# Leptospirosis Prevalence and Risk Factors Among Patients Presenting With Fever to 4 Healthcare Sites in Sub-Saharan Africa and South East Asia: An International Multisite Observational and Nested Case-Control Study

John A Crump, Polycarp Mogeni, Sara A Ajanovic, Justina M Bramugy, Mabvuto Chimenya, Edward W Green, Sham Lal, David C W Mabey, Mayfong Mayxay, Paul N Newton, et al.

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## To cite this version:

John A Crump, Polycarp Mogeni, Sara A Ajanovic, Justina M Bramugy, Mabvuto Chimenya, et al. "Leptospirosis Prevalence and Risk Factors Among Patients Presenting With Fever to 4 Healthcare Sites in Sub-Saharan Africa and South East Asia: An International Multisite Observational and Nested Case-Control Study." Journal of Infectious Diseases, In press, pp.jiaf464. 10.1093/infdis/jiaf464.pasteur-05444093

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https://pasteur.hal.science/pasteur-05444093v1

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# Leptospirosis Prevalence and Risk Factors Among Patients Presenting With Fever to 4 Healthcare Sites in Sub-Saharan Africa and South East Asia: An International Multisite Observational and Nested Case-Control Study

John A. Crump,<fnref n="1" />,º Polycarp Mogeni,²,º Sara A. Ajanovic,³,⁴,⁵,º Justina M. Bramugy,³ Mabvuto Chimenya,⁶ Edward W. Green,⁶,⁷ Sham Lal,²,º

David C. W. Mabey,²,º Mayfong Mayxay,⁸,⁹,¹⁰,¹¹,º Paul N. Newton,⁸,⁹,º Ioana D. Olaru,²,¹²,¹³,º Heidi Hopkins,²,º and Mathieu Picardeau¹⁴,º; on behalf of the Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Consortium

**Background.** We investigated the prevalence, diversity, and risk factors for acute leptospirosis in the Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) study.

**Methods.** Febrile patients aged ≥2 months in Laos, Malawi, Mozambique, and Zimbabwe underwent a standardized clinical and exposure assessment. Acute and convalescent serum were tested by *Leptospira* microscopic agglutination test (MAT) and acute plasma by *Ifb1* polymerase chain reaction. A ≥4-fold rise in antibody titer, or a single reciprocal titer ≥800, or *Leptospira* PCR positive defined confirmed leptospirosis. The identity of possible infecting strains was investigated by MAT and sequencing of PCR products.

**Results.** Of 7851 febrile participants enrolled, 134 (1.7%) had confirmed leptospirosis: 88 (4.6%) in Laos, 17 (1.0%) Malawi, 7 (0.3%) Mozambique, and 22 (1.2%) Zimbabwe, and 23 (0.8%) had supportive evidence of leptospirosis. Participants with leptospirosis had greater odds of headache (adjusted odds ratio [aOR] 2.20, *P* < .001), rash (aOR 1.45, *P* < .001), conjunctivitis (aOR 3.33, *P* < .001), and jaundice (aOR 1.75, *P* < .001); and had greater odds of being older (aOR 1.02 per year, *P* < .001), working in rice fields (aOR 6.24, *P* < .001), drinking river water (aOR 5.11, *P* = .001). Predominant reactive *Leptospira* serogroups were Ballum and Icterohemorrhagiae at African sites, and Australis in Laos. Identified species were *Leptospira borgpetersenii*, *L. interrogans*, and *L. kirschneri*.

**Conclusions.** Leptospirosis was a cause of febrile illness at all sites. Some clinical features helped to identify patients with leptospirosis. Interventions related to rice field work and river exposure may prevent disease. Diverse *Leptospira* serogroup reactivity was observed and may suggest potential hosts.

**Keywords.** fever; *Leptospira*; leptospirosis; prevalence; risk factors.

Leptospirosis is a zoonotic bacterial infection established as a major cause of fever in Asia [1] and increasingly recognized as an important cause of fever in Africa [2, 3]. Leptospirosis commonly presents as a febrile illness in humans that is difficult to distinguish clinically from other causes of fever and may be

severe and fatal [4]. Reference standard laboratory diagnosis relies on microscopic agglutination testing (MAT) of paired sera, nucleic acid amplification tests, and bacterial isolation rarely available in low-resource endemic areas [5]. *Leptospira* includes diverse pathogenic and intermediate pathogenic strains capable of causing human illness across a wide range of *Leptospira* species, serogroups, and serovars. Nonhuman animals are the reservoirs of *Leptospira* spp. associated with human illness. Infected animals may remain well or experience a range of clinical manifestations from infertility, abortion, and poor milk yield, and become carriers, continuing to excrete *Leptospira* in their urine, in turn contaminating water and moist soil [5]. Humans are infected following direct contact with the urine of animals shedding *Leptospira* or indirectly by contacted with urine-contaminated environments [6]. The organism

<footnote n="1">¹Centre for International Health, University of Otago, Dunedin, New Zealand; ²London School of Hygiene and Tropical Medicine, London, United Kingdom; ³Centro de Investigação em Saúde de Manhãça, Maputo, Mozambique; ⁴ISGlobal—Barcelona Institute for Global Health, Barcelona, Spain; ⁵University of Barcelona, Barcelona, Spain; ⁶Malawi-Liverpool-Wellcome Trust Clinical Research Programme, Blantyre, Malawi; ⁷Department of Clinical Sciences, Liverpool School of Tropical Medicine, Liverpool, United Kingdom; ⁸Lao-Oxford-Mahosot Hospital-Wellcome Trust Research Unit (LOMWRU), Microbiology Laboratory, Mahosot Hospital, Vientiane, Lao PDR; ⁹Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom; ¹⁰Institute of Research and Education Development, University of Health Sciences, Ministry of Health, Vientiane, Lao PDR; ¹¹Saw Swee Hock School of Public Health, National University of Singapore, Singapore; ¹²Biomedical Research and Training Institute, Harare, Zimbabwe; ¹³Institute of Medical Microbiology, University Hospital Münster, Münster, Germany; and ¹⁴Unité Biologie des Spirochètes, French National Reference Center for Leptospirosis, WHO Collaborating Center for Reference and Research on Leptospirosis, Institut Pasteur, Paris, France</footnote>

<footnote>Received 11 June 2025; accepted 28 August 2025; published online 16 October 2025
Correspondence: John A. Crump, MD, Centre for International Health, University of Otago, PO Box 56, Dunedin 9054, Otago, New Zealand (john.crump@otago.ac.nz).</footnote>

<footnote>**The Journal of Infectious Diseases®**</footnote>

<footnote>© The Author(s) 2025. Published by Oxford University Press on behalf of Infectious Diseases Society of America.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
https://doi.org/10.1093/infdis/jiaf464</footnote>
enters the human host via mucous membranes and broken skin. *Leptospira* have a complex and diverse ecology with variation in dominant host species by location and over time. Rodents are recognized as major reservoirs of *Leptospira* worldwide [5] and activities exposing persons to rodent urine, including residence in an urban slum, proximity to open sewers, exposure to floodwaters, and rice paddy work, are associated with increased risk for disease [5-7]. Risk factors for leptospirosis are less well understood in mainland Africa than elsewhere. While rodents are likely to be important hosts, cattle exposure was associated with increased odds for disease at some locations [8-11].

The lack of widespread access to reference diagnostic tests for leptospirosis, their complexity, and the limited investment in leptospirosis as a neglected tropical zoonosis mean that our understanding of its role as a cause of febrile illness and associated risk factors is incomplete [12, 13]. The Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) study [14] sought to describe treatable and preventable causes of febrile illness among inpatients and outpatients at multiple sites with little or no data on leptospirosis and other causes of fever using reference standard diagnostics.

