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This review focuses on the most reliable and up-to-date methods for diagnosing trypanosomoses, a group of diseases of wild and domestic mammals, caused by trypanosomes, parasitic zooflagellate protozoans mainly transmitted by insects. In Africa, the Americas and Asia, these diseases, which in some cases affect humans, result in significant illness in animals and cause major economic losses in livestock. A number of pathogens are described in this review, including several Salivarian trypanosomes, such as Trypanosoma brucei sspp. (among which are the agents of sleeping sickness, the human African trypanosomiasis [HAT]), Trypanosoma congolense and Trypanosoma vivax (causing “Nagana” or animal African trypanosomosis [AAT]), Trypanosoma evansi (“Surra”) and Trypanosoma equiperdum (“Dourine”), and Trypanosoma cruzi , a Stercorarian trypanosome, etiological agent of the American trypanosomiasis (Chagas disease). Diagnostic methods for detecting zoonotic trypanosomes causing Chagas disease and HAT in animals, as well as a diagnostic method for detecting animal trypanosomes in humans (the so-called “atypical human infections by animal trypanosomes” [a-HT]), including T. evansi and Trypanosoma lewisi (a rat parasite), are also reviewed. Our goal is to present an integrated view of the various diagnostic methods and techniques, including those for: (i) parasite detection; (ii) DNA detection; and (iii) antibody detection. The discussion covers various other factors that need to be considered, such as the sensitivity and specificity of the various diagnostic methods, critical cross-reactions that may be expected among Trypanosomatidae, additional complementary information, such as clinical observations and epizootiological context, scale of study and logistic and cost constraints. The suitability of examining multiple specimens and samples using several techniques is discussed, as well as risks to technicians, in the context of specific geographical regions and settings. This overview also addresses the challenge of diagnosing mixed infections with different Trypanosoma species and/or kinetoplastid parasites. Improving and strengthening procedures for diagnosing animal trypanosomoses throughout the world will result in a better control of infections and will significantly impact on “One Health,” by advancing and preserving animal, human and environmental health. Graphical Abstract

The emergence of the mobile colistin-resistance genes mcr-1 has attracted significant attention worldwide. This study aimed to investigate the genetic features of mcr-1 -carrying plasmid among carbapenem-resistant Enterobacterales (CRE) isolates and the potential genetic basis governing transmission. Seventeen mcr -harboring isolates were analyzed based on whole genome sequencing using short-read and long-read platforms. All the mcr-1 -carrying isolates could be conjugatively transferred into a recipient Escherichia coli UB1637. Among these 17 isolates, mcr-1 was located on diverse plasmid Inc types, consisting of IncX4 (11/17; 64.7%), IncI2 (4/17; 23.53%), and IncHI/IncN (2/17; 11.76%). Each of these exhibited remarkable similarity in the backbone set that is responsible for plasmid replication, maintenance, and transfer, with differences being in the upstream and downstream regions containing mcr-1 . The IncHI/IncN type also carried other resistance genes ( bla TEM-1B or bla TEM-135 ). The mcr-1 -harboring IncX4 plasmids were carried in E. coli ST410 (7/11; 63.6%) and ST10 (1/11; 9.1%) and Klebsiella pneumoniae ST15 (1/11; 9.1%), ST336 (1/11; 9.1%), and ST340 (1/11; 9.1%). The IncI2-type plasmid was harbored in E. coli ST3052 (1/4; 25%) and ST1287 (1/4; 25%) and in K. pneumoniae ST336 (2/4; 50%), whereas IncHI/IncN were carried in E. coli ST6721 (1/2; 50%) and new ST (1/2; 50%). The diverse promiscuous plasmids may facilitate the spread of mcr-1 among commensal E. coli or K. pneumoniae strains in patients. These results can provide information for a surveillance system and infection control for dynamic tracing.

