Pathogenic Leptospira in dogs and rodents in Tha Wang Pha, Thailand - Prevalence, diversity and linked environments

This study reveals that free-roaming dogs and diverse rodent species in rural Tha Wang Pha, Thailand, serve as reservoirs for multiple pathogenic *Leptospira* species, including the first confirmed detection of *L. weilii* in Thai rodents, highlighting a complex transmission cycle that bridges animal habitats and human settlements and underscores the need for integrated One Health surveillance.

The Gist

Imagine a hidden game of "tag" happening in the rural villages of Northern Thailand. The tagger is a tiny, invisible bacteria called Leptospira, which causes a serious illness called leptospirosis. Usually, we think of this disease as something humans catch from dirty water or floodwaters. But this study asks a crucial question: Who is actually passing the tag around?

The researchers went into the villages of Nan Province to find the "players" in this game. They looked at two main groups: dogs (the neighborhood pets that roam free) and small wild mammals (like rats, mice, and hedgehog-like creatures called gymnures).

Here is the story of what they found, broken down simply:

1. The Suspects: A Crowd of Tiny Carriers

The team caught and tested nearly 400 small wild animals. They found that about 10% of them were carrying the bacteria in their kidneys.

• The "Super-Spreaders": It wasn't just one type of rat. They found the bacteria in 12 different species of small mammals. Some were common house rats, others were wild forest rats, and even a few strange-looking hedgehog-like animals.
• The Weight Factor: They noticed something interesting: the heavier (and likely older) the animal was, the more likely it was to be infected. Think of it like a library book that has been checked out many times; the longer an animal lives, the more chances it has to catch the bacteria and keep it.

2. The Bridge: The Free-Roaming Dog

This is where the story gets really important. The researchers found that 8.4% of the local dogs also had the bacteria in their blood.

• The "Bridge" Metaphor: Imagine the wild animals live in the forest and the rice fields, while humans live in the village houses. The dogs are the bridge. They run back and forth between the wild woods (where the infected rats live) and the human homes.
• Because these dogs roam freely, they can pick up the bacteria from the wild animals and bring it right to your doorstep, potentially passing it to humans. They are the "middleman" in the transmission chain.

3. The New Discovery: A New "Player" in the Game

The scientists didn't just count the infections; they looked at the type of bacteria. They found three different "flavors" (species) of Leptospira:

1. L. interrogans: A very common type found in many places.
2. L. borgpetersenii: Another common type.
3. L. weilii: This is the big news. This is the first time scientists have ever confirmed this specific type of bacteria in wild rodents in Thailand. Before this, we only knew it from farm animals (like cows and pigs) or dogs. Finding it in wild rats means the "menu" of bacteria in Thailand is more diverse than we thought.

4. The Environment: Where the Game Happens

You might think the bacteria is only found in muddy flood zones. The study found it in forests, rice fields, and upland farms.

• Surprisingly, they did not find the bacteria in the small wild animals living inside human houses. The wild rats in the houses were mostly clean.
• However, the bacteria was everywhere else: in the forests and the agricultural lands. This suggests that the "danger zone" isn't just the village center, but the edges where nature meets farming.

The Big Picture: Why Should We Care?

Think of the disease cycle like a relay race.

• The Wild Animals are the first runners, holding the baton (the bacteria) in the forest.
• The Dogs are the second runners. They grab the baton from the wild animals and run it into the village.
• The Humans are the final runners, getting tagged when they touch the dogs or the contaminated environment.

The Takeaway:
To stop the race (and the disease), we can't just look at humans. We need a "One Health" approach, which means treating human health, animal health, and the environment as one connected team.

• Vaccinating dogs could be a powerful way to cut the bridge between the wild and the village.
• Watching the wild animals helps us know where the "hotspots" are.
• Understanding the new bacteria types helps doctors prepare better treatments.

In short, this study shows that in rural Thailand, the fight against leptospirosis requires watching the rats in the forest, the dogs in the village, and the water they all share. It's a complex puzzle, but now we have more pieces to solve it.

Technical Summary

1. Problem Statement

Leptospirosis remains a significant public health issue in Thailand, with incidence rates rising again after a period of decline. While the disease is zoonotic and transmitted through contact with urine from infected carrier animals, the specific transmission cycles involving different animal reservoirs in rural Thailand are not fully understood. Previous studies have often focused on single species (e.g., only rodents or only dogs) or relied heavily on serology rather than molecular detection. There is a critical gap in understanding the interaction between free-roaming dogs and terrestrial micromammals (rodents and other small mammals) and how these species contribute to the transmission of pathogenic Leptospira to humans in rural settings.

2. Methodology

The study was conducted in the Tha Wang Pha district of Nan Province, Northern Thailand, involving two primary host groups: domestic dogs and terrestrial micromammals.

