Description of a foodborne disease outbreak caused by Streptococcus equi subsp. Zooepidemicus
Streptococcus equi subsp. Zooepidemicus (SEZ) are β-haemolytic gram-positive bacteria.
It belongs to group C of Lancefield, and it shares up to 98% of homologues DNA sequences with Streptococcus equi (S. equi) (Pelkonen et al., 2013).
SEZ is an opportunistic pathogen able to infect domestic and wildlife animals (Priestnall et al., 2011). In horses, SEZ is commonly found in respiratory and reproductive tract of healthy animals. In dogs, can cause fever, dyspnoea, pneumonia, and haemorrhagic nasal discharge. In some cases, it can lead the animal to death. In pigs and primates are commonly reported polyarthritis, bronchopneumonia, pleuritis, pericarditis, endocarditis, and meningitis. In ruminants can be isolated in animals with mastitis (Corpa et al., 2018).
Although rare, in human the infection usually happens following contacts with infected animals, especially horses, but foodborne outbreaks were reported due to ingestion of unpasteurized milk and milk products (Bordes-Benìtez et al., 2006, Kerdsin et al., 2022). The disease is quite severe, causing meningitis and septicaemia.
Correlation between clinical cases and source of the infection is possible thanks to DNA extraction and sequencing of the genome by Next Generation Sequencing (NGS). Multilocus Sequence Typing (MLST) enable characterization of the isolated colonies and Sequence type determination (Pelkonen et al., 2013). Even more discriminant is the analysis of the Single Nucleotide Polymorphism (SNP) (EFSA BIOHAZ, 2019).
The case: outbreak determination, epidemiological investigation and Streptococcus equi subsp. Zooepidemicus
Between November 2021 and June 2022, in the province of Pescara, 37 cases of Streptococcus equi subsp. Zooepidemicus were notified. Among them, five people died because of meningitis. Clinical signs included septicaemia, pharyngitis, arthritis, uveitis, and endocarditis. Cases of the disease involved patients aged 6 to 98 years and no significant gender differences were reported. Different bodies of the Local Authority (LA) were involved to resolve the emergence. Among them, the Food Hygiene and Nutrition Service (Servizio di Igiene degli alimenti e della Nutrizione-SIAN), the Veterinary Service (Servizi Veterinari-SSVV), and the Hygiene and Public Safety Unit (Servizio Igiene e Sanità Pubblica-SIESP).
In total, 21 strains isolated from 19 clinical cases, blood, cerebrospinal and synovial fluid, were collected during the epidemic.
The strains were sent to the Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise (IZSAM) to be identified by Matrix-Assisted Laser Desorption - Time of Flight (MALDI-TOF) (Bruker Biotyper, Germany), and confirmed by MLST analysis of the sequenced genome by Next-generation sequencing (NGS) technique.
From the analysis the strain causing the outbreak belonged to ST61. Correlation between strains was possible through SNPs analysis using CFSAN pipeline. As reference a hybrid genome of a clinical case was obtained with Illumina and MINION technology.
The Local Authority carried out an epidemiological investigation to identify the source of infection. During the epidemiological investigation, patients reported consumption of local dairy products, namely raw milk products. The investigations carried out in the territory allowed to identify 8 dairies in the area that produced products of the type indicated. Therefore, after a first visit, two sampling seasons were scheduled.
During the first session, 9 samples of raw cow’s and/or sheep’s milk, 25 samples of pasteurized and unpasteurized dairy products and 8 samples of water were taken. All samples were sent to the IZSAM for microbiological analysis.
Streptococcus equi subsp. Zooepidemicus was isolated in a sample of cow’s milk mass taken from one of the sampled dairies. Samples were then taken at the only dairy which was positive at the first sampling. The presence in bulk milk was thus confirmed. In addition, SEZ was also isolated in aged cheeses of 37 and 70 days. Whole genome sequencing confirmed similarity and correlation with clinical strains by SNP analysis subsp. Zooepidemicus (SEZ) are β-haemolytic gram-positive bacteria (Figure 1). It belongs to group C of Lancefield, and it shares up to 98% of homologues DNA sequences with Streptococcus equi (S. equi) (Pelkonen et al., 2013).
