BCG activation of trained immunity is associated with induction of cross reactive COVID-19 antibodies in a BCG vaccinated population.

BACKGROUND: Pakistan is endemic to a diverse set of parasitic, mycobacterial and viral diseases. The recognition of BCG Trained Immunity (TI) led us to postulate that the continued presence of BCG-TI may play a protective role, previously reported for both infectious and noninfectious conditions. Most of the previous studies have addressed the issue of BCG-TI in the paediatric populations. This study addressed the key issue of maintenance of BCG-TI in a wider age range (adolescent and adults) to identify the strength and quality of the immune responses. OBJECTIVE: To assess the BCG-induced recall responses in healthy individuals by cytokines secreted from the TI network and its potential role in providing cross-protection against COVID-19 and other viral infections. STUDY DESIGN: In this cross-sectional study, healthy young adults and adolescents (n = 20) were recruited from 16-40 years of age, with no prior history of TB treatment, autoimmune, or chronic inflammatory condition. METHODS: BCG-induced cytokine responses were assessed using prototypic markers for cells of the TI network [macrophages [M1 (TNFα, IFNγ), M2 (IL10)], NK (IL2), Gamma delta (γδ) T (IL17, IL4)] and SARS CoV2 IgG antibodies against RBD using short-term (12 hrs.) cultures assay. RESULTS: Significant differences were observed in the magnitude of recall responses to BCG with macrophage cytokines showing the highest mean levels of TNFα (9148 pg/ml) followed by IL10 (488 pg/ml) and IFNγ (355 pg/ml). The ratio of unstimulated vs.BCG-stimulated cytokines was 132 fold higher for TNFα, 40 fold fo r IL10, and 27 fold for IFNγ. Furthermore, SARS-CoV-2 antibodies were also detected in unstimulated plasma which showed cross reactivity with BCG. CONCLUSION: The presence of cross reactive antibodies to SARS-CoV-2 and the relative ratio of pro- and anti-inflammatory cytokines secreted by activated TI cellular network may play a pivotal role in protection in the early stages of infection as observed during the COVID-19 pandemic in the younger age groups resulting in lower morbidity and mortality.

Predictors of Respiratory Syncytial Virus, Influenza Virus, and Human Metapneumovirus Carriage in Children Under 5 Years With WHO-Defined Fast-Breathing Pneumonia in Pakistan.

BACKGROUND: Pneumonia is a leading cause of morbidity and mortality in children < 5 years. We describe nasopharyngeal carriage of respiratory syncytial virus (RSV), human metapneumovirus (hMPV), and influenza virus among children with fast-breathing pneumonia in Karachi, Pakistan. METHODS: We performed a cross-sectional analysis of nasopharyngeal swabs from children aged 2-59 months with fast-breathing pneumonia, enrolled in the randomized trial of amoxicillin versus placebo for fast-breathing pneumonia (RETAPP) (NCT02372461) from 2014 to 2016. Swabs were collected using WHO standardized methods, processed at the Aga Khan University, Pakistan. Viral detection was performed using LUMINEX xTAG respiratory viral panel assay and logistic regression identified clinical and sociodemographic predictors. FINDINGS: Of the 1000 children tested, 92.2% (n = 922) were positive for viral carriage. RSV, hMPV, and influenza virus were detected in 59 (6.4%), 56 (6.1%), and 58 (6.3%) children and co-infections in three samples (two RSV-hMPV and one influenza-hMPV). RSV carriage was common in infants (56%), we observed a higher occurrence of fever in children with hMPV and influenza virus (80% and 88%, respectively) and fast breathing in RSV (80%) carriage. RSV carriage was positively associated with a history of fast/difficulty breathing (aOR: 1.96, 95% CI 1.02-3.76) and low oxygen saturation (aOR: 2.52, 95% CI 1.32-4.82), hMPV carriage was positively associated with a complete vaccination status (aOR: 2.22, 95% CI 1.23-4.00) and body temperature ≥ 37.5°C (aOR: 2.34, 95% CI 1.35-4.04) whereas influenza viral carriage was associated with body temperature ≥ 37.5°C (aOR: 4.48, 95% CI 2.53-7.93). CONCLUSION: We observed a high nasopharyngeal viral carriage among children with WHO-defined fast-breathing pneumonia in Pakistan. Fever, difficulty in breathing, hypoxia and vaccination status are important clinical predictors for viral nonsevere community-acquired pneumonia.

