Vaginal Lactobacilli Supernatants Protect from Herpes Simplex Virus Type 1 Infection in Cell Culture Models.

A healthy vaginal microbiota hosts Lactobacillus as the most predominant genus. Lactobacilli play a role in human health through the production of diverse antimicrobial substances that can act against human pathogens or modulate the immune system. Previous reports highlighted the ability of vaginal lactobacilli to counteract viruses causing STIs, e.g., HIV-1 and HSV-2. In this report, we analyze the activity of supernatants of vaginal lactobacilli against HSV-1 infection, which is becoming increasingly relevant as a STI. We show that the supernatants of two vaginal Lactobacillus species (i.e., L. crispatus and L. gasseri) were active at neutralizing HSV-1 infection in two different cell lines of human and simian origin. Specifically, we demonstrate that L. crispatus strains are the most effective in antiviral activity, as evidenced by the comparison with a vaginal pathogen taken as reference. The effect was specific and not attributable to the generic toxicity of the supernatants to the cells. Our results pave the way for the development of probiotics to limit the impact of HSV-1 infection on women's health.

The Use of ICD-9-CM Coding to Identify COVID-19 Diagnoses and Determine Risk Factors for 30-Day Death Rate in Hospitalized Patients in Italy: Retrospective Study.

BACKGROUND: In Italy, it has been difficult to accurately quantify hospital admissions of patients with a COVID-19 diagnosis using the Hospital Information System (HIS), mainly due to the heterogeneity of codes used in the hospital discharge records during different waves of the COVID-19 pandemic. OBJECTIVE: The objective of this study was to define a specific combination of codes to identify the COVID-19 hospitalizations within the HIS and to investigate the risk factors associated with mortality due to COVID-19 among patients admitted to Italian hospitals in 2020. METHODS: A retrospective study was conducted using the hospital discharge records, provided by more than 1300 public and private Italian hospitals. Inpatient hospitalizations were detected by implementing an algorithm based on specific International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) code combinations. Hospitalizations were analyzed by different clinical presentations associated with COVID-19 diagnoses. In addition, 2 multivariable Cox regression models were performed among patients hospitalized "due to COVID-19" from January 1 to December 31, 2020, to investigate potential risk factors associated with 30-day death and the temporal changes over the course of the pandemic; in particular, the 30-day death rates during the first and the second waves were analyzed across 3 main geographical areas (North, Center, and South and Islands) and by discharge wards (ordinary and intensive care). RESULTS: We identified a total of 325,810 hospitalizations with COVID-19-related diagnosis codes. Among these, 73.4% (n=239,114) were classified as "due to COVID-19," 14.5% (n=47,416) as "SARS-CoV-2 positive, but not due to COVID-19," and 12.1% (n=39,280) as "suspected COVID-19" hospitalizations. The cohort of patients hospitalized "due to COVID-19" included 205,048 patients, with a median age of 72 years and a higher prevalence of male patients (n=124,181, 60.6%). The overall 30-day death rate among hospitalized patients due to COVID-19 was 9.9 per 1000 person-days. Mortality was lower for women (hazard ratio [HR]=0.83; P<.001) and for patients coming from high migration pressure countries, especially Northern Africans (HR=0.65; P<.001) and Central and Eastern Europeans (HR=0.66; P<.001), compared to patients coming from Italy and high-income countries. In the southern regions and the Islands, mortality was higher compared to the northern regions (HR=1.17; P<.001), especially during the second wave of COVID-19 among patients with a transfer to intensive care units (HR=2.52; P<.001). CONCLUSIONS: To our knowledge, the algorithm is the first attempt to define, at a national level, selection criteria for identifying COVID-19 hospitalizations within the HIS. The implemented algorithm will be used to monitor the pandemic over time, and the patients selected in 2020 will be followed up in the next years to assess the long-term effects of COVID-19.

Limosilactobacillus vaginalis Exerts Bifidogenic Effects: A Novel Postbiotic Strategy for Infant Prebiotic Supplementation.

