Neuropilin-1 is a novel host factor modulating the entry of hepatitis B virus.

BACKGROUND & AIMS: Sodium taurocholate cotransporting polypeptide (NTCP) has been identified as the cellular receptor for hepatitis B virus (HBV). However, hepatocytes expressing NTCP exhibit varying susceptibilities to HBV infection. This study aimed to investigate whether other host factors modulate the process of HBV infection. METHODS: Liver biopsy samples obtained from children with hepatitis B were used for single-cell sequencing and susceptibility analysis. Primary human hepatocytes, HepG2-NTCP cells, and human liver chimeric mice were used to analyze the effect of candidate host factors on HBV infection. RESULTS: Single-cell sequencing and susceptibility analysis revealed a positive correlation between neuropilin-1 (NRP1) expression and HBV infection. In the HBV-infected cell model, NRP1 overexpression before HBV inoculation significantly enhanced viral attachment and internalization, and promoted viral infection in the presence of NTCP. Mechanistic studies indicated that NRP1 formed a complex with LHBs and NTCP. The NRP1 b domain mediated its interaction with conserved arginine residues at positions 88 and 92 in the preS1 domain of the HBV envelope protein LHBs. This NRP1-preS1 interaction subsequently promoted the binding of preS1 to NTCP, facilitating viral infection. Moreover, disruption of the NRP1-preS1 interaction by the NRP1 antagonist EG00229 significantly attenuated the binding affinity between NTCP and preS1, thereby inhibiting HBV infection both in vitro and in vivo. CONCLUSIONS: Our findings indicate that NRP1 is a novel host factor for HBV infection, which interacts with preS1 and NTCP to modulate HBV entry into hepatocytes. IMPACT AND IMPLICATIONS: HBV infection is a global public health problem, but the understanding of the early infection process of HBV remains limited. Through single-cell sequencing, we identified a novel host factor, NRP1, which modulates HBV entry by interacting with HBV preS1 and NTCP. Moreover, antagonists targeting NRP1 can inhibit HBV infection both in vitro and in vivo. This study could further advance our comprehension of the early infection process of HBV.

A study on the mechanism of Beclin-1 m6A modification mediated by catalpol in protection against neuronal injury and autophagy following cerebral ischemia.

OBJECTIVE: Catalpol (CAT) has various pharmacological activities and plays a protective role in cerebral ischemia. It has been reported that CAT played a protective role in cerebral ischemia by upregulaing NRF1 expression. Bioinformatics analysis reveals that NRF1 can be used as a transcription factor to bind to the histone acetyltransferase KAT2A. However, the role of KAT2A in cerebral ischemia remains to be studied. Therefore, we aimed to investigate the role of CAT in cerebral ischemia and its related mechanism. METHODS: In vitro, a cell model of oxygen and glucose deprivation/reperfusion (OGD/R) was constructed, followed by evaluation of neuronal injury and the expression of METTL3, Beclin-1, NRF1, and KAT2A. In vivo, a MCAO rat model was prepared by means of focal cerebral ischemia, followed by assessment of neurological deficit and brain injury in MCAO rats. Neuronal autophagy was evaluated by observation of autophagosomes in neurons or brain tissues by TEM and detection of the expression of LC3 and p62. RESULTS: In vivo, CAT reduced the neurological function deficit and infarct volume, inhibited neuronal apoptosis in the cerebral cortex, and significantly improved neuronal injury and excessive autophagy in MCAO rats. In vitro, CAT restored OGD/R-inhibited cell viability, inhibited cell apoptosis, LDH release, and neuronal autophagy. Mechanistically, CAT upregulated NRF1, NRF1 activated METTL3 via KAT2A transcription, and METTL3 inhibited Beclin-1 via m6A modification. CONCLUSION: CAT activated the NRF1/KAT2A/METTL3 axis and downregulated Beclin-1 expression, thus relieving neuronal injury and excessive autophagy after cerebral ischemia.

ZNF148 inhibits HBV replication by downregulating RXRα transcription.

