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The emergence of infectious diseases presents a significant global health, economic, and security risk. Climate change can unexpectedly lead to the spread of pathogens, vectors, or hosts into new areas, contributing to the rise of infectious diseases. Surveillance plays a crucial role in monitoring disease trends and implementing control strategies. In this study, we document the first discovery of Heterobilharzia americana, a parasitic schistosome of mammals and its intermediate hosts Galba cubensis and Galba humilis along the banks of the Colorado River in California. We conducted multiple samplings of snails from various locations in the region with a previous history of canine schistosomiasis. Nucleotide sequencing of the multiple regions of the snails' and parasites' DNA revealed the coexistence of G. cubensis and G. humilis, both infected with H. americana. Phylogenetic analyses further validate the presence of H. americana in California, suggesting a wider distribution than previously reported. Our findings have implications for public health, veterinary medicine, and biodiversity conservation, contributing to developing effective control strategies to prevent the spread of this emerging infectious disease. [ABSTRACT FROM AUTHOR]

Indoplanorbis exustus is a freshwater gastropod belonging to the family Planorbidae. This snail is widely distributed across the tropics and plays an important role as the intermediate host for trematodes. However, relatively little is understood regarding the genetic relationship between I. exustus and trematodes. The goals of this study were to investigate the current transmission status of trematode cercariae in I. exustus in Thailand and to examine the genetic diversity, genetic structure, and demographic history of I. exustus. We collected 575 I. exustus from 21 provinces across six regions of Thailand and investigated cercarial infections by using the shedding method. I. exustus from two provinces were infected with cercarial trematodes, and two types of cercarial stages were molecularly identified as furcocercous cercaria and xiphidiocercariae. Phylogenetic tree analysis based on 28S rDNA and ITS2 sequences demonstrated that furcocercous cercaria and xiphidiocercariae were closely clustered with a clade of Euclinostomum sp. and Xiphidiocercariae sp., respectively. Phylogenetic and network analyses of I. exustus haplotypes based on the COI, 16S rDNA, and ITS1 genes demonstrated four main clades. Only snails in clade A were distributed in all regions of Thailand and harbored trematode cercariae. The level of genetic diversity of I. exustus was relatively high, but most populations were not genetically different, thus suggesting the appearance of gene flow within the I. exustus populations. Overall, the haplotype network was star-shaped, thus suggesting the recent demographic expansion of populations. This result was also supported by the unimodal mode of the mismatch distribution graph and the large negative values of the neutrality tests. Therefore, the I. exustus snail was likely another freshwater snail of the invasive species in Thailand. This information will aid in monitoring the spread of the parasitic trematodes carried by I. exustus from different populations. [ABSTRACT FROM AUTHOR]

Sand flies (Diptera: Psychodidae) are significant medical vectors of pathogens that cause illness in humans and animals. The most significant of these is the genus Leishmania (Kinetoplastida: Trypanosomatidae). Leishmaniasis is an important vector-borne disease. They are spread throughout the Old and New Worlds by the bites of female phlebotomine sand flies. Despite this importance, certain provinces where autochthonous leishmaniasis has historically existed and is re-emerging are lacking current data regarding their sand fly fauna. To provide a current update on the local sand fly fauna, a comprehensive entomological survey encompassing eight provinces was conducted subsequent to an analysis of past data regarding the species composition and distribution of sand flies in the area. Thus, the purpose of this study was to ascertain the species composition in various locations across eight provinces spread across four regions in Thailand. In order to gather information about the distribution of the phlebotomine sand fly, researchers used modified Centers for Disease Control and Prevention light traps between February 2021 and February 2022. At these 30 trappings in each province, we conducted 240 trapping nights in total. Collected sand flies were morphologically identified. The adult sand flies collected in this study were 8314 and represented 24 species of four genera, including 16 of Sergentomyia, 6 of Phlebotomus, 1 of Idiophlebotomus, and 1 of Chinius. Among these species, Sergentomyia anodontis had the largest population (25.71%), and population peaks were observed in Uthaithani province (51.81%). The collected sand flies' male-to-female ratio was 1.21:1.00 (4,556:3,758). The study also discovered that Phlebotomus argentipes, a vector of Leishmania spp., was captured in Phayao province, and Phlebotomus major major was majorly found in Uthaithani province. The widespread distribution of sand fly populations in this study may indicate the epidemiologic importance of vector ecology in this habitat. Our results confirm the presence of several sand fly species in different provinces of Thailand and provide valuable information about the distribution and behavior of phlebotomine sand flies, which can be used to develop effective control strategies for the prevention of sand fly-borne diseases. In several Thai provinces, the study also tracked the nocturnal movements of both male and female sand flies. The results showed that phlebotomine sand flies are more active at night, peaking just after dusk. [ABSTRACT FROM AUTHOR]

