Prevalence and Epidemiological Profile of VDRL Seroreactivity Among Blood Donors in the Brazilian Amazon

1. Introduction

The introduction should briefly place the study in a broad context and highlight why it is Syphilis is an infectious disease caused by the bacterium Treponema pallidum, which can be transmitted through sexual contact or from mother to child. When left untreated, it may lead to severe systemic complications over time [1]. Although rare, syphilis transmission can also occur through blood transfusion; this risk is mitigated by rigorous blood donor screening and by the pathogen’s limited viability outside the human body [2].

In accordance with blood safety standards proposed by the World Health Organization, Brazil has established technical regulations for hemotherapy procedures under Consolidation Ordinance No. 5/2017, which defines guidelines and standards for blood donor selection, collection, clinical and laboratory screening, processing, storage, distribution, transfusion, surveillance, and control of blood and its components nationwide [3].

Syphilis detection in blood banks is performed through a combination of non-treponemal and treponemal tests, with the Venereal Disease Research Laboratory (VDRL) test being the most widely used non-treponemal assay. It is based on a slide flocculation reaction, allowing the titration of circulating antibodies for serological screening and monitoring of disease activity. In primary syphilis, VDRL demonstrates a diagnostic sensitivity of approximately 75%, reinforcing the need for its interpretation in conjunction with confirmatory treponemal tests [4,5].

Endemic conditions of VDRL seroreactivity among blood donors constitute a significant problem related to the wastage of blood units during donor screening [6]. It is estimated that 1.6 million blood units have been discarded due to the presence of markers, including those associated with Treponema species, the etiological agent of syphilis [7].

Another challenge associated with blood donation is the significant concern regarding the risk of acquiring infections through transfusion. It is estimated that there is a 1% probability of adverse events, including transfusion-transmitted infections, for each unit of blood transfused [8].

One strategy employed in prevalence studies involves the investigation of transfusion-transmitted infections (TTIs), such as syphilis, enabling improved donor selection approaches that enhance transfusion safety [9].

In this context, the objective of this study was to analyze VDRL seroreactivity in blood donation screening, considering the sociodemographic profile, temporal trends, prevalence rates, and geographic distribution of cases deemed ineligible for syphilis in the 1st Social Protection Region of the State of Pará, Brazil, from 2017 to 2023.

2. Materials and Methods

This was an observational, descriptive epidemiological study with a quantitative approach and a retrospective design, based on secondary data, including temporal trend analysis for the period from 2017 to 2023.

2.1. Data Collection

2.1.1. Setting and Coverage Area

Data collection was conducted at the hemotherapy service of the State of Pará, located in Belém, using information from donations collected at the main facility, external campaigns, and collection units within the regional blood network. These units included donation centers located in the shopping malls Pátio Belém and Castanheira, in Belém, and Metrópole, in Ananindeua. The study coverage area corresponded to Management Region I: the 1st Social Protection Region, which includes the municipalities of Belém, Ananindeua, Benevides, Marituba, and Santa Bárbara do Pará.

2.1.2. Data Source

Secondary data were obtained from the hemotherapy foundation database system, SBS.Web (National Hemotherapy Services Management System). The selected data included the total number of blood donations in the 1st Social Protection Region (SPR), the date of the most recent positive VDRL serological screening, the donor’s place of residence (city and neighborhood), and sociodemographic variables such as age, education level, sex, marital status, and race, for the period from 2017 to 2023.

2.1.3. Data Quality Assurance

The data used in this study were obtained from the SBS.Web system, routinely used by the Pará State Hemotherapy and Hematology Foundation to record and monitor activities related to blood donation. This database compiles essential information for each donation, including donor identification, collection date, serological screening results, and the donor’s municipality or neighborhood of residence, thereby reducing the likelihood of incomplete or inconsistent records.

The dataset extracted for the purposes of this study underwent an exploratory verification process, including an assessment of the internal consistency of sociodemographic variables, dates, and laboratory results. No inconsistencies or errors were identified that would justify additional exclusions, allowing the full use of all eligible records in the analysis.

