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Rabies is a type of acute fetal encephalitis caused by rabies virus (RABV). While it becomes incurable after symptom onset, it can be prevented by post-exposure prophylaxis (PEP) during the long incubation period. While preclinical diagnosis aids the appropriate PEP administration, it is mostly non-feasible owing to the absence of viremia or a specific antibody response during the incubation period. Here, we attempted to identify a serum biomarker for the preclinical diagnosis of rabies. Using the serum from a mouse inoculated intramuscularly (i.m.) with 5×105 focus-forming units (FFU) of recombinant RABV expressing red firefly luciferase (1088/RFLuc) immediately before symptom onset, we conducted two-dimensional differential gel electrophoresis, followed by mass spectrometry, and confirmed that apolipoprotein A1 (ApoA1) was upregulated. ELISA showed that the serum ApoA1 and specific antibody levels increased during the incubation period and on the day of symptom onset. Since a lower infectious dose can be used to induce the unstable and long incubation period generally observed in natural infection, we examined the ApoA1 level in mice inoculated i.m. with 103 FFU of 1088/RFLuc by monitoring viral dynamics using in vivo imaging. The serum ApoA1 and specific antibody levels were upregulated in 50% and 58.3% of mice exhibiting robust RABV replication, respectively, but not in mice exhibiting weak RABV replication. In addition, it was reported that ApoA1 was found to be a biomarker for neuronal damage. Additional biomarker candidates will be needed for the effective preclinical diagnosis of rabies. This article is protected by copyright. All rights reserved.

The study aimed to evaluate pork loin quality after short-term (ten-day) storage in a supercooling refrigerator. Pork loin samples were stored for 10 days in a traditional refrigerator (control) and a commercially available supercooling refrigerator (SC). Pork quality measurements included meat pH, meat color, water holding capacity (drip loss and cooking loss), tenderness (hardness), and a sensory evaluation. Temperature changes of 0.45 ± 0.2°C and 0.02 ± 0.25°C occurred in the control and the SC, respectively, during 10 days of storage. The temperature in the SC chamber did not remain below freezing point, failing to meet expectations. Regarding the pork quality measurements, only the drip losses in the control and the SC were significantly different (4.45% vs. 2.59%, p < 0.01) after 10 days of storage. There were no significant differences between the two types of refrigerator in terms of the other measurements. Additionally, the overall acceptability of the pork loin did not vary significantly between the control and the SC when the sensory evaluation was performed. Therefore, a commercial SC could prove beneficial in terms of water holding capacity during the short-term storage of meat. Further research should be performed to evaluate quality changes that occur during long-term storage of meat in SC s and evaluate a wide range of meat, such as beef and chicken.

Background The Philippines is one of the major endemic countries for canine rabies in Southeast Asia. However, detailed description and analysis of laboratory-confirmed animal rabies are limited. Highly accurate surveillance requires a thorough understanding of the target area-specific problems and obstacles. Therefore, we aim to describe and analyze the rabies suspect animals in Central Luzon, Philippines, to clarify the characteristics of management and clinical signs by conducting interviews with the owners. Methods We prospectively collected information on the rabies suspect animals submitted to the Regional animal laboratory in Central Luzon through passive laboratory-based rabies surveillance between 1st April 2019 and 30th September 2020. We performed active interviews directly or telephonically with the owner. The direct fluorescent antibody test was performed on the hippocampus, brain stem, and cerebellum for laboratory confirmation. Descriptive statistics were used to characterize the number of rabies cases according to management methods and characteristics of suspected animals during the observation period. Clinical symptoms of suspected rabid animals were analyzed by univariate logistic regression analysis. Results There were 292 sample submissions during the study period. Of these, 160 were positive for dFAT. Samples of pet animals (85.3%) provided by owners or their acquaintances (59.2%) accounted for the majority of laboratory confirmed cases. Case mapping showed that more rabies-suspected cases were sent from areas near the regional laboratory than from those far from the laboratory, despite the incidence of rabies being high in these areas. The management and clinical symptoms of 227 animal cases showed that most owners were managing their animals at home and were allowing them to roam outside (69.6%) and be unvaccinated (78.9%). Rabid animals were more likely to manifest aimless running, restlessness, and agitation. Conclusions Our study provided some features of animals with laboratory-confirmed rabies in Central Luzon. However, most of the samples were submitted from areas near the rabies diagnosis laboratory, and the number of samples submitted from remote areas was low. To improve the surveillance capacity, it is necessary to increase sample submissions from remote areas.

This study aimed to investigate the influence of Astragalus membranaceus (AM), Adenophora triphylla (AT), and Ulmus pumila (UP) extracts on the quality traits, palatability, and storage stability of sous-vide (SV) cooked chicken breasts. Chicken breasts were marinated in AM, AT, or UP extracts for 1 h, and then consistently cooked at a constant temperature of 60°C for 2 h. SV cooked chicken breasts with the UP extract exhibited lower lightness and higher yellowness values on the surface region compared to those with the AM and AT extracts (p 0.05), although the UP group had lower color acceptability (p 0.05). Owing to these results regarding overall sensory acceptability, samples from the UP group were more preferred by the trained panelists compared to samples from the control group (p