Infant microbiota shaping strictly influences newborns' well-being and long-term health, and babies born by cesarean-section and formula-fed generally show low microbial gut diversity and are more prone to develop various disorders. The supplementation with beneficial microbes of vaginal origin or derivatives (postbiotics, including heat-inactivated cells) represents a valid strategy to drive the correct gut microbiota shaping. Here, we explored for the first time the bifidogenic activity of a heat-killed vaginal strain (Limosilactobacillus vaginalis BC17), in addition to the assessment of its safety. L. vaginalis BC17 whole genome was sequenced by Nanopore technology and highlighted the absence of antibiotic resistance genes and virulence factors, indicating the strain safety profile for human health. MIC values confirmed that L. vaginalis BC17 is susceptible to widely employed antibiotics. Heat-killed BC17 cells significantly enhanced the planktonic growth of Bifidobacterium spp. For the first time, stimulating effects were observed also toward biofilm formation of bifidobacteria and their pre-formed biofilms. Conversely, heat-killed BC17 cells exerted antibacterial and anti-biofilms activities against Gram-positive and Gram-negative pathogens. Lyophilized heat-killed BC17 cells were formulated in a sunflower oil suspension (1010 heat-killed cell/g) intended for infant oral intake. This possessed optimal technological (i.e., re-dispersibility and stability) and functional properties (i.e., bifidogenic activity) that were maintained even after pre-digestion in acidic conditions.

Trends and perioperative mortality in gastric cancer surgery: a nationwide population­based cohort study.

This study aimed to investigate changes and perioperative mortality over a 6-year period within the Italian Hospital Information System among patients with gastric cancer (GC) who underwent gastrectomies and to identify risk factors associated with 90-day mortality. Additionally, nationwide differences between high and low-volume hospitals were evaluated. A nationwide retrospective study was conducted using patient hospital discharge records (HDRs) based on the International Classification of Diseases, 9th Revision, Clinical Modification (ICD-9-CM) classification. The HDRs were linked to the National Tax Registry records using deterministic record linkage. The data were obtained from the Italian National Outcomes Evaluation Programme (PNE). Multivariate logistic regression was used to examine risk factors for 90-day mortality among patients with GC who underwent partial or total gastrectomies over the period from 2018 to 2020 with adjustment for comorbidities. Overall, the number of patients with GC who underwent total or partial gastrectomies steadily decreased in Italy from 5765 in 2015 to 4291 in 2020 (p < 0.001). The use of the laparoscopic approach more than doubled from 2015 (10.8%) to 2020 (26.3%), with a concomitant conversion rate from laparoscopy to open surgery decreasing from 7.7 to 5.8%. The 30 and 90-day mortality rates remained stable over time (p > 0.05). Low-volume hospitals had higher inpatient, early, and late mortality compared to high-volume hospitals (5.9% vs 3.8%, 6.3% vs 3.8%, and 11.8% vs 7.9%, respectively; p < 0.001). Multivariate logistic regression analysis showed that an advanced age (adjusted odds ratio: 3.72; 95% [CI]: 3.15-4.39; p < 0.001), an open approach (adjusted-OR: 1.69, 95% CI: 1.43-1.99, p < 0.001) and a total gastrectomy (adjusted-OR: 1.44, 95% CI: 1.27-1.64, p < 0.001) were independent risk factors for 90-day mortality. Additionally, patients with GC who referred to high-volume hospitals were 26% less likely to die within 90 days after a gastrectomy than those who underwent surgery in low-volume hospitals. During the 6-year period, surgeons implemented a minimally invasive approach to reduce the conversion over time. Centralisation was associated with better outcomes while advanced age, an open approach, and total gastrectomy were identified as risk factors for 90-day mortality.

Impact of COVID-19 Diagnosis on Mortality in Patients with Ischemic Stroke Admitted during the 2020 Pandemic in Italy.

AIMS: The impact of the COVID-19 pandemic on the event rate of patients with ischemic stroke has been poorly investigated. We sought to evaluate the impact of the COVID-19 infection on mortality in patients with ischemic stroke admitted during the 2020 pandemic in Italy. METHODS: We analyzed a nationwide, comprehensive, and universal administrative database of patients who were admitted for ischemic stroke during and after the national lockdown for the COVID-19 infection in 2020, and the equivalent periods over the previous 5 years in Italy. The 2020 observed hospitalization and mortality rates of stroke patients with and without COVID-19 infection were compared with the expected rates, in accordance with the trend of the previous 5 years. RESULTS: During the period of observation, 300,890 hospitalizations for ischemic stroke occurred in Italy. In 2020, 41,302 stroke patients (1102 with concomitant COVID-19 infection) were admitted at 771 centers. The rate of admissions for ischemic stroke during the 2020 pandemic was markedly reduced compared with previous years (percentage change vs. 2015: -23.5). Based on the 5 year trend, the 2020 expected 30 day and 1 year mortality rates were 9.8% and 23.9%, respectively, and the observed incidence of death rates were 12.2% and 26.7%, respectively (both p < 0.001). After multiple corrections, higher rates of mortality were observed among patients admitted for stroke with a concomitant COVID-19 diagnosis. CONCLUSIONS: During the COVID-19 pandemic in 2020 in Italy, the rate of hospitalizations for ischemic stroke was dramatically reduced, although both the 30 day and 1 year mortality rates increased compared with the previous 5 year trend.