To provide insights into variation in the role of acute leptospirosis as a cause of febrile illness, the diversity of infecting *Leptospira* species and serogroups, clinical features, clinical diagnosis, behaviors, and exposures associated with disease, we undertook a secondary analysis of FIEBRE data focused on participants with and without leptospirosis. We extended this work by examining the predominant reactive *Leptospira* serogroups associated with human illness in FIEBRE sites and their potential hosts.

## METHODS

The overall design of FIEBRE has been described in detail elsewhere [14]. A brief summary is provided below, including detailed methods for the present analysis.

### Study Sites

FIEBRE was conducted at 4 sites: Vientiane Provincial Hospital, Phonghong, Lao People's Democratic Republic (Laos); Chikwawa District Hospital, Malawi; Manhiça District Hospital and General Macamo Hospital, Maputo, Mozambique; and Sally Mugabe Central Hospital, Parirenyatwa Hospital, Chitungwiza General Hospital, and 3 primary care clinics in Harare, Zimbabwe.

### Enrollment, Follow-up, Data Collection, and Sample Collection for Febrile Participants

Febrile outpatients and inpatients aged ≥2 months were eligible for enrollment if they had a tympanic or axillary temperature of ≥37.5°C at presentation and had not been hospitalized or undergone surgery in the previous month. Outpatients were eligible if they were residents within a defined catchment area

around the health facility at the time of enrollment. For outpatients aged ≥15 years, those without symptoms of either lower respiratory infection or diarrheal disease were eligible. For outpatients aged ≥2 months to <15 years, those without symptoms of diarrheal disease were eligible. After provision of informed consent, demographic information and a standardized clinical and exposure history were taken, and physical examination was performed on all participants and recorded on a case report form (Supplementary Appendix). The exposure history included questions relevant to risk for leptospirosis, such as exposure to surface water, rice fields, and animals, including livestock.

Whole blood and ethylenediaminetetraacetic acid (EDTA) blood were collected on day 0, and participants were asked to return 28 days after enrollment (acceptable range: 26-48 days) for collection of convalescent whole blood. Serum and plasma were separated, aliquoted, and stored at -70°C. Samples were shipped on dry ice to the London School of Hygiene and Tropical Medicine, London, United Kingdom, for distribution on dry ice to reference laboratories.

## Laboratory Methods

### Leptospira Serology

At the Unité Biologie des Spirochètes, French National Reference Center for Leptospirosis, WHO Collaborating Centre for Reference and Research on Leptospirosis, Institut Pasteur, Paris, France, acute and convalescent serum was first screened by *Leptospira fainei* serovar Hurstbridge IgM enzyme-linked immunosorbent assay (ELISA) [15]. Serum of participants screening positive by ELISA on the acute or convalescent sample proceeded to testing with the *Leptospira* standard MAT. The MAT panels were based on the World Health Organization recommended list of globally representative reference *Leptospira* strains [16] adjusted to incorporate African and Asian regional isolates (Supplementary Appendix).

### Leptospira Nucleic Acid Amplification Testing and Speciation

Acute plasma collected in EDTA from all participants was tested by polymerase chain reaction (PCR) to the *Ifb1* pathogenic *Leptospira* target [17]. Among samples positive by *Leptospira Ifb1* PCR, PCR products were sequenced for *Leptospira* speciation [18, 19].

### Study Definitions

For the purpose of this analysis and consistent with widely accepted case definitions [20], we defined confirmed acute leptospirosis as a participant with ≥4-fold rise in antibody titer between acute and convalescent sample or a single reciprocal titer ≥800, or *Leptospira* PCR positive for the *Ifb1* gene target. We defined supportive evidence of leptospirosis as a participant with a single reciprocal titer ≥200 but <800. The predominant reactive serogroup was defined as the serogroup for the reacting serovar with the highest MAT titer.
## Potential Host Animal Species of *Leptospira*

To generate hypotheses about potential host animals of *Leptospira* by site, we interrogated the Dr Leopold Kirschner database of *Leptospira* species and serovar isolations and detections from animals worldwide at the serogroup level [21, 22]. We did this for leading predominant reactive *Leptospira* serogroups by study site for participants with serologically confirmed leptospirosis at the level of country and United Nations region.

## Sample Size

The sample size for this analysis was driven by that of the parent FIEBRE study and is described in detail elsewhere [14].

## Statistical Analyses

We used median and interquartile range (IQR) to summarize continuous variables, while categorical variables were summarized using percentages. We used range to summarize antibody titers by serogroup. To investigate clinical predictors and exposures associated with leptospirosis, we used separate logistic regression models to assess the associations between confirmed or supportive evidence of leptospirosis and the sociodemographic and clinical variables, adjusted for sex and age without data imputation. To account for the possibility of differences in predominant exposure pathways between Africa and Asia, we conducted subgroup analyses separately for data from African sites and from the Asian site. We used bivariable analyses, retaining significant covariates at *P* < .1 and backward exclusion of nonsignificant covariates with *P* > .05 to arrive at a final parsimonious model. The variance inflation factor was used to examine potential collinearity. Age and sex were considered a priori confounders and were included in multivariable models. Uncertainty was reported as 95% confidence intervals (CIs). To explore the possibility of variation in predominant transmission routes by continent, and particularly the role of ruminant livestock as sources for leptospirosis at African sites [8–10], we undertook subgroup analyses separately for data from Africa and Laos. All data analyses were performed using STATA version 18.0 (Stata Corp, College Station, Texas).

## Research Ethics

Ethics approval was obtained from the Lao National Ethics Committee for Health Research Committee and the Oxford Tropical Research Ethics Committee, United Kingdom, for Laos; the University of Malawi College of Medicine Research and Ethics Committee and the Liverpool School of Tropical Medicine Research Ethics Committee for Malawi; Comité Institucional de Medical Research Bioética para a Saúde do Centro de Investigação em Saúde de Manhiça and the Comité Nacional de Bioética em Saúde de Moçambique for Mozambique; and the Medical Research Council of

Zimbabwe for Zimbabwe. The study was also approved by the research and ethics committee of the London School of Hygiene and Tropical Medicine. Written informed consent was obtained from all study participants or their parents or guardians.

## Role of the Funding Source

The funder had no role in study design; in the collection, analysis, and interpretation of data; in the writing of the report; nor in the decision to submit the paper for publication.

## RESULTS

Of 7851 febrile participants enrolled across study sites from 22 June 2018 to 31 March 2021, 142 had missing serology and PCR results, and 7413 (94.4%) had at least day 0 or day 28 serum samples available for testing for leptospirosis, of which 4682 (59.6%) had paired day 0 and day 28 sera available, and 7273 (92.6%) had plasma available for PCR analysis (Figure 1).

Of 7851 febrile participants, 134 (1.7%) had confirmed leptospirosis: 89 (1.1%) by ≥4-fold rise in *Leptospira* MAT titer between the acute and convalescent sample, 13 (0.2%) by a single *Leptospira* MAT reciprocal titer ≥800, and 38 (0.5%) by *Leptospira* lfb1 PCR. Among those confirmed by *Leptospira* lfb1 PCR, the detected *Leptospira* species are shown in Table 1. Confirmed leptospirosis was identified in 88 (4.6%) participants from Laos, 177 (1.0%) from Malawi, 7 (0.3%) from Mozambique, and 22 (1.2%) from Zimbabwe (Table 2). Of 89 participants with a ≥4-fold rise in *Leptospira* MAT titer between the acute and convalescent sample and a *Leptospira* lfb1 PCR result, 6 (6.7%) were also *Leptospira* lfb1 PCR positive. Of 12 participants with a single *Leptospira* MAT reciprocal titer ≥800 and a *Leptospira* lfb1 PCR result, 0 (0.0%) were *Leptospira* lfb1 PCR positive. Of 22 participants positive by *Leptospira* lfb1 PCR and paired sera tested by *Leptospira* MAT, 6 (27.3%) also had a ≥4-fold rise in *Leptospira* MAT titer between the acute and convalescent sample. Of 16 participants who were positive by *Leptospira* lfb1 PCR and a serum sample tested by *Leptospira* MAT, 0 (0.0%) had a single *Leptospira* MAT reciprocal titer ≥800. In Laos 14 (15.9%) of 88 cases of confirmed leptospirosis were diagnosed by PCR, whereas in Africa sites 24 (52.2%) of 46 cases of confirmed leptospirosis were diagnosed by PCR.

