Interaction and assembly of the DNA replication core proteins of Kaposi's sarcoma-associated herpesvirus.

IMPORTANCE: Eukaryotic DNA replication is a highly regulated process that requires multiple replication enzymes assembled onto DNA replication origins. Due to the complexity of the cell's DNA replication machinery, most of what we know about cellular DNA replication has come from the study of viral systems. Herein, we focus our study on the assembly of the Kaposi's sarcoma-associated herpesvirus core replication complex and propose a pairwise protein-protein interaction network of six highly conserved viral core replication proteins. A detailed understanding of the interaction and assembly of the viral core replication proteins may provide opportunities to develop new strategies against viral propagation.

Stock Market Forecasting Based on Spatiotemporal Deep Learning.

This study introduces the Spacetimeformer model, a novel approach for predicting stock prices, leveraging the Transformer architecture with a time-space mechanism to capture both spatial and temporal interactions among stocks. Traditional Long-Short Term Memory (LSTM) and recent Transformer models lack the ability to directly incorporate spatial information, making the Spacetimeformer model a valuable addition to stock price prediction. This article uses the ten minute stock prices of the constituent stocks of the Taiwan 50 Index and the intraday data of individual stock on the Taiwan Stock Exchange. By training the Timespaceformer model with multi-time-step stock price data, we can predict the stock prices at every ten minute interval within the next hour. Finally, we also compare the prediction results with LSTM and Transformer models that only consider temporal relationships. The research demonstrates that the Spacetimeformer model consistently captures essential trend changes and provides stable predictions in stock price forecasting. This article proposes a Spacetimeformer model combined with daily moving windows. This method has superior performance in stock price prediction and also demonstrates the significance and value of the space-time mechanism for prediction. We recommend that people who want to predict stock prices or other financial instruments try our proposed method to obtain a better return on investment.

Enzymatic reactions of AGO4 in RNA-directed DNA methylation: siRNA duplex loading, passenger strand elimination, target RNA slicing, and sliced target retention.

RNA-directed DNA methylation in plants is guided by 24-nt siRNAs generated in parallel with 23-nt RNAs of unknown function. We show that 23-nt RNAs function as passenger strands during 24-nt siRNA incorporation into AGO4. The 23-nt RNAs are then sliced into 11- and 12-nt fragments, with 12-nt fragments remaining associated with AGO4. Slicing recapitulated with recombinant AGO4 and synthetic RNAs reveals that siRNAs of 21-24 nt, with any 5'-terminal nucleotide, can guide slicing, with sliced RNAs then retained by AGO4. In vivo, RdDM target locus RNAs that copurify with AGO4 also display a sequence signature of slicing. Comparing plants expressing slicing-competent versus slicing-defective AGO4 shows that slicing elevates cytosine methylation levels at virtually all RdDM loci. We propose that siRNA passenger strand elimination and AGO4 tethering to sliced target RNAs are distinct modes by which AGO4 slicing enhances RNA-directed DNA methylation.

Ranking the environmental factors of indoor air quality of metropolitan independent coffee shops by Random Forests model.

Independent coffee shops are the alternative workplaces for people working remotely from traditional offices but are not concerned about their indoor air quality (IAQ). This study aimed to rank the environmental factors in affecting the IAQ by Random Forests (RFs) models. The indoor environments and human activities of participated independent coffee shops were observed and recorded for 3 consecutive days including weekdays and weekend during the business hours. The multi-sized particulate matter (PM), particle-bound polycyclic aromatic hydrocarbons (p-PAHs), total volatile organic compounds (TVOCs), CO, CO2, temperature and relative humidity were monitored. RFs models ranked the environmental factors. More than 20% of the 15-min average concentrations of PM10, PM2.5, and CO2 exceeded the World Health Organization guidelines. Occupant density affected TVOCs, p-PAHs and CO2 concentrations directly. Tobacco smoking dominated PM10, PM2.5, TVOCs and p-PAHs concentrations mostly. CO concentration was affected by roasting bean first and tobacco smoking secondly. The non-linear relationships between temperature and these pollutants illustrated the relative low concentrations happened at temperature between 22 and 24 °C. Tobacco smoking, roasting beans and occupant density are the observable activities to alert the IAQ change. Decreasing CO2 and optimizing the room temperature could also be the surrogate parameters to assure the IAQ.

