Is Oral Vaccination against Escherichia coli Influenced by Zinc Oxide?

BACKGROUND: Although zinc oxide has been banned at therapeutic doses in the EU, its use is still legal in most countries with industrial pig farming. This compound has been shown to be very effective in preventing E. coli-related diseases. However, another strategy used to control this pathogen is vaccination, administered parenterally or orally. Oral vaccines contain live strains, with F4 and F18 binding factors. Since zinc oxide prevents E. coli adhesion, it is hypothesised that its presence at therapeutic doses (2500 ppm) may alter the immune response and the protection of intestinal integrity derived from the vaccination of animals. METHODS: A group of piglets were orally vaccinated at weaning and divided into two subgroups; one group was fed a feed containing 2500 ppm zinc oxide (V + ZnO) for the first 15 days post-vaccination (dpv) and the other was not (V). Faeces were sampled from the animals at 6, 8, 11, 13, and 15 dpv. Unvaccinated animals without ZnO in their feed (Neg) were sampled simultaneously and, on day 15 post-vaccination, were also compared with a group of unvaccinated animals with ZnO in their feed (ZnO). RESULTS: Differences were found in E. coli excretion, with less quantification in the V + ZnO group, and a significant increase in secretory IgA in the V group at 8 dpv, which later equalised with that of the V + ZnO group. There was also some difference in IFNα, IFNγ, IL1α, ILß, and TNFα gene expression when comparing both vaccinated groups (p < 0.05). However, there was no difference in gene expression for the tight junction (TJ) proteins responsible for intestinal integrity. CONCLUSIONS: Although some differences in the excretion of the vaccine strain were found when comparing both vaccinated groups, there are no remarkable differences in immune stimulation or soluble IgA production when comparing animals orally vaccinated against E. coli in combination with the presence or absence of ZnO in their feed. We can conclude that the immune response produced is very similar in both groups.

Effect of ß-Mannanase Addition during Whole Pigs Fattening on Production Yields and Intestinal Health.

The presence of ß-mannans in feed can produce a futile and chronic immune stimulation in fattening pigs. In this trial, a 1-4-endo-D-ß-mannanase was added to the feed (HC) during growth and fattening (0.03% of Hemicell HT) and physical performance and pathological data were recorded, and intestinal integrity and immune activation were studied by molecular biomarkers, compared to a control group (CON). The treatment diet was reduced in energy content by 40 Kcal/kg NE. From each group, 113 and 112 animals housed in 8 pens were individually identified and weighed three times: at 7th, 63rd and 116th days in feed. The FCR was calculated for groups of two pens and ADG individually. There was no difference in ADG (CON = 0.836, HC = 0.818) nor in FCR between groups (p = 0.486). During growth, there was a higher frequency of normal feces in HC and there were also no differences in the frequency of gastric lesions. A significant increase in Claudin, Occludin, IFN-γ and IL8 was observed in the CON in feces and a significant decrease in IL-6 in HC. In tissues, there were differences for IL-12p40, TNF-alpha in jejunum (increased CON) and TGF-ß in ileum and jejunum, (decreased HC). The economic performance was EUR 4.7 better in the treated group. In conclusion, the addition of 1-4-endo-D-ß--mannanase to the feed with a 1.6% reduction in net energy compared to the control, allowed the animals to perform as well as the animals on the higher energy diet, with lower prevalence of diarrhea.

Oral and Parenteral Vaccination against Escherichia coli in Piglets Results in Different Responses.