2.2. Serological Screening and Diagnostic Algorithm

Based on Brazilian hemotherapy guidelines and regulations, screening of blood donors for syphilis in hemotherapy services includes non-treponemal testing, with VDRL used as the initial screening tool. Donors with reactive VDRL results are considered temporarily or permanently ineligible for blood donation.

Although confirmatory treponemal tests are routinely performed for donor counseling and clinical referral, the individual results of these tests were not available in the SBS.Web database used in this study. Therefore, the present analysis was restricted to VDRL seroreactivity recorded in the routine blood screening system.

2.3. Study Population and Sample

The study population comprised all blood donations with VDRL seroreactivity that met clinical screening eligibility criteria, performed at the headquarters of the Pará State Hemotherapy and Hematology Foundation and its collection network.

Population sampling was conducted using a census-based approach, including all blood donations with VDRL seroreactivity deemed ineligible for syphilis during screening, within the hemotherapy network under the jurisdiction of the Pará State hemotherapy center, from 2017 to 2023.

2.4. Inclusion Criteria

Donations eligible at clinical screening and collection, with positive VDRL serology leading to ineligibility for syphilis during laboratory screening, performed at the headquarters of the Pará State Hemotherapy and Hematology Foundation and its blood collection network, with recorded residence in the municipalities of Ananindeua, Belém and its islands, Benevides, Barcarena, Marituba, and Santa Bárbara do Pará, as well as their respective neighborhoods, from 2017 to 2023.

2.5. Exclusion Criteria

Based on the census sampling approach and the absence of incomplete records, no participants were excluded from this study.

2.6. Data Analysis

Descriptive statistics were used to evaluate the sociodemographic profile of the study population. The quantitative variable age was analyzed using measures of central tendency (mean and standard deviation) and through its absolute and relative frequency distribution. Categorical variables, including sex, marital status, education level, race, reason for donation, and the municipalities and neighborhoods involved, were analyzed based on frequency distribution, with absolute and relative proportions.

To assess the categorical frequencies observed for the variables age group, sex, marital status, education level, and race/skin color, the chi-square goodness-of-fit test was applied to determine differences in the distribution of categories within each variable [10].

To analyze variation in the mean age of the study population between 2017 and 2023, analysis of variance (ANOVA) was used. To identify which specific years showed statistically significant differences in mean age distribution, Tukey’s post hoc test was applied [11].

To evaluate trends in VDRL seroreactivity cases over the period from 2017 to 2023, linear regression analysis was performed (Montgomery, Peck, Vining, 2021). The prevalence rate of VDRL seroreactivity among all blood donations was calculated using the following formula [12].

$$\mathrm{P}\mathrm{r}\mathrm{e}\mathrm{v}\mathrm{a}\mathrm{l}\mathrm{e}\mathrm{n}\mathrm{c}\mathrm{e}\mathrm{R}\mathrm{a}\mathrm{t}\mathrm{e}=\left({\displaystyle \frac{\begin{array}{c}(\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{V}\mathrm{D}\mathrm{R}\mathrm{L}\mathrm{s}\mathrm{e}\mathrm{r}\mathrm{o}\mathrm{r}\mathrm{e}\mathrm{a}\mathrm{c}\mathrm{t}\mathrm{i}\mathrm{v}\mathrm{i}\mathrm{t}\mathrm{y}\mathrm{u}\mathrm{n}\mathrm{f}\mathrm{i}\mathrm{t}\\ \mathrm{f}\mathrm{o}\mathrm{r}\mathrm{s}\mathrm{y}\mathrm{p}\mathrm{h}\mathrm{i}\mathrm{l}\mathrm{i}\mathrm{s}\mathrm{b}\mathrm{y}\mathrm{m}\mathrm{u}\mathrm{n}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{p}\mathrm{a}\mathrm{l}\mathrm{i}\mathrm{t}\mathrm{y}/\mathrm{y}\mathrm{e}\mathrm{a}\mathrm{r})\end{array}}{\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{n}\mathrm{u}\mathrm{m}\mathrm{b}\mathrm{e}\mathrm{r}\mathrm{o}\mathrm{f}\mathrm{d}\mathrm{o}\mathrm{n}\mathrm{o}\mathrm{r}\mathrm{s}\mathrm{f}\mathrm{r}\mathrm{o}\mathrm{m}\mathrm{e}\mathrm{a}\mathrm{c}\mathrm{h}\mathrm{m}\mathrm{u}\mathrm{n}\mathrm{i}\mathrm{c}\mathrm{i}\mathrm{p}\mathrm{a}\mathrm{l}\mathrm{i}\mathrm{t}\mathrm{y}/\mathrm{y}\mathrm{e}\mathrm{a}\mathrm{r}}}\right)\times 100$$

To assess the association and similarity in the temporal behavior of VDRL seroreactivity prevalence rates across the municipalities studied, Pearson’s correlation test was used [13].