Reaction Mechanisms of Pol IV, RDR2, and DCL3 Drive RNA Channeling in the siRNA-Directed DNA Methylation Pathway.

In eukaryotes with multiple small RNA pathways, the mechanisms that channel RNAs within specific pathways are unclear. Here, we reveal the reactions that account for channeling in the small interfering RNA (siRNA) biogenesis phase of the Arabidopsis RNA-directed DNA methylation pathway. The process begins with template DNA transcription by NUCLEAR RNA POLYMERASE IV (Pol IV), whose atypical termination mechanism, induced by nontemplate DNA base-pairing, channels transcripts to the associated RNA-dependent RNA polymerase RDR2. RDR2 converts Pol IV transcripts into double-stranded RNAs and then typically adds an extra untemplated 3' terminal nucleotide to the second strands. The dicer endonuclease DCL3 cuts resulting duplexes to generate 24- and 23-nt siRNAs. The 23-nt RNAs bear the untemplated terminal nucleotide of the RDR2 strand and are underrepresented among ARGONAUTE4-associated siRNAs. Collectively, our results provide mechanistic insights into Pol IV termination, Pol IV-RDR2 coupling, and RNA channeling, from template DNA transcription to siRNA strand discrimination.

Sharing the load: Mex67-Mtr2 cofunctions with Los1 in primary tRNA nuclear export.

Eukaryotic transfer RNAs (tRNAs) are exported from the nucleus, their site of synthesis, to the cytoplasm, their site of function for protein synthesis. The evolutionarily conserved ß-importin family member Los1 (Exportin-t) has been the only exporter known to execute nuclear export of newly transcribed intron-containing pre-tRNAs. Interestingly, LOS1 is unessential in all tested organisms. As tRNA nuclear export is essential, we previously interrogated the budding yeast proteome to identify candidates that function in tRNA nuclear export. Here, we provide molecular, genetic, cytological, and biochemical evidence that the Mex67-Mtr2 (TAP-p15) heterodimer, best characterized for its essential role in mRNA nuclear export, cofunctions with Los1 in tRNA nuclear export. Inactivation of Mex67 or Mtr2 leads to rapid accumulation of end-matured unspliced tRNAs in the nucleus. Remarkably, merely fivefold overexpression of Mex67-Mtr2 can substitute for Los1 in los1Δ cells. Moreover, in vivo coimmunoprecipitation assays with tagged Mex67 document that the Mex67 binds tRNAs. Our data also show that tRNA exporters surprisingly exhibit differential tRNA substrate preferences. The existence of multiple tRNA exporters, each with different tRNA preferences, may indicate that the proteome can be regulated by tRNA nuclear export. Thus, our data show that Mex67-Mtr2 functions in primary nuclear export for a subset of yeast tRNAs.

Multiple Layers of Stress-Induced Regulation in tRNA Biology.

tRNAs are the fundamental components of the translation machinery as they deliver amino acids to the ribosomes during protein synthesis. Beyond their essential function in translation, tRNAs also function in regulating gene expression, modulating apoptosis and several other biological processes. There are multiple layers of regulatory mechanisms in each step of tRNA biogenesis. For example, tRNA 3' trailer processing is altered upon nutrient stress; tRNA modification is reprogrammed under various stresses; nuclear accumulation of tRNAs occurs upon nutrient deprivation; tRNA halves accumulate upon oxidative stress. Here we address how environmental stresses can affect nearly every step of tRNA biology and we describe the possible regulatory mechanisms that influence the function or expression of tRNAs under stress conditions.

In vivo biochemical analyses reveal distinct roles of ß-importins and eEF1A in tRNA subcellular traffic.

Bidirectional tRNA movement between the nucleus and the cytoplasm serves multiple biological functions. To gain a biochemical understanding of the mechanisms for tRNA subcellular dynamics, we developed in vivo ß-importin complex coimmunoprecipitation (co-IP) assays using budding yeast. Our studies provide the first in vivo biochemical evidence that two ß-importin family members, Los1 (exportin-t) and Msn5 (exportin-5), serve overlapping but distinct roles in tRNA nuclear export. Los1 assembles complexes with RanGTP and tRNA. Both intron-containing pre-tRNAs and spliced tRNAs, regardless of whether they are aminoacylated, assemble into Los1-RanGTP complexes, documenting that Los1 participates in both primary nuclear export and re-export of tRNAs to the cytoplasm. In contrast, ß-importin Msn5 preferentially assembles with RanGTP and spliced, aminoacylated tRNAs, documenting its role in tRNA nuclear re-export. Tef1/2 (the yeast form of translation elongation factor 1α [eEF1A]) aids the specificity of Msn5 for aminoacylated tRNAs to form a quaternary complex consisting of Msn5, RanGTP, aminoacylated tRNA, and Tef1/2. Assembly and/or stability of this quaternary complex requires Tef1/2, thereby facilitating efficient re-export of aminoacylated tRNAs to the cytoplasm.

