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Corresponding author: Dr. Le Thi Phuong Mai, Department of Public Health, National Institute of Hygiene and Epidemiology, 1 Yersin St., Hai Ba Trung Dist.
Fairly high seroprevalence of Leptospira was found in Vietnam.
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The diversity of Leptospira serovars circulates in all geographical zones.
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More females were observed to be infected than males.
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Effective measures for disease prevention may be imperatively needed.
Abstract
Background
To our knowledge, this study is the first report on the seroprevalence of human leptospirosis and its epidemiological profile in 3 different geographical and climatic zones of Vietnam.
Methodology
A hospital-based surveillance in 11 public hospitals in 3 provinces in Vietnam enrolled 3,815 patients with suspected leptospirosis. Two consecutive enzyme-linked immunosorbent assay IgM and a single microscopic aggregation test were applied at a 1:100 to 1:800 dilution for probable or confirmed cases.
Results
The findings showed that of the 3,815 suspected cases, 68 (1.8%) were Leptospira-confirmed and 248 (6.5%) probable cases, whereas more than a third were positive for acute ELISA-IgM sera. Furthermore, 20 different serovars were found, of which Wolffi (14.2%), Hebdomadis (13.8%), and Icterohaemorrhagiae (12.6%) were the most predominant. The ratio of probable and confirmed cases of leptospirosis between females and males was 1.4:1, and their clinical manifestation was not specific. Cases were more likely to be detected in groups that are farmers, pet raising or livestock farming, of working age, practicing either wading in mud or walking barefoot, or exposed to heavy rainfall.
Conclusions
Analysis of human leptospirosis has indicated fairly high seroprevalence and diversity of Leptospira serovars circulating in all studied geographical zones in Vietnam. The findings suggest an imperative need for effective measures of disease prevention, especially in high-risk groups.
Leptospirosis is a zoonosis of worldwide distribution caused by the spirochete Leptospira spp. Humans are incidentally infected, usually after direct contact with urine of infected animals or indirect contact with contaminated soil and water, where Leptospira spp. can survive for a long period (
). Manifestation of human leptospirosis can vary from asymptomatic to very severe or life-threatening. However, the disease is commonly misdiagnosed as other febrile illnesses such as dengue, scrub typhus, and chikungunya (
The disease results in 1.03 million cases annually (95% CI 434,000-1,750,000) and 58,900 deaths (95% CI 23,800-9,900) worldwide. Southeast Asia is one of the regions with the highest observed estimates of disease morbidity and mortality, in which Vietnam has been classified into the high-risk incidence group (
Leptospirosis was first reported in the Mekong Delta in Vietnam in 1998, accounting for 18.8% seropositive agglutination titers ranging from l/l00 to l/3200. A later report synthesized data from a hospital study in the period from 1993-2001, indicating a leptospirosis prevalence of 2%-8%. Several individual studies, either cross-sectional or hospital-based, have shown that the seroprevalence of leptospirosis in Vietnam is high, ranging from more than 10%-80% depending on location, but Leptospira-circulating serogroups have not been updated (
). Vietnam's infectious disease reporting system, which is supervised by the General Department of Preventive Medicine of the Ministry of Health, identifies 42 diseases, including leptospirosis. However, it is a clinically-based reporting system, resulting in underreporting. According to the Infectious Disease Statistic Yearbooks from 2014-2018, none or less than 20 cases annually have been reported (
). The purpose of the present study was to determine the seroprevalence of human leptospirosis and describe its epidemiological profile in 3 different geographical and climatic zones of Vietnam.
Methodology
Study site
The hospital-based surveillance was set up in 3 provinces, namely Thai Binh, Ha Tinh and Can Tho, representing 3 different geographical and climatic zones in Vietnam. Thai Binh is located in the center of the Red River Delta, in the north of Vietnam, and has a subtropical humid climate. Ha Tinh lies in the central coast region, with very dry summers and many typhoons. Can Tho is located in the south of Vietnam, with interlacing canals and tropical dry and wet seasons.
