Silicon and iron nanoparticles protect rice against lead (Pb) stress by improving oxidative tolerance and minimizing Pb uptake.

Lead (Pb) is toxic to the development and growth of rice plants. Nanoparticles (NPs) have been considered one of the efficient remediation techniques to mitigate Pb stress in plants. Therefore, a study was carried out to examine the underlying mechanism of iron (Fe) and silicon (Si) nanoparticle-induced Pb toxicity alleviation in rice seedlings. Si-NPs (2.5 mM) and Fe-NPs (25 mg L-1) were applied alone and in combination to rice plants grown without (control; no Pb stress) and with (100 µM) Pb concentration. Our results revealed that Pb toxicity severely affected all rice growth-related traits, such as inhibited root fresh weight (42%), shoot length (24%), and chlorophyll b contents (26%). Moreover, a substantial amount of Pb was translocated to the above-ground parts of plants, which caused a disturbance in the antioxidative enzyme activities. However, the synergetic use of Fe- and Si-NPs reduced the Pb contents in the upper part of plants by 27%. It reduced the lethal impact of Pb on roots and shoots growth parameters by increasing shoot length (40%), shoot fresh weight (48%), and roots fresh weight (31%). Both Si and Fe-NPs synergistic application significantly elevated superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and glutathione (GSH) concentrations by 114%, 186%, 135%, and 151%, respectively, compared to plants subjected to Pb stress alone. The toxicity of Pb resulted in several cellular abnormalities and altered the expression levels of metal transporters and antioxidant genes. We conclude that the synergistic application of Si and Fe-NPs can be deemed favorable, environmentally promising, and cost-effective for reducing Pb deadliness in rice crops and reclaiming Pb-polluted soils.

Modulation of metal transporters, oxidative stress and cell abnormalities by synergistic application of silicon and titanium oxide nanoparticles: A strategy for cadmium tolerance in rice.

Heavy metals, especially cadmium (Cd), cause severe toxicity symptoms in crop plants. Applying nanoparticles (NPs) as nano-fertilizers is a novel approach to mitigating plants' Cd stress. However, knowledge about the combinational use of silicon (Si) and titanium dioxide (TiO2) NPs to mitigate Cd stress, especially in rice, must be highlighted. TiO2-NPs (15 mg L-1) and Si-NPs (2.5 mM) were applied alone and in combination to rice plants grown without (control; no Cd stress) and with (100 µM) Cd concentration. Results revealed that compared to the control plants, root length, shoot length, shoot fresh weight, and root dry weight of rice seedlings were significantly decreased by 25.43%, 26.64%, 34.13%, and 29.87% under Cd exposure. However, the synergistic effect of TiO2- and Si-NPs increased rice plants' shoot length, root length, root dry weight, and shoot fresh weight by 24.62%, 29.81%, 36.16%, and 33.07%, respectively, under the Cd-toxicity. The concentration of malondialdehyde (MDA) and H2O2 were amplified due to Cd stress, which leads to damage to the subcellular structures. Si and TiO2-NPs co-application improved the anti-oxidative enzymatic activities (catalase, peroxidase, superoxide dismutase) and an elevated concentration of non-enzymatic glutathione in Cd-exposed rice. The Cd accumulation was condensed by 21.37% and 19.7% in the shoot, while 48.31% and 45.65% in root tissues under Si-NPs + Cd and TiO2-NPs + Cd treatments compared to Cd-alone treated seedlings, respectively. The expression patterns of metal transporters, such as OsNramp1 and OsHMA3, were the highest when rice plants were cultivated under Cd stress and significantly reduced when treated with sole and combined Si- and TiO2-NPs treatments. In conclusion, combining Si- and TiO2-NPs significantly improved the antioxidant enzymatic activities, chlorophyll contents, biomass production, and reduced cellular damage. Despite limitations, our findings guide future research, addressing risks, optimizing concentrations, and assessing long-term effects that can balance agricultural progress with environmental sustainability.

Identifying and elucidating the resistance of Staphylococcus aureus isolated from hospital environment to conventional disinfectants.

