Whole Blood Holding Time Prior to Plasma Processing Alters microRNA Expression Profile

MicroRNAs (miRNAs) can exhibit aberrant expression under different physiological and pathological conditions. Therefore, differentially expressed circulating miRNAs have been a focus of biomarker discovery research. However, the use of circulating miRNAs comes with challenges which may hinder the reliability for their clinical application. These include varied sample collection protocols, storage times/conditions, sample processing and analysis methods. This study focused on examining the effect of whole blood holding time on the stability of plasma miRNA expression profiles. Whole blood samples were collected from healthy pregnant women and were held at 4°C for 30 min, 2 h, 6 h or 24 h prior to processing for plasma isolation. Plasma RNA was extracted and the expression of 179 miRNAs were analyzed. Unsupervised principal component analysis demonstrated that whole blood holding time was a major source of variation in miRNA expression profiles with 53 of 179 miRNAs showing significant changes in expression. Levels of specific miRNAs previously reported to be associated with pregnancy-associated complications such as hsa-miR-150-5p, hsa-miR-191-5p, and hsa-miR-29a-3p, as well as commonly used endogenous miRNA controls, hsa-miR-16-5p, hsa-miR-25-3p, and hsa-miR-223-3p were significantly altered with increase in blood holding time. Current protocols for plasma-based miRNA profiling for diagnostics describe major differences in whole blood holding periods ranging from immediately after collection to 26 h after. Our results demonstrate holding time can have dramatic effects on analytical reliability and reproducibility. This highlights the importance of standardization of blood holding time prior to processing for plasma in order to minimize introduction of non-biological variance in miRNA profiles.

Cryptosporidium and Giardia in Livestock in Tigray, Northern Ethiopia and Associated Risk Factors for Infection: A Cross-Sectional Study

The occurrence and species/genotypes of Cryptosporidium and Giardia duodenalis infecting young livestock in selected districts of Tigray, Ethiopia were investigated, along with risks associated with infection. A total of 757 faecal samples were collected from calves, lambs, and goat kids from four rural districts in Tigray, and also from calves in periurban Mekelle, Tigray's main city, and analysed for Cryptosporidium oocysts and Giardia cysts. Farmers answered questionnaires regarding potential risk factors at sample collection. Immunofluorescent antibody staining was used for parasite detection, and PCR at selected genes and sequencing of positive samples was used for molecular characterisation. The occurrence of Cryptosporidium infection was 10, 9, and 4% in calves, lambs, and goat kids, respectively; equivalent figures for Giardia infection were 39, 32, and 21%. Molecular characterisation of Cryptosporidium isolates revealed C. ubiquitum, subtype XIIa in all three host species; C. ryanae in calves and goat kids; C. andersoni and C. bovis were identified only in calves, and C. xiaoi was identified in lambs. For Giardia, Assemblage E predominated in all host species, but among calf isolates we also identified a few potentially zoonotic genotypes (assemblages A (AI) and Assemblage B). Periparturient care was shown to be a particularly relevant risk factor for infection, and infections were less likely to occur under extensive management systems. Our major findings were widespread occurrence of both parasites in livestock, and the apparent lack of the most common zoonotic species. Our results are discussed in relation to other relevant studies. As our study was conducted in Tigray, further investigation in different settings in Ethiopia could provide relevant information on transmission and zoonotic potential. In addition, given the dependency on healthy animals for the livelihoods of the population of Tigray, investigation of the effect of these common parasites on livestock productivity is important.