The available E. coli vaccines involve two main types (inactivated and live non-pathogenic) and two routes of administration (oral and parenteral) but the mechanism by which both vaccines and routes of administration work is not yet fully elucidated. The influence of a parenteral vaccine (PV) and an oral one (OV) was studied by analyzing the gene expression of biomarkers indicating cellular infiltration (calprotectin, CAL), tight junction proteins (occludin OCL, and zonulin ZON) that maintain intestinal paracellular integration and two proinflammatory (IFN-γ) and anti-inflammatory (TGF-ß) mediator cytokines, as well as histomorphology and IgA production cell density. Differences were observed in CAL, more infiltrated in orally vaccinated animals; OCL also increased in orally vaccinated animals, and higher density of IgA-producing cells in ileum for orally vaccinated groups. Cytokine expression is also different; and there is a lower mRNA for IFN-γ in the parenteral than in the oral vaccinated animals. Finally, the villus height-to-crypt depth ratio was higher in the orally vaccinated groups. The data collectively show clear and different effects derived from the use of each type of vaccine, route of administration and regimen. The results suggest a more rapid and direct effect of oral vaccination and a state of suppression in the absence of a second oral stimulus by the pathogen.

Real-world treatment patterns and clinical outcomes in patients with AML in Japan who were ineligible for first-line intensive chemotherapy.

Acute myeloid leukemia (AML) predominantly affects elderly adults, and its prognosis worsens with age. Treatment options for patients in Japan ineligible for intensive chemotherapy include cytarabine/aclarubicin ± granulocyte colony-stimulating factor (CA ± G), azacitidine (AZA), low-dose cytarabine (LDAC), targeted therapy, and best supportive care (BSC). The country's aging population and the evolving treatment landscape are contributing to a need to understand treatment pathways and associated outcomes. This retrospective chart review evaluated outcomes in patients across Japan with primary/secondary AML who were ineligible for intensive chemotherapy and began first-line treatment or BSC between 01/01/2015 and 12/31/2018. The primary endpoint was overall survival (OS); secondary endpoints included progression-free survival (PFS) and healthcare resource utilization (HRU). Of 199 patients (58% > 75 years), 121 received systemic therapy (38 CA ± G, 37 AZA, 7 LDAC, 39 other) and 78 received BSC. Median OS was 5.4, 9.2, 2.2, 3.8, and 2.2 months for CA ± G, AZA, LDAC, other systemic therapy, and BSC, respectively; median PFS was 3.4, 7.7, 1.6, 2.3, and 2.1 months, respectively. HRU rates were uniformly high, with > 80% patients hospitalized in each cohort. The poor clinical outcomes and high HRU among Japanese AML patients who are ineligible for intensive chemotherapy highlight an unmet need for novel therapies.

Healthcare resource utilization trends in patients with acute myeloid leukemia ineligible for intensive chemotherapy receiving first-line systemic treatment or best supportive care: A multicenter international study.

OBJECTIVES: This retrospective chart review examined real-world healthcare resource utilization (HRU) in patients with AML ineligible for intensive therapy who received first-line systemic therapy or best supportive care (BSC). METHODS: Data were collected anonymously on patients with AML who initiated first-line hypomethylating agents (HMA), low-dose cytarabine (LDAC), other systemic therapy, or BSC. HRU endpoints included hospitalizations, outpatient consultations, transfusions, and supportive care. RESULTS: Of 1762 patients included, 46% received HMA, 11% received LDAC, 17% received other systemic therapy, 26% received BSC; median treatment durations were 118, 35, 33, and 57 days, respectively. Most patients were hospitalized, most commonly for treatment administration, transfusion, or infection (HMA 82%, LDAC 93%, other systemic therapy 83%, BSC 83%). A median number of hospitalizations were 2-6 across systemic groups and two for BSC, with median durations of 8-18 days. Transfusion rates and outpatient consultations were highest for HMA (80% and 79%) versus LDAC (57% and 53%), other systemic therapy (57% and 63%), and BSC (71% and 66%). Antivirals/antibiotics and antifungals were used more frequently than growth factors (72-92%, 34-63%, and 7-27%, respectively). CONCLUSION: Patients with AML ineligible for intensive therapy have high HRU; novel therapies are needed to alleviate this burden.

Real-world treatment patterns and clinical outcomes in patients with AML unfit for first-line intensive chemotherapy.