The rabies virus is one of the most neurotropic of all viruses infecting mammals. During the terminal phases of infection, the virus spreads to peripheral tissues, including the skin. The external skin of the nose, called the nasal planum, is a sensory organ where numerous nerve bundles and terminal nerves are distributed. Therefore, the nasal planum is expected to serve as a postmortem diagnostic material. However, the distribution of rabies virus antigens in the nasal planum in rabid animals has not yet been studied. In this study, the nasal planum was obtained from 45 rabid dogs. In all rabid dogs, the viral antigen was detected in the peripheral nerve tissues, Merkel cells, and squamous cells. The viral antigen in the epidermis exhibited three patterns: first, a diffuse positive pattern from the basal layer to the squamous layer; second, a reticular positive pattern along the cell membrane in the squamous layer; and third, a basal layer pattern of the epidermis. In the dermis, viral antigens were detected more often in lamellated corpuscles just beneath the rete pegs. These results suggest that the nasal planum could serve as a useful alternative source for postmortem diagnosis in rabies endemic countries.

This study investigated the combined effects of cooking temperature and time on the meat and eating quality characteristics of the sous-vide chicken breast. For the control group, chicken breast samples were cooked in a convection oven until the internal temperature reached 71°C. Each sample for sous-vide cooking was vacuum packaged and then cooked under continuous thermocontrolled conditions in a water bath at 6 combinations of cooking temperature (60 and 70°C) and time (1, 2, and 3 h). Sous-vide cooked chicken meat at 60°C for 1 h (SV60-1h) showed lower cooking loss (6.58 vs. 26.5%, P

We previously reported a novel diagnostic method using follicle-sinus complexes (FSCs) in the muzzle skin for postmortem diagnosis of rabies in dogs. However, whether this method works in other animal species remains unclear. Here, FSCs were collected from a wolf, a red fox, 2 bats, and a cat, and examined for the presence of viral antigen, viral mRNA, and viral particles. Viral antigen and viral mRNA were confirmed in Merkel cells (MCs) in FSCs of all species. Electron microscopy performed using only samples from wolf and cat confirmed viral particles in MCs of FSCs. These results suggested that this novel diagnostic method using FSCs might be useful for detection of rabies not only in domestic but also wild animals.

A 3-month-old male Scottish Fold kitten with pleural fluid and low ratio of albumin to globulin (A/G ratio) was brought to our small animal hospital. Since RNA from the type I feline coronavirus (FCoV) were detected in drained pleural fluid, the cat was tentatively diagnosed with effusive feline infectious peritonitis (FIP). Following the administration of itraconazole and prednisolone, the A/G ratio increased, and the pleural fluid mostly disappeared. The fecal FCoV levels temporarily decreased. However, the cat showed neurological manifestations and was eventually euthanized due to status epilepticus after 38 days of treatment. In conclusion, itraconazole partly exerted a beneficial effect in a cat with FIP. However, further investigation of a possible role of itraconazole in FIP treatment is warranted.

A 7-month-old castrated French bulldog was presented with a left-sided mandibular tumor. The initial tumor biopsy diagnosis was ameloblastoma. The owner brought this dog the Kitasato University Veterinary Teaching Hospital for more detailed examination and treatment. Computed tomography revealed a tumor on the left lateral mandibular gingiva from the caudal third of the incisor tooth to the right canine tooth, associated with severe amorphous osteolysis of the mandibular bone. The tumor was surgically excised and diagnosed as papillary squamous cell carcinoma. Currently, 2514 d (6.9 y) since the operation, the dog is healthy, without recurrence. Key clinical message: Although papillary squamous cell carcinoma is rare, many cases have been reported in the oral cavity of medium-to large-sized dogs. Based on this report, papillary squamous cell carcinoma can occur in small dogs such as young French bulldogs and a good prognosis can be achieved with proper resection.

Inspired by the analogy between the application process of cosmetics and large amplitude oscillatory shear (LAOS), we suggest a novel predictive model for the spreadability of cosmetic formulations via LAOS analysis and machine learning techniques. Rheological measurements of cosmetic formulations, including the transient elastic and viscous moduli from the sequence of physical process (SPP) analysis, were selected as features for the predictive models, and the spreadability of each formulation that is quantitatively rated by trained panels was set up as the target variable. First, multiple linear regression prediction models are derived, and it was shown that the LAOS-SPP parameters were more effective features than other rheological parameters that were conventionally related to spreadability of cosmetics. Additionally, a non-linear prediction model was developed based on the random forest regressor algorithm, considering the possibility of the nonlinear correlation between rheological measurements and spreadability. The random forest regressor model showed better performance than the linear regression model, and the LAOS-SPP parameters were found to be more effective features for the random forest regressor model as in the multiple linear regression model. The correlation between the LAOS-SPP parameters and the spreadability is interpreted in terms of the rheological transition during rubbing process of cosmetics. Our findings indicate the importance of the nonlinear rheological behavior in the texture perception mechanism of cosmetics, and how rheological measurements can be combined with machine learning techniques to solve complicated problems.

In the present study, follicle-sinus complexes (FSCs) were harvested from the muzzle skin of 123 dogs with suspected canine rabies, and the sensitivity and specificity of FSC analysis were compared with those of brain tissue immunohistochemistry analysis. In the FSCs, viral antigen was detected from Merkel cells. Sensitivity was 97.3%, specificity was 100%, and the coefficient κ was 0.88. These results reconfirm that FSCs are very useful for the postmortem diagnosis of canine rabies, and suggest that 5 FSCs can yield results that are almost equivalent to those derived from brain tissue analysis in rabid dogs.

