The Effects of Honeysuckle (Lonicera caerulea L.) Berry Iridoid-Anthocyanin Extract on the Intestinal and Muscle Histopathology in Mice during Experimental Trichinellosis.

The aim of the study was to determine the effect of iridoid-anthocyanin extract from honeysuckle (Lonicera caerulea L.) (LC) berries on histopathological changes in the intestines and muscles during experimental trichinellosis in mice. The LC extract was administered to uninfected mice (LC group) and Trichinella-spiralis-infected mice (T+LC) orally at a dose of 2 g/kg bw, six times at 24 h intervals, from day 3 prior to infection to day 3 post-infection (dpi). Jejunum samples were collected on 5, 7, 14, and 21 dpi, and their histological assessment involved the villus height to crypt depth ratio (VH/CD), goblet cell (GC) number, and morphological changes. In the T. spiralis-infected muscles, the extent of inflammatory infiltration on the 14th and 21st dpi was assessed. LC in the infected mice restored the VH/CD ratio to control values on 14 dpi. A beneficial effect of the LC extract on the villus height was also observed 14 dpi in the LC and T+LC groups. No differences in the extent of inflammatory infiltration in the muscles between the T+LC and T groups were observed. In conclusion, the iridoid-anthocyanin extract from honeysuckle berry contributed to alleviating the symptoms of the intestinal phase of T. spiralis infection.

The relationship between the intensity of Gasterophilus intestinalis larvae infection and the serum and salivary humoral immune response in horses.

Infection with Gasterophilus intestinalis (botfly) larvae often occurs in horses. The aim of the study was to isolate the larvae of G. intestinalis and evaluate the serum and salivary humoral immune response using self-developed ELISA in G. intestinalis infected horses. Blood serum or saliva samples were taken from 125 infected horses and 54 uninfected slaughtered horses. The antigens from G. intestinalis larvae were used for development of ELISA in order to evaluate the intensity of G. intestinalis IgG, IgM, and IgA antibody reactivity in the serum or saliva of naturally infected horses and horses without larvae in the gastrointestinal tract (control group). Serum antibodies against second and third larvae's stadium antigens reacted significantly more intensively in infected than in healthy horses in IgG (p ≤ 0.001; p ≤ 0.05, respectively) and IgA (p ≤ 0.05;p ≤ 0.001, respectively) classes. Salivary IgG and IgA specific's antibody reactivity was significantly higher in horses with moderate (p ≤ 0.01) and severe infection (p ≤ 0.001) compared to the healthy horses. The determination of the G. intestinalis IgG and IgA antibody activity in saliva and serum may be used for detecting horses moderately and severely infested with larvae.

Evaluation of Immunotropic Activity of Iridoid-Anthocyanin Extract of Honeysuckle Berries (Lonicera caerulea L.) in the Course of Experimental Trichinellosis in Mice.

Our experiment determined the immunotropic activity of a natural, iridoid-anthocyanin extract from honeysuckle berry (Lonicera caerulea L.) (LC). The extract was administered to mice infected with Trichinella spiralis, orally at a dose of 2 g/kg bw, six times at 24 h intervals (from day 3 prior to the infection to day 3 post-infection (dpi) with T. spiralis. At 5, 7, 14, and 21 dpi, samples of blood, spleen, and mesenteric lymph nodes (MLN) were collected, and isolated lymphocytes were analyzed by flow cytometry. The splenocyte proliferation was estimated with MTT testing, and the intensity of intestinal and muscle infection was also studied. LC stimulated the local immune system by inducing lymphocyte proliferation in the spleen 7 dpi and altered the percentage and absolute count of B (CD19+) and T (CD3+, CD8+) cells 7, 14, and 21 dpi in the peripheral blood. LC extract affected the dynamics of expulsion of adult Trichinella from the intestines and prolonged the intestinal phase of the infection but did not change the number of larvae in the muscles. These results suggest that Lonicera caerulea L. fruit extract modulates murine cellular immune response during intestinal phase of T. spiralis infection but shows no antiparasitic activity.

Cryptosporidium spp. in dogs and cats in Poland.

