A possible dose-response equation: Viral load after plasma infusion in COVID-19 patients and anti-SARS-CoV-2 antibody titers in convalescent plasma.

BACKGROUND: Clinical trials of convalescent plasma therapy for coronavirus disease 2019 (COVID-19) are extensive, but the relationship between antibody titers, infused volume of plasma and virus clearance in patients remains unknown. This study proposed a possible estimating equation for clinical use of high antibody titer convalescent plasma. METHODS: A total of 38 patients were recruited in the Guanggu District Maternal and Child Health Hospital of Hubei Province from March 1 to 30, 2020. COVID-19 convalescent plasma was collected and high-titer (≥1:640) anti-S-RBD units used. The SARS-CoV-2 nucleic acid viral load was measured 24 h before and 72 h after convalescent plasma infusion. RESULTS: Convalescent plasma therapy was associated with reduced viral load in patients with moderate and severe severity. The viral negative rate at 72 h was 65.8%. The disappearance of viral nucleic acid in study patients was positively correlated with infuscate antibody titer and volume (r = 0.3375, p = 0.04). A possible estimation equation was as follows: Log10 (Reduction in viral load) = 0.18 + 0.001 × (Log2 S-RBD antibody titer × Plasma infusion volume) (r = 0.424, p = 0.009). In a single case, the viral nucleic acid persisted 14 days after the fourth plasma infusion. CONCLUSIONS: This study proposes a potential dose-response equation that adds a convenient way to estimate the dose of convalescent plasma product. It is beneficial to facilitate the rational allocation of plasma with high antibody titers and provide an individualised use strategy for convalescent plasma therapy.

Analysis of viral load in different specimen types and serum antibody levels of COVID-19 patients.

BACKGROUND: COVID-19 has caused a global pandemic and the death toll is increasing. However, there is no definitive information regarding the type of clinical specimens that is the best for SARS-CoV-2 detection, the antibody levels in patients with different duration of disease, and the relationship between antibody level and viral load. METHODS: Nasopharyngeal swabs, anal swabs, saliva, blood, and urine specimens were collected from patients with a course of disease ranging from 7 to 69 days. Viral load in different specimen types was measured using droplet digital PCR (ddPCR). Meanwhile, anti-nucleocapsid protein (anti-N) IgM and IgG antibodies and anti-spike protein receptor-binding domain (anti-S-RBD) IgG antibody in all serum samples were tested using ELISA. RESULTS: The positive detection rate in nasopharyngeal swab was the highest (54.05%), followed by anal swab (24.32%), and the positive detection rate in saliva, blood, and urine was 16.22%, 10.81%, and 5.41%, respectively. However, some patients with negative nasopharyngeal swabs had other specimens tested positive. There was no significant correlation between antibody level and days after symptoms onset or viral load. CONCLUSIONS: Other specimens could be positive in patients with negative nasopharyngeal swabs, suggesting that for patients in the recovery period, specimens other than nasopharyngeal swabs should also be tested to avoid false negative results, and anal swabs are recommended. The antibody level had no correlation with days after symptoms onset or the viral load of nasopharyngeal swabs, suggesting that the antibody level may also be affected by other factors.

Effect of Convalescent Plasma Therapy on Time to Clinical Improvement in Patients With Severe and Life-threatening COVID-19: A Randomized Clinical Trial.