• Sample Collection:
  • Dogs: 95 serum samples were collected from apparently healthy, free-roaming owned dogs in four villages (V4–V7) between 2019 and 2023.
  • Micromammals: 399 individuals from 13 rodent species and one gymnure (Hylomys suillus) were trapped across eight villages (V1–V8) between 2012 and 2018. Trapping occurred in four distinct landscapes: forests, uplands (non-flooded agricultural), lowlands (flooded agricultural), and human settlements.
• Laboratory Analysis:
  • Detection: DNA was extracted from dog serum and rodent kidney tissues. Pathogenic Leptospira was detected using a probe-specific reverse transcription real-time PCR (RT-qPCR) targeting the rrs (16S) gene.
  • Genotyping: Positive samples underwent conventional PCR amplification of the secY, rrs, and lfb1 genes, followed by Sanger sequencing to identify specific pathogenic species.
  • Phylogenetics: Maximum-likelihood trees were constructed to compare local strains with global and regional sequences from GenBank.
• Statistical Analysis:
  • A Generalized Linear Mixed Model (GLMM) with a binomial error distribution was used to assess the impact of environmental (landscape, season) and host-related (species, sex, standardized body mass) factors on infection status. Random effects accounted for village and year interactions.

3. Key Results

Prevalence and Host Diversity:

• Micromammals: 10.0% (40/399) tested positive for pathogenic Leptospira. Infection was found in 12 different species.
  • Bandicota indica and Rattus exulans were the most numerous infected species (20% of all positive cases each).
  • However, the highest prevalence rates were observed in Berylmys berdmorei (27.3%), Berylmys bowersi (20.0%), Bandicota indica (17.0%), and Hylomys suillus (16.7%).
  • Infected micromammals were found in agricultural (upland/lowland) and forest habitats but were absent in human settlements, except for R. exulans, which dominated that habitat but showed lower infection rates compared to wild habitats.
• Dogs: 8.4% (8/95) of dogs tested positive for Leptospira DNA in their serum.

Genetic Diversity and Novel Findings:

• Sequencing was successful for 11 out of 40 positive micromammal samples, identifying three pathogenic species:
  1. Leptospira interrogans (5 isolates): Found in B. bowersi, B. indica, and H. suillus.
  2. Leptospira borgpetersenii (2 isolates): Found in Mus species.
  3. Leptospira weilii (4 isolates): Found in B. indica and Berylmys spp.
• Novelty: This study reports the first confirmed detection of L. weilii in rodents in Thailand. Previously, L. weilii in Thailand had only been identified in livestock (cattle, pigs) and dogs.
• Phylogenetics: The identified strains clustered closely with sequences from humans, dogs, and rodents in Thailand and Cambodia, suggesting shared transmission cycles.

Risk Factors (GLMM Analysis):

• Body Mass: Standardized body mass was the only significant predictor of infection ($p=0.03$). Heavier individuals (relative to their species average) had a higher probability of infection, suggesting age or body condition as a risk factor.
• Environment: No statistically significant association was found between infection status and landscape type (forest, upland, lowland, settlement), season, or sex, though there was a non-significant trend toward higher prevalence in agricultural lands.
• Spatial Variation: Significant variation was attributed to the "village $\times$ year" interaction, indicating that risk is highly localized and temporal.

4. Key Contributions

1. First Detection of L. weilii in Thai Rodents: The study expands the known host range of L. weilii in Thailand from livestock/dogs to wild micromammals, suggesting a more complex transmission cycle than previously thought.
2. Multi-Species Reservoir Characterization: It confirms that a diverse array of micromammals (not just Rattus species) serve as reservoirs, with specific species like Berylmys showing high infection intensities.
3. The "Bridge Host" Hypothesis: The study provides evidence supporting the role of free-roaming dogs as epidemiological bridges. Dogs frequent both human settlements and wild/agricultural habitats where infected rodents reside, potentially facilitating spillover to humans.
4. Methodological Rigor: By combining RT-qPCR with multi-gene sequencing and robust statistical modeling (GLMM), the study moves beyond simple prevalence surveys to identify specific drivers of infection (e.g., body mass).

5. Significance and Implications

• Public Health: The presence of pathogenic Leptospira in healthy dogs and diverse wild rodents highlights a persistent zoonotic risk in rural Thailand. The finding that L. weilii (often associated with livestock) is present in wild rodents suggests potential cross-species transmission in shared environments like water sources and grazing areas.
• One Health Approach: The results underscore the necessity of integrated surveillance. Controlling leptospirosis requires addressing the interface between wildlife, domestic animals (dogs/livestock), and humans.
• Intervention Strategies:
  • Vaccination: Routine vaccination of free-roaming dogs is suggested as a potential intervention, though efficacy depends on matching local serovars.
  • Surveillance: Future efforts should focus on molecular surveillance across all host types (wildlife, livestock, dogs) to map the complete transmission cycle, particularly for under-explored species like L. kirschneri and L. wolffii.
  • Risk Communication: Communities should be educated about the risks of free-roaming dogs interacting with contaminated environments and the importance of avoiding contact with water sources in high-risk agricultural zones.

In conclusion, the paper reveals a complex, dynamic epidemiological landscape for leptospirosis in Northern Thailand, driven by diverse rodent reservoirs and facilitated by free-roaming dogs acting as transmission bridges to human populations.