Multidrug resistance among uropathogenic clonal group A E. Coli isolates from Pakistani women with uncomplicated urinary tract infections.

OBJECTIVE: Multi-drug resistance (MDR) has notably increased in community acquired uropathogens causing urinary tract infections (UTIs), predominantly Escherichia coli. Uropathogenic E. coli causes 80% of uncomplicated community acquired UTIs, particularly in pre-menopausal women. Considering this high prevalence and the potential to spread antimicrobial resistant genes, the current study was conducted to investigate the presence of clinically important strains of E. coli in Pakistani women having uncomplicated cystitis and pyelonephritis. Women belonging to low-income groups were exclusively included in the study. Seventy-four isolates from urine samples were processed, phylotyped, and screened for the presence of two Single Nucleotide Polymorphisms (SNPs) particularly associated with a clinically important clonal group A of E. coli (CgA) followed by antibiotic susceptibility testing and genome sequence analysis. RESULTS: Phylogroup B2 was most prevalent in patients and 44% of isolates were positive for the presence of CgA specific SNPs in Fumarate hydratase and DNA gyrase subunit B genes. Antibiotic susceptibility testing showed widespread resistance to trimethoprim-sulfamethoxazole and extended-spectrum beta-lactamase production. The infection analysis revealed the phylogroup B2 to be more pathogenic as compared to the other groups. The genome sequence of E. coli strain U17 revealed genes encoding virulence, multidrug resistance, and host colonization mechanisms. CONCLUSIONS: Our research findings not only validate the significant occurrence of multidrug-resistant clonal group A E. coli (CgA) in premenopausal Pakistani women suffering from cystitis and pyelonephritis but also reveal the presence of genes associated withvirulence, and drug efflux pumps. The detection of highly pathogenic, antimicrobial-resistant phylogroup B2 and CgA E. coli strains is likely to help in understanding the epidemiology of the pathogen and may ultimately help to reduce the impact of these strains on human health. Furthermore, the findings of this study will particularly help to reduce the prevalence of uncomplicated UTIs and the cost associated with their treatment in women belonging to low-income groups.

Prevalence of Type III Secretion System (T3SS) and Biofilm Development in Genetically Heterogeneous Clinical Isolates of Pseudomonas aeruginosa from Nigeria.

Pseudomonas aeruginosa infection in seriously ill patients is a major concern due to its ability to form biofilm and secrete effector toxins. There is little information on the prevalence of T3SS effector toxins and biofilm production in clinical isolates of P. aeruginosa from Nigeria. The goal of this study is to evaluate the prevalence of T3SS toxins and biofilm production among isolates from selected tertiary hospitals in Nigeria. This study examined 430 clinical isolates from our previous work, comprising 181 MDR (multidrug-resistant) and 249 non-MDR isolates. Biofilm production and type III secretion toxins were determined using colorimetric microtiter plate assay and polymerase chain reaction, respectively. Carbapenem-resistant isolates were typed using REP-PCR and BOX-PCR. Biofilm production was detected in 386/430 (89.8%) of the isolates. Out of 386 biofilm producers, 167 (43.3%) were multidrug-resistant isolates. PCR identified four T3SS virulence types among 430 isolates, including 78 (18.1%) exoU+/exoS- isolates, 343 (79.8%) exoU-/exoS + isolates, 5 (1.2%) exoU+/exoS + isolates, and 4 (0.9%) exoU-/exoS- isolates. Both REP- and BOX-PCR consist of eight clusters. On the REP-PCR dendrogram, ExoU+/ExoS- isolates majorly occupied cluster IV. Clusters IV, VII, and VIII consist of isolates from wounds on BOX-PCR dendrogram. There was a positive association between strong biofilm production and multidrug resistance in our P. aeruginosa isolates. This study identified multidrug-resistant, biofilm-producing P. aeruginosa strains that secrete cytotoxic effectors which are significant virulence factors in P. aeruginosa. This poses a severe risk to our healthcare system and highlights the importance of continuous surveillance to prevent infectious disease outbreaks.

A prospective study on linking diarrheagenic E. coli with stunted childhood growth in relation to gut microbiome.

