Monitoring of Paenibacillus larvae in Lower Austria through DNA-Based Detection without De-Sporulation: 2018 to 2022.

American foulbrood is caused by the spore-forming Paenibacillus larvae. Although the disease effects honey bee larvae, it threatens the entire colony. Clinical signs of the disease are seen at a very late stage of the disease and bee colonies are often beyond saving. Therefore, through active monitoring based on screening, an infection can be detected early and bee colonies can be protected with hygiene measures. As a result, the pressure to spread in an area remains low. The cultural and molecular biological detection of P. larvae is usually preceded by spore germination before detection. In this study, we compared the results of two methods, the culture detection and RT-PCR detection of DNA directly isolated from spores. Samples of honey and cells with honey surrounding the brood were used in a five-year voluntary monitoring program in a western part of Lower Austria. DNA-extraction from spores to speed up detection involved one chemical and two enzymatic steps before mechanical bashing-beat separation and additional lysis. The results are comparable to culture-based methods, but with a large time advantage. Within the voluntary monitoring program, the proportion of bee colonies without the detection of P. larvae was high (2018: 91.9%, 2019: 72.09%, 2020: 74.6%, 2021: 81.35%, 2022: 84.5%), and in most P. larvae-positive bee colonies, only a very low spore content was detected. Nevertheless, two bee colonies in one apiary with clinical signs of disease had to be eradicated.

The role of molecular-genetic techniques in BVDV eradication in Lower Austria.

A voluntary bovine viral diarrhoea virus (BVDV) control programme, which later became a compulsory eradication programme, based on the Swedish model was introduced in Lower Austria in 1997. The persistently infected animals were detected by Ag-ELISA and all samples were re-tested by the improved single-tube RT-PCR, employing panpestivirus primers targeting the 5'-UTR of the virus genome. In 2010, the BVDV eradication programme, which became compulsory from 2004, reached the final stage with only five remaining infected herds in which BVDV was difficult to eradicate. To resolve the problem in those herds, a molecular epidemiology approach was used. No differences in the spectrum of BVDV-1 subgenotypes at the beginning and at the final stage of eradication programme were found. The genetic study revealed the importance of human risk factor when finishing an eradication programme. Molecular epidemiology was also used to analyse BVDV isolates associated with re-introductions to BVDV-free herds.

Analysis of BVDV isolates and factors contributing to virus transmission in the final stage of a BVDV eradication program in lower Austria.

The BVDV eradication program in Lower Austria according to the Swedish model started in 1997 as a voluntary strategy and became obligatory for all herd owners in 2004. In this paper we analyse BVDV isolates found in PI animals of the last infected cattle farms at the genetic level and describe possible risk factors for virus transmission in the final stage of the BVDV eradication program in Lower Austria, where only five infected herds were identified in 2010. Viral isolates from 23 farms on which PI animals had been detected in 2009 (n = 10), 2010 (n = 5), 2008 (n = 5) and in 2006 (n = 3) were analysed at a genetic level at the 5'-UTR. All isolates analysed by phylogenetic analysis fall into five BVDV-1 subgenotypes, i.e., b, e, f, g, and h. No new subgenotypes were identified when compared to isolates analysed at the beginning of the BVDV eradication program. The phylogenetic analysis also revealed three groups of herds with genetically identical isolates; the first group comprised two herds, the second seven herds and the third four herds. Analysis of several factors such as distance between herds, purchase of cattle, work of farmers, veterinarians, assistants of milk recording associations and animal carriers indicated the latter as the most critical factor for transmitting the same viruses. Seven herds of two groups of identical BVDV isolates were served by the same cattle carrier, six herds shared the veterinarian and five herds had the same assistant of the milk recording association who could be the risk factor for the introduction of BVDV into the herd. The analysis helped to stop virus transmission. Therefore, from early 2012 up to now, no new infection has occurred in Lower Austria. The results indicated that biosecurity is a very important factor to prevent the spreading of viruses in the final stage of the BVDV eradication program.