Malaria remains one of the most important communicable diseases. A rapid, simple and accurate method is a crucial part of malaria diagnosis. The aim of this study was to reevaluate the microwave irradiation method to extract DNA from Plasmodium falciparum and compare with six other existing DNA extraction methods such as QIAamp DNA mini kit (Qiagen), FTA elute card, phenol-chloroform, Chelex, Chelex without proteinase-K and Rapid boiling. Two different P. falciparum isolates were used: (i) Laboratory strains with 0.3% parasitemia and (ii) clinical isolate with 0.6% parasitemia. Each DNA extraction method was validated for the presence of P. falciparum by a routine nested and real time PCR. In order to evaluate the sensitivity of the DNA extraction by microwave, double serial dilution of P. falciparum from in vitro culture at parasitemia that ranged from 0.0001 to 0.17% were used to extract the DNA by microwave and the P. falciparum DNA was then detected by nested and real-time PCR. The nested and real-time PCR were able to detect. P. falciparum DNA at the parasitemia level as low as 0.0003% and 0.0001%, respectively. Our results can reproduce the results from earlier studies and reveal microwave as a rapid and simple tool to extract P. falciparum DNA and subsequent molecular diagnosis of malaria.

OBJECTIVES: For the first time, Boliwong, an indigenous community in the Philippines, was surveyed for the prevalence of Cryptosporidium from April to December 2017. METHODS: Cryptosporidium oocysts were detected in samples from the river, creek, and water pumps via immunomagnetic separation techniques, and from human and animal concentrated faecal samples using the modified Ziehl-Neelsen technique. RESULTS: Seven of the 24 water samples (29.2%) were positive for Cryptosporidium, with the highest concentration (0.8 oocyst/L) detected in the creek. Of 35 fecal samples from different animal groups, 8 (21.6%) were positive for Cryptosporidium oocysts. The highest intensity of oocyst shedding was detected in dogs (χ2=8.00). Of the 137 human fecal samples, 39 (28.5%) were infected with Cryptosporidium. In this study, 3 risk factors were found to be associated with infection: (1) location (crude odds ratio [cOR], 16.39; 95% confidence interval [CI], 2.11 to 127.41; p=0.008), (2) drinking water from the natural spring (cOR, 0.29; 95% CI, 0.11 to 0.82; p<0.05), and (3) using an open pit as a sanitary toilet facility (cOR, 2.44; 95% CI, 1.14 to 5.20; p<0.05). When the cOR was adjusted, using an open pit as a sanitary toilet facility remained a significant risk factor of infection (adjusted OR, 0.41; 95% CI, 0.19 to 0.90; p<0.05). CONCLUSIONS: There is a potentially emerging Cryptosporidium zoonosis in Boliwong, Lagawe, Philippines. It is recommended that the toilet facilities and the water system in the community be rehabilitated to avoid any possible disease outbreak. Health education is also needed in the community to maintain proper hygiene and sanitation practices.
Animals ; Cryptosporidium* ; Disease Outbreaks ; Dogs ; Drinking Water ; Epidemiology ; Health Education ; Humans ; Hygiene ; Immunomagnetic Separation ; Natural Springs ; Odds Ratio ; Oocysts ; Philippines* ; Prevalence ; Public Health ; Risk Factors ; Rivers ; Sanitation ; Toilet Facilities ; Water

