Abstract In the present report, we describe a case of sclerosing orbital pseudotumor in an 11‐year‐old castrated male American Shorthair cat. Ophthalmic exam showed lagophthalmos, retracted right upper eyelid, and resistant to retropulsion in his right eye. Under magnetic resonance imaging (MRI) scans, increased volume of the extraocular muscles (EOMs) of the right eye was prominent. Immunosuppressive dosage of prednisolone partially ameliorated the clinical signs, but some clinical signs were still gradually progressive or persistent. In the second MRI scan, decreased diameter of the thickened right extraocular muscles was found. After the third MRI scan, enucleation of the right eye was performed due to substantial adverse effects of systemic steroid therapy. Histopathological examination revealed no evidence of neoplastic transformation nor infection. Feline restrictive orbital myofibroblastic sarcoma (FROMS) was therefore excluded, suggesting unknown causes of extensive fibrotic changes in the right orbit of the affected cat.

Porcine circovirus type 2 (PCV2) is the major cause of serious porcine circovirus-associated diseases (PCVADs) causing significant economic losses in the pig industry. The PCV2 vaccines have proved effective by lowering serum and tissue viral loads. However, little is known about its effects on the cellular functions of viral target cells in pigs. For this, blood samples from 38 vaccinated and 34 non-vaccinated, 12–14 week-old, clinically healthy and PCV2-infected pigs in a commercial farm were sampled and anticoagulated. The viral load in plasma and the cellular proliferation and mRNA expression levels of IL-1 [Formula: see text], IL-2, IL-4, IL-8, IL-10 and IFN-[Formula: see text] of peripheral blood mononuclear cells (PBMCs) with or without concanavalin A (Con A) and PCV2 stimulation in vitro were then investigated by real-time polymerase chain reaction and flow cytometry. Compared with the non-vaccinated pigs, the average viral loads in the plasma and cultivated PBMCs were reduced in the PCV2-vaccinated pigs. Significant differences in the aspects of increased cell proliferation, decreased average viral loads in PBMCs three days after Con A stimulation, increased mRNA expression levels of IL-2 and IFN-[Formula: see text] after re-exposure to PCV2, and decreased levels of IL-1 [Formula: see text], IL-4 and IL-8 after Con A or PCV2 stimulation were also noted in the PCV2-vaccinated pigs. The results suggested that the PCV2-vaccinated pigs were likely to develop better cell-mediated immunity than the non-vaccinated pigs.

AbstractThe toxic effects of aflatoxin B1 (AFB1) on the physiological functions of swine alveolar macrophages (SAM) were investigated. Freshly isolated SAM were incubated with various AFB1 concentrations (1.6 × 10–1 – 1.6 × 105 nmol/L) and time periods, and their phagocytic ability, synthesis of DNA, RNA and protein, and cell activation by lipopolysaccharide (LPS), were analysed. Results demonstrated that a significant (p < .05) reduction (60%) in Staphylococcus aureus uptaken by SAM appeared 3 hr after AFB1 (>16 nmol/L) treatment. The synthesis of DNA, RNA and protein were markedly reduced, among which DNA and protein synthesis were affected more noticeably. The activation of SAM by LPS was significantly (p < .05) suppressed when the concentration of AFB1 reached 1.6 × 103 nmol/L. In general, most of the analysed effects were more prominent as AFB1 concentration or incubation period increased. Taken together, AFB 1could elicit significant adverse effects on the physiological functions of SAM. Exposure of pigs to aflatoxin‐contaminated feed may increase their susceptibility to various secondary infections.

