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BACKGROUND: Infectious bovine keratoconjunctivitis (IBK; pinkeye) is generally considered to be caused by corneal infections with Moraxella bovis. Previous studies demonstrated that M. bovis cytotoxin-specific mucosal immune responses in the bovine eye can be stimulated by intranasal vaccination with a recombinant M. bovis cytotoxin subunit adjuvanted with polyacrylic acid. METHODS: A randomized controlled field trial (two-arm parallel design with blinding) was conducted in beef steers in Northern California to determine if this vaccine could prevent naturally occurring IBK and/or reduce morbidity rates associated with this disease. Beef steers were vaccinated intranasally on days 0 and 21 with either a recombinant M. bovis cytotoxin subunit adjuvanted with polyacrylic acid (Vaccine group) or adjuvant alone (Control group). Eye examinations were performed on all steers every 7 days for 16 weeks to document the occurrence of IBK and to determine sizes of corneal ulcers. Serum and tear samples were collected on days 0, 42, and 112 from a subset of animals to measure changes in systemic and ocular immune responses to M. bovis cytotoxin. RESULTS: The cumulative proportion of steers that developed IBK after 16 weeks did not differ between groups. Variables related to disease severity were numerically lower in steers that received the experimental vaccine. IBK-affected Vaccine group steers had a significantly lower number of observation weeks with severe ulcers versus Control group steers. Cytotoxin-specific tear IgA was significantly higher in Vaccine group compared to Control group steers on day 112. Conclusion: Although the proportion of animals that developed corneal ulcers associated with IBK did not differ between groups, the lowered metrics of disease severity in vaccinated steers suggests that intranasal vaccination with recombinant M. bovis cytotoxin can reduce the severity of IBK in cattle.

BACKGROUND AND AIM: Infectious bovine keratoconjunctivitis (IBK) causes a significant economic loss to cattle industries in many countries, including Kazakhstan. Although Moraxella bovis is recognized as an etiologic agent of IBK, other bacterial and viral agents have been suspected to play a role in the pathogenesis of this disease. This study aimed to evaluate samples collected from the eyes of IBK-affected cattle in Eastern Kazakhstan at different stages of IBK for the presence of Mor. bovis, Moraxella bovoculi, Mycoplasma bovis, Mycoplasma bovoculi, and Bovine Herpes Virus Type 1 (BHV-1) and to characterize Mor. bovoculi pilA gene sequence diversity from Mor. bovoculi positive samples. MATERIALS AND METHODS: Individual ocular swabs (n = 168) were collected from cattle that had clinical signs of IBK during the summer of 2022 on farms in the Abay region of Kazakhstan. Eye lesion scores (1, 2, and 3) were assigned depending on the degree of ocular damage. Infectious bovine keratoconjunctivitis-associated organisms were detected using a multiplex real-time polymerase chain reaction assay. The Mor. bovoculi pilA gene was sequenced from Mor. bovoculi positive samples. RESULTS: Mycoplasma bovis and BHV-1 were not detected in any of the collected samples. Mycoplasma bovoculi was identified in the majority of samples overall, usually in mixed infection with Moraxella spp. Moraxella bovoculi was detected in 76.2% of animals and predominated in animals with eye lesion scores 2 and 3. Mycoplasma bovoculi was detected only in association with Mor. bovis and/or Mor. bovoculi in animals with eye lesion scores 2 and 3. Moraxella bovis was found in 57.7% of animals and was always identified in association with another organism. Sequencing of the pilA gene in 96 samples from Mor. bovoculi positive samples identified five PilA groups. The majority belonged to PilA group A. However, three new PilA groups were identified and designated PilA groups N, O, and P. CONCLUSION: The results indicate a high prevalence of Myc. bovoculi and Mor. bovoculi in eyes of cattle with IBK on livestock farms in Eastern Kazakhstan. Additional novel Mor. bovoculi PilA groups were identified.

