Hepatitis B virus core protein (HBc) has become a hot spot in drug carrier protein research due to its natural particle self-assembly ability and ease of modification. The truncation of the C-terminal polyarginine domain (CTD, aa 151-183) of HBc does not affect the self-assembly of the particles. However, it does affect the internal and external charges of the particles, which may subsequently affect drug encapsulation. Thus, the truncated C-terminal polyarginine domain (CTD) of HBc and the inserted RGD peptide were selected to construct and express three HBc variants (RH) encapsulated with ICG (RH/ICG) with different C-terminal lengths to compare the stability and drug activity of their nanoformulations. RH160/ICG was found to have a great advantages in encapsulation efficiency and biological imaging. Compared with other HBc variants, RH160/ICG significantly improved encapsulation efficiency, up to 32.77%±1.23%. Cytotoxicity and hemolysis assays further demonstrated the good biocompatibility of RH160/ICG. Cell uptake and in vivo imaging experiments in mice showed that RH160/ICG could efficiently deliver ICG in tumor cells and tumor sites with good imaging effect. This research provides a new direction for further expanding the diagnosis and treatment application of ICG and development of HBc-based nanoparticle drug carrier platform.
Animals ; Hepatitis B/drug therapy* ; Hepatitis B Core Antigens ; Indocyanine Green/chemistry* ; Mice ; Nanoparticles/chemistry* ; Viral Core Proteins

Objective: To understand the epidemiological characteristics of one large HIV molecular transmission cluster in Jiaxing city, Zhejiang province, 2017 in order to select those people under high-risk and providing basis for programs on prevention. Methods: During 2017, newly diagnosed HIV/AIDS cases in this city were recruited. Plasma samples were collected from subjects, followed by RNA extraction, RT-PCR and nest-PCR for pol gene amplification, before being sequenced and aligned. Mega 6.0 software was used to construct phylogenetic tree, and Cytoscape 3.6.0 software was used to identify HIV molecular transmission clusters. Cases within the large transmission clusters were investigated, using a field-epidemiology-questionnaire. Data related to socio-demographics and previous sexual behaviors were collected and EpiData 3.0 and SPSS 20.0 software were used. Results: In the large transmission cluster with subtype identified as CRF07_BC, in Jiaxing, 2017, 26 cases of the total 30 cases were investigated. A total of 80.8% (21/26) could be identified as newly infected within the last two years and 30.8%(8/26) could be identified as newly infected within the last one year, including 22 cases infected locally. Among several infected cases who were at age 45 years or older, they admitted that they had experienced unprotected sexual contacts in local city for long time and having had more than 10 disclosed sexual contacts within the last two years at the local venues. Conclusions: This molecular cluster had been formed and scaled up quickly in recent two years, it has played an important role in promoting and scaling up the HIV transmission. Three cases identificed as high risk played an importantrde role in scaling up this cluster.
Amplified Fragment Length Polymorphism Analysis ; China/epidemiology* ; Genes, pol ; Genotype ; HIV Infections/transmission* ; HIV-1/isolation & purification* ; Humans ; Molecular Epidemiology ; Phylogeny ; Polymerase Chain Reaction ; RNA, Viral/blood* ; Sexual Behavior ; pol Gene Products, Human Immunodeficiency Virus

Porcine endogenous retroviruses (PERVs) integrate into germline DNA as proviral genome that enables vertical transmission from parents to their offspring. The provirus usually survives as part of the host genome rather than as an infectious agent, but may become pathogenic if it crosses species barriers. Therefore, replication-competent PERV should be controlled through selective breeding or knockout technologies. Two microRNAs (miRNAs), dual LTR1 and LTR2, were selected to inhibit the expression of PERV in primary porcine kidney cells. The inhibition efficiency of the miRNAs was compared based on their inhibition of different PERV regions, specifically long terminal repeats (LTRs), gag, pol, and env. Gene expression was quantified using real-time polymerase chain reaction and the C-type reverse transcriptase (RT) activity was determined. The messenger RNA (mRNA) expression of the PERV LTR and env regions was determined in HeLa cells co-cultured with primary porcine kidney cells. The mRNA expression of the LTR, gag, pol, and env regions of PERV was dramatically inhibited by dual miRNA from 24 to 144 h after transfection, with the highest inhibition observed for the LTR and pol regions at 120 h. Additionally, the RT activity of PERV in the co-culture experiment of porcine and human cells was reduced by 84.4% at the sixth passage. The dual LTR 1+2 miRNA efficiently silences PERV in primary porcine kidney cells.
Coculture Techniques ; DNA ; Endogenous Retroviruses ; Gene Expression ; Genome ; HeLa Cells ; Humans ; Kidney ; MicroRNAs ; Parents ; Proviruses ; Real-Time Polymerase Chain Reaction ; RNA, Messenger ; RNA-Directed DNA Polymerase ; Selective Breeding ; Terminal Repeat Sequences ; Transfection