INTRODUCTION: Cryptosporidium spp. are common protozoan parasites of animals and humans. Due to their zoonotic potential it is important to know their species and prevalence in dogs and cats. OBJECTIVE: The aim of the study was to determine the occurrence and molecular characteristics of Cryptosporidium spp. in dogs and cats in Poland. MATERIAL AND METHODS: A total of 365 faecal samples (264 dogs and 101 cats) collected from animals living in Poland were analyzed using the Ziehl-Neelsen staining method and genus-specific PCR assay to amplify the Cryptosporidium 18S rRNA gene. RESULTS: Cryptosporidium were found in 11 out of the 365 examined stool samples (3%). PCR analysis identified Cryptosporidium in 9 out of 264 canine stool samples (3.4%) and 2 out of 101 feline specimens (2%). DNA sequencing confirmed the presence of C. canis and C. parvum in dogs and C. felis in cats. CONCLUSIONS: This is the first molecular characterization of Cryptosporidium spp. infection in dogs and cats in Poland.

Effects of Selected Prebiotics or Synbiotics Administered in ovo on Lymphocyte Subsets in Bursa of the Fabricius, Thymus, and Spleen in Non-Immunized and Immunized Chicken Broilers.

The effects of in ovo-delivered prebiotics and synbiotics on the lymphocyte subsets of the lymphoid organs in non-immunized 7-day-old broiler chickens and in non-immunized, sheep red blood cells (SRBC)-immunized, and dextran (DEX)-immunized 21- and 35-day-old birds were studied. The substances were injected on the 12th day of egg incubation: Prebiotic1 group (Pre1) with a solution of inulin, Prebiotic2 group (Pre2) with a solution of Bi2tos (non-digestive transgalacto-oligosaccharides), Synbiotic1 group (Syn1) with inulin and Lactococcus lactis subsp. lactis IBB SL1, and Synbiotic2 group (Syn2) with Bi2tos and Lactococcus lactis subsp. cremoris IBB SC1. In 7-day-old chicks, a decrease in T splenocytes was noticed in all groups. The most pronounced effect in 21- and 35-day-old birds was an increase in TCRγδ+ cells in Syn1 and Syn2 groups. A decrease in bursal B cells was observed in DEX-immunized Pre1 group (21-day-old birds), and in the Syn1 group in non-immunized and SRBC-immunized 35-day-old birds. An increase in double-positive lymphocytes was observed in Pre1 (35-day-old birds) and Pre2 (immunized 21-day-old birds) groups. In Pre1 and Syn1 groups (21- and 35-day-old), an increase in B splenocytes and a decrease in T splenocytes were observed. We concluded that Syn1 was the most effective in the stimulation of the chicken immune system.

Genotypes of Giardia duodenalis in Household Dogs and Cats from Poland.

BACKGROUND: Giardia duodenalis is a widespread protozoan parasite affecting humans and many species of animals, including dogs and cats. Due to its zoonotic potential, it is important to know the frequency of this parasite in companion animals. The aim of this study was to determine current epidemiological status of G. duodenalis in household dogs and cats. METHODS: In this study, 293 fecal samples from pet dogs and cats were collected from January 2017 to July 2019 and tested for G. duodenalis by PCR (using ß-giardin gene). The animals were divided into groups depending on their age, breed and fecal consistency. RESULTS: The examination allowed for detection of G. duodenalis in 6.0% of canine and 3.9% of feline fecal samples. The highest frequency was revealed in young (under one-year old) dogs. Sequencing confirmed the presence of assemblages C and D in dogs and A and F in cats. CONCLUSION: The study showed current frequency of G. duodenalis in dogs and cats and also revealed the occurrence of host-specific assemblages as well as zoonotic assemblage A.

Evaluation of the protective effect of immunization spf DBA/2J mice with selected bacterial, recombinant Hsp60 antigens during Salmonella Enteritidis challenge.

Salmonella Enteritidis is one of the most common causes of food poisoning in humans. Many attempts have been made to develop an effective vaccine against S. Enteritidis for use in poultry, but experiments aimed at the complete elimination of this pathogen from poultry farms have not provided satisfactory results. The development of new generation vaccines against salmonellosis, such as subunit vaccines based on heat shock proteins (HSPs), is strongly justified. The high immunogenicity of Hsp60 isolated from Procaryota, including Salmonella, has been suggested by the presence of IgG anti-Hsp60 antibodies in mice immunized with these proteins. The aim of the studies was to evaluate the protective effects of immunization with recombinant Hsp60 from selected gram-negative bacteria (S. Enteritidis, Escherichia coli, Pasteurella multocida, Histophilus somni) in spf DBA/2 J mice experimentally infected with S. Enteritidis. The study demonstrated that double subcutaneous immunization of mice with a dose of 10 µg rHsp60 induced a specific immune response of IgG antibodies in tested animals. The median lethal dose (LD50) for the murine model spf DBA/2 J orally infected with S. Enteritidis was estimated at 6.84 × 105 cfu/animal. Mice immunized with rHsp60 from gastrointestinal pathogens (S. Enteritidis and E. coli) showed better survival after experimental infection with a 3 × LD50 dose from S. Enteritidis, compared to animals immunized with proteins obtained from respiratory pathogens (P. multocida and H. somni). However, the log-rank analysis did not show significant differences in the survival rates between rHsp60-immunized mice and controls. S. Enteritidis was not isolated any less frequently from internal organs and faeces of rHsp60-immunized mice than from controls. Nevertheless, the level of haptoglobin (but not IL-6) was increased in all mice in which the presence of the pathogen was observed. Bacterial Hsp60 is an interesting candidate for a subunit vaccine, but its use in livestock animals must be further investigated.