This study aimed to investigate the influence of Astragalus membranaceus (AM), Adenophora triphylla (AT), and Ulmus pumila (UP) extracts on the quality traits, palatability, and storage stability of sous-vide (SV) cooked chicken breasts. Chicken breasts were marinated in AM, AT, or UP extracts for 1 h, and then consistently cooked at a constant temperature of 60°C for 2 h. SV cooked chicken breasts with the UP extract exhibited lower lightness and higher yellowness values on the surface region compared to those with the AM and AT extracts (p0.05), although the UP group had lower color acceptability (p0.05). Owing to these results regarding overall sensory acceptability, samples from the UP group were more preferred by the trained panelists compared to samples from the control group (p

As the representative targeted anticancer drug for colon cancer patients, cetuximab is the EGFR targeted therapeutic antibody and used for treatment of KRAS wild type cancers. Even some patient with KRAS wt gene did not respond cetuximab. However, there is no treatment available for cetuximab-resistant patient group, which is almost 50% of KRAS WT gene holders. Recently, our team identified cetuximab primary resistant related proteins named as mtRTK (mutant receptor tyrosine kinase) by array analysis based cetuximab responder or non-responder colon cancer patient tissues. We investigated mtRTK’s oncogenic potential as a novel anti-cancer target. A large proportion of colon cancer patients (36.2% Caucasian, 56.9% Korean) expressed the mtRTK was identified, using the sequencing analysis of patient samples. Based on these results, our efforts have led to the discovery of WM-S1, mtRTK inhibitor, which is the first mtRTK inhibitor in clinical development. The potent enzyme inhibitor showed a high anticancer activity confirmed in Patient-Derived Cells (PDC) and Patient-Derived Xenograft (PDX) animal models expressing the mutation. In preclinical studies demonstrate that WM-S1 is well tolerated in rats and dogs. Furthermore, WM-S1 has potent anticancer activities for various solid tumor (NSCLC, cholangiocarcinoma, etc.) including activated mtKRAS colon cancer expressing the mtRTK. Currently we are investigating WM-S1 in a phase 1a trial in AUS, which is the first mtRTK inhibitor in clinical development. Meanwhile, the mtRTK inhibitor WM-S1 drives antitumor immunity (with anti-PD-L1) in NSCLC. Combinational approaches with immunotherapy showed that synergistic effect of WM-S1 and anti-PD1 monoclonal antibody, suppressing tumor growth by 75% in anti-PD1 resistance NSCLC-derived humanized mouse model. A phase 1b trial is expected to develop WM-S1 through not only indication expansion but also combination therapy with immuno-checkpoint inhibitors in the USA, AUS and KOR from Q2 2022. In conclusion, mtRTK is a potential oncogenic driver mutation in various solid tumor. A first-in-class anticancer agent WM-S1 targeting mtRTK can be promising therapeutic agents for cetuximab-resistant colon cancer patients regardless of KRAS mutation status and other cancers. Citation Format: Joseph Kim, Jai-Hee Moon, Kyung-Mi Lee, Hyun Ryu, Eun Hye Park, Sang Hee Kim, Jeong Seok Kim, Young Ok Ko, Yong Seok Kim, Hyo Jin Kim, Tae Young Kim Kim, Moon Seong Yoo, Soll Jin, Seongrak Kim, Yoon Sun Park, Min Ki Lee, Mi So Lee, Ji Hyun Go, Yu Geun Ji, Jun Hyung Lee, Haneul Lee, Min Hwa Kim, Eun Hee Ko, Yeo Jin Lee, Seung-Mi Kim, Joon-yee Jeong, Yeon-seoung Choi, Seung-geon Bae, Jinwoo Lee, Won Jun Lee, Min-Kyeong Kim, Ji min Shin, Dong-in Koh, Sun-Chul Hur, Chun-Ho Park, Hyun Ho Lee, Dong-Hoon Jin. WM-S1, the novel small molecule inhibitor of mutant RTK/receptor tyrosine kinase, for the treatment of cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6219.

Implementation of lateral flow devices (LFDs) for rabies antigen detection is expected to improve surveillance through the efficient detection of rabid animals in resource-limited settings; however, the use of LFDs for diagnosis remains controversial because some commercially available kits show low sensitivity. Therefore, we compared the diagnostic efficacy of three LFDs (ADTEC, Bionote, and Elabscience kits) paralleled with the direct fluorescent antibody test (dFAT) using fresh samples and investigated the diagnostic accuracies. To do so, we evaluated rabies-suspected samples submitted to the Regional Animal Disease Diagnostic Laboratory III, Philippines. Furthermore, we conducted real-time RT-PCR and sequencing to measure the accuracy of field laboratory diagnosis. The total number of animals submitted during this study period was 184 cases, including negative control samples. Of these, 53.9% (84 cases) were positive in the dFAT. Dogs were the most common rabies-suspected animal (n = 135). The sensitivities of the ADTEC and Bionote kits were 0.88 (74 cases) and 0.95 (80 cases), respectively. The specificity of both kits was 1.00 (100 cases). Furthermore, the sensitivity and specificity of the ADTEC kit after directly homogenizing the samples in assay buffer without dilution in phosphate-buffered saline (ADTEC kit DM) were 0.94 (79 cases) and 1.00 (100 cases), respectively. By contrast, there were no positive results using the Elabscience kit among all dFAT-positive samples. The sensitivity and specificity of LFDs make these tests highly feasible if properly used. Therefore, LFD tests can be used to strengthen the surveillance of rabies-infected animals in endemic and resource-limited settings., Author summary Implementation of lateral flow devices (LFDs) for rabies antigen detection is expected to improve surveillance through the efficient detection of rabid animals in resource-limited settings because they are not only accurate but rapid, user-friendly, and low-cost tools. Although LFDs may positively contribute to accurate data reporting, the use of LFDs for definitive diagnosis remains controversial because some LFDs showed inadequate ability. In this study, we evaluated the diagnostic accuracy of three LFDs (ADTEC, Bionote, and Elabscience kits) paralleled with the direct fluorescent antibody test (dFAT) and real-time RT-PCR using fresh samples and investigated discrepancies among these tests. Of these, half of the submitted samples were positive in the dFAT. The sensitivity and specificity of the simplified ADTEC method (ADTEC kit DM) and Bionote kits were comparable with those of dFAT, but no positive reactions were observed using the Elabscience kit. Despite controversial discussions regarding the use of LFDs, we demonstrated that the diagnostic accuracy of ADTEC and Bionote kits was satisfactory under regional diagnostic laboratory conditions. Our results support the potential of LFD tests for in-field diagnosis of rabies in endemic countries and the feasibility for practical use when they are properly used.