The median (IQR) age of all participants with confirmed leptospirosis was 27 (17–41) years compared with 18 (4.8–33) years for those without confirmed leptospirosis (*P* < .001). Of pediatric participants, 31 (0.9%) had confirmed leptospirosis, and the median (IQR) age was 8 (5–11) years. Among adult participants, 103 (2.5%) had confirmed leptospirosis, and the median (IQR) age was 35 (23–46) years. Among all participants, 62 (46.3%) with confirmed leptospirosis were female compared with 4006 (52.9%) of those without confirmed leptospirosis (*P* = .128).
<figure><img src="image_3.png" /><figcaption>**Figure 1.** Study flow diagram for acute leptospirosis prevalence and risk factors in the Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) study, 2018–2021.</figcaption></figure>

<table><caption>**Table 1.** *Leptospira* Species Identified Among Ifb1 PCR-Positive Participants in the Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Study, 2018–2021</caption><thead><tr><th>Site</th><th>Species</th><th>N Positive Age &lt;15 y</th><th>N Positive Age ≥15 y</th><th>N Positive All Ages</th></tr></thead><tbody><tr><td rowspan="2">Laos</td><td>L. interrogans</td><td>3</td><td>10</td><td>13</td></tr><tr><td>L. kirschneri</td><td>0</td><td>1</td><td>1</td></tr><tr><td rowspan="2">Malawi</td><td>L. borgpetersenii</td><td>0</td><td>1</td><td>1</td></tr><tr><td>L. interrogans</td><td>2</td><td>3</td><td>5</td></tr><tr><td rowspan="2">Mozambique</td><td>L. interrogans</td><td>2</td><td>0</td><td>2</td></tr><tr><td>L. kirschneri</td><td>0</td><td>4</td><td>4</td></tr><tr><td rowspan="2">Zimbabwe</td><td>L. borgpetersenii</td><td>0</td><td>2</td><td>2</td></tr><tr><td>L. interrogans</td><td>6</td><td>4</td><td>10</td></tr><tr><td rowspan="2">Overall</td><td>L. borgpetersenii</td><td>0</td><td>3</td><td>3</td></tr><tr><td>L. interrogans</td><td>13</td><td>17</td><td>30</td></tr><tr><td></td><td>L. kirschneri</td><td>0</td><td>5</td><td>5</td></tr></tbody></table>

Of 7851 febrile participants, 23 (0.3%) had supportive evidence of leptospirosis. Supportive evidence of leptospirosis was identified in 18 (2.3%) participants from Laos, 1 (0.3%) from Malawi, 1 (0.1%) from Mozambique, and 3 (0.5%) from Zimbabwe (Table 2).

## Clinical Characteristics and Predictors of Leptospirosis

Clinical characteristics of participants with and without confirmed or supportive evidence of leptospirosis are shown in Table 3. On bivariable analysis of participants with confirmed

or supportive evidence of leptospirosis compared with those without the median (IQR) admission temperature was 38.0°C (37.6°C, 38.5°C) versus 38.1°C (37.7°C, 38.9°C) (odds ratio [OR] 0.77 per °C, *P* = .026), median (IQR) oxygen saturation 96.0% (95.0%, 98.0%) versus 97.0% (95.0%, 98.0%) (OR 0.96 per %, *P* = .008), median (IQR) respiratory rate was 22.0 (20.0, 25.0) versus 22.0 (20.0, 28.0) per minute (OR 0.96 per breath per minute, *P* = .001), headache was reported by 114 (78.6%) versus 3440 (66.1%) (OR 1.9, *P* = .002), conjunctivitis was observed in 4 (2.5%) versus 57 (0.8%) (OR 3.40,
<table><caption>**Table 2.** Confirmed or Supportive Evidence of Leptospirosis by Study Site, Age Group, and Case Definition Criterion for Patients Presenting With Fever for Health Care in the Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Study, 2018–2021</caption><thead><tr><th rowspan="2">Age Group, y</th><th colspan="3">≥4-Fold Rise In<br>Leptospira MAT Titer<br>Between Paired Sera</th><th colspan="3">Single Leptospira MAT<br>Titer ≥800</th><th colspan="3">Lfb1 PCR Positive</th><th colspan="3">Total Confirmed<br>Leptospirosis</th><th colspan="3">Single Reciprocal Titer<br>≥200 But &lt;800</th><th colspan="3">Total Confirmed or<br>Supportive Evidence of<br>Leptospirosis</th></tr><tr><th>N</th><th>n</th><th>(%)</th><th>N</th><th>n</th><th>(%)</th><th>N</th><th>n</th><th>(%)</th><th>N</th><th>n</th><th>(%)</th><th>N</th><th>n</th><th>(%)</th><th>N</th><th>n</th><th>(%)</th></tr></thead><tbody><tr><td>Laos</td><td>&lt;15</td><td>241</td><td>9 (3.73)</td><td>352</td><td>5 (1.42)</td><td>654</td><td>3 (0.46)</td><td>717</td><td>16 (2.23)</td><td>352</td><td>5 (1.42)</td><td>717</td><td>21 (2.93)</td><td>352</td><td>5 (1.42)</td><td>717</td><td>21 (2.93)</td></tr><tr><td>≥15</td><td>735</td><td>59</td><td>(8.03)</td><td>438</td><td>4 (0.90)</td><td>1197</td><td>11 (0.92)</td><td>1206</td><td>72 (5.97)</td><td>438</td><td>13 (2.97)</td><td>1206</td><td>85 (7.05)</td><td>438</td><td>13 (2.97)</td><td>1206</td><td>85 (7.05)</td></tr><tr><td>All ages</td><td>976</td><td>68</td><td>(6.97)</td><td>790</td><td>9 (1.14)</td><td>1851</td><td>14 (0.76)</td><td>1923</td><td>88 (4.58)</td><td>790</td><td>18 (2.28)</td><td>1923</td><td>106 (5.51)</td><td>790</td><td>18 (2.28)</td><td>1923</td><td>106 (5.51)</td></tr><tr><td>Malawi</td><td>&lt;15</td><td>674</td><td>2 (0.30)</td><td>244</td><td>1 (0.41)</td><td>863</td><td>2 (0.23)</td><td>930</td><td>5 (0.54)</td><td>244</td><td>0 (0.00)</td><td>1020</td><td>5 (0.49)</td><td>244</td><td>0 (0.00)</td><td>1020</td><td>5 (0.49)</td></tr><tr><td>≥15</td><td>681</td><td>8 (1.17)</td><td>132</td><td>0 (0.00)</td><td>739</td><td>4 (0.54)</td><td>819</td><td>12 (0.54)</td><td>132</td><td>1 (0.76)</td><td>819</td><td>13 (1.59)</td><td>132</td><td>1 (0.76)</td><td>819</td><td>13 (1.59)</td></tr><tr><td>All ages</td><td>1355</td><td>10 (0.74)</td><td>376</td><td>1 (0.27)</td><td>1602</td><td>6 (0.37)</td><td>1749</td><td>17 (1.47)</td><td>376</td><td>1 (0.27)</td><td>1749</td><td>18 (1.03)</td><td>376</td><td>1 (0.27)</td><td>1749</td><td>18 (1.03)</td></tr><tr><td>Mozambique</td><td>&lt;15</td><td>561</td><td>0 (0.00)</td><td>546</td><td>0 (0.00)</td><td>1122</td><td>2 (0.18)</td><td>1157</td><td>2 (0.17)</td><td>546</td><td>0 (0.00)</td><td>1157</td><td>2 (0.17)</td><td>546</td><td>0 (0.00)</td><td>1157</td><td>2 (0.17)</td></tr><tr><td>≥15</td><td>653</td><td>2 (0.31)</td><td>356</td><td>0 (0.00)</td><td>998</td><td>4 (0.40)</td><td>1016</td><td>5 (0.49)</td><td>356</td><td>1 (0.28)</td><td>1016</td><td>6 (0.59)</td><td>356</td><td>1 (0.28)</td><td>1016</td><td>6 (0.59)</td></tr><tr><td>All ages</td><td>1214</td><td>2 (0.16)</td><td>902</td><td>0 (0.00)</td><td>2120</td><td>6 (0.28)</td><td>2173</td><td>7 (0.32)</td><td>902</td><td>1 (0.11)</td><td>2173</td><td>8 (0.37)</td><td>902</td><td>1 (0.11)</td><td>2173</td><td>8 (0.37)</td></tr><tr><td>Zimbabwe</td><td>&lt;15</td><td>408</td><td>1 (0.25)</td><td>372</td><td>1 (0.27)</td><td>703</td><td>6 (0.85)</td><td>828</td><td>8 (0.97)</td><td>372</td><td>1 (0.27)</td><td>828</td><td>9 (1.09)</td><td>372</td><td>1 (0.27)</td><td>828</td><td>9 (1.09)</td></tr><tr><td>≥15</td><td>729</td><td>8 (1.10)</td><td>291</td><td>2 (0.69)</td><td>997</td><td>6 (0.60)</td><td>1036</td><td>14 (1.35)</td><td>291</td><td>2 (0.69)</td><td>1036</td><td>16 (1.54)</td><td>291</td><td>2 (0.69)</td><td>1036</td><td>16 (1.54)</td></tr><tr><td>All ages</td><td>1137</td><td>9 (0.79)</td><td>663</td><td>3 (0.45)</td><td>1700</td><td>12 (0.71)</td><td>1864</td><td>22 (1.18)</td><td>663</td><td>3 (0.45)</td><td>1864</td><td>25 (1.34)</td><td>663</td><td>3 (0.45)</td><td>1864</td><td>25 (1.34)</td></tr><tr><td>Total</td><td>&lt;15</td><td>1844</td><td>12 (0.64)</td><td>1514</td><td>7 (0.46)</td><td>3342</td><td>13 (0.39)</td><td>3632</td><td>31 (0.85)</td><td>1514</td><td>6 (0.40)</td><td>3632</td><td>37 (1.02)</td><td>1514</td><td>6 (0.40)</td><td>3632</td><td>37 (1.02)</td></tr><tr><td>≥15</td><td>2798</td><td>77 (2.75)</td><td>1217</td><td>6 (0.49)</td><td>3931</td><td>25 (0.64)</td><td>4077</td><td>103 (2.53)</td><td>1217</td><td>17 (1.40)</td><td>4077</td><td>120 (2.94)</td><td>1217</td><td>17 (1.40)</td><td>4077</td><td>120 (2.94)</td></tr><tr><td>All ages</td><td>4682</td><td>89 (1.90)</td><td>2731</td><td>13 (0.48)</td><td>7273</td><td>38 (0.52)</td><td>7709</td><td>134 (1.75)</td><td>2731</td><td>23 (0.84)</td><td>7709</td><td>157 (2.04)</td><td>2731</td><td>23 (0.84)</td><td>7709</td><td>157 (2.04)</td></tr></tbody></table>