To determine the inhibitory effect of endophytic fungi from Dysosma versipellis on HIV-1 IN-LEDGF/p75 interaction,the protein-protein interaction between human immunodeficiency virus type 1( HIV-1) integrase and lens epithelial growth factor p75 protein( LEDGF/p75) was used as a target. The homogeneous time-resolved fluorescence( HTRF) technique was used in the inhibitory activity assay. The results showed that eight endophytic fungi with anti-IN-LEDGF/p75 interaction activity were screened out from fifty-three strains with different morphological characteristic. Among them,106 strain showed strong inhibitory activity against HIV-1 IN-LEDGF/p75 interaction with IC50 value of 5. 23 mg·L-1,and was identified as a potential novel species of Magnaporthaceae family by the analyses of ITS-rDNA,LSU and RPB2 sequences data. This study demonstrated that potential natural active ingredients against the HIV-1 IN-LEDGF/p75 interaction exist in the endophytic fungi of D. versipellis. These results may provide available candidate strain resources for the research and development of new anti-acquired immunodeficiency syndrome drugs.
Berberidaceae ; microbiology ; Endophytes ; Fungi ; chemistry ; HIV Integrase ; metabolism ; HIV-1 ; drug effects ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Protein Binding

Objective: To understand prevalence and transmission of transmitted drug resistance (TDR) among HIV infected men who have sex with men (MSM) in Tianjin from 2014 to 2017. Methods: A total of 225 blood samples were collected from HIV infected MSM in Tianjin from 2014 to 2017. Pol gene fragments were obtained by viral RNA extraction and nested PCR amplification. Phylogenetic and drug resistance analyses were conducted. Results: A total of 205 samples were successfully sequenced and analyzed. Based on pol sequences, 53.2% (109/205), 28.8% (59/205), 10.2% (21/205), 4.9% (10/205) and 2.9% (6/205) of the samples were positive for HIV subtypes CRF01_AE, CRF07_BC, B, CRF55_01B and unique recombinant forms (URFs). Twenty transmission clusters, including 75 sequences, were identified and 62.5% (10/16) of sequences with TDR were in 5 clusters. The prevalence of TDR was 7.8% between 2014 and 2017. The annual prevalence rate increased from 3.9% (2/51) in 2014, 5.7% (3/53) in 2015, 9.6% (5/52) in 2016 to 12.2%(6/49) in 2017, the difference was not significant (χ(2)=2.504, P=0.127). CRF01_AE and B strains had high TDR prevalence (3.4%, 7/205) and (2.9%, 6/205), respectively. The TDR mutation was mainly NNRTIs, the TDR prevalence was 6.3% (13/205). In contract, the TDR prevalence of NRTIs and PIs were 1.5% (3/205) and 1.0% (2/205) respectively. Conclusion: Results from this study suggested that the prevalence of HIV-1 TDR strains in MSM was serious in Tianjin. It is necessary to take effective prevention and control measures.
China ; Drug Resistance, Viral/genetics* ; Genes, pol ; Genotype ; HIV Infections/transmission* ; HIV Reverse Transcriptase/genetics* ; HIV Seropositivity/genetics* ; HIV-1/isolation & purification* ; Homosexuality, Male/statistics & numerical data* ; Humans ; Male ; Mutation ; Phylogeny ; Polymerase Chain Reaction ; Prevalence ; RNA, Viral/genetics* ; pol Gene Products, Human Immunodeficiency Virus/genetics*

We recently reported an unconventional mechanism by which miRNAs inhibit HIV-1 viral production. This occurs when miRNAs bind nonspecifically to the viral structural protein Gag, interfering with viral RNA-mediated Gag assembly at the plasma membrane. Consequently, misassembled viral complexes are redirected into the endocytic pathway where they are delivered to lysosomes for degradation. In this study, we demonstrate that autophagy is a critical mediator of the viral degradation pathway and that this pathway is not HIV-1 specific. Misassembled viral complexes were found to colocalize extensively with LC3 and p62 in late endosomes/lysosomes, demonstrating a convergence of autophagy with functional degradative compartments. Knocking down autophagosome formation machineries reduced this convergence, while treatment with autophagy-inducer rapamycin enhanced the convergence. Furthermore, similar autophagy-dependent nonspecific miRNA inhibition of murine leukemia virus (MLV) assembly was shown. Overall, these results reveal autophagy as a crucial regulator of the retroviral degradation pathway in host cells initiated by nonspecific miRNA-Gag interactions. These findings could have significant implications for understanding how cells may regulate retroviral complex assembly by miRNA expression and autophagy, and raise the possibility that similar regulations can occur in other biological contexts.
Autophagy ; Cell Membrane ; metabolism ; Gene Products, gag ; genetics ; metabolism ; HEK293 Cells ; HIV-1 ; metabolism ; Humans ; Lysosomes ; metabolism ; MicroRNAs ; genetics ; metabolism ; Virus Assembly