Low-Level Light Therapy Protects Red Blood Cells Against Oxidative Stress and Hemolysis During Extracorporeal Circulation.

Aim: An activation of non-specific inflammatory response, coagulation disorder, and blood morphotic elements damage are the main side effects of the extracorporeal circulation (ECC). Red-to-near-infrared radiation (R/NIR) is thought to be capable of stabilizing red blood cell (RBC) membrane through increasing its resistance to destructive factors. We focused on the development of a method using low-level light therapy (LLLT) in the spectral range of R/NIR which could reduce blood trauma caused by the heart-lung machine during surgery. Methods: R/NIR emitter was adjusted in terms of geometry and optics to ECC circuit. The method of extracorporeal blood photobiomodulation was tested during in vivo experiments in an animal, porcine model (1 h of ECC plus 23 h of animal observation). A total of 24 sows weighing 90-100 kg were divided into two equal groups: control one and LLLT. Blood samples were taken during the experiment to determine changes in blood morphology [RBC and white blood cell (WBC) counts, hemoglobin (Hgb)], indicators of hemolysis [plasma-free hemoglobin (PFHgb), serum bilirubin concentration, serum lactate dehydrogenase (LDH) activity], and oxidative stress markers [thiobarbituric acid reactive substances (TBARS) concentration, total antioxidant capacity (TAC)]. Results: In the control group, a rapid systemic decrease in WBC count during ECC was accompanied by a significant increase in RBC membrane lipids peroxidation, while in the LLLT group the number of WBC and TBARS concentration both remained relatively constant, indicating limitation of the inflammatory process. These results were consistent with the change in the hemolysis markers like PFHgb, LDH, and serum bilirubin concentration, which were significantly reduced in LLLT group. No differences in TAC, RBC count, and Hgb concentration were detected. Conclusion: We presented the applicability of the LLLT with R/NIR radiation to blood trauma reduction during ECC.

Effects of iridoid-anthocyanin extract of Cornus mas L. on hematological parameters, population and proliferation of lymphocytes during experimental infection of mice with Trichinella spiralis.

The influence of iridoid-anthocyanin aqueous extract of cornelian cherry fruits (CM) on hematological parameters, lymphocyte subsets and proliferation during Trichinella spiralis infection in mice was investigated. CM (100 mg/kg) was administered orally to T. spiralis-infected mice six times within a period encompassing three days prior to the infection and three days after the infection (dai). CM increased the percentage of CD3+, CD4+ cells and CD4+/CD8+ ratio and decreased total count of CD8+ and CD19+ splenocytes (5th dai). An increase in total count of CD4+, CD3+, CD19+ splenocytes was observed (21st dai). CM elevated the percentage of CD4+ cells (7th dai) and CD4+/CD8+ ratio (21st dai) in MLN. CM increased (14th dai) and then reduced (21st dai) the percentage of CD8+ MLN lymphocytes and decreased total count of MLN CD8+ cells (21st dai) and B cells (14th dai). An activation of lymphocyte proliferation in spleen and simultaneous decrease in MLN on 5th dai was observed. An increase in red blood cells parameters (5th dai) and in leukocyte count (7th dai) was found. A rise in platelet count was noticed both on 5th and 7th dai. Moreover, the number of adult T. spiralis on 5th dai in mice receiving CM extract was lower than in the control mice. These results suggested that iridoid-anthocyanin aqueous extract of CM stimulated murine immune response during T. spiralis infection.

The evaluation of immunogenic impact of selected bacterial, recombinant Hsp60 antigens in DBA/2J mice.