Clinical research nurses perspective on recruitment challenges and lessons learnt from a large multi-site observational study

Background Recruitment of large numbers of study participants within a designated time frame for multi-site clinical research studies is a significant challenge faced by researchers. If a study does not manage to recruit targeted number of participants, it could have a significant impact on the statistical significance of the research. Purpose This paper highlights the challenges of recruitment for a large multi-site UK-based tuberculosis observational study ‘PREDICT’. Methods It uses a case study analysis from the research nurses perspective, and descriptive information retrieved from non-recruitment log forms to understand reasons for potential recruits not participating. Results Some of the main challenges to recruitment included patients not attending their clinic appointments, time required to obtain site-specific permissions and courier timings for blood sample collection. This paper also outlines key reasons for potential recruits who did not participate. Some of the common barriers to participation for non-recruited participants were work and family commitments, additional blood tests and language barriers. Conclusion Successful strategies which were implemented to overcome some of the challenges during the study are presented. This paper, therefore, aims to present the challenges faced, lessons learnt and successful strategies implemented to inform the planning of similar longitudinal studies of this scale in future.

Comparison of oral metabolome profiles of stimulated saliva, unstimulated saliva, and mouth-rinsed water

Saliva includes a substantial amount of biological information, which has enabled us to understand the relationship between oral metabolites and various oral and systemic disorders. However, collecting saliva using a controlled protocol is time-consuming, making saliva an unsuitable analyte in large cohort studies. Mouth-rinsed water (MW), the water used to rinse the mouth, can be collected easily in less time with less difference between subjects than saliva and could be used as an alternative in oral metabolome analyses. In this study, we investigated the potential of MW collection as an efficient alternative to saliva sample collection for oral metabolome profiling. MW, stimulated saliva, and unstimulated saliva were collected from 10 systemically healthy participants. The samples were subjected to metabolome analysis using capillary electrophoresis time-of-flight mass spectrometry, and the types and amounts of metabolites in the samples were compared. Qualitatively, MW contained the same metabolites as unstimulated and stimulated saliva. While the quantity of the metabolites did not drastically change between the sampling methods, all three reflected individual differences, and the features of MW were the same as those of the unstimulated saliva. Overall, these results suggest that MW may be an appropriate alternative to saliva in oral metabolome profile analysis.

PENGARUH KEPEMILIKAN PUBLIK, UKURAN PERUSAHAAN, UMUR LISTING, JUMLAH DEWAN KOMISARIS INDEPENDEN DAN PROFITABILITAS TERHADAP KETEPATAN WAKTU CORPORATE INTERNET REPORTING PADA PERUSAHAAN SEKTOR KEUANGAN YANG TERDAFTAR DI BURSA EFEK INDONESIA

This research aims to determine and analyze the effect of public ownership, company size, age of listings, number of independent commissioners, and profitability on the timeliness of corporate internet reporting in financial sector companies listed on the Indonesia Stock Exchange. The sample used in this study is financial sector companies listed on the Indonesia Stock Exchange with the sample collection technique used purposive sampling with the amount of final sample to 50 companies that meet the criteria. The data used are secondary data from the official page on the Indonesia Stock Exchange. The analytical tool used in this study is logistic regression analysis using Microsoft Excel and SPSS 16. The result of this study indicate that partially the variable of public ownership, age of listings, number of independent commissioners, and profitability has no effect on the timeliness of corporate internet reporting. While the company size have a significant effect on the timeliness of corporate internet reporting. And the simultaneously that the variable of public ownership, company size, age of listings, number of independent commissioners, and profitability have a significant effect on the timeliness of corporate internet reporting. Keywords : Public Ownership, Company Size, Age of Listings, Number of Independent Commissioners, and Profitability, Timeliness of Corporate Internet Reporting.

Rapid response petrology for the opening eruptive phase of the 2021 Cumbre Vieja eruption, La Palma, Canary Islands

How and why magmatic systems reactivate and evolve is a critical question for monitoring and hazard mitigation efforts during initial response and ongoing volcanic crisis management. Here we report the first integrated petrological results and interpretation provided to monitoring authorities during the ongoing eruption of Cumbre Vieja, La Palma, Canary Islands, Spain. The first eruptive products comprised simultaneous Strombolian fountain-fed lava flows and tephra fall from near-continuous eruption plumes. From combined field, petrographic and geochemical analyses conducted in the 10 days following sample collection, we infer low percentage mantle melts with a variably equilibrated multimineralic crystal-cargo and compositional fractionation by winnowing during eruptive processes. Hence ‘rapid response’ petrology can untangle complex magmatic and volcanic processes for this eruption, which combined with further study and methodological improvement can increasingly assist in active decision making.