For the evaluation of hierarchical clustering among municipalities, based on the similarity of their VDRL seroreactivity prevalence rates, dendrogram analysis was performed [14].

All descriptive and inferential statistical analyses were conducted using R^® software, version 4.4.2 (R Core Team, 2024). For all inferential analyses, a significance level (alpha error) of 5% (p-value < 0.05) and a 95% confidence interval (CI) were adopted.

2.7. Ethical and Legal Considerations

As this study involved secondary data from human subjects, the project was submitted to the Certificate of Ethical Consideration (CAAE), under registration number 79115224.4.0000.517, with the National Research Ethics Commission. The requirement for informed consent (ICF) was waived, and a Data Use Agreement (DUA) was adopted. The Research Ethics Committee issued a favorable opinion under registration number 6,837,370.

3. Results

The flowchart of the record selection process, developed in accordance with the recommendations of STROBE Statement for observational studies, identified 362,705 blood donations recorded in the SBS.Web system from 2017 to 2023. All donations that met clinical eligibility criteria and presented reactive results in the non-treponemal VDRL test, resulting in ineligibility for syphilis during laboratory screening, were included in the analysis. No records with indeterminate results or missing essential fields were identified, resulting in a final analytical sample of 3730 VDRL-seroreactive donations (Figure 1).

Between 2017 and 2023, a total of 362,705 blood donations were recorded. Of these, 1.02% (3730) showed VDRL seroreactivity, indicating ineligibility for syphilis during screening. The mean age of the study population was 37.6 (±11.5) years, with a higher concentration of individuals aged 26 to 44 years, showing a highly significant difference compared to other age groups (p-value = 0.0001), accounting for 2103 (56.38%) of the total.

Regarding sex, a relatively balanced distribution was observed among VDRL-seroreactive donations, with 1803 (48%) female and 1,927 (51%) male donors. However, a statistically significant difference was observed for males (p-value = 0.042). Concerning marital status, most individuals were single, totaling 2667 (71.50%), with a highly significant difference (p-value = 0.0001) compared to other categories.

In terms of education level, the majority had completed secondary education, corresponding to 2302 (61.72%), with a highly significant difference (p-value = 0.0001) compared to other levels. Regarding race, a significant difference was observed (p-value = 0.0001), with a predominance of self-reported mixed-race (brown) individuals, totaling 3,065 (82.17%) cases (Table 1).

In the analysis of age distribution by sex in the study population, the highest concentration of individuals was observed between 20 and 40 years for both sexes, with a slight predominance of males. The mean age was 37.2 years for men and 36.1 years for women; nevertheless, a statistically significant difference in age distribution between sexes was observed (Figure 2).

The distribution of mean age among VDRL-seroreactive blood donations deemed ineligible for syphilis between 2017 and 2023 shows statistically significant variations, as indicated by the results of the ANOVA test (p < 2.2e−16), with a decreasing age trend observed over the study period (Figure 3).

Using Tukey’s multiple comparisons test (Tukey HSD), the points of greatest variation in mean age across the years were identified and graphically represented for each pair of years through confidence intervals. This analysis confirms a progressive shift toward a younger age profile in the study population over the analyzed period (Figure 4).

At the top of the distribution is the municipality of Belém, with 2532 (67.9%) recorded cases of VDRL seroreactivity, followed by Ananindeua, with 899 (24.1%) cases. The municipality of Marituba accounted for 188 (5%) records, followed by Benevides, with 63 (1.7%) cases. Barcarena recorded 38 cases (1%), while Santa Bárbara do Pará had the lowest number, with 10 occurrences (0.3%) (Figure 5).

