Molecular detection and characterization of Acanthamoeba infection in dogs and its association with keratitis in Korea.

Acanthamoeba infection is associated with keratitis in humans; however, its association with keratitis in dogs remains unclear. To investigate this possibility, we collected 171 conjunctival swab samples from dogs with eye-related diseases (65 with keratitis and 106 without keratitis) at Chungbuk National University Veterinary Teaching Hospital, Korea, from August 2021 to September 2022. Polymerase chain reaction identified 9 samples (5.3%) as Acanthamoeba positive; of these, 3 were from dogs with keratitis (4.6%) and 6 were from dogs without keratitis (5.7%). Our results indicated no significant association between Acanthamoeba infection and keratitis, season, sex, or age. All Acanthamoeba organisms found in this study had the genotype T4, according to 18S ribosomal RNA analysis. Acanthamoeba infection in dogs might have only a limited association with keratitis.

Outbreak of severe diarrhea due to zoonotic Cryptosporidium parvum and C. xiaoi in goat kids in Chungcheongbuk-do, Korea.

Severe diarrhea was reported in goat kids in Chungcheongbuk-do, Korea, from 2021 to 2023, and Cryptosporidium infection was suspected. To confirm the cause of this outbreak, fecal samples were collected from goat farms where diarrhea had been reported and analyzed for Cryptosporidium infection using a molecular assay. A total of 65 fecal samples, including 37 from goats with diarrhea and 28 from goats without diarrhea, were collected from six goat farms. Forty-eight of the goats were kids (<2 months) and 17 were adults (>1 year). Cryptosporidium was identified in 53.8% (35/65) of total samples. Overall, 86.5% (32/37) of the diarrheic fecal samples tested positive; however, Cryptosporidium was not detected in any fecal sample from non-diarrheic adult goats. Therefore, cryptosporidiosis was significantly associated with diarrhea in goat kids, and adult goats were not responsible for transmission of Cryptosporidium to them. Phylogenetic analysis and molecular characterization revealed two Cryptosporidium species, namely, C. parvum (n = 28) and C. xiaoi (n = 7). In the C. parvum-positive samples, gp60 gene analysis revealed three zoonotic subtypes-IIaA18G3R1, IIdA15G1, and IIdA16G1. To the best of our knowledge, this study is the first to identify C. parvum IIaA18G3R1 and IIdA16G1 in goats, as well as the first to identify C. xiaoi in goats in Korea. These results suggest that goat kids play an important role as reservoir hosts for different Cryptosporidium species and that continuous monitoring with biosecurity measures is necessary to control cryptosporidiosis outbreaks.

Tick Populations and Molecular Analysis of Anaplasma Species in Ticks from the Republic of Korea.

The present study was performed to survey the dominant tick populations and molecularly determine the pathogenic agents of anaplasmosis in ticks from Gyeongsang, Republic of Korea. A total of 3825 questing ticks were collected by the flagging method from 12 sites near animal farms in Gyeongsang from March to October 2021. A molecular genomic study was performed with ticks stored in 70% ethanol to detect Anaplasma genes by the previously described method. The monthly incidence of ticks varied by developmental stages, i.e., nymphs, adults, and larvae, and each of their populations peaked in May, March, and October, respectively. The predominant tick species were Haemaphysalis longicornis, Haemaphysalis sp., Haemaphysalis flava, Ixodes nipponensis, and Amblyomma testudinarium in order. To determine the Anaplasma infection rate, collected ticks were pooled into 395 groups. The minimum infection rate (MIR) of Anaplasma was 0.7% (27 pools). That of A. phagocytophilum was highest (23 pools, MIR 0.6%), followed by A. phagocytophilum-like Anaplasma spp. clade B (2 pools, MIR 0.1%), A. bovis (1 pool, MIR 0.1%), and A. capra (1 pool, MIR 0.1%), respectively. In this study, five species of ticks, including unidentified Haemaphysalis species, were collected in 12 survey sites in Gyeongsang, but their prevalence was somewhat different according to the tick species and survey sites. Further, the incidence rate (6.8%) of 4 Anaplasma spp. was not as high in tick pools. However, the results of this study may offer a basis for future epidemiological research and risk assessment of tick-borne diseases.

Molecular epidemiology of Theileria species in ticks and its potential threat to livestock in the Republic of Korea.

The purpose of this study was to investigate molecular epidemiology of Theileria spp. in ticks in Korea and assess their potential threat from wildlife animals to domestic animals. A total of 21152 hard ticks were collected from Chungcheong and Jeolla provinces of Korea from March to October 2021. Tick species were identified by microscopy and Theileria spp. were screened by nested PCR targeting 18S rRNA. Haemaphysalis spp. were the most abundant tick species, followed by H. longicornis, H. flava, Amblyomma testudinarium, and Ixodes nipponensis. Of the collected ticks, 6914 ticks (541 pools) were screened, and PCR showed 211 positive pools (39.0%; MIR: 3.05). The PCR and phylogenetic analysis identified two Theileria species, T. luwenshuni and Theileria sp., with T. luwenshuni (162/211, 76.78%; MIR: 2.34) being more abundant than Theileria sp. (36/211, 17.06%; MIR: 0.52); co-infection of the two species were noted (13/211, 6.16%; MIR: 0.19). Among the tick species, H. longicornis, especially nymphs, showed the highest prevalence. Regarding season, the highest prevalence was observed in May. Considering the tick and Theileria species identified in this study, H. longicornis nymph and cervine play a critical role in maintaining Theileria spp. in Korea and could be a potential threat to domestic animals, including deer and goats. In addition, there are significant correlations among tick distribution, region, season, and prevalence of Theileria.

