Ocular Brugia pahangi Filariasis Complicated by Severe Macular Damage in Thailand: Case Report and Literature Review.

Extralymphatic filariasis caused by filaria of zoonotic origins has been frequently reported in Thailand over recent years. Here, we report the first case of ocular filariasis in a 7.5-year-old Thai boy who initially presented with progressive conjunctival redness and blurred vision in his right eye. A small, slender, coiled worm was found and surgically removed from the right anterior chamber. Histopathological examination illustrated predominant eosinophilic inflammation surrounding the parasite, which showed smooth and thin cuticle, prominent lateral chords, flat and broad muscle cells, one intestine, and two reproductive tubes with unsegmented ova, typically characteristic of a female adult Brugia filarial nematode. The parasite was also molecularly identified as B. pahangi, based on mitochondrial cytochrome c oxidase subunit I sequence analysis. The patient was then empirically prescribed albendazole, systemic prednisolone, and topical methylprednisolone. Unfortunately, his vision did not recover after 2 months due to severe maculopathy, most likely resulting from parasitic infestation and subsequent vitreous inflammation. To the best of our knowledge, this is the first case of ocular infestation by B. pahangi with visual complications that occurred outside a filariasis-endemic area of Thailand. Furthermore, this report provides clinical data on preceding cases of B. pahangi filariasis formally reported in southeast Asian countries, including Thailand and Malaysia, which facilitate a better understanding of the epidemiology of this sporadic zoonotic infection for effective disease elimination.

Molecular Identification of Host Blood Meals and Detection of Blood Parasites in Culicoides Latreille (Diptera: Ceratopogonidae) Collected from Phatthalung Province, Southern Thailand.

Five hundred and fifty-nine female biting midges were collected, and seventeen species in six subgenera (Avaritia, Haemophoructus, Hoffmania, Meijerehelea, Remmia, and Trithecoides) and two groups (Clavipalpis and Shortti) were identified. The dominant Culicoides species was C. peregrinus (30.94%), followed by C. subgenus Trithecoides. From blood meal analysis of engorged biting midges, they were found to feed on cows, dogs, pigs, and avians. The majority of blood preferences of biting midges (68%; 49/72) displayed a mixed pattern of host blood DNA (cow and avian). The overall non-engorged biting midge field infectivity rate was 1.44 % (7/487). We detected Leucocytozoon sp. in three Culicoides specimens, one from each species: C. fulvus, C. oxystoma, and C. subgenus Trithecoides. Crithidia sp. was found in two C. peregrinus specimens, and Trypanosoma sp. and P. juxtanucleare were separately found in two C. guttifer. More consideration should be paid to the capacity of biting midges to transmit pathogens such as avian haemosporidian and trypanosomatid parasites. To demonstrate that these biting midges are natural vectors of trypanosomatid parasites, additional research must be conducted with a greater number of biting midges in other endemic regions.

Development and evaluation of a visible reverse transcription-loop-mediated isothermal amplification (RT-LAMP) for the detection of Asian lineage ZIKV in field-caught mosquitoes.

The Zika virus (ZIKV) infection is an emerging and re-emerging arbovirus infection that is transmitted to humans through the bite of infected mosquitoes. Early detection of ZIKV in mosquitoes is one of the prerequisite approaches for tracking the spread of the virus. Therefore, this study aims to develop and validate a visual reverse transcription-loop-mediated isothermal amplification (RT-LAMP) method called ZIKV-RT-LAMP, for detecting ZIKV in field collected mosquito samples from Thailand. A single-tube ZIKV-RT-LAMP assay was developed to detect Asian lineage ZIKV RNA. The detection limit and cross-reactivity of ZIKV were investigated. The hemi-nested RT-PCR (hn-RT-PCR) and the colorimetric LAMP kit (cLAMP kit) were performed as reference assays. The detection limit of the ZIKV-RT-LAMP assay was 10-6 ffu/ml or pfu/ml, making it highly specific and 100 times more sensitive than the hn-RT-PCR and cLAMP kits. The ZIKV-RT-LAMP assay detected the Asian lineage of ZIKV RNA without cross-reactivity with other arthropod-borne viruses. The sensitivity and specificity of the ZIKV-RT-LAMP assay were 92.31% and 100%, respectively. The ZIKV-RT-LAMP is a simple, rapid, and inexpensive method for detecting ZIKV in field-caught mosquitos. In the future, extensive surveys of field-caught mosquito populations should be conducted. Early detection of ZIKV in field-caught mosquitoes provides for prompt and effective implementation of mosquito control strategies in endemic areas.