Quality Control Pathways for Nucleus-Encoded Eukaryotic tRNA Biosynthesis and Subcellular Trafficking.

tRNAs perform an essential role in translating the genetic code. They are long-lived RNAs that are generated via numerous posttranscriptional steps. Eukaryotic cells have evolved numerous layers of quality control mechanisms to ensure that the tRNAs are appropriately structured, processed, and modified. We describe the known tRNA quality control processes that check tRNAs and correct or destroy aberrant tRNAs. These mechanisms employ two types of exonucleases, CCA end addition, tRNA nuclear aminoacylation, and tRNA subcellular traffic. We arrange these processes in order of the steps that occur from generation of precursor tRNAs by RNA polymerase (Pol) III transcription to end maturation and modification in the nucleus to splicing and additional modifications in the cytoplasm. Finally, we discuss the tRNA retrograde pathway, which allows tRNA reimport into the nucleus for degradation or repair.

Separate responses of karyopherins to glucose and amino acid availability regulate nucleocytoplasmic transport.

The importin-ß family members (karyopherins) mediate the majority of nucleocytoplasmic transport. Msn5 and Los1, members of the importin-ß family, function in tRNA nuclear export. tRNAs move bidirectionally between the nucleus and the cytoplasm. Nuclear tRNA accumulation occurs upon amino acid (aa) or glucose deprivation. To understand the mechanisms regulating tRNA subcellular trafficking, we investigated whether Msn5 and Los1 are regulated in response to nutrient availability. We provide evidence that tRNA subcellular trafficking is regulated by distinct aa-sensitive and glucose-sensitive mechanisms. Subcellular distributions of Msn5 and Los1 are altered upon glucose deprivation but not aa deprivation. Redistribution of tRNA exportins from the nucleus to the cytoplasm likely provides one mechanism for tRNA nuclear distribution upon glucose deprivation. We extended our studies to other members of the importin-ß family and found that all tested karyopherins invert their subcellular distributions upon glucose deprivation but not aa deprivation. Glucose availability regulates the subcellular distributions of karyopherins likely due to alteration of the RanGTP gradient since glucose deprivation causes redistribution of Ran. Thus nuclear-cytoplasmic distribution of macromolecules is likely generally altered upon glucose deprivation due to collapse of the RanGTP gradient and redistribution of karyopherins between the nucleus and the cytoplasm.

Identification and characterization of two novel spliced genes located in the orf47-orf46-orf45 gene locus of Kaposi's sarcoma-associated herpesvirus.

UNLABELLED: The orf47-orf46-orf45 gene cluster of Kaposi's sarcoma-associated herpesvirus (KSHV) is known to serially encode glycoprotein L (gL), uracil DNA glycosylase, and a viral tegument protein. Here, we identify two novel mRNA variants, orf47/45-A and orf47/45-B, alternatively spliced from a tricistronic orf47-orf46-orf45 mRNA that is expressed in the orf47-orf46-orf45 gene locus during the early stages of viral reactivation. The spliced gene products, ORF47/45-A and ORF47/45-B, consist of only a partial region of gL (ORF47), a unique 7-amino-acid motif, and the complete tegument protein ORF45. Like the ORF45 protein, ORF47/45-A and ORF47/45-B expressed in cells sufficiently activate the phosphorylation of p90 ribosomal S6 kinase (RSK) and extracellular signal-regulated protein kinase (ERK). However, unlike ORF45, both ORF47/45-A and ORF47/45-B contain a signal peptide sequence and are localized at the endoplasmic reticulum (ER). Additionally, we found that ORF47/45-A and ORF47/45-B have an extra function that mediates the upregulation of GRP78, a master regulator of ER homeostasis. The important event regarding GRP78 upregulation can be observed in all tested KSHV-positive cell lines after viral reactivation, and knockdown of GRP78 in cells significantly impairs viral lytic cycle progression, especially at late lytic stages. Compared with some other viral glycoproteins synthesized through the ER, our results strongly implicate that the ORF47/45 proteins may serve as key effectors for controlling GRP78 expression and ER homeostasis in cells. Taken together, our findings provide evidence showing the reciprocal association between the modulation of ER homeostasis and the progression of the KSHV lytic cycle. IMPORTANCE: Emerging evidence has shown that several viruses appear to use different strategies to control ER homeostasis for supporting their productive infections. The two parts of this study identify two aspects of the association between the regulation of ER homeostasis and the progression of the KSHV lytic cycle. The first part characterizes the function of two early lytic cycle proteins, ORF47/45-A and ORF47/45-B, on the activation of a major ER chaperone protein, GRP78. In addition to the ability to promote GRP78 upregulation, the ORF47/45 proteins also activate the phosphorylation of RSK and ERK. The second part reveals that upregulation of GRP78 is essential for the progression of the KSHV lytic cycle, especially at late stages. We therefore propose that activation of GRP78 expression by viral proteins at the early lytic stage may aid with the protection of host cells from severe ER stress and may directly involve the assembly or release of virions.