Study population and data collection
From October 2018 to October 2019, effective surveillance was set up at 11 public hospitals, 4 of which are provincial hospitals and the other 7 district hospitals in the 3 previously mentioned provinces (Figure 1). All patients who visited the outpatient and inpatient departments of these hospitals were evaluated with regard to inclusion criteria. We included suspected cases, defined as presence of fever or a history of fever in the last 5 days associated with at least 2 of the following conditions: (i) myalgia, (ii) headache, (iii) jaundice, and (iv) conjunctival suffusion. In each study hospital, patients were first asked for demographic information, disease history, and exposure to risk factors, and then clinical signs and symptoms. Subsequently, a pair of serum samples was taken within 7 to 14 days.
Serum samples were subjected to either the enzyme-linked immunosorbent assay (ELISA) or microscopic agglutination test (MAT) for leptospirosis confirmation. We tested for IgM ELISA in the first sample. If it was positive, we also did MAT on the first sample. If either the first ELISA-IgM sample or MAT was negative, we tested for ELISA-IgM in the second sample. As for MAT, we used a set of 25 serovars, which were referred from The Institut Pasteur de Nouvelle-Calédonie (Table 1). A 2-fold serial dilution was used from 1:100 up to 1:800, considering that the highest positive dilution is the serum titer.
A confirmed positive case was defined if a 4-fold increase in titer was observed between the 2 consecutive antileptospiral IgM ELISA reactions in acute and convalescent phase samples, with a titer of 20 IU/ml or more, and if the single MAT was 1:400 for at least 1 of these serovars, whereas the dilution at 1:100 was considered a probable case (Figure 2). All tests were performed at the National Institute of Hygiene and Epidemiology and the provincial Centers for Disease Control in the 3 study provinces.
The research protocol was approved by the Institutional Ethics Committee of the National Institute of Hygiene and Epidemiology, Hanoi, Vietnam on April 12, 2018. Written informed consent was obtained from all study participants. If a participant was less than 18 years old, informed consent was obtained from the parent or guardian.
Results
Seroprevalence of leptospirosis and clinical manifestation
A total of 3,815 suspected cases were enrolled in the study, in almost equal proportions from the 3 provinces. Of the 3,815 suspected cases, 68 (1.8%) and 248 (6.5%) cases were Leptospira-confirmed and probable, respectively, while more than a third were positive for acute ELISA-IgM sera. Among the 3 provinces, the highest seroprevalence was in Thai Binh province with 7% and 2.3%, followed by Ha Tinh (8.5% and 2%), and then Can Tho (7.1% and 1%), for probable and confirmed cases, respectively (Figure 3).
Figure 3Suspected cases and seroprevalence of leptospirosis by province
Throughout the 3 provinces, 20 different serovars were found using MAT with a titer cutoff of 1:100 to 1:400, indicating diversification of Leptospira serovars circulating in Vietnam. Of the 20 serovars, 17 were detected in Ha Tinh, 16 in Thai Binh, and 12 in Can Tho. The predominant serovars, in general, were Wolffi (14.2%), Hebdomadis (13.8%), and Icterohaemorrhagiae (12.6%), but varied particularly between the 3 provinces. In Thai Binh, the highest proportion of serovars were Castellonis (12.3%), Djasiman (12.3%), and Wolffi (12.3%); in Ha Tinh, Icterohaemorrhagiae (19%) and Wolffi (15%); and in Can Tho, Hebdomadis (23.6%) and Wolffi (15.3%). It should be noted that approximately 2.8% of samples (ranging from 2% to 3.7% depending on province) reflected multiple interaction of different serovars, which were defined as unknown serovars (Figure 4). Almost all probable and confirmed cases presented with headache (97.8%), high fever (97%), and myalgia (92.4%), while typical symptoms of leptospirosis such as jaundice (14.6%) and conjunctival suffusion (18.4%) were less commonly recorded. Approximately one-third to nearly half of these had cough (47.5%), joint pain (42.7%), nausea (36.7%), and chills (32%) (Figure 5). No deaths were recorded among the enrolled patients.
In the 3 provinces included in the study, the aggregate results showed that cases of leptospirosis were detected more in women (57.9%) than in men (42.1%), with a ratio of 1.4:1. Generally, about two-thirds (63.3%) of the individuals belonged to 2 aggregated age categories of 24 to 40 and 40 to 60 years. However, this was not the case in Ha Tinh province in patients with leptospirosis in the older age group of 40 to ≥60 years. Regarding occupation, 53.5% and 83.2% of the cases in Thai Binh and Ha Tinh, respectively, were farmers, whereas this occupation group represented only 29.1% of those in Can Tho (Table 2).