BACKGROUND: Staphylococcus aureus is a nosocomial pathogen, detection and elucidation of its resistance mechanisms to conventional disinfectants may aid in limiting its spread on environmental surfaces in health care settings. In the current study, disinfectant susceptibility of S. aureus strains isolated from the hospital environment as well as possible associations between the presence of disinfectant-resistance genes and reduced susceptibility to disinfectants was investigated. METHODS: A total of 245 samples were collected from the hospital environmental surfaces. The minimum inhibitory (MIC) and bactericidal concentrations (MBC) of disinfectants against S. aureus isolates were determined using the micro-broth dilution method. The qac genes (qacA, qacE, and qacΔE1) were detected by PCR and confirmed by sanger sequencing. RESULTS: A total of 47 S. aureus strains were isolated, with more than 85% of them showing methicillin resistance. The qacA, qacE, and qac∆E1 genes were found in 23.4%, 29.7%, and 4.2% isolates respectively. All the isolates with qac genes had higher MIC and MBC values to selected disinfectants. CONCLUSIONS: Significant methicillin resistant S. aureus (MRSA) contamination in the hospital environment was detected. Furthermore, higher qac gene frequencies were found in MRSA isolates that also correlated with higher MIC/MBC values to different disinfectants. The study proposes that hospitals should develop policies to determine disinfectant MICs against the common environmental isolates to contain the spread of resistant strains.

Effectiveness of international virtual training on biorisk management in the context of COVID-19.

Introduction: The COVID-19 pandemic has resulted in enormous increases in laboratory activities to keep pace with diagnostic testing and research efforts. However, traditional training, technical assistance, and capacity-building approaches were disrupted by the travel and movement restrictions put in place to control the spread of the disease. To address the needs of laboratorians and managers to conduct laboratory activities safely and securely during the pandemic, a highly interactive virtual training (IVT) workshop on biorisk management during COVID-19 was conducted through active learning strategies that connected speakers with participants. The objective of the training was to increase the basic knowledge and standards of biosafety and biosecurity practices, risk assessment, and control measures with reference specifically to the context of the COVID-19 pandemic and apply a rigorous evaluation methodology to assess the effectiveness of the IVT. The training covered a broad range of topics and encompassed national to international guidelines. Methods: Participants were selected through official channels at the national level, focusing on institutions within Pakistan. The sessions included lectures from international experts in biorisk management concepts, and incorporated poll questions as well as pre- and post-tests and feedback on the speakers' knowledge and presentation skills, to increase interactivity. The pre- and post-test comprised similar multiple-choice questions and provided to every participant to ascertain the impact of the training on awareness and knowledge of biorisk management topics and concepts, and results were compared using paired t-tests. For feedback on the speakers, participants were asked to submit their ratings measured on a five-point Likert scale. The reliability of the Likert scale was estimated using Cronbach's alpha. Analyses were performed using Microsoft Excel and SPSS version 23. Results: In total, 52 individuals from different laboratories across Pakistan and Pakistani students from abroad (China) as well participated in at least one session of the IVT. The participants' pre- and post-test scores showed a significant increase in knowledge and awareness (p < 0.001). The obtained Cronbach's alpha score was >0.8, indicating high reliability of the generated feedback on the IVT approach and speakers. Conclusion: The IVT on biosafety and biosecurity in the context of the COVID-19 pandemic proved beneficial for laboratory professionals and could be a useful model to continue in the future for raising awareness and knowledge.

A modified hand washing method for resource limited settings.

The Good Microbiological Practices & Procedures (GMPP) is the most significant risk control measure as per the fourth edition of the WHO laboratory biosafety manual. Among GMPP, one of the best practices is hand washing. WHO and other public health agencies have published several guidance documents on hand washing, that describe closing the tap using a disposable paper towel/tissue paper at the end of hand washing as one of the critical steps. In resource-limited settings, where disposable paper towels cannot be provided at all times, the staff is left with ambiguous instructions on how to close the tap. In this paper, a modified hand washing method is documented that doesn't necessitate the use of disposable paper towels. In this method, both hands and faucets remain in contact with soap for at least 40-60 s. The method was validated by the use of Glo Germ. A survey questionnaire was also designed and conducted for the lab staff (n = 12) of the two laboratories, where this method was implemented, to assess whether this hand washing method brought any improvement in their hand washing practices and implementation. All (100%) of the survey respondents reported that this method of hand washing is more applicable and implementable than the WHO-recommended hand washing technique. Eighty three percentage reported that this modified method of hand washing raised their hand washing compliance. The authors suggest that this hand washing method can be used in resource-limited laboratory settings as an effective GMPP to ensure infection control.

