Morphometric and Molecular Analysis of Five-Spine Epidinium Morphotypes Taken from the Rumen of European Bison, Bison bonasus.

An important feature of ruminal ciliates is their phenotypic plasticity, which makes their identification difficult. The common manifestation of the phenotypic plasticity in rumen ciliates is a change in their cell size and caudal spination. We analyzed various morphotypes of Epidinium with five caudal processes (spines) taken from the rumen of European bison (Bison bonasus). In the study, the cluster analysis and K-means analysis of morphometric data could not distinguish very similar morphotypes of Epidinium with five caudal processes. However, the morphotype of E. parvicaudatum prevailed (70%). The DNA of four individual E. parvicaudatum was isolated successfully from formaldehyde-preserved samples. The partial 18S rDNA gene sequences (about 350-400 bp) were identical to Epidinium sequences in GenBank (E. caudatum, a one-spine morphotype, and E. cattanei, a five-spine morphotype). It can be assumed that these short sequences cannot distinguish the differences between the Epidinium morphospecies. Complete gene sequences from various hosts and various molecular markers are necessary to reveal the validity of the Epidinium five-spine species. In conclusion, classical morphology should be supplemented with molecular data when more morphotypes of the rumen ciliate species are present in samples.

The Application of MALDI-TOF MS for a Variability Study of Paenibacillus larvae.

In recent decades, the significant deterioration of the health status of honey bees has been observed throughout the world. One of the most severe factors affecting the health of bee colonies worldwide is American foulbrood disease. This devastating disease, with no known cure, is caused by the Gram-positive spore-forming bacteria of Paenibacillus larvae species. At present, DNA-based methods are being used for P. larvae identification and typing. In our study, we compare two of the most advanced DNA-based technologies (rep-PCR and 16S rRNA analyses) with MALDI-TOF MS fingerprinting to evaluate P. larvae variability in Central Europe. While 16S rRNA analysis presents a very limited variation among the strains, MALDI-TOF MS is observed to be more efficient at differentiating P. larvae. Remarkably, no clear correlation is observed between whole-genome rep-PCR fingerprinting and MALDI-TOF MS-based typing. Our data indicate that MALDI-TOF protein profiling provides accurate and cost-effective methods for the rapid identification of P. larvae strains and provides novel perspectives on strain diversity compared to conventional DNA-based genotyping approaches. The current study provides a good foundation for future studies.

Can the foregut nematode Haemonchus contortus and medicinal plants influence the fecal microbial community of the experimentally infected lambs?

The abomasal parasitic nematode Haemonchus contortus can influence the abomasal microbiome of the host. On the other hand, no information occurs on the influence of the parasite on the hindgut microbiome of the host. We evaluated the impact of Haemonchus contortus on the fecal microbial community of the experimentally infected lambs treated with a mixture of medicinal herbs to ameliorate the haemonchosis. Twenty-four female lambs were divided into four groups: infected animals (Inf), infected animals supplemented with a blend of medicinal herbs (Inf+Herb), uninfected control animals (Control), and uninfected animals supplemented with medicinal herbs (C+Herb). Inf and Inf+Herb lambs were infected orally with approximately 5000 L3 larvae of a strain of H. contortus susceptible to anthelmintics (MHco1). Herb blend (Herbmix) consisted of dry medicinal plants of Althaea officinalis, Petasites hybridus, Inula helenium, Malva sylvestris, Chamomilla recutita, Plantago lanceolata, Rosmarinus officinalis, Solidago virgaurea, Fumaria officinalis, Hyssopus officinalis, Melisa officinalis, Foeniculum vulgare, and Artemisia absinthium. Each animal was fed meadow hay and a commercial concentrate (600 + 350 g DM/d). Inf+Herb and C+Herb lambs were fed Herbmix (100 g DM/d and animal). Treatment lasted for 50 days. The fecal microbial fermentation parameters (short-chain fatty acids, ammonia, and pH) were evaluated at intervals of 0, 20, 32, and 50 days. The fecal eubacterial populations were evaluated by denaturing gradient gel electrophoresis (DGGE) at day 32 when H. contortus infection was the highest. No substantial effects of the H. contortus infection and the herbal treatment on fecal microbial fermentation parameters and fecal eubacterial populations were observed. Evaluation of DGGE patterns by Principal component analysis pointed to the tendency to branch the C+Herb group from the other experimental groups on Day 32. The results indicate that hindgut microbial activity was not disturbed by H. contortus infection and herbal treatment.

Natural chemotherapeutic alternatives for controlling of haemonchosis in sheep.

BACKGROUND: Parallel in vitro and in vivo experiments were designed to evaluate promising chemotherapeutic alternatives for controlling haemonchosis in ruminants. In vitro anthelmintic activities (egg hatch test - EHT; larval development test - LDT) of aqueous and methanolic herbal extracts Mix1 and Mix2 were investigated. The in vivo effects of dietary supplementation with Mix1 and Mix2 on the parasitological status, inflammatory response, antioxidant parameters and microbial community of the lambs infected experimentally with Haemonchus contortus were investigated. Lambs were divided into four groups for the in vivo study: uninfected control lambs (C), infected lambs (I), infected lambs supplemented with Mix1 (I + Mix1) and infected lambs supplemented with Mix2 (I + Mix2). The experimental period was 70 days. RESULTS: The number of eggs per gram (EPG) of feces was quantified 22, 30, 37, 44, 51, 58, 65 and 70 days post-infection, and mean abomasal worm counts were assessed 70 days post-infection. Quantitative analyses identified 57.3 and 22.2 mg/g phenolic acids, 41.5 and 29.5 mg/g flavonoids and 1.4 and 1.33 mg/g protoberberine-type alkaloids in Mix1 and Mix2, respectively. The methanolic extracts of the herbal mixtures in both in vitro tests had higher anthelmintic effects (P < 0.01) than the aqueous extracts, but the effects did not differ significantly between Mix1 and Mix2 (P > 0.05). I + Mix1 and I + Mix2 lowered mean EPGs between 44 and 70 d by 58.1 and 51.6%, respectively. The level of IgG antibodies against H. contortus increased significantly after infection in each infected group. CONCLUSION: These results represent the first monitoring of the in vitro anthelmintic effects of herbal mixtures on H. contortus. The in vivo experiment indicated that the anthelmintic effect was not sufficient for the elimination of parasites, but this herbal treatment may affect the host over a longer term, reducing the parasitic infection in the host.