Cellular localization of Visudyne® as a function of time after local injection in an in vivo model of squamous cell carcinoma: an investigation into tumor cell death

Objective To evaluate the effects of time on cellular localization of Visudyne^® after local injection. Animals Twenty athymic nude mice. Procedures A squamous cell carcinoma (SCC) cell line (A‐431) was injected into right and left dorsolumbar subcutaneous tissue of each mouse, representing treatment (T) and control (C) tumors. In experiment 1 (Exp 1; n = 10) and 2 (Exp 2; n = 10), the T tumors received a local injection of Visudyne^® (0.1 mg/cm³), and C tumors received an equal dose of 5% dextrose in water (D5W). Mice were randomly subdivided into two groups (A and B; n = 5 per group). Mice in Exp 1A and B were sacrificed 1 and 30 min after local injection, respectively. Experiment 1A and B tumors were evaluated by fluorescence microscopy to determine drug localization. Experiment 2A and B tumors were exposed to LED illumination 1 and 30 min after injection, respectively, and evaluated by transmission electron microscopy (TEM) to determine ultrastructural tumor cell damage. Results Fluorescence was detected within the cytoplasm of T tumors in both Exp 1A and B. Significance was detected in fluorescence intensity between T1 min vs. T30 min (P = 0.03) and between T1 min and C1 min tumors (P = 0.01), respectively. Tumors in Exp 2A and B demonstrated evidence of apoptotic cell death. Conclusions Fluorescence microscopy demonstrated higher Visudyne^® concentration within SCC cytoplasm of 1 min compared with 30‐min tumors. Transmission electron microscopy results revealed that tumors treated by photodynamic therapy (PDT) within 30 min of local injection undergo cellular apoptosis.     

Proximal mandibular nerve block, using electrolocation, for rostral mandibulectomy in a geriatric dog

We describe a case of proximal mandibular nerve block with ropivacaine, using electrolocation, for perioperative pain management in a geriatric dog undergoing rostral mandibulectomy. The patient did not require intraoperative analgesia or analgesic supplementation for 8 h after the end of the surgery.     

Idiopathic sterile pyogranuloma in three domestic cats

Pyogranulomatous inflammation has been extensively described in cats, in particular in cases of feline infectious peritonitis and also associated with Mycobacteria, Actinomyces, Nocardia, Rhodococcus and fungal infections. Idiopathic sterile pyogranulomatous dermatitis has also been described. In this case series we describe the clinical presentation, histopathology and outcome of three cases of feline idiopathic sterile steroid-responsive pyogranuloma with different presentation and different locations of the lesion, but with the common feature of having a mass with no superficial skin involvement.     

Phenotypic diversity and evolution of farmer varieties of bread wheat on organic farms in Europe

The contribution of farmers to the creation and maintenance of genetic diversity is beginning to receive more recognition in developed countries. Although the use of landraces and historic varieties has largely disappeared in countries with industrialized agricultural systems and formal seed markets, certain varieties with particular cultural significance have been continuously cultivated by farmers and other varieties have been preserved ex situ in genebanks. Recently, associations of organic farmers have become involved in the conservation and use of landraces and historic varieties (called farmer varieties in this article) because these varieties possess agronomic and quality traits that they have not found in modern varieties. In this study, eight farmer varieties of bread wheat from Europe selected by participating farmers were evaluated in on-farm trials during 3 years of cultivation. Each variety was grown on each farm, and phenotypic changes in each variety were measured the third year in a replicated split-plot trial on each farm comparing the version of each variety the farmer had multiplied to a sample of each variety from the region of origin. All varieties, including the two modern pureline varieties used as checks, showed statistically significant phenotypic changes over 3 years of multiplication. However, the magnitude of these changes was moderate and did not call into question varietal identity or distinctness. In addition, some traits of putative agronomic and adaptive importance, such as grain weight per spike and thousand kernel weight, did not respond positively to natural selection (environmental conditions and management practices) which suggests the necessity of farmer selection to maintain and improve varietal performance.     

Mixtures of organic amendments and biochar promote beneficial soil microbiota and affect Fusarium oxysporum f. sp. lactucae,Rhizoctonia solani and Sclerotinia minor disease suppression

The use of soil fumigants and fungicides to control soilborne pathogens is reduced due to awareness of their negative drawbacks. Long-term application of such agrochemicals negatively affects soil microbiota and reduces natural soil suppressiveness. We investigated long-term impacts of biochar and not-pyrogenic organic amendment (OAs: manure, alfalfa straw and glucose), on disease suppression compared with conventional management in three pathosystems: Fusarium oxysporum f. sp. lactucae (FOL)–lettuce, Rhizoctonia solani–tomato and Sclerotinia sclerotiorum–lettuce, by conditioning soil for 2 years. Soil analyses included pH, electrical conductivity, organic carbon, total nitrogen, C:N ratio, N-NO₃⁻, N-NH₄⁺, cation exchange capacity, available phosphorus, Ca²⁺, Mg²⁺, K⁺, Na⁺, fluorescein diacetate (FDA), and total and active limestone. Soil microbiota was characterized by combining BIOLOG EcoPlates with next-generation sequencing of 16S and 18S rRNA genes. Soil amended with OAs generally suppressed disease by S. sclerotiorum and FOL compared to fumigants and synthetic fertilizers. However, the incidence of R. solani infection was lower in soil treated with synthetic fertilizers than soil amended with OAs. EC, pH, C:N, N-NO₃⁻, N-NH₄⁺, FDA and BIOLOG were significantly correlated with disease, depending on pathosystem. Bacterial richness and diversity, presence of some genera like Acidobacteria, Chloracidobacteria, Solibacteres, Anaerolineae, Nitrospira and Deltaproteobacteria were negatively related to disease incidence of FOL and S. sclerotiorum, whereas damping-off caused by R. solani was negatively affected by the presence of Flavobacterium and Chitinofagha. Therefore, we concluded that long-term application of OAs can effectively improve soil suppressiveness and reduce disease incidence against root pathogens, although the effects vary considerably depending upon pathosystem.     