Francisella tularensis, a highly infectious bacterium, is an etiological agent of the zoonotic disease tularemia. It is widely distributed in the Northern Hemisphere, including Japan. Here, we have determined the complete genome sequences of two strains of F. tularensis subsp. holarctica bv. japonica isolated from hares in 2008 and 2009., Francisella tularensis, a highly infectious bacterium, is the etiological agent of the zoonotic disease tularemia. It is widely distributed in the Northern Hemisphere, including Japan. Here, we have determined the complete genome sequences of two strains of F. tularensis subsp. holarctica bv. japonica isolated from hares in 2008 and 2009.

The aim of this study was to compare the histochemical and meat quality characteristics between the normal and white-striping (WS) pectoralis major muscles. Additionally, this study investigated the effects of oven cooking (OV) and sous-vide (SV) cooking methods on objective texture parameters and sensory quality characteristics of the normal and WS chicken breast meats. Results showed that the WS condition broilers had higher body and breast weights (P < 0.001), and a greater area of muscle fiber than the normal broilers (P < 0.001). The WS fresh fillets exhibited a lower preference of visual appearance compared to the normal fillets (P < 0.05), although no differences were detected in the characteristics of meat quality between the groups (P > 0.05). After cooking, there was greater cooking loss, Warner-Bratzler shear force, and texture parameter analysis-hardness values for breast fillets cooked by OV treatment at 180°C for reached core temperature to 71°C compared to the fillets cooked by SV treatment at 60°C for 2 h (P < 0.05), whereas the normal and WS groups were exhibited similar values within each cooking methods (P > 0.05). Regarding sensory quality characteristics, WS breast fillets cooked by SV (SV+WS) were rated as tenderer and juicier, and given a higher overall acceptability score compared to normal and WS fillets cooked by OV (P < 0.05). However, owing to a lesser developed flavor in SV+WS fillets, the panelists assigned a lower overall acceptability rating in these fillets compared to SV+Normal fillets (P < 0.05). Overall, the SV cooking can be an effective method for improving the sensory quality characteristics of WS and normal chicken breast.

A 7-year-old male cavalier King Charles spaniel was presented with a cervical subcutaneous mass. The mass had a multilobular growth pattern and each individual lobule contained ductal epithelial cells and myoepithelial cells surrounding the ductal cells. Immunohistochemically, the ductal epithelial cells were positive for cytokeratin (CK) 7 and CAM5.2 and the myoepithelial cells were positive for α-smooth muscle actin and p63. Both types of cells were positive for CK14, a myoepithelial cell marker. Ultrastructurally, the ductal epithelial cells were attached by desmosomes and contained abundant intracytoplasmic tonofilaments. Some ductal epithelial cells contained myofilaments characteristic of myoepithelial cells. Based on these findings, a diagnosis of epithelial-myoepithelial carcinoma was made. It is presumed that the tumour originated from an intercalated duct in a parotid salivary gland.

We established a system for the recovery of a segmented recombinant rabies virus, the virus genome RNA of which was divided into two parts: segment 1 encoding the nucleoprotein, phosphoprotein, matrix protein, and glycoprotein genes, and segment 2 encoding the large RNA-dependent RNA polymerase gene. The morphology of the segmented recombinant rabies virus was bullet-like in shape with a length of approximately 130 nm, which is shorter than the 200-nm long non-segmented recombinant rabies virus. The segmented recombinant rabies virus was maintained for at least 18 passages. The virus multiplication rate of the segmented recombinant rabies virus was lower than that of the non-segmented recombinant rabies virus during the passages, and the relative amounts of virus genome RNAs for segment 1 and segment 2 differed in the supernatant of the segmented recombinant rabies virus infected cells. These results suggest that the segmented recombinant rabies virus packages either segment 1 or segment 2 into each virus particle. Thus, co-infection with segmented recombinant rabies virus particles packaging segment 1 or segment 2 may be necessary for the production of progeny virus.

