Mitochondrial genome evidence suggests Cooperia sp. from China may represent a distinct species from Cooperia oncophora from Australia

Cooperia spp. are parasitic nematodes parasitizing in small intestine of ruminants with a worldwide distribution. Infection of ruminants with Cooperia species can cause severe enteritis, causing significant socio-economic losses to the livestock industry. However, it is yet to know whether there is genetic diversity in mitochondrial (mt) DNA sequences of Cooperia nematodes from different geographic regions. The objective of the present study was to examine sequence difference in mt genomes between Cooperia sp. from China and other Cooperia species. We determined the sequences of the internal transcribed spacer (ITS-1 and ITS-2) of nuclear ribosomal DNA (rDNA) of 11 Cooperia specimens collected from the small intestine of a Tianzhu White yak in Gansu Province, northwestern China, which had 99% similarity with that of C. oncophora from Brazil (GenBank accession Number: AJ544290) in ITS-1, and 99% similarity with those from Denmark (AB245040), Scotland and Australia (AJ000032) in ITS-2, indicating that specimens used in the present study should at least represent parasites in Cooperia. We then determined the complete mt genome sequences of one representative specimen of Cooperia sp. from China (CspC), compared the mt DNA sequences with that of C. oncophora from Australia (COA, GQ888713), and conducted phylogenetic analysis with selected nematodes using both maximum likelihood (ML) and Bayesian inference (BI) methods based on both concatenated 12 PCGs, rrnL and rrnS sequences and partial cox2 sequences. The complete mt genome sequence of CspC (KY769271) is 13, 583 bp in length, which is 91 bp shorter than that from COA. The sequence difference over the entire mt genome between CspC and COA was 12.2% in nucleotide and 6.3% in inferred amino acids, with nad4L and nad1 being the most variable and the most conserved PCGs, respectively. Phylogenetic analysis indicated that CspC and COA were closely-related but distinct taxa. The determination of mt genome sequences for Cooperia sp. from China also provides novel resources for further studies of taxonomy, systematics and population genetics of Cooperia from different geographical locations.     

Histopathological changes during experimental infections of calves with Cooperia punctata

Eleven male two-month-old Holstein calves were used to determine the pathological changes induced by a Cooperia punctata infection. After weaning, ten calves received a single oral dose of 45,000 C. punctata infective larvae. One calf remained as a non-infected control. Groups of two calves were killed on days 7, 14, 21, 28 and 35 post-infection (p.i.) for determination of worm burdens and histopathological evaluation. The small intestine was sub-divided into three sections of approximately equal length, and representative samples of mucosa were fixed in 10% formalin, cut, and stained with haematoxylin-eosin. Samples of intestinal contents and mucosal digests were taken and fixed in 10% formalin for an estimation of total worm burdens. An increase in the number of adult parasites and a decrease in the number of larvae were observed with time (P<0.001). A higher concentration of worms was found in the first segment of the small intestine during the five weeks of observation. Histology showed larvae in the intestinal mucosa on day 7 p.i., with a discrete increase in the cellular response. Adult worms and a marked cellular infiltrate with eosinophils and neutrophils were present on day 21 p.i., and these persisted until day 35 p.i. Microcysts resulting from worm destruction were observed from day 21 p.i.     

Immune expulsion of the trichostrongylid Cooperia oncophora is associated with increased eosinophilia and mucosal IgA

Previous experiments have shown that a primary infection with 100000 infective larvae of the trichostrongylid Cooperia oncophora allows discrimination between different type of responder animals based on the speed by which the parasite is expelled from the host. In most of the animals (intermediate responders) the expulsion occurs 35-42 days after infection. This experiment was carried out to investigate which mechanisms contribute to the clearance of the parasite from the intestine. Sequential necropsy of the animals 14, 28 and 42 days after infection together with a segmental division of the small intestine, allowed us to characterise essential components associated with development of immunity and expulsion of the parasite from its niche. The results show that during the patent phase of the infection the parasite preferentially resides in the proximal gut. Forty-two days after infection ongoing expulsion is characterised by a migration of the worms to the more distal part of the intestine. Expulsion of the adult worm population appears to be mast-cell independent and is associated with a significant increase in parasite-specific mucous IgA and IgG1 as well as with an influx of eosinophils in the intestinal lamina propria. Although we did not observe a specific lymphocyte recruitment into the intestinal mucosa, the accumulation of eosinophils seems to be mediated by CD4+ cells. We measured significant negative correlations between the number of eosinophils and the expulsion rate of the parasite expressed by sex ratio and ratio eggs per gram faeces. Parasite-specific mucosal IgA levels were negatively correlated to the fecundity of the worms, expressed as number of eggs per female worm. Our results describe the involvement of both eosinophils and mucosal IgA in the regulation of C. oncophora expulsion and suggest the development of a Th2 effector immune response.     