Effects of Extra-Long-Acting Recombinant Bovine FSH (bscrFSH) on Cattle Superovulation

Over the last few years, several commercial FSH products have been developed for cattle superovulation (SOV) purposes in Multiple Ovulation and Embryo Transfer (MOET) programs. The SOV response is highly variable among individuals and remains one of the main limiting factors in obtaining a profitable number of transferable embryos. In this study, follicle stimulating hormone (FSH) from different origins was included in two SOV protocols, (a) FSH from purified pig pituitary extract (NIH-FSH-p; two doses/day, 12 h apart, four consecutive days); and (b) extra-long-acting bovine recombinant FSH (bscrFSH; a single dose/day, four consecutive days), to test the effects of bscrFSH on the ovarian response, hormone profile levels, in vivo embryo production and the pluripotency gene expression of the obtained embryos. A total of 68 healthy primiparous red Angus cows (Bos taurus) were randomly distributed into two experimental groups (n = 34 each). Blood sample collection for progesterone (P4) and cortisol (C) level determination was performed together with ultrasonographic assessment for ovarian size, follicles (FL) and corpora lutea (CL) quantification in each SOV protocol (Day 0, 4, 8, and 15). Moreover, FSH profiles were monitorised throughout both protocols (Day 0, 4, 5, 6, 7, 8, 9, 10, and 15). In vivo embryo quantity and quality (total structures, morulae, blastocysts, viable, degenerated and blocked embryos) were recorded in each SOV protocol. Finally, embryo quality in both protocols was assessed by the analysis of the expression level of crucial genes for early embryo development (OCT4, IFNt, CDX2, BCL2, and BAX). P4 and cortisol concentration peaks in both SOV protocols were obtained on Day 15 and Day 8, respectively, which were statistically different compared to the other time-points (p < 0.05). Ovarian dimensions increased from Day 0 to Day 15 irrespective of the SOV protocol considered (p < 0.05). Significant changes in CL number were observed over time till Day 15 irrespective of the SOV protocol applied (p < 0.05), being non- significantly different between SOV protocols within each time-point (p > 0.05). The number of CL was higher on Day 15 in the bscrFSH group compared to the NIH-FSH-p group (p < 0.05). The number of embryonic structures recovered was higher in the bscrFSH group (p = 0.025), probably as a result of a tendency towards a greater number of follicles developed compared to the NIH-FSH-p group. IFNt and BAX were overexpressed in embryos from the bscrFSH group (p < 0.05), with a fold change of 16 and 1.3, respectively. However, no statistical differences were detected regarding the OCT4, CDX2, BCL2, and BCL2/BAX expression ratio (p > 0.05). In conclusion, including bscrFSH in SOV protocols could be an important alternative by reducing the number of applications and offering an improved ovarian response together with better embryo quality and superior performance in embryo production compared to NIH-FSH-p SOV protocols.

Frequency of Genital Tuberculosis in Patients Undergoing Diagnostic Laparoscopy for Infertility

OBJECTIVES: To determine the frequency of genital tuberculosis in patients undergoing diagnostic laparoscopy for infertility. METHODOLOGY: It is a descriptive (cross-sectional) prospective study. The study was performed within the duration of six months i.e., February 12th, 2020 to August 12th, 2020 at the Department of Obstetrics and Gynecology, Khyber Teaching Hospital, Peshawar. A total of 196 patients were observed by using 47.1% proportion of genital tuberculosis in infertility, 95% confidence interval with margin of error 7%, using software of WHO for sample size calculation. Further, sample collection was performed using non-probability (consecutive) sampling techniques. RESULTS: The mean age of the patients in our study was 30 years (SD+3.92). Fifty seven percent of cases were having primary infertility and 43% of patients were having secondary infertility. The incidence of genital tuberculosis undergoing infertility was 45%. CONCLUSION: Our study concludes that the incidence of genital TB in infertility in our setup was 45%.