Introduction.Moraxella bovoculi is frequently isolated from the eyes of cattle with infectious bovine keratoconjunctivitis (IBK; pinkeye). As with M. bovis, which has been causally linked to IBK, M. bovoculi expresses an RTX (repeats in the structural toxin) cytotoxin that is related to M. bovis cytotoxin. Pilin, another pathogenic factor in M. bovis, is required for corneal attachment. Seven antigenically distinct pilin serogroups have been described in M. bovis.Hypothesis/Gap Statement. Multiple different serogroups exist amongst type IV pilin encoded by M. bovis, however, it is not known whether M. bovoculi exhibits a similar degree of diversity in type IV pilin that it encodes.Aim. This study was done to characterize a structural pilin (PilA) encoded by M. bovoculi isolated from cases of IBK to determine if diversity exists amongst PilA sequences.Methodology. Ninety-four isolates of M. bovoculi collected between 2002 and 2017 from 23 counties throughout California and from five counties in four other Western states were evaluated.Results. DNA sequencing and determination of deduced amino acid sequences revealed ten (designated groups A through J) unique PilA sequences that were ~96.1-99.3 % identical. Pilin groups A and C matched previously reported putative PilA sequences from M. bovoculi isolated from IBK-affected cattle in the USA (Virginia, Nebraska, and Kansas) and Asia (Kazakhstan). The ten pilin sequences identified were only ~74-76 % identical to deduced amino acid sequences of putative pilin proteins identified from the previously reported whole-genome sequences of M. bovoculi derived from deep nasopharyngeal swabs of IBK-asymptomatic cattle.Conclusions. Compared to the diversity reported between structural pilin proteins amongst different serogroups of M. bovis, M. bovoculi PilA from geographically diverse isolates derived from IBK-affected cattle are more conserved.

As is the case for controlling other infectious livestock diseases, the most successful efforts to control infectious bovine keratoconjunctivitis (IBK) will include consideration of the host, the environment, herd management, and ongoing surveillance even after the immediate crisis has passed. Research over many years has led to the discovery of a variety of antibiotic treatments and antibiotic regimens that can be effective against IBK. The discoveries of Mor bovoculi and reports of IBK associated with Mycoplasma spp without concurrent Mor bovis or Mor bovoculi have raised new questions into the roles that other organisms may play in IBK pathogenesis.

Infectious bovine keratoconjunctivitis (IBK) caused by Moraxella bovis is the most common eye disease of cattle. The pathogenesis of M. bovis requires the expression of pili that enable the organism to attach to the ocular surface and an RTX (repeats in the structural toxin) toxin (cytotoxin or hemolysin), which is cytotoxic to corneal epithelial cells. In this pilot study, ocular mucosal immune responses of steers were measured following intranasal (i.n.) vaccination with a recombinant M. bovis cytotoxin adjuvanted with polyacrylic acid. Beef steers were vaccinated with either 500 μg (n = 3) or 200 μg (n = 3) of recombinant M. bovis cytotoxin plus adjuvant. Control group steers (n = 2) were vaccinated with adjuvant alone, and all steers were given a booster on day 21. Antigen-specific tear IgA and tear IgG, tear cytotoxin-neutralizing antibody responses, and serum cytotoxin-neutralizing antibody responses were determined in samples collected prevaccination and on days 14, 28, 42, and 55. Changes in tear antigen-specific IgA levels from day 0 to days 28, 42, and 55 were significantly different between groups; however, in post hoc comparisons between individual group pairs at the tested time points, the differences were not significant. Our results suggest that i.n. vaccination of cattle with recombinant M. bovis cytotoxin adjuvanted with polyacrylic acid effects changes in ocular antigen-specific IgA concentrations. The use of intranasally administered recombinant M. bovis cytotoxin adjuvanted with polyacrylic acid could provide an alternative to parenteral vaccination of cattle for immunoprophylaxis against IBK.

A randomized, blinded, controlled field trial was conducted during summer 2006 in a northern California, USA, herd of beef cattle to evaluate the efficacy of a recombinant Moraxella bovoculi cytotoxin subunit vaccine to prevent naturally occurring infectious bovine keratoconjunctivitis (IBK; pinkeye). A convenience sample comprised of 127 steers were administered a subcutaneous dose of either adjuvant alone (ISCOM matrices; control group) or recombinant M. bovoculi cytotoxin carboxy terminus adjuvanted with ISCOM matrices (MbvA group) and were boostered 21 days later. The steers were examined once weekly for 15 weeks for evidence of IBK. No significant difference in the cumulative proportion of corneal ulcerations was detected between groups. Compared to the control calves, the MbvA vaccinates had significantly higher increases in serum neutralizing titers to M. bovoculi hemolysin between week 0 and week 6. The prevalence of M. bovis isolations was higher from ulcerated eyes of calves vaccinated with MbvA as compared to control calves. Vaccination of calves against the carboxy terminus of M. bovoculi RTX toxin resulted in significant increases in serum hemolysin neutralizing titers and may modulate organism type cultured from ulcerated eyes of calves in herds where both M. bovis and M. bovoculi exist. Use of M. bovoculi antigens alone in vaccines to prevent IBK may not be beneficial in herds where IBK is associated with both M. bovoculi and M. bovis.