Revision of the African species of the genus Cooperia Ransom, 1907 (nematoda,trichostrongylidae)

Summary Fourteen of the 20 species recorded from Africa are resdescribed and illustrated. Of the remaining six species, four, namely, Cooperia borgesi, C. hippotragusi, C. minor and C. reduncai are considered species inquirendae and one, C. africana, is a possible synonym of C. punctata. The validity of C. oncophora and C. surnabada is discussed. A key to 16 species is given.Part of a thesis approved by the University of London for the award of a Ph.D. degree.Part of a thesis approved by the University of London for the award of a Ph.D. degree.     

Prevalence of Cooperia punctata, C. pectinata and C. oncophora infections in dairy calves in Brazil

Cooperia is the most prevalent helminth parasitizing calves in Brazil. Three species of this genus occur most often: C. punctata, C. pectinata and C. oncophora. Six calves from dairy farms in the south of the State of Minas Gerais aged six to 15 months were killed and necropsied each month over a period of two years. The Cooperia species were identified, counted, and the numbers related to the calves' age. The worm burdens due to the three species of Cooperia were statistically different. C. punctata was the most prevalent species and had a positive correlation with the age of the calves; C. pectinata appeared with lower intensity but was always present, and C. oncophora was not found in calves older than 11 months. These results show the existence of different degrees of resistance to Cooperia species among calves as the three species did not behave similarly. It seems to be an acquired resistance. C. punctata appears to be less immunogenic than C. pectinata and C. oncophora. As C. punctata and C. pectinata are more pathogenic than C. oncophora, it seems that this pathogenicity can be related to immunogenic aspects associated with the species.     

The influence of weather and egg contamination on the development of third-stage larvae of Cooperia oncophora on pasture.

The influence of weather and egg contamination on the dynamics of herbage contamination with infective larave of Cooperia oncophora was investigated on artificially contaminated grass plots and in a grazing experiment with 24 first-year grazing calves from May to October 1987 in Lower Saxony, Germany. On the experimental plots the larval translation was highest at the beginning of July and in the second part of September, following high mean weekly temperatures. Between July and September peak recovery of larvae from herbage occurred 4 weeks after contamination. A seasonal pattern of larval translation similar to that on the experimental plots could be demonstrated on the grazed pastures when the number of larvae per m2 of pasture had been adjusted to the previous egg output by means of a contamination index. The resulting 'relative larval density' is regarded as a good indicator for larval development on pasture. From July to September the larval population on pasture resulted mainly from the egg contamination 2-3 weeks earlier. The short persistence of the infective larvae on herbage was probably due to the frequent and heavy rainfall throughout the season, causing a passive washout of larvae into the soil. On single pastures the larval density started to increase within 1 week after the calves had first contact with these fields. The impact of the calves on the distribution of larvae is discussed.     

T-cell mediated immune responses in calves primary-infected or re-infected with Cooperia oncophora: similar effector cells but different timing

Cooperia oncophora is the most prevalent intestinal nematode of cattle occurring in Western Europe. Primary infection with 100000 third stage infective larvae (L3) induces acquired immunity in a high proportion of the animals but there is little information on immunity against re-infection. In the current experiment, the contribution of the T-cell mediated immunity in protection against re-infection with C. oncophora was investigated in detail. Priming elicited long-lasting protective immunity that was evidenced by a significantly decreased worm burden and egg excretion in primed animals compared to challenge control animals. Lymphocyte proliferation tests with excretory/secretory products (ESP) of C. oncophora and with three distinct ESP fractions indicated an enhanced reactivity in primed animals and suggested that by fractionating of ESP we selected for proteins involved in protective immunity against re-infection with C. oncophora. Phenotypic analysis of T cell subsets at diverse anatomical locations revealed that the enhanced reactivity of lymphocytes from peripheral blood and lymph nodes of the infected animals coincided with a significantly increased frequency of CD4(+) cells at these locations but a deceased frequency of CD4(+) cells in the lamina propria. These findings were independent of the immune status of the animals but more pronounced in the primed animals than in the challenge control animals. In addition we demonstrated that primary and secondary infections with C. oncophora were associated with two waves of eosinophils and that the kinetics of this cell population differed as a result of priming. Based on the observed correlations we propose that the early increase of eosinophils is T cell independent and merely a consequence of inflammation in the parasitised gut. In contrast, the second wave of eosinophils depends upon CD4(+) cells and correlations with parasitological parameters at this time point support a role of eosinophils as effector cells against adult stages of C. oncophora.     

In vitro interaction of Brazilian strains of the nematode-trapping fungi Arthrobotrys spp. on Panagrellus sp. and Cooperia punctata

In vitro tests were carried out to verify the activity of 26 Brazilian isolates of predatory fungi of the genus Arthrobotrys on a free-living nematode (Panagrellus sp.) and on infective larvae of Cooperia punctata, a parasitic gastrointestinal nematode of cattle. The results showed that the free-living nematode Panagrellus sp. was the most preyed upon, compared to C. punctata, for all the fungal treatments. Also, variable predatory capacity was observed for different fungal isolates belonging to the same genus when applied to different nematode species.