Feline leptospirosis prevalence worldwide: A systematic review and meta-analysis of diagnostic approaches

Background and Aim: Leptospirosis in felids (domestic and wild cats) presents an ongoing challenge in our understanding. Numerous studies have reported the detection of Leptospira spp. in these feline populations, highlighting their potential as zoonotic carriers. This systematic review and meta-analysis aimed to provide insight into the global prevalence of leptospirosis in domestic and wild cats. Materials and Methods: We conducted extensive searches across five databases (PubMed, Scopus, Web of Science, Science Direct, and Google Scholar) following the Preferred Reporting Items for Systematic Reviews and Meta-analyses Protocols guidelines. Random-effect meta-analyses were performed using R software version 4.3.0 to estimate pooled prevalence rates. Subgroup meta-analyses were conducted based on continents, diagnostic methods, sample types, and wildcat genera. Results: A total of 71 articles on leptospirosis in domestic cats and 23 articles on leptospirosis in wild cats met the eligibility criteria. Our findings indicated a significantly higher pooled seroprevalence of leptospirosis in domestic cats compared with infection prevalence (9.95% [95% confidence interval (CI), 7.60%–12.54%] vs. 4.62% [95% CI, 2.10%–7.83%], p = 0.01). In contrast, no significant difference was observed in pooled seroprevalence and infection prevalence among wild cats (13.38% [95% CI, 6.25%–21.93%] vs. 2.9% [95% CI, 0.00%-18.91%], p = 0.21). A subgroup meta-analysis of domestic cats revealed significant differences in seroprevalence across continents, sample types, and diagnostic methods. On the contrary, wild cats had no significant differences in any of the subgroups. Conclusion: Leptospira spp. have evidently been exposed to both domestic and wild cats, highlighting their potential roles as reservoir hosts for leptospirosis. These findings highlight the importance of considering felids as a possible public health threat.


Introduction
Leptospirosis is one of the most widespread zoonotic and waterborne diseases on a global scale [1].This infectious ailment affects a broad spectrum of animals, including rats, horses, cows, pigs, dogs, sea lions, and even felids such as cats [2].However, the symptoms of leptospirosis are rarely observed in cats, with clinical manifestations predominantly observed in young cats [2].It has been postulated that cats may show resistance to leptospirosis, particularly in their propensity for rodent predation, even though there is a lower likelihood of developing clinical symptoms of leptospirosis.This suggested resistance is also linked to the acidity level of cat urine, purportedly diminishing the viability of Leptospira spp.[3,4].Although clinical signs of Leptospira spp.exposure are infrequently exhibited in cats, serological evidence of Leptospira spp.exposure has been documented in domestic cats (Felis catus) presenting sub-clinical symptoms and apparently healthy cats [5].This raises concerns about the potential transmission of Leptospira spp.from cats to the surrounding environment [5].However, the precise role of cats in leptospirosis pathogenesis still needs to be further understood [2].Cats may act Copyright: Andityas, et al.Open Access.This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/ by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Available at www.veterinaryworld.org/Vol.17/February-2024/3.pdf as reservoir hosts for leptospirosis [2,3,[6][7][8][9], indefinitely sustaining the circulation of infectious agents in the environment and serving as a transmission source to other animals.Due to the absence of clinical symptoms in many cats, they are also called carrier hosts [2,10,11].
Notably, leptospirosis has been observed not only in domestic cats but also in various wild feline species, such as jaguars (Panthera onca), mountain lions (Puma concolor), and bobcats (Lynx rufus) [12,13].These findings may raise public health concerns, especially as human activities encroached on wildlife habitats, leading to increased interactions between animals and humans, thereby amplifying the potential for pathogen transmission [14].In addition, some wild animals reside in captivity within zoological parks and circuses situated near urban areas, which further increases the risk of human exposure to zoonotic agents, including

Selection criteria
All extracted articles underwent a comprehensive evaluation using the Rayyan-Intelligent Systematic Review platform (www.rayyan.ai) to check for duplication and screen the articles to ensure the integrity of the research process.Two reviewers, MA and DMN within the Rayyan platform, independently selected the manuscript through title and abstract screening.The screening process was performed on the basis of several inclusion criteria, such as a study population consisting of both domestic and wild cats, and the outcomes should specifically report the prevalence of leptospirosis.Case studies, surveys, or cross-sectional studies were eligible for inclusion.However, articles reporting experimental studies, cohort studies, or casecontrol studies were excluded from consideration.In addition to these criteria, articles were included only if they provided detailed information on sample sizes for each feline species, types of samples collected, and diagnostic methods.The precise questions used as screening guidelines have been described in the protocol.