Shigella Detection and Molecular Serotyping With a Customized TaqMan Array Card in the Enterics for Global Health (EFGH): Shigella Surveillance Study.

Background: Quantitative polymerase chain reaction (qPCR) targeting ipaH has been proven to be highly efficient in detecting Shigella in clinical samples compared to culture-based methods, which underestimate Shigella burden by 2- to 3-fold. qPCR assays have also been developed for Shigella speciation and serotyping, which is critical for both vaccine development and evaluation. Methods: The Enterics for Global Health (EFGH) Shigella surveillance study will utilize a customized real-time PCR-based TaqMan Array Card (TAC) interrogating 82 targets, for the detection and differentiation of Shigella spp, Shigella sonnei, Shigella flexneri serotypes, other diarrhea-associated enteropathogens, and antimicrobial resistance (AMR) genes. Total nucleic acid will be extracted from rectal swabs or stool samples, and assayed on TAC. Quantitative analysis will be performed to determine the likely attribution of Shigella and other particular etiologies of diarrhea using the quantification cycle cutoffs derived from previous studies. The qPCR results will be compared to conventional culture, serotyping, and phenotypic susceptibility approaches in EFGH. Conclusions: TAC enables simultaneous detection of diarrheal etiologies, the principal pathogen subtypes, and AMR genes. The high sensitivity of the assay enables more accurate estimation of Shigella-attributed disease burden, which is critical to informing policy and in the design of future clinical trials.

Environmental surveillance for COVID-19 using SARS-CoV-2 RNA concentration in wastewater - a study in District East, Karachi, Pakistan.

Background: Wastewater-based surveillance is used to track the temporal patterns of the SARS-CoV-2 virus in communities. Viral RNA particle detection in wastewater samples can indicate an outbreak within a catchment area. We describe the feasibility of using a sewage network to monitor SARS-CoV-2 trend and use of genomic sequencing to describe the viral variant abundance in an urban district in Karachi, Pakistan. This was among the first studies from Pakistan to demonstrate the surveillance for SARS-CoV-2 from a semi-formal sewage system. Methods: Four sites draining into the Lyari River in District East, Karachi, were identified and included in the current study. Raw sewage samples were collected early morning twice weekly from each site between June 10, 2021 and January 17, 2022, using Bag Mediated Filtration System (BMFS). Secondary concentration of filtered samples was achieved by ultracentrifugation and skim milk flocculation. SARS-CoV-2 RNA concentrations in the samples were estimated using PCR (Qiagen ProMega kits for N1 & N2 genes). A distributed-lag negative binomial regression model within a hierarchical Bayesian framework was used to describe the relationship between wastewater RNA concentration and COVID-19 cases from the catchment area. Genomic sequencing was performed using Illumina iSeq100. Findings: Among the 151 raw sewage samples included in the study, 123 samples (81.5%) tested positive for N1 or N2 genes. The average SARS-CoV-2 RNA concentrations in the sewage samples at each lag (1-14 days prior) were associated with the cases reported for the respective days, with a peak association observed on lag day 10 (RR: 1.15; 95% Credible Interval: 1.10-1.21). Genomic sequencing showed that the delta variant dominated till September 2022, while the omicron variant was identified in November 2022. Interpretation: Wastewater-based surveillance, together with genomic sequencing provides valuable information for monitoring the community temporal trend of SARS-CoV-2. Funding: PATH, Bill & Melinda Gates Foundation, and Global Innovation Fund.

Building up a genomic surveillance platform for SARS-CoV-2 in the middle of a pandemic: a true North-South collaboration.

