Search

SUMMARYThis is the first extensive report on the identification and characterization of Avibacterium paragallinarum (AVP) isolates obtained from outbreaks of infectious Coryza (IC) in layer flocks from Sonora State in Mexico with an IC vaccination history. Isolates obtained from IC outbreaks during the years 2007 up to and including 2019 were identified by conventional PCR test and 16S rRNA gene analysis, serotyped by Page serotyping and genotyped by the recently described partial sequence analysis of the HPG2 region. Furthermore, antimicrobial susceptibility profiles were determined by a recently improved MIC test. The conventional PCR test and the 16S rRNA analyses confirmed the isolates as AVP. Serotyping results showed the involvement of isolates belonging to serotypes A, B and C in the IC outbreaks. Genotyping of the HPG2 region revealed the presence of Sequence Type (ST)1, ST4 and ST11 of which the latter is also identified in Europe. The Minimal Inhibitory Concentration (MIC) susceptibility test showed that all tested isolates were susceptible for the majority of tested antimicrobials including erythromycin and tetracycline which are important antibiotics for the treatment of IC. Vaccination failures due to inadequate vaccination application procedures should be ruled out in the control of IC in Sonora State.

This is the first extensive report on the identification and characterization of

Background: Vaccination of farmed minks against canine distemper virus (CDV) has proved to be very effective. In the Netherlands, vaccination of farmed minks against CDV was mandatory until the closure of the local agricultural product boards at the end of 2014. Objectives: To describe the first documented outbreaks of CD in Dutch mink farms since the closure of the agricultural product boards, as well as an outbreak in Belgium, with special attention to genotyping of the isolates. Methods: A full post-mortem was performed on three carcasses per submission from farms A–C and on two carcasses from farm D. Molecular detection with subsequent typing was performed on eleven samples originating from four different farms. To assess genetic diversity partial sequences of the H gene of CDV were compared based on phylogenetic analysis. Results: In 2017, there was a sudden series of CD outbreaks affecting four mink farms in the Netherlands (A–C) and Belgium (D). Gross, histologic and immunohistochemical findings were similar. There was a degree of genetic similarity between the viruses on farms A and D (98.5%) and between the viruses on farms B and C (97.3%), but the viruses from farms A and D belonged to a different clade than the viruses from farms B and C. Higher mortalities were reported in white and pastel minks. Conclusions: Findings indicated that the difference in severity of the outbreaks was partially related to the genetic composition of the farm populations. Vaccination against CDV on Dutch and Belgian mink farms seems warranted.

This study reports the results of diagnostic and molecular typing methods for 18

Protocol cDNA synthesis for PRRSV ORF 2-7 PCR: Protocol RT-reaction For preparation of the RT mix the next components were combined in a pre-mix: Component pre-mix n = 1 n = 13 (PRRSV) RNA 7 µL Primer stock 10 µM Poly dT 2 µL 26 µL 10 mM dNTP mix 1 µL 13 µL For each sample 2 µL poly dT primer and 1 µL dNTP mix were mixed with 7 µL RNA. Samples were incubated for 5 minutes at 65°C (denaturation) and subsequently cooled on melting ice for at least 1 minute. A cDNA synthese mix was prepared : Component n = 1 n= 13 10X RT buffer 2 µL 26 µL 25 mM MgCL2 4 µL 52 µL 0.1 M DTT 2 µL 26 µL RNaseOUT (40 U/µL) 1 µL 13 µL SuperScript III RT (200 U/µL) 1 µL 13 µL 10µL cDNA syntheses mix was added to 10 µL RNA/pre-mix . After mixing, the tubes were centrifuged for a few seconds. Samples were incubated for 75 minutes at 50°C (cDNA syntheses) and subsequently for 5 minutes at 85°C (termination). After that, the samples were cooled on melting ice. Samples were centrifuged for a few seconds and to each cDNA sample 1 µL RNAse H was added and incubated for 20 minutes at 37°C. The samples were used directly in the PRRSV ORF 2-7 PCR or stored at -70°C.