Lecithin and Chitosan as Building Blocks in Anti-Candida Clotrimazole Nanoparticles.

The main focus when considering treatment of non-healing and infected wounds is tied to the microbial, particularly bacterial, burden within the wound bed. However, as fungal contributions in these microbial communities become more recognized, the focus needs to be broadened, and the remaining participants in the complex wound microbiome need to be addressed in the development of new treatment strategies. In this study, lecithin/chitosan nanoparticles loaded with clotrimazole were tailored to eradicate one of the most abundant fungi in the wound environment, namely C. albicans. Moreover, this investigation was extended to the building blocks and their organization within the delivery system. In the evaluation of the novel nanoparticles, their compatibility with keratinocytes was confirmed. Furthermore, these biocompatible, biodegradable, and non-toxic carriers comprising clotrimazole (~189 nm, 24 mV) were evaluated for their antifungal activity through both disk diffusion and microdilution methods. It was found that the activity of clotrimazole was fully preserved upon its incorporation into this smart delivery system. These results indicate both that the novel carriers for clotrimazole could serve as a therapeutic alternative in the treatment of fungi-infected wounds and that the building blocks and their organization affect the performance of nanoparticles.

Exopolysaccharides from vaginal lactobacilli modulate microbial biofilms.

BACKGROUND: Exopolysaccharides (EPS) secreted by beneficial lactobacilli exert a plethora of positive activities, but little is known about their effects on biofilms of opportunistic vaginal pathogens and especially on biofilms of lactobacilli themselves. Here, the EPS produced by six vaginal lactobacilli, belonging to Lactobacillus crispatus (BC1, BC4, BC5) and Lactobacillus gasseri (BC9, BC12, BC14) species were isolated from cultural supernatants and lyophilized. RESULTS: Lactobacillus EPS were chemically characterized in terms of monosaccharide composition by liquid chromatography (LC) analysis coupled to UV and mass spectrometry (MS) detection. Moreover, the ability of EPS (0.1, 0.5, 1 mg/mL) to stimulate the biofilm formation of lactobacilli and to inhibit the formation of pathogens' biofilms was evaluated by crystal violet (CV) staining and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Isolated EPS (yields 133-426 mg/L) were heteropolysaccharides mainly composed of D-mannose (40-52%) and D-glucose (11-30%). For the first time we demonstrated that Lactobacillus EPS were able to stimulate in a dose-dependent manner (p < 0.05) the formation of biofilms of ten strains belonging to L. crispatus, L. gasseri and Limosilactobacillus vaginalis species, in terms of cell viability (84-282% increase at 1 mg/mL) and especially biofilm biomass (40-195% increase at 1 mg/mL), quantified with MTT assay and CV staining, respectively. EPS released from L. crispatus and L. gasseri were found to better stimulate the biofilms of the same producer species rather than that of other species, including producing strains themselves and other strains. Conversely, the biofilm formation of bacterial (Escherichia coli, Staphylococcus spp., Enterococcus spp. and Streptococcus agalactiae) and fungal (Candida spp.) pathogens was inhibited. The anti-biofilm activity was dose-dependent and was more marked for L. gasseri-derived EPS (inhibition up to 86%, 70%, and 58% at 1 mg/mL, 0.5 mg/mL, and 0.1 mg/mL, respectively), whilst L. crispatus-derived EPS resulted overall less efficient (inhibition up to 58% at 1 mg/mL and 40% at 0.5 mg/mL) (p < 0.05). CONCLUSIONS: Lactobacilli-derived EPS favour the biofilm formation of lactobacilli preventing, at the same time, that of opportunistic pathogens. These results support the possible employment of EPS as postbiotics in medicine as a therapeutic/preventive strategy to counteract vaginal infections.