Importance: Convalescent plasma is a potential therapeutic option for patients with coronavirus disease 2019 (COVID-19), but further data from randomized clinical trials are needed. Objective: To evaluate the efficacy and adverse effects of convalescent plasma therapy for patients with COVID-19. Design, Setting, and Participants: Open-label, multicenter, randomized clinical trial performed in 7 medical centers in Wuhan, China, from February 14, 2020, to April 1, 2020, with final follow-up April 28, 2020. The trial included 103 participants with laboratory-confirmed COVID-19 that was severe (respiratory distress and/or hypoxemia) or life-threatening (shock, organ failure, or requiring mechanical ventilation). The trial was terminated early after 103 of a planned 200 patients were enrolled. Intervention: Convalescent plasma in addition to standard treatment (n = 52) vs standard treatment alone (control) (n = 51), stratified by disease severity. Main Outcomes and Measures: Primary outcome was time to clinical improvement within 28 days, defined as patient discharged alive or reduction of 2 points on a 6-point disease severity scale (ranging from 1 [discharge] to 6 [death]). Secondary outcomes included 28-day mortality, time to discharge, and the rate of viral polymerase chain reaction (PCR) results turned from positive at baseline to negative at up to 72 hours. Results: Of 103 patients who were randomized (median age, 70 years; 60 [58.3%] male), 101 (98.1%) completed the trial. Clinical improvement occurred within 28 days in 51.9% (27/52) of the convalescent plasma group vs 43.1% (22/51) in the control group (difference, 8.8% [95% CI, -10.4% to 28.0%]; hazard ratio [HR], 1.40 [95% CI, 0.79-2.49]; P = .26). Among those with severe disease, the primary outcome occurred in 91.3% (21/23) of the convalescent plasma group vs 68.2% (15/22) of the control group (HR, 2.15 [95% CI, 1.07-4.32]; P = .03); among those with life-threatening disease the primary outcome occurred in 20.7% (6/29) of the convalescent plasma group vs 24.1% (7/29) of the control group (HR, 0.88 [95% CI, 0.30-2.63]; P = .83) (P for interaction = .17). There was no significant difference in 28-day mortality (15.7% vs 24.0%; OR, 0.59 [95% CI, 0.22-1.59]; P = .30) or time from randomization to discharge (51.0% vs 36.0% discharged by day 28; HR, 1.61 [95% CI, 0.88-2.95]; P = .12). Convalescent plasma treatment was associated with a negative conversion rate of viral PCR at 72 hours in 87.2% of the convalescent plasma group vs 37.5% of the control group (OR, 11.39 [95% CI, 3.91-33.18]; P < .001). Two patients in the convalescent plasma group experienced adverse events within hours after transfusion that improved with supportive care. Conclusion and Relevance: Among patients with severe or life-threatening COVID-19, convalescent plasma therapy added to standard treatment, compared with standard treatment alone, did not result in a statistically significant improvement in time to clinical improvement within 28 days. Interpretation is limited by early termination of the trial, which may have been underpowered to detect a clinically important difference. Trial Registration: Chinese Clinical Trial Registry: ChiCTR2000029757.

[Extraction of polysaccharide from Penthorum chinense].

OBJECTIVE: To study the extraction technology of polysaccharide from Penthorum chinense. METHODS: Single factor test and orthogonal experiment design methods L9 (3(3)) were applied to analyze the influence of solid-liquid rate, extraction temperature and time on the extraction percent of polysaccharide from Penthorum chinense. RESULTS: The temperature significantly affected the extraction percent of the soluble polysaccharide, and the optimum conditions for the extraction technology were as follows: solid-liquid rate at 1:15, temperature at 95 degrees C, time for 3h. Under this condition, the extraction percent of polysaccharide from Penthorum chinense was 1.12%, RSD = 1.21%. CONCLUSION: The research can provide a theoretical foundation for further development and utilization of polysaccharide from Penthorum chinense.

Optimizing the shoot proliferation protocol of Penthorum chinense by axillary buds.

Four parameters, three hormones and sucrose, at seven concentrations, were designed for shoot proliferation of Penthorum chinense by uniform design. The obtained data were used for building two quadratic polynomial equations by partial least square to determine optimum concentrations of four factors. Experiments for verification confirmed that no significant difference existed between the predicted and the validated values in shoot number and length based on all inoculated explants.

Analysis on the association between sperm DNA fragmentation index and conventional semen parameters, blood microelements and seminal plasma ROS in male patients with infertility.