Abbreviations: Laos, Lao People's Democratic Republic; MAT, standard microscopic agglutination test; PCR, polymerase chain reaction.
<table><caption>**Table 3.** Clinical Characteristics of Participants With and Without Confirmed Leptospirosis or Supportive Evidence of Leptospirosis by Bivariable and Logistic Regression, Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Study, 2018–2021</caption><thead><tr><th rowspan="2"></th><th colspan="2">Confirmed or Supportive N = 157</th><th colspan="2">Neither N = 7551</th><th colspan="3">Unadjusted Model</th><th colspan="3">Adjusted Model**</th></tr><tr><th>n</th><th>(%)</th><th>n</th><th>(%)</th><th>OR</th><th>95% CI</th><th>P-value</th><th>aOR</th><th>95% CI</th><th>P-value</th></tr></thead><tbody><tr><td>Sex</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Female</td><td>75</td><td>(47.8)</td><td>3993</td><td>(52.9)</td><td>Ref.</td><td></td><td></td><td>Ref.</td><td></td><td></td></tr><tr><td>Male</td><td>82</td><td>(52.2)</td><td>3568</td><td>(47.1)</td><td>1.23</td><td>(.89,1.68)</td><td>.205</td><td>1.33</td><td>(.75, 2.38)</td><td>.332</td></tr><tr><td>Age years, median (IQR)</td><td>27.0</td><td>(17.0-43.0)</td><td>18.0</td><td>(4.7-33.0)</td><td>1.02</td><td>(1.01, 1.03)</td><td>&lt;.001</td><td>1.01</td><td>(1.00, 1.02)</td><td>.107</td></tr><tr><td>Health facility admission status</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Outpatient</td><td>80</td><td>(51.0)</td><td>4320</td><td>(57.2)</td><td>Ref.</td><td></td><td></td><td>Ref.</td><td></td><td></td></tr><tr><td>Inpatient</td><td>77</td><td>(49.0)</td><td>3232</td><td>(42.8)</td><td>1.29</td><td>(.94, 1.76)</td><td>.118</td><td>1.43</td><td>(.75, 2.74)</td><td>.278</td></tr><tr><td>Admission temperature °C, median (IQR)</td><td>38.0</td><td>(37.6-38.5)</td><td>38.1</td><td>(37.7-38.9)</td><td>0.77</td><td>(.61, .97)</td><td>.026</td><td></td><td></td><td></td></tr><tr><td>Systolic blood pressure mmHg, median (IQR)</td><td>110.0</td><td>(100.0-120.0)</td><td>110.0</td><td>(97.0-120.0)</td><td>1.00</td><td>(1.00, 1.00)</td><td>.990</td><td></td><td></td><td></td></tr><tr><td>Diastolic blood pressure mmHg, median (IQR)</td><td>70.0</td><td>(60.0-80.0)</td><td>70.0</td><td>(60.0-79.0)</td><td>0.99</td><td>(.98, 1.01)</td><td>.384</td><td></td><td></td><td></td></tr><tr><td>O<sub>2</sub> saturation %, median (IQR)</td><td>96.0</td><td>(95.0-98.0)</td><td>97.0</td><td>(95.0-98.0)</td><td>0.97</td><td>(.94, .99)</td><td>.008</td><td></td><td></td><td></td></tr><tr><td>Respiratory rate per minute, median (IQR)</td><td>22.0</td><td>(20.0-25.0)</td><td>22.0</td><td>(20.0-28.0)</td><td>0.96</td><td>(.93, .98)</td><td>.001</td><td></td><td></td><td></td></tr><tr><td>Diarrhea</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>33</td><td>(89.2)</td><td>3290</td><td>(91.8)</td><td>Ref.</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>4</td><td>(10.8)</td><td>292</td><td>(8.2)</td><td>1.37</td><td>(.48, 3.88)</td><td>.559</td><td></td><td></td><td></td></tr><tr><td>History of fever days, median (IQR)</td><td>3.0</td><td>(2.0-4.0)</td><td>2.0</td><td>(2.0-4.0)</td><td>1.00</td><td>(.97, 1.04)</td><td>.914</td><td></td><td></td><td></td></tr><tr><td>Cough</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>108</td><td>(68.8)</td><td>4847</td><td>(64.7)</td><td>Ref.</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>49</td><td>(31.2)</td><td>2639</td><td>(35.3)</td><td>0.83</td><td>(.59, 1.17)</td><td>.294</td><td></td><td></td><td></td></tr><tr><td>Vomit</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>128</td><td>(81.5)</td><td>6372</td><td>(85.1)</td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>29</td><td>(18.5)</td><td>1115</td><td>(14.9)</td><td>1.29</td><td>(.86, 1.95)</td><td>.215</td><td></td><td></td><td></td></tr><tr><td>Headache</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>31</td><td>(21.4)</td><td>1765</td><td>(33.9)</td><td>Ref.</td><td></td><td></td><td>Ref.</td><td></td><td></td></tr><tr><td>Yes</td><td>114</td><td>(78.6)</td><td>3440</td><td>(66.1)</td><td>1.89</td><td>(1.26, 2.82)</td><td>.002</td><td>2.20</td><td>(1.79, 2.70)</td><td>&lt;.001</td></tr><tr><td>Musculoskeletal pain</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>59</td><td>(49.2)</td><td>1995</td><td>(51.4)</td><td>Ref.</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>61</td><td>(50.8)</td><td>1890</td><td>(48.6)</td><td>1.09</td><td>(.76, 1.57)</td><td>.637</td><td></td><td></td><td></td></tr><tr><td>Abdominal pain</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>118</td><td>(75.6)</td><td>5588</td><td>(76.6)</td><td>Ref.