During rabies virus infections, the minor salivary glands are one of the important organs for virus replication and excretion into the oral cavity. However, details of pathological findings and viral antigen distribution in the minor salivary glands remain poorly understood. In this study, we conducted pathological tests on the tongues of 71 rabid dogs in the Philippines; the minor salivary glands (von Ebner's glands, lingual glands), circumvallate papilla, autonomic ganglia, and skeletal muscles were evaluated. Inflammatory changes were observed in the von Ebner's glands of 20/71 dogs, in the circumvallate papilla of 10/71, and in the tongue muscle of 1/71. Conversely, no morphological changes were observed in the lingual glands and autonomic ganglia. Viral antigens were detected via immunohistochemistry-based methods in the cytoplasm of the acinar epithelium in the von Ebner's glands of all 71 dogs. Virus particles were confirmed in the intercellular canaliculi and acinar lumen via electron microscopy. In the autonomic ganglia, viral antigens were detected in 67/71 rabid dogs. Viral antigens were detected in the taste buds of all 71 dogs, and were distributed mainly in type II and III taste bud cells. In tongue muscle fibers, viral antigens were detected in 11/71 dogs. No virus antigens were detected in lingual glands. These findings suggest that rabies virus descends in the tongue along the glossopharyngeal nerve after proliferation in the brain, and von Ebner's glands and taste buds are one of the portals of virus excretion into the saliva in rabid dogs.

Recently, we reported that follicle-sinus complexes (FSCs) in the muzzle skin are useful for postmortem diagnosis of rabid dogs. Here, we compared the sensitivity and specificity of detecting the viral antigen in the brain and FSCs of 226 suspected rabid dogs, and assessed whether the FSC harbored the virus genome and particles. The viral antigen was detected in 211 of 226 samples with 100% sensitivity and specificity. Viral RNA and particles were observed in the cytoplasm of Merkel cells (MCs). These results suggest that MCs are targets of virus infection and FSCs are useful material for diagnosing rabies.

The number of lung cancer patients is increasing every year worldwide. For these patients' treatment, not only anticancer drugs but also immune anticancer drugs are rapidly being developed. So far, the PD-1 antibody is widely used in treating NSCLC patients, and various pharmaceutical companies are continuously developing it. However, in PD-L1 low or negative patients, new drugs are required because there are no appropriate drugs. Of course, the PD-1 antibody is sometimes used, but the response rate is low, so new therapeutic drugs are needed.We identified a new target for PD-L1 low or negative patients using PD-L1 negative or positive lung cancer tissues. The new target's code name is "Target protein A." Target protein A expressed higher in tumor than normal. As a result of analyzing caucasian lung cancer patients' tissues by immunohistochemistry, 36% of cases expressed "Target protein A". Moreover, we also confirmed that target protein A was higher in tumors than normal through the TCGA database. Based on these results, we developed an immune anticancer drug (WM-A1) targeting Target protein A and demonstrated that it induces cancer cell death by inhibiting the binding with the binding partner protein B. Meanwhile, to clarify the relationship between PD-L1 and Target protein A, we analyzed the expression between the two proteins in lung cancer cell lines and proved that they exhibit reverse correlation. Among PD-L1 low or negative lung cancer patient tissues, about 32.75% of tissues expressed Target protein A. Conversely, among the patient samples expressing Target protein A, the PD-L1 low or negative population was 63%. We checked a similar trend through the TCGA database. For the efficacy of WM-A1, we confirmed that WM-A1 induced T cell-mediated cell death in a co-culture system with human PBMC and secreted major cytokines. In addition, we revealed that the group transplanted with a cell line expressing Target protein A in the PBMC humanized model showed high immuno-anticancer efficacy. Furthermore, we demonstrated that WM-A1 increased T cell activation and related cytokine expression. However, there was no reactivity to the PD-1 antibody because it expressed Target protein A and did not express PD-L1.Therefore, we suggest that WM-A1 is a new therapeutic antibody for the PD-L1 low or negative patient population. Based on these findings, we are planning a preclinical study (GLP-Tox) in 2021. #Corresponding author: Dong-Hoon Jin*These authors (Hye jin Son and Minki Lee) contributed equally to this work. Citation Format: Hye jin Son, Minki Lee, Seongrak Kim, Jai-Hee Moon, Hana Kim, Wonhwa Shin, Joseph Kim, Mi So Lee, Daeun Kim, Seunggeon Bae, Joonyee Jeong, Seung-Woo Hong, Chun-Ho Park, Hyun Ho Lee, Dong-Hoon Jin. New therapeutic antibody ("WM-A1") for treatment of low or no PD-L1 NSCLC patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1216.

Most studies on rabies virus pathogenesis in animal models have employed fixed rabies viruses, and the results of those employing street rabies viruses have been inconsistent. Therefore, to clarify the pathogenesis of street rabies virus (1088 strain) in mice, 106 focus forming units were inoculated into the right hindlimb of ddY mice (6 weeks, female). At 3 days postinoculation (DPI), mild inflammation was observed in the hindlimb muscle. At 5 DPI, ganglion cells in the right lumbosacral spinal dorsal root ganglia showed chromatolysis. Axonal degeneration and inflammatory cells increased with infection progress in the spinal dorsal horn and dorsal root ganglia. Right hindlimb paralysis was observed from 7 DPI, which progressed to quadriparalysis. However, no pathological changes were observed in the ventral horn and root fibers of the spinal cord. Viral antigen was first detected in the right hindlimb muscle at 3 DPI, followed by the right lumbosacral dorsal root ganglia, dorsal horn of spinal cord, left red nuclei, medulla oblongata and cerebral cortex (M1 area) at 5 DPI. These results suggested that the 1088 virus ascended the lumbosacral spinal cord via mainly afferent fibers at early stage of infection and moved to cerebral cortex (M1 area) using descending spinal tract. Additionally, we concluded that significant pathological changes in mice infected with 1088 strain occur in the sensory tract of the spinal cord; this selective susceptibility results in clinical features of the disease.