Genetic characterization of chikungunya virus isolates from Aedes aegypti mosquitoes collected during a recent outbreak in Bangkok, Thailand.

Chikungunya virus (CHIKV) is a mosquito-borne emerging pathogen that is transmitted to humans through the bite of female Aedes mosquitoes. CHIKV infection has become a major public health concern worldwide, as it has a significant impact on the healthcare system. Since 2004, the virus has emerged in Africa and subsequently spread to countries located near the Indian Ocean, including India, and to Europe, the Americas, and Asia. In Thailand, a large CHIKV outbreak occurred during 2008-2009 and was caused by a virus originating from the east/central/south African (ECSA) CHIKV genotype. Since then, the ECSA genotype of CHIKV has continued to circulate and has caused sporadic cases in different areas in Thailand. Approximately 20,000 reported cases have been confirmed by the Bureau of Epidemiology, Ministry of Public Health, Thailand, from January 1, 2018 to July 31, 2020. However, the causes of this CHIKV re-emergence remain unclear. To obtain a better understanding of CHIKV circulation during the recent outbreak in Bangkok, Thailand, complete genome analysis of CHIKV isolates from field-caught mosquitoes collected in outbreak areas was performed. A total of 28 Ae. aegypti samples (21 females and 7 males) were collected, and individual mosquitoes were used for CHIKV detection and isolation. Eleven of 28 (39.29%) female and three of 28 (10.71%) male mosquitoes were positive for CHIKV by E1 nested RT-PCR. Four CHIKV isolates were successfully isolated from four female Ae. aegypti mosquitoes. Based on complete genome analysis, several amino acid substitutions were identified in the protein coding region. The E1:K211E and E2:V264A mutations in the background of the E1:226A mutation were observed in all four CHIKV isolates. An important observation was the presence of one amino acid substitution, leading to an E1:K245R change. This mutation was found in all four CHIKV isolates from mosquitoes in this study and in Thai patients described previously. Additionally, phylogenetic analysis indicated that the four CHIKV isolates belonged to the Indian Ocean clade of the ECSA genotype. The results obtained in this study provide detailed information on the molecular characteristics and evolution of currently circulating CHIKV strains in Thailand, which are useful for developing prevention and control strategies.

Molecular analysis of mitochrondrial cytb of Pediculus humanus capitis in Thailand revealed potential historical connection with South Asia.

BACKGROUND: Pediculus humanus capitis or head louse is an obligate ectoparasite and its infestation remains a major public health issue worldwide. Molecular analysis divides head lice into six clades and intra-clade genetic differences have been identified. Several hypotheses have been formulated to elucidate the discrepancies of the variety of head lice among different regions of the world. It is currently concluded that head lice distribution might be associated with human migration history. This study aims to investigate genetic data of human head lice in Thailand. We believe that the analysis could help establish the correlation between local and global head lice populations. METHOD: We investigated mitochondrial cytochrome b (cytb) gene of the collected 214 head lice to evaluate genetic diversity from 15 provinces among 6 regions of Thailand. The head lice genes were added to the global pool for the phylogenetic tree, Bayesian tree, Skyline plot, and median joining network construction. The biodiversity, neutrality tests, and population genetic differentiation among the 6 Thailand geographic regions were analyzed by DNAsp version 6. RESULTS: The phylogenetic tree analysis of 214 collected head lice are of clade A and clade C accounting for roughly 65% and 35% respectively. The Bayesian tree revealed a correlation of clade diversification and ancient human dispersal timeline. In Thailand, clade A is widespread in the country. Clade C is confined to only the Central, Southern, and Northeastern regions. We identified 50 novel haplotypes. Statistical analysis showed congruent results between genetic differentiation and population migration especially with South Asia. CONCLUSIONS: Pediculosis remains problematic among children in the rural areas in Thailand. Cytb gene analysis of human head lice illustrated clade distribution and intra-clade diversity of different areas. Our study reported novel haplotypes of head lice in Thailand. Moreover, the statistic calculation provided a better understanding of their relationship with human, as an obligate human parasite and might help provide a better insight into the history of human population migration. Determination of the correlation between phylogenetic data and pediculicide resistance gene as well as residing bacteria are of interest for future studies.

Early detection of novel Leishmania species DNA in the saliva of two HIV-infected patients.

