Effect of hydrogen peroxide on the oxidative burst of neutrophils in pigs and ruminants

Background and Aim: Neutrophils represent between 20% and 75% of white blood cells in animals and play a key role in an effective immune response. The generation of reactive oxygen species (ROS) is commonly referred to as an oxidative burst and is crucial under healthy and disease conditions. Interestingly, ROS are emerging as regulators of several neutrophil functions, including their oxidative burst. The present study aimed to investigate the effect of hydrogen peroxide on the oxidative burst of neutrophils, collected from domestic animal species (namely, pig, cattle, and sheep), and exposed to different stimuli. Materials and Methods: A total of 65 slaughtered animals were included in the present study: Twenty-two pigs, 21 cattle, and 22 sheep. Blood samples were collected at bleeding and neutrophils were then purified using ad hoc developed and species-specific protocols. Neutrophils were treated with hydrogen peroxide at micromolar-to-millimolar concentrations, alone, or combined with other stimuli (i.e., opsonized yeasts, and phorbol 12-myristate 13-acetate). The generation of ROS was evaluated using a luminol-derived chemiluminescence (CL) assay. For each animal species, data were aggregated and reported as mean area under curve±standard deviation. Finally, data were statistically analyzed by one-way ANOVA, followed by Tukey's post hoc test. Results: Exposure of bovine and ovine neutrophils to hydrogen peroxide alone resulted in a dose-dependent enhancement of the CL response, which was significantly stronger at its highest concentration and proved particularly prominent in sheep. Opsonized yeasts and phorbol 12-myristate 13-acetate both proved capable of stimulating the generation of ROS in all animal species under study. Hydrogen peroxide negatively modulated the oxidative burst of neutrophils after exposure to those stimuli, observed response patterns varying between pigs and ruminants. Porcine neutrophils, pre-exposed to micromolar concentrations of hydrogen peroxide, showed a decreased CL response only to opsonized yeasts. Conversely, pre-exposure to hydrogen peroxide reduced the CL response of ruminant neutrophils both to yeasts and phorbol 12-myristate 13-acetate, the effect being most prominent at 1 mM concentration. Conclusion: These results indicate that hydrogen peroxide is capable of modulating the oxidative bursts of neutrophils in a species-specific and dose-dependent manner, substantial differences existing between pigs and ruminants. Further investigation is required to fully comprehend such modulation, which is crucial for the proper management of the generation of ROS under healthy and disease conditions

In situ observation of stress relaxation in epitaxial graphene

Upon cooling, branched line defects develop in epitaxial graphene grown at high temperature on Pt(111) and Ir(111). Using atomically resolved scanning tunneling microscopy we demonstrate that these defects are wrinkles in the graphene layer, i.e. stripes of partially delaminated graphene. With low energy electron microscopy (LEEM) we investigate the wrinkling phenomenon in situ. Upon temperature cycling we observe hysteresis in the appearance and disappearance of the wrinkles. Simultaneously with wrinkle formation a change in bright field imaging intensity of adjacent areas and a shift in the moire spot positions for micro diffraction of such areas takes place. The stress relieved by wrinkle formation results from the mismatch in thermal expansion coefficients of graphene and the substrate. A simple one-dimensional model taking into account the energies related to strain, delamination and bending of graphene is in qualitative agreement with our observations.Comment: Supplementary information: S1: Photo electron emission microscopy and LEEM measurements of rotational domains, STM data of a delaminated bulge around a dislocation. S2: Movie with increasing brightness upon wrinkle formation as in figure 4. v2: Major revision including new experimental dat

Testing a model of pacific oysters’ (Crassostrea gigas) growth in the adriatic sea: Implications for aquaculture spatial planning

Assessing the potential biomass yield is a key step in aquaculture site selection. This is challenging, especially for shellfish, as the growth rate depends on both trophic status and water temperature. Individual ecophysiological models can be used for mapping potential shellfish growth in coastal areas, using as input spatial time series of remotely sensed SST and chlorophyll-a. This approach was taken here to estimate the potential for developing oyster (Crassostrea gigas) farming in the western Adriatic Sea. Industry relevant indicators (i.e., shell length, total individual weight) and days required to reach marketable size were mapped using a dynamic energy budget model, finetuned on the basis of site-specific morphometric data collected monthly for a year. Spatially scaled-up results showed that the faster and more uniform growth in the northern Adriatic coastal area, compared with the southern one, where chlorophyll-a levels are lower and summer temperatures exceed the critical temperature limit for longer periods. These results could be used in planning the identification of allocated zones for aquaculture, (AZA), taking into account also the potential for farming or co-farming C. gigas. In perspective, the methodology could be used for getting insights on changes to the potential productivity indicators due to climatic changes

