Flow cytometry as a tool for the rapid enumeration of 1-µm microplastics spiked in wastewater and activated sludge after coagulation-flocculation-sedimentation.

Considering the limited literature and the difficulty of quantifying 1-µm micro-nanoplastics (1-µm MNP) in complex aqueous matrices such as wastewater and sludge, the removal rate of these very small particles in wastewater treatment plants (WWTP) represents a major challenge. In this study, coagulation-flocculation-sedimentation (CFS) with aluminum salts was investigated to evaluate the removal of 1-µm MNPs spiked in tap water, raw wastewater, pre-settled wastewater, and activated sludge. Quantification of 1-µm MNP was performed using the high-throughput flow cytometry (FCM) analysis which takes only a few minutes and produces results with high accuracy and reproducibly. The results indicated that the 1-µm MNPs were highly stable in pure water and unable to settle rapidly. In raw wastewater, sedimentation without coagulants removed less than 4% of 1-µm MNP. Conversely, CFS treatment showed a significant improvement in the removal of 1-µm MNP from wastewater. At dosages of 0.3-3 mg Al3+/L, the removal of MNPs in wastewater reached 30% and no flocs were observed, while floc formation was visible with increased dosages of 3-12 mg Al3+/L, obtaining MNP removal greater than 90%. CFS in activated sludge with a solids content of 5800 mg MLSS/L registered the highest removal efficiency (95-99%) even for dosages of 0.3-60 mg Al3+/L and pH dropping to 5. However, activated sludge showed extremely high removal efficiency of MNPs (97.3 ± 0.9%) even without coagulants. The large, dense flocs that constitute activated sludge appear particularly efficient in capturing 1-µm MNPs during the sedimentation process even in the absence of coagulants.

A Case of Acrodermatitis Dysmetabolica in a Child Affected by Citrullinemia Type I: When Early Diagnosis and Timely Treatment Are Not Enough.

An infant with a prenatal diagnosis of citrullinemia, who started standard treatment at birth (L-arginine; sodium benzoate and a personalized diet characterized by a low protein intake and supplementation of essential nutrients and amino acids), presented at 4 months of age with extended, progressive, and severe skin lesions consistent with acrodermatitis dysmetabolica. Guidelines for the diagnosis and management of urea cycle disorders underline that a low-protein diet places patients at risk of essential fatty acids, trace elements, and vitamin deficiency. At hospital admission, our patient had normal levels of zinc and alkaline phosphatases. The plasmatic amino acid profile revealed a severe and generalized deficiency. In particular, the serum levels of arginine, valine, and isoleucine were very low and the dermatitis did not improve until the blood levels of these amino acids increased. In our patient, skin lesions happened despite an early diagnosis of citrullinemia and timely treatment due to compliance issues as a consequence of linguistic barriers.

Antitissue transglutaminase antibodies' normalization after starting a gluten-free diet in a large population of celiac children-a real-life experience.

INTRODUCTION: Few data are available regarding the trend of IgA anti-transglutaminase antibodies (TGA-IgA) in children with celiac disease (CD) on a gluten-free diet (GFD). Our aim is to examine the normalization time of CD serology in a large pediatric population, and its predictors. MATERIAL AND METHODS: We retrospectively evaluated the normalization time of TGA-IgA and its predictive factors (age, sex, ethnicity, symptoms, associated diabetes/thyroiditis, Marsh stage, TGA-IgA and endomysial antibody levels at diagnosis, diet adherence), in 1024 children diagnosed from 2000 to 2019 in three pediatric Italian centers, on a GFD. RESULTS: TGA-IgA remission was reached in 67,3%, 80,7%, 89,8% and 94,9% after 12, 18, 24 and 36 months from starting a GFD, respectively (median time = 9 months). TGA-IgA >10´upper limit of normal at diagnosis (HR = 0.56), age 7-12 years old (HR = 0.83), poor compliance to diet (HR = 0.69), female sex (HR = 0.82), non-Caucasian ethnicity (HR = 0.75), and comorbidities (HR = 0.72) were independent factors significantly associated with longer time to normalization. CONCLUSIONS: Our population is the largest in the literature, with the majority of patients normalizing CD serology within 24 months from starting a GFD. We suggest a special attention to patients with comorbidities, language barriers or age 7-12 years for a proper management and follow-up.

Pyelonephritis in Pediatric Uropathic Patients: Differences from Community-Acquired Ones and Therapeutic Protocol Considerations. A 10-Year Single-Center Retrospective Study.