BACKGROUND: Leishmaniasis caused by two new species of Leishmania; L. siamensis and L. martiniquensis have been recently described in Thailand. The disease has mainly been documented in AIDS patients from southern Thailand. In this study, polymerase chain reaction (PCR) was used to determine HIV-Leishmania co-infection in southern Thailand. METHODS: One ml of saliva and 3 ml of EDTA blood were collected from HIV-infected patients for PCR detection of Leishmania DNA, cloning and sequencing. The positive PCR samples were then cultured on Schneider's insect medium. RESULTS: Three out of 316 saliva samples collected from HIV-infected patients were found to be positive for Leishmania DNA (0.95%). Among the positive samples, one patient was observed with disseminated cutaneous lesions and also tested positive via saliva, whole blood and buffy coat in PCR. The second case presenting with nodular lesions also gave a positive saliva test via PCR two months prior to buffy coat. This diagnosis was confirmed by microscopic examination and a culture of biopsy samples from a nodule. The last case was an asymptomatic Leishmania infection which tested PCR positive only in saliva with a consecutive sample collection conducted for three months. CONCLUSIONS: The prevalence of Leishmania infection in HIV infected patients within this study is 0.95%. Leishmania DNA was detected in saliva by PCR prior to blood and buffy coat of two HIV infected patients. Early detection of Leishmania DNA in saliva would be beneficial for the follow up of asymptomatic Leishmania infected patients, the early treatment of leishmaniasis and for surveillance survey purpose. However, full evaluation of sensitivity and specificity of this technique with a large cohort of patients is required before deployment.

Disseminated dermal leishmaniasis caused by Leishmania siamensis in a systemic steroid therapy patient.

Leishmania siamensis infection was recently reported from Thailand. Clinical presentation of L. siamensis infections is generally related to human immunodeficiency virus infection. Herein, disseminated dermal L. siamensis infection in a systemic steroid therapy patient from Myanmar is described.

Detection of Leishmania siamensis DNA in saliva by polymerase chain reaction.

Polymerase chain reaction was used to detect Leishmania siamensis DNA from clinical samples collected from six leishmaniasis patients during 2011-2012. The samples used in this study came from bone marrow, blood, buffy coat, saliva, urine, and tissue biopsy specimens. Saliva was a good source for L. siamensis DNA by polymerase chain reaction. L. siamensis DNA was also found in saliva of an asymptomatic case-patient. Levels of L. siamensis DNA in saliva decreased until being undetectable after treatment. These levels could be used as a marker to evaluate efficacy of the treatment. A larger study is needed to evaluate this method as a screening and survey tool to study the silent background of Leishmania infection among the at-risk population.

Resolution of eosinophilia after treatment of cutaneous gnathostomiasis.

The present study was to investigate the dynamics of eosinophil in peripheral blood of patients with cutaneous gnathostomiasis before and after worm removal. The total of 28 proven cases of cutaneous gnathostomiasis treated by albendazole were included in the present study. The absolute eosinophil count (AEC) was higher than 500/ul during infestation in almost all the patients, the positive rate was 89%, and significantly decreased to normal level after receiving albendazole and worm removal within 3 months in 96%. In conclusion, an increas of AEC is another important hallmarks of cutaneous gnathostomiasis and this parameter could be the earlier indicator for responsiveness to treatment.

Decrease of mosquito salivary gland proteins after a blood meal: an implication for pathogenesis of mosquito bite allergy.

Salivary gland protein profiles ofAedes aegypti (L.) and Culex quinquefasciatus (Say) pre- and post-blood feeding were analyzed. SDS-PAGE studies before blood feeding of Ae. aegypti demonstrated 8 major polypeptide bands of 20, 35, 37, 42, 45, 47, 70 kDa and a high molecular weight band >118 kDa, whereas those of Cx. quinquefasciatus demonstrated 9 major polypeptide bands of 20, 26, 36, 38, 45, 47, 49 kDa and 2 high molecular weight bands >118 kDa. After a blood feeding, salivary gland polypeptides of Ae. aegypti at 35, 37, 45, 47, 70 kDa and high molecular weight band >118 kDa were depleted, while the polypeptide bands of 20, 26, 36, 38 kDa were depleted in Cx. quinquefasciatus. The presented study suggests that these major polypeptides were introduced into vertebrate hosts when a mosquito took a blood meal. Further investigation in molecular, biochemical and immunological aspects of these salivary gland polypeptides may provide information for better understanding in the role of these proteins in mosquito bite allergy.

Treatment of cutaneous gnathostomiasis with ivermectin.