Stunted growth is an emerging global challenge affecting children under the age of 5 years in low- and middle-income countries. Despite such a high global prevalence of stunting, the mechanism of pathogenesis and the role of associated gut microbiota is poorly understood. The present study was designed to investigate the association of pathogenic strains of E. coli with the residential gut microbiota of stunted growth children. A total of 64 stool sample were collected from children aged ≤ 5 years, and were processed for isolation and molecular characterization of diarrheagenic E. coli. Selected stool samples (n = 39 including three normal controls) were then analysed for microbial community profiling using 16S ribosomal RNA (rRNA) gene sequencing. Furthermore, associations between changes in the microbiota in the presence of different E. coli strains was explored. Pathotyping of the isolated E. coli (n = 64) has shown that 39.68% belonged to one of the five pathotypes of E. coli whilst the remaining ones were non-typeable. Amongst the different pathotypes, EPEC was found to be the most prevalent (52%; n = 13), followed by EAEC (20%; n = 5), EIEC (12%; n = 3), EHEC (8%; n = 2) and ETEC 2 (8%; n = 2). Phylogrouping analysis has shown that majority of the strains belonged to B2 (28.12%). Microbial diversity is shown to be significant and varied when the samples are organized under the recovered phylogroups. Moreover, based on predictive metabolism, the colonization of these strains were found to be significantly associated with energy utilization pathways such as Denovoprine-2 and glyoxylate-by. Differential analysis has shown that Escherichia-Shigella and Enterococcus were altered for the children with stunted growth.

Diverse Repertoire and Relationship of Exopolysaccharide Genes in Cold-Adapted Acinetobacter sp. CUI-P1 Revealed by Comparative Genome Analysis.

This study focused on the exploration of microbial communities inhabiting extreme cold environments, such as the Passu and Pisan glaciers of Pakistan, and their potential utilization in industrial applications. Among the 25 initially screened strains, five were found to be suitable candidates for exopolysaccharide (EPS) production, with strain CUI-P1 displaying the highest yield of 7230.5 mg/L compared to the other four strains. The purified EPS from CUI-P1 was tested for its ability to protect probiotic bacteria and E. coli expressing green fluorescence protein (HriGFP) against extreme cold temperatures, and it exhibited excellent cryoprotectant and emulsification activity, highlighting its potential use in the biotechnological industry. Furthermore, the genome of Acinetobacter sp., CUI-P1 comprised 199 contigs, with a genome size of 10,493,143bp and a G + C content of 42%, and showed 98.197% nucleotide identity to the type genome of Acinetobacter baumannii ATCC 17978. These findings offer promising avenues for the application of EPS as a cryoprotectant, an essential tool in modern biotechnology.

Gaseous and soil OCPs and PCBs along the Indus River, Pakistan: spatial patterns and air-soil gradients.

This study presents first-hand information on the occurrence of persistent organic pollutants (POPs) in the ambient air and surface soil along the Indus flood-plain, Pakistan. The sampling campaign was conducted at 15 site locations during 2014-15, along the Indus River (approximately 1300 km). Composite surface soil samples (N = 15) and passive air samples (N = 15) were collected for the estimation of gaseous POPs as well as air-soil exchange to evaluate the POP emission and distribution or dispersion patterns, source tracking, and contribution of the local and regional sources towards POP accumulation in the Indus River system. Among the studied POPs, levels of DDTs and PCBs were noticeably higher in ambient air (50-560 and 10-1100 pg m-3) and in soil (0.20-350 and 1.40-20 ng g-1), respectively. Regarding the spatial patterns, higher DDT concentrations (ng g-1) were detected in the air and soil samples collected from the wet mountain zone (WMZ) (p < 0.05), followed by the alluvial riverine zone (ARZ), low-lying mountain zone (LLZ), and frozen mountain zone (FMZ). The PCB data did not exhibit significant differences (p > 0.05) for the air samples, while PCB concentrations were significantly higher (p < 0.05) in soil from the LLZ, which may be associated with rapid urbanization and industrial activities in this area. The air-soil exchange of DDTs and PCBs showed net volatilization at most of the studied sites except for a few samples from the FMZ and WMZ. Results of this study about air-soil exchange gradients indicate the long range regional atmospheric transport (LRAT) of POPs to the colder areas (FMZ) of Pakistan, where these act as a secondary source of POPs in these areas.

Colibactin possessing E. coli isolates in association with colorectal cancer and their genetic diversity among Pakistani population.