ORF50-dependent and ORF50-independent activation of the ORF45 gene of Kaposi's sarcoma-associated herpesvirus.

The ORF45 gene of Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a multifunctional tegument protein. Here, we characterize the transcriptional control of the ORF45 gene and show that its promoter can be activated by ORF50 protein, a latent-lytic switch transactivator. The ORF45 promoter can also be induced by sodium butyrate (SB), a histone deacetylase inhibitor, in the absence of ORF50 protein. Although SB induces the ORF45 gene independently of ORF50, its full activation may require the presence of ORF50. Deletion and point mutation analyses revealed that two RBP-Jκ-binding sites in the ORF45 promoter confer the ORF50 responsiveness, whereas NF-Y and Sp1-binding sites mediate the response to SB. Direct binding of NF-Y, Sp1, or RBP-Jκ protein to the ORF45 promoter is required for the promoter activation induced by SB or by ORF50. In conclusion, our study demonstrates both ORF50-dependent and ORF50-independent transcriptional mechanisms operated on the activation of the ORF45 gene.

A rapid and sensitive non-radioactive method applicable for genome-wide analysis of Saccharomyces cerevisiae genes involved in small RNA biology.

Conventional isolation and detection methods for small RNAs from yeast cells have been designed for a limited number of samples. In order to be able to conduct a genome-wide assessment of how each gene product impacts upon small RNAs, we developed a rapid method for analysing small RNAs from Saccharomyces cerevisiae wild-type (wt) and mutants cells in the deletion and temperature-sensitive (ts) collections. Our method implements three optimized techniques: a procedure for growing small yeast cultures in 96-deepwell plates, a fast procedure for small RNA isolation from the plates, and a sensitive non-radioactive northern method for RNA detection. The RNA isolation procedure requires only 4 h for processing 96 samples, is highly reproducible and yields RNA of good quality and quantity. The non-radioactive northern method employs digoxigenin (DIG)-labelled DNA probes and chemiluminescence. It detects femtomole levels of small RNAs within 1 min exposure time. We minimized the processing time for large-scale analysis and optimized the stripping and reprobing procedures for analyses of multiple RNAs from a single membrane. The method described is rapid, sensitive, safe and cost-effective for genome-wide screens of novel genes involved in the biogenesis, subcellular trafficking and stability of small RNAs. Moreover, it will be useful to educational laboratory class venues and to research institutions with limited access to radioisotopes or robots.

Saccharomyces cerevisiae possesses a stress-inducible glycyl-tRNA synthetase gene.

Aminoacyl-tRNA synthetases are a large family of housekeeping enzymes that are pivotal in protein translation and other vital cellular processes. Saccharomyces cerevisiae possesses two distinct nuclear glycyl-tRNA synthetase (GlyRS) genes, GRS1 and GRS2. GRS1 encodes both cytoplasmic and mitochondrial activities, while GRS2 is essentially silent and dispensable under normal conditions. We herein present evidence that expression of GRS2 was drastically induced upon heat shock, ethanol or hydrogen peroxide addition, and high pH, while expression of GRS1 was somewhat repressed under those conditions. In addition, GlyRS2 (the enzyme encoded by GRS2) had a higher protein stability and a lower K(M) value for yeast tRNA(Gly) under heat shock conditions than under normal conditions. Moreover, GRS2 rescued the growth defect of a GRS1 knockout strain when highly expressed by a strong promoter at 37 °C, but not at the optimal temperature of 30 °C. These results suggest that GRS2 is actually an inducible gene that may function to rescue the activity of GRS1 under stress conditions.