Next-generation sequencing technology has revolutionised pathogen surveillance over the last two decades. However, the benefits are not equitably distributed, with developing countries lagging far behind in acquiring the required technology and analytical capacity. Recent declines in the cost associated with sequencing-equipment and running consumables have created an opportunity for broader adoption. During the COVID-19 pandemic, rapid diagnostics development and DNA sequencing revolutionised the ability to diagnose and sequence SARS-CoV-2 rapidly. Socioeconomic inequalities substantially impact the ability to sequence SARS-CoV-2 strains and undermine a developing country's pandemic preparedness. Low- and middle-income countries face additional challenges in establishing, maintaining and expanding genomic surveillance. We present our experience of establishing a genomic surveillance system at the Aga Khan University, Karachi, Pakistan. Despite being at a leading health sciences research institute in the country, we encountered significant challenges. These were related to collecting standardised contextual data for SARS-CoV-2 samples, procuring sequencing reagents and consumables, and challenges with library preparation, sequencing and submission of high-quality SARS-CoV-2 genomes. Several technical roadblocks ensued during the implementation of the genomic surveillance framework, which were resolved in collaboration with our partners. High-quality genome sequences were then deposited on open-access platforms per the best practices. Subsequently, these efforts culminated in deploying Pakistan's first SARS-CoV-2 phyllo surveillance map as a Nextstrain build. Our experience offers lessons for the successful development of Genomic Surveillance Infrastructure in resource-limited settings struck by a pandemic.

Promoting Science Literacy and Awareness across the Globe: the Role of Scientists as Science Ambassadors.

Science literacy has many personal and societal benefits that allows for better informed decision-making. Although the importance of science literacy is recognized globally, there are many challenges associated with its promotion. Scientists are more frequently engaging with nonscientific audiences through public outreach activities and with increasing support from institutions and professional societies. This is especially true regarding microbiologists and other related professionals since the start of the global 2019 coronavirus disease pandemic heightened the need to convey novel and rapidly evolving scientific information to lay audiences. The means by which professionals engage with these audiences affect the efficacy of the relay of scientific information. One method of engagement is the "ambassador approach," which aims to establish dialogue among different groups of people and scientists. In this perspective article, we discuss this approach, highlighting activities for the promotion of science literacy organized by the American Society for Microbiology Ambassador Program and similar programs of other scientific societies. We discuss the benefits and challenges of implementing an ambassador approach, propose potential improvements that could be made to existing programs promoting science literacy, and ultimately advocate for increased implementation of science ambassador programs.

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.

Synthesis, Characterization, Photocatalysis, and Antibacterial Study of WO3, MXene and WO3/MXene Nanocomposite.

Tungsten oxide (WO3), MXene, and an WO3/MXene nanocomposite were synthesized to study their photocatalytic and biological applications. Tungsten oxide was synthesized by an easy and cost-effective hydrothermal method, and its composite with MXene was prepared through the sonication method. The synthesized tungsten oxide, MXene, and its composite were characterized by X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM), Fourier transform infrared (FTIR), energy-dispersive X-ray analysis (EDX), and Brunauer-Emmett-Teller (BET) for their structural, morphological, spectral, elemental and surface area analysis, respectively. The crystallite size of WO3 calculated from XRD was ~10 nm, the particle size of WO3 was 130 nm, and the average thickness of MXene layers was 175 nm, which was calculated from FESEM. The photocatalytic activity of as-synthesized samples was carried out for the degradation of methylene blue under solar radiation, MXene, the WO3/MXene composite, and WO3 exhibited 54%, 89%, and 99% photocatalytic degradation, respectively. WO3 showed maximal degradation ability; by adding WO3 to MXene, the degradation ability of MXene was enhanced. Studies on antibacterial activity demonstrated that these samples are good antibacterial agents against positive strains, and their antibacterial activity against negative strains depends upon their concentration. Against positive strains, the WO3/MXene composite's inhibition zone was at 7 mm, while it became 9 mm upon increasing the concentration. This study proves that WO3, MXene, and the WO3/MXene nanocomposite could be used in biological and environmental applications.

Tigecycline population pharmacokinetics in critically ill patients with decompensated cirrhosis and severe infections.

