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This study investigates the distribution of the Lymnaea (Radix) spp. in Pak Chong district, Nakhon Ratchasima province, northeast Thailand, where a vast cattle farming area is located and has a high prevalence of Fasciola spp. in the cattle. By random selection, 1,414 snails were collected from the natural and man-made ponds. The snails were recorded for morphology and processed for DNA isolation. The snail species were investigated by conventional PCR using a 16S rDNA-specific primer. The result demonstrated that all collected snails were R. (L.) rubiginosa. Moreover, the infection of Fasciola gigantica in the snails was investigated by PCR using a cytochrome c oxidase I (COX1)-specific primer. The results illustrated that the overall prevalence was 22.5% (318/1414), with the highest prevalence in the Nong Sa Rai subdistrict at 73.6% (81/110), which is the highest prevalence of Fasciola gigantica in the snail host that has ever been reported. The lowest prevalence existed in the Pong Ta Long subdistrict at 3.7% (4/109). Our results corresponded to the previous report on the Fasciola spp. infection in the cattle from this area, and the geographical analysis revealed that the most suspected factor would be the earth dam located in these subdistricts, where many animals live freely during the day. Our findings could be helpful for further parasite control and could trigger the study of the biology and associated factors in the future.

Background and Aim: Gastrointestinal (GI) nematode infection remains an important problem in livestock, particularly cattle. The infection may lead to serious health complications and affect animal products. The objective of this study was to investigate GI nematode infection and its associated risk factors in dairy and beef cattle farmed in Pak Chong District of Nakhon Ratchasima Province, northeast Thailand. Materials and Methods: Fecal specimens were collected from 101 dairy cattle and 100 beef cattle. Formalin-ethyl acetate concentration techniques were used to process the samples and the samples were observed under a light microscope. Samples were subjected to molecular identification of specific genera using conventional polymerase chain reaction and DNA sequencing. Results: The overall prevalence of GI nematode infection was 33.3%. The strongyle nematode was the most significant GI nematode in this area with a prevalence of 28.4%. The prevalence of strongyle nematodes was 58.0% in beef cattle and only 7.9% in dairy cattle. Trichuris spp. was another nematode found in both types of cattle with an overall prevalence of 5.0% with 9.0% in beef cattle and 1.0% in dairy cattle. The results of the epidemiological study indicate that the age of cattle, food, water sources, farming system, and housing floor are the most important risk factors. Among the strongyle nematodes, Ostertagia spp. was the most prevalent (82.0%), followed by Haemonchus spp. (62.3%) and Trichostrongylus spp. (8.2%), respectively. Conclusion: Infection with GI nematodes still exists in this area, particularly in beef cattle. Our reported data may benefit local parasitic control policies in the future.

Abstract Background Helminth infection is a global health issue that not only causes acute helminthiasis but long-term infection may lead to complicated symptoms as well as severe complications. The World Health Organization cooperated with the Ministry of Public Health in many countries, particularly where high prevalence, spending a lot of resources for limiting the infection. In Thailand, the incidence of parasitic helminth infections was continuously declined in the last few decades according to several campaigns for parasitic elimination. However, the rural community in the northeast of Thailand where the highest prevalence of the country still needs to be monitored. This present study aims to report the current prevalence of parasitic helminth infections in Nakhon Ratchasima and Chaiyaphum provinces where sharing a huge area of the northeastern region of Thailand but only a few studies have been published. Methods The stool specimens were collected from 11,196 volunteers and processed by modified Kato-Katz thick smear, PBS-ethyl acetate concentration techniques, and PCR. The epidemiological data were collected, analyzed, and used for generating of parasitic hotspots. Results The results indicated that O. viverrini remains the major parasite in this area with a total prevalence of 5.05% followed by Taenia spp., Hookworms, T. trichiura, and Echinostoma spp., respectively. Mueang district of Chaiyaphum province has the highest prevalence especially O. viverrini with a prevalence of 7.15% that higher than the latest national surveillance. Interestingly, the prevalence of O. viverrini was hugely reported (more than 10%) in five subdistricts. The geographic localization of O. viverrini infections revealed that a lot of water reservoirs such as the lakes or branches of the river in the two-most prevalent subdistricts. Our finding indicated that gender and age were insignificantly different. Conclusion This finding suggested that the parasitic helminth infection in the rural areas of northeast of Thailand remains high and the housing location is a major contributing factor for the parasitic infection.

