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. 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_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_TH) was likely to be effective in killing Ae. aegypti larvae given the mortality rates above 60% at 72 h and 96 h. 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_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.

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.

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, 2 species of EPNs, H. zealandica and S. kushidai, and 2 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. Disc 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.

Objective: To evaluate the efficacy of symbiotic bacteria, Xenorhabdus indica, Xenorhabdus stockiae, Photorhabdus luminescens subsp. 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 10⁷-10⁸ CFU/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.

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 ofH. ligurriensreared 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 ofH. ligurriensto 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.

To survey the Angiostrongylus cantonensis (A. cantonensis) or the rat lungworm in a rat, definitive host, and in a giant African land snail (Achatina fulica), the intermediate host, in Phitsanulok, Thailand. Rats and giant African land snails were captured from Tha Pho sub-district, Phitsanulok, Thailand. Rats were killed and examined for adult A. cantonensis. The artificial digestion method following Baermann technique were used for isolation third stage larvae of A. cantonensis. Sixty-two rats were captured and they were identified as Rattus argentiventer, Rattus rattus (R. rattus), Bandicota savilei, and Bandicota indica but only one animal (R. rattus) of 62 rats (1.61%) was positive with adult worm of A. cantonensis. The third stage larvae of A. cantonensis were examined on 307 Angiostrongylus fulica snails. It was found that the overall infection rate was 12.38% (38 infected out of 307 Achatina snails). This study demonstrates that A. cantonensis is available in the natural hosts of Phitsanulok. This suggests that the transmissions of this parasite to human may occur in this region.