Despite increasing control measures, numerous parasitic and infectious diseases are emerging, re-emerging or causing recurrent outbreaks particularly in Asia and the Pacific region, a hot spot of both infectious disease emergence and biodiversity at risk. We investigate how biodiversity affects the distribution of infectious diseases and their outbreaks in this region, taking into account socio-economics (population size, GDP, public health expenditure), geography (latitude and nation size), climate (precipitation, temperature) and biodiversity (bird and mammal species richness, forest cover, mammal and bird species at threat). We show, among countries, that the overall richness of infectious diseases is positively correlated with the richness of birds and mammals, but the number of zoonotic disease outbreaks is positively correlated with the number of threatened mammal and bird species and the number of vector-borne disease outbreaks is negatively correlated with forest cover. These results suggest that, among countries, biodiversity is a source of pathogens, but also that the loss of biodiversity or its regulation, as measured by forest cover or threatened species, seems to be associated with an increase in zoonotic and vector-borne disease outbreaks.

Phylogenetic and population genetic analyses were conducted on tick specimens collected from cattle in northern, northeastern, central, and southern regions of Thailand. Morphological identification indicated these ticks consisted of three species, Rhipicephalus microplus from all four regions, R. sanguineus from the northern and northeastern regions, and a Haemaphysalis species only collected from the northeastern region. Analysis of cytochrome c oxidase subunit I gene ( COI ) sequences identified R. microplus clades A and C, while clade B was not detected in this study. The same analysis indicated specimens morphologically identified as Haemaphysalis were H. bispinosa, confirming previous reports of their prevalence in northeastern Thailand. H. bispinosa showed low haplotype and nucleotide diversity, suggesting either a bottleneck or founder effect. Both R. microplus clades displayed high haplotype diversity and low nucleotide diversity, a pattern associated with population expansion. Genetic structural analysis revealed significant genetic differences in R. microplus clade A, especially between mainland (northern, northeastern, and central regions) and peninsular (southern region) populations, which indicated limited gene flow between these areas while suggesting movement of these ticks across the mainland. The sequence analyses described in this report enhance understanding of the natural history of ticks in Thailand and are expected to guide and strengthen tick control strategies across Southeast Asia.

Leptospirosis is the most common bacterial zoonoses and has been identified as an important emerging global public health problem in Southeast Asia. Rodents are important reservoirs for human leptospirosis, but epidemiological data is lacking. We sampled rodents living in different habitats from seven localities distributed across Southeast Asia (Thailand, Lao PDR and Cambodia), between 2009 to 2010. Human isolates were also obtained from localities close to where rodents were sampled. The prevalence of Leptospira infection was assessed by real-time PCR using DNA extracted from rodent kidneys, targeting the lipL32 gene. Sequencing rrs and secY genes, and Multi Locus Variable-number Tandem Repeat (VNTR) analyses were performed on DNA extracted from rat kidneys for Leptospira isolates molecular typing. Four species were detected in rodents, L. borgpetersenii (56% of positive samples), L. interrogans (36%), L. kirschneri (3%) and L. weilli (2%), which were identical to human isolates. Mean prevalence in rodents was approximately 7%, and largely varied across localities and habitats, but not between rodent species. The two most abundant Leptospira species displayed different habitat requirements: L. interrogans was linked to humid habitats (rice fields and forests) while L. borgpetersenii was abundant in both humid and dry habitats (non-floodable lands). L. interrogans and L. borgpetersenii species are widely distributed amongst rodent populations, and strain typing confirmed rodents as reservoirs for human leptospirosis. Differences in habitat requirements for L. interrogans and L. borgpetersenii supported differential transmission modes. In Southeast Asia, human infection risk is not only restricted to activities taking place in wetlands and rice fields as is commonly accepted, but should also include tasks such as forestry work, as well as the hunting and preparation of rodents for consumption, which deserve more attention in future epidemiological studies.