BACKGROUND: Progressive hepatitis B virus (HBV) infection can result in cirrhosis, hepatocellular cancer, and chronic hepatitis. While antiviral drugs that are now on the market are efficient in controlling HBV infection, finding a functional cure is still quite difficult. Identifying host factors involved in regulating the HBV life cycle will contribute to the development of new antiviral strategies. Zinc finger proteins have a significant function in HBV replication, according to earlier studies. Zinc finger protein 148 (ZNF148), a zinc finger transcription factor, regulates the expression of various genes by specifically binding to GC-rich sequences within promoter regions. The function of ZNF148 in HBV replication was investigated in this study. METHODS: HepG2-Na+/taurocholate cotransporting polypeptide (HepG2-NTCP) cells and Huh7 cells were used to evaluate the function of ZNF148 in vitro. Northern blotting and real-time PCR were used to quantify the amount of viral RNA. Southern blotting and real-time PCR were used to quantify the amount of viral DNA. Viral protein levels were elevated, according to the Western blot results. Dual-luciferase reporter assays were used to examine the transcriptional activity of viral promoters. ZNF148's impact on HBV in vivo was investigated using an established rcccDNA mouse model. RESULTS: ZNF148 overexpression significantly decreased the levels of HBV RNAs and HBV core DNA in HBV-infected HepG2-NTCP cells and Huh7 cells expressing prcccDNA. Silencing ZNF148 exhibited the opposite effects in both cell lines. Furthermore, ZNF148 inhibited the activity of HBV ENII/Cp and the transcriptional activity of cccDNA. Mechanistic studies revealed that ZNF148 attenuated retinoid X receptor alpha (RXRα) expression by binding to the RXRα promoter sequence. RXRα binding site mutation or RXRα overexpression abolished the suppressive effect of ZNF148 on HBV replication. The inhibitory effect of ZNF148 was also observed in the rcccDNA mouse model. CONCLUSIONS: ZNF148 inhibited HBV replication by downregulating RXRα transcription. Our findings reveal that ZNF148 may be a new target for anti-HBV strategies.

Kinesin Family Member-18A (KIF18A) Promotes Cell Proliferation and Metastasis in Hepatocellular Carcinoma.

BACKGROUND & AIMS: Kinesin family member 18A (KIF18A) is notable for its aberrant expression across various cancer types and its pivotal role is driving cancer progression. In this study, we aim to investigate the intricate molecular mechanisms underlying the impact of KIF18A on the progression of HCC. METHODS: Western blotting assays, a quantitative real-time PCR and immunohistochemical analyses were performed to quantitatively assess KIF18A expression in HCC tissues. We then performed genetic manipulations within HCC cells by silencing endogenous KIF18A using short hairpin RNA (shRNA) and introducing exogenous plasmids to overexpress KIF18A. We monitored cell progression, analyzed cell cycle and cell apoptosis and assessed cell migration and invasion both in vitro and in vivo. Moreover, we conducted RNA-sequencing to explore KIF18A-related signaling pathways utilizing Reactome and KEGG enrichment methods and validated these critical mediators in these pathways. RESULTS: Analysis of the TCGA-LIHC database revealed pronounced overexpression of KIF18A in HCC tissues, the finding was subsequently confirmed through the analysis of clinical samples obtained from HCC patients. Notably, silencing KIF18A in cells led to an obvious inhibition of cell proliferation, migration and invasion in vitro. Furthermore, in subcutaneous and orthotopic xenograft models, suppression of KIF18A sgnificantly redudce tumor weight and the number of lung metastatic nodules. Mechanistically, KIF18A appears to facilitate cell proliferation by upregulating MAD2 and CDK1/CyclinB1 expression levels, with the activation of SMAD2/3 signaling contributing to KIF18A-driven metastasis. CONCLUSION: Our study elucidates the molecular mechanism by which KIF18A mediates proliferation and metastasis in HCC cells, offering new insights into potential therapeutic targets.

Molecular detection and genetic diversity of Rickettsia spp. in pet dogs and their infesting ticks in Harbin, northeastern China.