Acute myeloid leukemia (AML) predominantly affects the elderly, and prognosis declines with age. Induction chemotherapy plus consolidation therapy is standard of care for fit patients; options for unfit patients include hypomethylating agents (HMA), low-dose cytarabine (LDAC), targeted therapies, and best supportive care (BSC). This retrospective chart review evaluated clinical outcomes in unfit patients with AML who initiated first-line treatment or BSC 01/01/2015-12/31/2018. Overall survival (OS), progression-free survival (PFS), time-to-treatment failure (TTF), and response rates were assessed. Of 1762 patients, 1310 received systemic therapies: 809 HMA, 199 LDAC, and 302 other therapies; 452 received BSC. Median OS was 9.9, 7.9, 5.4, and 2.5 months for HMA, LDAC, other, and BSC, respectively. Median PFS was 7.5, 5.3, 4.1, and 2.1 months for HMA, LDAC, other, and BSC, respectively; median TTF was 4.9, 2.1, 2.2, and 2.1 months, respectively. Our findings highlight the unmet need for novel therapies for unfit patients.

Field validation of biocontrol strategies to control brown rot on stone fruit in several European countries.

BACKGROUND: Brown rot caused by Monilinia spp. is the most significant disease of stone fruit. New approaches to fruit production have necessitated the development of control strategies that are more eco- and consumer-friendly. An efficient field strategy to control brown rot was previously designed based on the application of two biocontrol agents (BCAs), Bacillus amyloliquefaciens CPA-8 (CPA-8) or Penicillium frequentans 909 (Pf909), with calendar-based treatment. In the present study, the strategy was validated on different stone fruit hosts in four producing countries over two seasons. RESULTS: The results obtained were reported according to three different scenarios: Scenario 1, in which there was no presence of disease in the field; Scenario 2, in which high disease pressure occurred in the field and treatments (biologicals or chemicals) were not effective; and Scenario 3, with low or medium to high disease presence. The results were successful because, in general, BCA strategies were shown to control brown rot to a similar extent as chemicals strategies. We found that most of the trials conducted in this study were classed under Scenario 3 (62.5%), with only 12.5% and 25% of the trials classed under Scenarios 1 and 2, respectively. CONCLUSION: These novel findings allowed the formulation of CPA-8 and Pf909 as valuable tools for farmers to produce stone fruits more competitively and meet consumer demand for safer and more environmentally friendly products. © 2021 Society of Chemical Industry.

Balance between resilient fruit surface microbial community and population of Monilinia spp. after biopesticide field applications of Penicillium frequentans.

The microbial variability on the host plant surface must be maintained because population diversity and quantity are essential to avoid disease development. It would be necessary to examine the patterns and mechanisms associated with the massive and reiterative introduction of a microbial pest control agent. The effect of inundative releases of biopesticide formulations containing Penicillium frequentans for the control of Monilinia spp. populations, and the effect on fruit surface microbiota on 18 stone fruit field experiments located in four European countries for more than two crop seasons against brown rot were studied. P. frequentans was monitored after application in order to assess whether it was persistent or not in the environment. Hydrolysis of fluorescein diacetate and denaturing gradient gel electrophoresis were used to study the effects of P. frequentans on fungal and bacterial non-target populations on fruit surface. The effect of P. frequentans formulations on the populations of Monilinia spp. on fruit was also assessed in different orchards. P. frequentans population on stone fruit surfaces showed ranged from 100 to 10,000 CFU cm-2, and postharvest recovered populations were more than 10-100-fold higher than preharvest recovered populations. The population of P. frequentans varied among orchards and years, rather than by the type of formulation. P. frequentans formulation reduced Monilinia spp. population and brown rot and latent infections caused by this pathogen both before and at harvest, while stabilizing or increasing antagonist populations and avoiding non-target microorganisms. However, fungicides reduced significantly the microbial activity on nectarine surfaces.

Dispersion, persistence, and stability of the biocontrol agent Penicillium frequentans strain 909 after stone fruit tree applications.

