Vaccine against Streptococcus suis Infection in Pig Based on Alternative Carrier Protein Conjugate.

Streptococcus suis is a serious pathogen in the pig industry with zoonotic potential. With respect to the current effort to reduce antibiotic use in animals, a prophylactic measure is needed to control the disease burden. Unfortunately, immunization against streptococcal pathogens is challenging due to nature of the interaction between the pathogen and the host immune system, but vaccines based on conjugates of capsular polysaccharide (CPS) and carrier protein were proved to be efficient. The main obstacle of these vaccines is manufacturing cost, limiting their use in animals. In this work, we tested an experimental vaccine against Streptococcus suis serotype 2 based on capsular polysaccharide conjugated to chicken ovalbumin (OVA) and compared its immunogenicity and protectivity with a vaccine based on CRM197 conjugate. Ovalbumin was selected as a cheap alternative to recombinant carrier proteins widely used in vaccines for human use. We found that the ovalbumin-based experimental vaccine successfully induced immune response in pigs, and the IgG antibody response was even higher than after immunization with capsular polysaccharide-CRM197 conjugate. Protectivity of vaccination against infection was evaluated in the challenge experiment and was found promising for both conjugates.

Resistance of Streptococcus suis Isolates from the Czech Republic during 2018-2022.

A determination of susceptibility/resistance to antimicrobials via serotype was carried out in 506 field isolates of Streptococcus suis, originating from pig farms in the Czech Republic in the period 2018-2022. A very high level of susceptibility of S. suis isolates was found to amoxicillin, in combination with clavulanic acid and sulfamethoxazole potentiated with trimethoprim. None of the tested isolates were resistant to these antimicrobial substances. Only two isolates were found to be intermediately resistant to enrofloxacin in 2020. With regard to ceftiofur, one isolate was intermediately resistant in 2020 and 2022, and two isolates were intermediately resistant in 2018 and 2021. A low level of resistance was detected to ampicillin (0.6% in 2021) and to florfenicol (1.15% in 2019; 1.3% in 2022). With regard to penicillin, a medium level of resistance was detected in 2018 (10.6%), but a low level of resistance was found in the following years (7.0% in 2019; 3.1% in 2020; 3.3% in 2021; 3.9% in 2022). On the contrary, a high or very high level of resistance was found to tetracycline (66.0% in 2018; 65.1% in 2019; 44.35% in 2020; 46.4% in 2021; 54.0% in 2022). Using molecular and serological methods, serotype 7 (16.4%) was determined to be predominant among S. suis isolates, followed by serotypes 1/2, 2, 9, 4, 3, 1, 29, 16, and 31 (10.7%; 8.5%; 5.7%; 5.5%; 4.5%; 4.3%; 3.6%; 3.4%; 3.4%, respectively). Other serotypes were identified among the investigated strains either rarely (up to 10 cases) or not at all. A relatively high percentage of isolates were detected as non-typeable (79 isolates; 15.6%). Dependence of resistance upon serotype assignment could not be proven in all but serotype 31, wherein all isolates (n = 17) were resistant or intermediately resistant to clindamycin, tilmycosin, tulathromycin, and tetracycline. The resistance to clindamycin and tetracycline may be related to the high consumption of these antibiotics on pig farms at present or in previous years. Macrolides (tilmicosin and tulathromycin) and tiamulin are not suitable for the treatment of streptococcal infections, but are used on pig farms to treat respiratory infections caused by gram-negative bacteria, so they were included in the study.

Experimental Evidence of the Long-Term Survival of Infective African Swine Fever Virus Strain Ba71V in Soil under Different Conditions.

The survival of African swine fever virus (ASFV) on different matrices and its infectivity in wild as well as domestic swine is still a matter of interest. ASFV is resistant to environmental effects; this fact is enhanced by the presence of organic material. Therefore, the aim of this work was to determine the ability of laboratory ASFV to survive in soil at different temperatures (4 and 22 °C) and with and without the presence of blood using culture procedures. The suitability of the procedure for determining the viability and titre of the ASFV field strain by the hemadsorption method was also verified, when a higher decrease in virus infectivity in the case of clay compared with peat was demonstrated. The stability of the virus was clearly temperature-dependent, the infectious virus was detected after 112 days, and the viral DNA was still detected in the matrix 210 days after inoculation in a relatively high and stable concentration (between 106 and 107 genome equivalents/mL). Based on this knowledge, soil and other environmental samples could provide rapid and reliable information on the disease outbreak and serve as indicators of the risk posed by the affected locality.

Characterisation of Streptococcus suis Isolates in the Czech Republic Collected from Diseased Pigs in the Years 2018-2022.

As in other countries, in the Czech Republic, Streptococcus suis infection in pigs is considered an economically significant disease for the pig industry, though little is known about its population structure. We collected S. suis isolates from 144 farms in the years 2018-2022. All samples were taken from animals suffering from symptoms indicating possible S. suis infection. Serotyping revealed the presence of 23 different serotypes, and 18.94% were non-typable strains. The most common was S7 (14.96%), while other serotypes had frequencies of less than 10%. Sequence typing identified 56 different sequence types, including 31 newly assigned sequence types together with 41 new alleles in genes in the MLST schema. A large portion of isolates (25.70%) were of unknown sequence type. The most common sequence types were ST29 (14.77%) and ST28 (10.04%); the other sequence types had frequencies of less than 10%. In total, 100 different combinations of serotypes and sequence types were identified. Among them, S7ST29 was found in 72 isolates, representing 13.63% of all isolates, and was significantly associated with the central nervous system. Many other isolates of particular serotype and sequence type combinations were found in a few cases, and a number of isolates were non-typable.