OBJECTIVES: Physiopathological changes in advanced cirrhosis could alter tigecycline pharmacokinetics (PK), thus affecting serum drug concentrations and compromising target attainment. We aimed to describe tigecycline PK in patients with decompensated cirrhosis and severe bacterial infections, identify the sources of PK variability and assess the performance of different dosing regimens to optimize the PK/pharmacodynamic (PD) target. METHODS: Serum concentrations and covariates were obtained from patients with severe infections under tigecycline treatment. A population PK analysis was performed using non-linear mixed-effects modelling and the final model was used to simulate tigecycline exposure to assess the PTA. RESULTS: Twenty critically ill patients were enrolled in the study. Data were best described by a two-compartment linear model. Mean ± SD parameter estimates for clearance (CL), intercompartmental clearance (Q), central and peripheral volumes of distribution (V1 and V2) were 14.8 ± 11 L/h, 38.4 ± 24 L/h, 63.7 ± 14 L and 233 ± 30 L, respectively. MELD score significantly influenced tigecycline CL, and total serum proteins significantly affected V1. Monte Carlo simulations showed that tigecycline elimination is hampered as MELD score values increase, consequently requiring lower drug doses. Patients with hypoproteinaemia would have lower peak tigecycline concentrations but similar steady-state concentrations compared with patients with normoproteinaemia. CONCLUSIONS: Our study confirms that tigecycline dose adjustment is needed in severe hepatic dysfunction and suggests using the MELD score for dose optimization since it is identified as a covariate that significantly influences tigecycline CL. Dosing regimens are recommended to reach several PK/PD targets considering this clinical variable and any MIC within the susceptibility range.

Rectal colonization by resistant bacteria increases the risk of infection by the colonizing strain in critically ill patients with cirrhosis.

BACKGROUND & AIMS: It remains unclear whether rectal colonization with multidrug-resistant organisms (MDROs) is prevalent and predisposes to infections by the same pathogens in patients with cirrhosis. METHODS: Two series of critically ill patients were evaluated. In the Barcelona cohort, 486 consecutive patients were prospectively evaluated, 129 with and 357 without cirrhosis (2015-2016). Rectal swabs were performed at admission and weekly thereafter (until intensive care unit [ICU] discharge) to detect MDRO colonization. Risk factors for colonization and infection by MDROs were evaluated. A retrospective cohort from Frankfurt (421 patients with cirrhosis; 2010-2018) was investigated to evaluate MDRO rectal colonization in another epidemiological scenario. RESULTS: In the Barcelona cohort, 159 patients were colonized by MDROs (32.7%), 102 (64.2%) at admission and 57 (35.8%) during follow-up. Patients with cirrhosis showed higher rates of rectal colonization at admission than those without cirrhosis (28.7% vs. 18.2%, p = 0.01) but similar colonization rates during ICU stay. Extended-spectrum beta-lactamase-Enterobacterales were the most frequent MDROs isolated in both groups. Colonization by MDROs independently increased the risk of infection by MDROs at admission and during follow-up. Risk of new infection by the colonizing strain was also significantly increased in patients with (hazard ratio [HR] 7.41) and without (HR 5.65) cirrhosis. Rectal colonization by MDROs was also highly prevalent in Frankfurt (n = 198; 47%; 131 at admission [66.2%] and 67 [33.8%] during follow-up), with vancomycin-resistant enterococci being the most frequent colonizing organism. Rectal colonization by MDROs was also associated with an increased risk of infection by MDROs in this cohort. Infections occurring in MDR carriers were mainly caused by the colonizing strain. CONCLUSION: Rectal colonization by MDROs is extremely frequent in critically ill patients with cirrhosis. Colonization increases the risk of infection by the colonizing resistant strain. LAY SUMMARY: Rectal colonization by multidrug-resistant organisms (MDROs) is a prevalent problem in patients with cirrhosis requiring critical care. The pattern of colonizing bacteria is heterogeneous with relevant differences between centers. Colonization by MDROs is associated with increased risk of infection by the colonizing bacteria in the short term. This finding suggests that colonization data could be used to guide empirical antibiotic therapy and de-escalation policies in patients with cirrhosis.

Changes in the gut microbiota and risk of colonization by multidrug-resistant bacteria, infection, and death in critical care patients.