Spatial variations in the biochemical potential of okra [Abelmoschus esculentus L. (Moench)] leaf and fruit under field conditions.

Okra (Abelmoschus esculentus L. (Moench) plays a significant role in humans nutrition because its fresh leaves, stems, flowers, pods and seeds, are used for multiple purposes. The present study attempted to determine the spatial variations in biochemical attributes of osmoprotectants and the oxidative defense system of okra plants. Samples of soil and okra plants (leaves and fruits) were collected from three different locations: Faisalabad region-1 (7 JB-I), Faisalabad region-2 (7 JB-II) and Pindi Bhattian. Chlorophyll contents, glycine betaine (GB), ascorbic acid (AsA), total phenolics, hydrogen peroxide (H2O2), proline, and malondialdehyde (MDA) contents were analyzed in the leaves and fruits of okra plants. Soil analyses showed that pH, electrical conductivity (EC), phosphorus (P), potassium (K), iron (Fe), and saturation of soil were higher in Faisalabad region 2, while organic matter, sand, Zn, and Cu were higher in the Pindi Bhattian region. The results from okra leaves showed that Pindi Bhattian had higher chlorophyll a, GB and H2O2 contents, while Faisalabad region 1 had a higher ratio of chlorophyll a/b compared to the other regions. However, Faisalabad regions 2 and 1 had higher leaf phenolic contents, Faisalabad regions 1 and 2 showed higher leaf proline contents, and Faisalabad region 2 possessed higher AsA and MDA contents. Analyses of okra fruits showed that Faisalabad region 2 had higher chlorophyll a and total chlorophyll contents, while Faisalabad region 1 had higher chlorophyll b contents. Faisalabad region 2 and Pindi Bhattian had higher ratios of chlorophyll a/b, and Faisalabad region 1 showed higher phenolic, AsA, H2O2, and MDA contents of okra fruit, whereas the Faisalabad regions exhibited higher proline and GB contents than the Pindi Bhattian region. Overall, okra leaves and fruits showed better responses in the Faisalabad regions, and these results may be used to screen for okra cultivars with better tolerance under different environmental conditions.

Assessment of biosafety implementation in clinical diagnostic laboratories in Pakistan during the COVID-19 pandemic.

Laboratory diagnostic capacity is crucial for an optimal national response to a public health emergency such as the COVID-19 pandemic. Preventing laboratory-acquired infections and the loss of critical human resources, especially during a public health emergency, requires laboratories to have a good biorisk management system in place. In this study, we aimed to evaluate laboratory biosafety and biosecurity in Pakistan during the COVID-19 pandemic. In this cross-sectional study, a self-rated anonymous questionnaire was distributed to laboratory professionals (LPs) working in clinical diagnostic laboratories, including laboratories performing polymerase chain reaction (PCR)-based COVID-19 diagnostic testing in Punjab, Sindh, Khyber Pakhtunkhwa, and Gilgit-Baltistan provinces as well as Islamabad during March 2020 to April 2020. The questionnaire assessed knowledge and perceptions of LPs, resource availability, and commitment by top management in these laboratories. In total, 58.6% of LPs performing COVID-19 testing reported that their laboratory did not conduct a biorisk assessment before starting COVID-19 testing in their facility. Only 31% of LPs were aware that COVID-19 testing could be performed at a biosafety level 2 laboratory, as per the World Health Organization interim biosafety guidelines. A sufficiently high percentage of LPs did not feel confident in their ability to handle COVID-19 samples (32.8%), spills (43.1%), or other accidents (32.8%). These findings demonstrate the need for effective biosafety program implementation, proper training, and establishing competency assessment methods. These findings also suggested that identifying and addressing gaps in existing biorisk management systems through sustainable interventions and preparing LPs for surge capacity is crucial to better address public health emergencies.