Heat Shock Proteins (HSP) are highly conserved proteins that are widely spread throughout all organisms. They function in the cytoplasm as chaperones; however, they could be expressed on the cell surface. It has been shown that Hsp60 obtained from gram-negative bacteria are able to stimulate cells of the acquired and innate immune system. The aim of this study was the evaluation of the immunogenic properties of recombinant Hsp60 proteins derived from four common pathogenic bacteria: Escherichia coli, Histophilus somni, Pasteurella multocida and Salmonella Enteritidis. The analysis of the humoral immune response in DBA/2J mice hyperimmunized with selected rHsp60 revealed high levels of IgG rHsp60-antibody with the predominance of the IgG1 subclass, in the reaction with both homologous and heterologous antigens. The presence of IgG2a and IgG2b was also observed; however, no antibodies of subclass IgG3 were detected. The comparison of plasma IgG antibody reactivity of mice immunized with two different doses of rHsp60 (10/20 µg) showed that the lower dose was sufficient to induce a strong humoral response. The reactivity of the IgG rHsp60-antibody with whole bacterial cells showed a significantly higher reaction with H. somni compared with other pathogens. It was demonstrated that the addition of all rHsp60 with polymyxin B to the culture medium stimulated splenocytes isolated from hyperimmunized mice to release IL-1ß and IL-6. As a strong stimulator of the immune system, bacterial-origin Hsp60 seems to be an interesting potential component of subunit vaccines aimed at the development of protection for animals during infections caused by gram-negative bacteria.

Zoonotic microsporidia in dogs and cats in Poland.

This study investigated the prevalence, genetic diversity, and zoonotic concerns of microsporidia in household dogs and cats in Poland. A total of 126 (82 dogs and 44 cats) fecal specimens were analyzed for the presence of specific DNA of Enterocytozoon bieneusi and Encephalitozoon spp. using a nested PCR protocol amplifying the internal transcribed spacer region of the rRNA gene. Microsporidia were found in 10 (7.9%) out of the 126 examined stool samples. Of the 82 dogs, 4 (4.9%) and 2 (2.4%) were positive for E. bieneusi (genotypes D and PtEbIX) and Encephalitozoon cuniculi genotype II, respectively. Of the 44 cats, 4 (9.1%) were positive for E. bieneusi (genotypes PtEbIX and eb52). Additionally, one cat (2.3%) was concurrently infected with E. bieneusi (PtEbIX) and E. cuniculi (genotype II). Considering that all detected microsporidia in dogs and cats have been previously associated with human microsporidiosis, companion animals may be a potential source of microsporidia infections in humans.

Molecular identification of Giardia duodenalis isolates from domestic dogs and cats in Wroclaw, Poland.

INTRODUCTION: Giardia duodenalis (G. intestinalis) is a common protozoan causing gastrointestinal disorders in many species of mammals. The genus of Giardia has high molecular diversity. Dogs and cats, in addition to their typical infection with assemblages C, D and F, may be a reservoir of zoonotic assemblages (A and B). OBJECTIVE: The aim of this study was a genetic characteristic of Giardia isolates of dogs and cats from the area of Wroclaw (Poland). MATERIALS AND METHOD: A total of 128 and 33 faecal samples from dogs and cats, respectively, were analyzed by routine coprological methods. The animals were diagnosed on the presence of G. duodenalis antigens in faeces soluble with the use of SNAP Giardia (IDEXX Laboratories) immunosorbent assay. 27 DNA isolates of Giardia were subjected to molecular identification (PCR-RFLP). RESULTS AND CONCLUSIONS: The prevalence of G. duodenalis was 21.1% (27/128) in dogs and 15.1% (5/33) in cats. In dogs, C assemblage was present in 18 (81%) positive stool samples, D assemblage in 2 (9%) samples, B assemblage present in one (4.5%), and mixed assemblages (C and D) occurred in one (4.5%) sample. F assemblage was found in 4 (80%) cats' positive stool samples and A assemblage occurred in one case (20%). Confirmation of the presence of A and B zoonotic assemblages suggests that infected pets can be a threat to human health. This study describes for the first time the presence of mixed infections within host-specific C and D assemblages in dogs in Poland.

Occurrence of tapeworms of the family Anoplocephalidae in herds of dairy cattle in Lesser Poland and in Lower Silesia, Poland.

This study was performed to determine the tapeworm infection of grazing cattle in 11 dairy herds in Lower Silesia and Lesser Poland (Galicia). Rectal faecal samples were examined microscopically for tapeworm eggs by Willis-Shlaaf's flotation, Telemann's sedimentation (for fatty stools) and decantation method. Out of 182 cattle, 10 (5.5%) were found to be infected with tapeworms. The prevalence in 5 of 11 examined herds varied from 15.5 to 30.5% and was highest for animal being in the first lactation. Tapeworm eggs were detected in animals being in the first and second lactation (about three- to four-years-old cows). All the qualitative methods were effective in the diagnosis of tapeworm infection in cattle.