Xenorhabdus and Photorhabdus can produce a variety of secondary metabolites with broad spectrum bioactivity against microorganisms. We investigated the antibacterial activity of Xenorhabdus and Photorhabdus against 15 antibiotic-resistant bacteria strains. Photorhabdus extracts had strong inhibitory the growth of Methicillin-resistant Staphylococcus aureus (MRSA) by disk diffusion. The P. akhurstii s subsp. akhurstii (bNN168.5_TH) extract showed lower minimum inhibitory concentrations (MIC) and minimal bactericidal concentrations (MBC). The interaction between either P. akhurstii subsp. akhurstii (bNN141.3_TH) or P. akhurstii subsp. akhurstii (bNN168.5_TH) or P. hainanensis (bNN163.3_TH) extract in combination with oxacillin determined by checkerboard assay exhibited partially synergistic interaction with fractional inhibitory concentration index (FICI) of 0.53. Time-killing assay for P. akhurstii subsp. akhurstii (bNN168.5_TH) extract against S. aureus strain PB36 significantly decreased cell viability from 105 CFU/ml to 103 CFU/ml within 30 min (P < 0.001, t-test). Transmission electron microscopic investigation elucidated that the bNN168.5_TH extract caused treated S. aureus strain PB36 (MRSA) cell membrane damage. The biosynthetic gene clusters of the bNN168.5_TH contained non-ribosomal peptide synthetase cluster (NRPS), hybrid NRPS-type l polyketide synthase (PKS) and siderophore, which identified potentially interesting bioactive products: xenematide, luminmide, xenortide A-D, luminmycin A, putrebactin/avaroferrin and rhizomide A-C. This study demonstrates that bNN168.5_TH showed antibacterial activity by disrupting bacterial cytoplasmic membrane and the draft genome provided insights into the classes of bioactive products. This also provides a potential approach in developing a novel antibacterial agent. [ABSTRACT FROM AUTHOR]

Background: Freshwater snails serve as intermediate hosts for a variety of trematodes that cause illness in the human and animal populations. Several species of freshwater snails in Thailand have been found to have larval trematode infections. We aimed to investigate a freshwater snail in Phitsanulok Province and report on its current status of larval trematode infection. Methods: Freshwater snails were collected from six localities (rice field and irrigation canal) by handpicking and using a count per unit of time sampling approach. The snails were identified by their external shell morphology. The shedding method was applied to observe the cercariae, which were photographed under a light microscope to determine their morphological types. Results: A total of 211 snails were classified into seven genera. The most abundant snail species was Lymnaea sp., representing 31.3% of the sample, followed by Physella sp., Bithynia sp., Pomacea canaliculata, Filopaludina martensi, Indoplanorbis exustus, and Melanoides tuberculata, in that order. From the sample, 21 snails (9.95%), including Bithynia sp., Lymnaea sp., I. exustus, and M. tuberculata, were infected with cercarial trematodes, which could be categorized into four types, namely amphistome, parapleurolophocercous, echinostome, and xiphidiocercaria. Amphistome emerged from Bithynia sp., and I. exustus was the most common cercaria to be recovered, representing 80.9% of all infected snails. Conclusion: This study presents the current prevalence of cercariae in infected snails within the studied area. It is important to manage intermediate host snails in order to restrict trematode life cycle completion. [ABSTRACT FROM AUTHOR]

Summary: Entomopathogenic nematodes (EPN) Steinernema and Heterorhabditis with symbionts with Xenorhabdus and Photorhabdus bacteria, respectively, are reported as biocontrol agents for insect control. The objectives of this study were to identify EPN and their symbiotic bacteria in national parks of Phitsanulok Province, Thailand, and to test how Xenorhabdus isolates attack Aedes aegypti larvae. We collected 810 soil samples from four national parks. The juvenile stage of EPN was isolated from soil samples using a baiting technique with Galleria mellonella followed by a White trap. Partial regions of 28S rDNA and internal transcript spacer were sequenced to identify EPN, and recA sequencing was used to discriminate between Xenorhabdus and Photorhabdus. We found that 74 of the 810 soil samples (9.1%) were positive for the EPN. The EPN were molecularly identified as S. surkhetense , S. longicaudum , H. indica and Heterorhabditis sp. SGmg3, and their symbiotic bacteria were identified as X. stockiae , X. griffiniae , X. indica , X. vietnamensis , P. luminescens subsp. akhurstii , and P. temperata subsp. temperata. Xenorhabdus griffiniae showed potential larvicidal activity against A. aegypti (91% mortality at 72 and 96 h after exposure). This study demonstrates the diversity of EPN and symbiotic bacteria in national parks of Thailand and the potential to use X. griffiniae as a biocontrol agent to kill A. aegypti larvae. [ABSTRACT FROM AUTHOR]

