Decreased SMP30 Expression Is Related With EMT in the Kidneys of Two Siberian Tigers With CKD.

BACKGROUND/AIM: Chronic kidney disease (CKD) is one of the most common causes of mortality in wild non-domestic felidae. The molecular mechanism regulating renal fibrosis in nephropathy is not fully understood especially in the felidae. This study aimed to elucidate senescence marker protein 30 (SMP30) expression patterns and its relationship with epithelial-mesenchymal transition (EMT) by immunostaining in two necropsied Siberian tigers (Panthera tigris altaica) with CKD. MATERIALS AND METHODS: Two kidney samples from male Siberian tigers were fixed and tissue sections were stained for histopathological assay. RESULTS: In CKD, renal tubular epithelial cells lost their tubular structures surrounded by severe interstitial fibrosis and were detached from the basement membrane. These damaged cells resembled the morphology of mesenchymal cells and showed much lower SMP30 expression compared with intact tubular epithelial cells. These cells also expressed vimentin, which is specifically expressed by mesenchymal cells, and through double staining, it was observed that vimentin was expressed in the tubular epithelial cells where SMP30 was not expressed. In addition, double-positive expression of pan-cytokeratin (pan-CK) and vimentin was found in damaged epithelial cells with mesenchymal features. CONCLUSION: We demonstrated possible evidence to understand the role of SMP30 as a new pivotal factor and the possibility of decreased SMP30 as a potential indicator of EMT at the end stage of CKD.

Molecular Detection and Genotyping of Theileria spp. in Deer (Cervidae) in Korea.

Major clinical symptoms of Theileria infection include fever, anemia, anorexia, jaundice, and decreased milk production. Although several studies have been conducted on tick-borne pathogens, including Theileria in Korea, only a few have focused on Theileria infection in deer, including the Korean water deer. Blood samples from 160 deer were collected and subjected to DNA extraction and polymerase chain reaction (PCR). Next, PCR-positive samples were sequenced and analyzed by constructing a phylogenetic tree. The results showed that the overall infection rate of Theileria was 8.1% (13/160). Infection rates of 100% were observed in the northern and southern regions. However, the study's limitation was its small sample size, wherein five and one samples were analyzed from the northern and southern regions, respectively. The central region exhibited the lowest infection rate of 2.9% (4/140). Infection rates also differed based on seasons, with the highest (18.4%, 9/49) being observed in spring, followed by that in summer (8.9%, 4/45). However, no infection was observed during autumn and winter. A phylogenetic analysis indicated that the PCR-positive samples contained Theileria luwenshuni, which usually infects small ruminants, such as goats and sheep.

The anti-platelet activity of panaxadiol fraction and panaxatriol fraction of Korean Red Ginseng in vitro and ex vivo.

Background: The anti-platelet activity of the saponin fraction of Korean Red Ginseng has been widely studied. The saponin fraction consists of the panaxadiol fraction (PDF) and panaxatriol fraction (PTF); however, their anti-platelet activity is yet to be compared. Our study aimed to investigate the potency of anti-platelet activity of PDF and PTF and to elucidate how well they retain their anti-platelet activity via different administration routes. Methods: For ex vivo studies, Sprague-Dawley rats were orally administered 250 mg/kg PDF and PTF for 7 consecutive days before blood collection via cardiac puncture. Platelet aggregation was conducted after isolation of the washed platelets. For in vitro studies, washed platelets were obtained from Sprague-Dawley rats. Collagen and adenosine diphosphate (ADP) were used to induce platelet aggregation. Collagen was used as an agonist for assaying adenosine triphosphate release, thromboxane B2, serotonin, cyclic adenosine monophosphate, and cyclic guanosine monophosphate (cGMP) release. Results: When treated ex vivo, PDF not only inhibited ADP and collagen-induced platelet aggregation, but also upregulated cGMP levels and reduced platelet adhesion to fibronectin. Furthermore, it also inhibited Akt phosphorylation induced by collagen treatment. Panaxadiol fraction did not exert any anti-platelet activity in vitro, whereas PTF exhibited potent anti-platelet activity, inhibiting ADP, collagen, and thrombin-induced platelet aggregation, but significantly elevated levels of cGMP. Conclusion: Our study showed that in vitro and ex vivo PDF and PTF treatments exhibited different potency levels, indicating possible metabolic conversions of ginsenosides, which altered the content of ginsenosides capable of preventing platelet aggregation.

