15-64 years of age, of which 19,136 (95%) were tested for HIV, 614 (3.2%) tested positive, and 119 (21%) of HIV-positive persons were unaware of their HIV status. Progress towards the UNAIDS first 90 target (90% of people living with HIV would be aware of their HIV status by 2020) substantially differed by administrative region of the country. In the bivariate analysis using log binomial regression, three regions (Oromia, Addis Ababa, and Harari), male gender, and young age (15-24 years) were significantly associated with awareness of HIV positive status. In multivariate analysis, the same variables were associated with awareness of HIV-positive status. CONCLUSION: One-fifth of the HIV-positive urban population were unaware of their HIV-positive status. The number of unaware HIV-positive individuals has a different distribution than the HIV prevalence. National and regional planning and monitoring activities could address this potentially substantial source of undetected HIV infection by increasing HIV testing among young people, men and individuals who do not use condoms.

Disease progression and mortality with untreated HIV infection: evidence synthesis of HIV seroconverter cohorts, antiretroviral treatment clinical cohorts and population-based survey data.

INTRODUCTION: Model-based estimates of key HIV indicators depend on past epidemic trends that are derived based on assumptions about HIV disease progression and mortality in the absence of antiretroviral treatment (ART). Population-based HIV Impact Assessment (PHIA) household surveys conducted between 2015 and 2018 found substantial numbers of respondents living with untreated HIV infection. CD4 cell counts measured in these individuals provide novel information to estimate HIV disease progression and mortality rates off ART. METHODS: We used Bayesian multi-parameter evidence synthesis to combine data on (1) cross-sectional CD4 cell counts among untreated adults living with HIV from 10 PHIA surveys, (2) survival after HIV seroconversion in East African seroconverter cohorts, (3) post-seroconversion CD4 counts and (4) mortality rates by CD4 count predominantly from European, North American and Australian seroconverter cohorts. We used incremental mixture importance sampling to estimate HIV natural history and ART uptake parameters used in the Spectrum software. We validated modelled trends in CD4 count at ART initiation against ART initiator cohorts in sub-Saharan Africa. RESULTS: Median untreated HIV survival decreased with increasing age at seroconversion, from 12.5 years [95% credible interval (CrI): 12.1-12.7] at ages 15-24 to 7.2 years (95% CrI: 7.1-7.7) at ages 45-54. Older age was associated with lower initial CD4 counts, faster CD4 count decline and higher HIV-related mortality rates. Our estimates suggested a weaker association between ART uptake and HIV-related mortality rates than previously assumed in Spectrum. Modelled CD4 counts in untreated people living with HIV matched recent household survey data well, though some intercountry variation in frequencies of CD4 counts above 500 cells/mm3 was not explained. Trends in CD4 counts at ART initiation were comparable to data from ART initiator cohorts. An alternate model that stratified progression and mortality rates by sex did not improve model fit appreciably. CONCLUSIONS: Synthesis of multiple data sources results in similar overall survival as previous Spectrum parameter assumptions but implies more rapid progression and longer survival in lower CD4 categories. New natural history parameter values improve consistency of model estimates with recent cross-sectional CD4 data and trends in CD4 counts at ART initiation.

A Comprehensive Approach to Assuring Quality of Laboratory Testing in HIV Surveys: Lessons Learned From the Population-Based HIV Impact Assessment Project.

BACKGROUND: Conducting HIV surveys in resource-limited settings is challenging because of logistics, limited availability of trained personnel, and complexity of testing. We described the procedures and systems deemed critical to ensure high-quality laboratory data in the population-based HIV impact assessments and large-scale household surveys. METHODS: Laboratory professionals were engaged in every stage of the surveys, including protocol development, site assessments, procurement, training, quality assurance, monitoring, analysis, and reporting writing. A tiered network of household, satellite laboratories, and central laboratories, accompanied with trainings, optimized process for blood specimen collection, storage, transport, and real-time monitoring of specimen quality, and test results at each level proved critical in maintaining specimen integrity and high-quality testing. A plausibility review of aggregate merged data was conducted to confirm associations between key variables as a final quality check for quality of laboratory results. RESULTS: Overall, we conducted a hands-on training for 3355 survey staff across 13 surveys, with 160-387 personnel trained per survey on biomarker processes. Extensive training and monitoring demonstrated that overall, 99% of specimens had adequate volume and 99.8% had no hemolysis, indicating high quality. We implemented quality control and proficiency testing for testing, resolved discrepancies, verified >300 Pima CD4 instruments, and monitored user errors. Aggregate data review for plausibility further confirmed the high quality of testing. CONCLUSIONS: Ongoing engagement of laboratory personnel to oversee processes at all levels of the surveys is critical for successful national surveys. High-quality population-based HIV impact assessments laboratory data ensured reliable results and demonstrated the impact of HIV programs in 13 countries.

Prevalence of nonsuppressed viral load and associated factors among HIV-positive adults receiving antiretroviral therapy in Eswatini, Lesotho, Malawi, Zambia and Zimbabwe (2015 to 2017): results from population-based nationally representative surveys.

INTRODUCTION: The global target for 2020 is that ≥90% of people living with HIV (PLHIV) receiving antiretroviral therapy (ART) will achieve viral load suppression (VLS). We examined VLS and its determinants among adults receiving ART for at least four months. METHODS: We analysed data from the population-based HIV impact assessment (PHIA) surveys in Eswatini, Lesotho, Malawi, Zambia and Zimbabwe (2015 to 2017). PHIA surveys are nationally representative, cross-sectional household surveys. Data collection included structured interviews, home-based HIV testing and laboratory testing. Blood samples from PLHIV were analysed for HIV RNA, CD4 counts and recent exposure to antiretroviral drugs (ARVs). We calculated representative estimates for the prevalence of VLS (viral load <1000 copies/mL), nonsuppressed viral load (NVL; viral load ≥1000 copies/mL), virologic failure (VF; ARVs present and viral load ≥1000 copies/mL), interrupted ART (ARVs absent and viral load ≥1000 copies/mL) and rates of switching to second-line ART (protease inhibitors present) among PLHIV aged 15 to 59 years who participated in the PHIA surveys in Eswatini, Lesotho, Malawi, Zambia and Zimbabwe, initiated ART at least four months before the survey and were receiving ART at the time of the survey (according to self-report or ARV testing). We calculated odds ratios and incidence rate ratios for factors associated with NVL, VF, interrupted ART, and switching to second-line ART. RESULTS: We included 9200 adults receiving ART of whom 88.8% had VLS and 11.2% had NVL including 8.2% who experienced VF and 3.0% who interrupted ART. Younger age, male sex, less education, suboptimal adherence, receiving nevirapine, HIV non-disclosure, never having married and residing in Zimbabwe, Lesotho or Zambia were associated with higher odds of NVL. Among people with NVL, marriage, female sex, shorter ART duration, higher CD4 count and alcohol use were associated with lower odds for VF and higher odds for interrupted ART. Many people with VF (44.8%) had CD4 counts <200 cells/µL, but few (0.31% per year) switched to second-line ART. CONCLUSIONS: Countries are approaching global VLS targets for adults. Treatment support, in particular for younger adults, and people with higher CD4 counts, and switching of people to protease inhibitor- or integrase inhibitor-based regimens may further reduce NVL prevalence.