Xenorhabdus and Photorhabdus are gram negative bacteria that can produce several secondary metabolites, including antimicrobial compounds. They have a symbiotic association with entomopathogenic nematodes (EPNs). The aim of this study was to isolate and identify Xenorhabdus and Photorhabdus species and their associated nematode symbionts from Northeastern region of Thailand. We also evaluated the antibacterial activity of these symbiotic bacteria. The recovery rate of EPNs was 7.82% (113/1445). A total of 62 Xenorhabdus and 51 Photorhabdus strains were isolated from the EPNs. Based on recA sequencing and phylogeny, Xenorhabdus isolates were identified as X. stockiae (n = 60), X. indica (n = 1) and X. eapokensis (n = 1). Photorhabdus isolates were identified as P. luminescens subsp. akhurstii (n = 29), P. luminescens subsp. hainanensis (n = 18), P. luminescens subsp. laumondii (n = 2), and P. asymbiotica subsp. australis (n = 2). The EPNs based on 28S rDNA and internal transcribed spacer (ITS) analysis were identified as Steinernema surkhetense (n = 35), S. sangi (n = 1), unidentified Steinernema (n = 1), Heterorhabditis indica (n = 39), H. baujardi (n = 1), and Heterorhabditis sp. SGmg3 (n = 3). Antibacterial activity showed that X. stockiae (bMSK7.5_TH) extract inhibited several antibiotic-resistant bacterial strains. To the best of our knowledge, this is the first report on mutualistic association between P. luminescens subsp. laumondii and Heterorhabditis sp. SGmg3. This study could act as a platform for future studies focusing on the discovery of novel antimicrobial compounds from these bacterial isolates. [ABSTRACT FROM AUTHOR]

Cryptozona siamensis, one of the most widespread land snails, is native to Thailand, and plays a key role as an agricultural pest and intermediate host for Angiostrongylus spp. However, its genetic diversity and population structure has not yet been investigated, and are poorly understood. Therefore, a genetic analysis of the C. siamensis population in Thailand was conducted, based mitochondrial 16S rRNA (402 bp) and COI (602 bp) gene fragment sequences. Cryptozona siamensis randomly collected from 17 locations in four populations across Thailand, between May 2017 and July 2018. Fifty-eight snails were used to examine the phylogeny, genetic diversity, and genetic structure. The maximum likelihood tree based on the 16S rRNA and COI fragment sequences revealed two main clades. A total of 14 haplotypes with 44 nucleotide variable sites were found in the 16S rRNA sequences, while 14 haplotypes with 57 nucleotide variable sites were found in the COI sequences. The genetic diversity of C. siamensis in term of the number of haplotypes and haplotype diversity, was found to be high but the nucleotide diversity showed low levels of genetic differentiation for the COI sequence as also noted with the 16S rRNA sequence. The population genetic structure of C. siamensis revealed genetic difference in most populations in Thailand. However, low genetic difference in some populations may be due to high gene flow. This study provides novel insights into the basic molecular genetics of C. siamensis. [ABSTRACT FROM AUTHOR]

Xenorhabdus and Photorhabdus, symbiotically associated with entomopathogenic nematodes (EPNs), produce a range of antimicrobial compounds. The objective of this study is to identify Xenorhabdus and Photorhabdus and their EPNs hosts, which were isolated from soil samples from Saraburi province, and study their antibacterial activity against 15 strains of drug-resistant bacteria. Fourteen isolates (6.1%), consisting of six Xenorhabdus isolates and eight Photorhabdus isolates, were obtained from 230 soil samples. Based on the BLASTN search incorporating the phylogenetic analysis of a partial recA gene, all six isolates of Xenorhabdus were found to be identical and closely related to X. stockiae. Five isolates of Photorhabdus were found to be identical and closely related to P. luminescens subsp. akhurstii. Two isolates of Photorhabdus were found to be identical and closely related to P. luminescens subsp. hainanensis. The remaining isolate of Photorhabdus was found to be identical to P. asymbiotica subsp. australis. The bacterial extracts from P. luminescens subsp. akhurstii showed strong inhibition the growth of S. aureus strain PB36 (MSRA) by disk diffusion, minimal inhibitory concentration, and minimal bactericidal concentration assay. The combination between each extract from Xenorhabdus/Photorhabdus and oxacillin or vancomycin against S. aureus strain PB36 (MRSA) exhibited no interaction on checkerboard assay. Moreover, killing curve assay of P. luminescens subsp. akhurstii extracts against S. aureus strain PB36 exhibited a steady reduction of 105 CFU/ml to 103 CFU/ml within 30 min. This study demonstrates that Xenorhabdus and Photorhabdus, showed antibacterial activity. This finding may be useful for further research on antibiotic production. [ABSTRACT FROM AUTHOR]