Diversity of Mosquitoes Collected from the Southern Areas of Khyber Pakhtunkhwa Pakistan

OBJECTIVES: The objective of the study was to assess the diversity of mosquitoes in various towns of the southern belt of KPK. METHODOLOGY: This was a descriptive study that was conducted in numerous towns of Western belt of Khyber Pakhtunkhwa. The study areas were Darra Adam Khel, District Kohat, District Karak, District Banu, District D.I. Khan and newly merged districts Mir Ali and Miranshah. From each study site, the samples were collected randomly. The sample collection was done through survey and area visits whereas; the identification process was done in a parasitology laboratory of Hayatabad-Peshawar. Sampling was done from June 2016 to May 2017. RESULTS: A total of 2150 adult mosquitoes were gathered and collected from 42 different locations of the southern belt of KPK and were identified. Based on their identification, 5 genera of the mosquitoes were recognized which were Culex, Anopheles, Psorophora, Aedes and Uranotenia. Culex was found to be most dominant in all the visited areas with a percentage of 12.65 in Miranshah followed by 11.81 in Mirali, 7.16 in Karak, 6.88 in Darra Adam Khel, 6.69 in D.I.Khan, 6.41 in Kohat and 5.11 in Bannu respectively. The results of our findings also revealed the presence of Culex genera in all the habitats and remained the dominant genera among the others followed by Anopheles and Psorophora. Aedes was found in the habitat of plants and grasses etc. whereas Uranotenia was found in marsh/swampy areas as well as in plants/grasses habitat only. CONCLUSION: The outcomes reveal that a climate shifting and extensive urbanization process is enforcing the diversity of mosquitos’ fauna in the southern belt of KPK.

An integrated lab-on-a-chip device for RNA extraction, amplification and CRISPR-Cas12a-assisted detection for COVID-19 screening in resource-limited settings

In response to the ongoing COVID-19 pandemic and disparities of vaccination coverage in low-and middle-income countries, it is vital to adopt a widespread testing and screening programme, combined with contact tracing, to monitor and effectively control the infection dispersion in areas where medical resources are limited. This work presents a lab-on-a-chip platform, namely IFAST-CRISPR, as an affordable, rapid and high-precision molecular diagnostic means for SARS-CoV-2 detection. The herein proposed sample-to-answer platform integrates RNA extraction, amplification and CRISPR-Cas-based detection with lateral flow readout in one device. The microscale dimensions of the device containing immiscible liquids, coupled with the use of silica paramagnetic beads and GuHCl, streamline sample preparation, including RNA concentration, extraction and purification, in 15 min with minimal hands-on steps. By combining RT-LAMP with CRISPR-Cas12 assays targeting the nucleoprotein (N) gene, visual identification of ≥ 470 copies mL-1 genomic SARS-CoV-2 samples was achieved in 45 min, with no cross-reactivity towards HCoV-OC43 nor H1N1. On-chip assays showed the ability to isolate and detect SARS-CoV-2 from 1,000 genome copies mL-1 of replication-deficient viral particles in 1 h. This simple, affordable and integrated platform demonstrated a visual, faster, and yet specificity and sensitivity-comparable alternative to the costly gold-standard RT-PCR assay, requiring only a simple heating source. Further investigations on multiplexing and direct interfacing of the accessible Swan-brand cigarette filter for saliva sample collection could provide a complete work flow for COVID-19 diagnostics from saliva samples suitable for low-resource settings.