Data extraction
Data were extracted by MA and subsequently reviewed by DMN.Inconsistencies or discrepancies that arose during this process have been carefully resolved through discussion and consensus.For inclusion in the meta-analysis, articles were required to provide comprehensive information regarding sample sizes for each feline species, types of samples collected, and diagnostic methods employed.The extracted data encompassed essential details, including author names, publication years, study locations (both country and continent), animal species involved, sample sizes and types, method of detection, and number of leptospirosis cases identified.All extracted data were collated into pre-designed Microsoft Excel sheets (Microsoft Corp., Redmond, WA, USA).

Quality assessment of individual studies
The quality assessment was performed using the Joanna Briggs Institute Critical Appraisal Checklist for studies reporting prevalence data [25].In response to the specific criteria outlined in the checklist, each study's quality was categorized as "yes," "no," "unclear," or "not applicable."The overall results were subsequently classified into three tiers based on the scores obtained: high (≥7), medium (4-6), and poor (≤3).This rigorous quality assessment process ensured that only studies meeting stringent quality criteria were included in our meta-analysis.

Statistical analysis
Statistical analysis was conducted using the "Meta" and "Metaphor" packages in R 4.3.0software (Comprehensive R Archive Network, Vienna, Austria) [26].The Freeman-Tukey double arcsine transformation was applied to test statistical significance.Effect sizes were evaluated based on pooled prevalence and 95% confidence interval (CI).Heterogeneity was assessed using the I 2 index, Cochrane's Q test, and the corresponding p-value.When moderate or high heterogeneity (I 2 > 50%) was detected, a random-effects model was employed, whereas a fixed-effects model was chosen in the presence of low heterogeneity.
The pooled prevalence analysis yielded seroprevalence and infection prevalence.Seroprevalence indicates indirect detection by measuring the presence of antibodies in cats, whereas infection prevalence indicates direct isolation and detection of Leptospira spp.from samples.In addition, the analysis was stratified to differentiate between domestic and wildcat populations due to lifestyle, environment, and disease exposure.This division allowed for a clearer understanding of the prevalence of leptospirosis.
Subgroup meta-analyses were performed based on continent, sample type, diagnostic method, and infection status.The genus was used for subgroup meta-analysis in the case of wild cats.We performed a meta-regression and cumulative meta-analysis to assess the trends of leptospirosis in domestic and wild cats based on publication years.Sensitivity analysis was also performed to verify the robustness of pooled prevalence in domestic and wild cats using leave-one-out meta-analysis, which indicates the disproportional influence of each study on the results.The Egger's test and funnel plot [27] were used to determine publication bias.The global distribution of leptospirosis in cats was displayed using QGIS v.3.2.0 (https://qgis.org/en/site/).

Literature search
A total of 1387 studies were initially identified for inclusion in this systematic review and meta-analysis.After the initial screening procedure, 124 articles met the eligibility criteria and progressed to further evaluation.Of these, 32 articles were subsequently excluded mainly due to unavailability of full-text access (n = 26) and inappropriate outcome measures (n = 6).As a result, a total of 91 studies were ultimately included in the meta-analysis.
The dataset used for the meta-analysis comprised 13,034 samples collected from domestic cats in 71 studies and 1034 samples collected from wild cats in 23 studies (Tables -1 and 2) [3, 5-13, 16-21, 28-101].Figure-1 illustrates the detailed process of study retrieval, screening, and collating studies on the prevalence of leptospirosis in domestic and wild cats.

Prevalence of leptospirosis in domestic cats: Analysis
The prevalence of leptospirosis in domestic cats was subjected to subgroup analysis based on continent, sample type, and diagnostic method.Significant differences were observed in seroprevalence (p = 0.01) across different continents as well as in seroprevalence between sample types (p < 0.01) and diagnostic methods (p < 0.01).However, there were no significant differences in infection prevalence among continents (p = 0.19).