Streptococcus suis Isolates-Serotypes and Susceptibility to Antimicrobials in Terms of Their Use on Selected Repopulated Czech Pig Farms.

Streptococcus suis represents a primary health problem (such as meningitis, septicemia and arthritis in piglets and fatteners) in the swine industry worldwide and also an emerging zoonotic pathogen. In the Czech Republic, many pig farms repopulated their herds over the past decades to reduce morbidity and minimize treatment. The study analysed serotypes, sequence types and antimicrobial susceptibility in 39 S. suis isolates obtained from organs of diseased pigs from selected 16 repopulated farms with a history of S. suis-associated diseases and routine antimicrobial treatment with tulathromycin and/or amoxicillin. The analysis revealed diversity of collected isolates with regular occurrence of more than three serotypes per farm. The serotypes identified were 1/2 and 7, each in six isolates, followed by serotype 2 and 3 found in five isolates each, other serotypes were less frequent. Seven isolates were not typable by multiplex PCR and we also found sequence type of unknown type in thirteen isolates. The majority of S. suis isolates were resistant to clindamycin (n = 31), tetracycline (n = 29) and tilmicosin and tulathromycin (n = 28). On the other hand, with the exception of two isolates that were intermediately susceptible to penicillin and ampicillin, all isolates were susceptible to all three tested subgroups of beta-lactam antibiotics.

Wild boar as a potential reservoir of zoonotic tick-borne pathogens.

The wild boar (Sus scrofa) population has increased dramatically over the last decades throughout Europe and it has become a serious pest. In addition, the common habitat of wild boar and of the tick, Ixodes ricinus, indicates the potential of wild boar to play a role in epidemiology of epizootic and zoonotic tick-borne pathogens, including Anaplasma phagocytophilum. In Europe, epidemiological cycles and reservoirs of A. phagocytophilum, including its zoonotic haplotypes, are poorly understood. In this study, we focused on detection and further genetic characterization of A. phagocytophilum and piroplasmids in 550 wild boars from eleven districts of Moravia and Silesia in the Czech Republic. Using highly sensitive nested PCR targeting the groEL gene, the DNA of A. phagocytophilum was detected in 28 wild boars (5.1 %) representing six unique haplotypes. The dominant haplotype was found in 21 samples from 7 different districts. All detected haplotypes clustered in the largest clade representing the European ecotype I and the dominant haplotype fell to the subclade with the European human cases and strains from dogs and horses. Nested PCR targeting the variable region of the 18S rRNA gene of piroplasmids resulted in one positive sample with 99.8 % sequence identity to Babesia divergens. The presence of these two pathogens that are primarily circulated by I. ricinus confirms the local participation of wild boar in the host spectrum of this tick and warrants experimental studies to address wild boar as a reservoir of zoonotic haplotypes of A. phagocytophilum.

Fatal toxoplasmosis in wild European brown hares (Lepus europaeus) in tularaemia endemic areas of the Czech Republic: Poses risk of infection for humans?

Toxoplasma gondii may cause fatal infection in European brown hares (Lepus europaeus). However, the role of this parasite in terms of mortality rate in tularaemia endemic areas, amount of parasites in affected organs and circulating genotypes, is still unknown. In total, 36 hares (killed or found dead) were submitted for pathomorphological examination as a part of the national tularaemia and brucellosis monitoring. Tissue samples (lung, heart, liver, spleen and kidney) were tested by quantitative real-time PCR targeting 529 bp region of T. gondii. Genotyping was performed by a 15 microsatellite markers method in a single multiplex PCR assay. The same tissues of hares were simultaneously used for the bacteriological cultivation. Toxoplasma gondii was detected by qPCR in the tissues of two hares. Spleen and lungs of one infected hare have been found harbouring up to ~7 millions of T. gondii parasites per gram of tissue. Both positive samples were characterized as T. gondii type II, one archetypal clonal type II and the other one a type II variant (W35 = 244). Bacteria Francisella tularensis was proved in pooled samples of three hares but without coinfection with T. gondii; all hares were negative for Brucella suis. Toxoplasma gondii has significant impact on mortality of European brown hares in tularaemia endemic areas and parasite load within the animal tissues may present high risk of human infection.

Resolution of Streptococcus suis Serotypes 1/2 versus 2 and 1 versus 14 by PCR-Restriction Fragment Length Polymorphism Method.

Streptococcus suis is an important pathogen of pigs but is also transmissible to humans, with potentially fatal consequences. Among 29 serotypes currently recognized, some are clinically and epidemiologically more important than others. This is particularly true for serotypes 2 and 14, which have a large impact on pig production and also on human health. Conventional PCR-based serotyping cannot distinguish between serotype 1/2 and serotype 2 or between serotype 1 and serotype 14. Although serotype 1/2 and serotype 2 have a very similar cps locus, they differ in a single-nucleotide substitution at nucleotide position 483 of the cpsK gene. Similarly, serotypes 1 and 14 have a very similar cps locus but also differ in the same nucleotide substitution of the cpsK gene. Fortunately, this cpsK 483G→C/T substitution can be detected by BstNI restriction endonuclease. A PCR-restriction fragment length polymorphism (RFLP) detection method amplifying a fragment of the cpsK gene digested by BstNI restriction endonuclease was developed and tested in reference strains of these serotypes and also in field isolates.