Table 2Characteristics of probable and confirmed cases
Statistical analysis of probable and confirmed cases showed that more than half (53.3%) or even nearly all (92%) of them in Ha Tinh were frequently exposed to rubbish and experienced heavy rain, but not many experienced flooding. Most (74.7%) had raised pets or were involved in livestock farming. Similarly, approximately 62.7% had seen rats at home.
Regarding some risk behaviors for leptospirosis infection, outdoor and water-sport activities (15.2%) were not common among probable and confirmed cases. Furthermore, two-thirds of them commonly practiced wading in mud (66.5%) and walking barefoot (66.8), with the highest rate observed in Ha Tinh. Not using personal protective equipment and consuming uncooked food and drinks were reported in approximately 38% and 40% of cases, respectively. However, this varied by province, of which Can Tho had the highest prevalence (Table 2).
Discussion
The present study included a cohort of 3,815 patients admitted in 11 public hospitals in Vietnam, of which 68 patients were laboratory-confirmed with leptospirosis and 248 were probable cases according to MAT and ELISA-IgM tests. To our knowledge, this is the first hospital-based surveillance of leptospirosis in 3 different geographical regions of Vietnam. The country has been considered endemic to leptospirosis, with very high seroprevalence in many areas, ranging from 2% (using MAT) to 82.3% (using IgG ELISA), depending on different population groups (
). In our study, up to one-third of the subjects presented IgM antibodies and 6.5% of them were confirmed cases. Variation in disease prevalence was observed across the 3 geographical areas, similar to other studies. A multicenter study in India revealed that leptospirosis accounts for approximately 3.27% to 28.16% of cases of acute febrile illness responsible for hospital attendance. Data reported to the Bureau of Epidemiology, Ministry of Public Health, Thailand showed an average annual incidence rate from 5.3/100,000 to 6.6/100,000 population. Other countries in the region reported various figures of leptospirosis, ie, 14/100,000 population in Sri Lanka, 4.8/1 million population in the Philippines, 4.1/1 million population in Taiwan, and 18.7% seropositive in Indonesia (
). Our study found slightly but not significantly different seroprevalence among the 3 provinces. The highest prevalence of leptospirosis was in Thai Binh, which is the most populous of the 3 provinces, with a population density of 1,174 inhabitants per square kilometer, whereas Can Tho and Ha Tinh have lower densities of 859 and 215 inhabitants per square kilometer, respectively. Approximately more than 80% of the population live in rural areas in Thai Bình and Ha Tinh provinces, while this figure is only 30% in Can Tho (
The most predominant serovars found in our study were Wolffi (14.2%), Hebdomadis (13.8%), and Icterohaemorrhagiae (12.6%), which differ from serovars identified in previous studies in Vietnam such as Bataviae, Panama, Icterohaemorrhagiae, Australis, Pyrogenes, Louisian, and Hurstbridge. The predominant serovars varied among the 3 provinces included in this study. Interestingly, Bataviae, Panama, and Pyrogenes, which were previously most prevalent in the southern provinces of Vietnam, have been replaced by Hebdomadis and Wolffi. Diversity of Leptospira has been documented in many other areas throughout the world. Thirteen different genotypes were identified in 36 blood samples in the French West Indies, and approximately 10 serogroups were found in an African study (
Regarding clinical features, leptospirosis can cause a wide range of symptoms and even non-specific manifestations. However, an abrupt onset of fever, chills, conjunctival suffusion, headache, myalgia, and jaundice, in addition to historical exposures, can lead to suggestive diagnosis of leptospirosis (
). Four symptoms, including conjunctival suffusion, headache, myalgia, and jaundice, were used for the selection of suspected cases; only 2 (headache and myalgia) were commonly recorded, leading to confusion with other entities such as influenza and dengue fever. In addition, other frequently recorded symptoms in leptospirosis cases such as cough, joint pain, nausea, and chills are common in influenza and unknown febrile diseases. As a result, none of these patients were clinically diagnosed with leptospirosis by medical doctors in study hospitals until laboratory tests were confirmed. Almost all patients were treated with 1 or 2 antibiotics, of which cephalosporin accounted for 57.5%, while 9.3% and 10.6% were penicillin and doxycycline, respectively. All patients fully recovered (data not shown).