Background The microenvironment within solid malignant tumors, including feline mammary gland carcinomas (FMGCs), is commonly hypoxic, possibly due to the lack of functional blood vessels in rapidly proliferating neoplastic tissue. Malignant cells can undergo genetic and adaptive changes that prevent them from dying due to oxygen deprivation through expressions of hypoxia-inducible factor 1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF). Therefore, HIF-1α and VEGF are ideal biomarkers for cancer therapy and prognostic evaluation. The aims of this study were to evaluate the expression of HIF-1α and VEGF in feline mammary carcinomas and analyze their correlations with clinical and pathological factors, such as clinical stage, histologic grading, regional metastasis, and overall survival rate. Results Paraffin-embedded tissue samples collected from 72 cats with FMGCs were retrospectively studied. Histologic pattern and histologic grading (Elston and Ellis grading system) of these FMGCs were determined. Our data indicated that grade II tubulopapillary carcinomas (43/72, 59.7%) prevailed in this study, and most FMCGs showed apparent necrosis, squamous metaplasia, and intratumoral stromal response. According to the results of immunohistochemical (IHC) stainings performed in tissue microarrays (TMAs), HIF-1α and VEGF overexpressions were respectively noted in 69.4% (50/72) and 77.8% (56/72) of FMGC cases. Chi-square test showed no correlation of HIF-1α overexpression with clinical and pathological factors. VEGF overexpression was significantly correlated with histologic pattern ( p = 0.021), stromal response ( p = 0.048), squamous metaplasia ( p = 0.001), and lymphovascular invasion ( p = 0.007). However, neither HIF-1α nor VEGF overexpression was correlated with histologic grading and metastasis. Of 38 cats with 1-year follow-up, IHC stainings of HIF-1α and VEGF were performed on whole tissue sections. The results showed that overexpression of HIF-1α was significantly correlated with the overall survival rate ( p < 0.05) (log-rank test), whereas there was no significant correlation between VEGF overexpression and overall survival rate. Conclusions This study suggests that the overexpression of HIF-1α may indicate poor prognosis/overall survival rate in cats with FMGCs. Developing compounds that inhibit HIF-1α may be a potential approach to FMGC treatment.

A 16-year-old male Russian blue cat was presented with acute onset of paraparesis of the forelimbs that progressed to tetraparesis. Neurological examination revealed non-ambulatory tetraparesis with decreased postural reactions in all four limbs. Magnetic resonance imaging revealed multifocal nerve root swelling on the right at C6/C7 and C7/T1, while ultrasonography demonstrated swelling of the right brachial plexus. To understand the cause of the nerve swelling, the right musculocutaneous nerve arising from the brachial plexus and the pectoralis muscle were biopsied. Histologically, there was evidence of neurolymphomatosis (neurotropic lymphoma) with Wallerian degeneration and denervation atrophy of myofibres. The neoplastic lymphoid cells expressed CD79a, CD20 and CD56. Based on these findings, a diagnosis of B-cell neurolymphomatosis was made. Expression of CD56, synonymous with neural cell adhesion molecule, is rare in B-cell lymphomas and has not been reported in feline B-cell lymphomas or feline neurolymphomatosis. CD56 expression was suspected to have played an important role in neurotropism of the neoplastic cells in this case.

A one-year-old male Maltese terrier presented with mild ataxia and disorientation for 4 months. Over time, clinical signs progressed from paraparesis to non-ambulatory tetraparesis, voice change and dysphagia. Histological examination revealed concurrent leukoencephalomyelitis and polyneuritis. Infectious etiologies, including dengue, Japanese encephalitis, Zika, canine distemper, pseudorabies, rabies, toxoplasmosis, neosporosis, leishmaniasis, and encephalitozoonosis, were ruled out by PCR and/or immunohistochemical (IHC) staining. IHC tested on neurological tissues highlighted a heterogeneous population of infiltrating T and B lymphocytes admixed macrophages. Therefore, this case was diagnosed with current leukoencephalomyelitis and polyneuritis, resembling combined central and peripheral demyelination (CCPD), an autoimmune inflammatory demyelinating disease affecting both the CNS and PNS in humans.

