Prevention of Herpes Simplex Virus Induced Stromal Keratitis by a Glycoprotein B-Specific Monoclonal Antibody

The increasing incidence of acyclovir (ACV) and multidrug-resistant strains in patients with corneal HSV-1 infections leading to Herpetic Stromal Keratitis (HSK) is a major health problem in industrialized countries and often results in blindness. To overcome this obstacle, we have previously developed an HSV-gB-specific monoclonal antibody (mAb 2c) that proved to be highly protective in immunodeficient NOD/SCID-mice towards genital infections. In the present study, we examined the effectivity of mAb 2c in preventing the immunopathological disease HSK in the HSK BALB/c mouse model. Therefore, mice were inoculated with HSV-1 strain KOS on the scarified cornea to induce HSK and subsequently either systemically or topically treated with mAb 2c. Systemic treatment was performed by intravenous administration of mAb 2c 24 h prior to infection (pre-exposure prophylaxis) or 24, 40, and 56 hours after infection (post-exposure immunotherapy). Topical treatment was performed by periodical inoculations (5 times per day) of antibody-containing eye drops as control, starting at 24 h post infection. Systemic antibody treatment markedly reduced viral loads at the site of infection and completely protected mice from developing HSK. The administration of the antiviral antibody prior or post infection was equally effective. Topical treatment had no improving effect on the severity of HSK. In conclusion, our data demonstrate that mAb 2c proved to be an excellent drug for the treatment of corneal HSV-infections and for prevention of HSK and blindness. Moreover, the humanized counterpart (mAb hu2c) was equally effective in protecting mice from HSV-induced HSK when compared to the parental mouse antibody. These results warrant the future development of this antibody as a novel approach for the treatment of corneal HSV-infections in humans.     

Catecholaminergic Gene Polymorphisms Are Associated with GI Symptoms and Morphological Brain Changes in Irritable Bowel Syndrome

Background

Genetic and environmental factors contribute to the pathophysiology of irritable bowel syndrome (IBS). In particular, early adverse life events (EALs) and the catecholaminergic system have been implicated.

Aims

To investigate whether catecholaminergic SNPs with or without interacting with EALs are associated with: 1) a diagnosis of IBS, 2) IBS symptoms and 3) morphological alterations in brain regions associated with somatosensory, viscerosensory, and interoceptive processes.

Methods

In 277 IBS and 382 healthy control subjects (HCs), 11 SNPs in genes of the catecholaminergic signaling pathway were genotyped. A subset (121 IBS, 209 HCs) underwent structural brain imaging (magnetic resonance imaging [MRI]). Logistic and linear regressions evaluated each SNP separately and their interactions with EALs in predicting IBS and GI symptom severity, respectively. General linear models determined grey matter (GM) alterations from the SNPs and EALs that were predictive of IBS.

Results

1) Diagnosis: There were no statistically significant associations between the SNPs and IBS status with or without the interaction with EAL after adjusting for multiple comparisons. 2) Symptoms: GI symptom severity was associated with ADRA1D rs1556832 (P = 0.010). 3) Brain morphometry: In IBS, the homozygous genotype of the major ADRA1D allele was associated with GM increases in somatosensory regions (FDR q = 0.022), left precentral gyrus (q = 0.045), and right hippocampus (q = 0.009). In individuals with increasing sexual abuse scores, the ADRAβ2 SNP was associated with GM changes in the left posterior insula (q = 0.004) and left putamen volume (q = 0.029).

Conclusion

In IBS, catecholaminergic SNPs are associated with symptom severity and morphological changes in brain regions concerned with sensory processing and modulation and affect regulation. Thus, certain adrenergic receptor genes may facilitate or worsen IBS symptoms.     

Effectiveness and safety of a long-acting,once-daily,two-phase release formulation of methylphenidate (Ritalin<Superscript>®</Superscript> LA) in school children under daily practice conditions

Long-acting (LA) preparations of methylphenidate allow for once-daily dosing; however, pharmacokinetics may vary and depend on food intake. The objective was to evaluate effectiveness of a two-phase release formulation (Ritalin^® LA) under daily practice conditions. This was a prospective, multicenter, observational study in Germany. Eligibility and dosing were determined by the physician based on the drug label. Outcomes included changes over 3 months of treatment in assessments of effect duration, clinical global impression (CGI), and quality of life (ILK). In 101 sites, 262 patients (197 boys, 63 girls, and two unknown) with a mean age of 10.9 years were enrolled; 50 were treated for the first time; 212 switched medication to Ritalin^® LA. After 3 months, CGI improved in 59.4 % of patients, and well-being overall was rated as good by 61.0 % of parents and 63.7 % of children. Based on parents’ assessment, the proportion of children suffering from strong disease burden decreased from 40.7 to 15.1 %. In 123 insufficient responders to previous ADHD medications, benefit from Ritalin^® LA was above average and effect duration was significantly prolonged as compared to pretreatment. Overall, 28 patients (10.7 %) had treatment-related adverse events with one case being serious; 23 patients (8.8 %) discontinued therapy, 7 (2.7 %) due to poor treatment response; and 212 patients (81 %) continued treatment beyond the study. In line with clinical trial data, Ritalin^® LA provides significant benefit also under routine practice conditions.     