Aedes aegypti is an insect vector that transmits several viruses affecting humans worldwide. Entomopathogenic nematodes (EPNs) and their symbiotic bacteria are organisms with the potential to control many insects. In this study, we did a survey aimed to identify EPNs and their symbiotic bacteria and evaluate the larvicidal activity of bacteria against Ae. aegypti. We collected 540 soil samples from 108 sites in Phitsanulok Province, lower northern Thailand. Baiting techniques and White traps were used to isolate EPNs from soil samples. By sequencing of 28S rDNA and internal transcribed spacer regions, 51 EPN isolates were identified as Steinernema surkhetense (35 isolates), Heterorhabditis indica (14 isolates) and Heterorhabditis sp. SGmg3 (two isolates). Based on sequencing of a partial region of the recA gene, 35 isolates of Xenorhabdus were identified as Xenorhabdus stockiae, and 20 Photorhabdus isolates were identified as Photorhabdus luminescens subsp. akhurstii (10 isolates), P. luminescens subsp. hainanensis (seven isolates) and P. asymbiotica subsp. australis (three isolates). Screening for larvicidal activity of bacteria against Ae. aegypti was performed in the laboratory. Xenorhabdus WB5.4 and Xenorhabdus WB12.5, which were closely related to X. stockiae, resulted in high mortality of Ae. aegypti (99.99% and 70%, respectively) at 96 hr after exposure. Comparing with control groups, mortality of Ae. aegypti larvae was low (1.11%–6.67%) after exposure for 24–96 hr. Our findings showed the potential of X. stockiae for controlling Ae. aegypti. Further studies are needed to elucidate the mechanisms through which these bacteria kill Ae. aegypti larvae. [ABSTRACT FROM AUTHOR]

Angiostrongyliasis is a parasitic disease caused by nematodes of the genus Angiostrongylus. Distribution of this worm corresponds to the dispersal of its main intermediate host, the giant African land snail Achatina fulica. Genetic characterization can help identify parasitic pathogens and control the spreading of disease. The present study describes infection of A. fulica by Angiostrongylus, and provides a genetic outlook based on sequencing of specific regions. We collected 343 land snails from 22 provinces across six regions of Thailand between May 2017 and July 2018. Artificial digestion and Baermann’s technique were employed to isolate Angiostrongylus larvae. The worm and its intermediate host were identified by sequencing with specific nucleotide regions. Phylogenetic tree was constructed to evaluate the relationship with other isolates. A. fulica from Chaiyaphum province was infected with A. cantonensis, whereas snails collected from Phrae and Chiang Rai provinces were infected with A. malaysiensis. The maximum likelihood tree based on 74 A. fulica COI sequences revealed monophyletic groups and identified two haplotypes: AF1 and AF2. Only AF1, which is distributed in all regions of Thailand, harbored the larvae of A. cantonensis and A. malaysiensis. Two mitochondrial genes (COI and cytb) and two nuclear regions (ITS2 and SSU rRNA) were sequenced in 41 Angiostrongylus specimens. The COI gene indicated that A. cantonensis was closely related to the AC10 haplotype; whereas the cytb gene revealed two new haplotypes: AC19 and AC20. SSU rRNA was useful for the identification of A. cantonensis; whereas ITS2 was a good genetic marker for differentiating between A. cantonensis and A. malaysiensis. This study provides genetic information about the parasite Angiostrongylus and its snail intermediate host. The data in this work may be useful for further study on the identification of Angiostrongylus spp., the genetic relationship between intermediate host and parasite, and control of parasites. [ABSTRACT FROM AUTHOR]

Summary: Bacteria of the genera Photorhabdus and Xenorhabdus produce a plethora of natural products to support their similar symbiotic life cycles. For many of these compounds, the specific bioactivities are unknown. One common challenge in natural product research when trying to prioritize research efforts is the rediscovery of identical (or highly similar) compounds from different strains. Linking genome sequence to metabolite production can help in overcoming this problem. However, sequences are typically not available for entire collections of organisms. Here, we perform a comprehensive metabolic screening using HPLC‐MS data associated with a 114‐strain collection (58 Photorhabdus and 56 Xenorhabdus) across Thailand and explore the metabolic variation among the strains, matched with several abiotic factors. We utilize machine learning in order to rank the importance of individual metabolites in determining all given metadata. With this approach, we were able to prioritize metabolites in the context of natural product investigations, leading to the identification of previously unknown compounds. The top three highest ranking features were associated with Xenorhabdus and attributed to the same chemical entity, cyclo(tetrahydroxybutyrate). This work also addresses the need for prioritization in high‐throughput metabolomic studies and demonstrates the viability of such an approach in future research. [ABSTRACT FROM AUTHOR]

We investigated the nocturnal activity of cave-dwelling sand flies at different time intervals and determined their species composition and seasonal variation. Sand flies were captured on one night each month using CDC light traps from 18:00-06:00 with the collecting bag being changed every two h between February, 2010 and January, 2011. A total of 18,709 individuals, including 10,740 males and 7,969 females, was collected. The overall ratio between male and female specimens was 1:0.74. The collected specimens included 14 species from four genera, Chinius, Idiophlebotomus, Phlebotomus, and Sergentomyia. Sergentomyia phadangensis was the most abundant species (comprising 31.9% of the collected individuals), followed by Se. anodontis (22.8%) and Ph. mascomai (18.2%). The highest number of specimens was collected in July (15.6%), followed by May (15.5%) with the peak of collection recorded at the time interval of 00:01-02:00, followed by 22:01-00:00. However, there were no significant differences observed among time intervals of sand fly collections (p=0.154). Observations of the nocturnal activity of male and female sand flies throughout the night suggest that phlebotomine sand flies show the greatest activity level after midnight. [ABSTRACT FROM AUTHOR]