Application of 3D-printed prosthetic lower beak in an Oriental stork (Ciconia boyciana).

To our knowledge, ours is the first case of applying a 3D-printed prosthetic beak to an Oriental stork (Ciconia boyciana). A stork in captivity underwent several surgeries for beak fractures, but the lower-mandible fractures failed to be repaired. Therefore, we applied a patient-specific beak prosthesis of titanium alloy and nylon. Because the prosthetic beak could not be maintained due to mandible and soft-tissue inflammation, the stork was euthanized. Still, we confirmed typical behavior and feeding for ~3 months after surgery. This report highlights some of the challenges we encountered and identifies process improvements required for a more successful surgery.

Molecular epidemiology of Toxoplasma gondii in cattle in Korea.

Toxoplasmosis is a major public health concern, with raw or undercooked meat being the primary source of human infection. Knowledge regarding the molecular epidemiology of Toxoplasma gondii in cattle destined for human consumption in Korea is lacking. The present study aimed to genetically characterize the infectious strains of T. gondii. Overall, 455 cattle blood samples from 84 farms in the Gyeongnam-do (Province) were randomly collected in 2017. Nested PCR analysis revealed that only 3 (0.7%) samples were infected with T. gondii. The B1 gene sequence of T. gondii was observed to be similar (97.3-99.6%) to that of other T. gondii isolates. This is the first study to perform the molecular detection of T. gondii in cattle in Korea. Although the prevalence of infection was low, our findings suggest that cattle present a potential public health issue. It may be crucial to recognize the importance of T. gondii infection in cattle meat.

Molecular analysis of Rickettsia spp. and related tick-borne pathogens detected in dogs in Korea.

Ticks are widespread in nature and serve as primary vectors for several tick-borne pathogens (TBPs). Ticks and TBPs cause considerable harm to humans and animals and have emerged as a major global public health concern. Domestic dogs are the major reservoirs of zoonotic agents owing to their constant interaction with humans. This study aimed to assess the prevalence and risk factors of canine TBPs, such as Rickettsiales, Coxiella burnetii, hepatozoa, and Borrelia spp., using molecular analyses. A total of 906 dogs were examined and 4 TBPs were identified: Anaplasma phagocytophilum (5; 0.6%), Hepatozoon canis (9; 1.0%), Candidatus Rickettsia longicornii (2; 0.2%), and Rickettsia tamurae (1; 0.1%). Ehrlichia spp., C. burnetii, and Borrelia spp. were not detected. To the best of our knowledge, this is the first study to perform a phylogenetic analysis of Candidatus R. longicornii and R. tamurae in dogs. These findings can help determine the potential public health risks by enhancing our understanding of the geographical and vector distributions of TBPs in Korea.

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.

Sudden death caused by Aeromonas hydrophila in a Crab-eating Macaque (Macaca fascicularis) associated with captivity stress.

Aeromonas hydrophila is a facultative anaerobic gram-negative bacterium regarded as an opportunistic pathogen in animals. A 17-year-old female crab-eating macaque (Macaca fascicularis) died after experiencing anorexia and depression for several days. The carcass was severely emaciated, and the sternum was exposed under subcutaneous lesions in the thorax. Many abnormal pathological lesions were found, including tracheal inflammation, pulmonary inflammatory emphysema, yellowish discoloration of the liver, enlargement of the gall bladder, necrosis of the heart, congested bilateral kidneys, and enlargement of the adrenal glands. The stomach was empty, mucosal ulcerations were found, and the duodenum was congested. Giemsa staining revealed rod-shaped organisms in the whole blood smear and major organs, which were identified as A. hydrophila. The animal had experienced stress, and decreased immune system function possibly contributed to the infection.