OBJECTIVES: This study aimed to investigate whethehr the diversity and composition of the intestinal microbiota determines the risk of multidrug-resistant organism (MDRO) acquisition, infection, and mortality in patients admitted to a liver intensive care unit (ICU). METHODS: This prospective study included patients admitted to a 12-bed ICU between July and December 2018. Rectal swabs to detect MDRO intestinal colonization were obtained at ICU admission and weekly thereafter during the ICU stay. The 16S rRNA gene sequencing was performed on 138 rectal swabs from 62 patients. We evaluated the potential association between gut microbiota composition and diversity and the risk of MDRO colonization, infection, and hospital mortality. RESULTS: Of the patients studied, 19 of 62 (30.65%) presented with MDRO colonization at admission, 16 (25.81%) were colonized during their stay, and 27 (43.55%) were not colonized; 45 of 62 patients (72.58%) developed an infection, and mortality was 29.03% (18 of 62). Higher bacterial diversity and abundance of Bacillales Family XI incertae sedis and Prevotella families were associated with a lower risk of colonization by MDRO, infection, and death (linear discriminant analysis effect size score >4), whereas the Enterococcaceae family was associated with an increased risk of infection and death (linear discriminant analysis effect size score >4). The LASSO regression and multivariate analysis identified Family XI incertae sedis to be associated with a lower risk of infection (OR: 0.997; 95% CI, 0.996-0.999; p = 0.001) and microbial evenness index to be associated with lower mortality risk (OR: 0.68; 95% CI, 0.49-0.95; p = 0.02). DISCUSSION: Microbial diversity and abundance of certain bacterial taxa could have prognostic value in patients admitted to a critical care unit. Larger perspective studies should address the value of these markers in clinical practice.

Impact of enteropathogens on faltering growth in a resource-limited setting.

Introduction: Environmental enteropathy is an important contributor to childhood malnutrition in the developing world. Chronic exposure to fecal pathogens leads to alteration in intestinal structure and function, resulting in impaired gut immune function, malabsorption, and growth faltering leading to environmental enteropathy. Methods: A community-based intervention study was carried out on children till 24 months of age in Matiari district, Pakistan. Blood and fecal specimens were collected from the enrolled children aged 3-6 and 9 months. A real-time PCR-based TaqMan array card (TAC) was used to detect enteropathogens. Results: Giardia, Campylobacter spp., enteroaggregative Escherichia coli (EAEC), Enteropathogenic Escherichia coli (EPEC), Enterotoxigenic Escherichia coli (ETEC), and Cryptosporidium spp. were the most prevailing enteropathogens in terms of overall positivity at both time points. Detection of protozoa at enrollment and 9 months was negatively correlated with rate of change in height-for-age Z (ΔHAZ) scores during the first and second years of life. A positive association was found between Giardia, fecal lipocalin (LCN), and alpha 1-Acid Glycoprotein (AGP), while Campylobacter spp. showed positive associations with neopterin (NEO) and myeloperoxidase (MPO). Conclusion: Protozoal colonization is associated with a decline in linear growth velocity during the first 2 years of life in children living in Environmental enteric dysfunction (EED) endemic settings. Mechanistic studies exploring the role of cumulative microbial colonization, their adaptations to undernutrition, and their influence on gut homeostasis are required to understand symptomatic enteropathogen-induced growth faltering.

COVID-19 Deliveries: Maternal Features and Neonatal Outcomes.

Integrated, data-driven criteria are necessary to evaluate delivery outcomes in pregnancies affected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) during the ongoing COVID-19 pandemic. This study analyzed maternal demographics, clinical characteristics, treatments, and delivery outcomes of 85 ethnically diverse, adult pregnant women who tested positive for SARS-CoV-2 at the time of delivery. Median maternal and gestational ages were 27 years (interquartile range [IQR]: 23-31) and 39 weeks (IQR: 37.3-40.0), respectively. Of the 85 SARS-CoV-2-positive participants, 67 (79%) had no COVID-19 symptoms at the time of routine COVID-19 admission testing, 14 (16%) reported mild COVID-19 symptoms, and 4 (5%) presented severe COVID-19 symptoms that required hospitalization. Patients in the severe COVID-19 group had significantly longer hospitalizations than those with nonsevere COVID-19 (7 [IQR: 4.5-9.5] vs 2 [IQR: 2-3] days; P<0.01). Neonatal outcomes included 100% live births with a median 1-minute Apgar score of 8 and 15% preterm births. No neonatal deaths or vertical transmissions were reported, and all neonatal intensive care unit admissions were related to prematurity. Overall, maternal symptom prevalence and peripartum complication rates were low, suggesting a generally good prognosis for pregnant women with SARS-CoV-2 infections at the time of delivery.

Gut integrity and duodenal enteropathogen burden in undernourished children with environmental enteric dysfunction.