Colorectal cancer (CRC) is the third most prevalent cause of tumorigenesis and several pathogenic bacteria have been correlated with aggressive cases of cancer i.e., genotoxin (colibactin) producing Escherichia coli (E. coli). This study was designed to investigate the genetic diversity of clb+clb+ E. coli strains and their association with CRC. Pathogenic E. coli isolates from colorectal biopsies were characterized based on phylotypes, antibiotic resistance pattern, and (Enterobacterial Repetitive Intergenic Consensus Sequence-based Polymerase Chain Reaction) ERIC-PCR. Furthermore, isolates were screened for the presence of the Pks (polyketide synthase) Island specifically targeting colibactin genes A and Q. The selective clb+clb+ isolates were subjected to cytotoxicity assay using Human embryonic kidney (HEK) cell lines. We revealed that 43.47% of the cancer-associated E. coli isolates were from phylogroup B2 comparatively more pathogenic than rest while in the case of healthy controls no isolate was found from B2. Moreover, 90% were found positive for colibactin and pks (polyketide synthase) island, while none of the healthy controls were found positive for colibactin genes. All healthy and cancer-associated isolates were tested against 15 antibiotic agents, we observed that cancer-associated isolates showed a wide range of resistance from 96% against Nalidixic acid to 48% against Doxycycline. Moreover, E. coli isolates were further genotyped using ERIC-PCR, and selected clb+clb+ E. coli isolates were subjected to cytotoxicity assay. We recorded the significant cytotoxic activity of clb+clb+ E. coli phylogroup B2 isolates that might have contributed towards the progression of CRC or dysbiosis of healthy gut microbiota protecting against CRC pathogenesis. Our results revealed a significant p<0.023 association of dietary habits and hygiene p<0.001with CRC. This is the first study to report the prevalence of E. coli phylogroups and the role of colibactin most virulent phylogroup B2 among Pakistani individuals from low socioeconomic setup.

Pathogenic microbe detection in placental tissues supports placental pathobiome association with preterm birth risk in Pakistani women: A brief snapshot.

Preterm birth (PTB) poses great risk to neonatal health in Pakistan with few tertiary health care facilities. Role of intrauterine microbiome in maintaining healthy pregnancy has been highlighted. However, there is ongoing debate whether a true placental microbiome exist. We analyzed placental and vaginal microbiome through V3-V4 16srRNA sequencing and observed increased abundance of proteobacteria, with concomitant decline in the firmicutes population in preterm vagina. Simplistic placental microflora included many environmental microbes with PTB placenta carrying pathogenic microbes like ureaplasma and mycoplasma species. We observed contribution of environmental, vaginal and skin contamination in term versus pathobiome signatures in preterm placenta.

Freely dissolved organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs) along the Indus River Pakistan: spatial pattern and risk assessment.

Freely dissolved OCPs and PCBs were measured by using polyethylene passive samplers at 15 sites during 2014 throughout the stretch of the Indus River to investigate the spatial pattern and risk assess. Levels (pg/L) of dissolved ∑OCPs and ∑PCBs ranged from 34 to 1600 and from 3 to 230. Among the detected OCPs, dissolved DDTs (p,p'-DDE, followed by p,p'-DDT) predominated with levels of 0.48 to 220 pg/L. The order of occurrence for other studied OCPs was as follows: HCB, endosulfans, chlordanes, and HCHs. Spatially, dissolved (pg/L) ∑OCPs varied (p < 0.05) as the following: surface water of the alluvial riverine zone (ARZ) showed the highest levels (114) followed by the frozen mountain zone (FMZ) (52.9), low-lying zone (LLZ) (28.73), and wet mountain zone (WMZ) (14.43), respectively. However, our zone-wise PCB data did not exhibit significant differences (p > 0.05). Principal component analysis/multilinear regression results showed pesticide usage in the crop/orchard fields and health sector, electric and electronic materials, and widespread industrial activities as the main source of OCPs and PCBs along the Indus River. Our results showed that OCPs and PCBs contaminated water intake, playing an important role towards the considerable cancer/non-cancer risk (HI and CR values) along the Indus River Flood-Plain.

Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform.

Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and RT-LAMP assays were performed on extracted RNA of seven wastewater samples from COVID-19 hotspots. RT­LAMP assay was also performed on wastewater samples without RNA extraction. Current detection of SARS-CoV-2 is mainly by RT-qPCR of ORF (ORF1ab) and N genes so we targeted both to find the best target gene for SARS-CoV-2 detection. We also performed RT-LAMP with/without RNA extraction inside microfluidic device to target both genes. Positivity rates of RT-qPCR and RT-LAMP performed on extracted RNA were 100.0% (7/7) and 85.7% (6/7), respectively. RT-qPCR results revealed that all 7 wastewater samples were positive for N gene (Ct range 37-39), and negative for ORF1ab, suggesting that N gene could be the best target gene for SARS-CoV-2 detection. RT-LAMP of N and ORF (ORF1a) genes performed on wastewater samples without RNA extraction indicated that all 7 samples remains pink (negative). The color remains pink in all microchannels except microchannels which subjected to RT-LAMP for targeting N region after RNA extraction (yellow color) in 6 out of 7 samples. This study shows that SARS-CoV-2 was successfully detected from wastewater samples using RT-LAMP in microfluidic chips. This study brings the novelty involving the use of wastewater samples for detection of SARS-CoV-2 without previous virus concentration and with/without RNA extraction.

In Silico and In Vitro Analysis of Helicobacter pullorum Type Six Secretory Protein Hcp and Its Role in Bacterial Invasion and Pathogenesis.

Helicobacter pullorum is a human zoonotic pathogen transmitted through poultry where it is associated with vibrionic hepatitis and colitis. Hemolysin co-regulated protein (Hcp) is an important structural as well as effector protein of type six secretory system; however, its role in H. pullorum invasion and pathogenesis has not been elucidated. In this study, we predicted the Helicobacter pullorum Hcp (HpuHcp) structure and identified Campylobacter jejuni Hcp (CjHcp) as its nearest homologue. Analysis of the predicted structure shows several common bacterial Hcp motifs like Protein kinase C phosphorylation site, Casein kinase II phosphorylation site, N-myristoylation site, cAMP-and cCGMP-dependent protein kinase phosphorylation site, N-glycosylation site. The presence of unique microbodies C-terminal targeting signal domain was present in HpuHcp which was seen for the first time in CjHcp. This could indicate that Hcp is a structural protein as well as a secretory protein. Moreover, the presence of a deamidase domain, similar to the tecA of Burkholderia cenocepacia an opportunistic pathogen, may help in bacterial internalization as it depolymerises the membranous actin by deamidation of the host cell Rho GTPases cdc42 and Rac1, which was supported by increased invasion of hepatocytes by Hcp-positive isolates.

Antimicrobial Resistance Pattern in E. coli Isolated from Placental Tissues of Pregnant Women in Low-Socioeconomic Setting of Pakistan.

Infections during pregnancy can culminate in adverse pregnancy outcomes, including preterm birth (PTB). Pakistan is among the top ten nations with high PTB-associated neonatal mortality rates, where access to prenatal as well as neonatal care is only afforded by the privileged few. Societal stigma further discourages women seeking healthcare for minor infections. Microbial pathogens associated with genitourinary infections can lead to gestational complications culminating in earlier onset of labor. In this study, association of Escherichia coli (E. coli) with PTB in Pakistani women of low-socioeconomic status is examined. 57 paired vaginal swabs and placenta samples from mothers with full term and preterm deliveries were collected and processed for isolation and molecular characterization of extraintestinal pathogenic E. coli (ExPEC). ExPEC isolated from vaginal swabs and placenta showed phylotype B2 being most prevalent (Vagina n = 3 (9), 33%) (Placenta n = 4 (12), 33%) in preterm cases followed by phylotype B1 (Vagina n = 2 (9), 22%) (Placenta n = 3 (12), 25%) and untypeable strains. Antibiotic susceptibility profiling showed a large percentage of resistant isolates to multiple antibiotics, including carbapenem and included extended-spectrum beta-lactamase (ESBL) producers. Our study is the first to report different phylotypes of E. coli from placental tissues in preterm deliveries which may be a cause for concern for maternal and neonatal health. ExPEC from vaginal swabs and placental of females delivering preterm shows the pathogenic phylotype B2 dominance with a large percentage of isolates resistant to multiple antibiotics, including carbapenem and included ESBL producers. The placental isolates may indicate ascending infection from vagina or urinary tract which may lead to preterm birth.

Lactobacillus species signature in association with term and preterm births from low-income group of Pakistan.