Reliable diagnostic tools are needed to choose the appropriate treatment and proper control measures for animal trypanosomoses, some of which are pathogenic. Trypanosoma cruzi , for example, is responsible for Chagas disease in Latin America. Similarly, pathogenic animal trypanosomoses of African origin (ATAO), including a variety of Trypanosoma species and subspecies, are currently found in Africa, Latin America and Asia. ATAO limit global livestock productivity and impact food security and the welfare of domestic animals. This review focusses on implementing previously reviewed diagnostic methods, in a complex epizootiological scenario, by critically assessing diagnostic results at the individual or herd level. In most cases, a single diagnostic method applied at a given time does not unequivocally identify the various parasitological and disease statuses of a host. These include “non-infected”, “asymptomatic carrier”, “sick infected”, “cured/not cured” and/or “multi-infected”. The diversity of hosts affected by these animal trypanosomoses and their vectors (or other routes of transmission) is such that integrative, diachronic approaches are needed that combine: (i) parasite detection, (ii) DNA, RNA or antigen detection and (iii) antibody detection, along with epizootiological information. The specificity of antibody detection tests is restricted to the genus or subgenus due to cross-reactivity with other Trypanosoma spp. and Trypanosomatidae, but sensitivity is high. The DNA-based methods implemented over the last three decades have yielded higher specificity and sensitivity for active infection detection in hosts and vectors. However, no single diagnostic method can detect all active infections and/or trypanosome species or subspecies. The proposed integrative approach will improve the prevention, surveillance and monitoring of animal trypanosomoses with the available diagnostic tools. However, further developments are required to address specific gaps in diagnostic methods and the sustainable control or elimination of these diseases. Graphical Abstract

During 2016-2018, we conducted surveillance for Japanese encephalitis virus (JEV) in mosquitoes and pigs in Japan, Thailand, the Philippines, and Indonesia. Phylogenetic analyses demonstrated that our isolates (genotypes Ia, Ib, III, IV) were related to JEV isolates obtained from the same regions many years ago. Indigenous JEV strains persist in Asia.

Toxoplasma gondii, a pathogen of significant concern in animal production, companion animal health, and public health, particularly affects immunocompromised individuals and pregnant women. Current diagnostic techniques employ both direct and indirect methods, with serological assays widely used for detecting T. gondii infections in humans and animals. In this study, the TIM-barrel structure of Br2 β-glucosidase was engineered to create 10 chimeric multi-epitope proteins for T. gondii serological detection. Indirect ELISA screening identified three promising candidate proteins, V4Z, SFF, and S7V-V4Z-SFF, with sensitivities ranging from 71–86% and specificities ranging from 68–76%. Among these, ELISA-V4Z achieved the highest concordance with the reference IFAT method (Kappa = 0.58, 95% CI = 0.32–0.84) and demonstrated a moderate positive predictive value (PPV, 67%) and strong negative predictive value (NPV, 90%). These results suggest that the V4Z chimeric protein demonstrated the strongest performance among the tested candidates for T. gondii detection, exhibiting the highest sensitivity and specificity along with moderate agreement with the reference IFAT. However, its overall diagnostic performance remains limited. These findings highlight the need for further refinement and validation to enhance its diagnostic potential and assess its applicability for broader serological testing.

Background As the largest group of mammalian species, which are also widely distributed all over the world, rodents are the natural reservoirs for many diverse zoonotic viruses. A comprehensive understanding of the core virome of diverse rodents should therefore assist in efforts to reduce the risk of future emergence or re-emergence of rodent-borne zoonotic pathogens. Results This study aimed to describe the viral range that could be detected in the lungs of rodents from Mainland Southeast Asia. Lung samples were collected from 3284 rodents and insectivores of the orders Rodentia, Scandentia, and Eulipotyphla in eighteen provinces of Thailand, Lao PDR, and Cambodia throughout 2006–2018. Meta-transcriptomic analysis was used to outline the unique spectral characteristics of the mammalian viruses within these lungs and the ecological and genetic imprints of the novel viruses. Many mammalian- or arthropod-related viruses from distinct evolutionary lineages were reported for the first time in these species, and viruses related to known pathogens were characterized for their genomic and evolutionary characteristics, host species, and locations. Conclusions These results expand our understanding of the core viromes of rodents and insectivores from Mainland Southeast Asia and suggest that a high diversity of viruses remains to be found in rodent species of this area. These findings, combined with our previous virome data from China, increase our knowledge of the viral community in wildlife and arthropod vectors in emerging disease hotspots of East and Southeast Asia. ADx5T_Q3aYxBw7CRZGTwQ6 Video abstract