We conducted this study to analyze the sperm DNA fragmentation index, conventional semen parameters, blood microelements and seminal plasma reactive oxygen species (ROS) in patients with male infertility to determine the association between each of the above male physiological parameters and DNA fragmentation index and infertility. Eighty cases of infertile males and 20 cases of normal males with children were divided into the infertility and control groups, respectively. Sperm DNA fragmentation index, conventional semen parameters, serum microelement content and seminal plasma ROS levels were detected, and the existing correlation between sperm DNA fragmentation index and the various physiological parameters were studied. The sperm DNA fragmentation index had no correlation with conventional sperm parameters. Our results demonstrated that zinc, lead and magnesium ions in the serum microelements were correlated with sperm DNA fragmentation (p<0.05). Upon an increase in zinc and lead serum concentration, there was a subsequent increase in sperm DNA fragmentation (p=0.008). Furthermore, when magnesium ion increased, it also caused an increase in sperm DNA fragmentation (p<0.05). The seminal plasma ROS of infertile males was higher than that of males with children (p<0.05). Our results suggest that sperm DNA fragmentation index is closely associated with the infertility rate and microelements of serum and seminal plasma ROS can impact the formation of sperm DNA fragmentation. Therefore, the sperm DNA fragmentation index can serve as an important parameter to assess male infertility.

[Application of carrying capacity assessment in spatial allocation of regional population: a case of Changzhou City of East China].

Based on the spatial differences in urban ecosystem carrying capacity, and by using geographic information system (GIS) spatial analysis technology and system dynamics (SD) model, this paper ascertained the moderate population size in urban area and its subareas, and presented an alternative method for quantitatively allocate the population. This method was applied in Changzhou, a city with more than three million populations in East China. A SD model consisted of population, economy, land and environment modules for the city was established and scenario analysis was made. The results showed that under the requirements of satisfying both economic development and environmental protection, the total population in the city in 2050 would reach nearly 4 million, and according to the differences in the urban ecosystem carrying capacity, a spatial allocation pattern of 2.4 million populations in main city and 150000-400000 populations in each of 5 new towns was proposed. This study could provide scientific support for the spatial allocation of population and economy in country land development and management.

[Comparative study of menstruation in 240 healthy women at various altitudes].

To assess the relation of menstrual physiologic functions and altitudes, the authors made a comparative study of 240 healthy women with four diagnostic methods on traditional Chinese medicine and obtained their menstrual changes, picture of tongue and type of pulse in Xining (2260m) and Yushu Zhaduo (4300m). The results suggested that the menarche which occurred before 15 years old (P less than 0.005), the menstruation cycle of 16-25 days (P less than 0.05), and moderate quantity menstruation (P less than 0.005) are found more in the Xining group than in the Zhaduo group; the menstrual phase in the Xining group lasted 4-7 days and 1-3 in the Zhaduo group. The incidence of abdominal pain, cold, and lumbago increased significantly in the Zhaduo group and the symptoms were serious. The age of menostasia after 46 was 100% in the Xining group, whereas 27.3% of the Zhaduo group had menostasia at the age of 36-45. The colour of tongue varied from darker red to pink with increasing altitudes (P less than 0.005). Meanwhile, cases of rapid pulse increased (P less than 0.005). It is concluded that the physiology of menstruation for healthy women has obviously changed under high altitude hypoxic environment. The mechanism of pathology could be explained by dysfunction of Zang and Fu, deficiency of Qi, stagnation of blood and Qi stasis due to cold. Treatment and prevention of menstrual disorders of women at high altitudes could be carried out by invigorating Qi, regulating blood, promoting the flow of Qi, by warming the channel and regulating Zang and Fu, etc.

Identification and characterization of Escherichia coli DNA helicase II mutants that exhibit increased unwinding efficiency.