Delivery of a Chlamydial Adhesin N-PmpC Subunit Vaccine to the Ocular Mucosa Using Particulate Carriers

Trachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), remains the world’s leading preventable infectious cause of blindness. Recent attempts to develop effective vaccines rely on modified chlamydial antigen delivery platforms. As the mechanisms engaged in the pathology of the disease are not fully understood, designing a subunit vaccine specific to chlamydial antigens could improve safety for human use. We propose the delivery of chlamydia-specific antigens to the ocular mucosa using particulate carriers, bacterial ghosts (BGs). We therefore characterized humoral and cellular immune responses after conjunctival and subcutaneous immunization with a N-terminal portion (amino acid 1–893) of the chlamydial polymorphic membrane protein C (PmpC) of Ct serovar B, expressed in probiotic Escherichia coli Nissle 1917 bacterial ghosts (EcN BGs) in BALB/c mice. Three immunizations were performed at two-week intervals, and the immune responses were evaluated two weeks after the final immunization in mice. In a guinea pig model of ocular infection animals were immunized in the same manner as the mice, and protection against challenge was assessed two weeks after the last immunization. N-PmpC was successfully expressed within BGs and delivery to the ocular mucosa was well tolerated without signs of inflammation. N-PmpC-specific mucosal IgA levels in tears yielded significantly increased levels in the group immunized via the conjunctiva compared with the subcutaneously immunized mice. Immunization with N-PmpC EcN BGs via both immunization routes prompted the establishment of an N-PmpC-specific IFNγ immune response. Immunization via the conjunctiva resulted in a decrease in intensity of the transitional inflammatory reaction in conjunctiva of challenged guinea pigs compared with subcutaneously and non-immunized animals. The delivery of the chlamydial subunit vaccine to the ocular mucosa using a particulate carrier, such as BGs, induced both humoral and cellular immune responses. Further investigations are needed to improve the immunization scheme and dosage.     

An Evaluation of the Performance and Economics of Membranes and Separators in Single Chamber Microbial Fuel Cells Treating Domestic Wastewater

The cost of materials is one of the biggest barriers for wastewater driven microbial fuel cells (MFCs). Many studies use expensive materials with idealistic wastes. Realistically the choice of an ion selective membrane or nonspecific separators must be made in the context of the cost and performance of materials available. Fourteen membranes and separators were characterized for durability, oxygen diffusion and ionic resistance to enable informed membrane selection for reactor tests. Subsequently MFCs were operated in a cost efficient reactor design using Nafion, ethylene tetrafluoroethylene (ETFE) or polyvinylidene fluoride (PVDF) membranes, a nonspecific separator (Rhinohide), and a no-membrane design with a carbon-paper internal gas diffusion cathode. Peak power densities during polarisation, from MFCs using no-membrane, Nafion and ETFE, reached 67, 61 and 59 mWm^-2, and coulombic efficiencies of 68±11%, 71±12% and 92±6%, respectively. Under 1000Ω, Nafion and ETFE achieved an average power density of 29 mWm^-2 compared to 24 mWm^-2 for the membrane-less reactors. Over a hypothetical lifetime of 10 years the generated energy (1 to 2.5 kWhm^-2) would not be sufficient to offset the costs of any membrane and separator tested.     

High Transmitter CD4+ T-Cell Count Shortly after the Time of Transmission in a Study of African Serodiscordant Couples

Background

2013 WHO guidelines recommend starting ART at CD4+ T-cell counts ≤500 cells/μL. We present the T-cell counts from adult Africans with HIV shortly following transmission to their sexual partners.

Methods

HIV-discordant couples in Zambia, Uganda and Rwanda were followed prospectively and received couples counseling and condoms. HIV uninfected partners were tested for HIV at least quarterly and HIV-infected partners received HIV care and referral for ART per national guidelines. Upon diagnosis of incident HIV infection in the previously HIV-uninfected partner, a blood sample was collected from both partners to measure CD4+ T-cells and perform viral linkage. The estimated date of infection (EDI) of the incident case was calculated based on testing history. EDI was unknown for suspected transmitting partners.

Results

From 2006–2011, 4,705 HIV-discordant couples were enrolled in this cohort, and 443 cases of incident HIV infection were documented. Virus linkage analysis was performed in 374 transmission pairs, and 273 (73%) transmissions were linked genetically. CD4 counts in the transmitting partner were measured a median of 56 days after EDI (mean:90.5, min:10, max:396). The median CD4 count was 339 cells/μl (mean:386.4, min:15, max:1,434), and the proportion of partners with a CD4+ T-cell count above 500/μl was 25% (95% CI:21, 31).

Conclusions

In our cohort of discordant couples, 73% of HIV transmissions occurred within the relationship, and the transmitter CD4+ T cell count shortly after the transmission event was frequently higher than the WHO 2013 ART-initiation guidelines.     

Engineering the Pichia pastoris methanol oxidation pathway for improved NADH regeneration during whole-cell biotransformation

Industrial biocatalytic reduction processes require the efficient regeneration of reduced cofactors for the asymmetric reduction of prochiral compounds to chiral intermediates which are needed for the production of fine chemicals and drugs. Here, we present a new engineering strategy for improved NADH regeneration based on the Pichia pastoris methanol oxidation pathway. Studying the kinetic properties of alcohol oxidase (AOX), formaldehyde dehydrogenase (FLD) and formate dehydrogenase (FDH) and using the derived kinetic data for subsequent kinetic simulations of NADH formation rates led to the identification of FLD activity to constitute the main bottleneck for efficient NADH recycling via the methanol dissimilation pathway. The simulation results were confirmed constructing a recombinant P. pastoris strain overexpressing P. pastoris FLD and the highly active NADH-dependent butanediol dehydrogenase from S. cerevisiae. Employing the engineered strain, significantly improved butanediol production rates were achieved in whole-cell biotransformations.