Prebiotic Activity of Vaginal Lactobacilli on Bifidobacteria: from Concept to Formulation.

The gut of babies born vaginally is rapidly colonized by Bifidobacterium spp. after birth, while in infants born by cesarean section (C-section), the presence of bifidobacteria drops dramatically, increasing the risk of developing gastrointestinal disorders. Considering that newborns naturally come into contact with maternal lactobacilli as they pass through the birth canal, the aim of this work is to exploit for the first time the bifidogenic activity exerted by the cell-free supernatants (CFSs) from lactobacilli of vaginal origin, belonging to the species Lactobacillus crispatus, Lactobacillus gasseri, Limosilactobacillus vaginalis, and Lactiplantibacillus plantarum. CFSs were recovered after 7 h, 13 h, and 24 h of fermentation and assessed for the ability to stimulate the planktonic growth and biofilms of Bifidobacterium strains belonging to species widely represented in the gut tract. A bifidogenic effect was observed for all CFSs; such activity was maximal for CFSs recovered in exponential phase and was strongly dependent on the species of lactobacilli. Importantly, no stimulating effects on an intestinal Escherichia coli strain were observed. CFSs from L. vaginalis BC17 showed the best bifidogenic profile since they increased bifidobacterial planktonic growth by up to 432% and biofilm formation by up to 289%. The CFS at 7 h from BC17 was successfully formulated with a hyaluronic acid-based hydrogel aimed at preventing and treating breast sores in lactating women and exerting bifidogenic activity in infants born mainly by C-section. IMPORTANCE Bifidobacteria in the gut tract of infants play crucial roles in the prevention of gastrointestinal diseases and the maturation of the immune system. Consequently, strategies to trigger a bifidogenic shift in the infant gut are highly desirable. Evidences suggest that the presence of a maternal vaginal microbiota dominated by health-promoting lactobacilli and the development of a bifidobacterium-enriched gut microbiota in newborns are interconnected. In this context, we found out that the cell-free supernatants from lactobacilli of vaginal origin were able to effectively stimulate the proliferation of Bifidobacterium spp. grown in free-floating and biofilm forms. The cell-free supernatant from Limosilactobacillus vaginalis BC17 showed excellent bifidogenic behavior, which was preserved even after its incorporation into a nipple formulation for lactating women. Lactobacilli derivatives, such as cell-free supernatants, have gained increasing interest by virtue of their safer profile than that of living cells and can be proposed as an ecosustainable approach to favor gut colonization of infants by bifidobacteria.

Tailored anti-biofilm activity - Liposomal delivery for mimic of small antimicrobial peptide.

The eradication of bacteria embedded in biofilms is among the most challenging obstacles in the management of chronic wounds. These biofilms are found in most chronic wounds; moreover, the biofilm-embedded bacteria are considerably less susceptible to conventional antimicrobial treatment than the planktonic bacteria. Antimicrobial peptides and their mimics are considered attractive candidates in the pursuit of novel therapeutic options for the treatment of chronic wounds and general bacterial eradication. However, some limitations linked to these membrane-active antimicrobials are making their clinical use challenging. Novel innovative delivery systems addressing these limitations represent a smart solution. We hypothesized that incorporation of a novel synthetic mimic of an antimicrobial peptide in liposomes could improve its anti-biofilm effect as well as the anti-inflammatory activity. The small synthetic mimic of an antimicrobial peptide, 7e-SMAMP, was incorporated into liposomes (~280 nm) tailored for skin wounds and evaluated for its potential activity against both biofilm formation and eradication of pre-formed biofilms. The 7e-SMAMP-liposomes significantly lowered inflammatory response in murine macrophages (~30 % reduction) without affecting the viability of macrophages or keratinocytes. Importantly, the 7e-SMAMP-liposomes completely eradicated biofilms produced by Staphylococcus aureus and Escherichia coli above concentrations of 6.25 µg/mL, whereas in Pseudomonas aeruginosa the eradication reached 75 % at the same concentration. Incorporation of 7e-SMAMP in liposomes improved both the inhibition of biofilm formation as well as biofilm eradication in vitro, as compared to non-formulated antimicrobial, therefore confirming its potential as a novel therapeutic option for bacteria-infected chronic wounds.

