Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Background: and Purpose: Recognizing cognitive decline patterns in mild cognitive impairment (MCI) is crucial for early screening and preventive interventions. However, studies on the trajectory of individual cognitive functions in MCI are limited. Thus, the purpose of this study was to identify subtypes and stages of cognitive decline in MCI using a machine learning method. Methods: A total of 944 subjects consisting of those who were cognitively normal and those with MCI were enrolled. Fifteen neuropsychological tasks were used in the analysis.The optimal number of subtypes was determined based on the cross-validation information criterion. Fifteen stages of cognitive trajectory were estimated for each subtype. Results: The following three subtypes were identified: amnestic-verbal subtype, dysexecutive subtype, and amnestic-visual subtype. Of 723 (76.6%) subjects who had reached stage 1 at least, amnestic-verbal subtype accounted for the most (n=340, 47.0%), followed by dysexecutive subtype (n=253, 35.0%) and amnestic-visual subtype (n=130, 18%). The amnestic-verbal subtype had significantly more males (amnestic-verbal: 41.8%, dysexecutive: 31.2%, and amnestic-visual: 28.5%), younger subjects (amnestic-verbal: 72.01 years, dysexecutive: 74.43 years, and amnestic-visual: 75.06 years), higher educational years (amnestic-verbal: 11.06 years, dysexecutive: 9.53 years, and amnestic-visual: 9.79 years), lower Clinical Dementia Rating sum of boxes (amnestic-verbal: 1.40, dysexecutive: 1.61, and amnestic-visual: 1.71), and lower Korean-Instrumental Activities of Daily Living score (amnestic-verbal: 0.20, dysexecutive: 0.27, and amnestic-visual: 0.26). Conclusions Three types of MCIs were extracted using SuStaIn. Pathways of MCI deterioration could be different. The amnestic type could be bisected based on whether episodic verbal or visual memory is degraded first.

Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Background: and Purpose: Recognizing cognitive decline patterns in mild cognitive impairment (MCI) is crucial for early screening and preventive interventions. However, studies on the trajectory of individual cognitive functions in MCI are limited. Thus, the purpose of this study was to identify subtypes and stages of cognitive decline in MCI using a machine learning method. Methods: A total of 944 subjects consisting of those who were cognitively normal and those with MCI were enrolled. Fifteen neuropsychological tasks were used in the analysis.The optimal number of subtypes was determined based on the cross-validation information criterion. Fifteen stages of cognitive trajectory were estimated for each subtype. Results: The following three subtypes were identified: amnestic-verbal subtype, dysexecutive subtype, and amnestic-visual subtype. Of 723 (76.6%) subjects who had reached stage 1 at least, amnestic-verbal subtype accounted for the most (n=340, 47.0%), followed by dysexecutive subtype (n=253, 35.0%) and amnestic-visual subtype (n=130, 18%). The amnestic-verbal subtype had significantly more males (amnestic-verbal: 41.8%, dysexecutive: 31.2%, and amnestic-visual: 28.5%), younger subjects (amnestic-verbal: 72.01 years, dysexecutive: 74.43 years, and amnestic-visual: 75.06 years), higher educational years (amnestic-verbal: 11.06 years, dysexecutive: 9.53 years, and amnestic-visual: 9.79 years), lower Clinical Dementia Rating sum of boxes (amnestic-verbal: 1.40, dysexecutive: 1.61, and amnestic-visual: 1.71), and lower Korean-Instrumental Activities of Daily Living score (amnestic-verbal: 0.20, dysexecutive: 0.27, and amnestic-visual: 0.26). Conclusions Three types of MCIs were extracted using SuStaIn. Pathways of MCI deterioration could be different. The amnestic type could be bisected based on whether episodic verbal or visual memory is degraded first.