Congenital Oral Squamous Cell Carcinoma in a Suckling Piglet

A 3-week-old suckling piglet spontaneously died after septicemic colibacillosis. At postmortem examination, bulging and ulcerated lesions were seen, affecting the oral mucosa on the inner surface of the lower lip. After histopathological investigation, the diagnosis of congenital oral squamous cell carcinoma was made. To the best of our knowledge, this is the first case of congenital oral squamous cell carcinoma ever described. A relationship has been shown or suggested between papillomavirus infection and oral squamous cell carcinoma in humans and animals. However, next-generation sequencing study did not demonstrate any papillomavirus sequences in the case reported herein

Occurrence, antimicrobial susceptibility, and pathogenic factors of Pseudomonas aeruginosa in canine clinical samples

Background and Aim: Pseudomonas aeruginosa is a relevant opportunistic and difficult to treat pathogen due to its widespread environmental diffusion, intrinsic resistance to many classes of antimicrobials, high ability to acquire additional resistance mechanisms, and wide range of pathogenic factors. The present study aimed to investigate the prevalence of P. aeruginosa in canine clinical samples, the antimicrobial susceptibility against antipseudomonal antibiotics, and the presence of extracellular pathogenic factors of the isolates, as well as their ability to produce biofilm. Materials and Methods: Overall, 300 clinical specimens from dogs with pyoderma or abscesses (n=58), otitis (n=59), and suspected bladder infection (n=183) were analyzed by standard bacteriological methods. P. aeruginosa isolates were tested for their antimicrobial susceptibility by disk and gradient diffusion methods to determine the minimum inhibitory concentrations. The ability of the isolates to produce biofilm was investigated by a microtiter plate assay, while virulence genes coding for elastase (lasB), exotoxin A (toxA), alkaline protease (aprA), hemolytic phospholipase C (plcH), and exoenzyme S (ExoS) were detected by polymerase chain reaction method. Results: A total of 24 isolates of P. aeruginosa were found in clinical specimens (urine n=3, skin/soft tissue n=6, and ear canal n=15). No resistance was found to ceftazidime, gentamicin, aztreonam, and imipenem (IMI), while low levels of resistance were found to enrofloxacin (ENR) (4.2%) and piperacillin-tazobactam (8.3%). However, 41.7% and 29.2% of the isolates showed intermediate susceptibility to ENR and IMI, respectively. Disk and gradient diffusion methods showed high concordance. The majority of the isolates revealed a weak (33.3%) or intermediate (45.8%) ability to form biofilm, while the strong biofilm producers (20.8%) derived exclusively from the ear canal samples. All isolates (100%) were positive for lasB, aprA, and plcH genes, while exoS and toxA were amplified in 21 (87.5%) and 22 (91.7%) isolates, respectively. Conclusion: In the present study, P. aeruginosa isolates from canine clinical samples were characterized by low levels of antimicrobial resistance against antipseudomonal drugs. However, the high presence of isolates with intermediate susceptibility for some categories of antibiotics, including carbapenems which are not authorized for veterinary use, could represent an early warning signal. Moreover, the presence of isolates with strong ability to produce biofilm represents a challenge for the interpretation of the antimicrobial susceptibility profile. In addition, the high prevalence of the extracellular pathogenic factors was indicative of the potential virulence of the isolates

Aspergillus section fumigati pneumonia and oxalate nephrosis in a foal

Equine pulmonary aspergillosis is a rare deep mycosis often due to the hematogenous spread of hyphae after gastrointestinal tract disease. We describe herein the main clinic-pathological findings observed in a foal, which spontaneously died after showing diarrhea and respiratory distress. Necropsy and histopathological investigations allowed to diagnose pulmonary aspergillosis, which likely developed after necrotic typhlitis-colitis. Biomolecular studies identified Aspergillus section Fumigati strain as the causative agent. Notably, severe oxalate nephrosis was concurrently observed. Occasionally, oxalate nephropathy can be a sequela of pulmonary aspergillosis in humans. The present case report suggests that the renal precipitation of oxalates can occur also in horses affected by pulmonary aspergillosis and could likely contribute to the fatal outcome of the disease

Vibrio parahaemolyticus control in mussels by a Halobacteriovorax isolated from the Adriatic sea, Italy