</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>38</td><td>(24.4)</td><td>1708</td><td>(23.4)</td><td>1.05</td><td>(.73, 1.52)</td><td>.782</td><td></td><td></td><td></td></tr><tr><td>Rash or skin lesions</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>144</td><td>(91.7)</td><td>7072</td><td>(94.0)</td><td>Ref.</td><td></td><td></td><td>Ref.</td><td></td><td></td></tr><tr><td>Yes</td><td>13</td><td>(8.2)</td><td>449</td><td>(6.0)</td><td>1.42</td><td>(.80, 2.53)</td><td>.231</td><td>1.45</td><td>(1.16, 1.82)</td><td>.001</td></tr><tr><td>Skin blanching</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>13</td><td>(100.0)</td><td>430</td><td>(96.6)</td><td>Ref.</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Yes</td><td>0</td><td>(0.0)</td><td>15</td><td>(3.4)</td><td>1.00</td><td></td><td></td><td></td><td></td><td></td></tr></tbody></table>
<table><caption>Table 3. Continued</caption><thead><tr><th rowspan="2"></th><th colspan="2">Confirmed or Supportive N = 157</th><th colspan="2">Neither N = 7551</th><th colspan="3">Unadjusted Model</th><th colspan="3">Adjusted Model**</th></tr><tr><th>n</th><th>(%)</th><th>n</th><th>(%)</th><th>OR</th><th>95% CI</th><th>P-value</th><th>aOR</th><th>95% CI</th><th>P-value</th></tr></thead><tbody><tr><td>Conjunctivitis</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>153</td><td>(97.5)</td><td>7462</td><td>(99.2)</td><td>Ref.</td><td>...</td><td>.019</td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>4</td><td>(2.5)</td><td>57</td><td>(0.8)</td><td>3.42</td><td>(1.23, 9.55)</td><td></td><td>3.33</td><td>(2.55, 4.36)</td><td>&lt;.001</td></tr><tr><td>Jaundice</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>147</td><td>(93.6)</td><td>7329</td><td>(97.4)</td><td>Ref.</td><td>...</td><td>.005</td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>10</td><td>(6.4)</td><td>195</td><td>(2.6)</td><td>2.56</td><td>(1.33, 4.93)</td><td></td><td>1.75</td><td>(1.39, 2.19)</td><td>&lt;.001</td></tr><tr><td>Pediatric Early Warning Score, median (IQR)</td><td>14.0</td><td>(10.0-20.0)</td><td>11.0</td><td>(7.0-18.0)</td><td>1.01</td><td>(.98, 1.05)</td><td>.512</td><td></td><td></td><td></td></tr><tr><td>Universal Vital Assessment Score, median (IQR)</td><td>0.0</td><td>(0.0-0.0)</td><td>0.0</td><td>(0.0-2.0)</td><td>0.65</td><td>(.52, .81)</td><td>&lt;.001</td><td></td><td></td><td></td></tr><tr><td>Logistic Organ Dysfunction Score, median (IQR)</td><td>0.0</td><td>(0.0-0.0)</td><td>0.0</td><td>(0.0-0.0)</td><td>0.53</td><td>(.13, 2.08)</td><td>.361</td><td></td><td></td><td></td></tr><tr><td>Antimalarials administered</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>153</td><td>(98.1)</td><td>6463</td><td>(86.8)</td><td>Ref.</td><td>...</td><td>.001</td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>3</td><td>(1.9)</td><td>980</td><td>(13.2)</td><td>0.13</td><td>(.04, .41)</td><td></td><td>0.10</td><td>(.02, .44)</td><td>.002</td></tr><tr><td>Any antimicrobial administered</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>10</td><td>(76.9)</td><td>751</td><td>(88.2)</td><td>Ref.</td><td>...</td><td>.223</td><td></td><td></td><td>...</td></tr><tr><td>Yes</td><td>3</td><td>(23.1)</td><td>100</td><td>(11.8)</td><td>2.25</td><td>(.61, 8.32)</td><td></td><td></td><td></td><td>...</td></tr><tr><td>Died by day 28<sup>a</sup></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>136</td><td>(88.9)</td><td>6293</td><td>(85.8)</td><td>Ref.</td><td>...</td><td>.761</td><td></td><td></td><td>...</td></tr><tr><td>Yes</td><td>5</td><td>(3.3)</td><td>266</td><td>(3.6)</td><td>0.87</td><td>(.35, 2.14)</td><td></td><td></td><td></td><td>...</td></tr><tr><td>Lost to follow-up<sup>b</sup></td><td>12</td><td>(7.8)</td><td>779</td><td>(10.6)</td><td>0.71</td><td>(.39, 1.29)</td><td>.265</td><td></td><td></td><td>...</td></tr><tr><td>Died in hospital<sup>c</sup></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>149</td><td>(94.9)</td><td>7279</td><td>(96.4)</td><td>Ref.</td><td>...</td><td>.997</td><td></td><td></td><td>...</td></tr><tr><td>Yes</td><td>2</td><td>(1.3)</td><td>98</td><td>(1.3)</td><td>1.00</td><td>(.24, 4.08)</td><td></td><td></td><td></td><td>...</td></tr><tr><td>Referred<sup>d</sup></td><td>6</td><td>(3.8)</td><td>175</td><td>(2.3)</td><td>1.67</td><td>(.73, 3.84)</td><td>.223</td><td></td><td></td><td>...</td></tr></tbody></table>