The occurrence of VDRL seroreactivity cases deemed ineligible for syphilis, among the most affected neighborhoods within the municipalities of the 1st Social Protection Region, was most prominent in the municipality of Ananindeua. The Coqueiro neighborhood led with 298 (8%) cases, followed by Águas Lindas with 88 (2.4%) and Icuí-Guajará with 83 (2.2%), all within the same municipality.

The remaining locations with the highest frequencies were concentrated in the municipality of Belém, particularly in the neighborhoods of Jurunas, with 192 (5.1%) cases, and Guamá, with 190 (5.1%). Tapanã and Pedreira reported identical figures, with 138 (3.7%) cases each, while Sacramenta, with 109 (2.9%), Terra Firme, with 99 (2.7%), Marambaia, with 96 (2.6%), and Condor, with 91 (2.4%), showed intermediate values. The neighborhoods of Marco, with 89 (2.4%), Telégrafo, with 86 (2.3%), and Cremação, with 85 (2.3%), presented slightly lower but still relevant figures.

In addition to these 14 neighborhoods, a substantial number of donors deemed ineligible for syphilis were identified among blood donations originating from other neighborhoods, totaling 1,948 records, which corresponds to 61.4% of all cases (Figure 6).

Belém was the only municipality that showed a statistically significant increasing trend in VDRL seroreactivity cases among blood donations deemed ineligible for syphilis (p-value = 0.02). Ananindeua, Benevides, and Marituba showed an upward trend in the number of cases, although without statistical significance. In contrast, Barcarena and Santa Bárbara do Pará did not show an increasing trend, indicating stability in case numbers or an insufficient historical series to detect a trend (Figure 7, Table 2).

In the municipalities of Ananindeua and Belém, prevalence rates showed a percentage increase between 2017 and 2021, followed by relative stability in 2022 and a decline in 2023. In the municipalities of Marituba, Benevides, Barcarena, and Santa Bárbara do Pará, prevalence rates exhibited a fluctuating pattern throughout the entire study period (Figure 8).

The correlations between Ananindeua × Belém (0.92), Belém × Marituba (0.92), and Ananindeua × Marituba (0.83) indicate a strong positive linear association between the prevalence rates of these municipalities, suggesting similar epidemiological behavior over the analyzed period. As one rate increases, the other tends to increase proportionally, reinforcing the similarity in their epidemiological patterns throughout the study period (Figure 9).

In the assessment of similarity and the hierarchical correlation structure among municipalities in the 1st Social Protection Region (SPR), based on the prevalence rates of VDRL seroreactivity in blood donations deemed ineligible for syphilis, Belém and Ananindeua showed the highest similarity and were grouped first. Marituba was subsequently incorporated into this group, suggesting that its epidemiological characteristics are very similar to those observed in Belém and Ananindeua. Barcarena was later added to this cluster, indicating a moderately lower level of similarity. Benevides joined the grouping at a more distant hierarchical level, suggesting greater differences compared to the previously grouped municipalities. In contrast, Santa Bárbara do Pará remained isolated and was incorporated into the overall clustering only at the highest hierarchical level, highlighting its greater distinction relative to the other locations (Figure 10).

4. Discussion

Studies conducted in the Americas indicate a marked increase in syphilis rates, particularly among adolescents and young adults, across different genders and sexual orientations [15]. In northeastern Brazil, investigations of the epidemiological profile of patients with positive VDRL results in a network of private laboratories found that the highest incidence occurs between 22 and 32 years of age [16].

In studies assessing the incidence of positive Venereal Disease Research Laboratory (VDRL) results among patients attended at a clinical analysis laboratory in São José dos Campos, São Paulo State (SP), reactive results were most prevalent in the age groups of 21 to 29 years, representing 36.99% of cases, and 30 to 39 years, corresponding to 32.88% of reactive cases [17], which supports the findings of the present study.

The slight male predominance among VDRL-seroreactive blood donations may reflect both the greater participation of men in the donor population and differential exposure patterns, as described in studies correlating VDRL seroreactivity with the clinical-epidemiological profile of patients referred for outpatient treatment, in which 67.74% of cases were male [18].