AIM/PURPOSE OF STUDY: It is estimated that around 15 million babies are born prematurely every year and approximately one million children die each year due to complications of preterm birth (PTB). Many survivors face a lifetime of disability, including learning disabilities and visual and hearing problems. The current study aimed to characterize Lactobacillus species isolated from vaginal swabs and determine their antibiotic susceptibility. MATERIAL AND METHODS: 40 term and 20 preterm samples were processed by culturing on MRS agar and initial identification was carried out using sugar fermentation reactions and 16S rRNA PCR. Moreover, Lactobacillus species from preterm and term cases using paired samples, i.e. vaginal swabs and placenta tissues from 8 preterm delivering mothers were further recruited for metagenomics study to possibly detect uncultured Lactobacillus species known to cause PTB. RESULTS: 40% samples from preterm delivering mothers lack any Lactobacillus species whereas in contrast vaginal microflora of all term delivering mothers carry one or more species of Lactobacillus. L. crispatus (46% in term group and 25% preterm group) was found to be the most abundant group followed by L. jensenii (25% in both groups) and L. gasseri (19% in term group and 10% in preterm group). The antimicrobial susceptibility profile suggests that preterm isolates were least resistant to linezolid (20% resistance rate) and trimethoprim/sulfamethoxazole (22% resistant rate) compared to term isolates (>60% for each drug group). Furthermore, the metagenomics data for paired samples (8 VS and 8 PT) from extreme PTB suggested that Lactobacillus Iners was the main difference between term and preterm deliveries. Moreover, overall lack of lactobacillus species or presence of rogue Lactobacillus species such as L. iners and L. vaginilis is associated with PTB. ERIC-PCR analysis using Lactobacillus crispatus revealed that all the pre-term samples are closely related and fall in same cluster while all the term samples fall in different cluster. CONCLUSION: The study not only provides a baseline data of distinct signatures of associated lactobacillus species with the PTB which may be further transmitted to new born infants but also developing further therapeutic interventions to better manage the PTB.

Evaluation of Vibrio cholerae O1 El Tor variants circulating in Sindh Pakistan.

OBJECTIVE: To investigate the existence of genetically diverse vibrio cholerae variant strains in a rural Sindh district, and to find out the phylogenetic relationship of indigenous vibrio cholerae strains. METHODS: The cross-sectional study was conducted from April 2014 to May 2016 in Khairpur, Pakistan, and comprised stool samples/rectal swabs collected from the main and city branches of the Khairpur Medical College Teaching Hospital, and the Pir Abdul Qadir Shah Jeelani Institute of Medical Sciences, Gambat. The samples were identified using standard microbiological, biochemical, serological techniques and polymerase chain reaction targeting the ompW gene. Whole genome sequencing and bioinformatics tool MUMmer 3.2.3 was used to compare indigenous and contemporary vibrio cholerae strains circulating in the province of Sindh. Neighbour-joining tree method was used to construct the phylogenic tree. RESULTS: Of the 360 samples, 76(21.11%) were found positive for vibrio cholera strains. The species-specific ompW gene was amplified at the correct size of 588bp. The isolates belonged to serogroup Inaba, O1, biotype El Tor. Unique sequences with same genomic coordinates showed that test strains were not similar to the reference sequence. Conserved genome sequences showed that 12 Out of 16 (75%) of the test strains were similar to each Other except the 3 strains isolated from Khairpur and 1 from Karachi. Multiple sequence alignment of the regions translated into protein showed that 13 out of 16 (81.25%) test strains were similar except 2 strains from Khairpur and 1 From Karachi. The phylogenetic tree showed that all isolated strains descended from the same ancestor along with the reference strain. CONCLUSIONS: Vibrio cholerae O1 El Tor variant existed in Khairpur.

Incidence and Molecular Characterization of Carbapenemase Genes in Association with Multidrug-Resistant Clinical Isolates of Pseudomonas aeruginosa from Tertiary Healthcare Facilities in Southwest Nigeria.