Entomopathogenic nematodes (EPNs) that are symbiotically associated with Xenorhabdus and Photorhabdus bacteria can kill target insects via direct infection and toxin action. There are limited reports identifying such organisms in the National Park of Thailand. Therefore, the objectives of this study were to identify EPNs and symbiotic bacteria from Nam Nao National Park, Phetchabun Province, Thailand and to evaluate the larvicidal activity of bacteria against Aedes aegypti and Ae. albopictus. A total of 12 EPN isolates belonging to Steinernema and Heterorhabditis were obtained form 940 soil samples between February 2014 and July 2016. EPNs were molecularly identified as S. websteri (10 isolates) and H. baujardi (2 isolates). Symbiotic bacteria were isolated from EPNs and molecularly identified as P. luminescens subsp. akhurstii (13 isolates), X. stockiae (11 isolates), X. vietnamensis (2 isolates) and X. japonica (1 isolate). For the bioassay, bacterial suspensions were evaluated for toxicity against third to early fourth instar larvae of Aedes spp. The larvae of both Aedes species were orally susceptible to symbiotic bacteria. The highest larval mortality of Ae. aegypti was 99% after exposure to X. stockiae (bNN112.3_TH) at 96 h, and the highest mortality of Ae. albopictus was 98% after exposure to P. luminescens subsp. akhurstii (bNN121.4_TH) at 96 h. In contrast to the control groups (Escherichia coli and distilled water), the mortality rate of both mosquito larvae ranged between 0 and 7% at 72 h. Here, we report the first observation of X. vietnamensis in Thailand. Additionally, we report the first observation of P. luminescens subsp. akhurstii associated with H. baujardi in Thailand. X. stockiae has potential to be a biocontrol agent for mosquitoes. This investigation provides a survey of the basic diversity of EPNs and symbiotic bacteria in the National Park of Thailand, and it is a bacterial resource for further studies of bioactive compounds. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the efficacy of symbiotic bacteria, Xenorhabdus indica, Xenorhabdus stockiae, Photorhabdus luminescens subs p. akhurstii and Photorhabdus luminescens subsp. hainanensis as a larvicide against Aedes aegypti and Aedes albopictus . Methods: Larvae (L3-L4) of Aedes aegypti and Aedes albopictus were given 2 mL of a suspension 107-108CFU/mL of each symbiotic bacterium. Distilled water and Escherichia coli ATCC®25922 were used as the control. The mortality rate of the larval mosquitoes was observed at 24, 48, 72 and 96 h. The experiment was performed in triplicates. Results: The larvae of both Aedes species started to die at 24 h exposure. Aedes aegypti showed the highest mortality rate (87%-99%), 96 h after exposure to Xenorhabdus stockiae (bNBP22.2_TH). The mortality rate of Aedes albopictus was between 82%and 96%at 96 h after exposure to Xenorhabdus indica (bKK26.2_TH). Low effectiveness of distilled water and Escherichia coli ATCC®25922 were observed in both Aedes larvae, with a mortality rate of 2%to 12%. Conclusions: The study confirms the oral toxicity of Xenorhabdus and Photorhabdus bacteria against Aedes spp. Xenorhabdus stockiae and Xenorhabdus indica may be an alternative agent for control Aedes spp. This is basic information for further study on the mechanism of action on Aedes larvae or application to control mosquito larvae in the community. [ABSTRACT FROM AUTHOR]

Hemipyrellia ligurriens (Diptera: Calliphoridae) is a forensically important blow fly species presented in many countries. In this study, we determined the morphology of all stages and the developmental rate of H. ligurriens reared under natural ambient conditions in Phitsanulok province, northern Thailand. Morphological features of all stages based on observing under a light microscope were described and demonstrated in order to use for identification purpose. Moreover, development time in each stage was given. The developmental time of H. ligurriens to complete metamorphosis; from egg, larva, pupa to adult, took 270.71 h for 1 cycle of development. The results from this study may be useful not only for application in forensic investigation, but also for study in its biology in the future. [ABSTRACT FROM AUTHOR]