Since 2010, newly identified variants of porcine epidemic diarrhoea virus (PEDV) have caused high mortality in neonatal piglets which has devastated the swine industry. The spike (S) glycoprotein of PEDV contains multiple neutralizing epitopes and is a major target for PEDV neutralization and vaccine development. To understand the antigenicity of the new PEDV variant, we characterized the neutralizing epitopes of a new genotype 2b PEDV isolate from Taiwan, PEDV Pintung 52 (PEDV-PT), by the generation of neutralizing monoclonal antibodies (NmAbs). Two NmAbs, P4B-1, and E10E-1-10 that recognized the ectodomain of the full-length recombinant PEDV S protein and exhibited neutralizing ability against the PEDV-PT virus were selected. Recombinant truncated S proteins were used to identify the target sequences for the NmAbs and P4B-1 was shown to recognize the C-terminus of CO-26K equivalent epitope (COE) at amino acids (a.a.) 575-639 of the PEDV S. Interestingly, E10E-1-10 could recognize a novel neutralizing epitope at a.a. 435-485 within the S1A domain of the PEDV S protein, whose importance and function are yet to be determined. Moreover, both NmAbs could not bind to linearized S proteins, indicating that only conformational epitopes are recognized. This data could improve our understanding of the antigenic structures of the PEDV S protein and facilitate future development of novel epitope-based vaccines.

Background Chronic inflammation has been implicated in sarcomagenesis. Among various factors, activation of nuclear factor-kappa B (NF-κB) signaling pathway has been documented being able to target genes associated with tumor progression and up-regulate the expression of tumor-promoting cytokines and survival genes in several human solid tumors. Feline injection sites sarcomas (FISS) are malignant entities derived from the mesenchymal origin. The disease has been considered to be associated with vaccine adjuvant, aluminum, which serves as a stimulus continuously inducing overzealous inflammatory and immunologic reactions. To understand the contribution of NF-κB in FISS, detection of activated NF-κB in paraffin-embedded specimens, in vitro establishment of primary cells derived from FISS, and evaluation of the effects of the NF-κB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), on primary tumor cells were conducted. Results In this study, nuclear expression of NF-κB p65 was detected in 83.3% of FISS cases and not correlated with tumor grading, sex, and age. Primary cells derived from FISS in three cats exhibiting same immunohistochemical characteristics as their original tumor were successfully established. The NF-κB inhibitor, DHMEQ, was able to prevent nuclear translocation of NF-κB p65, inhibit cell proliferation, migration, and colonization in dosage-dependent manners, and induce cell apoptosis in these primary FISS cells. Conclusions High expression rate of nuclear NF-κB p65 in FISS cases and dose-dependent inhibitory effects on the growth of FISS primary cells treated with NF-κB inhibitor suggested that NF-κB might be a potential molecular therapeutic target for FISS.

Plasmacytoid and rhabdoid variants of urothelial carcinomas (UCs) of the urinary bladder have been described in humans with plasma cell–like or rhabdoid cellular appearance and aggressive clinical outcome. Canine UC of the bladder is generally classified as papillary/nonpapillary and infiltrating/noninfiltrating with limited information regarding other histological patterns. We report 3 cases of UC of the urinary bladder showing a unique discohesive cellular morphology with malignant behavior resembling the human plasmacytoid and rhabdoid variants of UC, which may raise some difficulties in diagnosis. Epithelial-mesenchymal transition and reduced E-cadherin expression were revealed by immunohistochemistry in 2 cases, possibly explaining the discohesive and invasive behavior of the tumor cells. The findings broaden the morphological spectrum as well as the distinct clinical features of canine UC of the urinary bladder.

