Abstract

The repeated occurrence of anthrax in grazing animals should be a reminder of a widespread presence of Bacillus anthracis spores in the environment. Its rapid diagnosis is critical to protect public health. Here, we report a case of anthrax in cattle that was investigated using conventional and molecular methods. In 2015, six cows suddenly died within three days and the number of dead animals increased to a total of 12 within two weeks. At necropsy, anthrax was suspected. Therefore, spleen tissue samples were collected (from 6/12 animals) and laboratory tests (microscopy, cultivation, and real-time PCR) performed. The results of tissue staining for microscopy and cultivation were in congruence, while B. anthracis real-time PCR outperformed both. Spleen tissues from all six animals were real-time PCR-positive, while B. anthracis was successfully cultivated and detected by microscopy from the spleen of only three animals. Additionally, the ear tissue from another (1/12) cow tested positive by real-time PCR, supporting the suitability of ear clippings for molecular confirmation of B. anthracis. Genotyping of the isolates using multiple-locus variable-number tandem repeat analysis (MLVA) revealed a common source of infection as all three typed isolates had an indistinguishable MLVA genotype, which has not been observed previously in Europe. The results indicate that molecular testing should be selected as the first-line tool for confirming anthrax outbreaks in animals to ensure timely protection of public health.

Highlights

  • In the spleen samples from 6/12 animals subjected to laboratory tests, the presence of B. anthracis was confirmed by at least one method: real-time PCR was positive in all six samples, in contrast to microscopy and cultivation, where only three samples

  • Real-time PCR was performed for the blood sample after DNA extraction, and it was positive for B. anthracis, but only pag was detected

  • Anthrax is a rare disease in Slovenia and Europe, but a prompt response to outbreaks in animals is crucial to minimize the risk of zoonotic transmission

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Summary

Introduction

The true worldwide incidence of anthrax is not known; epizootics occur each year, resulting in the death of hundreds to thousands of animals and transmission of the disease to humans. It is estimated that between 2000 and 20,000 human anthrax cases occur worldwide yearly [2,3]. Anthrax is not a major human or animal health issue in developed countries; in Europe, it is a rare disease with only a few cases reported annually. Between 2007 and 2019, 97 confirmed human anthrax cases were reported in Europe, ranging from one to 32 cases per year [4]. We describe the most recent animal anthrax cases in Slovenia and compare the performance of methods available in our laboratory

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