• The prevalence of entomopathogenic nematode is 5.1% in agricultural soil of northern thailand. • Steinernema guangdongense, Steinernema lamjungense, Steinernema surkhetense, Steinernema siamkayai, Heterorhabditis bacteriophora, and Heterorhabditis indica are molecularly identified. • Heterorhabditis bacteriophora shows the highest efficacy against Aedes aegypti and Culex quinquefasciatus. Entomopathogenic nematodes (EPNs) of the genera Steinernema and Heterorhabditis have been considered to be effective biological control agents for several insects. In this study, we isolated and identified EPNs from soil samples in agricultural areas of northern Thailand and evaluated their efficacy for controlling larvae of three mosquito vector species, Aedes aegypti, Ae. albopictus and Culex quinquefasciatus. A total of 51 of 1,000 soil samples were positive (5.1% prevalence) for EPNs, which were identified through sequencing of the rDNA and ITS to 37 Steinernema isolates (3.7%) and 14 Heterorhabditis isolates (1.4%). For the bioassay, the larvae of mosquitoes were exposed to Steinernema surkhetense (eALN6.3_TH), Steinernema lamjungense (eALN11.5_TH), Heterorhabditis indica (eACM14.2_TH) and Heterorhabditis bacteriophora (eALN18.2_TH). Heterorhabditis bacteriophora showed the highest efficacy against Ae. aegypti and Cx. quinquefasciatus. At 96 h after exposure, the mortality rates were 60.0 and 91.7%, respectively. The EPNs were observed in the dead mosquito larvae, which were mostly found in the thorax followed by the head and abdomen. Some EPNs were dead with melanization, and some were able to survive in the cavity of mosquito larvae. Our results show the low prevalence of EPN in agricultural areas of Thailand. Moreover, H. bacteriophora may be considered an alternative biocontrol agent for managing and controlling these vector mosquitoes. [Display omitted] [ABSTRACT FROM AUTHOR]

The giant African land snail, Achatina fulica , is a pulmonate land snail of the Achatinidae family. The snail is native of East Africa and has spread to tropical and subtropical areas in different parts of the world. Achatina fulica is an important agricultural pests and the intermediate host for Angiostrongylus spp., especially Angiostrongylus cantonensis , which causes eosinophilic meningoencephalitis in humans. However, information on the genetic diversity and phylogenetic relationships among A. fulica populations in Thailand is limited. The goal of this study was to evaluate the genetic diversity and analyze haplotype network relationships between A. fulica populations in Thailand and other areas of the world. We collected A. fulica from 22 provinces in 6 regions of Thailand and analyzed a phylogenetic tree and haplotype network based on a fragment of 239 base pairs of the 16S rRNA sequence. The maximum likelihood, neighbor joining, and Bayesian inference trees for the 133 A. fulica 16S rRNA sequences revealed only one group, closely related only to A. fulica haplotype C , in all regions of Thailand. The haplotype network analysis showed that haplotype C is associated with F, P, E, Q, H and D. The genetic distance between the haplotype and A. fulica isolates in Thailand varies from 0 and 0.024, with a total of 21 variable sites. Haplotype C is predominant in Thailand and associated with other haplotypes from several areas. As inferred from 16S rDNA sequences, this study demonstrated low genetic diversity in A. fulica in Thailand. Low genetic diversity in the population can increase susceptibility to A. cantonensis infection, which may be associated with the current distribution of A. cantonensis. [Display omitted] • We present low genetic diversity of Achatina fulica based on mitochondrial 16S rRNA gene. • Haplotype network indicates similarity of genetic diversity of A. fulica, in all the regions of Thailand. • Haplotype C of A. fulica have dispersed across Thailand and several regions worldwide. • Expansion of the A. fulica population in Thailand may facilitate the spread of A. cantonensis to all areas of the country. [ABSTRACT FROM AUTHOR]

A new species of entomopathogenic nematode, Steinernema adamsi n. sp., was recovered from the soil of a longan tree (Dimocarpus sp.) in Mueang Lamphun District, Thailand, using baiting techniques. Upon analysis of the nematode's morphological traits, we found it to be a new species of Steinernema and a member of the Longicaudatum clade. Molecular analyses of the ITS rDNA and D2D3 of 28S rDNA sequences further confirmed that S. adamsi n. sp. is a new species of the Longicaudatum clade, which is closely related to Steinernema guangdongense and Steinernema longicaudam. Using morphometric analysis, the infective juveniles measure between 774.69 and 956.96 μm, males have a size range of 905.44 to 1,281.98 μm, and females are within the range of 1,628.21 to 2,803.64 μm. We also identified the symbiotic bacteria associated with the nematode based on 16S sequences as Xenorhabdus spp. closely related toXenorhabdus griffiniae. Furthermore, we have successfully assessed a cryopreservation method for the long-term preservation of S. adamsi n. sp. Successful cryopreservation of this new species will allow for the longer preservation of its traits and will be valuable for its future use. The discovery of this new species has significant implications for the development of effective biological control agents in Thailand, and our work contributes to our understanding of the diversity and evolution of entomopathogenic nematodes., (© American Society of Parasitologists 2023.)