Background Silver nanoparticles (AgNPs) have been widely used in many commercial products due to their excellent antibacterial ability.The AgNPs are released into the environment, gradually accumulate in the ocean, and may affect animals at high trophic level, such as cetaceans and humans, via the food chain.Hence, the negative health impacts caused by AgNPs in cetaceans are of concern.Cytokines play a major role in the modulation of immune system and can be classified into two types, Th1 and Th2.Th1/Th2 balance can be evaluated by the ratios of their polarizing cytokines (i.e., interferon [IFN]-γ/ Interleukin [IL]-4), and animals with imbalanced Th1/Th2 response may become more susceptible to certain kinds of infection.Therefore, the present study evaluated the in vitro cytokine responses of cetacean peripheral blood mononuclear cells (cPBMCs) to 20 nm citrate-AgNPs (C-AgNP 20 ) by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR).Methods Blood samples were collected from 6 captive common bottlenose dolphins (Tursiops truncatus).The cPBMCs were isolated and utilized for evaluating the in vitro cytokine responses.The cytokines evaluated included IL-2, IL-4, IL-10, IL-12, interferon (IFN)-γ, and tumor necrosis factor (TNF)-ɑ.The geometric means of two housekeeping genes (HKGs), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and β2-microglobulin (B2M), of each sample were determined and used to normalize the mRNA expression levels of target genes. ResultsThe ratio of late apoptotic/necrotic cells of cPBMCs significantly increased with or without concanavalin A (ConA) stimulation after 24 h of 10 μg/ml C-AgNP 20 treatment.At 4 h of culture, the mRNA expression level of IL-10 was significantly decreased with 1 μg/ml C-AgNP 20 treatment.At 24 h of culture with 1 μg/ml C-AgNP 20 , the mRNA expression levels of all cytokines were significantly decreased, with the exceptions of IL-4 and IL-10.The IFN-γ/IL-4 ratio was significantly decreased at 24 h of culture with 1 μg/ml C-AgNP 20 treatment, and the IL-12/IL-4 ratio was significantly decreased at 4 or 24 h of culture with 0.1 or 1 μg/ml C-AgNP 20 treatment, respectively.Furthermore, the mRNA expression level of TNF-α was significantly decreased by 1 μg/ml C-AgNP 20 after 24 h of culture. DiscussionThe present study demonstrated that the sublethal dose of C-AgNP20 (≤ 1 μg/ml) had an inhibitory effect on the cytokine mRNA expression levels of cPBMCs with the evidence of Th2 cytokine bias and significantly decreased the mRNA expression level of TNF-α.Th2 cytokine bias is associated with enhanced immunity against parasites but decreased immunity to intracellular microorganisms.TNF-α is a contributing factor for the inflammatory response against the infection of intracellular pathogens.In

Silver, such as silver nanoparticles (AgNPs), has been widely used in commercial products and may be released into the environment. The interaction between Ag deposition and biological systems is raising serious concerns because of one health consideration. Cetaceans, as the top predators of the oceans, may be exposed to Ag/Ag compounds and suffer negative health impacts from the deposition of these compounds in their bodies. In the present study, we utilized autometallography (AMG) to localize the Ag in the liver and kidney tissues of cetaceans and developed a model called the cetacean histological Ag assay (CHAA) to estimate the Ag concentrations in the liver and kidney tissues of cetaceans. Our results revealed that Ag was mainly located in hepatocytes, Kupffer cells and the epithelial cells of some proximal renal tubules. The tissue pattern of Ag/Ag compounds deposition in cetaceans was different from those in previous studies conducted on laboratory rats. This difference may suggest that cetaceans have a different metabolic profile of Ag, so a presumptive metabolic pathway of Ag in cetaceans is advanced. Furthermore, our results suggest that the Ag contamination in cetaceans living in the North-western Pacific Ocean is more severe than that in cetaceans living in other marine regions of the world. The level of Ag deposition in cetaceans living in the former area may have caused negative impacts on their health condition. Further investigations are warranted to study the systemic Ag distribution, the cause of death/stranding, and the infectious diseases in stranded cetaceans with different Ag concentrations for comprehensively evaluating the negative health effects caused by Ag in cetaceans.