In a multicenter Brazilian study, both the prevalence and incidence of RPR-positive syphilis were associated with lower educational attainment, defined as secondary education or less, likely due to limited access to health information and preventive measures among donors with lower education levels [19].

When analyzing marital status in this study, a predominance of single donors was observed, followed by married individuals. This finding is consistent with results from studies investigating syphilis seroreactivity among blood donors, in which the majority of participants (81.9%) were single [20].

The high proportion of single individuals is consistent with findings from a study conducted in Cuba involving 120 participants with positive syphilis serology, in which 76.7% did not have a stable relationship. In this context, it is assumed that individuals without a stable partner may engage with a greater number of sexual partners, increasing the likelihood of exposure to sexually transmitted infections (STIs) [21,22].

Considering the substantial and significant proportion of mixed-race (brown) individuals in this study compared to other categories, this evidence aligns with prevalence studies of syphilis serology in the Americas. These studies found that, when analyzing risk markers such as sex, age, marital status, education, income, and residence, all variables except sex, were associated with syphilis seroreactivity, regardless of race [23].

Racial disparities in healthcare access have been increasingly documented in recent years, including in the context of sexually transmitted infections (STIs). However, it is important to emphasize that skin pigmentation itself is not a predictive factor for STI pathogenicity; rather, it is more closely associated with underlying socioeconomic inequalities [24].

The substantial proportion of spontaneous blood donations with VDRL seroreactivity, resulting in ineligibility for syphilis during screening, corroborates findings from studies conducted in the East (Xi’an, China), which reported an increasing trend in syphilis prevalence among voluntary blood donors [25].

These elevated rates of voluntary donors deemed ineligible to donate blood due to indications of syphilis contrast with the recommendations of the World Health Organization, which emphasize that achieving a safe and adequate blood supply depends on regular, voluntary, and altruistic donors, who present a lower risk of transfusion-transmitted infections (TTIs) compared to family/replacement and paid donors [26].

To understand the higher cumulative frequency of VDRL seroreactive cases in the municipalities of Ananindeua and Belém, as well as their concentration in the fourteen neighborhoods with the highest case predominance, these findings suggest a possible influence of unfavorable urban and socioeconomic contexts on the territorial distribution of cases.

In this context, it is important to consider the Diagnostic Report of the Municipality of Belém and its Metropolitan Region, prepared in 2024 by the Fundação de Amparo e Desenvolvimento da Pesquisa, affiliated with the Federal University of Pará. The report indicates that these areas are part of the 47% of the Belém Metropolitan Region classified as having socioeconomic inequality, characterized by the presence of precarious settlements, referred to as “subnormal agglomerations” by the Brazilian Institute of Geography and Statistics. These precarious housing areas vary across metropolitan municipalities, ranging from 5% in Barcarena to 61% in Marituba [27].

The conditions in these neighborhoods, characterized by low socioeconomic status, increase the likelihood of facing barriers to accessing healthcare. Poverty, limited access to adequate housing, education, and employment, as well as socioeconomic discrimination, may increase the risk of acquiring diseases such as syphilis, potentially leading to delays in diagnosis and appropriate treatment [28].

The significant upward trend in VDRL seroreactive cases among blood donations deemed ineligible for syphilis in the municipality of Belém (p-value = 0.02), as an urban area accounting for 67.9% of the study population, reflects patterns similar to global data showing increasing syphilis notifications. These trends may be associated with determining factors such as the expansion of the sex industry, engagement in unprotected sexual practices, a false perception of curability of sexually transmitted infections, and limited awareness of the asymptomatic nature of the disease [29].

In Brazil, the rising trend in syphilis cases may be attributed to factors such as expanded testing, particularly the use of rapid tests, reduced condom use among the population, reluctance among healthcare professionals to administer penicillin in Primary Health Care (PHC), and improvements in reporting within the hemovigilance system [30].

High proportions of syphilis notifications among blood donors were also identified following laboratory assessments conducted across the State of Pará in 2020, when 563 cases of syphilis were recorded, exceeding all blood donations deemed ineligible due to HIV, hepatitis B and C, HTLV-1/2, and malaria [31].