Environmental enteric dysfunction (EED) is a subclinical condition of intestinal inflammation, barrier dysfunction and malabsorption associated with growth faltering in children living in poverty. This study explores association of altered duodenal permeability (lactulose, rhamnose and their ratio) with higher burden of enteropathogen in the duodenal aspirate, altered histopathological findings and higher morbidity (diarrhea) that is collectively associated with linear growth faltering in children living in EED endemic setting. In a longitudinal birth cohort, 51 controls (WHZ > 0, HAZ > -1.0) and 63 cases (WHZ< -2.0, refractory to nutritional intervention) were recruited. Anthropometry and morbidity were recorded on monthly bases up to 24 months of age. Dual sugar assay of urine collected after oral administration of lactulose and rhamnose was assessed in 96 children from both the groups. Duodenal histopathology (n = 63) and enteropathogen analysis of aspirate via Taqman array card (n = 60) was assessed in only cases. Giardia was the most frequent pathogen and was associated with raised L:R ratio (p = 0.068). Gastric microscopy was more sensitive than duodenal aspirate in H. pylori detection. Microscopically confirmed H. pylori negatively correlated with HAZ at 24 months (r = -0.313, p = 0.013). Regarding histopathological parameters, goblet cell reduction significantly correlated with decline in dual sugar excretion (p< 0.05). Between cases and controls, there were no significant differences in the median (25th, 75th percentile) of urinary concentrations (µg/ml) of lactulose [27.0 (11.50, 59.50) for cases vs. 38.0 (12.0, 61.0) for controls], rhamnose [66.0 (28.0, 178.0) vs. 86.5 (29.5, 190.5)] and L:R ratio [0.47 (0.24, 0.90) vs. 0.51 (0.31, 0.71)] respectively. In multivariable regression model, 31% of variability in HAZ at 24 months of age among cases and controls was explained by final model including dual sugars. In conclusion, enteropathogen burden is associated with altered histopathological features and intestinal permeability. In cases and controls living in settings of endemic enteropathy, intestinal permeability test may predict linear growth. However, for adoption as a screening tool for EED, further validation is required due to its complex intestinal pathophysiology.

The Clinical Presentation of Culture-positive and Culture-negative, Quantitative Polymerase Chain Reaction (qPCR)-Attributable Shigellosis in the Global Enteric Multicenter Study and Derivation of a Shigella Severity Score: Implications for Pediatric Shigella Vaccine Trials.

BACKGROUND: Shigella is a leading cause of childhood diarrhea and target for vaccine development. Microbiologic and clinical case definitions are needed for pediatric field vaccine efficacy trials. METHODS: We compared characteristics of moderate to severe diarrhea (MSD) cases in the Global Enteric Multicenter Study (GEMS) between children with culture positive Shigella to those with culture-negative, quantitative polymerase chain reaction (qPCR)-attributable Shigella (defined by an ipaH gene cycle threshold <27.9). Among Shigella MSD cases, we determined risk factors for death and derived a clinical severity score. RESULTS: Compared to culture-positive Shigella MSD cases (n = 745), culture-negative/qPCR-attributable Shigella cases (n = 852) were more likely to be under 12 months, stunted, have a longer duration of diarrhea, and less likely to have high stool frequency or a fever. There was no difference in dehydration, hospitalization, or severe classification from a modified Vesikari score. Twenty-two (1.8%) Shigella MSD cases died within the 14-days after presentation to health facilities, and 59.1% of these deaths were in culture-negative cases. Age <12 months, diarrhea duration prior to presentation, vomiting, stunting, wasting, and hospitalization were associated with mortality. A model-derived score assigned points for dehydration, hospital admission, and longer diarrhea duration but was not significantly better at predicting 14-day mortality than a modified Vesikari score. CONCLUSIONS: A composite severity score consistent with severe disease or dysentery may be a pragmatic clinical endpoint for severe shigellosis in vaccine trials. Reliance on culture for microbiologic confirmation may miss a substantial number of Shigella cases but is currently required to measure serotype specific immunity.

Meropenem population pharmacokinetics in patients with decompensated cirrhosis and severe infections.