Notes. FIEBRE enrollment criteria excluded some potential participants with diarrhea and lower respiratory infection syndromes, as described in the Methods section. Multivariable models were adjusted for age and sex, regardless of significance in the univariable regression models. Pediatric Early Warning Score, Universal Vital Assessment score, and Logistic Organ Dysfunction Score are not available for children and adults only, respectively.

Abbreviations: OR, odds ratio; aOR, adjusted odds ratio; 95% CI, 95% confidence interval; Ref., referent; IQR, interquartile range.

\*\*Multivariable models were adjusted for age and sex, regardless of significance in the univariable regression models.

<sup>a</sup>Any death within 28 days of enrollment.

<sup>b</sup>Could not be traced at day 28 after enrollment.

<sup>c</sup>Any death among an inpatient participant occurring during the enrollment admission.

<sup>d</sup>In-hospital death status among inpatient participants could not be ascertained due to transfer to a nonstudy healthcare facility.
<table><caption>Table 4. Exposures Among Participants With and Without Confirmed Leptospirosis or Supportive Evidence of Leptospirosis by Bivariable and Logistic Regression, Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Study, 2018–2021</caption><thead><tr><th rowspan="2"></th><th colspan="2">Confirmed Or<br>Supportive N = 157</th><th colspan="2">Neither N = 7551</th><th colspan="3">Unadjusted Model</th><th colspan="3">Adjusted Model**</th></tr><tr><th>n</th><th>(%)</th><th>n</th><th>(%)</th><th>OR</th><th>95% CI</th><th>P-value</th><th>aOR</th><th>95% CI</th><th>P-value</th></tr></thead><tbody><tr><td>Sex</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Female</td><td>75</td><td>(47.8)</td><td>3993</td><td>(52.9)</td><td>Ref.</td><td>...</td><td></td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Male</td><td>82</td><td>(52.2)</td><td>3558</td><td>(47.1)</td><td>1.23</td><td>(.89, 1.68)</td><td>.205</td><td>1.09</td><td>(.61, 1.94)</td><td>.772</td></tr><tr><td>Age years, median (IQR)</td><td>26.5</td><td>(17.0-43.0)</td><td>18.0</td><td>(4.7-33.0)</td><td>1.02</td><td>(1.01, 1.03)</td><td>&lt;.001</td><td>1.02</td><td>(1.01, 1.02)</td><td>&lt;.001</td></tr><tr><td>Lived or worked in close contact with cattle in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>137</td><td>(87.8)</td><td>7048</td><td>(94.1)</td><td>...</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>19</td><td>(12.2)</td><td>439</td><td>(5.9)</td><td>2.23</td><td>(1.36, 3.63)</td><td>.001</td><td>...</td><td>...</td><td>...</td></tr><tr><td>Lived or worked in close contact with goats in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>153</td><td>(97.5)</td><td>7105</td><td>(94.8)</td><td>Ref.</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>4</td><td>(2.5)</td><td>391</td><td>(5.2)</td><td>0.48</td><td>(.18, 1.29)</td><td>.144</td><td>...</td><td>...</td><td>...</td></tr><tr><td>Lived or worked in close contact with pigs in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>149</td><td>(94.9)</td><td>7291</td><td>(97.2)</td><td>...</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>8</td><td>(5.1)</td><td>207</td><td>(2.8)</td><td>1.89</td><td>(.92, 3.90)</td><td>.085</td><td>...</td><td>...</td><td>...</td></tr><tr><td>Lived or worked in close contact with cattle, goats, or pigs in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>132</td><td>(84.6)</td><td>6710</td><td>(89.7)</td><td>Ref.</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>24</td><td>(15.4)</td><td>769</td><td>(10.3)</td><td>1.59</td><td>(1.02, 2.47)</td><td>.040</td><td>...</td><td>...</td><td>...</td></tr><tr><td>Waded, swum or bathed in pond water, lake water, or stream water in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>113</td><td>(72.0)</td><td>6822</td><td>(91.1)</td><td>Ref.</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>44</td><td>(28.0)</td><td>667</td><td>(8.9)</td><td>3.98</td><td>(2.79, 5.69)</td><td>&lt;.001</td><td>...</td><td>...</td><td>...</td></tr><tr><td>Worked in rice fields in the past month</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>96</td><td>(61.1)</td><td>6903</td><td>(92.2)</td><td>Ref.</td><td>...</td><td></td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>61</td><td>(38.9)</td><td>580</td><td>(7.8)</td><td>7.56</td><td>(5.43, 10.54)</td><td>&lt;.001</td><td>6.24</td><td>(3.25, 11.98)</td><td>&lt;.001</td></tr><tr><td>Source of drinking water in the dry season: river</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>154</td><td>(98.1)</td><td>7489</td><td>(99.7)</td><td>Ref.</td><td>...</td><td></td><td>Ref.</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>3</td><td>(1.9)</td><td>21</td><td>(0.3)</td><td>6.95</td><td>(2.05, 23.54)</td><td>.002</td><td>5.11</td><td>(2.00, 13.09)</td><td>.001</td></tr><tr><td>Source of drinking water in the dry season: pond</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>157</td><td>(100.0)</td><td>7508</td><td>(100.0)</td><td>Ref.</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>0</td><td>(0.0)</td><td>2</td><td>(&lt;0.1)</td><td>1.00</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Source of drinking water in the dry season: river or pond</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>No</td><td>154</td><td>(98.1)</td><td>7487</td><td>(99.7)</td><td>Ref.</td><td>...</td><td></td><td>...</td><td>...</td><td>...</td></tr><tr><td>Yes</td><td>3</td><td>(1.9)</td><td>23</td><td>(0.3)</td><td>6.34</td><td>(1.88, 21.34)</td><td>.003</td><td>...</td><td>...</td><td>...</td></tr></tbody></table>

Notes. Multivariable models were adjusted for age and sex regardless of significance in the univariable regression models.

Abbreviations: OR, odds ratio; aOR, adjusted odds ratio; 95% CI, 95% confidence interval; Ref., referent; IQR, interquartile range.

**Multivariable models were adjusted for age and sex, regardless of significance in the univariable regression models.

$P=.019$), the median (IQR) Universal Vital Assessment score among adults was 0.0 (0.0-0.0) versus 0 (0.0-2.0) (OR per unit 0.65, $P<.001$), jaundice was seen in 10 (6.3%) versus 195 (2.6%) (OR 2.56, $P=.005$), and antimalarials were administered in 3 (1.9%) versus 980 (13.2%) (OR 0.13, $P=.001$). A multivariable analysis found headache (adjusted OR [aOR] 2.20, $P<.001$), rash or skin lesions (aOR 1.45, $P=.001$), conjunctivitis (aOR 3.33, $P<.001$), and jaundice (aOR 1.75, $P<.001$) remained associated with confirmed or supportive evidence of leptospirosis, whereas administration of antimalarials

was associated with reduced odds of confirmed or supportive evidence of leptospirosis (aOR 0.10, $P=.002$). Of those with confirmed or supportive evidence of leptospirosis, 2 (1.3%) of 151 with vital outcome data died in hospital.