Analysis of leptospirosis prevalence in wild cats
Prevalence was also analyzed in subgroups of the continent, sample type, diagnostic method, and genus in wild cats.Notably, there were no significant differences in the results of these analyses among the subgroups.

Risk of publication bias and quality assessment of individual studies
Funnel plots for both domestic and wildcats, as illustrated in Figure -4, revealed no significant indications of publication bias.Furthermore, the Egger's test showed no evidence of publication bias between domestic and wildcats (p = 0.08 and p = 0.50, respectively).To assess the risk of bias, 47 studies were found to be of high-quality, whereas 24 studies in the field of domestic cat studies had a moderate quality rating.Wildcat studies comprised 16 high-quality studies and seven studies with moderate-quality ratings (https:// osf.io/b3u8j/).

Discussion
This study emphasized the seroprevalence and infection prevalence of leptospirosis in both domestic and wild cats.Our results reveal that domestic cats exhibited a lower seroprevalence than wildcats (9.95% vs. 13.38%).However, the infection prevalence in domestic cats was higher than in wild cats (4.62% vs. 2.9%).Among wildcats, Puma spp.showed the highest prevalence, followed by Felis spp., Lynx spp., Panthera spp., and Leopardus spp. in descending order.Several risk factors may contribute to variations in the seroprevalence and infection prevalence of feline leptospirosis, including the proximity of cats to human settlements, residence in flood-prone areas, close interaction with other animals, and the presence of rodents as potential vectors [28,29].The prevalence of leptospirosis in domestic cats may vary according to the lifestyle of the cat and geographical location [3,28,30].Domestic cats that have access to the outdoors are susceptible to leptospirosis due to their predilection to prey on rodents, direct engagement with contaminated water sources, and shared habitation with farm animals that serve as potential reservoirs, excreting the leptospiral bacteria in their urine [2].The transmission of Leptospira spp.bacterial infections is linked not only to water sources containing the pathogen but also to the dietary habits of wildcats living in natural environments [102].Bobcats consume various mammalian prey, including rodents, lagomorphs, white-tailed deer (Odocoileus virginianus), and other ungulates [102][103][104].The Eurasian lynx (Lynx lynx) preys on rodents, ungulates, and European roe deer (Capreolus capreolus), with rodents being their alternative food source [105].It is important to acknowledge that rodents are reservoir hosts for leptospirosis [106], which highlights the potential risk of leptospirosis in wild cats.Furthermore, wildcats living in captivity are suspected to have Leptospira spp.infection due to suboptimal hygiene practices [16].
A detailed subgroup analysis at the continental level revealed variations in the 95% CIs for the seroprevalence and infection prevalence of leptospirosis in domestic and wild cats.Notably, our findings underscored that leptospirosis prevalence in Asia is significantly high in both domestic and wildcat populations.This observed pattern of high prevalence reflects human leptospirosis in the Asia-Pacific region, where the disease remains highly endemic with reported incidence rates ranging from 1 to over 10 cases/100,000 individuals [107].Leptospirosis morbidity is estimated to be high in several Asian countries (especially Southeast and South Asia).For example, India reported an estimated incidence rate of 19.7 cases/100,000 population, while Indonesia reported a rate of 39.2 cases/100,000 population [108].
The prevalence of human leptospirosis is most pronounced in tropical countries, mainly due to conducive social and environmental risk factors that facilitate disease transmission.Major outbreaks of leptospirosis are often associated with factors such as flooding, inadequate sanitation, climate change, and the presence of high-maintenance host populations, including domestic and wild animals.These hosts play a pivotal role in the dissemination of diseases within these regions [107,109].As a result, approximately 73% of global leptospirosis cases and fatalities are reported in these tropical regions [108].