Introduction: Cholangiocarcinoma (CCA) is a highly progressive tumor. The standard chemotherapy varies in its effectiveness, with generally low efficacy. So, the discovery of novel chemotherapy is still required. The objective of this preliminary study was to determine the cytotoxic effects induced by three kinds of bamboo mushrooms (Dictyophora indusiata or Chinese bamboo mushroom; Ch-DTP, Short skirt bamboo mushroom (Thai isolate); Th-DTP, and orange skirt bamboo mushroom; Or-DTP) on CCA cells. Materials and methods: CCA cell lines, including CL-6, HuCCT1, HuH28, and OUMS normal fibroblast cells, were treated with various concentrations of DTP extracts. The MTT assay was used to determine cytotoxicity, and cell morphology was observed by using phase-contrast microscopy. Results and discussion: The results suggested that Ch-DTP effectively killed all three CCA cell lines in both low (0.3 mg/mL) and high (0.6 mg/mL) doses, but Th-DTP and Or-DTP had significantly reduced cell viability only at high doses (p

Objective: To determine the prevalence of mosquito larvae in campus areas and the infection rate of endosymbiotic bacteria, Wolbachia in mosquito larvae. Method: The mosquito larvae samples were collected in residential areas and academic buildings of Suranaree University of Technology located in Northeastern Thailand during 2017-2018. Mosquito species identification was performed using GLOBE mosquito protocols and Rattanarithikul & Panthusiri’s keys. The gene encoding for the surface protein of Wolbachia was amplified by PCR and confirmed by DNA sequencing. Results: Armigeres sp. is the highest proportion of mosquito larvae followed by Culex spp., Aedes albopictus, Aedes aegypti, and Toxorynchites spp., respectively. Aedes aegypti have breeding sites mostly in the containers found indoors, whereas the main breeding sites of Aedes albopictus were found in both outdoors and indoors. The House Index and Breteau Index for Aedes spp. was more than 5% and 20%, respectively, in both areas, indicating that these areas are dengue sensitive. The highest proportion of Wolbachia infection was found in the larvae of Culex spp. (86.21%), followed by Aedes albopictus (69.23%) and rarely detected in Aedes aegypti (9.09%). Conclusion: The present study reported the first natural infection of Wolbachia in mosquito larvae in Thailand. Our result suggested that the mosquito species containing higher proportion of Wolbachia are less likely to be vectors for dengue. Therefore, Wolbachia transfection in mosquito larvae could be applied as a biocontrol for dengue and other mosquito-borne disease prevention.

There is an inverse relationship between the high incidence of helminth infection and the low incidence of inflammatory disease. Hence, it may be that helminth molecules have anti-inflammatory effects. Helminth cystatins are being extensively studied for anti-inflammatory potential. Therefore, in this study, the recombinant type I cystatin (stefin-1) of Fasciola gigantica (rFgCyst) was verified to have LPS-activated anti-inflammatory potential, including in human THP-1-derived macrophages and RAW 264.7 murine macrophages. The results from the MTT assay suggest that rFgCyst did not alter cell viability; moreover, it exerted anti-inflammatory activity by decreasing the production of proinflammatory cytokines and mediators, including IL-1β, IL-6, IL-8, TNF-α, iNOS, and COX-2 at the gene transcription and protein expression levels, as determined by qRT-PCR and Western blot analysis, respectively. Further, the secretion levels of IL-1β, IL-6, and TNF-α determined by ELISA and the NO production level determined by the Griess test were decreased. Furthermore, in Western blot analysis, the anti-inflammatory effects involved the downregulation of pIKKα/β, pIκBα, and pNF-κB in the NF-κB signaling pathway, hence reducing the translocation from the cytosol into the nucleus of pNF-κB, which subsequently turned on the gene of proinflammatory molecules. Therefore, cystatin type 1 of F. gigantica is a potential candidate for inflammatory disease treatment.