OBJECTIVES: Meropenem pharmacokinetics (PK) may be altered in patients with cirrhosis, hampering target attainment. We aimed to describe meropenem PK in patients with decompensated cirrhosis and severe bacterial infections, identify the sources of PK variability and assess the performance of different dosing regimens to optimize the PK/pharmacodynamic (PD) target. METHODS: Serum concentrations and covariates were obtained from patients with severe infections under meropenem treatment. A population PK analysis was performed using non-linear mixed-effects modelling and the final model was used to simulate meropenem exposure to assess the PTA. RESULTS: Fifty-four patients were enrolled in the study. Data were best described by a one-compartment linear model. The estimated typical mean value for clearance (CL) was 8.35 L/h and the estimated volume of distribution (V) was 28.2 L. Creatinine clearance (CLCR) and MELD score significantly influenced meropenem CL, and acute-on-chronic liver failure (ACLF) significantly affected V. Monte Carlo simulations showed that a lower meropenem dose would be needed as CLCR decreases and as the MELD score increases. Patients with ACLF would have lower peak meropenem concentrations but similar steady-state concentrations compared with patients with no ACLF. CONCLUSIONS: Our study identified two new covariates that influence meropenem PK in patients with decompensated cirrhosis in addition to CLCR: MELD score and ACLF. Dosing regimens are recommended to reach several PK/PD targets considering these clinical variables and any MIC within the susceptibility range.

Epidemiology of Shigella infections and diarrhea in the first two years of life using culture-independent diagnostics in 8 low-resource settings.

Culture-independent diagnostics have revealed a larger burden of Shigella among children in low-resource settings than previously recognized. We further characterized the epidemiology of Shigella in the first two years of life in a multisite birth cohort. We tested 41,405 diarrheal and monthly non-diarrheal stools from 1,715 children for Shigella by quantitative PCR. To assess risk factors, clinical factors related to age and culture positivity, and associations with inflammatory biomarkers, we used log-binomial regression with generalized estimating equations. The prevalence of Shigella varied from 4.9%-17.8% in non-diarrheal stools across sites, and the incidence of Shigella-attributable diarrhea was 31.8 cases (95% CI: 29.6, 34.2) per 100 child-years. The sensitivity of culture compared to qPCR was 6.6% and increased to 27.8% in Shigella-attributable dysentery. Shigella diarrhea episodes were more likely to be severe and less likely to be culture positive in younger children. Older age (RR: 1.75, 95% CI: 1.70, 1.81 per 6-month increase in age), unimproved sanitation (RR: 1.15, 95% CI: 1.03, 1.29), low maternal education (<10 years, RR: 1.14, 95% CI: 1.03, 1.26), initiating complementary foods before 3 months (RR: 1.10, 95% CI: 1.01, 1.20), and malnutrition (RR: 0.91, 95% CI: 0.88, 0.95 per unit increase in weight-for-age z-score) were risk factors for Shigella. There was a linear dose-response between Shigella quantity and myeloperoxidase concentrations. The burden of Shigella varied widely across sites, but uniformly increased through the second year of life and was associated with intestinal inflammation. Culture missed most clinically relevant cases of severe diarrhea and dysentery.

Genomewide Association Study of Severe Covid-19 with Respiratory Failure.

BACKGROUND: There is considerable variation in disease behavior among patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (Covid-19). Genomewide association analysis may allow for the identification of potential genetic factors involved in the development of Covid-19. METHODS: We conducted a genomewide association study involving 1980 patients with Covid-19 and severe disease (defined as respiratory failure) at seven hospitals in the Italian and Spanish epicenters of the SARS-CoV-2 pandemic in Europe. After quality control and the exclusion of population outliers, 835 patients and 1255 control participants from Italy and 775 patients and 950 control participants from Spain were included in the final analysis. In total, we analyzed 8,582,968 single-nucleotide polymorphisms and conducted a meta-analysis of the two case-control panels. RESULTS: We detected cross-replicating associations with rs11385942 at locus 3p21.31 and with rs657152 at locus 9q34.2, which were significant at the genomewide level (P<5×10-8) in the meta-analysis of the two case-control panels (odds ratio, 1.77; 95% confidence interval [CI], 1.48 to 2.11; P = 1.15×10-10; and odds ratio, 1.32; 95% CI, 1.20 to 1.47; P = 4.95×10-8, respectively). At locus 3p21.31, the association signal spanned the genes SLC6A20, LZTFL1, CCR9, FYCO1, CXCR6 and XCR1. The association signal at locus 9q34.2 coincided with the ABO blood group locus; in this cohort, a blood-group-specific analysis showed a higher risk in blood group A than in other blood groups (odds ratio, 1.45; 95% CI, 1.20 to 1.75; P = 1.48×10-4) and a protective effect in blood group O as compared with other blood groups (odds ratio, 0.65; 95% CI, 0.53 to 0.79; P = 1.06×10-5). CONCLUSIONS: We identified a 3p21.31 gene cluster as a genetic susceptibility locus in patients with Covid-19 with respiratory failure and confirmed a potential involvement of the ABO blood-group system. (Funded by Stein Erik Hagen and others.).