Using a combination of both ethyl methanesulfonate and site-directed mutagenesis, we have identified a region in DNA helicase II (UvrD) from Escherichia coli that is required for biological function but lies outside of any of the seven conserved motifs (T. C. Hodgman, Nature 333:22-23, 1988) associated with the superfamily of proteins of which it is a member. Located between amino acids 403 and 409, alterations in the amino acid sequence DDAAFER lead to both temperature-sensitive and dominant uvrD mutations. The uvrD300 (A406T) and uvrD301 (A406V) alleles produce UV sensitivity at 44 degrees C but do not affect sensitivity to methyl methanesulfonate (MMS). In contrast, the uvrD303 mutation (D403AD404A) causes increased sensitivity to both UV and MMS and is dominant to uvrD+ when present at six to eight copies per cell. Several of the alleles demonstrated a strong antimutator phenotype. In addition, conjugal recombination is reduced 10-fold in uvrD303 strains. Of all of the amino acid substitutions tested, only an alanine-to-serine change at position 406 (uvrD302) was neutral. To determine the biochemical basis for the observed phenotypes, we overexpressed and purified the UvrD303 protein from a uvrD delta294 deletion background and characterized its enzymatic activities. The highly unusual UvrD303 protein exhibits a higher specific activity for ATP hydrolysis than the wild-type control, while its Km for ATP binding remains unchanged. More importantly, the UvrD303 protein unwinds partial duplex DNA up to 10 times more efficiently than wild-type UvrD. The DNA binding affinities of the two proteins appear comparable. Based on these results, we propose that the region located between amino acids 403 and 409 serves to regulate the unwinding activity of DNA helicase II to provide the proper balance between speed and overall effectiveness in the various DNA repair systems in which the protein participates.

Induction of hyporesponsiveness to intact foreign protein via retroviral-mediated gene expression: the IgG scaffold is important for induction and maintenance of immune hyporesponsiveness.

IgG molecules can be highly tolerogenic carriers for associated antigens. Previously, we reported that recipients of bone marrow or lipopolysaccharide-stimulated B-cell blasts, both of which were retrovirally gene-transferred with an immunodominant peptide in-frame with the variable region of a murine IgG heavy chain, were rendered profoundly unresponsive to that epitope. To further investigate whether tolerance to larger molecules can be achieved via this approach and whether the IgG scaffold is important for induction and maintenance of immunological tolerance, we engineered two retroviral constructs encoding the cI lambda repressor (MBAE-1-102 and MBAE-1-102-IgG) for gene transfer. Our results show that recipients of bone marrow or peripheral B cells, transduced with the MBAE-1-102-IgG recombinant, are hyporesponsive to p1-102. In addition, the self-IgG scaffold enhanced the induction and maintenance of such an immune hyporesponsiveness. Thus, our studies demonstrate that in vivo-expressed IgG heavy chain fusion protein can be processed and presented on the appropriate MHC class II, resulting in hyporesponsiveness to that antigen and offering an additional therapeutic approach to autoimmune diseases.

Conserved motifs II to VI of DNA helicase II from Escherichia coli are all required for biological activity.

There are seven conserved motifs (IA, IB, and II to VI) in DNA helicase II of Escherichia coli that have high homology among a large family of proteins involved in DNA metabolism. To address the functional importance of motifs II to VI, we employed site-directed mutagenesis to replace the charged amino acid residues in each motif with alanines. Cells carrying these mutant alleles exhibited higher UV and methyl methanesulfonate sensitivity, increased rates of spontaneous mutagenesis, and elevated levels of homologous recombination, indicating defects in both the excision repair and mismatch repair pathways. In addition, we also changed the highly conserved tyrosine(600) in motif VI to phenylalanine (uvrD309, Y600F). This mutant displayed a moderate increase in UV sensitivity but a decrease in spontaneous mutation rate, suggesting that DNA helicase II may have different functions in the two DNA repair pathways. Furthermore, a mutation in domain IV (uvrD307, R284A) significantly reduced the viability of some E. coli K-12 strains at 30 degrees C but not at 37 degrees C. The implications of these observations are discussed.