A Survey of Helminths of Dogs in Rural and Urban Areas of Uzbekistan and the Zoonotic Risk to Human Population.

Dogs are very popular pets that can be infected with a wide diversity of endo- and ectoparasites, some of which have zoonotic potential. The aim of the present study was to determine the diversity and prevalence of helminths in rural and urban dogs in Tashkent, Samarkand and Karakalpakstan regions of Uzbekistan. A total of 399 dogs from rural and urban areas were examined by necropsy between November 2016 and March 2022. All helminth species were morphologically identified. A total of 31 species belonging to the classes Trematoda (3), Cestoda (9), Nematoda (18) and Acanthocephala (1) were identified in 378 dogs (94.7%). Twenty-one species are indicated for the helminth fauna of urban dogs and 31 species for rural dog populations. From the 31 species of helminths identified 18 species are zoonotic and four of them (Echinicoccus granulosus, Dipylidium cani-num, Toxocara canis, Dirofilaria repens) have a significant epidemiological importance. The study showed that the prevalence and diversity of helminths in dogs in rural areas of Uzbekistan is higher than in urban dogs.

Phylogenetic Characteristics of Echinococcus granulosus Sensu Lato in Uzbekistan.

Echinococcosis occurs mainly in areas with heavy livestock farming, such as Central Asia, America, and Australia. Echinococcus granulosus sensu lato (s.l.) infection causes echinococcosis in intermediate hosts, such as sheep, cattle, goats, camels, and horses. Numerous cases of echinococcosis occur in Uzbekistan as stock farming is a primary industry. Epidemiological and genetic studies of E. granulosus s.l. are very important for mitigating its impact on public health and the economy; however, there are no such studies on E. granulosus s.l. in Uzbekistan. In the present study, to determine which genotypes exist and are transmitted, we isolated Echinococcus sp. from definitive hosts (one isolate each from jackal and dog) and intermediate hosts (52 isolates from humans and 6 isolates from sheep) in Uzbekistan and analyzed the isolates by sequencing 2 mitochondrial DNA components (cox1 and nad1). The results showed that all of isolates except one belonged to the E. granulosus sensu stricto (s.s.) G1 and G3 genotypes. Phylogenetic analysis based on cox1 sequences showed that 42 isolates from humans, 6 isolates from sheep, and one isolate from jackal were the G1 genotype, whereas the remaining 8 isolates from human and the one isolate from dog were the G3 genotype. These results suggest that the G1 and G3 genotypes of E. granulosus s.s. are predominant in Uzbekistan, and both wild animals and domestic animals are important for maintaining their life cycle. Only one isolate from human sample was confirmed to be E. eqiinus (G4 genotype), which is known to be for the first time.

Prevalence of Intestinal Helminth Infections in Dogs and Two Species of Wild Animals from Samarkand Region of Uzbekistan.

This study aimed to determine the prevalence of intestinal helminth parasitic infections and associated risk factors for the human infection among the people of Samarkand, Uzbekistan. Infection status of helminths including Echinococcus granulosus was surveyed in domestic and wild animals from 4 sites in the Samarkand region, Uzbekistan during 2015-2018. Fecal samples of each animal were examined with the formalin-ether sedimentation technique and the recovery of intestinal helminths was performed with naked eyes and a stereomicroscope in total 1,761 animals (1,755 dogs, 1 golden jackal, and 5 Corsac foxes). Total 658 adult worms of E. granulosus were detected in 28 (1.6%) dogs and 1 (100%) golden jackal. More than 6 species of helminths, i.e., Taenia hydatigena, Dipylidium caninum, Diplopylidium nolleri, Mesocestoides lineatus, Toxocara canis, and Trichuris vulpis, were found from 18 (1.0%) dogs. Six (T. hydatigena, Toxascaris leonina, Alaria alata, Uncinaria stenocephala, D. caninum, and M. lineatus) and 2 (D. nolleri and M. lineatus) species of helminths were also detected from 5 Corsac foxes and 1 golden jackal, respectively. Taeniid eggs were found in 2 (20%) out of 10 soil samples. In the present study, it was confirmed that the prevalences of helminths including E. granulosus are not so high in domestic and wild animals. Nevertheless, the awareness on the zoonotic helminth infections should be continuously maintained in Uzbekistan for the prevention of human infection.

Therapeutic effects of Echinococcus granulosus cystic fluid on allergic airway inflammation.