Background: and Purpose: Recognizing cognitive decline patterns in mild cognitive impairment (MCI) is crucial for early screening and preventive interventions. However, studies on the trajectory of individual cognitive functions in MCI are limited. Thus, the purpose of this study was to identify subtypes and stages of cognitive decline in MCI using a machine learning method. Methods: A total of 944 subjects consisting of those who were cognitively normal and those with MCI were enrolled. Fifteen neuropsychological tasks were used in the analysis.The optimal number of subtypes was determined based on the cross-validation information criterion. Fifteen stages of cognitive trajectory were estimated for each subtype. Results: The following three subtypes were identified: amnestic-verbal subtype, dysexecutive subtype, and amnestic-visual subtype. Of 723 (76.6%) subjects who had reached stage 1 at least, amnestic-verbal subtype accounted for the most (n=340, 47.0%), followed by dysexecutive subtype (n=253, 35.0%) and amnestic-visual subtype (n=130, 18%). The amnestic-verbal subtype had significantly more males (amnestic-verbal: 41.8%, dysexecutive: 31.2%, and amnestic-visual: 28.5%), younger subjects (amnestic-verbal: 72.01 years, dysexecutive: 74.43 years, and amnestic-visual: 75.06 years), higher educational years (amnestic-verbal: 11.06 years, dysexecutive: 9.53 years, and amnestic-visual: 9.79 years), lower Clinical Dementia Rating sum of boxes (amnestic-verbal: 1.40, dysexecutive: 1.61, and amnestic-visual: 1.71), and lower Korean-Instrumental Activities of Daily Living score (amnestic-verbal: 0.20, dysexecutive: 0.27, and amnestic-visual: 0.26). Conclusions Three types of MCIs were extracted using SuStaIn. Pathways of MCI deterioration could be different. The amnestic type could be bisected based on whether episodic verbal or visual memory is degraded first.

Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Background: and Purpose: Recognizing cognitive decline patterns in mild cognitive impairment (MCI) is crucial for early screening and preventive interventions. However, studies on the trajectory of individual cognitive functions in MCI are limited. Thus, the purpose of this study was to identify subtypes and stages of cognitive decline in MCI using a machine learning method. Methods: A total of 944 subjects consisting of those who were cognitively normal and those with MCI were enrolled. Fifteen neuropsychological tasks were used in the analysis.The optimal number of subtypes was determined based on the cross-validation information criterion. Fifteen stages of cognitive trajectory were estimated for each subtype. Results: The following three subtypes were identified: amnestic-verbal subtype, dysexecutive subtype, and amnestic-visual subtype. Of 723 (76.6%) subjects who had reached stage 1 at least, amnestic-verbal subtype accounted for the most (n=340, 47.0%), followed by dysexecutive subtype (n=253, 35.0%) and amnestic-visual subtype (n=130, 18%). The amnestic-verbal subtype had significantly more males (amnestic-verbal: 41.8%, dysexecutive: 31.2%, and amnestic-visual: 28.5%), younger subjects (amnestic-verbal: 72.01 years, dysexecutive: 74.43 years, and amnestic-visual: 75.06 years), higher educational years (amnestic-verbal: 11.06 years, dysexecutive: 9.53 years, and amnestic-visual: 9.79 years), lower Clinical Dementia Rating sum of boxes (amnestic-verbal: 1.40, dysexecutive: 1.61, and amnestic-visual: 1.71), and lower Korean-Instrumental Activities of Daily Living score (amnestic-verbal: 0.20, dysexecutive: 0.27, and amnestic-visual: 0.26). Conclusions Three types of MCIs were extracted using SuStaIn. Pathways of MCI deterioration could be different. The amnestic type could be bisected based on whether episodic verbal or visual memory is degraded first.

Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Patients with cancer experience psychological and social problems; in particular, older patients with cancer face many difficulties during the cancer treatment process owing to aging and underlying diseases. Furthermore, the lives of individuals may be impacted by the COVID-19 pandemic. Therefore, the purpose of this study is to describe the experiences of older patients with cancer during the cancer diagnosis and treatment process amid the COVID-19 pandemic. Methods: This study employed a qualitative, descriptive phenomenological approach to explore and analyze the experiences of the participants. The participants in this study consisted of patients aged over 65 who were diagnosed with cancer, and data were collected from May 4 to June 30, 2022 through in-depth individual interviews. The collected data were analyzed using Giorgi’s phenomenological analysis. Results: Participants were six older patients with cancer with an average age of 69.66 years. Five theme clusters and 15 themes were generated. The theme clusters were “psychological adaptation to cancer diagnosis,” “receiving social and medical support,” “difficulty in the treatment process,” “fear of the COVID-19 pandemic,” and “living through difficulties.” Conclusion: This study revealed that older patients with cancer demonstrated a unique psychology about how to accept the shock of the diagnosis and the resulting coping patterns and feelings of depression. Additionally, it was verified that older patients with cancer, susceptible to infections due to their advanced age and weakened immunity, also face an increased vulnerability to COVID-19. Therefore, a support system tailored to the characteristics of older patients with cancer should be established.