In our study, women were more likely to be infected than men, whereas the incidence of leptospirosis is much higher in men than in women in many publications. The data in Germany from 1997-2005 showed that males were infected with leptospirosis 2.6 times more frequently than females. The ratio between male and female patients was 9:1 in Albania, and 96% of cases in Seychelles were males (
). A possible explanation for the higher rate of leptospirosis in women observed in this study is that women in Vietnam account for a higher proportion of the labor force than the global and regional average. According to the International Labour Organization, in 2019, 70.9% of Vietnamese women of working age were part of the country's workforce and 85.9% of them were employed in subsistence agriculture (
). Among our probable and confirmed leptospirosis cases, approximately two-thirds work as farmers and most have raised pets or livestock. Working age groups are at higher risk for disease. Similarly, farming occupation was found to be the dominant risk factor for Leptospira infection in Indonesia, and Malaysian cattle farmers in Malaysia were at 5.9 times greater risk of leptospirosis. In New Zealand, the 25-44 year-old age group accounted for 56.9% of leptospirosis cases, with the highest proportion being livestock farming workers (64%) (
Mitoriana Porusia A. FDA, Windi Wulandari, Duangruedee Chotklang ‘Risk factors of leptospirosis incidence in agricultural area’, International Journal of Public Health Science,10 (6).
). In addition, most patients with leptospirosis practiced either wading in mud or barefoot walking, which were documented as risk behaviors associated with the disease (
). Although increased precipitation is positively associated with patients with leptospirosis or severe illness, some studies indicated that leptospirosis can occur throughout the year (
Temporal analysis of the relationship between leptospirosis, rainfall levels, and seasonality, Santa Catarina, Brazil, 2005-2015. 60. Revista do Instituto de Medicina Tropical de Sao Paulo,
2018: e39
). Most confirmed cases in our study, especially those in Can Tho, were exposed to heavy rainfall, suggesting a positive association.
In conclusion, analysis of the characteristics of human leptospirosis has indicated fairly high seroprevalence and diversity of Leptospira serovars that circulate in all studied geographical zones in Vietnam. The findings suggest an imperative need for effective measures of disease prevention, especially in high-risk groups.
Disclaimer
The funders of this study had no role in the decision to conduct this study or the analysis, findings, or writing of this manuscript.
Conflict of interest statement
The authors declare no conflict of interest.
Ethical issue
The project was approved by the Institutional Review Board of the National Institute of Hygiene and Epidemiology on April 12, 2018.
Financial support
This work was completed under the ECOMORE project, which was financed by the Agence Française de Développement.
Acknowledgments
The authors thank the Centers for Disease Control in Thai Binh, Ha Tinh, and Can Tho provinces for coordinating field work, the Institut Pasteur de Nouvelle-Calédonie, especially Dr. Cyrille Goarant, for providing technical support, and the Institut Pasteur de Paris for coordinating all the works of the project, including this study.
Authors' contributions
Le Thi Phuong Mai provided conceptualization and wrote the main text of the manuscript.
Tran Ngoc Phuong Mai and Luu Phuong Dung analyzed the data.
Tran Ngoc Phuong Mai, Van Dinh Tran, Hoang Hai, Nguyen Tu Quyet, and Phan Dang Than conducted investigation and data collection.
Pham Thanh Hai, Luong Minh Hoa, and Do Bich Ngoc tested samples.
Nguyen Thi Thu and Nguyen My Hanh prepared tables and figures.
All authors reviewed and provided comments to the manuscript.
Reference
Biscornet L.
de Comarmond J.
Bibi J.
et al.
An Observational Study of Human Leptospirosis in Seychelles.
The American journal of tropical medicine and hygiene.2020; 103: 999-1008
Temporal analysis of the relationship between leptospirosis, rainfall levels, and seasonality, Santa Catarina, Brazil, 2005-2015. 60. Revista do Instituto de Medicina Tropical de Sao Paulo,
2018: e39 (-e39)
Mitoriana Porusia A. FDA, Windi Wulandari, Duangruedee Chotklang ‘Risk factors of leptospirosis incidence in agricultural area’, International Journal of Public Health Science,10 (6).
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