OBJECTIVES: For the first time, Boliwong, an indigenous community in the Philippines, was surveyed for the prevalence of Cryptosporidium from April to December 2017.METHODS: Cryptosporidium oocysts were detected in samples from the river, creek, and water pumps via immunomagnetic separation techniques, and from human and animal concentrated faecal samples using the modified Ziehl-Neelsen technique.RESULTS: Seven of the 24 water samples (29.2%) were positive for Cryptosporidium, with the highest concentration (0.8 oocyst/L) detected in the creek. Of 35 fecal samples from different animal groups, 8 (21.6%) were positive for Cryptosporidium oocysts. The highest intensity of oocyst shedding was detected in dogs (χ2=8.00). Of the 137 human fecal samples, 39 (28.5%) were infected with Cryptosporidium. In this study, 3 risk factors were found to be associated with infection: (1) location (crude odds ratio [cOR], 16.39; 95% confidence interval [CI], 2.11 to 127.41; p=0.008), (2) drinking water from the natural spring (cOR, 0.29; 95% CI, 0.11 to 0.82; p<0.05), and (3) using an open pit as a sanitary toilet facility (cOR, 2.44; 95% CI, 1.14 to 5.20; p<0.05). When the cOR was adjusted, using an open pit as a sanitary toilet facility remained a significant risk factor of infection (adjusted OR, 0.41; 95% CI, 0.19 to 0.90; p<0.05).CONCLUSIONS: There is a potentially emerging Cryptosporidium zoonosis in Boliwong, Lagawe, Philippines. It is recommended that the toilet facilities and the water system in the community be rehabilitated to avoid any possible disease outbreak. Health education is also needed in the community to maintain proper hygiene and sanitation practices.
Animals ; Cryptosporidium ; Disease Outbreaks ; Dogs ; Drinking Water ; Epidemiology ; Health Education ; Humans ; Hygiene ; Immunomagnetic Separation ; Natural Springs ; Odds Ratio ; Oocysts ; Philippines ; Prevalence ; Public Health ; Risk Factors ; Rivers ; Sanitation ; Toilet Facilities ; Water

OBJECTIVES: For the first time, Boliwong, an indigenous community in the Philippines, was surveyed for the prevalence of Cryptosporidium from April to December 2017. METHODS: Cryptosporidium oocysts were detected in samples from the river, creek, and water pumps via immunomagnetic separation techniques, and from human and animal concentrated faecal samples using the modified Ziehl-Neelsen technique. RESULTS: Seven of the 24 water samples (29.2%) were positive for Cryptosporidium, with the highest concentration (0.8 oocyst/L) detected in the creek. Of 35 fecal samples from different animal groups, 8 (21.6%) were positive for Cryptosporidium oocysts. The highest intensity of oocyst shedding was detected in dogs (χ2=8.00). Of the 137 human fecal samples, 39 (28.5%) were infected with Cryptosporidium. In this study, 3 risk factors were found to be associated with infection: (1) location (crude odds ratio [cOR], 16.39; 95% confidence interval [CI], 2.11 to 127.41; p=0.008), (2) drinking water from the natural spring (cOR, 0.29; 95% CI, 0.11 to 0.82; p<0.05), and (3) using an open pit as a sanitary toilet facility (cOR, 2.44; 95% CI, 1.14 to 5.20; p<0.05). When the cOR was adjusted, using an open pit as a sanitary toilet facility remained a significant risk factor of infection (adjusted OR, 0.41; 95% CI, 0.19 to 0.90; p<0.05). CONCLUSIONS There is a potentially emerging Cryptosporidium zoonosis in Boliwong, Lagawe, Philippines. It is recommended that the toilet facilities and the water system in the community be rehabilitated to avoid any possible disease outbreak. Health education is also needed in the community to maintain proper hygiene and sanitation practices.

Acanthamoeba, one of free-living amoebae (FLA), remains a high risk of direct contact with this protozoan parasite which is ubiquitous in nature and man-made environment. This pathogenic FLA can cause sight-threatening amoebic keratitis (AK) and fatal granulomatous amoebic encephalitis (GAE) though these cases may not commonly be reported in our clinical settings. Acanthamoeba has been detected from different environmental sources namely; soil, water, hot-spring, swimming pool, air-conditioner, or contact lens storage cases. The identification of Acanthamoeba is based on morphological appearance and molecular techniques using PCR and DNA sequencing for clinico-epidemiological purposes. Recent treatments have long been ineffective against Acanthamoeba cyst, novel anti-Acanthamoeba agents have therefore been extensively investigated. There are efforts to utilize synthetic chemicals, lead compounds from medicinal plant extracts, and animal products to combat Acanthamoeba infection. Applied nanotechnology, an advanced technology, has shown to enhance the anti-Acanthamoeba activity in the encapsulated nanoparticles leading to new therapeutic options. This review attempts to provide an overview of the available data and studies on the occurrence of pathogenic Acanthamoeba among the Association of Southeast Asian Nations (ASEAN) members with the aim of identifying some potential contributing factors such as distribution, demographic profile of the patients, possible source of the parasite, mode of transmission and treatment. Further, this review attempts to provide future direction for prevention and control of the Acanthamoeba infection.
Acanthamoeba ; Amoeba ; Animals ; Asia, Southeastern ; Asian Continental Ancestry Group ; Encephalitis ; Humans ; Keratitis ; Nanoparticles ; Nanotechnology ; Parasites ; Plants, Medicinal ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Soil ; Swimming Pools ; Water