The temporal evaluation of prevalence rates of VDRL seroreactivity among blood donations considered ineligible in this study, in the municipalities of Ananindeua and Belém, shows a pattern consistent with the increasing trend of syphilis described at the national level. This pattern aligns with the data reported in the 2024 Syphilis Epidemiological Bulletin [32], indicating agreement between the local scenario and the broader epidemiological context observed in Brazil.

Data from the 9th Hemotherapy Production Bulletin, referring to the year 2020, reported syphilis prevalence rates of 1.08% in Brazil and 1.15% in the State of Pará [31]. This represents the highest prevalence among the serological markers investigated, highlighting syphilis as a significant challenge to transfusion safety, including in regional contexts such as the 1st Social Protection Region, where rates are comparable to national and state averages.

Based on the profiles of the municipalities of Barcarena, Benevides, Marituba, and Santa Bárbara do Pará, a recognized statistical limitation in epidemiological analysis emerges; therefore, caution is required when interpreting fluctuations observed in VDRL seroreactivity prevalence rates in municipalities with low case frequencies. These differences should be interpreted carefully, as prevalence rates based on small numbers of cases are more prone to variability or fluctuations in blood donations over time [33].

This suggests that, in the presence of widely fluctuating population sizes and a stable or minimally varying numerator over time, substantial impacts on the variation of occurrence rates may arise throughout the analyzed period [34].

This situation may be related to data scarcity, smaller population sizes compared to more urbanized regions, or lower demand for blood donors. It makes the analysis of prevalence rates more challenging, as less populated regions may present disproportionately high values even with a small number of recorded cases—a phenomenon known in epidemiology as the “small numbers problem” or statistical fluctuation [35].

The greater similarity observed among more urbanized municipalities, such as Ananindeua, Belém, and Marituba, can be interpreted in light of Tobler’s First Law of Geography, which states that geographically proximate phenomena tend to exhibit greater similarity [36].

This circumstance is consistent with studies describing the prevalence of endemic diseases in regions or communities that are geographically proximate and socially connected, which may be associated with population movement, territorial integration, and shared socioeconomic conditions through effects referred to as “social spillover” [37].

Studies conducted in Brazilian metropolitan regions have reported a territorial concentration of conditions related to sexually transmitted infections, characterized by high population density, social inequalities, and intense urban mobility, favoring the formation of epidemiological clusters in integrated urban areas [38,39].

In light of this study, and considering the entire discussion regarding the historical pattern of cases and the prevalence rates of VDRL seroreactivity among blood donations deemed ineligible for syphilis in the municipalities of the 1st Social Protection Region, it is important to emphasize that epidemiological studies have traditionally considered blood donors as representative of the general population, in which donor prevalence data are subject to real-time surveillance and screening within high-risk groups [40].

Furthermore, it is noteworthy that asymptomatic patients referred from other departments, such as the local hemotherapy service during routine screening, show an increasing number of VDRL-positive results, reflecting a potential burden of infection in the local population and demanding greater attention from public authorities to infectious disease control programs [18].

Study Limitations

The limitations of this study include the use of secondary data derived from the information system of the regional public hemotherapy service. Although routine data evaluation and auditing procedures are in place, the possibility of inconsistent or incomplete records cannot be excluded.

VDRL test reactivity does not distinguish between active syphilis infection, previously treated infection, or nonspecific reactivity. This limitation may lead to misclassification bias, requiring careful consideration when interpreting the estimates presented in this study.

Therefore, the prevalence estimates identified should be interpreted with caution, particularly when used as indirect indicators of population exposure and transfusion risk.

5. Conclusions

A reduction in the mean age of the study population was observed between 2017 and 2023, with a predominance of VDRL-seroreactive donations among male, single individuals with secondary education and self-reported mixed-race (brown) status, primarily concentrated in the municipalities of Belém and Ananindeua. A significant upward trend was identified in Belém, while other municipalities exhibited fluctuating temporal patterns, with greater similarity observed among integrated urban areas. The territorial concentration of cases in socially vulnerable neighborhoods underscores the importance of using these findings to support blood network planning, strengthen serological screening, and enhance epidemiological surveillance strategies and health education initiatives.