Supaluk Popruk

Background: Cryptosporidium spp. and Giardia duodenalis are important parasitic protozoa infecting humans and a wide range of animal species. Some species and assemblages have zoonotic potential and pose health risks to both humans and animals. Objectives: This study aimed to molecularly detect and characterize Cryptosporidium spp. and G. duodenalis and assess their zoonotic potential in pigs, cattle, cats and wildlife. Materials and Methods: Nested PCR and sequence analyses of the small subunit ribosomal RNA ( ssu rRNA) gene for Cryptosporidium spp. and ssu rRNA, β‐giardin ( bg ), glutamate dehydrogenase ( gdh ) and triosephosphate isomerase ( tpi ) genes for G. duodenalis were used for molecular detection and characterization. Results: The PCR positivity rate for Cryptosporidium was 17.9% (12/67) in pigs, whereas the corresponding rates for G. duodenalis were 6.0% (4/67) in pigs and 2.5% (2/80) in cats. Neither parasite was detected in cattle (0/61) or wildlife (0/144). Cryptosporidium scrofarum , a potentially zoonotic species, was the only Cryptosporidium species identified. Among the four Giardia ‐positive pig samples, assemblages B and E were each identified in two samples. Co‐infection with C. scrofarum and assemblage E was detected in one pig. Feline‐associated assemblage F was identified in two cat samples. Conclusions: Potentially zoonotic C. scrofarum and zoonotic G. duodenalis assemblage B were detected in pigs, together with livestock‐associated assemblage E. Feline‐associated assemblage F was identified in cats. These findings provide historical, sample‐specific molecular data on the animals examined.

Background and Aim: Toxoplasma gondii is a globally important zoonotic protozoan, and felids serve as definitive hosts. Although molecular studies have demonstrated substantial genetic diversity of T. gondii in feline populations, information from Thailand remains limited, particularly for stray cats in central and western regions. This retrospective study aimed to determine the prevalence of T. gondii DNA and characterize multilocus genotype profiles in archived DNA extracted from stray-cat fecal samples collected in Nakhon Nayok and Kanchanaburi provinces, Thailand, in 2013. Materials and Methods: A total of 120 archived fecal DNA samples were analyzed, comprising 80 samples from Nakhon Nayok and 40 from Kanchanaburi. Conventional polymerase chain reaction (PCR) targeting the B1 gene was used for molecular detection. Positive amplicons were confirmed by Sanger sequencing and compared with GenBank reference sequences using Basic Local Alignment Search Tool. Sequence-confirmed samples were subsequently analyzed using multilocus nested PCR-restriction fragment length polymorphism (Mn-PCR-RFLP). Reference strains RH, PRU, and VEG represented clonal types I, II, and III, respectively. Differences in PCR positivity between provinces were assessed using Fisher’s exact test. Results: Thirteen of 120 samples (10.8%) were positive for T. gondii DNA, including 8/80 (10.0%) from Nakhon Nayok and 5/40 (12.5%) from Kanchanaburi; the difference was not significant (p = 0.758). B1 sequences showed 94.52%–98.70% identity with reference sequences. Multilocus genotyping was successful in four of 13 positive samples (30.8%). Two type I profiles and one type III profile were identified in Kanchanaburi, whereas one probable type II profile was identified in Nakhon Nayok. No mixed, atypical, or ToxoDB#9 profiles were detected among the four successfully characterized samples. Conclusion: This study provides preliminary historical molecular evidence of classical T. gondii clonal-lineage profiles in stray-cat fecal DNA from central and western Thailand. However, the limited genotyping success prevents estimation of genotype frequencies or regional distribution. Detection of T. gondii DNA in feces should not be interpreted as evidence of viable oocysts, active shedding, infectivity, or current public health risk. Larger prospective studies using fresh samples, quantitative PCR, and higher-resolution genotyping are warranted. Keywords: archived DNA, feline toxoplasmosis, molecular epidemiology, multilocus genotyping, stray cats, Thailand, Toxoplasma gondii, toxoplasmosis.