Previous studies showed that Echinococcus granulosus infection reduces allergic airway inflammation in experimentally infected hosts and the cystic fluid of E. granulosus is known to activate regulatory T (CD4+CD25+Foxp3+T, Treg) cells. To evaluate the effects of cystic fluid of E. granulosus on allergic airway inflammation, we investigated the regulation of the inflammatory reaction by cystic fluid using an allergic airway inflammation animal model. Cystic fluid was administered to C57BL/6 mice seven times every other day, after which allergic airway inflammation was induced using ovalbumin and aluminum. The airway resistance, number of eosinophils and other immune cells in the bronchoalveolar lavage fluid, and levels of Th2 and Th17-related cytokines were significantly reduced by cystic fluid pre-treatment in allergic airway inflammation-induced mice. The number IL-4+CD4+ T cells decreased, the number of Treg cells increased in the lung-draining lymph nodes and spleen of cystic fluid pre-treated mice. In conclusion, E. granulosus-derived cystic fluid may alleviate the Th2 allergic airway inflammatory response via Treg cells. Further studies of the immune regulation of cystic fluid may lead to the development of therapeutic agents for immune disorders.

A novel antihypertension agent, sargachromenol D from marine brown algae, Sargassum siliquastrum, exerts dual action as an L-type Ca2+ channel blocker and endothelin A/B2 receptor antagonist.

We isolated the novel vasoactive marine natural products, (5E,10E)-14-hydroxy-2,6,10-trimethylpentadeca-5,10-dien-4-one (4) and sargachromenol D (5), from Sargassum siliquastrum collected from the coast of the East Sea in South Korea by using activity-guided HPLC purification. The compounds effectively dilated depolarization (50mMK+)-induced basilar artery contraction with EC50 values of 3.52±0.42 and 1.62±0.63µM, respectively, but only sargachromenol D (5) showed a vasodilatory effect on endothelin-1 (ET-1)-induced basilar artery contraction (EC50=9.8±0.6µM). These results indicated that sargachromenol D (5) could act as a dual antagonist of l-type Ca2+ channel and endothelin A/B2 receptors. Moreover, sargachromenol D (5) lowered blood pressure in spontaneous hypertensive rats (SHRs) 2h after oral treatment at a dose of 80mg/kg dose and the effect was maintained for 24h. Based on our ex vivo and in vivo experiments, we propose that sargachromenol D (5) is a strong candidate for the treatment of hypertension that is not controlled by conventional drugs, in particular, severe-, type II diabetes-, salt-sensitive, and metabolic disease-induced hypertension.

Fasciola hepatica: Infection Status of Freshwater Snails Collected from Gangwon-do (Province), Korea.

Fasciola hepatica is a trematode that causes zoonosis, mainly in cattle and sheep, and occasionally in humans. Few recent studies have determined the infection status of this fluke in Korea. In August 2015, we collected 402 samples of freshwater snails at Hoenggye-ri (upper stream) and Suha-ri (lower stream) of Song-cheon (stream) in Daegwalnyeong-myeon, Pyeongchang-gun in Gangwon-do (Province) near many large cattle or sheep farms. F. hepatica infection was determined using PCR on the nuclear ribosomal internal transcribed spacer 2 (ITS-2). Among the 402 samples, F. hepatica 1TS-2 marker was detected in 6 freshwater snails; thus, the overall prevalence in freshwater snails was 1.5%. The prevalence varied between collection areas, ranging from 0.0% at Hoenggye-ri to 2.9% at Suha-ri. However, F. gigantica ITS-2 was not detected in the 6 F. hepatica-positive samples by PCR. The nucleotide sequences of the 6 F. hepatica ITS-2 PCR-positive samples were 99.4% identical to the F. hepatica ITS-2 sequences in GenBank, whereas they were 98.4% similar to F. gigantica ITS-2 sequences. These results indicated that the prevalence of F. hepatica in snail intermediate hosts was 1.5% in Gangwon-do, Korea; however the prevalence varied between collection areas. These results may help us to understand F. hepatica infection status in natural environments.

Antimalarial Activity of C-10 Substituted Triazolyl Artemisinin.

We synthesized C-10 substituted triazolyl artemisinins by the Huisgen cycloaddition reaction between dihydroartemisinins (2) and variously substituted 1, 2, 3-triazoles (8a-8h). The antimalarial activities of 32 novel artemisinin derivatives were screened against a chloroquine-resistant parasite. Among them, triazolyl artemisinins with electron-withdrawing groups showed stronger antimalarial activities than those shown by the derivatives having electron-donating groups. In particularly, m-chlorotriazolyl artemisinin (9d-12d) showed antimalarial activity equivalent to that of artemisinin and could be a strong drug candidate.