A case of rickets in an artificially raised white-tailed eagle (Haliaeetus albicilla) chick at a zoo.

A two-week-old white-tailed eagle presented with an inability to stand and flex its limbs. Despite hatching naturally and owing to lack of parental attention, the bird was raised indoors by zookeepers with no access to sunlight. Palpation and radiographic examination of the bilateral tibiotarsus and femur bone revealed pronounced deformation and curvature, and bilateral decreased bone densities, respectively. The reduced calcium concentration in the blood was treated with calcium gluconate injections and calcium-supplemented feeds. Chopped mouse tails were fed directly, and whole pink-skinned nude mice were fed weekly. The zookeeper also gently massaged the bird and dressed it with a bandage. Sunlight exposure was provided daily. Saliva containing chicken feed was obtained from the mother. The bird could stand properly after four weeks of treatment, and the blood calcium concentration was restored to normal levels.

Successful repair of coracoid and femur fractures in a Eurasian eagle owl (Bubo bubo) and its post-release survival.

To our knowledge, this is the first case of concurrent diaphyseal left coracoid and left femoral fractures in a Eurasian eagle owl and its post-release survival in Korea. The femur was surgically repaired using an external skeletal fixator-intramedullary (IM) pin tie-in method, and the coracoid was repaired solely with an IM pin on day 6 after femur surgery. The eagle owl underwent a gradual rehabilitation process. The bird was successfully rehabilitated and released 101 d after initial presentation. The bird was monitored using a wildlife tracking device and was confirmed to have survived for over 5 mon in the wild.

Molecular Detection and Genetic Diversity of Blastocystis in Korean Dogs.

Blastocystis is a genus of unicellular heterokont parasites belonging to a group of organisms known as Stramenopiles, which includes algae, diatoms, and water molds. Blastocystis includes several species that habitat in the gastrointestinal tracts of organisms as diverse as humans, farm animals, birds, rodents, reptiles, amphibians, fish, and cockroaches. It is important to public health and distributed globally, but its prevalence in dogs in Korea has not been reported to date. Here, we collected 787 canine fecal samples and assessed Blastocystis infection by age, sex, region, season, and diarrhea symptoms. We determined Blastocystis subtypes using phylogenetic analyses based on 18S rRNA gene sequences. We identified, 10 Blastocystis positive samples (1.3%). A higher proportion of infected dogs was asymptomatic; however, infection rates did not significantly differ according to region, age, sex, and season. Phylogenetic analysis showed that the Blastocystis sp. identified belonged to 4 subtypes (STs), ST1, ST5, ST10, and ST14, thus revealed the genetic diversity of Blastocystis sp. in dogs Korean. This is first report on the presence of Blastocystis sp. in dogs Korean. This study revealed a lower infection rate than expected and differed from previous studies in STs. Further studies are warranted to observe the national infection status of Blastocystis in dogs and the genetic characteristics of this genus.

Enhanced Asymptomatic Systemic Infection Caused by Plesiomonas shigelloides in a Captive Gray Wolf (Canis lupus).

A 7-year-old male gray wolf was found dead at a zoo during exhibition. To determine the cause of death, histological and gross necropsy diagnoses and a molecular analysis were performed. The gross necropsy revealed a swollen abdomen, hemorrhagic exudates around the mouth, splenomegaly, a discolored liver, a congested kidney, hemorrhagic ascites, and dark gray-colored membranes and air bubbles in the fundus of the stomach. Rod-shaped bacteria were found in the liver parenchyma and hemorrhagic ascites using Giemsa staining. The nucleotide sequencing of the cultured bacteria identified the causative agent as Plesiomonas shigelloides, which is rarely responsible for systemic infections. This study describes a rare case and the first reported systemic gastrointestinal infection due to P. shigelloides in a zoo animal.

Enhanced Virulence of Aeromonas hydrophila Is Induced by Stress and Serial Passaging in Mice.