The detection of cat leptospirosis can be performed either by direct or indirect methods.Direct methods involve the identification or isolation of Leptospira spp.agent from clinical specimens, whereas indirect methods focus on leptospirosis antibody testing [110].For indirect detection, MAT and serum testing were the most commonly used methods.MAT, in particular, is considered the gold standard for detecting leptospirosis, capable of identifying various serovars of Leptospira spp.[111].Our findings indicate that the cutoff MAT titer values in domestic cats ranged from 1:20 to 1:200, whereas those in wild  cats ranged from 1:50 to 1:100.It should be noted that cats usually respond to leptospirosis, both experimentally and in natural infections, with low antibody titers ranging from 1:30 to 1:400 [31].In view of the fact that leptospirosis vaccines for cats are not currently available, antibodies measured in MAT from cat serum reflect the immune response to Leptospira spp.Infection [32].However, MAT's utility may be limited during the early stages of the disease due to the lack of specific antibodies in the immune system or low antibody titers, which can lead to false negative results [112].In addition, the diagnosis of infection can become more complex if the host has been previously exposed by Verma et al. [113] and Goris and Hartskeerl [114] to different serogroups.As a result, direct methods provide more favorable results in the early stages of the disease.The most widely utilized direct diagnostic method for detecting cat leptospirosis is PCR using urine samples.This method offers distinct advantages for identifying Leptospira spp.during the initial phases of the disease when antibody titers may not be sufficient for indirect methods and during acute leptospirosis.However, PCR using urine samples can be challenging because it is not always practical and often requires catheterization [115].To improve diagnostic accuracy, it is recommended to combine both direct and indirect evidence methods.For example, PCR-enzymelinked immunosorbent assay technique can serve as an alternative to MAT, yielding higher accuracy and more reliable results [116].
Meta-regression, which employs a single data point from each study to estimate prevalence trends [117], and cumulative meta-analysis, which utilizes pooled evidence from studies to track the evolution of evidence over time with new research publications [118] offer valuable insights into disease trends.Our meta-regression analysis revealed a significant decrease in the prevalence of leptospirosis in domestic cats over time, whereas wild cats showed no significant trend change over time.
On the basis of a cumulative meta-analysis, the prevalence of leptospirosis in domestic cats was originally reported at 4.86% in 1957.In 2013, this value increased steadily to 19.18% and then decreased to 7.59% in 2023.On the other hand, leptospirosis in wild cats was first reported in 1988 with a prevalence of 25%, which is gradually approaching 11.19% in 2022.This fluctuating trend in feline leptospirosis over time is similar to trends observed in human leptospirosis [119,120].In humans, an upward trend was noted during 1997-2012, which was linked to environmental disruptions, such as floods and heavy rainfall, facilitating the dispersion of pathogens across broader areas [119].This trend reflects an increase in the incidence of leptospirosis in domestic cats in 2013, potentially reflecting increased exposure to pathogens.
Despite this study's insights, certain limitations warrant recognition.Variations in the number of cat leptospirosis studies between continents raise the possibility of bias in prevalence, particularly in regions with fewer studies, such as Australia and North America.In addition, some categories within subgroups had to be excluded due to a lack of available studies.For this reason, caution should be exercised when interpreting the results of certain subgroup analysis.In line with our primary objective of assessing the overall pooled prevalence of leptospirosis in domestic and wild cats, the variability in serovars was not investigated.We acknowledge the need for further studies to elucidate the specific serovars of Leptospira spp. to enhance our understanding of leptospirosis in cats.

Conclusion
Our study highlights that both domestic and wild cats are susceptible to Leptospira spp.bacterial exposure.Although the seroprevalence and infection prevalence of leptospirosis in these feline populations are relatively low, the risk of transmission of Leptospira spp. to the surrounding environment should still be considered.Therefore, leptospirosis in domestic and wild cats should be regarded as a public health concern in view of its potential for zoonotic transmission.

Figure- 1 :
Figure-1: PRISMA flow diagram of selected articles included in this study.

Figure- 3 :
Figure-3: Scatterplot of meta-regression analysis to assess trends in leptospirosis prevalence over the years in (a) domestic cats and (b) wild cats.a b

Figure- 4 :
Figure-4: Funnel plots of standard error to assess publication bias across prevalence studies in (a) domestic cats and (b) wild cats.a b

Table - 1
: Characteristics of the eligible studies based on the seroprevalence and infection prevalence of leptospirosis in domestic and stray cats.

Table - 2
: Characteristics of the eligible studies based on the seroprevalence and infection prevalence of leptospirosis in wild cats.

Table - 3
: Pooled prevalence and subgroup analyses of leptospirosis domestic and wild cats.