BACKGROUND: Pet dogs are important companion animals that share the environment within households, and play an important role in local community life. In addition, pet dogs also are reservoirs of zoonotic agents, including Rickettsia spp., thus increasing the risk of rickettsial infections in humans. It's meaningful to investigate the epidemiology of rickettsial agents in pet dogs, and make contribute to the surveillance of rickettsioses in human in China. RESULTS: In this study, a total of 496 pet dogs' blood samples and 343 ticks infested in pet dogs were collected, and the presence and prevalence of Rickettsia were determined by amplifying the partial gltA and 17-kDa genes, with an overall positive rate of 8.1 % in blood samples and 14.0 % in tick samples. In addition, the rrs, gltA, groEL, and ompA genes of rickettsial were also recovered to determine the species of Rickettsia detected furtherly. Sequencing blast and phylogenetic analyses revealed the presence of three human pathogenic Rickettsia species (Rickettsia raoultii, Candidatus Rickettsia tarasevichiae and Rickettsia felis) in samples associated with pet dogs. Moreover, all the sequences of Rickettsia that we obtained presented close relationship with others available in GenBank, and Rickettsia raoultii was the most predominant Rickettsia species infected in pet dogs' blood samples or in tick samples. CONCLUSIONS: This study provides the molecular epidemiology data about the Rickettsia spp. infection associated with pet dogs in urban areas of Harbin city. Three rickettisae species pathogenic to humans were identified from pet dogs' blood and the infested ticks in urban areas of Harbin city. Considering the intimate relationship between human and pets, these results indicate the potential transmission risk of human rickettisal infections from pet dogs through ectoparasites, and also highlighting that more attention should be paid to rickettsial infection in pet dogs and the infested ticks from the "One health" perspective.

Epidemiology and genotypic diversity of feline bocavirus identified from cats in Harbin, China.

Feline bocavirus (FBoV) is a globally distributed linear, single-stranded DNA virus infect cats, currently classified into three distinct genotypes. Although FBoV can lead to systemic infections, its complete pathogenic potential remains unclear. In this study, 289 blood samples were collected from healthy cats in Harbin, revealing an overall FBoV prevalence of 12.1%. Notably, genotypes 1 and 3 of FBoV were found co-circulating among the cat population in Harbin. Additionally, recombination events were detected, particularly in the newly discovered NG/104 and DL/102 strains. Furthermore, negative selection sites were predominantly observed across the protein coding genes of FBoV. These findings suggest a co-circulation of genetically diverse FBoV strains among cats in Harbin, indicate that purifying selection is the primary driving force shaping the genomic evolution of FBoV, and also underscore the importance of comprehensive surveillance efforts to enhance our understanding of the epidemiology and evolutionary characteristics of FBoV.

Circulation of four species of Anaplasmataceae bacteria in ticks in Harbin, northeastern China.

Ticks play an important role in the evolution and transmission of Anaplasmataceae bacteria which are agents of emerging infectious diseases. In this study, a total of 1286 adult ticks belonging to five species were collected from cattle, goats, horses and vegetation in Harbin area, Heilongjiang province, northeastern China. The tick-borne Anaplasmataceae bacteria were identified by amplifying and sequencing the 16S rRNA (rrs) and heat shock protein-60 encoding (groEL) genes. The results showed that Ixodes persulcatus was dominant (38.8%, 499/1283) among the five tick species, and Anaplasmataceae bacteria were detected in all tick species with an overall prevalence of 7.4%. Four species of Anaplasmataceae bacteria (Anaplasma phagocytophilum, Anaplasma ovis, Anaplasma bovis, and "Candidatus Neoehrlichia mikurensis"), which are pathogenic to humans and/or animals, were identified from tick samples by phylogenetic analyzes of the rrs and groEL gene sequences. Interestingly, the cluster 1 strains were first identified in Asian, and a novel cluster was also detected in this study. These data revealed the genetic diversity of Anaplasmataceae bacteria circulating in ticks in Harbin area, highlighting the need to investigate these tick-borne pathogens and their risks to human and animal health.