Covid-19 diagnosis and mortality in patients with non-ST-elevation myocardial infarction admitted in Italy during the national outbreak.

INTRODUCTION: We sought to assess the clinical impact of Covid-19 infection on mortality in patients with Non-ST elevation myocardial infarction (NSTEMI) admitted during the national outbreak in Italy. METHODS: We analysed a nationwide, comprehensive, and universal administrative database of consecutive NSTEMI patients admitted during lockdown for Covid-19 infection (March,11st - May 3rd, 2020) and the equivalent periods of the previous 5 years in Italy. The observed rate of 30-day and 6-month all-cause mortality of NSTEMI patients with and without Covid-19 infection during the lockdown was compared with the expected rate of death according to the trend of the previous 5 years. RESULTS: During the period of observation, 48.447 NSTEMI hospitalizations occurred in Italy. Among these, 4981 NSTEMI patients were admitted during the 2020 outbreak: 173 (3.5%) with and 4808 (96.5%) without a Covid-19 diagnosis. According to the 5-year trend, the 2020 expected rate of 30-day and 6-month all-cause mortality was 6.5% and 12.2%, while the observed incidence of death was 8.3% (p = 0.001) and 13.6% (p = 0.041), respectively. Excluding NSTEMI patients with a Covid-19 diagnosis, the 6-month mortality rate resulted in accordance with the prior 5-year trend. After multiple corrections, the presence of Covid-19 diagnosis resulted one of the independent predictors of all-cause mortality at 30 days [adjusted odds ratio (OR) 4.3; 95% confidence intervals (CI) 2.90-6.23; p < 0.0001] and 6 months (adjusted OR 3.5; 95% CI: 2.43-5.03; p < 0.0001). CONCLUSIONS: During the 2020 national outbreak in Italy, a concomitant diagnosis of Covid-19 in NSTEMI was associated with a significantly higher rate of mortality.

Impact of COVID-19 Diagnosis on Mortality in Patients with ST-Elevation Myocardial Infarction Hospitalized during the National Outbreak in Italy.

Background. We sought to assess the clinical impact of COVID-19 infection on mortality in patients with ST-elevation myocardial infarction (STEMI) admitted during the national outbreak in Italy. Methods. We analysed a nationwide, comprehensive, and universal administrative database of consecutive STEMI patients admitted during lockdown for COVID-19 infection (11 March−3 May 2020) and the equivalent periods of the previous 5 years in Italy. The observed rate of 30-day and 6-month all-cause mortality of STEMI patients with and without COVID-19 infection during the lockdown was compared with the expected rate of death, according to the trend of the previous 5 years. Results. During the study period, 32.910 STEMI hospitalizations occurred in Italy. Among these, 4048 STEMI patients were admitted during the 2020 outbreak: 170 (4.2%) with and 3878 (95.8%) without a COVID-19 diagnosis. According to the 5-year trend, the 2020 expected rates of 30-day and 6-month all-cause mortality were 9.2% and 12.6%, while the observed incidences of death were 10.8% (p = 0.016) and 14.4% (p = 0.017), respectively. Excluding STEMI patients with a COVID-19 diagnosis, the mortality rate resulted in accordance with the prior 5-year trend. After multiple corrections, the presence of COVID-19 diagnosis was an independent predictor of all-cause mortality at 30 days [adjusted odds ratio (OR) 4.5; 95% confidence intervals (CI) 3.09−6.45; p < 0.0001] and 6 months (adjusted OR 3.6; 95% CI: 2.47−5.12; p < 0.0001). Conclusions.During the 2020 national outbreak in Italy, COVID-19 infection significantly increased the mortality trend in patients with STEMI.

Lactobacilli extracellular vesicles: potential postbiotics to support the vaginal microbiota homeostasis.