Two cases of ectopic dirofilariasis by Dirofilaria immitis in subconjunctival and subcutaneous tissues in dogs.

We report two cases of ectopic dirofilariasis caused by Dirofilaria immitis in the subconjunctival and subcutaneous tissues of dogs. In Case 1, a 10-year-old female poodle suffered from a subconjunctival mass in the left eye that was refractory to antibiotic and steroid treatments. The mass was removed surgically, and a whitish nematode was identified within the mass. In Case 2, a stray 3-year-old male greyhound was rescued and transferred to a local veterinary hospital. During care, two nematodes were observed on the left hind paw and leg. The observed worms were not typical of D. immitis; therefore, the species was confirmed using molecular methods. Phylogenetic analysis revealed high genetic identity with other previously reported D. immitis strain. Subcutaneous and subconjunctival dirofilariasis have been mainly caused by D. repens. To the best of our knowledge, this is the first report of subconjunctival ectopic dirofilariasis by D. immitis in a dog, and the first report of subcutaneous localization in Korea. Therefore, in endemic regions, ectopic dirofilariasis caused by D. immitis should be considered as a differential diagnosis in subconjunctival and subcutaneous masses.

Distribution of Gastrointestinal Parasitic Infection in Domestic Pigs in the Republic of Korea: Nationwide Survey from 2020-2021.

This study aimed to examine the distribution of gastrointestinal parasitic infections in domestic pigs in the Republic of Korea. From May 2020 to October 2021, 364 pig fecal samples were collected from 75 farms in 7 Provinces and microscopically examined. A total of 170 (46.7%) pigs were infected with at least one of the following parasites: Balantioides coli, strongyles, Ascaris suum, Trichuris suis, and coccidia. By parasite species, B. coli, strongyles, A. suum, T. suis, and coccidia oocysts or eggs were detected in 144 (39.6%), 24 (6.6%), 14 (3.8%), 4 (1.1%), and 1 (0.3%) samples, respectively. One hundred fifty-four, 15, and 1 cases showed single, double, and triple infections, respectively. Of the swine fecal samples from 75 farms, 69 specimens (92.0%) were infected with 1 or more parasites. All surveyed farms across the country exhibited a positive rate of over 30%, among which the highest positive rate was 65.0% in Chungcheongnam-do, and Jeollabuk-do was followed by 61.9%. Winter showed a statistically lower prevalence than other seasons. This study showed that gastrointestinal parasites are prevalent in pigs in Korea, although the diversity of parasites is low.

Pomegranate (Punica granatum) Juice Shows Antioxidant Activity against Cutaneous Leishmaniasis-Induced Oxidative Stress in Female BALB/c Mice.

Leishmania species are parasites that multiply within phagocytes and cause several clinical diseases characterized by single or multiple ulcerations. One of the complications that can induce tissue damage and the resulting scars is caused by secondary bacterial infections. Studies to find new, effective, and safe oral drugs for treating leishmaniasis are being conducted since several decades, owing to the problems associated with the use of antimonials available. Previously, the antiparasitic and antioxidant properties of Punica granatum (pomegranate, P. granatum) have been reported. Therefore, in the present study, we aimed to investigate the antileishmanial activity of pomegranate aqueous juice in vitro and in female BALB/c mice. A 3-(4.5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in Leishmania major promastigotes and alterations in the antioxidant status, liver function, and skin histological changes in L. major-infected mice orally treated with pomegranate juice alone and in combination with the antibiotic ciprofloxacin, were used to investigate the in vitro and in vivo antileishmanial activity of pomegranate juice, respectively. Oral P. granatum juice treatment significantly reduced the average size of cutaneous leishmaniasis lesions compared with that of the untreated mice. This antileishmanial activity of P. granatum was associated with enhanced antioxidant enzyme activities. Histopathological evaluation proved the antileishmanial activity of P. granatum, but did not reveal changes in the treated animals, compared to the positive control. In conclusion, P. granatum shows high and fast antileishmanial activity probably by boosting the endogenous antioxidant activity.

Anti-Leishmanial Activity (In Vitro and In Vivo) of Allicin and Allicin Cream Using Leishmania major (Sub-strain Zymowme LON4) and Balb/c Mice.

BACKGROUND: Leishmania is a unicellular protozoan parasite that produces several human diseases, ranging from localized self-healing cutaneous lesions to deadly visceral infections. OBJECTIVE: The effect of allicin on the growth of Leishmania major (L. major) promastigotes was evaluated under in vitro conditions. Moreover, the efficacy of a topical allicin cream was examined in BALB/c (Bagg albino, laboratory-bred strain of the House Mouse) mice with cutaneous leishmanial lesions compared to the currently used drug, sodiumstibogluconate (pentostam). METHODS: Cytotoxiciy and promastigote proliferation were measured. Different concentrations (50, 100, 150, and 200 µM) of liquid allicin were tested on L. major promastigotes twice: after 24 and 48 hours using an MTT colorimetric assay. In the in vivo condition, the efficacies of allicin cream and liquid allicin at two concentrations (0.15 µM/mouse and 0.30 µM/mouse) were evaluated. Serum factors of the control and treated groups were tested to evaluate the toxic effects of allicin on the liver and kidney. RESULTS: Allicin at a concentration of 50 µM inhibited the growth of Leishmania promastigotes. Topical application of allicin cream reduced lesion sizes in mice. No significant differences in biochemical analysis were observed between the control and treated groups. CONCLUSIONS: Allicin has antileishmanial effects under in vitro and in vivo conditions and may be used in clinical applications.