ULTRASONOGRAPHIC AND PATHOLOGIC FEATURES OF INTESTINAL SMOOTH MUSCLE HYPERTROPHY IN FOUR CATS

The ultrasonographic findings for four cats with intestinal smooth muscle hypertrophy are described. In two cats, intestinal smooth muscle hypertrophy was associated with chronic enteritis. In the remaining two cats, intestinal smooth muscle hypertrophy affected the intestinal tract proximal to stenosis due to alimentary lymphoma and an intestinal foreign body, respectively. Moderate increased thickness of the affected intestinal wall, measuring 7-8 mm, was evident on abdominal ultrasonographic examination of all subjects. In addition, the ultrasonographic five-layered feature of the intestinal wall was maintained, and only the muscular layer appeared thickened. Abdominal ultrasound allowed a presumptive diagnosis of intestinal smooth muscle hypertrophy that was confirmed histologically in all cats.     

Veterinary ocular microbiome: Lessons learned beyond the culture

Ocular pathogens cause many painful and vision‐threatening diseases such as infectious keratitis, uveitis, and endophthalmitis. While virulent pathogens and pathobionts play important roles in disease pathogenesis, the scientific community has long assumed disruption of the ocular surface occurs prior to microbial colonization and subsequent infection. While nonpathogenic bacteria are often detected in corneal and conjunctival cultures from healthy eyes, cultures also frequently fail to yield growth of common ocular pathogens or nonpathogenic bacteria. This prompts the following question: Is the ocular surface populated by a stable microbial population that cannot be detected using standard culture techniques? The study of the microbiome has recently become a widespread focus in physician and veterinary medicine. Research suggests a pivotal symbiotic relationship with these microbes to maintain healthy host tissues, and when altered is associated with various disease states (“dysbiosis”). The microbiota that lives within and on mammalian bodies have long been known to influence health and susceptibility to infection. However, limitations of traditional culture methods have resulted in an incomplete understanding of what many now call the “forgotten organ,” that is, the microbiome. With the introduction of high‐throughput sequencing, physician ophthalmology has recognized an ocular surface with much more diverse microbial communities than suspected based on traditional culture. This article reviews the salient features of the ocular surface microbiome and highlights important future applications following the advent of molecular techniques for microbial identification, including characterizing ocular surface microbiomes in our veterinary species and their potential role in management of infectious and inflammatory ocular diseases.     

GeneScanning analysis of Ig/TCR gene rearrangements to detect clonality in canine lymphomas

The diagnosis of canine lymphoma is achieved using morphological and immunological methods. In a certain percentage of cases, difficulties in making a definitive diagnosis of lymphoproliferative disorders may occur despite extensive immunophenotyping. Therefore, additional diagnostics, such as molecular assessment of Ig/TCR gene rearrangements clonality, may confirm the final diagnosis. Polyacrylamide gel electrophoresis and heteroduplex analysis have already been proven to be suitable for detecting clonality but are cumbersome and labor-intensive. In the present study, GeneScanning analysis of PCR products originating from different primer sets targeting different regions of Ig and TCR was validated in improving sensitivity as well as in reducing the turnaround time of gene rearrangement assays. GeneScanning exploits 5' fluorescently labelled primers for the automated and fast analysis of PCR products either as singleplex or multiplex runs. Initially, the assay was set up using DNA purified from normal tissues (n=6), hyperplastic/reactive tissues (n=10) and a small set of immunophenotyped lymphoma samples (n=12). The optimized methods were then used in a large set of 96 canine lymphoma samples. Normal and hyperplastic/reactive lymphoid tissues showed typically polyclonal or, occasionally, oligoclonal PCR products. Lymphoma samples showed monoclonal peaks arranged as a single or, occasionally, a double narrow base peak sometimes embedded in a polyclonal background. In all immunophenotyped cases, an Ig or TCR clonal finding corresponded to B- and T-cell lymphomas, respectively. Overall, 94/96 (97.9%) samples showed clonal Ig/TCR clonal rearrangements among which clonal Ig was found in 61/96 (63.5%) of samples and clonal TCR in 33/35 Ig negative samples (34.4% of all cases). In one out of ten randomly chosen cases, both Ig and TCR clonal gene rearrangements were found. Among the factors affecting assay accuracy, DNA quality has been shown to be critical and the amplification of DNA controls of different size are recommended to evaluate DNA integrity. Frozen material such as that which remained inside the hub of the needle used for diagnostic procedures is optimal for the analysis herein described. In conclusion, GeneScanning represents a versatile tool for routinely assessing Ig/TCR clonal rearrangements and supporting the diagnostic protocol of canine lymphomas.