Aeromonas hydrophila was isolated from an African black-footed penguin (Spheniscus demersus) that died while in zoo captivity. At necropsy, the virulence of A. hydrophila appeared to be enhanced by stress, so was assessed in the presence of in vitro and in vivo stressors and serial passaging in mice. Virulence genes from the isolate were amplified by PCR. In vitro assays were conducted to test the hemolytic activity, cytotoxicity, and effect of stress hormones on A. hydrophila virulence. In vivo assays were conducted to test the stress effect on mortality of A. hydrophila-infected mice and virulence in mice. Two virulence genes coding for hemolysin (ahh1) and aerolysin (aerA) were detected, and the cytotoxic potential of the isolate was demonstrated in baby hamster kidney and Vero cells. Some or all mice inoculated with A. hydrophila and exposed to stress hormones (epinephrine and norepinephrine) or low temperature died, while mice inoculated with A. hydrophila and exposed to fasting or agitation stressors or no stressors survived. We concluded that stress can be fatal in mice experimentally infected with A. hydrophila and that serial passaging in mice dramatically enhances the virulence of A. hydrophila.

Genetic Diversity and Zoonotic Potential of Blastocystis in Korean Water Deer, Hydropotes inermis argyropus.

Blastocystis is a protozoan parasite commonly detected in the intestinal tract of humans and animals. It has been actively studied worldwide; however, information on Blastocystis is limited in Korea. Because there is an increasing concern about the contact between wildlife and domestic animals or humans, we assessed the infection status and zoonotic potential of Blastocystis in Korean water deer (KWD, Hydropotes inermis argyropus) using genotyping and phylogenetic analysis. A total of 125 fresh fecal samples were collected from KWD which were killed by vehicles on highways or roadsides in this study. Among the 125 samples, 51 (40.8%) were PCR positive. We performed nucleotide sequencing and phylogenetic analysis of 26 of the 51 PCR-positive samples. By analyzing Blastocystis 18S rRNA, two subtypes (ST4 and ST14) were identified in this study. Of the 26 samples analyzed, 25 were identified as ST14 and one as ST4. Infection of ST14 in humans has not been reported. Although only one ST4 sample was detected in this study, ST4 has zoonotic potential without showing ruminant specificity. Thus, continuous attention should be provided to the potential of transmission between wildlife and domestic animals and humans.

Morphological and molecular diagnosis of Eimeria sp. that caused fatality in a red-necked wallaby (Macropus rufogriseus) in Korea.

A red-necked wallaby (Macropus rufogriseus) in the Republic of Korea, introduced from Australia, died in 12 d after exhibiting anorexia and diarrhea. Postmortem examination revealed that the wallaby died due to coccidiosis by Eimeria sp. Morphologically, Eimeria sp. identified closely resembled E. mykytowyczi. The 18S rRNA sequence analysis showed that Eimeria sp. identified in this study has a 98.5% identity with that in Australian red kangaroo (M. rufus). However, owing to insufficient molecular information on marsupial-specific Eimeria, exact species could not be determined. Phylogenetically, Eimeria sp. identified in this study belonged to clade five of the marsupial group.

Identification of Moraxella lacunata from pulmonary abscesses in three zoo herbivores.

Although Moraxella lacunata causes conjunctivitis, keratitis, endocarditis, and otolaryngitis in humans, its infection is rare in animals. We report three cases of asymptomatic pulmonary abscesses caused by M. lacunata in zoo herbivores, including two elks (Cervus canadensis) and a common eland (Taurotragus oryx). In all cases, macroscopic findings included coalescence of lung lobes and severe pulmonary abscesses filled with cheese-like materials in cysts. Microscopic findings included pneumonia characterized by marked fibrin exudates in alveolar spaces and infiltration of inflammatory cells. M. lacunata was identified in bacterial cultures from pulmonary abscesses using biochemical API 20NE system. M. lacunata is rarely isolated from zoo animals; however, herein, we describe the first report of pulmonary abscesses caused by M. lacunata infection.