Entomopathogenic nematodes (EPNs) are insect parasitic nematodes of the genera Het-erorhabditis and Steinernema . These nematodes are symbiotically associated with the bacteria, Photorhabdus and Xenorhabdus, respectively. National parks in Thailand are a potentially rich resource for recovering native EPNs and their symbiotic bacteria. The objectives of this study are to isolate and identify EPNs and their bacterial flora from soil samples in four national parks in Thailand and to evaluate their efficacy for controlling mosquito larvae. Using a baiting method with a Galleria mellonella moth larvae and a White trap technique, 80 out of 840 soil samples (9.5%) from 168 field sites were positive for EPNs. Sequencing of an internal transcribed spacer resulted in the molecular identification of Heterorhabditis nematode isolates as H. indica , H. baujardi and Heterorhabditis SGmg3, while using 28S rDNA sequencing, Steinernema nematode species were identified as S. guang-dongense , S. surkhetense , S. minutum , S. longicaudum and one closely related to S. yirgalemense . For the symbiotic bacterial isolates, based on recA sequencing, the Photorhabdus spp. were identified as P. luminescens subsp. akhurstii , P. luminescens subsp. hainanensis and P. luminescens subsp. australis . Xenorhabdus isolates were identified as X. stockiae , X. indica , X. griffiniae , X. japonica and X. hominickii . Results of bioassays demonstrate that Photorhabdus isolates were effective on both Aedes aegypti and Culex quinquefasciatus . Therefore, we conclude that soil from Thailand's national parks contain a high diversity of entomopathogenic nematodes and their symbiotic bacteria. Photorhabdus bacteria are larvicidal against culicine mosquitoes and may serve as effective biocontrol agents.

The rat lungworm Angiostrongylus cantonensis is globally known to be the cause of oeosinophilic meningitis in humans. Another congener, Angiostrongylus malaysiensis , is closely related to A. cantonensis and has been described as a potential human pathogenic parasite. These 2 worms are similar in terms of life cycle, host range and morphological and genetic information. However, there are limited studies about their genetic diversity based on the 66-kDa protein-encoding gene. The objective of this study was to explore the 66-kDa protein sequence variation of A. cantonensis and A. malaysiensis collected from Thailand. Two adult and 53 third-stage larval specimens of Angiostrongylus from 4 geographic locations in Thailand were molecularly identified using the 66-kDa protein gene. The phylogenetic trees (Bayesian inference tree and maximum-likelihood tree) showed that Angiostrongylus formed a monophyletic clade with a clear separation between A. cantonensis and A. malaysiensis . The genetic distance between A. cantonensis and A. malaysiensis varies from 0.82 to 2.86%, with a total of 16 variable sites. The analysis of genetic diversity revealed 1 and 5 new haplotypes of A. cantonensis and A. malaysiensis , respectively, and showed genetic differences between the populations of A. cantonensis and A. malaysiensis . The haplotype networks of A. cantonensis and A. malaysiensis populations in Thailand are similar to those of populations in some countries, indicating the range expansion of genomic origin between populations in different areas. In conclusion, the 66-kDa protein gene was a good genetic marker for studying genetic diversity and discriminating between A. cantonensis and A. malaysiensis .

Pomacea is a freshwater snail in family Ampullariidae that is native to South and Central America. This snail is among the more important intermediate hosts for Angiostrongylus cantonensis and agricultural pests. Herein, we investigated the prevalence of A. cantonensis larvae and the genetic diversity of Pomacea samples collected across Thailand based on mitochondrial cytochrome c oxidase subunit I ( COI ) gene sequences. The larval-infection rate was 1.7% in Pomacea canaliculata specimens collected from the Uttaradit Province of northern Thailand. We randomly selected specimens of P. canaliculata and P. maculata for genetic analysis. We analyzed 244 COI sequences, including 49 sequences from samples collected from Thailand and a publicly accessible database of snails in their native and non-native ranges. A maximum-likelihood tree of P. canaliculata and P. maculata revealed two main clades. The genetic diversity analysis identified seven P. canaliculata haplotypes and six P. maculata haplotypes, and showed genetic differences between the populations of P. canaliculata and P. maculata . The haplotype networks of P. canaliculata and P. maculata populations in Thailand are similar to those of populations in multiple countries, indicating that this species spread widely to many parts of the world.

Background: Aedes aegypti is a potential vector of West Nile, Japanese encephalitis, chikungunya, dengue and Zika viruses. Alternative control measurements of the vector are needed to overcome the problems of environmental contamination and chemical resistance. Xenorhabdus and Photorhabdus are symbionts in the intestine of entomopathogenic nematodes (EPNs) Steinernema spp. and Heterorhabditis spp. These bacteria are able to produce a broad range of bioactive compounds including antimicrobial, antiparasitic, cytotoxic and insecticidal compounds. The objectives of this study were to identify Xenorhabdus and Photorhabdus isolated from EPNs in upper northern Thailand and to study their larvicidal activity against Ae. aegypti larvae., Results: A total of 60 isolates of symbiotic bacteria isolated from EPNs consisted of Xenorhabdus (32 isolates) and Photorhabdus (28 isolates). Based on recA gene sequencing, BLASTN and phylogenetic analysis, 27 isolates of Xenorhabdus were identical and closely related to X. stockiae, 4 isolates were identical to X. miraniensis, and one isolate was identical to X. ehlersii. Twenty-seven isolates of Photorhabdus were closely related to P. luminescens akhurstii and P. luminescens hainanensis, and only one isolate was identical and closely related to P. luminescens laumondii. Xenorhabdus and Photorhabdus were lethal to Ae aegypti larvae. Xenorhabdus ehlersii bMH9.2&#95;TH showed 100% efficiency for killing larvae of both fed and unfed conditions, the highest for control of Ae. aegypti larvae and X. stockiae (bLPA18.4&#95;TH) was likely to be effective in killing Ae. aegypti larvae given the mortality rates above 60% at 72 h and 96 h., Conclusions: The common species in the study area are X. stockiae, P. luminescens akhurstii, and P. luminescens hainanensis. Three symbiotic associations identified included P. luminescens akhurstii-H. gerrardi, P. luminescens hainanensis-H. gerrardi and X. ehlersii-S. Scarabaei which are new observations of importance to our knowledge of the biodiversity of, and relationships between, EPNs and their symbiotic bacteria. Based on the biological assay, X. ehlersii bMH9.2&#95;TH begins to kill Ae. aegypti larvae within 48 h and has the most potential as a pathogen to the larvae. These data indicate that X. ehlersii may be an alternative biological control agent for Ae. aegypti and other mosquitoes.