A Mentorship Program to Develop Biorisk Manuals for Laboratories in Pakistan.

Introduction: Institutions implementing a biorisk management system need to establish comprehensive guidance to support the implementation of biosafety and biosecurity practices. A biorisk manual describes how a biorisk management system will be implemented in an organization and includes topics such as facility-specific policies and procedures to safely and securely handle, store, and dispose of biological agents and toxins in adherence with international guidance. Methods: To promote the adoption of biosafety and biosecurity in Pakistan, the Pakistan Biological Safety Association and Health Security Partners developed a biorisk manual writing project in 2019 in partnership with experts from the BioRisk Association of the Philippines 2015, Inc. This project helped 13 researchers and laboratory professionals in Pakistan develop biorisk manuals for their institutions. The project comprised two phases: an in-person group training on how to develop a laboratory biorisk manual, and 10 months of additional remote mentoring assistance for the development and finalization of the biorisk manual tailored to each institution's specific needs. By the end of the project, 12 of the 13 participants had customized biorisk manuals for their institutions in place. In January 2022, a survey was conducted among the workshop participants to learn how successful they were in implementing the developed manual in their institutions. Results: Participants reported varying degrees of successful implementation. They also suggested that the biosafety and biosecurity associations should engage top management at institutions to strengthen administrative support and provide a sufficient workforce to promote implementation.

A Cross-Sectional Survey to Assess Biorisk Management System in Research and Diagnostic Laboratories in Khyber Pakhtunkhwa, Pakistan.

Financial, cultural, and managerial hurdles have made biosafety and biosecurity measures difficult in resource-constrained countries like Pakistan. Because of increasing awareness of biorisk management, diagnostic and research laboratories have made major advances in biosafety and biosecurity in the recent decade. As a result, identifying and addressing gaps in biorisk management has never been more critical. The purpose of this study was to assess the current situation of personal protective equipment (PPE), biosafety behavior, waste management, biosafety and biosecurity measures, training and safety, and health services in diagnostic and research laboratories across Pakistan's Khyber Pakhtunkhwa (KP) province. We adapted the WHO Laboratory Assessment tool (2012) and CWA 15793 (Biorisk management guidelines) for conducting a cross-sectional survey, which was distributed among various laboratories in KP. The survey included 30 laboratories, including 11 diagnostic and 19 research laboratories. In comparison to diagnostic laboratories, biorisk management practices in research laboratories were better in terms of PPE, biosafety behavior, waste management, biosafety measures, biosecurity measures, trainings, and safety and health services. KP laboratories' biorisk management practices have improved over time, according to our findings. However, we were able to identify inadequacies that would require considerable improvements to the current setups based on the WHO and CWA 15793 recommendations. Organizations can tailor their biosafety measures and training to address identified gaps using the presented KP snapshot.

Awareness and Attitudes of Research Students Toward Dual-use Research of Concern in Pakistan: A Cross-sectional Questionnaire.

Dual-use research poses a significant challenge for scientists in the biomedical field and for global health security in general. As the scientific knowledge and materials required for the development of biological agents become progressively more accessible and inexpensive, there is an increased need to understand and improve the governance of scientific research. Prevention of the misuse of facilities, equipment, agents, and scientific knowledge requires high levels of awareness of the concept of dual-use research, starting with early-career scientists and graduate students. In this study, the attitudes and level of awareness of postgraduate students in Pakistan toward the issues surrounding dual-use research were assessed through a survey containing both quantitative and qualitative questions in 32 universities in 4 provinces, federal area, and Azad Jammu and Kashmir regions of Pakistan; 933 students responded. Most (58.2%) had never heard of dual-use research of concern (DURC), while 18.5% had heard the term but were unsure of its meaning. Irrespective of prior knowledge, a higher percentage of students (68.6%) felt an obligation to report research misuse. Considering the need for DURC training, 94.1% of the respondents agreed that the principal investigator should take the responsibility to train students on DURC at the start of a research project. When experimental results having dual-use potential, 69.1% indicated they would publish with limited protocol, with 43.5% indicating they would publish the limited protocol only if there was a way for scientists to access their data. The survey results revealed limited DURC awareness among researchers across Pakistan. However, the respondents, although not formally educated about DURC, were quite aware of its impact. The information gained in this survey will be valuable in addressing country-specific awareness and training needs.