Silver nanoparticles (AgNPs) have been extensively used in commercial products, including textiles, cosmetics, and health care items, due to their strong antimicrobial effects. They also may be released into the environment and accumulate in the ocean. Therefore, AgNPs are the major source of Ag contamination, and public awareness of the environmental toxicity of Ag is increasing. Previous studies have demonstrated the bioaccumulation (in producers) and magnification (in consumers/predators) of Ag. Cetaceans, as the apex predators of ocean, may have been negatively affected by the Ag/Ag compounds. Although the concentrations of Ag/Ag compounds in cetacean tissues can be measured by inductively coupled plasma mass spectroscopy (ICP-MS), the use of ICP-MS is limited by its high capital cost and the requirement for tissue storage/preparation. Therefore, an autometallography (AMG) method with an image quantitative analysis by using formalin-fixed, paraffin-embedded (FFPE) tissue may be an adjuvant method to localize Ag distribution at the suborgan level and estimate the Ag concentration in cetacean tissues. The AMG positive signals are mainly brown to black granules of various sizes in the cytoplasm of proximal renal tubular epithelium, hepatocytes, and Kupffer cells. Occasionally, some amorphous golden yellow to brown AMG positive signals are noted in the lumen and basement membrane of some proximal renal tubules. The assay for estimating the Ag concentration is named the Cetacean Histological Ag Assay (CHAA), which is a regression model established by the data from image quantitative analysis of the AMG method and ICP-MS. The use of AMG with CHAA to localize and semi-quantify heavy metals provides a convenient methodology for spatio-temporal and cross-species studies.

Background Silver nanoparticles (AgNPs) have been widely used in many commercial products due to their excellent antibacterial ability. The AgNPs are released into the environment, gradually accumulate in the ocean, and may affect animals at high trophic level, such as cetaceans and humans, via the food chain. Hence, the negative health impacts caused by AgNPs in cetaceans are of concern. Cytokines play a major role in the modulation of immune system and can be classified into two types, Th1 and Th2. Th1/Th2 balance can be evaluated by the ratios of their polarizing cytokines (i.e., interferon [IFN]-γ/ Interleukin [IL]-4), and animals with imbalanced Th1/Th2 response may become more susceptible to certain kinds of infection. Therefore, the present study evaluated the in vitro cytokine responses of cetacean peripheral blood mononuclear cells (cPBMCs) to 20 nm citrate-AgNPs (C-AgNP20) by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Methods Blood samples were collected from 6 captive common bottlenose dolphins (Tursiops truncatus). The cPBMCs were isolated and utilized for evaluating the in vitro cytokine responses. The cytokines evaluated included IL-2, IL-4, IL-10, IL-12, interferon (IFN)-γ, and tumor necrosis factor (TNF)-ɑ. The geometric means of two housekeeping genes (HKGs), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and β2-microglobulin (B2M), of each sample were determined and used to normalize the mRNA expression levels of target genes. Results The ratio of late apoptotic/necrotic cells of cPBMCs significantly increased with or without concanavalin A (ConA) stimulation after 24 h of 10 μg/ml C-AgNP20 treatment. At 4 h of culture, the mRNA expression level of IL-10 was significantly decreased with 1 μg/ml C-AgNP20 treatment. At 24 h of culture with 1 μg/ml C-AgNP20, the mRNA expression levels of all cytokines were significantly decreased, with the exceptions of IL-4 and IL-10. The IFN-γ/IL-4 ratio was significantly decreased at 24 h of culture with 1 μg/ml C-AgNP20 treatment, and the IL-12/IL-4 ratio was significantly decreased at 4 or 24 h of culture with 0.1 or 1 μg/ml C-AgNP20 treatment, respectively. Furthermore, the mRNA expression level of TNF-α was significantly decreased by 1 μg/ml C-AgNP20 after 24 h of culture. Discussion The present study demonstrated that the sublethal dose of C-AgNP20 (≤ 1 μg/ml) had an inhibitory effect on the cytokine mRNA expression levels of cPBMCs with the evidence of Th2 cytokine bias and significantly decreased the mRNA expression level of TNF-α. Th2 cytokine bias is associated with enhanced immunity against parasites but decreased immunity to intracellular microorganisms. TNF-α is a contributing factor for the inflammatory response against the infection of intracellular pathogens. In summary, our data indicate that C-AgNP20 suppresses the cellular immune response and thereby increases the susceptibility of cetaceans to infection by intracellular microorganisms.