Practical Algorisms for PCR-RFLP-Based Genotyping of Echinococcus granulosus Sensu Lato.

Echinococcus granulosus sensu lato (s.l.) is a causative agent of cystic echinococcosis or cystic hydatid disease in humans and domestic and wild animals. The disease is a serious health problem in countries associated with poverty and poor hygiene practices, particularly in livestock raising. We introduced a practical algorism for genotyping the parasite, which may be useful to many developing countries. To evaluate the efficiency of the algorism, we genotyped 3 unknown strains isolated from human patients. We found that unknowns 1 and 3 were included in G1, G2, and G3 genotypes group and unknown 2 was included in G4 genotype (Echinococcus equinus) according to the algorisms. We confirmed these results by sequencing the 3 unknown isolates cox1 and nad1 PCR products. In conclusion, these new algorisms are very fast genotype identification tools that are suitable for evaluating E. granulosus s.l. isolated from livestock or livestock holders, particularly in developing countries.

Plerocercoids of Nybelinia surmenicola (Cestoda: Tentacularidae) in Squids, Todarodes pacificus, from East Sea, the Republic of Korea.

A visceral helminth of the squid, Todarodes pacificus, is reported from the East Sea, the Republic of Korea. Total 39 squid samples were purchased from a fish market in Jumunjin-eup, Gangneung-si (City) from August 2014 to July 2015 and were examined for helminth parasites with naked eyes and under a stereomicroscope after opening the abdominal cavity with a pair of scissors. Whitish larval worms were mainly found in the stomach and abdominal cavity of the squid. They were detected in 25 (64.1%) out of 39 squids examined, and the infection density was 7 larvae per infected squid. Spatula-shaped larvae were 8.2×2.0 mm in average size, round to slightly flattened anteriorly, with round hatching posteriorly, and had characteristic 4 tentacles with numerous hooklets in the scolex. The larvae were identified as the plerocercoid stage of Nybelinia surmenicola by their morphological features. This finding represents a new host record and the first report of N. surmenicola infection in T. pacificus squids from the east coast of Korea.

Geographical Distribution and Seasonal Indices of Chigger Mites on Small Mammals Collected on the East Coast of the Republic of Korea.

The geographical distributions and relative population densities of scrub typhus vector mites collected from small mammals were determined for 5 locations on the east coast of the Republic of Korea. Collection sites included Goseong, Gangneung, and Hoengseong in Gangwon province and Uljin and Yeongdeok in Gyeongbuk province. A total of 275 small mammals including members of Rodentia (rodents) and Soricomorpha (shrews, such as Crocidura lasiura ) belonging to 4 genera and 4 species were captured in the field from 2012 to 2013. Apodemus agrarius was collected most frequently (220, 80%), followed by C. lasiura (25, 9.1%), Mus musculus (15, 5.5%), and Myodes regulus (15, 5.5%). A total of 23,436 larval chigger mites (Family Trombiculidae) belonging to 3 genera and 8 species (Leptotrombidium pallidum, Leptotrombidium scutellare, Leptotrombidium palpale, Leptotrombidium orientale, Leptotrombidium zetum, Neotrombicula tamiyai, Neotrombicula japonica, and Euschoengastica koreaensis) were collected from the small mammals. The predominant chigger species collected during the spring and fall seasons from A. agrarius were L. pallidum (57.6%), L. palpale (14.5%), and L. scutellare (7.9%). Leptotrombidium scutellare was collected only along the southeastern coast at Yeongdeok, Gyeongbuk province. The geographical distribution of scrub typhus vectors and reservoir hosts are important aspects of understanding the epidemiology of the disease as well as the potential impacts of climate change and health risks.

Karyological studies of Biomphalaria tenagophila (d'Orbigny, 1835) (Gastropoda: Planobidae) from Rio de Janeiro, Brazil.

The karyotypes of Biomphalaria tenagophila collected from Rio de Janeiro, Brazil were studied using the air-drying method. Somatic cells of this species had 2n=36. The 18 chromosome pairs were identified and classified into 3 groups. The diploid cell has 7 pairs of metacentric, 8 pairs of submetacentric, and 3 pairs of subtelocentric chromosomes. Observed chromosomes ranged from 2.4 to 6.4 µm, and the total length was 122.3 µm. This is the first report on the chromosome of B. tenagophila.