The effect of a simple traditional exercise programme (Baduanjin exercise) on sleep quality of older adults: a randomized controlled trial.

BACKGROUND: Sleep disturbance is a major problem for older adults, calling for the development of alternative medicine techniques to help improve sleep quality in this population. OBJECTIVES: The purpose of this study was to explore the effectiveness of a Baduanjin exercise program on sleep quality in Taiwanese elderly. DESIGN: A randomized controlled trial, longitudinal research design was employed. SETTING AND PARTICIPANTS: This study was conducted in an urban area of northern Taiwan. The inclusion criteria for participants were as follows: (1) community-dwelling older adults age 60 years or older; (2) no regular exercise within 6 months; (3) able to communicate; and (4) independent in self-care. Subjects were excluded if they had (1) depression tendency as demonstrated by the Chinese version of the Geriatric Depression Score of eight or higher; (2) impaired mobility; or (3) unstable health status. A total of 202 older people were screened for this study, 87 of whom were eligible according to the inclusion criteria. Of these, 55 completed the 12-week study. METHODS: Fifty-five participants were randomly assigned to the exercise group (27) or the control group (28). Those in the exercise group received 12 weeks of Baduanjin exercise training, while those in the control group had no intervention. The Pittsburg Sleep Quality Index was administered to subjects at four time points: before the intervention, and at the 4th, 8th, and 12th week after intervention. RESULTS: Subjects in the Baduanjin exercise group had significantly improved overall sleep quality, subjective sleep quality, sleep latency, sleep duration, sleep efficiency, and daytime dysfunction after 12 weeks of intervention (p<0.001), while those in control group showed no significant difference in sleep quality. Compared with the control group, the Baduanjin exercise group reported significantly better sleep quality after four weeks of intervention which was maintained throughout the 12-week exercise period. CONCLUSIONS: This study confirmed that the Baduanjin exercise program can improve sleep quality for Taiwanese community-dwelling older adults. We recommend application of this simple, gentle exercise program in older persons to improve their sleep quality.

Rapid and economic DNA extraction from a single salmon egg for real-time PCR amplification.

Salmon eggs are common in Japanese sushi and other seafood products; however, certain fish eggs are used as counterfeit salmon eggs which are found in foods and processed products. This study develops a simple, rapid, and cost-effective method for DNA extraction, filtration (FT) and dilution (DL) protocols from a single salmon egg with good DNA quality for real-time PCR amplification. The DNA amount, DNA quality, and real-time PCR performance for different dilutions and different lengths of PCR amplicons were evaluated and compared with the common Qiagen tissue kit (QTK) and Chelex-100-based (CX) protocols. The extracted DNA from a single salmon egg using the FT or DL protocol can be applied in phylogenic research, food authentication and post-marketing monitoring of genetically modified (GM) food products.

Regulation of tRNA bidirectional nuclear-cytoplasmic trafficking in Saccharomyces cerevisiae.

tRNAs in yeast and vertebrate cells move bidirectionally and reversibly between the nucleus and the cytoplasm. We investigated roles of members of the beta-importin family in tRNA subcellular dynamics. Retrograde import of tRNA into the nucleus is dependent, directly or indirectly, upon Mtr10. tRNA nuclear export utilizes at least two members of the beta-importin family. The beta-importins involved in nuclear export have shared and exclusive functions. Los1 functions in both the tRNA primary export and the tRNA reexport processes. Msn5 is unable to export tRNAs in the primary round of export if the tRNAs are encoded by intron-containing genes, and for these tRNAs Msn5 functions primarily in their reexport to the cytoplasm. The data support a model in which tRNA retrograde import to the nucleus is a constitutive process; in contrast, reexport of the imported tRNAs back to the cytoplasm is regulated by the availability of nutrients to cells and by tRNA aminoacylation in the nucleus. Finally, we implicate Tef1, the yeast orthologue of translation elongation factor eEF1A, in the tRNA reexport process and show that its subcellular distribution between the nucleus and cytoplasm is dependent upon Mtr10 and Msn5.