The first clinical cases of Haemoproteus infection in a snowy owl (Bubo scandiacus) and a goshawk (Accipiter gentilis) at a zoo in the Republic of Korea.

This study reports two clinical cases of avian haemosporidian infection caused by a Haemoproteus sp., involving a snowy owl (Bubo scandiacus) and a goshawk (Accipiter gentilis), at a zoo. The snowy owl died after presenting with anorexia, depression and lethargy. A blood smear with Wright's staining confirmed Haemoproteus infection. Necropsy of the snowy owl revealed hypertrophy of the internal organs, including the liver, gallbladder, kidney and adrenal glands. The goshawk showed anorexia, depression and a lowered head position, and was diagnosed with a Haemoproteus infection based on a blood smear. The goshawk was completely cured by treatment with a combination of atovaquone and proguanil hydrochloride. Both cases showed decreased erythrocytes, hemoglobin and hematocrit values on complete blood count.

Sudden death of an Indian peafowl (Pavo cristatus) at a zoo due to non-pigmented Serratia marcescens infection.

A 16-year-old female Indian peafowl (Pavo cristatus) died two days after recognition of conjunctivitis in the right eye, anorexia and depression. Gross necropsy revealed a thick pseudomembrane under the eyelid and hydropericardium. Histopathological examination revealed hepatocellular necrosis, sinusoidal and vascular congestion and infiltrated inflammatory cells. Infiltration by inflammatory cells was noted in the epicardium. The lungs had mild interstitial pneumonia with the extensive congestion within the capillaries of the air sacs. Tubular interstitial congestion and necrosis was noted in the kidneys. Bacterial culture and nucleotide sequencing of the inflammatory specimens identified the causative agent as Serratia marcescens, an uncommon bacterium in birds. In summary, this study describes the sudden death of an Indian peafowl due to S. marcescens infection, which is rarely seen in animals.

Sudden death of a Siamese crocodile (Crocodylus siamensis) due to systemic aspergillosis.

A 4-year-old female Siamese crocodile (Crocodylus siamensis) housed at a zoo died without any prior clinical signs. During necropsy, numerous scattered, well-demarcated, yellowish-white, firm nodules were observed throughout the liver and lungs. Microscopic examination with periodic acid-Schiff staining revealed granulomatous inflammation in the liver and lungs. Liver granulomas were characterized by the presence of a connective tissue barrier and hyphae, and the centers of the granulomas showed signs of necrosis. Lung samples showed characteristics similar to those observed in the liver samples. The fungus was identified as Aspergillus fumigatus based on its appearance on Sabouraud dextrose agar, microscopic examination with lactophenol cotton blue staining and genetic sequencing. Therefore, zoo veterinarians should pay close attention to fungal infections in captive animals.

Novel Detection of Coxiella spp., Theileria luwenshuni, and T. ovis Endosymbionts in Deer Keds (Lipoptena fortisetosa).

We describe for the first time the detection of Coxiella-like bacteria (CLB), Theileria luwenshuni, and T. ovis endosymbionts in blood-sucking deer keds. Eight deer keds attached to a Korean water deer were identified as Lipoptena fortisetosa (Diptera: Hippoboscidae) by morphological and genetic analyses. Among the endosymbionts assessed, CLB, Theileria luwenshuni, and T. ovis were identified in L. fortisetosa by PCR and nucleotide sequencing. Based on phylogeny, CLB 16S rRNA sequences were classified into clade B, sharing 99.4% identity with CLB from Haemaphysalis longicornis in South Korea. Although the virulence of CLB to vertebrates is still controversial, several studies have reported clinical symptoms in birds due to CLB infections. The 18S rRNA sequences of T. luwenshuni and T. ovis in this study were 98.8-100% identical to those in GenBank, and all of the obtained sequences of T. ovis and T. luwenshuni in this study were 100% identical to each other, respectively. Although further studies are required to positively confirm L. fortisetosa as a biological vector of these pathogens, strong genetic relationships among sequences from this and previous studies suggest potential transmission among mammalian hosts by ticks and keds.