Background Infrared radiation (IR) exposure generates reactive oxygen species and induces matrix metalloproteinase-1 expression in human skin. Moreover, while not as acute as ultraviolet radiation, repeated infrared irradiation can result in the photoaging of skin. Broad-spectrum sunscreens can protect skin from IR, but no human in vivo test methods for the evaluation of sunscreens' IR protection effect have been developed. We aimed to develop such a method. Materials and methods We included 155 Korean subjects in our three-part clinical study. The IR reflectance of subjects' skin was measured using a benchtop model of an IR light source and a reflectance measuring probe. We measured the IR reflectance in relation to skin color and hydration level to set up our experimental conditions. We then calculated the infrared protection factors (IPFs) of cosmetic emulsions as the IR reflectance ratio between cosmetic sunscreen-applied skin and non-sunscreen-applied skin and assessed the relationship between IPFs and the amount of sunscreen ingredients. Finally, this method was validated using several commercial sunscreen cosmetics. Results Skin color and hydration level did not influence the IR reflectance of subjects' skin. The IPFs of cosmetic sunscreens showed a positive correlation with the amount of inorganic sunscreen ingredients. Conclusion In this study, we developed a simple, fast, and ethically acceptable human in vivo test method for evaluating the IPFs of cosmetic sunscreens.

Staphylococcus aureus is an important bacterial pathogen causing bovine mastitis, but little is known about the virulence factor and the inflammatory responses in the mammary infection. Staphylococcal enterotoxin C (SEC) is the most frequent toxin produced by S. aureus, isolated from bovine mastitis. To investigate the pathogenic activity of SEC in the inflammation of the mammary gland and the immune responses in an animal model, mouse mammary glands were injected with SEC, and the clinical signs, inflammatory cell infiltration, and proinflammatory cytokine production in the mammary glands were assessed. SEC induced significant inflammatory reactions in the mammary gland, in a dose-dependent manner. SEC-injected mammary glands showed a severe inflammation with inflammatory cell infiltration and tissue damage. In addition, interleukin (IL)-1&beta, and IL-6 production in the SEC-injected mammary glands were significantly higher than those in the PBS control glands. Furthermore, the SEC-induced inflammation and tissue damage in the mammary gland were specifically inhibited by anti-SEC antibody. These results indicated, for the first time, that SEC can directly cause inflammation, proinflammatory cytokine production, and tissue damage in mammary glands, suggesting that SEC might play an important role in the development of mastitis associated with S. aureus infection. This finding offers an opportunity to develop novel treatment strategies for reduction of mammary tissue damage in mastitis.

H9N2 avian influenza virus causes sporadic human infection. Since humans do not possess acquired immunity specific to this virus, we examined the pathogenicity of an H9N2 virus isolated from a human and then analyzed protective effects of a vaccine in cynomolgus macaques. After intranasal challenge with A/Hong Kong/1073/1999 (H9N2) (HK1073) isolated from a human patient, viruses were isolated from nasal and tracheal swabs in unvaccinated macaques with mild fever and body weight loss. A formalin-inactivated H9N2 whole particle vaccine derived from our virus library was subcutaneously inoculated to macaques. Vaccination induced viral antigen-specific IgG and neutralization activity in sera. After intranasal challenge with H9N2, the virus was detected only the day after inoculation in the vaccinated macaques. Without vaccination, many bronchus-associated lymphoid tissues (BALTs) were formed in the lungs after infection, whereas the numbers of BALTs were smaller and the cytokine responses were weaker in the vaccinated macaques than those in the unvaccinated macaques. These findings indicate that the H9N2 avian influenza virus HK1073 is pathogenic in primates but seems to cause milder symptoms than does H7N9 influenza virus as found in our previous studies and that a formalin-inactivated H9N2 whole particle vaccine induces protective immunity against H9N2 virus.

The direct fluorescent antibody test (dFAT) on fresh brain tissues is the gold standard for rabies virus antigen detection in dogs. However, this method is laborious and holds a high risk of virus exposure for the experimenter. Skin biopsies are useful for the diagnosis of humans and animals. In mammals, the tactile hair, known as the follicle-sinus complex (FSC), is a specialized touch organ that is abundant in the muzzle skin. Each tactile hair is equipped with more than 2,000 sensory nerve endings. Therefore, this organ is expected to serve as an alternative postmortem diagnostic material. However, the target cells and localization of rabies virus antigen in the FSCs remain to be defined. In the present study, muzzle skins were obtained from 60 rabid dogs diagnosed with rabies by dFAT at the Research Institute of Tropical Medicine in the Philippines. In all dogs, virus antigen was clearly detected in a part of the outer root sheath at the level of the ring sinus of the FSCs, and the majority of cells were positive for the Merkel cell (MC) markers cytokeratin 20 and CAM5.2. Our results suggest that MCs in the FSCs of the muzzle skin are a target for virus replication and could serve as a useful alternative specimen source for diagnosis of rabies.