## Exposures Associated With Leptospirosis

Exposures reported by participants with and without confirmed or supportive evidence of leptospirosis are shown in Table 4. On bivariable analysis of participants with confirmed or supportive evidence of leptospirosis compared with those
<table><caption>**Table 5.** Predominant Reactive Serogroup and Potential Regional *Leptospira* Reservoirs Among Participants With Serologically Confirmed Leptospirosis Based on ≥4-Fold Rise in Titer Between Acute and Convalescent Serum, Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Study, 2018–2021</caption><thead><tr><th>Site</th><th>Serogroup</th><th>N Positive</th><th>Acute Titer Range</th><th>Convalescent Titer Range</th><th>Median Titer</th><th>Potential Regional Reservoirs<sup>a</sup></th></tr></thead><tbody><tr><td rowspan="10">Laos</td><td>Australis</td><td>19</td><td>0–200</td><td>200–12 800</td><td>400</td><td>Cattle, rat, dog, pig, horse, nutria, toad</td></tr><tr><td>Sejroe</td><td>17</td><td>0–200</td><td>200–3200</td><td>400</td><td>Cattle, dog, rat, vole</td></tr><tr><td>Bataviae</td><td>14</td><td>0–200</td><td>200–1600</td><td>400</td><td>Rat, frog, cat</td></tr><tr><td>Icterohemorrhagiae</td><td>12</td><td>0–200</td><td>200–6400</td><td>800</td><td>Rat, cattle, dog, pig, vole, civet, raccoon, mouse</td></tr><tr><td>Grippotyphosa</td><td>9</td><td>0–0</td><td>200–1600</td><td>200</td><td>Rat, frog, cattle, dog, flying squirrel, goat, mouse, hedgehog, sheep, tick from cattle</td></tr><tr><td>Autumnalis</td><td>3</td><td>0–0</td><td>800–1600</td><td>1600</td><td>Rat, cattle, dog, toad, hedgehog, pig</td></tr><tr><td>Celledoni</td><td>3</td><td>0–0</td><td>400–12 800</td><td>800</td><td>Rat</td></tr><tr><td>Pomona</td><td>2</td><td>0–0</td><td>400–800</td><td>600</td><td>Civet, cattle, mouse, goose, cat, dog, pig, rat, squirrel, fox</td></tr><tr><td>Hebdomadis</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Cattle, mouse, rat, raccoon, vole</td></tr><tr><td rowspan="6">Malawi</td><td>Icterohemorrhagiae</td><td>5</td><td>0–0</td><td>200–6400</td><td>200</td><td>Rat, cattle, goose, fox</td></tr><tr><td>Sejroe</td><td>2</td><td>0–0</td><td>400–6400</td><td>3400</td><td>Rat, cattle</td></tr><tr><td>Australis</td><td>1</td><td>0–0</td><td>6400</td><td>6400</td><td>Rat, cattle</td></tr><tr><td>Mini</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>None</td></tr><tr><td>Pyrogenes</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Rat, cattle</td></tr><tr><td>Mozambique</td><td>Australis</td><td>2</td><td>0–0</td><td>200–200</td><td>200</td><td>Rat, cattle</td></tr><tr><td rowspan="3">Zimbabwe</td><td>Autumnalis</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Rat</td></tr><tr><td>Icterohemorrhagiae</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Rat, cattle, goose, fox</td></tr><tr><td>Ballum</td><td>6</td><td>0–0</td><td>200–6400</td><td>300</td><td>Rat, mouse</td></tr><tr><td rowspan="3">Zimbabwe</td><td>Icterohemorrhagiae</td><td>3</td><td>0–0</td><td>200–800</td><td>400</td><td>Rat, cattle, goose, fox</td></tr><tr><td>Pomona</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Cattle</td></tr><tr><td>Pyrogenes</td><td>1</td><td>0–0</td><td>200</td><td>200</td><td>Rat, cattle</td></tr></tbody></table>

without the median (IQR) age was 26.5 (17.0, 43.0) versus 18.0 (4.7, 33.0) years (OR 1.02 per year, *P* < .001), living or working in close contact with cattle in the past month was reported by 19 (12.2%) versus 439 (5.9%) (OR 2.23, *P* = .001), living or working in close contact with cattle, goats, or pigs in the past month by 24 (15.4%) versus 769 (10.3%) (OR 1.59, *P* = .040), wading, swimming, or bathing in pond water, lake water, or steam water in the past month by 44 (28.0%) versus 667 (8.9%) (OR 3.98, *P* < .001), working in rice fields in the past month by 61 (38.9%) versus 580 (7.8%) (OR 7.56, *P* < .001), a river as the source of drinking water in the dry season by 3 (1.9%) versus 21 (0.3%) (OR 6.95, *P* = .002), and a river or pond the source of drinking water in the dry season by 3 (1.9%) versus 23 (0.3%) (OR 6.34, *P* = .003), respectively. On multivariable analysis, working in rice fields in the past month (aOR 6.24, *P* < .001) and having a river as the source of drinking water in the dry season (aOR 5.11, *P* = .001) remained associated with confirmed or supportive evidence of leptospirosis.

Of participants at African sites, 4 (8.5%) of 47 with confirmed or supportive evidence of leptospirosis and 80 (1.4%) of 5673 without worked in rice fields. On multivariable analysis of exposures at African sites, working in rice fields in the past month (aOR 9.10, *P* < .001) was associated with confirmed or supportive evidence of leptospirosis, and living and working

in close contact with cattle in the past month (aOR 1.56, *P* = .309) was not. In contrast, among participants at the Asian site, 47 (53.4%) of 88 with confirmed or supportive evidence of leptospirosis and 500 (27.6%) of 1809 without worked in rice fields. On multivariable analysis of exposures at the Laos site, working in rice fields in the past month (aOR 2.94, *P* = .001) and having a river as the source of drinking water in the dry season (aOR 9.25, *P* = .009) were associated with confirmed or supportive evidence of leptospirosis.

### Predominant Reactive *Leptospira* Serogroups, Species, and Potential Host Range

Table 5 shows the predominant reactive *Leptospira* serogroup among participants with serologically confirmed leptospirosis based on ≥4-fold rise in titer between acute and convalescent serum by study site. In Laos, participants with serologically confirmed leptospirosis were predominantly reactive to *Leptospira* serogroups Australis, Sejroe, Bataviae, Icterohemorrhagiae, Grippotyphosa, Autumnalis, Celledoni, Pomona, and Hebdomadis in descending order of prevalence. Strains representing these serogroups have been isolated from a wide range of host species in Asia, including but not limited to rodents, ruminant livestock, dogs, and pigs [21, 22]. In Malawi, participants with

<footnote><sup>a</sup>References by host are available in Dr Leopold Kirschner's database [21, 22].</footnote>
serologically confirmed leptospirosis were predominantly re-
active to *Leptospira* serogroups Icterohemorrhagiae, Sejroe,
Australis, Mini, and Pyrogenes; in Mozambique to
*Leptospira* serogroups Australis, Autumnalis, and
Icterohemorrhagiae; and in Zimbabwe to *Leptospira* se-
rogroups Ballum, Icterohemorrhagiae, Pomona, and
Pyrogenes. Strains representing these serogroups have been
predominantly isolated from rats and cattle in Africa
(Table 5) [21, 22]. Among the identified *Leptospira* species
*Leptospira borgpetersenii*, *Leptospira interrogans*, and
*Leptospira kirschneri*, variation was observed between study
sites, also consistent with variation in host species (Table 1).

DISCUSSION

Using a standardized study design and laboratory strategy, we
showed that leptospirosis was a common cause of febrile illness
at a site in Laos, and was present but less common at sites in
Malawi, Mozambique, and Zimbabwe. In terms of clinical fea-
tures, headache, rash or skin lesions, conjunctivitis, and jaun-
dice were associated with increased odds of confirmed or
supportive evidence of leptospirosis. Exposures, including
working in rice fields in the past month and having a river as
the source of drinking water in the dry season, were associated
with increased odds of confirmed or supportive evidence of lep-
tospirosis. *L. interrogans*, *L. kirschneri*, and *L. borgpetersenii*
were confirmed in participants with positive *lfb1* PCR. A wider
diversity of *Leptospira* predominant reactive serogroups and
potential host species was observed in Laos compared with in
African sites. Rats and cattle feature as common potential hosts
across all study sites.

Leptospirosis often presents as a nonspecific febrile illness
that is difficult to distinguish from other causes of fever by clin-
ical history and physical examination alone [5]. We found that
headache, rash or skin lesions, conjunctivitis, and jaundice, all
recognized clinical features of leptospirosis, were associated
with increased odds of the disease. However, while these fea-
tures support a diagnosis of leptospirosis, none is sufficiently
sensitive nor specific to be reliable for leptospirosis diagnosis
alone. This highlights the need for wider access to reference di-
agnostics for leptospirosis, and better still, for developing
point-of-care tests of sufficient accuracy to be useful for patient
care [23, 24].

Working in rice fields in the past month and measures of
surface water exposure were associated with increased odds
of leptospirosis across FIEBRE study sites. Notably, an analysis
of risk factors for leptospirosis restricted to African sites con-
firmed the role of rice field exposure at those sites but found
no significant association with living and working in close con-
tact with cattle despite other African studies identifying an im-
portant role for cattle exposure in leptospirosis risk [8, 10].
While rice field exposure was considerably less common among

participants at African sites compared with the Asian site, this
finding highlights the potential impact on leptospirosis risk of
increases in irrigated rice farming in Africa [25]. The lack of as-
sociation of leptospirosis with cattle exposure in African sites
highlights the diverse ecology of the disease by location, even
within the same region.