The porcine epidemic diarrhea virus (PEDV) poses a great threat to the global swine industries and the unreliable protection induced by the currently available vaccines remains a major challenge. We previously generated a genogroup 2b (G2b) PEDV Taiwan Pintung 52 (PEDVPT) strain, PEDVPT-P96, and determined its promising host immune response against the virulent PEDVPT-P5 strain. To study the attenuation determinants of PEDVPT-P96 and establish a PEDVPT-P96-based recombinant vector as a vaccine platform for further antigenicity modification, iPEDVPT-P96, a full-length cDNA clone of PEDVPT-P96, was established. Comparing to the parental PEDVPT-P96 virus, the iPEDVPT-P96 virus showed efficient replication kinetics with a delayed decline of viral load and similar but much more uniform plaque sizes in Vero cells. In the 5-week-old piglet model, fecal viral shedding was observed in the PEDVPT-P96-inoculated piglets, whereas those inoculated with iPEDVPT-P96 showed neither detectable fecal viral shedding nor PEDV-associated clinical signs. Moreover, inoculation with iPEDVPT-P96 elicited comparable levels of anti-PEDV specific plasma IgG and fecal/salivary IgA, neutralizing antibody titers, and similar but less effective immunoprotection against the virulent PEDVPT-P5 challenge compared to the parental PEDVPT-P96. In the present study, an infectious cDNA clone of an attenuated G2b PEDV strain was successfully generated for the first time, and the in vitro and in vivo data indicate that iPEDVPT-P96 is further attenuated but remains immunogenic compared to its parental PEDVPT-P96 viral stock. The successful development of the iPEDVPT-P96 cDNA clone could allow for the manipulation of the viral genome to study viral pathogenesis and facilitate the rapid development of effective vaccines.

Devastating outbreaks of porcine epidemic diarrhea (PED) started in China in late 2010 and rapidly spread to North America and Asia causing severe diarrhea and high mortality in neonatal piglets, indicating that a new generation of vaccine against porcine epidemic diarrhea virus (PEDV) is urgently needed. In the present study, to mimic the native spike (S) glycoprotein, a stable cell line producing the trimeric ectodomain of S glycoprotein of the PEDV Pintung-52 (PEDV-PT) strain was successfully established by incorporating T4 bacteriophage foldon sequence of fibritin trimerization domains at the C-terminal end and replacing the signal peptide of S protein with the tissue plasminogen activator signal peptide sequence at the N terminal end. The trimeric structure, bio-reactivity to PEDV-specific antibodies, and the N-glycosylation level of the recombinant S protein were characterized. To induce systemic and mucosal immunity, conventional 5-week-old piglets were immunized with the trimeric S glycoprotein combined with the B subunit of Escherichia coli heat-labile enterotoxin (LTB) by the intramuscular (IM) route. As compared with the control group, all piglets in the S protein-LTB immunized (IM PEDV S-LTB) group generated systemic PEDV S-specific IgG and neutralizing antibody in blood but a low level of fecal PEDV-specific IgA and limited protection against challenge of PEDV-PT strain. Our results suggest that the recombinant PEDV trimeric S glycoprotein could be a potential subunit vaccine candidate against PEDV, but IM immunization with LTB as the adjuvant provided insufficient protection. The development of a vaccine regimen for inducing mucosal immunity is an important task for generating a successful subunit vaccine against PEDVs.