Non-O1Vibrio cholerae bacteremia in an infant, first case report from Pakistan.

To the best of our knowledge, Non-O1 Vibrio cholerae (NOVC) bacteremia has never been documented in Pakistan. This case report is the first reported case of bacteremia in an infant due to NOVC in Pakistan. A neonate was admitted to a hospital with fever and no history of diarrhea. The isolate was identified biochemically and serologically and was sensitive to all the drugs tested as per CLSI 2014 guidelines.

A cornucopia of screening and diagnostic techniques for human papillomavirus associated cervical carcinomas.

Cervical carcinoma is one of the major consequences of human papillomavirus (HPV) infections. Although HPV infections of cervix do not always progress to cancer, 90% cases of cervical cancer have been found associated with high risk HPV (hrHPV) infection. Usually, HPV infection is asymptomatic; however, this asymptomatic infection can cause abnormal changes in cervix ultimately leading to cancer development. These changes can be detected by the application of screening tests at regular time intervals. For this purpose, morphological, cytological, and DNA based techniques are available. Nevertheless, abnormal screening tests have only the predictive value for precancerous lesions and thus require further evaluation which is usually done by using diagnostic techniques. So far, colposcopy and histological examination alone were considered as the gold standards for cervical cancer diagnosis. Currently, some tests based on expression level of host cell biomarkers are also being used along with histology for diagnostic purpose. Albeit, these tests have significant specificity and sensitivity values but they are unable to suggest a particular viral genotype involved in infection. Diagnostic methods such as PCR, HPV genotyping assays, microarray, and mRNA based assays are useful to predict the genotypes as well as the quantity of viral load in a host cell. Similarly, these diagnostic procedures have high specificity and sensitivity ranges. However, only few of them are practiced commonly, as approval of these tests as routine diagnostic tests requires clinical validation and cost effectiveness.

Matrix-assisted refolding and purification of placenta-derived recombinant human interleukin-6 produced in Escherichia coli.

Biological activity of human interleukin-6 (IL-6) is associated with a vast number of diseases such as rheumatoid arthritis, sepsis, and severe inflammatory diseases. In this study, human IL-6 cDNA was isolated from a cDNA library that was constructed with mRNA derived from human placental tissues. Subsequently, the complete human IL-6 cDNA was cloned and expressed in BL21DE3 cells. The recombinant human IL-6 (rhIL-6) protein was expressed in a form of an insoluble inclusion body. Inclusion bodies were solubilized under denaturing conditions and purified by immobilized metal affinity chromatography with gradual on-column refolding by the gradient elution method (from 6 to 0 M urea). The protein was purified to apparent homogeneity of about 99% with a yield of 50 mg/L. The purity was assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), size exclusion high-performance liquid chromatography, and Western blotting analysis. The bioactivity was assessed by proliferation assay of TF-1 cells in a dose-dependent manner. The present study confirms the expression of the placenta-derived IL-6 gene in a prokaryotic expression system and matrix-assisted on-column refolding and purification of rhIL-6 by immobilized metal affinity chromatography.

Phylogenomic analysis of polyketide synthase genes in actinomycetes: structural analysis of KS domains and modules of polyketide synthases.

Polyketides are complex and diverse secondary metabolites, synthesised by large multifunctional enzymes, Polyketide Synthases (PKS). The phylogenomic analysis of ß-ketosynthase (KS) domains and PKSs within actinomycetes suggests the contribution of point mutations, gene duplications, horizontal gene transfer and homologous recombination in the evolution of PKSs. PKS genealogy suggested the ancestral module structure with KS-AT-ACP domain composition. KS domains showed similar core and highly variable loop regions at the dimer interface, which seems to affect the selectivity of the primer unit. In PKS modules, the linker regions comprise a significant fraction of the module. The reducing domains (ketoreductase and dehydrogenase) protrude out from the central axis of the module and also responsible for extreme variability in the final products. Thus, phylogenomic and structural analysis of PKSs can assist in the artificial reprogramming of PKSs.