Preparation PCR mix PRRSV ORF 2-7 PCR: Reagent n = 1 n =65 10x Accuprime PCR buffer (I) 5 µL 325 µL Fw primer 10 µM (1 or 2 or 3)* 1 µL 65 µL Rv primer 10 µM EU 15098R** 1 µL 65 µL Accu prime TaqDNA polymerase High fidelity 0.5 µL 32.5 µL Nuclease free water 40.5 µL 2632.5 µL Template 2 µL PCR protocol: PCR protocol: Step Temp (°C) Time Denaturation / Activation 94°C 15 secDenaturation 94°C 15 secAnnealing 55°C 30 secElongatie 68°C 60 sec per kB 6 min Extra Elongatie 68°C 2 min per kB 12 min 45 cycli Cooling 4 °C ∞ Gel electrophoreses PCR products were separated on a 0.8 1X TBE agarose gel with 0.05% ethidium bromide. 5 µL amplicon was mixed with 2 µL loading buffer. Runtime was 1 hour at 100 Volt. To estimate the fragment sizes of the PCR products a High DNA Mass Ladder was includen in every gelelectrophoresis run. (6 DNA fragments of 10,000; 6000; 4000; 3000; 2000; and 1000 bp were expected). For each reaction 4 µL High DNA Mass ladder was used. The expected PRRS ORF 2-7 DNA fragment was ~3375 bp.

Magmax pathogen RNA/DNA kit 200 µL input: serum protocol

This is the first extensive report on the identification and characterization of Avibacterium paragallinarum (AVP) isolates obtained from outbreaks of infectious coryza (IC) in IC-vaccinated layer flocks from Sonora State in Mexico. Isolates obtained from IC outbreaks during the years 2007, 2014, 2015, 2017, and 2019 were identified by conventional PCR test and 16S rRNA gene analysis, serotyped by Page serotyping and genotyped by the recently described partial sequence analysis of the HPG2 region. Furthermore, antimicrobial susceptibility profiles were determined by a recently improved minimal inhibitory concentration (MIC) test. The conventional PCR test and the 16S rRNA analyses confirmed the isolates as AVP. Serotyping results showed the involvement of isolates belonging to serotypes A, B, and C in the IC outbreaks. Genotyping of the HPG2 region revealed the presence of sequence type (ST)1, ST4, and ST11, of which the latter has also been identified in Europe. The MIC susceptibility test showed that all tested isolates were susceptible for the majority of tested antimicrobials, including erythromycin and tetracycline, which are important antibiotics for the treatment of IC. The IC situation in Sonora State, Mexico, is complex because of the presence of serotypes A, B, and C. This finding emphasizes the importance of biosecurity in combination with the application of the most optimal vaccination programs in the control of IC in Sonora State, Mexico.

A 30-year-old female donkey with pituitary pars intermedia dysfunction (PPID) was referred to the Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, for losing weight despite good appetite and treatment for PPID. Twelve days after returning home, the mare developed nasal discharge, and the next day it was found recumbent and was only able to stand up with manual assistance. The next day the mare showed severe tetraparesis, ataxia, and hypotonia of anus, tail and bladder, and it became completely recumbent. Equine herpesvirus myeloencephalopathy was suspected, and to confirm the diagnosis a deep nasal swab was taken. Because of the poor prognosis the mare was euthanased. The swab scored very strong positive for equine herpesvirus-1 (EHV-1) and virus isolation was performed to enable further characterisation of the virus. The identity of the virus was again confirmed as EHV-1 with type-specific monoclonal antibodies, but sequencing identified the virus as asinine herpesvirus-3 (EHV-8).

Background The emerging skin disease fur animal epidemic necrotic pyoderma (FENP) has been attributed to infection with Arcanobacterium phocae (ABP). The exact pathogenesis and risk factors of FENP have yet to be elucidated. Animals Three mink from each of three different mink farms (A–C) with postvaccination skin wounds at the vaccination site and six mink from an unaffected mink farm (D) that had used the same vaccine batch and vaccination site (hind leg). Methods and results All minks from farms A–C had severe necrotizing to necropurulent dermatitis where they were vaccinated intramuscularly in the hind leg. ABP was the sole bacterium cultured from six of nine wounds. Using 16S–23S rDNA intergenic spacer region and BOX-PCR, the ABP isolates from these wounds were indistinguishable from isolates originating from several cases of FENP. Conclusions and clinical importance This is the first report of FENP-like lesions at the site of vaccination, in the days following the procedure, associated with ABP. At farms with FENP vaccination, procedures should be considered carefully.