BACKGROUND: Lactobacillus species dominate the vaginal microflora performing a first-line defense against vaginal infections. Extracellular vesicles (EVs) released by lactobacilli are considered mediators of their beneficial effects affecting cellular communication, homeostasis, microbial balance, and host immune system pathways. Up to now, very little is known about the role played by Lactobacillus EVs in the vaginal microenvironment, and mechanisms of action remain poorly understood. RESULTS: Here, we hypothesized that EVs can mediate lactobacilli beneficial effects to the host by modulating the vaginal microbiota colonization. We recovered and characterized EVs produced by two vaginal strains, namely Lactobacillus crispatus BC5 and Lactobacillus gasseri BC12. EVs were isolated by ultracentrifugation and physically characterized by Nanoparticle Tracking Analysis (NTA) and Dynamic Light Scattering (DLS). EVs protein and nucleic acids (DNA and RNA) content was also evaluated. We explored the role of EVs on bacterial adhesion and colonization, using a cervical cell line (HeLa) as an in vitro model. Specifically, we evaluated the effect of EVs on the adhesion of both vaginal beneficial lactobacilli and opportunistic pathogens (i.e., Escherichia coli, Staphylococcus aureus, Streptococcus agalactiae, and Enterococcus faecalis). We demonstrated that EVs from L. crispatus BC5 and L. gasseri BC12 significantly enhanced the cellular adhesion of all tested lactobacilli, reaching the maximum stimulation effect on strains belonging to L. crispatus species (335% and 269% of average adhesion, respectively). At the same time, EVs reduced the adhesion of all tested pathogens, being EVs from L. gasseri BC12 the most efficient. CONCLUSIONS: Our observations suggest for the first time that EVs released by symbiotic Lactobacillus strains favor healthy vaginal homeostasis by supporting the colonization of beneficial species and preventing pathogens attachment. This study reinforces the concept of EVs as valid postbiotics and opens the perspective of developing postbiotics from vaginal strains to maintain microbiota homeostasis and promote women's health.

Vaginal Lactobacillus Impair Candida Dimorphic Switching and Biofilm Formation.

Lactobacillus spp. generally dominate the vaginal microbiota and prevent pathogen adhesion and overgrowth, including Candida spp., by various mechanisms. Although Candida spp. can be commensal, in certain conditions they can become pathogenic, causing vulvovaginal candidiasis. The insurgence of candidiasis is related to the expression of Candida virulence factors, including morphologic switching and biofilm formation. Germ tubes, pseudohyphae, and hyphae promote Candida tissue invasion, biofilms increase persistence and are often resistant to antifungals and host immune response. Here, we explored the inhibitory activity of vaginal Lactobacillus strains belonging to Lactobacillus crispatus, Lactobacillus gasseri, Limosilactobacillus vaginalis, and Lactiplantibacillus plantarum species towards Candida virulence factors. With the aim to investigate the interrelation between mode of growth and functionality, supernatants were collected from lactobacilli planktonic cultures and, for the first time, from adherent ones, and were evaluated towards Candida dimorphic switching and biofilm. Candida biofilms were analyzed by multiple methodologies, i.e., crystal violet staining, MTT assay, and confocal microscopy. Lactobacillus supernatants reduce Candida switching and biofilm formation. Importantly, L. crispatus supernatants showed the best profile of virulence suppression, especially when grown in adherence. These results highlight the role of such species as a hallmark of vaginal eubiosis and prompt its employment in new probiotics for women's health.

The National Outcomes Evaluation Programme in Italy: The Impact of Publication of Health Indicators.

In Italy the National Outcomes Evaluation Programme, (P.N.E.) is the most comprehensive comparative evaluation of healthcare outcomes at the national level. The aim of this report is to describe the P.N.E. and some of the most relevant results achieved. The P.N.E. analysed 184 indicators on quality of care in 2015-2020 period. The data sources are the Italian Health Information Systems. The indicators reported were: proportion of surgery within 2 days after hip fracture in the elderly (HF), 30-day mortality after hospital admission for acute myocardial infarction (AMI), proportion of reoperations within 90 days of breast-conserving surgery and proportion of primary caesarean deliveries. Risk adjustment methods were used to take into account patients' characteristics. From 2010 to 2020 the proportion of interventions within 2 days after HF increased from 31.3% to 64.6%, the AMI 30-day mortality decreased from 10.4% to 8.3%, the proportion of reinterventions within 90 days of breast-conserving surgery decreased from 12.0% to 5.9% and the proportion of primary caesarean deliveries decreased from 28.4% to 22.7%. Results by area of residence showed heterogeneity of healthcare quality. We observed a general improvement in different clinical areas not always associated with a reduction of heterogeneity among areas of residence.

Estimating the Risk of Maternal Death at Admission: A Predictive Model from a 5-Year Case Reference Study in Northern Uganda.