This study evaluated the application of a Halobacteriovorax isolated from water of the Adriatic Sea (Italy) in controlling V. parahaemolyticus in mussels (Mytilus galloprovincialis). Two 72 h laboratory-scale V. parahaemolyticus decontamination experiments of mussels were performed. The test microcosm of experiment 1 was prepared using predator/prey free mussels experimentally contaminated with Halobacteriovorax/V. parahaemolyticus at a ratio of 103 PFU/105 CFU per ml, while that of experiment 2 using mussels naturally harbouring Halobacteriovorax that were experimentally contaminated with 105 CFU per ml of V. parahaemolyticus. For experiment 1, was also tested a control microcosm only contaminated with 105 CFU per ml of V. parahaemolyticus. Double layer agar plating and pour plate techniques were used to enumerate Halobacteriovorax and V. parahaemolyticus, respectively. 16 S rRNA analysis was used to identify Halobacteriovorax. For both experiments in the test microcosm the concentration of prey remained at the same level as that experimentally added, i.e. 5 log for the entire analysis period. In experiment 1, V. parahaemolyticus counts in mussels were significantly lower in the test microcosm than the control with the maximum difference of 2.2 log at 24 h. Results demonstrate that Halobacteriovorax can modulate V. parahaemolyticus level in the mussels. The public impact of V. parahaemolyticus in bivalves is relevant and current decontamination processes are not always effective. Halobacteriovorax is a suitable candidate in the development of a biological approach to the purification of V. parahaemolyticus in mussels

Insights into the oral bacterial microbiota of sows

The investigation of bacterial microbiota represents a developing research field in veterinary medicine intended to look for correlations between animal health and the balance within bacterial populations. The aim of the present work was to define the bacterial microbiota of the oral cavity of healthy sows, which had not been thoroughly described so far. In total, 22 samples of oral fluid were collected and analyzed by 16S-rRNA gene sequencing. CLC Genomics Workbench 20.0 (QIAGEN Digital Insights, Aarhus, Denmark) was then used to examine the results. The predominant orders were Lactobacillales, Clostridiales, and Corynebacteriales. Lactobacillaceae, Corynebacteriaceae, Moraxellaceae, Aerococcaceae, and Staphylococcaceae were the most represented families. As regards the most abundant genera, Lactobacillus, Corynebacterium, Acinetobacter, Staphylococcus, Rothia, Aerococcus, and Clostridium can be pointed out as the bacterial core microbiota. Sows were also divided into “gestating” and “lactating” groups, and mild differences were found between pregnant and lactating sows. The data herein described represent an original contribution to the knowledge of the porcine bacterial microbiota. Moreover, the choice of sows as experimental animals was strategic for identifying the adult microbial community. These data provide a basis for further studies on the oral bacterial microbiota of pigs

Sequence homology at the breakpoint and clinical phenotype of mitochondrial DNA deletion syndromes

Mitochondrial DNA (mtDNA) deletions are a common cause of mitochondrial disorders. Large mtDNA deletions can lead to a broad spectrum of clinical features with different age of onset, ranging from mild mitochondrial myopathies (MM), progressive external ophthalmoplegia (PEO), and Kearns-Sayre syndrome (KSS), to severe Pearson syndrome. The aim of this study is to investigate the molecular signatures surrounding the deletion breakpoints and their association with the clinical phenotype and age at onset. MtDNA deletions in 67 patients were characterized using array comparative genomic hybridization (aCGH) followed by PCR-sequencing of the deletion junctions. Sequence homology including both perfect and imperfect short repeats flanking the deletion regions were analyzed and correlated with clinical features and patients\u27 age group. In all age groups, there was a significant increase in sequence homology flanking the deletion compared to mtDNA background. The youngest patient group (\u3c6 years old) showed a diffused pattern of deletion distribution in size and locations, with a significantly lower sequence homology flanking the deletion, and the highest percentage of deletion mutant heteroplasmy. The older age groups showed rather discrete pattern of deletions with 44% of all patients over 6 years old carrying the most common 5 kb mtDNA deletion, which was found mostly in muscle specimens (22/41). Only 15% (3/20) of the young patients (\u3c6 years old) carry the 5 kb common deletion, which is usually present in blood rather than muscle. This group of patients predominantly (16 out of 17) exhibit multisystem disorder and/or Pearson syndrome, while older patients had predominantly neuromuscular manifestations including KSS, PEO, and MM. In conclusion, sequence homology at the deletion flanking regions is a consistent feature of mtDNA deletions. Decreased levels of sequence homology and increased levels of deletion mutant heteroplasmy appear to correlate with earlier onset and more severe disease with multisystem involvement. © 2010 Sadikovic et al