Pseudomonas aeruginosa, resistant to multiple antibacterial agents including carbapenems, is of great global public health concern. There is limited data available regarding incidence of Metallo-Beta Lactamase producing P. aeruginosa, their molecular basis of resistance in particular carbapenem resistance and any genetic relatedness among circulating clinical isolates in Southwest Nigeria. Four hundred and thirty P. aeruginosa isolates were collected from seven tertiary care hospitals (predominantly from wound, ear, and urinary tract infections) and verified by PCR targeting oprI and oprL. Antibiotic susceptibility using 16 selected antibiotics and MBL screening was performed. The integrons (class 1, 2 and 3) and carbapenemase genes- blaGES, blaNMC-A, blaBIC-1, blaSME, blaIMP, blaVIM, blaSPM, blaNDM, blaAIM, blaDIM, blaSIM, blaGIM, blaOXA-48, blaOXA-58 were detected by PCR and were sequenced. Quantitative real-time polymerase chain reaction was used to quantify expression levels of eight efflux pump genes, ampC cephalosporinase and outer membrane porin, oprD. The isolates were genotyped using Enterobacterial Repetitive Intergenic Consensus sequence Polymerase Chain Reaction (ERIC-PCR). Four hundred and thirty P. aeruginosa isolates were subjected to antibiotic susceptibility testing, revealing that 109 (25.4%) isolates were multidrug-resistant, 47 (10.9%) were extensively drug-resistant and 25 (5.8%) were pandrug-resistant. MBL was seen in 17.0% (73/430) isolates. MBL-encoding genes; blaVIM-5 and blaNDM-1 were detected in 86.3% (63/73) isolates, with blaVIM-5 and blaNDM-1 in 35.6% (26/73) and 38.4% (28/73), respectively, whereas co-occurrence of blaVIM-5 and blaNDM-1 was found in 12.3% (9/73). Forty-one (56.2%) carbapenem-resistant P. aeruginosa strains carried class 1 integrons, while co-occurrence of class 1 and 2 integrons was seen in 12.3%. qPCR results indicated that MexXY-OprM was highly expressed pump in 58.9%, ampC upregulated in 26.0%, while oprD porin was downregulated in 65.8% isolates. ERIC-PCR results suggest that carbapenem-resistant strains exhibit genetic heterogeneity. The high incidence of MBL-encoding genes and integrons in diversified clinical P. aeruginosa from southwestern Nigeria is of great concern. The co-occurrence of blaVIM-5 and blaNDM-1 as well as resistance in general manifesting a gradient based on genotypic variation suggests that there is a strong need for efficient surveillance programs and antibiotic stewardship.

Prevalence and Associated Risk Factor of COVID-19 and Impacts of Meteorological and Social Variables on Its Propagation in Punjab, Pakistan.

The current study identifies the spatial distribution of COVID-19 cases and its association with meteorological and social variables in Punjab (densely populated province of Pakistan). To identify the COVID-19 propagation, the weekly growth, recovery, and deaths rate have also been calculated. The geographic information system (GIS) has used to determine COVID-19 impacts on gender (male/female), age groups, and causalities over an affected population (km-2) for the period of 11th March to 12th August, 2020 in each district of province. Our results show that 43 peak days (where daily positive cases were above 900) have been observed in Punjab during 27th May to 8th July, 2020. The high population density districts, i.e., Lahore and Islamabad, have been affected (five persons per square kilometers) due to COVID-19, whereas the maximum death tolls (> 50 persons per millions) have also been observed in these urban districts. The meteorological variables (temperature, humidity, heat index, and ultraviolet index) show negative significant relationship to basic reproduction number (R0), whereas daily COVID-19 cases are positively correlated to aerosols concentration at 95% confidence level. The government intervention (stringency index) shows a positive impact to reduce the COVID-19 cases over the province. Keeping in view the COVID-19 behavior and climatology of the region, it has been identified that the COVID-19 cases may likely to increase during the dry period (high concentration of aerosols) i.e., October-December, 2020 and post-spring season (April to June), 2021 in urban areas of Pakistan. This study provides an overview on districts vulnerability that would help the policy makers, health agencies to plan their activities to reduce the COVID-19 impacts.

Design and Prototyping of Genetically Encoded Arsenic Biosensors Based on Transcriptional Regulator AfArsR.