In a randomized open study, we compared the efficacy of a single dose of oral ivermectin (200 microg/kg) and oral albendazole (400 mg/day for 21 days) for the treatment of cutaneous gnathostomiasis. Thirty-one patients were randomly assigned to receive ivermectin (n = 17) or albendazole (n = 14). Thirteen of 17 patients who received ivermectin responded, 3 relapsed, and 1 was unresponsive (cure rate = 76%). Thirteen of 14 patients who received albendazole responded very well and did not relapse. Only one patient was unresponsive (cure rate = 92%; P > 0.05). No major side effects were observed in both groups. We concluded that a single dose of ivermectin (200 microg/kg) is less effective than albendazole (400 mg/day for 21 days) for treatment of cutaneous gnathostomiasis, but there was no statistically significant difference (P > 0.05).

Study of specific IgG subclass antibodies for diagnosis of Gnathostoma spinigerum.

Gnathostoma spinigerum infection is endemic in Thailand and many Asian countries. Current diagnosis is the skin test and enzyme-linked immunosorbent assay (ELISA) for IgG antibody against the G. spinigerum third-stage larvae (L3), but cross-reactivity is common. We evaluated the sensitivity and specificity of anti-G. spinigerum L3 IgG subclass antibodies for diagnosis of 43 patients with gnathostomiasis. The majority of patients with gnathostomiasis (91%) had eosinophilia. While the anti-G. spinigerum L3 IgG1 antibody provided the highest sensitivity (98%), the anti-G. spinigerum L3 IgG2 antibody had the highest specificity (88%). The ELISA that detected anti-G. spinigerum L3 IgG1 antibody could be a reliable laboratory screening test, while anti-G. spinigerum L3 IgG2 antibody could be used to confirm the diagnosis.

Prevalence of parasitic infections among Thai patients at the King Chulalongkorn Memorial Hospital, Bangkok, Thailand.

Parasitic diseases are still considered to be a major public health problem. Most patients with parasitic infections are asymptomatic and therefore remain undetected. Asymptomatic parasitic infections are usually discovered by routine parasite examination. To determine the result of parasite examination at the Parasitology Unit, Out Patient Department, King Chulalongkorn Memorial Hospital, Bangkok, Thailand, the authors collected the data of individuals examined for parasite infections from June to December 1997. A total of 6,231 Thais provided the data for analysis. Evidence of parasitic infections was found in 557 (8.94%) cases. The disease was most prevalent in males (57.3%), and in the age group >15-30 years old (11.13%). The population from the Northeast of Thailand was found to harbor parasites with the highest prevalence rate (17.03%), while it was 11.90 per cent in the northern group. The parasitic prevalence rates in the West, East, South and Central regions were 10.60 per cent, 8.90 per cent, 7.74 per cent, and 4.92 per cent, respectively. The parasite most commonly identified was Strongyloides stercoralis (33.39%), while giardiasis was the most common protozoan infection (14.36%). The highest infection rates of S. stercoralis, hookworms, Opisthorchis viverrini, and Gnathostoma spinigerum were found in northeasterners. People from the North of Thailand were infected mostly with G. lamblia. People of working-age from northeastern as well as northern regions harbored pathogenic parasites with high prevalence rates. To prevent parasitic infections, health education for these high risk groups should be provided.

Prevalence and intensity of third stage Gnathostoma spinigerum larvae in swamp eels sold in three large markets in Bangkok, Thailand.

Gnathostoma spinigerum is a common human tissue parasite in Thailand. The swamp eel is the major intermediate or paratenic host for this parasite. The high prevalence of the infective third stage larvae (L3) of this parasite in the rainy season has been noted in previous studies. During June 1999 (rainy season), we performed a cross-sectional survey of the prevalence and the intensity of G. spinigerum L3s in the livers of swamp eels that were obtained from three large Bangkok markets (Klong Toey, Pran Nok, and Tevej). Of a total of 785 livers, G. spinigerum L3s were found in 97: an infection rate of 12%. The prevalence rates in Klong Toey, Pran Nok, and Tevej markets were 13%, 10% and 14% respectively. There was no significant difference in the proportion of infected eels between the markets (p > 0.05). The intensity of L3 burden in the livers ranged from 1-17 larvae, with an average of 2.60 +/- 0.24 larvae (mean +/- SE). In this study a high prevalence of G. spinigerum L3s was found; there was no significant difference in the intensity of larvae in swamp eels between markets. These findings emphasize the importance of public health education: people need to be aware of the dangers of consuming raw or undercooked food.