Cryptococcal meningoencephalitis, the most common form of cryptococcosis, is caused by the opportunistic fungal pathogen, Cryptococcus neoformans. Molecular strategies used by C. neoformans to invade the central nervous system (CNS) have been the focus of several studies. Recently, the role of a novel secreted metalloprotease (Mpr1) in the pathogenicity of C. neoformans was confirmed by studies demonstrating that Mpr1 mediated the migration of fungal cells into the CNS. Given this central function, the aim here was to identify the molecular determinants of Mpr1 activity and resolve their role in the migration of cryptococci across the blood-brain barrier (BBB). The Mpr1 protein belongs to an understudied group of metalloproteases of the M36 class of fungalysins unique to fungi. They are generally synthesized as propeptides with fairly long prodomains and highly conserved regions within their catalytic core. Through structure-function analysis of Mpr1, our study identified the prodomain cleavage sites of Mpr1 and demonstrated that when mutated, the prodomain appears to remain attached to the catalytic C-terminus of Mpr1 rendering a nonfunctional Mpr1 protein and an inability for cryptococci to cross the BBB. We found that proteolytic activity of Mpr1 was dependent on the coordination of zinc with two histidine residues in the active site of Mpr1, since amino acid substitutions in the HExxH motif abolished Mpr1 proteolytic activity and prevented the migration of cryptococci across the BBB. A phylogenetic analysis of Mpr1 revealed a distinct pattern likely reflecting the neurotropic nature of C. neoformans and the specific function of Mpr1 in breaching the BBB. This study contributes to a deeper understanding of the molecular regulation of Mpr1 activity and may lead to the development of specific inhibitors that could be used to restrict fungal penetration of the CNS and thus prevent cryptococcal meningoencephalitis-related deaths.

ABSTRACT Cryptococcus spp. cause life-threatening fungal infection of the central nervous system (CNS), predominantly in patients with a compromised immune system. Why Cryptococcus neoformans has this remarkable tropism for the CNS is not clear. Recent research on cerebral pathogenesis of C. neoformans revealed a predominantly transcellular migration of cryptococci across the brain endothelium; however, the identities of key fungal virulence factors that function specifically to invade the CNS remain unresolved. Here we found that a novel, secreted metalloprotease (Mpr1) that we identified in the extracellular proteome of C. neoformans (CnMpr1) is required for establishing fungal disease in the CNS. Mpr1 belongs to a poorly characterized M36 class of fungalysins that are expressed in only some fungal species. A strain of C. neoformans lacking the gene encoding Mpr1 (mpr1Δ) failed to breach the endothelium in an in vitro model of the human blood-brain barrier (BBB). A mammalian host infected with the mpr1Δ null strain demonstrated significant improvement in survival due to a reduced brain fungal burden and lacked the brain pathology commonly associated with cryptococcal disease. The in vivo studies further indicate that Mpr1 is not required for fungal dissemination and Mpr1 likely targets the brain endothelium specifically. Remarkably, the sole expression of CnMPR1 in Saccharomyces cerevisiae resulted in a robust migration of yeast cells across the brain endothelium, demonstrating Mpr1’s specific activity in breaching the BBB and suggesting that Mpr1 may function independently of the hyaluronic acid-CD44 pathway. This distinct role for Mpr1 may develop into innovative treatment options and facilitate a brain-specific drug delivery platform. IMPORTANCE Cryptococcus neoformans is a medically relevant fungal pathogen causing significant morbidity and mortality, particularly in immunocompromised individuals. An intriguing feature is its strong neurotropism, and consequently the hallmark of cryptococcal disease is a brain infection, cryptococcal meningoencephalitis. For C. neoformans to penetrate the central nervous system (CNS), it first breaches the blood-brain barrier via a transcellular pathway; however, the identities of fungal factors required for this transmigration remain largely unknown. In an effort to identify extracellular fungal proteins that could mediate interactions with the brain endothelium, we undertook a proteomic analysis of the extracellular proteome and identified a secreted metalloprotease (Mpr1) belonging to the M36 class of fungalysins. Here we found that Mpr1 promotes migration of C. neoformans across the brain endothelium and into the CNS by facilitating attachment of cryptococci to the endothelium surface, thus underscoring the critical role of M36 proteases in fungal pathogenesis.