Glioblastoma-specific anticancer activity of pheophorbide a from the edible red seaweed Grateloupia elliptica.

The chlorophyll-related compound pheophorbide a (Pa) was successively purified from an edible red seaweed, Grateloupia elliptica, using silica, octadecyl silica column chromatography and reversed phase-high-performance liquid chromatography, as well as the cell cycle inhibitory and apoptotic effects of Pa being investigated in U87MG glioblastoma cells. The Pa exhibited strong anticancer effects in the absence of direct photo-irradiation against various cancer cell lines, including U87MG, SK-OV-3, and HeLa cells. Among the cancer cells, the strongest anticancer activity of Pa exhibited on U87MG cells with IC50 values of 2.8 µg/ml. In addition, Pa specifically had cytostatic activity on glioblastoma cells rather than human umbilical vein endothelial cells. Analysis of the cell cycle distribution showed that Pa induced G0/G1 arrest of U87 MG cells. In addition, arrested cells induced late apoptosis and DNA degradation under dark condition. These results suggest that Pa isolated from G. elliptica is a potential glioblastoma-specific anticancer agent without side effects on normal cells.

5E- and 5Z-farnesylacetones from Sargassum siliquastrum as novel selective L-type calcium channel blockers.

A specific blocker of L-type Ca(2+) channels may be useful in decreasing arterial tone by reducing the open-state probability of L-type Ca(2+) channels. The aim of the present study was to evaluate the farnesylacetones, which are major active constituents of Sargassum siliquastrum, regarding their vasodilatation efficacies, selectivities toward L-type Ca(2+) channels, and in vivo antihypertensive activities. The application of 5E-(farnesylacetone 311) or 5Z-farnesylacetone (farnesylacetone 312) induced concentration-dependent vasodilatation effects on the basilar artery that was pre-contracted with depolarization and showed an ignorable potential role of endothelial-derived nitric oxide. We also tested farnesylacetone 311 or 312 to determine their pharmacological profiles for the blockade of native L-type Ca(2+) channels in basilar arterial smooth muscle cells (BASMCs) and ventricular myocytes (VMCs), cloned L- (α1C/ß2a/α2δ), N- (α1B/ß1b/α2δ), and T-type Ca(2+) channels (α1G, α1H, and α1I). Farnesylacetone 311 or 312 showed greater selectivity toward the L-type Ca(2+) channels among the tested voltage-gated Ca(2+) channels. The ranked order of the potency for farnesylacetone 311 was cloned α1C≒L-type (BASMC)≒L-type (VMCs)>α1B>α1H>α1I>α1G and that for farnesylacetone 312 was cloned α1C≒L-type (BASMCs)≒L-type (VMCs)>α1H>α1G>α1B>α1I. The oral administration of the farnesylacetone 311 (80mg/kg) conferred potent, long-lasting antihypertensive activity in spontaneous hypertensive rats, but it did not alter the heart rate.

Anti-human rhinoviral activity of polybromocatechol compounds isolated from the rhodophyta, Neorhodomela aculeata.

An extract of the red alga, Neorhodomela aculeata, exhibited antiviral activity against human rhinoviruses. Bioassay-guided purification was performed to yield six compounds, which were subsequently identified as lanosol (1) and five polybromocatechols (2-6) by spectroscopic methods, including 1D and 2D NMR and mass spectrometric analyses. Structurally, all of these compounds, except compound 5, contain one or two 2,3-dibromo-4,5-dihydroxyphenyl moieties. In a biological activity assay, compound 1 was found to possess antiviral activity with a 50% inhibitory concentration (IC50) of 2.50 µg/mL against HRV2. Compound 3 showed anti-HRV2 activity, with an IC50 of 7.11 µg/mL, and anti-HRV3 activity, with an IC50 of 4.69 µg/mL, without demonstrable cytotoxicity at a concentration of 20 µg/mL. Collectively, the results suggest that compounds 1 and 3 are candidates for novel therapeutics against two different groups of human rhinovirus.

Penetration of the oral mucosa by parasite-like sperm bags of squid: a case report in a Korean woman.

We report a case of oral stings by spermatophores of the squid Todarodes pacificus . A 63-yr-old Korean woman experienced severe pain in her oral cavity immediately after eating a portion of parboiled squid along with its internal organs. She did not swallow the portion, but spat it out immediately. She complained of a pricking and foreign-body sensation in the oral cavity. Twelve small, white spindle-shaped, bug-like organisms stuck in the mucous membrane of the tongue, cheek, and gingiva were completely removed, along with the affected mucosa. On the basis of their morphology and the presence of the sperm bag, the foreign bodies were identified as squid spermatophores.