The capacity of dispersion, persistence, and stability from biocontrol agents is essential before these organisms can be developed into a commercial product. Interactions that microorganisms establish with stone fruit trees may be beneficial in the exploitation of trees in agriculture as crop production. The natural background levels of Penicillium frequentans strain 909 dispersion, persistence, and stability were assessed after tree applications and postharvest conditions. A fingerprinting-based approach to trace genetic stability of P. frequentans along stored time and its release in the field was developed. P. frequentans was successfully traced and discriminated. This strain was dispersed well in treated trees, persisting in the ecosystem up to 2 weeks and staying genetically stable after 36 months of storage. However, the dispersal of P. frequentans was very limited on around untreated trees and soil. P. frequentans dispersed randomly into the air, and its presence reduced from the first day to disappear completely at 15-21 days after application. Great losses of P. frequentans and its increased dispersal in open field conditions probably resulted from rainfall. Biological control strategies with Pf909 were discussed.

Surfactant effects on wettability of Penicillium frequentans formulations to improve brown rot biocontrol.

BACKGROUND: Penicillium frequentans can be used in the management of brown rot caused by Monilinia spp. Competition is the primary mode of biocontrol activity of P. frequentans, which must therefore cover most of fruit surface to avoid pathogen infection. Our objective was to optimize the efficacy of P. frequentans by maximizing fruit surface coverage and retention with the antagonist formulation by surfactant incorporation. RESULTS: Sixteen surfactants were assessed for the management of brown rot at 3-5 different concentrations. Nine surfactants increased the droplet surface up to 2.5 times compared with water on an inert surface, with or without the presence of P. frequentans in each drop. Eight surfactants increased P. frequentans on the fruit surface, enhancing colony forming units after run off or lateral spray application uptake by 50% compared to the control without surfactants. But only some doses of sodium carboxymethyl cellulose, gelatin, Tween 20, sorbitan alkyl esters, synthetic latex, polyethylene glycol isotridecyl ether, and hydroxypropyl methylcellulose could show the same covered fruit surface after run off or lateral spray application. There were also no phytotoxic side-effects on five different species of stone fruit. CONCLUSIONS: The efficacy of P. frequentans dry conidia can be enhanced by optimizing the composition of the formulation with surfactants. © 2018 Society of Chemical Industry.

Adaptive conditions and safety of the application of Penicillium frequentans as a biocontrol agent on stone fruit.

Penicillium frequentans (Pf909) reduces brown rot caused by Monilinia spp. in stone fruit. The registration of a microbial biocontrol agent in Europe requires information on the risks and safety of a biological product. This study focused on the impact of the physical environment on Pf909 survival and growth, Pf909 mycotoxin production on fruit surface, and the Pf909 resistance to commercial antifungal compounds used in animal and human medicine. The effect of temperature (4 to 37°C), water activity (0.999 to 0.900), pH (3 to 11), light intensity (0, 90 and 180lm) and photoperiod (0/24, 12/12, 16/8, 24/0; light/dark) on mycelial growth and sporulation of Pf909 were monitored for 10days in vitro on culture medium. Antifungal activity of antifungal compounds on mycelial growth of Pf909 was also measured by a quantitative micro spectrophotometric assay after 72h of incubation. The presence or absence of four non-volatile mycotoxins (patulin, penicillic acid, ochratoxin A and citrinin) on nectarine surfaces treated with Pf909 was determined by HPLC. Growth rate was significantly influenced by water activity, temperature and light exposure conditions. Pf909 showed maximum growth and sporulation at 22°C and 25°C, in wet conditions (0.999 water activity), with a pH5.6 to 9, and in darkness or a short light photoperiod. Our results showed all antifungal compounds used reduced significantly Pf909 mycelial growth at tested commercial doses. HPLC data analysis showed that 7days after biofungicide (Pf909) application there were no mycotoxin products on the surface of nectarine. Finally, no phylogenetic relationship has been shown among Pf909 and other species of Penicillium that produce mycotoxins. In conclusion, from an ecological point of view, Pf909 would establish and survive actively over a broad range of climatic conditions. The probability of risks to human and animal health is considered very low.