Genetically encoded biosensors based on engineered fluorescent proteins (FPs) are essential tools for monitoring the dynamics of specific ions and molecules in biological systems. Arsenic ion in the +3 oxidation state (As3+) is highly toxic to cells due to its ability to bind to protein thiol groups, leading to inhibition of protein function, disruption of protein-protein interactions, and eventually to cell death. A genetically encoded biosensor for the detection of As3+ could potentially facilitate the investigation of such toxicity both in vitro and in vivo. Here, we designed and developed two prototype genetically encoded arsenic biosensors (GEARs), based on a bacterial As3+ responsive transcriptional factor AfArsR from Acidithiobacillus ferrooxidans. We constructed FRET-based GEAR biosensors by insertion of AfArsR between FP acceptor/donor FRET pairs. We further designed and engineered single FP-based GEAR biosensors by insertion of AfArsR into GFP. These constructs represent prototypes for a new family of biosensors based on the ArsR transcriptional factor scaffold. Further improvements of the GEAR biosensor family could lead to variants with suitable performance for detection of As3+ in various biological and environmental systems.

Vaginal microbiome: normalcy vs dysbiosis.

It has been long understood that the vaginal microflora is crucial in maintaining a normal physiological environment for the host and its involvement is deemed indispensable for reproductive success. A global concept of normalcy vs. dysbiosis of vaginal microbiome is debatable as women of different races have a unique vaginal microflora with regional variations. Vaginal microflora is a dynamic microenvironment affected by gestational status, menstrual cycle, sexual activity, age, and contraceptive use. Normal vaginal flora is dominated by lactobacilli especially in women of European descent vs. African American women. These microbes confer the host vagina protection from potentially pathogenic microbes that may lead to urinary tract infections and sexually transmitted diseases. Changes in the vaginal microbiota including reduced lactobacilli abundance and increased facultative and anaerobic organism populations result in bacterial vaginosis, that predisposes the host to several conditions like low birth weight and increased risk of contracting bacterial infections. On the other hand, the vaginal microbiome is also reshaped during pregnancy, with less microbial diversity with a dominance of Lactobacillus species. However, an altered vaginal microbiota with low lactobacilli abundance especially during pregnancy may result in induction of excessive inflammation and pre-term labor. Since the vaginal microbiome plays an important role during embryo implantation, it is not surprising that bacterial vaginosis is more common in infertile women and associated with reduced rates of conception. Probiotic has great success in treating bacterial vaginosis and restoring the normal microbiome in recent. This report, reviewed the relationships between the vaginal microbiome and women's reproductive health.

COVID-19 Crisis Creates Opportunity towards Global Monitoring & Surveillance.

The spectrum of emerging new diseases as well as re-emerging old diseases is broadening as infectious agents evolve, adapt, and spread at enormous speeds in response to changing ecosystems. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a recent phenomenon and may take a while to understand its transmission routes from less traveled territories, ranging from fomite exposure routes to wastewater transmission. The critical challenge is how to negotiate with such catastrophic pandemics in high-income countries (HICs ~20% of the global population) and low-and middle-income countries (LMICs ~ 80% of the global population) with a total global population size of approximately eight billion, where practical mass testing and tracing is only a remote possibility, particularly in low-and middle-income countries (LMICs). Keeping in mind the population distribution disparities of high-income countries (HICs) and LMICs and urbanisation trends over recent years, traditional wastewater-based surveillance such as that used to combat polio may help in addressing this challenge. The COVID-19 era differs from any previous pandemics or global health challenges in the sense that there is a great deal of curiosity within the global community to find out everything about this virus, ranging from diagnostics, potential vaccines/therapeutics, and possible routes of transmission. In this regard, the fact that the gut is the common niche for both poliovirus and SARS-CoV-2, and due to the shedding of the virus through faecal material into sewerage systems, the need for long-term wastewater surveillance and developing early warning systems for better preparedness at local and global levels is increasingly apparent. This paper aims to provide an insight into the ongoing COVID-19 crisis, how it can be managed, and what measures are required to deal with a current global international public health concern. Additionally, it shed light on the importance of using wastewater surveillance strategy as an early warning practical tool suitable for massive passive screening, as well as the urgent need for microfluidic technology as a rapid and cost-effective approach tracking SARS-CoV-2 in wastewater.

Apoptosis induced by SARS-CoV-2: can we target it?

Some viruses are known to be associated with increased apoptosis. Apoptotic cell death triggered by these viruses has a complex role in host antiviral immunity, and might facilitate the viral clearance or act as a mechanism for virus-induced tissue injury and disease progression. The induction of apoptosis is a hallmark of SARS-CoV-2 infection. Accumulating evidence suggests that there is a direct relationship between apoptosis rate and COVID-19 pathogenicity/severity. Targeting virus-induced apoptosis could be a promising strategy in the treatment of SARS-CoV-2 virus infection.