Background: Uganda is one of the countries in the Sub-Saharan Africa with a very high maternal mortality ratio estimated at 336 deaths per 100,000 live births. We aimed at exploring the main factors affecting maternal death and designing a predictive model for estimation of the risk of dying at admission at a major referral hospital in northern Uganda. Methods: This was a retrospective matched case-control study, carried out at Lacor Hospital in northern Uganda, including 130 cases and 336 controls, from January 2015 to December 2019. Multivariate logistic regression was used to estimate the net effect of the associated factors. A cumulative risk score for each woman based on the unstandardised canonical coefficients was obtained by the discriminant equation. Results: The average maternal mortality ratio was 328 per 100,000 live births. Direct obstetric causes contributed to 73.8% of maternal deaths; the most common were haemorrhage (42.7%), sepsis (24.0%), hypertensive disorders (18.7%) and complications of abortion (2.1%), whereas malaria (23.5%) and HIV/AIDS (20.6%) were the leading indirect causes. The odds of dying were higher among women who were aged 30 years or more (OR 1.12; 95% CI, 1.04-1.19), did not attend antenatal care (OR 3.11; 95% CI, 1.36-7.09), were HIV positive (OR 3.13; 95% CI, 1.41-6.95), had a caesarean delivery (OR 2.22; 95% CI 1.13-4.37), and were referred from other facilities (OR 5.57; 95% CI 2.83-10.99). Conclusion: Mortality is high among mothers referred late from other facilities who are HIV positive, aged more than 30 years, lack antenatal care attendance, and are delivered by caesarean section. This calls for prompt and better assessment of referred mothers and specific attention to antibiotic therapy before and after caesarean section, especially among HIV-positive women.

Human Lactobacillus Biosurfactants as Natural Excipients for Nasal Drug Delivery of Hydrocortisone.

The inclusion of a chemical permeation enhancer in a dosage form is considered an effective approach to improve absorption across the nasal mucosa. Herein we evaluated the possibility of exploiting biosurfactants (BS) produced by Lactobacillus gasseri BC9 as innovative natural excipients to improve nasal delivery of hydrocortisone (HC). BC9-BS ability to improve HC solubility and the BS mucoadhesive potential were investigated using the surfactant at a concentration below and above the critical micelle concentration (CMC). In vitro diffusion studies through the biomimetic membrane PermeaPad® and the same synthetic barrier functionalized with a mucin layer were assessed to determine BC9-BS absorption enhancing properties in the absence and presence of the mucus layer. Lastly, the diffusion study was performed across the sheep nasal mucosa using BC9-BS at a concentration below the CMC. Results showed that BC9-BS was able to interact with the main component of the nasal mucosa, and that it allowed for a greater solubilization and also permeation of the drug when it was employed at a low concentration. Overall, it seems that BC9-BS could be a promising alternative to chemical surfactants in the nasal drug delivery field.

Lactobacillus Biofilms Influence Anti-Candida Activity.

Lactobacilli are the dominant members of the healthy human vaginal microbiota and represent the first defense line from pathogen infection, including vulvovaginal candidiasis. Biofilm is the predominant microbial growth form in nature, and the formation of biofilms inside the human body has important implications in health and disease. In particular, the formation of biofilm by members of the human resident microbiota is desirable, as it can improve microbial persistence and influence functionality. In the present study, we investigated the capability of 16 vaginal Lactobacillus strains (belonging to Lactobacillus crispatus, Lactobacillus gasseri, Lactobacillus vaginalis, and Lactobacillus plantarum species) to form biofilms, and we correlated their mode of growth to anti-Candida activity. L. plantarum strains were the best biofilm producers, and high variability was registered in the level of biofilm formation among L. crispatus and L. gasseri strains. Culture supernatants derived from Lactobacillus biofilm and planktonic growth were tested toward a panel of Candida clinical isolates (Candida albicans, Candida glabrata, Candida lusitaniae, Candida tropicalis, Candida krusei, and Candida parapsilosis) and their metabolome assessed by 1H-NMR. L. crispatus and L. plantarum strains exhibited the best fungistatic profile, and biofilms enhanced their anti-Candida activity; on the contrary, L. gasseri strains were more effective when grown in a planktonic mode. Biofilm/planktonic mode of growth also affects Lactobacillus metabolism, mainly influencing nitrogen and amino acid pathways, and anti-Candida activity is instead strictly related to carbohydrate metabolism. The present study underlined the strict interdependence between microbial mode of growth, metabolism, and functional properties. Biofilm formation by members of the healthy human microbiota represents a crucial issue in the field of microbial physiology and host-microbiota interactions, beyond supporting the development of new antimycotic strategies based on probiotics grown in adherence.

