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.     

An atypical case of respiratory actinobacillosis in a cow

A not pregnant 4-year-old Jersey cow was presented with the sudden appearance of respiratory noise, nasal discharge and moderate respiratory difficulty. Upon physical examination a snoring-like noise, extended head and neck position, exaggerated abdominal effort, bilateral nasal discharge and left prescapular lymph node enlargement were noted. Sub-occlusion of the initial portion of the respiratory tract was suspected. Radiographic and endoscopic examinations revealed a pedunculate mass on the dorsal aspect of the rhinopharynx, which was removed with endoscopically assisted electrosurgery. Histologic examination revealed a chronic pyogranulomatous inflammation with eosinophilic club-like bodies surrounding small colonies of rod-shaped bacteria. Results of histochemical staining were consistent with Actinobacillus-like bacteria and a diagnosis of respiratory actinobacillosis was reached. Surgery and antibiotic therapy were resolutive, as demonstated by an endoscopic check at the second month after surgery, even without the association of the traditional iodine cure, which is regarded as the treatment of choice for actinobacillosis.     

Incidence of Poisonings in Domestic Carnivores in Italy

Veterinary Research Communications -     

Effects of the radiometric properties of innovative biodegradable mulching materials on snapdragon cultivation

Research on the application of innovative biodegradable materials for substrate mulching in greenhouse floricultural cultivation was carried out in Southern Italy. A transparent biodegradable film and a transparent biodegradable spray coating were compared with a commercial transparent low density polyethylene film. The biodegradable film was a starch based extruded material, the sprayed coating was created from a polysaccharide solution. The materials were used to mulch the peat and perlite growing media contained in trays during snapdragon cultivation in order to increase the substrate temperature and hasten early growth. The climate variables inside and outside the greenhouse and the growing medium temperature under the mulching materials were recorded. Biometrical data, such as mean blooming time, plant height and spike length, were collected. The laboratory radiometric tests showed that the biodegradable mulching materials had a lower solar transmissivity in comparison to the low density polyethylene film. The capacity to reduce the long wave infrared thermal losses was higher for the biodegradable mulches: the long wave infrared transmissivity coefficient was 53.9% for the low density polyethylene film, 12.1% for the biodegradable film and 8.2% for the spray coating. The increase of the growing medium temperature allowed the harvest of the snapdragon grown in the mulched trays 7 days before that of the flowers grown in the un-mulched trays. At the end of the crop cycle the disposal of the biodegradable mulching materials was carried out fragmenting and mixing them with plant residue and the growing medium. The residue biodegraded in almost 1 month for the spray coating and in about 12 months for the biodegradable film.     

Inferomedial placement of a single-entry subpalpebral lavage tube for treatment of equine eye disease

The objective of this study was to describe method of placement, and frequency and severity of complications associated with a subpalpebral lavage system placed in the medial aspect of the equine inferior eyelid. The inferomedial subpalpebral lavage (ISPL) tube is positioned deep in the medial aspect of the inferior conjunctival fornix so that the footplate lies flat between the lower eyelid and the anterior surface of the nictitans. Retrospective data from the placement of 92 ISPL systems placed in 86 horses during a 31-month period were examined. Tube placement was performed using sedation and regional anesthesia only in 59% of horses. The median duration of tube placement was 19 days (range: 1-61 days). Seventy-one horses were treated for up to 55 days following discharge from hospital with an ISPL tube in place. No complications were reported with 59% of ISPL systems. Non-ocular complications were found in 38% of ISPL systems and included tube displacement from the conjunctival fornix (18%), suture loss requiring resuturing of the system to the horse's head (14%), and damage necessitating replacement of the injection port (6%). Ocular complications were recorded in 3% of horses and were limited to inferior eyelid swelling. Vision was retained in 88% of horses. The ISPL system is easily and safely placed, and well tolerated for extended periods. It appears to be associated with infrequent and minor complications when compared with placement of subpalpebral lavage tubes in the superior eyelid.     

Identification of a universal Group B streptococcus vaccine by multiple genome screen

Group B Streptococcus (GBS) is a multiserotype bacterial pathogen representing a major cause of life-threatening infections in newborns. To develop a broadly protective vaccine, we analyzed the genome sequences of eight GBS isolates and cloned and tested 312 surface proteins as vaccines. Four proteins elicited protection in mice, and their combination proved highly protective against a large panel of strains, including all circulating serotypes. Protection also correlated with antigen accessibility on the bacterial surface and with the induction of opsonophagocytic antibodies. Multigenome analysis and screening described here represent a powerful strategy for identifying potential vaccine candidates against highly variable pathogens.