Perinatal hepatitis B virus transmission in Lao PDR: A prospective cohort study.

BACKGROUND: Mother-to-child transmission of hepatitis B virus (HBV) is the main cause of new infections worldwide. We aimed at assessing the percentage of infants successfully immunized in two major hospitals in Vientiane, Lao PDR where HB immune globulin (HBIg) is not available. METHODS: We studied a prospective cohort of chronically HBV infected pregnant women and their infants until 6 months post-partum from January 2015 to March 2017. All infants received HB vaccine at birth and 6, 10 and 14 weeks thereafter, and HBV status was assessed at 6 months of age. HBV surface gene sequencing was performed in infected mother-infant pairs. RESULTS: Of 153 mothers with HB surface antigen (HBsAg), 60 (39%) had detectable serum HBe antigen (HBeAg). HBeAg positive pregnant women were younger than those negative (median age 26 versus 28 years; p = 0.02) and had a significantly higher HBV viral load at delivery (median 8.0 versus 4.0 log10 IU/mL, p <0.001). Among the 120 infants assessed at 6 months of age, 5 (4%) were positive for HBsAg and had detectable HBV viral load by polymerase chain reaction. All were born to mothers with HBeAg and viral load >8.5 log10 IU/mL. However, only four (3.3%, 95% CI 0.5% to 7.0%) had a virus strain closely related to their mother's strain. HBV surface gene mutations were detected in 4 of the 5 infected infants. Anti-HBs antibody levels were below 10 IU/L in 10 (9%) uninfected infants at 6 months of age. CONCLUSIONS: Mother-to-child transmission occurred less frequently than expected without the use of HBIg. Adding HBIg and/or maternal antiviral prophylaxis may have prevented some of these infections. The observation of unsatisfactory levels of anti-HBs antibodies in 9% of the uninfected infants at 6 months highlights the need for improvement of the universal immunization procedures.

In vitro analysis of the susceptibility of HIV-1 subtype A and CRF01_AE integrases to raltegravir.

The antiviral efficacy of raltegravir (RAL) has been proven against human immunodeficiency virus type 1 (HIV-1) subtypes B and C but remained to be determined against other subtypes. Therefore, the enzymatic activities as well as RAL resistance of HIV-1 subtype A and CRF01_AE integrases (INs) were investigated. Previously published subtype A and CRF01_AE IN sequences from RAL-naïve patients were aligned to generate consensus sequences for both IN subtypes. Subtype A and CRF01_AE INs encoded by these consensus sequences as well as the corresponding enzymes harbouring the N155H resistance mutation were expressed and purified. Enzymatic activities of subtype A and CRF01_AE INs were analysed with regard to typical 3'-end processing (3'-P) and strand transfer (ST) activities both in the presence and absence of RAL and were compared with subtype B IN as well as with the corresponding INs harbouring the N155H resistance mutation. Subtypes B, A and CRF01_AE INs showed similar 3'-P and ST activities. In the presence of RAL, the three wild-type INs exhibited ST activity IC50 values (50% inhibitory concentrations) of 86.3 ± 32.5, 158.3 ± 99.0 and 100.0 ± 65.7 nM, respectively. Analysis of 3'-P activity in the presence of RAL revealed IC(50) > 10 µM for all three enzymes. The three INs harbouring the N155H mutation presented in vitro low but similar resistance levels to RAL. In conclusion, INs from HIV-1 subtypes B, A and CRF01_AE showed similar responses to RAL in vitro, suggesting the potency of this antiretroviral drug to treat HIV-1 subtype A- and CRF01_AE-infected patients.

Role of Hsp104 in the propagation and inheritance of the [Het-s] prion.

The chaperones of the ClpB/HSP100 family play a central role in thermotolerance in bacteria, plants, and fungi by ensuring solubilization of heat-induced protein aggregates. In addition in yeast, Hsp104 was found to be required for prion propagation. Herein, we analyze the role of Podospora anserina Hsp104 (PaHsp104) in the formation and propagation of the [Het-s] prion. We show that DeltaPaHsp104 strains propagate [Het-s], making [Het-s] the first native fungal prion to be propagated in the absence of Hsp104. Nevertheless, we found that [Het-s]-propagon numbers, propagation rate, and spontaneous emergence are reduced in a DeltaPaHsp104 background. In addition, inactivation of PaHsp104 leads to severe meiotic instability of [Het-s] and abolishes its meiotic drive activity. Finally, we show that DeltaPaHSP104 strains are less susceptible than wild type to infection by exogenous recombinant HET-s(218-289) prion amyloids. Like [URE3] and [PIN(+)] in yeast but unlike [PSI(+)], [Het-s] is not cured by constitutive PaHsp104 overexpression. The observed effects of PaHsp104 inactivation are consistent with the described role of Hsp104 in prion aggregate shearing in yeast. However, Hsp104-dependency appears less stringent in P. anserina than in yeast; presumably because in Podospora prion propagation occurs in a syncitium.

Prion and non-prion amyloids of the HET-s prion forming domain.

HET-s is a prion protein of the fungus Podospora anserina. A plausible structural model for the infectious amyloid fold of the HET-s prion-forming domain, HET-s(218-289), makes it an attractive system to study structure-function relationships in amyloid assembly and prion propagation. Here, we report on the diversity of HET-s(218-289) amyloids formed in vitro. We distinguish two types formed at pH 7 from fibrils formed at pH 2, on morphological grounds. Unlike pH 7 fibrils, the pH 2 fibrils show very little if any prion infectivity. They also differ in ThT-binding, resistance to denaturants, assembly kinetics, secondary structure, and intrinsic fluorescence. Both contain 5 nm fibrils, either bundled or disordered (pH 7) or as tightly twisted protofibrils (pH 2). We show that electrostatic interactions are critical for the formation and stability of the infectious prion fold given in the current model. The altered properties of the amyloid assembled at pH 2 may arise from a perturbation in the subunit fold or fibrillar stacking.

Methods for the in vivo and in vitro analysis of [Het-s] prion infectivity.

Prions have been described in mammals and fungi. The [Het-s] infectious genetic element of the filamentous fungus Podospora anserina is the prion form of the HET-s protein. This protein is involved in the control of a cell death reaction termed heterokaryon incompatibility. The infectious form of HET-s corresponds to a self-perpetuating amyloid. The purpose of the present paper is to describe the techniques that can be used to analyse [Het-s] prion propagation in vivo and HET-s amyloid aggregation in vitro. In addition, we report several methods that can be used to infect Podospora with recombinant HET-s amyloid.