Influence of Lactobacillus Biosurfactants on Skin Permeation of Hydrocortisone.

One of the most widely used strategies to improve drug diffusion through the skin is the use of permeation enhancers. The aim of this work was to investigate the effect of two biosurfactants (BS), produced by Lactobacillus crispatus BC1 and Lactobacillus gasseri BC9, on the skin permeation profile of hydrocortisone (HC, model drug). HC aqueous solubility and in vitro diffusion studies through porcine skin were performed in the presence of BC1-BS and BC9-BS at concentrations below and above critical micellar concentrations (CMC). Moreover, skin hydration tests and differential scanning calorimetry (DSC) analysis were performed to further investigate BS interaction with the outermost layer of the skin. Both BS increased HC solubility, especially at concentrations above their CMC. At concentrations below the CMC, drug permeation through the skin was improved, as the result of a dual effect: a) the formation of a superficial lipophilic environment, as confirmed by the reduction in skin hydration and b) the interaction between BS and the stratum corneum (SC), as demonstrated by the DSC curves. From the obtained data, it appears that BC1-BS and BC9-BS could represent new promising green excipients for drug permeation enhancement through the skin.

Anti-Candida Activity of Hyaluronic Acid Combined with Lactobacillus crispatus Lyophilised Supernatant: A New Antifungal Strategy.

Vulvovaginal candidiasis (VVC) and recurrencies are common in reproductive-aged women. The emergence of Candida strains resistant to conventional antimycotic drugs prompted the search for alternative therapies. Hyaluronic acid (HA), a uniform and linear glycosaminoglycan, has been proposed as an anti-Candida agent. Vaginal lactobacilli and their derivatives, including cell free culture supernatants (CFS), represent potential strategies for the treatment of Candida infections. In the present paper, the anti-Candida potential of HA and lyophilised CFS (LCFS), obtained from the vaginal strain Lactobacillus crispatus BC5, was investigated. HA and LCFS proved to be active towards a panel of clinical Candida isolates belonging to different species in a dose dependent manner and their association maintained the antifungal activity. Notably, also Candida species generally resistant to conventional antifungals resulted sensitive. A vaginal matrix based on microcrystalline cellulose and containing effective doses of both agents was developed and characterised. This vaginal formulation showed mucoadhesive ability and almost abrogated Candida albicans growth. In conclusion, HA and LCFS from L. crispatus BC5 are thus good candidates to design a new therapeutic strategy to counteract VVC, and the proposed vaginal matrix represents a promising prototype.

Lactobacillus crispatus BC1 Biosurfactant Counteracts the Infectivity of Chlamydia trachomatis Elementary Bodies.

Lactobacilli-derived biosurfactants (BS) have shown promising effects as antimicrobial molecules. Since Lactobacillus crispatus plays a crucial role in maintaining vaginal eubiosis, BS from this species could represent novel therapeutic agents to counteract sexually transmitted pathogens, such as Chlamydia trachomatis (CT). The aim of the present study was to assess the inhibitory effects of a BS produced by the vaginal strain L. crispatus BC1 on the infectivity of CT elementary bodies (EBs). For concentrations ranging between 1 and 0.5 mg/mL at 60-min contact time, L. crispatus BC1 BS displayed a highly significant anti-CT activity, with about 50% reduction of EB infectivity towards HeLa cells. To identify the components responsible for chlamydial inhibition, a panel of selected fatty acids, including those present in BS lipopeptidic structure, was tested against CT EBs. Pentadecanoic acid, myristic acid, ß-hydroxy-myristic acid, and ß-hydroxy-palmitic acid were able to significantly reduce EBs infectivity up to 5-0.5 µg/mL, concentrations that resulted to be non-toxic for HeLa cells. These data can contribute to the understanding of the biological role of lactobacilli in the vaginal niche, as well as to promote the application of their produced BS as an innovative and antibiotic-sparing anti-chlamydial strategy.