Photorhabdus and Xenorhabdus are symbiotic with entomopathogenic nematodes (EPNs) of the genera Heterorhabditis and Steinernema , respectively. These bacteria produce several secondary metabolites including antimicrobial compounds. The objectives of this study were to isolate and identify EPNs and their symbiotic bacteria from Mae Wong National Park, Thailand and to evaluate the antibacterial activities of symbiont extracts against drug resistant bacteria. A total of 550 soil samples from 110 sites were collected between August 2014 and July 2015. A total of EPN isolates were obtained through baiting and White trap methods, which yielded 21 Heterorhabditis and 3 Steinernema isolates. Based on molecular identification and phylogenetic analysis, the most common species found in the present study was P. luminescens subsp. akhurstii associated with H. indica . Notably, two species of EPNs, H. zealandica and S. kushidai , and two species of symbiotic bacteria, X. japonica and P. temperata subsp. temperata represented new recorded organisms in Thailand. Furthermore, the association between P. temperata subsp. temperata and H. zealandica has not previously been reported worldwide. Disk diffusion, minimal inhibitory concentration, and minimal bactericidal concentration analyses demonstrated that the crude compound extracted by ethyl acetate from P. temperata subsp. temperata could inhibit the growth of up to 10 strains of drug resistant bacteria. Based on HPLC-MS analysis, compound classes in bacterial extracts were identified as GameXPeptide, xenoamicin, xenocoumacin, mevalagmapeptide phurealipids derivatives, and isopropylstilbene. Together, the results of this study provide evidence for the diversity of EPNs and their symbiotic bacteria in Mae Wong National Park, Thailand and demonstrate their novel associations. These findings also provide an important foundation for further research regarding the antimicrobial activity of Photorhabdus bacteria.

Entomopathogenic nematodes (EPNs) are used successfully for biological control of subterranean larval pests leading to reduced environmental contamination if chemical control measures are employed. Their diversity and distribution in Thailand are unclear, so the present study sought to obtain a better understanding these EPN populations in the lower northern region of Thailand. We collected 930 soil samples from 186 sites of Kamphaeng Phet, Nakhon Sawan, Phetchabun, Phichit, Phitsanulok, Sukhothai, Tak, Uthai Thani, and Uttaradit Provinces, Thailand from December 2011 to November 2012. Galleria mellonella was used as host for isolating and propagating EPNs. Seventy soil samples (7.5%) yielded EPNs of two genera, Steinernema (3.0%) and Heterorhabditis (4.5%). The majority of the isolated EPNs were found in loam at 26°C-33°C and pH values of 5.0-7.0. Molecular identification from partial 28S rDNA sequences revealed S. websteri, isolated from soil samples from Nakhon Sawan and Uthai Thani. Phylogenetic analysis of these EPNs showed they are closely related to S. websteri JC1032. The identification that S. websteri was the predominant EPN should enable its application for biological control in the local prevailing soil conditions.

The aim of this study was to find novel proteins expressed from an Angiostrongylus cantonensis adult female worm cDNA library for serodiagnosis of angiostrongyliasis. An immuno-dominant clone, fAC22, was identified by immunoscreening with pooled positive sera from proven angiostrongyliasis patients. The clone contained an open reading frame of 2,136 bp encoding a 80.5 kDa protein with a predicted isoelectric point of 5.8. The deduced amino acid sequence (712 amino acids) contained the conserved domain of Small mutS related (Smr) superfamily protein, with similarity with the Smr domain protein of Brugia malayi. The fusion His-tagged 81 kDa recombinant protein expressed as inclusion body in Escherichia coli was solubilized and purified by Ni-affinity chromatography for use in immunoblot analysis. Its sensitivity, specificity, positive and negative predictive values in immunodiagnostic test was 93.5, 91.5, 79.0 and 97.5%, respectively. Although some cross-reactivity of the antigen was observed among gnathostomiasis, bancroftian filariasis, ascariasis, echinococcosis, paragonimiasis and opisthorchiasis, sera from 14 other infections were all negative. These data indicate its possible application in immunodiagnosis of clinically suspected angiostrongyliasis. Key words: Angiostrongylus cantonensis,eosinophilic meningitis, recombinant fusion protein, immunodiagnosis