Background: Giardia duodenalis remains a neglected parasitic infection in Thailand, with uncertain national prevalence and limited data on genetic assemblages. This systematic review and meta-analysis aimed to estimate the pooled prevalence of G. duodenalis infections in Thailand and assess the distribution of assemblages to inform surveillance and control strategies. Methods: The protocol for this study was registered in PROSPERO (CRD42024629406). A comprehensive search was conducted across six databases: EMBASE, Ovid, PubMed, Scopus, Nursing & Allied Health Premium, and the Thai-Journal Citation Index (TCI). The methodological quality of included studies was assessed using the Joanna Briggs Institute (JBI) critical appraisal checklist for observational studies. A meta-analysis using random-effects models estimated the pooled prevalence of G. duodenalis infections and the distribution of assemblages. Results: Sixty-three studies comprising of 47,989 participants were included. The prevalence of G. duodenalis infection varied substantially by diagnostic method, ranging from 1.40% using direct smear microscopy to 1.99% using concentration techniques, 3.66% with culture-based methods, 13.44% with immunoassays, and 8.98% with molecular techniques ( P < 0.0001). Prevalence also differed geographically, with the highest estimates in Western Thailand (12.22%) and the lowest in Northeastern Thailand (0.54%). Vulnerable groups, including orphans (15.14%) and preschool children (11.73%), carried a disproportionate burden of infection. Among the 240 samples analyzed by molecular methods, Assemblage B was the most common (55.0%), followed by Assemblage A (32.5%), mixed A/B infections (12.08%), and a single case of Assemblage F (0.42%). Conclusion: G. duodenalis remains endemic in Thailand, with transmission concentrated in socially vulnerable groups and regions with higher environmental risk. The predominance of Assemblage B and the underuse of molecular diagnostics highlight critical gaps in understanding transmission pathways. These findings suggest that control strategies in Thailand should prioritize high-risk populations and incorporate molecular epidemiology into routine surveillance to more effectively guide prevention efforts. Clinical trial: Not applicable.

Cryptosporidium spp. and Giardia duodenalis are significant intestinal protozoan parasites affecting humans and animals worldwide. These infections are transmitted through the faecal–oral route, by contaminated water, food or close contact with infected hosts. The zoonotic risk of transmission from reptiles is considered low; yet, limited research exists regarding Cryptosporidium and Giardia infections in these animals. This study investigated the presence of Cryptosporidium spp. and G. duodenalis in reptile faecal samples from a breeding farm in the Chonburi Province, Eastern Thailand. The test population included 363 iguanas, 50 tortoises, 79 bearded dragons and nine snakes. DNA extraction from faecal samples was performed, followed by nested PCR for Cryptosporidium and G. duodenalis , and all positive samples were sequenced for molecular characterisation. Cryptosporidium and G. duodenalis were detected in 13 and 25 out of 501 faecal samples, respectively. Specifically, seven of 363 iguana samples and six of 79 bearded dragon samples tested positive for Cryptosporidium . Cryptosporidium avium was found in 11 samples, and one sample revealed Cryptosporidium serpentis ; the remaining sample could not be identified at the species level. For G. duodenalis , 15 of 363 iguanas tested positive, along with six of 79 bearded dragons, two of 50 tortoises and two of nine snakes. Only G. duodenalis assemblage B was identified in positive samples. This study provides epidemiological information on the infection rate and distribution of species of Cryptosporidium and Giardia in a breeding reptile farm in Chonburi Province, Eastern Thailand, and the findings are discussed in the context of the scientific literature. Summary: This study represents one of the largest surveys on Cryptosporidium and Giardia duodenalis in reptiles. Cryptosporidium avium , typically associated with avian hosts, was the predominant Cryptosporidium species identified in iguanas and bearded dragons. Giardia duodenalis assemblage B, known for its zoonotic potential, was the primary assemblage found in iguanas.

The escalating global incidence of antimicrobial resistance poses a significant public health challenge. In response, exploring alternative antimicrobial agents, particularly derived from plants, becomes crucial to alleviate the selective pressure exerted by conventional antibiotics. The aim of this study was to characterize the composition of essential oil extracted from Litsea cubeba fruits and to evaluate its antimicrobial potential, along with its major compound, across solid, liquid, and vapor phases. The antimicrobial activity was assessed against a diverse range of human pathogenic Gram-positive bacteria (n=8), Gram-negative bacteria (n=34), filamentous fungi (n=2), and yeast (n=1). Disk diffusion, broth macrodilution, and vapor-phase diffusion methods were employed. This study found that all phases of L. cubeba essential oil and purified limonene exhibited broad-spectrum bactericidal and fungicidal activities (solid-phase: inhibition zone diameter (IZD) 19 mm vs 14 mm; liquid-phase: minimum inhibitory concentration (MIC) 2.0 mg/mL vs 4.0 mg/mL; vapor-phase: IZD 90 mm vs 45 mm), with superior efficacy against filamentous fungi and yeast compared to bacteria (solid-phase: IZD 90 mm vs 17.5 mm; liquid-phase: MIC 2.0 mg/mL vs 0.06 mg/mL; vapor-phase: IZD 90 mm vs 12.5 mm; all p-values<0.05). Among bacteria, solid-phase L. cubeba essential oil demonstrated increased activity against Staphylococcus saprophyticus and Acinetobacter lwoffii whereas liquid-phase L. cubeba essential oil had optimal activity against Streptococcus agalactiae and Elizabethkingia meningoceptica. Notably, Trichophyton rubrum, Nannizzia gypsea, and Candida albicans displayed high susceptibility to all phases of L. cubeba essential oil. These findings highlight the potential activity of L. cubeba essential oil, across its various phases, as a promising alternative antimicrobial agent against medically significant pathogens, providing essential baseline information for further exploration and development of L. cubeba essential oil in the pursuit of combating antimicrobial resistance.