Objective To examine the acanthamoebicidal effects of ethyl acetate, aqueous and butanol fractions of dried flower buds of Lonicera japonica (L. japonica) Thunb. (Flos Lonicerae) in vitro. Methods Acanthamoeba triangularis isolates were obtained from environmental water samples and identified by PCR. They were exposed to ethyl acetate, water and butanol fractions of L. japonica Thunb. at concentrations ranging from 0.5 mg/mL to 1.5 mg/mL. The extracts were evaluated for growth inhibition at 24, 48 and 72 h, respectively. Chlorogenic acid at a concentration of 1 mg/mL was examined for inhibition of encystment. Results Ethyl acetate fraction at a concentration of 1.5 mg/mL evoked a significant reduction of trophozoite viability by 48.9% after 24 h, 49.2% after 48 h and 33.7% after 72 h chlorogenic acid, the major active constituent of L. japonica Thunb. at the concentration of 1 mg/mL reduced the cysts/trophozoite ratio by 100% after 24 h, 84.0% after 48 h and 72.3% after 72 h. This phenolic compound at concentration of 1 mg/mL concurrent with 0.6% hydrogen peroxide inhibited hydrogen peroxide-induced encystment by 92.8% at 72 h. Conclusions Results obtained from this study show that ethyl acetate fraction at 1.5 mg/mL is the most potent fraction of L. japonica Thunb. and its major constituent chlorogenic acid showed the remarkable inhibition of encystment at a concentration of 1 mg/mL.

Objective: To investigate the influence of ABCB1 polymorphisms on the plasma level of efavirenz in Thai adult cases infected with HIV-1. Methods: A single nucleotide polymorphism of ABCB1 3435C>T (rs1045642) in the gene encoding ABCB1 was genotyped using real-time PCR-based alleles in 149 HIV-infected Thai adults receiving efavirenz treatment. Plasma concentrations of efavirenz were measured by high-performance liquid chromatography 12 hr after administration. The relationship between plasma efavirenz concentrations and ABCB1 3435C>T polymorphisms was analyzed. Results: Logistic regression analysis showed no significant predictors of high plasma efavirenz concentration in relation to age, gender, body weight, CD4 count and plasma HIV-1 RNA, blood biochemical parameters, antiretroviral duration or ABCB1 3435C>T polymorphisms, except for height (OR=0.902, 95% CI: 0.835-0.973) (P<0.05). The minor allele frequency of ABCB1 3435C>T was 0.446. The frequency of the heterozygous mutant ABCB1 3435C/ T was 53.02% (n=79), ABCB1 3435T/T homozygous mutant was 18.12% (n=27) and the wild type ABCB1 3435C/C genotype was 28.86% (n=43). The overall median plasma concentration of efavirenz in 149 HIV-infected Thai cases was 2.41 mg/L [IQR: (1.46-4.12) mg/L]. The plasma concentration of efavirenz was higher in cases with ABCB1 3435T/T homozygous mutant [2.73 mg/L, IQR: (2.02-4.19) mg/L] and ABCB1 3435C/T heterozygous mutant [2.29 mg/L, IQR: (1.41-4.28) mg/L] genotypes compared to the wild type ABCB1 3435C/C homozygous [2.1 mg/L, IQR: (1.37-3.53) mg/L]. However, there was no statistically significant difference in the efavirenz concentration between the different genotypes (P>0.05). Conclusions: There is no statistical significance for a tendency toward higher plasma efavirenz concentration in the ABCB1 3435T/T and ABCB1 3435C/T genotypes. No parameters of physiological characteristics in this study except for height were found to be predictors of high plasma efavirenz concentration in Thai HIV-1 infected cases.