The diversity of *Leptospira* predominant reactive serogroups was higher in Laos than in African sites, possibly in part because the higher number of participants with serologically confirmed disease with serologically confirmed disease in Laos. Additionally, the Laos MAT panel was supplemented with local *Leptospira* strains, whereas we had limited ability to enrich the African MAT panel with local strains since relatively very few *Leptospira* strains have been isolated from African countries [21, 22]. This was in turn associated with a wider range of potential host species in Laos than in the African sites. While using predominant patterns of *Leptospira* serogroup reactivity provides only a crude insight into potential animal hosts of infecting strains [26], the findings generate hypotheses for future studies in these locations to investigate specific animal contacts as risk factors for human infection.

Our study had a number of limitations. MAT was only done
after screening of acute and convalescent serum by L. fainei se-
rovar Hurstbridge IgM ELISA, the sensitivity of which is uncer-
tain for the diagnosis of leptospirosis particularly in Africa.
Since the case definition for confirmed leptospirosis relied in
part on the availability of a convalescent serum sample for
MAT, it is likely that the case fatality ratio of leptospirosis
was underestimated among those who died before a convales-
cent sample was collected. PCR to lfb1 detected a larger propor-
tion of confirmed leptospirosis infections relative to MAT
among participants enrolled at African sites relative to the
Asian site. This may reflect unmeasured differences in pre-
enrollment antimicrobial use in Laos compared with the
African sites, or perhaps the African MAT panel did not suffi-
ciently represent the diversity of *Leptospira* strains circulating
at the African sites compared with the Asian site. More work
is needed to increase available isolates of pathogenic and inter-
mediate pathogenic *Leptospira* from African countries to im-
prove the performance of serologic diagnosis and gain a
more complete picture of predominant reactive serogroup di-
versity. While our risk factor questionnaire included questions
relevant to leptospirosis, it was not comprehensive and was in-
sufficient to gain a nuanced picture of leptospirosis risk factors
by site. The use as controls of participants presenting to health-
care facilities rather than those from the community may have
introduced bias. Since leptospirosis was less common in
Malawi, Mozambique, and Zimbabwe than Laos, our power
to detect risk factors at African sites was reduced.

We confirm that leptospirosis is an important cause of fever
in Laos and was present but less common at all 3 African sites
studied. Since a specific diagnosis of leptospirosis cannot be
made by clinical means alone, ongoing efforts to develop accu-
rate diagnostic tests suitable for use in low-resource settings are
needed. Rice field exposure is a major risk factor for infection,
even in Africa, where rice field exposure was not as common,
highlighting the need for focused prevention in rice field work-
ers, especially as irrigated rice farming expands in Africa. We
highlight the need to redouble efforts to isolate *Leptospira*
strains in Africa to improve serologic diagnosis and identify
hypotheses for more detailed risk factor studies.

**Supplementary Data**

Supplementary materials are available at *The Journal of Infectious Diseases* online (http://jid.oxfordjournals.org/). Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author.

Notes

**Acknowledgments.** The authors thank study team members for their work on the FIEBRE study. They thank Victoria Gould for managing sample aliquoting and shipment to reference laboratories and Michael Lawrence for administrative support of the FIEBRE study at the London School of Hygiene and Tropical Medicine. They thank Amandine Houvert, Vallier Sordoillet, Farida Zinini, Céline Lorioux, Jean-Francois Mariet, and Farah Martin, Unité Biologie des Spirochètes, French National Reference Center for Leptospirosis, WHO Collaborating Center for Reference and Research on Leptospirosis, Institut Pasteur. The study was presented in part at the 71st American Society of Tropical Medicine and Hygiene annual meeting, Seattle, Washington, 30 October to 3 November 2022 (Abstract LB-5320).

**Author contributions.** H. H., J. A. C., M. M., P. N. N., D. C. W. M., and other FIEBRE Consortium members designed the parent study, and H. H. and D. C. W. M. provided overall coordination. P. N. N., M. M., E. W. G., M. C., J. M. B., S. A. A., I. D. O., and H. H. were involved in study activities at sites. J. A. C. conceived the analysis for the present paper. M. P. undertook leptospirosis diagnostic testing. H. H. and S. L. participated in data cleaning and management. J. A. C. and P. M. accessed and verified the underlying data. P. M. performed statistical analyses. J. A. C. wrote the first draft of the manuscript. All authors had full access to all data, reviewed subsequent drafts, and agreed to submission of the final manuscript.

**Data availability.** The de-identified dataset, along with the
corresponding data dictionary that defines each field in the
set, will be freely available online with no restrictions once all
protocol objectives are completed via the London School of

Hygiene and Tropical Medicine's Compass and can be accessed
and downloaded at https://datacompass.lshtm.ac.uk/.

**Febrile Illness Evaluation in a Broad Range of Endemicities (FIEBRE) Consortium.** Benjamin Amos, PhD, Elizabeth A. Ashley, MB BS, PhD, Oliver Baerenbold, PhD, Stéphanie Baghoumina, Núria Balanza, MSc, Tsitsi Bandason, MSc, Quique Bassat, MD, PhD, Tapan Bhattacharyya, PhD, Stuart D. Blacksell, PhD, Zumilda Boca, Christian Bottomley, PhD, John Bradley, PhD, Clare I. R. Chandler, PhD, Vilada Chansamouth, Joseph Chipanga, BSc, Anelsio Cossa, Ethel Dauya, Catherine Davis, Justin Dixon, PhD, Somyoth Douangphachanh, Audrey Dubot-Pérès, PhD, Michelle M. Durkin, Nicholas A. Feasey, PhD, Rashida A. Ferrand, MB BS, PhD, Colin Fink, Elizabeth J. A. Fitchett, MRCPCH, Alessandro Gerada, MD MRCP FRCPath, Stephen R. Graves, MB BS, PhD, Becca L. Handley, Coll D. Hutchison, Risara Jaksuwan, Jessica Jervis, Jayne Jones, Kevin C. Kain, MD, Suzanne H. Keddie, PhD, Khamxeng Khounpaseuth, Katharina Kranzer, PhD, Khamfong Kunlaya, Pankaj Lal, Xavier de Lamballerie, David G. Lalloo, FRCP, Manophab Luangraj, MD, Yoel Lubell, PhD, Eleanor MacPherson, PhD, Sengchanh Manichan, Tegwen Marlais, PhD, Florian Maurer, MD, Michael Miles, Campos Mucasse, Chelsea Nguyen, Vilayouth Phimolsarnnousith, Chrissy h. Roberts, PhD, Amphone Sengduangphachanh, Siho Sengsavang, Molly Sibanda, Somvai Singha, John Stenos, Ampai Tanganuchitcharnchai, MSc, Hira Tanvir, James E. Ussher, MB ChB, PhD, Marta Valente, Marie A. Voice, PhD, Manivanh Vongsouvath, Msopole Wamaka, L. Joseph Wheat, MD, Shunmay Yeung, MB BS, PhD.

**Financial support.** The Febrile Illness Evaluation in a Broad Range of Endemicities study is funded by the UK government; the views expressed, however, do not necessarily reflect the UK government's official policies. P. N. N. is supported by the Wellcome Trust. For the purpose of Open Access, the author has applied a CC BY public copyright license to any Author Accepted Manuscript version arising from this submission. The funder of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the manuscript. I. D. O. received support through a Wellcome Trust Clinical PhD Programme awarded to the London School of Hygiene and Tropical Medicine (grant number 203905/Z/16/Z).

**Potential conflicts of interest.** All authors: No reported conflicts.

All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.

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