Figure 1. Streptococcus equi subsp. Zooepidemicus growth on Blood Agar
SEZ is an opportunistic pathogen able to infect domestic and wildlife animals (Priestnall et al., 2011). In horses, SEZ is commonly found in respiratory and reproductive tract of healthy animals. In dogs, can cause fever, dyspnoea, pneumonia, and haemorrhagic nasal discharge. In some cases, it can lead the animal to death. In pigs and primates are commonly reported polyarthritis, bronchopneumonia, pleuritis, pericarditis, endocarditis, and meningitis. In ruminants can be isolated in animals with mastitis (Corpa et al., 2018).
Although rare, in human the infection usually happens following contacts with infected animals, especially horses, but foodborne outbreaks were reported due to ingestion of unpasteurized milk and milk products (Bordes-Benìtez et al., 2006, Kerdsin et al., 2022). The disease is quite severe, causing meningitis and septicaemia.
Correlation between clinical cases and source of the infection is possible thanks to DNA extraction and sequencing of the genome by Next Generation Sequencing (NGS). Multilocus Sequence Typing (MLST) enable characterization of the isolated colonies and Sequence type determination (Pelkonen et al., 2013). Even more discriminant is the analysis of the Single Nucleotide Polymorphism (SNP) (EFSA BIOHAZ, 2019).
Conclusions
In conclusion, the deep epidemiological investigation and microbiological analysis of the samples allowed isolation of the pathogen and its identification and characterization. More discriminant technique, such as NGS, enabled to trace back the source of infection quickly, limiting the human transmission and safeguarding public health safety.
The coordination between different bodies of the LA was fundamental to reach these results. The laboratories at IZSAM supported the LA since the beginning of the outbreak, and the collaborations is continuous, in order to develop a method able to isolate, identify and characterize the pathogen, to confirm the correlation between strains isolated from clinical cases and food products, and to discriminate between strains belonging to the outbreak and those that are not.
References
- Bordes-Benítez A, Sánchez-Oñoro M, Suárez-Bordón P, García-Rojas AJ, Saéz-Nieto JA, González-García A, Alamo-Antúnez I, Sánchez-Maroto A, Bolaños-Rivero M. Outbreak of Streptococcus equi subsp. zooepidemicus infections on the island of Gran Canaria associated with the consumption of inadequately pasteurized cheese. Eur J Clin Microbiol Infect Dis. 2006 Apr;25(4):242-6
- Corpa JM, Carvallo F, Anderson ML, Nyaoke AC, Moore JD, Uzal FA. Streptococcus equi subspecies zooepidemicus septicemia in alpacas: three cases and review of the literature. J Vet Diagn Invest. 2018 Jul;30(4):598-602
- EFSA BIOHAZ Panel (EFSA Panel on Biological Hazards), Koutsoumanis, K, Allende, A, Alvarez-Ordóñez, A, Bolton, D, Bover-Cid, S, Chemaly, M, Davies, R, De Cesare, A, Hilbert, F, Lindqvist, R, Nauta, M, Peixe, L, Ru, G, Simmons, M, Skandamis, P, Suffredini, E, Jenkins, C, Malorny, B, Ribeiro Duarte, AS, Torpdahl, M, da Silva Felício, MT, Guerra, B, Rossi, M and Herman, L, 2019. Scientific Opinion on the whole genome sequencing and metagenomics for outbreak investigation, source attribution and risk assessment of food-borne microorganisms. EFSA Journal 2019;17(12):5898, 78 pp.
- Kerdsin, A., Chopjitt, P., Hatrongjit, R., Boueroy, P., & Gottschalk, M. (2022). Zoonotic infection and clonal dissemination of Streptococcus equi subspecies zooepidemicus sequence type 194 isolated from humans in Thailand. Transboundary and Emerging Diseases. 69, e554– e565
- Pelkonen S, Lindahl SB, Suomala P, Karhukorpi J, Vuorinen S, Koivula I, Väisänen T, Pentikäinen J, Autio T, Tuuminen T. Transmission of Streptococcus equi subspecies zooepidemicus infection from horses to humans. Emerg Infect Dis. 2013 Jul;19(7):1041-8
- Priestnall S, Erles K. Streptococcus zooepidemicus: an emerging canine pathogen. Vet J. 2011 May;188(2):142-8. doi: 10.1016/j.tvjl.2010.04.028. Epub 2010 May 31.
De Angelis Maria Elisabetta, Bosica Serena
Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise “G. Caporale”
Unit of hygiene in food technology and animal feeds