Vasodilatation effect of farnesylacetones, active constituents of Sargassum siliquastrum, on the basilar and carotid arteries of rabbits.

Two farnesylacetones, 311 and 312, major active constituents of Sargassum siliquastrum collected from the coast of the East Sea in Korea, showed a moderate vasodilatation effect on the basilar arteries of rabbits. Therefore, treatment with farnesylacetones 311 and 312 may selectively accelerate cerebral blood flow through dilatation of the basilar artery.

Sargaquinoic acid and sargahydroquinoic acid from Sargassum yezoense stimulate adipocyte differentiation through PPARalpha/gamma activation in 3T3-L1 cells.

We screened active compounds from natural marine products able to increase PPARalpha/gamma transcriptional activity. Sargaquinoic acid (SQA) and sargahydroquinoic acid (SHQA) from Sargassum yezoense were identified as novel PPARalpha/gamma dual agonists. The binding affinity of SQA with PPARgamma was higher than that of the specific PPARgamma agonist troglitazone, leading to an activation of PPARgamma transcriptional activity. In parallel, treatment of 3T3-L1 cells with SQA and SHQA led to an increase in adipocyte differentiation and increased expression of adipogenic marker genes such as aP2, PPARgamma, resistin, adiponectin, C/EBPalpha and Glut4. Collectively, our data suggest that SQA and SHQA are novel PPARalpha/gamma dual agonists and may be beneficial for reducing insulin resistance through regulation of adipogenesis.

Selective vasodilatation effect of sargahydroquinoic acid, an active constituent of Sargassum micracanthum, on the basilar arteries of rabbits.

Sargahydroquinoic acid (2), a major active constituent of Sargassum micracanthum collected from the coast of the East Sea in Korea, showed a selective vasodilatation effect on the basilar arteries of rabbits. Therefore, treatment with sargahydroquinoic acid may selectively accelerate cerebral blood flow through dilatation of the basilar artery without lowering systemic blood pressure.

Genetic comparison of liver flukes, Clonorchis sinensis and Opisthorchis viverrini, based on rDNA and mtDNA gene sequences.

To clarify the genetic relationships between Clonorchis sinensis and Opisthorchis viverrini, patterns of inter-/intraspecific polymorphism were compared for four markers with nuclear ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA) in liver flukes C. sinensis from Korea (Kimhae) and China (Shenyang and Nanning) and O. viverrini from Laos (Savannakhet). Intra- and interspecific variations in the 18S, ITS2, and 28S rDNA and mitochondrial cytochrome c oxidase subunit I (mtCOI) of mtDNA gene sequences were low and nearly identical. Three isolates of C. sinensis showed a high similarity (99-100%). No variation was detectable in the ITS2 sequence for the C. sinensis from Korea and China. ITS2 region sequences of O. viverrini vs C. sinensis showed 95% identity and differed at 28 nucleotide positions. Pairwise sequence divergence with three C. sinensis isolates and O. viverrini ranged from 0 to 3.94% in mtCOI gene. The mtCOI sequences were more highly conserved relative to the ITS2 sequences. These genetic data from different geographical areas showed that the liver flukes are not variable and are virtually identical almost despite belonging to entirely different genera.

The inhibitory effects of eckol and dieckol from Ecklonia stolonifera on the expression of matrix metalloproteinase-1 in human dermal fibroblasts.

In order to develop new anti-photoaging agents, we examined the inhibitory effects of 29 seaweed extracts on transcriptional activities of NF-kappaB and AP-1, and MMP-1 expression. The extracts from 3 species of Alariaceae, Eisenia bicyclis, Ecklonia cava and Ecklonia stolonifera, have showed strong inhibition of both NF-kappaB and AP-1 reporter activity, which were well correlated with their abilities to inhibit MMP-1 expression. In addition, MMP-1 expression was dramatically attenuated by treatment with eckol or dieckol which were purely isolated from E. stolonifera, indicating that these compounds are active principles to inhibit MMP-1 expression in human dermal fibroblasts. Taken together, our data demonstrate the inhibitory effect of eckol and dieckol from Ecklonia species on MMP-1 expression in human dermal fibroblasts and provide a possibility to develop as an agent for the prevention and treatment of skin aging.