An outbreak of reproductive failure in a pig farm in Taiwan was investigated. Coinfection with porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) was diagnosed in a stillborn pig by histopathology, polymerase chain reaction, and immunohistochemistry, and should be considered as a cause of reproductive failure.

An adult female California king snake (Lampropeltis getula californiae) housed in Taipei Zoo was presented with a 2-week history of anorexia, fatigue and abdominal swelling. Exploratory laparotomy revealed a gastric mass with two circular perforations and multiple mottled white to beige protuberances along the mucosal surface. Histologically, the gastric mass showed an invasive, transmural growth of epithelial cells arranged in nests, lobules, acini and sheets in the mucosa and submucosa that progressively transformed into signet ring cells in the muscularis externa and subserosa. All of the neoplastic cells expressed pan-cytokeratin immunohistochemically. Based on the World Health Organization histological criteria, a diagnosis of diffuse-type gastric mucinous and signet ring cell adenocarcinoma was made.

Silver nanoparticles (AgNPs) have been extensively used and are considered as an emerging contaminant in the ocean. The environmental contamination of AgNPs is expected to increase greatly over time, and cetaceans, as the top ocean predators, will suffer the negative impacts of AgNPs. In the present study, we investigate the immunotoxicity of AgNPs on the leukocytes of cetaceans using several methods, including cytomorphology, cytotoxicity, and functional activity assays. The results reveal that 20 nm Citrate-AgNPs (C-AgNP20) induce different cytomorphological alterations and intracellular distributions in cetacean polymorphonuclear cells (cPMNs) and peripheral blood mononuclear cells (cPBMCs). At high concentrations of C-AgNP20 (10 and 50 μg/ml), the time- and dose-dependent cytotoxicity in cPMNs and cPBMCs involving apoptosis is demonstrated. C-AgNP20 at sub-lethal doses (0.1 and 1 μg/ml) negatively affect the functional activities of cPMNs (phagocytosis and respiratory burst) and cPBMCs (proliferative activity). The current study presents the first evidence of the cytotoxicity and immunotoxicity of AgNPs on the leukocytes of cetaceans and improves our understanding of environmental safety concerning AgNPs. The dose-response data of AgNPs on the leukocytes of cetaceans are invaluable for evaluating the adverse health effects in cetaceans and for proposing a conservation plan for marine mammals.

The group 2b (G2b) porcine epidemic diarrhea virus (PEDV) that emerged in 2013 has since caused devastating diseases and economic loss. The full-length genome of the G2b Taiwan PEDV-Pintung 52 (PEDV-PT) strain and its intestinal tropism by evaluating the pathological changes in the original PEDV-PT infected field piglet and orally inoculation of either 10, 103, or 105 50% tissue culture infective dose/mL (TCID50/mL) of the plaque-purified PEDV-PT-Passage 5 (P5) in 7-day-old conventional piglets were analyzed. Phylogenetic analysis of the full-length genome indicated that the G2b Taiwan PEDV-PT strain was closely related to the North American G2b PEDV strains. Some pathological features of the G2b Taiwan PEDV-PT infection, including the absence of lesions and antigen signal in the crypt epithelial cells of the jejunum and ileum and in the villus enterocytes of the duodenum and colon, were different from those of infections by the North American G2b PEDV strains. This difference in the intestinal tropism of the G2b Taiwan PEDV-PT strain highlights the importance of studying the pathogenicities of different PEDV variants. Moreover, similar distributions of PEDV antigens and lesions in the G2b Taiwan PEDV-PT infected field piglet and its plaque-purified isolate, PEDV-PT-P5, inoculated piglets indicating that the plaque-purified PEDV-PT-P5 viral stock could facilitate the preclinical evaluation of vaccines and other interventions aimed at preventing the G2b PEDV infection.