Elsevier

Veterinary Parasitology

Volume 288, December 2020, 109277
Veterinary Parasitology

Acute phase protein pattern and antibody response in pigs experimentally infected with a moderate dose of Trichinella spiralis, T. britovi, and T. pseudospiralis

https://doi.org/10.1016/j.vetpar.2020.109277Get rights and content

Highlights

  • APPs in pigs infected with a moderate dose of various Trichinella spp. were measured.

  • Increases of Pig-MAP, CRP and Hp levels were mild, fugacious and irrelevant.

  • Kinetics of anti-Trichinella IgG in particular experimental groups was evaluated.

  • Anti-Trichinella IgM was detected on day 30 pi only in T. spiralis infected swine.

Abstract

The aim of the present study was to evaluate the acute-phase protein (APP) response in three groups of pigs experimentally infected with a moderate infective dose, i.e. 1000 muscle larvae (ML) of Trichinella spiralis, 3000 ML of Trichinella britovi, and 2000 ML of Trichinella pseudospiralis. Over a 62-day period of infection, we examined the serum level and kinetics of the haptoglobin (Hp), C-reactive protein (CRP), serum amyloid A (SAA), and pig major acute-phase protein (pig-MAP). In addition, to better understand the immune response of pigs experimentally infected with three different species of Trichinella, the kinetics of IgG and IgM antibodies against excretory-secretory (ES) antigens of Trichinella ML were also investigated. In order to assess anti-Trichinella IgG dynamics, we used a commercial and an in-house ELISA based on both heterologous (T. spiralis) and homologous (T. spiralis, T. britovi, and T. pseudospiralis) Trichinella species ES antigens. Among the four APPs analyzed, the concentration of CRP and pig-MAP significantly increased only in T. britovi-infected swine when compared with control pigs. This took place as early as 6 days post-infection (dpi). Hp was the only APP whose concentration significantly increased in pigs infected with T. pseudospiralis, this occurring as late as on day 62 pi. Despite the statistical differences found, increases in pig-MAP, CRP, and Hp levels were rather mild and transitory; none of these proteins were found to be elevated in the serum of all experimental groups of pigs at the same time point after infection. Specific IgG antibodies against ES antigens of Trichinella ML were first detected by the commercial and in-house T. spiralis ML ES-antigen ELISAs on days 30, 36 and 36 pi in pigs experimentally infected with T. spiralis, T. britovi, and T. pseudospiralis, respectively. However, seroconversion in pigs experimentally infected with T. britovi was detected slightly earlier (30 dpi) when the ELISA based on homologous rather than heterologous ES antigens was applied. In serum samples from pigs infected with T. spiralis, statistically significant increases in the level of specific IgM antibodies against T. spiralis ML ES antigens were first detected on day 30 pi and after this time, their concentration began to decrease. No changes in the level of anti-Trichinella IgM were observed in T. britovi- or T. pseudospiralis-infected pigs throughout the entire period of the experiment.

Introduction

Trichinella spp., a tissue-dwelling parasitic nematode, is capable of inducing a zoonotic disease (trichinellosis) detrimental to human health and life. Many previous reports have described the parasite’s life cycle in detail; in a simplified way: the infection occurs after consumption of raw or undercooked meat infected with first-stage (L1) invasive muscle larvae (ML) of Trichinella. Globally, pork meat and/or its products are identified as one of the most important sources of human infections and in this context, backyard pigs, free-ranging pigs or any herds kept under non-controlled management conditions currently pose the greatest risk to public health (Murrell and Pozio, 2011; Pozio, 2014).

To date, within the genus of Trichinella, ten species (T. spiralis, T. nativa, T. britovi, T. pseudospiralis, T. murrelli, T. nelsoni, T. papuae, T. zimbabwensis, T. patagoniensis, and Trichinella T13) and three additional genotypes (Trichinella T6, Trichinella T8, and Trichinella T9) have been recognized, molecularly characterized, scientifically described and classified into two clades: encapsulated and non-encapsulated (Pozio and Zarlenga, 2005; Krivokapich et al., 2012; Sharma et al., 2019). Among the aforementioned species and genotypes, T. spiralis shows the highest adaptation to domestic pigs, and this phenomenon has been reflected in the long-term studies conducted by the International Trichinella Reference Center. According to this research, 91 % of all European isolates of T. spiralis tested originated from pigs or wild boars (Pozio, 2019). In addition, infections caused by T. britovi and T. pseudospiralis were also confirmed in pigs in Europe. However, it should be emphasized that natural T. pseudospiralis infections in swine were limited to several outbreaks in some European countries, namely, Slovakia, Croatia, Bosnia and Herzegovina, and Spain (Hurníková et al., 2005; Beck et al., 2009; Santrac et al., 2015; Zamora et al., 2015).