The diagnostic criteria for sequential rapidly destructive coxarthrosis remain unclear and this condition is rarely reported in patients with human immunodeficiency virus (HIV). Here, we report a case of an HIV-infected 73-year old female who experienced hip joint destruction. The patient was diagnosed with HIV in 2012 (at age 68 years) and began continuous treatment with nucleoside reverse transcriptase and protease inhibitors. Twenty-nine months after her HIV diagnosis, the patient experienced osteonecrosis of the right hip and underwent a total hip arthroplasty (THA). Twelve months post right-hip THA, X-ray results showed good outcomes. Eight months later (20 months post THA), however, osteolysis of the left femoral head was detected upon radiological exam and THA of the left hip was performed; chronic inflammation and fibrosis were identified in the resultant biopsy. Favorable results were obtained at 3 months after the second surgery.
Arthroplasty, Replacement, Hip ; Biopsy ; Diagnosis ; Female ; Femur Head ; Fibrosis ; Head ; Hip ; Hip Joint ; HIV ; Humans ; Humans ; Inflammation ; Osteoarthritis, Hip ; Osteolysis ; Osteonecrosis ; Protease Inhibitors ; RNA-Directed DNA Polymerase

OBJECTIVE: Respiratory viral infection of the neonatal period is highly contagious. Rapid and accurate diagnosis is important for proper treatment and prevention. However, the existing diagnostic method, respiratory virus cell culture, takes a long time to diagnose. Recent development of rapid diagnostic methods such as multiplex reverse transcriptase polymerase chain reaction (RT-PCR) enable early detection and effective treatment of respiratory viral infections. We compared the efficiency of multiplex RT-PCR and R-mix virus culture for rapid detection of respiratory viruses. METHODS: We retrospectively analyzed the clinical features and results of R-mix virus culture and multiplex RT-PCR with nasopharyngeal aspiration specimens in 117 newborns admitted to neonatal intensive care unit suspected of infectious diseases. RESULTS: R-mix virus culture was positive in 29 cases (24.8%) and RT-PCR in 86 cases (73.5%). R-mix virus culture and multiplex RTPCR were identical in 54 cases (positive 26 cases, negative 28 cases). Among 75 cases that showed different results, 60 showed negative result in R-mix virus culture and positive result in multiplex RT-PCR, and three showed positive result in R-mix virus culture and negative result in multiplex RT-PCR. Different viruses were detected in the remaining 12 cases by both methods. CONCLUSION: Multiplex RT-PCR is faster than R-mix virus culture and has the advantage of identifying new respiratory viruses. On the other hand, Multiplex RT-PCR is more susceptible to false positives and mixed infections than R-mix virus culture, so more attention is required when interpreting test results.
Cell Culture Techniques ; Coinfection ; Communicable Diseases ; Diagnosis ; Hand ; Humans ; Infant, Newborn ; Intensive Care, Neonatal ; Methods ; Retrospective Studies ; Reverse Transcriptase Polymerase Chain Reaction* ; RNA-Directed DNA Polymerase*

There are high levels of co-incidence of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2) in porcine tissue. This study established a duplex nested reverse transcriptase polymerase chain reaction (RT-PCR) method that targets the genomic RNA of type 2 PRRSV and the mRNA of PCV2 in infected tissues. The method amplified discriminative bands of 347 bp and 265 bp specific for type 2 PRRSV and PCV2, respectively. The limits of detection of the duplex nested RT-PCR were 10(1.5) TCID₅₀/mL for type 2 PRRSV and 10² infected cells/mL for PCV2. The kappa statistic, which measures agreement between methods, was 0.867, indicating a good level of agreement. This RNA-based duplex RT-PCR approach can be another way to detect type 2 PRRSV and PCV2 simultaneously and with improved convenience.
Circovirus ; Limit of Detection ; Methods ; Porcine Reproductive and Respiratory Syndrome* ; Porcine respiratory and reproductive syndrome virus* ; Reverse Transcriptase Polymerase Chain Reaction* ; RNA ; RNA, Messenger ; RNA-Directed DNA Polymerase*

The advent of oral antiviral agents has revolutionized hepatitis B treatment. It has led to decreased incidence and mortality related to hepatocellular carcinoma. However, although nucleos(t)ide analogs (NA) are fast and potent in inhibiting hepatitis B virus (HBV) polymerase and reverse transcriptase activity, complete cure of the virus is not possible. The complete eradication of HBV requires the covalently-closed-circular DNA (cccDNA) to be eliminated. Novel gene editing methods, such as zing finger nucleases, transcription activator-like effector nucleases, and the clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9) system, designed to target specific DNA sequences has great potential for therapeutic application. Among these, the CRISPR/Cas9 system may be the most feasible approach to eradicate HBV cccDNA. Further studies are needed to develop a more efficient and safer method of delivery using the CRISPR/Cas9 system to achieve complete cure of chronic hepatitis B.
Antiviral Agents ; Base Sequence ; Carcinoma, Hepatocellular ; Clustered Regularly Interspaced Short Palindromic Repeats ; DNA ; DNA, Circular ; Fingers ; Hepatitis B virus* ; Hepatitis B* ; Hepatitis B, Chronic ; Hepatitis* ; Incidence ; Methods ; Mortality ; RNA-Directed DNA Polymerase