Rabies is a zoonotic disease caused by the rabies virus. While the salivary glands are important as exit and propagation sites for the rabies virus, the mechanisms of rabies excretion remain unclear. Here, we investigated the histopathology of the salivary glands of rabid dogs and analyzed the mechanism of excretion into the oral cavity. Mandibular and parotid glands of 22 rabid dogs and three control dogs were used. Mild to moderate non-suppurative sialadenitis was observed in the mandibular glands of 19 of the 22 dogs, characterized by loss of acinar epithelium and infiltration by lymphoplasmacytic cells. Viral antigens were detected in the mucous acinar epithelium, ganglion neurons and myoepithelium. Acinar epithelium and lymphocytes were positive for anti-caspase-3 antibodies and TUNEL staining. In contrast, no notable findings were observed in the ductal epithelial cells and serous demilune. In the parotid gland, the acinar cells, myoepithelium and ductal epithelium all tested negative. These findings confirmed the path through which the rabies virus descends along the facial nerve after proliferation in the brain to reach the ganglion neurons of the mandibular gland, subsequently traveling to the acinar epithelium via the salivary gland myoepithelium. Furthermore, the observation that nerve endings passing through the myoepithelium were absent from the ductal system suggested that viral proliferation and cytotoxicity could not occur there, ensuring that secretions containing the virus are efficiently excreted into the oral cavity.

A 15-year-old, female mixed-breed dog presented with abdominal distention. An exploratory laparotomy revealed a large left ovarian mass (20 × 15 × 12 cm). Histopathological examination of the mass revealed a mixed myxoid liposarcoma and a well-differentiated leiomyoma. Four months after surgical removal of the mass, the dog died due to multiorgan metastasis. The metastasis was composed solely of the liposarcoma component. The liposarcoma component was Alcian Blue- and Oil red O-positive, and demonstrated immunoreactivity with S-100, adipophilin and vimentin. Electron microscopy revealed that the tumor cell cytoplasms were packed with lipid vacuoles and dilated rough endoplasmic reticulum. To our knowledge, this is the first report of myxoid liposarcoma and leiomyoma co-existing in a canine ovary.

A two-year-old castrated male Pomeranian dog was referred with the chief complaints of coughing and subcutaneous emphysema. On physical examination, the crepitant areas were palpable. When auscultated, the right chest was absent of respiratory sound, while the sound of the opposite side was enhanced. Radiographs presented pneumothorax and pneumomediastinum. On computed tomography, hypoattenuated bulla-like lesion at right middle lung lobe and trapped air in mediastinum were shown. After patient stabilization, surgery for excision of affected lobe was performed. During follow-up period, there were no recurrence and complication on radiographic examination. Based on clinical and pathological findings, the dog was diagnosed as congenital lobar emphysema.

Rabies is a fatal zoonotic disease endemic in developing countries of Asia and Africa. Recently, the direct rapid immunohistochemical test (DRIT) was recommended by the World Health Organization (WHO) and the World Organization for Animal Health (OIE) as a diagnostic test for rabies. Therefore, a biotinylated polyclonal antibody (pAb) against the rabies lyssavirus (RABV) nucleoprotein was developed using a plasmid cDNA vaccine derived from a challenge virus standard 11 strain. A preliminary evaluation on the efficacy of this reagent in recognizing the Philippine RABV strain was tested using banked canine hippocampal tissue samples with DRIT and the results were compared to dFAT. The effects of acetone and formalin fixation on DRIT were also assessed through immunoreactivity scores of the specimens. Of the 142 samples examined, 104 tested positive and 38 negative using both dFAT and DRIT, showing 100% agreement between the two diagnostic procedures. Moreover, no false positive or false negative results were observed using acetone and formalin fixation. Thus, locally prepared biotinylated pAb from plasmid cDNA can be used for DRIT, especially in resource-limited laboratories in the Philippines. However, these results should be confirmed with a more thorough evaluation of this technique, and the range of detection needs to be further evaluated in a larger panel of animal samples and on other lyssaviruses.

The new point of view in vehicle of Roof Rack is proposed by Nylon-Continuous Fiber Composites Structure, which is a new development method based on the principle of environment resistance material, plastic injection, new fabric compression, engineering design, test and evaluations. As a result of the application of new roof rack against aluminium roof rack on roof, this paper classifies Increased of environment resistance materials in compressed continuous Fiber fabric and plastic injection, body mountings, Shapes of cross section and internal rib, customers clamping of roof rack, positions of cross bar introduces Reduction of weight, cost and Whistle Noise, improvement of liberty of colors.

In this study, G proteins of the rabies virus (RABV) Kyoto strain were detected in the cytoplasm but not distributed at the cell membrane of mouse neuroblastoma (MNA) cells. G proteins of CVS-26 were detected in both the cell membrane and perinuclear space of MNA cells. We found that N-glycosylation of street RABV G protein by the insertion of the sequon Asn(204) induced the transfer of RABV G proteins to the cell surface membrane. Fixed RABV budding from the plasma membrane has been found to depend not only on G protein but also on other structural proteins such as M protein. However, the differing N-glycosylation of G protein could be associated with the distinct budding and antigenic features of RABV in street and fixed viruses. Our study of the association of N-glycan of G protein at Asn(204) with the transport of RABV G protein to the cell surface membrane contributes to the understanding of the evolution of fixed virus from street virus, which in turn would help for determine the mechanism underlying RABV budding and enhanced host immune responses.