Modern veterinary practice, especially when it is involved in the swine production process, employs active measures and standardized tools to monitor and evaluate the health status of animals at the herd level. One such approach which has been developed over the past few years includes the assessment of the level of selected serum acute-phase proteins (APPs). The acute-phase response is defined as an innate immune reaction which appears shortly after tissue injuries in the course of, among others, trauma, inflammation, or tumor growth. Generally, acute-phase proteins are synthesized in the liver and their production is driven by pro-inflammatory molecules forming two groups; namely, IL1- and IL-6-type cytokines (Petersen et al., 2004). From a diagnostic point of view, out of many positive APPs that have been thoroughly studied in pigs, four play an important roles, and they are haptoglobin (Hp), C-reactive protein (CRP), serum amyloid A (SAA), and pig major acute-phase protein (pig-MAP). Several previous reports showed an increase of some of these proteins in the serum of swine suffering from various viral (H3N2 or H1N1 swine influenza A virus, PRRSV, or ASFV), bacterial (Streptococcus suis, Bordetella bronchiseptica, Actinobacillus pleuropneumoniae, or Mycoplasma hyorhinis) or parasitic (Toxoplasma gondii) diseases (Heegaard et al., 1998; Jungersen et al., 1999; Magnusson et al., 1999; Sorensen et al., 2006; Carpintero et al., 2007; Pomorska-Mól et al., 2011, 2012a,b, 2014; Saco et al., 2016). In addition, some research results, most often published as case reports, showed a specific CRP level increase in the serum of humans infected with T. spiralis, T britovi, or Trichinella T9 (Clausen et al., 1996; Tint et al., 2009; Paraličová et al., 2013; Gvozdenovic et al., 2014; Popović-Dragonjić and Kocić, 2018; Tada et al., 2018). Contrary to these studies, an elevated level of CRP was found in the serum of Sprague Dawley rats experimentally infected with Nippostrongylus brasiliensis but not with T. spiralis (Stadnyk et al., 1990). At the same time it should be noted that APP reactant level measurement does not make it possible to diagnose a specific disease but only indicates pathological (inflammatory) changes; however, it is important to distinguish which of them should be taken into account when APP increase occurs. To the best of our knowledge, there is no published literature regarding the effect of infection with various Trichinella species on the level of selected APPs in serum of infected swine. Furthermore, our previous study using a combination of two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry showed that infection with three different species of Trichinella was able to induce changes in the serum proteomic profile of experimentally infected pigs (Gondek et al., 2020). Unfortunately, several serum proteins were not successfully identified in that trial.

The present study evaluates the acute-phase protein pattern and antibody response in pigs experimentally infected with a moderate dose (i.e. 1000–3000 ML) of T. spiralis, T. britovi, and T. pseudospiralis.

Section snippets

Parasites, animals and study design

We examined the serum concentration and kinetics of the positive acute-phase proteins Hp, CRP, SAA and pig-MAP through a 62-day period of infection. In addition, to better understand the immune response of pigs experimentally infected with three different species of Trichinella, the kinetics of IgG and IgM antibodies against ES antigens of Trichinella muscle larvae were also evaluated. Previous data showed a slight supremacy of homologous ES antigens derived from the infecting species of

Intensity of Trichinella larvae infection in muscles of pigs experimentally infected with T. spiralis, T. britovi, and T. pseudospiralis

The results for determination of the intensity of Trichinella ML infection (lpg) in the muscles of pigs experimentally infected with T. spiralis, T. britovi, and T. pseudospiralis are shown in Fig. 1.

Muscle larvae of Trichinella were isolated from all pigs experimentally infected with T. spiralis, T. britovi, and T. pseudospiralis. Trichinella larvae were not detected in muscles sampled from control pigs. Our studies confirmed that T. spiralis shows the highest infectivity to domestic pigs,

Discussion

Our recent study showed that infection with T. spiralis, T. britovi, and T. pseudospiralis is able to induce changes in the serum proteomic profile of experimentally infected pigs. It was also found that proteins involved in lipid metabolism, immune response, blood coagulation, collagen production or muscle differentiation were differentially expressed in porcine serum depending on the Trichinella species infecting the animal (Gondek et al., 2020).

Among the four APPs analyzed, the concentration

CRediT authorship contribution statement

Michał Gondek: Conceptualization, Methodology, Funding acquisition, Writing - original draft, Project administration, Data curation, Resources. Przemysław Knysz: Methodology, Software. Małgorzata Pomorska-Mól: Methodology, Writing - review & editing. Monika Ziomek: Writing - review & editing. Justyna Bień-Kalinowska: Methodology, Writing - review & editing.

Declaration of Competing Interest

The authors report no declarations of interest.

Acknowledgments

This research was partly funded by the National Science Centre in Poland: grant number DEC-2017/01/X/NZ6/00582 and grant number UMO-2015/18/E/NZ6/00502.

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