Efficacy of Albumin Treatment for Patients with Cirrhosis and Infections Unrelated to Spontaneous Bacterial Peritonitis.

BACKGROUND & AIMS: We performed a randomized trial to determine whether albumin should be administered to patients with infections unrelated to spontaneous bacterial peritonitis (SBP). METHODS: We performed a multicenter, open-label trial in which 118 patients with cirrhosis, non-SBP infections, and additional risk factors for poor outcome were randomly assigned to receive antibiotics plus albumin (study group; n = 61) or antibiotics alone (control group; n = 57). The primary outcome was in-hospital mortality; secondary outcomes were effect of albumin on disease course. RESULTS: There were no significant differences at baseline between groups in results from standard laboratory tests, serum markers of inflammation, circulatory dysfunction, or liver severity scores. However, the combined prevalence of acute on chronic liver failure (ACLF) and kidney dysfunction was significantly higher in the study group (44.3% vs 24.6% in the control group; P = .02), indicating greater baseline overall severity. There was no significant difference in the primary outcome between groups (13.1% in the study group vs 10.5% in the control group; P = .66). Circulatory and renal functions improved in only the study group. A significantly higher proportion of patients in the study group had resolution of ACLF (82.3% vs 33.3% in the control group; P = .03). A significantly lower proportion of patients in the study group developed nosocomial infections (6.6% vs 24.6% in the control group; P = .007). CONCLUSIONS: In a randomized trial of patients with advanced cirrhosis and non-SBP infections, in-hospital mortality was similar between those who received albumin plus antibiotics vs those who received only antibiotics (controls). However, patients given albumin were sicker at baseline and, during the follow-up period, a higher proportion had ACLF resolution and a lower proportion had nosocomial infections. ClinicalTrials.gov no: NCT02034279.

Plasma levels of circulating DNA are associated with outcome, but not with activation of coagulation in decompensated cirrhosis and ACLF.

Acute-on-chronic liver failure (ACLF) is a recently (re)defined syndrome of acute decompensation of cirrhosis that presents with extrahepatic organ failure(s) and poor outcome. Given the prominent role of inflammation and activation of coagulation in ACLF, we hypothesized that ACLF might be characterized by the generation of neutrophil extracellular traps (NETs), that could drive both activation of coagulation and progression of organ failure. METHODS: We measured markers of circulating DNA, activation of coagulation, inflammation, and oxidative stress in 52 patients with acute decompensation (AD) of cirrhosis and 57 patients with ACLF on admission, and compared levels with 40 healthy controls. RESULTS: All analytes were higher in patients compared to controls. Plasma levels of cell-free DNA, but not of the specific NET marker myeloperoxidase-DNA complexes were higher in patients with ACLF compared to AD cirrhosis. In addition, TAT complexes (coagulation), IL-6 (inflammation), and TBARS (oxidative stress) were higher in ACLF compared to AD. Markers for activation of coagulation were not associated with circulating DNA, IL-6, or TBARS. In contrast, levels of circulating DNA, IL-6, and TBARS were higher in patients with more severe disease, higher in patients with organ failure, and higher in patients that died within 30 days of admission. Importantly, myeloperoxidase-DNA levels did not differ between patients with complications and poor outcome. CONCLUSIONS: Collectively, we show that cell-free DNA, inflammation, and oxidative stress are associated with outcomes in AD and ACLF, but not with activation of coagulation. Our data argue against a role of NETs in activation of coagulation and in progression of organ failure in patients with AD and ACLF. LAY SUMMARY: Acute-on-chronic liver failure is a devastating syndrome that can follow acute decompensation of chronic liver disease. Herein, we demonstrate that these patients accumulate DNA released from dying cells in their blood, and that the quantity of this DNA is related to the outcome of disease. We also show that outcome of disease is not related to recently described neutrophil extracellular traps, which have been shown in animal models to play vital roles in the progression of liver diseases.