Most studies on rabies virus pathogenesis in animal models have employed fixed rabies viruses, and the results of those employing street rabies viruses have been inconsistent. Therefore, to clarify the pathogenesis of street rabies virus (1088 strain) in mice, 106 focus forming units were inoculated into the right hindlimb of ddY mice (6 weeks, female). At 3 days postinoculation (DPI), mild inflammation was observed in the hindlimb muscle. At 5 DPI, ganglion cells in the right lumbosacral spinal dorsal root ganglia showed chromatolysis. Axonal degeneration and inflammatory cells increased with infection progress in the spinal dorsal horn and dorsal root ganglia. Right hindlimb paralysis was observed from 7 DPI, which progressed to quadriparalysis. However, no pathological changes were observed in the ventral horn and root fibers of the spinal cord. Viral antigen was first detected in the right hindlimb muscle at 3 DPI, followed by the right lumbosacral dorsal root ganglia, dorsal horn of spinal cord, left red nuclei, medulla oblongata and cerebral cortex (M1 area) at 5 DPI. These results suggested that the 1088 virus ascended the lumbosacral spinal cord via mainly afferent fibers at early stage of infection and moved to cerebral cortex (M1 area) using descending spinal tract. Additionally, we concluded that significant pathological changes in mice infected with 1088 strain occur in the sensory tract of the spinal cord; this selective susceptibility results in clinical features of the disease.

Tularemia, a highly infectious zoonotic disease caused by Francisella tularensis, occurs sporadically in Japan. However, little is known about the prevalence of the disease in wild animals. A total of 632 samples obtained from 150 Japanese black bears, 142 Japanese hares, 120 small rodents, 97 rats, 53 raptors, 26 Japanese monkeys, 21 Japanese raccoon dogs, 20 masked palm civets, and three Japanese red foxes between 2002 and 2010 were investigated for the presence of antibodies to F. tularensis by competitive enzyme-linked immunosorbent assay (cELISA) and the commonly used microagglutination (MA) test. Seropositive cELISA and MA results were obtained in 23 and 18 Japanese black bears, three and two Japanese raccoon dogs, and two and one small rodents, respectively. All MA-positive samples (n=21) were also positive by cELISA. Six of seven samples that were only positive by cELISA were confirmed to be antibody-positive by western blot analysis. These findings suggest that cELISA is a highly sensitive and useful test for serosurveillance of tularemia among various species of wild animals. Because this is the first study to detect F. tularensis-seropositive Japanese raccoon dogs, these could join Japanese black bears as sentinel animals for tularemia in the wild in Japan. Further continuous serosurveillance for F. tularensis in various species of wild animals using appropriate methods such as cELISA is important to assess the risks of human exposure and to improve our understanding of the ecology of F. tularensis in the wild.

This study introduces a simple method to functionalize porous polymer microspheres with in situ synthesis of metal nanoparticles via mussel-inspired polydopamine chemistry. Highly open porous poly(methyl methacrylate) (PMMA) microspheres are prepared by a single oil-in-water emulsion solvent evaporation technique using Pluronic F127 as an extractable porogen. The internal pore surfaces of the prepared microspheres are coated with polydopamine via the oxidative polymerization of dopamines in an aqueous solution. The deposited polydopamine mediates the spontaneous reduction of silver nitrates into solid silver nanoparticles (Ag NPs) within the pores of the prepared microspheres, resulting in porous PMMA microspheres decorated with Ag NPs. Anti-bacterial experiments show that the Ag NP-decorated PMMA microspheres can be used as an excellent anti-bacterial platform. This study suggests that highly open porous microspheres can be used as a template to synthesize functional metal–polymer hybrid materials using the mussel-inspired polydopamine chemistry in an aqueous solution under ambient conditions.

Feline pituitary tumors are rare. An 8-year-old male Japanese domestic cat presented with anorexia and emaciation. The cat died 17 days after admission from progressive neurological symptoms. At necropsy, a pituitary tumor measuring 25 × 18 × 15 mm was found. Microscopically, the tumor was divided into multiple lobules and had grown invasively into the adjacent brain tissue and sphenoid bone. Tumor cells had pleomorphic nuclei with prominent centrally located nucleoli and abundant amphophilic polygonal cytoplasm. Immunohistochemically, the tumor cells stained with anti-adrenocorticotropic hormone (ACTH), α-melanin-stimulating hormone (MSH) and β-endorphin antibodies. Ultrastructurally, the cytoplasm of the tumor cells contained various sized secretory granules. Based on these pathological findings, this tumor was diagnosed as pituitary carcinoma originated from pars intermedia cells.

A malignant epithelioid schwannoma of the oral cavity was diagnosed in an 8-year-old domestic short-hair cat. The mass was located in the gingiva of the upper left premolar to molar region and showed multinodular growth patterns. The mass comprised epithelioid cells arranged in densely packed sheets. Tumor cells had large, round to oval nuclei with prominent nucleoli and an abundant eosinophilic cytoplasm. Immunohistochemically, most of the tumor cells were positive for S-100 protein, glial fibrillary acidic protein and vimentin, but all lacked melanoma-associated antigen and muscle and neuroendocrine markers. Stains for type IV collagen showed linear immunoreactivity around single cells and groups of cells. Ultrastructurally, tumor cells were separated by a well-defined basement membrane, and interdigitating cell processes were observed. To our knowledge, this is the first report of feline malignant epithelioid schwannoma.