Molecular detection and genetic characterization of bovine hepacivirus identified in ticks collected from cattle in Harbin, northeastern China.

Bovine hepacivirus (BovHepV) is a member of the genus Hepacivirus of the family Flaviviridae, which can cause acute or persistent infections in cattle. Currently, BovHepV strains identified in cattle populations worldwide can be classified into two genotypes with eight subtypes in genotype 1. BovHepV has been identified in a wide geographic area in China. Interestingly, the viral RNA of BovHepV has also been detected in ticks in Guangdong province, China. In this study, Rhipicephalus microplus tick samples were collected in Heilongjiang province, northeastern China, and BovHepV was screened with an overall positive rate of 10.9%. Sequence comparison and phylogenetic analysis showed that the BovHepV strains detected in this study belong to the subtype G. This is the first report about the detection of BovHepV in ticks in Heilongjiang province, China, which expands our knowledge that ticks may be a transmission vector of BovHepV.

Epidemiology and genotypic diversity of canine circovirus identified in pet dogs in Harbin, China.

Canine circovirus (CanineCV) is a single-stranded DNA virus that circulates in dogs and wild carnivores around the world. It has been suggested to be associated with diseases of respiratory and gastrointestinal systems, though its pathogenic potential remains unclear. Currently, CanineCV is divided into six genotypes (genotype 1-6), and genotypes 2, 3, and 4 have been described in China. In this study, 359 blood samples from pet dogs with or without clinical signs were collected in Harbin city. After PCR screening, a total of 34 samples were tested positive for CanineCV, and nine full-length genome sequences were recovered from positive samples. Pairwise sequence comparison showed that they shared 82.4-99.3% genome-wide identity with other CanineCVs available in GenBank. Additionally, recombination events were detected, all of which were determined to be associated with sequences obtained in China. The reconstructed phylogenetic tree based on the recombination-free complete genome sequences revealed that the complete genome sequences generated herein were clustered into genotypes 1 and 3. Furthermore, purifying selection was the dominant evolutionary pressure acting on the genomes of CanineCV. These results expand the knowledge about the genetic diversity of CanineCV circulating in China, and also promote us to better understand the evolution of CanineCV.

Epidemiology and Genetic Diversity of Bartonella in Rodents in Urban Areas of Guangzhou, Southern China.

Bartonella spp. are gram-negative bacteria that can infect a wide spectrum of mammals. Rodents are considered to be the natural reservoir of many Bartonella species that are transmitted by various blood-sucking arthropods. The close contact between rodents and humans in urban areas increased the chance of transmitting rodent-borne Bartonella to humans. Investigation of the epidemiological characteristics of Bartonella infection in rodents is of great significance for the prevention and control of human Bartonellosis. In this study, rodents were captured to monitor the prevalence of Bartonella in urban areas of Guangzhou city. Six official or candidate species of Bartonella, including two confirmed zoonotic species, were detected with an overall prevalence of 6.4% in rodents captured herein. In addition, Rattus norvegicus was the predominant host species for Bartonella infection, and B. queenslandensis was the dominant species circulating in rodents in these areas. These results provide insights into the prevalence and genetic diversity of Bartonella species circulating in rodents in the urban areas of Guangzhou, and also urged the surveillance of rodent-associated Bartonella species in these areas.

Pathogenic Leptospira Species Are Widely Disseminated among Wild Rodents in Urban Areas of Guangzhou, Southern China.

Leptospirosis is a neglected zoonotic disease with global importance caused by pathogenic Leptospira. Rodents are considered the most significant reservoirs for both human and animal infection. Historically, Guangzhou has been an endemic region of human leptospirosis. Although the incidence in humans has significantly decreased in the past decades in China, the epidemiology of pathogenic Leptospira in wild rodents is of great significance for the prevention and control of human leptospirosis. In this study, a total of 296 wild rodents were trapped in urban areas of Guangzhou, in southern China, in 2020. Three pathogenic Leptospira species, i.e., Leptospira interrogans, L. borgpetersenii, and L. kirschneri, were detected by nested PCR in this wild rodent population with an overall prevalence of 9.5%. Additionally, L. interrogans was detected in three of the four captured rodent species, and the relative high prevalence suggests that L. interrogans probably represents the preponderant species of the pathogenic Leptospira circulating in Guangzhou. Taken together, this study reveals a high genetic diversity of pathogenic Leptospira disseminated among wild rodents in the urban areas of Guangzhou and emphasizes that the risk for the occurrence of human leptospirosis in Guangzhou remains high.