Pyelonephritis (PN) represents an important cause of morbidity in the pediatric population, especially in uropathic patients. The aim of the study is to demonstrate differences between PNs of uropathic patients and PNs acquired in community in terms of uropathogens involved and antibiotic sensitivity; moreover, to identify a proper empiric therapeutic strategy. A retrospective study was conducted on antibiograms on urine cultures from PNs in vesicoureteral reflux (VUR) patients admitted to pediatric surgery department and from PNs in not VUR patients admitted to Pediatric Emergency Unit between 2010 and 2020. We recorded 58 PNs in 33 patients affected by VUR and 112 PNs in the not VUR group. The mean age of not VUR patients at the PN episode was 1.3 ± 2.6 years (range: 20 days of life-3 years), and almost all the urine cultures, 111 (99.1%), isolated Gram-negative bacteria and rarely, 1 (0.9%), Gram-positive bacteria. The Gram-negative uropathogens isolated were Escherichia coli (97%), Proteus mirabilis (2%), and Klebsiella spp. (1%). The only Gram-positive bacteria isolated was an Enterococcus faecalis. As regards the antibiograms, 96% of not VUR PNs responded to beta-lactams, 99% to aminoglycosides, and 80% to sulfonamides. For the VUR group, mean age was 3.0 years ± 3.0 years (range: 9 days of life-11 years) and mean number of episodes per patient was 2.0 ± 1.0 (range: 1-5); 83% of PNs were by Gram-negatives bacteria vs. 17% by Gram-positive: the most important Gram-negative bacteria were Pseudomonas aeruginosa (44%), Escherichia coli (27%), and Klebsiella spp. (12%), while Enterococcus spp. determined 90% of Gram-positive UTIs. Regimen ampicillin/ceftazidime (success rate: 72.0%) was compared to ampicillin/amikacin (success rate of 83.0%): no statistically significant difference was found (p = 0.09). The pathogens of PNs in uropathic patients are different from those of community-acquired PNs, and clinicians should be aware of their peculiar antibiotic susceptibility. An empiric therapy based on the association ampicillin + ceftazidime is therefore suggested.

SARS-CoV-2 from faeces to wastewater treatment: What do we know? A review.

SARS-CoV-2, the virus that causes COVID-19, has been found in the faeces of infected patients in numerous studies. Stool may remain positive for SARS-CoV-2, even when the respiratory tract becomes negative, and the interaction with the gastrointestinal tract poses a series of questions about wastewater and its treatments. This review aims to understand the viral load of SARS-CoV-2 in faeces and sewage and its fate in wastewater treatment plants (WWTPs). The viral load in the faeces of persons testing positive for SARS-CoV-2 was estimated at between 5·103 to 107.6 copies/mL, depending on the infection course. In the sewerage, faeces undergo dilution and viral load decreases considerably in the wastewater entering a WWTP with a range from 2 copies/100 mL to 3·103 copies/mL, depending on the level of the epidemic. Monitoring of SARS-CoV-2 in sewage, although no evidence of COVID-19 transmission has been found via this route, could be advantageously exploited as an early warning of outbreaks. Preliminary studies on WBE seem promising; but high uncertainty of viral loads in wastewater and faeces remains, and further research is needed. The detection of SARS-CoV-2 in sewage, based on RNA sequences and RT-PCR, requires a shared approach on sample pre-treatment and on-site collection to ensure comparable results. The finding of viral RNA in stools does not imply that the virus is viable and infectious. Viability of CoVs such as SARS-CoV-2 decreases in wastewater - due to temperature, pH, solids, micropollutants - but high inactivation in WWTPs can be obtained only by using disinfection (free chlorine, UVC light). A reduction in the quantity of disinfectants can be obtained by implementing Membrane-Bioreactors with ultrafiltration to separate SARS-CoV-2 virions with a size of 60-140 nm. In sludge treatment, thermophilic digestion is effective, based on the general consensus that CoVs are highly sensitive to increased temperatures.

Effects of hydrodynamic cavitation, low-level thermal and low-level alkaline pre-treatments on sludge solubilisation.

WAS is a polluting and hazardous waste generated in WWTPs that must be treated to prevent pollution and human health risks. Anaerobic digestion is the most used process for sludge stabilization. However, it must be improved in terms of both speed and extend of degradation. With the purpose of reducing the energy and chemical consumption linked to sludge treatment, in this study, different anaerobic digestion pre-treatments such as low-level mechanical (hydrodynamic cavitation, 2 bar), low-level thermal (50 °C) and low-level alkaline (NaOH, KOH and Ca(OH)2, pH 10) methods, and a combination thereof, were tested as strategies to improve sludge solubilisation. When the pre-treatments were used alone, the alkaline pre-treatment showed the highest sludge solubilisation. Among the alkaline reagents tested, NaOH and KOH led to higher DDPCOD (41.6 and 39.4%), while only 8.4% was achieved by using Ca(OH)2. However, the low-level hydrodynamic cavitation assisted thermo-alkaline pre-treatment was the most efficient in terms of both sludge solubilisation (DDPCOD = 53.0%) and energy efficiency (EE = 64.5 mgΔSCOD kJ-1). The synergetic effects of the combined pre-treatment were also confirmed by the highest release of EPS. Furthermore, cytometric analyses showed that the main mechanism involved in sludge solubilisation for the investigated pre-treatments was flocs disintegration rather than cell lysis.

Assessment of activated sludge viability with flow cytometry.

The aim of the study was to evaluate the applicability of fluorescent dyes and multiparameter flow cytometry for the rapid and direct viability/activity assessment of activated sludge samples taken from wastewater treatment plants. Viability and activity of the biomass were estimated respectively through cellular membrane integrity, staining with SYBR Green I and Propidium Iodide, and through fluorogenic dyes capable of detecting enzymatic activity, as FDA and BCECF-AM. A procedure has been developed to disaggregate sludge flocs before dyes staining and cytometric analysis. The developed procedure allows a high recovery of bacteria with good accuracy and repeatability, and minimize the damage of the cells suspension obtained from the disaggregation of the flocs. These measurements were applied to estimate the two main parameters required to define the biological activated sludge process: the endogenous decay rate and the specific growth rate in exponential phase with high F/M ratio. Oxygen utilization rate measurements (OUR) were conducted to conventionally monitor the activity of the biomass. The preliminary data are encouraging and support the possibility to investigate bacteria dynamics on wastewater treatment plants.