Bovine neosporosis is among the main causes of abortion in cattle worldwide, causing serious economic losses in the beef and dairy industries. A highly sensitive and specific diagnostic method for the assessment of the epidemiology of the disease, as well as it surveillance and management, is imperative, due to the absence of an effective treatment or vaccine against neosporosis. In the present study, the immunodiagnostic performance of Neospora caninum peroxiredoxin 2 (NcPrx2), microneme 4 (NcMIC4), and surface antigen 1 (NcSAG1) to detect IgG antibodies against N. caninum in cattle were evaluated and compared with that of the indirect fluorescent antibody test (IFAT). The results revealed that NcSAG1 had the highest sensitivity and specificity, with values of 88.4% and 80.7%, respectively, followed by NcPrx2, with a high sensitivity of 87.0% but a low specificity of 67.0%, whereas NcMIC4 showed sensitivity and specificity of 84.1% and 78.9%, respectively, when compared with IFAT. A high degree of agreement was observed for NcSAG1 (k = 0.713) recombinant protein, showing the highest diagnostic capability, followed by NcMIC4 (k = 0.64) and NcPrx2 (k = 0.558). The present study demonstrates that NcSAG1 is helpful as an antigen marker and also demonstrates the potential immunodiagnostic capabilities of NcPrx2 and NcMIC4, which could serve as alternative diagnostic markers for detecting N. caninum infection in cattle. These markers may find utility in future treatment management, surveillance, and risk assessment of neosporosis in livestock or other animal host species. Further research should be directed toward understanding the in vivo immune response differences resulting from immunization with both recombinant proteins.

Giardia duodenalis is a significant protozoan that affects humans and animals. An estimated 280 million G. duodenalis diarrheal cases are recorded annually. Pharmacological therapy is crucial for controlling giardiasis. Metronidazole is the first-line therapy for treating giardiasis. Several metronidazole targets have been proposed. However, the downstream signaling pathways of these targets with respect to their antigiardial action are unclear. In addition, several giardiasis cases have demonstrated treatment failures and drug resistance. Therefore, the development of novel drugs is an urgent need. In this study, we performed a mass spectrometry-based metabolomics study to understand the systemic effects of metronidazole in G. duodenalis. A thorough analysis of metronidazole processes helps identify potential molecular pathways essential for parasite survival. The results demonstrated 350 altered metabolites after exposure to metronidazole. Squamosinin A and N-(2-hydroxyethyl)hexacosanamide were the most up-regulated and down-regulated metabolites, respectively. Proteasome and glycerophospholipid metabolisms demonstrated significant differential pathways. Comparing glycerophospholipid metabolisms of G. duodenalis and humans, the parasite glycerophosphodiester phosphodiesterase was distinct from humans. This protein is considered a potential drug target for treating giardiasis. This study improved our understanding of the effects of metronidazole and identified new potential therapeutic targets for future drug development.

Blastocystis is one of the most common enteric protozoa that inhabits the intestinal tract of humans and different animals. Moreover, it has a worldwide geographic distribution. Its main mode of transmission is via the fecal-oral route. At present, 26 subtypes are widely distributed across both humans and animals. The current study aimed to determine the prevalence and subtype distribution of Blastocystis among school-aged children living on the Thai-Myanmar border, Ratchaburi province, Thailand. In total, 508 samples were collected from children at six schools. The prevalence of Blastocystis infection was amplified and sequenced in the 600 bp barcode region of the small-subunit ribosomal RNA (SSU rRNA). The overall prevalence of Blastocystis infection was 3.35% (17/508). ST3 (11/17) was the most predominant subtype, followed by ST1 (5/17) and ST2 (1/17). A phylogenetic tree was constructed based on the Tamura92+G+I model using the maximum-likelihood algorithm. Surprisingly, all sequences of the ST3-positive samples were closely correlated with the cattle-derived sequence. Meanwhile, all sequences of the Blastocystis ST1-positive samples were closely correlated with the human-derived sequence. Nevertheless, further studies should be conducted to validate the zoonotic transmission of Blastocystis. Based on our findings, personal hygiene and sanitation should be improved to promote better health in children in this area.