A Highly Divergent Hepacivirus Identified in Domestic Ducks Further Reveals the Genetic Diversity of Hepaciviruses.

Hepaciviruses represent a group of viruses that pose a significant threat to the health of humans and animals. During the last decade, new members of the genus Hepacivirus have been identified in various host species worldwide, indicating the widespread distribution of genetically diversified hepaciviruses among animals. By applying unbiased high-throughput sequencing, a novel hepacivirus, provisionally designated Hepacivirus Q, was discovered in duck liver samples collected in Guangdong province of China. Genetic analysis revealed that the complete polyprotein of Hepacivirus Q shares 23.9-46.6% amino acid identity with other representatives of the genus Hepacivirus. Considering the species demarcation criteria for hepaciviruses, Hepacivirus Q should be regarded as a novel hepacivirus species of the genus Hepacivirus within the family Flaviviridae. Phylogenetic analyses also indicate the large genetic distance between Hepacivirus Q and other known hepaciviruses. Molecular detection of this novel hepacivirus showed an overall prevalence of 15.9% in duck populations in partial areas of Guangdong province. These results expand knowledge about the genetic diversity and evolution of hepaciviruses and indicate that genetically divergent hepaciviruses are circulating in duck populations in China.

Research note: genetic diversity of duck circoviruses circulating in partial areas of Guangdong province, southern China.

Duck circovirus (DuCV) is the smallest known virus in waterfowl that infects both domestic and wild duck. Infected ducks often show stunted growth and immunosuppression, which increases the rate of secondary infection with other pathogens. In this study, 270 liver tissue samples were collected to screen the presence of DuCV in Guangdong province, China, and the complete genome sequences were recovered and systematically analyzed. Genetic analyses revealed that sequences determined in this study shared 81.6% to 100.0% genome-wide pairwise identity with previously identified DuCV genomes. Phylogenetic analyses showed that 2 DuCV genotypes with a high infection rate were co-circulating in duck population in Guangdong province, and extensive recombination events have occurred during the evolution of DuCV. Our results expand upon the knowledge regarding the genetic diversity and evolution of DuCV, and also indicate that extensive genetically divergent DuCV are co-circulating in the duck populations in Guangdong, southern China.

Extensive genetic diversity and recombination events identified in goose circoviruses circulating in partial areas of Guangdong province, Southern China.

Circoviruses represent a group of small viruses with circular single-strand DNA genome that infect a wide range of both domesticated and wild animals. Domesticated geese infected with circovirus have been confirmed in many parts of the world, and is considered to cause immunosuppression and facilitate the secondary infections caused by other pathogens. In the present study, extensive genetically diversified goose circoviruses (GoCVs) were identified in the liver samples of domesticated geese from Guangdong province, southern China. Genetic analysis revealed that the sequences generated in this study shared 81.5 to 99.7% genome-wide pairwise identity with previously identified GoCV genomes. More importantly, nine recombination events were identified among all known complete genomome sequences of GoCV including those obtained herein, and the majority was determined associate with the sequences identified from Guangdong province, suggesting that recombination is the primary driver for the diversification of GoCVs. Additionally, purifying selection was the dominant evolutionary pressure acting on the genomes of GoCVs, and the ORF C1 gene of GoCV showed a higher genetic variation than ORF V1 gene. These results expand the knowledge about the genetic diversity and evolution of GoCV, and also indicate extensive genetically divergent GoCV strains were co-circulating in goose population in partial areas of Guangdong province, southern China.