Disability and pain management methods of Taiwanese arthritic older patients.

AIM: This study aims to investigate the prevalence of disability, factors influencing disability and pain self-management techniques employed by older arthritis patients in Taiwan. BACKGROUND: Arthritis is the most common chronic disease in older people. It may result in pain, stiffness and joint deformity, which ultimately lead to disability. Understanding the factors influencing disability is needed to help arthritis sufferers to achieve optimal health status. DESIGN: A cross-sectional design was employed for this study. METHODS: One hundred and fifty-one older persons diagnosed with either rheumatoid arthritis or osteoarthritis were interviewed in the rheumatology clinics located in two medical centres in northern Taiwan. Six questionnaires were administered: Barthel Index, Instrumental Activity of Daily Living, Rapid Assessment of Disease Activity in Rheumatology, the Chinese version of Pain Management Inventory, Geriatric Depression Scale and Life Satisfaction Index. RESULTS: Disability was found in 11% of Taiwanese individuals diagnosed with either rheumatoid arthritis or osteoarthritis. Those in disability reported more severe disease activity, pain, depression and lower life satisfaction. Hierarchical multiple regression analysis revealed that 31-46% of the total variance of disability could be explained by age, gender, marriage, joint pain score, diagnosis, disease activity, depression and pain management. Patients with rheumatoid arthritis had significantly higher levels of disability, disease activity during the preceding six months, more depression and less life satisfaction than patients with osteoarthritis. CONCLUSION: Higher disability was explained by older age, female, unmarried, diagnosed with rheumatoid arthritis, more joint pain, more disease severity, more depression and more use of pain management strategies in arthritis patients. RELEVANCE TO CLINICAL PRACTICE: Nurses are urged to recognise the individual differences among the factors that are thought to contribute most to disability. An individualised, multidimensional and comprehensive treatment plan with informational support is essential to maximise pain management skills of arthritic older people to achieve improvement in pain, level of disability and mental health.

A novel growth hormone 1 gene-derived probe for Oncorhynchus masou formosanus distinguished from the Oncorhynchus subspecies.

Traditional mitochondrial 16S rRNA is commonly used in many species identification studies. However, it is difficult to apply to the phylogenetic studies among the Oncorhynchus subspecies, which is a crucial need for management purposes for Oncorhynchus masou formosanus, Taiwan salmon. In this study, we have developed an improved species identification method for Taiwan salmon distinguished with other Oncorhynchus subspecies tested by exploiting PCR for growth hormone (GH) 1 gene. By comparing DNA sequences for GH1 from 11 species of Oncorhynchus subspecies we designed novel PCR primers that exploit differences between Taiwan salmon and other Oncorhynchus subspecies. Therefore, the technique is an important tool in the management of populations of the endangered land-locked Taiwan salmon preventing from their possible hybrids with other Oncorhynchus subspecies once tested.

Forecasting the declining rate of chronic hepatitis-B carrier status at a Taiwanese university: twenty years after implementation of an universal HBV vaccination program in Taiwan.

BACKGROUND: Prior to the introduction of universal hepatitis B virus (HBV) vaccination in Taiwan in 1984, 15-20% of the general population were chronic HBV carriers. METHODS: We forecasted and quantified the declining HBV carrier rate 20 years subsequent to the implementation of universal HBV vaccination in Taiwan. At a Taiwanese university, 28,763 freshmen tested for serum HBsAg level were divided into ten age cohorts by date of birth, from July 1976 to June 1986 inclusive. Comparisons of HBsAg carrier rates according to gender were examined with the Z test. Regression methods and a time series model were applied to our sample to forecast trends in changes to the HBsAg carrier rate for the next five years. RESULTS: Regression analysis demonstrated a trend toward declining HBsAg-positive carrier rates. The HBsAg carrier rate for male students decreased from 16.8% (for those born between July 1976 and June 1977) to 2.2% (for those born between July 1985 and June 1986). The carrier rate for their female counterparts over the same period declined from 12.2% to 2.4%. The HBsAg carrier rate for male participants was significantly greater than that of their female counterparts for certain years during the test period. The results of time series analysis suggests the HBsAg carrier status rate will approach zero for students born after July 1987 (expected to enrol in the university in 2006). CONCLUSIONS: Our data demonstrate that in order for the HBV carrier rate to approximate zero, universal vaccination programs need to continue for at least 21 years.